Clean Water Act Analytical Methods

Whole Effluent Toxicity Methods

""Whole Effluent Toxicity (WET) refers to the aggregate toxic effect to aquatic organisms from all pollutants contained in a facility's wastewater (effluent). It is one way we implement the Clean Water Act's prohibition of the discharge of toxic pollutants in toxic amounts. WET tests measure wastewater's effects on specific test organisms' ability to survive, grow and reproduce.
The WET methods listed below are specified at 40 CFR 136.3, Table IA. WET test methods consist of exposing living aquatic organisms (plants, vertebrates and invertebrates) to various concentrations of a sample of wastewater, usually from a facility's effluent stream. WET tests are used by the National Pollutant Discharge Elimination System (NPDES) permitting authority to determine whether a facility's permit will need to include WET requirements.
All manuals include guidelines on laboratory safety, quality assurance, facilities and equipment, dilution water, effluent sampling methods and holding times and temperatures, data analysis, report preparation, and organism culturing and handling.

Acute Toxicity to Freshwater and Marine Organisms

This manual describes test procedures for effluents and receiving waters and includes guidelines on test species selection and mobile toxicity test laboratory design. The acute toxicity tests generally involve exposure of any of 20 test organisms to each of five effluent concentrations and a control water. The test duration ranges from 24 to 96 hours. The following methods are described in the manual (5th edition, 2002).

Acute Toxicity WET Methods

No. Method Title
Freshwater methods:
2000.0 Fathead Minnow, Pimephales promela, and Bannerfin shiner, Cyprinella leedsi
2002.0 Daphnia, Ceriodaphnia dubia
2019.0 Rainbow trout, Oncorhynchus mykiss, and Brook trout, Salvelinus fontinalis
2021.0 Daphnia puplex and Daphnia magna
Marine/Estuarine methods:
2004.0 Sheepshead minnow, Cyprinodon variegatus
2006.0 Silverside, Menidia beryllina, Menidia menidia, and Menidia peninsulae
2007.0 Mysid, Americamysis bahia

Chronic Toxicity to Freshwater Organisms

This manual (4th edition, 2002) describes four- to seven-day methods for estimating the chronic toxicity of effluents and receiving waters to three species.

Chronic Toxicity - Freshwater WET Methods

No. Method Title
1000.0 Fathead minnow, Pimephales promelas, larval survival and growth
1001.0 Fathead minnow, Pimephales promelas, larval survival and teratogenicity
1002.0 Daphnia, Ceriodaphnia dubia, survival and reproduction
1003.0 Green alga, Selenastrum capricornutum, growth

Chronic Toxicity to Marine and Estuarine Organisms

This manual (3rd edition, 2002) describes five short-term (one-hour to nine-day) methods for estimating the chronic toxicity of effluents and receiving waters to four species. These rules do not apply to discharges into marine waters of the Pacific Ocean.

Chronic Toxicity - Marine and Estuarine WET Methods

No. Method Title
1004.0 Sheepshead minnow, Cyprinodon variegatus, larval survival and growth
1005.0 Sheepshead minnow, Cyprinodon variegatus, embryo-larval survival and teratogenicity
1006.0 Inland silverside, Menidia beryllina, larval survival and growth
1007.0 Mysid, Americamysis bahia, survival, growth and fecundity
1008.0 Sea urchin, Arbacia punctulata, fertilization

Top of Page