Brugia malayi Gene Expression in Response to the Targeting of the Wolbachia Endosymbiont by Tetracycline Treatment

Abstract

Background Brugia malayi, like most human filarial parasite species, harbors an endosymbiotic bacterium of the genus Wolbachia. Elimination of the endosymbiont leads to sterilization of the adult female. Previous biochemical and genetic studies have established that communication with its endobacterium is essential for survival of the worm.Methodology/Principal findingsWe used electron microscopy to examine the effects of antibiotic treatment on Wolbachia cell structure. We have also used microarray and quantitative RT-PCR analyses to examine the regulation of the B. malayi transcripts altered in response to the anti-Wolbachia treatment. Microscopy of worms taken from animals treated with tetracycline for 14 and 21 days (14 d and 21 d) demonstrated substantial morphologic effects on the Wolbachia endobacterium by 14 d and complete degeneration of the endobacterial structures by 21 d. We observed upregulation of transcripts primarily encoding proteins involved in amino acid synthesis and protein translation, and downregulation of transcripts involved in cuticle biosynthesis after both 7 d and 14 d of treatment. In worms exposed to tetracycline in culture, substantial effects on endobacteria morphology were evident by day 3, and extensive death of the endobacteria was observed by day 5. In a detailed examination of the expression kinetics of selected signaling genes carried out on such cultured worms, a bimodal pattern of regulation was observed. The selected genes were upregulated during the early phase of antibiotic treatment and quickly downregulated in the following days. These same genes were upregulated once more at 6 days post-treatment.Conclusions/SignificanceUpregulation of protein translation and amino acid synthesis may indicate a generalized stress response induced in B. malayi due to a shortage of essential nutrients/factors that are otherwise supplied by Wolbachia. Downregulation of transcripts involved in cuticle biosynthesis perhaps reflects a disruption in the normal embryogenic program. This is confirmed by the expression pattern of transcripts that may be representative of the worms' response to Wolbachia in different tissues; the early peak potentially reflects the effect of bacteria death on the embryogenic program while the second peak may be a manifestation of the adult worm response to the affected bacteria within the hypodermis.

A feedback loop betweenWolbachiaand theDrosophila gurkenmRNP complex influencesWolbachiatiter

“…4C,D). However, all bacteria examined in wild-type and grk nurse cells were encompassed by smooth, contiguous membrane typical of Wolbachia (Stouthamer et al, 1999;Serbus et al, 2008;Werren et al, 2008), unlike the whorled, fragmented appearance observed for Wolbachia killed by tetracycline (Ghedin et al, 2009). The bacterial cytoplasm also appeared homogenous, lacking the electron-dense accumulations visible in Wolbachia after heat shock or when heavily loaded with phage particles (Bordenstein et al, 2006;Zhukova et al, 2008).…”

Wolbachia infection increases recapture rate of field-released Drosophila melanogaster

“…Wolbachia has now been shown to affect host mating competitiveness , metabolic rate (Evans et al 2009), locomotor activity (Evans et al 2009;Fleury et al 2000), responsiveness to food cues (Peng et al 2008), fecundity (Fleury et al 2000;Weeks et al 2007), and longevity (Min and Benzer 1997), as well as provide nutritional supplementation (Brownlie et al 2009;Ghedin et al 2009) and protection against pathogens (Hedges et al 2008;Kambris et al 2009;Moreira et al 2009;Osborne et al 2009). …”

Wolbachia endosymbionts and human disease control

“…Not unexpectedly, in microarray experiments, following antibiotic treatment, Wolbachia genes almost exclusively showed down-regulation [42]. Three Wolbachia genes (hypothetical, shortchain alcohol dehydrogenase and stress induced morphogen) were up-regulated following treatment [41], however, it is not yet clear as to the significance of these observations. Broadly similar changes were observed in L. sigmodontis; expression of genes involved in translation, transcription, protein folding/sorting, structure, motility, metabolism, signaling and immunomodulation were affected by Wolbachia clearance [43].…”

“…In a microarray experiment conducted with B. malayi, genes in certain classes (e.g., signaling) showed a bimodal pattern of regulation in response to antibiotic treatment [41]. Since antibiotics affect embryogenesis in advance of worm viability, the authors postulated that early changes in transcript levels reflect disruption of the embryo program while later transcriptional changes are the result of reduction of the Wolbachia load in the hypodermis.…”