Aquatic Hemiptera in Southwest Cameroon: Biodiversity of Potential Reservoirs of Mycobacterium ulcerans and Multiple Wolbachia Sequence Types Revealed by Metagenomics

Seraphine N Esemu,Lucy M Ndip,Roland N Ndip,Benjamin L Makepeace,Achah J Kfusi,Rory J Post,Alistair C Darby,Catherine S Hartley,Xiaofeng Dong

doi:10.3390/d11120225

Abstract

Buruli ulcer (BU), caused by Mycobacterium ulcerans, is a neglected tropical disease associated with freshwater habitats. A variety of limnic organisms harbor this pathogen, including aquatic bugs (Hemiptera: Heteroptera), which have been hypothesized to be epidemiologically important reservoirs. Aquatic Hemiptera exhibit high levels of diversity in the tropics, but species identification remains challenging. In this study, we collected aquatic bugs from emerging foci of BU in the Southwest Region of Cameroon, which were identified using morphological and molecular methods. The bugs were screened for mycobacterial DNA and a selection of 20 mycobacteria-positive specimens from the families Gerridae and Veliidae were subjected to next-generation sequencing. Only one individual revealed putative M. ulcerans DNA, but all specimens contained sequences from the widespread alpha-proteobacterial symbiont, Wolbachia. Phylogenetic analysis placed the Wolbachia sequences into supergroups A, B, and F. Circularized mitogenomes were obtained for seven gerrids and two veliids, the first from these families for the African continent. This study suggests that aquatic Hemiptera may have a minor role (if any) in the spread of BU in Southwest Cameroon. Our metagenomic analysis provides new insights into the incursion of Wolbachia into aquatic environments and generated valuable resources to aid molecular taxonomic studies of aquatic Hemiptera.

Highlights

Mycobacterium ulcerans is a slow-growing environmental pathogen that infects the skin and subcutaneous tissues, causing Buruli ulcer (BU; known as Bairnsdale/Daintree ulcer in Australia) [1].BU is an emerging, treatable but neglected skin infection that manifests as slowly developing, unspecificDiversity 2019, 11, 225; doi:10.3390/d11120225 www.mdpi.com/journal/diversityDiversity 2019, 11, 225 indolent nodules, papules, or induration by oedema, which can progress to necrotizing skin ulcers [2].BU treatment outcomes can be good if the condition is diagnosed early, while late diagnosis can lead to extreme patient suffering and severe complications necessitating surgery or even amputation of limb(s) [3,4]
Emergence of BU has been linked to the acquisition of a megaplasmid, pMUM, by Mycobacterium marinum, a far less virulent environmental pathogen associated with aquatic habitats [7]
While our original goal to characterize environmental strains of M. ulcerans from aquatic bugs in this region was not achievable due to a virtual absence of the pathogen in our specimens, we demonstrate that serendipitously, they harbored surprisingly diverse Wolbachia sequence types

Summary

Introduction

Mycobacterium ulcerans is a slow-growing environmental pathogen that infects the skin and subcutaneous tissues, causing Buruli ulcer (BU; known as Bairnsdale/Daintree ulcer in Australia) [1].BU is an emerging, treatable but neglected skin infection that manifests as slowly developing, unspecificDiversity 2019, 11, 225; doi:10.3390/d11120225 www.mdpi.com/journal/diversityDiversity 2019, 11, 225 indolent nodules, papules, or induration by oedema, which can progress to necrotizing skin ulcers [2].BU treatment outcomes can be good if the condition is diagnosed early, while late diagnosis can lead to extreme patient suffering and severe complications necessitating surgery or even amputation of limb(s) [3,4]. Mycobacterium ulcerans is a slow-growing environmental pathogen that infects the skin and subcutaneous tissues, causing Buruli ulcer (BU; known as Bairnsdale/Daintree ulcer in Australia) [1]. BU is an emerging, treatable but neglected skin infection that manifests as slowly developing, unspecific. Emergence of BU has been linked to the acquisition of a megaplasmid, pMUM, by Mycobacterium marinum, a far less virulent environmental pathogen associated with aquatic habitats [7]. The megaplasmid encodes for polyketide synthases and accessory enzymes for the biosynthesis of a macrolide toxin, mycolactone [8], which is responsible for tissue damage and local immune suppression in BU [9]. Phylogenomic evidence indicates that all mycolactone-producing mycobacteria (MPM) form a monophyletic group comprising three clonal lineages, which should be considered ecovars of M. ulcerans rather than distinct species

Methods

Results

Discussion

Conclusion