Bochow, Shaun (2016) Characterisation of Cherax quadricarinatus densovirus: the first virus characterised from Australian freshwater crayfish. PhD thesis, James Cook University.

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Abstract

Early disease investigations in Cherax spp. were observational in nature. Investigators found a variety of pathogens including fungi, ciliates, platyhelminthes, nematodes and viruses. To date, the viral flora identified have been studied using histopathology and transmission electron microscopy. There have been two transmission trials carried out using Cherax spp. infecting viruses, a Reo-like virus and a parvovirus-like virus. Despite multiple detections of viruses in Cherax spp., there has been no molecular characterisation of any of these pathogens. Of the various viral flora identified in Cherax spp., four were classified as parvovirus-like. These include Cherax destructor systemic parvo-like virus, Cherax quadricarinatus gill parvo-like virus, spawner-isolated mortality like virus and Cherax quadricarinatus parvo-like virus (now Cherax quadricarinatus densovirus, CqDV). The focus of this project was to characterise, for the first time, a Cherax spp. infecting virus, CqDV. The isolate was obtained from the staff at the Queensland Government's former Tropical and Aquatic Animal Health Laboratory. Infected tissue was used to recreate the disease at James Cook University. As well as confirming the results of Bowater et al. (2002), we extended these results, characterising the tissue tropism of CqDV using quantitative real-time PCR (qPCR). We revealed that CqDV preferentially targets ectodermal tissues, consistent with histopathology observations, and that infection is systemic. CqDV was detected in the heart and muscle, most likely from infected haemocytes in the haemocoel. The genome of CqDV was sequenced using primer walking. The CqDV genome is 6,334 nucleotides in length (GenBank: KP410261) and has four open reading frames (ORFs), three (non structural proteins, NS3, NS1 and NS2) on the sense strand and one (viral protein, VP) on the anti-sense strand, indicating an ambisense organisation. Bioinformatics analysis of the ORFs identified highly conserved motifs characteristic of the family Parvoviridae, including endonuclease and helicase motifs in the NS1 protein and a phospholipase A2 motif in the VP. Phylogenetic analysis firmly placed CqDV in the subfamily Densovirinae, genus Ambidensovirus, species Decapod ambidensovirus, virus variant Cherax quadricarinatus densovirus. The CqDV isolate grouped with the blattodean-infecting densoviruses but its genome was architecturally similar to the Lipidopteran ambidensovirus 1 group. The CqDV genome shared 75 % amino acid homology across the entire genome with sea star associated densovirus (SSaDV). Although CqDV and SSaDV are phylogenetically similar, they are geographically and environmentally distinct from each other, infecting two different orders, a unique feature not observed in the Densovirinae to date. Sequencing of the transcriptome revealed four introns in the NS ORFs. These introns could produce four new hypothetical proteins. These included two truncated isoforms of NS3, and one truncated isoform each of NS1 and NS2. The new hypothetical proteins were all in frame. Finally CqDV's ability to cause disease in another commercially cultured crayfish, C. destructor was assessed. Although qPCR was able to detect CqDV in C. destructor + CqDV treatment group, no clinical signs of disease were observed. Future work should investigate if CqDV is replicating in C. destructor and whether or not it can be reisolated and is still infectious to C. quadricarinatus. Future work should also focus on characterising the protein expression of CqDV. CqDV's ability to cause disease in echinoderms and SSaDV's ability to cause disease in freshwater crayfish should also be assessed.

Item ID:	48889
Item Type:	Thesis (PhD)
Keywords:	ambidensovirus, aquaculture, Cherax quadricarinatus, comparative genomics, crayfish, densovirinae, densovirus, freshwater crayfish, genome annotation, parvovirus, phylogeny
Related URLs:	https://researchonline.jcu.edu.au/40676/
Additional Information:	Publications arising from this thesis are available from the Related URLs field. The publications are: Chapter 5: Bochow, Shaun, Condon, Kelly, Elliman, Jennifer, and Owens, Leigh (2015) First complete genome of an Ambidensovirus; Cherax quadricarinatus densovirus, from freshwater crayfish Cherax quadricarinatus. Marine Genomics, 24 (3). pp. 305-312.
Date Deposited:	15 Jun 2017 04:50
FoR Codes:	06 BIOLOGICAL SCIENCES > 0605 Microbiology > 060506 Virology @ 25% 06 BIOLOGICAL SCIENCES > 0604 Genetics > 060408 Genomics @ 50% 06 BIOLOGICAL SCIENCES > 0699 Other Biological Sciences > 069999 Biological Sciences not elsewhere classified @ 25%
SEO Codes:	92 HEALTH > 9201 Clinical Health (Organs, Diseases and Abnormal Conditions) > 920109 Infectious Diseases @ 90% 83 ANIMAL PRODUCTION AND ANIMAL PRIMARY PRODUCTS > 8301 Fisheries - Aquaculture > 830101 Aquaculture Crustaceans (excl. Rock Lobster and Prawns) @ 10%
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