Development and validation of a PCR-based method to differentiate human sex in blood-fed Aedes aegypti (Diptera: Culicidae)

Monitoring mosquito host choice to identify high-risk groups for different vector-borne diseases is important to devise vector control strategies and disease management. The present study was conducted to develop and validate a PCR-based method to identify human sex in blood-fed Aedes aegypti mosquitoes. Several human genes present in both the X and Y chromosomes were screened and diagnostic PCR primers were successfully designed and amplified for the human STS gene. The limit of detection of this PCR assay was carried out on Ae. aegypti fed with human blood up to 5 days (120 hours) post blood-meal under laboratory condition. The efficiency of this PCR assay was evaluated in field-collected Ae. aegypti mosquitoes and compared with other existing methods. The developed PCR primers can successfully amplify and distinguish human sex in mosquitoes up to 72 hours after a blood meal, with an amplified product of 627bp and 298bp for male (XY) and 627bp for female (XX) blood-fed mosquitoes. Further, validation of this assay in field-collected Ae. aegypti mosquitoes revealed that this assay could detect human sex in mosquito blood meal substantially more efficiently (X-2 = 4.5, p = 0.034) than other PCR based assay. The newly developed PCR assay highly specific to human DNA and can distinguish male and female DNA for up to 72 hours. This assay can be is used for identifying highrisk groups and extended to other medically important hematophagous insects to assess their role in disease transmission and epidemic preparedness.