Prevalence of Salmonella typhi among patients with febrile illness in rural and peri-urban populations of Vellore district, as determined by nested PCR targeting the flagellin gene

Fever is one of the most common illnesses in all age groups in India. Typhoid fever is a continuing problem in developing countries such as India, which has poor sanitation facilities. The diagnosis of typhoid fever is still made by conventional culture-based isolation and identification. Serologic diagnostic tests, though widely used, have many deficiencies. Our objective was to investigate a molecular nested polymerase chain reaction (nPCR) technique to detect Salmonella typhi among patients with febrile illness in rural and peri-urban communities in Vellore district (Tamil Nadu, India). nPCR targeting the flagellin gene (fliC) was carried out using HotStar Taq DNA polymerase on DNA extracted from the buffy coat fraction of blood samples. Blood culture was done in a completely automated blood culture system, BacT/Alert(R), on prospectively collected blood samples. Relevant clinicopathologic data were obtained. nPCR was found to have a lower limit of detection of 0.01 colony-forming units per milliliter. The prevalence of typhoid fever as estimated by nPCR was 4.7% in pyrexia of unknown origin (PUO) in the rural/peri-urban communities of Vellore district. The prevalence of S. typhi as estimated by blood culture was 2.0%, which was lower than the nPCR estimation. nPCR had sensitivity and specificity of 100% and 97.3%, respectively. The observed agreement between blood culture and nPCR was 0.973 and the Kappa coefficient was 0.59 (p < 0.0001). The frequency of typhoid fever as detected by nPCR was 4.35% among rural patients and 5.32% among peri-urban individuals. nPCR on DNA extracts of buffy-coat samples using HotStar Taq was found to be a valuable and specific technique for diagnosis of typhoid fever.