As part of a continuing survey, effluents from five mills in Canada were tested in the laboratory for their potential to affect fish reproduction. The study included effluents from two thermomechanical pulp (TMP) mills, two kraft pulp mills, and one mill that used both chemical and mechanical pulping. The laboratory test used adult fathead minnows and involved a 21-day exposure to each effluent. All the effluents were tested at 2 and 20% concentration. The effluent from 1 of the kraft mills was also tested at 40% concentration. The endpoints of the test included, egg production, gonad size, sex steroids, secondary sexual characteristics, and vitellogenin concentration in males, considered to be an indicator of estrogenicity. The results of this study were similar to the results of our previous survey. None of the effluents produced noteworthy changes at 2% concentration. At 20% concentration, only the effluent from the multiprocess mill produced a significant reduction in eggs, which was considered to be the most important indicator of reproductive performance. Some effluents did produce an increase and/or a decrease in a variety of endpoints other than egg production, but the most consistent response was an induction of vitellogenin in males exposed to three of the five effluents tested. In summary, these results indicate that most mill effluents up to 20 or 40% concentration do not affect the overall reproductive capacity of minnows in the laboratory. However, the mill effluents do seem to contain substances that cause vitellogenin induction.

The regulatory Environmental Effects Monitoring (EEM) program in Canada has found that pulp and paper mill effluents reduce the gonad size of wild fish. In such cases, there is a need for Investigation of Cause (IOC) and Investigation of Solution (IOS) studies to find the cause(s) for these responses as well as cost-effective mitigation strategies. IOC and IOS work requires the availability of practical (in terms of time and effluent volume needs) and biologically meaningful tests for assessing fish reproduction in the laboratory. One available fish reproduction test uses adult fathead minnow (Pimephales promelas) and involves a two- to three-week pre-exposure period and a three-week exposure to a test substance. Work was undertaken to determine if both the pre-exposure and effluent-exposure phases could be reduced, leading to a shorter test with a lower effluent volume requirement. For this purpose, effluent samples were taken from two thermomechanical (newsprint) pulp mills and used in tests with ≤7 days of pre-exposure and 5 to 6 days of effluent exposure. Egg production was monitored in each test, and in tests with effluents from one mill, other endpoints - such as gonad somatic indices and whole-body sex steroid levels, as well as vitellogenin activity in males - were measured. The shortened fish reproduction test in this study provided results similar to tests of longer duration, specifically in terms of egg production, and was able to distinguish between pulp and paper mill effluents of different potencies. The test was also found to be directly applicable for toxicity identification evaluation work as it was clearly able to distinguish between the solids and soluble fractions of a biotreated newsprint mill effluent. As such, the test is a promising tool for EEM-related IOC and IOS work.