The Human Blood Fluke, Schistosoma mansoni, Harbors Bacteria Throughout the Parasite's Life Cycle.

While symbiotic relationships between invertebrates and bacteria have been extensively described, studies of microbial communities inhabiting parasitic worms remain scarce. Exploring the microbiota associated with helminths responsible for major infectious diseases will inform on parasite biology, host-pathogen interactions, and disease pathophysiology. We investigated the presence of microorganisms inhabiting tissues of the human parasite Schistosoma mansoni. In situ hybridization using a pan-bacterial 16S rRNA gene probe revealed bacteria colonizing key developmental stages that were successfully removed after antibiotic treatment of live parasites. Understanding the composition and function of the S. mansoni-associated microbiota may lead to the development of novel microbiome-targeting control strategies.

The role of the host gut microbiome in the pathophysiology of schistosomiasis.

The pathophysiology of schistosomiasis is linked to the formation of fibrous granulomas around eggs that become trapped in host tissues, particularly the intestines and liver, during their migration to reach the lumen of the vertebrate gut. While the development of Schistosoma egg-induced granulomas is the result of finely regulated crosstalk between egg-secreted antigens and host immunity, evidence has started to emerge of the likely contribution of an additional player-the host gut microbiota-to pathological processes that culminate with the formation of these tissue lesions. Uncovering the role(s) of schistosome-mediated changes in gut microbiome composition and function in granuloma formation and, more broadly, in the pathophysiology of schistosomiasis, will shed light on the mechanisms underlying this three-way parasite-host-microbiome interplay. Such knowledge may, in turn, pave the way towards the discovery of novel therapeutic targets and control strategies.

Gastrointestinal worms and bacteria: From association to intervention.

A plethora of studies, both experimental and epidemiological, have indicated the occurrence of associations between infections by gastrointestinal (GI) helminths and the composition and function of the host gut microbiota. Given the worldwide risk and spread of anthelmintic resistance, particularly for GI parasites of livestock, a better understanding of the mechanisms underpinning the relationships between GI helminths and the gut microbiome, and between the latter and host health, may assist the development of novel microbiome-targeting and other bacteria-based strategies for parasite control. In this article, we review current and prospective methods to manipulate the host gut microbiome, and/or to exploit the immune stimulatory and modulatory properties of gut bacteria (and their products) to counteract the negative impact of GI worm infections; we also discuss the potential applications of these intervention strategies in programmes aimed to aid the fight against helminth diseases of humans and livestock.

The gut microbial metabolic capacity of microbiome-humanized vs. wild type rodents reveals a likely dual role of intestinal bacteria in hepato-intestinal schistosomiasis.

Increasing evidence shows that the host gut microbiota might be involved in the immunological cascade that culminates with the formation of tissue granulomas underlying the pathophysiology of hepato-intestinal schistosomiasis. In this study, we investigated the impact of Schistosoma mansoni infection on the gut microbial composition and functional potential of both wild type and microbiome-humanized mice. In spite of substantial differences in microbiome composition at baseline, selected pathways were consistently affected by parasite infection. The gut microbiomes of infected mice of both lines displayed, amongst other features, enhanced capacity for tryptophan and butyrate production, which might be linked to the activation of mechanisms aimed to prevent excessive injuries caused by migrating parasite eggs. Complementing data from previous studies, our findings suggest that the host gut microbiome might play a dual role in the pathophysiology of schistosomiasis, where intestinal bacteria may contribute to egg-associated pathology while, in turn, protect the host from uncontrolled tissue damage.

Parasitic helminths and the host microbiome - a missing 'extracellular vesicle-sized' link?

Infections by gastrointestinal (GI) helminths have been associated with significant alterations of the structure of microbial communities inhabiting the host gut. However, current understanding of the biological mechanisms that regulate these relationships is still lacking. We propose that helminth-derived extracellular vesicles (EVs) likely represent key players in helminth-microbiota crosstalk. Here, we explore knowledge of helminth EVs with an emphasis on their putative antimicrobial properties, and we argue that (i) an enhanced understanding of the mechanisms governing such interactions might assist the discovery and development of novel strategies of parasite control, and that (ii) the identification and characterisation of helminth molecules with antimicrobial properties might pave the way towards the discovery of novel antibiotics, thus aiding the global fight against antimicrobial resistance.

Helminth Microbiota Profiling Using Bacterial 16S rRNA Gene Amplicon Sequencing: From Sampling to Sequence Data Mining.

Symbiont microbial communities play important roles in animal biology and are thus considered integral components of metazoan organisms, including parasitic worms (helminths). Nevertheless, the study of helminth microbiomes has thus far been largely overlooked, and symbiotic relationships between helminths and their microbiomes have been only investigated in selected parasitic worms. Over the past decade, advances in next-generation sequencing technologies, coupled with their increased affordability, have spurred investigations of helminth-associated microbial communities aiming at enhancing current understanding of their fundamental biology and physiology, as well as of host-microbe interactions. Using the blood fluke Schistosoma mansoni as a key example of parasitic worms with complex life cycles involving multiple hosts, in this chapter we (1) provide an overview of protocols for sample collection and (2) outline an example workflow to characterize worm-associated microbial communities using high-throughput sequencing technologies and bioinformatics analyses of large-scale sequence data.

Gut-microbiota-derived extracellular vesicles: Overlooked mediators in host-helminth interactions?

Helminth infections impact the composition of the mammalian gut microbiota; however, the mechanisms underpinning these interactions are, thus far, unknown. In this article, we propose that microbiota-derived extracellular vesicles might represent key players in host-helminth-microbiome crosstalk, and outline future directions to elucidate their role(s) in host-parasite relationships.

Experimental infection with the hookworm, Necator americanus, is associated with stable gut microbial diversity in human volunteers with relapsing multiple sclerosis.

BACKGROUND: Helminth-associated changes in gut microbiota composition have been hypothesised to contribute to the immune-suppressive properties of parasitic worms. Multiple sclerosis is an immune-mediated autoimmune disease of the central nervous system whose pathophysiology has been linked to imbalances in gut microbial communities. RESULTS: In the present study, we investigated, for the first time, qualitative and quantitative changes in the faecal bacterial composition of human volunteers with remitting multiple sclerosis (RMS) prior to and following experimental infection with the human hookworm, Necator americanus (N+), and following anthelmintic treatment, and compared the findings with data obtained from a cohort of RMS patients subjected to placebo treatment (PBO). Bacterial 16S rRNA high-throughput sequencing data revealed significantly decreased alpha diversity in the faecal microbiota of PBO compared to N+ subjects over the course of the trial; additionally, we observed significant differences in the abundances of several bacterial taxa with putative immune-modulatory functions between study cohorts. Parabacteroides were significantly expanded in the faecal microbiota of N+ individuals for which no clinical and/or radiological relapses were recorded at the end of the trial. CONCLUSIONS: Overall, our data lend support to the hypothesis of a contributory role of parasite-associated alterations in gut microbial composition to the immune-modulatory properties of hookworm parasites.

Vaccination against the brown stomach worm, Teladorsagia circumcincta, followed by parasite challenge, induces inconsistent modifications in gut microbiota composition of lambs.

BACKGROUND: Growing evidence points towards a role of gastrointestinal (GI) helminth parasites of ruminants in modifying the composition of the host gut flora, with likely repercussions on the pathophysiology of worm infection and disease, and on animal growth and productivity. However, a thorough understanding of the mechanisms governing helminth-microbiota interactions and of their impact on host health and welfare relies on reproducibility and replicability of findings. To this aim, in this study, we analysed quantitative and qualitative fluctuations in the faecal microbiota composition of lambs vaccinated against, and experimentally infected with, the parasitic GI nematode Teladorsagia circumcincta over the course of two separate trials performed over two consecutive years. METHODS: Two trials were conducted under similar experimental conditions in 2017 and 2018, respectively. In each trial, lambs were randomly assigned to one of the following experimental groups: (i) vaccinated/infected, (ii) unvaccinated/infected and (iii) unvaccinated/uninfected. Faecal samples collected from individual animals were subjected to DNA extraction followed by high-throughput sequencing of the V3-V4 region of the bacterial 16S rRNA gene and bioinformatics and biostatistical analyses of sequence data. RESULTS: Substantial differences in the populations of bacteria affected by immunisation against and infection by T. circumcincta were detected when comparing data from the two trials. Nevertheless, the abundance of Prevotella spp. was significantly linked to helminth infection in both trials. CONCLUSIONS: Despite the largely conflicting findings between the two trials, our data revealed that selected gut microbial populations are consistently affected by T. circumcincta infection and/or vaccination. Nevertheless, our study calls for caution when interpreting data generated from in vivo helminth-microbiome interaction studies that may be influenced by several intrinsic and extrinsic host-, parasite- and environment-related factors.

Baseline Gut Microbiota Composition Is Associated With Schistosoma mansoni Infection Burden in Rodent Models.

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.

Helminths, hosts, and their microbiota: new avenues for managing gastrointestinal helminthiases in ruminants.

INTRODUCTION: Evidence is emerging of complex interactions occurring between gastrointestinal (GI) parasites of ruminants and the resident gut flora, with likely implications for the pathophysiology of worm infection and disease. Similarly, recent data point toward the occurrence of a GI nematode (GIN)-specific microbiota, with potential roles in worm fundamental physiology and reproduction. Parasite-microbiota relationships might represent potential targets for the development of novel parasiticides. AREAS COVERED: In this article, we review current knowledge of the role(s) that host- and helminth-associated microbiota play in ruminant host-parasite relationships, and outline potential avenues for the control of GIN of farmed ruminants via the manipulation of resident microbial species with putative functions in infection establishment, host-immune modulation, and/or parasite fitness and survival. EXPERT OPINION: In order for this knowledge to be translated into practical applications, we argue that several aspects of the nematode-microbiota cross-talk must be addressed, including (i) the causality of interactions between the parasite, the gut microbiota, and the host immune system, (ii) the modes of action of dietary prebiotics and probiotics, (iii) the mechanisms by which diet supplementation aids the development of resistance/tolerance to GI helminth infections and (iv) the composition of the GIN microbiome and its role(s) in parasite biology and physiology.

Infection with the sheep gastrointestinal nematode Teladorsagia circumcincta increases luminal pathobionts.

BACKGROUND: The multifaceted interactions between gastrointestinal (GI) helminth parasites, host gut microbiota and immune system are emerging as a key area of research within the field of host-parasite relationships. In spite of the plethora of data available on the impact that GI helminths exert on the composition of the gut microflora, whether alterations of microbial profiles are caused by direct parasite-bacteria interactions or, indirectly, by alterations of the GI environment (e.g. mucosal immunity) remains to be determined. Furthermore, no data is thus far available on the downstream roles that qualitative and quantitative changes in gut microbial composition play in the overall pathophysiology of parasite infection and disease. RESULTS: In this study, we investigated the fluctuations in microbiota composition and local immune microenvironment of sheep vaccinated against, and experimentally infected with, the 'brown stomach worm' Teladorsagia circumcincta, a parasite of worldwide socio-economic significance. We compared the faecal microbial profiles of vaccinated and subsequently infected sheep with those obtained from groups of unvaccinated/infected and unvaccinated/uninfected animals. We show that alterations of gut microbial composition are associated mainly with parasite infection, and that this involves the expansion of populations of bacteria with known pro-inflammatory properties that may contribute to the immunopathology of helminth disease. Using novel quantitative approaches for the analysis of confocal microscopy-derived images, we also show that gastric tissue infiltration of T cells is driven by parasitic infection rather than anti-helminth vaccination. CONCLUSIONS: Teladorsagia circumcincta infection leads to an expansion of potentially pro-inflammatory gut microbial species and abomasal T cells. This data paves the way for future experiments aimed to determine the contribution of the gut flora to the pathophysiology of parasitic disease, with the ultimate aim to design and develop novel treatment/control strategies focused on preventing and/or restricting bacterial-mediated inflammation upon infection by GI helminths. Video Abstract.

MICHELINdb: a web-based tool for mining of helminth-microbiota interaction datasets, and a meta-analysis of current research.

BACKGROUND: The complex network of interactions occurring between gastrointestinal (GI) and extra-intestinal (EI) parasitic helminths of humans and animals and the resident gut microbial flora is attracting increasing attention from biomedical researchers, because of the likely implications for the pathophysiology of helminth infection and disease. Nevertheless, the vast heterogeneity of study designs and microbial community profiling strategies, and of bioinformatic and biostatistical approaches for analyses of metagenomic sequence datasets hinder the identification of bacterial targets for follow-up experimental investigations of helminth-microbiota cross-talk. Furthermore, comparative analyses of published datasets are made difficult by the unavailability of a unique repository for metagenomic sequence data and associated metadata linked to studies aimed to explore potential changes in the composition of the vertebrate gut microbiota in response to GI and/or EI helminth infections. RESULTS: Here, we undertake a meta-analysis of available metagenomic sequence data linked to published studies on helminth-microbiota cross-talk in humans and veterinary species using a single bioinformatic pipeline, and introduce the 'MICrobiome HELminth INteractions database' (MICHELINdb), an online resource for mining of published sequence datasets, and corresponding metadata, generated in these investigations. CONCLUSIONS: By increasing data accessibility, we aim to provide the scientific community with a platform to identify gut microbial populations with potential roles in the pathophysiology of helminth disease and parasite-mediated suppression of host inflammatory responses, and facilitate the design of experiments aimed to disentangle the cause(s) and effect(s) of helminth-microbiota relationships. Video abstract.

Secreted cathepsin L-like peptidases are involved in the degradation of trapped antibodies on the surface of Echinostoma caproni.

Antibody trapping is a recently described strategy for immune evasion observed in the intestinal trematode Echinostoma caproni, which may aid to avoiding the host humoral response, thus facilitating parasite survival in the presence of high levels of local-specific antibodies. Parasite-derived peptidases carry out the degradation of trapped antibodies, being essential for this mechanism. Herein, we show that cathepsin-like cysteine endopeptidases are active in the excretory/secretory products (ESPs) of E. caproni and play an important role in the context of antibody trapping. Cysteine endopeptidase activity was detected in the ESPs of E. caproni adults. The affinity probe DCG-04 distinguished a cysteine peptidase band in ESPs, which was specifically recognized by an anti-cathepsin L heterologous antibody. The same antibody localized this protein in the gut and syncytial tegument of adult worms. Studies with cultured parasites showed that in vivo-bound antibodies are removed from the parasite surface in the absence of peptidase inhibitors, while addition of cathepsin L inhibitor prevented their degradation. These results indicate that cathepsin L-like peptidases are involved in the degradation of surface-trapped antibodies and suggest that cysteine peptidases are not only crucial for tissue-invading trematodes, but they can be equally relevant at the parasite-host interface in gut-dwelling flukes.

Helminth-microbiota cross-talk - A journey through the vertebrate digestive system.

The gastrointestinal (GI) tract of vertebrates is inhabited by a vast array of organisms, i.e., the microbiota and macrobiota. The former is composed largely of commensal microorganisms, which play vital roles in host nutrition and maintenance of energy balance, in addition to supporting the development and function of the vertebrate immune system. By contrast, the macrobiota includes parasitic helminths, which are mostly considered detrimental to host health via a range of pathogenic effects that depend on parasite size, location in the GI tract, burden of infection, metabolic activity, and interactions with the host immune system. Sharing the same environment within the vertebrate host, the GI microbiota and parasitic helminths interact with each other, and the results of such interactions may impact, directly or indirectly, on host health and homeostasis. The complex relationships occurring between parasitic helminths and microbiota have long been neglected; however, recent studies point towards a role for these interactions in the overall pathophysiology of helminth disease, as well as in parasite-mediated suppression of inflammation. Whilst several discrepancies in qualitative and quantitative modifications in gut microbiota composition have been described based on host and helminth species under investigation, we argue that attention should be paid to the systems biology of the gut compartment under consideration, as variations in the abundances of the same population of bacteria inhabiting different niches of the GI tract may result in varying functional consequences for host physiology.

Helminths and microbes within the vertebrate gut - not all studies are created equal.

The multifaceted interactions occurring between gastrointestinal (GI) parasitic helminths and the host gut microbiota are emerging as a key area of study within the broader research domain of host-pathogen relationships. Over the past few years, a wealth of investigations has demonstrated that GI helminths interact with the host gut flora, and that such interactions result in modifications of the host immune and metabolic statuses. Nevertheless, whilst selected changes in gut microbial composition are consistently observed in response to GI helminth infections across several host-parasite systems, research in this area to date is largely characterised by inconsistent findings. These discrepancies are particularly evident when data from studies of GI helminth-microbiota interactions conducted in humans from parasite-endemic regions are compared. In this review, we provide an overview of the main sources of variance that affect investigations on helminth-gut microbiota interactions in humans, and propose a series of methodological approaches that, whilst accounting for the inevitable constraints of fieldwork, are aimed at minimising confounding factors and draw biologically meaningful interpretations from highly variable datasets.

RORγt+ Treg to Th17 ratios correlate with susceptibility to Giardia infection.

Infections with Giardia are among the most common causes of food and water-borne diarrheal disease worldwide. Here, we investigated Th17, Treg and IgA responses, and alterations in gut microbiota in two mouse lines with varying susceptibility to Giardia muris infection. Infected BALB/c mice shed significantly more cysts compared with C57BL/6 mice. Impaired control of infection in BALB/c mice was associated with lower Th17 activity and lower IgA levels compared with C57BL/6 mice. The limited metabolic activity, proliferation and cytokine production of Th17 cells in BALB/c mice was associated with higher proportions of intestinal Foxp3+RORγt+ regulatory T cells and BALB/c mice developed increased RORγt+ Treg:Th17 ratios in response to G. muris infection. Furthermore, G. muris colonization led to a significantly reduced evenness in the gut microbial communities of BALB/c mice. Our data indicate that differential susceptibility to Giardia infections may be related to RORγt+ Treg controlling Th17 activity and that changes in the microbiota composition upon Giardia infection partially depend on the host background.

Classic Models for New Perspectives: Delving into Helminth-Microbiota-Immune System Interactions.

Whilst a wealth of data indicate that infections by gastrointestinal helminths are accompanied by significant alterations in the composition of the vertebrate gut flora, little is known of the immune-molecular mechanisms that regulate host-parasite-microbiota interactions. 'Traditional' experimental models of gastrointestinal helminthiases, in which the role(s) of each of the components of this triad can be tested, provide an opportunity to advance research in this area. In this article, we propose the Echinostoma caproni-mouse system as a potentially useful tool for studies of the role of the host gut microbiota in preventing pathology and inducing parasite clearance via interleukin (IL)-25, an epithelial-derived alarmin with key roles in antihelminth immunity and maintenance of gut homeostasis.

Adaptation of the secretome of Echinostoma caproni may contribute to parasite survival in a Th1 milieu.

Echinostoma caproni (Trematoda: Echinostomatidae) is an intestinal trematode, broadly employed to study the host-dependent mechanisms that govern the evolution of intestinal helminth infections. Resistance against E. caproni homologous secondary infections has been reported in mice and appears to be related to the generation of a local Th2 response, whereas Th1 responses promote the development of chronic primary infections. Herein, the ability of E. caproni to modulate its secretome according to the host environment is investigated. A two-dimensional differential in gel electrophoresis (2D-DIGE) analysis was performed to elucidate changes in the excretory/secretory products of E. caproni adults after primary and secondary infections in mice. A total of 16 protein spots showed significant differences between groups, and 7 of them were successfully identified by mass spectrometry. Adult worms exposed to a primary infection appear to upregulate proteins involved in detoxification (aldo-keto reductase), stress response (GroEL), and enhancement of parasite survival (acetyl-CoA A-acetyltransferase and UTP-glucose-1-phosphate urydyltransferase). In contrast, any protein was found to be significantly upregulated after secondary infection. Upregulation of such proteins may serve to withstand the hostile Th1 environment generated in primary infections in mice. These results provide new insights into the resistance mechanisms developed by the parasites to ensure their long-term survival.