DNA from whole mosquitoes was prepared with the Promega Wizard Genomic DNA Purification Kit according to the manufacturer’s instructions. Bacterial 16 S rDNA (Escherichia coli position 341–805) were amplified by using general bacterial primers 341F (CCTACGGGNGGCWGCAG) and 805R (GACTACHVGGGTATCTAATCC)26 (link). This primer pair matches approximately 90% of all good-quality bacterial sequences and covers all phyla in the Ribosomal Database Project release 10.25. Each DNA sample was individually PCR-amplified with Ready to go PCR beads (GE Health Care) by initial denaturation at 95 °C for 5 min followed by 35 cycles of [40 s at 95 °C, 40 s at 53 °C and 1 min at 72 °C] followed by a final 7-min extension at 72 °C. In a second PCR was added 1 of 50 flanking barcode sequence pairs to run samples in parallel27 (link) using the same conditions as above, but only for 10 cycles of iteration.
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