Figure 2

Comparison of Wolbachia marker sensitivity by PCR. (A) The three Wolbachia markers wsp, IS5 and ARM were tested on the following specimens: New world Drosophila species from the Drosophila willistoni group including D. paulistorum Amazonian (AM1, AM2), and Centroamerican (CA1, CA2) semispecies. Orinocan semispecies (OR) served as Wolbachia positive control; Ds ^- as Wolbachia negative control. B = blank. Quality of DNA was assessed with universal primer set 12SCFR, 12SCRR targeting the mitochondrial 12S rRNA gene [20, 21]. Expected amplicon sizes for Wolbachia positive control (OR) are 631 bp (wsp), 752 bp (IS5), 315 bp (ARM) and 399 bp (12S rRNA). (B) Same markers as above were tested on additional samples including hybrids: A/O hybrid plus parents AM and OR; Glossina Gs/Gm hybrid plus parental strains Gsw and Gmm (Additional file 2). Drosophila New world members include D. willistoni Dw ⁺ and Dw ^-. Old world species are D. melanogaster Dm ⁺; D. simulans Ds ⁺ and Ds ^-. B = blank. Note: IS5 primer set does not produce amplicons in all three Glossina samples due to complete absence of this IS element in symbionts of tsetse flies (see discussion).