Biological test method for toxicity tests using early life stages of rainbow trout: chapter 8

Section 8: Reporting Requirements

• 8.1 Minimum Requirements for a Test-specific Report
  • 8.1.1 Test Substance
  • 8.1.2 Test Organism
  • 8.1.3 Test Facilities and Apparatus
  • 8.1.4 Control/Dilution Water
  • 8.1.5 Test Method
  • 8.1.6 Test Conditions and Procedures
  • 8.1.7 Test Results
• 8.2 Additional Reporting Requirements
  • 8.2.1 Test Substance
  • 8.2.2 Test Organism
  • 8.2.3 Test Facilities and Apparatus
  • 8.2.4 Control/Dilution Water
  • 8.2.5 Test Method
  • 8.2.6 Test Conditions and Procedures
  • 8.2.7 Test Results

Each test-specific report must indicate if there has been any deviation from any of the "must" requirements delineated in Sections 2 to 7 of this biological test method, and, if so, provide details as to the deviation. The reader must be able to establish from the test-specific report whether the conditions and procedures preceding and during the test rendered the results valid and acceptable for the use intended.

Section 8.1 provides a list of the items that must be included in each test-specific report. Section 8.2 gives a list of items that must either be included in the test-specific report, provided separately in a general report, or held on file for a minimum of five years. Specific monitoring programs or related test protocols might require selected test-specific items listed in Section 8.2 to be included in the test-specific report, or might relegate certain test-specific information as "data to be held on file" (e.g., details about the test substance and/or explicit procedures and conditions during sample collection, handling, transport, and storage).

Procedures and conditions that are common to a series of ongoing tests (e.g., routine toxicity tests for monitoring or compliance purposes) and consistent with specifications in this document, may be referred to by citation or by attachment of a general report which outlines standard laboratory practice.

Details on the conduct and findings of the test, which are not conveyed by the test-specific report or general report, must be kept on file by the laboratory for a minimum of five years, so that the appropriate information can be provided if an audit of the test is required. Filed information might include:

• a record of the chain-of-continuity for samples tested for regulatory or monitoring purposes;
• a copy of the record of acquisition for the sample(s);
• certain chemical analytical data on the sample(s);
• bench sheets for the observations and measurements recorded during the test;
• bench sheets and warning chart(s) for the reference toxicity tests;
• detailed records of the source and health of the broodstock;
• all pertinent information on the procedures for stripping, handling, packaging, and storage of gametes and on the subsequent fertilization process; and
• information on the calibration of equipment and instruments.

Original data sheets should be signed or initialled and dated by the laboratory personnel conducting the tests.

8.1 Minimum Requirements for a Test-specific Report

Following is a list of items that must be included in each test-specific report.

8.1.1 Test Substance

• brief description of sample type (e.g., chemical or chemical product, effluent, elutriate, leachate, or receiving water), if and as provided to the laboratory personnel;
• information on labelling or coding, for each sample or subsample;
• date of sample/subsample collection; date and time sample(s)/subsample(s) received at test facility;
• dates or days during test when individual samples or subsamples used;
• for effluent or leachate, measurement of temperature of sample or, if multiple subsamples, one only of these subsamples, upon receipt at test facility;
• measurements of dissolved oxygen and pH of sample or subsample of wastewater or receiving water, just before its preparation and use in toxicity test; and
• date of elutriate generation and description of procedure for preparation; dates or days during an elutriate test when individual samples or subsamples are used.

8.1.2 Test Organism

• species and common name;
• source of gametes or broodstock; number of female and male broodstock used for fertilization;
• brief description of procedure for checking sperm motility;
• brief description (including time interval) of procedure for fertilization of gametes;
• time interval from completion of fertilization until exposure of all groups of eggs to test solutions; and
• any unusual appearance or treatment of gametes or eggs, before their use in the test.

8.1.3 Test Facilities and Apparatus

• name and address of test laboratory;
• name of person(s) performing the test; and
• brief description of test chamber(s) and associated apparatus (e.g., incubation units; pumps or other apparatus if flow-through test).

8.1.4 Control/Dilution Water

• type(s) and source(s) of water used as control and dilution water; and
• type and quantity of any chemical(s) added to control or dilution water.

8.1.5 Test Method

• citation of biological test method used (i.e., as per this document);
• brief mention and description of test options chosen (e.g., E, EA, or EAF test; static-renewal or flow-through test);
• design and description if specialized procedure (e.g., renewal of test solutions at intervals other than daily; preparation and use of elutriate; preparation and use of solvent and, if so, solvent control);
• brief description of procedure(s) in those instances in which a sample, subsample, or test solution has been filtered or adjusted for hardness or pH;
• brief description of frequency and type of all observations and all measurements made during test; and
• name and citation of program(s) and methods used for calculating statistical endpoints.

8.1.6 Test Conditions and Procedures

• design and description if any deviation from or exclusion of any of the procedures and conditions specified in this document;
• number and concentration of test solutions, including controls; volume and depth of solution in each test chamber;
• number of individuals per test chamber, and number of replicates per concentration;
• brief statement concerning presence or absence of pre-aeration or aeration; if any, give rate and duration used to aerate sample or test solutions before and during exposure of test organisms;
• manner and rate of exchange (i.e., L/g·d) of test solutions;
• dates when test was started and ended; and
• all required measurements of temperature, pH, and DO (mg/L and percent saturation) in sample and test solutions including controls, before and during the test.

8.1.7 Test Results

• average number and percentage of nonviable embryos in each replicate and concentration, including the controls (E test), seven days after fertilization; EC50 and 95% confidence limits, and EC25;
• average number and percentage of nonviable alevins in each replicate and concentration, including the controls, seven days after 50% hatch in the controls (EA test); EC50 and confidence limits, and EC25;
• average number and percentage of nonviable individuals at time of 50% control swim-up, in each replicate and concentration, including the controls (EAF test); EC50 and confidence limits, and EC25;
• number of dead fry in each concentration after 30 days of exposure with feeding, and number that started the exposure (EAF test); LC50 and confidence limits;
• average dry weight of fry surviving the 30-d exposure with feeding in each replicate and concentration (EAF test); IC25 and 95% confidence limits;
• narrative statement(s) on delayed hatching and deformed alevins in each concentration (EA test); description of any apparent differences from control;
• narrative statement(s) on deformed alevins, delayed swim-up, and abnormal behaviour of fry in each concentration (EAF test); description of any apparent differences from control;
• the results of E tests with the reference toxicant(s), conducted concurrently, together with the geometric mean value (± 2 SD) for the same reference toxicant(s) as derived at the test facility in previous tests; and
• anything unusual about the test, any deviation from these procedures, any problems encountered, any remedial measures taken.

8.2 Additional Reporting Requirements

Following is a list of items that must be either included in the test-specific report or the general report, or held on file for a minimum of five years.

8.2.1 Test Substance

• identification of person(s) who collected and/or provided the sample or subsamples;
• records of sample/subsample chain-of-continuity and log-entry sheets; and
• conditions (e.g., temperature, in darkness, in sealed container) of samples/subsamples upon receipt and during storage.

8.2.2 Test Organism

• history of broodstock (e.g., fecundity; records of any disease and treatments);
• procedures and observations during gamete collections (e.g., degree of ripeness of females; motility of sperm); and
• conditions and procedures during transport and storage of gametes, motility of sperm preceding fertilization, details regarding procedures for fertilization.

8.2.3 Test Facilities and Apparatus

• description of systems for regulating light and temperature within the test facility;
• description of system for providing air and regulating air flow to test chambers;
• detailed description and/or illustration of test chambers and associated apparatus (e.g., size, shape, type of material, design); and
• description of procedures used to clean or rinse test apparatus.

8.2.4 Control/Dilution Water

• sampling and storage details if the control/dilution water was "upstream" receiving water;
• details on any pre-treatment of water (e.g., filtration, sterilization, chlorination/dechlorination, temperature adjustment, de-gassing, aeration); and
• ancillary water-quality variables (e.g., dissolved metals, ammonia, pesticides, suspended solids, humic and fulvic acids) measured before and/or during the toxicity test.

8.2.5 Test Method

• description of laboratory's previous experience with the test option chosen and associated procedures herein;
• procedure used in preparing and storing stock and/or test solutions of chemicals; description and concentration(s) of any solvent used;
• methods used for chemical analyses of sample or test solutions, with citations; details concerning sampling, sample preparation and storage, before chemical analyses; and
• use and description of preliminary or range-finding test.

8.2.6 Test Conditions and Procedures

• photoperiod, light source, and intensity at surface of test solutions;
• description of any thinning of swim-up fry to start the second phase of an EAF test (manner, numbers, timing);
• description of food source, type, and ration (quantity and frequency of feeding) used to feed swim-up fry during the EAF test;
• any other chemical measurements on sample, stock solutions, or test solutions (e.g., chemical concentration, suspended solids content, conductivity, hardness, and alkalinity), before and/or during the test;
• appearance of sample or test solutions; changes in appearance noted during test; and
• conditions and procedures for measuring the EC50 for reference toxicant(s) used in the E test.

8.2.7 Test Results

• additional observations and numerical data supporting narrative statements of effect during an EA or EAF test for which there are no formal observations (see Sections 4.4 and 8.1.7), including information pertaining to: proportions of nonviable embryos (EA and EAF tests); delayed hatching (EA and EAF tests); mortality of alevins after hatching (EA and EAF tests); deformed alevins (EA and EAF tests); delayed swim-up (EAF test); and abnormal behaviour of fry (EAF test);
• results for range-finding test (if conducted);
• warning chart showing the most recent and historic results for toxicity tests with the reference toxicant(s);
• graphical presentation of data; and
• results of any acute lethality tests conducted concurrently using rainbow trout fry or fingerlings and a portion of the sample or test solutions.