CEFAS: A pathway to mitigating infection and disease caused by emergent Enterocytozoon hepatopenaei (Microsporidia) in the whiteleg shrimp (Penaeus vannamei)
The University of Exeter’s College of Life and Environmental Sciences, in partnership with the Centre for Environment, Fisheries and Aquaculture Science (Cefas), are inviting applications for a fully-funded PhD studentship to commence in September 2016 (or as soon as possible thereafter). For eligible students the studentship will cover UK/EU tuition fees plus an annual tax-free stipend of £14,198 for 3.5 years. The student would be based in Biosciences in the College of Life and Environmental Sciences at the Streatham Campus in Exeter.
Dr Bryony Williams, University of Exeter
Dr Grant Stentiford, Cefas
Dr Ornchuma Itsathitphaisarn, Mahidol University, Thailand
The Microsporidia are a diverse parasite phylum infecting all major animal lineages in all environments. Parasites of economically important taxa (such as shrimp) are close relatives of those infecting humans, suggesting that most human infections are in fact zoonotic, obtained via interaction with water and the food chain (1). Recently, a high profile example has emerged in Asian shrimp farming. Enterocytozoon hepatopenaei (EHP) was first detected in 2009 as a low-prevalence/low intensity infection of Thai tiger shrimp, and described by researchers at Cefas and Mahidol University (2). It is the closest relative to the human pathogen Enterocytozoon bienuesi infecting human patients with HIV/AIDS. It has since emerged as a high prevalence/high intensity disease in whiteleg shrimp farmed in China, Indonesia, Malaysia, Vietnam and India. It is associated with severe growth retardation and mortality. This pathogen has emerged very rapidly, is associated with the well publicised condition termed ‘Early Mortality Syndrome’ (EMS), and is set to cause multi hundred million-dollar losses in Asian shrimp production. Infections are now at a critical level with some farms reporting 100% prevalence in shrimp ponds. Given its emerging importance and potential for spread, EHP fulfils many of the criteria for listing by the OIE and other regional bodies responsible for controlling spread of emergent aquatic pathogens (3) and urgent study is required to devise strategies to mitigate its effects on the shrimp industry.
Identifying host-parasite interactions: The student will exploit existing genomic resources for Enterocytozoon hepatopenaei to better understand how the pathogen interacts with its shrimp host. The student will identify the EHP putative secretome using bioinformatics and screen this for potential effector families and for proteins that are predicted to be localised to the surface of the cell/spore. These putative ‘host interactors’ will then be cloned and expressed and used as bait to pull down their targets from host tissues, with the aim of identifying interaction targets in the host during infection. These shrimp proteins will be identified using protein mass spectrometry using the publically available Litopenaeus vannamei (White leg shrimp) EST database as a reference. These interactions will be confirmed using a yeast II hybrid assay and other proteomic approaches such as co-immmunoprecipitation.
Identifying potential naturally resistant mutants: After identifying the interaction partners in the host and pathogen, multiple Litopenaeus vannamei (infected and uninfected) isolates will be PCR screened for diversity at these genomic sites. Here we will look for evidence of natural variation in these genes and explore whether changes at these sites are imparting a natural resistance to microsporidian infection.
Testing host receptor mutations using CRISPR-CAS9: In the future it may become more common to design and consume genetically modified seafood and the generation of GM disease resistant shrimp may soon become desirable. The work above generates essential information about the types of targets that could be knocked down in order to generate EHP resistant white-leg shrimp. One technology that may be used for this purpose is the CRISPR-CAS9 system. The whiteleg shrimp produces zygotes by external fertilisation and these represent a potential target for transformation with the CRISPR–CAS9 constructs. The student will work with collaborators in Thailand at CENTEX Shrimp/BIOTEC to generate CRISPR-CAS9 modified shrimps with knock-down of proteins involved in microsporidian interactions with the aim of setting up infection assays with EHP.
This award provides annual funding to cover UK/EU tuition fees and a tax-free stipend. For students who qualify to pay UK/EU tuition fees, the award will cover the tuition fees in full plus a tax-free stipend of £14,198 per year for 3.5 years.
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