Abstract
Parasitic helminth infections have a considerable impact on global human health as well as animal welfare and production. Although co-infection with multiple parasite species within a host is common, there is a dearth of tools with which to study the composition of these complex parasite communities. Helminth species vary in their pathogenicity, epidemiology and drug sensitivity and the interactions that occur between co-infecting species and their hosts are poorly understood. We describe the first application of deep amplicon sequencing to study parasitic nematode communities as well as introduce the concept of the gastro-intestinal “nemabiome”. The approach is analogous to 16S rDNA deep sequencing used to explore microbial communities, but utilizes the nematode ITS-2 rDNA locus instead. Gastro-intestinal parasites of cattle were used to develop the concept, as this host has many well-defined gastro-intestinal nematode species that commonly occur as complex co-infections. Further, the availability of pure mono-parasite populations from experimentally infected cattle allowed us to prepare mock parasite communities to determine, and correct for, species representation biases in the sequence data. We demonstrate that, once these biases have been corrected, accurate relative quantitation of gastro-intestinal parasitic nematode communities in cattle fecal samples can be achieved. We have validated the accuracy of the method applied to field-samples by comparing the results of detailed morphological examination of L3 larvae populations with those of the sequencing assay. The results illustrate the insights that can be gained into the species composition of parasite communities, using grazing cattle in the mid-west USA as an example. However, both the technical approach and the concept of the ‘nemabiome’ have a wide range of potential applications in human and veterinary medicine. These include investigations of host-parasite and parasite-parasite interactions during co-infection, parasite epidemiology, parasite ecology and the response of parasite populations to both drug treatments and control programs.
Highlights
Helminth infections have a considerable global impact on human and animal health [1,2,3]
We show that the approach is extremely robust and quantitatively accurate when applied to field populations harvested from cattle feces and discuss how the approach is applicable to study the biodiversity of parasite populations in other domestic animal and wildlife hosts as well as in humans
The ITS-2 rDNA was amplified and sequenced from 15 individual L3 larvae taken from the characterized laboratory passaged isolates of C. punctata, C. oncophora, N. helvetianus, T. axei, T. colubriformis, H. placei, O. ostertagi and O. radiatum to assess intra- and inter-species variation
Summary
Helminth infections have a considerable global impact on human and animal health [1,2,3]. The use of next-generation sequencing technologies is currently revolutionizing the diagnosis of bacterial and viral infections as well as opening up new areas of research including the exploration of the ‘microbiome’ [7,8] Such approaches have not yet been applied to metazoan parasite communities, even though mixed infections are the norm rather than the exception in both humans and animals [9]. Applying “microbiome” type approaches would allow the investigation of interactions between the complex assemblages of parasite species and their hosts and their consequences for transmission dynamics, immunity and parasite ecology Such approaches have massive potential as new tools for diagnosis and surveillance as well as for investigating the response of parasite populations to drug treatments and other control strategies
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