A global genotyping survey of Strongyloides stercoralis and Strongyloides fuelleborni using deep amplicon sequencing.

Strongyloidiasis is a neglected tropical disease caused by the human infective nematodes Strongyloides stercoralis, Strongyloides fuelleborni fuelleborni and Strongyloides fuelleborni kellyi. Previous large-scale studies exploring the genetic diversity of this important genus have focused on Southeast Asia, with a small number of isolates from the USA, Switzerland, Australia and several African countries having been genotyped. Consequently, little is known about the global distribution of geographic sub-variants of these nematodes and the genetic diversity that exists within the genus Strongyloides generally. We extracted DNA from human, dog and primate feces containing Strongyloides, collected from several countries representing all inhabited continents. Using a genotyping assay adapted for deep amplicon sequencing on the Illumina MiSeq platform, we sequenced the hyper-variable I and hyper-variable IV regions of the Strongyloides 18S rRNA gene and a fragment of the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene from these specimens. We report several novel findings including unique S. stercoralis and S. fuelleborni genotypes, and the first identifications of a previously unknown S. fuelleborni infecting humans within Australia. We expand on an existing Strongyloides genotyping scheme to accommodate S. fuelleborni and these novel genotypes. In doing so, we compare our data to all 18S and cox1 sequences of S. fuelleborni and S. stercoralis available in GenBank (to our knowledge), that overlap with the sequences generated using our approach. As this analysis represents more than 1,000 sequences collected from diverse hosts and locations, representing all inhabited continents, it allows a truly global understanding of the population genetic structure of the Strongyloides species infecting humans, non-human primates, and domestic dogs. Author summary Strongyloidiasis is a neglected tropical disease caused by the human infective worms (nematodes) Strongyloides stercoralis, Strongyloides fuelleborni fuelleborni and Strongyloides fuelleborni kellyi. Little is known about the genetic diversity of these nematodes and the possibility of geographically isolated genetic types is a particularly interesting research question, given the unique life cycle of these worms which includes both sexual and asexual stages. We extracted DNA from human, dog and primate feces containing Strongyloides, collected from several countries representing all inhabited continents. Using next-generation sequencing, we analyzed three Strongyloides DNA fragments from these specimens; two fragments of the 18S gene and one from the cytochrome c oxidase subunit 1 (cox1) gene. Using this approach, we discovered some unique S. stercoralis and S. fuelleborni genotypes, and identified a previously unknown S. fuelleborni infecting humans within Australia. We compared our data to all 18S and cox1 sequences of S. fuelleborni and S. stercoralis available in public databases and identified several patterns relating to the global distribution of certain genotypes. This knowledge could allow us to infer the origin of human Strongyloides infections in the future, and assess the role certain animals (non-human primates and dogs) might play in the transmission of Strongyloides to humans.