Identification of Aedes aegypti and its Respective Life Stages by Real-Time PCR

Abstract : An Aedes aegypti specific fluorogenic probe hydrolysis (TaqMan) PCR assay was developed for real-time screening using a field-deployable thermocycler. Laboratory-based testing of Ae. aegypti, Ae. aegypti (Trinidad strain); Culex pipiens; Culex pipiens quinquefasciatus; Anopheles stephensi; Ochlerotatus taeniorhynchus individual adult mosquitoes and mixed pools (n=10) demonstrated 100% concordance in both in vitro sensitivity (6/6) and specificity (10/10). A single adult Aedes aegypti was identified in a pool of 100 non-Aedes aegypti mosquitoes. The limit of detection of Aedes aegypti egg pools was 5 individual eggs. Field-testing was conducted in central Honduras. An Aedes aegypti and Culex spp. panel of individual and mixed pools (n = 30) of adult mosquitoes, pupae, and larvae demonstrated 100% concordance in sensitivity (22/22) and 97% concordance in specificity (29/30) with one false positive. Field-testing of an Aedes aegypti and Culex spp. blind panel (n = 16) consisting of individual and mixed pools of adult mosquitoes, pupae, and larvae demonstrated 90% concordance in sensitivity (9/10) and 88% concordance in specificity (14/16).