The draft Metal Mining Effluent Regulation (MMER) requires that all Canadian metal mines produce effluent that is non-acutely lethal to rainbow trout when tested in accordance with Environment Canada test methods. Mine operations will also be required to monitor the acute lethality of effluent to Daphnia magna. If a rainbow trout test produces mortality of more than 50% of the test organisms in 100% effluent, the sample is considered to “fail” the acute lethality test. In the event of a toxicity failure, the draft MMER requires that the mine implement a plan to investigate the cause of acute lethality. The reliability of data generated by these tests is, therefore, an important issue in the context of maintaining confidence in the use of these tests as a basis for assessing regulatory compliance.

The Toxicological Investigations of Mining Effluents (TIME) Network was established with representation from governments, industry, environmental non-governmental organizations, the consulting community, and academia, to address toxicological issues related to the amended Metal Mining Effluent Regulation (MMER). During the first TIME workshop, held in November 1999, several potential projects were prioritized, including the development of a guidance document for acute lethality testing of mine effluents. Concerns have been expressed by industry in the past regarding the variability and repeatability of effluent toxicity test results. Therefore, this guidance document has been prepared for industry personnel, aquatic toxicity testing laboratories, and regulatory authorities to aid in the understanding of key aspects of acute lethality testing and to provide guidance aimed at maximizing data reliability.

An overview of the current state of knowledge pertaining to this topic is provided in the document, including: a historical background of aquatic toxicology in Canada, the current regulatory framework in which toxicity testing is conducted, common metal mining contaminants and their potential impact on effluent toxicity, a literature review of test method variability, and a summary of test system deviations observed in a review of metal mining effluent toxicity data.

A literature review based primarily on toxicity test methods used in the United States (i.e., U.S. EPA methods) provided some insight into the potential sources of variability associated with biological test methods in general. Analyst proficiency and judgment, as well as test organism condition and health were considered to be the largest sources of variability. Additionally, a strong QA/QC program was considered essential in helping to control test method deviations, which can lead to test variability.

Variability associated with test results specifically conducted using the Environment Canada test methods was evaluated using data sets obtained from the Canadian Association for Environmental Analytical Laboratories (CAEAL) (proficiency testing (PT) program) and from nine volunteer laboratories that provided reference toxicant test results to the Ontario Ministry of the Environment (OMOE). Coefficients of variation (CVs) were estimated using variance components analysis for all intra-laboratory (within lab) reference toxicant data. The within-laboratory CVs for rainbow trout reference toxicity tests were as follows: 13.3% using phenol as a reference toxicant, and 38.5% using dissolved zinc as a reference toxicant. The within-laboratory CVs for D. magna reference toxicity tests were: 8.7% using sodium chloride as a reference toxicant, and 33.3% using dissolved zinc as reference toxicant. Inter-laboratory (among-lab) CVs were estimated from the CAEAL PT data set also using variance components analyses. These CVs were estimated using the date and results from the testing of the CAEAL PT sample. This analysis yielded 52 CVs for the rainbow trout PT data set. CVs ranged from 8.0 to 60.4% with a median CV = 15.7%. Similarly, 28 CVs were estimated from the D. magna CAEAL PT data set. The CVs ranged from 7.5 to 53.1% with a median CV = 12.9%. Overall, the magnitude of variability observed in these biological test methods is within the range of (and in some cases, lower than) the variability observed in analytical chemistry methods.

The main portion of the document provides guidance on aspects of the Environment Canada General and Reference Methods relating to: sample collection and handling (including collection of split-samples), test organism culture and holding, test method requirements, statistical analyses, and reporting requirements, all for the purpose of maximizing data reliability. All parties involved with the testing program have critical roles to play, whether collecting the sample, performing the tests, or reviewing the test for compliance with the MMER.

Information on laboratory accreditation programs in Canada, laboratory assessments, and their importance in reducing test method variability is also provided for background in the understanding of toxicity laboratory quality assurance. In addition, guidance is provided to mine personnel for the selection of a competent ecotoxicity laboratory, as well as the implementation of test report evaluations of acute lethality data, and second-party laboratory assessments.

The guidance document improves upon, and provides greater detail on the specific guidance already provided in the rainbow trout and D. magna Reference Method documents (Environment Canada, 2000a,b). It will assist mine personnel in the collection and submission of samples and the evaluation of the resulting toxicity test reports, and it will enhance the efforts of laboratories to produce highly reliable data. Furthermore, the guidance document will also be of assistance to a broad range of stakeholders with an interest in acute lethality testing. This document does not supersede current government guidance, policy, or regulation including Environment Canada’s Reference Methods EPS 1/RM/13 and EPS 1/RM/14 (Environment Canada 2000a,b).