Next generation sequencing improves the resolution of detecting mixed host blood meal sources in field collected arboviral mosquito vectors

Abstract

Accurate knowledge of blood meal hosts of different mosquito species is critical foridentifying potential vectors and establishing the risk of pathogen transmission. Wecompared the performance of Miseq next generation sequencing approach relative toconventional Sanger sequencing approach in identification of mosquito blood mealsusing genetic markers targeting the 12S rRNA and cytochrome oxidase I (COI) genes.We analysed the blood meals of three mosquito vector species (Aedes aegypti,Aedessimpsonis.l. andCulex pipienss.l.) collected outdoors, and compared the frequency ofsingle- versus multiple-blood feeding. Single host blood meals were mostly recovered forSanger-based sequencing of the mitochondrial 12S rRNA gene, whereas Miseq sequenc-ing employing this marker and the COI marker detected both single and multiple bloodmeal hosts in individual mosquitoes. Multiple blood meals (two or more hosts) whichmostly included humans were detected in 19%–22.7% ofAe. aegyptisamples. Most sin-gle host blood meals for this mosquito species were from humans (47.7%–57.1%) anddogs (9.1%–19.0%), with livestock, reptile and rodent hosts collectively accounting for4.7%–28.9% of single host blood meals. The frequency of two or more host blood mealsinAe. simpsonis.l. was 26.3%–45.5% mostly including humans, while single host bloodmeals were predominantly from humans (31.8%–47.4%) with representation of rodent,reptile and livestock blood meals (18.2%–68.2%). Single host blood meals fromCx.pipienss.l. were mostly from humans (27.0%–39.4%) and cows (11.5%–27.36%). Multipleblood meal hosts that mostly included humans occurred in 21.2%–24.4% ofCx. pipienss.l. samples. Estimated human blood indices ranged from 53%–76% forAe. aegypti,32%–82% forAe. simpsonis.l. and 26%–61% forCx. pipienss.l. and were consistentlylower for Sanger-based sequencing approach compared to Miseq-based sequencingapproach. These findings demonstrate that Miseq sequencing approach is superior toSanger sequencing approach as it can reliably identify mixed host blood meals in a singlemosquito, improving our ability to understand the transmission dynamics of mosquito-borne pathogens