Development of a Real-time PCR Assay for the Detection and Quantification of Red Seabream Iridovirus (RSIV).

Caipang, Christopher Marlowe A.; Hirono, Ikuo; Aoki, Takashi

doi:10.3147/jsfp.38.1

Cited by 60 publications

(38 citation statements)

References 25 publications

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“…TaqMan real-time PCR assays are commonly utilised in diagnostic laboratories to provide rapid, highly sensitive and sequence-specific results. Universal megalocytivirus SYBR Green qPCRs have previously been developed by Caipang et al (2003), Gias et al (2011) and Rimmer et al (2012), but, while more expensive, TaqMan qPCRs are generally more specific and reproducible than SYBR Green assays (Gunson et al 2006, Beld et al 2007, Purcell et al 2011.…”

Section: Discussionmentioning

confidence: 99%

“…The Megalocytivirus TaqMan qPCR, based on a SYBR Green method (Caipang et al 2003), was developed for the detection of RSIV but has also been shown to detect ISKNV-like viral nucleic acid in our laboratory. All samples were initially screened by this Megalocytivirus TaqMan qPCR ( Table 1) The qPCR assays were performed in a 7500 Fast Real-Time PCR System (Life Technologies) and analysed with the 7500 software.…”

Section: Molecular Detection and Sequence Analysismentioning

confidence: 99%

See 1 more Smart Citation

Molecular confirmation of infectious spleen and kidney necrosis virus (ISKNV) in farmed and imported ornamental fish in Australia

Mohr¹,

Moody²,

Williams³

et al. 2015

Dis. Aquat. Org.

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Section: Discussionmentioning

confidence: 99%

Section: Molecular Detection and Sequence Analysismentioning

confidence: 99%

Molecular confirmation of infectious spleen and kidney necrosis virus (ISKNV) in farmed and imported ornamental fish in Australia

Mohr¹,

Moody²,

Williams³

et al. 2015

Dis. Aquat. Org.

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“…Bacterial genomic DNA was extracted using the previously described method (Caipang et al 2003), and the 16S rDNA was amplified using the eubacterial universal primers (Bianciotto et al 2003). The PCR products were sequenced using Big Dye Terminator ver 3.1 (Applied Biosystems, USA), and comparative sequence analysis was carried out on the sequences available in NCBI (National Center for Biotechnology Information) employing nucleotide BLAST (Basic Local Alignment Search Tool).…”

Section: Molecular Characterization Of the Isolated Phytase-producingmentioning

confidence: 99%

Responses of Atlantic cod Gadus morhua head kidney leukocytes to phytase produced by gastrointestinal-derived bacteria

Lazado¹,

Caipang²,

Gallage³

et al. 2009

Fish Physiol Biochem

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This study identified phytase-producing bacteria that were previously isolated from the gastrointestinal tract of Atlantic cod, Gadus morhua and determined its effect on head kidney leukocytes. Out of the 216 bacterial strains tested, the two phytase producers were identified as Pseudomonas sp. and Psychrobacter sp. based on their 16S rDNA sequence. Crude phytase from these two bacterial strains was produced employing the shake flask method. Even though the total protein of the crude phytase was not significantly different for the two bacteria, the phytase activity of the crude enzyme produced by Pseudomonas sp. (97.1±16.7 U) was significantly higher than that of the enzyme from Psychrobacter sp. (75.9±2.4 U). When cod head kidney leukocytes were incubated with the crude phytase (50 μg ml(-1)), it resulted in enhanced cell proliferation, higher myeloperoxidase, and acid phosphatase activities. Extracellular responses-respiratory burst activity and hydrogen peroxide production were not enhanced by the crude enzyme. As a consequence, the growth of two pathogenic bacteria Aeromonas salmonicida and Vibrio anguillarum was not suppressed by the supernatants obtained from head kidney leukocytes incubated with the crude bacterial phytase. Thus, the enzyme from phytase-producing intestinal bacteria of Atlantic cod can stimulate intracellular head kidney leukocyte activities but not the production of extracellular substances that are involved in antibacterial response. These have implications on the potential use of bacterial phytase as feed supplement to boost cellular immune response of the fish and could be employed as a health management strategy in culture systems.

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“…Several detection techniques have been established and applied to detect RSIV, including quantitative realtime PCR (Caipang et al, 2003), conventional PCR (Oshima et al, 1996;Oshima et al, 1998;Kurita et al, 1998), Giemsa staining (Inoue et al, 1992) and immunofluorescence assay using a monoclonal antibody (Nakajima and Sorimachi, 1995). Among these methods, real-time PCR is effective for detection and quantification of the virus.…”

Section: Introductionmentioning

confidence: 99%

Rapid and sensitive detection of iridovirus by loop-mediated isothermal amplification (LAMP)

Hwang¹,

Suh²,

Park³

et al. 2015

Afr. J. Microbiol. Res.

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Red seabream iridovirus (RSIV), a member of the Iridoviridae family, is the causative pathogen of some of the most explosive epidemics of emerging viral diseases in many Asian countries, leading to huge economic losses in aquaculture. Rapid molecular detection for surveillance or diagnosis has been a critical component in reducing the prevalence of RSIV infection. In the present study, a novel and highly specific loop-mediated isothermal amplification (LAMP) assay for the sensitive and rapid detection of RSIV infection in fishes was developed. Using a set of synthesized primers matching a specific region of the RSIV genome (GenBank accession no.: AB666336.1), the efficiency and specificity of the LAMP assay were optimized in terms of the reaction temperature and DNA polymerase concentration, as they are the main determinants of the sensitivity and specificity of the LAMP assay. In particular, we demonstrated that our assay could be applied to efficiently detect RSIV infection in red sea bream. Our results provide a simple and convenient method for the detection of viral infection in aquatic organisms.

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Development of a Real-time PCR Assay for the Detection and Quantification of Red Seabream Iridovirus (RSIV).

Cited by 60 publications

References 25 publications

Molecular confirmation of infectious spleen and kidney necrosis virus (ISKNV) in farmed and imported ornamental fish in Australia

Molecular confirmation of infectious spleen and kidney necrosis virus (ISKNV) in farmed and imported ornamental fish in Australia

Responses of Atlantic cod Gadus morhua head kidney leukocytes to phytase produced by gastrointestinal-derived bacteria

Rapid and sensitive detection of iridovirus by loop-mediated isothermal amplification (LAMP)

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