Diagnostic methods for detection & isolation of dengue viruses from vector mosquitoes

Dengue is a deadly mosquito-borne infection warranting urgent attention for its containment particularly in the tropical and subtropical countries. In the absence of a vaccine or any specific drug for its treatment, an early diagnosis is considered indispensable to prevent any casualty. Detection of viruses in human sera particularly in endemic areas is cumbersome, difficult and also not desirable. Therefore, as an alternative approach, detection of the dengue virus antigen in mosquitoes has provided a reliable tool to (i) comprehend the types of viruses circulating in nature; and (ii) help in designing vector-specific control strategies. A mélange of diagnostic techniques are currently available with some advantages or disadvantages. Traditionally, cell cultures and suckling mice have been employed for virus isolations. While the virus isolation method in baby mice is time consuming, slow and expensive, the mosquito cell cultures offer a good degree of specificity. Mosquito inoculation techniques have been reported for detection and propagation of flaviviruses. Though this technique is sensitive for routine virological confirmation of dengue fever, it requires large number of infected mosquitoes, besides being time consuming. Insect bioassays (Toxo-IFA) are generally cumbersome requiring special facilities and are not suitable for large-scale epidemiological surveillance. ELISA has been shown to be a rapid and sensitive alternative to insect bioassays for monitoring arboviruses in wild populations. Reverse transcriptase polymerase chain reaction (RT-PCR) is a recent molecular diagnostic technology used for detecting virus infections in mosquitoes, which gives rapid results but is expensive and prone to contamination. This review describes the development of various techniques involved in detection and isolation of dengue viruses in mosquitoes. Definite diagnosis of the impending dengue epidemic can be made using ELISA for virological surveillance system on dengue virus antigen in the mosquito vectors. Therefore, ELISA offers a potential tool and a convenient system for quickly screening large number of samples up to the serotype level which can be employed effectively and efficiently for large scale dengue surveillance programmes on wild caught mosquito vectors. ELISA positive samples can be screened further by Toxo-IFA system for virus isolation. On the other hand, techniques like mosquitoes cell culture, mosquito inoculation (Toxo-IFA) and RT-PCR techniques can be employed for dengue virus amplification.