Evaluation of a rapid polymerase chain reaction based identification technique for Vibrio cholerae isolates

Roux, Wouter J Le; Masoabi, D.; Wet, C. M. E. de; Venter, Stephanus

doi:10.2166/wst.2004.0059

Cited by 6 publications

(3 citation statements)

References 8 publications

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“…The specificity of these primers was confirmed using DNA probes (Nandi et al, 2000). Le Roux et al (2004) have evaluated and identified a combination of three primers for the detection of V. cholerae ompW gene in environmental isolates. Their work has shown that the PCR approach is more specific than the API 20E and VITEK 32 systems in identifying environmental V. cholerae strains.…”

Section: Pcr Detection Of Unique Gene Fragmentsmentioning

confidence: 97%

The role, isolation and identification of Vibrio species on the quality and safety of seafood

N¹

2012

Biotechnol. Mol. Biol. Rev.

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Seafoods in their natural environments are associated with a variety of microorganisms. Fish shelf life reduction results from microbial metabolism, mainly by Gram negative bacteria that produce chemical compounds responsible for bad odour, texture and taste. Shelflife is estimated by performing total viable bacterial counts at ambient and refrigeration temperatures. The type and number of bacteria present on seafood depends on the microbial composition of the surrounding waters, on the intrinsic factors, extrinsic factors, processing, and implicit factors and on the microbial interactions within the fish itself. Although, sea food safety assessment is preferably determined by detecting indicator organisms; such as Enterobacteriaceae and coliforms, none of these groups fulfil all requirements that guarantee food safety necessitating direct detecting of relevant pathogens. Vibrio species are part of the bacteria genera associated with seafoods borne diseases. Prompt and accurate detection and identification methods of pathogens are imperative to determine the product compliance with seafood microbiological criteria. Although cultural methods have long been used in detecting human pathogens including Vibrio species in fish, these methods are time consuming and sometimes inaccurate. Also some pathogens have the propensity to change into the Viable but non culturable (VBNC) state in unfavourable environments. The use of molecular methods is hampered by drawbacks, such as inter species 16S rRNA sequence similarity and that some strains carry multiple copies of the 16S rRNA gene. A combination of classical, numerical taxonomy and Multi locus sequence analysis (MLSA) methods are promising to give absolute resolution between closely related Vibrio species.

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Section: Pcr Detection Of Unique Gene Fragmentsmentioning

confidence: 97%

The role, isolation and identification of Vibrio species on the quality and safety of seafood

N¹

2012

Biotechnol. Mol. Biol. Rev.

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“…These systems have been used to identify biothreat agents such as Bacillus anthracis (20), Yersinia spp. (16,183), Vibrio cholerae (176), and other pathogens (219). BiOLOG has recently introduced a "dangerous pathogen" supplement (DP Database) to its MicroLog system, although no published evaluations of the system are available currently.…”

Section: Automated Biochemical Testsmentioning

confidence: 99%

Current and Developing Technologies for Monitoring Agents of Bioterrorism and Biowarfare

et al. 2005

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SUMMARY Recent events have made public health officials acutely aware of the importance of rapidly and accurately detecting acts of bioterrorism. Because bioterrorism is difficult to predict or prevent, reliable platforms to rapidly detect and identify biothreat agents are important to minimize the spread of these agents and to protect the public health. These platforms must not only be sensitive and specific, but must also be able to accurately detect a variety of pathogens, including modified or previously uncharacterized agents, directly from complex sample matrices. Various commercial tests utilizing biochemical, immunological, nucleic acid, and bioluminescence procedures are currently available to identify biological threat agents. Newer tests have also been developed to identify such agents using aptamers, biochips, evanescent wave biosensors, cantilevers, living cells, and other innovative technologies. This review describes these current and developing technologies and considers challenges to rapid, accurate detection of biothreat agents. Although there is no ideal platform, many of these technologies have proved invaluable for the detection and identification of biothreat agents.

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“…It is associated but not limited to estuarine systems. V. cholerae is known to persist in inland freshwater systems (le Roux et al, 2004). Efforts to manage and mitigate cholera outbreaks invariably start with bacteriological monitoring of aquatic ecosystems, a procedure that is considered essential for the tracking of V. cholerae O1 (Binsztein et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

The use of a high resolution melt real-time polymerase chain reaction (PCR) assay for the environmental monitoring of Vibrio cholerae

Roux¹

2011

Afr. J. Microbiol. Res.

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A real-time polymerase chain reaction (PCR) assay utilizing high resolution melt (HRM) curve analysis was developed and tested for the monitoring of Vibrio cholerae in water samples. The assay utilized previously published primers that are specific to regions of the V. cholerae ompW and ctxAB genes, allowing it to differentiate between toxigenic and non-toxigenic strains. The ompW and ctxAB primers amplify target regions of 588 and 564 bp in length (respectively) and the amplicons could be accurately identified using HRM curve analysis. High resolution melt curve analysis provided additional accuracy for the determination of amplicon melting temperatures, and allowed amplification of the two targets in a multiplex reaction. Two laboratories employed the assay to analyse 178 water samples obtained from diverse environmental water sources, for the presence of V. cholerae. The assay was found to be a rapid, highly accurate, sensitive and cost effective method for the detection and distinction between toxigenic and non-toxigenic V. cholerae strains in water.

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Evaluation of a rapid polymerase chain reaction based identification technique for Vibrio cholerae isolates

Cited by 6 publications

References 8 publications

The role, isolation and identification of Vibrio species on the quality and safety of seafood

The role, isolation and identification of Vibrio species on the quality and safety of seafood

Current and Developing Technologies for Monitoring Agents of Bioterrorism and Biowarfare

The use of a high resolution melt real-time polymerase chain reaction (PCR) assay for the environmental monitoring of Vibrio cholerae

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