1、Designation: E1604 94 (Reapproved 2007)E1604 12Standard Guide forBehavioral Testing in Aquatic Toxicology1This standard is issued under the fixed designation E1604; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revision, the year of last r
2、evision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () indicates an editorial change since the last revision or reapproval.1. Scope1.1 This guide covers some general information on the selection and application of behavioral methods useful for determiningthe
3、sublethal effects of chemicals to fish, amphibians, and macroinvertebrates.1.2 Behavioral toxicity occurs when chemical or other stressful conditions, such as changes in water quality or temperature,induce a behavioral change that exceeds the normal range of variability (1).2 Behavior includes all o
4、bservable, recordable, ormeasurable activities of a living organism and reflects genetic, neurobiological, physiological, and environmental determinants (2).1.3 Behavioral methods can be used in biomonitoring, the determination of no-observed-effect and lowest-observed-effectconcentrations, and the
5、prediction of hazardous chemical impacts on natural populations (3).1.4 Behavioral methods can be applied to fish, amphibians, and macroinvertebrates in standard laboratory toxicity tests, testsof effluents, and sediment toxicity tests.1.5 The various behavioral methods included in this guide are ca
6、tegorized with respect to seven interdependent, functionalresponses that fish, amphibians, and macroinvertebrates must perform in order to survive. These functional responses includerespiration, locomotion, habitat selection, feeding, predator avoidance, competition, and reproduction (4). These resp
7、onses can bedocumented visually or through video or acoustic imagery. Electronically recorded information can be derived through manualtechniques or through the use of digital image analysis software. (5, 6, 7)1.5.1 The functional responses are not necessarily mutually exclusive categories. For inst
8、ance, locomotion, of some form ofmovement, is important to all behavioral functions.1.6 Additional behavioral methods for any category may be added when new tests are developed as well as when methods areadapted to different species or different life stages of an organism.1.7 This guide is arranged
9、as follows:Section NumberScope 1Referenced Documents 2Terminology 3Summary of Guide 4Significance and Use 5Interferences 6Test Facility 7Water Supply 8Safety Precautions 9Test Material 10Test Organisms 11Responses Measured 12Behavioral Test Method Selection Criteria 13Experimental Design 14Acceptabi
10、lity of Test 15Calculation of Test Results 16Report 171.8 The values stated in SI units are to be regarded as the standard.1 This guide is under the jurisdiction of ASTM Committee E47 on Biological Effects and Environmental Fateand is the direct responsibility of Subcommittee E47.01 onAquatic Assess
11、ment and Toxicology.Current edition approved Oct. 1, 2007Dec. 1, 2012. Published October 2007February 2013. Originally approved in 1994. Last previous edition approved in 20022007 asE1604-94(2002).E1604-04(2007). DOI: 10.1520/E1604-94R07.2 The boldface numbers in parentheses refer to the list of ref
12、erences at the end of this standard.This document is not an ASTM standard and is intended only to provide the user of an ASTM standard an indication of what changes have been made to the previous version. Becauseit may not be technically possible to adequately depict all changes accurately, ASTM rec
13、ommends that users consult prior editions as appropriate. In all cases only the current versionof the standard as published by ASTM is to be considered the official document.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States11.9 This sta
14、ndard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibilityof the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatorylimitations prior to use. Specific precautionary state
15、ments are given in Section 9.2. Referenced Documents2.1 ASTM Standards:3E729 Guide for Conducting Acute Toxicity Tests on Test Materials with Fishes, Macroinvertebrates, and AmphibiansE1241 Guide for Conducting Early Life-Stage Toxicity Tests with FishesE1383 Guide for Conducting Sediment Toxicity T
16、ests with Freshwater Invertebrates (Withdrawn 1995)43. Terminology3.1 DefinitionsThe words “must,” “should, “may,” “can,” and “might” have very specific meanings. “Must” is used to expressan absolute requirement, that is, to state that the test ought to be designed to satisfy the specified condition
17、, unless the purpose ofthe test requires a different design. “Must” is used only in connection with the factors that directly relate to the acceptability ofthe test. “Should” is used to state that the specified condition is recommended and ought to be met if possible. Although theviolation of one “s
18、hould” is rarely a serious matter, violation of several will often render the results questionable. Terms such as“is desirable,” “is often desirable,” and “might be desirable” are used in connection with less important factors. “May” is used tomean “is (are) allowed to,” “can” is used to mean “is (a
19、re) able to,” and “might” is used to mean “could possibly.” Thus the classicdistinction between “may” and “can” is preserved, and “might” is never used as a synonym for either “may” or “can.”3.1 DefinitionsThe words “must,” “should, “may,” “can,” and “might” have very specific meanings. “Must” is us
20、ed to expressan absolute requirement, that is, to state that the test ought to be designed to satisfy the specified condition, unless the purpose ofthe test requires a different design. “Must” is used only in connection with the factors that directly relate to the acceptability ofthe test. “Should”
21、is used to state that the specified condition is recommended and ought to be met if possible. Although theviolation of one “should” is rarely a serious matter, violation of several will often render the results questionable. Terms such as“is desirable,” “is often desirable,” and “might be desirable”
22、 are used in connection with less important factors. “May” is used tomean “is (are) allowed to,” “can” is used to mean “is (are) able to,” and “might” is used to mean “could possibly.” Thus the classicdistinction between “may” and “can” is preserved, and “might” is never used as a synonym for either
23、 “may” or “can.”3.2 Definitions of Terms Specific to This Standard:3.2.1 behaviorthe complex of observable, recordable, or measurable activities of a living organism.3.2.2 behavioral toxicitythe phenomenon observed when a behavioral response varies beyond the range of normal as a resultof exposure t
24、o chemical or other stressors.4. Summary of Guide4.1 The potential toxicity of chemical substances in water, food, or sediments is assessed by measuring the behavior of fish,amphibians, and macroinvertebrates during exposure, using static, flow-through, or food exposure systems. The behavioralrespon
25、se of organisms exposed to chemical substances in water, effluents, food, or sediments is compared with the behavioralresponses of control organisms. The behavioral responses measured during toxicity tests are highly sensitive to sublethal exposure.The behavioral measures are relevant to essential l
26、ife functions that fish, amphibians, and macroinvertebrates often must performin order to survive and include respiration, locomotion, habitat selection, feeding, predator avoidance, competition, andreproduction. Data are obtained to determine the effects of toxic substances on behavior from short (
27、for example, 1 h) or long-term(partial to full life cycle) exposures.5. Significance and Use5.1 Protection of a species requires the prevention of detrimental effects of chemicals on the survival, growth, reproduction,health, and uses of individuals of that species. Behavioral toxicity provides info
28、rmation concerning sublethal effects of chemicalsand signals the presence of toxic test substances.5.1.1 The behavioral responses of all organisms are adaptive and essential to survival. Major changes in the behavioralresponses of fish, amphibians, and macroinvertebrates may result in a diminished a
29、bility to survive, grow, or reproduce and causesignificant changes in the natural population (58).5.2 The results from behavioral toxicity tests may be useful for measuring injury in the assessment of damages resulting fromthe release of hazardous materials (69).5.3 Behavioral toxicity test methods
30、may be useful for long-term monitoring of effluents (710).3 For referencedASTM standards, visit theASTM website, www.astm.org, or contactASTM Customer Service at serviceastm.org. For Annual Book of ASTM Standardsvolume information, refer to the standards Document Summary page on the ASTM website.4 T
31、he last approved version of this historical standard is referenced on www.astm.org.E1604 1225.4 The results from behavioral toxicity data can be used to predict the effects of exposure on fish, amphibians, and aquaticinvertebrates likely to occur in field situations as a result of exposure under sim
32、ilar conditions, including the avoidance of exposureby motile organisms (811).5.5 The results from behavioral toxicity tests might be an important consideration for assessing the hazard of materials toaquatic organisms. Such results might also be used when deriving water quality criteria for fish an
33、d aquatic invertebratesorganisms.5.6 The results from behavioral toxicity tests can be used to compare the sensitivities of different species, relative toxicity ofdifferent chemical substances on the same organism, or effect of various environmental variables on the toxicity of a chemicalsubstance.5
34、.7 The results from behavioral toxicity tests can be used to predict the effects of long-term exposure.5.8 The results of behavioral toxicity tests can be useful for guiding decisions regarding the extent of remedial action neededfor contaminated aquatic and terrestrial sites.5.9 The behavioral char
35、acteristics of a particular organism must be understood and defined before a response can be used asa measure of toxicity. The range of variability of any behavioral response of unexposed organisms is influenced by genetic,experiential, physiological, and environmental factors. Thus it is important
36、to avoid selecting test organisms from populations thatmay vary significantly in these factors.5.10 The results of behavioral toxicity tests will depend on the behavioral response measured, testing conditions, water quality,species, genetic strain, life stage, health, and general condition of test o
37、rganisms. Therefore, the behavioral response may beaffected by the test environment.6. Interferences6.1 A number of factors can suppress, elicit, or alter behavioral responses and thus influence behavioral test results andcomplicate data interpretation. The following factors should be considered in
38、the experimental protocol or in the discussion ofresults when measuring behavioral responses during toxicity tests:6.1.1 The pretest handling of test organisms resulting from collection, transfer, and maintenance of the culture environment canaffect the response observed during exposure to toxic sub
39、stances.6.1.2 The health, nutritional state, and physical condition of the organism can influence the test.6.1.3 Behavioral responsiveness may vary by species, genetic strain, population, gender, and developmental stage of theorganism.6.1.4 Prior exposure to hazardous materials, environmental stress
40、es, and pathogens can affect the behavioral responses.6.1.5 Social status, such as dominance or sex of the individuals tested, and experiential factors, such as prior experience withpredator or prey species, can influence the behavioral response. Individuals tested in isolation may respond different
41、ly than whentested in groups.6.1.6 Cyclical changes (circadian, seasonal, annual, and reproductive) in behavioral responses can occur.6.1.7 The behavioral response can be affected by apparatus design and by the procedural sequence of the measurement method.6.1.8 Behavioral responses will vary accord
42、ing to the extent to which test organisms acclimate to the physical variables of thetesting environment, including water quality, temperature, water flow, light, cover, and substrate, as well as their recovery fromhandling, acceptance of diet, and adjustment to novel testing chambers.6.1.9 Behaviora
43、l responses to toxic substances may subside over time.7. Test Facility7.1 FacilitiesThe facility should include a constant temperature area for culturing and testing. Test and culture chambers maybe placed in a temperature-controlled recirculating water bath or in a constant-temperature area. Air us
44、ed for aeration should befree of fumes, oil, and water and can require filters to remove oil, water, and bacteria. The test facility should be well-ventilatedand free of fumes. Enclosures may be necessary to ventilate test chambers.7.1.1 Culture and animal care facilities should not be in a room in
45、which toxicity tests are conducted, stock solutions or testsolutions are prepared, or equipment is cleaned.7.1.2 A timing device should be used to provide a light:darkness cycle. A15 to 30-min transition period, allowing for a gradualchange in light intensity when the lights are turned on or off, ma
46、y be desirable for reducing stress caused by instantaneousillumination or darkness.7.2 Construction MaterialsConsistent with specifications delineated, for example, in Guide E1241, equipment and facilitiesthat come into contact with stock solutions, test solutions, food, sediment, air, or water, int
47、o which the test organisms are placed,should not contain substances that can be leached or dissolved in amounts that affect the test organisms adversely. The materialsshould be chosen to minimize sorption of test materials.7.3 Water and Air Delivery SystemsThe water delivery system used in flow-thro
48、ugh testing can be one of several designs. Thesystem should be capable of delivering equal volumes of water at an equal rate of flow to each replicate treatment container.E1604 123Various metering systems, using different combinations of siphons, pumps, solenoids, valves, etc., have been used succes
49、sfully tocontrol the flow rates of water and toxic substances (see Guide E1241).7.3.1 The metering system should be calibrated before the test by determining the flow rate of water and air through each testchamber. The general operation of the metering system should be visually checked daily throughout the test. The water deliverysystem should be adjusted during the test if necessary. At any particular time during the test, flow rates through any two testchambers should not differ by more than 10 %.7.4 Test ChambersIn a behavioral toxicity test with fish, amphibians, and m
