Thematic Issue on Horizontal Gene Transfer An assessment of the potential of herbivorous insect gut bacteria to develop competence for natural transformation

Whereas the capability of DNA uptake has been well established for numerous species and strains of bacteria grown in vitro, the broader distribution of natural transformability within bacterial communities remains largely unexplored. Here, we investigate the ability of bacterial isolates from the gut of grass grub larvae (Costelytra zealandica (White); Coleoptera: Scarabaeidae) to develop natural genetic competence in vitro. A total of 37 mostly species-divergent strains isolated from the gut of grass grub larvae were selected for spontaneous rifampicin-resistance. Genomic DNA was subsequently isolated from the resistant strains and exposed to sensitive strains grown individually using established filter transformation protocols. DNA isolated from wild-type strains was used as a control. None of the 37 isolates tested exhibited a frequency of conversion to rifampicinresistance in the presence of DNA at rates that were significantly higher than the rate of spontaneous mutation to rifampicin-resistance in the presence of wild-type DNA (the limit of detection was approximately < 1 culturable transformant per 109 exposed bacteria). To further examine if conditions were conducive to bacterial DNA uptake in the grass grubs gut, we employed the competent bacterium Acinetobacter baylyi strain BD413 as a recipient species for in vivo studies. However, no transformants could be detected above the detection limit of 1 transformant per 103 cells, possibly due to low population density and limited growth of A. baylyi cells in grass grub guts. PCR analysis indicated that chromosomal Acinetobacter DNA remains detectable by PCR for up to 3 days after direct inoculation into the alimentary tract of grass grub larvae. Nevertheless, neither transforming activity of the DNA recovered from the alimentary tract of grass grubs larvae nor competence of bacterial cells recovered from inoculated larvae could be shown.