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The Journal of veterinary medical science2014; 76(9); 1271-1275; doi: 10.1292/jvms.14-0140

Development of a loop-mediated isothermal amplification method for detecting Streptococcus equi subsp. zooepidemicus and analysis of its use with three simple methods of extracting DNA from equine respiratory tract specimens.

Abstract: Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) is a dominant pathogenic bacterium in equine pneumonia. We developed a specific loop-mediated isothermal amplification (LAMP) method, which targets the gene encoding sorbitol-6-phosphate 2-dehydrogenase (sorD), for detecting S. zooepidemicus and examined the clinical efficacies of its use in combination with each of 3 DNA extraction methods easily used by veterinary practitioners, namely the Loopamp PURE DNA Extraction Kit, InstaGene Matrix and a conventional boiling method. The LAMP method plus the Loopamp PURE DNA Extraction Kit gave higher rates of positivity than the other combinations in both clinical and spiked samples containing clinically significant concentrations (>1 × 10(4) CFU/ml) of S. zooepidemicus.
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Publication Date: 2014-05-28 PubMed ID: 24871644PubMed Central: PMC4197157DOI: 10.1292/jvms.14-0140Google Scholar: Lookup
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  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

The research focuses on the development of a specific method, known as loop-mediated isothermal amplification (LAMP), for detecting a primary cause of pneumonia in horses, Streptococcus equi subsp. zooepidemicus. The study also investigates the effectiveness of this method combined with three different ways of extracting DNA.

Underlying Research Objectives

  • The primary aim of this research is to develop a loop-mediated isothermal amplification (LAMP) method specifically for the detection of Streptococcus equi subsp. zooepidemicus, a primary pathogenic bacterium causing pneumonia in horses.
  • Furthermore, the study intends to evaluate the effectiveness of this new method in conjunction with three easily applied DNA extraction procedures that are usable by veterinary practitioners. These procedures are the Loopamp PURE DNA Extraction Kit, InstaGene Matrix, and a conventional boiling method.

Research Method and Process

  • The LAMP method developed in the study targets the gene encoding sorbitol-6-phosphate 2-dehydrogenase (sorD) – a specific gene identified in S. zooepidemicus.
  • The effectiveness of this method was evaluated both in clinical applications and theoretical samples spiked with significantly high concentrations (>1 × 10(4) CFU/ml) of S. zooepidemicus.
  • Comparisons of the results were made between the LAMP method combined with each of the three DNA extraction techniques.

Findings and Conclusion

  • The outcome of the research revealed that the combination of the LAMP method and the Loopamp PURE DNA Extraction Kit resulted in higher positivity rates for the detection of S. zooepidemicus. This was the case for both actual clinical specimens and spiked samples containing significantly high concentrations of the bacteriophage.
  • This suggests that this particular combination may prove the most effective strategy for detecting the bacterium in a veterinary practice setting.

Cite This Article

APA
Kinoshita Y, Niwa H, Katayama Y. (2014). Development of a loop-mediated isothermal amplification method for detecting Streptococcus equi subsp. zooepidemicus and analysis of its use with three simple methods of extracting DNA from equine respiratory tract specimens. J Vet Med Sci, 76(9), 1271-1275. https://doi.org/10.1292/jvms.14-0140

Publication

The Journal of veterinary medical science
ISSN: 1347-7439
NlmUniqueID: 9105360
Country: Japan
Language: English
Volume: 76
Issue: 9
Pages: 1271-1275

Researcher Affiliations

Kinoshita, Yuta
  • Microbiology Division, Epizootic Research Center, Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotuke, Tochigi 329-0412, Japan.
Niwa, Hidekazu
    Katayama, Yoshinari

      MeSH Terms

      • Animals
      • Colony Count, Microbial / veterinary
      • DNA, Bacterial / chemistry
      • DNA, Bacterial / genetics
      • Horse Diseases / diagnosis
      • Horse Diseases / microbiology
      • Horses
      • Limit of Detection
      • Nucleic Acid Amplification Techniques / methods
      • Nucleic Acid Amplification Techniques / veterinary
      • Pneumonia / diagnosis
      • Pneumonia / microbiology
      • Pneumonia / veterinary
      • Polymerase Chain Reaction / veterinary
      • Sensitivity and Specificity
      • Streptococcal Infections / diagnosis
      • Streptococcal Infections / microbiology
      • Streptococcal Infections / veterinary
      • Streptococcus equi / genetics
      • Streptococcus equi / isolation & purification

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      Citations

      This article has been cited 6 times.
      1. Garner C, Stephen C, Pant SD, Ghorashi SA. Comparison of PCR-HRM, colorimetric LAMP and culture based diagnostic assays in the detection of endometritis caused by Streptococcus equi subsp. zooepidemicus in mares. Vet Res Commun 2023 Jun;47(2):495-509.
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      2. Uchida-Fujii E, Kinoshita Y, Niwa H, Maeda T, Nukada T, Ueno T. High prevalence of Mycoplasma equirhinis in Thoroughbred horses with respiratory symptoms in autumn 2018. J Vet Med Sci 2021 Dec 9;83(12):1907-1912.
        doi: 10.1292/jvms.21-0163pubmed: 34732605google scholar: lookup
      3. Jelocnik M, Nyari S, Anstey S, Playford N, Fraser TA, Mitchell K, Blishen A, Pollak NM, Carrick J, Chicken C, Jenkins C. Real-time fluorometric and end-point colorimetric isothermal assays for detection of equine pathogens C. psittaci and equine herpes virus 1: validation, comparison and application at the point of care. BMC Vet Res 2021 Aug 19;17(1):279.
        doi: 10.1186/s12917-021-02986-8pubmed: 34412635google scholar: lookup
      4. Inoshima Y, Takasu M, Ishiguro N. Establishment of an on-site diagnostic procedure for detection of orf virus from oral lesions of Japanese serows (Capricornis crispus) by loop-mediated isothermal amplification. J Vet Med Sci 2017 Jan 10;78(12):1841-1845.
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