Genetic diversity among laboratory and field populations of Chironomus riparius was investigated using microsatellite DNA analysis. Individuals of midge cultures reared in 10 different laboratories showed a clear reduction in the number of alleles per locus and in the level of heterozygosity compared to two natural populations sampled in southern Germany. To reconstruct the rate of genetic impoverishment under laboratory conditions, genetic diversity was monitored in a C. riparius laboratory population for 23 generations. Additionally, 11 populations originating from different laboratories were crossed, and genetic diversity was compared among pure, crossed, and natural populations. The results demonstrate that C. riparius strains used in standard toxicity tests exhibit low levels of genetic variability. In addition, our experiments show that refreshment of stocks with individuals from other laboratories is not sufficient to restore genetic variation. Potential consequences of genetic impoverishment and improvements for toxicological bioassays are discussed.