Biomonitoring using benthic macroinvertebrates has been used to assess water quality in Europe since the early 20th century. Most methods use community-level measurements, and the use of single-species responses has been limited, despite their potential benefits as sensitive, early warning indicators. Here we evaluate a single-species in situ assay in which the response is feeding inhibition of the freshwater amphipod Gammarus pulex. The assay was deployed in uncontaminated reference sites to quantify background variability in feeding rates and to elucidate sources of this variation. The ability of the assay to detect impacts of point-source discharges was assessed and the ecological relevance of the assay determined by comparing assay responses to aspects of community structure and functioning. Water temperature accounted for 76% of the variation in the feeding rate of animals deployed at uncontaminated sites, and summer deployments had a >90% power to detect a 30% inhibition in feeding. Inhibition of the situ feeding rate of G. pulex deployed downstream of a variety of point-source discharges ranged from 27 to 99.6%. Gammarus pulex is an important detritivore in stream communities, and a strong positive correlation existed between in situ feeding rate measured over 6 d and leaf decomposition measured over 28 d. A positive correlation also existed between in situ feeding and macroinvertebrate diversity and a biotic index. The G. pulex in situ feeding assay is a short-term sublethal biomonitor of water quality that is indicative of community- and ecosystem-level responses occurring over longer time periods. It is robust, responsive, and relevant.