Division of Virology, Defiance Research and Development Establishment, Jhansi Road, Gwalior-474002, India.
Abstracts of the papers presented at the International Conference of Indian Virological Society on “Emerging and Re-emerging viral Diseases of the Tropics and Subtropics” at Indian Agricultural Research Institute, New Delhi, India, December 11–14, 2007.
The development of a One-step SYBR Green I based real-time RT-PCR assay is reported for detection and quantification of Chikungunya virus (CHIKV) in acute phase patient serum samples and cell culture supernatant by targeting the E1 structural gene. A linear relationship was obtained between the virus concentration and cycle threshold (Ct) value over a range of 107 to 0.1PFU/ml. The Ct values obtained against the known concentration of serially diluted CHIK virus was used for construction of standard curve. The reported assay was found to be 10 fold more sensitive compared to conventional RT-PCR with a detection limit of 0.1 PFU/ml. The feasibility of this reported assay system for clinical diagnosis was validated with 51 suspected clinical serum samples of the CHIKV epidemic in Southern India, 2006. The comparative evaluation with acute phase patient serum samples revealed the higher sensitivity of real-time RT-PCR assay by picking up 6 additional samples with low copy number of template. None of the healthy serum samples analyzed in this study showed amplification. The quantification of the viral load in the acute phase serum samples was also determined employing standard curve, which varies from 0.1 to 107 PFU/ml. These findings demonstrated that the reported assay has the potential usefulness for clinical diagnosis due to simultaneous detection and quantification of chikungunya virus in acute phase patient serum samples.
