ABSTRACT
CONTEXT: Melioidosis and Brucellosis are important endemic infections among people in India, especially in rural settings. Conventional detection techniques have several limitations. Only a few studies exist on the prevalence of Melioidosis and Brucellosis in rural area especially in India. AIM: We sought to evaluate detection of Burkholderia pseudomallei and Brucella spp. among patients presenting febrile illness. MATERIAL AND METHODS: Previously described polymerase chain reaction (PCR) assays for both pathogens were evaluated with Deoxyribonucleic acid extracts of buffy coat samples collected from 301 patients recruited prospectively. Data was not amenable to statistical analysis. RESULTS: The PCR showed specific amplification and no non-specific amplification with heterologous Gram-negative bacilli. The lower limit of detection of the assay for B. pseudomallei was determined to be 1 colony-forming unit /mL and for Brucella it was 1.95 × 10(3) plasmids per microliter. Blood culture in automated blood culture system was negative for all the samples. This prospective study carried out in southern India for the first time. PCR for Brucella was positive in 1% of the patient samples whereas 0.3% was positive for B. pseudomallei. CONCLUSION: The finding of Brucella and Burkholderia infections in our populations leads us to suggest that tests for Brucella and B. pseudomallei should also form part of a diagnostic platform for patients with Pyrexia of unknown origin in tropical developing countries.
ABSTRACT
BACKGROUND AND OBJECTIVE: 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). METHODS: 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. RESULTS: 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. CONCLUSION: 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.
