In spite of the extensive contribution of intestinal pathology to the pathophysiology of schistosomiasis, little is known of the impact of schistosome infection on the composition of the gut microbiota of its mammalian host. Here, we characterised the fluctuations in the composition of the gut microbial flora of the small and large intestine, as well as the changes in abundance of individual microbial species, of mice experimentally infected with Schistosoma mansoni with the goal of identifying microbial taxa with potential roles in the pathophysiology of infection and disease. Bioinformatic analyses of bacterial 16S rRNA gene data revealed an overall reduction in gut microbial alpha diversity, alongside a significant increase in microbial beta diversity characterised by expanded populations of Akkermansia muciniphila (phylum Verrucomicrobia) and lactobacilli, in the gut microbiota of S. mansoni-infected mice when compared to uninfected control animals. These data support a role of the mammalian gut microbiota in the pathogenesis of hepato-intestinal schistosomiasis and serves as a foundation for the design of mechanistic studies to unravel the complex relationships amongst parasitic helminths, gut microbiota, pathophysiology of infection and host immunity.

Over 250 million people suffer from schistosomiasis, a tropical disease caused by parasitic flatworms known as schistosomes. Humans become infected by free-swimming, water-borne larvae, which penetrate the skin. The earliest intra-mammalian stage, called the schistosomulum, undergoes a series of developmental transitions. These changes are critical for the parasite to adapt to its new environment as it navigates through host tissues to reach its niche, where it will grow to reproductive maturity. Unravelling the mechanisms that drive intra-mammalian development requires knowledge of the spatial organisation and transcriptional dynamics of different cell types that comprise the schistomulum body. To fill these important knowledge gaps, we perform single-cell RNA sequencing on two-day old schistosomula of Schistosoma mansoni. We identify likely gene expression profiles for muscle, nervous system, tegument, oesophageal gland, parenchymal/primordial gut cells, and stem cells. In addition, we validate cell markers for all these clusters by in situ hybridisation in schistosomula and adult parasites. Taken together, this study provides a comprehensive cell-type atlas for the early intra-mammalian stage of this devastating metazoan parasite.

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. This report presents novel findings revealing (a) the genome sequence of the food-borne trematode Fasciola hepatica (the liver fluke) isolated from sheep, which stands out among neglected tropical diseases due to its zoonotic impact on both human and animal health and (b) the first instance (and the genome) of the rickettsial endobacterium of the genus Neorickettsia in F. hepatica. Using stage-specific gene expression data, we identified liver fluke proteins likely involved in host-parasite interactions, and using immunolocalization, we confirmed Neorickettsia in organs and tissues of the adult trematode. The presence of the bacteria in fluke reproductive tissues and eggs suggests a possible mechanism for vertical transmission, and the presence of bacteria in the oral sucker used to anchor flukes to the lining of the biliary tract suggests a potential mechanism for horizontal transmission to the mammalian host. This is of interest because related Neorickettsia cause severe, even deadly, illness in a variety of species, including humans. This is the first report to localize Neorickettsia endobacteria within the tissues of adult F. hepatica. The discoveries in our manuscript have wide impact for the fields of both the pathophysiology and evolution of Fasciola and related FBTs, and the transmission strategies of Neorickettsia.

Schistosomiasis is the most important helminthiasis worldwide in terms of morbidity and mortality. Most of the infections occurs in Africa, which about two thirds are caused by Schistosoma haematobium. The infection with S. haematobium is considered carcinogenic leading to squamous cell carcinoma (SCC) and urothelial carcinoma of the urinary bladder. Additionally, it is responsible for female genital schistosomiasis leading to infertility and higher risk of human immunodeficiency virus (HIV) transmission. Remarkably, a recent outbreak in Corsica (France) drew attention to its potential re-mergence in Southern Europe. Thus far, little is known related to host-parasite interactions that trigger carcinogenesis. However, recent studies have opened new avenues to understand mechanisms on how the parasite infection can lead cancer and other associated pathologies. Here, we present a historical perspective of schistosomiasis, and review the infection-associated pathologies and studies on host–parasite interactions that unveil tentative mechanisms underlying schistosomiasis-associated carcinogenesis.

In spite of growing evidence supporting the occurrence of complex interactions between Schistosoma and gut bacteria in mice and humans, no data is yet available on whether worm-mediated changes in microbiota composition are dependent on the baseline gut microbial profile of the vertebrate host. In addition, the impact of such changes on the susceptibility to, and pathophysiology of, schistosomiasis remains largely unexplored. In this study, mice colonized with gut microbial populations from a human donor (HMA mice), as well as microbiota-wild type (WT) animals, were infected with Schistosoma mansoni, and alterations of their gut microbial profiles at 50 days post-infection were compared to those occurring in uninfected HMA and WT rodents, respectively. Significantly higher worm and egg burdens, together with increased specific antibody responses to parasite antigens, were observed in HMA compared to WT mice. These differences were associated to extensive dissimilarities between the gut microbial profiles of each HMA and WT groups of mice at baseline; in particular, the gut microbiota of HMA animals was characterized by low microbial alpha diversity and expanded Proteobacteria, as well as by the absence of putative immunomodulatory bacteria (e.g. Lactobacillus). Furthermore, differences in infection-associated changes in gut microbiota composition were observed between HMA and WT mice. Altogether, our findings support the hypothesis that susceptibility to S. mansoni infection in mice is partially dependent on the composition of the host baseline microbiota. Moreover, this study highlights the applicability of HMA mouse models to address key biological questions on host-parasite-microbiota relationships in human helminthiases.

Background. At least 250 million people worldwide suffer from schistosomiasis, caused by Schistosoma worms. Genome sequences for several Schistosoma species are available, including a high-quality annotated reference for Schistosoma mansoni. There is a pressing need to develop a reliable functional toolkit to translate these data into new biological insights and targets for intervention. CRISPR-Cas9 was recently demonstrated for the first time in S. mansoni, to produce somatic mutations in the omega-1 (ω1) gene. Methods. We employed CRISPR-Cas9 to introduce somatic mutations in a second gene, SULT-OR, a sulfotransferase expressed in the parasitic stages of S. mansoni, in which mutations confer resistance to the drug oxamniquine. A 262-bp PCR product spanning the region targeted by the gRNA against SULT-OR was amplified, and mutations identified in it by high-throughput sequencing. Results. We found that 0.3-2.0% of aligned reads from CRISPR-Cas9-treated adult worms showed deletions spanning the predicted Cas9 cut site, compared to 0.1-0.2% for sporocysts, while deletions were extremely rare in eggs. The most common deletion observed in adults and sporocysts was a 34 bp-deletion directly upstream of the predicted cut site, but rarer deletions reaching as far as 102 bp upstream of the cut site were also detected. The CRISPR-Cas9-induced deletions, if homozygous, are predicted to cause resistance to oxamniquine by producing frameshifts, ablating SULT-OR transcription, or leading to mRNA degradation via the nonsense-mediated mRNA decay pathway. However, no SULT-OR knock down at the mRNA level was observed, presumably because the cells in which CRISPR-Cas9 did induce mutations represented a small fraction of all cells expressing SULT-OR. Conclusions. Further optimisation of CRISPR-Cas protocols for different developmental stages and particular cell types, including germline cells, will contribute to the generation of a homozygous knock-out in any gene of interest, and in particular the SULT-OR gene to derive an oxamniquine-resistant stable transgenic line.