The Enterocytozoonidae: the emergence of a microsporidian clade into the aquatic and terrestrial food-chain

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Microsporidia are a group of over 1000 described parasitic microbes with species infecting all major animal lineages in all environments. One particularly important family of microsporidia are the Enterocytozoonidae. This is an intriguing group with members that infect hosts from both aquatic and terrestrial habitats. It includes two high profile members: Enterocytozoon bieneusi, which is one of the most common causes of AIDS-associated diarrhoea and the tiger shrimp pathogen Enterocytozoon hepatopenaei. The latter is associated with severely retarded shrimp growth, which results in considerable economic losses for millions of people in the aquaculture industry. Additionally, E. hepatopenaei has recently been shown to have jumped host causing severe disease in the edible Whiteleg Shrimp. The Enterocytozoonidae, and particularly E. hepatopenaei, given its emerging economic importance, need further study and this studentship aims to use environmental DNA to address the following two questions in Enterocytozoonidae biology:The first aim of the studentship is to answer the question of how E. hepatopenaei has spread across shrimp farms in Asia. Though initially reported from Thailand, E. hepatopenaei has also been found in China, Indonesia, Malaysia, Vietnam and India. This transmission may be mediated by movement of infected shrimp from farm to farm, or the infection may be acquired from intermediate hosts and/or contaminated feed. This project proposes to use molecular data to determine how transmission has occurred and therefore inform aquaculture policies to prevent further spread. Recent collaborative work between the proposed project partners has generated genome sequence data for E. hepatopenaei which is currently being used to design better diagnostic PCR primers to understand the prevalence of the pathogen. This project proposes to expand this work by sequencing genomes of E. hepatopenaei collected from multiple geographic locations in Thailand, from China and from both Whiteleg and Tiger shrimp hosts. The generated molecular data will be used to reconstruct evolutionary relationships between the strains and infer movement between geographic localities and hosts. The second aim addresses the question of how many undiscovered Enterocytozoonidae parasites are present in the environment. Humans and shrimps are two important hosts for the Enterocytozoonidae and both may be acquiring these infections from, as yet, unidentified intermediate hosts. This component of the project aims to identify reservoirs or carriers for important Enterocytozoonidae and further species that may have the capacity to cause human or animal disease. We will collect invertebrate samples from environments at the interface of human and aquatic animal populations, for example, domestic sewage, riverine, estuarine and coastal habitats and from shrimp ponds in Thailand. We will then extract the DNA from all these organisms along with that of their associated parasites and use state of the art gene sequencing technology to sequence the same gene for each parasite in the sample. By studying the diversity of the gene sequences we can identify the microsporidian parasites present in the original sample. When we detect our pathogens of interest (E. bieneusi and E. hepatopenaei) or close relatives of these, we will label the amplified DNA sequences and use these as probes to determine the identity of intermediate hosts.The proposed work will be supervised as a collaboration between: Bryony Williams (University of Exeter) who will provide will provide equipment, training and expertise for genomics and molecular work; CASE partner Grant Stentiford (CEFAS) who will provide training in sample collection, fieldwork and identification of intermediate hosts; Kallaya Dangtip and Ornchuma Itsathiphaisarn (Center of Excellence for Shrimp Molecular Biology and Biotechnology, Mahidol University, Thailand) who will provide shrimp and shrimp pond samples.