1.1''This standard guide describes methods for conducting laboratory toxicity tests with early life stages of freshwater mussels including glochidia and juvenile mussels in water-only and effluent exposures (Annex A1). Future revisions to this standard may describe methods for conducting toxicity tests with endpoints of reproduction, behaviors, and biomarkers.

1.2''Freshwater mussels (order Unionida) are one of the most imperiled groups of animals in the world, and environmental contamination has been linked as a contributing factor to the decline of mussel populations (Lydeard et al. 2004 (1); Strayer et al. 2004 (2); Haag 2012 (3); Lopes-Lima et al. 2017 (4)).2 Three critical life stages (glochidia, juvenile mussels, and adults) have been used in toxicity assessments and the toxicity studies are separated according to the medium of exposure (water, sediment, and host fish (Ingersoll et al. 2007 (5)). Recent studies on early life stages of mussels have demonstrated that the mussels are among the most sensitive freshwater species to a variety of contaminants, including ammonia, some metals (for example, aluminum, copper, nickel, and zinc), and major ions (for example, chloride, nitrate, potassium, and sulfate) (Bringolf et al. 2007 (6); Newton et al. 2007 (7); Wang et al. 2007ab, 2010, 2011ab, 2016, 2017ab, 2018abc, 2020ab (8-20); Cope et al. 2008 (21); Gillis et al. 2008, 2010, 2011, 2021 (22-25); Miao et al. 2010 (26); Salerno et al. 2020 (27)). These studies indicate that environmental guideline values for individual chemicals established for the protection of aquatic organisms may not be adequately protective of sensitive stages of freshwater mussels. For example, when freshwater mussel toxicity data were included in an update to the United States Environmental Protection Agency (USEPA) ambient water quality criteria (WQC) for ammonia, the acute criterion decreased by about a 1.4 fold and the chronic criterion decreased by 2.4 fold (USEPA 2013) (28).

1.3''Summary of Life History of Freshwater Mussels:''