Background & objectives: An outbreak of acute encephalitis syndrome was reported from Vidarbha region of Maharashtra State, India, during July 2012. to confirm the precise role of spp. in CHPV transmission. species, computer virus isolation, Vero E6 cell line Chandipura computer virus (CHPV) belongs to genus and was discovered during an outbreak of dengue-chikungunya-like illness in Nagpur district, Maharashtra, India, in 1965 from a patient with febrile illness1. Prevalence CX-5461 kinase inhibitor of CHPV in India was established by Mouse monoclonal to MPS1 sporadic cases reported from different parts of the country, computer virus isolations from humans and sandflies as well as presence of antibodies in humans and vertebrates2. However, CHPV, as a computer virus of public health importance was realized only when an outbreak of encephalitis with high case fatality rate (CFR) among children was reported from central India in 20033,4. The outbreak was characterized by high morbidity followed by rapid deterioration of cases and death in three Says of India, sandflies from India and from Africa during arbovirus investigations7. During July 2012, an outbreak of acute encephalitis syndrome (AES) with high case fatality was reported from several districts of Vidarbha region of Maharashtra viz. Nagpur, Bhandara, Chandrapur, Wardha, spp. during the recent July 2012 AES outbreak in Maharashtra. Material & Methods Sandfly collection was done in 13 villages/localities in the four districts of Maharashtra to determine their role in CHPV transmission (Table). Collection was made using hand held mouth aspirators from indoor and outdoor resting places. Oral consent from house owners was obtained to inspect their houses and peri-domestic areas for sandfly collection. Emphasis was given to collect sandflies CX-5461 kinase inhibitor from households, from where cases were reported. Majority of the houses had un-plastered brick/mud walls which are ideal for sandfly breeding. Collections were made from the damp/dark places of living rooms, kitchen, bathrooms, toilets and cattle sheds attached to the houses. The adult sandflies were transported alive to National Institute of Virology (NIV), Pune, and identified following the keys provided by Lewis8. Pools were prepared according to genera, gender and locality. Table Details of sandflies collected from Vidarbha region for computer virus isolation Open in a separate window Individual pools of sandflies were triturated in a small volume (0.5-1 ml) of chilled minimum essential medium (MEM, Sigma, USA), with pre-chilled, sterile mortars and pestles as described by Sudeep spp. and 17 to spp. (Table). Twenty nine pools of the former were prepared according to sex and locality and processed for computer virus isolation. sandflies could not be processed for computer virus isolation as none of them could be brought alive to the laboratory. In the first passage, CPE in Vero E6 cells was observed with three pools at 48 h post-infection (PI). However, in the 2nd passage, only one sample exhibited CPE and the CX-5461 kinase inhibitor other two failed. Distinct CPE was observed at 7 h PI in Vero E6 cell line. The isolate was obtained from a pool comprising only two female sandflies collected from Chachar village in Nagpur district (Table). RT-PCR studies targeting the N-gene confirmed the agent as CX-5461 kinase inhibitor CHPV as a 527 bp band corresponding to the N-gene was observed. A distinct band identical to positive control could be detected in the study (Fig.). Sequencing of the PCR product showed 10-12 nuecleotide changes in the new isolate in comparison to earlier CHPV.