Heritable endosymbionts of Drosophila

Abstract

Although heritable microorganisms are increasingly recognized as widespread in insects, no systematic screens for such symbionts have been conducted in Drosophila species (the primary insect genetic models for studies of evolution, development, and innate immunity). Previous efforts screened relatively few Drosophila lineages, mainly for Wolbachia. We conducted an extensive survey of potentially heritable endosymbionts from any bacterial lineage via PCR screens of mature ovaries in 181 recently collected fly strains representing 35 species from 11 species groups. Due to our fly sampling methods, however, we are likely to have missed fly strains infected with sex ratio-distorting endosymbionts. Only Wolbachia and Spiroplasma, both widespread in insects, were confirmed as symbionts. These findings indicate that in contrast to some other insect groups, other heritable symbionts are uncommon in Drosophila species, possibly reflecting a robust innate immune response that eliminates many bacteria. A more extensive survey targeted these two symbiont types through diagnostic PCR in 1225 strains representing 225 species from 32 species groups. Of these, 19 species were infected by Wolbachia while only 3 species had Spiroplasma. Several new strains of Wolbachia and Spiroplasma were discovered, including ones divergent from any reported to date. The phylogenetic distribution of Wolbachia and Spiroplasma in Drosophila is discussed.

Figure 1.—

(A) Phylogenetic distribution of drosophilid species screened for endosymbiotic bacteria. Phylogenetic relationships among most of the species groups and genera in the subfamily Drosophilinae are based on Markow and O'Grady (2006). Capitalized taxon names represent genera other than the genus Drosophila. Noncapitalized names are species groups within the genus Drosophila; the subgenus to which they belong is also indicated. For each taxon, numbers represent (from left to right, respectively) number of species screened mainly for Wolbachia in previous studies, number of species screened for all groups of bacteria (i.e., ovary extracts) in this study, number of species screened specifically for Wolbachia and Spiroplasma (i.e., whole flies) in this study, the combined total number of species screened in this study, the total species screened for Wolbachia on the basis of previous and this study. (B) Species found to be positive for Wolbachia or Spiroplasma from (left to right) previous studies, this study, the combined total, first for Wolbachia and then for Spiroplasma. Previous studies: ^aBourtzis et al. (1996); ^bCharlat et al. (2004); ^cClark et al. (2005); ^dDyer and Jaenike (2005); ^eGiordano et al. (1995); ^fHaine et al. (2005); ^gJaenike et al. (2003); ^hMiller and Riegler (2006); ⁱMontenegro et al. (2005); ^jMontenegro et al. (2006); ^kSalzberg et al. (2005a,b); ^lVavre et al. (1999); ^mVeneti et al. (2004); ⁿWerren and Jaenike (1995); ^oWerren et al. (1995a,b); ^pWilliamson and Poulson (1979); ^qWilliamson et al. (1999); ^rZhou et al. (1998).

Figure 2.—

Consensus of trees based on 16S rRNA gene of Wolbachia lineages inferred by maximum likelihood and Bayesian analyses. Numbers indicate the range of Bayesian posterior probabilities obtained under different sets of taxa and characters. Recognized Wolbachia supergroups are indicated by letters A–F. Taxon names indicate the host species and are followed by the GenBank accession number. Orange labels are haplotypes observed in this study but reported before, red labels had never been reported before, and blue labels are other Wolbachia strains associated with Drosophila reported in previous studies. Most analyses were based on a 1374-bp data set that included only taxa for which at least 1340 bp were available, but a subset of analyses included a taxon with a shorter sequence. Lineages from the D and C supergroups were removed sequentially in a subset of the analyses.

Figure 3.—

Maximum likelihood phylogeny of Spiroplasma 16S rRNA gene based on 439 characters. Haplotypes associated with Drosophila found in this study are indicated in red (hap 1–4), while the one found in a previous study is indicated in blue. Tree was rooted with Spiroplasma ixodetis. Numbers next to nodes indicate Bayesian posterior probabilities (>50%). Numbers next to taxon labels correspond to GenBank accessions, and host organism is indicated in parentheses.