United States
Yironmental
sncy
EP/V600/R-00/017
May 2000
"rww.epa.gov/ncerqa
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NATIONAL CENTER FOR ENVIRONMENTAL RESEARCH
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Table of Contents
Introduction vii
Section 1. Molecular Genetics Research
Genetic Diversity in California Native Fish Exposed to Pesticides 3
Susan Anderson, Andrew Whitehead, Bernie May, Mark Bagley,
Kathryn Kuivila, Barry Wilson, David Hinton
Molecular Detection of Anaerobic Bacteria as Indicator
Species for Fecal Pollution in Water 5
Katharine G. Field
Intraspecies Genetic Diversity Measures of Environmental Impacts 7
Dan E. Krone, G. Allen Burton, Keith Grasman
Demographic and Genetic Factors Affecting Population Viability of Lupinus perennis,
an Indicator Species of Oak Savanna 9
Helen J. Michaels, R.J. Mitchell
Are Genetic Diversity and Genetic Differentiation Bioindicators of Contaminant Impact on
Natural Populations? Fundulus heteroclitus as a Model Estuarine Species 11
Michael C. Newman, Margaret Mulvey, Michael A. Unger, Wolfgang K. Vogelbein
Multilevel Indicators of Ecosystem Integrity in Alpine Lakes of the Sierra Nevada 12
James T. Oris, Sheldon I. Guttman, A. John Bailer, John E. Reuter,
Glenn C. Miller
Ecosystem Monitoring via Genetic Diversity Surveys of Dandelions
Using VNTR Multilocus DNA Probes 14
Steven Rogstad, Brian Keane
Section 2. Multiscale and Landscape Indicators
Foliar Chemistry as an Indicator of Forest Ecosystem Status, Primary Production,
and Stream Water Chemistry 19
John Aber, Richard Hallett, Mary Martin, Marie-Louise Smith, Scott Ollinger, Scott Bailey
Environmental Factors That Influence Amphibian Community Structure
and Health as Indicators of Ecosystem Integrity 21
Val Beasley, Luanda Johnson, Carl Richards, Patrick Schoff, Rebecca Cole, Camilla Lleske,
Anna Schotthoefer, Cathy Johnson, Joseph Murphy, Marvin Piwoni
Modeling Ozone Flux to Forests Across an Ozone Concentration Gradient in the
Sierra Nevada Mountains, CA 23
Allen Goldstein, Jeanne Panek
Effects of Forest Fragmentation on Community Structure and Metapopulation
Dynamics of Amphibians 24
Lucinda Johnson, Catherine Johnson, Randall Boone, John Gross
Land Use and Geomorphic Indicators of Biotic Integrity in Piedmont Streams 25
DavidS. Leigh, B.J. Freeman, M.C. Freeman, E.A. Kramer, M.J. Paul
C.M. Pringle, A.D. Rosemond, R. Cifaldi, A. Roy, D. Walters
in
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Table of Contents (continued)
Developing Effective Ecological Indicators for Watershed Analysis 26
Duncan T. Patten, Wayne Minshall, Rick Lawrence, Andrew Marcus
Development and Evaluation of Multiscale Mechanistic indicators of Regional Landscapes 27
Carl Richards, Luanda B. Johnson, George E. Host
Development and Testing of a Multi-Resource Landscape-Scale Ecological Indicator: Forest
Fragmentation, Structure, and Distribution Relative to Topography 28
Steven W. Seagle, Philip A. Townsend
Ecological Indicators for Large River-Floodplain Landscapes 30
Monica G. Turner, E.H. Stanley, M.D. Dixon, R.E, Freeman, S.E. Gergel,
H. Kong, J.R. Miller, J. West
Characterization of the Ecological Integrity of Commercially Grazed Rangelands
Using Remote Sensing-Based Ecological Indicators 32
NeilE. West, Robert A. Washington-Allen, R. Douglas Ramsey
Section 3. Aquatic Indicators
Using Bioindicators To Develop a Calibrated Index of Regional Ecological
Integrity for Forested Headwater Ecosystems 37
Robert P. Brooks, Robert S. Mulvihill, Terry Master, Timothy J. O'Cornell
Stream Plethodontid Assemblage Response (SPAR) Index: Development,
Application, and Verification in the MAHA 38
Robert P. Brooks, Gian L. Rocco
Soil Enzyme Stability as an Ecological Indicator 39
Richard P. Dick
Integrative Indicators of Ecosystem Condition and Stress Across Multiple Trophic Levels
in the San Francisco Estuary 40
Richard C. Dugdale
Developing an Indicator for Nutrient Supply in Tropical and Temperate Estuaries,
Bays, and Coastal Waters Using the Tissue Nitrogen and
Phosphorus Content of Macroalgae 42
Peggy Fong
Microbial Indicators of Biological Integrity and Nutrient Stress for Aquatic Systems 44
James P. Grover, Thomas H. Chnanowski
Foraminifera as Ecosystem Indicators: Phase 1. A Marine Benthic Perturbation Index;
Phase 2. Bioassay Protocols 46
Pamela Hallock, Heidi Crevison, Thomas Dix, Helen Talge, Dana Williams
Biogeochemical Indicators of Watershed Integrity and Wetland Eutrophication 48
K. Ramesh Ready, W.F. DeBusk, A. Ogram, W. Graham, MM. Fisher,
E.F. Lowe, LW. Keenan
Development and Evaluation of Ecosystem Indicators for Urbanizing Midwestern Watersheds 49
Anne Spade, Jonathan M. Harbor, Midhat Hondzo, Bernard A. Engel
IV
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Table of Contents (continued)
An Integrative Aquatic System Indicator 51
Richard S. Stemberger, Eric K. Miller
Effects of Interacting Stressors in Agricultural Ecosystems: Mesocosm and Field
Evaluation of Multilevel Indicators of Wetland Responses 53
Stephen T. Threlkeld
Index of Authors 55
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Introduction
The mission of the United States Environmental Protection Agency (EPA) is to protect public health and
safeguard and improve the natural environment—the air, water, and land upon which life depends. Achievement of
this mission requires the application of sound science to the assessment of environmental problems and to the
evaluation of possible solutions. The National Center for Environmental Research (NCER) at EPA is committed to
providing the best products in high-priority areas of scientific research through significant support for long-term
research.
The Office of Research and Development's (ORD) Ecological Research Strategy identifies monitoring
research focused on biological indicator development at the molecular, community, and landscape levels of biological
organization as a high-priority research program. These indicators will be used for the monitoring of ecosystem
condition as well as for exposure evaluation. The development of new.characterization methods and the improvement
of multiscale monitoring designs also are high-priority research components. This research represents the extramural
component of ORD's Environmental Monitoring and Assessment Program (EMAP).
In 1997, NCERQA issued a Request for Applications (RFA) on Ecosystem Indicators. The purpose of this
solicitation was to support research leading to the development of techniques and indicators that characterize and
quantify the integrity and sustainability of ecosystems at local, regional, national, and/or global scales. In 1998 and
1999, the RFAs focused on molecular and landscape indicators, while maintaining their previous emphasis on integrity
and sustainability. In 2000, the program's focus switched to estuarine indicators, with the expectation of funding
groups of researchers on the East Coast, West Coast, Gulf of Mexico, and Great Lakes. Additionally, in FY 2000,
new research on the development of ecosystem classification systems and associated reference conditions will be
funded.
Annual progress reviews such as this allow investigators to interact with one another and discuss progress
and findings with EPA and other interested parties. If you have any questions regarding the program, please contact
the program manager, Barbara Levinson, by telephone at (202) 564-6911, or by e-mail at levinson.barbara@epa.gov.
vu
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Section 1.
Molecular Genetics Research
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Genetic Diversity in California
Native Fish Exposed to Pesticides
Susan Anderson ', Andrew Whitehead ', Bernie May 2, Mark Bagley 2, Kathryn Kuivtta 3, Barry Wilson *,
and David Hinton 2
'University of California at Davis, Bodega Marine Laboratory, Bodega Bay, CA; University of California at
Davis, Davis, CA; and3U.S. Geological Survey, Sacramento, CA
The overall goal of this project is to determine the
effect pesticide exposure may have on genetic variation
in a California native fish. Objective 1 is to examine
populations of Sacramento sucker (Catostomus occi-
dentalis) exposed to landscape-scale pesticide releases to
determine whether changes hi genetic diversity are as-
sociated with indicators of pesticide exposure and are
distinguishable from natural genetic variation. Molecu-
lar techniques to be utilized include Amplified Fragment
Length Polymorphisms (AFLP) and microsatellites.
Objective 2 is to compare the AFLP technique with the
Randomly Amplified Polymorphic DNA (RAPD) tech-
nique to determine which produces the most informative
and reproducible DNA fingerprints. Objective 3 is to
evaluate potential linkages between any observed
changes in genetic variation and fitness parameters in
individuals and populations.
Pesticide exposure patterns are being characterized
as one step in separating natural genetic variation from
pesticide-related variation. An existing pesticide-use
database is being used in combination with Geographic'
Information System (GIS) applications, as well as pest-
icide analysis in rivers and creeks, to quantify historical
patterns of exposure, and thereby, to select sampling
sites.
Fish are being sampled from sites inferred to be
impacted by agriculture and from upstream clean ref-
erence populations (see Figure 1). Fin clips are archived
from individual fish for genomic analysis. AFLP and
microsatellite analyses will be performed on archived fin
clips, and the resulting genomic patterns will be com-
pared among exposed and reference populations.
Biomarkers indicating.genetic damage (DNA strand
breaks) and enzyme inhibition (acetylcholinesterase
activity) in fish exposed to pesticides are being examined
at selected reference and exposed sites as an additional
method of discriminating pesticide-exposed populations
from reference populations. An assessment of the infor-
mativeness and reliability of RAPD and AFLP finger-
printing methodologies has been completed for pop-
ulation genetic analyses using fish with a well-established
pedigree. Numbers of segregating bands were compared
and tested for differences hi reproducibility.
Whether exposed and reference populations differ
in average sensitivity to pesticide exposure in laboratory
experiments will be tested using fitness parameters and
biomarker responses as endpoints. Correlations between
genotype and tolerance also will be tested.
A definitive study design was devised using data on
pesticide exposure and information regarding fish avail-
ability. Seven hundred fin-clip samples of Sacramento
sucker from multiple watersheds were archived. At this
time, field sampling for AFLP and microsatellite anal-
yses are approximately 75 percent complete. The DNA
strand break assay has been further developed this year.
The acetylcholinesterase enzyme inhibition assay also
has been optimized with an examination of variation hi
response among tissues and characterization of different
isoforms.
In comparing the relative merits of RAPD and
AFLP techniques, it was found that the number of segre-
gating bands is higher using RAPD than AFLP, but that
the reproducibility of RAPD bands is far lower. Only i
percent of the AFLP bands were judged to be irrepro-
ducible. In contrast, 16.1 percent of the RAPD bands
were irreproducible. At present, there is no reason to
believe that any criterion that can be developed will
increase the reliability of RAPD methodology to a level
that is comparable to AFLP analysis.
GIS analyses coupled with chemical analyses have
resulted in the selection of sampling locations that to-
gether represent a strong field experimental design. This
design allows for field replication, extensive character-
ization of background genetic variation, and examina-
tion of genetic differences over a very large geographic
scale.
This study is unique because genetic variation as-
sociated with contaminant exposure has not been eval-
uated in fish populations on a large geographic scale.
The RAPD technique is significant because it is used in
ongoing U.S. EPA Environmental Monitoring and As-
sessment Program (EMAP) studies; however, the newly
available AFLP technique permits examination of more
of the genome per unit effort and has been demonstrat-
ed to be more informative and reproducible.
Tasks for the year 2000 are directed entirely at
progress on Objective 1. The first task is completion of
approximately one-half of the AFLP and microsatellite
analyses. The second task is evaluation of DNA strand
breaks and acetylcholinesterase enzyme inhibition in lab-
oratory exposures to diazinon as well as in field caging
studies.
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Molecular Detection of Anaerobic
Bacteria as Indicator Species for Fecal Pollution in Water
Katharine G. Field
Department of Microbiology, Oregon State University, Corvallis, OR
Fecal pollution in water is a threat to ecosystem
integrity that also poses health risks to humans. Often
the problem is not mitigated because the source of the
pollution cannot be determined. For example, runoff
from nonpoint sources such as farm manure and failing
septic systems may be implicated. The standard indica-
tors for fecal pollution, fecal coliforms, do not distin-
uish between human and animal sources.
An indicator system based on molecular markers
from the anaerobic bacterial group Bacteroides-Prevo-
tella was developed as part of this research project. The
indicators are not grown, but instead, molecular markers
amplified from filtrate from water samples are mea-
sured. Using this method, human fecal pollution is dis-
tinguished from cow fecal pollution in estuarine and
river waters (see Figure 1).
The objectives are to: (1) develop additional
markers from other important polluting species; (2) iden-
tify the indicator strains or species; and (3) make the
system quantitative, allowing for estimation of the pro-
portions of different sources of fecal pollution.
Fecal genetic markers are amplified out of feces or
water samples by polymerase chain reaction, using fluo-
rescently tagged primers specific for 16S rDNA genes
from Bacteroides-Prevotella. The amplification pro-
ducts are digested with restriction enzymes and rapidly
screened on a DNA automated sequencer hi GeneScan™
mode, which estimates the proportions of each fragment
based on relative fluorescence. Fluorescence data are
converted into an electropherogram diagnostic for spe-
cific bacterial constituents. Work on this research pro-
ject using this approach resulted in the identification of
two Bacteroides-Prevotella diagnostic markers for hu-
man feces and three markers for cow feces.
Starting with fecal DNAs from other species, the
same approach will be used to test for the cross-occur-
rence of markers among different species, and to devel-
op markers from other species. 16S rDNA clone li-
braries will be prepared from Bacteroides-Prevotella
amplification products and the marker strains in the
clone libraries will be located and phylogenetically iden-
tified. The proportional contribution from cattle, hu-
man, and other sources will be measured by means of
real-time quantitative PCR with a Perkin-Elmer Taq-
Man.
The project is in its first year and significant pro-
gress has been made towards the first two objectives.
Fecal samples from 10 additional species have been test-
ed for the presence of the human and cow markers. Of
these, other ruminant species had some of the cow mark-
ers. None had human markers. Markers for additional
species currently are being developed. 16S rDNA clone
libraries were prepared from PCR products from Bact-
eroides-Prevotella-specific primers, and cow and human
marker sequences were identified by row-and-column
PCR followed by GeneScan™ analysis. When the clones
were sequenced, it was found that each marker repre-
sented a gene cluster rather than a single sequence. The
human marker clones were related toBacteroides unifor-
mis and B. vulgatus.. The cow markers comprised two
novel gene clusters within Bacteroides but were unre-
lated to any previously characterized microorganisms.
Source identification of fecal pollution is a long-
standing problem. Results from this research promise to
provide a solution that will be rapid and economical.
The next steps will be to continue developing Bacter-
oides-Prevotella markers from important polluting spe-
cies, such as swine and waterfowl, and to survey the fre-
quency of occurrence of the markers in individuals and
populations, to make the assay quantitative. In addition,
PCR primers specific to each individual marker are
being developed to make the assay easier and more af-
fordable for water-quality laboratories that do not have
access to a DNA automated sequencer.
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Figure 1. T-RPLP analysis of amplified Saaeroides-Prevotella 16S rRNA genes disdnguishes human from cow fecal pollution. Solid lines
represent combined human fecal DNAs; dotted lines represent combined cow fecal DNAs. The arrows indicate cow- or human-
specific genetic markers.
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Intraspecies Genetic Diversity
Measures of Environmental Impacts
Dan E. Krone, G. Allen Burton, and Keith Grasman
Department of Biological Sciences, Wright State University, Dayton, OH
This project's primary focus is an assessment of the
suitability of RAPD-PCR (randomly amplified polymor-
phic DNA-polymerase chain reaction) genetic diversity
measures as an alternative or adjunct for toxicity and
bioaccumulation tests of ecological risk in a wide variety
of resource types.
Initial efforts will assess the amenability of a set of
organisms from a range of trophic levels to RAPD-PCR
analysis and will identify organisms suitable for use at a
variety of sites. Subsequent work will use these organ-
isms to determine the impact of a broad range of envi-
ronmental stressors upon overall genetic diversity levels
within exposed populations.
The initial stages of this project have focused on
refining the RAPD-PCR DNA profiling technique and
determining which native organisms at a variety of local
aquatic and terrestrial sites would be most suitable -for
analyses. Profiles from six different aquatic species
(crayfish, snails, fathead minnows [Pimephales pro-
melas], mosquitofish [Gatnbusia affinis], damsel flies,
and Hyalella azteca) and five terrestrial species (pill
bugs, earth worms, spiders, garlic mustard, and violets)
have been generated and evaluated. All organisms stud-
ied to date have proved amenable to highly reproducible
RAPD-PCR typing.
The important roles played by the species consi-
dered to date, in addition to their nonmigratory natures,
normally high population densities, broad distribution,
and sexual reproduction make them very well-suited for
genetic diversity studies. The extremely high genetic
similarity seen in garlic mustard (probably due to its
recent introduction and invasive nature) both within and
between collection sites makes it unsuitable for sub-
sequent work. All other organisms display statistically
significant differences in genetic diversity between pop-
ulations collected at contaminated and reference sites
(see Figure 1 for an example hi crayfish). Populations
collected at reference sites generally have had the highest
levels of diversity, and these measures are correlated
with the diversity measures of other species collected at
the same sites as well as with other measures of
ecosystem health, including BBI (the Index of Biotic
Integrity) and ICI (the Invertebrate Community Index).
Initial surveys of aquatic systems and their closely
associated terrestrial sites have been made at Dick's
Creek hi Middletown, Ohio, and the Little Scioto River
in Marion, Ohio. Analyses of the DNA profiles from
the populations of organisms will be completed and used
to determine which will be most suitable for use at other
sites.
In Year 2 of the proposed study, genetic diversity
levels of the most informative sentinel species will be
considered hi surveys of a third aquatic system that has
been contaminated with a different set of pollutants.
Specifically, organisms will be collected at one reference
and three contaminated sites along the Clark Fork River
in southwest Montana.
This river transects a variety of agricultural areas
along its 120-mile reach. Mining, milling, and smelting
activities have occurred extensively throughout its
headwater and tributary streams, resulting in substantial
heavy metal contamination of the watershed. The
system is particularly interesting, due to its harboring of
a pollution sensitive and high-quality benthic macro-
invertebrate community despite extreme metal toxicity
levels.
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Genetic Similarity
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Figure 1. Index of Biotic Integrity (ffll) values are inversely correlated with RAPD-PCR based measures of genetic similarity. Average
pairwise similarity of each organism relative to all others collected at its site are plotted against the independently obtained EBI value
(Ohio EPA, personal communication) for that site (N=144, r=-0.770, p< <0.001) and a solid line shows the best fit linear
regression. Crayfish collected from the impacted and reference Ottawa River sites are displayed as triangles (a), those collected
from sites along the Little Scioto River are displayed as circles (o) and those collected from Elk Creek and its reference stream,
Dick's Creek, are shown as squares (D). Large Xs correspond to the mean pairwise genetic similarity of crayfish at each site (N=8,
r=-0.804, p<0.01) and a dashed line corresponds to the best fit linear regression for those points.
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Demographic and Genetic Factors Affecting Population
Viability of Lupinus perennis. an Indicator Species of Oak Savanna
Helen J. Michaels1 andRJ. Mitchell2
'Department of Biological Sciences, Bowling Green State University, Bowling Green, OH; Department of
Biology, University of Akron, Akron, OH
Plants in small and sparse populations often have
low reproductive success, indicating that fragmentation
of populations by human activities may prevent pop-
ulations from being self-sustaining. This research is in-
tended to elucidate whether and how population decline
develops for a model organism, Perennial Lupine (Lup-
inus perennis: Fabaceae). This plant species, an im-
portant indicator species for the imperiled Oak Openings
Savanna community of the Great Lakes ecosystem, not
only suffers from habitat loss and fragmentation, but also
is the only host plant for three endangered butterfly
species. Thus, understanding how demographic and
genetic factors contribute to decline of this species will
improve understanding of this community and recovery
efforts for imperiled butterfly species.
The causes of declines in plant reproductive suc-
cess hi small and sparse populations include decrease in
pollinator services, loss of genetic diversity, inbreeding
depression, and combinations of these factors. This re-
search seeks to determine the importance of each factor,
utilizing a blend of observational and experimental tech-
niques and merging precise data from novel DNA-based
indicators of genetic diversity with classical ecological
data on reproductive ecology.
The observational studies focus on how existing
variation in population size and density affect the factors
above. Experimental studies involve a reciprocal trans-
plant between large and small populations to confirm that
population size is the cause of the observed patterns.
These data will not only improve the understanding of
fundamental biological principles in small populations,
but also will be informative for designing Oak Savanna
management strategies.
Pollinator visitation increased significantly in dense
areas within populations, but did not differ significantly
among populations varying in size. Significant inbreed-
ing depression of seed set and offspring fitness was ex-
hibited hi a pilot experiment. A previously unnoticed
seed predator (Megalotomus quinquespinosus; Hemipt-
era; Alydidae) that mimics ant behavior and appearance
was detected. Feeding damage from this bug disrupts
Lupine seed dormancy, potentially affecting plant de-
mography and population health. Several DNA micro-
satellite markers with which to assess inbreeding rates
have been developed.
The pollinator visitation data indicates that eco-
logical factors may influence population reproductive
success on a fine scale, such that within-population den-
sity variation may be as or more important than pop-
ulation size itself (see Figure 1). Should these trends be
substantiated in subsequent studies, management activ-
ities may need to consider plant density as well as popu-
lation size to improve the status of L. perennis, the spe-
cies that depend upon it, and the overall health of the
Oak Savanna ecosystem.
The pilot study of inbreeding depression has helped
refine the methods and experimental design to be em-
ployed in the larger study during Year 2. The results
also suggest that inbreeding depression will be detect-
able. The occurrence of an ant-mimicking seed predator
appears to be unknown to most scientists and managers
interested in Lupine and the Karner Blue. Work to dis-
seminate this information is underway through conver-
sations and a manuscript in preparation.
For Year 2 of the project (2000-2001), the tasks to
be completed include: assessing genetic variation (iso-
lating DNAs, running gels), analyzing the mating system
(planting seeds, isolating DNAs, running gels), planting
phytometers in the field, continuing pollinator observa-
tions, inbreeding depression pollinations, planting in-
breeding depression seeds (fall), and analyzing and writ-
ing up research results. During Year 3 (2001-2002), it
is planned to assess phytometer success, complete the
mating system gels, assess inbreeding depression seed-
lings, and further analyze and write up the research
results.
More information can be found at the following
Web Sites: http://www.uakron.edu/biology/mitchell/
lupine.html and at http://www.bgsu.edu/departments/
biology/people/faculty/michaels/research. html.
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i
IO
Sparse Dense
Lupine Abundance Class
Figure 1. Inflorescence visitation rates by bumble bees to observation plots in 1999 (mean ± se). Lupine density significantly affected
visitation rate (F,^ = 9.6, P < 0.0027).
10
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Are Genetic Diversity and Genetic
Differentiation Bioindicators of Contaminant Impact
on Natural Populations? Fundulus heteroclitus as a Model Estuarine Species
Michael C. Newman, Margaret Mulvey, Michael A. Unger, and Wolfgang K. Vogelbein
Virginia Institute of Marine Science, The College of William & Mary, Gloucester Point, VA
Molecular genetic traits of the mummichog are
being evaluated as bioindicators of population-level ef-
fects of pollution. Mummichogs were sampled from
nine localities along the Elizabeth River in Virginia.
Sediment PAH concentrations span four orders of mag-
nitude among these sites. Localities vary also in sed-
iment concentrations of chlorinated hydrocarbons, tri-
butyl tin, and metals. The prevalence of proliferative
liver lesions was high in fish from one locality, the
Atlantic Wood (AW) site, with a PAH concentration of
371,212 ng/g of sediment.
Molecular genetic data currently are being used to
address several predictions regarding population re-
sponse to contaminants: (1) populations residing in con-
taminated habitats are genetically distinct from populat-
ions in neighboring clean sites; (2) populations at pol-
luted sites exhibit lower genetic diversity than populat-
ions taken from clean sites; and (3) the genetic structure
of mummichog in the Elizabeth River reflects the
mosaic of highly contaminated and clean habitat.
Genetic data consist of allozyme genotypes and
DNA sequences for a 450 base pair segment of the
d-loop of the mitochondria! genome. Preliminary anal-
yses of allozyme data do not indicate correlation with
level of contamination. Analyses suggest that overall
genetic variability did not differ with PAH concen-
tration. Highly polluted localities (AW, RS) were not
genetically depauperate relative to neighboring relatively
nonpolluted localities (SC, JC). Indeed, mummichog
from the heavily PAH-contaminated AW site show high
diversity in d-loop haplotypes.
Mummichogs from the AW site share the common
haplotype found at other locations along the Elizabeth
River as well as several highly divergent haplotypes (see
Figure 1). Genetic analyses suggest that mummichog
may move among localities more than previously report-
ed.
Hypotheses regarding contaminant tolerance and
individual and populations performance relative to gen-
etic variability and contaminants will be examined in the
next phase of the project. Additional survey work of
mummichog inhabiting contaminated and clean habitats
may be undertaken to evaluate the generality of these
results.
Base change =™
() number observed
Numbers designate haplotypes
© Inferred
Figure 1. Relationships among d-loop haplotypes and distribution among Elizabeth River sites.
11
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Multilevel Indicators of Ecosystem
Integrity in Alpine Lakes of the Sierra Nevada
James T. Oris ', Sheldon I. Guttman ', A. John Bailerl, John E. Renter2, and Glenn C. Miller3
'Miami University, Oxford, OH;2 University of California, Davis, CA; 3 University of Nevada-Reno, Reno, NV
The overall objective of the project is to develop
protocols for environmental assessments of alpine lakes
in the Sierra Nevada with a range of human impacts.
These assessments will be conducted over the range of
levels of biological organization (molecular to eco-
system), utilizing currently available assessment tech-
niques and with the addition of two new ecological in-
dicators.
The use of population genetics analysis as a re-
sponse indicator and the use of molecular biomarkers of
exposure to contaminants as a diagnostic indicator will
be tested for incorporation into monitoring and assess-
ment programs for surface waters. These indicators will
provide information concerning the status of population
diversity and stability as well as the exposure to non-
persistent, nonbioaccumulative contaminants. This in-
formation is missing from current monitoring and
assessment protocols.
Over the 3-year project period, standard environ-
mental assessments of a select group of alpine lakes with
a defined range of human impacts will be conducted.
There will be a total of 16 assessment sites (see Figure
1). Those selected for assessment include four minimal-
ly impacted areas (Castle Lake, Eagle Lake, Marlette
Lake, and Upper Angora Lake), eight areas with a range
of moderate impacts (Fallen Leaf Lake, Gold Lake,
Lake Tahoe at Sand Harbor, Prosser Reservoir,
Spaulding Reservoir, Stampede Reservoir, Topaz Lake,
and Twin Lakes), and four highly impacted areas (Boca
Reservoir, Dormer Lake, Lake Tahoe at Tahoe City, and
Lake Tahoe at South Lake Tahoe).
In addition to the standard assessment, population
genetic assessments will be conducted in fish and in-
vertebrates at these same sites. Allozyme electropbo-
retic analysis and Randomly Amplified Polymorphic
DNA (RAPD) analyses will be conducted on two organ-
isms common to the lakes of the region (fish: Lahontan
redside; invertebrate: Signal crayfish). Contaminant
exposure assessments also will be conducted in fish using
molecular biomarkers of exposure in the gills of fish.
Five markers indicative of exposure to a wide variety of
chemical contaminants (persistent and nonpersistent) and
that can account for interactions among complex mix-
tures of contaminants will be measured over time in the
assessment areas using caged rainbow trout.
These additional techniques then will be applied to
current assessment protocols. Because a group of lakes
with defined levels of human impacts will be examined,
the discriminatory ability of the assessment techniques
may be analyzed using the current protocols compared to
the protocols with the two new indicators added.
It is hypothesized that because current protocols do
not account for genetic diversity or nonpersistent con-
taminants, the addition of these new indicators will
greatly enhance the monitoring and assessment programs
for surface waters.
12
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| Castle Lake(1),
North, 180 miles
North
to
Sacramento
Assessment Sites:
1. Castle Lake
2. Marietta Lake
3. Eagle Lake
4. Upper Angora Lake
5. Gold Lake
6. Stampede Reservoir
7. Prosser Reservoir
9. Spaulding Reservoir
9. Topaz Lake
10. Twin Lakes
11. Fallen Leaf Lake
12. Sand Point, Lake Tahoe
13. Boca Reservoir
14. Donner Lake
15. Tahoe City Marina, Lake Tahoe
16. Ski Run Marina, Lake Tahoe
to/
Sacramento
Hwy 395
to
Los Angeles
Figure 1. Map of assessment site locations within the Sierra Nevada.
13
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Ecosystem Monitoring via Genetic Diversity
Surveys of Dandelions Using VNTR Multilocus DNA Probes
Steven Rogstad and Brian Keane
Department of Biological Sciences, University of Cincinnati, Cincinnati, OH
New methodologies in molecular genetics may
provide novel types of ecological indicators for mon-
itoring the integrity of natural ecosystems and the sus-
tainability of ecosystems that are affected by anthropo-
genic influences. One type of ideal ecological indicator
would be an organism that grows in a wide variety of
habitats that could easily be used to monitor for the
presence of mutagens or anthropogenic factors that alter
population genetics.
Dandelions ^Taraxacum offidnale: Weber; Aster-
aceae) are a model ecological indicator organism
because: (1) dandelions have an extremely wide ecolog-
ical amplitude, growing almost worldwide from sea-level
to alpine biomes, and from the tropics to north-temperate
habitats; (2) dandelions grow rapidly, and populations
can be easily manipulated and monitored; (3) it has been
documented that a variety of pollutants can be seques-
tered in dandelion tissues; (4) dandelion seeds are pro-
duced asexually, rendering the detection of mutations
easy; and (5) preliminary data have been generated using
variable-number-tandem-repeat (VNTR) DNA probes
that suggest that mutation rates and population genetic
diversity parameters can be analyzed successfully with
dandelions.
The specific hypothesis to be tested is that dande-
lions can be used as a sensitive ecological indicator
species via comparative analyses of VNTR genetic mark-
ers used to examine anthropogenic changes hi genetic
diversity of both mutation rates and population genetics,
at pollution-impacted versus nonimpacted sites.
Dandelions will be sampled from 12 sites: 6 rel-
atively pristine sites versus 6 sites with chronic, high
levels of contaminants. To determine whether rates of
mutation differ between these two types of sites, leaf
tissue and 8-10 seeds will be collected from each of 10
plants per site. Dandelion seeds are produced via aga-
mospermy, a nonsexual process in which all seed-
lings from a maternal plant should be genetically
identical to that maternal plant.
Preliminary results demonstrate that although die
previous statement is true, a low level of mutation (about
0.005% of bands transmitted) is detectable. By survey-
ing the parent-to-offspring transmission of tens of thou-
sands of VNTR markers, whether mutation rates differ
between pristine versus contaminated sites is being test-
ed (see Figure 1).
These markers also will be used to examine wheth-
er populations differ in genetic diversity. Initial results
demonstrate that very local populations harbor a large
amount of genetic diversity, aldiough certain clones may
be widely distributed. Soil and dandelion tissue have
been analyzed from more than 20 sites for metals, and
12 of these sites covering a range of contamination are
being selected to compare dandelion mutation rates and
population genetics.
It is expected that a determination will be made as
to whether VNTR markers in dandelions can be used as
sensitive indicators of anthropogenic changes in pop-
ulation genetic diversity due to either altered mutation
rates or stressor-induced selection. If mutation rates or
genetic diversity in dandelions are correlated with
pollution history, dandelions provide an easily utilized
biomonitor to survey the ecological integrity and sustain-
ability of a wide range of habitats across multiple spatial
scales throughout the world.
Besides completing the above projects, further
experiments growing dandelions hi controlled media
spiked with mixtures of common pollutants will be con-
ducted to investigate whether mutation rates increase
with increasing pollutant concentrations. Further, stud-
ies have been initiated to determine whether dandelions
from the most heavily polluted sites have higher fitness
when competing with dandelions from more pristine sites
when both are grown in polluted and nonpolluted soils.
14
-------
«. •«•*** *** *** '»'<* *.-» - ****
Jpnp: *Hi*» iW Y* * • *•'•"' " *' ' * • •''
t •,"
Figure 1. Maternal plaiu-io-oft'spring transmission of VNTR genetic markers in dandelion detected with the TTCCA ( = core sequence) PCR-
STR probe. Markers for two different maternal parents are shown (lanes A and B), with 10 clonal offspring of each maternal
parent also shown (5 offspring on each side of each parent). More than J10 VNTR markers are shown across both parents, ami
parents clearly differ in the markers transmitted to their clonal offspring. One new mutant marker is detectable in the eleventh lane
from the left (marked with a < ). If an autoradiograph is interpreted to have a transmission of approximate^ .^> markers lo 20
offspring (= MX) markers transmitted), with one novel marker, the detectable mutation rate is 1/601 = 0.00166 for ihai particular
gel-probe autoradiograph. Three micrograms of genomic ON A per lane were digested with a five fold excess of Taq\ restriction
endonuclease.
15
-------
Section 2.
Multiscale and Landscape Indicators
-------
Foliar Chemistry as an Indicator of Forest
Ecosystem Status. Primary Production, and Stream Water Chemistry
John Aber, Richard Hallett, Mary Martin, Marie-Louise Smith, Scott OUinger, and Scott Bailey
Complex Systems Research CenterfEOS, University of'New Hampshire, Durham, NH
Monitoring the biogeochemical status of forest and
stream ecosystems is a key component of assessing
environmental quality in the Northeastern United States.
Any monitoring system that requires spatially continuous
capabilities will need to use some form of remote sens-
ing. Because forest canopies are the only portion of the
system accessible to optical reflectance remote sensing
instruments, they offer the most likely target surface for
monitoring forest health in this spatial mode.
It was hypothesized that forest productivity, soil
mineralogy, and foliar chemistry at the whole-stand (not
individual tree) level are all tightly linked to the bio-
geochemical status of a forest ecosystem. It was further
hypothesized that the concentration of cations in forest
canopies will be measurable by high spectral resolution
remote sensing, as has been demonstrated for nitrogen
and lignin, and that watershed-level stream chemjstry,
reflecting soil mineralogy, also will be predictable from
watershed-level values of canopy chemistry derived by
remote sensing.
The study area is the White Mountain National
Forest (WMNF), a 300,000 ha area in northern New
Hampshire. At the intensive plot scale, the long-term
sampling program at the Bartlett Experimental Forest
and the Hubbard Brook Experimental Forest, New
Hampshire, will be used and augmented to examine and
attempt to predict interannual variations in foliar chem-
istry as well as woody and foliar production.
At the regional scale, canopy chemistry, soil min-
eralogy, and forest productivity will be measured at a
series of existing experimental and monitoring research
sites. Work on the spatially continuous monitoring scale
across the White Mountain region will include the devel-
opment of algorithms for predicting canopy cation con-
centrations using data from NASA's Airborne Visible-
Infrared Imaging Spectrometer (AVIRIS). At the
watershed scale, 50 streams will be sampled in the
WMNF covering a range hi estimated mineralogical
richness. A nested approach will be used to determine
the optimum scale at which stream water chemistry may
be predicted. In addition, high spatial resolution (3-4 m)
spectral data will be collected for sampled watersheds
via low altitude AVIRIS data acquisition.
Preliminary findings include: (1) strong relation-
ships between whole-stand level foliar canopy N, NPP,
and forest floor C:N ratios exist; (2) foliar Ca can be
mapped across the WMNF (300,000 ha) using remote
sensing technology (see Figure 1); and (3) a CIS model
has been developed for the WMNF that predicts element
content of glacial till—the predictions for till Ca have
been compared to plot level foliar Ca data, and a signifi-
cant relationship exists.
Remote sensing technology can be used to estimate
foliar chemistry (N, Ca) at a landscape scale. With this
information, forest productivity estimates can be made.
Soil mineralogical characteristics also can be modeled at
the landscape scale. Relationships between stream water
chemistry, mineralogy, foliar chemistry, and forest pro-
ductivity can be determined with this information. This
program then would establish the scientific basis for
developing a satellite- or aircraft-based remote sensing
program for monitoring forest health and stream water
quality.
The next steps are to: (1) continue the collection
and analysis of stream water from 50 streams across the
WMNF; (2) process and analyze new AVIRIS imagery
from a low altitude platform for selected water-
sheds—AVIRIS low altitude data (3-4 m resolution) for
the summer of 2000 has been requested through NASA's
Airborne Science Program; (3) collect data from new
plots across the WMNF to validate foliar and glacial till
element concentration maps; and (4) use soil and foliar
element coverages to develop relationships with mea-
sured stream water chemistry and forest productivity.
19
-------
a
a.
(0
O
>»
a
o
c
ro
O
"O
E
Q.
CO
{£.
10000
9000
8000 -
7000 -
6000 -
5000 -
4000 -
< 3000
R2 = 0.76
a<.001
Actual vs Predicted
1:1 Line
Regression Line
3000 4000 5000 6000 7000 8000 9000 10000
Measured Canopy Ca (ppm)
Figure 1. AVIRIS predicted plot level canopy Ca concentration plotted against measured canopy Ca concentrations.
20
-------
Environmental Factors That Influence Amphibian
Community Structure and Health as Indicators of Ecosystem Integrity
Val Beasley ', Lucinda Johnson 2, Carl Richards3, Patrick Schoff2, Rebecca Cole ", Camilla Lieske*,
Anna Schotthoefer', Cathy Johnson 2, Joseph Murphy ', and Marvin Piwonis
'College of Veterinary Medicine, University of Illinois, Urbana, 1L; 2Natural Resources Research Institute,
University of Minnesota, Duluth, MN; Minnesota Sea Grant, Duliah, MN; "USGS National Wildlife Health
Center, Madison, WI; Illinois Waste Management and Research Center, Champaign, IL
The overall goal of this research is to assess the
relative influence of landscape patterns, biotic interact-
ions, water quality, and contaminants on amphibian
health and community structure. The following will be
evaluated: (1) the relative influence of wetland- and
watershed-scale factors (i.e., landscape and ecological
data) on amphibian community structure and health; and
(2) whether amphibian community structure and health
are indicative of ecological integrity.
During 1998, anuran community structure, wetland
habitat, and landscape characteristics were quantified at
64 sites distributed throughout northern Illinois, south-
ern-to-central Wisconsin, and southern-to-central Min-
nesota (see Figure 1). Site-specific surveys also were
conducted to determine the incidence and types of anuran
malformations and deformations.
Landscape data were summarized within a 2 km
and 10 km radius/buffer surrounding each wetland.
Macroinvertebrates were identified to the family level,
and dominant vegetation was characterized for each wet-
land. Anuran data were summarized for 1998 and 1999,
individual species-habitat relationships were examined,
and site occupancy models were developed for several
frog species based on local and regional characteristics.
Logistic regression models were developed to predict the
occurrence of the wood frog, spring peeper, and gray
tree frog. Models incorporating data at several spatial
scales were better predictors of frog presence/absence
than those incorporating variables from only one spatial
scale. During 1999, 36 wetlands in central Minnesota
were analyzed for anuran and macroinvertebrate comm-
unity structure, wetland vegetation, and physical/chem-
ical attributes, including habitat structure, water, and
sediment quality. Parasitological studies were conducted
on tadpoles, metamorphs, adult frogs, and gastropods.
In separate studies, full necropsies were performed on
485 frogs. A comprehensive set of tissues was collect-
ed, fixed in buffered formalin, and processed for histo-
pathologic examination.
Malformations or deformations were identified in
13 of 559 frogs from 8 wetlands hi 1998, and in 7 of 544
frogs from 6 sites hi 1999. Among the malformations
were webbed skin between the femur and tibio-fibula,
skin lacking spots, and hemimely (partial absence) of the
hind limb. Parasites identified included: Echinostoma
spp., Rhabdias ranae, Haematolechus sp., Megalodiscus
sp., and Ribeiroia sp. Echinostoma spp. were found in
the kidneys of frogs from all sites at prevalences of
40-100 percent (mean 82%) and intensities of 8-321
echinostomes/frog (mean 90).
In pathological studies on 485 frogs, intersex was
identified in 6 percent of all frogs (from 11 sites). At
least one lesion was identified in 55 percent of frogs.
Condition scores from slightly poor to emaciated were
noted hi 8 percent of frogs from 12 sites. Parasitism
was noted in the body cavity, GI tract, kidney, liver,
lung, muscle, and skin. The most common abnormality
for body cavity, kidney, liver, and lung was inflam-
mation (52-84%); for GI tract it was no digesta (50%);
for musculoskeletal system it was trauma (52%); and for
spleen it was splenomegaly (73%). Diagnoses and sex
determinations will be confirmed histologically.
Examinations of metamorphs, identification of
parasites, histopathology studies, and chemical analyses
from the 1999 field season will be completed in the
coming months. Additional ecological, water quality,
contaminant, and anuran community structure and health
data will be collected hi 2000 before the assessment of
the potential use of amphibian community structure and
health as indicators of wetland conditions.
21
-------
i AMPHIBIAN COMMUNITY
STUDY AREAS
I
| Amphibian Community
> Study Sin Location*
j • Ptimliltn
EMtvtn Bn>u
-------
Modeling Ozone Flux to Forests Across an Ozone
Concentration Gradient in the Sierra Nevada Mountains. CA
Allen Goldstein and Jeanne Panek
Department of Environmental Science, Policy, and Management, University of California, Berkeley, CA
Tropospheric ozone is a pollutant that is responsi-
ble for forest damage worldwide. Extensive ozone mon-
itoring networks currently measure ozone concentrat-
ions throughout the United States and Europe; how-
ever, the physiologically relevant measure of ozone for
forest health is not concentration, but rather ozone flux,
the amount of ozone that actually enters the foliage.
A model is being developed to estimate ozone flux
from ozone concentration utilizing routinely measured
ozone and meteorology. It is hoped that this model will
be adapted for monitoring networks. The utility of 613C
as a proxy for stomatal conductance in the estimation of
ozone deposition also is being explored.
In 1997 and 1998, an initial study was conducted at
Blodgett Forest to investigate the influence of typical
Mediterranean summer drought on the uptake of ozone
and general physiological response of ponderosa pine.
Fluxes of ozone, C02, water, and energy were measured
by eddy covariance from May through October. Using
this eddy flux data, typically reported ozone exposure
metrics were compared against direct measurements of
ozone deposition to document that ozone metrics are
poor predictors of ozone uptake in California pine eco-
systems.
In May 1999, the field campaign was expand-
ed—sites were established at three additional locations
along an ozone injury gradient in the Sierra Nevada
Mountains, California. Site locations and characteristics
are presented in Table 1. These sites take advantage of
existing ozone monitoring (National Park Service and
California Air Resources Board Environmental Protect-
ion Agency) and an ongoing forest damage assessment
project. Year-round continuous measurements of eco-
system-scale ozone, carbon dioxide, water, and energy
fluxes started in May at Blodgett Forest. Measurements
of leaf-level physiology were made monthly at each site
from May through September and included diurnal net
photosynthesis, stomatal conductance and transpiration,
predawn and afternoon water potential, photosynthesis
response curves to light and carbon, and dark res-
piration. Foliage was collected for starch 813C analysis
every month. At the end of the season, foliage samples
were collected for cellulose 613C measurement
Another study at Blodgett Forest to gain further
insights into the limitations imposed by typical summer
drought on the uptake of carbon and ozone in the pon-
derosa pine ecosystem was completed. Two sites were
set up—one control and one watered—in a poaderosa
pine plantation. Carbon uptake in 1-year-old control fo-
liage was reduced compared to the watered treatment
during the 3 measurement days following treatment by
39 percent, 35 percent, and 30 percent, respectively, per
unit leaf area. Stomatal conductance was lower at the
control site, leading to a reduction in estimated ozone
deposition (ozone concentration times stomatal conduc-
tance) of 36 percent, 46 percent, and 41 percent of the
watered site, respectively. This experiment demon-
strated that site moisture is one of the most important
factors controlling ozone uptake in California forests.
Expected changes in climate will profoundly affect the
ozone uptake by California forest ecosystems.
The field campaign will continue during the grow-
ing seasons of 2000 and 2001. Models that estimate sto-
matal conductance and ozone flux into foliage will be
evaluated and compared for their appropriateness in the
Sierra Nevada ponderosa pine ecosystem. The selected
model will be developed further to apply to this project's
specific needs and compare modeled ozone deposition
with direct measurements of canopy-scale ozone depo-
sition at Blodgett Forest.
Table 1. Site locations and characteristics.
„.„ II i .• 1 ci Elevation 1 „,,, O/one"
Site Location Slope . . Oil' , . ,
II 1 r (m) | (|>pb)
Sequoia/King's Canyon National Park
Yosemite National Park
Blodgett Forest Research Station
Control Site
Watered Site
White Cloud
N36°33'55" W118°46'36"
N37°42143"W119°42'19"
N38°53'43" W120°37'58"
N38°53'43" W120°37'58"
N39°19'00" W120°50'45"
30%
10%
2%
1%
10%
1920
1220
1315
1315
1326
41.3
14.7
N/A
N/A
27.3
63"
41"
49", 57C
Same as Control
62'
'Ozone Injury Index (OH) is derived from a combination of the primary effects of ozone on pine (Arbaugh et al.,1998).
224 hour mean ozone concentration (in ppb) from June 1-October 31 were: (a) from Arbaugh et al.,1998, (b) from our measurements in 1998,
and (c) from our measurements in 1999.
23
-------
Effects of Forest Fragmentation on Community
Structure and Metapopulation Dynamics of Amphibians
Lucinda Johnson ', Catherine Johnson ', Randall Boone 2, and John Gross 2
'Natural Resources Research Institute, University of Minnesota, Duluth, MN; 2Natural Resource Ecology
Laboratory, Colorado State University, Fort Collins, CO
The goals of this research project are to quantify
the effects of forest fragmentation on amphibian com-
munity structure and metapopulation dynamics in vernal
pools, and to develop appropriate ecosystem indicators
for vernal pool ecosystems at multiple spatial scales.
The objectives of this study are to: (1) quantify the
manner and extent to which forest fragmentation influ-
ences amphibian community structure of vernal pool
ecosystems; (2) assess the extent to which landscape- and
local-scale features reflect fundamental structural prop-
erties of vernal pool habitats and their biotic com-
munities and, conversely, the extent to which indices of
biotic integrity (e.g., amphibian community structure)
reflect local and landscape properties; (3) develop pre-
dictive models that integrate landscape-scale factors with
pond-scale attributes to quantify key compositional-and
structural attributes of the amphibian community, and
derive ecosystem indicators at multiple spatial scales;
and (4) develop predictive models to quantify the extent
to which forest fragmentation influences the metapop-
ulation dynamics of woodland amphibians and predict
the consequences of landscape change on these metapop-
ulations.
Recent evidence of declines and an increased rate
of malformations in amphibian populations have prompt-
ed much interest and research into the potential anthro-
pogenic stressors associated with these occurrences.
Results of some of this research indicate that fragment-
ation may have a significant effect on the metapop-
ulation dynamics of amphibian communities, which
could result in declines and regional extinctions of pop-
ulations. Forest fragmentation results in changes to for-
est landscapes and habitats that may have both direct and
indirect effects on local and regional' amphibian com-
munities (e.g., direct habitat loss, disruption of dispersal
corridors, altered habitat structure, and microclimate
changes).
It was hypothesized that forest fragmentation can
be directly related to changes in landscape structure and
local habitats (e.g., vernal pools) that have a measure-
able effect on the integrity of amphibian communities.
Landscape, local habitat, and biotic community variables
will be quantified to examine the effects of forest frag-
mentation on pond-breeding woodland amphibians at
three spatial scales: the landscape scale, the local or
"pond-scape" scale (including ponds and surrounding
terrestrial habitat), and the aquatic pond habitat. Rela-
tionships among these hierarchically nested scales will be
quantified using an integrated series of empirical models.
This approach of identifying relationships across scales
and integrating them into a modeling framework will
allow the development of multiscale ecological indicators
of the effect of forest fragmentation on vernal pool eco-
systems and on regional amphibian communities.
It also was hypothesized that indices of community
diversity and structure and amphibian species traits can
be used to assess effects of forest fragmentation on ver-
nal pool systems and regional amphibian communities.
Metapopulation models will be developed to assess the
effects of fragmentation on local and regional popula-
tions, and to elucidate the mechanisms by which forest
fragmentation may influence the composition and persis-
tence of vernal pool communities. This is a new project,
and to date there are no results to report.
24
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Land Use and Geomorphic
Indicators of Biotic Integrity in Piedmont Streams
David S. Leigh, B.J. Freeman, M.C. Freeman, E.A. Kramer, M.J. Paul, C.M. Pringle, A.D. Rosemond,
R. Cifaldi, A. Roy, and D. Walters
Geography Department, University of Georgia, Athens, GA
The overall goal and objective of this project is to
define the predictive capabilities of scale-variable at-
tributes of land cover (CIS-based) and geomorphology as
risk assessment indicators of biotic integrity of stream
ecosystems on the southern Piedmont.
The watershed under investigation is the upper
Etowah River Basin north of Atlanta, GA. Given vari-
ous aspects of historical landscape change, this research
is investigating the following three ancillary questions:
(1) Do physical stressors and the corresponding eco-
logical response vary as a function of land cover in the
watershed? (2) Is this relationship consistent within
watersheds of vastly different sizes? (3) Do antecedent
land cover conditions (>50 years ago) influence the
physical stressor and ecological response relationship?
The approach to answer these questions involves
two main projects over a 3-year period. The first pro-
ject involves a comprehensive field survey of the geo-
morphic condition, habitat, water quality, and biolog-
ical integrity in 30 streams draining watersheds of three
distinct size classes of about 15, 50, and 100 km2.
These watersheds have variable land cover of 50-100
percent forest, based on the 1993 multiresolution land
characteristic (MRLC) databases. The stream reaches
are surveyed for a length of at least 15 stream widths.
Regression analysis is used to develop predictive models
of biotic conditions from geomorphic indicators, habitat
assessment, water quality, and land cover characteristics.
The second project involves a detailed analysis of geo-
morphic conditions, stream habitat, and biotic integrity
of five streams (out of the original 30) that appear as
unexplained residuals in the predictive models, which
will be compared to five other streams that fit the
models. This will involve an expansion of 10 surveyed
reaches to a length of at least 1 km, and further assess-
ment of the reach-scale versus watershed-scale controls
on biotic integrity.
During January 2000, raw data from all aspects of
the field survey were finalized and are being analyzed
along with the land cover data. The preliminary findings
suggest that reach-scale geomorphic and habitat assess-
ments are somewhat better indicators of biotic integrity
than the 1993 watershed-scale land cover characteristics.
For example, the average particle size of the stream bed
(in phi units) is highly correlated with the relative
abundance of pool species of fish, exhibiting a correla-
tion coefficient of 0.82 (see Figure 1). In contrast, the
best correlation between pool species and land cover is
found with percent forest, exhibiting a correlation co-
efficient of-0.48, and all other land cover classes are not
significantly correlated.
The significance of the preliminary findings may be
that objective reach-scale assessments of physical char-
acteristics are better indicators of biotic conditions than
watershed land cover. More analysis of linkages be-
tween the watershed-scale land cover and reach-scale
physical conditions are needed to explore causal relation-
ships and develop multiple regression models that may
provide the best indicators of biotic conditions, however.
The next steps involve continued analysis of the raw data
from the 30 sites and identification of sites for the
second phase of the analysis.
Upper Etowah Watersheds (10-150 km2)
LO-
TS
0.9-
0.8-
0.7-
±1
CO
CD 0.6-
J2 0.5
1^0.4-1
CD
'o
CD
Q.
52 0.2-1
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0.0
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•24 -26
•1413'21
•15
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-6
-3312
•20
-8
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bldr
cobble
gravel
-9
-! r
-7 -6 -5 -4 -3 -2
Mean Phi of Channel Bed
sand
Figure I. Relationship between mean Phi particle size and pool species (relative abundance) in streams of the upper Etowah River Watershed
(10-150 km-basins).
25
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Developing Effective Ecological
Indicators for Watershed Analysis
Duncan T. Patten ', Wayne Minshall2, Rick Lawrence 3, and Andrew Marcus3
'Yellowstone Ecosystem Studies, Bozeman, MT; 2Idaho State University, Pocmello, ID; ^Montana State University,
Bozeman, MT
This project is designed to develop improved
indicators and innovative techniques for assisting and
monitoring ecological integrity at the watershed level in
the Western United States. The specific objectives of
this study are to develop practical, scientifically valid
indicators that: (1) span multiple resource categories;
(2) are relatively scale independent; (3) address different
levels of biological organization; (4) can be rapidly and
cost-effectively monitored by remote sensing; and (5) are
sensitive to a broad range of anthropogenic and natural
environmental stressors.
The study is based, in part, on the hypothesis that
streams and riparian areas often reflect the ecological
integrity of the associated watersheds. Due to a "funnel
effect," these areas are the accumulation zones of en-
vironmental disturbances occurring in the watershed (see
Figure 1). For example, logging, overgrazing, and for-
est fires influence sediment erosion rates that directly
influence downstream stream and riparian conditions.
Monitoring of key indicators in these accumulation zones
may provide an efficient, cost-effective way to evaluate
and monitor the ecological integrity and sustainability of
the surrounding watershed.
The Upper Yellowstone River and its tributaries
will be used because the watersheds of these streams in-
corporate a broad range of environmental conditions
from relatively pristine in certain watersheds within
Yellowstone National Park, to highly disturbed by forest
fires and land uses such as mining, logging, and agri-
culture in other watersheds. Identification, assessment,
and validation of effective indicators will involve the in-
tegration of results from research at various scales, in-
cluding: (1) analysis of hyperspectral and traditional
multispectral imagery from both aerial and satellite plat-
forms; (2) field surveys of stream morphology and ripar-
ian habitat associated with remote sensing to assess in-
dicators; and (3) intensive site-specific stream sampling
of macroinvetebrate communities to validate the effec-
tiveness of these indicators. Use and evaluation of re-
mote sensing technologies is the primary research meth-
odology. All indicators chosen must be able to be mon-
itored by remote sensing.
Remote sensing techniques, which will enable se-
lection of indicators to be used for rapid, cost-effective
ecological monitoring on the regional and local scale,
also will be used to help identify key ecological indicat-
ors in both streams and riparian areas. These also will
be correlated with ecological indicators of disturbances
in the surrounding watersheds, resulting in a set of ef-
fective ecological indicators and the development of in-
novative techniques for efficient watershed-level analysis
at various scales in the Western United States.
As this project is hi its initial phase, the sequential
steps to be taken over the duration of this project in-
clude: (1) coarse watershed characterization for initial
site selection; (2) stream and riparian site selection;
(3) ground evaluation and selection of indicators at
stream and riparian sites; (4) fine watershed char-
acterization using remotely sensed data to determine
stressors on study sites; (5) collection of remotely sensed
data for study sites; (6) validation of watershed condition
using aquatic biota; and (7) analysis of riparian and
streams conditions as indicators of watershed condition,
and remote sensing as measurement of indicators.
, -
r
WATERSHED CONDITIONS
LAND USES
(Stressors)
T HYDROLOGY Surface & Ground Water
Channel and ^ I Hydraulics
Floodplain P*" j Fluvial Processes
Alterations y
CHANNEL Channel changes
GEOMORPHOLOGY
Indicators of Stream
"Health" Identified by
Sediment
Dynamics
Ground Measurements
RIPARIAN VEGETATIONS—
Indicators of Riparian
"Health" Identified by
Ground Measurements
VALIDATION OF WATERSHED
"HEALTH" (STREAM WATER
QUALITY — Aquatic Insects)
Figure 1. Conceptual model of interrelationships among watershed conditions, stream and riparian indicators, and watershed "health"
indicators (aquatic insects), showing drivers and stressors. Remote sensing will be used for watershed characterization and
evaluated as a technological tool for measuring ecosystem indicators in the upper Yellowstone River Watershed, the study area of
this project which represents a western North American landscape.
26
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Development and Evaluation of Multiscale
Mechanistic Indicators of Regional Landscapes
Carl Richards ', Luanda B. Johnson 2, and George E. Host2
'Minnesota Sea Grant College Program, University of Minnesota, Duluth, MN; 2Natural Resources Research
Institute, University of Minnesota, Duluth, MN
This project aims to develop suites of ecological
indicators that cross spatial scales, mechanistically re-
flect ecosystem states and processes, are statistically ro-
bust, and are applicable across regional landscapes.
These indicators also will be based on readily accessible
information available in a real-time framework.
The specific objectives are to: (1) develop pre-
dictive models that integrate landscape-scale factors with
reach-scale physical and chemical stream attributes to
quantify key compositional and structural attributes of
stream biota and derive ecosystem indicators at multiple
spatial scales; (2) evaluate the appropriate scale of ter-
restrial and aquatic data necessary to resolve regional
and local aquatic resource questions; (3) improve the
ability to distinguish and quantify natural variation in
indicators from that derived from anthropogenic stress-
ors; (4) assess the extent to which regional and local-
scale indices reflect fundamental ecosystem processes
and structural properties of stream habitats and biota;
and (5) quantify confidence limits and evaluate the geo-
graphic transferability of regional- and local-scale in-
dicators.
A multitiered sampling and modeling strategy was
used to integrate data collected at regional, local, and
site scales. These data will be used to identify indicators
at each scale that reflect critical ecosystem processes or
state variables related to the integrity and sustainability
of those ecosystems. These data also will be used to de-
velop and test indicators representing fundamental driv-
ing variables and processes at multiple spatial scales, and
integrate them into a system for identifying positive or
negative trends in ecosystem health.
The basic configuration of the landscape varied
substantially between the two primary study regions. The
grain of the landscape was finer in Michigan with
respect to both surficial geology and land cover char-
acteristics. Watersheds, in Minnesota were largely ho-
mogeneous with respect to these categories, while Mich-
igan watersheds exhibit more variation.
Strong (albeit seasonal) relationships were found
between landscape-scale features and dissolved nutrient
concentrations; however, results of nutrient bioassays
indicated that nutrients rarely limited primary produc-
tion, and landscape-scale variables describe only a mod-
erate amount of the variability in net primary production
(less than 50%). This trend also was observed hi macro-
invertebrate communities that were weakly related to
landscape-scale attributes but strongly related to local
(reach-scale) attributes.
Different parameters of interest are driven by dif-
ferent scales and factors within the landscape. Land-
scape-scale information can be very useful hi predicting
differences in stream water nutrient concentrations. In
contrast, it is difficult to predict major patterns of mac-
roinvertebrate assemblages or primary production with-
out understanding "the reach-scale characteristics that
have strongest influence on these variables; however, the
reach-scale variables are themselves partially governed
by landscape-scale variables. The project will continue
by examining other stream assemblages, and employing
correlative and mechanistic analyses to identify relation-
ships among ecological indicators operating at local and
regional scales. This approach will be directed by the
conceptual model presented in Figure 1. A critical part
of the next phase will be model evaluation, particularly
with respect to placing confidence limits on predictive
models. Following this will be an integration of indivi-
dual models into a larger predictive framework.
Watershed Characterization
Land Use/Land Cover
Landscape Pattern
Riparian Conditions
Hydrotogic Sub-Model
N, P, and Sediment Loads
Reach-Scale Characteristics
Physical Habitat
Water Quality
Ecosystem Prediction Sub-Model
NPP and Organic Matter Retention
Community Composition and Structure
Community Characterization
Species Richness/Diversity
Fish IBI
Macroinvcrtcbrate iBl
Figure 1. Conceptual model illustrating the organization of mechanistic factors operating at multiple spatial scales. Components summarize
the development of predictive models, ecosystem attributes, and fundamental ecosystem processes. Indicators are derived at all
spatial scales.
27
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Development and Testing
of a Multi-Resource Landscape-Scale Ecological Indicator:
Forest Fragmentation, Structure, and Distribution Relative to Topography
Steven W. Seagle and Philip A. Townsend
Appalachian Laboratory, University of Maryland Center for Environmental Science, Frostburg, MD
The goal of this project is to understand how water
quality and avian habitat quality vary across landscapes
as a function of forest distribution and topography (see
Figure 1). More specifically, the project seeks to deter-
mine whether water and avian habitat quality covary so
that they can be simultaneously extrapolated across the
Mid-Atlantic Highlands using existing databases of to-
pography, forest cover, and remotely sensed forest struc-
ture.
Efforts thus far have focused on: (1) taking an in-
tensive spring/summer field season census of forest
interior bird diversity and reproduction; (2) collecting
field data on forest structure within intensive field sites;
and (3) obtaining and analyzing radar and Landsat
satellite imagery. Field sites were chosen to represent
the climatic and topographic variation of the Mid-
Atlantic Highlands. Twenty 10-ha study sites were est-
ablished to equally represent dry and wet topographic
positions within the Appalachian Plateau and Ridge-and-
Valley provinces. Intensive studies on these sites will
form the empirical basis to test the hypotheses con-
cerning avian habitat quality, and then provide the stat-
istical basis to extrapolate across the Mid-Atlantic High-
lands.
During 1999, each study site was intensively sur-
veyed for breeding bird reproductive success. An array
of forest structural data also was collected for each site.
Preliminary results from the avian survey indicate that
forest interior bird reproductive success is higher on the
Appalachian Plateau than in the Ridge-and-Valley, and
within each province reproductive success is greater at
lower slope positions. These results support the hypo-
thesis that reproductive success, and thus habitat quality,
is related to topography, with wetter provinces and
topographic positions having higher success. Radar im-
ages of the intensive field sites are being analyzed to
identify predictors of variation in forest structural char-
acteristics. It is planned to collectively use degree of
forest fragmentation, structural diversity, and topograph-
ic position to map the quality of avian habitat across the
Mid-Atlantic Highlands.
Amount of forest cover within multiuse watersheds
is negatively correlated with the export of nitrogen from
the watershed. Watersheds within the Mid-Atlantic are
being analyzed to test the hypothesis that topographic
position of forest improves the correlation of forest cover
with water quality. If this hypothesis is correct, and pre-
liminary findings regarding the effect of topography on
avian habitat quality withstand further statistical scrutiny,
then forest cover and forest spatial distribution will to-
gether form a powerful indicator of both water and avian
habitat quality that can be mapped over large areas from
existing databases.
Efforts currently are focused on three tasks. First,
the relationship between forest structure and character-
istics of remotely sensed radar images are being anal-
yzed. This relationship will help refine predictions of
avian habitat quality. Second, processing of Landsat
land cover data and integration of forest cover with topo-
graphical data are aimed at developing simple measures
that describe the relationship between forest spatial pat-
tern and topography. These measures will be calculated
at the watershed level and correlated with water quality
parameters. Finally, a second field season to extend the
understanding of spatial variation in avian reproduction
and forest structure is being prepared.
28
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AQUATIC
RESOURCE
INDICATOR
COMPONENTS
TERRESTRIAL
RESOURCE
( QUALITY OF
I SURFACE WATER
FOREST
FRAGMENTATION
FOREST
TOPOGRAPHIC
DISTRIBUTION
FOREST
VERTICAL
STRUCTURE
FOREST INTERIOR
BIRD REPRODUCTION
RELATIONSHIP BEING
TESTED
RELATIONSHIP BASED ON
EXISTING DATA
Figure 1. Multiple characteristics of forests in the Mid-Atlantic are being examined as indicators of both surface water quality and forest
Ointerior bird reproductive success. Hypothesized relationships (dark arrows) are being tested through either reanalysis of existing
databases or field studies. In contrast, the effect of fragmentation on forest-interior bird reproduction is already quantified (white
arrow). As indicator components, both forest fragmentation and forest topographic distribution are readily mapped from existing
databases. The importance of forest vertical structure for forest bird diversity (white arrow) is well-known, but the relationship is
difficult to extrapolate from existing data. Thus, remotely sensed radar imagery is being examined for extrapolating forest
structure over large areas.
29
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Ecological Indicators
for Large River-FIoodpIain Landscapes
Monica G. Turner, E.H. Stanley, M.D. Dixon, R.E. Freeman, S.E. Gergel, H. Kang, J.R. Miller,
and J. West
Department of Zoology, University of Wisconsin, Madison, WI
Identifying landscape indicators that are well-
correlated with specific ecological functions remains a
crucial research need. Ecological indicators (population,
community, ecosystem, and landscape) are being devel-
oped and tested along reaches of the Wisconsin River.
A conceptual framework of this research is presented in
Figure 1. Two questions are being addressed: (1) Which
landscape metrics are most useful for monitoring pop-
ulation, community, and ecosystem processes in large
river-floodplain landscapes? (2) What are the constraints
on extrapolating relationships between landscape metrics
and ecological processes in large river-floodplain land-
scapes?
Spatially extensive field sampling is being com-
bined with landscape indicators to predict ecological
variables over broad scales. Field sampling (n = 220
plots) was conducted during May-July of 1999 within six
10-15 km study reaches. Vegetation (trees, shrubs,
saplings, tree seedlings, and herbaceous cover), soils,
and litter accumulation were sampled within each 10 x
20 m plot, and soil cores were obtained to estimate po-
tential denitrificadon, A census of birds was taken twice
at each plot using 8-minute point counts.
Preliminary results suggest that microbiological
indices can be explained by ecological processes that
operate at different spatial scales. At the landscape
scale, microbial activity appears to be influenced by land
cover, whereas the presence of levees may be a more
important determinant at the scale of individual tran-
sects. At finer scales, different enzyme activities were
found in soils under different types of trees. Microbial
processes are important for the ecological integrity of
floodplain ecosystems; these results suggest that changes
in land use patterns, levee construction or removal, or
tree species can affect these processes. Initial analyses
of 4,870 bird observations (70 species) revealed sim-
ilarity among the bird communities associated with each
study reach, with an average of 70-80 percent of the
species shared between reaches on a pairwise basis.
However, numbers of individual birds were quite
variable, with the three southernmost reaches containing
approximately 30 percent more birds than the three
northernmost reaches (e.g., the number of woodpecker
species and individuals decreased from south to north).
Some species generally associated with northern
Wisconsin habitats were only detected at the southern-
most reaches, suggesting a strong influence of land use
or river flow modification. From a total of 39 tree spe-
cies encountered, some floodplain species (e.g.,Acer-
saccharinum, Fraxinus pennsylvanica) were more abun-
dant in the less human-modified southern reaches,
whereas some upland species (Populus tremuloides,
Quercus velutina) were more abundant in the more
modified northern reaches.
During the coming year, the following activities
are planned: (1) analyzing the field data obtained in six
river reaches during the 1999 field season (bird
censuses, vegetation sampling, denitrification, and
microbial activity) in relation to landscape indicators;
(2) interpreting historical and recent aerial photography
for study reaches in the 1930s, 1960s, and 1990s;
(3) conducting the second season of field sampling; and
(4) beginning to address Question 2 by predicting and
testing ecological indicators in new study landscapes,
determining whether there are thresholds in landscape
pattern beyond which ecological processes change
qualitatively, and assessing the sensitivity of ecological
indicators to landscape changes.
30
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Societal
Values
Water Quality
Flood Control
Integrity /Biodiversity
of Watershed
Assessment
Endpolnt
Proportion of river-
floodplain landscape
with acceptable
Integrity and
biodiversity
• i t i^r^V i
Proportion of river-
floodplain landscape
with acceptable
ability to retain and
purify water
Ecological
Processes
•••IM^Hi i
Indicators
Community and
Population
Succession
Species composition
Species recruitment
i a . i . i i i . i i i i
Tree species richness
Relative abundance of
tree species
Recruitment of riparian
pioneer trees
Forest age structure
Invasive species
Forest birds
, ,
Landscape
Landscape structure
Landscape change
Topographic position
, , i . , , i , i i i i i
No. o! cover types
Area of each
cover type
Patch size, shape and
distribution
Edge and road density
Riparian width & gaps
Adjacency probabilities
i i
Ecosystem
Nitrogen dynamics
Organic matter
Soil processes
t 1 1 i t 1 i 1 t a i 1 1
Litter accumulation
Coarse woody debris
Soil organic matter
Denitrilication
Soil nitrogen
Soil carbon
Aboveground net
primary production
I t I i 1
Figure 1. Conceptual framework for research in the Wisconsin River floodpiain landscape showing relationships among societal values,
assessment endpoints, ecological processes, and indicators.
31
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Characterization of the Ecological Integrity of Commercially
Grazed Rangelands Using Remote Sensing-Based Ecological Indicators
Neil E. West, Robert A. Washington-Allen, and R. Douglas Ramsey
Department ofRangeland Resources, Utah State University, Logan, UT
The purpose of this retrospective study was to
characterize the ecological integrity of a semi-arid land-
scape using satellite remote sensing and Geographic In-
formation System (CIS) technologies.
The ecological basis of the research was the tran-
sition threshold hypothesis. A transition threshold is de-
fined as the boundary in space and time between two se-
rai states (e.g., the change of a grassland to woodland).
A transition is the process (e.g., climate or grazing in-
tensity) that brings about a change in state. An ecolog-
ical risk assessment framework was used to determine
the assessment endpoints of rangeland degradation as a
change in plant growth form composition, a decrease in
plant productivity, a reduction in soil quality, accelerated
soil erosion, and a change in landscape composition and
pattern.
Measurement endpoints, which relate to these five
assessment endpoints, were derived from 27 years
(1972-1998) of wet and dry season Landsat satellite im-
agery and 2 years of annual scenes (1985 and 1986).
Measurement endpoints were the soil-adjusted vegetation
index (SAVI), a surrogate for vegetation parameters and
soil quality; the soil stability index (SSI), a surrogate for
soil erosion; and thematic maps, where landscape met-
rics were used to measure changes in landscape structure
and configuration. The study site was 54,000 ha of the
sagebrush steppe portion of the 108-year-old mixed
grazing operation (wildlife and beef cattle) Deseret Land
& Livestock Ranch (DLL) in northeastern Utah.
A GIS database of site biological, physical, and ad-
ministrative characteristics, including historical and cur-
rent ranch management records, was developed. The
data were analyzed at the waterpoint, paddock, and land-
scape scales using graphical timeseries and multiple re-
gression analyses. Sagebrush steppe is a two-phase
mosaic of shifting dominance between shrubland and
grassland due to herbivory, fire, and climate change. It
was hypothesized that at the landscape-level, SAVI
would respond to climate change (i.e., La Nina/El
Nino). It also was hypothesized that at the community
level, coincident periods of drought and intense grazing
would lead to a landscape dominated by shrubland and
increased erosion.
Two validation studies were conducted that confir-
med that SAVI, an indicator of phenology, was consis-
tent with field data and that interannual SAVI can dis-
criminate between grazing effects. It was found that at
the landscape-scale, the ranch had become more frag-
mented and was an unstable limit cycle responding to El
Nino (wetting) and La Nina (drying), respectively.
At the community-level, shrubland dominated since
1974, grassland was declining, and there is no indication
of increased erosion. Three-dimensional surface anal-
ysis suggests that in terms of SAVI response, increased
grazing and climate are factors in shrub increase, but
grazing appears to be the main factor in grass cover
decline (see Figure 1).
Ecosystem measures such as SAVI can miss
fundamental changes in composition that are important
to ranch management. Limited field datasets from fed-
eral agencies had assessed the ranch in good con-
dition. However, these datasets were temporally limited
(6 years versus the 27 years of imagery), inconsistent, or
measured inappropriate indicators. The following tasks
remain to be completed: validation of the SSI, accuracy
assessment of dry season thematic maps, comparison of
metrics within 20 paddocks, and completion of the
timeseries piosphere study.
32
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T*
' ««
„ ,H .
- •«•
-•*
'•»
'•x
50
45
40
JS
30
!S
JC
Figure 1. The top left figure suggests a fold catastrophe and is the relationship between the independent factors: number of cows and the
Utah Region 5 Palmer Drought Severity Index (PDSI), and the dependant factor: landscape-level lagged wet season mean soil-
adjusted vegetation index (SAVI). PDSI < 0 indicates drought conditions. The same independent factors are related to
community measures of grass cover (top right) and shrub cover (bottom center).
33
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Section 3.
Aquatic Indicators
-------
Using Bioindicators To Develop a Calibrated Index
of Regional Ecological Integrity for Forested Headwater Ecosystems
Robert P. Brooks ', Robert S. MulvihiU2, Terry Master3, and Timothy J. O'ConneU'
'Perm Stale Cooperative Wetlands Center, University Park, PA; 2Powdermill Nature Reserve, Carnegie Museum of
Natural History, Rector, PA; 3East Stroudsburg University, East Stroudsburg, PA
The objective of this project is to develop a region-
al index of ecological integrity for forested headwaters
in the Mid-Atlantic States, with particular emphasis on
the Louisiana waterthrush (Seiurus motadlla), referred
to as "LOWA." LOWA are forest-breeding warblers
that also are riparian obligate. They are dependent both
on large patches of mature forest and high-quality
instream conditions. The index is intended to integrate
and calibrate existing indicators of integrity that address
stream health and upland forest condition individually,
and at different scales.
Beginning in 1998, LOWA were monitored on 23
forested headwater streams in three physiographic pro-
vinces of Pennsylvania (see Figure 1). These streams
represent either: (1) high water quality in large patches
of interior forest; (2) high water quality in a relatively
fragmented forest setting; or (3) low water quality in
large patches of interior forest. Established indices of
biotic integrity (IBIs) will be applied to the same streams
where LOWA reproductive success already has been
measured. The degree to which attributes of LOWA re-
productive success can link small scale IBIs, such as
macroinvertebrate IBIs, to landscape scale IBIs (i.e., the
Bird Community Index) is being investigated.
Early analysis confirms that LOWA population
variables correspond with large-scale condition as de-
fined by the Bird Community Index. Correlates to in-
stream condition are more complex to generate, and the
results are pending. For example, the 1998 macroin-
vertebrate characterization for the Central study area in-
cluded 9,862 individuals in 180 genera, 83 families, 18
orders, 9 classes, and 4 phyla.
In 1999, unusually high nest predation rates in all
three study areas occurred. Nest predation rates were
elevated on both reference and impacted streams. In the
Central study area, the evidence suggests that mink
(Mustela vison) are the culprit predators. It is speculat-
ed that mink have become locally (and perhaps region-
ally) abundant following a succession of mild winters.
Preliminary results of this research already have
displayed the importance of " offstream" wetland seeps
and springs as vital to the foraging efficiency of LOWA
nesting on acidified streams. On acidified streams with
abundant offstream foraging opportunities, LOWA are
reproducing at a rate comparable to that observed hi the
most "pristine" streams.
This work also highlights the need to conduct re-
search at large scales and across physiographic bound-
aries. For example, had all of the sites been located in
one region in 1999, it would not be possible to demon-
strate that the elevated nest predation observed was a
widespread phenomenon. The 2000 field season is the
third and final season on this project. In the upcoming
year, the IBI will be developed, and its relationship to
the land cover pattern and the habitat attributes that have
been measured from the ground will be examined.
reference
degraded
Eastern
Central
Study Area
Western
Figure 1. Mean habitat suitability index (HSI) scores for Louisiana Waterthrush on reference and degraded streams in three physiographic
provinces of Pennsylvania.
37
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Stream Plethodontid Assemblage Response (SPAR)
Index: Development. Application, and Verification in the MAHA
Robert P. Brooks and Gian L. Rocco
Penn State Cooperative Wetlands Center, Environmental Resources Research Institute, Pennsylvania State
University, University Park, PA
Amphibians generally are considered to be valu-
able response indicators (Vitt et al. 1991, Dunson et al.
1992). In small lotic systems that largely exclude pre-
dacious fish, small, lungless salamanders of the Family
Plethodontidae, can exist in surprisingly high numbers,
functioning as both invertebrate predator and vertebrate
prey.
By virtue of their diverse and complex life his-
tories and abundant and stable populations, stream sala-
manders offer the opportunity of serving as important
ecological indicators for the assessment of headwaters,
especially where traditional indicator species assem-
blages (fish, macroinvertebrates) may be poorly devel-
oped or absent.
In a study of 14 Central Appalachians headwaters
in Pennsylvania, Rocco and Brooks (in preparation)
document significant stream plethodontid responses to
acidified and degraded stream conditions, attesting to
their potential as bioindicators. Various useful metrics,
based on species composition, abundance, and lifestage,
also are proposed. In consideration of the effectiveness,
relative ease, and low cost of the sampling technique,
stream plethodontid assemblage responses (SPAR), used
alone or in combination with other small stream assess-
ment criteria, promises to be an effective assessment and
monitoring tool for headwaters.
At the moment, however, the ability to interpret
SPAR in similarly impaired streams across a larger
geographical area is hampered by the relatively small
area sampled. A reference base ideally would be geo-
graphically widespread, represent the entire range of
conditions, and consist of a representative and unbiased
sample of the population of headwaters in the region of
interest.
To address this need, the primary objectives of this
research are to: (1) describe the range and variability of
SPAR across commonly encountered gradients of anthro-
pogenic degradation (e.g., stream acidification, forest
and riparian corridor fragmentation and degradation,
pollution) in the Mid-Atlantic Highlands Area (MAHA);
(2) develop and adjust SPAR for use in MAHA head-
waters; and (3) evaluate the reliability and resolution of
SPAR by application and verification.
The development, application, and verification of
SPAR within MAHA will entail a two-phase process.
Phase I aims to document and study plethodontid assem-
blage responses to as many different stream conditions
as possible, across a large geographical area. This data-
set will enable the development of the SPAR index for
the MAHA. In Phase II, a random set of headwaters of
unknown condition will be sampled and its impairment
determined by the SPAR index. Once applied, an inde-
pendent assessment of the stream by traditional methods
(macroinvertebrates, water quality, surrounding land-
scape) will allow the verification of the reliability and
consistency of the index, and if necessary, its cal-
ibration. The use of volunteers during the second phase
will permit evaluation of the method for nonspecialists
and long-term amphibian monitoring efforts.
The response composition of free-ranging, nat-
urally occurring stream salamander populations in de-
graded and nondegraded watersheds will be studied by
intensive sampling. Relevant abiotic and biotic var-
iables at the plot, stream reach, and watershed will be
measured and correlated to SPAR metrics.
The expected findings include: (1) description of
stream salamander assemblage response along disturb-
ance and pollution gradients commonly encountered in
the MAHA Region; (2) identification of species most
responsive to the environmental degradation investi-
gated; (3) recommendations on the use of stream sala-
mander assemblages in the assessment of headwaters;
and (4) development and testing of stream salamander
metrics that could be implemented separately or in con-
junction with other criteria when performing small
stream assessments.
Ultimately, it is hoped that this work leads to the
improvement of existing headwater assessment protocols
for the MAHA by the addition of a bioindicator that is
abundant, widespread, ecologically important to several
trophic levels, and interfaces between aquatic and
terrestrial components of riparian areas.
38
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Soil Enzyme Stability
as an Ecosystem Indicator
Richard P. Dick
Department of Soil Science, Oregon State University, Corvallis, OR
Sustainability assessment of ecosystems to assist
land managers and policymakers in promoting long-term
sustainability is a national priority, but quantifying envi-
ronmental sustainability remains an elusive goal. One
approach is to use the soil as an indicator of ecosystem
"health."
Soil enzyme activity assays are advantageous as
potential indicators because they are: (1) operationally
practical; (2) sensitive integrative "biological finger-
prints" of past soil management; and (3) apparently re-
lated to soil aggregation, linking enzymes with soil tilth.
In the first year of the project, the initial screening
of three promising soil enzyme assays as indicators took
place at three experimental sites in Oregon (located in
divergent forest and agroecosystems) where detailed
management history is known and soils vary widely in
soil "health" (i.e., biological activity and organic matter
content) because of past soil management. This included
refinement of sample handling and laboratory protocols
to increase sensitivity and reproducibility of the indica-
tors. Samples were taken in early May, June, and Sep-
tember of 1999 to assess the seasonal temporal var-
iability of these indicators. It was found that the forest
soils generally have much higher activities than agri-
cultural soils. This makes sense, because agricultural
soils are regularly disturbed. Forest soils, even with
recent logging, would only have had surface disturbance
once, and even then, there would not be tillage. There
is a very consistent pattern in the agricultural soils when
within-site treatment effects are examined, in that those
receiving greater C inputs (as cover cropping, manure,
and green manure) have higher activities than the plots
receiving lower C inputs.
Across agricultural soils, the biological measure-
ments such as ergosterol, fungi biomass, and biomass C
are very consistent with soil enzyme activities. Bacterial
counts, however, show very few significant differences
as a function of soil management in either forest or agri-
cultural soils. This suggests that soil enzyme activities
are good indicators of the soil biological component and
are reflecting the differences due to fungal biomass.
The second year of the project will involve expan-
ding to a wider range of soil types and soil management
systems. A detailed data analysis will be performed and
the potential for relative soil quality indicators that are
independent of soil type will be investigated.
39
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Integrative Indicators of Ecosystem Condition and Stress
Across Multiple Trophic Levels in the San Francisco Estuary
Richard C. Dugdale
Romberg Tiburon Center for Environmental Studies, San Francisco State University, Tiburon, CA
The goal of this project is to evaluate a set of po-
tential indicators of ecosystem condition for the San
Francisco Estuary (SFE). The objectives of the research
and field effort are to: (1) devise indicators of ecosys-
tem condition; (2) investigate their relationships to
stressors, including salinity, turbidity, nutrients, and in-
troduced species; and (3) assess the utility of these in-
dicators.
This research is being conducted on the open-water
ecosystem of the SFE, including the portion of the
landscape from freshwater to marine ecosystems. These
potential indicators represent key population and individ-
ual-level processes hi a variety of trophic levels. The
benthic community structure also is being examined for
possible indicators of stress. All indicators are relatively
simple to measure, significant to population ecology,
expected to be sensitive to stress levels, and likely to be
transportable to other aquatic habitats.
These indicators are: (1) nutrient status and pro-
ductivity performance of phytoplankton; (2) relative con-
tribution of diatoms to algal biomass and productivity;
(3) reproductive rate of common copepod (zooplankton)
species; (4) condition indices of larval herring; (5) con-
dition indices of the dominant benthic bivalves; (6) sev-
eral measures of benthic community structure; and
(7) expression of stress proteins in the indicator animal
species.
This project is very new, funded in October 1999,
so only preliminary findings are available. The field
work has been initiated with monthly cruises to three
sites hi the SFE. Measurements of temperature, salinity,
nutrients, and algal biomass as chlorophyll, paniculate
nitrogen, and paniculate carbon were made. NO3 and
NH, uptake (fractionated by size class) with samples
incubated under natural turbidity and clarified water
conditions were obtained during each cruise.
Comparison nutrients and chlorophyll concen-
trations from December 1999 with previous obser-
vations in Central Bay suggest relatively low values,
probably resulting from an exceptionally dry fall.
Comparison of nutrient and chlorophyll concentrations
from December 1999 hi Central Bay with the two new
up-estuary stations, San Pablo and Suisun Bays, shows
nutrients, especially Si(OH)4, increasing and chlorophyll
decreasing in the freshwater direction. Copepod (zoo-
plankton) reproductive rates (see Figure 1) were
measured by collecting with gentle net tows, diluting the
catch in surface bay water, and incubating individual
females for 24 hours in 125 mL polycarbonate bottles
filled with bay water with eggs strained out.
The first herring larval collections have just been
made at the three sampling stations. The contents of
each tow were split, with half preserved in formalin and
half in ethanol for measurements of body characteristics
including length, weight, head width, and total eye diam-
eter.
The ethanol-preserved samples are stored for fu-
ture studies outside the scope of this project. Sites have
been selected for the benthos project at each of the three
study areas and collections made for condition and
glycogen analyses. Work on stress-induced proteins has
been impacted by the departure of the post-doc involved,
who has taken a faculty position at another institution.
Planning for the prosecution of this aspect of the project
is underway with the possibility of arrangements for
collections to be made at our institution and the analyses
at the individual's new post. Other options are being
explored as well.
40
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o
c
CD
3
CT
CD
2
0
4812
Eggs per Female
16
20
24
Figure 1. Histograms of egg production rates from copepods (Acartia) at Stations 2 (top) and 3 (bottom) during the first cruise. A different
species, not shown here, was most abundant at Station 1. Vertical lines indicate medians. Differences in egg production rates such
as these will be analyzed for relationships with salinity, temperature, chlorophyll, and other environmental variables.
41
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Developing an Indicator for Nutrient Supply
in Tropical and Temperate Estuaries, Bays, and Coastal Waters
Using the Tissue Nitrogen and Phosphorus Content of Macroalgae
Peggy Fong
Department of Organismic Biology, Ecology, and Evolution, University of California, Los Angeles, CA
Coastal eutrophication that results from increases
in nutrient supply is a critical problem worldwide,
causing major changes in marine populations and com-
munities. The overall goal of this project is to develop
an indicator that quantifies nutrient supply to tropical and
temperate marine ecosystems using the tissue nitrogen
(N) and phosphorus (P) content of macroalgae. To com-
plement the present suite of indicators used to measure
eutrophication, this indicator is targeted to be especially
useful in systems where nutrients are supplied in pulses
or those where nonpoint sources of nutrients such as
groundwater or fluxes from the benthos are important.
The five specific objectives are to: (1) continue to
identify and test potential species for use as indicators;
(2) establish relationships between timing and magnitude
of nutrient supply and accumulation of N and P in algal
tissue; (3) establish quantitative relationships between
environmental conditions, N and P supply, and tissue N
and P; (4) develop a numerical simulation model based
on experimental results that may be used as a "standard
curve" for the indicator to hindcast nutrient supply in the
field; and (5) field test the indicator.
Macroalgae from two regions in the Caribbean
(Southwest Puerto Rico and Caribbean Panama) and two
in the Eastern Tropical Pacific (Gulfs of Panama and
Chiriquf, Panama) were collected to determine if the
nitrogen (N) and phosphorus (P) content of their tissues
reflected differences in nutrient availability in these sys-
tems, making them good indicator species. Large dif-
ferences were found in the tissue P content of Hypnea
musciformis, an exotic red alga that has invaded most
tropical areas. H. musciformis from the Caribbean had
one-half the amount of P than in the ETP. There also
were significant differences in N content, but these were
of a lower magnitude. Similarly, Dictyota spp. in the
Carribean had only one-half the tissue P content found in
the ETP; however, tissue N of Dictyota spp. was not
different among these ocean areas, suggest ting it may
not be useful as an indicator of P (see Figure 1).
The differences hi P content in algal tissue between
the Caribbean and ETP may reflect the greater per-
centage of P-adsorbing carbonates hi Caribbean sed-
iments. Tissue P content also was significantly different
among regions: Gulf of Panam£ > Gulf of Chiriquf >
Caribbean Panama > Puerto Rico. This contrasts with
tissue N where the pattern was Gulf of Panama > Gulf
ofChiriqui = Puerto Rico > Caribbean Panama. While
greater N and P in the Gulf of Panama may be attributed
to seasonal upwelling, the larger N content in Puerto
Rico compared to Caribbean Panama may be due to
anthropogenic inputs.
The tissue N and P content of macroalgae may
provide useful insight into nutrient availability in tropi-
cal systems. Furthermore, cosmopolitan species such as
H. musciformis may be especially good indicators of
nutrient regime.
42
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Hypnea
Dictyota
ETP
Caribbean
Figure 1. Differences were found between Hypnea musciformis and Dicnola spp. sampled from the Eastern Tropical Pacific (ETP) and ihe
Caribbean in terms of: a) tissue N, b) tissue P, and c) N to P ratio.
43
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Microbial Indicators of Biological
Integrity and Nutrient Stress for Aquatic Systems
James P. Grover and Thomas H. Chrzanowski
Department of Biology, University of Texas at Arlington, Arlington, TX
This project explores connections between bio-
logical integrity and nutrient supply and limitation, fo-
cusing on the microbial component of lakes and reser-
voirs. Several hypotheses are being tested concerning
patterns and correlations among indicators of biological
integrity that are based on nutrients (i.e., carbon,
nitrogen, phosphorus, and trace minerals) and microbial
communities. The temporal and interregional variability
of these indicators also are being assessed.
Two reservoirs in north Texas are being examined
year-round, and two lakes in the Experimental Lakes
Area (Ontario, Canada) are being studied during the ice-
free growing season (see Figure 1). One of the latter
lakes is pristine, while the other has been experimentally
eutrophied with phosphorus additions. Much of this
work involves measures of seston element composition
and measures of changes in microbial communities fol-
lowing experimental manipulation of nutrients (bio-
assays).
Seasonal patterns have been analyzed and compar-
isons made among lakes for the first year of sampling.
It was hypothesized that indicators based on the C:N:P
composition of seston would agree with growth re-
sponses in bioassay experiments.
In Texas lakes, temporal correlations between
indicators of P-limitation and seston C:P and N:P ratios
were all in the hypothesized directions. Correlations
between indicators of N-limitation often were opposite to
predictions. Indicators of algal and bacterial nutrient
limitation were positively correlated in Texas lakes, as
hypothesized. Bioassay experiments in Canadian lakes
were generally uninformative, largely due to high var-
iability and a number of unexpected negative effects
from nutrient enrichment.
Physiological profiles of bacterial communities
based on the use of 95 carbon substrates support a
hypothesis that bacterial community structure shifts in
response to seasonal changes in nutrient limitation.
Principal components analyses reveal relatively high
response to amino and carboxylic acids in cool seasons,
and to carbohydrates in warm seasons. The amplitude
of this pattern was stronger in the experimentally eutro-
phied Canadian Jake than in the oligotrophic reference
lake.
These results address the relative utility of bioassay
experiments and measures of seston C:N:P composition
as indicators of nutrient limitation. The two approaches
perform more consistently when P-limitation is diag-
nosed than when N-limitation is diagnosed. Indicators
based on seston C:N:P ratios appear to be more widely
applicable than those based on bioassay experiments,
because the latter performed poorly in Canadian lakes.
On the other hand, where applicable (in Texas lakes),
bioassay experiments yield insights to microbial pop-
ulations that may be useful in modeling and lake man-
agement. Physiological profiling of bacteria suggests
seasonal shifts in use of carbon substrates, which may be
linked to seasonal changes known for other limnetic
organisms. These seasonal patterns appear to be more
pronounced in eutrophic than in oligotrophic lakes, and
may help characterize lakes of different trophic status.
As the additional data from Years 2 and 3 are
analyzed, interannual variations in the indicators and the
patterns of correlation summarized above will be as-
sessed. To obtain greater insight from indicators based
on seston composition, size-fractionated sampling of
seston in Canada during 1999 was instituted, though data
are not yet completely analyzed.
44
-------
Experimental Lakes Area
Two regions, two lakes in each region.
North Texas
Microbiological analysis:
Algal abundance and composition
Bacterial abundance
Bacterial composition (Biolog)
Bacterial productivity
Protozoan grazing
Dilution bioassays:
Algae: N, P, trace nutrients
Bacteria: N, P, organic C
Depth Profiles
Temperature, DO
.c
TSL
8
, ,
f 1
i /
'''
/'
/
*
Samples for:
Seston C:N:P
Dissolved
nutrients
Particulate
nutrients
Figure 1. Microbial indicators of biological integrity and nutrient stress for aquatic systems.
45
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Foraminifera as Ecosystem Indicators:
Phase 1. A Marine Benthic Perturbation Index; Phase 2. Bioassav Protocols
Pamela Hallock, Heidi Crevison, Thomas Dix, Helen Talge, and Dana Williams
Department of Marine Science, University of South Florida, St. Petersburg, PL
The objectives of this project are to develop pro-
cedures for the routine use of foraminifera as indicators
of biological integrity in field and laboratory appli-
cations. In Phase 1, sediment cores are being used to
test an index for assessing perturbations of marine ben-
thic ecosystems based on changes in key taxa of fora-
minifera. In Phase 2, protocols for assessing stress re-
sponses in foraminifera induced by elevated temperature
and visible and ultraviolet radiation are being developed.
A prototype index to quantify change in benthic
ecosystems that can be applied to historical, sediment-
core, and surface-sediment datasets has been developed.
The model ranks relative abundances of key foramin-
iferal taxa and morphogroups, and total abundances,
generating an index value when foraminiferal assem-
blages are compared temporally or spatially. Tests of
the index using sediment cores from Florida Keys reefs
and Tampa Bay are in progress.
Bleaching has been observed in populations of
Amphistegina in all subtropical oceans since 1991. Field
sampling in the Florida Keys in 1997-1999 revealed
continued stress, with a significant increase in the sea-
sonal bleaching peak in 1998. Stress symptoms in Am-
phistegina can be documented using visual, physio-
logical, and cytological techniques. Visual rankings,
which can be verified by cytological analyses using elec-
tron microscopy and image analysis, remain the most
cost-effective method of documenting stress responses.
Protocols for measuring ATP to determine viability and
metabolic activities were adapted and results compared
with visual and cytological assessments. High variability
and standardization limitations indicate that ATP analysis
is not cost-effective.
Exposure to elevated intensities of photosynthet-
ically active radiation (400-700 nm), alone or in combi-
nation with added UVB (280-320 nm), induced symbi-
ont loss in A. gibbosa similar to that seen in field pop-
ulations. Comparison of responses to two spectrally dif-
ferent visible light sources revealed that, although ex-
perimental intensities of blue and white light were the
same in photons (7.76 fiE/m2s), the higher-energy blue
light (7.21 x 10'5 W/m2) both induced faster growth and
more symbiont loss than the lower-energy white light
(5.69 x 10-5 W/m2) (see Figure 1).
Mass bleaching devastated corals nearly worldwide
in 1998. Bleaching in A. gibbosa in the Florida Keys
also peaked in the summer of 1998. A key difference
between bleaching in corals and foraminifers is that coral
bleaching correlates most consistently with elevated sea
surface temperatures, which can induce photo inhibitory
stress, while bleaching in Amphistegina appears to be
directly associated with photoinhibitory stress. Other
researchers using molecular biomarkers have found evi-
dence of stress in corals 3 months prior to visible bleach-
ing, indicating that corals may be responding to the same
stresses that are bleaching the foraminifera. Recog-
nizing similarities and differences between these two
physiologically similar, though taxonomically different
symbiotic systems, should facilitate understanding the
global decline of coral reefs. Visual techniques remain
the most dependable and cost-effective methods of doc-
umenting and analyzing bleaching symptoms and shell
breakage in Amphistegina spp.
Goals for the year 2000 are to standardize visual
protocols using digital photography and image analysis,
while continuing to document responses to photic and
temperature stress. Contacts also have been established
with scientists applying molecular biomarker protocols
to a variety of reef organisms, with the goal of deter-
mining whether these techniques can be applied to Am-
phistegina. The results of the core analyses and stess ex-
periments will be prepared for publication.
46
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Blue Hi
Blue Low -^-White Hi -+r- White Low
0.95
% Symbiont Loss
0.60
7/6/99 7/11/99 7/16/99 7/21/99 7/26/99 7/31/99 8/5/99 8/10/99
Figure 1. Influence of spectral quality on growth and symbiont loss. Mean diameter of 45 A. gibbosa individuals exposed to two inteasities
(7.76 and 2.81 ^E/nr) of two spectrally different light-sources (blue lamp-photons are concentrated in the higher energy portion of
the electromagnetic spectrum, white lamp-photons are more concentrated in the lower energy portion). Shaded boxes show the
percentages of individuals showing visible symbiont loss after 4 weeks (grown in 10 cm petri dishes at 25" C in nutrient-enriched
Erdschreiber medium).
-17
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Biogeochemical Indicators
of Watershed Integrity and Wetland Eutrophication
K. Ramesh Reddy \ W.F. DeBusk ', A. Ogram ', W. Graham ', M.M. Fisher2, E.F. Lowe 2,
and L. W. Keenan 3
!Soil and Water Science Department, Agricultural and Biological Engineering Department, University of Florida,
Gainsville, FL; 2St. Johns River Water Management District, Palatka, FL
The central hypothesis of this research is that rates
of biogeochemical cycling of C, N, and P in wetlands
can be used to indicate the ecological integrity of wet-
lands, and that the concentrations of certain forms of
these elements can accurately predict the rates of ecolog-
ically important processes.
The objectives of this research are to: (1) identify
the key biogeochemical processes impacted by nutrient
loading and measure the rates of these processes along
the nutrient gradient; (2) develop relationships between
a "process" and its related, easily measurable "indica-
tor" ; (3) determine the spatial and temporal distribution
of easily measurable indicators for a test wetland ecosys-
tem; (4) determine the spatial variations in biogeochem-
ical processes, and develop spatial maps for various
processes to determine the extent of impact and risk as-
sessment; and (5) validate the predictability of empirical
relationships by making independent measurements of
biogeochemical processes in different wetland ecosys-
tems. It is planned to test the hypotheses presented
above in the Blue Cypress Marsh Conservation Area
(BCMCA), located within Upper St. Johns River Basin,
FL. Some areas of the BCMCA have been impacted
over the years as a result of nutrient loading from adja-
cent uplands, resulting in distinct nutrient and vegetation
gradients. The BCMCA provides the benefit of estab-
lished gradients of high-nutrient (impacted) to low-
nutrient systems (unimpacted), to test the hypotheses of
this proposal.
The research will focus on key biogeochemical
processes and microbial communities regulating the fate
of nutrients in the soil and overlying litter layer at var-
ious spatial and temporal scales. This will be accom-
plished by conducting a series of laboratory and field ex-
periments.
Statistical and modeling tools will be used for
analysis and synthesis of the biogeochemical process and
indicator data, including: (1) descriptive statistics re-
lated to frequency distribution and central tendency of
indicator values to make comparisons between areas of
low and high anthropogenic impact; (2) multivariate
analyses to evaluate the relationship between biogeo-
chemical indicators, biogeochemical processes, and eco-
logical integrity; (3) geostatistical analysis to compare
the characteristic spatial patterns and structure of biogeo-
chemical indicators; and (4) temporal analysis of in-
dicators in low- and high-impact areas.
The first stage of this research is expected to result
in the confirmation of the utility of simple chemical
measures as reliable predictors of rates of biogeochem-
ical processes. The study also will produce statistical
models describing these relationships.
Such models would provide valuable research and
management tools. Because these measures would be
rapid and inexpensive, investigations of biogeochemical
processes could be expanded providing a better under-
standing of these fundamental mechanisms of eco-
systems.
48
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Development and Evaluation of Ecosystem
Indicators for Urbanizing Midwestern Watersheds
Anne Spade, Jonathan M. Harbor, Midhat Hondzo, and Bernard A. Engel
Purdue University, West Lafayette, IN
Urbanization is thought to have negative impacts on
stream ecosystems, and yet the actual causal relation-
ships between land use change and stream community
response have not been well studied. This project focus-
es on the development of predictive indicators of urban-
ization that are applicable to Midwestern watersheds and
stream ecosystems.
The objectives of this project are to: (1) quantify
the impacts of urbanization on hydrologic regimes, water
quality, and habitat structure of stream ecosystems using
paired experimental watersheds, and to develop linked
models that accurately predict these impacts; (2) use the
linked models as a virtual laboratory within which to
generate and test indicators of urbanization and hydrol-
ogic change in terms of responses of fish and macro-
invertebrate communities; and (3) use these models and
indicators to assess the response of stream communities
to alternative urbanization scenarios with extension to
larger watersheds in the region.
The research examines seven watersheds in central
Indiana that are in transition from rural to urban. Link-
ages between increased urban runoff, altered channel
morphology, water quality effects, and reduced biotic
integrity are being evaluated for three sites on each
stream. Percent urbanization of the watershed, derived
from 1997 SPOT satellite imagery, is used in a runoff
model (L-THIA) to predict flow regime at each site.
Intensive water quality sampling at selected sites pro-
vides a dataset for the development and testing of a
physically-based water quality model. Stream cross-
section measurements are used to determine the critical
discharge (Qcrit) required to mobilize substrate.
Statistics describing the long-term exceedance of
Qcri, characterize the level of habitat disturbance at the
site. Physical habitat at each site has been measured
using standard procedures, including a qualitative habitat
evaluation index (QHEI). Macroinvertebrate, periphy-
ton, and fish collections at each site are used to char-
acterize stream community structure.
A dynamic hydrology model has been developed
that can simulate cross-sectional averaged velocities,
shear stress velocities, and water depth variability during
storm peaks. Water temperature dynamics and nutrient
transport also have been modeled with satisfactory re-
sults. Stream habitat measurements (QHEI) and bio-
logical collections have shown significant differences
across the range of urbanization.
A suite of functional biological metrics currently
are being evaluated. This work will provide a sound
basis for the use of specific indicators as tools in re-
gional planning of watershed development. The risk
analysis portion of the work will provide a probabilistic
measure as to whether a potential urbanization scenario
can achieve stream water quality and biological targets.
The hydrologic and water quality models developed thus
far will be extended for use with dissolved oxygen and
nutrient transport, and will be linked to the L-THIA run-
off model. Stream biota and habitat quality, which al-
ready have been correlated to general land use (see
Figures 1-3), will be further correlated to channel mor-
phometry and flow variability.
Once this is accomplished, a risk analysis can be
conducted on the effects of various runoff patterns on the
stream community metrics.
Urbanization leads to
increased Imperviousness
Increased runoff volante and frequency I
Increased channel scour
Channel Incision
Channel widening
Increased energy + destabilized
channel, degraded habitat (QHEI)
Figure 1. A conceptual model of stream channel changes associated with urban watersheds.
49
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Q)
EP
CO
_C
o
Natural
Urban
Time
Figure 2. Hydrographs in urban streams are expected to show increased mean flow as well as increased frequency of storm peak flow.
80
LU
I
a
60
• Middle and Lower Sites
R2 = 0.72
A Upper Sites
R2 = 0.00
5 10 15 20 25 30
Watershed imperviousness, %
Figure 3. Significant degredation of fish habitat quality, as measured by QHEI, has been observed in streams undergoing urbanization in the
region near Indianapolis, IN.
50
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An Integrative
Aquatic Ecosystem Indicator
Richard S. Stemberger and Eric K. Miller
Department of Biological Sciences and Environmental Studies Program, Dartmouth College, Hanover, NH
This project uses measures of the relative supply of
carbon (C), nitrogen (N), and phosphorus (P) to inte-
grate watershed, lake, and pelagic zooplankton assem-
blages in a multitier ecological indicator for monitoring
lake integrity. Complex species assemblages are aggre-
gated into simple consumer guilds that reflect relative
elemental N and P intracellular requirements of species.
Element supply ratios relate a variety of potential risks
to lake ecosystem function such as loss of the cool-water
refuge, bioaccumulation of toxins, algal turbidity, acidi-
fication, and UV-B toxicity. These risks are associated
with zooplankton assemblage structure along the N:P
ratio gradient.
Objectives of the project include the following:
1. Evaluate metrics based on the C, N, and P sup-
ply of the total dissolved plus seston fraction of lake
water as indicators of the character of lake zooplankton
assemblage.
2. Establish the strength of the linkage between
the relative C, N, and P supply from the watershed to
the C, N, and P supply to the lake. The hypothesis that
N:P, C:N, and C:P ratios hi lake water and watershed
C, N, and P supply to lakes are significantly influenced
by landscape factors related to human land use activity,
forest ecosystem composition, and regional air pollution
gradients will be tested (see Figure 1 for conceptual
model).
3. Conduct a landscape characterization analysis
of features in the lake-watershed basins which, hi con-
junction with concurrent watershed-associated stream
and groundwater measurements, will be used to develop
proxy measures for expected relative C, N, and P supply
conditions for lake water.
4. Establish zooplankton assemblage-derived var-
iables that reflect risks to ecosystem function, struct-
ure, or human health as a function of the elemental sup-
ply gradient.
5. Conduct a sensitivity analysis of the zooplank-
ton metrics using existing and extensive temporal and
spatial datasets. These analyses will establish statistical
confidence and power to detect change for measures of
lake integrity.
In the first field season, a total of 36 visits to 26
lakes in the Adirondack Mountain and Saint Lawrence
Valley regions of New York and Vermont were con-
ducted. Fifteen of 75 selected tributary riparian zone
locations were sampled for field characterizations that
included vegetation survey, leaf-litter sampling, and soil
characterization and sampling.
The foundation data layers (topographic factors,
land cover/land use, soils, hydrography, climate, and
atmospheric deposition) of a regional CIS that will be
used for the characterization of the study watersheds
have been acquired, converted to grid data, reprojected,
.and resampled as necessary. The generation of riparian
zone overlays is in progress.
The EMAP dataset was analyzed for sensitivity of
various zooplankton metrics to distinguish differences
among lakes on both regionwide and subregional scales.
Sensitivity of metrices increased in one or more sub-
regions relative to regionwide estimates for all metrics
tested. Richness and population metrics of calanoid
copepods and large cladocerans had 80-90 percent of
then: variance associated with the lake component rel-
ative to variance from temporal and interaction sources.
Metric performance improved at spatial scales, reflecting
underlying historical biogeographic associations.
Patterns hi the structure of zooplankton assem-
lages of the EMAP lakes in relation to effects of pre-
dators and nutrient treatments in experimental meso-
cosms also were evaluated. Fish predation reduced the
body sizes of comprising taxa in the mesocosms, but N
and P addition determined whether the assemblages were
calanoid (high N:P ratio), or cladoceran and rotifer (low
N:P) dominant. Nutrient effects were immediate and
substantial.
The factors tested in the experiments are identical
to those identified in principal component analysis (PC A)
of natural lakes—total N, total P, N:P ratio, and young
fish. The results from these independent but mutually
supporting studies greatly strengthens the underlying
importance of nutrient ratios in controlling structure of
zooplankton assemblages and further strengthens the
foundations for nutrient-based zooplankton indicators.
51
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200
Calanoid & Omnivorous
c Cladocerans
£400
o , ; 75
c
o
Large & Small
Cladocerans,
Copepods, Rotifers
40-50
Cyciopoids-
Microzooplankton
o
5-20
Microzooplankton
1.0
Oligotrophy
Mesotrophy
Eutrophy
Figure 1. Plot of N and P spate showing how the zooplankton-NP ratio indicator is interpreted within different classes of terrestrial
watershed vegetation cover and land use. Open circles indicate lakes having Chi a values > 12 ng/L . These systems are
associated with P values > than 1 nmole/L and reflect riparian, urban, or agricultural P pollution. Dashed line in upper right
panel delimits the N, P space (50 jimoles L-l of N and 1 mole/L P) that characterizes the majority of Northeastern lakes. Broken
vertical lines in lower panel delimit lake trophic state based on total-P criteria: 0 = oligotrophic lakes (< 10 ^ig/L total P),
M = mesotrophic lakes (> 10 < 30 ng/L total P), and E = eutrophic lakes (> 30 fig/L total P). Note all zooplankton groups may
occur within each trophic state category and that total biimiass will increase wnh increasing P.
52
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Effects of Interacting Stressors
in Agricultural Ecosystems: Mesocosm and Field
Evaluation of Multilevel Indicators of Wetland Responses
Stephen T. Threlkeld
Department of Biology, University of Mississippi, University, MS
The primary objective is to evaluate indicators of
molecular, cellular, population, community, and ecosys-
tem responses to multiple, potentially interacting, natural
and anthropogenic stressors at different spatial and tem-
poral scales in agricultural wetlands.
The indicators are chosen to represent a selection
of mechanism-based and system-level integrative char-
acteristics that might be amenable to cost-effective rou-
tine monitoring. The null hypothesis is that indicators
that effectively characterize ecosystem responses to sin-
gle stressors are also scale- and interaction-independent
(i.e., useful even when there are multiple, interacting
stressors with diverse operational scales). In the pro-
ject's first year, attention was focused on the devel-
opment of a Geographic Information Systems (GIS) data-
base for the watershed of the Little Tallahatchie River hi
northern Mississippi. Physical features, agricultural land
use practices, and potential pesticide loads in the basin
have been characterized.
These datasets were incorporated into the GIS data-
base that will be used to randomly select approximately
150 field sites from about 18,000 potential sampling
areas (ponds, oxbow lakes, impoundments, and inter-
mittent stream segments) that have been identified.
Physical features have been mapped from 1:100,000 and
1:24,000 from U.S. Geological Survey data. Features
not visible on 1:100,000 data (already in digital form)
were digitized from 1:24,000 maps (e.g., intermittent
streams, small ponds).
Land use practices have been characterized from
data obtained through county Farm Services Agency of-
fices that maintain separate records for each farm in
their jurisdiction. For each square mile of the study
area, crop data has been gathered for all major and
minor crops planted in 1998.
Potential pesticide loads were calculated for the
following chemicals: chlorpyrifos, atrazine, methyl para-
thion, and monosodium methane arsonate—which com-
prise the majority of pesticides applied to major crops in
the region (soybeans, cotton, corn, broadcast grains)—by
multiplying the chemical application rate (amount of
chemical per acre) by the number of acres in a square
mile planted in a crop for which a particular chemical is
used.
Application rates were obtained from resource
information at the Lafayette County (Mississippi) Ex-
tension Office. This information incorporates data from
companies that produce the chemicals as well as data
pertinent to Mississippi that may affect application (e.g.,
weather patterns, soil types). From these chemical dis-
tributions, watershed areas have been identified that are
most likely to have wetlands with agrichemical load-
ings typical of the single or multiple stressor treatments
planned for the mesocosm experiment (see Figure 1),
which is to commence in May 2000.
Many of the chemical combinations to be used in
the mesocosm experiment (even the four chemical com-
binations) are represented in the GIS by multiple sections
with numerous ponds that can be used for field eval-
uation. In the first year, the analytical and sampling pro-
tocols to be used in the upcoming mesocosm experiment
have been further developed.
53
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Pesticide Loading Combinations for the.
Little Tallahatchie Drainage Basin
Mississippi, USA
Values represent combinatioins ot high loading only!!
M£TP.CPF.»KJATRI15)
METP.CPf.sodUSMAlS)
AIKIGH(3)
AJ. but KMT V?UM OTty
BOTO, bwwxhattract 1
2 0 2 4 6 8 10 12 14 16 18 20 Miles
Figure 1. Pesticide loading combinations for the Little Tallahatchie River basin. Different shading patterns identify square mile sections of the drainage basin that are
characterized by chemical loadings (single and multiple pesticide combinations) that also will be used in the mesocosm experiment.
-------
Index of Authors
Aber, J., 19
Anderson, S., 3
Beasley, V., 21
Brooks, R.P., 37, 38
Dick, R.P., 39
Dugdale, R.C.,40
Field, K.G., 5
Fong.P.,42
Goldstein, A., 23
Grover, J.P., 44
Hallock, P., 46
Johnson, L., 24
Krane, D.E., 7
Leigh, D.S., 25
Michaels, H.J., 9
Newman, M.C., 11
Oris, J.T., 12
Patten, D.T., 26
Reddy, K.R., 48
Richards, C., 27
Rogstad, S., 14
Seagle, S.W., 28
Spacie, A., 49
Stemberger, R.S., 51
Threlkeld, S.T.,53
Turner, M.G., 30
West, N.E., 32
55
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