Isolation of Chandipura virus (Vesiculovirus: Rhabdoviridae) from Sergentomyia species of sandflies from Nagpur, Maharashtra, India.

BACKGROUND & OBJECTIVES: An outbreak of acute encephalitis syndrome was reported from Vidarbha region of Maharashtra s0 tate, India, during July 2012. Anti-IgM antibodies against Chandipura virus (CHPV) were detected in clinical samples. Sandfly collections were done to determine their role in CHPV transmission. METHODS: Twenty nine pools of Sergentomyia spp. comprising 625 specimens were processed for virus isolation in Vero E6 cell line. Diagnostic RT-PCR targeting N-gene was carried out with the sample that showed cytopathic effects (CPE). The PCR product was sequenced, analysed and the sequences were deposited in Genbank database. RESULTS: CPE in Vero E6 cell line infected with three pools was detected at 48 h post infection. However, virus could be isolated only from one pool. RT-PCR studies demonstrated 527 nucleotide product that confirmed the agent as CHPV. Sequence analysis of the new isolate showed difference in 10-12 nucleotides in comparison to earlier isolates. INTERPRETATION & CONCLUSIONS: This is perhaps the first isolation of CHPV from Sergentomyia spp. in India and virus isolation during transmission season suggests their probable role in CHPV transmission. Further studies need to be done to confirm the precise role of Sargentomyia spp. in CHPV transmission.

Preliminary findings on Bagaza virus (Flavivirus: Flaviviridae) growth kinetics, transmission potential & transovarial transmission in three species of mosquitoes.

BACKGROUND & OBJECTIVES: Bagaza virus (BAGV), a flavivirus synonymous with Israel turkey meningoencephalitis virus, has been found to circulate in India. BAGV has recently been held responsible for inducing febrile illness in humans and causing unusually high mortality to wild birds in Spain. A study was therefore, undertaken to determine its replication kinetics in certain mosquitoes and to determine vector competence and potential of the mosquitoes to transmit BAGV experimentally. METHODS: Aedes aegypti, Culex tritaeniorhynchus and Cx quinquefasciatus mosquitoes were inoculated with BAGV; samples were harvested every day and titrated in BHK-21 cell line. Vector competence and experimental transmission were determined by examining the saliva of infected mosquitoes for virus and induction of sickness in suckling mice, respectively. RESULTS: Cx. tritaeniorhynchus and Ae. aegypti mosquitoes yielded 5 log10 and 4.67 log10 TCID50/ml of virus on day 3 post-infection (PI), respectively while Cx. quinquefasciatus yielded a titre of 4 log10 TCID50/ml on day 4 PI. BAGV was detected in saliva of all the infected mosquitoes demonstrating their vector competence. Experimental transmission of BAGV to infant mice as well as transovarial transmission was demonstrated by Cx. tritaeniorhynchus but not by Ae. aegypti and Cx. quinquefasciatus mosquitoes. INTERPRETATION & CONCLUSIONS: Replication of BAGV to high titres and dissemination to saliva in three most prevalent mosquitoes in India is of immense public health importance. Though no major outbreak involving man has been reported yet, BAGV has a potential to cause outbreaks in future.

Venereal transmission of Chandipura virus by Phlebotomus papatasi (Scopoli).

Experiments were conducted in the laboratory on Phlebotomus papatasi to determine the possible role of males in maintaining or sustaining the Chandipura virus (CHPV) activity in nature. This study indicated that infected males are capable of passing on the virus to female sand flies while mating. The infection rate was found to be 12.5% in uninfected females when mated with infected males. The occurrence of venereal transmission of this virus may have epidemiologic importance in the natural cycle of CHPV.

Vertical and venereal transmission of Chandipura virus (Rhabdoviridae) by Aedes aegypti (Diptera: Culicidae).

Experiments in the laboratory documented vertical and venereal transmission of Chandipura virus (CHPV) in Aedes aegypti (L.). The minimum filial infection rate among the progeny of infected females was 1.2%; the rate among male and female progeny was 0.9 and 1.4%, respectively. The venereal infection rate of CHPV among inseminated females was 32.7%. Our study indicates the possible occurrence of vertical and venereal transmission of CHPV in insect vectors.

Experimental studies on the vector potential of certain Culex species to West Nile virus.

Experimental studies were carried out to determine the vector potential of four species of mosquitoes to West Nile (WN) virus, viz. Culex tritaeniorhynchus, Cx. vishnui, Cx. bitaeniorhynchus and Cx. univittatus. All the four species of mosquitoes successfully transmitted and supported the growth of WN virus. The study indicated that the four species of mosquitoes could act as potential vectors of WN virus in nature.

Activity of dengue-2 virus and prevalence of Aedes aegypti in the Chirimiri colliery area, Madhya Pradesh, India.

Repeated outbreaks of a suspected viral fever in Chirimiri colliery area, Madhya Pradesh were reported since 1990. The area consists of an agglomeration of sprawling settlements at varying altitudes of 816 to 890 m and it has partial sylvan cover. During a 1992 outbreak 25 patients' sera were tested, of which 13 showed seropositivity to dengue (DEN) by MAC-ELISA test; DEN-2 was isolated from Aedes aegypti collected from two of the eight settlements of the area. The principal vector, Ae. aegypti, was prevalent in all the settlements studied; Breteau indices (BI) varied between 2.5 and 125.0; adult house indices (AHI) between 0 and 60.0%; Ae. albopictus and Ae. vittatus occurred in considerable numbers; Ae. aegypti bred in more containers with nonpotable water than those with potable water; the breeding of this species was noted in a maximum number of cement tanks while mud pots were predominant among the available containers. Paired comparisons between relative prevalence indices showed significant correlation and regression coefficients. Significant association of Ae. aegypti breeding with the households having tap water supply was noted, the relative risk declining with the people's use of well water either exclusively or in combination with other sources of water supply. It was also collected in the nonresidential areas. The role of ecological factors in the maintenance and spread of Ae. aegypti and dengue in these settlements is discussed.

Experimental studies on the susceptibility of Aedes vittatus to dengue viruses.

Ae. vittatus mosquitoes were infected by oral route and by intrathoracic inoculation with dengue (DEN) viruses and tested for the presence of dengue virus antigen in their head squashes and salivary glands by indirect immunofluorescence. The results indicate that this species was susceptible to all four types of DEN viruses and supported the growth of DEN-2 virus.

Susceptibility of four species of mosquitoes to Chandipura virus and its detection by immunofluorescence.

Susceptibility of Culex tritaeniorhynchus, Cx. Bitaeniorhynchus, Cx. quinquefasciatus, and Aedes aegypti to Chandipura (CHP) virus was compared after parental inoculation of the mosquitoes. Virus detection was done by indirect immunofluorescence (IF). CHP antigen in head squashes of all the four species was seen at 24 hr post infection (p.i.). The mosquitoes supported the virus growth and transmission by bite to 2 days old suckling Swiss albino mice. Ae. aegypti which was found the most susceptible mosquito species for CHP virus can be used as a substitute for laboratory mice.

Isolation of Japanese encephalitis virus from mosquitoes collected in Karnataka state, India from 1985 to 1987.

Detection and isolation of Japanese encephalitis (JE) virus using mosquito inoculation and immunofluorescence techniques were attempted from female mosquitoes collected in JE endemic areas of Kolar and Mandya districts of Karnataka state, India, from 1985 to 1987. 65,388 mosquitoes consisting of 19 species in 1541 pools were processed. Of these, 18 pools showed the presence of JE virus antigen. JE virus was isolated from 9 pools, 3 of Culex gelidus, 2 of C. tritaeniorhynchus, and one each of C. quinquefasciatus, C. fuscocephala, C. vishnui and Anopheles peditaeniatus. Isolation of JE virus from C. gelidus, C. fuscocephala, C. quinquefasciatus and An. peditaeniatus is reported for the first time in India.