During the first stage of my project, I focused on the identification novel genes of both the host and the parasite potentially involved on the interplay of whipworms with both gut epithelium and immune cells. To do this, I reviewed the literature and data on susceptibility to intestinal inflammation challenges of existing mutant mouse lines. In addition, I performed transcriptomic and imaging experiments studying the very first events after whipworm infection of mice (3 hours, one and 3 days). These studies revealed early interactions of the larvae with goblet cells, a specific epithelial cell type that produces mucus. They also revealed the expression of whipworm proteases and protease inhibitors that are potentially required for mucus degradation, suggesting the interaction with goblet cells is critical for the invasion and colonization of the gut.
In the second stage of the fellowship, new mutant mouse lines were generated for novel host genes identified as part of work by the Wellcome Trust-funded Infection and Immunity Immunophenotyping (3i) Consortium. I evaluated the role of these host genes on the development of immune responses to whipworms and their expulsion by infecting 300 mutant mouse lines with a high dose of T. muris. Through this screen, I identified 10 genes conferring either enhanced resistance or susceptibility to whipworms, namely Adal, Fam160a1, Klk5, Wac, Irak1, IL-27, Arpc1b, and the members of the IL-10 receptor family, Il-10, Il-10ra and Il-10rb.
Through the last phase of my project, I focused on experiments to deeply investigate the role on immunity to whipworm infections of different members of the IL-10 receptor family. Specifically, I found that IL-10 signaling, but not the related cytokines IL-22 or IL-28, promotes intestinal colonization resistance against opportunistic pathogens and controls immunopathology during whipworm infection. In addition, to further understand the early interactions of whipworms with the gut epithelium, I am: 1) attempting to generate mutant whipworms to study the function of parasite genes during early infection; 2) doing proteomics and mucus degradation experiments with whipworm larvae; and 3) performing gene expression analysis of single gut epithelial cells to investigate the initiation of the host responses to whipworm infection. Finally, as an alternative model and to replace the use of mice for these studies, I developed a novel in vitro model using mouse and human miniature (mini)-guts and inject them with mouse and human whipworm larvae. Mini-guts are 3D cell clusters generated from gut tissue that have similar characteristics and function to the gut. Using this new model, I am currently using microscopy and transcriptomic technologies to better understand the processes of invasion and establishment of the larvae inside the epithelium.
To disseminate the results of this research project, during the length of this fellowship, I presented my findings on international scientific conferences and internal seminars and retreats at the Sanger Institute and the 3i consortium, resulting in both a poster and a presentation prize. Moreover, all sequencing data obtained in this study has been made accessible prior to publication through the European Nucleotide Archive (www.ebi.ac.uk/ena/) and WormBase Parasite (parasite.wormbase.org/). The results of the challenge of mutant mouse strains are also publically available at www.immunophenotype.org. In addition, mutant mouse lines produced during this project are available upon request (www.sanger.ac.uk/mouseportal/). Finally, I am currently preparing manuscripts describing the main findings of this project, which will soon be submitted to peer-reviewed journals.
