Universal reverse-transcriptase real-time PCR for infectious hematopoietic necrosis virus (IHNV)

Diseases of Aquatic Organisms

Kyle A. Garver¹  Laura Sprague¹  Rachel L. Thompson¹  William N. Batts¹  James R. Winton¹  Maureen K. Purcell¹  Corie Sampson¹  Laura M. Hawley¹ 

关键词: Infectious hematopoietic necrosis virus;    HNV;    Real-time PCR;    Diagnostic validation;    Field validation;    Steelhead trout;    Reproducibility;    Genogroup;   

DOI  :  10.3354/dao02644

学科分类：生物科学（综合）

来源: Inter-Research

【 摘 要 】

ABSTRACT: Infectious hematopoietic necrosis virus (IHNV) is an acute pathogen of salmonid fishes in North America, Europe and Asia and is reportable to the World Organization for Animal Health (OIE). Phylogenetic analysis has identified 5 major virus genogroups of IHNV worldwide, designated U, M, L, E and J; multiple subtypes also exist within those genogroups. Here, we report the development and validation of a universal IHNV reverse-transcriptase real-time PCR (RT-rPCR) assay targeting the IHNV nucleocapsid (N) gene. Properties of diagnostic sensitivity (DSe) and specificity (DSp) were defined using laboratory-challenged steelhead trout Oncorhynchus mykiss, and the new assay was compared to the OIE-accepted conventional PCR test and virus isolation in cell culture. The IHNV N gene RT-rPCR had 100% DSp and DSe and a higher estimated diagnostic odds ratio (DOR) than virus culture or conventional PCR. The RT-rPCR assay was highly repeatable within a laboratory and highly reproducible between laboratories. Field testing of the assay was conducted on a random sample of juvenile steelhead collected from a hatchery raceway experiencing an IHN epizootic. The RT-rPCR detected a greater number of positive samples than cell culture and there was 40% agreement between the 2 tests. Overall, the RT-rPCR assay was highly sensitive, specific, repeatable and reproducible and is suitable for use in a diagnostic setting.

【 授权许可】

Unknown