Investigations of Parasite-Host-Microbiota Interactions Using Cutting-edge Genomics and bioinformatics technologies

Lead Research Organisation: University of Cambridge
Department Name: Veterinary Medicine

Abstract

The increased incidence of allergic and autoimmune disorders in developed countries has been attributed to a decline in childhood exposure to pathogens that causes inappropriate immune responses to harmless stimuli. Thus, several attempts have been made to exploit this 'hygiene hypothesis' by reintroducing some of these infectious agents, particularly intestinal worms, into patients as novel immunotherapy. However, the precise mechanisms by which worms modulate the immune system are unclear. Recent studies have established a link between infections by gastrointestinal parasitic helminths and modifications of the composition of the gut microbiota and relative abundance of selected microbial species, thus prompting the hypothesis that the ability of gastrointestinal parasites to modulate the immune response of the vertebrate hosts may be in part associated to changes in the commensal makeup. However, thus far, data on the interactions between parasitic helminths, vertebrate hosts and their commensal microbiota is lacking. Elucidating the complex relationships between the parasite, the host and the gut microbiota is crucial, because of an increasing need for an integrated approach to understand host---pathogen interactions that, in turn, underpin the translation of experimental findings to the clinic. Recent technological advances provide unique opportunities to investigate various aspects of these interactions. For instance, high---throughput sequencing platforms, together with bioinformatics techniques that allow timely processing of large sequence outputs, enable the identification of host genes and microbial populations and/or species associated with the establishment of a parasite in the gastrointestinal tract of its definitive host.
The specific research aims of this project will be: (1) to determine helminth---induced changes in gut microbiota following experimental infection of vertebrate hosts; (2) to conduct analyses of transcriptional regulation following parasite infection using next---generation sequencing technologies and bioinformatics; (3) to establish the role of the gut microbiota and the relative contribution of helminth---induced modulation of host gene expression in parasite---mediated suppression of inflammation; (4) to select candidate bacterial species for the development of novel helminth---based therapies of gut inflammatory disorders.
This interdisciplinary project will integrate key areas of parasitology, microbiology, immunology, genetics, molecular biology and bioinformatics and will provide essential skills in these areas. This project is timely, since its outcomes will not only have broad implications for future studies of the biology and origin of parasitism itself, but will also pave the way to investigations aimed at exploiting the immunoevasive strategies of parasites for the development of helminth---based therapeutics for chronic inflammatory disorders.

Publications

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Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/M011194/1 01/10/2015 31/03/2024
1643688 Studentship BB/M011194/1 01/10/2015 30/09/2019 Timothy Jenkins
 
Description Understanding the impact that parasitic helminths exert on the host gut microbiota is central to a better grasp of host-parasite interactions and the fundamental molecular mechanisms that govern essential biological processes and, ultimately, could assist in identifying novel treatment approaches against parasitic helminths, as well as helminth-borne
treatments of allergic and autoimmune conditions.
The present thesis (i) investigated the consequences of natural multi- or mono-species infections by helminth parasites on the composition of the human gut microbiota (Chapters 2 and 3), (ii) elucidated the longitudinal impact of experimentally controlled mono-species helminth infections on the human gut microbiota (Chapter 4), and, finally, (iii) examined what impact an extra-intestinal (EI) helminth infection has on the host microbiome in a murine model of human schistosomiasis (Chapter 5). The objectives of the present chapter were (a) to summarise the fundamental research achievements, (b) to discuss the findings and implications that can be drawn from this research in relation to host-parasite interactions, and (c) to provide an outlook on opportunities and prospects for future investigations.
Overall, this thesis has described the results from bioinformatic analyses of ~43,305, 300 paired-end reads generated via 16S rRNA sequencing of human faeces (Chapters 2-5). The majority of the raw reads (Chapters 2, 3, and 5) have been deposited and made publicly available on public databases (i.e., Mendeley data and the European Nucleotide Archive) and present a substantial resource for future investigations within the field of parasitology, but also for broader investigations of the gut microbiota.
Knowledge of helminths' impact on host microbiota is pivotal to untangle the complex network of host-parasite interactions. However, until recently, such investigations had to rely on culture-based techniques for the profiling of gut microbial changes 1. This only allowed highly limited and biased investigations into such microbiota shifts. However, recent innovations in next generation sequencing and 'omics' technologies have made quantitative and less or non-biased approaches, such as 16S rRNA sequencing, affordable and thus widely accessible 2. These technological advancements have vastly improved our ability to investigate gut microbiota changes and gain insights into microbial ecosystem interactions within the host gut 3-5. At the commencement of this thesis, little data existed on the impact of parasitic helminth infections on the host microbiome. Indeed, only four studies had been published on the impact of parasitic helminths infections on the human gut microbiota at that time 6-9 and, while they established that such infections could significantly alter the host microbiota, few clear trends could be identified consistently across these studies. This lack of consistency could largely be traced back to the variability in study designs, helminth species, and data analyses techniques, amongst others, between those studies 10. Although a complete elimination of confounding factors is improbable in investigations of human helminth infections, it is possible to minimise and carefully account for such influences 10. Hence, in this thesis consistent DNA extraction, sequencing library preparation, and data analysis techniques have been applied to a range of human-helminth infection scenarios, as well as a murine model of human helminth infection. The aim was to apply technical consistency to draw more confident inferences from the data, while also allowing the detection of helminth induced host microbiota changes, which are consistent across different studies. Indeed, though many of the microbiota changes observed across the studies presented in this thesis appeared to be specific to the host-helminth system that was being investigated, some intriguing consistencies emerged. Firstly, low level, long term, and single species subclinical infections were associated with increased gut microbial diversity within the host and seemed to promote a stable and healthy gut microbial composition (Chapters 3 and 4). Notably, these findings are supported by data from other studies that examined the effects of experimental single species (Necator americanus) infections in coeliac patients and reported increased gut microbial diversity following helminth administration 7,11. Contrarily, acute heavy burden infections, associated with pathology, appeared to have the opposite effect, i.e. reducing the overall diversity of the host's gut microbiota and associated with the presence and proliferation of potentially pro-inflammatory bacteria and/or opportunistic pathogens (Chapter 5). Indeed, heavy parasite burden infections are common practice in most studies investigating rodent models of helminth infections and have been frequently reported to lead to significant decreases in gut microbiota diversity 12-14. Meanwhile, I was unable to detect a significant difference in alpha diversity in subjects naturally infected with different species of STHs (Chapter 1). Other studies of natural and/or mixed infections have been lacking a consistency in trends of host gut microbial diversity, with some reporting increased 8,15 and decreased 9 levels of diversity, or no change at all 16,17. This is likely linked to the significant differences in the study cohorts and their geographic locations, as well as the varied types of infections and species of parasite involved (reviewed by 10). Notably, I also found that murine infections with parasitic (EI) helminths can have a significant impact on the host microbiota, even before eggs traverse through the host gut epithelial layer (Chapter 5). Though the gut microbiota perturbations significantly increased upon egg laying and consequent disruption of the gut epithelial layer, these data demonstrated that host microbiome changes can be indirectly induced by EI helminths, likely due to the parasites' strong immunomodulatory properties (reviewed by 18,19).
Together, these data suggest that both GI and EI parasitic helminth infections have the potential to detrimentally impact the hosts they infect, besides the direct pathology they induce, but also adds further weight to the idea of a therapeutic and controlled use of helminths in the context of helminth therapy. Indeed, considering the beneficial effects parasitic helminths may have on the host gut microbiota, together with the mounting evidence towards an intrinsic link between autoimmune diseases and the gut microbiome, infection-associated changes on the microbial composition of the host gut might represent an additional route via which helminths could exert a therapeutic effect on patients suffering from such conditions 6,7,11, in addition to the release of ESPs with immunomodulatory properties (reviewed by 20).
Besides the trends in gut microbial diversity, further patterns emerged when assessing the changes in specific bacterial taxa during helminth infections. In this thesis, I found that study cohorts characterised by increased levels of gut microbial diversity and which appeared to have an overall healthier microbiome than the cohort they were compared to, presented an increased relative abundance of the bacterial family Leuconostocaceae (uninfected cohort Chapter 2 and infected cohort Chapter 3) and the genus Turicibacter (uninfected cohort Chapter 5 and infected cohort Chapter 3), with the phylum/class Tenericutes/Mollicutes (infected Chapter 4) also proving interesting, due to the plethora of evidence from other studies indicating they could present a key taxon involved in disease progression of autoimmune conditions 21-25. On the other hand, a decrease in gut microbial alpha diversity appeared to be associated with an increased relative abundance of the bacterial family Enterobacteriaceae (uninfected cohort Chapter 3 and infected cohort Chapter 2), the genus Akkermansia (infected cohort Chapter 2 and infected cohort Chapter 5), and the family Lachnospiraceae, particularly the bacterial genus Dorea (uninfected cohort Chapter 4, infected cohort Chapter 5).


References
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Chapters refer to the PhD Thesis: Exploring the impact of gastrointestinal parasitic helminths on the human microbiome using advanced biomolecular and bioinformatics technologies
Exploitation Route Regrettably, within my thesis the relevance of the identified bacteria could merely be inferred from evidence present in the literature and the investigation of the functional importance of these taxa fell outside the scope of this thesis. However, the bacteria identified here present excellent targets for future investigations aiming at untangling the gut microbial nuances underlying host-helminth interactions. Furthermore, these bacteria are likely not just important in the context of parasitic helminth infections, but also fall within the broader scope of human microbiome health 26 and, thus, could be of interest to studies focusing on specific bacteria involved in gut microbiome stability and gut epithelial health, as well as investigating gut inflammation.

Reference
26 Lloyd-Price, J., Abu-Ali, G. & Huttenhower, C. The healthy human microbiome. Genome Med 8, 51, doi:10.1186/s13073-016-0307-y (2016).
Sectors Healthcare,Pharmaceuticals and Medical Biotechnology