The English case-control Infectious Intestinal Disease Study (1993-1996) failed to detect an enteric pathogen or toxin in 49% of cases of gastroenteritis. In the present study, polymerase chain reaction (PCR) assays were applied to DNA and cDNA generated from 4,627 faecal samples from cases and controls archived during the original study for the detection of norovirus, rotavirus, sapovirus, Campylobacter spp., Salmonella spp., enteroaggregative Escherichia coli, Cryptosporidium spp., and Giardia spp. The percentage of archived samples from cases and from controls in which at least one agent (or toxin) was detected increased from 53% in the original study to 75% and from 19 to 42%, respectively, after the application of PCR assays. Among cases, the following percentages of enteric pathogens were detected: norovirus 36%, rotavirus A 31%, sapovirus 4%, Salmonella spp. 6%, Campylobacter jejuni 13%, Campylobacter coli 2%, other Campylobacter spp. 8%, enteroaggregative E. coli 6%, Giardia spp. 2%, and Cryptosporidium spp. 2%. The present study provides additional insight into the aetiology of infectious intestinal disease in England and highlights the occurrence of viral infections in cases as well as in asymptomatic individuals. Other notable findings include the frequent presence of Campylobacter spp. other than C. jejuni or C. coli, the high frequency of multiple agents in 41% of cases and in 13% of controls, and the variation in the aetiology and rate of infection found for different age groups. The results demonstrate the greater sensitivity of PCR-based methods compared to current conventional methods.