Development of a multiplex RT-PCR assay for simultaneous detection of the major viruses that affect rainbow trout (Oncorhynchus mykiss)

Pinheiro, Ana Cristina De Aguiar Saldanha; Volpe, Enrico; Principi, Donatella; Prosperi, Santino; Ciulli, Sara

doi:10.1007/s10499-015-9912-9

Cited by 13 publications

(13 citation statements)

References 31 publications

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“…Rapid, sensitive, and specific diagnostic methods are important for preventing the spread of pathogens in aquaculture (Castro et al ; Pinheiro et al ). SAD may result in high mortalities in cultured turbot and flounder.…”

Section: Discussionmentioning

confidence: 99%

Identification and Detection of the Pathogenic Bacteria Responsible for Swollen Abdomen Disease in Cultured Turbot, Scophthalmus maximus, and Flounder, Paralichthys olivaceus

Xue

Sun

2017

J World Aquaculture Soc

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Swollen abdomen disease (SAD) seriously threatens the aquaculture of turbots and flounders. Two dominant bacterial strains, FS1 and FS2, were isolated from the livers and kidneys of fish with diagnosed SAD. Applications of biochemical analyses, sequence analyses of 16S ribosomal RNA gene and heat shock protein 60 gene revealed two distinct pathogenic bacterial species, Edwardsiella tarda and Vibrio alginolyticus. These two hypothesized SAD‐causing pathogens were validated by challenge trials on flounder, Paralichthys olivaceus. Challenges with E. tarda and V. alginolyticus demonstrated lethal dose 50 (LD50) values at 1.51 × 105 colony‐forming units (CFU) and 1.05 × 105 CFU, respectively. To develop a rapid SAD diagnosis method in flounders and turbots, a multiplex polymerase chain reaction (PCR) assay method was developed to simultaneously detect E. tarda and V. alginolyticus. Our multiplex PCR assay successfully detected as low as 105 CFU/mL E. tarda and V. alginolyticus in flounders and turbots. No other common fish pathogens were detected with the multiplex PCR, suggesting a high specificity of this assay. The multiplex PCR assay developed in this study showed a great reliability in detecting SAD‐causing bacterial pathogens. Further optimization of this assay may contribute to the development of a novel SAD diagnosis tool in aquaculture.

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Section: Discussionmentioning

confidence: 99%

Identification and Detection of the Pathogenic Bacteria Responsible for Swollen Abdomen Disease in Cultured Turbot, Scophthalmus maximus, and Flounder, Paralichthys olivaceus

Xue

Sun

2017

J World Aquaculture Soc

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“…In order to investigate the presence of IPNV, RNA was extracted from approximately 25 mg of tissue (visceral specimens) using NucleoSpin RNA II (Macherey Nagel) according to the manufacturer's instructions. The RNA samples were subjected to reverse transcriptase–polymerase chain reaction (RT–PCR) analysis to detect IPNV with a protocol previously described (Pinheiro, Volpe, Principi, Prosperi, & Ciulli, ).…”

Section: Methodsmentioning

confidence: 99%

Evidence of fish and human pathogens associated with doctor fish (Garra rufa, Heckel, 1843) used for cosmetic treatment

Volpe

Mandrioli

Errani

et al. 2019

Journal of Fish Diseases

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Doctor fish (Garra rufa, Heckel, 1843) are increasingly used for cosmetic treatment raising particular concerns regarding the potential transmission of infections to clients. Investigations of microbial causes undertaken in two outbreaks of mortality among G. rufa used for cosmetic treatment revealed the presence of multiple bacteria, including both fish and human pathogens such as Aeromonas veronii, A. hydrophila, Vibrio cholerae, Shewanella putrefaciens, Mycobacterium marinum and M. goodii. This range of bacteria indicates an intense microbial proliferation involving multiple pathogens, most likely induced by the poor health condition of the fish. Most of the detected pathogens are well‐known agents of zoonosis. Indeed, M. goodii is an emerging nosocomial human pathogen that has never been detected in fish to date, nor in other animals. This first detection of M. goodii associated with fish infection points out a new zoonotic potential for this pathogen. These findings point out that handling, poor environmental conditions and the presence of fish pathogens, that can compromise the immune system of fish, can result in a mixed microbial proliferation and increase the spread of waterborne bacteria, including zoonosis agents. Accordingly, the microbiological surveillance of fish used for cosmetic treatment is extremely important, particularly in association with mortality outbreaks.

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“…Recently, Pinheiro et al . ( 28 ) have developed a multiplex RT-PCR (mRT-PCR) assay for the single-assay simultaneous detection of four major rainbow trout viruses, namely viral haemorrhagic septicaemia virus (VHSV), infectious haematopoietic necrosis virus (IHNV), infectious pancreatic necrosis virus (IPNV), and SDV. The described method was able to efficiently detect the viral RNA in infected cell culture supernatants and in tissue samples, highlighting the presence of single infections as well as co-infections in rainbow trout samples.…”

Section: Diagnostic Methodsmentioning

confidence: 99%

Salmonid alphavirus (SAV)

Deperasińska

Schulz

Siwicki

2018

Journal of Veterinary Research

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Salmonid alphavirus (SAV), genus Alphavirus, family Togaviridae, is a single-stranded RNA virus affecting Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss). It is known to be responsible for pancreas disease (PD) and sleeping disease (SD) which are increasing problems, causing high fish mortality and economic losses in the European aquaculture industry. Pancreas disease was first described in Atlantic salmon in Scotland in 1976 and a similar disease caused by the closely related sleeping disease virus was first described in rainbow trout in France. There have also been reports of salmonid alphavirus infections from other European countries, including Ireland, England, Norway, Germany, Italy, and Spain. Salmonid alphaviruses have been classified into six subtypes (SAV1–6). SAV1 and SAV4–6 cause pancreas disease in Atlantic salmon in Ireland or Scotland, SAV2 is the causative agent of sleeping disease in rainbow trout, and SAV3 has been detected in Atlantic salmon in Norway. The aim of this paper was to summarise current knowledge of infections caused by salmonid alphavirus and diagnostic methods including the newest techniques, and to briefly describe prevention from SAV infections by vaccination.

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Development of a multiplex RT-PCR assay for simultaneous detection of the major viruses that affect rainbow trout (Oncorhynchus mykiss)

Cited by 13 publications

References 31 publications

Identification and Detection of the Pathogenic Bacteria Responsible for Swollen Abdomen Disease in Cultured Turbot, Scophthalmus maximus, and Flounder, Paralichthys olivaceus

Identification and Detection of the Pathogenic Bacteria Responsible for Swollen Abdomen Disease in Cultured Turbot, Scophthalmus maximus, and Flounder, Paralichthys olivaceus

Evidence of fish and human pathogens associated with doctor fish (Garra rufa, Heckel, 1843) used for cosmetic treatment

Salmonid alphavirus (SAV)

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