A Salmonella-specific DNA fragment of the Salmonella typhimurium LT2 chromosome has been isolated. The fragment (2.3 kilobases (kb)) was used as a probe in a colony hybridization assay, where 185 strains of 93 different Salmonella serovars were correctly identified as belonging to Salmonella. The specificity of the probe was evaluated in colony hybridization assays on pure cultures of non-Salmonella bacteria and on specimens with an indigenous flora. Sixty-three strains of 34 non-Salmonella Gram-negative species did not hybridize to the fragment. By DNA hybridization to faecal samples from calves, pigs and chickens, and samples of animal feed, three samples out of 10 positive by traditional culture methods gave negative results by hybridization, 45 samples were negative in both methods, while one sample was positive only in the hybridization assay. From this sample, Salmonella livingstone was isolated by a replica plate hybridization technique. The probe therefore proved 100% specific for the genus Salmonella. The 2.3 kb fragment may form the basis of hybridization assays for specific detection of Salmonella in food, environmental and clinical specimens.