Vertebrate Host Identification from Mosquito Blood-Meals

We carried out blood-meal analysis on each individually extracted blood-fed mosquito to determine its vertebrate host. We used primers for cytochrome b (cyt b) and 16S rRNA markers to resolve the vertebrate host source of the blood-meals [25 (link)]. Total nucleic acid (1 μl) from each blood-fed mosquito was used as template in 10-μl PCRs containing 2 μl of 5X HOT FIREPol^® EvaGreen^® qPCR Mix (Solis BioDyne, Estonia) and 10 pmoles of each forward/reverse primer. Thermo-cycling and high-resolution melting (HRM) analysis were carried out in a Rotor-Gene Q real-time PCR thermo-cycler (Qiagen, Hilden Germany) as previously described [26 (link)]. DNA extracted from human, cattle, sheep, goat, pig, camel, and chicken samples served as positive controls in each of the runs. Rotor-Gene Q software 2.1.0 was used to select representative amplicons for post-PCR clean (Exo 1-rSAP combination, Biolabs, UK) and sequencing at Macrogen (The Netherlands).

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Chiuya T., Masiga D.K., Falzon L.C., Bastos A.D., Fèvre E.M, & Villinger J. (2021). A survey of mosquito-borne and insect-specific viruses in hospitals and livestock markets in western Kenya. PLoS ONE, 16(5), e0252369.

Publication 2021

5X HOT FIREPol® EvaGreen® qPCR Mix Rotor-Gene Q real-time PCR

16s rrna Blood Blood analysis Camel Cattle Chicken Cytochrome b Gene Goat Human Mosquito Nucleic acid Primer Q gene Real time pcr Sheep Vertebrate

Corresponding Organization : International Centre of Insect Physiology and Ecology

Other organizations : International Livestock Research Institute, University of Liverpool

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Variable analysis

independent variables

Vertebrate host source of the blood-meals

dependent variables

Vertebrate host source determined by cytochrome b (cyt b) and 16S rRNA markers

control variables

DNA extracted from human, cattle, sheep, goat, pig, camel, and chicken samples as positive controls

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