Identification of Novel Genes and Proteoforms in Angiostrongylus costaricensis through a Proteogenomic Approach.

RNA sequencing (RNA-Seq) and mass-spectrometry-based proteomics data are often integrated in proteogenomic studies to assist in the prediction of eukaryote genome features, such as genes, splicing, single-nucleotide (SNVs), and single-amino-acid variants (SAAVs). Most genomes of parasite nematodes are draft versions that lack transcript- and protein-level information and whose gene annotations rely only on computational predictions. Angiostrongylus costaricensis is a roundworm species that causes an intestinal inflammatory disease, known as abdominal angiostrongyliasis (AA). Currently, there is no drug available that acts directly on this parasite, mostly due to the sparse understanding of its molecular characteristics. The available genome of A. costaricensis, specific to the Costa Rica strain, is a draft version that is not supported by transcript- or protein-level evidence. This study used RNA-Seq and MS/MS data to perform an in-depth annotation of the A. costaricensis genome. Our prediction improved the reference annotation with (a) novel coding and non-coding genes; (b) pieces of evidence of alternative splicing generating new proteoforms; and (c) a list of SNVs between the Brazilian (Crissiumal) and the Costa Rica strain. To the best of our knowledge, this is the first time that a multi-omics approach has been used to improve the genome annotation of A. costaricensis. We hope this improved genome annotation can assist in the future development of drugs, kits, and vaccines to treat, diagnose, and prevent AA caused by either the Brazil strain (Crissiumal) or the Costa Rica strain.

The Differences in the Susceptibility Patterns to Triclabendazole Sulfoxide in Field Isolates of Fasciola hepatica Are Associated with Geographic, Seasonal, and Morphometric Variations.

Triclabendazole (TCBZ) resistance is an emerging problem in fascioliasis that is not well understood. Studies including small numbers of parasites fail to capture the complexity of susceptibility variations between and within Fasciolahepatica populations. As the first step to studying the complex resistant phenotype−genotype associations, we characterized a large sample of adult F. hepatica with diverging TCBZ susceptibility. We collected parasites from naturally infected livestock slaughtered in the Cusco and Cajamarca regions of Peru. These parasites were exposed to TCBZ sulfoxide (TCBZ.SO) in vitro to determine their susceptibility. We used a motility score to determine the parasite's viability. We titrated drug concentrations and times to detect 20% non-viable (susceptible conditions) or 80% non-viable (resistant conditions) parasites. We exposed 3348 fully motile parasites to susceptible (n = 1565) or resistant (n = 1783) conditions. Three hundred and forty-one (21.8%) were classified as susceptible and 462 (25.9%) were classified as resistant. More resistant parasites were found in Cusco than in Cajamarca (p < 0.001). Resistant parasites varied by slaughterhouse (p < 0.001), month of the year (p = 0.008), fluke length (p = 0.016), and year of collection (p < 0.001). The in vitro susceptibility to TCBZ.SO in wildtype F. hepatica was associated with geography, season, and morphometry.

Genomes of Fasciola hepatica from the Americas Reveal Colonization with Neorickettsia Endobacteria Related to the Agents of Potomac Horse and Human Sennetsu Fevers.

Food borne trematodes (FBTs) are an assemblage of platyhelminth parasites transmitted through the food chain, four of which are recognized as neglected tropical diseases (NTDs). Fascioliasis stands out among the other NTDs due to its broad and significant impact on both human and animal health, as Fasciola sp., are also considered major pathogens of domesticated ruminants. Here we present a reference genome sequence of the common liver fluke, Fasciola hepatica isolated from sheep, complementing previously reported isolate from cattle. A total of 14,642 genes were predicted from the 1.14 GB genome of the liver fluke. Comparative genomics indicated that F. hepatica Oregon and related food-borne trematodes are metabolically less constrained than schistosomes and cestodes, taking advantage of the richer millieux offered by the hepatobiliary organs. Protease families differentially expanded between diverse trematodes may facilitate migration and survival within the heterogeneous environments and niches within the mammalian host. Surprisingly, the sequencing of Oregon and Uruguay F. hepatica isolates led to the first discovery of an endobacteria in this species. Two contigs from the F. hepatica Oregon assembly were joined to complete the 859,205 bp genome of a novel Neorickettsia endobacterium (nFh) closely related to the etiological agents of human Sennetsu and Potomac horse fevers. Immunohistochemical studies targeting a Neorickettsia surface protein found nFh in specific organs and tissues of the adult trematode including the female reproductive tract, eggs, the Mehlis' gland, seminal vesicle, and oral suckers, suggesting putative routes for fluke-to-fluke and fluke-to-host transmission. The genomes of F. hepatica and nFh will serve as a resource for further exploration of the biology of F. hepatica, and specifically its newly discovered trans-kingdom interaction with nFh and the impact of both species on disease in ruminants and humans.

Genomic diversity in Onchocerca volvulus and its Wolbachia endosymbiont.

Ongoing elimination efforts have altered the global distribution of Onchocerca volvulus, the agent of river blindness, and further population restructuring is expected as efforts continue. Therefore, a better understanding of population genetic processes and their effect on biogeography is needed to support elimination goals. We describe O. volvulus genome variation in 27 isolates from the early 1990s (before widespread mass treatment) from four distinct locales: Ecuador, Uganda, the West African forest and the West African savanna. We observed genetic substructuring between Ecuador and West Africa and between the West African forest and savanna bioclimes, with evidence of unidirectional gene flow from savanna to forest strains. We identified forest:savanna-discriminatory genomic regions and report a set of ancestry informative loci that can be used to differentiate between forest, savanna and admixed isolates, which has not previously been possible. We observed mito-nuclear discordance possibly stemming from incomplete lineage sorting. The catalogue of the nuclear, mitochondrial and endosymbiont DNA variants generated in this study will support future basic and translational onchocerciasis research, with particular relevance for ongoing control programmes, and boost efforts to characterize drug, vaccine and diagnostic targets.