Phylogenetic Analysis of Wolbachia from Folsomia Candida (Collembola: Isotomidae) via Sequences from WspB and Ribosomal Protein Genes

Abstract

Folsomia candida (Collembola: Isotomidae), a springtail that occurs commonly in soils throughout the world, is widely used in soil pollution and ecotoxicological studies. F. candida typically reproduces by parthenogenesis, which is widely believed to result from Wolbachia infection. Wolbachia is a cytoplasmically transmitted alpha‐proteobacterium that induces diverse reproductive alterations in its invertebrate hosts. Wolbachia is currently divided into six supergroups (A to F). Wolbachia from F. candida (wCan) is defined as a member of supergroup E based on sequences of 16S rDNA and ftsZ genes. Unexpectedly, however, no additional gene sequences have been described since wCan was first discovered in 1999. We have observed that recovery of undegraded DNA from springtails can be difficult. The objective of this study is to obtain additional gene sequences for wCan phylogenetic studies. We optimized recovery of DNA by lysing springtail eggs in the presence of 20mM EDTA, followed by extraction of undegraded DNA from agarose gels after electrophesis. Characterization of a nuclease that co‐purifies with springtail DNA is in progress. We used purified genomic DNA as template to amplify Wolbachia surface protein B (wspB) gene and five ribosomal protein genes by polymerase chain reaction (PCR). A concatenated phylogenetic tree based on these six gene sequences from wCan, together with corresponding sequences from nine additional Wolbachia strains, shows a closer relationship with supergroup B relative to supergroup A. Our study expands the DNA sequencing information of wCan and sheds light on its phylogeny.