United States
Environmental Protection
Agency
Office of
Radiation Programs
Washington, D.C. 20460
EPA 520/1-85-016
February 1986
Radiation
x>EPA
Effects of Radiation on
Aquatic Organisms and
Radiobiological Methodologies
for Effects Assessment
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EPA520/1-85-OT6
Effects of Radiation on Aquatic
Organisms and Radiobiological
Methodologies for Effects Assessment
S.L Anderson
F.L Harrison
Work Performed Under
IAG DW 89930414-01
February 1986
U.S. Environmental Protection Agency
Office of Radiation Programs
Washington, D.C. 20460
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FOREWORD
biological effects to marine organisms.
This report, prepared by Lawrence Liverraore National
Laboratory, summarizes the literature on effects to aquatic
organisms from acute and chronic exposure to ionizing radiation
fol Sor?lllt?? pa?nophysiology, reproduction, development and
genetic effects. Methodologies for the study of radiobiological
effect are discussed, and recommendations for future research
Ire provfSed As such, these "commendations reflect only the
views and opinions of the authors and have not beenT.af°P*^r7
the Agency as necessary to meet any ocean disposal regulatory
'objectives.
The Office of Radiation Programs (ORP) will use this report
to determine the direction of any future effects testing Program.
The methodologies described in the report may also be useful to
those studying pollutants other than radiation.
The Agency invites all readers of this report to send any
comments or suggestions to Mr. David E. Janes, Director, Analysis
and Support Division..Office of Radiation Programs CANR-461),
Environmental Protection Agency, Washington, D.C. Z04b0.
^^
ers, Acting Dire<
Sheldon Meyers, Acting Director
Office of Radiation Programs (ANR-458)
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ABSTRACT
This report reviews the results of studies on biological effects of
ionizing radiation on aquatic organisms on the bases of biological effects
(mortality, pathophysiology, reproduction, development, and genetics) and
radiation regime (acute and chronic). The extensive data base is summarized
in tables, but only the studies demonstrating effects from exposure to low
levels of radiation are discussed critically because mortality as an
endpoint is unsuitable for assessing the sublethal effects that may be
expected from ocean disposal of low-level radioactive waste.
Results of studies of the effect of acute radiation on mortality
indicate that the range of lethal levels in adult fish is 375 to 55,000 rad
and that mortality of fish embryos has been demonstrated as low as 16 R.
For invertebrates, lethal doses range from 210 rad to above 50,000 rad.' For
both fish and invertebrates, U>50 observation times are not standardized,
and results may be modifed by other factors such as lifestage and temperature.
Research using pathophysiological endpoints on fish and invertebrates
has mainly served to characterize lethal radiation syndromes. Further
research in this area should focus on immune responses.
One of the few areas in aquatic radiobiology for which there are data
on the effects of chronic, low-dose irradiation regimes is the study of
reproductive effects in fish. Effects have been shewn from 0.59 rad/d to
12 rad/d. These data indicate that the cellular sensitivity of some fishes
is not less than that in mammals.
Developmental defects in embryos and embryo death induced by exposure
below 100 R have been demonstrated for three species of fish. The lowest
effect-level was 16 R. Much of the existing data on chronic irradiation of
fish embryos is not useful because errors exist in the dosimetry for studies
using radionuclides in the test water. However, an increase in opercular
defects in Oncorhynchus tshawytscha was found at 0.5 R/d. Little information
exists on developmental effects in invertebrates. However, decreased
hatching rate of the snail Physa hyterostropha was observed at 10 rad/h.
iii
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Genotoxic effects in fish and invertebrates have been observed at low
doses. Increased frequency of chromosomal aberrations in cultured fish
cells has been observed after 50 R exposures. In the marine worm Neanthes
arenaceodentata, significant increases in sister-chromatid exchange and
frequency of chromosomal aberrations have been observed at 60 rad and
200 rad. An increased frequency of major eye malformations in trout embryos
occurred after sperm were exposed to 25 rad. The data on genotoxic effects
further demonstrate that the cellular-level sensitivity of certain aquatic
organisms may not be significantly different from that of mammals.
Because the information on effects of low levels of radiation on
aquatic organisms is limited, information on terrestrial animals is also
discussed to provide greater perspective on potential effects at the dose
rates in question. In addition, the limited information on effects on
populations and communities from chronic irradiation is reviewed.
We conclude that no data are available demonstrating that significant
detrimental effects on aquatic organisms occur at the maximal levels of
contamination (0.240 rad/d at the end of the pipeline at Windscale) reported
in the ocean due to waste-disposal activity. However, there are not
sufficient data on aquatic organisms in the literature to determine
threshold dose rates at which important radiobiological effects would be
observed. Furthermore, the certainty of any prediction would be limited by
our incomplete understanding of the effects of potentially modifying
factors, especially temperature, on the responses of the organisms to low
dose rates or total doses.
Important gaps in knowledge needed by regulators on the biological
effects of ionizing radiation on aquatic organisms were identified, as
follows: (1) few reliable studies at chronic and low dose rates have been
conducted, (2) few studies have been done on marine organisms, (3) very few
studies have been done on marine invertebrates, (4) information on modifying
factors such as temperature, species specificity, and cell kinetics is very
scarce, and (5) the long-term effects of low-level radiation on fertility in
fish and invertebrates have not been adequately characterized.
General recommendations for future research include additional studies
on marine invertebrates and reproductive and genotoxic endpoints in marine
fishes and invertebrates. Further studies are also needed on the effects of
modifying environmental factors, such as temperature. The additional
methodologies for studying radiobiological effects include quantification
of DNA single-strand breaks, unscheduled DNA synthesis, and premature
chromosome condensation. Possible monitoring approaches that were given
include using biological dosimetry such as .quantification of chromosomal
aberrations in a tissue that integrates effects and using the radiation-
induced electron paramagnetic resonance signals in calcified tissues.
iv
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CONTENTS
Abstract . . . . . iii
List of Tables vii
List of Figures -jx
Introduction ........ 1
Stages in the Development and Modification of Radiation Injury 3
Development of Radiation Injury 4
Modification of Radiation Effects 6
Principles of Human Radiation Protection and Their
Relation to Aquatic Toxicology 9
Effects on Aquatic Animals of Acute and Chronic Exposure to
Ionizing Radiation • 11
Mortality ....... 11
Criteria for Establishing a Lethal Dose ......... 11
Mortality of Fish from Acute Radiation Exposure .... .12
Mortality of Invertebrates from Acute Radiation Exposure ..... 20
Mortality of Fish and Invertebrates from Chronic
Radiation Exposure 24
Conclusions on Mortality Data 24
Pathophysiology 25
Pathophysiological Effects in Fish from Acute
Radiation Exposure '.-.-.'•.. 25
Pathophysiological Effects in Invertebrates from Acute
Radiation Exposure 32
Pathophysiological Effects in Fish and Invertebrates from
Chronic Radiation Exposure 32
Conclusions on Pathophysiology Data 33
Reproduction . 35
Reproductive Effects in Fish from Acute Radiation Exposure .... 35
Reproductive Effects in Invertebrates from"Acute Radiation
Exposure 37
Reproductive Effects in Fish and Invertebrates from Chronic
Radiation Exposure 39
Conclusions on Reproduction Data 43
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Development
Developmental Effects from Acute and Chronic
Irradiation of Fish ... > ... « « 44
Developmental Effects from Acute and Chronic
Irradiation of Invertebrates ........ . ..... . . . • 50
Conclusions on Development Data • 52
Genetics 53
General Considerations in Aquatic Genotoxicity Research ...... 54
Induction of Chromosomal Aberrations in Fish ........ , . . 57
Induction of Mutations in Fish ...... 60
Induction of Chromosomal Aberrations in Invertebrates ....... 63
Induction of Mutations in Invertebrates . . 65
Conclusions on Genetics Data ... ............ • • ••« 66
Comparison of Radiation Effects on Aquatic Organisms to Those
on Terrestrial Organisms •••••• 5
Mortality and Pathophysiology ....... . . . . • • •'.• « « •'...• 67
fiQ
Reproduction °3
Development .-. . . . . . . . . . . . ; . .......... • « «
Genetics • 69
Low-Level Radiation Effects and Human Health ........... 73
Conclusions on Effects Levels 73
Effects on Populations and'Communities from Chronic Exposure _
to Ionizing Radiation 74
Value of Single-Species Toxicity Tests in the Evaluation of
Effects on Populations and CommunUies '4
Effects of Ionizing Radiation on. Populations of Aquatic
Organisms . * .
Provisional Dose Assessments for Deep-Sea Animals 81
Additional Methodologies for the Study o'f Radiobiological Effects ... 84
Monitoring Approaches . . . 85
Recommendations for Future Research on Biological
Effects and Biomonitoring Strategies b/
OQ
Acknowledgments ........... °3
References and Bibliography ' • 90
Appendix I. List of Major Review Articles :. 120
Appendix II. Reference Summary Table ............ 121
vv
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LIST OF TABLES
1. Induction of mortality in fish from exposure to acute levels
of radiation 13
2. Induction of mortality in invertebrates from exposure to acute
levels of radiation 21
3. Induction of mortality in fish from chronic exposure to
radiation. . ........'. . 25
4. Induction of mortality in invertebrates from chronic exposure
to radiation 26
5. Induction of pathophysiological changes in fish from exposure
to acute levels of radiation. ......... 27
6. Induction of pathophysiological changes in invertebrates from
exposure to acute levels of radiation. '; ...... 33
7. Induction of pathophysiological changes in fish from
chronic exposure to radiation 34
8. Induction of reproductive changes in fish from exposure to
acute levels of radiation
36
9. Induction of reproductive changes in invertebrates from
exposure to acute levels of radiation. . . 38
10. Induction of reproductive changes in fish from chronic
exposure to radiation
. 40
11. Induction of reproductive changes in invertebrates from
chronic exposure to radiation 41
12. Induction of developmental changes in fish from
exposure to acute levels of radiation 45
13. Induction of developmental changes in fish from chronic
exposure to radiation 43
14. Induction of developmental changes in invertebrates from
exposure to acute levels of radiation. ... 51
15. Induction of developmental changes in invertebrates from
chronic exposure to radiation 52
16. Induction of chromosomal aberrations in fish from
acute and chronic exposure to radiation 58
vii
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17. Induction of mutations in fish from acute and chronic
exposure to radiation 61
18. Induction of chromosomal aberrations in invertebrates from
acute and chronic exposure to radiation
21,
22,
19. Induction of mutations in invertebrates from acute exposure
to radiation ' <
20. Summary of btological effects observed at low dose rates and
low total doses in aquatic'organisms as compared to selected
data for terrestrial organisms . . . .
Induction 'of effects on populations of aquatic organisms
from chronic exposure to radiation
Estimates of the radiation dose rates (nSv/h) to benthic deep-
sea organisms from natural background and the peak dose rates
predicted from dumping low-level radioactive wastes
64
66
70
79
82
viii
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LIST OF FIGURES
2.
3.
4.
5.
6.
Stages in the development and modification of radiation
injury
Important endpoints in cytogenetic research and their
implications. ...... ........
Comparison of yields of diGentries and centric rings in
fish (A. splendens, A), toad (B. marinus, Q), and man
(H. sapiens Q) (Woodhead,, 19767"
Generalized effect levels for selected biological endpoints . . .
The relationship between error of effect-level estimates
and stages of investigation (Cairns, 1983)
Ambient toxicity correlation between Ceriodaphnia young
per female and ecological survey data (Mount et al., 1984)
5
56
59
68
75
77
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INTRODUCTION
Safe disposal of wastes,, whether nuclear or nonnuclear, is the ultimate
concern of effective waste management. Wastes presently discharged directly
into U.S. marine waters are conservatively estimated to exceed 50 million tons
per year. Of this amount, approximately 80% consists of dredged materials, 1055
consists of industrial wastes, 9% consists of sewage sludge, and the remaining
1% consists of miscellaneous wastes (Bierman et al_., 1984). Because the total
quantity of these wastes is increasing as the result of increases in
population, consideration is being given to alternative disposal strategies
and sites.
In the U.S., packaged, low-level, solid radioactive wastes were disposed
of off both the Atlantic and Pacific coasts (Joseph et_al_., 1971). Since the
onset of the nuclear age, radioactive wastes have been disposed of on land and
in the ocean. In addition, the testing of nuclear weapons has contributed
measurable quantities of radionuclides to the ocean. Although these practices
were discontinued by 1970, little effort was made until recently to determine
the subsequent fate and distribution of the radionuclides in these wastes.
Information now available indicates that some man-made radionuclides from
ocean disposal are present in the bottom sediments, but there is little or no
accumulation by organisms in the human food chain (Dyer, 1976; Noshkin et_ al_.,
1978).
When reviewing requests for permits, U.S. Environmental Protection Agency
(EPA) administrators are required to (1) determine that ocean "dumping will
not unreasonably degrade or endanger human health, welfare, or amenities, or
the marine environment, ecological systems, or economic potentialities" and
(2) establish regulations and criteria to implement a permit program (Marine
Protection, Research, and Sanctuaries Act, 1972). In 1977, the EPA published
final regulations and criteria for ocean dumping. The regulations define the
types of environmental information that will be required by EPA to determine
criteria for ocean disposal. Evaluation of the potential environmental hazard
from a waste-disposal operation involves combining information on the
potential biological effects of wastes at given concentrations and knowledge
of the fate of the waste material in the receiving water. Fate must be
studied to predict the partitioning of a waste material in the environment and
to predict the area impacted by disposal operations. Biological effects must
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be studied to determine the contaminant concentrations constituting threshold-
effect levels (level below which lesions are repaired or not realized at the
organismal level and no detrimental effects observed).
A problem encountered in defining biological effects of radioactive waste
is that traditional bioassays that use mortality as an endpoint are unsuitable
for assessing the sublethal effects that may be expected from ocean disposal
of low-level radioactive waste. Most mortality tests are limited to
observations over a short time period from which long-term assessments of
impact are to be inferred. This extrapolation of short-term results to a
long-term impact is highly risky because the response elicited from high levels
of radiation is different from that to low levels of radiation. Furthermore,
many of the species that have been selected for standard testing are
inappropriate for evaluating the effects on biota indigenous to the sites
proposed for radioactive waste disposal. It is also difficult to determine
the uncertainty associated with estimates of effects from exposure to low
levels of radiation, because little is known about, how specific estimates are
modified by changes in experimental parameters such as temperature.
The purpose of this document is to summarize the biological effects of
radiation exposure on aquatic organisms and to describe state-of-the-art
methods for assessing those effects. Several comprehensive reviews of
published data on the effects of radiation on aquatic organisms have been
published (Polikarpov, 1966; Tempieton et ^1_., 1971; Chipman, 1972; Ophel,
1976; Blaylock and Trabalka, 1978; Egami and Ijiri, 1979; Woodhead, 1984).
Full bibliographic information on these review papers is given in Appendix I.
Consequently, it is not necessary to give a detailed account of the entire
literature. We will synthesize the conclusions on effect levels and
generalizations on approaches developed by these authors and provide data
relevant to the concern of the Office of Radiation Programs of the EPA, I.e.,
biological effects that might be expected from the disposal of low-level
radioactive wastes into marine environments.
The organization of the report is as follows. First, we discuss the
stages in development and modification of radiation injury and explain some
differences between the principles of human radiation protection and those for
the protection of aquatic life. Then, we discuss effects of acute and chronic
exposure to radiation on fishes and aquatic invertebrates. The biological
endpoints considered are mortality, pathophysiological changes, reproductive
effects, developmental effects, and genetic effects. The endpoints are
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discussed with respect to their sensitivity and factors that may modify the
response. For each endpoint9 selected results on terrestrial organisms are
provided for comparison of the relative sensitivities of terrestrial and
aquatic organisms. Tables provide representative data for each effect
category. Critical comments are made, where appropriate, only for the studies
demonstrating very low levels of observable effects. Next, population-level
studies are analyzed, and the value of single-species toxicity tests in the
evaluation of effects on populations and communities is discussed. Then,
additional methodologies for the study of radiobiological effects are
described, as are monitoring approaches. Finally, recommendations for future
research are presented. A table providing the kinds of information included
in a group of references reviewed for this report is given in Appendix II.
All of these articles are listed in the references, but not all are cited in
the text. Hence, the reference list is also a bibliography for wtrich
annotations are given in Appendix II.
Bec'ause evaluation of the biological effects of radiation exposure is our
primary goal, we have considered discussions of the fate of radionuclides in
the ocean to be beyond the scope of this work. Hence, discussion of radiation
regimes in the ocean is minimal, and representative dose rates are only given
for general comparison of effect levels. In a final analysis of effect
levels, biological responses to radioactivity from both internal and external
emitters and the relative biological effectiveness of the specific
radionuclides in the waste in question must be considered. Radiation units
used, in citing previously published work, are those of the author(s). In all
other instances, the international units, gray (1 Gy = 100 rad) and Sievert
(1 Sv = 100 rem), are used. No attempt has been made to formulate a stringent
definition of acute versus chronic exposure or high versus low dose rates.
Effects of radiation exposure on marine bacteria and plants are not discussed.
STAGES IN THE DEVELOPMENT AND MODIFICATION OF RADIATION INJURY
Before specific effects of radiation exposure in fishes and invertebrates
are discussed, we will describe and explain the stages in.the development of
radiation injury and the nature of radiation syndromes. The types of questions
that must be considered in an analysis of radiation effects are:
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• What biological molecules or structures are damaged by radiation?
• What types of injury occur, and how are they produced?
• What time interval is required for the effects of radiation to be
realized?
• What effects occur at high (potentially lethal) doses versus low
(sublethal) doses?
• At what levels of biological organization do different factors that
modify the effects of radiation act?
Because these topics are explained in detail in texts on radiation biology
such as Arena (1971), only a brief discussion of these topics are given here.
DEVELOPMENT OF RADIATION INJURY
The development of radiation injury occurs at three levels of biological
organization: the molecular level, the cellular level, and the organismal
level (Fig. 1). When an organism has been exposed to ionizing radiation,
energy absorbed in tissues induces atomic changes. These changes may occur
through ionization or excitation. Free radicals are produced through
ionization, and other types of molecular configurations are produced through
excitation. Consequently, molecular effects of ionizing radiation may occur
through direct ionization or excitation of an important molecule (e.g., DNA or
critical enzymes), or effects may be produced indirectly when the molecule of
interest receives energy by transfer from another molecule or is acted on by
free radicals (Arena, 1971). When molecular changes are induced in DNA,
single- and double-strand breaks result in such effects as mutation,
chromosomal breakage, chromosomal rearrangement, and potentially, point
mutations (Guerrero et al_., 1984). These effects may occur in somatic or germ
cells. When they occur in germ cells, effects are sometimes heritable. When
molecules other than DNA are damaged, cell death may also occur due to such
factors as membrane damage and enzymatic breakdown of cellular structures.
The types of damage realized at the cellular and organismal levels depend
on the absorbed dose. For mammals, the causes of death from acute radiation
exposure are described below. For massive doses of 100,000 rad (1000 Gy) or
more, death occurs during irradiation or immediately thereafter because of
inactivation of important biological molecules causing "molecular death."
Doses of approximately 10,000 rads (1000 Gy) cause death within a day or two
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Exposure to radiation
Energy1
absorption!
I
Ionized and electronically
excited molecules
Direct action
ioni / \ Indii
r \
Indirect action
Molecular changes!
(especially DNA lesions):
Somatic!
mutations'
X
Delayed somatic;
effects |
Genetic disease)
death
Death of organism;
or infertility
Modification of injury
development by:
RBE, dose rate
Oxygen
Temperature
DNA repair
Cell repopulation
Cell cycle effects
Figure 1. Stages in the development and modification of radiation injury.
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due to central nervous system damage. Between 1000 and 10,000 rads (10 and
100 Gy), death generally occurs in 3 to 5 d or less from damage to cells in
the gastrointestinal tract. Loss of these cells and resultant changes in
permeability of the gut cause severe imbalance of fluids and electrolytes and
bacteremia, which results in death. Doses of 300 to 900 rads (3-9 Gy) cause
cell death in blood-forming (hematopoietic) organs. When death occurs at
these doses, it is often linked to either secondary infections because of
decreased immunity or hemorrhage from loss of platelets (Casarett, 1968).
The major somatic and heritable effects of low-level radiation exposure
have been identified for mammals (NRC, 1980; UNSCEAR, 1977). The principal
effect of low-level radiation exposure on somatic tissue is cancer induction
in a variety of organs and tissues. Oocyte death (possibly from membrane
damage) and abnormal development of the embryo and fetus have also been
observed below 10 rad (0.1 Gy) (NRC, 1980; UNSCEAR, 1977). Concern over
effects on germ cells of low-level radiation exposure centers on induction of
increased frequency of heritable genetic diseases. This may occur from
mutation, chromosome breakage, chromosome rearrangement, and faulty segregation
of chromosomes at metaphase (aneuploidy). Effects such as these may require
from hours to years to become evident.
MODIFICATION OF RADIATION EFFECTS
Modification of the effects of radiation can occur at all stages presented
in Fig. 1. It is known that the relative biological effectiveness (RBE) of a
radiation type has a large effect on the amount of damage produced. Concerning
the type of radiation and energy, the most important property affecting RBE is
the linear energy transfer (LET). LET is defined as the amount of energy (in
keV) dissipated by an ionizing particle per micrometer of path (Arena, 1971).
Because of differences in ionization density, the following general hierarchy
of biological'effectiveness has been established: alpha particles > protons =
neutrons > beta particles = gamma rays (ICRP, 1979).
Many responses to radiation exposure are modified by dose rate; a higher
total dose is required at low dose rates to produce the same effect observed
at high dose rates. This factor has been discussed in detail by Bedford
(1982). Because dose rate is an important modifying factor, in this review we
will consider chronic, low-level radiation effects separately from those
produced as a result of acute exposure.
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DMA repair operates at the molecular level and is an important modifier
of radiation response. This factor may have particular significance at low
dose rates. It'is widely debated whether threshold dose rates of radiation
exist. However, some repair pathways are known to be error prone and, thus,
repair of a lesion does not necessarily imply return to status quo (Schendel,
1981). Furthermore, it is possible that DNA repair enzymes are induced during
low-level, chronic exposures (Tuschl et^al_., 1983). The relative abilities of
eukaryotic organisms to repair DNA are unknown. Little is known about DNA
repair in aquatic invertebrates (Ejima and Shiroya, 1982), but certain
DNA-repair pathways have been examined in fish (Achey et jf[., 1979; Ishikawa
et al_., 1978; Mano e^ al_., 1980, 1982; Mitani, 1983; Mitani and Egami, 1982;
Mitani et jil_., 1982; Regan and Cook, 1967; Regan j2t jil_., 1983; Walton et al.,
1983; Woodhead and Achey, 1979; Woodhead et^ al_., 1978).
Tissue-oxygen concentration is one of the most widely studied modifiers
of radiation effects (McNally, 1982). It has been shown that the radio-
resistance of most systems increases approximately two to fourfold under anoxic
conditions. These modifying effects are probably produced at the molecular
level. Theories proposed to explain the relationship between oxygen tension
and radiosensitivity are that fewer peroxyl and hydroperoxyl radicals are
formed at low oxygen tensions and that oxygen may inhibit the DNA repair
process (Casarett, 1968). This factor has not been studied with any aquatic
organism despite its demonstrated importance in mammalian systems and its
possible relevance to deep-ocean disposal of radioactive waste. Oxygen
tensions vary greatly between deepwater sites, and this could modify many
effects such as gamete death and chromosomal damage.
The effect of temperature on the modification of radiation response occurs
at the molecular and cellular levels. Cold temperature is known to slow the
development of lethal biochemical lesions and lengthen cell-cycle times, but
it may also slow repair processes. The effect of temperature are discussed in
more detail in the following section on mortality.
Radiation response is known to depend on the stage in cycling of the cell
and on the rate at which cycling is occurring. Cell survival is known to be
affected by the stage that is irradiated; S phase, the period in which DNA
synthesis occurs, generally is the most radioresistant (McNally, 1982). Types
and amounts of chromosomal aberrations induced by radiation also vary with
cell-cycle stage at radiation (Wolff, 1968). Moreover, one of the basic tenets
of radiation biology is the law of Bergonie and Tribondeau, which states that,
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in general, the radiosensitivity of cells is directly proportional to their
rate of division and inversely proportional to their degree of differentiation.
In other words, cell killing occurs selectively in cells that are rapidly
dividing and not highly .differentiated. This is because one of the prime
mechanisms for cell killing is the induction of chromosomal aberrations that
cause changes 'in the genetic complement and in gene expression in the damaged
cell. Cell death from aberrations is also due to mechanical interference of
the migration of the chromosomes during cell division. Many of these effects
will not be realized until the cell divides, and thus cells that are cycling
are more sensitive than those that are not. Furthermore, the main reason that
relatively less cell killing occurs in highly differentiated cells is that they
rarely or never divide. These principles can be applied to radiation effects
• " :
on aquatic organisms, and they imply that life stage and growth rate of an
organism may have a tremendous effect on its radiosensitivity.
When cells are damaged by radiation, they may be replaced by normal cells.
This effect is termed cell repopulation, and it is an important modifier of the
apparent radiation response at the organismal level. The practical extension
of this principle is that when an organism or tissue has the ability to replace
damaged cells, effects of radiation at the cellular level will not be realized
at the organismal level unless they are in a specialized tissue that cannot be
repopulated or unless repopulation occurs slowly. Therefore, the amount of
tissue specialization in an organism and/or the ability of tissues to
differentiate and dedifferentiate may modify radiation response at the
organismal level.
Finally, a distinction on the radiosensitivity of dividing and non-
dividing cells must be made. Rapidly dividing cells are most sensitive to
acute doses of radiation because of increased probability of cell death.
However, slowly dividing cells may be most sensitive to chronic irradiation
because they may accumulate chromosomal damage from integration of the dose.
Thus, to assess effects in cells exposed to low, chronic doses of radiation for
dosimetry or monitoring purposes, it is best to select a non-dividing cell
system and then induce the cells to divide so that the damage can be assessed.
Effects on DNA in non-dividing cells can also be studied using methods that do
not require analysis of metaphase cells, such as quantification of unscheduled
DNA synthesis.
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PRINCIPLES OF HUMAN RADIATION PROTECTION AND THEIR RELATION TO AQUATIC
TOXICOLOGY
Radiation standards to protect humans were developed for different
concerns than those to protect aquatic life. The primary concern for human
populations was to protect individuals, their progeny, and mankind as a whole
against cancer and genetic diseases. The primary concern for aquatic life was
to protect ecosystems and their economic potential by maintaining populations
of the species indigenous to those ecosystems. Some basic principles of human
radiation protection are presented here, and they are discussed as they relate
to the approaches used to protect aquatic organisms. The intention of this
discussion is to provide a link between the human-radiation-biology community
and the aquatic-toxicology community.
In human radiation protection, a distinction is made between stochastic
and non-stochastic effects. These are defined as follows (ICRP, 1979):
"Stochastic" effects are those for which the probability of an
effect occurring, rather than its severity, is regarded as a
function of dose, without threshold. "Non-stochastic" effects are
those for which the severity of the effect varies with the dose, and
for which a threshold may therefore occur. At the dose range
involved in radiation protection, hereditary effects are regarded as
being stochastic. Some somatic effects are stochastic; of these,
carcinogenesis is considered to be the chief somatic risk of
irradiation at low doses and therefore the main problem in radiation
protection.
The goals of human radiation protection, are (1) to prevent non-stochastic
effects by setting limits at sufficiently low values so that no threshold dose
would be reached and (2) to limit the probability of stochastic effects by
keeping all justifiable exposures as low as is reasonably achievable;
achievement of the latter can be assisted through risk assessment.
Effects on health may be caused by stochastic and non-stochastic effects
in exposed individuals and stochastic effects in subsequent generations. The
detriment in a population is the expectation of harm based on the probability
and severity of the effect. The detriment to health, G, in a group of P
persons is given by
G = P I Pi ^
i .
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where p. is the probability of suffering the effect i, and g.. is a
weighting factor for the severity of the effect (ICRP, 1977).
To predict the probability of suffering a stochastic effect, linear
extrapolation is used, based on a cautious assumption of proportionality. For
non-stochastic effects, the probability of suffering a given effect is
determined by the shape of dose-response curves and threshold doses. The fact
that many effects are age and tissue specific is also considered, as are the
characteristics of the exposure.
In aquatic toxicology, little is known about frequencies and significance
of stochastic effects such as cancer induction and hereditary change.
Information to protect aquatic ecosystems is obtained by examining primarily
non-stochastic effects. Bioassays to assess short-term changes in mortality
rate in different life-history stages or changes in reproductive success are
performed. In most cases, the shape of dose-response curves for specific
effects have not been well characterized. Consequently, it is not unusual for
those in the regulatory community to estimate application factors by such
methods as the formulation of acute:chronic ratios (Roop and Hunsaker, 1985).
Hence, protection of aquatic life is usually attempted by combining results of
standard bioassays with estimates of necessary safety factors. Biomonitoring
is sometimes implemented to determine the adequacy of predictions formulated
from toxicity testing.
Because the assumption of linearity used in human radiation protection
for stochastic effects renders the term "No Observable Effect Level (NOEL)"
used in aquatic toxicology unacceptable, and because we know so little about
the shapes of dose-response curves for relevant endpoints, we refer to low-
effect levels in this report. In a few cases when general aquatic toxicology
is discussed, we use the term NOEL as it would be used in that discipline. The
pathophysiologic, reproductive, and developmental effects discussed in this
report are all considered non-stochastic effects. Only a few studies discussed
in the section on genetic effects provide information on stochastic effects.
10
-------
EFFECTS ON AQUATIC ANIMALS OF ACUTE AND CHRONIC
EXPOSURE TO IONIZING RADIATION
MORTALITY
In aquatic toxicology, acute lethal levels of toxicants are usually
determined for three purposes. First, they are used to determine relative
sensitivities of organisms to lethal effects of a toxic agent. Second, they
are used to specify maximum levels of a toxicant permitted for short times in
a given environment. Third, they are used to set permissible chronic-exposure
levels. The third purpose is addressed in this section. Lethal doses of
acute ionizing radiation have been determined for a variety of fishes and
aquatic invertebrates. However, information on lethality alone is of limited
use in estimating biological effect levels of chronic, low-level, ionizing
radiation. Reasons for this conclusion are given in this and subsequent
sections.
When lethal levels of a toxicant are established for the purpose of
setting maximum chronic exposure levels, safety factors termed "application
factors" are used to set a "safe" value below the acute lethality limit..
Determination of an appropriate application factor is often subjective.
Therefore, it is important to understand the role of factors that modify the
level at which mortality will be observed so that an indication of the
potential error associated with a predicted safe value can be assessed.
In the section that follows, the criteria for determining a lethal dose
are discussed. Next, representative mortality data for various species of
aquatic invertebrates and fishes are presented. These are discussed with
regard to specifes specificity and other factors that may modify the levels at
which lethal effects are observed.
Criteria for Establishing a Lethal Dose
The LD5Q is the dose that kills 50% of the organisms tested in a
specified amount of time. In acute radiation studies, the dose is given over
a short time period of seconds or minutes, and an observation period is
specified. In mammalian studies, the standard observation time is 30 or
60 d. Therefore, a lethal dose criterion termed the 1-050/30 or LD50/60 is
established.
11
-------
In poikilotherms (animals, such as fishes and invertebrates, that do not
maintain a constant internal body temperature), cell-cycle times are generally
much longer and more variable than in mammals. This means that certain types
of radiation damage will be realized more slowly in poikilotherms than in
mammals. This does not necessarily mean that the cells involved are more
radioresistant. Consequently, a 30- to 60-d observation period is generally
inadequate. However, there is no standard time period that is suitable for all
poikilotherms, and this accounts for the large differences in observation
times for the experimental data reported.
Mortality of Fish from Acute Radiation Exposure
Results given in Table 1 demonstrate that the range of lethal levels for
adult fish is 375 to 55,000 rad and that 50% mortality of fish embryos has been
demonstrated as low as 16 R. Thus, the range of existing data on fish LDgQ
is over 3 orders of magnitude. Modifying factors that have been examined
include temperature, salinity, lifestage, RBE, dose rate, and species
specificity. However, the role of many modifying factors is still poorly
defined, and few studies on marine fish exist. Data provided in Table 1
represent the majority of the information in the literature on fish mortality
from acute ionizing radiation.
One of'the most important factors affecting ID™ values is the experiment
duration. It has already been mentioned that 30 d is inadequate for most
poikilotherms. However, most of the studies in the literature (Table 1) were
terminated at 30 d. Shechmeister et_ al_. (1962) demonstrated a dose-dependent
decrease in lifespan of the goldfish Carassius auratus from 100 to
10,000 rad. These results demonstrate that lethal effects of acute irradiation
can be observed at increasingly lower doses, the longer the organism is
observed.
In five studies cited in Table 1 (Blaylock and Mitchell, 1969; Egami,
1969a; Etoh and Egami, 1967; Hyodo, 1965a; Lockner et_al_-» 1972), the role of
temperature in modifying the lethal response to radiation was investigated.
These authors concur that low temperature slows the rate of development of
lethal lesions and thus postpones death. However, if the observation period
is sufficiently long, increased death rates from low doses may still be
detected. A finding of particular interest is that low temperature may negate
dose-rate effects; this implies that accumulation or integration of effects' at
12
-------
Table 1. Induction of mortality in fish from exposure to acute levels of radiation.
Lethal dose3
(% mortality, days)
16 R
(50%, 150 d)
58 R
(50%, 55 d)
90 rad
(50%, blastula to
metamorphosis)
300-500 R
(50%, 55 d)
375 r ad-neutron
(50%, 30 d)
1283 rad-x ray
(50%, 30 d)
1000 rad
(50%, 60 d)
2500 rad
(50%, 30 d)
1020 rad (25 ppt)e
1540 rad (15 ppt)
2050 rad (5 ppt)
Organism/1 if estage
Oncorhynchus kisutchc
(silver salmon)/l-cell
stage
Salmo gairdneriic
(rainbow trout)/l-cell
stage embryos
A
Pleuronectes platessa"
(plaice)/embryos
Salmo gairdneriic
(rainbow trout)/32-cell
to late eye embryos
Carassius auratusc
(goldf1sh)/adults
Carassius auratus0
(goldfish)/adults
Fundulus heteroclitusd
(mummichog)/adults
Radiation regime
x ray
12-2400 R
x ray
25-2570 R
x ray
30-150 rad
x ray
25-2570 R
Fast neutrons, 2 MeV
40 r ad/mi n
250-3000 rad
x ray
475-1663 rad
x ray
100-10,000 rad
60rn
Co
365 r ad/mi n
500-2000 rad
Comments Reference
Bonham and Welantier,
1%3
One-cell or blastodisc stage most Welander, 1954
sensitive
LD,.n at metamorphosis for plaice Ward et al., 1971
OU r r ___ __ i
larvae irradiated at blastula stage
'.•'•-
Irregular relationship of sensitivity Welander, 1954
and increased dose
Relative biological effectiveness (RBE) Etoh et al., 1974
discussed
Dose dependent decrease in 1 if espan Shechmeister
observed from 100-10,000 rad et.al., 1962
Increased sensitivity with Increased Angelovic et al.,
salinity and temperature 196.9
(50%» 30 d)
-------
Table 1. (Continued.)
Lethal dose3
(% mortality, days)
1050 R
(50*. 30 d)
1120 R
(50*. 30 d)
1200 rad
(5035, 30 d, 18°C)
55,000 rad
(5055, 30 d, 10°C)
1450 R
(505!, 30 d)
2350 R
(5055, 30 d)
2500 R
(5055, 60 d)
Organism/lifestage
Micropogon undulatus--
( — )/juveniles or
postlarvae
Fundulus heteroclitus
(mummichogj/juveniles or
postlarvae
Tinea vulgaris0
(tench)/adults
Mugil cephalus0
(mullet)/juveniles or
postlarvae
Poecilia reticulatac
(guppy)/l-week old
Oncorhynchus tschawytscha0
(chinook salmon)/embryos
Radiation regime
60Co
60Co
60Co
2 rad/s (18°C) and
250 rad/s (10°C)
0-60,000 rad
60Co
x ray
x ray
250-10,000 R
Comments Reference
White and Angelovic,
1966 (in Chipman, Is72)
i
White and Angelovic,
1966 (in Chipman, Iy72)
Note different dose rates and short Lockner et ^1_.,
time period (30 d); temperature effects 1972
observed
White and Angelovic,
1966 (in Chipman, Ia7ii)
Erickson, 1973
No significant mortality at 30 d; Welander et al.,
however, 5055 mortality was obtained 1948
in 40 d in all doses >_ 2500 R;
significant increases in mortality
occurred at 1000 R
-------
Table 1. (Continued.)
en
Lethal dose
(% mortality, days)
3000 R
(50%, 30 d)
3500 R
(50%, 30 d)
3699 rad
(50%, 30 d)
5000 R
(100%, 7 d)
5550 R
(50%, 30 d)
8000 rad
(100%, 30 d, 22°C)
Organism/lifestage Radiation regimeb
Lagodon rhomboidesc 60Co
(pinfish)/juveniles or
postlarvae
Eucinostomus sp.d 60Co
( "— )/juveniles or
postlarvae
Gambusia affinis affinisc 60Co
(mosquitofish)/adults ' 8.2 rad/sec
1500-6000 rad
Lagodon rhomboides0 Gamma ravs
(pinfish)/adults 385 R/h
5000 R
Paralichthys lethostigmad 60Co
(plaice)/juveniles or
postlarvae
Carassius auratusc x rav
(goldfish)/adults 0, 8000 rad
Comments
'
LD50/50 was 20°'° R
Tests at 20, 25, and 30°C showed that
sensitivity increased with increased
temperature; no evidence of increased
radioresistance of population exposed
chronically for 10 generations to
<10 rad/d
Changes'- in cellular components of
blood described
LD50/50 was 250° R
Low temperature inhibits the development
of intestinal damage, which would lead
Reference
White and Angelovic,
1966 (in Chipman, 1972)
'
White and Angelovic,
1966 (in Tempi eton
jEit^K, 1971)
Blaylock and
Mitchell, 1969
-Engel et al.,
1967
White and Angelovic,
1966 (in Templeton
£t jij_. , 1971)
Hyodo, 19b5a
8000 rad
83%, 30 d, 4°C)
to death
-------
Table 1. (Continued.)
Lethal dosea'
mortality, days) Organism/lifestage
Radiation regime
Comments
Reference
10,000 R
(41 .7%, 30 d)
Carassius auratusc x ray
(goldfish)/adults 1000-16,000 R
Mortality rate following partial-body
irradiation depended on exposure and
damage of the directly irradiated part,
as well as absorbed dose in the tissue
as a whole
Etoh et al., 1968
Pimephales promelasc
(fathead minnow)/
adults
Oryzias latipes
(medaka)/adults
Oryzias latipes0
(medaka)/adults
Oryzias latipesc
(tnedaka)/adults
60Co
200-3000 rad at 2.62 rad/min
600-6000 rad at 37.8 rad/min
2300-24,000 rad at 1730 rad/min
x ray
Single dose 100,000 R or
2 X 50,000 R. Single dose 4000 R
or 2 X 2000 R and 2000-4000 R
fractionated
x ray
1000-31,000 R (includes
numerous fractionations)
60Co
14 R/min-1000-4000 R
0.6, 0.8, 5.4, 10.5, and
105 R/min~3600-4800 R
Time to 5055 mortality:
3000 rad (2.62 rad/min) - 760 d
(37.8 rad/min) - 28 d
(1730 rad/min) - 17 d
If interval between fractionations is
3 d or longer, mortality from
radiation was reduced
Longer intervals between fractionations
diminished the effect of irradiation;
decreased effects due to fractionation
did not occur at low temperatures
Dose-rate effect observed but negated
at low temperature (6°C) (0% survival
at 3000-6000 R over 4 d at 6°C vs
80% at 25°C)
Cheeet.al_., 1979
Egami and Etoh,
1966
Etoh and Egami,
1967
Etoh and Egami,
1967
-------
Table 1. (Continued.)
Lethal dosea
(% mortality, days) Organism/1 ifestage
Radiation regime
Comments
Reference
Oryzias latipes0
(medaka)/adults
x ray
Many split doses
When 3000-R dose was split in half with
3 d between doses, 10% (24°C) and 40%
(15°C) mortality were observed at 30 d,
vs 80% (24°C and 15°C) mortality when
the dose was delivered continuously
Egami, 1969a
that caused • specified percent mortamy in
Radiation regime is presented as source, dose rate, and total dose; a dashed line indicates information was not available. For x-ray data a dose
rate is not given; factors important in determining x-ray dose rate are voltage, target material, .filtration, tube current, and target-to-object
c d1Stance. Units are those used by the author; R is the abbreviation for roentgen.
d Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
Organism is exclusively marine.
6 " •
ppt is the abbreviation for parts per thousand.
-------
low dose rates may occur (Etoh and Egami, 1967). Possible negation of dose-
rate effects may introduce large uncertainties in predictions of biological
effect levels in organisms in the deep ocean from data available on coastal
organisms. Accumulation of chromosomal damage from chronic exposure could
result in unusually high effects at low dose rates, but slowing of lesion
development and cell cycling due to cold temperature could impart tremendous
radioresistance. It is not known what the relative importance of these factors
would be.
Effect of low temperature on radiation injury has been examined in animals
other than fishes and aquatic invertebrates because biomedical researchers
have been interested in the role of metabolic rate on the development of
radiation-induced, molecular lesions. Some early findings were summarized in
Bacq and Alexander (1961), as follows:
If frogs irradiated with 3000 to 6000 r at 23°C are cooled to
5°C, 80 to 90 percent of the animals survive for more than 3 to
4 months after irradiation, whereas controls kept at 23°C die in 3
to 6 weeks. The lesion in the cooled animals is latent and it
they are warmed after 60 to 130 d they die. No lesion is found in
the ovarian ova of amphibians 12 d after total irradiation with
3000 to 5000 r if the animals are kept at 5°C, but if they are
kept at 22°C after exposure to 3000 r, all the ova are affected.
If the chilled and irradiated animals are warmed after 12 d the
lesions appear very quickly.
Lamarque and Gros irradiated the eggs of the silk worm (Bombyx
mori) and kept them in the cold; when they were warmed six months
TaTer, it was found that very few of the radiation lesions had
been repaired.
Similarly, hibernating mammals such as squirrels or marmots
are much less radiosensitive when irradiated and maintained in
hibernation. On warming 2 to 4 weeks after irradiation the animals
show the radiation response of animals exposed to x rays in the
non-hibernating state and die in about 20 d following a lethal
dose.
A logical extension of the observations of Bacq and Alexander (1961) is that
organisms that receive a radiation dose in cold deep water and then migrate to
warm surface waters could be at increased risk. More recently, cell kinetics
and cell survival at lowered temperatures have been examined in.detail, and
further evidence for increased radiosensitivity was obtained (e.g., Van Rijn
et_al_., 1985; and Szechter and Schwarz, 1977).
Salinity may also modify the level at which lethal effects of radiation
are observed. Angelovic et al_. (1969) found an LD5Q/30 for the mummichog
Fundulus heteroclitus of 2050 rad at a salinity of 5 ppt and 1020 rad at 25 ppt,
18
-------
It has been widely established (Ophel, 1976; Donaldson and Foster, 1957)
that radiosensitivity varies with age and that early lifestages are more
sensitive than adults. LD5Q values below 100 R have been reported for a
variety of fish species. They Include 16-R LD50/30 for single-cell embryos of
the silver salmon Oncorhynchus kisutch (Bonham and Welander, 1963), 90-rad
LDgQ for survival to metamorphosis in larvae of the piaice P1euronectes
platessa (Ward et _a]_., 1971), and 58-R LD5Q,55 for the rainbow trout Salmo
gairdnerii irradiated as single-cell embryos (Welander, 1954). These values
are low relative to the LD5Q values of above 1000 rad obtained for adult
fish in most of the studies reported in Table 1. Thus, the lethal effects of
radiation in fishes are highly dependent on lifestage at irradiation, and
differences in ID-- of more than two orders of magnitude may be expected for
different stages of the same species.
The relative biological effectiveness (RBE) of the radiation and dose
rate delivered by a radiation source may also alter the response. A high RBE
for neutrons in mammals has been demonstrated (Casarett, 1968). Similarly,
Etoh et^ al_. (1974) found that neutrons were four times more effective than
x rays in the induction of mortality in goldfish.
For a given total dose, the effects of decreasing the dose rate and
fractionating the dose have been studied in fish (Chee jjt j*l_., 1979; Egami,
1969a; Egami and Etoh, 1966; Etoh and Egami, 1967). Classic results, similar
to those in mammals, have been obtained, i.e., as dose rate is decreased and
as intervals between fractionated doses are increased, a greater total dose is
required to produce the same biological effects observed at high dose rates
with no fractionation.
Species-specific differences in radiation-induced lethality are difficult
to evaluate from the data available (Table 1) because of the differences among
studies in exposure conditions, lifestages, and observation periods. However,
the research of White and Angelovic (1966), reported in Chipman (1972) and
Templeton et jil_. (1971), shows that for six species of marine fish irradiated
with 60Co as juveniles or postlarvae, the LD /30 ranged from 1050 R to 5550 R.
Juveniles and postlarvae are actively growing organisms, but because not all
may be growing at the same rate, differences in sensitivity may be attributable
to differences in growth rate as well as to differences in species.
19
-------
Mortality of Invertebrates from Acute Radiation Exposure
The comparative radiosensitivity of living organisms has been discussed
in numerous reviews (e.g., Whicker and Schultz,- 1982). Although many factors
determined the radiosensitivity of an organism, it has been emphasized that,
among other factors, organismal radiosensitivity increases with increasing
complexity of the organism. Bacq and Alexander (1961) report LD50/3Q values
for some unicellular protists such as Paramecium sp_._ to be in excess of
100,000 R. LD5Q/30 values for many higher invertebrates (Table 2) range in
the thousands and ten thousands of rads (tens and hundreds of Gy). However,
values as low as 210 rad (Engel et al_., 1974) have been reported. While the
role of some modifying factors such as salinity and species specificity have
been examined (Engel et al_., 1974 and Table 2), the lower limit of radiation
inducing mortality in invertebrates is probably still undefined. In
particular, very little work has been done to determine the radiosensitivity
of gametes and early lifestages of invertebrate species. Other areas of
weakness are the short observation times used in many of the studies and the
absence of data on temperature effects.
Some work that has been completed on mortality of early lifestages of
invertebrates indicates greater radioresistance of embryos of invertebrates
than of fish. Ravera (1968), reported in Blaylock and Trabalka (1978), showed
LD values for the four-cell, trochophore, veliger, and hippostage embryos of
50 •
the gastropod Physa acuta to be 1075, 2350, 4900, and 10,750 R, respectively.
In contrast, Bacq and Alexander (1961) cite work on Drosophila that indicates
an LD™ for 3-h Drosophila embryos of 200 R. It must be re-emphasized at this
point that the results of such studies depend not only on the dose received but
also on many other inherent experimental parameters such as observation time,
mitotic delay at high doses, differences in cell-cycle time, and temperature.
Furthermore, similar radiosensitivities of related species cannot be assumed,
and rules on increasing radiosensitivity with phylogenetic position are not
absolute. Hoppenheit (1969), in Chipman (1972), has shown that the LD50/3Q of
the amphipod Gammarus duebeni (3900 R) is twice that of Gammarus zaddachi
(1700 R). In addition, the LD5Q/30 of the more phylogenetically advanced
crustacean Callinectes sapidus (blue crabs) is 56,600 R (Engel, 1967, in
Chipman, 1972). Differences in radiosensitivity between sexes have also
been observed. In the mollusc Urosalpinx cinerea, males (LD50/7o.of 15,000 R)
are twice as radiosensitive as females (LD50/7o of 30»000 R)*
20
-------
Table 2. Induction of mortality in invertebrates from exposure to acute levels of radiation.
ro
Lethal dose0
(% mortality, days) Organism/1 ifestage
Radiation regime
Comments
Reference
210 R (30 ppt)c
405 R (25 ppt)
450 R (20 ppt)
600 R (15 ppt)
(50%, 40 d)
1075 R
(50%, -)
1700 R
(50%, 30 d)
1700 R
(50%, 30 d)
2000-9000 R
(100%, 35-75 d)
3500 R
(50%, 30 d)
3900 R
(50%, 30 d)
5000 R
(50%, 80 d)
Palaemonetes pugiod — Salinity effects observed
(grass shrimp)/adults —
200-4800 R
Physa acutad — —
(freshwater snail)/
four-cell embryos
Gammarus salinus6
(amphipod)/ —
Gammarus zaddachi6
(amphipod)/ --
Hydra- ftrsca" -- . —
(coel enter ate)/ —
Gammarus duebeni6
(amphipod) /females
Gammarus duebeni6
(amphipod)/males
Crassostrea virginica6 60Co —
(oyster)/ ~ —
Engel et al., 1974
Ravera, 1968 (in Blaylock
and Trabalka, 197tt)
Hoppenheit, 19&9 (in
Chipman, 1972)
Hoppenheit, 1969 (in
Chipman, 1972)
Nikitin, 1938 (in
Polikarpov, 19bt>)
Hoppenheit, 1969 (in
Chipman, 1972)
Hoppenheit, 1969 (in
Chipman, 1972)
White and Angelovic, 1966
-(in Templeton et al., 1971)
-------
Table 2. (Continued.)
ro
ro
Lethal dose3
(% mortality, days)
5500 R
(100%, 75 d)
8000 R
(50*. 50 d)
10,000 R
(100%, 60 d)
10,000 R
(50%, 35 d)
10,000 R
(50%, 35 d)
10,000 R
(50%, 50 d)
12,000 R
(100%, 63 d)
20,000 R
(50%, 50 d)
28,000 R
(50%, 24 d)
Organism/lifestage Radiation regime Comments
Planaria polychroa""
(planarian)/ —
Uca pugnax6 Co
(fiddler crab) /females
Dendrocoelum lacteum"
(annelid)/ --
Calliopius laeviusculus6
(amphipod)/ --
Allorchestis angustus6
(amphipod)/ --
Arbacia punctulata6
(sea urchin)/ —
Pelmatohydra oligactis"
(coelenterate)/ —
Nassarius obsoletus Co
(mollusc)/males
Physa acutad x ray Behavioral changes before death
(freshwater snail )/adults 2000-220,000 R
Reference
Nikitin, 1938 (in
Polikarpov, 1966)
White and Angelovic, I960
(in Templeton et al_., Iy71)
Schmidt, 1946 (in
Polikarpov, 1966)
Bonham and Palumbo,
1951 (in Chipman, 197*)
Bonham and Palumbo,
1951 (in Chipman, 1972)
White and Angelovic, 1966
(in Templeton et^ al_., la71)
Polikarpov, 1957a,b (in
Polikarpov, 1966)
White and Angelovic, 196b
(in Templeton et^a]_'> 1971)
Ravera, 1967
-------
Table 2. (Continued.)
ro
Co
Lethal dosea
(% mortality, days)
30,000 R
(50%, 50 d)
40,000 R
(50%, 50 d)
50,000 rad
(50%, 73 djadults)
(50%, 45 d;juveniles)
50,000 R
(50%, 80 d)
56,600 R
(50%, 30 d)
Lethal dose is pxnre
Organism/1 if estage Radiation regime6
Urosalpinx cinerea6 60Co
(mollusc)/males
Urosalpinx cinerea6 60Co
(mollusc)/ females
Neanthes- arenaceodentata6 137Cs
(polychaete)/adults and 500 rad/min
Juveniles 100-100,000 rad
Mercenaria mercenaria6 60Co
(clam)/ -
Callinectes sapidus6
(blue crab)/adu'lts
Diaptomus clavipesd 60Co
(calanoid copepod)/ 7000 rad/min
adults and embryos 1000-100,000 rad
Comments Reference
White and Angelovic, 1966
(in Templeton et_£l_.; 1971)
White and Angelovic, 19b6
(in Tempi eton et_ al_., 1971)
Effects of cell reproduction were considered Anderson et al.,
in prep.
White and Angelovic, 1966
(in Tempi eton jrtaK, 1971)
Engel, 1967 (in
Chipman, 1972.)
Mean survival time was significantly Gehrs et al., 1975
decreased at 1000 rad; lowest dose LD50/3Q
cannot be determined due to short lifespan;
control death occurred prior to 30 d
-•••— "•" ^'^>"- '»»' <.ani.jr in a. speciriea numoer or days. Units are those used by the
authors; R is the abbreviation for roentgen.
Radiation regime is presented as source, dose rate, and total dose; a dashed line indicates information was not available. For x-ray dat, a
araet to ohi^ S-T '"T- *"***"* '" determining *~™y dose rate are Volta9e> ta^ Aerial, filtration, tube current, and
c target-to-object distance. Units are those used by the author; R is the abbreviation for roentgen.
ppt is the abbreviation for parts per thousand.
e Organism is freshwater, anadroraous, or estuarine as opposed to exclusively marine.
Organism is exclusively marine.
-------
Mortality of Fish and Invertebrates from Chronic Radiation Exposure
In a few studies, mortality in chronically irradiated fish and
invertebrates (see Tables 3 and 4) is examined. Donaldson and Bonham (1964)
reported no significant differences in mortality between the salmon
Oncorhynchus tschawytscha embryos irradiated with 0.5 R/d for approximately
20 d and the control salmon embryos; observations were conducted up to the
time of release of the smolts. Erickson (1973) reported no increase in
mortality of the guppy Poecilia reti oil ata exposed to 0.05 to 1 mCi/mL of
tritium (total doses of 340-4700 rad). Adults of the blue crab Callinectes
sapidus subjected to chronic gamma irradiation required dose rates greater
than 29 rad/h for 70 d to cause death (Engel, 1967), and juveniles of the clam
Hercenaria mercenaria exposed to 0.006-37.0 rad/h for 14 months exhibited only
decreases in reproduction and growth at the highest dose rate, 16-37 rad/h
(Baptist et_a]_., 1976).
Conclusions on Mortality Data
Although it is clear that many aspects of mortality due to radiation
injury in aquatic species remain obscure, it is not likely that observable
numbers of aquatic organisms will die from the radiation levels that are
present at current ocean dumpsites. The highest dose rate observed from the
release of low-level, radioactive waste in the ocean is 0.240 rad/d, which
occurred at the end of the pipeline at the Windscale reprocessing plant in the
U.K. (Woodhead, 1984). Such doses are not expected over large areas at ocean
dumpsites (NEA, 1980); however, we use this figure as the documented maximum
level for our discussion. At 0.240 rad/d, mortality has not been observed in
any aquatic organism (Tables 1-4). However, very few studies at chronic dose
rates have been conducted. It is likely that reproductive and genetic effects
will have more relevance at these levels. Nevertheless, it is not totally
inconceivable that chronic irradiation of sensitive lifestages such
as brooded embryos, which may be very slow in developing at cold temperatures,
could accumulate sufficient damage to cause death. How, or if, the biochemical
lesions would be realized at such low temperatures is not known. In
conclusion, the lowest observed lethal effect level is a 16-R LD50/150 for
single-cell silver salmon embryos (Bonham and Welander, 1963). Methods and
24
-------
Table 3. Induction of mortality in fish from chronic exposure to radiation.
Organism/1 ifestage
(Chinook salmon)/embryos
Poecilia reticulata^
(guppy)Xyoung fish
Radiation
regime3
Comments
Reference
Oncorhynchus tschawytschab b°Co
0.5 R/d
33-40 R
No significant dif-
ferences in mortality
Donaldson and
Bonham, 1964
Tritium No significant increase Erickson,
0.05-1 mCi/mL in mortality 1973
340-4700 rad
a Radiation regime is presented as source, dose rate, and total dose. Units
are those used by the author, R is the abbreviation for roentgen.
Organism is freshwater, anadromous, or estuarine as opposed to exclusively
marine.
analyses used in this study are suitable. However, it should be noted that the
LD50/150 of salmon embryos at slightly different incubation times (or
developmental stages) ranged from 16 R to 309 R.
PATHOPHYSIOLOGY
In this section on pathophysiological effects, we describe research
conducted on several types of somatic tissue. Research on germ tissue or
developing embryos is presented in the sections on reproduction and
development.
Pathophysiological Effects in Fish from Acute Radiation Exposure
Research has defined a variety of pathophysiological lesions in fish from
acute radiation exposure. In most such studies, endpoints that characterize
radiation syndromes in mammals have been examined. These include effects on
hemopoietic tissue (Welander et,al_., 1948; Eton et. al_., 1968; Cosgrove et al_.,
25
-------
Table 4. Induction of mortality in invertebrates from chronic exposure to
radiation.
Organism/1 ifestage
Radiation
regimea
Comments
Reference
Callinectes sapijusb 6°Co Decreased survival only Engel, 1967
(blue crab)/adults 3.2-29.0 rad/h at highest dose rate,
70 d 29.0 rad/h
Mercenar.ia
mercenaria
(clam)/juvenile
60Co Deleterious effects on Baptist
0.006-37.0 rad/h growth and survival only £t a]_.,
14 mo at highest dose rate of 1976
16-37 rad/h with only 10%
survival to 14 mo
Argopecten irradiensb 60Co No deleterious effects Baptist
(scallop)/juvenile 0.006-37.0 rad/h on growth and survival et_al_.,
3 mo 1976
a Radiation regime is presented as source, dose rate, and total dose.
are those used by the author.
Units
Organism is exclusively marine.
1975; Lockner et al_., 1972), effects on immunity (Preston, 1959; Shechmeister
et al_., 1962), or damage to the intestinal epithelium (Hyodo, 1965a; Etoh et,
IT.7"l968; Hyodo-Taguchi and Egami, 1969; Johnson et al_., 1970). However,
with the exception of studies relating to immunity, most of these effects have
been studied at lethal levels, and they mainly serve to characterize lethal
radiation syndromes in fish (Table 5). The few studies that document effects
below 1000 rad are discussed below.
Hemopoietic cells of fish are affected by sublethal doses of radiation.
Welander et a]_. (1948) observed a significant decrease in the number of
hemopoietic cells in embryos of the.chinook salmon Oncorhynchus tschawytscha
26
-------
Table 5. Induction of pathophysiological changes in fish from exposure to acute levels of radiation.
Endpoint
Organism/1ifestage
Susceptibility Carassius auratusb
to infection (goldfish)/adults
Immune response Plaicec/—
ro
•vl
Hemopoiesis
Hemopoiesis
Hemopoiesis
Hemopoiesis
vulgarisb
(tench)/adults
Ictalurus punctatusb
(channel catfish)/
.young
Gambusia affinisb
(mosquitofish)/adults
Lepomis macrochirusb
(bluegill)/adults
Radiation regime3
Comments
x ray 100& mortality was observed at 1000 rad when
1000, 2000 rad for Aeromonas study infected with. Aeromonas salmonicida compared
100, 1000 rad for Gyrodactylus study to 0% in uninfected irradiated (1000 rad) fish
and infected unirradiated fish
78% mortality was observed at 100 rad in fish
infected with Gyrodactylus sp.. whereas
unirradiated infected fish exhibited
approximately 15% mortality (day 20)
x ray
8000 R
60Co
2 rad/sec and 250 rad/sec
500-60,000 rad
60Co
305 rad/mi n
500-2500 rad
60rn
Co
360 rad/min
750-6000 rad
60Co.
160 rad/min,
1000, 3000 rad
Irradiated fish could still develop
leucocytes is in response to bacteremia;
this was demonstrated in two fish that died
at 45 and 51 d after exposure
At 18°C, 100 rad caused 50% decrease in
lymphocyte count; at 10°C, 16,000 rad are
required for the same effect
Reference
Shechmeister
et_al_.,
Preston,
Lockner £
al., 1972
Severe atrophy of hemopoietic tissue in kidney, Cosgrove et al_.,
spleen, and thymus in all fish at 500-2500 rad; 1975
recovery of hemopoietic system seen after 29 d
750 rad caused only slight depression of
hemopoiesis (endpoint not specified)
1000 rad caused atrophy in the hemopoietic
system of kidney within 3 d
Cosgrove et a\_.,
1975
Cosgrove jst
1976
-------
Table 5. (Continued.)
ro
oo
Endpoint
Hemopoiesis
Hemopoiesis and
glomerular
development
Concentration of
serum-protein
fractions
Blood coagulation
Blood coagulation
Damage to
intestinal
epithelium
Damage to
intestinal
epithelium
Organism/lifestage
Lagodon rhomboidesc
(pinfish)Xadults
Oncorhynchus tschawytscha
(chinook salmon)/
embryos
Lepomis macrochirus
(bluegill)/yearlings
Lagodon rhomboidesc
(pinfish)/adults
Fundulus sp.
(mummichog)/adults
Oncorhynchus kisutch
(coho salmon)/juveniles
Carassius auratus
(goldfish)/adults
Radiation regime3
Gamma ray
2000 and 5000 R
x ray
250-10,000 R
60rn
Co
167 R/min,
1000-3000 R
x ray
100-1600 R
x ray
100 R
x ray
140-8000 R
x ray
0, 8000 rad
Comments
Number of thrombocytes in irradiated fish
decreased linearly with time; rate of decrease
was greater in fish irradiated at higher dose;
number of leucocytes decreased also
Number of hemopoietic cells significantly
decreased at 500 R while at the same exposure,
development of glomeruli was only slightly
inhibited
An initial 50% decrease in beta globulins,
alpha globulins, and albumins occurred within
2-24 h after exposure; hemoconcentration
occurre'd after initial decrease
Increase in the coagulation time with exposures
above 100 R
Increase in coagulation1 time greatest at 24 h
and approached that of controls at 96 h
At 500 R, minor and temporary effects were
observed; doses of 1000 R and above caused
considerable cell damage with only partial
recovery
Fish irradiated at 8000 rad and held at 22°C
developed intestinal damage, which lead to death,
but those held at 4°C did not develop intestinal
Reference
Engel et al.,
iyo7
Welander et
al_., 1948
Ulrikson, 1973
Engel et al.,
19b5
Engel £t al. ,
1965
Johnson et al . ,
1970
.Hyodo, 1965a
warmer water
-------
TableS. (Continued.)
ro
CO
Endpoint
Damage to
intestinal
epithelium
Damage to
intestinal
epithelium
Organism/1 ifestage
Carassius auratusb
(goldfish)/adults
Oryzias latipesb
(medaka)/adults
Carassius auratusb
(goldfish)/adults
Radiation regime3
x ray
0, 8000 R
x ray
50-128,000 R
x ray
4000-32,000 R
Comments
Ratio of mitotic figures (3H-thynvidine
labeled:unlabeled) was lower in irradiated
fish intestine when compared to controls
Pathological changes in intestinal epithelium
responsible for acute radiation death at the
"dose-independent range" (3000-30,000 R).
Mitotic activity of epithelial cells of
Reference
Hyodo, 1965b
Hyodo-Taguchi
and Egami, 1969
Thyroid damage Various coral reef
fishes0
Thyroid follicle Salmo salarb
enlargement (Atlantic salmon)/
1arvae
Thymus volume
Oryzias latipes
(medaka)/adults and
embryos
Nuclear bomb test at
Enewetak Atoll
(131D
x ray
0, 350, 1000 R
Adults: y ray
50 R/min
200 and 500 R
Embryos: y ray
intestine completely inhibited; response
temperature dependent; histological changes
in intestine correlated with radiation death
Thyroid damage in coral reef fishes from bomb Gorbman, 1963
testing at Enewetak Atoll was demonstrated
histopathologically
Increased size.of thyroid gland follicles at Oganesyan,
1000 R; a-very slight increase was also observed 1973
at 350 R
Decreased thymus volume in adults showed Ghoneum and
recovery; decreased thymus volume in embryos Egami, 1980
showed no recovery at 4000 R
1000-4000 R
-------
Table 5. (Continued.)
CO
o
Endpoint
Organism/1 ifestage
Hitotic delay Carassius auratus
(goldfish)/adults
Inhibition of
DNA synthesis
Formation of
taste bud cells
Damage to skin,
intestine, gill,
and hemopoietic
tissue
Interrenal cell
hypertrophy
Oncorhynchus kisutch
(coho'salmon)/juveniles
Oryzias latipes
(medaka)/embryos and
adults
Carassius auratus
(goldfish)/adults
Carassius auratus
(goldfish)/adults
Radiation regime
x ray
1000, 2000, and 8000 R
x ray
1000, 2000, and 4000 R
137Cs
360 R/min (embryos)
250 R/min (adults)
1000 R
x ray
1000-16,000 R
x ray
250-128,000 R
Comments
Reference
At 3-6 h after exposure, the number of thymidine- Hyodo-Taguchi,
labeled intestinal epithelial cells in the 1970
irradiated groups'did not differ from controls;
after 24-48 h, the number of labeled cells in
the 8000-R group approached 0; in the 1000- and
2000-R groups an initial depression occurred with
recovery between 48 and 96 h
At all doses, DNA synthesis was depressed from
10-160 min; no longer time points or lower
doses were used
Partial inhibition of taste-bud-cell formation
in embryos; number of taste-bud cells in adults
declined but repopulation appeared to minimize
effects
Histological damage to intestinal and
hemopoietic tissue was demonstrated after
10 and 20 d at 1000 R; irradiation of the
tail region showed skin damage to be an
important cause of death; irradiation of
the head region caused gill damage, which
caused sodium loss through damaged tissue
Hypertrophy of interrenal cells was-produced in
fish at 1000-16,000 R; this effect was not seen
in hypophysectomized fish, and it was suggested
that ACTH secretion was stimulated by irradiation
Johnson et a!.,
1970
Gnoneum et al.,
19&3
Etoh et al., 1968
Aoki et _al_.,
19b6
-------
Table 5. (Continued.)
Endpoint
Organism/lifestage
Radiation regime3
Comments
Reference
CO
Osmoregulation Oncorhynchus kisutchb x ray
(coho salmon)/juveniles 1000-1200 rad
Osmoregulation
Osmoregulation
External
morphology
Anguilla anguilla
(eel)/adults
Anguilla anguil1ab
(eel)/adults
Eucinostomus sp.c
(mojarra)/post-larvae
60r
Co
1000 R
x ray
1000 R
x ray
400-3200 R
Osmoregulatory ability decreased at lethal Conte, 19bb
levels: (a) decreased survival of fish at doses
greater than 1000 rad when transferred from
hypoosmotic to hyperosmotic conditions (when
compared to those maintained under hypoosmotic
conditions); (b) increased plasma sodium, chloride,
and osmotic concentrations in surviving irradiated
fish maintained in salt water
Effect of increased salinity and irradiation Hansen, 1975
caused additive increase in lipid biosynthesis
in eel gills (assumes changes in membrane
permeability accompanied by changes in the
composition of the lipid moiety)
Radiation and salinity effects are decreased Hansen, 1980
in vitro, implying neurohormonal regulation of
these effects
Apparent decreased growth rate with increased Engel ^t aK,
exposure; changes in body proportions, size of 19b5
spines, and pigmentation.
1 Radiation regime is presented as source, dose rate, and total dose; a dashed line indicates information was not available. For x-ray data, a aose
rate is not given; factors important in determining x-ray dose rate are voltage, target material, filtration, tube current, and target-to-object
distance. Units are those used by the author; R is the abbreviation for roentgen.
Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
Organism is exclusively marine.
-------
after an x-ray exposure of 500 R. Cosgrove et^al_. (1975) noted severe
hemopoietic atrophy in kidney, spleen, and thymus of the channel catfish
Ictalurus punctatus after a 500-rad dose. Lockner et.al_. (1972) observed a
50% decrease in lymphocyte count when the tench Tinea vulgaris received
100 rad at 18°C, whereas at 10°C, 16,000 rad were required to produce the same
effect in the specified observation period.
Immunity of irradiated goldfish Carassius auratus was examined by ,
Shechmeister et a/L (1962). It was found that fish infected with Gyrodactylus
£p_. and receiving up to 100 rad from an x-ray source exhibited 78% mortality.
In contrast, unirradiated infected fish exhibited only 15% mortality.
Damage to intestinal epithelium is a characteristic acute lethal radiation
syndrome in mammals, and, in fishes, it has also been studied mostly at lethal
levels. However, minor and temporary damage to the intestinal epithelium of
coho salmon Oncorhynchus kisutch juveniles exposed to 500 R from a x-ray
source has been documented (Johnson et^al_., 1970). Exposures of 1000 R and
above caused considerable cell damage with only partial recovery.
Pathophysiological Effects in Invertebrates from Acute Radiation Exposure
Little or no work has been done to characterize the pathophysiological
effects of acute ionizing radiation on aquatic invertebrates (Table 6).
However, Engel et_al_. (1974) found that irradiation (200 to 4800 rad) and
increased salinity acted additively in affecting amino acid pools of the
estuarine shrimp Palaemonetes pugio. These results may indicate that
irradiation causes membrane damage that inhibits regulation of intracellular
amino-acid pools used in osmoregulation.
Pathophysiological Effects in Fish and Invertebrates from Chronic Radiation
Exposure
To our knowledge, no research has been conducted that characterizes
pathophysiological effects from chronic radiation exposure in aquatic
invertebrates. This discussion will, therefore, be restricted to results of
research conducted on fish (Table 7).
Cosgrove j2t jil_. (1975) examined the mosquitofish Gambusia affinis for
hemopoietic damage after exposure to 60Co at 0.5 to 5.43 rad/h for up to
128 d. They found no demonstrable damage to hemopoietic organs after 37 d at
32
-------
Table 6. Induction of pathophysiological changes in invertebrates from exposure
to acute levels of radiation.
Endpoint
Radiation
Organism/1ifestage regime3
Comments
Reference
Osmoregulation Palaemonetes pugio^"
(grass shrimp)/
adults
Radiation and Engel ^t
increased salinity al., 1974
200-4800 rad act additively,
affecting ami no
acid pools
Radiation regime is presented as source, dose rate, and total dose; a dashed
line indicates information was not available.. Units are those used by the
author.
Organism is freshwater, anadromous, or estuarine as opposed to exclusively
marine.
5.43 rad/h (4822 rad total dose). Mild hemopoietic atrophy in kidneys and
spleens in some fish was seen after 128 d at 1.5 and 3.0 rad/h (4608 rad and
9216 rad total doses).
The immune response of chronically irradiated rainbow trout (Salmo
gairdnerii) embryos has been examined by analyzing antibody synthesis against
Chondrococcus columnaris disease in fish exposed to tritiated water (Strand ^t
al_., 1973a). Developing embryos exposed to tritiated water for 20 d
demonstrated a depressed immune response when compared to control fish.
Conclusions on Pathophysiology Data
With the exception of studies relating to immunity, research on the
pathophysiological effects of radiation exposure in aquatic organisms has
primarily served to elucidate that the mechanisms of radiation-induced
mortality in aquatic organisms are similar to those observed in mammals. Thus,
they are mainly studies on the cellular manifestations of lethal doses.
Factors that modify these responses have not been widely studied (except for
the effects of changes in salinity), but we propose that future work in this
33
-------
Table 7. Induction of pathophysiological changes in fish from chronic exposure to radiation.
Endpoint
Immune response:
antibody synthesis
aqainst Chondrococcus
columnaris disease
Hemopoiesis
Organism/lifestage
Salmo gairdnerii
(rainbow trout)/
embryos
Gambusia af finis
(mosquitofish)/adults
Radiation regime*
Tritium
1 uCi/mL and 10 uCi/mL
20 d
60Co
0.5-5.43 rad/h
336-9216 rad
Conroents Reference
Developing embryos exposed to tritiated Strand ^t^K,
water demonstrated depressed immune 1973a
response
No demonstrable damage to hemopoietic Cosgrove £t _al_. ,
tissues after 37 d at 5.43 rad/h (4822 1975
rad); mild hemopoietic atrophy in kidney
and spleen in some fish after 128 d at
1.0 and 3.0 rad/h (4608 rad and 9216 rad)
co
a Radiation regime is presented as source, dose rate, and total dose (or exposure time). Units are those used by the author.
Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
-------
area would not be as productive as examining more sensitive endpoints such as
reproductive and genotoxic effects and the factors that modify them. However,
further studies on immunity in fishes may be warranted. In mammals, the
pathophysiologic effect of primary concern at low doses is tumor induction in
a variety of tissues. To our knowledge, only one study has reported data that
demonstrate radiation-induced tumor production in aquatic organisms (Anders et
al_., 1973a,b).
REPRODUCTION
Effects of ionizing radiation on fertility have been observed at
relatively low dose rates and low total doses in mammals, fishes, and aquatic
invertebrates. Our review focuses on these low-dose effects because other
research utilizing relatively high doses has been reviewed previously (see
Appendix I for references). Reproductive effects occur at lower doses than
those causing mortality or typical pathophysiologic lesions, excluding tumor
development in mammals. Effects of ionizing radiation on reproduction in
mammals include responses at very low doses. For example, the ID™ of mouse
oocytes has been reported to be 5 rad from tritium exposure and 8 R for x-ray
exposure. The reported LD5Q of monkey oocytes after tritium exposure is
8 rad. Complete sterilization was observed in female mice exposed 20 to 40 d
postconception to 8.4 R/d from a 137Cs source (in NRC, 1980). In discussing
reproductive effects, it is important to note that in many studies in the
aquatic literature, the induction of reproductive effects and dominant lethal
mutations are not separated. For example, if an organism is irradiated and
reductions in brood size are noted, they may be due to either gamete death or
to dominant lethal mutations expressed in early development.
Reproductive Effects in Fish from Acute Radiation Exposure
Effects of ionizing radiation on reproduction in teleosts have been
studied by several investigators (Table 8). However, Egami and co-workers
performed more investigations in this area than any other group. For the
purpose of this discussion, their work is divided into three categories.
First, mechanistic studies were conducted that use high dose rates and total
doses greater than 1000 rad (10 Gy). Because of the high doses used in these
studies, which were reviewed in Egami and Ijiri (1979), they are not listed in
35
-------
Table 8. Induction of reproductive changes in fish from exposure to acute levels of radiation.3 Entries are
ordered according to the lowest dose at which effects were observed. This dose is not necessarily the
lowest dose at which effects could have been observed.
Organism/1 ifestage
Radiation regime
Comments
References
CJ
Oncorhynchus tschawytscha
(chinook salmon)/embryos
Qryzias latipes
(medaka)/adult males
x ray
250-1000 R
x ray
250-2000 rad
Counts.of primordial germ cells in salmon
irradiated at 250 R were 10% of control
values
500 rad caused a significant decrease in
oviposition frequency, an increase in percent
of unfertilized eggs, and an increase in the
number of sterile fish
Welander ej: _a]_.,
1948
Hyodo-Taguchi,
1980
Salmo gairdnerii
(rainbow trout)/
29-d embryos
r«
Co
33.5 R/min
0, 600, 800 R
Sterility was induced at lowest exposure tested
(600 R) and at all observation times (60-150 d)
Konno, 1980
Research on the reproductive effects of acute irradiation in teleosts has been reviewed by Egami and Ijiri,
1979. Because most of this research employed doses above 1000 rad (10 Gy), and has been reviewed extensively it
is not listed above.
Radiation regime is presented as source, dose rate, and total dose. For x-ray data, a dose rate is not given;
factors important in determining x-ray dose rate are voltage, target material, filtration, tube current, and
target-to-object distance. Units are those used by the author; R is the abbreviation for roentgen.
Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
-------
Table 8 and are not discussed. The second type of study conducted was the
analysis of gamete death from acute doses below 1000 rad (10 Gy). These
studies were alsfo extensively reviewed in Egami and Ijiri (1979), but some are
listed in Table 8 and are discussed in this section. The third type of study
conducted has been the analysis of gamete death from doses delivered at
relatively low dose rates. Results of this research are listed in Table 9 and
are discussed in detail in the section on reproductive effects due to chronic
radiation exposure.
Effects of acute ionizing radiation on female germ cells of teleosts have
been observed after their exposures to doses as low as 250 R. Welander et. a]_.
(1948) found that counts of primordial germ cells in the chinook salmon
Oncorhynchus tschawytscha exposed to 250 R from an x-ray source were 10% of
control values. Konno (1980) exposed rainbow trout Salmo gairdnerii embryos
to a 60Co source (33.5 R/min) for total doses of 0, 600, and 800 R. Sterility,
which was induced at the lowest dose tested, was detected at all observation
periods (60 to 150 d).
Male germ cells of the medaka Oryzias latipes were studied extensively by
Egami and co-workers, who found a temporary reduction in testicular weights
after exposure to 100 to 2000 R (x-ray source). They also found that, in
sexually inactive fish exposed to 100 to 1000 R, certain spermatogonial types
and primary spermatocytes exhibited damage within 3 d (Egami and Ijiri, 1979).
Reproductive Effects in Invertebrates from Acute Radiation Exposure
Effects of acute irradiation on reproduction in aquatic invertebrates
occur over a dose range of at least two orders of magnitude (Table 9). Causes
for this broad range may not lie in actual species-specific differences in
gamete killing, but in differences in the gamete stage irradiated and in the
cell-repopulation capacity of different organisms.
In experiments by Ravera (1967), adults of the freshwater snail Physa
acuta received 2000 to 220,000 R from an x-ray source. A dose of 2000 R
reduced fertility and embryo viability. However, recovery from the damage to
germ tissue was evident in snails receiving 2000 and 10,000 R. At 10,000 R.,
the production of egg capsules and the number of eggs per capsule were reduced.
Moreover, all embryos produced during the first 50 d after irradiation from
snails receiving 10,000 R were not viable. A greater percentage of the embryos
37
-------
co
CO
Table 9. Induction of reproductive changes in invertebrates from exposure to acute levels of radiation. Entries are
ordered according to the lowest dose at which effects were observed. This dose is not necessarily the lowest
dose at which effects could have been observed.
Organism/1 if estage
Gammarus duebeni
(amphipod)/adults
Artemia salina0
(brine shrimp)/adults
Radiation
regiraea
x ray
65-740 R
x ray
500 R and
1000 R
Comments
Reduced egg-production rate at 220 R
Progressive decrease in sensitivity of oocytes
throughout prophase and into metaphase. Significant
reduction in hatching after 500-R exposure
References
Hoppenheit, 1973
Cervini and
Giavelli, 1965
Artemia salina
(brine shrimp)/adults
Physa acuta
(freshwater snail)/adults
Reduced fecundity at 1000 R, the lowest exposure tested Squire, 1970
1000-5000 R
x ray Reduced fecundity of the adults and reduced viability
2000-220,000 R and fertility of the eggs was observed at 2000 R;
temperature increase exacerbated radiation-induced effects
Neanthes arenaceodentatac
(polychaete)/adults and
juveniles
Physa acuta
(freshwater snail)/adults
Cs No oocyte development was observed in adults receiving
500 rad/min 50 Gy; reduced fecundity was observed between
100-100,000 rad 1 and 5 Gy
x ray Reduced fecundity of adults; 100,000 R abolished
2000-220,000 R reproductive capacity of adults
Ravera, 1967
Anderson _et aly,
in prep.
Ravera, 1966
a Radiation regime is presented as source, dose rate, and total dose; a dashed line indicates information was not
available. For x-ray data, a dose rate is not given; factors important in determining x-ray dose rate are voltage,
target material, filtration, tube current, and target-to-object distance. Units are those used by the author; R is the
abbreviation for roentgen.
15Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
c Organism is exclusively marine.
-------
produced after that time were viable. Ravera (1967) proposed that repopulation
of the gonad by undamaged germ cells could explain this effect. Reproductive
activity was completely abolished at 100,000 R.
Differential radiosensitivity between meiotic stages has been studied in
prophase and metaphase oocytes of Artemia salina. Cervini and Giavelli (1965)
showed that radiosensitivity of oocytes declined as prophase progressed.
Metaphase oocytes were also less sensitive than prophase oocytes.
Hoppenheit (1973) observed reduced egg-production rate in adults of the
amphipod Gammarus duebeni receiving as low as 220 R. However, this was offset
by higher survival of adult females and increased brood size.
Anderson .et jal_. (in preparation) found that brood size of the marine
polychaete worms Neanthes arenaceodentata was reduced after doses between
1 and 5 Gy were delivered to adult worms. In contrast, preliminary data on
irradiated juveniles indicated such effects occurred between 5 and 10 Gy,
suggesting that repopulation of the gonad by undamaged germ cells may occur.
In determining the comparative sensitivity of gametes in selected species,
it is important to know the gametogenic stage that was irradiated. However,
this is not well characterized in some reports. Furthermore, both oogenesis
and spermatogenesis are asynchronous in many aquatic organisms and, hence,
many stages may be irradiated simultaneously.
Reproductive Effects in Fish and Invertebrates from Chronic Radiation Exposure
Chronic, low-level radiation studies may be conducted over many
gametogenic stages and cycles. The lowest dose rate at which effects of
chronic radiation exposure on fertility of aquatic invertebrates and fishes
have been demonstrated (Tables 10 and 11) is between 0.59 R/d (Trabalka and
Allen, 1977) and 10 R/d (Bonham and Donaldson, 1972). This is comparable to
the range in which effects of chronic irradiation on fertility are first
observed in mammals. These data indicate that fish and invertebrate oocytes
are not more radioresistant than mammalian oocytes. However, when single acute
doses of radiation are used to study reproduction, lower organisms may have a
higher capacity to replace damaged cells.
At least five studies on fishes and aquatic invertebrates were conducted
in which the effect of chronic radiation exposure in the dose-rate range
mentioned above were examined (Tables 10 and 11). Trabalka and Allen (1977)
compared populations of the mosquitofish Gambusia affinis from the
39
-------
Table 10. Induction of reproductive changes in fish from chronic exposure to radiation. Entries are ordered according to the lowest cose
at which effects were observed. This dose is not necessarily the lowest dose at which effects could have been observed.
Organism/lifestage
Gambusia affinis
(mosquitofish)/all stages
Poecilia reticulata
(tropical guppy)/ 0-3 d
embryos
Poecilia reticulata
(tropical guppy)/ 0-3 d
embryos
Oryzias latipes
(medaka)/adult males
Oncorhynchus tschawytscha
(chinook salmon)/smolts
Radiation regime
White Oak Lake
0.59 R/d
lifetime
137Cs
4.08, 9.60, 30.48 rad/d
Total number of days varied
137Cs
6.0, 12.0, 40.8 rad/d
Total number of days varied
137Cs
1.3-84'.3 R/d
78-5058 R
rn
Co
0.5-50 R/d
40-4000 R
Comments
No decrease in fecundity observed at 0.59 R/d;
however, increased embryo mortality was noted.
Total fecundity was markedly reduced at all dose rates
because of a decrease in mean actual brood-size and an
increase in temporary and permanent infertility; the
lowest dose rate tested was 4.08 rad/d
One pair out of 6 and 5 out of 7 were sterile at 6.0
and 12.0 rad/d, respectively; total accumulated doses
were 4000 and 8000 rad; at 40.8 rad/d complete
sterility occurred at 5000 rad; this marked dose-
rate effect was not reflected in changes in fecundity
in fertile guppies
408 R delivered at 6.8 R/d caused a decreased
oviposition frequency, an increased percent of
unfertilized eggs, and an increase in number of
infertile fish
Gonadal development was retarded at 10 R/d or more
References
Trabalka and
Allen, 1977
Woodhead, 1977
Purdom and
Woodhead, 1973
Hyodo-Taguchi,
1980
Bonham and
Donaldson, 1972
a Radiation regime is presented as source, dose rate, and total dose (or exposure time). Units are those used by the author; R is the
abbreviation for roeritgen.
b Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
-------
Table 11. Induction of reproductive changes in invertebrates from chronic exposure to radiation. Entries are ordered according to the
lowest dose at which effects were observed. This dose is not necessarily the lowest dose at which effects could have been
observed.
Organism/1ifestage Radiation regime3
Comments
References
Physa heterostrophab White Oak Lake
(aquatic snail)/adults 0.65 rad/d
lifetime
Frequency of egg-capsule production was reduced in the irradiated
(0.65 rad/d) population, but an increase in number of eggs per
capsule occurred, resulting in similar rates of egg production
compared to controls
Cooley, 1973
£hjrsa heterostropha Co Egg capsule and egg production were completely stopped by 600 rad/d, Cooley and Miller,
(aquatic snail)/adults 24, 240, 600 rad/d were reduced significantly by 4 weeks and stopped completely.by 1971
24 weeks 19 weeks at 240 rad/d, and were only significantly lower than controls
during week 4 to 12 for the group receiving 24 rad/d
* Radiation regime is presented as source, dose rate, and total dose (or exposure time). Units are those used by the author.
Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
-------
radionuclide-contaminated White Oak Lake at the Oak Ridge National Laboratory
to those from a matched control site. They found no decrease in fecundity, but
an increase in embryo mortality of the fish from White Oak Lake. These results
were obtained by returning animals collected from radionuclide-contaminated and
control sites to the laboratory and observing subsequent reproduction under
controlled conditions. This study is more definitive than others conducted on
animals from White Oak Lake, but results are confounded by the fact that
contaminants other than radionuclides are present in White Oak Lake. Cooley
(1973) examined the reproductive biology of pond snails at White Oak Lake. He
also transplanted animals from White Oak Lake to the laboratory. He found
that frequency of egg-capsule production was reduced; however, an increased
number of eggs per capsule was also documented. It is interesting to note
that a prior laboratory study by Cooley and Miller (1971) documented clearcut
reproductive decline at 240 rad/d but not at 24 rad/d. Irradiation was
initiated on 45-d-old snails, and laboratory effects might have occurred at
lower levels if irradiation was extended over the entire lifetime of the
organism.
Three of the five studies mentioned were conducted exclusively in the
laboratory, where control of confounding variables is more readily achieved.
The most rigorous of these studies was that of Woodhead (1977), who examined
fecundity of the guppy Poecilia reticulata receiving 4.08, 9.60, and
30.48 rad/d. Total fecundity was significantly reduced at all dose rates.
Reductions in fecundity in this important and thorough study are probably due
both to reproductive effects (damage to gametes) and the induction of dominant
lethal mutations in gametes. In a similar study, Woodhead et_al_. (1983)
reported that Ameca spi endens is even more sensitive than the guppy. However,
this report is an abstract, and the details of the research cannot be
evaluated at this time. Hyodo-Taguchi (1980) observed an increased percentage
of unfertilized Oryzias latipes eggs after males used to inseminate the eggs
received 6.8 rad/d for 60 d. No statistically significant effects were
observed at 2.9 rad/d, the next lower dose rate used. Bonham and Donaldson
(1972) exposed chinook salmon Oncorhynchus tschawytscha embryos for the first
80 d of life to 0.5 to 50 R/d. Approximately 4 weeks after the irradiations
were completed, gonadal development was observed in smolts. They found that
gonadal development was first retarded in those receiving 10 or more R/d.
42
-------
Conclusions on Reproduction Data
Comparisons of the effects of acute irradiation on reproduction of fish
and invertebrates are often not valid because the gamete stage irradiated
differed between studies. Furthermore, even when the same stage was
irradiated, it was not established that the capacity for repopulation of
damaged cells was the same between organisms. Research on effects of chronic
irradiation on the gonads is of particular interest, however, because the
results show effect levels comparable to those observed in mammals. Dose
rates between 0.5 R/d to 10 R/d appear to define a critical range in which
detrimental effects on fertility are first observed in a variety of sensitive
organisms. Unfortunately, almost nothing has been done on this subject with
invertebrates. For aquatic organisms, nothing is known about whether
responses would occur at lower levels if effects of modifying environmental
factors were examined. The only definitive study on the effects of lifetime
irradiation on the reproduction of aquatic organisms is that of Woodhead
(1977). He demonstrated decreased fecundity in the guppy at 4.08 rad/d.
Finally, it must be re-emphasized that the dose rates mentioned above are
slightly higher than the highest dose rates observed in the ocean due to
waste-disposal activities (0.240 rad/d) and considerably higher than those
expected at deep ocean disposal sites.
DEVELOPMENT
Effects of radiation on the development of fish embryos has been the most
widely debated topic in aquatic radiobiology (Woodhead, 1984), and results on
irradiation of developing embryos by introduction of radionuclides directly
into the test water have been most contentious. Recently, this topic was
exhaustively reviewed by Woodhead (1984). Our purpose is to summarize some of
the general conclusions of Woodhead (1984) and to identify the studies
demonstrating effects at relatively low levels from (1) acute irradiation,
(2) chronic irradiation from a sealed source, and (3) chronic irradiation by
radionuclides in the water. Data on the effects of acute and chronic
irradiation of aquatic invertebrate embryos are also reviewed. Tables 12-15
contain representative data on these subjects. No data on studies employing
radionuclides in the water, other than tritium, are included.
43
-------
Developmental Effects from Acute and Chronic Irradiation of Fish
Careful dosimetry is important in all radiobiological experiments,
particularly so when radionuclides are present in the water rather than in a
sealed, external source or generated by an x-ray machine. Accumulation of
radionuclides in embryos and absorption of radionuclides onto membranes may
affect the dose received by an organism. Woodhead (1984) concluded that all
studies on fish embryos using radionuclides in the water (except those using
tritium because it is not bioconcentrated) contain minor to very serious flaws
in dosimetry. This means that, for such experiments, the dose received by
the organism cannot be characterized. Because of this criticism, results from
such studies are presented separately.
Different criteria'have been used to evaluate the effects of ionizing
radiation on developing fish embryos (Table 12): hatching success, embryo
mortality, and frequency of morphologically abnormal embryos and larvae. The
forms of the gross abnormalities in fishes include anophthalmia, cyclopia,
monophthalmia, microphthalmia, abdominal and caudal foreshortening, abdominal
lordosis, scoliosis, and abnormal fin structure. The relatively minor changes
in fishes include changes in standard length, eye diameter, head length,
pigmentation, and fin-ray number. From the differences in sensitivities at
successive early stages of development, it appears that a number of different
vulnerable processes must be involved in the production of maldevelopment and
that these processes are in progress at various times considerably prior to the
earliest histological evidence of organogenesis (McGregor.and Newcombe, 1968).
Several experiments have demonstrated a trend of decreasing
radiosensitivity with increasing development (Woodhead, 1984). For example,
Bonham and Welander (1963) determined the LD5Q at hatching and at 150 d
after fertilization for the silver salmon irradiated at the one-cell stage.
The minimum values were 30 R and 16 R, respectively, for fish irradiated at
the one-cell stage. A consistent decrease in radiosensitivity of the embryos
with increased age at the time of irradiation was observed in the later stages
with an LD5Q of 1871 R at hatching for irradiation in the late-eyed stage.
These differences in radiosensitivity are related to critical periods in
organogenesis and changes in mitotic activity.
Allen and Mulkay (1960) investigated the histological effects of exposure
of the paradise fish Macropodus opercularis to 1000 R at different stages of
development. The order of decreasing sensitivity of tissue and organ systems
44
-------
Table 12. Induction of developmental changes in fish from exposure to acute levels of radiation. Entries are ordered according to the
lowest dose at which effects were observed. This dose is not necessarily the lowest dose at which effects could have been
observed.
en
Organism/1ifestage
Radiation regimea
Comments
Pleuronectes platessac
(plaice)/embryos
Fundulus heteroclitus0
(mummi chog)/embryos
x ray
30-150 rad
x ray .
Oncorhynchus tschawytscha x ray
(chinook salmon)/embryos 250-10,000 R
Cyprinus carpiob
(carp)/one-cell embryos
60r
Co
7.1 rad/sec
50-2000 rad
References
Oncorhynchus kisutchb
(silver salmon )/one-cell
embryos
Salmo gardnerii
(rainbow trout)/embryos
x ray
12-2400 R
x ray
25-2570 R
LD50 at 16 R observed 150 d after fertilization
Retardation in growth seen, at exposures as low as 38 R
for those irradiated during earlv-eved staaes: > 200 R
Bonham and
Welander, 1963
Welander, 1954
required for changes in other stages; increased
frequency of abnormalities in irradiated embryos
LD50 at metamorphosis for plaice larvae irradiated
at blastula stage was 90 rad
Eggs at 300-400 R in the one-cell to two-cell stage
developed major malformations; fertilized eggs and
early cleavage stages showed greatest sensitivities
Development of the eye, gill epithelium, fin, and
chromatoph'ores were studied; slight differences in
pigmentation and retardation of gill-filament
development were noted at 500 R; otherwise, no
effects were observed below 1000 R
LD50 at natchin9 was 601 rad for 0 radiation and
501 rad for y radiation; no hatching at 1000 rad
Ward et al_., 1971
Rugh and '&
Clugston,
1955
Welander and
Donaldson, 1948
Blaylock and
Griffith, 1971
-------
Table 12. (Continued.)
Organism/lifestage
Radiation regime3
Comments
References
cr>
Salroo gairdnerii
(rainbow trout)/embryos
Oncorhynchus tschawytscha
(chinook salmon)/embryos
Oryzias latipes
(medaka)/fertilized ovum
and sperm through last
embryonic stage
Oryzias latipes
(medaka)/fertilized ovum
and sperm through last
embryonic stage
x ray
10, 100, and 1000 rad
x ray
250-10,000 R
137Cs
250, 33.33, and 1.7 R/min
2000 R
x ray
2000 R
No significant increase in malformations and mortality McGregor and
at 10 and 100 rads; significant increases at 1000 rad; Newcombe, 1968
malformations the same as those from irradiating gametes
Lowest exposure at which significantly decreased weight Welander ana
and length were observed was 1000 R Donaldson, 1948
Decreased % hatch and increased incubation time from Egami and Hama,
2000 R delivered at 250 R/min and 33.3 R/rain, none 1975
at 1.7 R/min; lowering the temperature during irradiation
negated dose-rate effects.
Changes in the hatch rate from a exposure to 2000 R Egami ana Hama,
depending on what stage irradiated (pattern irregular) 1975
Pleuronectes platessa0
(plaice)/embryos
x ray
25-4800 R
No significant reduction
and mortality
in % hatching, malformations,
Tempi eton, 1966
Radiation regime is presented as source, dose rate, and total dose; a dashed line indicates information was not available. For x-ray
data, a dose rate is not given; factors important in determining x-ray dose rate are voltage, target'material, filtration, tube
current, and target-to-object distance. Units are those used by the author; R is the abbreviation for roentgen.
Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
c Organism is exclusively marine.
-------
was the following: blood and hemopoietic tissue > eye > central nervous
system > germ cells > muscles > gut \ heart > ear and lateral line >
pronephros > olfactory organ > notochord > pigment cells. For all tissues and
organs, it was found that specific types of damage to a presumptive tissue
could be elicited by radiation prior to the appearance,of its identifiable
precursor cells. Also, in agreement with the law of Bergonie and Tribondeau,
the period of differentiation of a tissue and any period of increased mitotic
activity in a tissue or organ were found to result in increased
radiosensitivity (Woodhead, 1984).
These data on organismal and tissue radiosensitivity in fish embryos
raise two more important points relevant to our analysis of low-effect
levels. First, the embryo or larval stage at which the radiation dose is
received is a major determinant of what the effect level will be. Second, the
time period over which observations are made must be sufficiently long for the
induced effects to be observable.
The lowest effect levels observed from acute irradiation of developing
fish embryos are the levels observed by Bonham and welander (1963) described
above (Table 12). Thus, fish-embryo mortality has been documented from
exposures of 16 R. This low value is lower than minimum-effect levels for
other species. Blaylock and Griffith (1971) found the LD5Q at hatching for
carp embryos exposed as single-cell eggs to be 601 rad for 6 radiation and
501 rad for y radiation. An LD5Q of 90 rad at metamorphosis was
determined for plaice larvae, the only exclusively marine fish studied,
irradiated at blastula stage (Ward et_al_., 1971).
Very few studies have been conducted that use sealed sources to examine
the effects of chronic irradiation on fish embryos (Table 13). As a part of a
larger study on the effects of chronic irradiation on chinook salmon embryos,
Donaldson and Bonham (1964) found a significant increase in opercular defects
of smolt at exposures of 33 to 49 R given at 0.5 R/d from the moment of
fertilization.
In contrast to the paucity of studies using external sources to examine
the effects of chronic irradiation on fish embryos, numerous studies exist in
which radionuclides have been introduced directly into the test water.
However, Woodhead (1984) has concluded that these studies have contributed
47
-------
-PS-
CD
Table 13. Induction of developmental changes In fish fro® chronic exposure to rad1at1on.a Entries are ordered according to the lowest dose at
which effects were observed. This dose 1s not necessarily the lowest dose at which effects could have been observed.
Organism/lifestage
Oncorhynchus tschawytschac
(chinook salmon)/embryos
Oncorhynchus kisutch0
(coho salmon)/embryos
Oncorhynchus tschawytschac
(chinook salmon)/embryos
Oncorhynchus kisutchc
(coho salmon)/embryos
Poecilia reticulata0
Radiation regime
b°Co
0.5 R/d
33-40 R
60Co
0.5 R/d
33-40 R
Tritium
Comments
Significant increase in opercular defects in embryos given
0.5 R/d when compared to controls
In some phases, growth was superior in the irradiated (0.5 R/d)
group with no discernable pattern; a greater average weight in
the irradiated group was statistically significant
Total doses of 3400 and 4700 rad for young fish and of 380 rad
References
Donaldson and
Bonham, 1964
Donaldson and
Bonham, 1964
Erickson, 197:
(guppy)/embryos and young
Salmo qairdneriic
(rainbow trout)/embryos
Gasterosteus aculeatus0
(stickleback)/embryos
0.05 to 1 mCi/mL
340-4700 rad for young
fish and 90-2500 rad
for embryos
Tritium
0.01, 0.1, 1.0, and 10 yCi/mL
25 d
Tritium
0.5, 1.0, and 2.0 mCi/mL
980-3920 rad
increases in weight seen in males receiving 2500 rad as embryos;
increased proportion of males and earlier appearance of sex
characteristics in males receiving 90-rad total dose during
erobryogenesis"
No significant decrease in hatching; slight enhancement in
hatching at 0.01 and 0.1 uCi/mL; no significant increase
in malformations
Significant reduction in mean eye diameter in
1.0- and 2.0-mCi/mL water
Strand j2t al.,
1973b
Waiden, 1973
a Most studies on developing fish embryos directly exposed to radionuclides in the test water are not reported herein. They have been
critically reviewed by Woodhead (1984). and are discussed briefly in the text.
b Radiation regime is presented as source,, dose rate, and total dose (or exposure time). Units are those used by the author; R is the
abbreviation for roentgen.
c Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
-------
little to the field of aquatic radiobiology because of the dosimetry problems
described above. With regard to determination of low-effect levels, Woodhead
(1984) concluded the following:
Using the criterion of embryo survival (as measured by hatching
rate) after irradiation for the major part of embryogenesis, the
£fi 19725). Since Fedorova
(1972b) only gives data tor a single concentration of H, it is
not possible to be certain that 750 rad represents the lower limit
Sninthl9!!1!1"" eftec*on the hatching success of Coregonus peled.
»rt?i,f Icf ? 9^e" ^ the pa??r U can also be concluded that
actual dose to the embryo could be as much as 50% greater, i.e
1130 rad. Fedorova's data also provide confirmation that a
significant part of the total mortality is caused by a small
fraction of the total dose delivered in the early part of
embryogenesis, and also that the embryo becomes less sensitive
during the course of development.
ahnnvlh^ a^4°n1^ 2 sets of data on the Incidence of embryo
abnormality (Strand and co-authors, 1973b; Till, 1976 1978)
again the data of Strand and. co-authors, (19735) can only be
ed /6Serlat1opnS due to the clear lack of consistency
hW6eK ?S 2 exPer,!ments reported; the embryos receiving
absorbed d?use yie1ded a 1ower proportion of anomalous
than any other treatment, including the 2 control
poulations. Till (1976, 1978) observed that the lowest dose at
proport1on of abnormal
Once
49
-------
Woodhead (1984) also delineated the following improvements necessary for
future studies using radionuclides directly in the test water:
1. Full details must be given of the differential accumulation of the
radionuclide(s) by the eggs and the components of their environment,
as a function of time at each radionuclide concentration.
2. Estimates of the absorbed dose to the embryo and its variation
during development must be made, and full details of the derivation
given.
3. Sufficient replicates must be made to permit the relative
statistical significance of any observation to be determined.
4. A range of radionuclide concentrations must be employed so as to
allow the construction of a reliable dose-effect curve over the
range of 0 to 100% response. Experiments performed with this
degree of attention to detail will not be easy or straightforward.
Almost nothing is known about the effect of modifying environmental
factors on the response of fish embryos to radiation. However, the modifying
effects of dose rate and temperature on response of fish embryos to
irradiation were examined by Egami and Hama (1975). They found that 2000-R
exposures at early stages of development reduced hatching rate and increased
development time when the exposure rate was 250 R/min or 33.3 R/min, but not
at 1.7 R/min. Decreasing the temperature of the eggs during the exposure
period negated these dose-rate effects.
Developmental Effects from Acute and Chronic Irradiation of Invertebrates
Few studies exist that chronicle the effects of radiation on development
of invertebrate embryos (Tables 14 and 15). Blaylock and Trabalka (1978) cite
studies using radionuclides in the water. These will not be reviewed here
because the doses received by the test organisms were not documented
adequately. Studies by Raver a on Physa_ acuta (1968), cited in Blaylock and
Trabalka (1978), are described in the section on mortality. Similar to results
on fish, an increasing radiosensitivity with age was noted for this snail. The
50
-------
Table 14. Induction of developmental changes in invertebrates from exposure to acute levels of radiation.
Entries are ordered according to the lowest dose at which effects were observed. This dose is not
necessarily the lowest dose at which effects could have been observed.
Organism/1 ifestage
Radiation regimea
Comments
References
Diaptomus clavipes13
60
'Co
Significant decrease in % hatch at lowest
(calanoid copepod)/adults 7000 rad/min dose, 1000 rad
and embryos 1000-100,000 rad
Gehrs et al.,
1975
Physa acuta x ray
(freshwater snail)/embryos 200-10,000 R
Decrease in percent hatching seen at exposures Ravera, 1966
as low as 2000 R
en
Physa acuta
(freshwater snail )/embryos
Artemia salinac - 60Co
(brine shrimp)/cysts 12,000 rad/min
60,000-960,000 rad
LD5Q values for the four-cell, trochophore,
veliger, and hippostage embryos of the
gastropod Physa acuta were 1075, 2350, 4900.
and 10,750 R, respectively
Approximately 25% decrease in hatchability of
irradiated cysts at 360,000 rad
Ravera, 1968
(in Bl ay lock
and Trabalka,
1978)
Iwasaki, 1965
was not available. For x-ray data, a dose rate is not given; factors important in determining x-ray dose
rate are voltage, target material, filtration, tube current, and target-to-object distance. Units are those
_ used by the author; R is the abbreviation for roentgen.
Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
Organism is exclusively marine.
-------
Table 15. Induction of developmental changes in invertebrates from chronic
exposure to radiation.
Organism/1ifestage
Radiation
regimea
Comments
References
Physa heterostrophac
(aquatic snail)/
adults and embryos
60Co
1, 10, 25 rad/h
25 weeks
Significant decrease in
% hatching at 10 rad/h
but not at 1 rad/h; no
hatching at 25 rad/h;
growth of adults was
significantly increased
by 10 and 25 rad/h
Cooley and
Miller,
1971
a Radiation regime is presented as source, dose rate, and total dose (or
exposure time). Units are those used by the author.
b Organism is freshwater, anadromous, or estuarine as opposed to exclusively
marine.
lowest observed effect level was an LD5Q of 1075 R for embryos with four cell
stages. Gehrs et al_. (1975) administered 1000 to 100,000 rads to embryos of
the freshwater copepod Diaptomus clavipes. A significant decrease in the
hatching rate was observed at the lowest dose.
Conclusions on Development Data
Mortality of developing fish embryos from acute irradiation has been
observed at as low a dose as 16 R (Bonham and Welander, 1963; see mortality
section for further comments). Increased developmental abnormalities were
documented by Donaldson and Bonham (1964) after exposures of 0.5 R/d. They
initiated these 80-d experiments with one-cell chinook salmon embryos.
Similarly, .developmental effects have been observed in mammals at dose rates
as low as 2 to 3 R/d. In NRC (1980) are cited studies indicating increased
prenatal and postnatal mortality from exposures of 2.5 R/d in rats irradiated
from conception to term.
52
-------
Woodhead (1984) concluded that, with maximum dose rates (related to waste
disposal) of 0.240 rad/d observed in the ocean, fish embryos would not be at
increased risk. However, he also believed a final uncertainty remained, with
the lack of convincing work on the introduction of radionuclides directly into
the test water. We accept these conclusions, but also..feel that additional
research on the effects of modifying environmental factors is necessary to
determine the error associated with our current knowledge of low observed
effect levels. Detrimental effects in fish have been demonstrated at 0.5 R/d,
a value only twice the highest dose rates observed in the ocean. Considering
the findings (Egami and Hama, 1975) that low temperature may negate dose-rate
effects, it is conceivable that effect levels at low temperatures may be lower
than those discussed above.
Finally, we conclude that there is not sufficient information on chronic
or acute irradiation of developing embryos of aquatic invertebrates. More work
is needed in this area.
GENETICS
The genetic effects of ionizing radiation have been a principal concern
of researchers in mammalian radiobiology. Genetic effects are particularly
important because some effects are known to occur at low dose rates (NRC, 1980;
UNSCEAR, 1982; UNSCEAR, 1977). The major somatic and heritable genetic effects
of low-level radiation have been identified for mammalian populations (NRC,
1980; UNSCEAR, 1982; UNSCEAR, 1977). If one accepts the theory that mutational
events in somatic cells are primarily responsible for cellular transformation
and tumor formation, then the principal genetic effect of low-level radiation
in.somatic tissue is cancer induction. Concern over effects on germ cells of
low-level radiation centers on inducing increased frequency of heritable
genetic disease. This may occur from mutation, chromosome breakage,
chromosome rearrangement, or faulty segregation of chromosomes at metaphase
which may lead to aneuploidy. The significance of increased frequency of
neutral mutations is also the subject of considerable debate (Newcombe, 1971).
Gene mutations may have negative effects (cell death or genetic disease),
positive effects (increased population heterozygosity), or neutral effects
(those that cannot be characterized). Although evidence exists for increased
53
-------
population fitness due to positive or neutral mutations, most experts concur
that .increases in mutation rates should be considered detrimental (Newcombe,
1971; UNSCEAR, 1977;).
Impact'of increased frequency of heritable genetic diseases (due to gene
mutations, chromosomal damage, or aneuploidy) and their potential for
individual suffering has been of greater concern to those interested in human-
health effects than to those interested in environmental effects. However,
genetic effects on populations are of concern to those interested in both
human and environmental health. Many scientists predict that any mutation
causing deleterious effects in populations would rapidly be eliminated by
natural selection. This may be true, but it should be remembered that such a
statement does not presume no effect at the organismal level.
Dominant lethal mutations also occur as a result of radiation. These are
chromosomal or gene mutations that result from irradiation of developing sperm
or eggs and cause early embryo death. Such mutations have consequences at the
population level if a high percentage of embryos from a spawn are affected.
Viability of embryos produced from irradiated adult fish is an index of
dominant lethal mutations and is referred to earlier in this report.
Production of chromosomal aberrations, which may also result in cell
death, is a well-documented effect of radiation and is the result of breakage
with or without exchange of chromosomal material. When stable aberrations such
as trans!ocations occur in germ cells, they may be transmitted to subsequent
generations. Metaphase analysis of chromosomal aberrations in human
lymphocytes has been used for biological dosimetry on human populations.
Because the genetic effects of ionizing radiation are of central
importance in mammalian radiobiology, several investigators working with
aquatic organisms have realized the need for genetic-effects research on
aquatic organisms (Kligerman, 1980; Metal! 1, 1979; Harrison _et_al_., 1985).
The status of this research to date is summarized in the following sections.
General Considerations in Aquatic Genotoxicity Research
Selection of endpoints, model species, and other basic aspects of study
design are important general considerations in aquatic genotoxicity research.
To date, both mutations at a specific genetic locus and dominant lethal
mutations have been studied in aquatic organisms, and the induction of
chromosomal aberrations has been studied by metaphase chromosome analysis.
54
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Studies on the induction of specific locus mutations are limited by lack of
knowledge about the genetics of most aquatic organisms, as well as the long
time required and the great expense of conducting this type of experiment
(Kligerman, 1980). However, measuring dominant lethal mutations is relatively
easily achieved in a variety of fish and invertebrates in which gametes and
developing embryos may be readily obtained from irradiated adults.
A variety of cytogenetic endpoints are available to examine the effects
of radiation on aquatic organisms. These are summarized in Fig. 2. These
endpoints include analysis of chromosomal aberrations in metaphase and in
interphase by premature chromosome condensation, analysis of sister chromatid
exchange (SCE), the micronucleus assay, and the anaphase aberration assay.
Metaphase analysis of chromosomal aberrations is the most laborious, but it is
also the most highly recommended endpoint (Kligerman, 1979; Metalli, 1979).
Selection of species for cytogenetic studies has been discussed by
Kligerman (1979). He lists five criteria for a model organism for in vivo
cytogenetics:
1. A satisfactory karyotype
2. Tissues that yield adequate numbers of well-spread metaphases
3. Ability to withstand experimental conditions
4. Ease of obtaining and maintaining in the laboratory
5. Relatively small size
To this list, we must add a few more criteria. First, the cell kinetics
of the tissue to be studied should be known or be characterized in advance.
This is because at each cell division, certain percentages of aberrations or
micronuclei are lost from the cell. This should be taken into account when.
species or tissues are being considered for analysis. Because tissue kinetics
are often complex, it is also recommended that a single cell type within an
organism, rather than a mixed tissue, be used whenever possible. Finally, it
is preferable to select a tissue that does not have a restricted seasonal
availability. An example of such a tissue would be oogonial or spermatogonial
tissue in fish and invertebrates with highly seasonal reproductive cycles.
Satisfaction of each of these criteria in any one study may not be possible at
this time; however, these are important guidelines for future research.
55
-------
Cytogenetic Endpolnt Types of Damage
Advantages
Disadvantages
Analysis of
chromosomal
aberrations
Sister chromatld
exchange (SCE)
Hlcronucleus assay
Anaphase aberration
assay
Breakage or breakage •
and exchange of •
chromosomes
Exchange of chromosomal •
material between sister •
chromatids; significance •
is unknown, but SCE has
been correlated with
increased mutation
frequency
Chromosomal breakage and
spindle malfunction
causing laggard
chromosomes
Chromosomal breakage,
trans!ocations, spindle
malfunction, and chromo-
some stickiness
Very sensitive
Types of aberrations
induced can be identified
Very sensitive
Highly quantitative
Fewer cells required
than for metaphase
aberration analysis
Sensitive
Quantitative
Relatively simple
preparation and analysis
Relatively simple
preparation and
analysis
Tedious
Many cells required for
analysis
1 or 2 cell divisions must
occur in thymidine analogue;
this limits some field
applications
Biological meaning is still
uncertain
More sensitive to chemical
damage than radiation damage
Scoring may be complicated
in mixed tissues or cells
with large nuclei
Single-cell suspensions
are required
Knowledge of cell kinetics
is essential
Because anaphase figures
cannot be accumulated, use
of this assay is restricted
to rapidly dividing tissue
such as embryos
Different orientations of
anaphase figures may give
different results
Figure 2. Important endpoints in,Cytogenetic research and their implications
56
-------
Induction of Chromosomal Aberrations in Fish
Three studies were performed in which chromosomal aberration induction in
fish in vivo was examined (Table 16). In the first study, Kligerman et a]_.
(1975) introduced the fish Umbra limi as a model for the study of chromosomal
aberrations in fishes. This animal has a favorable karyotype of 22 large
metacentric and submetacentric chromosomes, and sufficient numbers of
metaphases from a variety of tissues are obtainable for analysis. They
administered a single exposure of radiation (325 R, x rays) to a group of fish
and compared aberration rates in this group to those in a control group. The
rate in the control was 0.03% aberrant metaphases per fish; in the
experimental group, it was 30% aberrant metaphases per fish.
Mong and Berra (1979) conducted a similar study on Umbra limj using
exposures of 350 R, 660 R, and 990 R from an x-ray source. They detected
9.2%, 13.8%, and 20.5% aberrant metaphases, respectively. In these two
studies, little attention was paid to important methodological details. For
example, in neither study was a clear breakdown of the types of aberrations
observed presented nor was it required that the entire chromosome complement
be visible, or that scored the cells be scored blind. Furthermore, mixed
tissues were analyzed without any discussion of cell kinetics or mitotic delay.
In the third in vivo study, Suyama et^al_. (1980) examined anaphase
translocation bridges induced by tritium exposure in embryos of the plaice
Limanda yokohamae. A significant increase over control values was detected in
the group exposed to 1 Ci/L for 22 h, but not at 0.1 Ci/L.
Two important in vitro studies (Table 16) were conducted on fish cells,
and significant effects were observed at the lowest radiations tested (50 R
and 75 rad). In the first study, Woodhead (1976) cultured Ameca splendens
embryonic tissue and delivered 75 to 1070 rad to these cells. In an attempt
to deal with cell kinetics, mitotic delay, and differential sensitivity at
different stages in the cell cycle, he sampled at 21, 45, and 69 h. He used
most of the characteristics of good cytogenetic study, such as blind scoring
and rigorous scoring criteria., in this work. In Fig. 3 is a summary of the
data and a comparison with other species. It is evident that the response
obtained was similar, but not identical, to that obtained by other
investigators for the toad Bufo marinus and human Homo sapiens cells.
57
-------
in
oo
Table 16. Induction of chromosomal aberrations in fish from acute and chronic exposure to radiation. Entries are ordered according to
the lowest dose at which effects were observed. This dose is not necessarily the lowest dose at which effects could have been
observed.
Organism/lifestage
Umbra limib
(central mudminnow)/
lymphocytes, in vitro
Ameca splendens (--)/
embryonic tissue,
in vitro
Umbra limi
(central mudminnow)/adults
Umbra limi
(central mudminnow)/adults
Limanda yokohamaec
(plaicej/embryos
Radiation regime3
x ray
50-200 R
50Co
560 ± 20 rad/min
75-1070 rad
x ray
325 R
x ray
350-990 R
Tritium
0.1, 1, 10 mCi/nt
22 h
Comments
Cultured lymphocytes were irradiated and examined for the presence
of dicentrics only; at the lowest exposure, 50 R, 16 dicentrics
were observed in 424 cells (0.038 dicentrics per cell); whereas none
were observed in 571 control cells
Aberrations significantly above control levels were detected at
the lowest dose, 75 rad, using embryonic tissue in vitro; response
level was comparable in sensitivity to that of other vertebrates
Chromosomal aberrations were observed in approximately 30% of
the metaphases examined per fish after an exposure of 325 R; the
control rate was 0.03%
9.2%, 13.8% and 20.5 % aberrant metaphases were detected
at 350 R, 660 R, and 990 R, respectively
Plaice embryos irradiated with 1 and 10 mCi/nt immediately after
fertilization showed significantly more mitoses with' translocation
bridges than did controls
References
Suyama and
Etoh, 1983
Woodhead, 1976
Kligerman et
a]_., 1975
Hong and Berra,
1979
Suyama £t al. ,
1980
a Radiation regime is presented as source, dose rate, and total dose (or exposure time). For x-ray data, a dose rate is not given;
factors important in determining x-ray dose rate are voltage, target material, filtration, tube current, and target-to-object
distance. Units are those used by the author; R is the abbreviation for roentgen.
b Organism is freshwater, anadromous, or estuarine as .opposed to exclusively marine.
c Organism is exclusively marine.
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In the second in vitro study, Suyama and Etoh (1983) examined x-ray-
induced dicentric yields in cultured lymphocytes of Umbra 1 imi. The dicentric
yields were significantly increased at 50 R, the lowest exposure tested.
However, only dicentrics were scored, and they were sometimes scored from
metaphase spreads in which only 20 or 21 of the 22 chromosomes were visible.
It is not clear whether or not the slides were coded for blind scoring.
Induction of Mutations in Fish
Induction of specific locus mutations and dominant lethal mutations have
been examined in a variety of studies on fish. However, in most of these
studies, relatively high exposures (500 to 1000 R) were used.
In studies on specific locus mutations, color-pattern mutation rates
(Purdom and Woodhead, 1973), changes in aggressive behavior (Holzberg and
Schroeder, 1972), and genetic regulation of melanophore genes (Anders e^al_.,
1973a,b) have been examined. The last study is the most notable because it
also is the only study we know of to demonstrate the induction of premelanomas
and melanomas in irradiated fish (Platypoecilus maculatus and Platypoecilus
variatus). These studies are summarized in Table 17. Effects were observed
from exposures between 500 and 1000 R. In view of the time required and
expense of conducting such large-scale breeding experiments, cytogenetic
endpoints and dominant lethal studies are more highly recommended for research
on aquatic organisms. Further information on mutation research in teleosts is
available in reviews by Schroeder (1979, 1980).
Rates of embryonic mortality and incidence of severe embryonic
abnormalities in embryos produced from irradiated adults or irradiated gametes
are indices of dominant lethal mutations. These are to be distinguished from
observations of these same effects in organisms irradiated as embryos; effects
from irradiation of embryos are indicative of developmental abnormalities.
Table 17 summarizes the many studies in which dominant lethal mutations
are examined (some studies in the section on reproduction have also involved
the analysis of dominant lethal mutations.) However, all except one used
relatively high exposures. The study that demonstrates the lowest effect
level is that of McGregor and Newcombe (1972a). They administered doses of 25
to 400 rad to rainbow trout Salmo gairdnerii sperm and examined the induction
of major eye malformations in F-, progeny. A significant increase was
observed at the lowest dose.
60
-------
Table 17. Induction of mutations in fish from acute and chronic exposure to radiation. Entries are ordered according to the lowest dose or dose
rate at which effects were observed. This dose is not necessarily the lowest dose at which effects could have been observed.
Organism/1 ifestage
Radiation regime-
Comments
Gambusia affinisb
(mosquitofish)/all stages
Salmo gairdnerii
(rainbow trout)/sperm
Salmo gairdneriib
(rainbow trout)/gametes
Poecilia reticulatab
(tropical guppy)/adults
References
White Oak Lake
0.59 rad/d
lifetime
60.
Co
64 rad/min
25-400 rad
x ray
200-20,000 rad
x ray
500 and 1000 R
Platypoecilus maculatusb and x ray
Platypoecilus variatusb 500-1500 R
(platyfish)/adults
Menidia menidiac
(Atlantic silverside)/sperm
Poecilia reticulatab
(tropical guppy)/adults
Cichlasonia nigrofasciatumb
(convict cichlidj/juveniles
x ray
800 R
60~
Co or x ray
1000-2000 rad
A significant increase in frequency of dead embryos in the irradiated
(0.59 rad/d) population versus control was demonstrated; this indicated
the presence of radiation-induced recessive lethal mutations maintained
in the population since initial studies in 1966
Major eye malformations were induced in F, progeny when sperm received
25 rad, the lowest dose tested; frequencies were linear between 25 and
400 rad, with a doubling dose of 54 rad; in contrast, decreased embryo
mortality was observed at 25 and 50 rad
Major malformations from exposure of eggs or sperm; at 200 rad, the
yield of malformations was 300 per million embryos per rad;
malformations appear to be associated with gross chromosomal changes
Vertebral column abnormalities were observed in F, progeny of adults
given 500 and 1000 R while germ cells were immature
Premelanomas and melanomas were induced at 500-1500 R in studies
on the genetic regulation of melanophore genes
Percent embryo mortality was 25 in control and 71 in eggs fertilized
with sperm exposed to 800 R
Color-pattern mutation rates were low in comparison with conventional
specific-locus rates in the mouse; however, very little data were
obtained (only 2 mutants at 1000 rad and 1 mutant at 2000 rad from
852 experimental animals)
x ray Aggressive behavior was reduced in FI male progeny of the Irradiated
500 R + 500 R, 24 h apart (1000 R) group
Trabalka and
Allen, 1977
McGregor and
Newcombe,
1972a,b
Newcombe and
McGregor, 19o7
Schroeder,
19o9a
Anders et
al_., 1973a,b
Engel £t al.,
1965
Purciom ana
Woodheao, 1*73
Holzberg and
Schroeder, 1972
-------
Table 17. (Continued.)
Organism/lifestage
Radiation regime"
Comments
ov
PO
Poecilia reticulata"
(tropical guppy)/juveniles
Poecilia reticulata0
(tropical guppy)/juveniles
Poecilia reticulata
(tropical guppy)/juveniles
and adults
Oryzias latipes
(medaka)/adults
Fundulus heteroclitus
(mummichog)/sperm
x ray
500 and 1000 R
x ray
1000 R
x ray
1000 R (oogonia and
spermato'gonia)
2 x 500 R-spermatozoa
x ray
2000-16,000 R
137CS
5000 R/min
500-150,000 R
In hybrid and inbred lines, litter size increased in the progeny of
irradiated parents; the frequencies of still births and post-natal
mortality were reduced in all the post-irradiation generations of the
inbred line whereas they were increased significantly in the F2
generation after spermatogonial irradiation at 1000 R in the hybrid
line; sex ratio was not affected by irradiation; the incidence of
mutations was higher in post-irradiation generations
Exposures of 1000 R to early spermatogonia did not increase the exchange
frequency between sex chromosomes, but irradiation of stem cells
provided a significantly higher incidence of crossing-over events
Synergistic interaction of recessive radiation-induced mutations was
studied; effects in post-irradiation F2 were greatest after exposure
of spermatozoa with 2 x 500 R followed by single doses of 1000 R to
spermatogonia and oogonia
Dominant lethal mutations were assessed by observing embryo mortality
in F-, progeny of irradiated adults; irradiation of sperm was more
effective in inducing dominant lethal mutations than irradiation of
oocytes; younger oocytes were more sensitive than older ones; effects
were observed at lowest exposure tested, 2000 R
Embryos developing from sperm exposed to 5000 R were stunted and
malformed (>50%); embryos exposed to high exposures were probably
haploid and developed by parthenogenesis
References
Schroeder,
19b9b
Schroeaer,
19b9c
Schroeder and
Holzberg,
Egami and
Hyoao-Taguchi,
1973
Lasher and
Rugh, 1962
a Radiation regime is presented as source, dose rate, and total dose (or exposure time). For x-ray data, a dose rate is not given; factors
important in determining x-ray dose rate are voltage, target material, filtration, tube current, and target-to-object distance. Units are those
used by the author; R is the abbreviation for roentgen.
b Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
c Organism is exclusively marine.
-------
Radiation-induced recessive lethal mutations have also been documented in
fish populations in White Oak Lake. Mosquitofish populations (Gambusia
affinis) irradiated for several generations produced embryos with a
significantly higher mortality percentage than those from a control population
(Trabalka and Allen, 1977). The dose rate administered to these organisms was
estimated to be 0.59 rad/d, although, as with other studies at White Oak Lake,
it is known that radionuclides are not the only contaminant in the lake.
Induction of Chromosomal Aberrations in Invertebrates
Induction of chromosomal aberrations in aquatic invertebrates was examined
in two sets of studies (Table 18). The first investigations were those of
Blaylock (1966a,b and 1973), who examined effects of irradiation on. polytene
chromosomes of chironomid midge larvae. Chironomus.tentans is an insect with
aquatic larval stages - the stages that were tested. Polytene chromosomes,
which do not require banding to visualize inversions and small deletions, are
particular to certain dipterari species. A 5-y study of chromosomal
polymorphism in chronically irradiated (0.63 rad/d) populations in White Oak
Lake demonstrated the presence of 10 inversions and 1 deletion that were never
observed in control populations. These aberrations were observed in only one
of the annual collections, and it was concluded that these mutations were
eliminated by natural selection. In contrast, six stable endemic inversions
were present in control and irradiated populations, and the frequency of these
endemic inversions was not increased in the irradiated populations. Although
no detailed population studies were conducted, it was known that C. tentans
was abundant in White Oak Lake after exposure to radiation of over 1000 times
background levels for over 100 generations.
The dose rates required to detect effects in the laboratory were higher
than those in the field. Blaylock (1973) exposed C. tentans larvae to tritium
(0.01 to 500 yCi/mL for 20 d) and found that aberrations were first detected
at dose rates of 38 rad/d with a minimum accumulated dose of 760 rad.
Harrison et_ al_. (1985) and Anderson et__al_. (in preparation) have made the
first attempts to examine chromosomal aberration induction in marine
invertebrates. Because no cell lines exist for any marine invertebrate,
studies were conducted in vivo using tissue prepared from embryo and juvenile
stages of the marine polychaete Neanthes arenaceodentata. Harrison et al.
(1985) examined the induction of chromosomal aberrations and SCE in embryos of
63
-------
Table 18. Induction of chromosomal aberrations in invertebrates from acute and chronic exposure to radiation. Entries are ordered according
to the lowest dose at which effects were observed. This dose is not necessarily the lowest dose at which effects could have been
observed.
Organism/1ifestage
Radiation regime
Comments
References
Chironorous tentans
(midge)/!arvae
Chironomus tentans
(midge)/larvae
Neanthes arenaceodentata
(polychaete)/!arvae
Neanthes arenaceodentata
(polychaete)/juveniles
White Oak Lake
0.65 rad/d
larval period
Tritium
0.01-500 uCi/nt
20 d
x ray
70 rad/min
8-380 rad
60Co
0.005 to 12 rad/h
0.1-300 rad
137Cs
500 rad/min
200-2000 rad
5-yr study of chromosomal polymorphism in chronically irradiated
(0.63 rad/d) population of C. tentans demonstrated the presence of 10
inversions and 1 deletion that were never observed in control populations;
these aberrations were observed only once and hence were probably
eliminated by natural selection; in contrast, 6 stable endemic inversions
were present in control and irradiated populations; the frequency of
these endemic inversions was not increased in the irradiated population
Aberrations were detected in salivary gland polytene chromosomes
at 125 yCi/mL and greater (760-3050 rad total dose or
38-153 rad/d)
Increased frequency of aberration with increase in dose; £ 200 rad
required to obtain significant change
Increased frequency of sister chromatid exchange; significant
increase observed at 60 rad
Increased frequency of chromosomal aberrations in juvenile worms
at 250 rad, the lowest dose yet tested
Blaylock, 1966a,b
Blaylock, 1973
Harrison et al.,
1985
Anderson et al.,
in prep
a Radiation regime is presented as source, dose rate, and total dose (or exposure time); for x-ray data, a dose rate is not given; factors
important in determining x-ray dose rate are voltage, target material, filtration, tube current, and target-to-object distance. Units are
those used by the author.
b Indicates organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
c Indicates organism is exclusively marine.
-------
N. arenaceodentata from acute irradiation; they found an increased frequency
of chromosomal aberrations with an increase in dose. Doses in excess of
200 rad were required to obtain a significant increase in aberration frequency.
An increased frequency of SCE with dose was observed, but it dropped off at
doses >100 rad; a significant; increase was seen at 60 .rad. Results of
research in progress with juvenile N. arenaceodentata show an aberration
frequency of approximately 0.3 per cell after 200-rad doses. In this study,
effect levels for the induction of mortality, decreased fecundity, and
chromosomal aberrations were compared. It was determined that significant
chromosomal aberration induction occurs at doses 2 to 3 orders of magnitude
below those causing death and at approximately the same doses as those that
cause reduction in fecundity.
Induction of Mutations in Invertebrates
To our knowledge, very little work has been done to examine the induction
in aquatic invertebrates of mutations by radiation. One investigator examined
induction of sterility in FI progeny of irradiated brine shrimp Artemia
salina (Table 19). No effects were observed in brine shrimp receiving 1000 R,
the lowest exposure tested; 3500 R induced 44& sterility in F, males.
In other studies on A. salina (Metal!i and Ballardin, 1962, 1972;
Ballardin and Metalli, 1965), diploid and tetraploid strains were exposed to
1000 R from an x-ray source, and hatching success was examined as an index of
dominant lethality. For the diploid strain, a reduction in hatching success
was seen in the second generation as well as the first, presumably due to the
expression of recessive lethal mutations. For the tetraploid strain, no such
reduction in hatching success occurred in the second generation. An estimated
mutation rate of 3.9 x 10" gamete" rad'1 was obtained for the diploid strain
in the first generation.
Conclusions on Genetics Data
Genotoxic effects of radiation exposure in fish and aquatic invertebrates
have just begun to be examined. Yet, these same effects in mammals are a prime
concern of mammalian radiobiologists. No genetic studies have addressed
chronic irradiation effects on fish, and only one chronic effects study has
been conducted on an aquatic invertebrate (Blaylock 1966a,b). Moreover, the
65
-------
Table 19. Induction of mutations in invertebrates from acute exposure to
radiation.
Organism/
1 ifestage
Radiation
regimea
Comments
References
Artemia salina
(brine shrimp)/
adults
137Cs
507 R/min
1000-10,000 R
44% sterility in F-, sons of
males at 3500 R was observed;
5000 R produced complete
sterility whereas 1000 R
produced no significant effect;
significant effects on survival
to adulthood also occurred at
3500 R and above
Squire, 1973
a Radiation regime is presented as source, dose rate, and total dose.
are those used by the author; R is the abbreviation for roentgen.
Units
Organism is exclusively marine.
invertebrate examined was the freshwater larva of an insect Chironomus
tentans. The lowest effect levels observed to date are (1) the observation by
Suyama and Etoh (1983) of a significant increase in dicentric yield in
cultured lymphocytes of the fish Umbra limi after 50-R exposures and (2) the
induction of major eye malformations in trout embryos after 25-R exposures to
sperm (McGregor and Newcombe, 1972a). It is important to note that data
obtained to date on genotoxic effects indicate that the sensitivity of some
aquatic organisms with respect to cell killing and DNA damage is not less than
that observed in mammals. No investigators have examined the role of
•modifying environmental factors on genotoxic effects.
Further analysis of the genetic effects of ionizing radiation on aquatic
organisms is needed. However, careful attention must be paid to selecting
endpoints and species to be examined. Moreover, greater attention to
experimental design and conduct is necessary to improve the standards of some
of the cytogenetic work.
66
-------
Metalli (1979) emphasized the need for relating studies of genetic effects
to endpoints such as reproduction to aid in extrapolating and generalizing
genetic data. It was also proposed that the analysis of dominant lethal
mutations and metaphase analysis of chromosomal aberrations were the endpoints
of choice for future study. He had some reservations about these endpoints,
however, and stated that under chronic exposure regimes, non-genetic factors
such as meiotic delay may affect the results obtained in dominant lethal
studies. He also questioned the expense of chromosomal aberration studies,
which were the most highly recommended type of study. In response, we suggest
that a portion of future cytogenetic research should use the induction of
micronucleis as an important endpoint, because it is less time consuming to
quantify than chromosomal aberrations (see Fig. 2 for comparing types of
effects detected).
Finally, it must be emphasized that further work in this area will
require a substantial research commitment. Even when optimal systems are
selected, chromosomal aberration and dominant lethal studies are labor
intensive. However, the importance of such work is difficult to contest.
COMPARISON OF RADIATION EFFECTS ON AQUATIC ORGANISMS
TO THOSE ON TERRESTRIAL ORGANISMS
Although there are not sufficient data on aquatic organisms to determine
the threshold dose rates below which effects would not be observed, we can
obtain an indication of what these dose rates might be by comparing the results
obtained from exposure of terrestrial animals to those obtained on aquatic
organisms. In Fig. 4, low-level effects on mammals are contrasted to those on
fishes and aquatic invertebrates, and in Table 20, the critical data on low-
level effects on aquatic animals and examples of selected low-level effects on
various species of terrestrial animals are provided.
MORTALITY AND PATHOPHYSIOLOGY
To our knowledge, no mortality due to acute irradiation below 10 rad
(0.1 Gy) occurs in any animal,, Excluding tumors, induction of acute
pathophysiologic lesions also occurs at higher doses. The significance of
such effects in the deep ocean is equivocal. Because of the long latency
period for cancer, long-lived species may be the only organisms at risk.
67
-------
Acute Dose Level
10 rads
100 rads
1000 rads
Mortality
Aquatic invertebrates
Fish
Mammals
Pathophysiology
Aquatic invertebrates
Fish
Mammals
Mammals (tumors)
Reproduction (oocyte sensitivity)
Aquatic invertebrates
Fish
Mammals
Development
Aquatic invertebrates
Fish
Mammals
Genotoxic Effects
Aquatic invertebrates
Fish
Mammals
Figure 4. Generalized effect levels for selected biological endpoints,
68
-------
REPRODUCTION
Gamete death has been demonstrated at low doses in mammals (Fig. 4 and
Table 20). It is also known that these response levels are highly species
specific. Despite the large amount of controversial data from research on
effects of radiation on developing fish embryos, little has been done to
define the shape of dose-response curves for gamete killing in fishes and
invertebrates. The research that has been done (Table 20) indicates that
effects may occur at levels on the order of 1 rad/d (1 x 10"2 Gy) or less.
DEVELOPMENT
The production of developmental abnormalities in fish embryos from low
doses of radiation has been the topic of considerable debate. Recently, this
area of research was reviewed critically by Woodhead (1984). He concluded that
effects observed on fish embryos at the lower dose levels from radionuclides
in the water are not verifiable. However, embryo death has been induced in
fishes with exposures as low as 16 R, and increased developmental abnormalities
have been found (with the use of sealed sources) in fish at 0.5 R/d. In
contrast to the extensive research on fish, few data are available on the
effects of radiation on the development of invertebrates. The data on mammals
demonstrate that effects of low levels (3 rad/d) of radiation occur in utero
(Table 20) and that they are keyed to critical periods in development.
GENETICS
The significance of effects on genetic material from exposure to low
levels of radiation has been debated widely by geneticists. Effects of
low-level radiation include increased gene mutation rates, chromosome breakage
and rearrangement, and aneuploidy. Production of chromosomal aberrations,
which may also result in cell death, is a well-documented effect of radiation
and is the result of breakage or combined breakage and exchange of chromosomal
material. When stable aberrations such as translocations occur in germ cells,
they may be transferred to subsequent generations. A significant increase in
chromosomal aberrations 'in mammalian lymphocytes in culture has been
demonstrated in people living in areas with natural high background
irradiation (0.11 to 0.34 R/y gamma-ray plus 0.001 to 1.6 R/y alpha ray
69
-------
Table 20. Summary of biological effects observed at low dose rates and low total doses in aquatic organisms as compared to
selected data for terrestrial organisms.
Dose rate or total dose
(Source)
Observed or projected effect
Reference
0.5 R/d (60Co)
0.59 R/d (mixed source,
White Oak Lake
sediment)
0.65 R/d (mixed source,
White Oak Lake
sediment)
4.08 rad/d (137Cs)
6.8 rad/d (137Cs)
>10 R/d (60Co)
Aquatic organisms
Increased incidence of opercular defects in salmon after 80 d of continuous
irradiation beginning immediately after fertilization; a significantly
greater average weight in the irradiated group was also observed
Increased embryo mortality in the mosquitofish Gambusia affinis relative to
those from an unirradiated field control site; no concomitant decrease in
fecundity was observed
Frequency of egg-capsule production was reduced in an irradiated population of
the aquatic snail Physa heterostropha; however an increased number of eggs per
capsule also occurred
Reduced total fecundity at lowest dose tested over the entire life cycle of
the guppy Poecilia reticulata
Increased percentage of unfertilized eggs from ricefish Oryzias latipes
adults irradiated for 120 d; no statistically significant effects were
observed at lower doses
Gonadal development retarded in Chinook salmon smolts irradiated for 80 d
after fertilization; lower exposures showed no such effects; the percent
return of migrating salmon was slightly higher in the group irradiated
at 0.5 R/d
Donaldson and
Bonham, 1964
Trabalka and
Allen, 1977
Cooley, 1973
Woodhead, 1977
Hyodo-Taguchi,
1980
Bonham and
Donaldson, 1972;
Donaldson and
Bonham, 1970
-------
Table 20. (Continued.)
Dose rate or total dose
(Source)
Observed or projected effect
Reference
Aquatic organisms (continued)
16 R (x ray)
25 rad (60Co)
60,,
25 and 50 rad (ouCo)
LDj-Q of silver salmon (Oncorhynchus kisutch) embryos irradiated in the
one-cell stage and observed 150 d after irradiation
Increased frequency of major eye malformations in trout embryos from
irradiation of trout sperm
Trout sperm at 25 and 50 rad produced decreased embryo mortality; increased
embryo mortality from 200-400 rad to sperm
Terrestrial organisms
a,b
Bonnam and
Welander, 1963
McGregor and
Newcombe, 1972a
McGregor and
Newcombe, 1972b
0.11-0.31 R (y rays) plus Increased frequency of chromosomal aberrations in human lymphocytes
0.001-1.6 R (a rays)
Pohl-Ruhling and
Fischer, 1979
0.043-0.43 R/d (y rays) Increased percent sterility in male dogs exposed to 0.043 or 0.086 R/d compared- Luckey, 1980
to control; whereas exposure to 0.43 R/d made male dogs sterile in one year
0.3 rad/d (HTO)
Decreased brain weight in Fo generation of rats irradiated from conception
to term
NRC, 1980
1 R/d (Y rays)
Decrease in percent survival in first 6 months of life in desert rodent
Perognathus formosus; thereafter, percent survival was increased relative
to controls
Whicker and
Schultz, 1982
TR/d (137Cs)
Significant increase in mutation rates in mouse spermatogonia versus controls UNSCEAR, 1977
-------
Table 20. (Continued.)
Dose rate or total dose
(Source)
Observed or projected effect
2 rad/d (y rays)
2.5 R/d (60Co)
3 rad/d (HTO)
No significant differences in sex ratios or egg distributions observed in
populations of the the lizard Uta stansburiana
Reduced brain, testis, and ovary weights in rats irradiated from conception
to term
Reference
Terrestrial Organisms (Continued)
1.1-1.37 rad/d (Y rays) Sterility induced in leopard lizard and whiptailed lizard from lifetime
irradiation
Whicker and
Schultz, 1982
Whicker and
Schultz, 1982
Prenatal and postnatal mortality observed in rats irradiated from conception NRC, 1980
to term
NRC, 1980
3.3 rad/d (HTO)
5 rad (HTO) and
8 R (x ray)
8 rad (HTO)
8.4 R/d (137Cs)
No effect on lifespan of rats irradiated from conception to term
LD50 of mouse oocytes
LDc0 of monkey oocytes
Complete sterilization of female mice irradiated 20-40 d postconception
NRC, 1980
NRC, 1980
NRC, 1980
NRC, 1980
presence of positive and negative effects at low dose rates. Other examples for terrestrial organisms are given in Luckey,
(1980); NRC (1980); UNSCEAR (1977); UNSCEAR (1982); and Whicker and Schultz (1982).
Many examples selected for the section on terrestrial organisms are cited from secondary references.
-------
exposure; Pohl-Ruhling and Fischer, 1979) and an increased dicentric yield in
cultured fish lymphocytes was demonstrated at 50 R, the lowest exposure
tested. Genotpxic effects of radiation may result in decreases in
reproductive success of an organism due to gamete killing and dominant lethal
mutations.
LOW-LEVEL RADIATION EFFECTS AND HUMAN HEALTH
The major somatic and heritable genetic effects of low-level radiation
have been identified for mammalian populations (NRC, 1980; UNSCEAR, 1977;
UNSCEAR, 1982). The principal effect of low-level radiation in somatic tissue
is probably cancer induction in a variety of organs and tissues. Also, gamete
death and developmental abnormalities to the embryo and fetus have also been
observed at low doses, below 10 rad.
Concern over effects on germ cells of low-level radiation centers on the
induction of increased frequency of heritable genetic disease. This may occur
from mutation, chromosome breakage, chromosome rearrangement, and faulty
segregation of chromosomes at metaphase which may result in aneuploidy. The
significance of increased frequency of neutral mutations is also the subject
of considerable debate.
CONCLUSIONS ON EFFECT LEVELS
Examination of the data on biological effects in aquatic invertebrates,
fishes, and mammals at dose rates of approximately 0;5 to 10 rad/d (5 x 10"3
to 1 x 10" Gy/d) indicates that those dose rates may define a range in which
a variety of low-level effects on reproduction, development and genetic
integrity are detectable in sensitive tissues and organisms. It is possible
that this range may encompass; threshold dose rates for important non-stochastic
effects. Furthermore, for aquatic organisms, it is apparent that little
information exists that is relevant to the role of factors modifying response
levels. Factors such as temperature, species specificity, and cell kinetics
may alter observable-effect levels by more than an order of magnitude and thus
affect the certainty of predicted effects. Finally, it should be emphasized
that the dose range at which data on effects are available is higher than
doses measured at currently existing radioactive waste-disposal sites, but the
low end of this dose range is only twice the highest dose rates recorded in
73
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the ocean due to radioactive waste disposal. Our section on population
effects includes a discussion of the importance of decreasing error in effect
estimates and the utility of single-species toxicity tests in assessing
environmental hazards.
EFFECTS ON POPULATIONS AND COMMUNITIES FROM CHRONIC EXPOSURE
TO IONIZING RADIATION
It is stated repeatedly in the aquatic radiobiology literature
(Templeton, 1976a; Whicker and Schultz, 1982; Woodhead, 1984) that effects on
individuals are not as important for aquatic species as they are for humans.
This is certainly true, but it must be remembered that it is general practice
in aquatic toxicology that no-effect levels are first determined at the
organismal level. That is, discharge criteria and pollution standards are set
from effect levels determined in single-species toxicity tests. These effect
levels are generally results from standard assays measuring mortality and
reproductive effects. Compensation arguments, such as those used in the
aquatic radiobiology literature (e.g., that the death of a percentage of a
brood is insignificant if the remaining percentage can support a population),
are important points of discussion, but, to our knowledge, they are not .
generally used to set environmental criteria and standards.
VALUE OF SINGLE-SPECIES TOXICITY TESTS IN THE EVALUATION OF EFFECTS ON
POPULATIONS AND COMMUNITIES
The adequacy of single-species toxicity tests in estimating environmental
hazard is one of the most important considerations in ecotoxicology. On this
subject, Cairns (1983) recently concluded that there has not yet been
sufficient study to determine the degree of reliability of these tests. He
believes there is a great need for more testing at the community level, but
also states that single-species toxicity tests are presently the major and
only reliable means of estimating environmental hazard.
Cairns (1983) presented a diagram (Fig. 5) showing how the error about an
environmental standard or criteria is decreased as more steps are taken to
test for no-effect levels. Multiple levels of testing are not required when
expected environmental concentrations are much lower than no-effect levels,
unless, for some reason, an extremely low level of error is necessary. This
74
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8!
li
•§!
M'j
1!
II
.2!
I
ii
o
o
Confidence intervals
• Highest test concentration;
producing no
biological effects.
Confidence intervals
• Highest expected
environmental
concentration.
i
2
i
3
i
4
i
5
I
6
Sequential tests of hazard assessment procedure
Figure 5. The relationship between error of .vffect-level estimates and stages
of investigation (Cairns, 1983). The dashed lines represent confidence
intervals around the line representing the highest expected concentration.
75
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is probably the situation that exists with regard to dumping low-level wastes
in the ocean. The recent NACOA report (1984) emphasized the need for societal
consensus on nuclear waste management and its effect on decision making
regarding dumping radioactive wastes into the ocean.
Many investigators performing ecotoxicological research have argued that
single-species toxicity tests are inadequate for estimating environmental
hazard and that ecological factors such as food density and community
interactions may greatly decrease the level at which environmental effects can
be detected (e.g., Gushing, 1979; Gray, 1979; Kooijman and Metz, 1984).
Perkins (1979) has argued the need for a variety of sublethal tests.
Other investigators have found that single-species tests are predictive
of community effects. Recent studies have compared - effect levels of chemical
contaminants using organismal-, population-, and community-level endpoints.
Hansen and Garton (1982) exposed laboratory stream communities to the
insecticide diflubenzuron for 5 months and assessed changes at the community
level using biomass and diversity estimates. They then conducted acute and
chronic toxicity tests on several fish and invertebrate species present in the
streams. It was found that single-species tests adequately predicted the
diflubenzuron concentrations affecting stream communities, but that the
concentrations eliciting responses in the most sensitive test species were
more than an order of magnitude lower than those resulting in the observed
community effects. Because of the short duration of the tests on communities,
however, they were not able to assess the sensitivity of important
population-level reproductive effects.
A recent EPA study (Mount et_ al_., 1984) showed that toxicity testing
using ambient waters was predictive of instream community response to pollution
in the Ottawa River. In this study, fish and benthic invertebrate samples were
taken at nine stations along the river. A sewage-treatment plant was located
between stations 2 and 3, a refinery between stations 3 and 4, and a chemical
plant between 4 and 5. In parallel, water samples were taken at these
stations, and the water was used to run reproductive bioassays on
Ceriodaphnia sp. A similar pattern (Fig. 6) was found between number of
benthic species represented at a station and the number of young per female
Ceriodaphnia sp. obtained in bioassays. This is a particularly interesting
result because the most classical combined organismal and population studies
in the aquatic radiobiology literature were conducted on Daphnia pulex; these
studies are described in the next section.
76
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TD
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(U
00
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S3
O
o
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o
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In summary, there is as yet no consensus on the use of community-level
testing in aquatic toxicology, and these tests are generally considered as
second-tier.tests, to be used after no-effect levels are determined at the
organismal level. Therefore, it is concluded that determination of no-effect
levels using relevant organismal-level endpoints, especially reproduction,
should remain the highest research priority for at least the next few years.
EFFECTS OF IONIZING RADIATION ON POPULATIONS OF AQUATIC ORGANISMS
A variety of perspectives on the effects of ionizing radiation on
populations of aquatic organisms is presented in reviews by Templeton (1976a),
Blaylock and Trabalka (1978), and Woodhead (1984). In a recent review, Whicker
and Schultz (1982) examined the effects of radiation on terrestrial and aquatic
populations. In this review, we will briefly discuss the results of
population-level studies on aquatic organisms (Table 21) and compare effect
levels to those observed for terrestrial organisms.
The classic studies of Marshall (1962, 1966, and 1967) are the first and
only studies in the aquatic radiobiology literature to use quantitative
population biology to study radiation effects. The basic findings of these
extensive studies are presented in Table 21. Marshall (1962) determined that
the dose rate at which the intrinsic rate of natural increase, r, was reduced
in chronically irradiated Daphnia pulex populations was 70 R/h. Birth rate
fell sharply at 50 R/h, whereas death rate was barely affected at these dose
rates. Marshall (1966.) found that for D. pulex populations chronically
irradiated for 55 weeks, populations exposed to 436 R/d or more became extinct.
Marshall (1962) established that the maximum tolerable exposure rate was
1330 R/d. The difference in response was due to food-supply limitation in the
second study. In a subsequent study, Marshall (1967) examined the effects of
exploitation on food-limited, irradiated D. pulex populations. He found that,
up to a certain level, exploitation had an ameliorating effect on radiation
stress. These studies would seem to indicate that very high chronic exposure
rates might be tolerated by invertebrate populations. However, evidence for
reproductive and genetic effects below these levels has been given by several
authors.
Studies conducted at White Oak Lake, which have been discussed in other
sections, demonstrated reproductive effects in snail and fish populations at
dose rates below 1 rad/d (Cooley, 1973; Trabalka and Allen, 1977) and genetic
78
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Table 21. Induction of effects on populations of aquatic organisms from chronic exposure to radiation. Entries are ordered according to the lowest
dose at which effects were observed. This dose is not necessarily the lowest dose at which effects could have.been observed.
Organism/lifestage
Oncorhynchus tschawytschab
(chinook salmon)/embryos
Gambusia affinis
(mosquitofish)/all stages
Chironomus tentansb
(midge)/larvae
PJiysa heterostropha
(aquatic snail)/adults
Daphnia pulex
(daphnia)/complete life
cycle
Radiation regime
60Co
0.5-1.3 R/d
40-104 R
White Oak Lake
0.59 rad/d
lifetime
White Oak Lake
0.65 rad/d
larval period
White Oak Lake
0.65 rad/d
lifetime
60Co
0-516 R/d; populations
irradiated 18.5 h/d for
55 weeks
Comments
At 0.5 R/d slightly higher rates of return in migrating salmon
were observed
No decrease in fecundity observed at 0.59 rad/d; a significant
increase in frequency of dead embryos in the irradiated
population versus control was demonstrated; this indicated
the presence of radiation-induced recessive lethal mutations
maintained in the population since initial studies in 1966
5-y study of chromosomal polymorphism in chronically irradiated
(0.65 rad/d) population of C^ tentans demonstrated the presence
of ten inversions and one deletion that were never observed in
control populations; these aberrations were observed only once and
hence were probably eliminated by natural selection; in contrast,
6 stable endemic inversions were present in control and irradiated
populations; the frequency of these endemic inversions was not
increased in the irradiated population
Frequency of egg-capsule production was reduced in. the irradiated
(0.65 rad/d) population, but an increase in number of eggs per
capsule occurred, resulting in similar rates of egg production
compared to controls
Populations exposed to 436 R/d or more became extinct
References
Bonham andvUonaloson'
19bb; Donaldson and
Bonham, 1970
Trabalka and Allen,
1977
Blaylock, 1966a,b
Cooley, 1973
Marshall, 1966
-------
Table 21. (Continued.)
oo,
o
Organism/I ifestage
Daphnia pulex
(daphnia)/complete life
cycle
Daphnia pulex
(daphnia)/complete life
cycle
Artemia salina0
(brine shrimp)/complete
life cycle
Radiation regime
60Co
22.8-75.9 R/h
19 h/d
20-35 d
Comments
Exposure rate at which intrinsic natural rate of increase was
reduced to zero was 70 R/h; birth rate fell sharply at 50 R/h
x ray Effects of exploitation on food-limited D. pulex were studied;
3.7-5.1, 108-162, exploitation decreased radiation effects observed in food-limited
270-378, and 468-684 R/d populations
80-100 weeks
x ray
1000, 2500, 3000 R/
few min (irradiated
3 times per y for 2 y)
Culture survival only at 1000 R; no life-span change at 1000 R,
but decrease in number of zygotes voided and number of mature
adults per pair
References
Marshall, 1962
Marshall, 1967
(irosch, 1962
* Radiation regime is presented as source, dose rate, and tota! dose (or exposure time). For x-ray data, a dose rate is not given; factors
important in determining x-ray dose rate are voltage, target material, filtration, tube current, and target-to-object Stance. Units are
used by the author; R is the abbreviation for roentgen.
b Organism is freshwater, anadromous, or estuarine as opposed to exclusively marine.
c Organism is exclusively marine.
-------
effects in Chironomus tentans populations at similar dose rates (Blaylock,
1966a,b). Effects at these low levels have not been determined in the
laboratory. However, reproductive effects between 1 and 10 rad/d have been
demonstrated in the laboratory (Woodhead, 1977; Hyodo-Taguchi, 1980).
Templeton (1976a) has stated that highly fecund fish species with rapid
development rates would be at less risk to radiation exposure than low-
fecundity fish species with slow development rates. This hypothesis has not
been tested on fish, but Turner et_aJL (1973) tested this hypothesis in
populations of lizards chronically irradiated in the field. Some species of
lizards became sterile after long-term irradiation at 1.1 to 1.3 R/d, whereas
other lizard species were unaffected by the same radiation exposure. These
differences in responses of the populations were attributed to differences in
population dynamics rather than differences in gamete sensitivity. It was
found that slow-developing, low-fecundity species were most affected by
chronic irradiation because higher doses could be accumulated. Sterility has
also been induced in dogs with chronic exposures of 0.43 R/d (Luckey, 1980).
Much evidence exists for Induction of hormesis from low levels of ionizing
radiation (Luckey, 1980). Intriguing results (Donaldson and Bonham, 1970),
indicating an increased return rate of salmon that had been irradiated as
embryos at 0.5 R/d versus unirradiated controls, are frequently given as
evidence of hormesis. However, in the same series of studies, Donaldson and
Bonham (1964) demonstrated an increase in opercular defects in embryos at the
same dose rate.
PROVISIONAL DOSE ASSESSMENTS FOR DEEP-SE'A ANIMALS
In risk-assessment procedures, knowledge of potential biological effects
and effect levels for a contaminant are combined with information about the
fate of the contaminant in the environment and within organisms. This report
focuses on the biological effects of low-level radiation on aquatic organisms.
However, currently used provisional dose assessments for deep-sea animals are
compared in this section to the biological effects levels we have described.
Woodhead and Pentreath (1983) made an assessment of the doses that may be
received from natural background radiation by deep-sea organisms (Table 22)
and compared these doses to those received by coastal organisms. Their
calculations indicate that the natural radiation background of deep-sea
organisms is likely to be as high and at least as variable as that of coastal
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Table 22. Estimates of the radiation dose rates (nSv/h) to benthic deep-sea
organisms from natural background and the peak dose rates predicted
from dumping low-level, radioactive wastes (OECD/NEA, 1985).
Fish
Large
crustaceans
Molluscs
Small
crustaceans
Background 8.5 x 101 to 9.9 x 102 to
9.3 x 102 to 3.3 x 102 to
1.4 x 10*
Past dumping 7.3 x 10
Past + future 1.0 x 102
dumping
r
Past + future 6.2 x 10^
dumping x 10
1
4.6 x 10*
3.8 x
5.0 x 10'
2.9 x 10*
3.4 x 10*
8.0 X I'O*
1.0 x 10
6.5 x 10
4.2 x
1.3 x
1.7 x 10*
9.7 x 10*
a Radiation dose based on the assumption that the radionuclides are
deposited in a rectangular box 40 km x 120 km x 75 m deep and include
the underlying sediments.
and shallow-water organisms, and they concluded that deep-sea organisms may
have evolved, therefore, in a radiation regime similar to that of coastal
organisms. However, extrapolation of results of radiation effects on coastal
organisms to those on deep-ocean organisms should be made with caution for the
following reasons:
1. Models used to estimate the radiation doses to deep-ocean organisms
were simplistic because insufficient data were available on
radionuclide (particularly alpha emitters) distribution in the
tissues (particularly gonadal tissue) of the organisms.
82
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2. Information on the dose-effect relationships for different classes
of deep-ocean organisms is not available. Therefore, it is not known
how the effect of dose would be modified by such environmental
factors as low temperature, low oxygen tension, high pressure, etc.
3. Evolution in a similar radiation regime may not result in similar
radiosensitivity. For example, the mouse and the rat have evolved in
similar radiation regimes, but the sensitivity of the rat and mouse
oocytes to radiation differ significantly.
As more information becomes available on the internal distribution within deep-
sea biota and the external distribution in the environment of radionuclides of
concern, model predictions can be refined, and the conservatism required to
ensure the protection of the environment can be reduced.
A considerable amount of attention has been given to the prediction of
the dose to organisms indigenous to the Northeast Atlantic Disposal Site,
which has been receiving low-level radioactive waste since 1949. The
feasibility of the continued use of this site has been under evaluation
(OECD/NEA, 1985). Radiation closes to bathypelagic and benthic deep-sea
organisms have been calculated assuming that the radionuclides are deposited
in a rectangular box 40 km x 120 km x 75 m deep, a box 250 km x 250 km x
500 m, and a box 2500 km x 3500 km x 1000 m deep. The boxes include the
underlying sediment. Results of the calculations for the smallest box and for
benthic organisms are given in Table 22. These results are the highest of the
three and their use would provide conservative estimates of potential
detrimental impact.
The highest dose rates predicted are those received by the benthic
molluscs at the site; these are greater than the maximum estimated for natural
background. As a result of using the most conservative model, the results of
calculations indicate that benthic molluscs would be. exposed to 6.5 x 104
nSv/h or 0.156 rem/d. According to data we have tabulated, this dose rate is
approximately an order of magnitude below the level at which the most subtle
reproductive and genotoxic effects are noted.
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ADDITIONAL METHODOLOGIES FOR THE STUDY OF RADIOBIOLOGICAL EFFECTS
Because the effects of radiation exposure in mammals have been of
widespread concern, many methodologies for studying radiobiological effects on
genetic material have been developed. Some of these approaches have never
been applied to the study of effects on aquatic organisms, and three, in
particular, may provide very useful information.
Analyzing the quantity of DNA single-strand breaks caused by radiation
exposure is possible because damaged DNA unwinds more rapidly than intact DNA
when cell lysates are exposed to alkaline conditions. DNA unwinding in
treated-cell lysates is monitored on a fluorometer. By using this and similar
techniques on appropriate tissues from aquatic organisms, investigators may be
able to detect effects from radiation exposures as low as 5 to 10 rad or
0.05 to 0.1 Gy (Birnboim and Gevcak, 1981). Furthermore, the relative rates
of strand breakage and repair could be studied in different aquatic organisms,
provided appropriate tissues are identified. The drawbacks of research in
this area are (1) that two to three years of development would be required to
adapt the technique and to identify the best tissues for-study and (2) that a
large proportion of DNA single-strand breaks are repaired (hence both repair
and recovery would have to be studied).
Unscheduled DNA synthesis (UDS), studied as an indicator of DNA repair
activity, is generally measured by analyzing 3H-thymidine uptake in exposed
cells or tissues. Uptake is usually quantified by autoradiography, although
other methods exist. Recently, Tuschl et al_. (1983) reported increased rates
of UDS in lymphocytes of persons occupationally exposed to greater than
0.014 rad/month over background levels. Development of this technique could
aid in the detection of genetic effects from low-level radiation exposure as
well as provide means for comparing relative rates of DNA repair among aquatic
organisms and between aquatic organisms and mammals. A minimum of two to
three years of development would be required before these kinds of comparisons
could be addressed.
The third technique, premature chromosome condensation, involves the
fusion of interphase cells from a test organism with a second type of cell
that is rapidly dividing. The effect is that interphase chromosomes may be
induced to condense prematurely such that chromosomal aberrations can be
visualized (Pantelias and Maillie, 1983). This technique has distinct
84
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relevance to invertebrate radiobiology because it enables the investigator to
directly analyze chromosomal damage without complications in interpretation
arising from interphase death of cells, mitotic delay, or cell repopulation.
The first step in the development of such an approach would be the selection
of appropriate tissues for fusion, This is a difficult first step because, to
our knowledge, fusion of marine invertebrate cells has only been conducted on
sea urchin embryos.
Other important assays of DNA damage exist that have not been mentioned
here. Developing any of these assays for application to aquatic organisms
will require a commitment to basic research before meaningful answers are
provided. However, such fascinating and important basic issues (which also
have practical predictive value) as the relative abilities of different
members of the animal kingdom to repair DNA await analysis.
MONITORING APPROACHES
The purpose of this review has been to examine the current state of
knowledge relevant to determining the potential biological effects of low
levels of ionizing radiation on aquatic organisms. The information provided
can be used by others to evaluate the potential hazard of dumping low-level,
radioactive waste into the ocean. An integral part of aquatic toxicological
hazard assessment is the determination of NOELs for a contaminant under
study. Exposure values are then incorporated into models to predict the
impact on a specific area of a given contaminant concentration. Next,
cost-benefit analyses are made to weigh the social and economic values of the
activity against the environmental risks. If a decision to resume dumping of
low-level radioactive wastes in the ocean is made on the basis of knowledge of
the effect levels, then a monitoring strategy would serve to check the
accuracy of predicted effects. Thus, development of monitoring methodologies
is a secondary goal, whereas the determination of no-effect or minimal-effect
levels is a primary goal in the decision-making process.
There are two general approaches to monitoring. The first is to directly
examine communities in impacted and control areas to determine whether
significant effects have occurred in the impacted area. Unfortunately, the
low biomass, high diversity, high patchiness, and extreme difficulty of doing
routine sampling in the deep sea make this option unfeasible. The second
85
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approach to monitoring is to determine the potential mechanisms for impact on
individual organisms and try to relate the magnitude of an observed response
to potential detrimental effects. To develop such an environmental dosimeter
for marine organisms, an endpoint of significance must be studied in a variety
of organisms under a variety of conditions to determine its potential
sensitivity and reproducibility. Furthermore, effect levels must be related
to other biological endpoints to determine the risk associated with any given
exposure.
The prime method for biological dosimetry due to radiation exposure in
human populations is metaphase analysis of chromosomal aberrations in
lymphocytes. Use of this analysis helps reconstruct the exposure level of the
person being examined. When this is linked with other knowledge about the
biological effects of such an exposure, biological risk estimates for this
person can be made.
To translate this method of environmental dosimetry to marine organisms,
considerable research on the basic cell biology of the organisms and tissue in
question is required. An understanding of the cell kinetics of a given system
is essential because a certain percentage of aberrations are lost at each cell
division and because radiation induces mitotic delay that affects the time at
which damage is seen. Other complicating factors such as interphase death of
damaged cells may also occur.
A second, very recent approach to biological dosimetry is the use of the
production from radiation of electron paramagnetic resonance (EPR) signals in
calcified tissue (McCreery et al.., 1984). The nature of the radiation-induced
EPR signal in calcified tissues,makes it amenable for use in biologic
dosimetry. This signal is associated with lattice defects or electron traps
within calcified material and has been examined in human bone and teeth that
have been irradiated in vitro. The signal is extremely stable and, at normal
temperatures, the time constant is approximately a million years. The response
to radiation appears to be dose related; the sensitivity is down to a total
dose of 10 to 20 rads; and data can be obtained with samples as small as 10 to
20 mg. This technique has been proposed for evaluating radiation exposure to
military personnel. However,, it should be equally applicable for detecting
radiation exposure of bone or teeth from deep-sea organisms or calcified
exoskeletons from deep-sea invertebrates. Of special interest would be the
response in brittle-star exoskeletons because brittle stars live in and on the
sediment layer, which would optimize their exposure to radiation.
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RECOMMENDATIONS FOR FUTURE RESEARCH ON BIOLOGICAL EFFECTS AND
BIOMONITORING STRATEGIES
Even though a large body of literature exists on the effects of ionizing
radiation on aquatic organisms, only a fraction of it addresses the specific
needs of decision makers with regard to ocean dumping of radioactive waste.
The general deficiencies in the data base are (1) few reliable studies at
chronic and low dose rates have been conducted, (2) few studies have been done
on marine organisms (which means that the comparative sensitivity of these
species with regard to selected endpoints is unknown), (3) very few studies
have been done on marine invertebrates, (4) information on modifying factors
such as temperature, species specificity, and cell kinetics is very scarce,
and (5) the long-term effects of low-level radiation on fertility in fish and
invertebrates have not been adequately characterized.
To characterize low- or threshold-effect levels in a time-effective and
cost-effective manner, the most relevant biological endpoints, species, and
modifying factors should be identified. Our recommendations are as follows:
1.
2.
3.
In view of evidence for radiation-induced sterility at long-term low
doses in both fish and mammals, further studies should be conducted
to determine gamete sensitivity and frequency of dominant lethal
mutations in a variety of species of marine fishes and invertebrates.
Information is needed on the effects of chronic irradiation on
marine invertebrates. This information should relate to parallel or
previous acute studies so that acute:chronic ratios can be
determined.
Results from research indicate that analysis of chromosomal
aberrations in fish and invertebrates may be a more sensitive
endpoint than gamete death. For this reason, further studies should
be conducted to characterize chromosomal aberration induction with
regard to the sensitivity of various species and the cell kinetics
of specific tissues. Use of the micronucleus assay should also be
considered.
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4. Because temperature can modify biological responses to radiation by
more than an order of magnitude (causing fractionated or chronic
doses to be more clearly additive) and because nothing has been done
on this effect in cold-adapted marine organisms, research should be
done to examine the role of low temperature on radiation damage at
low-level, chronic doses. Species that normally live in cold
environments should be selected, and a variety of endpoints,
including oocyte sensitivity, observed at different chronic and
acute doses to examine the possibility of a decrease in the
dose-rate effect.
If biological monitoring of ocean disposal sites becomes a high priority,
a research effort should be considered to develop and characterize a method of
biological dosimetry beyond the determination of body burdens. Because
studying parameters of community structure or population reproduction are
currently unfeasible in the deep ocean, we have sought other options and
identified two potential approaches:
First, examination of chromosomal aberrations in selected organisms has
potential for environmental dosimetry. However, development of this technique
with regard to ultimate sensitivity, reproducibility, and species selection is
a long-term project. In some cases, such an effort may be combined with the
reproductive studies recommended above to provide important information on the
relative sensitivities of these endpoints.
The second option identified is the application of electron paramagnetic
resonance (EPR) spectroscopy to evaluate radiation effects in calcified
tissues. The nature of radiation-induced EPR signal in calcified tissues makes
it amenable for use as a biologic dosimeter. It is proposed that calcified
exoskeletons of invertebrates or bones of fishes be used as the test material.
88
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ACKNOWLEDGMENTS
The authors wish to acknowledge M. Varela and R. Dyer at the U.S.
Environmental Protection Agency, Office of Radiation Programs for their
assistance in the development of this manuscript. Others who provided
scientific advice are also gratefully acknowledged, they are: L. Anspaugh,
B. Backus, A. V. Carrano, N. Nelson, and T. Straume. In addition, B. Strack
and G. Reed provided editorial assistance, and A. Fountain and J. Johansen
produced the manuscript.
89
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Van Rijn, J., 0. Van Den Berg, J.B.A. Kipp, D.H.J. Schamhart, and R. Van Wijk
(1985), "Effect of Hypothermia on Cell Kinetics and Response to
Hyperthermia and X Rays," Radiat. Res. 101, 292.
Wadley, G.W. and A. D. Welander (1971), "X-Rays and Temperature: Combined
Effects on Mortality and Growth of Salmon Embryos," Trans. Am. Fish. Soc.
100, 267.
Walden, S.O. (1973), "Effects of Tritiated Water on the Embryonic Development
of the Three- Spine Stickleback, Gasterosteus aculeatus Linnaeus," in Proc.
3rd Natl. Symp. Radionuclides in Ecosystems, CONF-710501, D.J. Nelson, Ed.
(U.S. Atomic Energy Commission, Washington, DC), pp. 1087-1090.
Walton, D.G., A.B. Acton, and H.F. Stich (1983), "DNA Repair Synthesis in
Cultured Mammalian and Fish Cells Following Exposure to Chemical Mutagens,"
Mutat. Res. 124, 153.
116
-------
Ward, E., S.A. Beach, and E.D. Dyson (1971), "The Effect of Acute
X-Irradiation on the Development of the Plaice, PIeuronectes platessa L.,"
J. Fish Biol. 3, 251 .
Watson, D.6. and W.L. Templeton (1973), "Thermo!umlnescent Dosimetry of
Aquatic Organisms," in Proc. 3rd Nat!. Symp. Radionuclides in Ecosystems,
CONF-710501, D.J. Nelson, Ed. (U.S. Atomic Energy Commission, Washington,
DC), pp. 1125-1130.
Welander, A.D. (1954), "Some Effects of X-Irradiation of Different Embryonic
Stages of the Trout (Salmo gairdnerii)." Growth 18. 227.
Welander, A.D., L.R. Donaldson, R.F. Foster, K. Bonham, and A.H. Seymour
(1948), "The Effects of Roentgen Rays on the Embryos and Larvae of the
Chinook Salmon," Growth 12, 203.
Whicker, F.W. (1980), "Ecological Effects of Transuranics in the Terrestrial '.
Environment," in Transuranic Elements in the Environment, W.C. Hansen, Ed.
(Office of Scientific and Technical Information Center, U.S. Department of
Energy, Oak Ridge, TN), pp. 701-713.
Whicker, F.W., and V. Schultz (1982), Radioecology: Nuclear Energy and the
Environment; Vol. II (CRC Press, Boca Raton, FL).
White, Jr., O.C., O.W. Angelovic, D.W.-Engel, and E.M. Davis (1967),
"Interactions of Radiation, Salinity, and Temperature on Estuarine
Organisms," in Ann. Rep. Bur. Comm. Fish. Radiobiol. Lab., Circ. 270 (U.S.
Fish Wildl. Sen/., Beaufort, NC), pp. 29-35.
Willis, D.L. (1980), "The Effect of Temperature on the Radiation Response of
the Rough-Skinned Newts, Taricha granulosa," in Radiation Effects on
Aquatic Organisms. N. Egami, Ed. (Gapan Scientific Societies Press, Tokyo;
University Park Press, Baltimore, MD), pp. 157-167.
117
-------
Willis, D.L. and W.L. Lappenbusch (1976), "The Radiosensitivity of the •--/•
Rough-Skinned Newt (Taricha granulosa)," in Proc. 4th Natl. Symp.
Radioecology, C.E. Cushing, Jr., Ed. (Dowden, Hutchinson, & Ross,
Stroudsburg, PA), pp. 363-375.
Wolf, K. and G.A. Mann (1980), "Poikilotherm Vertebrate Cell Lines and
Viruses: A Current Listing for Fishes," In Vitro 16, 16B.
Wolff, S. (1968), "Chromosome Aberrations and the Cell Cycle," Radiat. Res.
33_, 609.
Woodhead, A.D., P. Achey, R.B. Setlow, and E. Grist (1978), "Photoenzymatic
Repair of Ultraviolet-Irradiated DMA in the Cells of a Shark, Prionace
glauca," Comp. Biochem. Physio!. 60B, 205.
Woodhead, A.D. and P.M. Achey (1979), "Photoreactivating Enzyme in the Blind
Cave Fish, Anoptichthys jordani," Comp. Biochem. Physio!. 63B» 73.
Woodhead, A.D., R.B. Setlow, and R.W. Hart (1979), "Genetically Uniform
Strains of Fish as Laboratory Models for Experimental Studies of the
Effects of Ionizing Radiation," in Methodology for Assessing Impacts of
Radioactivity on Aquatic Ecosystems, Technical Report Series 190
(International Atomic Energy Agency, Vienna), pp. 317-333.
Woodhead, D.S. (1970), "The Assessment of the Radiation Dose to Developing
Fish Embryos Due to the Accumulation of Radioactivity by the Egg," Radiat.
Res. 43, 582.
Woodhead, D.S. (1973a), "Levels of Radioactivity in the Marine Environment
and the Dose Commitment to Marine Organisms," in Proc. Symp. Radioactive
Contamination of the Marine Environment (International Atomic Energy
Agency, Vienna), pp. 499-525.
Woodhead, D.S. (1973b), "The Radiation Dose Received by Plaice (Pleuronectes
platessa) from the Waste Discharged into the North-East Irish Sea from the
Fuel Reprocessing Plant at Windscale," Health Phys. 25, 115.
118
-------
Woodhead, D.S. (1976), "Influence of Acute Irradiation on Induction of
Chromosome Aberrations in Cultured Cells of the Fish Ameca splendens," in
Proc. Symp. Biol. Environ. Eff. of Low-Level Radiat., Vol. I (International,
Atomic Energy Agency, Vienna), pp. 67-76.
Woodhead, D.S. (1977), "The Effects of Chronic Irradiation on the Breeding
Performance of the Guppy, Poecilia reticulata (Osteichthyes: Teleostei),"
Int. J. Radiat. Biol. 32, 1.
Woodhead, D.S. (1979), "Methods of Dosimetry for Aquatic Organisms," in
Methodology for Assessing Impacts of Radioactivity on Aquatic Ecosystems,
Technical Report Series 190 (International Atomic Energy Agency, Vienna),
pp. 43-96.
Woodhead, D.S. and R.J. Pentreath (1983), "A Provisional Assessment of
Radiation Regimes in Deep Ocean Environments," in Wastes in the Oceans,
Vol. 3 (John Wiley, New York, NY), pp. 133-152.
Woodhead. D.S., C.J. Barker, and B.D. Rackham (1983), "The Effects of Chronic
Y-Irradiation on Experimental Fish Populations: A Preliminary Account",
Abstract in Biological Effects of Low-level Radiation (International Atomic
Energy Agency, Vienna), pp. 646-648.
Woodhead, D.W. (1984), "Contamination Due to Radioactive Materials," in
Marine Ecology, Vol. V, Part 3, 0. Kinne, Ed. (John Wiley and Sons, Ltd.,
Chichester, UK), 1618 pp.
119
-------
APPENDIX I
List of Major Review Articles
Blaylock, B.6. and J.R. Trabalka (1978), "Evaluating the Effects of Ionizing
Radiation on Aquatic Organisms," in Advances in Radiation Biology, Vol. 7,
J.T. Lett and H. Adler, Eds. (Academic Press, New York, NY), pp. 103-152.
Chipman, W.A. (1972), "Ionizing Radiation," in Marine Ecology, Vol. I, Pt. 3,
Ch. 11, 0. Kinne, Ed. (John Wiley/Interscience, New York, NY),
pp. 1579-1657.
Egami, N. and K.I. Ijiri (1979), "Effects of Irradiation on Germ Cells and
Embryonic Development in Teleosts," Int. Rev. Cytol. 59, 195.
Ophel, I.L. (1976), "Effects of Ionizing Radiation on Aquatic Organisms," in
Effects of Ionizing Radiation on Aquatic Organisms and Ecosystems,
Technical Report Series 172 (International Atomic Energy Agency, Vienna),
pp. 57-88.
Polikarpov, G.G. (1966), Radioecology of Aquatic Organisms (Reinhold,
New York, NY).
Templeton, W.L. (1976), "Effects of Ionizing Radation on Aquatic Populations
and Ecosystems," Technical Report Series 172 (International Atomic Energy
Agency, Vienna), pp. 89-119.
Templeton, W.L., R.E. Nakatani, and E.E. Held (1971), "Radiation Effects,"' in
Radioactivity in the Marine Environment, (National Academy of Sciences,
Washington, DC), 272.
Woodhead, D.W. (1984), Contamination Due to Radioactive Materials," in
Marine Ecology, Vol. V, Part 3, 0. Kinne, Ed. (John Wiley and Sons, Ltd.,
Chichester, UK), 1618.
120
-------
APPENDIX II
Reference Summary Table
To conduct a systematic critical examination of the literature, we first
classified articles according to effect endpoints and examined modifying
factors. After reviewing the nearly 200 articles presented in this summary
table, we selected only the most pertinent references for inclusion in the
text and tables. However, all papers are given in the list of references. We
then added further articles to the text and reference list, but did not add
them to this summary table.
121
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Appendix II. Reference summary table
General sorts
Effect endpoints
Modifying factors
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Author
Cosgrove, G.E. (BGB)
Cosgrove, G.E. (+)
Crenshaw, J.W.
Croute, F. (+)
Daniel, G.E. (HDP)
Donaldson, L.R. (KB)
Donaldson, L.R. (KB)
Dunster, H.J. (+)
Egami, N. (HE)
Egami, N.
Egami, N.
Egami, N.
Egami, N. (AH)
Egami, N. (AH)
Egami, N. (AH)
Egami, N. (K-II)
Egami, N. (AH-F)
Egami, N. (AH-F)
Egami, N. (+)
Egami, N. (Y H-T)
Egami, N.. (YH-T)
Emergy, R.M, (MCM)
Emery, R.M. (DCK)
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Appendix II. Reference summary table
General sorts
Effect endpoints
Modifying factors
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Author
Etoh, H. (NE)
Etoh, H. (+)
Etoh, H. (+)
Etoh, H. (IS)
Folsom, T.R. (TMB)
Frank, M.L.
Fujita, S. (NE)
Gehrs, C.W. (+)
Gesell, T.F. (+)
Ghoneum, M.M.H. (+)
Ghoneum, M.M.H. (NE)
Gorbman, A. (MSJ)
Green, E.L.
Grosch, D.S.
Grosch, D.S.
Guthrie, J.E. (RAB)
Guthrie, J.G. (RAB)
Hama, A. (NE)
Hama-Furukawa, A. (NE)
Hamaguchi, S.
Hansen, H.J.M.
Hansen, H.J.M.
Held, E.E.
Hetherington, J.A. (+)
Holton, R.L. (+)
Holzberg, S. (JHS)
Holzberg, S.
Hoppenheit, M.
Horsley, R.J. (AL)
Hyodo, Y.
Hyodo, Y.
Hyodo, Y.
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74
80
73
73
84
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76
83
80
63
68
62
66
71
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80
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80
60
76
73
72
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73
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Appendix II. Reference suraary table
Effect endpoints
Modifying factors
Author
Mc6regor, J.F. (HBN)
McGregor, J.F. (HBN)
McGregor, J.F. (HBN)
Metal! i, P.
Michibata, H. (NE)
Migalovskaya, V.N.
Neuhold, J.M. (RKS)
Newcombe, H.B. (JFM)
Newcombe, H.B.
Newcombe, H.B. (JFM)
Oganesyan, S.A.
Ophel, I.L.
Patel, B.
Patel, B. (S.P.)
Polikarpov, 6.6.
Preston, A.
Preston, A.
Preston, A. (OFJ)
Purdom, C.E.
Purdom, C.E. (DSW)
Ravera, 0.
Ravera, 0.
Rice, T.R. (JRB)
Rugh, R. (HC)
Scarborough, B.B. (R6A)
Schroeder, J.H.
Schroeder, J.H.
Schroeder, J.H.
Schroeder, J.H. (SH)
Schroeder, J.H.
Schroeder, J.H.
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72
72
79
77
73
67
67
71
72
73
79
75
79
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69
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Appendix II. Reference summary table
General sorts
Author
Setlow, R.B.
Shechmeister, I.L. (+)
Slobodkin, L.B. (SR)
Squire, R.D.
Squire, R.D.
Strand, J.A. (+)
Strand, J.A. (+)
Styron, C.E.
Suyama, I. (+)
Suzuki, J. (NE)
Templeton, W.L.
Templeton, W.L. (+)
Templeton, W.L.
Templeton, W.L.
Till, J.E. (+)
Trabalka, J.R. (CPA)
Tsytsugina, V.G.
Tsytsugina, V.G.
Ulrickson, -G.U.
Upton, A.C. (+)
Wadley, G.W. (ADW)
Wai den, S.J.
Ward, E. (.+)
Watson, D.G. (WLT)
Welander, A.D. (+)
Wei ander, A.D.
Whicker, F.W.
White, J.C. (+)
Willis, D.L.
Willis, D.L. (WLL)
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Appendix II. Reference summary table
Effect endpoints
Modifying factors
Author
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