Development and diagnostic validation of a one-step multiplex RT-PCR assay as a rapid method to detect and identify Nervous Necrosis Virus (NNV) and its variants circulating in the Mediterranean
Errani, Francesca (University of Bologna. Department of Veterinary Medical Sciences)
Volpe, Enrico (University of Bologna. Department of Veterinary Medical Sciences)
Riera-Ferrer, Enrique (Universitat Autònoma de Barcelona. Departament de Biologia Animal, de Biologia Vegetal i d'Ecologia)
Caffara, Monica (University of Bologna. Department of Veterinary Medical Sciences)
Padrós, Francesc 1965- (Universitat Autònoma de Barcelona. Departament de Biologia Animal, de Biologia Vegetal i d'Ecologia)
Gustinelli, Andrea (University of Bologna. Department of Veterinary Medical Sciences)
Fioravanti, Marialetizia (University of Bologna. Department of Veterinary Medical Sciences)
Ciulli, Sara (University of Bologna. Department of Veterinary Medical Sciences)

Data:	2022
Resum:	Nervous Necrosis Virus (NNV) represents one of the most threatening pathogens for Mediterranean aquaculture. Several NNV strains are currently co-circulating in the Mediterranean Basin with a high prevalence of the RGNNV genotype and the RGNNV/SJNNV reassortant strain and a more limited diffusion of the SJNNV genotype and the SJNNV/RGNNV reassortant. In the present study, a one-step multiplex RT-PCR (mRT-PCR) assay was developed as an easy, cost-effective and rapid diagnostic technique to detect RGNNV and the reassortant RGNNV/SJNNV strain and to distinguish them from SJNNV and the reassortant SJNNV/RGNNV strain in a single RT-PCR reaction. A unique amplification profile was obtained for each genotype/reassortant enabling their rapid identification from cell culture lysates or directly from brain tissues of suspected fish. The method's detection limit varied between 10 2. 3 and 10 3. 4 TCID ml -1 depending on viral strains. No cross-reacitivty with viruses and bacteria frequently associated with gilthead seabream, European seabass and marine environment was observed. The mRT-PCR was shown to be an accurate, rapid and affordable method to support traditional diagnostic techniques in the diagnosis of VNN, being able to reduce considerably the time to identify the viral genotype or the involvement of reassortant strains.
Ajuts:	European Commission 727610
Drets:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	Reverse transcriptase-polymerase chain reaction ; Polymerase chain reaction ; RNA sequencing ; RNA extraction ; Necrosis ; Genotyping ; RNA amplification ; Viral genome
Publicat a:	PloS one, Vol. 17 (august 2022) , ISSN 1932-6203

DOI: 10.1371/journal.pone.0273802
PMID: 36018889

16 p, 1.2 MB

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