United States
Environmental Protection
Agency
Environmental Research
Laboratory
Athens GA 30605
EPA-8 '9-039
April 1979
Fate and Biological
Effects of Cadmium
Introduced into
Channel
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RESEARCH REPORTING SERIES
Research reports of the Office of Research and Development, U.S. Environmental
Protection Agency, have been grouped into nine series. These nine broad cate-
gories were established to facilitate further development and application of en-
vironmental technology. Elimination of traditional grouping was consciously
planned to foster technology transfer and a maximum interface in related fields.
The nine series are:
1 Environmental Health Effects Research
2 Environmental Protection Technology
3. Ecological Research
4 Environmental Monitoring
5 Socioeconomic Environmental Studies
6. Scientific and Technical Assessment Reports (STAR)
7 Interagency Energy-Environment Research and Development
8. 'Special" Reports
9 Miscellaneous Reports
This report has been assigned to the ECOLOGICAL RESEARCH series. This series
describes research on the effects of pollution on humans, plant and animal spe-
cies, and materials. Problems are assessed for their long- and short-term influ-
ences Investigations include formation, transport, and pathway studies to deter-
mine the fate of pollutants and their effects. This work provides the technical basis
for setting standards to minimize undesirable changes in living organisms in the
aquatic, terrestrial, and atmospheric environments.
This document is available to the public through the National Technical Informa-
tion Service, Springfield, Virginia 22161.
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EPA-600/3-79-039
April 1979
FATE AND BIOLOGICAL EFFECTS OF CADMIUM
INTRODUCED INTO CHANNEL MICROCOSMS
by
John P. Giesy, Jr., Henry J. Kania, John W. Bowling,
Robert L. Knight, Susan Mashburn, and Susan Clarkin
Savannah River Ecology Laboratory
Institute of Ecology
University of Georgia
Drawer E
Aiken, South Carolina 29801
Interagency Agreement
IAG-D6-0369-1
between
U. S. Environmental Protection Agency
and
U. S. Department of Energy
Project Officer
Harvey W. Holm
Environmental Systems Branch
Environmental Research Laboratory
Athens, Georgia 30605
ENVIRONMENTAL RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U. S. ENVIRONMENTAL PROTECTION AGENCY
ATHENS, GEORGIA 30605
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DISCLAIMER
This report has been reviewed by the Environmental Research Laboratory,
U. S. Environmental Protection Agency, Athens, Georgia and approved for
publication. Approval does not signify that the contents necessarily reflect
the views and policies of the U. S. Environmental Protection Agency, nor does
mention of trade names or commercial products constitute endorsement or
recommendation for use.
ii
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FOREWORD
Environmental protection efforts are increasingly directed towards
preventing adverse health and ecological effects associated with specific
compounds of natural or human origin. As part of this Laboratory's research
on the occurrence, movement, transformation, impact, and control of envi-
ronmental contaminants, the Environmental Systems Branch studies complexes
of environmental processes that control the transport, transformation,
degradation, and impact of pollutants or other materials in soil and water
and assesses environmental factors that affect water quality.
Environmental concentrations of cadmium, which is known to be acutely
and chronically toxic to plants and animals, have increased significantly
since 1945 as a result of its widespread use in many industrial processes and
products. Efforts to limit human exposure must rest on a good understanding
of the cycling of the metal in fresh water, estuary, and marine ecosystems
and biota before its significance to water pollution can be assessed. Although
many studies have been conducted on the uptake of cadmium by organisms in the
laboratory, few studies have been made in complex environments. This report
describes the results of a study of the fate and biological effects of chronic
concentrations of the metal over a number of trophic levels during the entire
growing season in a complex, artificial aquatic ecosystem.
David W. Duttweiler
Director
Environmental Research Laboratory
Athens, Georgia
iii
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ABSTRACT
Cadmium was continuously input to aquatic microcosm channels resulting
in commit i -it ions of 5 and 10 |jg Cd/1. Cadmium accumulation into both biotic
and abiotic component* was determined. Biological effect* of cadmium were
determined hy monitoring atructuial and functional properties of the entire
system aw well a! structual changes in population! and compared to control
whi«h recHved no cadmium.
Cadmium inputs and output u equilibrated within approximately 20 daya of
initial cadmium inputs. However, approximately 20% of the cadmium leaving
the channels waa associated with par I iculatea. Community component a accumu-
lated cadmium proportional to cadmium exposure levels, Idquil ibrium Cd con-
rent rations of sediment!, periphyton, macrophytes, chlrunomida and mosquito
Hah rxpoaed to 10 (jg Cd/1 were 0.5'J, 55, 250, 40, and 40 MM Cd/g dry weight.
Cadmium wan rapidly eliminated from ail biotic components, with coneentra-
tiona returning to levels similar to those in control channel! within a few
weeks in thr aufwiuhs community to a few months in nrncrophytea. Organic
headpool sedimenta showed no significant decrease in cadmium content six
months aftei rtiasatlon of i admium inputs, Indicating that the abiotic half
time for contaminated environments is very long. Half times for elimination
from (harinel sediments were 72 and 38 daya (or "> and 10 (Jg/I inputs, respec-
tively after (M inputs were terminated,
Cadmium < <*used significant changes in both community itructure and
function. Some proto/oan, crustacean and insect taxa were completely elimi-
nated from channels receiving cadmium. Other taxa showed increased or de-
creased relative denultifb. Both macrophytc and periphyton growth was In-
hihiteii by these levels of cadmium exposure, Population and community re-
covery was rapid, with communities with rspld growth and Invasion potentials
indist inguiahalile from control systems within weeks of the time cadmium in-
puts weir stopped.
Methods of pei turhdi ion assessment at both the population and system
IrvH were compaied. Effects can be demonstrated at both levels ot organi-
sation, und system level parameters were sensitive to cadmium-Induced
changes, however, measurements of system level parameters were not helpful in
determining mechanisms of the cadmium effect!.
Microcosms of the scale and complexity atudied here are useful for
validating and verifying predictive models of the the fates of contaminants
and testing assessment utrategles but are not appropriate for toxlclty test-
ing, or determining mechanisms and coef f icients of uptake, elimination or
degradat ion.
iv
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Thla report was aubmltted In fulfillment of Interageney Agreement
No, IAG-D6-0369-1 by Che Savannah River Ecology Laboratory under the
aponaorahip of the U,8, Environmental Protection Agency. Thla report
covera the period May 12, 1975, to May 31, 1978, and work waa completed
a'a of May 31, 1978,
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CONTENTS
Foreword
Abstract iv
Figures viii
Tables xiii
Acknowledgment xv
Section
1. • Introduction 1
2. Conclusions 4
3. Recommendations 6
4. Facility Description 7
5. Water Chemistry 10
6. Sediments 14
7. Aufwuchs 17
8. Macrophy tes 40
9. Invertebrates 53
10. Fish 99
11. Leaf Decomposition Ill
12. System Responses 120
13. References 128
Appendix A Analytical Techniques 145
Appendix B Plants and Animals Collected from Channels during
Cadmium Study 148
Appendix C Published Information Resulting from Agreement
Prior to Publication of this Report 156
vii
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FIGURES
Number Page
1 Photograph of stream microcosm facility 8
2 Glass slides used for sampling aufwuchs 19
3 Mean Cd concentrations in aufwuchs collected from long term
glass slides incubated in the channels from the beginning
of Cd exposure 22
4 Mean Cd concentrations in aufwuchs collected from short term
glass slides incubated in the channels for the eight weeks
prior to sampling 23
5 Linear regression of Cd elimination from the aufwuchs community
colonizing glass slides 26
6 Mean aufwuchs biomass accrual on long term glass slides incu-
bated from the beginning of the experiment with confidence
intervals indicated 27
7 Mean aufwuch biomass accrual on channel walls with two standard
error confidence intervals indicated 28
8 Mean viable algal cell volume collected from long term glass
slides incubated from the beginning of the experiment with
two standard error confidence intervals indicated 29
9 Mean viable algal cell volume collected from channel walls
with two standard error confidence intervals indicated ... 30
10 Algal ratio for aufwuchs collected from both long term glass
slides and channel walls with two standard error confi-
dence limits reported from long term glass slide samples . . 32
11 Pigment ratio for aufwuchs collected from both long term
glass slides and channel walls with two standard errors
confidence limits reported for glass wall samples 33
viii
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Number Pa8e
12 Percent of algal community, collected from long term glass
slides and channel walls comprised of green algae with
two standard errors confidence limits reported for long
term glass samples 34
13 Diversity values for the algal community colonizing long term
glass slides and channel walls with two standard errors
confidence intervals indicated for long term glass
slide samples 35
14 Evenness values for the algal community colonizing long term
glass slides and channel walls with two standard errors
confidence intervals indicated for long term glass slide
samples 36
15 Macrophyte standing crop biomass as a function of distance
from the headpools, as of September, 1977 43
16 Macrophyte standing crop biomass as a function of distance
from the headpools, as of March, 1977 43
17 Cadmium concentrations in J. diffusissimus colonizing the
headpools, expressed on a dry weight bases ........ 46
18 Cadmium concentrations in J. d i ffusi s simus colonizing the
channels, expressed on a dry weight basis 47
19 Cadmium concentrations in J. diffusissimus transplanted to
the channels, expressed on a dry weight basis 48
20 Cadmium concentrations in C. heterophylla shoots colonizing
the channels, expressed on a dry weight basis 49
21 Cadmium concentrations in C. heterophylla roots colonizing
channels, expressed on a dry weight basis 50
22 Photograph showing Hester-Dendy type invertebrate samplers
suspended in channels 55
23 Photograph showing invertebrate sampler in plexiglass and
screen sampling box 56
24 Photograph showing polyurethane sponge microinvertebrates
samplers 60
25 Mean Cd concentrations in pooled samples of chironomids, ex-
pressed on a dry weight basis. 66
26 Density of chironomids in plate samplers 67
ix
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Number Page
27 Mean chironomid weights 68
28 Mean number of macroinvertebrates per sampler 73
29 Mean number of macroinvertebrate taxa per sampler with two
standard error confidence intervals indicated 74
30 Percent community composition of macroinvertebrate community
in control channels 76
31 Percent community composition of macroinvertebrate community
in channels receiving 5 pg Cd/1 77
32 Percent community composition of macroinvertebrate community
in channels receiving 10 (Jg Cd/1 78
33 Density of Ephemeroptera per sampler 79
34 Density of Ceratopogonidae per sampler 80
35 Density of Pi. aequiseta per sampler 31
36 Density of E. agilis per sampler 82
37 Total number of microinvertebrates observed per month in
polyurethane sponges 83
38 Simpson's diversity index. A, means calculated across sampler
by sampling period with two standard error confidence
intervals indicated. B, calculated by summation 85
39 Evenness of Simpson's diversity index. A, means calculated
across sampler by sampling period with two standard
error confidence intervals indicated. B, calculated
by summation 86
40 Shannon's diversity index. A, means calculated across sampler
by sampling period with two standard error confidence
intervals indicated. B, calculated by summation 87
41 Evenness of Shannon's diversity index. A, means calculated
across sampler by sampling period with two standard error
confidence intervals indicated. B, calculated by summa-
tion 88
42 Macintosh's diversity index. A, calculated across sampler
by sampling period with two standard error confidence
intervals indicated. B, calculated by summation 89
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Number Pa8e
43 Evenness of Macintosh's diversity index. A, calculated across
sampler by sampling period with two standard errors confi-
dence intervals indicated. B, calculated by summation . . 90
44 Probability of interspecific encounter diversity index. A,
calculated across sampler by sampling period with two
standard errors confidence intervals indicated. B, cal-
culated by summation 91
45 Evenness of probability of interspecific encounter diversity
index. A, calculated across sampler by sampling period with
two standard errors confidence intervals indicated. B, cal-
culated by summation 91
46 Renyi's generalized entropy series (a = 1) calculated across
sampler by sampling period, with two standard errors.
Confidence intervals indicated 92
47 Renyi's generalized entropy series (a = 1) calculated by
summation 93
48 Renyi's generalized entropy series (a = z) calcualted across
sampler by sampling period, with two standard error con-
fidence intervals indicated 94
49 Renyi's generalized entropy series (a = z) calculated by
summation 95
50 Mortality of large C. fluminea as a function of time 96
51 Mortality of small C. fluminea as a function of time 97
52 Cadmium accumulation by mosquitofish (G. affinis). n and
2 SE are indicated 103
53 Leaf litter pack, Type I 112
54 Leaf litter pack, Type II 113
55 Electron photomicrograph of the effect of Cd on microbial
colonization of P_. taeda. A. Control ..... 116
56 Electron photomicrograph of the effect of Cd on microbial
colonization of P_. taeda. B. 10 ug Cd/1 117
57 Electron photomicrograph of microbial colonization of
0^. nigra. A. Control 118
58 Electron photomicrograph of microbial colonization of
(£. nigra. B. 5 ug Cd/1 119
xi
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Number P:*w
rj'i Community metabolism. Gross primary production and
respiration with the shaded areas representing net
production 122
f/0 Aufwuchs accrual on short-term ^l.iss slides with two
standard error confidence intervals indicated 121
61 Algal cell volume accrual on short-term glass slides
with two standard error confidence intervals
indicated 124
62 Cadmium concentration in material exported from the
channel microcosms 125
xil
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TABLES
Mumber Page
1 Characteristics of microcosm sediments .......... . . . g
2 Mean water quality of treated well water ............ 9
3 Mean values (X ± SE) for chemical parameters measured at two
station* in each channel during the period of cadmium
inputs. Sample size is reported in parentheses. ...... 12
4 Mean cadmium concentrations in unfiltered water samples
(X ± SD, n = 10) ....................... 13
5 Mean organic content of channel sediments during study period
(X 1 SE, n s 16) ................ ..... 15
6 Mean Cd concentrations in pool sediments (X ± SE, n = 16) ... 15
7 Mean concentrations in channel sediments during Cd exposure
period (X ± SE, n = 18) .............. ..... 16
8 Uptake and elimination rates of cadmium by aufwuchs at two
treatment levels calculated using the Von Bertalantfy
model, c .......................... 24
9 Distribution and number of individual macrophytes in head
pools as of January 1976 ................ . . 42
10 Cadmium concentrations in macropbytes removed from channels
not receiving Cd ...................... 45
11 Mean Cd concentrations in insects, during the period of Cd
inputs ........................... 62
12 Mean Cd concentrations in insects during the period after Cd
inputs were terminated ..... .............. 63
13 Scheffe's S -procedure values for insect Cd concentrations at
each sampling period ... ................. 65
xlii
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Number Page
14 Mean Cd concentrations for P. hymenaea life cycle segments
by treatment, expressed on a dry weight basis (X ± 2 SE) . . 70
15 Mean % Cd in each life cycle segment of estimated numphs by
treatment (X ± 2 SE) 71
16 Effect of Cd on density of taxa in sponge samplers 84
17 Mean Cd concentrations in C. fluminea whole tissue expressed on
a dry weight basis 98
18 Linear models of the form p = mx + b of Cd uptake by G.
af finis 102
19 Non linear least squares fit of Cd accumulation by G.
affinis. Data fit to Q = Q (1 - e ) using the Gauss-
Newton iterative technique 104
20 Factorial main effects of Cd levels in food and water on whole
body concentrations of Cd in mosquitofish with 95% confi-
dence interval and F-test (P),n=5 105
21 Mean Cd concentration in mosquitofish under four treatment
combinations over time with 95% confidence interval,
n = 5 106
22 Simple effects and interaction term for week 8 which includes a
significant interaction between food and water with 95%
confidence interval and F-test (P), n=5 107
23 Bluegill and mosquitofish mortality between March and June,
1976 . . • 110
24 Initial leaf material in leaf litter packs 114
25 Effect of Cd on final biomass_of leaf material in leaf litter
packs exposed for 28 wk. (X ± 2 SD) 115
26 Cadmium concentration in leaf litter material exposed for 28
wk. (X ± 2 SD) 118
27 Summary of organic export from the channel microcosms during
and after cadmium input. Values are averages of.weekly
, _ , . . -2. , -1 Izb
averages in grams ash-free dry weight m • day
xiv
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ACKNOWLEDGEMENTS
We wish to thank the staff of the EPA's Environmental Research Labora-
tory, Athens, Georgia, and the University of Georgia's Savannah River Ecology
Laboratory for their assistance. Special recognition is due Drs. Harvey Holm
and Ray Lassiter of the EPA for their help in all phases of the study. J.
Cheatham, S. Giddings and R. Didgeon provided technical assistance throughout
the study. Ms. J. Coleman prepared ink drawings.
The channels, which were constructed with funds provided by the Environ-
mental Protection Agency in 1970, have been operated by personnel of the
Savannah River Ecology Laboratory since that time (Kania and Beyers, 1974;
Kania et al. , 1976). Operating funds have been provided by the EPA;, the DOE
and its predecessors have provided office space and laboratory facilities,
and a variety of crucial services.
XV
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SECTION I
INTRODUCTION
Cadmium is a relatively rare element, not found in a pure state in the
environment (Hiatt and Huff, 1975), making up an average of less than one-
half gram per ton of the earth's crust (Page and Berggen, 1973; Fassett,
1974). While the natural occurrence of Cd in the environment is quite small,
cadmium is used in many industrial processes and products such as storage
batteries, pigments, semiconductors, plastics, stabilizer compunds, alloys
and plating solutions and occurs as a contaminant in zinc ores, automobile
tire dust, fossil fuels and agricultural fertilizers (Friberg et al. , 1971;
Anon., 1975; Hiatt and Huff, 1975).
Because Cd is a trace contaminant in so many materials and is released
from so many diffuse sources, reduction or elimination of point source re-
leases may not significantly reduce the trend of increased Cd mobilization
with increased general human activity. The special difficulty with metals
such as Cd is their persistence. Unlike organic contaminants, metals do not
degrade in the environment and regardless of their source, most metallic
wastes eventually end up in surface and subsurface waters (Buhler, 1971).
Cadmium is a biologically nonessential element (Anon, 1971; Fassett,
1974; Rosenthal and Sperling, 1974) and is known to be acutely and chroni-
cally toxic to plants and animals (Lagerwerff and Spect, 1970; Flick e_t al. ,
1971; Burkitt et al., 1971; Cheremisinoff and Habib, 1971; Schroeder, 1974;
Hiatt and Huff, 1975; Chadwick, 1976; Giesy et aJL. , 1977). The human health
aspects of acute and chronic Cd poisoning in humans have been reviewed ex-
tensively (Anon., 1971; Fassett, 1974; Page and Bingham, 1973; Piscator,
1974; Schroeder, 1974; Friberg and Kjellstron, 1975; Fulkerson, 1975; Hiatt
and Huff, 1975; Anon., 1975; Perry et al., 1976). Beside the often cited
acute "itai itai disease", cadmium has been implicated as a possible carci-
nogen and mutagen and Cd exposure is correlated with cardiovascular disease,
renal disfunction and hypertension (Flick et al. , 1971; Perry et al., 1976).
Cadmium releases to the environment have increased drastically since 1945,
with a concomitant increase in the reported cases of Cd toxicity.
It is difficult to set standards for human exposure because the toxic
effects of Cd exposure are cumulative. At birth, human beings have essen-
tially no Cd in their tissues and gradually and continuously accumulate Cd,
particularly in red blood cells, kidney, liver, bone, pancreas and liver
(Wagner, 1971). Intake by humans is approximately 200-300 jjg Cd/day which
accumulates at a rate of approximately 3 |Jg/day and is eliminated very slowly
from the body (Hiatt and Huff, 1975). There is considerable evidence that
organisms, which have evolved under conditions of very low Cd exposure, deal
with Cd by sequestration, rather than excretion and elimination. In fact, a
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nonspecific metal binding protein, thionein, is present in the organs of many
animals, including humans (Hiatt and Huff, 1975; Anon, 1975). It is unknown
whether protection against Cd toxicity by thionein is limited or whether
protection is a function of accumulation rate. Rapid increases in Cd mobili-
zation to the biosphere, relative to geologic-evolutionary time may have
severe effects on organisms. The joint FAO/WHO Expert Committee on Food
Additives has concluded the "present day levels of cadmium in the human
kidney should not be allowed to rise further (Anon, 1975).
To achieve this goal, a good understanding of the geologic and biotic
cycling of Cd will be necessary. More work is needed on the cycling of
cadmium in fresh water, estuary and marine ecosystems before the significance
of water pollution can be assessed relative to both ecological effects and
man's food (Fulkerson, 1975). Fleischer et al. (1964) state, after a review
of the literature describing the levels of Cd in plants and animals, that
"Experimental studies of uptake over the lifetime of experimental animals are
required for a number of representative species and at least one food chain
study should be made in each of the three environments: terrestrial, fresh-
water and marine. Model ecosystems (Microcosms) might be the most appro-
priate systems for these studies." These same authors conclude, after a very
short review of the literature on ecological effects of cadmium that "Our
ignorance of the effects of cadmium in natural or polluted systems is almost
total."
While many studies have been conducted on acute toxic effects of Cd on
and uptake by organisms and specific physiological responses in the labora-
tory, few studies have been conducted in complex environments. When con-
ducted in complex systems, studies have generally addressed effects or uptake
independently, focusing on single populations or taxonomic groups, under con-
ditions such that the source term is not known and have not generally been
conducted over a sufficiently long period of time to be meaningful.
Research conducted in outdoor artificial stream channels can provide a
vital link between laboratory studies carried out under carefully controlled
conditions and field studies which can seldom be adequately controlled or
replicated. Artificial streams provide realistically complex biological
systems where replication is possible, a number of treatments can be inves-
tigated simultaneously, certain critical parameters can be readily con-
trolled, and the addition and removal of stresses can be readily effected
resulting in little or no environmental damage. Microcosms can be effec-
tively used to study both environmental transport (Draggan, 1976) and effects
of toxic materials in aquatic microcosms (Taub, 1976). The microcosms used
in this study were complex, self perpetuating functioning ecosystems, func-
tionally analogous to the littoral zone of softwater paludal systems.
This study was conducted to examine the fates and biological effects of
chronic Cd concentrations (5 and 10 M8/1) over a number of trophic levels,
during an entire growing season in a complex aquatic ecosystem. Cadmium
uptake and elimination and compartmentalization by the aufwuchs, macrophyte,
macroinvertebrate, fish sediment, and water components were measured and
models of Cd dynamics in these systems proposed. Rate of Cd elimination from
the contaminated system was also monitored. Biological effects were measured
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at the population, community and ecosystem level. A comparison of the rela-
tive sensitivity of each of these organizational levels to Cd-induced stress
is presented. Effects on macrophytes, aufwuchs, microinvertebrates, macro-
invertebrates, and fish populations and system level measures of Cd induced
effects were made and rate of recovery assessed. As well as basic informa-
tion on chronic Cd effects.
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SECTION II
CONCLUSIONS
1) When exposed to 5 or 10 |Jg Cd/1 sediments and all biologic compo-
nents monitored accumulated cadmium. Cadmium accumulation was approximately
porportional to exposure concentration. Equilibrium Cd concentrations of
sediments, periphyton, macrophytes, chironomids and mosquito fish exposed to
10 (Jg Cd/1 were 0.59, 55, 250, 40 and 40 (Jg Cd/g dry weight, respectively.
2) Cadmium concentrations in biotic components reached equilibrium
within 20 days. Cadmium accumulation in the aufwuchs andjJLish components
could be described by a model of the form C = C (1 - e ). Biological
elimination was rapid, with cadmium concentrations in the aufwuchs and macro-
invertebrate communities indistinguishable from background within 30 days of
cessation of cadmium exposure. Cadmium elimination from sediments was much
slower. Six months after Cd inputs were terminated there was no significant
decrease in Cd concentrations in organic headpool sediments. Half times for
elimination from sand substrata and detritus in the channels were 72 and 38
days for the 5 and 10 (Jg Cd/1 treatments respectively.
3) Cadmium caused effects at both the population and system levels.
Standing crops of both aufwuchs and macrophytes were depressed during the
time of cadmium exposure. A number of invertebrate taxa were eliminated due
to cadmium inputs, while others were released from competitive or predatory
influence and did well in systems receiving cadmium. Both 5 and 10 |Jg Cd/1
treatments were chronically toxic to crayfish and snails.
4) Invertebrate and algal population structure returned to background
levels within a few weeks of cadmium cessation, while fish and macrophyte
populations did not recover as rapidly. This is due to the more rapid growth
and colonization rates of algae and invertebrates.
5) Systems level structure and function were affected by cadmium.
Algal and macroinvertebrate species diversity, leaf litter decomposition, net
and gross production, community respiration, and P:R ratios were depressed
due to cadmium exposure.
6) Microcosms of the scale and complexity studied here are not appro-
priate for toxicity testing or determining mechanisms and coefficients of
uptake, elimination or degradation.
7) Microcosms of the scale and complexity studied here are appropriate
for verification and validation of predictive models of local fates of trace
contaminants but not of worldwide transport models.
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8) Comparison of both population and system levels of perturbation
assessment were composed. Effects can be demonstrated at. both levels of
organization and system level parameters were sensitive to cadmium induced
changes, however, measurements of system level parameters were not useful in
determining mechanisms of cadmium induced effects.
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SECTION III
BKCOMMKNDATIONH
I) Result* of recent studies of cadmium toxlclty to mlcrocrustaceans
(Gelsey et al,, 1977) should be considered when water quality criteria are
revised.
2) Kxposure of aquatic sediments to even low concentrations of cadmium
iH In elevated cadmium concentration in this component which le persls-
and should he avoided.
U Assessment of radmlum induced perturbations should not be monitored
solely by either strurtural or functional attributes of systems or popula-
tions.
4) Mlrror-osms of the type studied here are not appropriate for 1)
determining uptake mechanisms or roeff'Irlents, 2) acute or chronic toxicity
ifcsilnK, 3) screening possibly hazardous rheml
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SECTION IV
FACILITY DESCRIPTION
The microcosm facility used in thin study is located on the Department
of Energy's (DOE) Savannah fliver Plant (SRP), a 507 km reserve, Including
portions of Aiken, Barnwell and Allendale Counties in South Carolina, U.S.A.
The facility conalsts of six concrete block channel* (Fig. 1) 91,'' m
long, 0,6J m wide and Q.'tl m deep with concrete pools (1.5 m x 3.0 m x 0.92
m) at both enda of each channel supported by a concrete slab oriented on an
east-west foundation, For this study, the pools and channels were lined wJth
a 0.05 cm thick black, polyvlnyl chloride (PVC) film.
Washed quart/ sand waa distribute in the channela to a uniform depth of
0,05m, and a 8-10 cm layer of natural stream bed sediment obtained locally
was distributed In the pools. This resulted in u system similar to local
aquatic systems which have both land and silt bank substrata (Table 1),
Water for the channels was pumped from a well located near the facility
and a hydrated lime slurry was continuously pumped into the main water dis-
tribution system throughout the period of the study to produce inorganic
water quality similar to that of local upper coastal plain surfaie waters. A
single batch of lime was used throughout the study and an ana I yb is of the
treatment water is given in Table 2,
Flows were monitored by V-notch weirs on each head pool where water
entered the channel. A flow rate oi 95 i/min was maintained manually by an
input valve located at em h head pool, resulting in a wair-r depth of 20 on in
the.channels. The mean water retention time and current were 2 hr and 1. J x
10* m/8, respectively,
At the time water flow was commenced, the systems were seeded with ma-
terial saved from the control channela of 4 previous *tudy (Kania /-i al,,
1976) to rapidly establish biological communities known to be well adapted to
channel conditions, The ma
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Figure 1. Stream microcosm facility.
TABLE 1. CHARACTERISTICS OF MICROCOSM SEDIMENTS
Clay
Organic Matter
CEC
Head Pools
Location
Tail Pools
3.6
28.4
67.0 meq/lOOg
12.8
34. 3
75.6 meq/lOOg
Channels
Sand
Silt
37.2%
30.7
36 . 5%
16.7
99 . 8%
<0.1
0.02 meq/lOOg
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TABLE 2. MEAN WATER QUALITY OF TREATED WELL WATER
Total alkalinity
Hardness
PH
Specific conductance
Ionic strength (I)
Total dissolved solids
so,-2
4
Total P
Nitrogen (N02 + NO )
Ca
Cu
Co
Cd
Cr
Fe
K
Mg
Mn
Na
9.14 mg/£ as CaC03
11.08 mg/£ as CaCo3
6.5
31 |J mho/ cm
2.5 x 10 - 4
20.5 mg/£
1.9 mg/Z
2.9 (Jg/£
15.8 |Jg/£
03.17 mg/£
3.4 |jg/£
2.5 |Jg/£
0.023 jJg/£
0.3 pg/£
1.7 |Jg/£
1 . 1 pg/£
246 pg/iu
7.0 |Jg/£
1.8 mg/£
Instruments Model 375A). Cadmium input solutions for each channel were made
every two days with concentrations adjusted to compensate for pump variation
over time. The Cd levels established were 5 pg/1 in two channels and 10 pg/1
in another two. Since the six channels are structured in three pairs (Figure
1), the dosing arrangement was chosen so that there was both a northern and a
southern exposed channel for each treatment. Cadmium inputs were discon-
tinued on 18 March 1977, one full year after they were begun.
-------
SECTION V
WATER CHEMISTRY
INTRODUCTION
The physical and chemical state of trace metals is dependent upon water
quality and must be considered when availability and toxicity of metals to
aquatic organisms are assessed (McKee and Wolf, 1963; Hartung, 1973; Brown et
al. , 1974; Clubb et a_l. , 1975). Metal toxicity to aquatic organisms is
hardness dependent (Sprague, 1969). For example, water hardness has an
antagonistic effect on Cd toxicity to zooplankton due to Ca and Mg (McKee and
Wolfe, 1963). Similarly, Pickering and Henderson (1966) found increases in
Cd 96 h LC values with increasing water hardness for all fish tested and
Kinkade andtrdman (1975) reported that organisms accumulate Cd faster from
soft than hard water. The free divalent metal ion is generally the most
toxic form (Stiff, 1971; Brown et al. , 1974; Pagenkopf et al. , 1974). The
soft acidic waters of the southeastern United States have low inorganic
ligand concentrations. Thus, inorganic solubility product chemistry predicts
that Cd introduced into these waters would exist raainly.as free-divalent ca-
tigns (Cd ) or as hydrated ions (CdOH , B = 1.5 x 10 ; CdO ~ , B, = 5.8 x
10 ) Weber and Posselt, 1974). All reactions are rapid resulting in replen-
ishment as a particular ionic form is depleted. Giesy et al. (1977) found
very low LC values for zooplankton exposed to Cd in the well water studied
here.
METHODS
Water samples were collected monthly from each channel at a single
station, located 60 m downstream of the input weirs. Temperature was mea-
sured with a YSI model 44 TD Telethermometer, conductivity with a Beckman
Model RC19 Conductivity Bridge, and pH with a Orion Model 401 Specific Ion
Meter with a glass electrode. Alkalinity and hardness were measured using
standard EPA (1976) or APHA (1976) techniques.
Total phosphorus, nitrite and nitrate nitrogen, sulphate ion, inorganic
and organic carbon analyses were performed periodically on water samples
collected from the inputs and upstream and downstream locations in each
channel. Downstream stations were sampled two hours after upstream and down-
stream locations so that the same water mass was examined and changes could
be related to the biological activities in the channels.
Both organic and inorganic carbon analyses were made using a Beckman
Model 915 total organic carbon analyzer. Other chemical parameters were
measured using accepted EEA methods (1976) with the addition of a 5X concen-
tration step prior to PO. and Cl analyses.
10
-------
Samples for Cd analyses were taken from inputs to the head pools, out-
flows of the headpools, outflows to the channels, and outflows from the tail
pools, on a monthly basis. A more frequent sampling schedule was initiated
at the time the Cd inputs were terminated. Water samples were taken in 160
ml glass milk dilution bottles, which were used only for water sampling and
always for the same station. After each sample was mixed well, 10 mis were
transferred volumetrically to an acid rinsed polyethylene bottle and acidi-
fied with 200 |Jl of redistilled cone. MHO . A portion of the remaining
sample was filtered, and 10 mis of the filtrate transferred to acid rinsed
bottles and acidified. Details of the analytical procedures are given in
Appendix I.
An attempt was made to separate particulate and dissolved Cd by both
membrane and fiberglass filtration. Results of analyses on filtered samples
were in general the same or higher than unfiltered samples. A low level con-
tamination problem causing higher levels on filtered samples was never com-
pletely resolved.
Samples of the PVC plastic used to line the channels were suspended at
both ends of each channel to see if this material either adsorbed or released
into the water. Subsamples of these suspended sheets were periodically re-
moved, washed free of periphyton, dissolved in hot concentrated H_SO, , oxi-
dized with concentrated HNO,. and hydrogen peroxide and analyzed using stan-
dard flame AA techniques. There was no measurable uptake or loss of Cd from
the PVC film.
RESULTS AND DISCUSSION
Additions of hydrated lime satisfied the CO demand of the well water
and resulted in water similar to surface waters of the upper coastal plain
(Table 2). The ionic strength of treated well water was 2.5 x 10 resulting
in an activity coefficient for Cd of 0.97, using the extended Deby-Huckle
equation. Chloride ion concentrations remained constant at all stations
(Table 3). Sulphate levels increased between the upstream and downstream
stations. Total organic carbon levels were below detection limits at all
times at all stations. Total inorganic carbon levels decreased along the
length of the channels with no differences due to Cd. Total phosphorus
concentrations were low but constant at all positions, and there was a marked
reduction in NO-NO nitrogen level in both the head pools and the channels.
There were no effects on nitrogen concentrations due to Cd. However, con-
sidering only data from the months of June and July, 1976, the average nitro-
gen uptake in the control channels was 9.6 ± 0.3 JJg/1 compared to 4.3 ± I
|Jg/l in the 5 p/1 treatment and 5.0 ± 1 pg/1 in the 10 p/1 treatment.
The observed Cd concentrations were not significantly different from the
desired concentrations and there were no significant differences between
sampling stations within each treatment system (Table 4). After Cd inputs
were terminated, Cd concentrations in water quickly dropped and within four
days were in the range of the control channels (0.02 0.06 |Jg Cd/1) at all
stations. Cadmium is used as a pla'stisizer in PVC plastic so this material
has significant concentrations of Cd in it. However, there was no measurable
addition or removal of Cd due to the PVC film liner.
11
-------
TABLE 3. MEAN VALUES (X ± SE) FOR CHEMICAL PARAMETERS MEASURED AT TWO STATIONS IN EACH CHANNEL DURING THE
PERIOD OF CADMIUM INPUTS. SAMPLE SIZE IS REPORTED IN PARENTHESES
Control 5 \Jtg/S, 10 |Jg/£
Up Down Up Down Up Down
N
Cl~
(6)
SOj
N)
TOC
(12)
TIC
(12)
Total P
(16)
mg/£
mg/£
mg/A
mg/£
M8/*
2.910.1 2.910.1 2.910.1 2 . 9 *± 0 . 1 2.910.1 2.910.1
1.87 1 0.08 2.10 1 0.06 1.81 1 0.05 2.13 1 0.05 1.79 ± 0.07 2.19 1 0.06
<0.5 <0.5 <0.5 <0.5 <0.5 <0.5
7.1 1 0.1 4.5 1 0.2 6.6 1 0.2 4.4 1 0.2 6.8 1 0.2 4.5 ± 0.2
3.5 1 0.3 3.5 1 0.3 3.1 1 0.2 4.1 1 0.5 3.3 1 0.2 3.5 1 0.3
N02-N0 N pg/£ 10.4 ± 0.5 3.6 ± 1.0 8.910.7 5.210.6 9. 310. 6 4. 710. 6
(10)
-------
TABLE 4. MEAN CADMIUM CONCENTRATIONS IN UNFILTERED WATER
SAMPLES (X ± SD, n = 10)
NOMINAL TREATMENT
Sampling station 5 |Jg Cd/1 10 |Jg Cd/1
Input 4.75 ± 2.07 5.00 ± 1.91 9.36 ± 4.22 10.15 ± 5.72
Head pool weir 4.30 ± 1.92 4.17 ± 1.85 9.45 ± 4.27 9.63 ± 5.21
Channel outflow 4.12 ± 1.71 3.94 ± 2.20 8.76 ± 3.82 9.61 ± 4.01
Tail pool outflow 4.27 ± 1.82 3.99 ± 2.26 8.30 ± 3.91 10.06 ± 4.15
13
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SECTION VI
SEDIMENTS
INTRODUCTION
In aquatic systems, the sediments can act as both a sink for pollutants
and as a source for the release of these pollutants under appropriate con-
ditions. The behavior of cadmium in sediments and at the sediment-water
interface must be known if the cycling of the element in the aquatic environ-
ment is to be understood. The purpose of the sediment work in this study was
to compare the uptake and release of cadmium from two different sediment
types receiving,chronic known exposures to dissolved cadmium.
METHODS
t
Sediment samples were taken monthly from upstream and downstream sta-
tions in each channel and also from both head and tail pools. Cores were
used for Cd determinations and also, in the case of sediment from the chan-
nels, to derive an estimate of organic content. Analytical procedures are
presented in Appendix I. Several different sample collection techniques were
used. The consistency of the highly organic silty streams sediments placed
in the pools was such that cores could not be taken and sampling was done
with a large syringe-like device which caused mixing of the sediments with a
small amount of water when the plunger was withdrawn. Several sizes of
coring tubes were used for the sandy sediments of the channels, initially,
frozen cores were taken so that stratification of the cadmium within the
sediment could be determined. This procedure was abandoned when it became
evident that, unlike mercury (Kania et al. , 1976), Cd was distributed
through-out the sediment although more highly concentrated in the highly
organic upper layer.
RESULTS AND DISCUSSION
The first sediment samples were taken at the end of April 1976, approxi-
mately 40 days after Cd inputs to the channels were initiated. At that time,
the organic content of the channel sediments had already reached equilibrium
values which were unrelated to treatment or position, and remained unchanged
throughout the remainder of the study (Table 5).
Sediment Cd concentrations were at equilibrium and no further increase
in levels with time were observed at any station. This is consistent with
the findings of Huckabee and Blaylock (1974) who found, working with spiked
microcosums, maximum sediment Cd activity was reached after only two days.
Bunzl (1975) found that humic acids sorbed Cd with a half time
14
-------
TABLE 5. MEAN ORGANIC CONTENT OF CHANNEL SEDIMENTS DURING
STUDY PERIOD (x ± SD, n = 26)
TREATMENT % ORGANIC
control 0.50 ± 0.36
5 |Jg Cd/£ 0.43 ± 0.10
10 Mg Cd/£ 0.41 ± 0.11
of approximately 30 seconds, indicating that this'step would not be limiting
in uptake of Cd by our highly organic pool sediments. There were no signifi-
cant differences between upstream and downstream stations in the channels,
however, the levels in the tailpools were generally higher than those in the
head pools (Table 6). Cadmium concentrations in channel sediments (Table 7)
are only for the time period during cadmium input, since Cd levels in these
sand sediments decreased after the inputs were terminated with mean half
times of 72 and 38 days in the 5 Mg/£ an(* 1° H8/£ treatments respectively.
No significant decrease in sediment Cd concentrations of pool sediments were
observed after Cd inputs were terminated so measured Cd concentrations of
these sediments include samples taken during the 9 months following Cd input
termination.
Naturally occurring Cd was measured in both sand and organic sediments
(Tables 6 and 7). The highly organic (25-30% by weight as C) silty stream
sediments had much higher Cd concentrations than the sand. The organic sedi-
TABLE 6. MEAN CD CONCENTRATIONS IN POOL SEDIMENTS
(X ± SE, n = 16)
TREATMENT Cd CONCENTRATION
|jg Cd/g dry weight
Head Tail
control
5|Jg Cd/£
10 (jg Cd/S.
1.27 ± 0.10
8.33 ± 0.96
10.6 ± 1.6
1.50 ± 0.10
21.4 ± 1.8
22.3 ± 2.7
15
-------
TABLE 7. MEAN CD CONCENTRATIONS IN CHANNEL SEDIMENTS
DURING CD EXPOSURE PERIOD (X ± SE, n = 18)
TREATMENT CD CONCENTRATION
((jg Cd/g dry weight)
control 0.014 ± 0.003
5 (Jg Cd/£ 0.209 ± 0.019
10 [jg Cd/S. 0.591 ± 0.061
merits also accumulated much higher Cd concentrations when exposed to Cd. Cd
uptake by sand was linearly proportional to Cd exposure, while that of or-
ganic silt sediments was not. Tail pool sediments acquired greater concentra-
tions than did that in head pools.
Cadmium concentrations in control channels are in the range of values
reported by other investigators for background levels (Table 7). Fleisher et
al. (1974) reported on average value for 26 samples of unspecified lake
sediments of 11 pg/g. Forstner (197 ) gives background sediment values for 5
lakes ranging from not detectable to 2.5 M8/8- Shepard (1976) discusses
background levels of 0.3 - 6.2 pg/g.
The sediment cadmium concentrations resulting from the one year exposure
to 5 and 10 (Jg/g water concentrations (Tables 6 and 7) were very low compared
to values reported for contaminated field sites. Shephard (1976) found
levels as high as 1300 (Jg/8 in a lake contaminated by an electroplating plant
and Kneip et. al. (1974) reported levels of 3000 - 50,000 |Jg/g in an indus-
trially contaminated sediment. The sediment Cd concentrations observed in
our study were similar to those in the Derwent River, England (Harding and
Whitton, 1978).
The low values observed in our work may be due to the low pH (Table 2)
of the water. Murray and Mernke (1974) found virtually no Cd adsorption on
suspended sediments at a pH of less than 6.6. The higher concentrations of
Cd in the highly organic pool sediments is consistent with the finds of
Riffaldi and Levi-Minzi (1975) who found that Cd adsorption maxima and co-
efficients were well correlated with cation exchange capacity and organic
matter content. There is no immediately apparent reason why the tail pool
sediments acquired higher Cd concentrations than the head pools although
Korte et al. (1976) state that the percentageof clay in a sediment is the
most useful predictor of whether or not a soil will retain a particular
element. The tail pool sediments had a significantly greater clay content
than the head pools (Table 1).
16
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SECTION VII
AUFWUCHS
INTRODUCTION
As used in this report, the terra "aufwuchs" refers to the complex epi-
lithic, episalmic and epipelic assemblage of autotrophs and heterotrophs
which developed on aquatic substrata. The German term aufwuchs was proposed
by Ruttner (1953) to conote the community of both plants and animals attached
to but not penetrating aquatic substrata. The term has been used inter-
changeably with the English term "periphyton" (Hynes, 1970); however, strict-
ly speaking periphyton is only the autotrophic or plant component of the
aufwuchs. We have separated the discussion of density and diversity of the
micro and macro invertebrates from the general discussion of the aufwuchs
community. These organisms, however, are included in estimates of standing
biomass and metabolism. Qualitatively, the aufwuchs community includes
algae, fungi, bacteria, protozoans, and small invertebrates and may form a
mat up to a few centimeters in thickness depending on substratum orientation
and current velocity. In flowing water systems, phytoplankton are virtually
absent and the algae of the aufwuchs as well as macrophytic plants constitute
the basis of the autocthonous food web. In well-lighted streams this iri situ
carbon production can be substantial and therefore the effect of Cd on
aufwuchs dynamics is of considerable importance. The effect of Cd on the
heterophic, non-algal components of the aufwuchs community are also of impor-
tance, since these organisms provide the mechanism for rapid cycling of
nutrients and therefore sustained productivity in a lotic environment. The
aufwuchs communities of aquatic ecosystems have been used as sensitive indi-
cators of both chemical and physical stressors (Rodgers and Harvey, 1976)
since it is sessile and taxonomically diverse and involved in all of the
functional processes of ecosystems. For these reasons, the aufwuchs commu-
nity is a biological integrator of ecosystem information (Weber, 1973).
Wetzel (1975) states that the trophic structure above the producer - decom-
poser level, with all of its complexities, population fluctuations, metabo-
lism and behavior, has relatively minor input on the carbon flux of eco-
systems. The aufwuchs community is also the component with the greatest
capacity to sorb potential toxicants.
In this study, three questions concerning the interaction of low Cd
levels with aufwuchs were addressed: 1) What are the kinetics of Cd uptake
and elimination and how are steady state concentrations in the aufwuchs re-
lated to water Cd concentrations? 2) What effect does Cd have on the struc-
ture of the aufwuchs community as measured by standing crop, species compo-
sition, pigment ratios, and chorophyll to biomass ratios? 3) How do any
changes in the structure of the aufwuchs community affect system level func-
17
-------
tioning through primary productivity, metabolism, and export? In the scope
of the above questions we are also asking if an aquatic system can adapt to a
continuous toxin input and be dependent on it as an organizing influence.
METHODS
Community Structure
Two hundred 50 by 75 mm washed glass slides were placed in each channel
on 1 November, 1975. These slides (referred to as long-term glass slides)
were placed vertically on five notched racks holding twenty slides (Figure 2)
placed 9 and 85 m from the head pools. These two groups are referred to as
head samples and tail samples. Slides were held approximately 5 cm above the
stream bottoms with their long axis parallel to stream walls. At monthly
intervals four slides were randomly chosen from both the head and tail sta-
tions of each channel and placed in washed beakers. Both surfaces of the
slides (area = 7500 mm ) were carefully scraped and washed with a minimum
volume of water (20 to 200 ml depending on aufwuchs density) and various
determinations made as described below.
Beginning on 15 December 1975 and continuing at 8 week intervals,
smaller sets of slides (6 to 9 at each position) were placed in the channels
and allowed to colonize 30 days. Four slides were collected from each loca-
tion and aufwuchs removed. These samples (referred to as short-term glass
slides) were handled in the same manner as the long-term slides.
After scraping the slides a direct count of number of chironomids was
made using a binocular dissecting scope at low power (30X). Densities of
other invertebrates (copepods, caldocerans, ostracods, etc.) and large algae
(Eremosphaeria and desmids) were qualitatively noted.
By November 1976, it was clear that glass slides were underestimating
aufwuchs production on some substrata in the channels such as the walls,
where the mat was several centimeters thick. This underestimation was due to
some sloughing of aufwuchs as the slides were lifted from the water and also
due to less accumulation on the slides because of their location in the cen-
tral part of the streams where current was maximum. Because of these limita-
tions samples were scraped directly from the stream walls and processed in
the same manner as glass slide samples. Two samples (area = 587 mm ) were
collected from each wall in the head and tail portions of each channel on 8
November 1976, 12 January 1977, 30 March 1977, and 28 June 1977. These
samples were then processed for algal cell densities by taxa, biomass, chlo-
rophyll and Cd concentrations as described below.
2
On two occasions, six bottom samples (area 426 mm ) were collected
from each channel to determine algal cell densities, biomass, chlorophyll,
and Cd concentration. These samples were collected by inserting a piece of
plexiglass tubing vertically into the sediment to the PVC liner. The entire
core was placed in a clean flask and processed in a manner similar to glass
slide or wall samples.
18
-------
Of the diverse flora and Kiuri.t in .mfwu- nples, only the algal com-
ponent was identified and ennmet/i! I dt -nt i ) i i .11 i «.n to species was made
when possihle using standard taxonomic references, hut in several cases
common algae were not class i f i .ihl c to specirs due to lack of fruiting stages
or confusing taxonomic literature. Complflc descriptions of all common
species were made and there was little difficulty in distinguishing between
them in routine counts.
Figure 2. Glass slides used for sampling aufwu
-------
Scraped samples were blended for 30 seconds at low speed in a Waring
blender to break up clumps and provide a more homogenous suspension for sub-
sampling. Subsamples were taken by volumetric pipette for biomass, chloro-
phyll, and Cd concentration determination. A few milliliters of suspended
sample was preserved with Lugol's iodine solution (APHA, 1976) for algal
counts. Several aliquots of both living and preserved samples were examined
and no significant differences found in cell densities or taxa observed.
Also, samples were compared before and after blending and there was almost no
qualitative or quantitative difference observed. However, one rare species
(Eremosphaeria viridis) was known to be disrupted beyond recognition by
either blending or preservation with Lugol's solution.
Algae were counted using the drop-transect method (Voelenweider , 1969).
Samples were mixed, with a vortex mixer and 50 |Jl subsamples removed using
an automatic microliter pipette placed on a clean glass slide and covered
with a cover slip. Random transects were then counted using an ocular micro-
meter to delineate the width of the scanned field. All cells of living
algae (intact cells with pigments) were counted as units and the number of
cells per area of the original surface calculated using equation 1.
n / 2 484 (N) (M)
cells/mm = - ^^ — *— ^
0.05 (A) (T) (1)
2
where: 484 = area of coverslip, in mm
N = number of cells counted
M = total sample volume, ml
A = area scanned, mm
0.05 = volume of 1 drop, ml
T = area sampled, mm
For interspecific comparisons cell densities were converted to volume of
living cells. This calculation was made by measuring 50 cells of each
species and estimating the average volume per cell using regular geometrical
shapes including spheres, cylinders and ellipsoids.
Aufwuchs biomass was determined for a 10.0 ml aliquot from the mixed
slurry described above. Aliquots were placed into pre-fired, pre-weighed
crucibles, dried at 100° for 24 hr, cooled in a dessicator and reweighed.
Biomass in grams per square meter of the original substrate (glass slide,
wall or bottom) was calculated using equation 2.
Biomass =
20
-------
where: M = total sample volume, ml
W = sample dry weight, g
2
T = sample area, m
Concentrations of chlorophylls a and b, carotenoids and phaeophytin pig-
ment ratios were determined using the acetone extraction method of Strickland
and Parsons (1972). Ten ml of mixed algal solution was filtered at 0.5 ATM
through Gelman A-E glass fiber filters with a small amount of saturated MgSO^
added as a buffer. The filter was ground in a glass tissue homogenizer,
using a teflon pestle with several milliliters of 90% reagent grade acetone
until it was completely disassociated. The total volume was adjusted to 10
ml with 90% acetone. The grinding tube was kept in a ice-water bath through-
out the grinding period (approximately one minute). Blended samples were
allowed to extract in the dark at 4°C for 24 hours. At that time the samples
were centrifuged, decanted and re-centrifuged. Absorbance of clarified
samples was measured at 750, 663, 645, 630, and 480 run in a one centimeter
cell using a Beckman ACTA-CIII Spectrophotometer. Extracts were acidified
with one drop of 0.1 M HC1, and their absorbance remeasured at 750 and 663
nm. Absorbance values were corrected for turbidity with the 750 run absor-
bance and background absorbance. Chlorophylls a_ and b and carotenoids were
calculated according to Strickland and Parsons (1972). Acidification ratio
was calculated by dividing the corrected 663 acidified absorbance into the
663 unacidified value.
Dried periphyton samples were wet ashed in 30 ml porcelain crucibles
with 2 ml of concentrated HNO at 80 C for 1 to 3 hours or until all solid
material had dissolved and No. evolution ceased. The samples were cooled,
two ml of 30% HO added, and reheated until gas evolution ceased. Samples
were cooled to room temperature, diluted volumetrically using deionized
water, and stored in washed polyethylene bottles. Analytical methods for
cadmium are described in Appendix I.
Due to the heterogeneous assembly of organisms composing the aufwuchs
community in the channels, an estimate of non-algal material was desired.
Utilizing the estimation of algal volume and total biomass values suitably
converted to live volumes, a percentage live algae was calculated for each
sample. The live volume:biomass ratio was measured for a young growth of
filamentous algae by lightly centrifuging cells from suspension, measuring
their volume and determining dry weight. Dry weight (g) was 1.74 percent of
live volume (cm ). Percentage algal composition of the aufwuchs community
was determined by dividing the volume of green algae (Chlorophyta) or blue-
green algae (Cyanophyta) by total algal volume. Through the study these two
groups together represented more than 95% of the algae present.
Preliminary comparisons of data have been limited to differences between
treatments with replicate streams and head and tail locations pooled. At
each sampling date the three values have been graphed and a test statistic
calculated for comparison between any two of the means.
21
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RESULTS AND DISCUSSION
Cd Accumulation
Cadmium levels reported for aufwuchs are on a dry weight basis for the
entire community. At continuous Cd exposures of 0.05, 5.0 and 10.0 |Jg Cd/1,
steady state concentrations in aufwuchs from long-term glass slides were
approximately 3, 36, and 58 |jg Cd/g dry weight (Fig. 3).
Concentration factors for Cd by aufwuchs were 7100X when exposed to 5
ppb and 5800X when exposed to 10 ppb which are similar to concentration
factors reported by Gerhards and Weller (1977). Linear relationships between
Cd concentration in culture media and algae have been reported by several
workers (Gerhards and Weller, 1977; Payer et al., 1976; Kutagiri, 1975; and
Kerfoot and Jacobs; 1976) however, the Cd concentration factor in the control
channels was approximately 64,000 or 10 times the concentration factor for
the higher Cd exposure concentrations. Other workers have found concentra-
tion factors of 500X for Chlorella pyrenoidosa (Hart and Cook, 1975); > 2000X
for Analystis niclulans (Katagiri, 1975); 4000 - 6700X for marine diatoms
(Kerfoot and Jacobs, 1976); 1000 - 2000X for bacteria and fungi (Doyle et
al. , 1975) 80000X for mixed algae (Kumada et al., 1973) and 10000X for marine
phytoplankton (Knauer and Martin, 1973). Since Cd concentration factors are
LONG TERM GLASS SLIDES
o»
\
T3
O
D>
g
i-
cr
i-
z
UJ
o
z
o
o
o
N
DNlF|M|A|M IJIJ I A |S | 0 ) N | D|J I F|M I A |M I J
1976 1977
SAMPLING DATE
Figure 3. Mean Cd concentrations in aufwuchs collected from long term glass
slides incubated in the channels from the beginning of Cd exposure.
22
-------
related to available Cd in the medium and thus affected by the chemical and
physical form which is determined by particulates, dissolved organics, water
hardness (Kinkade and Erdman, 1975), orthophosphate levels (Motohashi and
Tsuchida, 1974) and undoubtably to other chemical parameters, which affect
the form in which Cd exists in water, it is not surprising to find a large
range of factors reported for various algae and fungi.
Initial Cd accumulation was not measured but a charge up curve was
observed with equilibrium reached within approximately 50 days (Figure 3).
Katagiri (1975) and Kerfoot and Jacobs (1976) reported Cd accumulation by
algae could be explained by a first order uptake model, which is consistent
with our data.
The Cd concentrations reached in short term accrual experiments (23
days) (Figure 4) were essentially the same as the ambient steady state Cd
concentrations observed on the long-term slides (Figure 3); indicating Cd
uptake by the aufwuchs community was rapid. After Cd inputs ceased, water
concentrations dropped to control levels within a few days. Cd concentra-
tions on short-term glass slides incubated in the former treatment streams
after input was stopped were not significantly different from controls.
100
_ 90
o>
T3 80
o
5 70
1 60
2 50
H
Z 40
UJ
o
z 30
O
o
T3 20
o
SHORT-TERM GLASS SLIDES
• • Control
a 6 5/x.cj Cd/L
D a lOuq Cd/L
N I D JIFIMIA MIJIJ A S 0 N D J F M| A | M| J | J A
1976
1977
SAMPLING DATE
Figure 4. Mean Cd concentrations in aufwuchs collected from short term
glass slides incubated in the channels for the eight weeks
prior to sampling.
23
-------
Cadmium uptake rate was calculated (Table 8) assuming a Von Bertalanfi growth
model to describe Cd accumulation by aufwuchs (equations 3 - 6). Cadmium
TABLE 8. STEADY STATE CD CONCENTRATIONS IN AUFWUCHS AND UPTAKE
AND ELIMINATION CONSTANTS
TREATMENT
(Mg Cl/£)
STEADY STATE
LEVEL
(Mg Cd g )
UPTAKE
RATEj
(pg Cd g day )
DECAY
CONSTANT
(day"1)
5
10
36
58
2.1
3.9
.06
.07
kQ
Qt = ;(
for steady state:
(3)
(4)
(5)
ss
J = Q k
ss
(6)
where:
Q = Cd concentration in aufwuchs
0 = steady state Cd concentration in aufwuchs
ss
J = Cd uptake rate
K = Cd elimination rate
accumulation in the benthic aufwuchs was also determined on two occasions
during Cd input using short term glass slides. The average values by treat-
ment were: controls 8, 5 pg Cd/£ - 75, and 10 pg Cd/A - 116 pg Cd/g of
ash-free dry weight. These values are.approximately double the values found
for the aufwuchs populations or vertical substrates and may be due to the
relatively slower flushing of biological material from this storage.
24
-------
Cadmium decay from the aufwuchs storage was followed in detail and data
from the walls and glass slides is combined in a semi-log plot in Figure 5.
The best linear fit for this data was found to be a single logarithmic decay
for each treatment. Assuming a linear decay model, half-life values of 11.8
and 10.4 days were found for 5 and 10 |jg Cd/£ channels respectively. Al-
though the control aufwuchs appeared to lose cadmium during this period, the
slope of the best-fit line is not significantly different from zero at the
95% confidence level.
Community Structure
After 20 months exposure to Cd, aufwuchs biomass was still increasing on
long-term glass slides (Figure 6) and channel walls (Figure 7). Ultimate
differences between standing crops on glass slides and walls indicate that
glass slides underestimate standing crops on an areal basis at high aufwuchs
densities. Before Cd input began, no appreciable differences were observed
in aufwuchs standing crops between streams. Within two months after Cd input
began, aufwuchs standing crops in channels receiving Cd were significantly
lower than those in control channels (Figure 6). Standing crop values in the
four treated channels remained similar to each other, but significantly lower
than controls for five months at which time within treatment variance began
to mask significant differences. After ten months of continuous Cd input,
mean aufwuchs standing crops were similar across all treatments. Benthic
aufwuchs samples measured after 11 months of Cd input had much greater bio-
mass levels (controls 157, 5 pg Cd/SL 177, and 10 pg Cd/£ - 172 g ash-free
dry weigh/ m ) than the vertical substrates and also showed no significant
differences between treatments.
Figures 8 and 9 summarize live algal volume on long-term glass slides
and walls indicating that the apparent recovery observed for total aufwuchs
biomass was not the result of algal recovery. Total algal volume declined in
the treated channels shortly after Cd input began and remained at constant
low levels throughout the rest of the study. Algal volume in the control
channels was significantly greater than in the treated channels throughout
the Cd input period, exhibiting a spring minimum and a late fall-early winter
maximum. -In_ bottom samples this trend was also ,pbs£rved with an average value
of 22 cm /m in the controls and 13 and 10 cm /m seen in the 5 and 10 pg
Cd/£ treatments, respectively.
Due to the similar effects of 5 and 10 |jg Cd/1 on algal volume, the
required Cd concentration to depress community algal standing crop is less
than 5 pg Cd/A. Klass et aJL. (1974) reported 6 pg Cd/£ reduced Scenedesmis
gradricauda growth and Katagiri (1975) observed growth inhibition of
Anacystis nidulans at 50 pg Cd/£. Conversely, Hart and Cook (1975) reported
growth stimulation of natural phytoplankton populations by 11 to 110 pg Cd/£.
This may have been an indirect effect due to reduced grazing by zooplankton,
which are very sensitive to Cd (Giesy et al., 1977; Marshall, 1977).
Aufwuchs standing crops showed little difference between treatments
after 10 months of Cd exposure, however, algal population densities were
significantly lower throughout the exposure period in channels receiving Cd.
25
-------
2.0 r
_i
o
o
2 1.0
o:
I-
z
LU
O
O
o
-o
O
0.0
15
D
D
20 25
MARCH
30
DAY
• Control
A 5/ig Cd/L
a 10 MQ Cd/L
APRIL
14
Figure 5. Linear regression of Cd elimination from the aufwuchs community colonizing
glass slides.
-------
N5
01
E
X.
o>
8'°
o
m
CO
I 5
LONG-TERM GLASS SLIDES
• • Control
*— * 5 /*g Cd/L
a o io/ig Cd/L
N DJFMAMJ
A S 0 N D
1976
SAMPLING DATE
1977
M
Figure 6. Mean aufwuchs biomass acrual on long term glass slides incubated from the
beginning of the experiment with confidence intervals indicated.
-------
N)
00
CO
E
70
60
CO
co 50
co
X
o
30
20
10
•—• Control
e>—^ 5figCd/L
0—° 10/igCd/L
WALL SAMPLES
1976
J
J
A
S
0
N \ D
J
F
M | A
M
J
J
1977
SAMPLING DATE
Figure 7. Mean aufwuchs biomass accrual on channel walls with two standard error
confidence intervals indicated.
-------
VD
CM
"
7.0
Q.O
5.0
£
^
uJ 4.0
§ 3.0
2.0
1.0
0.0
LONG-TERM GLASS SLIDES
•—• Control
A—a 5 fig Cd/L
a—a 10 fig Cd/L
, I I
N
D
J
F
M
A
M
J
J
A
S
0
N
D
J
F
M
1976
SAMPLING DATE
1977
Figure 8. Mean viable algal cell volume collected from long term glass slides incu-
bated from the beginning of the experiment with two standard error confi-
dence intervals indicated.
-------
CM
E
o
UJ
2.0
10.0
8.0
6.0
4.0
2.0
0.0
WALL SAMPLES
Control
Cd/L
Cd/L
1976
J
J
A
S
0
N
D
J
F
M
A
M
J
J
1977
SAMPLING DATE
Figure 9. Mean viable algal cell volume collected from channel walls with two
standard error confidence intervals indicated.
-------
This decrease in the relative importance of the algal component in the
aufwuchs community is reflected in a decreased chlorophyll £: aufwuchs biomass
ratio (Fig. 10) which declined after initial colonization, due to accrual of
photosynthetically inactive algae and heterotrophic organisms. This ratio
was relatively consistent in all channels throughout the study. The chloro-
phyll: biomass ratio varied seasonally with significantly lower values during
early summer and higher values during winter. Using an algal dry weight/
algal live volume X 100% value of 1.74%, determined for a relatively pure
culture of stream algae, we found that throughout the study live algae made
up 2% or less of the total aufwuchs dry weight.
Throughout most of the period of cadmium input, the aufwuchs communities
were visably different in color. Communities in control channels were green
to black while those in channels receiving Cd were orange-yellow. This ob-
servation was quantitatively verified by comparison of chlorophyll a to
carotenoid pigments (Figure 11). In the control channels, a fall-winter
maximum in the chlorophyll a:carotenoid ratio was observed corresponding to
the period of highest algal standing crop. Communities in channels treated
with Cd had significantly lower chlorophyll a:carotenoid ratios. Margalef
(1961) and Odum and Hoskins, (1957) found that lower chlorophyll a:carotenoid
ratios indicate a shortage of available nutrients. It is possible that
nutrient recycling was limited by Cd and, therefore, algal populations were
nutrient-starved. (See section XI). However, water nutrient levels were
generally higher in channels receiving Cd than in control channels. An ex-
planation for this paradox may be that most of the aufwuchs were actually ex-
posed to much lower soluble nutrient concentrations under the surface layer
of the mat and were dependent upon internal nutrient cycling.
Diatoms were rare in all samples from walls and glass slides regardless
of treatment although several diatom species were observed in protozoan
sponge samples. Nearly 100% of the algae were green (chlorophyta) or blue-
green (Cyanophyta). Algal dominance shifted towards blue-greens with suc-
cessional development (Figure 12). Samples from channel walls indicated a
significantly higher percentage of blue-green algae in the Cd-treated chan-
nels .
Initial colonization in all channels was dominated by Oscillatoria
geminata (filamentous blue-green), Geminella turfosa (filamentous green) and
Stigeoclonium elongatum (filamentous green). These three species as well as
several other filamentous greens and blue-greens, unicellular greens and
blue-greens, and five desmid species were common in all channels throughout
the study. A total list of algal species collected during the study is pre-
sented in Appendix (II).
Species diversity was calculated for the algal component of each
aufwuchs sample using the formula derived from information theory (Pielou,
1969). Diversity (H) initially decreased and then increased through the
spring and summer to a plateau (Figure 13). The trend observed for diversity
was largely due to changes in eveness of the algal population distribution as
opposed to colonization by larger numbers of species (Figure 14). A few
months after Cd input began, diversity values were significantly lowered in
31
-------
to
(O
o
E 7.0
o
60
ol
I 5.0
o
g" 4.0
O
ri 3.0
oc
2.0
< 1.0
0.0
LONG-TERM GLASS SLIDES
~A1SR5~
WALLS
Walls, Slides
Control
•- -m , D — o 10
Cd/L
\i i i I i i i I I II I III I i i I
N|D|J|F|M|A|M|J|J|A| s [ O|N|D|J |F|M|A|M|J
1976
1977
SAMPLING DATE
Figure 10. Algal ratio for aufwuchs collected from both long term glass slides and
channel walls with two standard error confidence limits reported for
long term glass slide samples.
-------
u>
OJ
4.0
O
c
Q>
•4—
O
k.
O
^
01
3.0
U
O 2.0
1.0
Q.
0.0
LONG-TERM GLASS SLIDES AND WALLS
Ill Ix III11IIIII 111 II I
Slides
Walls
• Control
^ 5 /tg Cd / L
o a 10 pg Cd / L • •
N
D
J
F
M
A
M
J
J
A
S
0
N
D
J
F
M
A
M
J
J
1976
SAMPLING DATE
1977
Figure 11. Pigment ratio for aufwuchs collected from both long term glass slides
and channel walls with two standard errors confidence limits reported
for glass wall samples.
-------
LONG-TERM GLASS SLIDES AND WALLS
u>
100
90
f? 80
J 70
O
g 60
O 50
Z 40
UJ
DC 30
UJ
°~ 20
10
0
Slides Walls
• • Control o o
6—* 5Mg Cd/L *-"*
o—o 10ig Cd/L •-•-•
: 1111 x i x x
V—
ill I
N D J FMA M J J A S ON D J FM AM J J
1976
1977
SAMPLING DATE
Figure 12. Percent of algal community, collected from long term glass slides and
channel walls comprised of green algae with two standard errors confi-
dence limits reported for long term glass samples.
-------
u>
4.0
3.0
2.0
en
a:
LJ
1.0
0.0
LONG-TERM GLASS SLIDES AND WALLS
I n 11111II inh mil i
1976
Sitetes Walls
• • Control o o
Cd/L * *
D o |0/*g Cd/L» •
SAMPLING DATE
1977
Figure 13. Diversity values for the algal community colonizing long term glass
slides and channel walls with two standard errors confidence intervals
indicated for long term glass slide samples.
-------
u>
CO
CO
LJ
2
LJ
>
U
I.Or
0.9
0.8
0.7
0.6
0.5
0.4
0.3
0.2
O.I
0.0
LONG-TERM GLASS SLIDES AND WALLS
I I I I I I III III 111 I I I
N D J F MA M J J A SONDJ F MA MJ J
1976
1977
SAMPLING DATE
Figure 14. Evenness values for the algal community colonizing long term glass slides
and channel walls with two standard errors confidence intervals indicated
for long term glass slide samples.
-------
the channels receiving 10 |Jg Cd/£. Diversity reduction was less extreme at 5
pg Cd/£ and disappeared after seven months of Cd exposure.
Interactions
During the first 18 months of stream succession, aufwuchs provided the
largest reduced carbon component for heterotrophic metabolism, and as indi-
cated in Figures 6 and 7 was continuing to increase in standing stock at the
end of the study. However, the total living algal contribution to the
aufwuchs had apparently levelled off by the first year at which time the
other autotrophic component of the streams (macrophytes) were rapidly in-
creasing. It is possible that total algal volume in the streams would have
continued to increase at a rate limited only by the increase in attachment
surface area created by macrophytic plants.
All other organisms residing in the streams were by necessity dependent
on algal and associated aufwuchs growth and would be expected to demonstrate
indirect responses to Cd's effect on the aufwuchs component. Although the
viable algal populations never showed recovery from Cd toxicity during the
year after Cd input, aufwuchs levels recovered to comparable levels in all
channels. The standing crops appear to have been largely limited by physical
forces such as water current, torrential rains, and severe winds acting on
them. In one respect we may now look at the densities of various consumers
and try to understand how they were limited by algal populations (they should
be lower in the treated channels) or by total aufwuchs biomass (initially
lower in treated channels but gradually recovering during Cd input). How-
ever, we also have quite a different possibility that one might easily over-
look. That is, the possibility that the consumers were directly affected by
Cd treatment levels and a large part of the algal response was an indirect
reaction to Cd, mitigated by decreased cycling capabilities of nutrients due
to lowered ecosystem complexity.
We can examine these two possibilities and try to determine if one is
more likely than the other based on the available data. The organisms found
in the channels that would be dependent upon algae for nutrition and upon
which the algae might be dependent for cycling of nutrients include: bac-
teria, fungi, protozoans, invertebrates and fish. For bacterial populations
we have no pertinent information except for the specific group of nitrogen
fixers. There was some indication that these organisms were adversely affec-
ted by Cd. Most likely this was a direct toxicity effect because of their
ability to fix N_ in the absence of an organic carbon source.
Fungi colonizing leaves in the tail pools were severely inhibited by Cd
treatment (See Section XI). Since these organisms were given a allocthanous
food source (dead tree leaves) their reaction to Cd would not be the result
of a lack of energy, but rather a direct toxicity effect.
Microinvertebrate populations showed variable results with micro-
crustaceas and Difflugia reduced by Cd treatment and flagellates, cilliates
and rotifers reaching higher population densities in Cd treated channels.
Decreasing microcrustacean population densities in Cd treated channels may
have been a result of reduced algal populations, since the single species of
37
-------
copepod and ostracod seen were vegetarian by nature. However the initial
total disappearance of these organisms when Cd was added to the channels may
more likely be the result of a direct toxicity effect which was mitigated by
successional evolution in the streams, allowing their partial recovery with
time. The increase in cilliate, flagellate, and roifer populations could not
be the result of lower algal populations but may have contributed to those
lower populations. Rather, as suggested elsewhere in this report, their
increased populations may have been the result of tolerance to Cd toxicity as
well as decreased predation and disease.
Macroinvertebrate populations also showed considerable differences be-
tween treatments and controls that may have affected algal populations. May-
flies and annelid worms were greatly decreased in the treatment channels and
because of their food habits being largely non-algal, it may be assumed that
their decreases were the result of direct Cd toxicity. The organisms which
feed heavily on algae, e.g. the dipterans (Chironomidae and Ceratopogonidae)
were stimulated to higher levels in the Cd streams in spite of lower algal
populations. This stimulation of herbivores due to lack of competition and
predators may have been largely responsible for the decreased algal popula-
tions observed.
Crayfish and Gambusia in Cd treated streams exhibited the greatest re-
sponse to direct Cd toxicity. These populations were severely reduced com-
pared to controls and indirect effects of these reductions were plainly seen.
Reduced populations of crayfish in the treated head pools resulted in greatly
increased macrophytic growth and a lowering of nutrient inputs to the treated
streams while the loss of Gambusia from the treated streams no doubt played a
big part in the increased herbivore populations.
So why then was the overall result of Cd input the reduction of algal
standing crop and consequently primary production (and respiration)? Algal
species capable of withstanding the direct toxicity effects of Cd were in the
treated streams as evidenced by luxuriant growths at isolated times and
locations. What were the effects of Cd that indirectly inhibited algae and
primary production? A likely explanation in need of further investigation is
the effect of Cd on the higher organisms, the heterotrophs or consumers, and
with the disruption of their balanced web of relationships by an exogenous
poison and the subsequent lowering of sustainable autocthonous energy fixa-
tion. Several studies with widely different organisms have shown that pri-
mary production may be maximum at levels of consumers greater than zero but
less than maximum possible populations (Cooper, 1973; Hargrave 1970; Flintard
and Goldman, 1975). Normally a system will be forced to adjust population
densities of the various consumers in a series of feedback steps, each lead-
ing to a slightly greater fixation of utilizable energy. In the time scale
of our study this natural system selection could be carried only so far
because the ultimate consumer (Cd) was being maintained at a fixed level
without feedback control. Natural systems, receiving a toxin such as Cd,
might indeed over a long enough time derive mechanisms controlling the
toxin's concentration. Possible examples of mechanisms already found for Cd
are sequestering of the element in non-living materials such as humics and
losses through sedimentation. Natural systems may be adapted to use environ-
38
-------
mental levels of Cd as a controller, or consumer in their tuned networks
resulting in maximum sustainable energy utilization. Thus the difficult
question of what attributes of a system should be protected (see Section
XIV).
39
-------
SECTION VIII
MACROPHYTES
INTRODUCTION
In shallow standing or sluggishly moving water, emergent macrophytes are
often a dominant feature of aquatic communities. These macrophytes exert a
strong influence on the community in several ways. Their most obvious con-
tribution is in the production of fixed carbon which is available to the
heterotrophic system components. These plants may also exert a strong con-
trolling influence by their utilization of available nutrients and light and
the release of soluble organic compounds which then affect the algal compo-
nents of the ecosystem. Also important is the physical habitat the macro-
phytes provide for invertebrates and small vertebrates. With respect to the
relative importance of macrophytes, Westlake (1975) states that the role of
macrophytes in the aquatic community lies more in their role in modifying and
diversifying habitats than in the supply of organic matter.
Aquatic macrophytes are able to concentrate Cd from both water and
sediments (Harding and Whitton, 1978) and may serve as a source of Cd to
herbivore populations.
Macrophyte populations were monitored to determine the effects of Cd on
colonization and growth as well as Cd accumulation by macrophytes.
METHODS
Clumps of Juncus diffusissimus plants growing in a pond adjacent to the
artificial streams facility were transplanted into the channels and tail
pools on March 15, 1976. Thirty clumps spaced at 2 m intervals were put into
the channels and 5 clumps were put into the tail pools, which at the time
contained crayfish. The plants introduced into the tail pools were quickly
eliminated by the crayfish except in one case (tail pool 6) where there was
100% mortality of the crayfish. This supports the hypothesis that the early
natural colonizing macrophytes in the channels were eliminated by the cray-
fish. The number of live shoots and height of the tallest shoot in each
clump placed in the channels was monitored monthly until October 1976.
"Live" shoots were considered to be those shoots extending to or upwards from
the water line and containing chlorophyll. Emergent and submergent shoot
samples were taken monthly from three clumps for Cd analysis. No attempt was
made to take root samples because of the damage this would have done to the
plants.
40
-------
By June of 1976, sufficient numbers of large J. diffusissimus had de-
veloped in the head pools for destructive sampling of these populations to be
initiated. Starting at that time, a single plant was removed monthly from
each pool, divided into shoot and root portions, dried, and subsamples taken
for digestion and subsequent Cd analysis. In September 1976, a similar sam-
pling program was initiated for J. diffusissimus naturally colonizing the
channels. Individual plants were removed from both ends of each channel and
prepared for Cd analysis. Callitriche heterophylla became relatively common
in the channels and pools by November 1976, and a monthly sampling program
for this species was begun at this time, again both upstream and downstream
stations in each channel were sampled.
By the time Cd input was terminated (March 1977), population densities
of naturally colonizing macrophytes were high enough so that plant biomass
sampling by quadrat analysis was feasible. A survey was made in. conjunction
with a large scale invertebrate sampling program. Ten 0.25 m sections of
sediment and associated plants were removed from each channel. A logarithmic
sampling distribution was used so that most samples were taken from the up-
stream reaches of the channels, which were most heavily colonized. All
plants were washed, sorted by species, counted, dried in a forced air oven at
100 C, and weighed to the nearest 0.1 g. From these data, macrophyte biomass
per unit area was calculated. The entire sampling procedure was repeated
using different quadrats in September 1977 in order to ascertain any changes
in macrophyte biomass that occurred after six months of recovery.
All samples taken for Cd analysis were rinsed free of sediment and
periphyton, and placed in plastic bags in which small holes had been punched.
After the samples were freeze dried to constant weight, subsamples of 0.05
-0.10 g were refluxed in previously fired (900 C, 1 hr.) porcelain crucibles
with 2 ml redistilled concentrated HNC* at 85 C on a hotplate until evolution
of NO- ceased. Samples were cooled to room temperature, treated with 1 ml
30% H^CL, heated until clear, cooled, and diluted with repeated deionized
water washings of the crucibles.
RESULTS AND DISCUSSION
In a previous study with the channel microcosms (Kania et al., 1976),
the emergent macrophyte J. diffusissimus became an increasingly important
component of the channel communities as time progressed, especially in the
upstream reaches. The growth of this rush at the heads of the control
channels was so extensive after two and one-half years that the water flows
became restricted and the systems had to be channelized. Because of the
seeding technique used for this study, the same Juncus was expected to again
become a dominant community member and the macrophyte analytical program was
designed with this in mind.
Shortly after the initial seeding of the channels in October 1975, two
types of macrophyte seedlings were observed in the channels. These plants
persisted only until December, however, and then disappeared. The disappear-
ance may have been caused by the feeding activity of the crayfish introduced
into the channels in December or the onset of cold weather, or the plants may
41
-------
simply not have been suited to the environment of the channels. By the end
of January 1976, the channels and tail pools were completely devoid of rooted
macrophytes of any kind although the head pools contained a relatively dense
growth of young macrophytes (Table 9). The persistence of macrophytes in the
head pools which had no crayfish indicates that crayfish were responsible for
the elimination of macrophytes in the channels and tail pools.
Data obtained during the first three months of sampling from the trans-
planted Juncus diffusissimus clumps indicated that the plants in the treated
channels may have been losing shoots faster than those in the control
channels.
TABLE 9. DISTRIBUTION AND NUMBER OF INDIVIDUAL MACROPHYTES IN HEAD POOLS
AS OF JANUARY 31, 1976
Juncus
diffusissimus
Gratiola
virginiana
Callitriche
heterophylla
Bacopa
caroliniana
Head
Pool
1
2
3
4
5
6
227
499
439
396
576
429
26
18
19
8
15
22
1
1
0
1
2
1
0
0
0
0
1
2
This phenomenon disappeared, however, during the following four month
period, during which all clumps produced new shoots in about equal numbers.
No differences in macrophyte growth rate were noted between treatment or con-
trol channels at any time. All measurements on the transplanted clumps were
discontinued after October 1976 because clumps were too large and overgrown
with algae to be effectively counted and measured.
By September 1977 when Juncus naturally colonizing the channels was
sampled for Cd, there were major differences between treated and untreated
systems with respect to macrophyte populations (Figure 15). Control channels
had many more plants than channels receiving Cd inputs. However, there were
no apparent differences between the 5 |J g/£ and 10 (j g/£ treatments. These
observations were confirmed by the macrophyte biomass sampling carried out in
March 1977 (Figure 16). Figure 16 also shows the distribution of the macro-
phytes in the channels with the greater population densities existing near
the input wiers. This same pattern was observed in a previous study (Kania
et al., 1976).
42
-------
300
200
100
Macrophyte Biomass -September 1977
Cd/L
j?
CJ
E
OS
100
400
OJ
300-
200-
100-
Ql-r
A' \
Cd/L
=f*^F
Control
20 40 60 80 100
DISTANCE FROM INPUT.m
Figure 15. Macrophyte standing crop biomass as
a function of distance from the head
pools, as of September 1977.
lOOr
Macrophyte Biomass - March 1977
i 1 1 1 1
0
DISTANCE FROMINPUT.m
Figure 16. Macrophyte standing crop biomass as
a function of distance from the head-
pools, as of March 1977
-------
The biomass in all systems approximately doubled in the six month period
with no cadmium inputs (Figure 15), with some trend toward recovery indicated
in at least the 10 p g/£ treatments (Figure 16). The macrophyte distribution
in the channels remained unchanged. The colonization pattern observed was
probably due to turbulence near the head and distance from seed source and
nutrient input.
Fleischer et al. (1974) state in their comprehensive review of Cd in the
environment that "reports of cadmium toxicity symptoms in plants grown under
field conditions have not been found." This is probably because the neces-
sary long term studies that include several species have not previously been
done. Because of the possibility of food chain transfer of Cd to humans a
number of laboratory uptake and plant toxicity studies have been done with
agricultural plants (John et al. , 1972; Page e_t al. , 1972; Francis and Rush,
1973; Haghiri, 1973; Turner, 1973; John et aTT, 1976; Petterson, 1976;
Koeppe, 1977; Reddy and Patrick, 1977; Wallace et a_l. , 1977) including forage
species (Bingham et al., 1976). These studies, which have in general been
carried out with extremely high levels of Cd, have demonstrated that the most
common response of plants to Cd is reduced growth (i.e. yield) although
chlorosis has also been reported, and that the levels of exposure required to
elicite a toxic response varies tremendously not only with species but even
with variety (John e_t al. , 1976) .
The amount of information on Cd effects on aquatic plant species is
limited. There have been two recent studies (which have included some in-
vestigation of effects) although they were primarily concerned with Cd uptake
by Spartina (Dunstan and Windom, 1975; Dunstan et al., 1975). These studies
showed that germination of Spartina seeds was not affected by Cd exposure
concentrations of up to 100 pg/iufc. In growth studies over an eight week
period, this concentration had no effect on growth rate or net primary pro-
duction. It is not clear as to whether the sensitivity of J. diffusissimus
to Cd as demonstrated in our study was simply a species difference or related
to the softwater medium in which the Cd was presented. Cearley and Coleman
(1973) working with the fresh water naiad Najas quadulepensis found that
plants exposed to Cd levels as low as 7 Mg/£ demonstrated reductions of
chlorophyll, turgor, and stolen development, although they do not relate the
extent of these responses to the doses used.
CADMIUM ACCUMULATION
The cadmium concentrations measured in macrophytes from the control
channels and head pools (Table 10) were similar to those reported by other
workers for freshwater macrophytes from uncontaminated areas (Cearley and
Coleman, 1973; Gommes and Muntau, 1976; Lee et a 1. , 1976) but are generally
higher than those reported for a marine form (Dunstan and Windom, 1975;
Dunstan et al. , 1975). The two species studied here appear to be very simi-
lar in their ability to concentrate Cd from dilute solutions (Figure 17, 18,
19, 20 and 21. In both species, roots were in all cases higher than the
leaves, (Figure 17, 18, 20 and 21) an observation made by others for aquatic
plants (Lee e_t al. , 1976; Gommes and Muntau, 1976).
44
-------
TABLE 10. CADMIUM CONCENTRATIONS IN MACROPHYTES REMOVED FROM
CHANNELS NOT RECEIVING CD.
Sample
Callitriche heterophylla
Natural colonizers
Leaves
Roots
Juncus diffusissimus
Natural colonizers
(channels)
Leaves
Roots
(Head pools)
Leaves
Roots
Transplants
Leaves
(Emergent)
(Submergent)
N x SD CV
ug/g %
10 1.05 1.94 185
28 8.55 6.96 81
54 1.48 0.96 65
55 6.19 3.51 56
32 0.70 0.92 131
31 2.43 1.44 59
94 0.61 0.84 138
94 0.75 0.98 130
45
-------
_t/. diffusissimus Colonizing Head Pools
_ 100
O
0
300
nj 200
o
O
o
O
100
LEAVES
JJASONDJ FMAMJJA
ROOTS
-a 5p.q Cd/L
Cd/L
/ */X
' / \
i \
I 1' n I
\
*—-j
\ V
J 1 J
A I S| 0
N | D
j |F
M
A
M
J
J
A
1976
1977
SAMPLING DATE
Figure 17. Cadmium concentrations in J. diffusissimus colonizing the headpools
expressed on a dry weight basis.
-------
100
~ 0
V.
-o
J. diffusissimus Colonizing Channels
LEAVES
Y^J^Y^Yo^^f
M 1 A IMIJ I J I A |
400
o>
2
- 300
a:
o 200
O
o
O
100
ROOTS
/
\
\
A
*r-+ 5fiq Cd/L
- D~O IO/igCd/L
\
\
\ \
SlOlNlDlJlFlMlAlMlJljlAl
1976
1977
SAMPLING DATE
Figure 18. Cadmium concentrations in J[. diffusissimus
colonizing the channels, expressed on a dry
weight basis.
47
-------
00
50 r
T3
O
O
I-
<
LU
O
30
20
O 10
O
O
0
J. diffusissimus Transplanted to Channels
WHOLE LEAF
5/ig Cd/L
Cd/L
,H
\
h /
A\
\ \
\
A I M { J I J | A \S\ OrN|D[J|F|M[A|M[J|J|A|
1976
1977
SAMPLING DATE
Figure 19. Cadmium concentrations in J_. diffusissimus transplanted to the channels,
expressed on a dry weight basis.
-------
I£U
1 100
Cd CONCENTRATION (/tg C
ro 01 ->i
01 O 01
a C. heterophylla
\ COLONIZING CHANNELS
-°V / \~o LEAVES
^v !
- "y s
N A (
NNX / \ 1 A- ^ 5/ig Cd/L
X / ^"-^ i ° D 10/i.g Cd/L
*r
1\
y/^^/^^^^^^r^^/'^f^/y^'^/f'. *" ^^
N|DJ FM AM J J A
1976 1977
SAMPLING DATE
Figure 20. Cadmium concentrations in C. heterophylla shoots colonizing the channels,
expressed on a dry weight basis.
-------
C. heterophylla
COLONIZING CHANNELS
^ 1000
\
0
a! 800
z
O
1- 600
*
UJ 400
O
o
-o 200
O
V I OUU.f O
J ROOTS
: 'n
W
N\
III A A C « /^ *J / 1
iji i o — 6 3 fig oo/L
|0 1
,'| D— o lOug Cd/L
1 ' \
P — o— ._A'i \
/ ! \ A
v_ g/ ^fNu/ \
xx-'x "*" i; ^x \^__
'* & \N ~~° — —o
\
7 -a a *
N | D | J F M|A M | J | J A
1976
1977
SAMPLING DATE
Figure 21. Cadmium concentrations in C. heterophylla roots colonizing channels,
expressed on a dry weight basis.
-------
J. diffusissimus plants transplanted into the channels were lower in Cd
throughout the study than the naturally colonizing forms that developed
there. The sediment in the pond from which the transplanted J. diffusissimus
were removed was much higher in organic matter content and clay than the
channel sediments. J. diffusissimus that colonized the highly organic natu-
ral sediments in the head pools had lower Cd concentrations than those in the
channels and quite similar to the transplanted plants. In the transplanted
J. diffusissimus there were no differences between submergent and emergent
portions of leaves with respect to Cd concentrations, and the concentrations
observed are not to any extent due to loosely adsorbed Cd or Cd associated
with unremoved periphyton.
No difference was observed between emergent and submergent portions of
leaves of transplanted J. diffusissimus so these have been averaged (Figure
19). In Cd treated systems, the naturally colonizing Callitriche hetero-
phylla and J. diffusissimus leaves and roots were at equilibrium at the time
sampling was initiated (Figure 17, 18, 20 and 21). This is not surprising
considering the rapid rate at which at least some aquatic plants can take up
this metal (Wolverton, 1975; Cearley and Coleman, 1973). We did not observe
the great species differences cited by other workers (Petterson, 1976;
Fleischer e_t al. , 1974), and in the channels, both species concentrated Cd to
a similar degree although the roots of C_. heterophylla were generally con-
tained greater Cd concentrations than did those of J. diffusissimus. This
may be due to the finer roots of C. heterophylla and thus greater potential
uptake surface.
In all samples roots contained greater Cd concentrations than did
leaves. Also, naturally colonizing Juncus in the head pools (Figure 17)
accumulated less Cd, than those growing in the channels (Figure 18). Cadmium
uptake patterns were different in these two regions in several ways; (1)
there was little difference in Cd uptakes by J. diffusissimus growing in the
pools exposed to 5 pg Cd/£ or 10 |Jg Cd/£. Conversely, Cd uptake by J.
diffusissimus growing in the channels was approximately proportional to that
which they were exposed, (2) there was a general increase in Cd levels in J.
diffusissimus roots growing in the headpools during exposure to Cd, while
there was a decrease in root Cd concentrations in plants growing in the
channels, (3) Cd concentrations in roots, especially those made after Cd in-
puts were terminated, were much more variable in samples from the head pools
than in those from the channels.
During the five month period after Cd inputs were stopped, Cd concen-
trations in plants growing in both the head pools and channels declined to
similar levels, even though sediment Cd levels in head pools remained con-
stant and greater than channel sediments. These results are consistent with
those of John et al. (1972) which demonstrated that plants grow on soils with
increased Cd adsorbing capacity and increased organic matter content had
lower Cd concentrations than those grown on soils with lower Cd sorbing
capacity (see Table 1).
Transplanted J. diffusissimus (Figure 19) never reached Cd levels as
great as those acquired by the naturally colonizing plants. Wolverton (1975)
51
-------
working with water hyacinths stated that mineral uptake rates per unit of dry
matter are greater for plants in a rapid growth phase, and decrease as the
plant ages. J. diffusissimus clumps transplanted into the channels in March
1976 contained at least one season's growth and after a small growth period
in the spring, did not measurably increase in size. Cadmiun concentrations
in the leaves of transplanted J. diffusissimus increased rapidly during the
month after Cd inputs were terminated. There is no apparent explanation for
this observation.
In all cases, Cd concentrations reported here for macrophytes from Cd
treated systems are high relative to other researchers. Lee et al. (1976)
reported root levels of 61 and 18 (Jg/g dry weight for Scirpus and Cyperus
roots growing in medium containing 500 pg/£ Cd and leaf levels of 3-20 and
20-65 Mg/g for nonrooted portions of these same species. Cearley and Coleman
(1976) report Cd levels of 60 \jg Cd/g ash for Najas exposed to 7 |Jg Cd/£.
Our high results may be the result of the extremely soft acid water of our
system.
52
-------
SECTION IX
INVERTEBRATES
INTRODUCTION
The importance of macroinvertebrates as essential components in aquatic
systems is well documented (Hynes, 1957; 1960; 1970; Weber, 1973; Cummins,
1973; Brinkhurst, 1974; Cummins, 1975; and Carins, 1977). Carins (1977)
states that, aquatic macroinvertebrates are important components in food webs
of aquatic systems, being primary and secondary consumers, and serving as
food sources for higher trophic levels. While Cummins (1973) states, the
role of macroinvertebrates in the overall structure and function of stream
and river ecosystems is the conversion of reduced carbon compounds derived
primarily from the surrounding land supplemented by in-stream carbon fixa-
tion, into temporary storage in their own tissue and into carbon dioxide.
Numerous surveys have used the aquatic communities of streams, rivers,
and lakes as indicators of water quality. The works of Gaufin and Tarzwell
(1952 and 1956) and others have demonstrated that the composition and distri-
bution of benthic invertebrate communities are useful tools in evaluating
perturbations in aquatic systems due to various types of pollutants.
Macroinvertebrates are especially well suited for such studies because:
1) their limited mobility does not allow for perturbation avoidance, 2) their
ubiquitous distribution in aquatic habitats, 3) the relative ease by which
they are collected and in many cases identified, 4) their fairly long life
cycles, which means that once a perturbation has affected a community's
composition and/or distribution, it generally requires an extended period of
time before new recruitment can reestablish the original community structure.
Cairns (1977) and Cairns et al. (1972) have suggested macroinvertebrates
(predominantly protozoans) are good indicators of environmental stressors.
Recently, much effort has been spent studying the acute toxicity, up-
take, and accumulation of heavy metals in aquatic organisms (Ravera et al-,
1973; Thorp and Lake, 1974; Karbe et al., 1975; Club et al., 1975*; 1975 ;
Nehring, 1976; and Enk and Mathis, 1977). However, only limited information
is presently available on the use of aquatic invertebrates in assessing the
effects of potentially harmful chronic levels of heavy metals in laboratory
as well as in natural systems.
The objective of this portion of our study was to determine the fates
and effects of chronic Cd exposures (5 and 10 jJg/Jfc) on benthic organisms
under controlled, semi-natural conditions. To accomplish these goals, steady
state levels and uptake and elimination rates of Cd were determined for a
53
-------
number of taxa. Partitioning and Cd dynamics due to growth and molting
cycles of organisms were also studied. Finally community measures were
employed to assess effects of Cd on the structure and function of the macro-
vertebrate community.
METHODS AND MATERIALS
Macroinvertebrates
Cadmium effects on the benthic community as well as individual benthic
populations were assessed by identification and enumeration of individuals
collected using artificial substratum samplers and random bottom samples.
The major portion of the macroinvertebrate survey relied heavily on multiple
plate artificial substratum samplers. Multiple-plate samplers were selected
because: (1) Multiple-plate samplers provide a practical means of collecting
macroinvertebrates in a system, where destructive bottom sampling would
devastate the system. (2) Plate samplers permitted quantitative comparison
between treatments as well as rapid and consistent sample handling. (3) Mul-
tiple-plate samples, although they may exclude some taxa, do collect a suffi-
cient diversity of benthic forms to be useful in relating benthic populations
to watr quality (Fullner, 1971).
Samplers utilized were modified from those described by Hester and Dendy
(1962), Fullner (1971) and EPA (1973). Samplers were constructed of 3.2 mm
double-tempered "masonite" cut into 7.6 cm square plates and 2.5 cm square
spacers. Each sampler consisted of 13 plates and 31 spacers. The "masonite"
plates and spacers were positioned on stainless steel rods, resulting in
three each of single spaced, double spaced, triple spaced, and quadruple
spaced plates. Each sampler had an effective sampling surface of 0.16 square
meters. Four stainless steel support racks, each supporting three samplers
were suspended at each sampling station (Figure 22). This arrangement allow-
ed sampling after both short-term and long-term exposure periods. The short-
term incubations were of six weeks duration (APHA, 1975; Weber, 1973). One
rack with three replicate samplers was removed from each sampling station at
six week intervals and replaced by a new set of samplers. Samplers were
preserved in 75% ethanol for subsequent sorting and enumeration. The long-
term sampling program required three sample racks of three replicate samplers
each at every sampling station. One sampler was removed every 12 weeks over
an eighteen month period and preserved for enumeration. This procedure
allowed us to collect samplers which had been exposed in the channels from
the first day of the macroinvertebrate program (September 1975), until the
end of the project.
Plate sampler removal was accomplished by enclosing each sampler in
chambers constructed of plexiglass and stainless steel screen (mesh size
0.589 mm) prior to removal, to minimize the loss of organisms from samplers
(Figure 23). Detritus and organisms collected were scraped from the samplers
and preserved in 75% ethanol. Sample volumes were reduced by filtering them
through #15 silk bottling cloth (mesh size 0.095 mm) before sorting.
-------
Samples were also collected by removing 0.25 m of bottom sediment en-
closed by two stainless steel screens (mesh size 0.589 mm) pressed into the
substratum. The bottom material removed was diluted with tap water and the
suspended material passed through a U. S. standard //30 sieve. Detritus and
associated organisms retained by the sieve were placed into a white enamel
pan, the living organisms were removed with forceps and preserved with 75%
ethanol. This procedure was repeated until the sand substratum produced no
Figure 22.
Hester-Dendy type invertebrate samples suspended
in channels.
55
-------
additional organisms. The remaining sand substratum, amalgamate and detrital
material was returned to the location from which it was removed.
Macroinvertebrates were sorted microscopically into taxa regardless of
size or instar. All representatives of each taxon were placed in labeled
vials and stored for future identification and enumeration. Samples were
sorted twice to assure complete collection.
»
V- MA*
Figure 23.
Invertebrate sampler in plexiglass and screen
sampling box.
56
-------
The sieving and sorting of macroinvertebrate samples was somewhat diffe-
rent than procedures described in Standard Methods (APHA, 1975) or in
Biological Field and Laboratory Methods (Weber, 1973). In these procedures,
as in most classical works, macroinvertebrates were defined as those inverte-
brates retained by a U. S. standard #30 sieve (0.595 mm mesh opening), while
all other invertebrates were considered microinvertebrates. These arti-
ficially created categories, as observed by Jonasson (1955), Mundie (1971),
and Mason e_t al. (1975), result in the selective retention of certain species
and, in general, the elimination of smaller instars (developmental states)
and/or taxa. Because of the limited literature available on long-term chron-
ic effects of trace elements on benthic communities, it was unclear as to
what role early developmental stages might play in overall community struc-
ture and function. Therefore, we have redefined macroinvertebrates to in-
clude not only those organisms retained on a U. S. sieve #30, but all recover-
able instars of those organisms, regardless of size. Also considered when
making this decision was the potential for increase in sample size. Johasson
(1955) found that by utilizing small mesh sieves he could achieve a 100 -
600% increase in numbers of individuals captured over collections made using
a sieve of 0.6 mm opening. He also observed that the 0.6 mm mesh sieve was
inefficient in collecting small Chironomid larvae. This result is crucial to
our study where the Chironomid larvae may comprise 75 100% of the benthic
population, depending on season and stage of channel colonization.
Macroinvertebrates collected in this study were generally identified to
genus and occasionally to species, the Chironomidae being the only exception.
The assemblages of Chironomidae collected by our techniques contained nume-
rous small instars. Therefore, due to the difficulties in taxonomy of close-
ly related groups and especially among younger instars, identifications were
made from a limited number of samples throughout the study.
Monthly insect samples were collected randomly from natural substratum
samples for all treatments. Because of the seasonal scarcity of particular
taxa and the prohibitive amount of time required to collect equal sample
sizes of each taxa, sample collection guidelines were necessary. The de-
cision was made to collect individuals of all available taxa within the time
interval required to collect 25 Chironomids. As a result, in all taxa except
Chironomidae, the number of individuals comprising a sample may vary both
within and between sampling periods for any treatment.
The organisms necessary to comprise a sample were taken randomly from
available substrata in each treatment, using a small nylon screen net (mesh
size 0.589). Samples were placed in plastic containers prior to sorting.
Organisms were sorted from small aliquots of substrata placed in enamel trays
and removed using stainless steel forceps. Upon removal, organisms were
rinsed with deionized water and placed into clean plastic vials containing
deionized water. Each taxon was placed in a separate vial and stored frozen
until digestion.
Immediately after emergence and prior to flight, adult dragonflies and
their corresponding exuvium were collected by hand, wherever possible, placed
in plastic bags, and stored frozen prior to preparation. This collecting
57
-------
method allowed for the direct comparison of adult and exuvium Cd concentra-
tions in individual dragonflies.
Insect samples except those of the larger dragonflies (adults and
nymphs) were digested as follows: samples were partially thawed, poured into
a fired porcelain crucible and completely thawed using deionized water.
Thawed organisms were then removed from the crucible using stainless steel
forceps and placed on acid washed, dried and tared platinum dishes. Platinum
dishes were then placed in small petri dishes and dried at 50 C in a drying
oven to obtain a constant dry weight. Dried samples were weighed on a Cahn
Model 2500 Electrobalance. All platinum dishes and samples were then placed
into acid washed 1 ml glass volumetric flasks for digestion. The taxa and
number of individuals per sample were recorded. Except for pooled samples of
Chironomids and Ceratopogonids, all other samples contained individual orga-
nisms. Insect sample digestion was accomplished using 60 pi of redistilled
cone. HNO and 20 pi of 30% H.O . Volumetric flasks containing samples were
heated in a water bath at 60 to 70 C (this procedure did not adversely
affect the platinum dishes). Digestion was determined to be complete by the
formation of a clear pale yellow solution in each flask. After complete
digestion, samples were allowed to cool and platinum dishes were removed with
an acid washed glass hook and rinsed with several drops of deionized water
over the flask. Sample volumes were adjusted to 1 ml with deionized water
and samples were ready for analysis.
Samples of larger insects, dragonfly adults, nymphs and exuvia were
freeze-dried to obtain constant dry weight and weighed. Samples were diges-
ted in fired porcelain crucibles using redistilled cone. HNO- (0.6 ml for
adults and nymphs; 0.4 ml for exuvia) and 30% H-0_ (0.2 ml for adults and
nymphs and none for exuvia) at 60 - 70 C on a hotplate. After complete
digestion, samples were rinsed into 5 ml glass volumetric flasks and brought
to volume using deionized water.
Monthly Corbicula fluminea samples were collected from each treatment,
by removing four transplanted organisms from the tail region of each channel.
The entire soft body of these organisms was dissected out of the shell using
a stainless steel surgical sealput, placed in a plastic bag and frozen prior
to digestion. C. fluminea samples were analyzed using both flame and flame-
less atomization techniques. Flame atomization was required for clams ex-
posed to 5 and 10 pg Cd/1 and flameless methods were used for control and
background organisms. Matrix interferences were encountered with flameless
atomization techniques and tissue Cd concentrations were determined using
standard addition techniques (see Appendix I).
C. fluminea samples were freeze-dried to obtain constant dry weight.
Samples were digested in fired porcelain crucibles using 1 and 2 ml of re-
distilled cone. HNO and 0.5 or 1 ml of 30% H-0 depending on tissue size
(large or small) on a hotplate at 50°C for 10 to 15 hr. Samples were diluted
to 25 ml with deionized water after cooling.
58
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Macroinvertebrates
The usual benthic sampling methods are biased toward collecting larger
invertebrate forms in aquatic systems. Even by using a smaller screen for
sorting purposes, a number of organisms relatively important in aquatic food
chains cannot be collected. We attempted to consider, at least in a qualita-
tive manner, Cd effects on these smaller invertebrate forms by utilizing an
aritificial substratum sampling method developed by Cairns (Cairns e_t al. ,
1969, Cairns and Ruthren, 1970). This method uses polyurethane sponges
suspended in the water as a habitat for colonization which can be sampled
with replication and a minimum disruption of the study area. This method has
been successfully used for studying the colonization and succession of fresh-
water protozoans (Youngue and Cairns, 1971) and also the response of proto-
zoan communities to chlorine stress (Cairns and Plafkin, 1975).
The sponges provided an ideal sampling substratum for a wide variety of
organisms in addition to protozoa. Indeed, the sponges could have been used
to sample the algae component of the eocsystem. Several algal species were
found only in samples from the sponges.
Rinsed polyurethane sponge cubes, 5 cm on a side (Figure 24) were sus-
pended at two locations in the channels and repeatedly squeezed to exclude
all air. Two new pairs of sponges were placed at each station monthly and
sampled two weeks later. Only one pair of sponges was examined, the other
provided a back-up capability in case of sample loss.
Sponges were squeezed dry by hand into a 500 ml beaker and immediately
mixed. Two-2 ml samples were taken and placed in counting chambers. The
total remaining volume of water was measured and a 100 ml aliquot preserved
with 5 ml of formalin. Samples were allowed to stand for approximately one
hour in the settling chamber. The total chamber volume was then examined at
a magnification of 56 X with a Wild M 40 inverted microscope and the total
number of larger forms identified and enumerated. This magnification allowed
the enumeration of the larger protozoans, rotifers, nematods, anelids, flat-
worms, insect larvae, ostracods, copepods, cladocerans and occasionally,
taridgrads, mites, and gastrotrichs. After the larger forms were enumerated,
the samples were examined at 560 X. Ten random fields were completely coun-
ted. Although some minute flagellated forms did not "settle" in the chamber,
these constituted a relatively small portion of the total microinvertebrate
population. No attempt was made to derive a species identification for all
of the multitude of forms observed because of the time constraints imposed by
the use of living materials, and the lack of taxonomic expertise; however,
consistently observed forms were identified.
Youngue and Cairns (1971) demonstrated that water contained in sponge
samplers may differ from the surrounding medium at least with respect to pH.
To determine if organisms inhabiting sponges suspended in the treated systems
were actually exposed to Cd, a series of sponges was submerged in Cd spiked
water for a period of two weeks. The Cd concentration of water squeezed from
the sponge was not significantly different from that in the surrounding
medium.
59
-------
Counts made in the sponge material were pooled into larger taxonomic
groups !><•( .ujse of the sm.i 1 1 numbers •>( r.ich species. Each time period was
analyzed separately by ANOVA techniques for treatment effects. There was no
significant (P 0.05) difference between upstream and downstream stations.
So data from these stations were pooled by treatment.
Figure 24. Polyurethane sponge microinvertebrate samplers,
•
-------
RESULTS AND DISCUSSION
Cd Accumulation
Information on Cd accumulation and elimination by aquatic macroinverte-
brates (ordinarily insects) is incomplete in this study due to the complexi-
ties and confounding properties of the natural environment. Factors which
were determined as being necessary to appropriately calculate uptake and
elimination rates along with concentration factors included: 1) the poten-
tial for multiple routes of exposure, uptake and elimination; 2) seasonal
effects on population level (emergence and recruitment), development cycle
(alterations in metabolism, size, shape and molting rate), feeding habits
such as selectivity and habitat selection; and 3) alterations in environ-
mental availability and exposure levels of the metal in question. All of
these reasons combined to make it quite apparent that in order to conduct a
comprehensive study of the fate of Cd in macroinvertebrate populations under
natural conditions one must conduct complete life history studies for each
group of organisms comprising the community. This requirement rapidly makes
the amount of time and effort required to conduct such a study on «» large
scale prohibitive. Therefore, what is presented is a genral overview of
several taxa in terms of Cd levels accumulated and eliminated, with more
specific data for several taxa (Diptera: Chironomidae and Odonata: Anisop-
tera; Libellulidae).
The taxonomic groups for which the most data are available are:
Ephemeroptera; Odonata, Anisoptera (Pantala hymenea) and Zyoptera (Ischnura
sp.); Coleoptera (Dytiscidae); and Diptera, Chironomidae and Ceratopoganidae
(Bezzia or Probezzia). Cadmium accumulation and elimination results for
these taxonomic groups are presented in tabular form (Tables 11 and 12), as
mean values calculated using all samples analyzed in each taxa for the
periods during and after Cd inputs. This method of presentation for uptake
and elimination data is probably inappropriate due to its disregard for po-
tentially important seasonal and developmental shifts in susceptibility,
uptake, and excretion of and potential for increasing Cd accumulation over
time. Therefore, Tables 11 and 12 are presented only as a generalized over-
view. Many of the potentially confounding problems affecting our findings
will be discussed in relation to specific observations made on midge or
dragonfly data.
Generalized Cd accumulation data (Table 11) suggests that aquatic insect
nymphs and adults do accumulate Cd and that accumulation is related to ex-
posure concentration in many cases. Fowler and Benazoun (1974) have pre-
viously reported a direct proportionality between uptake rate and environ-
mental concentration for Cd in the shrimp Lysmatai seticaudata and the mussel
Mytilus edulis, however in our study, this relationship for aquatic insects
is less exact. Those taxonomic groups obtaining the greatest Cd concentra-
tions were the midges, mayflies and damselflies. Our findings also suggest
there is no biomagnification of Cd with increasing trophic level. Detriti-
vores and/or herbivores always maintained higher Cd levels than did carni-
vores regardless of treatment. Schwimer (1973) has previously reported the
biodiminution of Cd in tidal invertebrates, from herbivores to predators.
However, these reports from aquatic communities appear to be just the reverse
61
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TABLE 11. MEAN CD CONCENTRATIONS IN INSECTS, DURING THE PERIOD
OF CD INPUTS
Range Number
Treatment
Taxa (jg/1 Cd
Ephemeroptera
*(detritivores
herbivores)
Odonata Anisoptera
^(Carnivores)
Odonata Zygoptera
^(Carnivores)
Coleoptera
^(Carnivores)
Chironomidae
*(detritivores
herbivores)
Ceratopogonidae
*(preditors &
scavengers)
0
5
10
0
5
10
0
5
10
0
5
10
0
5
10
0
5
10
Mg/8 Cd
dry wt
1.6
40.7
176.0
2.6
18. A
34.3
3.2
32.4
46.4
0.8
4.1
13.0
5.6
55.4
91.6
2.0
23.4
33.1
low
0.0
0.0
59.0
0.0
9.3
1.9
0.0
5.7
29.5
0.0
1.2
6.1
1.2
17.0
22.4
0.0
7.7
11.6
of
high Samples
5.8
96.8
324.8
5.7
38.3
188.4
11.0
61.1
93.4
2.6
9.3
25.6
64.7
190.2
345.5
5.6
56.9
56.1
16
4
8
24
7
10
33
13
7
9
7
4
36
40
44
9
7
8
Number
of
Organisms
18
4
11
24
8
10
34
15
9
7
7
4
391
421
491
52
41
49
^indicate a genral classification of trophic categories for dominant orga-
nisms occurring in taxonomic groups collected in our experimental system.
62
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TABLE 12. MEAN CD CONCENTRATIONS IN INSECTS DURING THE PERIOD
AFTER CD INPUTS WERE TERMINATED
Taxa
Ephemeroptera
*(detritivores &
hervivores)
Odonata Anisoptera
•'-'(Carnivores)
Odonata Zygoptera
^(Carnivores)
Coleoptera
"-(Carnivores)
Chironomidae
(detritivores &
herbivores)
Ceratopogonidae
(preditors &
scavengers
Treatment
Mg/1 Cd
0
5
10
0
5
10
0
5
10
0
5
10
0
5
10
0
5
10
Treatment
Mg/g Cd
dry wt
11.1
4.5
26.0
32.4
0.6
7.4
24.6
4.9
31.5
52.6
5.1
28.1
33.8
Range
low high
_
-
_
- -
-
0.0 13.2
3.0 44 . 2
19.3 67.8
0.2 1.0
0.1 24.5
0.7 45 . 2
1.6 12.1
6.4 107.2
10.1 158.7
1.6 9.6
21.1 33.7
10.1 64.1
Number
of
Samples
1
_
-
17
17
8
2
5
7
7
17
41
3
6
7
Number
of
Organisms
_
1
_
—
17
17
8
2
5
7
71
300
404
6
22
18
^indicate a general classification of trophic categories for dominant orga-
nisms occurring in taxonomic groups collected in our experimental system.
63
-------
for terrestrial invertebrate communities. Skinner e_t al. (1976) have report-
ed the biomagnification of Cd in the terrestrial food webs of a coal ash
basin.
Our results also indicate a wide range of variability in the "ability1'
to accumulate Cd both among and between taxonomic groups at any particular
time. Similar findings have been reported by Thorp and Lake (1973, 1974)
Nehring (1976) Nammingea and Wilhm (1977) Bryan (1976) and Renfro et. al..
(1974). Bryan (1976) states that although rates of uptake may be related to
the external concentration, there is no certainty that concentrations in the
organism will reflect those of the environment. Many researchers have attri-
buted this variability to some combination of: 1) fluctuations in uptake
and/or excretion rates; 2) external surface contamination, or 3) the presence
of gut contents in organisms sampled. Elwood, Hildebrand and Beauchamps
(1976) have reported that gut contents in Tipula sp. comprise approximately
22.1% or an individual's dry weight and over 50% of the total body burden in
70% of the 30 elements they analyzed. They concluded that gut content was an
extremely important source of potential error and should be of primary con-
siderations when determining body burdens and conducting studies on trophic
level transfers of elements in aquatic systems. The potential for surface
contamination of biological samples and its effect on the variability of de-
terminations has been alluded to by numerous authors as resulting from in-
adequate or prolonged rinsing of samples prior to digestion.
Generalized Cd elimination data (Table 12) indicate that Cd is elimi-
nated from the insect community over time for those populations or taxa
analyzed. However, due to the variability of the data resulting from: 1)
sporadic collection, 2) variable sample size, 3) effects of life cycle de-
velopment, and 4) new organism recruitment; little can be said about the
rates at which elimination may occur. Data suggest that of those taxa
analyzed, the Coleopterans and Ceratopoganids may have reacted somewhat
differently to the chronic Cd exposure than did other taxa.
Coleopteran data suggest that these organisms continued to accumulate Cd
even after Cd inputs were terminated. This result is probably an artifact
created by the relatively long life cycle of these organisms (in comparison
with other taxa sampled) and the method by which the mean values were calcu-
lated. As previously mentioned this method of pooling samples over long time
intervals does not account for Cd accumulation over time. Thus organisms
with extensive Cd exposures were included in the Cd elimination data, while
many earlier collected organisms with limited Cd exposures are included in
the Cd accumulation data, giving the appearance of continued Cd accumulation
after Cd inputs had been terminated.
Ceratopogonid Cd body burdens did not decrease after Cd inputs were
stopped, even though other community components displayed a fairly rapid
elimination of Cd after inputs were terminated.
Pooled monthly samples of chironomid Cd concentrations suggest a ten-
dency for significant differences in Cd accumulation between treatment and
control situations. Results of Schefcf's S procedure (Kirk, 1968) for the
64
-------
separation of means (Table 13) indicate that even though there are signifi-
cant differences in chironomid Cd body burdens between treatments and con-
trols, there is considerable similarity in Cd levels between treatments, thus
indicating that, in the case of chironomids, the fates of 5 (Jg/1 and 10 pg/1
Cd exposure are not significantly different considering the natural vari-
ability encountered.
Cadmium accumulation by chiromonoids appears extremely dependent on the
time of year during which samples are collected. Mean chironomid Cd concen-
TABLE 13. SCHEFF'S S-PROCEDURE VALUES FOR INSECT CADMIUM CONCENTRATIONS
MEANS AT EACH SAMPLING PERIOD
Treatment
Period
(month)
1976
June
July
August
September
October
November
December
1977
January
February
March
April
May
June
lOppb
183. 17a
218. 03a
97.97a
40.33a
38.84a
28.50a
40.94a
37.42a
44.28a
52.07a
113. 66a
87.10a
32.10a
5ppb
142. 92a
119.13b
58.57a
23.08b
I6.55b
26.64a
40.53a
32.85a
33.66ab
35 . 40a
58.58a
66.63a
I6.66ab
2.98b
2.26c
31.20a
1.30c
2.23c
2.01b
3.84b
2.08b
2.56b
2.55a
3.00a
7.47b
Oppb
(questionable
sample)
Means with the same superscript within a sampling period are not signifi-
cantly different (a = 0.05).
65
-------
trations calculated on a monthly basis (Figure 25) indicate that during the
period of April through July, Cd burdens were higher than at any other time
of year. This finding occurred on an annual basis both during and after Cd
inputs were terminated, suggesting a seasonal shift in the uptake or avail-
ability of Cd in the chironomids. Thorp and Lake (1974) reported indications
of potential seasonal differences in Cd toxicity to Paratya tasmaniensis
(Decapoda: Atyidae). Clubb, Gaufin and Lord (1975) have reported findings
which indicate that organisms collected and treated after November were less
sensitive to Cd toxicity than organisms collected and treated prior to Novem-
ber, suggesting that early developmental stages in insects are more sensitive
to Cd.
In an attempt to discern if the higher Cd body burdens observed during
the April through July period resulted from: 1) Cd accumulation over time,
250
Chironomid Pooled Samples
200
•o
o
3 150
z
o
< 125
CL
Z
UJ
o 100
5 75
50
25
15
5
0
J 1 J
A | S
0 | N
D | J
— • —
F
— •
M
A
M
— •
J
1976
1977
SAMPLING DATE
Figure 25. Mean Cd concentrations in pooled samples of
chironomids, expressed on a dry weight basis,
66
-------
2) shifts in surface to volume ratios, or 3) increased susceptibility to Cd
of early instars; chironomid population, biomass and Cd accumulation data
were used conjunctively.
The mean numbers of chironomids (Figure 26) indicates that during the
period April through July, when Cd body burdens were highest, the greatest
number of collectable organisms was also present. When combining the chiro-
nomid mean numbers data with mean individual dry weight data (Figure 27)
one finds that not only are the greatest numbers of organisms present during
April through July but that the individuals present are the smallest found
during the entire year. These results indicate that during the April through
July period chironomid populations are predominately comprised of early
instar individuals. Thus, the potential for increased Cd accumulation over
time due to extended periods of Cd exposure is eliminated.
Linear regressions using Cd concentration and mean individual dry
weights were performed in an attempt to discern if the high Cd concentrations
observed during the April through July period resulted from shifts in surface
to volume ratios of different size individuals or from increased susceptibil-
ity to Cd of early instars. The hypothesis being that Cd is a non-regulated
metal absorbed from solution by passive diffusion across a gradient created
by adsorption at the surface (Bryan, 1976), and one would expect shifts in
surface to volume ratios to influence the Cd level found in the organism.
700
£ 600
Q.
E
o
\ 500
k.
o>
JD
E
c 400
300
UJ
O
oo
200
100
Chironomidae
\\
\\\
• • Control
a -a 5^g Cd/L
o o 10 ftg Cd/L
/I \
Nov | Jon
FebJMar
Apr
Jun |Aug|Sep |Oct|Dec
Jan
Mar
Apr
Jun
1976
1977
SAMPLING DATE
Figure 26. Density of chironomids in plate samplers.
67
-------
Therefore, smaller individuals (early instars) having a greater surface to
volume ratio would be expected to exhibit higher Cd levels than larger indi-
viduals (later instars) possessing smaller surface to volume ratios. Results
of linear regressions between Cd concentration and mean individual dry weight
(a measure of size) were not significantly different from zero, suggesting
size and probably the external complexation properties of cutical were not of
primary importance in controlling Cd accumulation in chironomids. These
results tend to support the finding of other researchers that Cd suscepti-
bility is apparently increased in early developmental stages of aquatic
insect. Which in turn opens the door to a myriad of potential hypotheses
which need to be investigated. Some of the potential hypotheses worthy of
consideration in attempting to explain our findings are:
Individual Chironomid
10
.08
.06
04
02
.20 r
.00 LT
Control
a D 5pq Cd/L
a a 10^9 Cd/L
E
h-
X
o
LJ
Q
O
2
O
IT
X
O
z
LJ
5
.18
.16
.14
.12
.10
.08
.06
.04
.02
1976
1977
SAMPLING DATE
Figure 27. Mean chironomid weights.
68
-------
l) Cummins (1973) statement that earlier instars of herbivore-detritivore
type organisms rely primarily on detrital feeding. This means that the
potential of selective feeding habits in various life stages of the same
organism could play an extremely important role in controlling the concen-
trations of metals detected. 2) Bryan's (1976) statement that the perme-
ability of various species is of considerable importance in determining their
tolerance to metals. This statement is already supported in the literature
by the findings of Renfro et al. (1974) who found that some species of cru-
staceans were more susceptible to toxicants shortly after molting than later
in any particular life stage. This finding suggests that not only should a
researcher consider the exposure time an organism has had to a particular
toxicant but the number of molts or instars completed and the time elapsed
since the most recent molt prior to collection, when trying to determine the
fate of metals in macroinvertebrate populations. Also of importance here is
that during early stages of development, molting generally occurs more rapid-
ly and therefore may increase the interval of susceptibility there by increas-
ing the Cd uptake during this segment of development. 3) Oliver's (1971)
statement that the period of rapid growth of larval chironomids of univolline
species occurs during the warm period of the year. If one then assumes that
during periods of rapid growth the period of maximum enzyme activity occurs
and combines this with Brown's (1976) findings that heavy metals may activate
enzymes or enhance their activity at low concentrations, we have another high
probability mechanism which metals concentration could be controlled on a sea-
sonal or annual basis. 4) Another possible cause for seasonal fluctuation in
metals concentrations of individual invertebrate taxa in field studies re-
sults from the alterations in a population's species composition. Morrison
and Steele (1977) in their work with mollusks found that species within a
given taxonomic group but with different environmental habitats exhibit
widely varying Cd accumulation rates. This means in our case that the
changes in Cd concentrations may very well be the result of a species shift
in the population. However, due to the size of organisms collected during
the April through July period it was time prohibitive and impossible to
conduct species identifications. Our chironomid Cd data in conjunction with
the finding of Morrison and Steele (1977) previously mentioned indicates that
if one is interested in investigating the fate of a metal in the natural
situations it becomes very important to know not only who comprises the
community but also their habitats and habits.
Cadmium concentration measurements for the dragonfly Pantala hymenaea
indicate that this organism accumulates Cd during portions of the life cycle.
Cadmium concentrations determined for each life cycle segment (adult,
exuvium, nymph, and estimated nymph) for the two treatments utilized are
presented in Table 14. Cadmium concentrations in adult dragonflies are
proportional to treatment concentrations to which they were exposed, however
orders of magnitude greater. Important to note here is that Cd levels repor-
ted for P. hymenaea represent incorporated Cd, because of the manner in which
these organisms were collected they had no opportunity for adulthood Cd ex-
posure. Therefore, the values shown in Table (14) for adults could be con-
sidered as that segment of the Cd mass balance leaving the system due to
individual P. hymenaea emergence. However, the assumption should not be made
that similar values could be extrapolated based upon individual weight for
other taxons or species in the aquatic system.
69
-------
TABLE 14. MEAN CD CONCENTRATIONS FOR P. HYMENAEA LIFE CYCLE
SEGMENTS BY TREATMENT EXPRESSED ON A DRY WEIGHT
BASIS (X ± 2 SE).
5ppb lOppb
Segment
Adults
Exuvia
Nymphs
Estimated
Nymphs
N
4
5
6
4
1
23
17
5
.6
.8
.3
.2
± 0.4
± 10.3
± 8.9
± 2.2
N
15
28
8
15
3
33
19
8
.2
.9
.1
.7
± 0
± 5
± 6
± 1
.7
.3
.3
.1
Cadmium data based on analysis of final instar exuvia indicate that on a
per gram dry weight basis the exuvium of P. hymenaea has a higher Cd content
than any other segment of the life cycle analyzed (Table 14). However, Cd
levels accumulated do not appear proportional to treatment. Results indicate
a trend towards increasing Cd concentrations over time along with a decreas-
ing trend in exuvium dry weight over time, suggesting that surface sorbtions
may be the means of Cd accumulation by the exuvia as Bryan (1976) has pre-
viously suggested. However, linear regressions performed between Cd con-
centration and individual exuvium dry weight were not significantly different
from zero, suggesting that some other mechanism or mechanisms are involved in
affecting the accumulation of Cd by the exuvium. Many of these potential
mechanisms have been alluded to in earlier segments of the macroinvertebrate
discussion.
Cadmium data for P. hymenaea nymphs is based on a relatively small
sample size of highly variable Cd determinations. Results reported in Table
(14) indicate that the nymphs do accumulate Cd and that this accumulation is
not proportional to treatment. However, it should be noted that all nymph Cd
determinations were accomplished using whole organisms from which the gut
contents were not removed. Therefore, variable amounts of food containing Cd
probably has greatly influenced the variability of these samples.
Cadmium data for estimated nymphs derived by combining the pg Cd/ml
sample for adults and their respective exuvium cast off at emergence then
dividing by the combined dry weight of those same samples, suggests that Cd
accumulation is proportional to treatment and is considerably different than
levels determined for actual nymphs. The difference observed between actual
and estimated nymphs may very well represent the percentage of total Cd body
burden attributable to gut contents in P. hymenaea. However, further re-
search is required before any specific conclusions can be drawn.
While calculating the total Cd body burdens for estimated nymph values,
the percentage contributed by both adults and exuvia was also determined
(Table 15). Results indicate that there is no difference in the percentage
70
-------
TABLE 15. MEAN % CD IN EACH LIFE CYCLE SEGMENT OF ESTIMATED NYMPHS BY
TREATMENT (X ± 2 SE)
IQppb
Combined
Segment
Adults
Exuvia
N N N
4 32.3 ± 17.2 15 32.0 ± 6.1 19 32.1 ± 5.8
4 67.7 ± 17.2 15 68.0 ± 6.1 19 67.9 ± 5.8
of the total Cd body burden attributable to adult or exuvium, with increasing
treatment level. This finding suggests that the mechanisms of accumulation
and elimination are constant and not altered by treatment level (at least at
chronic levels). These data also bring to our attention the ability of the
exuvium (exoskeleton) to accumulate Cd and the potentially important roles
this structure may play in the toxicity and/or cycling of Cd.
The ability of the exuvium of P. hyroenaea to accumulate approximately
68% of the total Cd body burden is similar to literature values for other
organisms. Renfro et al. (1974) reported that 45% of the total Zn body
burden of shrimp was located in the exoskeleton while approximately 61% was
found in the exoskeleton of crabs. Renfro et al. (1974) concluded that the
occurrence and rate of molting in invertebrates could account for a consider-
able portion of the variability of their and other researchers studies and
that exoskeletons are of potential importance in the cycling of metals in the
environment, either through their actions as a metals sink or by adding in
the recycling or availability processes. Another question to be proposed and
investigated in relation to the Cd accumulating abilities of the exuvia P.
hymenaea is: does the exuvium act as a mechanism protecting a species from
Cd toxicity due to its ability to accumulate or absorb the metal? There is a
significant amount of literature on mechanisms in other invertebrate forms
which have the ability to complex metals and which have been hypothesized as
mechanisms for transporting and potential detoxifying metals. Bryan (1976)
lists several: 1) the ability of blood proteins to bind Zn in crayfish, 2)
the apparent storage of Cu in fine granules within the epidermal cells of
marine polychaeles, and 2) the presence of wandering leucocytes in mollusks
and their known importance in transporting and detoxifying metals. The
possibility does exist that the exoskeleton may be functioning in a similar
manner for P. hymenaea.
Population and Community Effects
Macroinvertebrates collected from the experimental channels were tole-
rant forms, typical of pond or sluggish waters (stream margin and littoral
zone) in the southeastern United States. The various benthic sampling
methods utilized during the 23 month study collected a total of 53 different
taxa of which only 14 were collected routinely. Macroinvertebrates consisted
71
-------
primarily of numerous chironomid species, mayflies, Callibaetis sp. and
Caenis sp., damselfly, Ischnura sp. and two genera of Ceratopogonidae,
Dasyhelea sp. and Bezzia sp. or Probezzia sp. Also present, but less abun-
dant, were several species of Anisoptera, Hemiptera, Coleoptera,
Trichcoptera, Lepidoptera and Annelidae. Appendix II lists all macroinverte-
brates collected from the treatments during the study period and the method
of collection. Macroinvertebrate colonization continued throughout the study
period with the continual recruitment of new species. Sampling emphasis was
placed on insect fauna, resulting in the possible omission of some non-insect
invertebrates.
Macroinvertebrates which colonized the channels were primarily insects
adapted for invading newly created bodies of water by flight. Benthic in-
vertebrate community development was allowed to proceed naturally. There-
fore, only a few organisms were collected from the channels prior to the
establishment of the periphyton community. Unlike most woodland or pastoral
streams where the dominant energy source results from allochtanous inputs
creating the development of heterotrophic systems, our artificial channel
system is highly autotrophic relying on periphyton and filimentous algal
forms as the energy basis for the establishment of higher trophic levels.
Because of this autrophic status and the physical structure (current veloc-
ity, water temp., sand substrate, etc.) one would not expect to find a number
of macroinvertebrate forms whose physiological or morphological development
and/or behavior has specialized them for the roll of processing larger or-
ganic material (leaves, macrophytes, etc.) converting it into partial sizes
and textures required by other components of the invertebrate community.
Such organic processors such as Trichoptera, Plecoptera and some of the
Ephemeroptera, Coleoptera and Diptera were rarely collected during this
study.
Those organisms first to establish permanent populations in the channels
were the midge larvae, which are known to dominate sandy substrata, folowed
shortly by a limited number of a variety of other organisms. The most impor-
tant of these initially rare taxa were: Pantala hymenaea, with a short life
cycle and common to temporary ponds (Corbet, 1962); Hesperocorixa sp., one of
the corixids, which as a group are acknowledged to be partially responsible
for the primary conversion of plant materials into animal food (Usinger,
1971) and Callibaetis sp., a mayfly found in small temporary woodland ponds
(Burks, 1953). These initial colonizers were followed sporadically by other
organisms throughout the study, of which the mayfly Caenis sp. , damselfly
Ischnura sp., dragonfly Erythrodiplax miniscula and the biting midges
Dasyhelea sp. and Bezzia or Probezzia sp. were the most important.
Colonization patterns observed in the channels were similar to those
observed in natural aquatic systems (Egglishaw, 1964; Hynes, 1970). Popula-
tions increased rapidly in late February or early March and peaked in April,
due to newly hatched, early instars. This large population, then, gradually
diminished through the summer due to predation, natural mortality and emer-
gence. In October there was a slight^increase in population levels resulting
from ovaposition. However, no significant alterations in this colonization
pattern could be attributed to Cd.
72
-------
The density of the entire macroinvertebrate community (Figure 28) al-
though a crude method of representing benthic community responses (Pennak and
Van Gerpen, 1947), indicated that with the exception of two points, September
1976 and April 1977, there were no significant differences among Cd treat-
ments. The September 1976 difference resulted from a tremendous increase in
the Pristina aequiseta populations in control channels, probably due to an
increase in dead and decomposing organic material created by the breaking-up
of filimentous algal mats covering the channels. The April 1977 difference
resulted from extremely rapid recruitment of new midge larvae in the channels
previously receiving 5 and 10 |Jg/l Cd as opposed to the slower recruitment
into control channels. This phenomenon may be due to a number of phenomena
including: differences in density or structure of other invertebrate popu-
lations, algal or macrophyte community colonization, or visual preference in
the selection of ovapositation sights as has been reported for some aquatic
insects.
Significant differences in the number of taxa colonizing multiplate
samplers attributable to Cd treatments occurred in only 3 of 14 sampling
periods (Figure 29). In all three cases, control channels had significantly
more taxa colonizing them than those receiving either 5 or 10 (Jg/1 Cd. Al-
though the number of these significant differences were few, they occurred in
1400 r
MACROINVERTEBRATES
Control
* 5/tg Cd/L
IO>igCd/L
Nov | Jon|Feb |Mor [Apr
Aug|Sep |Oct|Dec|Jon |Mor |Apr |Jun
1976
1977
SAMPLING DATE
Figure 28. Mean number of macroinvertebrates per sampler.
73
-------
SIK o-ssion during Cd inputs, and -it a particularly important time in the
foloni/ation pattern, late fall and winter. This period of the year is when
the henthic fauna IB >//-m-r a 1 J y most stable in both density and diversity.
Also of importance is that those individuals over-wintering in the system are
responsibJe for initiating the following springs recruitment and coloniza-
tion. Thus any type of effect which acts in an additive fashion with natural
seasonal affect.s to affect the size and composition of the over-wintering
bent hie community could seriously affect r.olonixation and successional de-
velopment in yea IB to < orne. The number of taxa in control channels (Figure
29) did not fluctuate as sharply as those in channels receiving r> or 10 pg/1
Cd inputs, indicating control channels were more stable and did not respond
•is rapidly to changes in environmental conditions as did channels receiving
Cd. This phenomena is probably the result of the control channels maintain-
ing higher algal productivities and greater macrophyte colonization during
the preceding part of the year, thus building up a greater organic and nu-
trient bar,/-, as well as diversity of habitat. This observation is supported
by our a 1^1 and macrophyte data and the works of Jones (1940; 1941; and
I'j'ih) who hypothesizes that insert larvae of a stream are largely affected by
the indirect effects of heavy metals pollution and that the principal in-
direct effect of such pollution is the formation of unstable physical condi-
tions due primarily to the elimination of algal and aquatic macropnytic
growth.
12
10
L_
~Ct.
E 8
o
CO
Is
4
2
0
I I I
a—
o—
Control
5fiq Cd/L
\0pq Cd/L
\ 11
- Q
Nov Jon Feb
Mar
Apr
Jun
AuglSep Oct I Dec JanJMor Apr Jun
1976
1977
SAMPLING DATE
figure 29. Mean number of macroinvertebratc- taxa per sampler with two
standard error confidence intervals indicated.
74
-------
Although there were no significant differences in organism densities due
to Cd treatments, the abundance of dominant taxa and shifts in relative com-
munity compositions revealed interesting trends. Between 88 and 100% of the
macroinvertebrate communities colonizing our system was accounted for by four
taxa; Chironomidae, Annelida (Pristina aequiseta), Copepoda (Euryclops
agilis) and Ephemeroptera (Figures 30, 31 and 32). The greatest fluctuations
in macroinvertebrate community composition resulted from shifts in the rela-
tive abundance of dominant taxa, while the relative composition of rarer taxa
remained constant. This pattern was reversed in channels receiving Cd, where
rarer taxa comprised a larger segment of the community. The rarer taxa in
these situations fluctuated considerably, while the dominant taxa maintained
more stable population levels during Cd inputs. This trend appears to have
gome relationship to environmental Cd concentration, with dominant taxa popu-
lations becoming more stable and rarer taxa populations exhibiting greater
instability as Cd concentration increases. Stability in the dominant taxa
group resulted primarily from the fact that as Cd treatment increased,
Chironomid abundance also increased, thus Chironomids comprised more and more
of the total invertebrate community. Our macroinvertebrate data indicated
that the presence of chronic Cd pollution at the 5 and 10 pg/1 level is not
shown by indicator species but by the dominance of tolerant species. Our
findings are supported by Hynes (1960) who states that no special fauna are
indicative of heavy metals pollution, although the surviving species may be
more abundant.
Population densities and percent community composition of Chironomidae,
Ephemeroptera, Ceratopogonidae, Annelida, and Copepoda were affected either
directly or indirectly by Cd. Chironomid population densities were always
less in control channels. However, there were only two points at which this
trend had mean values which were statistically different (Figure 26). This
observation was primarily due to habitat availability; resulting from more
periphyton covered sandy substrata and less decomposing organic material de-
position in channels receiving Cd inputs. Mean weight data for individual
chironomids (Figure 27) indicated that organisms taken from control channels
were generally lighter than were similar individuals taken from channels re-
ceiving Cd inputs. This trend continued during Cd input and is most probably
due to habitats and related environmental conditions. Oliver (1971) states
that larvae of many Chironomid species have the ability to grow and develop
as conditions permit. Our research, however, leaves us with no explanation
as to why chironomids collected from channels receiving 5 and 10 pg/1 Cd
should have achieved a greater individual body weight.
Abundance of individual mayfly genera indicated a slight shift in occur-
rence of the genus Caenis in controls vs. channels receiving Cd (Figure 33).
Caenis became more prevalent in the control channels as opposed to treatment
channels during Cd inputs. It should be noted that even though the trend did
occur during all sampling periods, at no time was there any significant (P <
0.05) differences observed. It is believed that this trend is again the
result of increased algal and macrophytic growth in control channels,
Ceratopogonidae (biting midges) became increasingly more abundant in
channels receiving 5 and 10 Mg/1 Cd (Figure 34). Increased prevalence of
75
-------
lOOr
90
80
70
Z
O
t= 60
O
0.
8 50
i-
z
Ud
UJ
Q,
30
20
10
5
Invertebrate Community Composition -O^g Cd/ L
I I Chironomidae
F7"! Annelida
I Ephemeroptera
I Others
Nov Jon Feb|Mof|Apr JunAug SepJQct Dec JonMorApr Jun
1976
1977
100 r
Invertebrate Community Composition - 0/^g Cd/L
O
H
V)
O
O
LJ
O
a:
UJ
a
95
90
87
• Zygoptera
HID Anisoptera
r~\ Diptera other than Chironomidae
Wh Hemiptera
tn Coleoptera
• Trichoptera
Nov| Jon|Feb|Mor|Apr [ Jun [ Aug|Sep|Oct | Dec[Jan [Mar[Apr pun
1976 1977
SAMPLING DATE
SAMPLING DATE
Figure 30. Percent community composition of macroinvertebrate community in control channels.
-------
lOOr
Invertebrate Community Composition- 5^9 Cd/L
O
8 95
a.
S
o
o
LU
O
cc
LU
a
90
87
f~l Zygoptera
Hj$ Anisoptera
Q Diptera other
than Chironomidoe
WL Hemiptera
D Coleoptera
• Jrichoptera
Nov
Sep OctJDecjJon Mor|AprlJun]
1976
1977
SAMPLING DATE
iOOr
90
Invertebrate Community Composition-5ug Cd/L
2
O
o
cr
UJ
a.
80
8 70
QL
5
O
o
60
50
40
30
0
Chironomida*
Annelida
Copepoda
Ephemeroptera
Others
Feb Mar Apr I Jun lAug ISeplOc t Dec Jan Mar Apr
1976
SAMPLING DATE
1977
Figure 31. Percent community composition of macroinvertebrate community in channels
receiving 5 yg Cd/1.
-------
100
90
so
O
QL
5
O
<-> 70
Invertebrate Community Composition- IC>g Cd/L
™
00
O
CK
60
50
40
O
I i Chironomidae
Q Annelida
IH Copepodo
Hi Ephemeroptera
Nov Jan Feb Mar Apr Jun Aug Sep Oct Dec Jon Mar Apr Jun
1976
SAMPLING DATE
1977
lOOr
8 95
0.
S
O
O
f-
z
LJ
O
a:
LU
a.
90 -
87
Invertebrate Community Composition — lO^g Cd/L
f~1 Zygoptera
H3 Anisoptero
Q Dipt era other
than Chironomidae
Iffifa Hemiptera
O Coleoptera
HI Trichoptera
Nov[Jon[Feb Mar Apr Jun AugjSep Oct Dec[Jan_ Mprj Apr|Jun
1976
SAMPLING DATE
1977
Figure 32. Percent community composition of macroinvertebrate community in channels
receiving 10 yg Cd/1.
-------
Ceratopogonidae in treatment channels resulted from the presence of one
genus, Dasyhelea sj> (Figure 34). Data taken from bottom sediment samples in
March 1977 indicated how prevalent these organisms became in channels receiv-
ing Cd inputs. In control channels a total of 6 Dasyhelea were collected
from 8 random bottom samples as opposed 484 and 661 individuals collected
from similar samples taken from 5 and 10 |Jg/l Cd treatments, respectively.
The increased Dasyhelea densities in control channels appears directly rela-
ted to decreased macrophyte colonization in channels receiving Cd inputs,
with a corresponding increased development of an algal mat covering the
channel bottom. Thomsen (1933) in Johannsen's (1969) book on Aquatic Diptera
indicates that the blanket algae of ponds is the preferred habitat of
Dasyhelea sp.
Ephemeroptera
20 r
10
Q.
E
a
en
.a
E
0
20
15
t 10
z
UJ
a
CO
z
<
o
tr
o
0
15
10
-• Control
•-* 5/ig Cd/L
-o I0>ig Cd/L
Nov Jon|Feb [Mar Apr Jun |Aug| Sep Oct Dec] Jon| Mor Apr Jun
Ca/libaetis and Baetis
•rv^-TT
Nov Jon Feb Mar Apr Jun Auq Sep |0ct Dec| Jon Mor Apr|Jun
Caenis
nf. n*.
Nov I Joi
Jon [ Feb [ Mor I Apr I Jun| AugjSep |Oct|Dec | Jon|Mor |Apr|jun|
1976
1977
SAMPLING DATE
Figure 33. Density of Ephemeroptera per sampler.
79
-------
Comparisons made between aquatic macrophyte biomass and macroinver-
tebrates abundance data, although sporadic and limited, indicated that the
presence of macrophytes influenced both the number and type of organism
present in many cases. Similar findings have been observed in natural aqua-
tic systems by Egglishaw, 1963 and by Cole, 1973. Populations exhibiting
distinct relationships to macrophyte colonization in our system were: 1)
Dasyhelea s£. , whose abundance decreased with increasing macrophytic coloni-
zation; 2) Erythrodiplax miniscula, who only began colonizing our system
during the second season after substantial macrophyte colonization had occurr-
ed in the upper portion of most streams; 3) Pantala hvmenaea, whose abundance
gradually declined during the second season as algal and macrophytic coloniza-
tion and growth expanded downstream. Therefore, any direct effect on macro-
phyte and/or periphyton colonization or growth attributed to Cd treatment
indirectly affected the macroinvertebrate community also.
12
10
o.
I 2
E
^
c
CO
z
UJ
0
CO
cr
O
Cera topoqonidae
\f
• • Control
&---.a 5/iq Cd/L
o o 10 ftg Cd/L
/ ;
/ /
/ ;
j.
0
12
10
8
6
4
2
n
•n f«o ff| •»" «*r»- •— 4|(3_~^Mjr- -A>
|Nov|Jan |Feb|Mar |Apr|Jun |Aug |Sep]Oct|Oec | Jan JMar
f-
Dasyhelea
.
;^^^^^^^^^^^^^^^^
-
P
/\
A/^ \
' / \ \
/^ N\ /
' / \ "s.
Aprjjun
f
1
1
I
i
/'
/
//
//
/ /
/ /
t
i
i
I
^n «n *^ *n «oB*^-«^X^X*=^n «c *k'' ~^*
|Nov|Jon |Feb|Mof[Apr|Jun|Aug|Sep|Oct [Dec |Jon |Mor |Apr|Jun
4 -
««n »o ^n «T1 T°T-—"— ~— _-• . mi . "l| '" . •ir-.-M
|Nov|Jan |Feb|Mor|Apr| Jun | Aug| Sep|0ct [Dec | Jan| Mof|Apr| Jun
Figure 34. Density of Ceratopogonidae per sampler.
80
-------
Microinvertebrates
Between 88 and 100% of the macroinvertebrate community was accounted for
by four taxa; Chironomids, Annelids (Pristina sp.) Copepods and Ephemerop-
terans (Figures 35 and 36). The greatest fluctuations in the composition of
macroinvertebrate community inhabiting the control channels resulted from
shifts in the relative importance of dominant taxa, while the relative compo-
sition of the rarer taxa remained relatively constant.
Annelids (Pristina sp.) (Figure 35) and Copepods (Figure 36) displayed
the most pronounced trends observed in all the invertebrates enumerated for
this section of the study. These observations would most likely have been
overlooked in conventional macroinvertebrate surveys or would only have been
observed if additional macroinvertebrate surveys were conducted at the same
time. The reason for these groups being reported in this section stems from
our previously mentioned changes in sampling techniques which allowed for the
collection and enumeration of these two groups.
Data on the Pristina, annelids and Copepods showed their abundances to
decrease noticeably with increasing Cd treatment. Pristina populations in
control channels were greater than in treatment channels (Figure 37). How-
ever, in only 2 of the 14 sample periods were the differences significant.
The results of Copepod data, where again, larger populations in controls as
opposed to treatment channels during the period of Cd inputs (Figure 36). In
o>
~a.
E
o
tn
1200
700
600
E 500
c
>- 400
\-
~ 300
LU
O
200
100
Pnst/na qequiseta
1178.71
Control
s^g cd/L
10/ig Cd/L
1976
Figure 35
1977
SAMPLING DATE
Density of P_. aequiseta per sampler
81
-------
9 of 14 sampling periods the controls had significantly larger populations of
Copepods than did treatments. Eight of these significantly different samp-
lings occurred during the Cd input period. Additional information on these
groups of organisms can be found in the microinvertebrate segment of the
report.
The major groups affected by Cd were flagellated and ciliated proto-
zoans, testate amoebae of the genus Difflugia, and ostracod and copepod
crustaceans (Table 16). Significant F values demonstrate a Cd effect but do
not indicate whether population densities were increased or decreased in the
treated systems. Cadmium reduced populations of the two crustaceans and the
amoeba Difflugia, the expected effect. However, densities of the flagellate
and ciliate protozoans in the channels receiving Cd were elevated relative to
those in control systems. In general the densities of microinvertebrates was
I50r Eu eye lops aailis
125 -
Q.
E
o
O)
JD
E
^
c
100 -
UJ
O
IT
O
NovUan Feb Mar Apr Jun Aug Sep Oct Dec Jon Mar Apr Jun
1976
1977
SAMPLING DATE
Figure 36. Density of IS. agilis per sampler.
82
-------
elevated in channels receiving Cd (Figure 37). Rotifer densities were also
elevated in channels receiving Cd in 11 of 12 samples; however, within treat-
ment variability was sufficiently great that only four statistically signifi-
cant F values were observed. This is significant in that Buikeraa et. al.
(1974) observed that rotifers might be a convenient organism for bioassay
work. The lowest Cd concentrations used in their studies, however, were much
greater than those used in our study.
It is difficult to compare protozoan densities from the sponge samplers
to other research work. No long term studies of the effects of metals on
this group have been conducted. In general, laboratory studies have been
carried out with relatively high metal concentrations and single species
(Gray and Ventilla, 1973; Milles, 1976; Bergquist, 1976; Giesy et al., 1977);
Lansing et al. , 1977) or with high metal concentrations and simple communi-
ties (Burbanck and Spoon, 1967; Ruthren and Cairns, 1973) exposed for short
time periods. The apparent stimulatory effect of Cd on ciliate (especially
Paramecium barsaria) and flagellate (especially Chlamydomonas sp.) protozoans
may be due to release from predation competition. Eucyclops agilis, the only
copepod observed in the samples, is essentially a vegetation (Fryer, 1957)
10,000 r
Microinvertebrotes
Control
Cd/L
10/tgCd/L
M I A IMIJ I JI A IS I 0
1976
1977
SAMPLING- DATE
Figure 37,
Total number of microinvertebrates observed per month
in polyurethane sponger.
83
-------
TABLE 16. EFFECT OF CD ON DENSITY OF TAXA IN SPONGE SAMPLERS.
# of months
# of significant sufficient data
Taxa F values (P < 0.05) available
Protozoa
Sarcodina (excluding Difflugia) 1 12
Difflugia sp (3) 11 12
Flagellata 6 12
Ciliata (excluding Paramecium
burSaria) 10 12
Paramecium bursaria 9 12
Platyhelminth.es
Turbellaria 4 11
Aschelminthes
Rotifera 4 12
Nematoda 1 12
Annelida 1 7
Arthropoda
Crustacea
Branchiopoda (Alonopsis
elongata) 2 12
Ostracoda 8 12
Copepoda (Eucyclops agilis) 10 12
Insecta
Diptera (Chironomidae) 1 8
and its increased densities in the control channels may have been responsible
for the reductions in Chlamydomonas populations in these control channels,
but E. agilis is too small to feed on P. barsaria and rotifers. That Cd has
a direct stimulatory effect, perhaps by controlling parasitic bacteria or
fungi, cannot be discounted.
Macroinvertebrate community diversity and evenness were calculated using
five different indices: Simpson's Index (Bowman et ,a_l. , 1971), (Figure 38);
evenness of Simpson's Index (Bowman et al. , 1971) (Figure 39); Shannon's
Index (H) calculated using log- (Figure 40); Evenness of Shannon's Index
(H/log (N-SPP) (Figure 41); Macintosh's Index (Pielou, 1969) (Figure 42);
Evenness of Macintosh's Index (Pielou, 1969) (Figure 43); Probability of
Interspecific Encounter (Hurlbert, 1971) (Figure 44), Evenness of Probability
of Interspecific Encounter (Hurlbert, 1971) (Figure 45); and Renzi's General-
84
-------
ized Entropy Series, first with a = 1 then as a = 2 (Hill, 1973) (Figures
46-49). The objective of this exercise was not to compare the accuracy of
these indices in distinguishing the potentially subtle effects of chronic Cd
exposure, but instead to determine which of these indices might best illus-
trate any subtle effects which might occur. The basis by which diversity and
evenness were calculated are: 1) as individual samples, the means of which
are plotted by treatment and sampling period in portion A of Figures 38-46
and 48; 2) by summation, here each sample is added to a running sum and di-
versity and evenness is calculated on the sum, portion B of Figures 38-46 and
51 represent values calculated on the composite total of all samples collec-
ted for each treatment and sample period. The reason for calculating diver-
sity and evenness on both basis was to see if sample size affected our abil-
ity to distinguish chronic effects.
Immediately after cadmium input was started there was a decrease in di-
versity in those channels, while the diversity in central channels increased,
due to continued colonization (Figures 38-49).
x
UJ
Q
CO
z
o
co
0.
CO
I I I I I I I I I I I I
NovI Jon |Feb| Mor [Apr | Jun| Aug| Sep|Oct|Dec | Jon|Mar | Apr |Jun
B.
Control
5^qCd/L
10/ig Cd/L
1.0 r
0.5
0.0
Nov|Jon |Feb|Mor|Apr|Juri[Aug|Sep |Oct|Dec |Jon| Mor|Apr| Jun|
1976 1977
SAMPLING DATE
Figure 38.
Simpson's diversity index. A, means calculated
across sampler by sampling period with two stand-
ard error confidence intervals indicated. B,
calculated by summation.
85
-------
Statistically significant (P < 0.05) differences in diversity and even-
ness occurred in 5 of 14 sampling periods. These differences occurred during
April through June, the spring and early summer emergence and recruitment
period, and again in October through January, the fall and winter minimum
population period. In all five cases the control channels had significantly
higher diversity and evenness values than did channels receiving either 5 or
10 |Jg/l Cd. There appeared to be no significant differences in the 5 and 10
pg/1 treatments. Our results indicate that significant differences occurred
only during the period of Cd input and not during the three months prior to
or the two months after Cd was input. Therefore, it appears that 5 and 10
(Jg/1 Cd does affect the invertebrate community sufficiently to affect both
diversity and evenness calculations. It should also be pointed that re-
gardless of the method or index employed the results were similar with only
the magnitude of the values calculated being affected.
Diversity and evenness indicate that channels receiving 5 and 10 pg/1 Cd
may be somewhat less stable than control channels as did the abundance and
community composition data hypothesis of Jones (1940, 1941, 1958) that in-
sects are probably indirectly affected by chronic levels of metals exposure
I I I I I i I I I I I 1 I I
£ °5
o
z
« 00
z
o
CO
a.
5
CO
u.
o
CO
CO
LJ
g '-°
Nov
Jon
Feb
Mor
Apr Jun Auq SepJQct [Dec |jon Mor Apr Jun|
B
• Control
* 5/iQ, Cd/L
c iQ/tg Cd/L
LU
05
0.0
Nov | Jon |Feb|Mor [Apr | Jun |Aug[ Sep|Oct|Dec | JonJMar [Apr] Jun
1976
1977
SAMPLING DATE
Figure 39. Eveness of Simpson's diversity index. A, means
calculated across sampler by sampling period with
two standard error confidence intervals indi-
cated. B, calculated by summation.
86
-------
and it is the effect on the surrounding environment which harms them the most
(direct effects in algae and macrophytes etc.). In our study, what we have
observed is not necessarily the effects of 5 and 10 |Jg/l Cd on existing algal
and macrophyte communities, but instead the retarding of their successional
development. Thus the 5 and 10 pg/1 Cd channels were maintained at earlier
successional stages than controls, and for this reason, probably were some-
what more unstable.
Peak diversity occurs approximately 30 days later in channels receiving
Cd than in those receiving no Cd regardless of calculation methods employed
(Figures 40-51). This is apparently the result of delayed development of
individuals, with a concomitant delay in hatching and thus community struc-
ture changes.
In i
I.Or
—• Control
—•* 5/tg Cd/L
10/tg Cd/L
Nov | Jon |Feb|Mor| Apr Jun AugSep|Oct Dec Jon | Mor|Apr|Jun
Nov|Jon [Feb|Mar]Apr
Oct Dec Jan
0.0
1976
SAMPLING DATE
1977
Figure 40. Shannon's diversity index A/ means calculated
across sampler by sampling period with two
standard error confidence intervals indicated.
B, calculated by summation.
87
-------
Molluscs
Due to the large amount of literature available on trace metal accumula-
tion by molluscs (Bertine and Goldberg, 1972; Wier and Watter, 1976 and
Pringle et al., 1968) and their increasing use as biomonitors of heavy metals
pollution, both gastropods and pelecypods were used in our study.
Two gastropods were proposed for study; the pulmonate, Helisoma
trivolvus and the prosobranch Campeloma lima. Through the use of these two
distinctly different physiological forms it was believed that additional
information regarding uptake and biological effects of Cd on gastropods under
natural, but controlled conditions could be obtained.
Helisoma trivolvus was selected for our initial work because its natural
habitat.is similar to the littoral habitat created in our experimental system
and they were easily collected in large numbers on the SRP. Also if H.
trivolvus could be successfully transplanted into our system with good sur-
vival, sufficient data would be acquired to enhance our chances of success
X
ui
0
z
V)
z
o
I
in
l.O
0.5
0.0
.111 I
I i I I I
[Nov]jan |Feb[Mar [Apr | Jun[Aug| Sep|0ct |Dec| Jon|Mar |Apr|Jun
B.
Control
a s cd/L
Cd/L
LU
z
LU
1.0
0.5
0.0
Nov|Jan |Feb|Mar |Apr|jun |Aug| SeplOct |Dec| JonlMar |Aprl Jun I
1976 1977
SAMPLING DATE
Figure 41.
Evenness of Shannon's diversity index. A,
means calculated across sampler by sampling
period with two standard error confidence
intervals indicated. B, calculated by sum-
mation .
88
-------
with C. lima. Camploma lima being a prosobranch snail, is a gill breather
which as a group are more limited to flowing or at least well oxygenated
waters than are the pulmonates such as H. trivolvus. Therefore, if H.
trivolvus would not survive our system successfully then the chances of C
lima success were slim.
Six hundred H. trivolvus were collected by hand from Par Pond on the
SRP, acclimated for two to four weeks, and 100 were transplanted into each
channel. Transplanted organisms were marked with a dot of fingernail polish
to distinguish them from offspring produced during the study. Initially, H.
trivolvus appeared to adapt well to our experimental system. However, two
weeks after their introduction, mass mortalities began occurring in all
channels, prior to Cd input. This phenomena continued after Cd input began
without alteration and by the end of three months no H. trivolvus or even
relic shells could be found in any of the experimental channels. As a result
of our experiences with H. trivolvus it was decided not to attempt to trans-
plant C. lima.
X
LU
If)
X
-------
The two pelecypods used in the study were Anodonta imbecilus (papershell
clam) and Corbicula fluminea (Asian clam). Anodonta imbecilus was selected
because it is common in the softwater ponds and reservoirs of the south-
eastern United States and because it could be collected readily. Corbicula
fluminea was chosen because it is an ubiquitous nuisance species throughout
the United States and is reported to be tolerant of environmental stresses,
allowing colonization to occur nearly anywhere adults and/or larvae can
migrate. All clams were collected by hand and acclimated for two to four
weeks prior to being transplanted into the channels. Thirty two A. imbecilus
of various sizes and 150 C. fluminea each of two size classes (1.7 cm and 2.6
cm shell length) were placed into the tail region of each channel. All
organisms were placed directly into compartmentalized areas of channel sedi-
ment where they were allowed to move freely.
Anodonta imbecilus adapted well to our experimental situation, moving
freely and filtering regularly. However, they did not survive due to heavy
crayfish predation. The crayfish, Procamberus acutus acutus, which we had
introduced into the channels, were observed crushing the paper thin outer
x
V)
o
o
o
1.0
05
x
LJ
Q
? 0.0
I I I I I i I 1 I I 1 1 I I
Nov
Jan
Feb
Mar
Apr
Jun
Aug
Sep
Oct
Dec
Jan
Mar
Aprjjun
a -- o
Control
5/iqCd/L
Cd/L
B.
u.
o
en 1.0
LU
z
UJ
a °-5
-
r\~* ^"^Dl
JF^\
-fr
t
0.0
Nov
Jan
Feb
Mar
Apr
Jun
Aug
Sep
Oct
Dec
Jan
Mar[Apr
Jun
1976
SAMPLING DATE
1977
Figure 43. Evenness of Macintosh's diversity index. A,
calculated across sampler by sampling period with
two standard errors confidence intervals indica-
ted. B, calculated by summation.
90
-------
o
u_
O °'° |Nov| Jon|Feb|Mor [Apr|Jun |AuglSep |0ct [ Dec| Jon|Mor | Apr | Jun|
W^*^2
^/
a.
CO
a:
LJ
B.
• • Control
tt & 5^g Cd/L
o a iQ/ig Cd/L
g i.o
m
<
m
£E
a
0.5
0.0
Nov|jon]Feb|Mor|Apr|Jun |Aug|Sep|Oct|Dec [ Jon|Mor |Apr| Jun j
1976 1977
SAMPLING DATE
Figure 44.
Probability of interspecific encoun-
ter diversity index. A, calculated
across sampler by sampling period
with two standard errors confidence
intervals indicated. B, calculated
by summation.
x
LJ
O
z
3
O
Q.
V)
cr
1.0
rl I I I I I I I I I I I I I
0.0
Nov|Jon |Feb |Mor \ Apr] Jun |Aug |Sep|Oct [Dec |jon |Mor [Apr |jun |
u.
o
m
<
CO
o
a:
Q.
B.
I Or
• • Control
a -A 5Mg Cd/L
a o \0fiq Cd/L
to
CO
LJ
0.5
> 0.0
Nov|Jon |Feb |Mor [Apr [Jun |Aug| Sep|0c11 Dec | Jon|Mor |Apr |Jun|
1976 1977
SAMPLING DATE
Figure 45.
Evenness of probability of inter-
specific encounter diversity index,
A, calculated across sampler by
sampling period with two standard
errors confidence intervals indi-
cated. B, calculated by summation,
-------
margins of the clam's shell, damaging the mantle flap and detaching portions
of the mantle from the shell. Once damaged, A. imbecilus died rapidly and
were eaten by the crayfish.
Corbicula fluminea reacted somewhat differently than did A imbecilus.
They were unaffected by the crayfish, but never adapted to the system. They
continued to show high mortality both before and after Cd exposure.
The abnormally high mortality of H. trivolvus and C. fluminea observed
during our study was attributed to water quality. Water quality of our ex-
perimental system was initially thought to be adequate to sustain small popu-
lations of locally occurring molluscus species. As a result of the abnormal
mortality rates observed early during the Cd study, however, a detailed
review of all water chemistry parameters was conducted. From the results of
A.
en
UJ
E
u
CO
o
CT
II
3.0
Q
U
M
2.0
ui
Z
UJ
'-5
1.0
0.5
0.0
Control
5^g Cd/L
10/ig Cd/L
Nov| Jon |Feb|Mor | Apr|jun |Aug |Sep |0ct [Dec |Jon |Mar|Apr| Jun|
1976
SAMPLING DATE
1977
Figure 46. Renyi's generalized Entropy series (a = 1)
calculated across sampler by sampling period,
with two standard errors confidence intervals
indicated.
92
-------
this review and concurrent testing of all water quality parameters, it was
determined that calcium levels in our system were not being maintained at
levels previously believed to exist. Wetzel 1974, states that calcium has
been implicated in numerous ways in the growth and population dynamics of
freshwater flora and fauna. Wilbur and Yonge (1964) indicate that the most
important chemical variable in determining the occurrance and distribution of
molluscus in the environment is calcium. Although we could find no infor-
mation about exact levels at which calcium determines the presence or absence
of molluscs, we could relate such measures as hardness and alkalinity (as
relative measures of total calcium) in our system to reported levels which
have appeared to limit molluscus distribution. Mean hardness, calcium, and
alkalinity values for our system were 29.0 mg/1, 10.8 mg.l, 9.9 mg.l, re-
spectively. Harman (1969) has reported only a few molluscs surviving at
levels around 21 ppm. While Harman (1970) and Pennak (1953) have both indi-
cated that a total alkalinity of 15 mgl appears to be essential for the
welfare of molluscan populations. Therefore, the results of other research-
ers suggests that our water quality is inappropriate to conduct studies
involving molluscs (with the possible exception of some of the Anodontinae
which are known to be softer water species). Even with the occurrence of
abnormally high mortality and the potential for water quality effects, C.
C 3.5
o
co 3.0
LU
o:
LU
w 2.5
o.
o
OL
LU
O
LU
N
2.0
1.5
cc. 1.0
LU
LU
CO
0.5
o.o
B.
//\
-H
^y
v ^
\
v
\
\
' \
\ 1 \
\ / \
\ \
\ /' \
. \ (
^\ I
\ r -\ '
V\
K /
l^-J
§
^ f
\\ /
\\ /
\\ /o
\Y ^
V"
-Jr
• • Control
A—^ 5 Mg cd/L
D o lO/igCd/L
y////////M//m^^^
Novj Jon [Feb Mar Apr Jun |Aug Sep
1976
SAMPLING DATE
1977
Figure 47. Renyi's generalized entropy series (a=l) calculated by
summation.
93
-------
fluminea populations displayed effects attributed to Cd exposure in both size
classes utilized. The tendency for mortalities in treated and control orga-
nisms to diverge at approximately 13 days after Cd inputs began, resulted in
significantly higher mortality levels for treated organisms than controls
(Figures 50 and 51). Median survival times for large-size-class treated and
control organisms were 20.8 and 29.2 days, respectfully. While small size
class organisms possessed median survival times of 17.5 and 27.5 days for
treated and control organisms. Therefore, results indicate that smaller
organisms were more susceptible to Cd and that Cd is probably acting as an
additive stressor in this case, causing treated organisms to die approxi-
mately 10 days before controls. Data also indicate that the total number of
deaths occurring in either size class is not significantly affected by treat-
ment.
A significant difference in mortality rates of small size class orga-
nisms at different treatment levels did occur during a 10 day interval begin-
ning three days after Cd inputs started (Figure 50). Results indicate that
during this interval higher mortalities occurred in 5 pg/1 treated organism
than in the 0 fJg/1 or 10 (JgA treated organisms. This observation may well
Cd
, / V
\ / ,***
Control
5/iq Cd/L
Cd/L
Nov| JonjFeb |Mor|Apr | Jun |Aug|Sep|Oct [ Dec |Jon|Mor [Apr | Jun
00
1976
SAMPLING DATE
1977
Figure 48- Renyi's generalized entropy series (a = z)
calculated across sampler by sampling period,
with two standard error confidence intervals
indicated.
94
-------
be explained by avoidance mechanism utilized by pelecypods where they just
close up, stop filtration processes and essentially go into anaerobic metabo-
lism maintaining only minimal bodily functions. This condition may be main-
tained until sensory receptors detect that the perturbation has passed or
until metabolic waste products reach a potentially hazardous level and the
organism has to start filtering again in order to eliminate waste. The
result of this behavior is increased toxicant exposure at lower ambient
toxicant levels. Harrison (personal communication) has observed similar
results in low level metals toxicity work using pelecypods.
The unexpected mortality of molluscs in our system severely affected our
efforts to collect Cd accumulation data. Therefore, no analyses are avail-
able for H. trivolvus or A. imbecilus Cd uptake and only limited data are
available for C. fluminea transplanted in the study (Table 17). Results
indicate C. fluminea, like other mollusks, concentrate Cd. The levels accu-
mulated appear to be in the same ratio as exposure levels; however, they are
orders of magnitude greater. Another point of interest is that although Cd
concentrations accumulated in 10 |Jg/l treated organisms are nearly double the
levels found in 5 |Jg/l treated organisms, the additional Cd body burden did
not significantly increase mortality.
CVJ
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CT
UJ
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a.
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cr
UJ
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or
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or
3.0 r
2.5
2.0
1.5
1.0
0.5
0.0
B.
• • Control
a •* 5/tg Cd/L
o o I0/*g Cd/L
D—
Nov Jon Feb Mar
///////////////////M
Cd On''
'W//////////////A
Apr Jun AugSep Oct
n[Mor |Apr] Jun
1976
SAMPLING DATE
1977
Figure 49. Renyi's generalized entropy series (a=z) calculated by
summation.
95
-------
150
140
130
120
110
100
co
2
52 90
o
tr 80
O
Z 70
Q
UL
O 60
UJ
Q
Estimated population decline
resulting from sampling
Corbicula fluminea Mortality
Large Size Class
Control
CH 5 8 10/ig Cd/L
U) 50
40
30
20
10
05 1.0 2.5 6.0 13.0 24.0 38.0
TIME (days)
Figure 50. Mortality of large C. fluminea as a function of
time.
96
-------
en
Estimated population decline
resulting from sampling
Corbicula fluminea Mortality
Small Size Class
^ Control
IH 5/xg Cd/L
CD IC>g Cd/L
150
140
130
120
I 10
w I0°
1 90
S
< 80
UJ
o
u. 70
O
H 60
I 50
40
30
20
10
0.5 1.0 2.5 6.0 13.0 24.0 38.0
TIME (days)
Figure 51. Mortality of small £. fluminea as a function of
time.
97
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TABLE 17. MEAN CD CONCENTRATION IN C. FLUMINEA WHOLE TISSUE EXPRESSED ON
A DRY WEIGHT BASIS
Treatment
Background
0 ppb
5 ppb
10 ppb
Treatment
Background
0 ppb
5 ppb
10 ppb
Large Size Class (Shell Length 2.6 cm)
A/16/76
M8 Cd/g
2.29
3.60
43.76
62.36
Small Size Class (Shell Length 1.7 cm)
4/16/76
Mg Cd/g
3.83
6.06
36.10
77.02
5/20/76
Mg Cd/g
5.24
62.88
5/20/76
Mg Cd/g
8.37
54.31
123.66
98
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SECTION X
FISH
INTRODUCTION
It has been determined that the occurrence of "itai itai" disease in Ja-
pan is correlated with fish which are able to concentrate Cd in their tissues
(Piscator, 1974; Perry et al, 1976). Therefore, there is a decided need to
assess the effects of Cd on fish populations since Cd in fish tissue can re-
sult in increased exposure to human. Measurements of acute toxicity in fish
have documented species dependent toxic thresholds varying from less than 10
ug/1 to greater than 55 M8/P (Kumada et a_l, 1973), from very low water con-
centrations (Cearly and Coleman, 1974; Eisler et al, 1972; Fowler, 1975;
Kneip and Lauer, 1973).
Most previous studies of Cd toxicity to and uptake by fish have been
conducted in short-term laboratory systems, or based on completely uncon-
trolled field sampling. Neither of these two types of information alone is
useful in constructing predictive models of the environmental behavior of Cd
or understanding of mechanisms of Cd accumulation. The microcosm system used
here was a unique opportunity to observe accummulation of Cd by fish from
continuous low water concentrations in a relatively natural complex habitat.
The mechanisms of Cd accumulation by fish must be understood before valid
models of Cd flux through fish populations or biotic communities can be
formulated and elemental cycling patterns predicted (Hardisty e_t al. , 1974;
Miettinen, 1975). Fish can potentially accumulate metals from both ingested
food items and directly from water. Kinkade and Erdman (1975) reported that
aquatic organisms accumulated Cd faster from soft than hard water. This
indicates that, in soft waters such as those of the southeastern United
States, Cd may be rapidly transported into biotic components of aquatic
communities. Two studies were conducted. One was to determine the relative
importance of food and water as vectors of Cd uptake to the mosquitofish, a
species ubiquitous in southeastern aquatic systems and a potentially impor-
tant compartment in aquatic and terrestrial food webs. The other study was
performed with bluegill and mosquitofish to determine long term Cd uptake
rates.
The two species used were the common bluegill (Lepomis machrochirus) and
the mosquitofish (Gambusia affinis). The bluegill is a carnivorous, warm wa-
ter species, commonly found in lakes, ponds and streams having a moderate
amount of vegetation, is frequently used as a bioassay organism and is a much
sought after human food source. The mosquitofish, common in many southern
lentic aquatic situations is a livebearer and a surface feeder (primarily
mosquito larvae).
99
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METHODS AND MATERIALS
Mosquitofish, (Gambusia affinis), were seined from Asphalt Pond, located
on the SRP. Two hundred fish were randomly placed in each channel by 1 March
1976. During the first month fish were in the channels, there was high mor-
tality, therefore dead fish were replaced until 18 March 1976, when Cd input
was initiated. The pectoral fins of these fish were clipped into the ray so
that they could be identified as initial fish, as opposed to fish born in the
channels. From April through September 1976, four live mosquitofish were
sampled monthly from each stream by dip net. The channels were checked daily
and dead fish collected. Both dead and live sampled fish were wet weighed
and frozen in plastic bags for future Cd analysis.
Bluegills (Lepomis macrochirus), were trapped from Rischer Pond located
on the SRP. On 19 and 20 January 1976, 50 bluegills were placed in each head
pool and tail pool. Dead bluegills were replaced until 18 March 1976 when Cd
input was initiated. On 4 March 1976 all bluegills were removed by electro-
fishing and replaced with 20 fish from Rischer Pond 9 March, 1976. In addi-
tion, 10 bluegills trapped from Par Pond, located on the SRP were added to
each tail pool. Beginning 20 April and lasting through September, 1976, a
single bluegill was sampled from each pool monthly. Both live sampled and
fish found dead were frozen in plastic bags for Cd analysis.
To determine the relative importance of Cd in food and water, a more
controlled study was conducted. Mosquitofish were maintained in cages and
fed either Cd contaminated food or clean food. Fish were acclimated for 72
hr in aquaria and separated by sex. Forty fish were randomly placed into
each of four 30 x 51 x 11.5 cm (water depth) 1/32 inch (0.08 cm) mesh stain-
less steel wire cages at a female-male ratio of 4:1 resulting in a total of
160 experimental organisms. Fish were acclimated in flowing well water for
168 hr. Cages were suspended in PVC-lined concrete troughs receiving well
water at a rate of 94.6 A/min resulting in a water velocity of 1.3 cm/s. Two
cages were suspended in flowing well water while two cages were suspended in
flowing well water containing 10 pg Cd/2. The Cd concentration was main-
tained by continuously metering in stock CdCl~ with daily calibrated persis-
talic pumps. Stainless steel screens were used to decrease cage volume after
each sampling to maintain constant fish to cage volume ratio throughout the
experiment. Fish were fed ajd libitum twice daily. Cages were cleaned daily,
30 minutes after feeding to remove excess food, feces and detritus.
Food consisted of Wardley's Basic Food Flakes (Wardley Products Co., Se-
caucus, New Jersey). Food flakes were blended to a fine powder and divided
into two portions. One portion was spiked with CdCl^, dried and reblended.
The resulting powder had the same consistency as the unspiked food. Unspiked
and spiked food had nominal Cd concentrations of 0.115 and 1.13 |Jg Cd/g dry
weight, respectively. One-tenth gram of spiked food was placed in 1 £ of
well water and allowed to stand for 5 minutes to determine the amount of Cd
lost before it was consumed. Food was then centrifuged from solution, using
a Sorvall SS-1 centrifuge equipped with a KBS continuous flow system
(Sorvall, Norwalk, Connecticut). The recovered residue was dried, weighed
analyzed for Cd.
100
-------
The experimental design was a 2 (food) by 2 (water) by 4 (time) three
way fully crossed design. Five fish were removed from each cage after 2, 4,
6 and 8 weeks for Cd analysis. Nominal Cd concentrations in the water were <
0.02 and 10 pg Cd/SL; while those in food were 0.1 and 1.0 pg Cd/g dry weight.
Data analyses were conducted with an IBM 360 model 195 computer using the
Statistical Analysis System (Service, 1972). Factorial effect means were
computed directly (Cochran and Cox, 1971). Significance of factorial main
effects was tested using 2-way analysis of variance (ANOVA) within time.
Comparisons of effect means within sampling were made using T-tests in
the absence of significant interaction terms (week 2, 4, and 6) and are pre-
sented with 95% confidence intervals. Comparisons of simple effect means in
the presence of a significant interaction term were made using Tukey's hon-
estly significant difference test and 95% confidence intervals for each sim-
ple effect mean are reported (Kirk, 1968).
Fish were freeze-dried, weighed and wet ashed in fired porcelain cruci-
bles using 2-4 ml redistilled HNO-, depending on sample weight. Samples were
heated to 70 C on a hot plate until NO evolution was negligible. Samples
were cooled, 1 m£H20 added and to 70° C until all NO. evolution ceased.
Fish food was digested in a similar manner.
The samples were allowed to cool to room temperature, diluted to 25 ml
with denionized HOH and stored in washed polyethylene bottles. Fish samples
were analyzed, using a Perkin-Elmer model 306 atomic absorption spectrophoto-
meter equipped with an HGA-2100 flameless atomizer and deuterium continuum
background corrector. Standard additions were performed and no significant
matrix interferences were found. (See Appendix I).
RESULTS AND DISCUSSION
Sampling fish was difficult because bluegills escaped from the pools
into the channels. Also, additional fish added to compensate did not retain
marks well and could not be identified from those that had been present for
longer periods of time. Also, birds often ate or partly destroyed bluegills
which died in the channels.
The mean Cd concentration in mosquitofish collected from Asphalt pond
was 0.45 + 0.16 |Jg Cd/g, dry weight (n = 6, + 2 SE). The mean initial Cd
concentrations in bluegills was 0.39 + 0.19 |Jg Cd/g, dry weight (n = 10, 2
SE), which is similar to that reported for southeastern bluegills (Giesy and
Wiener, 1977). Wet weight to dry weight ratios for mosquitofish and blue-
gills were 0.31 and 0.28 respectively. Mosquitofish rapidly accumulated Cd
from both the 5 and 10 (Jg Cd/£ treatments (Fig. 54). Fish exposed to 10 |jg
Cd/£ exhibited a significantly higher rate of accumulation.
This result is similar to that observed by Merlini et al. , (1973) for
Lepomis gibbosus. The Cd concentration in G. affinis tissue did not reach
equilibrium in either Cd treatment during the 6 month exposure. Mortality of
bluegill maintained in Cd treated channels was high and the bluegill popu-
lation became extinct at one time, so accumulation results are not presented
for this species.
101
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Although the mosquitofish had not reached an equilibrium value after 180
days of exposure, they did exhibit a leveling off trend (Fig. 52). For this
reason, uptake rates lor this population were calculated using the first 130
day:; of exposure. The rate of Cd accumulation by G. affinis on the first 130
days of exposure can he described by linear regression models (Table 18).
TABLE 18. LINEAR MODELS OF THE FORM y = mx + b of Cd UPTAKE BY G. AFFINIS
Slope (m)
Wt
Cd Exposure
5 Hg/L
0.14
Concentration
10 pg/L
0.23
SD of m
0.01
0.02
Intercept (pg Cd/g dry wt)
1.01
1.02
0.93
0.95
N
27.
19.
The results of this analysis indicate that the rate of accumulation of fish
exposed to 10 pg Cd/Jfc was approximately twice that of fish exposed to 5 (Jg
Cd/H. There was less lag in Cd accumulation in fish exposed to 10 pg/£ than
those exposed to 5
The fact that organisms achieve different equilibrium Cd body burdens
may be due to several mechanisms. A possible mechanism is a constant elimi-
nation rate (KQ) and donor controlled uptake rate (JJ (equations 7 and 8)
KQ
(7)
Uptake rate can be described by equation
where: ^ = J - KQ (8)
Q - steady state Cd concentration
102
-------
T = time
K = constant
Steady state concentrations (Q j can be calculated using equation (Q
ss
Cadmium accumulation by G. af finis residing in the channels receiving Cd
were fitted to the Von Bertalanfly model (using the Gauss-Newton it.rratiw
least squares Jjechniqua-p (Barr et aj. . , ,1378),) with derivatives of t.hf form
^2i= 1 - (e " J and ^ = (C )(T)(e ). The predicted equilibrium con-
centrations (Q ) are 35.7 and 61. 0 (jg Cd/g dry wt for fish exposed to 5 and
10 pg Cd/£ respectively (Table 19). The estimated uptake constants for fi.sh
exposed to 5 and 10 (jg Cd/A are 0.0058 and 0.0054, respectively. Uptake of
Cd by fish exposed to 5 and 10 pg Cd/£ fit quite well by the Von Bertanfly
model (Table 19).
A 3-way ANOVA was used as the preliminary test of signif U-.-jrir «• of food
and water sources of Cd in the cage study but the power of the U-st war; much
reduced due to the large differences in variances between treatments. Since
the primary aspect of this study was the main factorial effects and interac-
tions independent of time, differences in Cd body burdens were tested using
Cd Accumulation by 6. af finis
30
60 90 130
DAYS OF EXPOSURE
180
Figure £2.
Cadmium accumulation by mosquitofish (G. affinis)
n and 2 SE are indicated.
103
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TABLE 19. NON LINEAR LEAST SQUARES FIX OF CD ACCUMULATION BY G. AFFINIS.
DATA FIT TO Q = Q (1 - e ~K1) USING THE GAUSS-NEWTON ITERATIVE
TECHNIQUE. °
Source
df
5 Mg Cd/L
Sum of Squares
Parameter
Estimate
Asymptotic
Standard Error
Mean Square
Regression
Residual
Uncorrected total
2
33
35
9775.19
526.20
2708.59
4887.59
15.94
K
35.78
0.0058
10.89
0.0027
Source
df
Parameter
K
Estimate
61.00
0.0054
10 Mg Cd/A
Sum of Squares
Asymptotic
Standard Error
36.43
0.005
Mean Square
Regression
Residual
Uncorrected total
2
23
25
17216.53
1965.13
19181.66
8608.26
85.44
p < 0.001
104
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2-way ANOVAS within sampling period. While making the statistical tests
within sampling time more powerful and facilitating the reporting of the re-
sults of the analysis, the ability to compute rigorous statistical tests
across time was lost. Treatment effects on fish dry weight were tested using
the 3-way ANOVA since the variances were more similar and the factorial ef-
fects were independent. For ease of reference, mean Cd concentrations with
95% confidence intervals are reported for each treatment combination at each
sampling (Table 21) even though these values can be calculated from the fac-
torial main effect means (Tables 20 and 22).
There was no significant increase in Cd levels in fish maintained in
water containing less than 0.02 (Jg/£ Cd regardless of food ration (Tables 20,
21 and 22). Fish maintained in water containing 10 pg Cd/£ had significantly
higher Cd concentrations than those maintained in low Cd water after 2, 4, 6
and 8 wk (Tables 20 and 22). Cadmium concentrations due to waterborne Cd
plateaued between week 4 and 6 before sharply increasing between week 6 and
8.
Water was a highly significant source of Cd to mosquitofish throughout
the experiment (Tables 20, 21 and 22) indicating Cd is taken up directly
through the gills (Kumada et al. , 1973). Hiyama and Makoto (1964) reported
that the gills had the highest Cd concentration of any organ and suggested Cd
transport across gill membranes as an uptake mechanism. Mummichog also accu-
mulate Cd directly from seawater under continuous flow conditions (Eisler ejL
al., 1972).
TABLE 20. FACTORIAL MAIN EFFECTS OF CD LEVELS IN FOOD AND WATER ON WHOLE
BODY CONCENTRATIONS OF CD IN MOSQUITOFISH WITH 95% CONFIDENCE
INTERVAL AND F-TEST (P), n = 5.
Main Effects
Cd/g dry wt)
Week General Mean
(M)
2 8.85
4 13.97
6 16.07
Water
(W)
+15.1
(*)
+25.5
(j-a-\
„„)
+24.5
(**)
Food
(F)
+2.00
(NS)
-1.36
(NS)
-2.4
(NS)
Water Food
(WF)
+1.46
(NS)
-1.08
(NS)
-2.4
(NS)
95%
CI
+ 11.6
+ 7.5
+ 10.2
*p < 0.01
**p < 0.001
105
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TABLE 21. MEAN CD CONCENTRATION IN MOSQUITOFISH UNDER FOUR TREATMENT COMBI-
NATIONS OVER TIME, C.I. = 95%, N = 5.
Week
Water
Response
(MS Cd/g)*
Food
8
L
H
L
H
L
H
L
H
1.60 + 1.48
14.67 + 2.33
1.48 + 0.64
28.10 + 10.6
3.86 + 2.58
30.72 + 14.16
0.41 + 0.12
46.88 + 12.57
H
1.07 + 0.68
18.14 + 17.07
0.93 + 0.17
25.38 + 2.85
3.82 + 3.14
25.88 + 2.86
0.61 + 0.22
71.49 + 16.49
*dry weight
There was no significant difference between Cd concentrations in fish
fed high Cd level food and those fed low Cd food at either the high or low
water Cd concentration through week 6 (Tables 20 and 21). When 0.5 g of spi-
ked fish food was soaked in 1 S. water, 0.42 g was recovered. The recovered
food had a Cd concentration of 0.95 pg/g dry wt, or 84% of the added Cd.
Feeding was generally complete in 5 min so fish were exposed to a consider-
able amount of Cd via the food pathway. Food was a significant source of Cd
only after 8 weeks where the only significant interaction between food and
water sources occurred (Table 22). The significant interaction between food
and water Cd sources is indicative of non-additivity between these two fac-
tors. Consumption of Cd spiked food did not increase whole body Cd concen-
trations in fish maintained in low Cd water. The positive interaction term
indicates that more Cd was accumulated than could be explained by either
factor acting alone, which may have been due to physiological changes induced
by the previously accumulated Cd. This significant interaction may indicate
two uptake mechanisms which are integrated. Food may become an important
uptake vector only after a threshold body burden is reached causing a de-
106
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TABLE 22. SIMPLE EFFECTS AND INTERACTION TERM FOR WEEK 8 WHICH INCLUDES A
SIGNIFICANT INTERACTION BETWEEN FOOD AND WATER WITH 95% CONFIDENCE
INTERVAL AND F-TEST (P), n = 5.
Simple Main Effects
95% CI
F-test (P)
Food at low water
Food at high water
Water at low food
Water at high food
Food x water interaction
0.196
24.6
46.5
70.9
12.2
21.0 NS
21.0 *
21.0 ***
21.0 ***
*
*p > 0.05
***p > 0.0001
crease in the fishes ability to restrict Cd influx via gastro-intestinal
assimilation.
There were no significant differences in Cd concentration due to size or
sex in exposed or unexposed fish. The mean live weights and dry weights of
test fish did not change during the course of the experiment and were not
affected by any of the treatment combinations.
Fassett (1975) suggests that an organism will accumulate Cd as long as
there is a continuous supply and therefore will not reach equilibrium.
Investigations concerning organisms attaining equilibrium concentrations vary
depending on the type of system and organism. In a static system, Kinkade
and Erdman (1975) showed catfish and guppies to reach equilibrium in 7 days,
perhaps due to Cd depletion. In a flowthrough system, Cearley and Colemen
(1974) found that bluegills and bass reached equilibrium in 2 months, whereas
rainbow trout, when exposed to 1.0 pg Cd/£ attained equilibrium in 10-20
weeks (Kumada et al, 1973). After three months exposure to 10 |Jg Cd/Jd, mo-
squitofish had approximately 6 times more Cd in their tissues than did rain-
bow trout exposed to 10 pg Cd/8, for 10 weeks (Kumada et al., 1973). Hiyama
and Makoto (1964) found fish came to equilibrium with Cd in solution in 15
days but did not indicate whether this was under static or continuous condi-
tions. Sullivan et al., (1978) reported that fathead minnows came to equili-
brium with Cd in both laboratory and field experiments within 20 days.
Miettinen (1975) found that Cd administered in the diet of rainbow trout
(Salmo gairdneri) was rapidly eliminated with only 1% of the administered
dose remaining in the body after 42 days. Cadmium accumulation by white
catfish is greatest in the gastrointestinal tract with little Cd accumulated
107
-------
in skin and gills (Rowe and Massaro, 1974). Hardisty e_t al. , (1974) reported
that Cd in the tissues of marine fishes is related to the number of crus-
taceans in the diet, indicating food as an important pathway of Cd uptake.
When dace were fed Cd contaminated food the amount of Cd accumulated was
increased over water exposure concentrations alone (Kumada et al., 1973).
Fishes eliminate Cd through the kidneys, and fish removed from Cd containing
water are able to reduce their body burdens of Cd by excretion (Kumada et
al. , 1973). Cadmium accumulation has been linked with renal hypertension
(Schroeder, 1974). The rapid increase in Cd accumulation after an apparent
equilibrium in this study may be due to renal failure with a subsequent
inability to excrete Cd. Another mechanism which may be responsible for the
rapid increase in uptake after 6 weeks accumulation is induction of metal-
bothionein, a metal binding protein which prevents Cd binding to sulfhydryl
containing enzymes (Fassett, 1974). Small doses of Cd are able to induce
protection against subsequent massive doses (Fassett, 1974). Cearley and
Coleman (1974) suggested a mechanism of elimination which is triggered after
threshold concentrations are reached in excretory tissues such as kidney. In
contrast, Eisler (1972) suggested that Cd does not accumulate in fish because
it is actively excreted.
The relative importance of water and food as sources of Cd to fish may
be dependent on many factors such as food quality, relative Cd concentrations
in food and water, form of Cd in water and species of fish. This experiment
was conducted under strictly controlled conditions to minimize variability.
For the species studied, direct uptake from water is the more important
vector of accumulation. Future investigations should involve effects of
physical-chemical water parameters and use physiologically labeled food
sources such as prey items. Comparisons of uptake of several essential and
nonessential elements by a number of aquatic organisms are needed before
comprehensive models of cycling and fluxing processes can be described and
predictive models constructed.
Water quality is important in determining the availability of Cd to
biota (Giesy et al. , 1977). Wiener and Giesy (1978) found fish residing in
soft waters, such as those used in this study and common to many areas of the
eastern United States, have higher concentration ratios for Cd then fish re-
siding in harder waters. Since our research indicated that water was the
primary source of Cd to G. affinis, concentration factors (equation 9) are an
appropriate method of comparing relative availability between aquatic situa-
tions (Jinks and Eisenbud, 1972).
C
—
C (9)
w
where:
C = Cd concentration in the fish, |Jg/g dry weight,
C = Cd concentration in the water, M8/P-
C,, = concentration factor.
r
108
-------
For these comparisons of relative availability to be valid, the assump-
tion of equilibrium conditions must be met. While this condition is not
strictly met after 180 days of Cd exposure, Cd concentrations in the orga-
nisms seemed to be approaching an equilibrium. The concentration factors
after 180 days exposure were 4.9 and 3.8 for the 5 and 10 |Jg Cd/£ treatments,
respectively. From our data it is not clear whether the final equilibrium
concentrations in fish exposed to 5 and 10 |Jg Cd/J2 would be significantly
different. There is no significant difference between the Cd concentration of
fish exposed to 5 or 10 (Jg Cd/£ after 180 days, but this is due to the great
variability of these data. Hamelink (1976) reports that the variability
about the mean accumulation increases with time due to the inherant property
of a population of animals expressing their individuality. Also the uptake
and elimination processes involved tend to produce log-normal distributions
of non-essential elements in aquatic organisms, causing an over estimate of
population variability when represented as a mean and standard error (Giesy
and Wiener, 1977). If in fact the final equilibrium Cd concentrations of
fish populations exposed to similar Cd concentrations under different condi-
tions are similar, concentration factors will not be useful in assessing
relative availabilities between various systems.
Total metal concentrations may be the same and under different environ-
mental conditions exhibit different availabilities because of differences in
the actual concentrations of available metal. However, this was not the case
in the system studied here where Cd was present in the same form in the chan-
nels receiving both 5 and 10 |jg Cd/£. Thus to assess relative availabilities
of metals from different environments, the total concentrations in each envi-
ronment must be equal. Comparisons of concentration factors calculated for
fish studied in the channels to literature values would therefore be inappro-
priate.
Although this study was not designed as a toxicity bioassay and complete
recovery of dead organisms was not assured, some information on chronic toxi-
city in a complex situation of exposure via both food and direct exposure in
the water was gleaned. Mortality may be due to both direct toxicity and
secondary effects of Cd exposure to other components of the system. There
was little difference between mortality in control channels and those receiv-
ing 5 (Jg Cd/JH for either bluegill or mosquitofish, however, mortality in the
channel receiving 10 (Jg Cd/£ was approximately twice that of the other two
treatments (Table 23).
The bluegills initially placed in the channels were all dead within a
few weeks when exposed to Cd. These animals had been exposed to multiple
stressors. The fish had been starved to maintain a small size and were
transported to the channels on a warm day. The second attempted stocking of
bluegills captured in Par Pond on the SRP also exhibited high mortality when
exposed to Cd in the channels. Necropsies of dead fish revealed these ani-
mals were highly parasitized with metacercaris of Diplostoimilum scheuringi
(Trematoda). When unparasitized, unstarved fish were collected from Rischer
Pond, the initial mortality was much less. While this is not a rigorous test
of these effects, it does indicate that a number of environmental and physio-
logical parameters are important in determining Cd toxicity.
109
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TABLE 23. BLUEGILL AND MOSQUITOFISH MORTALITY BETWEEN MARCH AND JUNE 1976.
Treatment Bluegill Mosquitofish
0 17 21
5 29 23
10 53 55
Ball (1967) found acute mortality of rainbow trout at 10 (Jg Cd/£. He
also, however, found 96-hr LCLQ values of 1.0 pg Cd/£ for steelhead strout.
Giesy et aJL. , (1977) found LC1~ of Cd to mosquitofish in the well water used
in the artificial streams to be 0.9 and 2.2 at 30 and 28 C respectively.
When comparing literature concerning metals, toxicity in fish, it must be
noted that Cd toxicity in fish will vary depending on water hardness, pH,
alkalinity, temperature, dissolved oxygen and species (Giesy et al, 1977).
110
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SECTION XI
LEAF DECOMPOSITION
INTRODUCTION
Prior to human perturbation, most streams and rivers were densely cover-
ed with vegetation. Shielding from direct sunlight and the structure of
stream channels fostered the development of a heterotrophic based system. The
dominant energy source of small woodland streams is allochthanous input
(Petersen and Cummins, 1974). Only a small portion of the energy contained
in leaf material is directly available to aquatic animals (Barlocher and
Kendrick, 1974). The animal and microbial components of the streams commu-
nity have evolved to process these inputs, with the animal community relying
on micro-organisms to degrade recalcitrant plant substances such as lignin
and cellulose. The microbial proteins, fats and carbohydrates are then
readily available to animals which feed on them (Hargrave, 1970). Many
stream dwelling invertebrates prefer to eat partly decomposed, or condi-
tioned, rather than freshly fallen leaves (Kaushik and Hynes, 1971; Cummins,
1974) and may feed on the leaves to acquire highly nutritious fungal cells
(Barlocher and Kendrick, 1973). Because of the importance of fungi and
bacteria as intermediaries in leaf litter processing, their inhibition in
streams would mean a drastic change in community structure and decreased
secondary productivity. While the toxic and inhibitory properties of heavy
metals to aquatic microbes have been studied, little is known about the
effects of low levels of these toxicants on the colonization and leaf litter
decomposition by microbial communities.
This study was part of a program designed to determine the biological
effects of low levels of Cd (drinking water standards and below). Since leaf
litter processing is important in lotic aquatic systems, this function was
chosen as a critical function to be protected to maintain ecosystem integri-
ty.
METHODS AND MATERIALS
Cadmium effects on the heterotrophic community were studied using leaf
litter decomposition packs. Fresh leaf matrriil was placed in 0.3 cm mesh,
15.2 cm square stainless steel envelopes (Fig. 53-54). These envelopes were
tied at the top and sides such that 1.0 cm openings remained on each side.
Leaf material was placed into each envelope in the order given in Table 24.
Two each of Type I and Type II (Table 24) envelopes were suspended 10 cm
above the bottom in each of the tail pools.
Ill
-------
macroin-
at 85° C
Leaf material was incubated in the tail pools for 28 wk between 28 April
and 22 November 1976. Leaf material was removed and examined for
vertebrates. Total dry weight biomass was determined after drying
for 96 hr. Multiple undried samples of each leaf type were fixed in 2%
Glutaraldehyde-0.1 M cacodylate buffer. Leaf samples were dehydrated by
serially washing 15~ min in 70%, 85%, 95% and 100% (twice) ethyl alcohol.
Dehydrated material was critical point dried and mounted on aluminum stubs
and gold coated for scanning electron microscopy. Each species was examined
for fungal and bacterial colonization and permanent records made.
-» - ••»^.
m
• '
•S
Figure 53. Leaf litter pack, Type I
112
-------
Leaf material was wet
rophytes (this report).
ashed using the methods reported for aquatic
The experimental design was a randomized nested design with two treat-
ments (5 and 10 |Jg/l Cd) and a control. There were two replicates of each
treatment channel with two replicates of each leaf pack type in each channel
resulting in four replicates of each leaf pack type per treatment. Results
were analyzed by standard Analysis of Variance Techniques and significance of
differences between means tested, using Tukeys'-w procedure (Steel and
Torrie, 1960).
Figure 54. Leaf litter pack, Type II.
113
-------
TABLE 24. INITIAL LEAF MATERIAL IN LEAF LITTER PACKS.
Species Wet Weight Added
To Each Envelope
(g)
TYPE I
Pinus taeda L. 5.0
Sassafras albidum (Nutt.) Nees. 3.0
Quercus nigra L. 3.0
Quercus laurifolia Michx. 2.0
Prunus americana Marsh. 2.0
Acer rubrum L. 2.0
TOTAL 17.0
TYPE II
Acer rubrum L. 3.0
Quercus nigra L. 3.0
Prunus americana Marsh. 2.0
TOTAL 8.0
RESULTS AND DISCUSSION
Exposure to both 5 and 10 pg/1 Cd significantly reduced leaf decomposi-
tion of Type I and II leaf packs (Table 25). There was no significant differ-
ence in leaf decomposition between Cd-treated channels for either litter pack
type. The ratio between initial live weights and final dry weights of Type I
and II leaf packs were 7.4 and 7.3, respectively.
Visual inspection of leaf material removed from the leaf packs, after 28
wk incubation revealed that leaves in 5 and 10 (Jg/1 Cd had deteriorated much
less than those in control water. Leaf material in control packs was brown
in color and many of the leaves had only veins and petioles remaining. Leaves
in the Cd treatments were green and completely intact. Microscopic examina-
tion revealed the intact structure of leaf surfaces, including leaf hairs and
stomates. Within the controls, the order of resistance to decomposition,
-------
from least to greatest, was: S. albidum, P. americanum, A. rubrum, Q.
laurifolia, Q. nigra and P. taeda. Although overall decomposition was re-
duced by the presence of Cd, S. albidum and P. americana were the most succep-
tible to decomposition, in the presence of Cd.
TABLE 25. EFFECT OF CD ON FINAL BIOMASS OF LEAF MATERIAL IN
LEAF LITTER PACKS EXPOSED FOR 28 WK. (X + 2 SD.)
Treatment
Dry Weight
(8)
CONTROL
5 |Jg/L Cd
10 MgA Cd
Type I
2.3 + 0.18
4.0 + 0.06i
4.0 + 0.73'
Type II
1.1 + 0.08
1.7 + 0.191
1.7 + O.ll1
a,b
not significantly different from one another,
n = 4, a = 0.05.
Few macroinvertebrates were found in the leaf packs. Two species of
Odonata, Erythrodiplax minuscula Rambur and Ishnura sp., on species of snail,
Limnea sp. and one species of flatworm were recovered from the leaf packs
suspended in control channels. The only macroinvertebrate recovered from
leaf packs incubated in treatment channels were flatworms.
Both 5 and 10 |jg/l Cd inhibited microbial colonization of leaf surfaces
(Figs. 55-58). Examination of leaf surfaces, using scanning electron
nucroscop^ (.SEMJ revealed the surfaces of leaves which had been suspended in
treatment channels were almost devoid of microbial colonization, while the
surfaces of leaves from control channels were well colonized. There were no
apparent differences in colonization of the upper and lower leaf surfaces or
position along the axes of pine needles.
Relatively little Cd was accumulated by leaf material suspended in the
channels (Table 26). Uptake by leaf material was directly proportional to Cd
concentration in the water.
When assessing the impact of a toxicant on an ecosystem, effects on the
most susceptible component of that system should be determined. While par-
ticular components may not be of primary economic or aesthetic interest, they
may be directly related to the overall desirability or productivity of a
stable ecosystem. Such is the case of the aquatic microflora responsible for
leaf litter decomposition in streams.
115
-------
Many microorganisms are adapted to high Cd concentrations (Chopra, 1971;
Doyle e_t al. , 1975). A variety of fungi and bacteria have been shown to be
tolerant to high concentrations of heavy metals, relative to concentrations
which are toxic to other organisms (Asworth and Amin, 1964; Ashida, 1965).
Cadmium presumably acts by inhibiting oxygen uptake. Sulfhydryl bonds, such
as those in cysteine protect cells from Cd toxicity by binding Cd and pre-
venting it from affecting enzyme systems (Tynelka and Zylinska, 1974).
Cadmium does not affect Escherichia coli metabolism of C- glucose until a
Cd concentration of 6 mg/1 is reached (Zwarun, 1973) and 10 (Jg/1 Cd had no
effect on the viability of a natural population of heterotrophic bacteria
(Albright et a_l. , 1972). Thormann (1975) found that the most sensitive
estuarine bacteria were inhibited by 100 ppm Cd while the less sensitive
species were able to grow in 400 mg/1 Cd. Heavy metal resistant acti-
nomycetes and bacteria have been isolated from soil near a zinc smelter which
were capable of at least 50% of normal growth at 700 pm Zn (Jordan and
Lechevaler, 1975).
Figure. 55.
Electron photomicrograph of the effect of Cd on micro-
bial colonization of P. taeda. A. Control.
116
-------
Leaf surfaces are rapidly colonized by fungi and bacteria under natural
conditions (Iversen, 1973). Beech leaves, for instance, lost 90% of their
weight during one year (Iversen, 1973). The results of our study indicate
that low Cd concentrations can inhibit the functioning of decomposing micro-
organisms. Heavy metals such as copper, zinc and cadmium inhibit fungal
spore germination (Ruhling and Tyler, 1973). Metals from a smelter have been
found to disrupt microbial processes in terrestrial ecosystems and depress
leaf litter decomposition (Auerbach e_t al. , 1976), while metals such as Cd
may affect the fungi colonizing the phylloplane of leaf surfaces (Gingell et
al., 1976).
Natural microbial communities are more complex than the pure cultures
often used to assess toxic effects of metals in laboratory studies (Albright
et al., 1972). Assessment of toxic and inhibiting effects of low levels of
heavy metals should be conducted in more complex situations than pure cul-
tures, and substrates. Ramamoorthy and Kushner (1975) suggested that many
synthetic media may complex heavy metals which may result in an underestimate
of metal toxicity or inhibition which may occur under natural conditions.
Figure 56.
Electron photomicrograph of the effect of Cd on micro-
bial colonization of P_. taeda. B. 10 ug Cd/1.
117
-------
TABLE 26. CADMIUM CONCENTRATION IN LEAF LITTER MATERIAL
EXPOSED FOR 28 WK. (X + 2 SD).
Treatment
Type I
Cd Concentration
(pg/g dry weight)
Type II
Control
5 pg/L Cd
10 pg/L Cd
2.8
O . J
18.4
t
t
t
0
2
4
.04
.2
.5
i .
12.
23.
9
2
}
1
*
t
o
I
/
.01
.4
.8
Figure 57.
Electron photomicrograph of microbial colonization of
- nigra. A. Control.
118
-------
Batch pure culture bioassays do not represent the complex colonization pro-
cedure which may be the critical stage in microbial decomposition of leaf
material under natural conditions. Bioassays, to determine toxic or inhi-
bitory effects of compounds on processes as complex as microbial colonization
and decomposition of leaf material must be conducted under conditions which
account for the complete colonization process, species interactions and be of
sufficient duration to allow for an organismal adaptation to occur.
Figure 58. Electron photomicrograph of microbial colonization of
fi- nigra. B. 5 yg Cd/1.
119
-------
SECTION XII
SYSTEM RESPONSES
INTRODUCTION
Even a comprehensive knowledge of the biology of individual species does
not provide enough information to accurately predict the complex interactions
of communities. Maki and Johnson (1977a; 1977b) suggested the ratio of
primary production to respiration (P:R) as a sensitive measure of environ-
mental stress. Parameters which reflect structural and functional attributes
of entire integrated systems are required. Some organisms may be more sensi-
tive than others and some more important to overall system functioning than
others. There are often functionally analogous species which may be inter-
changed with little effect on overall system functioning. Thus, rational
assessment of impacts of potential environmental perturbations must be made
in the context of what effects they will have on the entire community, and
not what effect they will have on individual taxa.
The information presented to this point has been largely static, des-
cribing the condition of state variables of community and population struc-
ture at various times throughout the study. This section will report the
effect of Cd on the dynamic ecosystem functioning measured by autotrophic
production, system metabolism and system export. The last two parameters
serve as integrators of the system's overall response to Cd and therefore may
be most suitable in application of data from this study to other aquatic
systems. An important property of natural flowing water systems is the
export of organic material to downstream systems (Odum, 1957a). Also of im-
portance is the retention and movement of a toxin in dissolved or particulate
forms to downstream communities. Therefore, an effort was made to quantify
the particulate organic matter and associated Cd leaving the streams in the
effluent water. Systems level measures may prove to be more efficient and
economical for assessing gross pollutional effects on aquatic communities as
functioning natural units.
METHODS AND MATERIALS
Measurements of total community primary production and respiration were
made on 30 June 1976, 28 July 1976, 23 September 1976, 20 October 1976, 24
November 1976, 9 February 1977, with a 24-hour upstream-downstream oxygen
diurnal analysis (Odum, 1956). Water samples were removed from the streams
by siphon at two hour intervals and dissolved oxygen content determined using
a YSI Model 54 dissolved oxygen meter calibrated using the azide modification
of the Winkler method (APHA, 1975).
120
-------
In the spring of 1977 a semi-automatic method of collecting diurnal
oxygen diurnal data was put into service. This system utilized 12 solenoid
valves (one at the head and one at the tail of each channel), two YSI oxygen
probes and meters, two timer boxes, and a chart recorder with another timer
attached. At each end all six gravity-fed lines passed through solenoid
valves into a single common line feeding the water over the end of the probe.
Dissolved oxygen was monitored for ten minutes each hour. Signals from the
corresponding meters were fed into a timer that switched input to the recor-
der at five minute intervals. In this manner, five minute recordings of
dissolved oxygen concentrations at each location were recorded for each hour
during day and night. Probes were calibrated several times during each 24
hour period.
The 0_ concentration for the head station was subtracted from the 0~
concentration in the same water mass at the tail station to determine oxygen
changes. These values were corrected for diffusion by calculating percent
saturation and using equation (10).
D = kS (10)
where: .
D = diffusion rate, gO_/m /hr ?
k = diffusion coefficient (gO-/m /hr at 100% saturation)
S = saturation deficit
A positive diffusion value indicates 0 diffusion into the water and there-
fore changes in oxygen concentration are corrected by subtracting D(gO~/m ).
Values of K between 0.04 and 0.8 were measured in the streams using the
floating dome method of Copeland and Duffer (1968) modified by McKellar
(1970). In no case did diffusion correction alter the metabolism values by
more than 10% of their uncorrected values.
Corrected rate of change data was plotted and areas integrated by count-
ing squares. Nighttime respiration values were averaged and 24 hour respira-
tion (R2A^ was assumed to equal the average nighttime hourly rate times 24
hours. Gross photosynthesis (PG) was the area above this average R line and
net photosynthesis (PNet) equals P- - R0/ . P/R ratios were calculated as
P /P G 24
PG/R24'
Exported organic material and associated Cd were quantified from October
1976 until August 1977. All effluent water from each channel was passed
through a four inch ABS plastic pipe into a "T" intersection which contained
a motor driven stainless steel mixer blade. Material collected on the end
screens was washed into the sampling system daily. Mixed effluent was sub-
sampled from each channel at a rate of 4 liters per day with a peristaltic
pump. These subsamples were filtered on to pre-fired Gelman A-E glass fiber
filters, dried, weighed, ashed at 450 C and reweighed to obtain ash-free dry
weight of exported material. From the length of sampling, the volume of
water exported and the volume of the collected subsample, channel export was
calculated as grams per channel per day.
121
-------
Export material was collected off of the end screens for routine Cd
analysis. Several grams of material were collected from each screen, blen-
ded, and subsampled for analysis. Subsamples were placed in tared crucibles,
dried, weighed, ashed at 450° C, and reweighed. The ash material was dis-
solved with hot HNO« and
RESULTS AND DISCUSSION
H2°2
and then measured as reported in Appendix I.
Overall community metabolism (production and respiration) was measured
by the diurnal oxygen method and algal production alone was estimated by the
short-term accrual on glass slides described earlier.
Exposure to Cd significantly reduced gross production, net production
and respiration at all sampling dates (Fig. 59). Exposure to 5 |Jg Cd/£ re-
sulted in values intermediate between controls and 10 fjg Cd/£. Shortly after
Cd input was stopped, metabolism values of all the channels converged and
were not significantly different from one another. During the period of
maximum summer productivity, however, channels which formerly received Cd
were slightly depressed compared to the former controls (Fig. 59) .
~ I
(^ 0
•^ 4
O*
3 3
2 2
IJ I
O
5 o
10/ig Cd-L
lj|A|SlOlNlDlJ|F|M|A|M|j|J"l
|J|A|S|0|N|D|J|F|M|A|MlJ|J|
_;o_n_t_rol
POROSS
|J|A|S|0|N|D|J|F|M|A|M1J |~3"1
1976 1977
SAMPLING DATE
Figure 59. Community metabolism. Gross primary
production and respiration with the
shaded area representing net production.
122
-------
Net aufwuchs production and that of its algal component in grams dry
weight or live cell volume per square meter per 28-day colonization period
(Figs. 60 and 61) should under-estimate the net production of stable commu-
nities since populations on glass slides begin colonizing clean slides each
sampling period. However, net production estimated in this manner may give
qualitative information on seasonal changes as well as a quantitative evalu-
ation of Cd effects.
During the first eight months of Cd input, net aufwuchs production as
well as that of the algal component was significantly higher in the control
channels. Aufwuchs net production was greatest in the sample collected in
June, while algal production was greatest in the August sample (Fig. 60 and
61).
Net production measured by the community method reached a maximum in
June and July (Fig. 59). During the second summer, after Cd inputs had been
terminated, net aufwuchs and algal production were both near minimum values
and yet the community data showed high net production in all treatments.
This descrepancy is due to the paucity of vascular plants in the streams dur-
ing the first summer and their subsequent increase to standing crop dominance
by the second summer.
1.5
CM
E
i.o
O
CD
0.5
0.0
SHORT-TERM GLASS SLIDES
Control
5M9 Cd/L
-o I0/ig Cd/L
1976
JLT S'|O|N|D|J|F|M|A|M|JIJ|
1977
SAMPLING DATE
Figure 60. Aufwuchs accrual on short-term glass slides with two standard
error confidence intervals indicated.
123
-------
There was also a greater accumulation of detritus in the control chan-
nels. O'Neill et al. (1975) reported that this type of organic accumulation
in aquatic systems contributes significantly to the persistence of the eco-
system. Thus the Cd input to the channels may have had long range effects on
the succession and stability of the community which developed in the chan-
nels. The accumulation of reduced carbon within control channels was due to
the greater net productivity in these channels. Experiments on leaf litter
decomposition indicated that the microbial decomposer system was inhibited by
Cd. (See section XI). Since the inorganic nutrient inputs to the channel
microcosm systems was low (see section V) as in other southeastern aquatic
ecosystems, the ecosystem stability would be greatly affected by the rate of
nutrient remineralization.
Table 27 summarizes the exported organic matter by treatment before and
after Cd input. A significantly greater amount of carbon was exported from
the control channels than from the treated channels during Cd input. The two
treatments were not significantly different with respect to export. After Cd
inputs were terminated, significant differences between former treatments
disappeared. Day to day export values for all streams were very variable and
highly dependent on external energy sources such as rain and wind, and inter-
nal changes such as loss of bottom mats and aufwuchs sloughing.
Average Cd levels in export by treatment are presented in Figure 62 on a
dry weight basis. For calculation of Cd exported, these values may be con-
verted to an ash-free dry weight basis by multiplying by 0.73 (determined
3.Or
CM
E
\
ro
E
o
LJ
2
O
<
_l
<
2.0
1.0
0.0
SHORT-TERM GLASS SLIDES
I
Control
5/tgCd/L
a—a lO^g Cd/L
J
F
M
A
M
J
J| A
s
0
N
D
J
F
M
A
M | J
J
1976
1977
SAMPLING DATE
Figure 61. Algal cell volume accrual on short-term glass slides with two
standard error confidence intervals indicated.
124
-------
from 25 export samples with a coefficient of variation of 12%). Great varia-
bility on a day to day basis was observed and is related to the variety of
sources in the channels that contributed to export. These sources were ben-
thic aufwuchs, wall and glass slide aufwuchs, and macrophytic plants. Cad-
mium levels in export material was proportional to water Cd concentration
levels. Cd export levels decreased to control levels within five months after
the inputs were stopped.
Nutrient cycling has been identified as a measurable attribute of the
abstract concept of ecosystem stability (Webster et al., 1975) and changes in
nutrient cycling have been suggested as measures of changes in community
structure (Odum, 1969). The interaction between communities and the elements
moving through them can influence species composition, diversity, and sta-
bility (Pomeroy, 1975). This approach is attractive because the nutrient
dynamics of an aquatic system can be more easily measured than traditional
population and community measure. Thus monitoring of changes in nutrient
dynamics may be a sensitive system level parameter, reflecting environmental
changes. One of the most sensitive biogeochemical cycles has been found to
be the nitrogen cycle (R. Todd, personal communication). Unfortunately, the
gaseous phases possible in the nitrogen cycle make monitoring of nitrogen
fluxes difficult.
200 r
_ 150
•o
o
o>
a.
z
o
100
tr
ui
o
o
o
-o
o
50
EXPORT
rt/ff/ffff/f/r/f/fffft
\ \
Ml
I I
• • Control
ir—o 5^g Cd/L
0 ° 10 M9 Cd/L
NDJF VI A MJJAS ONDJ F M A Ml J J A
1976
1977
SAMPLING DATE
Figure 62. Cadmium concentration in material exported from
the channel microcosms.
125
-------
TABLE 27. SUMMARY OF ORGANIC EXPORT FROM THE CHANNEL MICROCOSMS DURING AND
AFTER CADMIUM INPUT. VALUES ARE AVERAGES OF WEEKLY AVERAGES IN
GRAMS ASH-FREE DRY WEIGHT m • day
Cadmium On
Control 5 ppb 10 ppb
X 27.0 20.9 19.4
S.E. 2.1 1.6 1.7
n 21.0 21.0 21.0
Cadmium Off
Control
X 27.0
S.E. 4.7
n 8.0
5 ppb
25.6
3.0
9.0
10 ppb
25.1
4.3
9.0
We measured upstream and downstream NO- + NO- levels in each channel
(see section V, water chemistry). Ammonia levels were not measured routinely
because they were below the detection limits of direct analysis and required
concentration, which due to contamination resulted in high variability. The
mean NO - NO- N concentrations decreased significantly over the length of
each channel. There was no significant difference (P > 0.90) between any of
the treatments. While we did observe demonstrable changes in many popula-
tion, community and system level parameters, these Cd-induced changes were
not reflected in changes in NO- NO fluxing in the channels. While this was
not a rigorous test of this system level parameter, it does indicate that
other measures were more sensitive to Cd stress in our channel microcosms.
A program to measure nitrogen fixation as a system level functional
parameter was attempted but because of systematic experimental errors, will
not be presented here.
_o
Total orthophosphates (PO, ) concentrations did not vary significantly
between channels and did not vary-between upstream and downstream sampling
stations (Table 3). Sulfate (SO, ) increased over the length of each chan-
nel due to aerial inputs but did not vary significantly due to Cd input
(Table 3).
126
-------
In general, we found nutrient cycling provided little indication of the
Cd stresses in the system studied here. Future studies of nutrien£ cycling
as a measure of stress induced changes should include measure of NH, and K .
In many respects the artificial streams used in this study were similar
to spring-fed streams which have been extensively studied elsewhere (Odum,
1957a; Odum, 1957b). Water of very low mineral and organic carbon content
was introduced constantly over developing plant communities. Many of the
species that thrived in the artificial streams are also found in naturally
occurring artesian fed streams in the area. Because of this similarity,
system measurements made in these channels may be compared to measurements
made in natural spring-fed streams and results of Cd input in the artificial
streams may be directly extrapolated to some natural systems.
In measuring several natural springs in Florida, Odum (1975a), reported
gross production values ranging from 0.7 to 64 g 0_/m /d with an average
summer value of 17 g 0 /m /d. Values of gross primary production from the
artificial streams is within this range, though they were well below the
average value at the end of the Cd study. An obvious difference is that the
Florida springs had had many years to develop their communities while the
artificial streams had values higher than 3 g 0_/m /d after only one year of
colonization. The upward slope of all curves during the second summer of
water input indicates that successional development was in an early stage and
higher metabolism values might be expected in the streams after a longer
colonization period.
The autotrophic character of the experimental channels (gross production
exceeding respiration) is typical of springs because of a lack of input of
organic matter for heterotrophic metabolism (Odum, 1956). However, the re-
sult of this system autotrophy must be a combination of net export of organic
material and net accrual of biomass. As has been stated elsewhere in this
report, the artificial streams were performing both roles with export and
biomass accrual both significantly lowered by 5 to 10 pg Cd/£.
The increased metabolism values for treated streams (Fig. 59) after Cd
input had stopped are very frustrating because of their correlation with the
end of Cd input and the beginning of the summer growth season as observed in
the curves for the control channels. Whether this enhanced metabolism re-
flects a growing adaptation to Cd toxicity or a rebound from the burden of
the metal on metabolic processes cannot be established from these data alone.
In light of aufwuchs data that showed similar levels of biomass between
treatments at the time of Cd shutoff but reduced populations of the algal
producers it would appear that no significant adaptation to Cd input was
occurring. Different species were being selected for tolerance but their
combined effect could not increase productivity to control values in the time
range of this study. Yet, in spite of the rigid control by trifling amounts
of Cd metal, recovery was almost instantaneous when the toxin was removed at
the onset of the prime growing season.
127
-------
REFERENCES
1. Albright, L. J., J. W. Wentworth and E. M. Wilson. 1972. Technique
for measuring metallic salt effects upon the indigenous heterotrophic
microflora of a natural water. Wat. Res. 6:1589-1596.
2. American Public Health Association. 1976. Standard Methods for the
Examination o_f Water and Wastewater, 14th ed. American Public Health
Association, American Water Works Association, Water Pollution Control
Federation. Publications Office, American Public Health Association.
1193 pp.
3. Anon. 1971. Metals focus shifts to cadmium. Environ. Sci. Techno1
754-755.
4. Anon. 1975. Cadmium in the Environment: Toxicity, Economy, Control.
Environmental Directorate, Organization for Economic Co-operation and
development, Paris, 88 pp.
5. Anon. 1976. Quality Criteria for Water. U.S. Environmental Protection
Agency, Washington, pp. 27-32.
6. Anon. 1976. Methods for Chemical Analysis of Water and Wastes. Envi-
ronmental Monitoring and Support Laboratory Environmental Research
Center, Cincinnati, Ohio. 298 pp.
7. Ashida, J. 1965. Adaptation of fungi to metal toxicants - Ann. Rev.
Phytopath. 3:153-174.
8. Asworth, L. J. and J. Amin. 1964. A mechanism for mercury tolerance
to fungi. Phytopath. 54:1459-1463.
9. Auerbach, S. I., D. E. Reichle and E. G. Struxness. 1976. Environmen-
tal Sciences Division Annual Progress Report, September 30, 1975, Oak
Ridge National Laboratory.
10. Awarun, A. A. 1973. Tolerance of Escherichia coli to cadmium. J. En-
viron. Qual. 2:353-355.
11. Barlocher, F. and B. Kendrick. 1973. Fungi in the diet of Gammarus
pseudolimnaeus (Amphipoda). Oikos 24:295-300.
12. Barlocher, F. and B. Kendrick. 1974. Dynamics of the fungal population
on leaves in a stream. J. Ecol. 62:761-792.
128
-------
13. Barr, A. J., J. H. Goodnight, J. P. Sail and J. T. Helwig. 1976. A
User's Guide to SAS-76. SAS Institute Incorporated, Raleigh, North
Carolina, 329 pp.
14. Bergquist, B. and E. C. Bove. 1976. Cadmium: Quantitative methodology
and study of the effect upon the locomotor rate of Tetrahymena pyritor-
mis. Acta Prot 15:471-483.
15. Bertine, K. K. and E. D. Goldberg. 1972. Trace Elements in clams,
mussels and shrimp. Limnology and Oceanography. 17:877-884.
16. Bingham, F. T., A. L. Page, R. J. Mahler, and T. J. Ganje. 1976. Yield
and Cadmium Accumulation of Forage Species in Relation to Cadmium Con-
tent of Sludge-amended Soil 1976. J. Environ. Qual. 5:57-60.
17. Bowman, K. 0., H. Hutcheson, E. P. Odum, and L. R. Shenton. 1971. Com-
ments on the distribution of indices of diversity, pp. 315-366 In:
Statistical Ecology Volume 3; Many Species Population Ecosystems and
Systems Analysis. G. A. Patil, E. C. Pielon and W. E. Waters (Eds.)
The Pennsylvania State University Press, University Park and Lonndon.
462 pp.
18. Briese, L. A. and J. P. Giesy. 1975. Determination of lead and cadmium
associated with naturally occurring organics extracted from surface
waters, using flameless atomic absorption. Atomic Abso Newslet. 14:133-
136.
19. Brown, G. W., Jr. 1976. Effects of polluting substances on enzymes of
aquatic organisms. Fish Res. Bd. Can. 33:2018.
20. Brown, V. M., T. L. Shaw, and D. G. Shurben. 1974. Aspects of water
quality and the toxicity of copper to rainbow trout. Water Res. 8:797-
803.
21. Brinkhurst, R. 0. 1974. The Benthos of Lakes. St. Martin's Press.
New York. 190 pp.
22. Bryan, G. W. 1976. Some Aspects of Heavy Metal Tolerance in Aquatic
Organisms. Society for Experimental Biology Seminar Series. 2:7-34.
Cambridge University Press.
23. Buhler, D. R. 1972. Environmental contamination of toxic metals. In:
Heavy Metals ^n the Environment. Oregon State University Water Resources
Research Institute. 20 pp.
24. Bunzl, K. 1974. Kinetics of ion exchange in soil organic matter III.
Differential ion exchange reactions of Pb ions in humic acid and peat.
J. Soil Sci. 25:517-532.
25. Burbank, W. D. and D. M. Spoon. 1967. The use of sessile ciliates
collected in plastic petri dishes for the rapid assessment of water
Pollution. J. Protozool. 14:739-744.
129
-------
26. Burkitt, A., P. Lester and G. Nickless. 1972. Distribution of heavy
metals in the vicinity of an industrial complex. Nature (Lond.) 238:
327-328.
27. Burks, B. D. 1953. The Mayflies, or Ephemeroptera of Illinois. Ento-
mological Reprint Specialists. P. 0. Box 77224, Dockweiler Station, Los
Angeles, California. 216 pp.
28. Cairns, J. 1969. Rate of species diversity restoration following
stress in freshwater protozoan communities. Univ. KANSAS Sci. Bull.
XLVIII 6:209-224.
29. Cairns, J., Jr. 1977. Quantificatin of biological integrity, pp. 171-
187 in, The Integrity of Water. Edited by R. K. Ballentine and L. J.
Guarraia. Environmental Protection Agency. Office of Water and Hazar-
dous Materials. Washington, D. C., U. S. Government Printing Office
Stock Number 055-001-01068-1.
30. Cairns, J., G. R. Lanza and B. C. Parker. 1972. Pollution related
structural and functional changes in aquatic communities with emphasis
on freshwater algae and protozoa. Proc. Acad. Nat. Sci. Phila. 124:
79-127-
31. Cairns, J. and J. L. Plafkin. 1975. Response of protozoan communities
exposed to chlorine stress. Arch. Protistenk. Bd. 117,5:47-53.
32. Cearley, J. E., and R. L. Coleman. 1973. Cadmium Toxicity and Accumu-
lation in Southern Naiad. Bull. Environ. Contain. Toxicol. 9(2): 100-101.
33. Cearley, J. E. and R. L. Coleman. 1974. Cadmium toxicity and biocon-
centration in large mouth bass and bluegill. Bull. Environ. Contam.
Toxicol. 11:46-51,
34. Chadwick, M. H. 1976. Cadmium in the environment. Biologist 23:23-29.
35. Cheremisinoft, P. N. and Y. H. Habib. 1972. Cadmium, Chronium, Lead,
Mercury: A plenary account of water pollution. Part I - Occurrence,
toxicity and detection. Wat. Sewage Wks. 7:73-86.
36. Chopra, I. 1971. Decreased uptake of Cd by a resistant strain of Sta-
phylococcus aureus. J. Gen. Microbiol. 63:265-267.
37. Ciaccio, L. 1973. Water and Water Pollution Handbook V.4. Marcel
Decker, New York. 1945 pp.
38. Clubb, R. W., A. R. Gaufin, and J. L. Lords. 19753. Synergism between
dissolved oxygen and cadmium toxicity in five species of aquatic in-
sects. Environ. Res. 9:285-
39. Clubb, R. W., A. R. Gaufin, and J. L. Lords. 1975 . Acute cadmium
toxicity studies upon nine species of aquatic insects. Environ. Res.
9:332-
130
-------
40. Cochran, W. G. and G. M. Cox. 1971. Experimental Designs. John Wiley,
New York, 612 pp.
41. Cole, R. A. 1973. Stream community response to nutrient enrichment.
J. Water Pollut. Cont. Fed. 45:1874.
42. Cook, S. E. K. 1976. Quest for an index of community structure sensi-
tive to water pollution. Environ. Pollut. 11:269-287.
43. Cooper, D. C. 1973. Enhancement of net primary productivity by herbi-
vore grazing in aquatic laboratory microcosms. Limnol. Oceanogr. 18:
31-37.
44. Copeland, B. J. and W. R. Duffer. 1964. Use of a clear plastic dome
to measure gaseous diffusion rates in natural waters. Limnol. Oceanogr.
9:494-499.
45. Corbet, P. S. 1962. A Biology of Dragonflies. H. F. and G. Witherby,
Ltd. 247 pp.
46. Cummins, K. W. 1973. Trophic relations of aquatic insects. Annu. Rev.
Entonol. 18:183-
47. Cummins, K. W. 1974. Structure and function of stream ecosystems.
Biosci. 24:631-641.
48. Cummins, K. W. 1975. Macroinvertebrates. In: River Ecology. B. A.
Whitton (Ed.) University of California Press. Berkeley and Los Angeles
725 pp.
49. Doyle, J. J., R. T. Marshall and W. H. Pfander. 1975. Effects of cad-
mium on the growth and uptake of cadmium by microorganisms. App. Micro-
biol. 29:562-564.
50. Draggan, S. 1976. The Microcosm as a tool for estimation of environ-
mental transport of toxic materials. Internat. J. Environ. Studies.
10:65-70.
51. Dunstan, W. H. and H. L. Windom. 1975. The Influence of Environmental
Changes in Heavy Metal Concentrations on Spartina alterniflora. In:
Estuarine Research, Vol. II: Geology and Engineering. Academic Press,
Inc., New York.
52. Dunstan, W. H., H. L. Windom, and G. L. Mclntire. 1975. The Role of
Spartina alterniflora in the flow of lead, Cadmium and Copper through
the Salt-marsh Ecosystem. Mineral Cycling in Southeastern Ecosystems
F. G. Howell, J. B. Gentry and M. H. Smith (Eds.) U. S. Energy Research
and Development Administration, pp. 250-256.
53. Egglishaw, H. J. 1964. The distributional relationsip between the
bottom fanna and plant detritus in streams. • J. Anim. Ecol. 33:463.
131
-------
54. Egloff, D. A. and W. H. Brakel. 1973. Stream pollution and a simpli-
fied diversity index. J. Wat. Pollut. Control Fed. 45:2269-2275.
55. Eisler, R., G. E. Zaroogian and R. J. Hennekey. 1972. Cadmium uptake
by marine organisms. J. Fish Res. Bd. Can. 29, 1367-1369.
56. Elwood, J. W., S. G. Hildebrand, and J. J. Beauchamp. 1976. Contribu-
tion of gnt contents to the concentration and body burden of elements
in Tipula spp. from a spring-fed stream. J. Fish. Res. Bd. Can. 33:1930
57 Enk, M. D. and B. J. Maths. 1977. Distribution of cadmium and lead in
a stream ecosystem. Hydrobiologia. 52:153-
58. Fassett, D. W. 1974. Cadmium. In: Metallic Contaminants and Human
Health. H. K. Lee (Ed.) Academic Press, New York, pp 97-124.
i
59. Fleischer, M., A. Sarofin, D. W. Fassett, P. Hammond, H. T. Shacklette,
I. C. T. Nisbet and S. Epstein. 1974. Environmental impact of cadmium:
A review by the panel on hazardous trace substances. Env. Health. Per-
spect. pp. 253-323.
60. Flick, D. F., H. F. Kraybill and J. M. Dimitrott. 1971. Toxic effects
of cadmium: A review. Environ. Res. 4:71-85.
61. Forstner, V. and G. Muller. 1973. Heavy metal accumulation in river
sediments: A response to environmental pollution. Geoforum 14:53-61.
62. Flint, R. W. and C. R. Goldman. 1975. The effects of a benthic grazer
on the primary productivity of the littoral zone of Lake Tahoe. Limnol.
Oceanogr. 20:935-944.
63. Fowler, B. 1975. Heavy metals in the environment. Ill: Overview.
Environ. Health Pros. 12:125-126.
64. Fowler, S. W. and G. Benayoun. 1974. Experimental studies on cadmium
flux through marine biota. Radioactivity in the Sea. No. 44. Interna-
tional Atomic Energy Agency, Vienna.
65. Francis, C. W., and S. G. Rush. 1973. Factors Affecting Uptake and
Distribution of Cadmium in Plants. In: Trace Substances In: Environ-
mental Health VII. D.D. Hemphill (Ed.).
66. Fribery, L., M. Piscator and G. Nordberg. 1971. Cadmium in the Envi-
ronment . CRC Press, Cleveland, 166 pp.
67. Friberg, L. and T. Kjellstrom. 1975. Cadmium in the Environment - III,
A toxicological and epidemiological appraisal. EPA-6501/2-75-049, U.S.
Environmental Protection Agency Office of Research and Development,
Washington.
68. Fryer, G. 1957. The food of some freshwater cyclopoid copepods and its
ecological significance. J. Animal Ecology. 26:263-286.
132
-------
69. Fulkerson, W. 1975. Cadmium - the Dissipated Element - Revisited. Un-
published Report Oak Ridge National Laboratory. 29 pp.
70. Fullner, R. W. 1971. A comparison of macroinvertebrates collected by
basket and modified multiple-plate samplers. J. Wat.. Poll. Cont. Fed.
43:494-
71. Gaufin, A. R. and C. M. Tarzwell. 1952. Aquatic Invertebrates as Indi-
cators of Stream Pollution. Public Health Reports 67(1):57-
72. Gaufin, A. R. and C. M. Tarzwell. 1956. Aquatic macro-invertebrates
communities as indicators of pollution in Lytle Creek. Sewage Indust.
Wast. 28:906-
73. Gaufin, A. R. 1973. Use of Aquatic Invertebrates in the assessment of
water quality. In: Biological Methods for the Assessment of Water
Quality. J. Cairns and K. L. Dickson (Eds.) American Society for test-
ing and materials, Baltimore, Md. 256 pp.
74. Gerhards, U. and H. Weller. 1977. The uptake of mercury, cadmium and
nickel by Chlorella pyrenoidosa. Z. Pflanzenphysiol Bd. 82:292-300.
75. Giesy, J. P., L. A. Briese and G. J. Leversee. 1978. Metal binding
capacity of selected maine surface waters. Environ. Geol. (In press).
76. Giesy, J. P., G. J. Leversee, and D. R. Williams. 1977. Effects of
Naturally Occurring Aquatic Organic fractions on cadmium toxicity to
Simocephalys serrulatus (Daphnidae) and Gambusia affinis (Poeciliidae).
Wat. Res. 11:1013-1020.
77. Giesy, J. P. and J. G. Wiener. 1977. Frequency distributions of trace
metal concentrations in five freshwater fishes. Trans. Amer. Fish Soc.
196:393-403.
78. Gile, J. D. and J. W. Gillett.1977.Terrestrial microcosm chamber evalua-
tions of substitute chemicals. Unpublished manuscript, USEDA, Coruallis.
79. Gingell, S. M., R. Campbell and M. H. Martin. 1976. The effect of
zinc, lead and cadmium pollution on the leaf surface microflora. En-
viron. Pollut. 11:25-37.
80. Gommes, R., and H. Muntan. 1976. Cadmium levels in biota and abiota
from Lake Maggiore. Directorate general scientific and technical infor-
mation and information management of commission of the European commu-
nities, EUR 54lle, 31 pp.
81. Gray, J. S. and R. J. Ventilla. 1973. Growth rate of Sediment-living
marine protozoa as a toxicity indicator for heavy metals. Amer. Biol.
118-121.
82. Goodman, D. 1975. The theory of diversity - stability relationships
in ecology. Quant. Rev. Biol. 50:237-266.
133
-------
83. Haghiri, F. 1973. Cadmium Uptake by Plants. J. Environ. Qual. 2(1):
93-96.
84. Hamelink, J. L. 1976. Current Bioconcentration test methods and the-
ory. In: Aquatic toxicology and hazard evaluation F. L. Mayer and
J. L. Hamelink (Eds.). ASTM Philadelphia, Pa.149-161 pp.
85. Harding, J. P. C. and B. A. Whitton. 1978. Zinc, Cadmium and Lead in
water, sediments and submerged plants of the Derwent Reservoir, Northern
England. Wat. Res. 12:307-316.
86. Hardisty, M. W., R. J. Huggins, S. Kartar and M. Sainsbury. 1974. Eco-
logical implications of heavy metal in fish from the Severy Estuary.
Mar. Poll. Bull. 5:12-15.
87. Hargrave, B. T. 1970. The utilization of benthic microflora by Hya-
lella azteca (Amphipoda). J. Anim. Ecol. 39:427-437.
88. Hargrave, B. T. 1970. The effect of a deposit-feeding amphipod on the
metabolism of benthic microflora. Limnol. Oceanogr. 15:21-30.
89. Harman, W. N. 1969. The effect of changing pH on the Unionidae. Nau-
tilus 83:69-70.
90. Harman, W. N. 1970. New distribution records and ecological notes on
central New York Unionacea. Amer. Nidi. Natur. 84:46-58.
91. Hart, B. A. and P. W. Cook. 1975. The effect of Cadmium on Freshwater
Phytoplankton. Water Resources Research Center, University of Vermont.
16 pp.
92. Hartung, R. 1973. Biological effects of heavy metal pollutants in
water. Adv. Exp. Med. Biol. 40:161-172.
93. Harve, G. N., B. Underdal and C. Christiansen. 1973. The content of
lead and some other heavy- elements in different fish species from a
Fjord in western Norway. In: Proc. Int. Symp. Environ, Health. Asp.
Lead. Amsterdam pp. 99-111.
94. Hem, J. D. 1972. Chemistry and occurrence of cadmium and zinc in sur-
face water and ground water. Water Reso. Res. 8:661-679.
95. Hester, F. E., and J. S. Dendy. 1962. A multiple-plate sampler for
aquatic macroinvertebrates. Trans. Amer. Fish. Soc. 91:420-
96. Hiatt, V. and J. E. Hugg. 1975. The Environmental impact of cadmium:
An overview. Int. J. Environ. Studies. 7:277-285.
97. Hill, M. 0. 1973. Diversity and evenness: a unifying notation and its
consequences. Ecol. 54:427.
134
-------
98. Hiyama, Y. and S. Makoto. 1964. On the concentration factors of radio-
active Cs, Sr, Cd, Zn and Ce in marine organisms. Rec. Oceanog. Works
Japan. 7:41-77.
99. Rolling, C. S. 1973. Resislience and stability of ecological systems.
Ann. Rev. Ecol. Syst. 4:1-24.
100. Hucabee, J. W. and B. G. Blaylock. 1974. Microcosm studies on the
transfer of Hg, Cd and Se from terrestrial to aquatic ecosystems. Eighth
Annual Conference on Trace Substances in Environmental Health. Univer-
sity of Missouri Columbia.
101. Hughes, B. D. 1978. The influence of factors other than pollution on
the value of Shannon's Diversity Index for benthic macro-invertebrates
in streams. Wat. Res. 12:359-364.
102. Hunter, W. R. 1964. Physiological Aspects of Ecology in Nonmarine
Molluscs, pp. 83-126. In; Physiology of Mollusca Vol. I. Edited by
K. M. Wilbur and C. M. Yonge. pp. 471.
103. Kurd, L. E., M. V. Mellinger, L. L. Wolf and S. J. McNaughton. 1971.
Stability and diversity at three trophic levels in terrestrial ecosy-
stems. Sci. 173:1134-1136.
104. Hurlbert, S. H. 1971. The nonconcept of species diversity: a critique
and alternative parameters. Ecol. 52:577-
105. Hynes, H. B. N. 1957. The use of invertebrates as indicators of river
pollution. Proceedings of the Linnean Society of London, Symposium of
Water Pollution 165-167 pp.
106. Hynes, H. B. N. 1970. The Ecology of Running Waters. Univ. Toronto
Press, Toronto. 555 pp.
107. Hynes, H. B. N. 1970. The ecology of stream insects. Ann. Rev. Ent.
15:25-
108. Hynes, H. B. N. 1971. The Biology of Polluted Waters. University of
Toronto Press. Toronto and Buffalo. 202 pp.
109. Iversen, T. M. 1973. Decomposition of autumn-shed beech leaves in a
springbrook and its significance for the fauna. Arch. Hydrobiol. 72:
305-312.
110. Jinks, S. M. and M. Eisenbud. 1972. Concentration factors in the en-
vironment. Radiation Data Rep. l3;243-247.
111. John, M. K., C. J. Van Laerhoven and H. H. Chuah. 1972. Factors Affect-
ing Plant Uptake and Phytotoxicity fo Cadmium Added to Soils. Environ.
Sci. and Tech. 6(12):1005-1009.
135
-------
112. John, M. K., and C. J. Van Laerhoven. 1976. Differential Effects of
Cadmium on Lettuce Varieties. Environ. Pollut. 10:163-173.
113. Jonasson, P. M. 1955. The efficiency of sieving techniques for samp-
ling freshwater bottom fauna. Oikos. 6:183.
114. Jones, J. R. E. 1940. A study of the zinc polluted River Ystwyth in
north Cardiganshire, Wales. Ann. App. Biol. 27:368.
115. Jones, J. R. E. 1941. The fauna of the River Dover, West Sales. J.
Anim. Ecol. 10:12-
116. Jones, J. R. E. 1958. An ecological study of the River Towy. J. Anim.
Ecol. 20:68-
117. Jordan, M. J. and M. P. Lechevalier. 1975. Effects of zinc-smelter
emissions on forest soil microflora. Can. J. Microbiol. 21:1855-1865.
118. Kania, H. J., R. L. Knight, and R. J. Beyers. 1976. Fate and Biologi-
cal Effects of Mercury Introduced into Artificial Streams. EPA-600/3-
76-060. U.S. Environmental Protection Agency, Athens, Ga. 129 pp.
119. Kaushik, N. K. and H. B. N. Hynes. 1971. The fate of the dead leaves
that fall into streams. Arch. Hydrobiol. 68:465-515. Petersen, R. C.
and K. W. Cummins. (1974). Leaf processing in a woodland stream.
Freshwat. Biol. 4:343-368.
120. Karbe, L., N. Antonacopoulos, and C. Schnier. 1975. The influence of
water quality on accumulation of heavy metals in aquatic organisms.
Verh. Internat. Verein. Limnol. 19:2094-
121. Katagiri, K. J. 1975. The effects of cadmium on Anacystis nidulans.
Ms. Thesis, University of Vermont, 53 pp.
122. Kerfoot, W. B. and S. A. Jacobs 1976 Cadmium accrual in combined waste-
water treatment aquaculture system. Environ. Sci. Techno1. 10:662-667.
123. Kinkade, M. L. and H+JE. Erdman. 1975. The influence of hardness com-
ponents (Ca and Mg in water on the uptake and concentration of cad-
mium in a simulated freshwater ecosystem. Environ. Res. 10:308-313.
124. Kirk, R. E. 1968. Experimental Design: Procedures for the Behavioral
Sciences. Wadsworth, Belmont, Calif. pp.
125. Klass, E., D. W. Rowe, and E. J. Massaro. 1974. The effect of cadmium
on population growth of the green algae Scenedesmus quadricauda Bull.
Environ. Cont. Toxicol. 12:442-445.
126. Kraner, G. A. and J. H. Martin. 1973. Seasonal variations of cadmium,
copper, manganese, lead, and zinc in water and phytoplankton in Monterey
Bay, California. Limnol. Oceanogr. 18:597-
136
-------
127. Kneip, T.+J. and H+JE. Eredman. 1975. The influence of hardness compo-
nents (Ca and Mg ) in water in the uptake and concentration of cad-
mium in a simulated freshwater ecosystem. Environ. Res. 10:308-313.
128. Kneip, T. J. and G. J. Laver. 19.73. Trace metal concentration factors
in aquatic ecosystems. Prog. Anal. Chem. 5:43-62.
129. Kneip, T. J., G. Re and T. Hernandez. 1974. Cadmium in an aquatic eco-
system: Distribution and effects. In: Trace Elements in Environmental
Health VIII. University of Missouri, Columbia, pp. 173-177.
130. Koeppe, D. E. 1977. The Uptake, Distribution, and Effect of Cadmium
and Lead in Plants. Sci. Total Environ. 7:197-206.
131. Kumada, H., S. Kimura, M. Vokote and Y. Matida. 1973. Acute and chro-
nic toxicity, uptake and retention of cadmium in freshwater organisms.
Bull. Freshwater Fish Res. Lab. Tokyo. 22:157-165.
132. Lagerwerff, J. V. and A. W. Spect. 1970. Contamination of roadside
soil and vegetation with cadmium, nickel, lead and zinc. Environ. Sci.
Technol. 4:583-586.
133. Lee, C. R., T. C. Sturgis, M. C. Landin. 1976. A Hydroponic Study of
Heavy Metal Uptake by Selected Marsh Plant Species. Report D-76-5.
Dept. of Army Corps of Engineers, Vicksburg, Miss., 47 pp.
134. Leigh, E. G., Jr. 1965. On the relation between the productivity,
Biomass, Diversity and stability of a community. Proc. Nat. Acad. Sci.
Zoology, 53:777-783.
135. Levins, R. 1966. The strategy of model building in population biology.
Amer. Sci. 54:421-431.
136. Lichtenstein, E. P., T. T. Liang and T. W. Fuhremann. 1978. A compart-
mentalized microcosm for studying the fate of chemicals in the environ-
ment. J. Agricul. and Food, (in press).
137. Lightheart, B., H. Bond and B. G. Volk. 1978. The use of soil/litter
microcosms with and without added pollutants to study certain components
of the decomposer community. Unpublished manuscript, U.S. E.P.A.,
Corvallis, Oregon.
138. Maki, A. W. and H. E. Johnson. 1977a. The influence of Larval Lampri-
cide (TFM: 3-trifluormethyl-4-nitrophenol) on Growth and Production of
two species of aquatic macrophytes, Elodea canadensis (Michx.) and Plan-
chon and Myriophyllum spicatym L. Bull. Environ. Contam. Toxicol. 17:57-
139. Maki, A. W. and H. E. Johnson. 1977b. Evaluation of a toxicant on the
metabolism of model stream communities. J. Fish Res. Bd. Can. 33:2740-
2746.
137
-------
140. Margalef, R. 1961. Communication of structure in planktonic popula-
tions. Limnol. Oceanogr. 6:124-128.
141. Marshall, J. S. 1976. Cong term effects of cadmium in Lake Michigan
water on average numbers and biomass in laboratory Daphnia populations
Annual Report, Argonne National Laboratory pp. 27-31.
142. Mason, W. T., P. A. Lewis, and P. L. Hudson. 1975. The influence of
sieve mesh size selectivity on benthic invertebrate indices of eutro-
phication. Verh. Internat. Verein. Limnol. 19:1550-
143. McKee, J. E. and H. W. Wolf. 1963. Water quality criteria. Calif. St.
Res. Cent. Bd., Publ. No. 3-A. 548 pp.
144. McKellar, H. N. 1975. Metabolism and Models of estuarine bay ecosystems
affected by a coastal power plant. Ph.D. Dissertation. University of
Florida, 269 pp.
145. Merlini, M., F. Argentesi, A. Brazzelli, B. Orcgioni and G. Pozzi. 1971.
The effects of sublethal amounts of cadmium and mercury on the metabo-
lism of zinc-65 by freshwater fish. Proc. Inter. Symp. Radio. Ecology
Applied to the Protection of Man and Hi's Environ. Rome, EUR 4800 dfie.
1327-1344 pp.
146. Miettinen, J. K. 1975. The accumulation and excretions of heavy metals
in organisms. In; Ecological Toxicity Research. Effects of Heavy Metal
and Organohalogen Compounds. A. D. Mclntyre and C. F. Mills (Eds.)>
Plenum Press, New York.
147. Mills, W. L. 1976. Water quality bioassay using selected protozoa.
J. Environ. Sci. Health 11:491-500.
148. Morrison, M. G. and R. Steele. 1977. Effects of environmental factors
on radiocadmium uptake by four species of marine bivalves. Marine
Biology. 40:303-
149. Motohashi, K. and T. Tsuchida. 1974. Uptake of cadmium by pure cul-
tured diatom, Skeletonema costatum. Bull. Plankton. Soc. Japan 21:55-59.
150. Mundie, J. H. 1971. Sampling benthos and substrate materials, down to
50 microns in size, in shallow streams. J. Fish. Res. Bd. Can. 28:849-
151. Murry, C. N. and S. Menke. 1974. Influence of soluble sewage material
on adsorption and desorption behavior of cadmium, cobalt, silver and
zinc in sediment - freshwater, sediment seawater systems. J. Oceanogr.
Soc. Japan. 30:216-221.
152. Namminga, H. and J. Wilhm. 1977. Heavy metals in water sediments and
chironomids. J. Water Pollut. Cont. Fed. 49:1725-
138
-------
153. Nash, R. G. and M. L. Beall, Jr. 1977. A microagroecosystem to monitor
the environmental fate of pesticides. Unpublished manuscript pesticide
degradation lab. Beltsville, Md.
154. Nehring, R. B. 1976. Aquatic insects as biological monitors of heavy
metal pollution. Bulletin of Environmental Contamination and Toxicology.
15;147-
155. Odum, E. P. 1962. Relationships between structure and function in eco-
systems. Jap. J. Ecol. 12:108-118.
156. Odum, E. P. 1969. The strategy of Ecosystem Development. Sci. 164:
262-270.
157. Odum, E. P. 1977. The emergence of ecology as a new integrative disci-
pline. Sci. 195:1289-1293.
158. Odum, H. T. 1956. Primary production in flowing waters. Limnol. Ocean-
ogr. 1:102-116.
159. Odum, H. T. 1957a. Primary production measurements in eleven Florida
springs and a marine turtle-grass community. Limnol. Oceanogr. 2:85-97.
160. Odum, H. T. 1957b. Trophic structure and productivity of Silver
Springs, Florida Ecol. Monographs 27:55-112.
161. Odum, H. T. and C. M. Hoskin 1957. Metabolism of a laboratory stream
microcosm. Inst. Marine Sci. Univ Texas 4:115-133.
162. Oliver, D. R. 1971. Life history of the chironomidae. Ann. Rev. Ent.
16:211.
163. O'Neill, R. V., W. F. Harris, B. S. Ausmys and D. E. Reichel. 1975. A
theoretical basis for ecosystem analysis with particular reference to
element cycling. In: Mineral cycling .in Southeastern Ecosystems. F. G.
Howell, J. B. Gentry and M. H. Smith (Eds.). U.S. Energy Research and
Development Administration, pp. 28-40.
164. Page, A. L., F. T. Bingham, and C. Nelson. 1972. Cadmium Absorption
and Growth and Various Plant Species as Influenced by Solution Cadmium
Concentration. J. Environ. Qual. 1:288-291.
165. Page, A. S. and T. F. Bingham. 1973. Cadmium residues in the environ-
ment. Residue Rev. 48:1-44.
166. Pagenkopf, G. K., R. C. Russo and R. V. Thorston. 1974. Effect of
complexation on toxicity of copper to fishes. J. Fish. Res. Bd. Can.
31;462-465.
167. Patrick, R. 1949. A proposed biological measure of stream conditions,
based on a survey of the Conestoga Basin, Lancaster County, Pa. Proc.
Acad. Nat. Sci. Phila. 101:277-341.
139
-------
168. Patrick, R. 1967. The effect of invasion rate, species pool and size
of an area on the structure of the diatom community. Proc. Nat. Acad.
Sci. 58:1335-1342.
169. Paus, P. E. 1971. The application of atomic absorption spectroscopy to
the analysis of natural waters. Atom. Abso. Newslet. 10:69-71.
170. Payer, H. D., K. H. Rankel, P, Schramel, E. Stengel, A. Bhuriratana, and
C. J. Boeder. 1976. Environmental influences on the accumulation of
lead, cadmium, mercury, antimony, arsenic, selenium, bromine and tin in
unicellular algae cultivated in Thailand and in Germany. Chemosphere
6:413-418.
171. Pennak, R. W. 1953. Freshwater Invertebrates of the United States.
Ronald Press, New York pp. 769.
172. Perry, H. M., G. S. Third and E. F. Perry. 1976. The biology of cad-
mium. Med. Clin. N. Amer. 60:759-769.
173. Petersen, R. C. and K. W. Cummins. 1974. Leaf processing in a woodland
stream. Freshwat. Biol. 4:343-368.
174. Petterson, 0. 1976. Heavy-Metal Ion Uptake by Plants from Nutrient
Solutions with Metal Ion, Plant Species and Growth Period Variations.
Plant and Soil 45:445-459.
175. Pickering, Q. H. and C. Henderson. 1966. The acute bioassay of some
heavy metals to different species of warmwater fishes. Air. Wat. Pollut.
Int. J. 10:453-463.
176. Pielou, E. C. 1966. The measurement of Diversity in Different Types of
Biological Collections. J. Theoret. Biol. 13:131-144.
177. Pielou, E. C. 1969. An Introduction to Mathematical Ecology. Wiley
and Sons, Inc. 286 pp.
178. Piscator, M. 1974. Epidemiological Aspects of Cadmium in the Environ-
ment. In: D. D. Hemphill (Ed.) Trace Substances in Environmental
Health-VII.
179. Pomeroy, L. R. 1975. Mineral cycling in marine ecosystems. In: Mine-
ral Cycling in Southeastern Ecosystems. F. G. Howell, J. B. Gentry and
M. H. Smith (Eds.). U.S. Energy Research and Development Administration
pp. 209-223.
180. Pringle, B. H., D. E. Hisseng, E. L. Katz and S. T. Mulawka. 1968.
Trace metal accumulation by estuarian mollusks. J. Sanitary Engineering
Division Proceedings of the American Society of Civil Engineers. June
455-475.
140
-------
181. Ramamoorthy, S. and D. J. Kushner. 1975. Binding of mercuric and other
heavy metal ions by microbial growth media. Microbial Ecol. 2:162-176.
182. Rattonetti, A. 1974. Determination of soluble cadmium, lead silver and
indium in rainwater and stream water with the use of flameless atomic
absorption. Anal. Chem. 46:739-742.
183. Ravera, 0., R. Gommers, and H. Muntan. 1973. Cadmium distribution in
aquatic environment and its effects on aquatic organisms. European
Colloquium Problems of the Contamination of Man and His Environment by
Mercury and Cadmium, Luxenbourg 3-5 July. 1973. C.E.C. EUR-5075,
1974:317-330.
184. Reddy, C. N. and W. H. Patrick, Jr. 1977. Effect of Redox Potential
and pH on the Uptake of Cadmium and Lead by Rice Plants. J. Environ.
Qual. 6:259-262.
185. Renfro, W. C., S. W. Fowler, M. Heyrand, and J. LaRosa. 1975. Relative
importance of food and water in long-term zinc biota. Journal of the
Fisheries Research Borad of Canada. 32:1339-
186. Riffaldi, R. and R. Levi-Minzi. 1975. Adsorption and disorption of Cd
on humic acid fraction of soils. Water Air Soil Pollut. 5:179-184.
187. Rodgers, J. H. and R. S. Harvey. 1976. The effect of current on peri-
phytic productivity as determined using carbon - 14 Wat. Reso. Bull.
12:1109-1118.
188. Rosenthall, H. and K. R. Sperling. 1974. Effects of cadmium on devel-
opment and survival of herring eggs. In; J. H. S. Blaxter (ed.). The
Early Life History of Fishes. Springer Venlag, New York. 765 pp.
189. Rowe, D. W. and E. J. Mssaro. 1974. Cadmium uptake and time dependent
alterations in tissue levels in the white catfish Ictalurus catus
(Pisces .-Ictaloridae). Bull. Environ. Contam. Toxicol. 11:244-249.
190. Ruhling, A. and G. Tyler. 1973. Heavy metal pollution and decomposi-
tion of spruce needle litter. Oikos 24:402-416.
191. Ruthven, J. A. and J. Cairns. 1970. Artificial microhabitat size and
the number of colonizing protozoan species. Trans. Amer. Microsc. Soc.
89;101-110.
•
192. Ruttner, F. 1963. Fundamentals of Limnology. Univ. Toronto Press,
Toronto. 295 pp.
193. Scanlon, J. W. 1975. Dangers to the Human Fetys from certain Heavy
metals in the environment. Rev. Environ. Health. 2:39-64.
194. Schroeder, H. A. 1974. The Poisons Around US, toxic metals in food,
air and water. Indiana University Press, Bloomington. 144 pp.
141
-------
195. Schwiraer, S. R. 1973. Trace metal levels in three subtidal inverte-
brates. Veliger 16:95-102.
196. Service, J. 1972. A User's Guide to the Statistical Analysis System
North Carolina State University, Raleigh, N.C.
197. Shephard, B. K. 1976. The aquatic chemistry of cadmium in a natural
and in a model aquatic system. M.S. thesis, Purdue University, 129 pp.
198. Simpson, E. H. 1949. Measurement of diversity. Nature (London) 163:
688.
199. Skinner, S. P., J. B. Gentry and J. P. Giesy. 1978. Cadmium dynamics
in terrestrial food webs of a coal ash basin. In: Environmental Che-
mistry and Cycling Processes. ERDA Symposium Series, CONF-760429.
(In press).
200. Sprague, J. B. 1969. Measurement of pollutant toxicity to fish. I.
Bioassay methods for acute toxicity. Wat. Res. 3:3793-3821.
201. Steel, R. G. D. and J. H. Torrie. 1960. Principles and Procedures of
Statistics with Special References to BjLological Sciences. McGraw-Hill,
New York. 481 pp.
202. Stiff, M. H. 1971. The chemical states of copper in polluted fresh-
water and a scheme of analysis to differentiate them. Water Res. 5:585-
589.
203. Strickland, J. D. H. and T. R. Parsons. 1972. A Practical Handbook of
Seawater Analysis. Fisheries Research Board of Canada, Ottawa, 310.
204. Sullivan, J. F., B. R. Murphy, G. J. Atchison and A. W. Mclntosh. 1978.
Time dependent cadmium uptake by fathead minnows (Pimephales promelas)
During field and laboratory exposure. Hydrobiologia. 57:65-68.
205. Taub, F. B. 1976. Demonstration of pollution effects in aquatic micro-
cosms. Intern. J. Environ. Studies. 10:23-33.
206. Thomsen, L. C. 1973. Aquatic Diptera part V. Ceratopogenidae. pp 57-
80 in, Aquatic Diptera; Eggs, Larvae, and Pupae of Aquatic Flies. O.A.
Johannsen. Entonological Reprint Specialists. Los Angeles, California.
207. Thorp, V. J. and P. S. Lake. 1973. Pollution of a Tasmanian river by
mine effluents. II. distribution of macroinvertebrates. Int. Revne.
ges. Hydrobiol. 58:885-
208. Thorp, V. J. and P. S. Lake. 1974. Toxicity bioassays of cadmium on
selected freshwater invertebrates and the interaction of cadmium and
zinc on the freshwater shrimp, Paratza tasmaniensis (Rick). Aust. Mari.
Freshwat. Res. 25:97-
142
-------
209. Thormann, D. 1975. Uber die wirkung von cadmium und Bleiauf die natpr-
liche heterotrophe Bakterienflora in Brackwasser des waser. A. Stuars.
Verpff. Inst. Meeresforsch. Bremerh. 15:237-267.
210. Turner, M. A. 1973. Effect of Cadmium Treatment on Cadmium and Zinc
Uptake by Selected Vegetable Species. J. Environ. Qual. 2:118-119.
211. Tynecka, Z. and W. Zylinska. 1974. Plasmid born resistance to some in-
organic ions in Staphylococcus aureus. Acta. Microbiol. Pol. Ser. A.
Microbiol. Gen. 6:83-92.
212. Usinger, R. L. 1971. Aquatic Hemiptera. pp 182-228 in, Aquatic Insects
of California, with keys to North American genera and California species.
Edited by R. L. Usinger. University of California Press, Berkeley, Los
Angeles, London.
213. Vollenweider, R. A. (ed.) 1969. A Mannual on Methods for Measuring
Primary Production in Aquatic Environments. IBP Handbook #12. London
213 pp.
214. Wagner, S. L. 1972. Arsenic and Cadmium in the Environment. In:
Heary Metals in the Environment, Oregon State University Water Re-
sources Research Institute.
215. Wallace, A., E. M. Romney, G. V. Alexander, S. M. Soufi, and P. M. Patel.
1977. Agron. J. 69(l):18-20.
216. Weber, C. I. 1973. Biological Field and Laboratory Methods for Measur-
ing the Quality of Surface Waters and Effluents. Environmental Monitor-
ing Series. Analytical Quality Control Laboratory. National Environ-
mental Research Center - Cincinnati. EPA-670/4-73-001. 176 pp.
217. Weber, W. J. and H. W. Poselt. 1974. Equilibrium models and precipita-
tion reactions for Cadmium (II). In: Aqueous Environmental Chemistry
of Metals. A. J. Rubin (Ed.) Ann Arbour Science, Ann Arbour, Michigan.
390 p.
218. Webster, J. R., J. B. Waide and B. C. Patten. 1975. Nutrient recycling
and the stability of ecosystems. In: Mineral Cycling in Southeastern
Ecosystems. F. G. Howell, J. B. Gentry and M. H. Smith (Eds.) U. S.
Energy Research and Development Administration, pp. 1-27.
219. Westlake, D. F. 1975. Macrophytes. In: Studies in Ecology V.Z B. A.
Whitton (Ed.) Blackwell Scientific Publications pp. 106-128.
220. Wetzel, R. G. 1975. Primary production. In: River Ecology, B. A.
Whitton (Ed.). Univ. Calif. Press, Los Angeles, pp. 230-247.
221. Wiener, J. G. and J. P. Giesy. 1978. Concentrations of Cd, Cu, Mn, Pb
and Zn in Resident and stocked fish in a highly organic, softwater pond.
J. Fish. Res. Bd. Can. (In press).
143
-------
222. Wier, C. F. and W. M. Walter. 1976. Toxicity of cadmium in the fresh-
water snail, Physa gyrina. Say. J. Environ. Onal. 5:359-362.
223. Wilhm, J. L. 1967. Comparison of some diversity indices applied to
populations of benthic macroinvertebrates in a stream receiving organic
wastes. Wat. Pol. Cont. Fed. 39:1673-1683.
224. Wilhm, J. and T. C. Dorris. 1968. Biological parameters for water qua-
lity criteria. Biosci. 18:477-481.
225. Wolverton, B. C. 1975. Water Hyacinths for Removal of Cadmium and Nic-
kel from Polluted Waters. NASA Technical Memorandum TM-X-72721 NASA
Nat. Space Tech. Lab., St. Louis, Mississippi 5 p.
226. Youngue, W. H. and J. Cairns. 1971. Colonization and succession of
freshwater protozoans in polyurethane foam suspended in a small pond in
North Carolina. Notulae Naturae No. 443, 13 pp.
144
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APPENDIX A
ANALYTICAL TECHNIQUES
The determination of trace levels of Cd in samples of water, biological
and geological materials requires a sensitive analytical technique (Paus,
1971). The literature is surfeited with articles devoted to Cd analysis in
environmental samples. It is not our purpose to review this literature here.
Friberg et al. (1975) present a thorough discussion of Cd analytical tech-
niques. Atomic absorption techniques can be used to analyze for trace
amounts of Cd in biological material (Harve et al. , 1973) and is also well
suited for the analysis of low levels of Cd usually found in natural waters
(Hem, 1972; Ciaccio, 1973; Rattonetti, 1974; APHA, 1975; Briese and Giesy,
1975; Giesy et al., 1978) and gives rapid determinations of these levels with
high reproducibility involving few interferences. Because of the composition
of various biological and geological matrices, there is no standardized
methodology suitable for all materials. Because of the obvious importance of
accurate Cd analyses for this project, a considerable amount of personnel
time and effort has been expended to develop an appropriate set of techniques
to sample, prepare and analyze biological and geological materials.
Cadmium determinations were made, using either a Perkin-Elmer Model 306
or Instrumentation Laboratories Model 351 atomic absorption spectrophoto-
meter. Both instruments are equipped with deuterium continuum background
correction systems and graphite atomizers for flameless operation. Flame
determinations were made using an air-acetylene fuel rich flame with the IL
instrument. Flameless determinations were made in normal and interrupted
modes using argon as a purge gas with the Perkin-Elmer instrument. We have
relied exclusively on flameless techniques for preliminary analyses because
of the low background levels present in the organisms introduced to the
channels. Later in the project, greater use was made of the more rapid flame
determinations as Cd levels increased.
Based on actual analytical results, the sensitivity for the determina-
tion of Cd by flame AA in our laboratory is 0.025 pg/ml where sensitivity is
defined as that concentration which gives an absorbance reading of 0.0044 (1%
A). Our detection limit in the flame mode is 0.005 (Jg/ml where detection
limit is defined as that concentration which gives a signal greater than 2S
above the background noise signal. Both the sensitivity and detection limi£
vary somewhat in time and with sample matrix.
The sensitivity for flameless atomic absorption is much more variable
than that for flame determinations. Since both interrupted or continuous
purge gas modes may be used, two sets of sensitivity and detection limits can
be calculated for the Perkin-Elmer instrument. Using a continuous flow of
145
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argon purge gas, 2.0 pg of Cd produces an absorbance of 0.0044 units. In the
interrupted mode, 1.0 pg Cd produces the same value. The interrupted mode of
operation is much more variable than continuous and is generally not requir-
ed. The amount of sample injected into the graphite rod atomizer varies but
is usually 10 or 20 pi. Using a 10 pi sample and the continuous purge mode,
the sensitivity is 2 X 10 mg/1 in solution. The detection limit varies so
much with matrix that it is impossible to give a general value.
The sensitivity and precision of Cd determinations by flameless AA have
been optimized for both plant and animal matrices. Drying, charring and
atomizing times and temperatures were systematically varied until a program
was found which maximized matrix destruction and minimized Cd loss due to
volatilization. Based on the results of this preliminary work, a drying of
slightly less than 100 C was used for all matrices. A 10 sec drying time
was used for 10 pi samples and a 40 sec period for 20 pi samples. Best
analytical results were attained when a pyrolysis program of 250-350 C
applied for 10-15 sec, regardless of sample size or matrix. The optimum
atomization conditions were found to be a temperature of 2000 C applied for
4-5 sec.
All determinations were corrected for. reagent blanks and compared
against commercially prepared certified standards. Matrix interferences were
evaluated in each material analyzed for Cd by the use of internal standards.
Background matrix interferences were also checked by determining the absor-
bance at an adjacent non-absorbing analytical wavelength adjacent to the
primary analytical line of 228.8 nm. There is no absorption due to Cd at
226.2 nm but this is near enough to the analytical line so that broad spec-
trum background absorbances can be determined.
Standard addition curves had the same slope as curves constructed from
standards in distilled water, indicating the selected charring and atomiza-
tion time and temperature regime removed all background interferences for
flameless Cd analysis in all matrices. Absorbances determined at the non-
absorbing wavelength of 226.2 nm resulted in absorbances of between 0.000 and
0.002, also indicating that background matrix interferences were absent in
all matrices.
Sample preparation and analytical procedures were tested by determining
Cd in bovine liver (BL) and standard orchard leaves (SOL) supplied by the
National Bureau of Standards (NBS). These matrices are analagous to other
animal and plant matrices and allow the evaluation of preparatory and analy-
tical techniques. Cadmium concentrations in BL and SOL are below the detec-
tion limits of our flame AA techniques. Using flameless methods, however, we
measured mean Cd concentrations of 0.31 pg/g dry weight in the BL (NBS certi-
fied value is 0.27 + .03 pg/g) and 0.13 pg/g dry weight in the SOL (NBS cer-
tified value is 0.11 + .02 pg/g).
During the processing of the NBS standards, a number of sources of Cd
contamination were discovered. Carry-over in glass and polyethylene reagent
and sample bottles is a problem at low Cd levels. Therefore, bottles that
have been used for high standards cannot be used to hold lower concentration
146
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Cd standards. No losses from samples or standards occurred during a 24 hr
period, but longer periods may cause sorption losses in low Cd standards.
Because of these losses, standards are made up from concentrated standards
daily. No losses from acidified samples occur over time.
Because of the ubiquity of Cd contamination and the low Cd concentra-
tions we worked with, special care was taken to restrict glassware contami-
nation. The use of marbles in flasks during the digestion process was found
to cause contamination and watch glasses were satisfactorily substituted.
Used glassware was immediately rinsed in tap water to remove residual
sample or standards and placed in a 1% bath of Contrad (American Hospital
Supply Co., McGaw, 111.) for 24 hr. This wash was followed by several dis-
tilled water rinses, a 24 hr. soak in distilled 1% HC1 and a minimum of 5
deionized water rinses.
The disposable plastic tips used with Eppendorf pipettes to introduce
samples and standards into graphite furnace atomizers vary in Cd contamina-
tion from lot to lot, and may introduce considerable error to low level sam-
ples. This contamination is reduced by rinsing the plastic tips several
times in 10% HNO before use. Even with rinsing, there may be occasional
anomalous Cd readings that can be attributed to the plastic tips. The sub-
stitution of an automatic Teflon delivery system eliminated this source of
contamination.
Reagents have also been found to require special selection and treat-
ment. Hydrochloric acid (HC1) and sulfuric acid (H SO,) are not used in the
digestion procedure because chlorides and sulfates are poor matrices for
atomic absorption analysis. Perchloric acid (HC10,) is not used because
perchloric acid solutions cannot be introduced into graphite rod atomizers
for flameless AA determinations. Redistilled reagent grade HNO was found to
be free of Cd contamination and satisfactory for use in the digestion pro-
cedure. Reagent grade H~0 may be somewhat contaminated with Cd, and correc-
tions must be made from aata obtained from reagent blanks.
147
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APPENDIX B
Plants and animals collected from channels during Cd study.
Phylum Chlorophyta
Sub-Phylum Chlorophyceae
Order Volvocales
Family Chlamydomonadaceae
Chlamydomonas spp.
Order Chlorococcales
Family Chlorococcaceae
Characium sp.
Chlorococcum humicola (Naegeli) Rabenhorst
Family Oocystaceae
Eremosphaera viridia DeBary
Family Scendesmaceae
Coelastrum sp.
Scenedesmus acutiformis Schroeder
Scenedesmus sp.
Order Ulotrichales
Family Ulotrichaceae
Hormidium subtile (Koetzing) Heering
Geminella turfosa (Skuja) Ramanathan
Family Microsporaceae
Microspora pachyderma (Wille) Lagerheim
Order Chaetophorales
Family Chaetophoraceae
Microthamnion strictissimum Rabenhorst
Stigeoclonium elongatum (Hassall) Kuetzing
Order Oedongoniales
Family Oedogoniaceae
Oedogonium sp.
Order Zygnematales
Family Zygnemataceae
Mougeotia spp.
Family Desmidiaceae
Cosmarium asphaerosphorum Nordstedt
C. laeve var. septentrionale Wille
C. pseudoconnatum var. ornatum Allorge
C. viride var. minor West
Euastrum sp.
Spaerozosma excavata Ralfs
Spondylosium planum West and West
Staurastrum alternans Brebisson
148
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Phylum Euglenophyta
Order Euglenales
Family Euglenaceae
Euglena mutabilis Schmitz
Phylum Chrysophyta
Sub-Phylum Chrysophyceae
Order Chromulinales
Family Chromulinaceae
Chromulina pseudonebulosa Pascher
Sub-Phylum Bacillariophyceae
Order Pennales
Family Naviculaceae
Navicula notha Wallace
Pinnularia sp.
Family Fragilariaceae
Synedra sp.
Family Epithemiaceae
Rhopalodia sp.
Phylum Cyanophyta
Order Chroococcales
Family Chroococcaceae
Merismopedia punctata Meyer
Order Oscillatoriales
Family Oscillatoriaceae
Oscillatoria geminata Meneghini
Order Nostocales
Family Nostocaceae
Anabaena sp.
Family Scytonemataceae
Microchaete sp.
Family Rivulariaceae
Calothrix parietina (Naegeli) Thuret
Phylum Pyrrhopyta
Class Dinophyceae
Family Glenodiniaceae
Glenodinium sp.
Phylum Spermatophyta
Family Callitrichaceae
Callitriche heterophylla
Family Juncaceae
Juncus acuminatus
Juncus diffusissimus
Family Lentibulariaceae
Utricularia biflora
Family Poaceae
Agrostis hyemalis
Family Polygonaceae
Polyganum hydropiperoides
149
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Family Scrophulariacease
Gratiola virginiana
Family Typhaceae
Typha latifolia
Collection Method
plate sample bottom sample
Phylum Arthropoda
Class Insecta
Order Ephemeroptera
Family Baetidae
Callibaetis sp. XX
Baetis sp. X
Family Caenidae
Caenis sp. X X
Family Leptophlebiidae
Paraliptophebia sp. X
Order Odonata
Suborder Anisoptera
Family Libellulidae
Pantala hymenaea X X
Erythrodiplax minusula X X
Pachydiplax longipennis X
Erythemis sp. X
Celithemis fasciata X
Suborder Zygoptera
Family Coenagrionidae
Argia sp. X X
Ischnura sp. X X
Order Hemiptera
Family Mesoveliidae
Mesovelia sp. X
Family Hebridae
Merragta sp. X
Family Gerridae
Gerris sp. X X
Family Veliidae
Microvelia sp. X X
Family Navcoridae
Pelocoris femoratus X
Family Nepidae
Ranatra sp. X
Family Notonectidae
Notonecta indica X X
Burnoa seimitra X
Family Corixidae
Hesperocorixa sp. X X
150
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Collection Method
plate sample bottom sample
Order Trichoptera
Family Hydroptilidae
Oxyethira sp.
Family Psychomyiidae
Polycentropus sp.
Order Lepdioptera
Family Pyralidae
Pyrausta sp.
Nymphula sp.
Order Coleoptera
Family Gyrinidae
Gyrinus sp.
Family Noteridae
Hydrocanthus iricolor
Family Haliplidae
Haliplus sp.
Family Dytiscidae
Bidessus sp.
Hydroporus sp.
Laccophilus sp.
Family Hydrophilidac
Berosus sp.
Enochrus sp.
Tropisternus sp.
Family Elmidae
Stenelmis sp.
Family Dryopidae
Helichus sp.
Order Diptera
Family Tipulidae
Helius
Limonia sp.
*Family Chironomidae
Subfamily Chironominae
Chironomus sp.
Cladotanytarsus sp.
Cryptrochironomus sp.
Polypedilum sp.
Rheotanytarsus sp.
Tanytarsus sp.
Subfamily Orthocladiinae
Cardiocladius sp.
Cricotopus sp.
Corynoneura sp.
Thienemanniella sp.
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
151
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Subfamily Pelopiinae
Ablabesmyia ornata
A. peleensis
Family Ceratopogonidae
Bezzia or Prubezzia sp.
Dasyhelea sp.
Family Tabanidae
Chrysopy sp.
Tabanus sp.
Phylum Plathelminthes
Class Turbellaria
Phylum Annelida
Class Hirudinae
Class Oligochaeta
Order Prosopora
Family Lumbriculidae
Order Pleisiopora
Family Naididae
Pristina sp.
Phylum Mollusca
Class Gastropoda
Subclass Pulmonata
Family Physidae
Physa sp.
Collection Method
plate sample bottom sample
X X
X
X
X
X
X
X
*only Identified from limited Number of Samples.
152
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Phylum Protozoa
Subphylum Plasmodroma
Class Mastigophora
Subclass Phytomastigina
Order Chrysomonadina
Suborder Eucrysomonadina
Family Chromulinidae
Chromulina sp.
Oikomonas sp.
Crysamveba sp.
Mallomonas sp.
Family Ochromonadidae
Ochromonas sp.
Suborder Rhizochrysidina
Rhizochrysis sp.
Order Cryptomonadida
Suborder Eucryptomonadina
Family Cryptomonadidae
Cryptomonas sp.
Cyathomonas truncata
Order Phytomonadida
Family Chlamydomonadidiae
Chlamydomonas sp.
Order Eyglemoidida
Family Euglenidae
Euglena spp.
Family Astasiidae
Astasia sp.
Family Anisonemidae
Anisonema spp.
Peranema sp.
Subclass Zoomatigia
Order Rhizomastigida
Family Mastigamvebidae
Mastigamoeba sp.
Class Sarcodena
Subclass Rhyopoda
Order Proteomyxida
Family Vampyrellidae
Nuclearia sp.
Vampyrella sp.
Hyalodiscus sp.
Reticulomyxa sp.
Order Amoebida
Family Amoebidae
Amoeba proteus
Amoeba discoides
Amoeba dubia
Amoeba spp.
153
-------
Vahlkampfia sp.
Hartmannella sp.
Order Testacida
Family Arcellidae
Arcella vulgaris
Family Difflugiidae
Difflugia urcevlata
Difflugia corona
Difflugia globosa
Difflugia lobostoma
Centropyxis sp.
Family Euglyphidae
Euglypha sp.
Subclass Actinopoda
Order Heliozoida
Family Actinophryidae
Actinophrys sol
Actinosphoerium sp.
Subphylum Ciliophora
Class Ciliata
Subclass Holotricha
Order Cyranostomatida
Family Holophryidae
Holophrya sp.
Lacrymaria sp.
Family Colepidae
Coleps sp.
Family Tracheliidae
Dileptus spp.
Family Loxodidae
Loxodes sp.
Family Chlamydodontidae
Chilodonella sp.
Order Trichostomatida
Family Colpodidae
Colpoda sp.
Order Hymenostomatida
Family Tetrahymenidae
Colpidium sp.
Family Parameciidae
Paramecium bursaria
Paramecium caudatum
Paramecium aurelia
Paramecium sp.
Subclass Spirotricha
Order Heterotrichida
Family Spirostomatidae
Spirostomum spp.
Blepharisma sp.
Family Stentoridae
Stentor sp.
154
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Order Oligotrichida
Family Halteriidae
Halteria sp.
Order Hypotrichida
Family Oxytrichidae
Uroleptus sp.
Urostyla sp.
Family Euplotidae
Euplotes spp.
Subclass Peritricha
Order Peritrichida
Vorticella sp.
155
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APPENDIX C
Published information supported in part by Interagency agreement EX-76C-09-
0819 between the U.S. Environmental protection agency and U.S. Department of
Energy.
1) Giesy, J. P., G. J. Leversee and D. R. Williams. 1977. Effects of
naturally occurring aquatic organic fractions on cadmium toxicity to
Simocephalus serrulatus (Daphnidae) and Gambusia affinis (Poeciliidae).
Water Res. 11:1013-1020.
2) Williams, D. R. and J. P. Giesy. 1978. Relative importance of food and
water sources to cadmium uptake by Gambusia affinis (Poeciliidae). En-
viron. Res. 16:326-332.
3) Giesy, J. P. 1978. Cadmium inhibition of leaf decomposition in an
aquatic microcosm. Chemosphere 6:467-475.
4) Thorp, J. H., J. P. Giesy and S. A. Wineriter. 1978. Effects of chro-
nic cadmium exposure on crayfish survival, growth, and tolerance to
elevated temperatures. Arch. Environ. Cont. Toxicol. (Submitted).
156
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TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)
1. REPORT NO.
EPA-600/3-79-039
3. RECIPIENT'S ACCESSION* NO.
4. TITLE AND SUBTITLE
Fate and Biological Effects of Cadmium Introduced
into Channel Microcosms
5. REPORT DATE
April 1979 issuing date
6. PERFORMING ORGANIZATION CODE
V.AUTHORIS) j. P> Giesy, Jr., H. J. Kania, J. W. Bowling,
R. L. Knight, S. Mashburn, and S. Clarkin
8. PERFORMING ORGANIZATION REPORT NO.
9. PERFORMING ORGANIZATION NAME AND ADDRESS
Savannah River Ecology Laboratory
University of Georgia
Aiken, South Carolina 29801
10. PROGRAM ELEMENT NO.
1HE775
11. CONTRACT/GRANT NO.
IAG-D6-0369-1
12. SPONSORING AGENCY NAME AND ADDRESS
Environmental Research Laboratory—Athens, GA
Office of Research and Development
U.S. Environmental Protection Agency
Athens, Georgia 30605
13. TYPE OF REPORT AND PERIOD COVERED
Final, 5/75-5/78
14. SPONSORING AGENCY CODE
EPA/600/01
15. SUPPLEMENTARY NOTES
16. ABSTRACT
Cadmium was continuously input to aquatic microcosm channels resulting in
concentrations of 5 and 10 yg Cd/1. Cadmium accumulation into both biotic and
abiotic components was determined. Biological effects of cadmium were determined
by monitoring structural and functional properties of the entire system as well
as structural changes in populations and compared to control systems, which
received no cadmium.
Cadmium inputs and outputs equilibrated within approximately 20 days of
initial cadmium inputs. However, approximately 20% of the cadmium leaving the
channels was associated with particulates. Community components accumulated
cadmium proportional to cadmium exposure levels. Cadmium was rapidly eliminated
from all biotic components, with concentrations returning to levels similar to
those in control channels within a few weeks in the aufwuchs community to a few
months in macrophytes. Organic headpool sediments showed no significant decrease
in cadmium content six months after cessation of cadmium inputs, indicating that
the abiotic half time for contaminated environments is very long. Half times
for elimination from channel sediments were 72 and 38 days for 5 and 10 yg/1
inputs, respectively, after Cd inputs were terminated.
17.
KEY WORDS AND DOCUMENT ANALYSIS
DESCRIPTORS
b.lDENTIFIERS/OPEN ENDED TERMS
c. COSATi Field/Group
Cadmium
Biology
Water pollution
Biological effects
06C
A8G
68D
18. DISTRIBUTION STATEMENT
RELEASE TO PUBLIC
19. SECURITY CLASS (ThisReport)
UNCLASSIFIED
21. NO. OF PAGES
173
20. SECURITY CLASS (This page)
UNCLASSIFIED
22. PRICE
EPA Form 2220-1 (9-73)
157
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