Introduction
Chikungunya virus (CHIKV), is transmitted to humans throughStegomyia mosquitoes. Kerala was the worst affected State in Indian during 2007 with a total of 55.8% of the reported Chikungunya fever cases in the country from the State (NVBDCP, India). Almost all the districts of Kerala were affected with the infection during 2006-2007. The factors attributed to this outbreak were attributed to crucial mutation “A226” acquired by the virus in 2007 and the abundance of the vector species Stegomyia albopicta in the region (Kumar,et al ., 2008). Stegomyia aegypti andStegomyia albopicta , the vectors for CHIKV were widely distributed and abundant during the pre and post monsoon season. The prevailing climate, terrain and agricultural practices in these districts were highly conducive for the breeding of St. albopicta and St.aegypti , the vector of Chikungunya and Dengue in Kerala. The worst affected region was the mid-highland regions which harbour the vast rubber plantation sectors in the State. The hilly and semi-forested districts of Kottayam and Pathanamthitta were the most affected districts during 2007 epidemic, which constituted the major portion in the rubber plantation sectors of Kerala. About 63.0% of people living in the rubber plantation areas (Kerala contributes 80.0% of the rubber production in India), was afflicted with this disease (Kumar et al ., 2011), St. albopicta acted as main vector species (Kumaret al ., 2008).
Incidence of Dengue fever, another mosquito borne arbo-viral disease is also on an increasing trend in Kerala (Kumar et al ., 2013).St. albopicta (Skuse) 1894 (Diptera: Culicidae), the predominantStegomyia species prevalent in Kerala was recorded as the vector species of both these arbo-viral diseases (Kumar et al ., 2008; Thenmozhi et al ., 2007). Innumerable Discarded Latex Collection containers (DLCCs), unused Fixed Latex Collection Containers (FLCCs), and tree holes, leaf axils of pine-apple plants and fallen leaves of areca-nut tree etc. were recorded the key breeding habitats of this species in rubber plantation sectors in Kerala (Kumar et al ., 2011; Sumodan, 2003) .
Dengue a vector – borne and emerging infectious disease is estimated to affect 50-100 million individuals each year in tropical and subtropical areas (Gubler, 2006; Halstead, 2007). Demographic and societal changes, such as population growth, unplanned urbanization, microevolution of the virus, climatic change and modern transportation, have greatly contributed to the increased incidence and geographical spread of Dengue virus infection in recent decades and distributed among about 120 countries globally. Dengue is caused by four serologically distinct types of Dengue virus. DENV-1, DENV-2, DENV-3 and DENV-4, belonging to the family Flaviviridae , genus Flavivirus . The viruses are transmitted to human beings through the bite of Stegomyiamosquitoes such as St. aegypti and St. albopicta which are considered as primary and secondary vectors of Dengue respectively (WHO, 2015) . The incidence of Dengue fever has increased significantly around the world in recent decades. About half of the global population lives under the risk of infection of this arbo-viral disease(WHO, 2009) .St. albopicta is becoming an increasingly important vector of Dengue because of its rapidly changing global distribution (Rezza, 2012) and it is generally believed to be a less efficient vector of arboviruses than St.aegypti . St. albopicta is highly adaptive and therefore can survive even in cooler temperate regions. Its spread to new areas is due to its tolerance to temperatures below freezing, hibernation and ability to shelter in microhabitats (WHO, 2015).
St. albopicta is (the Asian tiger mosquito), is an aggressive day-biting species known in different parts of the world including South East Asia (Russell et al., 1969) , and Southern China (Qui et al., 1981) . It belongs to the East and Southeast Asia, where it was originally lived at the edges of forests breeding in tree hole, containers and other natural reservoir (Rezza, 2012) .Worldwide trade in second hand tires transported to various places, which often contain water could be an ideal place for eggs and larvae of these mosquitoes, has been a key factor in the large-scale conquest of St. albopicta in new areas which easily adapts to new environments, even in a temperate climate. This expansion is creating new opportunities for viruses to circulate in new areas, becoming a common cause of epidemics in St. aegypti -free countries from Hawaii to Mauritius. HoweverSt. albopicta is considered as an inefficient vector of Dengue because not well adapted to urban domestic environments and is less anthropophilic than St.aegypti .
The entire population of India lives under the risk of Dengue. In India the first case of Dengue has been reported from Kolkata (previously Calcutta) during 1963. The number of Dengue cases had been an increasing trend since 2001 and the maximum number of suspected cases (28055) has been recorded during 2010. In Kerala Dengue fever was first recorded in Kottayam District in 1997 (Kalra and Prasittisuk, 2004). This district continues to contribute the maximum number of cases of Dengue fever in the State, next to Trivandrum District in the south Kerala every year. Topographically, Kottayam District is semi-forested region with hills and hillocks located at the foothills of the Western Ghats. This district is the abode of rubber plantations in the country and grows rubber in about 60% of its net sown area under agriculture. St. albopicta the vector of Dengue fever, is abundant throughout the plantation belt of Kerala, including Kottayam District (Kumar et al. , 2011). All the four Dengue serotype are prevalent in the State, however, Dengue 2 and 3 are the major serotypes prevalent in Kerala State (Kumar et al ., 2013).
Rubber plantations in the region could be classified as immature ones and mature ones based on the age and productivity of the trees, broadly. Latex collection is carried out by placing hemispherical plastic containers of either 450ml/600ml/900ml capacity, attached to the bark of the mature trees about 7 years old. A small portion of bark of the tree is removed in a horizontal oblique manner by a sharp knife in an interval of 2-5 days and the latex oozing out is collected into the containers fitted to the tree. Also rain guards are fitted to the latex yielding section of the bark to prevent rainwater interfering with the tapping process and collection of latex. The latex oozing out of the scar is removed about 3 hours after making the cuts in the bark. The container is kept in the tree itself after latex collection in upright position to collect the residual latex.
In absence of effective vaccines or prophylaxis against most of the arboviruses, control of these vector borne diseases heavily rely on vector control. Community oriented Integrated Vector Management (IVM) strategies involving inter-sectoral collaboration is the best approach proposed for the same. However, during outbreak situations, usage of adulticidal measures using insecticides are being resorted to, towards rapid management of the outbreaks (WHO, 2008). Recently, studies have indicated that Stegomyia species when infected by the intracellular proteobacteria called Wolbachia limits pathogen replication, in particular the enveloped, positive single-stranded RNA viruses such as dengue (DENV), yellow fever (YFV) and chikungunya (CHIKV) (Moreira et al ., 2009; Bian et al ., 2010; Walkeret al ., 2011). Based on this observation, a global campaign to eliminate dengue (http://www.eliminatedengue.com) was initiated to control dengue in countries such as Australia, Vietnam, Brazil, Colombia, Indonesia etc. by introduction of Wolbachia into the populations of these vector species. However, the mechanism of pathogen inhibition by Wolbachia is still being investigated, but this blocking phenomenon has been linked to priming of the host innate immune system and competition for limited resources between pathogens andWolbachia (Caragata et al., 2013). Wolbachia is found in many arthropods such as insects, mites and isopods and also in filarial nematodes. This natural insect symbiont is reported to be most concentrated in reproductive organs, but often have a wide tissue distribution within the host insect, including midgut and salivary glands. Wolbachia infection in St . albopicta was discovered by Wright and Barr in 1980 and is reported to be super infected with wAlbA and wAlbB strains. These strains are known to play a crucial role in dynamics, evolution and reproductive system of their host. Usually the arboviruses acquired through the blood meal must invade midgut cells and ultimately disseminate to the salivary glands to be transmitted, so that Wolbachia and the virus can potentially be present within the same cells. Wolbachia itself is not an infectious agent, but is maternally transmitted in the egg cytoplasm and is able to invade host populations by manipulating their reproduction. Hence, considerable scientific attention was directed at better understanding the impact of Wolbachia infections on the evolution and ecology of host populations (Charlat et al , 2003). Moreover, the self-spreading ability of Wolbachia makes it an attractive biocontrol agent to interrupt or reduce arbovirus transmission (Sinkins, 2013). However, the infection prevalence details of Wolbachia in natural populations of St. albopictus in endemic regions of the arbo-viral Diseases as Dengue and Chikungunya remains not well documented. Hence, we undertook a study to elucidate the natural infection status of this parasite in this vector species in a highly endemic state of Dengue Kerala.