RT-nested PCR detection of Mourilyan virus in Australian Penaeus monodon and its tissue distribution in healthy and moribund prawns

Cowley, Jeff A., McCulloch, Russell J., K. V., Rajendran, Cadogan, Lee C., Spann, Kirsten M. and Walker, Peter J. (2005) RT-nested PCR detection of Mourilyan virus in Australian Penaeus monodon and its tissue distribution in healthy and moribund prawns. Diseases of Aquatic Organisms, 66 2: 91-104. doi:10.3354/dao066091

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Author Cowley, Jeff A.
McCulloch, Russell J.
K. V., Rajendran
Cadogan, Lee C.
Spann, Kirsten M.
Walker, Peter J.
Title RT-nested PCR detection of Mourilyan virus in Australian Penaeus monodon and its tissue distribution in healthy and moribund prawns
Journal name Diseases of Aquatic Organisms   Check publisher's open access policy
ISSN 0177-5103
Publication date 2005
Sub-type Article (original research)
DOI 10.3354/dao066091
Open Access Status File (Publisher version)
Volume 66
Issue 2
Start page 91
End page 104
Total pages 14
Place of publication Oldendorf, Germany
Publisher Inter-Research
Language eng
Abstract Mourilyan virus (MoV) is a newly identified virus of Penaeus monodon prawns that is genetically related to the Uukuniemi virus and other phleboviruses of the Bunyaviridae. This paper describes an RT-nested PCR test that can reliably detect between 2 and 6 copies of a synthetic MoV RNA. Total RNA isolated from the lymphoid organ, gills and haemocytes of P. monodon with moderate infections gave comparable amplicon yields in the RT-PCR step of the test. However, in prawns with extremely low-level infections, haemocytes and gill tissue proved slightly more reliable in detecting MoV RNA following nested PCR. The distribution of MoV in tissues of healthy and moribund P. monodon was examined by in situ hybridisation (ISH) using a digoxigenin-labelled DNA probe to a similar to 0.8 kb M RNA segment cDNA insert in clone pMoV4.1. The DNA probe targeted a region in the MoV M RNA segment containing a coding sequence with homology to the C-terminus of the G2 glycoprotein of phleboviruses. In healthy prawns harbouring an unapparent MoV infection, ISH signal primarily occurred in the lymphoid organ, where it was more prominent in hypertrophied cells of 'spheroids' than within cells of normal tubules. ISH signal was also sometimes detected in cells of cuticular epithelium, segmental nerve ganglion and the antermal and tegmental glands. MoV was distributed widely throughout these and other cephalothoracic tissues of mesodermal and ectodermal origin in moribund P. monodon following experimental infection or collected from farm pond edges during disease episodes. Transmission electron microscopy of gill of moribund, captive-reared P. monodon identified spherical (similar to 85 nm diameter) to ovoid MoV particles (similar to 85 x 100 nm) in and around highly necrotic cells in which the nucleus and other organelles had disintegrated. MoV virions co-existed with rod-shaped virions of gill-associated virus and were often seen clustered within cytoplasmic vacuoles or associated with the outer rim of concentric ring-shaped structures comprised of endoplasmic membranes likely to represent degenerated Golgi.
Keyword Fisheries
Veterinary Sciences
Mourilyan virus
Uukuniemi virus
phlebovirus
bunyavirus
Penaeus monodon
penaeid shrimp
prawn
in situ hybridisation
PCR
Gill-associated Virus
Yellow-head Virus
Isolated Mortality Virus
Taura-syndrome Virus
Hematopoietic Necrosis Virus
In-situ Hybridization
Lymphoid Organ Virus
Peripheral Neuropathy
Electron Microscopy
Eastern Australia
Q-Index Code C1

Document type: Journal Article
Sub-type: Article (original research)
Collections: Excellence in Research Australia (ERA) - Collection
Clinical Medical Virology Centre Publications
 
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Created: Fri, 25 Jan 2008, 16:42:01 EST