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

Maureen K. Purcell; Rachel L. Thompson; Kyle A. Garver; Laura M. Hawley; William N. Batts; Laura Sprague; Corie Sampson; James R. Winton

doi:10.3354/dao02644

USGS · 70058654

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

Maureen K. Purcell·Rachel L. Thompson·Kyle A. Garver·Laura M. Hawley·William N. Batts·Laura Sprague·Corie Sampson·James R. Winton

View original source ↗Export metadata ↓

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.

Explore related subjects

Diseases of Aquatic Organisms Article

Keep this discovery

Explore connections, maps & timelines

BibTeX RIS

Maureen K. Purcell, Rachel L. Thompson, Kyle A. Garver, Laura M. Hawley, William N. Batts, Laura Sprague, Corie Sampson, James R. Winton. 2013. Universal reverse-transcriptase real-time PCR for infectious hematopoietic necrosis virus (IHNV). https://doi.org/10.3354/dao02644

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related USGS reports

USGS2026

First report of the flatworm Convolutriloba longifissura (Acoela) affecting a Caribbean coral species in a land-based nursery

Flatworms are widespread in the marine realm, but only a few species parasitize corals. Several of these are considered pests in the aquarium trade, and the invasive potential of these species is a concern. In the Caribbean there are no records to date of flatworms naturally occurring in coral nurseries and/or parasitizing corals in the wild. Here we report, for the first time, a flatworm identified as Convolutriloba longifissura affecting a Caribbean coral and include a phylogenetic and histological analyses. Flatworms were collected from nursery corals and artificial recruitment units (bioballs) suspended in the water column at Punta Cana reef. Sequencing and morphology of individuals from both indicate that C. longifissura is present in the wild as well as the nursery. Experiments were conducted to investigate the effect of flatworm exposure on signs of macroscopic stress (i.e. mucus production, paleness, bleaching, and tissue loss) on microfragments of Pseudodiploria clivosa . Standard histopathological techniques were applied to describe microscopic alterations of the coral exposed to C. longifissura. We found corals exposed to 30 individuals of C. longifissura had a 2.5-fold increased probability of developing macroscopic stress signs compared to controls after 14 d of exposure to different flatworm densities. While the actual impacts of these flatworms on coral restoration and microfragmentation programs or wild corals in the Caribbean remain to be determined, our results indicate that flatworms are no longer an exclusive problem of Indo-Pacific corals.

Punta Cana reef↗

USGS2025

Histological progression and bacterial load dynamics of Renibacterium salmoninarum in Chinook salmon Oncorhynchus tshawytscha

Renibacterium salmoninarum , the cause of bacterial kidney disease (BKD), severely impacts salmonid populations. Much of our understanding of the BKD pathology in salmonids comes from evaluating fatal infections in wild populations or spawning Pacific Northwest salmonids. Our study investigated the histological progression and bacterial load dynamics of R. salmoninarum infection in Chinook salmon following intraperitoneal injection to enhance understanding of the disease’s dynamics. Seventy presmolt salmon were injected with the ATCC-33209 isolate of R. salmoninarum and monitored over 10 wk. Histological signs of disease were observed in spleen and liver tissues at 1 wk post-injection and in kidney tissues at 4 wk post-injection, with signs of disease increasing over time. Additionally, histopathological analysis revealed splenic and hepatic capsulitis (coelomitis), individual hepatocyte necrosis, progressive granulomatous hepatitis, splenitis, and nephritis with necrosis. Gram staining confirmed the presence of Gram-positive bacteria within macrophages and extracellularly in infected tissues. Quantitative PCR revealed significant increases in bacterial loads in both kidney and liver tissues over the study period, with higher bacterial loads observed in the kidney tissue. Bacterial load was strongly correlated with disease stage, with peak bacterial burdens coinciding with the most severe histological changes. Our study provides a comprehensive account of R. salmoninarum infection and disease progression in Chinook salmon after intraperitoneal injection.

Diseases of Aquatic Organisms↗

USGS2024

Cytology in cnidaria using Exaiptasia as a model

A need exists for additional methods to examine cnidaria at the cellular level to aid our understanding of health, anatomy, and physiology of this important group of organisms. This need is particularly acute given that disease is emerging as a major factor in declines of ecologically important functional groups such as corals. Here we describe a simple method to process cnidarian cells for microscopic examination using the model organism Exaiptasia . We show that this organism has at least 18 cell types or structures that can be readily distinguished based on defined morphological features. Some of these cells can be related back to anatomic features of the animal both at the light microscope and ultrastructural level. The cnidome of Exaiptasia may be more complex than what is currently understood. Moreover, cnidarian cells, including some types of cnidocytes, phagocytize cells other than endosymbionts. Finally, our findings shed light on morphologic complexity of cell-associated microbial aggregates and their intimate intracellular associations. The tools described here could be useful for other cnidaria.

Diseases of Aquatic Organisms↗