Concurrent transcriptional profiling of Dirofilaria immitis and its Wolbachia endosymbiont throughout the nematode life cycle reveals coordinated gene expression.

Ashley N Luck,Jeremy M Foster,Michelle L Michalski,Christopher C Evans,Molly D Riggs,Barton E Slatko,Andrew R Moorhead

doi:10.1186/1471-2164-15-1041

Abstract

BackgroundDirofilaria immitis, or canine heartworm, is a filarial nematode parasite that infects dogs and other mammals worldwide. Current disease control relies on regular administration of anthelmintic preventives, however, relatively poor compliance and evidence of developing drug resistance could warrant alternative measures against D. immitis and related human filarial infections be taken. As with many other filarial nematodes, D. immitis contains Wolbachia, an obligate bacterial endosymbiont thought to be involved in providing certain critical metabolites to the nematode. Correlations between nematode and Wolbachia transcriptomes during development have not been examined. Therefore, we detailed the developmental transcriptome of both D. immitis and its Wolbachia (wDi) in order to gain a better understanding of parasite-endosymbiont interactions throughout the nematode life cycle.ResultsOver 215 million single-end 50 bp reads were generated from total RNA from D. immitis adult males and females, microfilariae (mf) and third and fourth-stage larvae (L3 and L4). We critically evaluated the transcriptomes of the various life cycle stages to reveal sex-biased transcriptional patterns, as well as transcriptional differences between larval stages that may be involved in larval maturation. Hierarchical clustering revealed both D. immitis and wDi transcriptional activity in the L3 stage is clearly distinct from other life cycle stages. Interestingly, a large proportion of both D. immitis and wDi genes display microfilarial-biased transcriptional patterns. Concurrent transcriptome sequencing identified potential molecular interactions between parasite and endosymbiont that are more prominent during certain life cycle stages. In support of metabolite provisioning between filarial nematodes and Wolbachia, the synthesis of the critical metabolite, heme, by wDi appears to be synchronized in a stage-specific manner (mf-specific) with the production of heme-binding proteins in D. immitis.ConclusionsOur integrated transcriptomic study has highlighted interesting correlations between Wolbachia and D. immitis transcription throughout the life cycle and provided a resource that may be used for the development of novel intervention strategies, not only for the treatment and prevention of D. immitis infections, but of other closely related human parasites as well.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-1041) contains supplementary material, which is available to authorized users.

Highlights

Dirofilaria immitis, or canine heartworm, is a filarial nematode parasite that infects dogs and other mammals worldwide
Third-stage larvae (L3) were obtained by gently crushing the infected mosquitoes, rinsing them onto a 32 μm mesh sieve set in a petri dish and soaking them in warm Hanks’ balanced salt solution, with larvae settling to the bottom of the dish for collection
Many reads map to rRNA, ~80% of all D. immitis predicted gene models were expressed in every life cycle stage examined (Table 1)

Summary

Introduction

Dirofilaria immitis, or canine heartworm, is a filarial nematode parasite that infects dogs and other mammals worldwide. We detailed the developmental transcriptome of both D. immitis and its Wolbachia (wDi) in order to gain a better understanding of parasite-endosymbiont interactions throughout the nematode life cycle. Dirofilaria immitis, the causative agent of canine heartworm disease, is a parasitic filarial nematode evolutionarily related to those responsible for human parasitic diseases such as lymphatic and cutaneous filariases. D. immitis requires an arthropod vector for transmission (in this case Aedes, Anopheles or Culex mosquitoes), as well as a mammalian host. The life cycle of D. immitis follows that of other filarial nematodes in that infected insects, mosquitoes in this case, introduce third-stage larvae (L3) into the vertebrate host during a blood meal. Circulating mf are ingested by a mosquito during another blood meal and molt twice within the vector before becoming infective L3 larvae [3]

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