Mycoplasma detection by triplex real-time PCR in bronchoalveolar lavage fluid from bovine respiratory disease complex cases

Jan B.W.J. Cornelissen*, Freddy M. de Bree, Fimme J. van der Wal, Engbert A. Kooi, Miriam G.J. Koene, Alex Bossers, Bregtje Smid, Adriaan F. Antonis, Henk J. Wisselink

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

4 Citations (Scopus)

Abstract

Background: In this study we evaluated the RespoCheck Mycoplasma triplex real-time PCR for the detection in bronchoalveolar lavage fluid (BALF) of Mycoplasma (M.) dispar, M. bovis and M. bovirhinis, all three associated with bovine respiratory disease (BRD). Primers and probes of the RespoCheck Mycoplasma triplex real-time PCR are based on the V3/V4 region of the 16S rRNA gene of the three Mycoplasma species. Results: The analytical sensitivity of the RespoCheck triplex real-time PCR was, as determined by spiking experiments of the Mycoplasma strains in Phosphate Buffered Saline, 300 colony forming units (cfu)/mL for M. dispar, and 30cfu/mL for M. bovis or M. bovirhinis. The analytical sensitivity of the RespoCheck Mycoplasma triplex real-time PCRwas, as determined on purified DNA, 10fg DNA per assay for M. dispar and 100fg fo rM. bovis and M. bovirhinis. The analytical specificity of the RespoCheck Mycoplasma triplex real-time PCR was, as determined by testing Mycoplasmas strains (n=17) and other bacterial strains (n=107), 100, 98.2 and 99.1% for M. bovis, M. dispar and M. bovirhinis respectively. The RespoCheck Mycoplasma triplex real-time PCR was compared with the PCR/DGGE analysis for M. bovis, M. dispar and M. bovirhinis respectively by testing 44 BALF samples from calves. Conclusion: In conclusion, the RespoCheck PCR assay can be a valuable tool for timely and accurate detection of three Mycoplasma species associated with in bovine respiratory disease.

Original languageEnglish
Article number97
JournalBMC Veterinary Research
Volume13
Issue number1
DOIs
Publication statusPublished - 2017

Fingerprint

Bovine Respiratory Disease Complex
bovine respiratory disease
Mycoplasma
Multiplex Polymerase Chain Reaction
Bronchoalveolar Lavage Fluid
Real-Time Polymerase Chain Reaction
quantitative polymerase chain reaction
Mycoplasma bovirhinis
Mycoplasma dispar
Mycoplasma bovis
Cattle Diseases
detection limit
analytical specificity
fluids
Polymerase Chain Reaction
DNA
rRNA Genes
assays

Keywords

  • Bovine Mycoplasma
  • Bovine respiratory disease
  • M. bovirhinis
  • M. bovis
  • M. dispar
  • RespoCheck
  • Triplex PCR

Cite this

@article{8f92e6f774744cc986da5e0b0c7940ac,
title = "Mycoplasma detection by triplex real-time PCR in bronchoalveolar lavage fluid from bovine respiratory disease complex cases",
abstract = "Background: In this study we evaluated the RespoCheck Mycoplasma triplex real-time PCR for the detection in bronchoalveolar lavage fluid (BALF) of Mycoplasma (M.) dispar, M. bovis and M. bovirhinis, all three associated with bovine respiratory disease (BRD). Primers and probes of the RespoCheck Mycoplasma triplex real-time PCR are based on the V3/V4 region of the 16S rRNA gene of the three Mycoplasma species. Results: The analytical sensitivity of the RespoCheck triplex real-time PCR was, as determined by spiking experiments of the Mycoplasma strains in Phosphate Buffered Saline, 300 colony forming units (cfu)/mL for M. dispar, and 30cfu/mL for M. bovis or M. bovirhinis. The analytical sensitivity of the RespoCheck Mycoplasma triplex real-time PCRwas, as determined on purified DNA, 10fg DNA per assay for M. dispar and 100fg fo rM. bovis and M. bovirhinis. The analytical specificity of the RespoCheck Mycoplasma triplex real-time PCR was, as determined by testing Mycoplasmas strains (n=17) and other bacterial strains (n=107), 100, 98.2 and 99.1{\%} for M. bovis, M. dispar and M. bovirhinis respectively. The RespoCheck Mycoplasma triplex real-time PCR was compared with the PCR/DGGE analysis for M. bovis, M. dispar and M. bovirhinis respectively by testing 44 BALF samples from calves. Conclusion: In conclusion, the RespoCheck PCR assay can be a valuable tool for timely and accurate detection of three Mycoplasma species associated with in bovine respiratory disease.",
keywords = "Bovine Mycoplasma, Bovine respiratory disease, M. bovirhinis, M. bovis, M. dispar, RespoCheck, Triplex PCR",
author = "Cornelissen, {Jan B.W.J.} and {de Bree}, {Freddy M.} and {van der Wal}, {Fimme J.} and Kooi, {Engbert A.} and Koene, {Miriam G.J.} and Alex Bossers and Bregtje Smid and Antonis, {Adriaan F.} and Wisselink, {Henk J.}",
year = "2017",
doi = "10.1186/s12917-017-1023-6",
language = "English",
volume = "13",
journal = "BMC Veterinary Research",
issn = "1746-6148",
publisher = "Springer Verlag",
number = "1",

}

TY - JOUR

T1 - Mycoplasma detection by triplex real-time PCR in bronchoalveolar lavage fluid from bovine respiratory disease complex cases

AU - Cornelissen, Jan B.W.J.

AU - de Bree, Freddy M.

AU - van der Wal, Fimme J.

AU - Kooi, Engbert A.

AU - Koene, Miriam G.J.

AU - Bossers, Alex

AU - Smid, Bregtje

AU - Antonis, Adriaan F.

AU - Wisselink, Henk J.

PY - 2017

Y1 - 2017

N2 - Background: In this study we evaluated the RespoCheck Mycoplasma triplex real-time PCR for the detection in bronchoalveolar lavage fluid (BALF) of Mycoplasma (M.) dispar, M. bovis and M. bovirhinis, all three associated with bovine respiratory disease (BRD). Primers and probes of the RespoCheck Mycoplasma triplex real-time PCR are based on the V3/V4 region of the 16S rRNA gene of the three Mycoplasma species. Results: The analytical sensitivity of the RespoCheck triplex real-time PCR was, as determined by spiking experiments of the Mycoplasma strains in Phosphate Buffered Saline, 300 colony forming units (cfu)/mL for M. dispar, and 30cfu/mL for M. bovis or M. bovirhinis. The analytical sensitivity of the RespoCheck Mycoplasma triplex real-time PCRwas, as determined on purified DNA, 10fg DNA per assay for M. dispar and 100fg fo rM. bovis and M. bovirhinis. The analytical specificity of the RespoCheck Mycoplasma triplex real-time PCR was, as determined by testing Mycoplasmas strains (n=17) and other bacterial strains (n=107), 100, 98.2 and 99.1% for M. bovis, M. dispar and M. bovirhinis respectively. The RespoCheck Mycoplasma triplex real-time PCR was compared with the PCR/DGGE analysis for M. bovis, M. dispar and M. bovirhinis respectively by testing 44 BALF samples from calves. Conclusion: In conclusion, the RespoCheck PCR assay can be a valuable tool for timely and accurate detection of three Mycoplasma species associated with in bovine respiratory disease.

AB - Background: In this study we evaluated the RespoCheck Mycoplasma triplex real-time PCR for the detection in bronchoalveolar lavage fluid (BALF) of Mycoplasma (M.) dispar, M. bovis and M. bovirhinis, all three associated with bovine respiratory disease (BRD). Primers and probes of the RespoCheck Mycoplasma triplex real-time PCR are based on the V3/V4 region of the 16S rRNA gene of the three Mycoplasma species. Results: The analytical sensitivity of the RespoCheck triplex real-time PCR was, as determined by spiking experiments of the Mycoplasma strains in Phosphate Buffered Saline, 300 colony forming units (cfu)/mL for M. dispar, and 30cfu/mL for M. bovis or M. bovirhinis. The analytical sensitivity of the RespoCheck Mycoplasma triplex real-time PCRwas, as determined on purified DNA, 10fg DNA per assay for M. dispar and 100fg fo rM. bovis and M. bovirhinis. The analytical specificity of the RespoCheck Mycoplasma triplex real-time PCR was, as determined by testing Mycoplasmas strains (n=17) and other bacterial strains (n=107), 100, 98.2 and 99.1% for M. bovis, M. dispar and M. bovirhinis respectively. The RespoCheck Mycoplasma triplex real-time PCR was compared with the PCR/DGGE analysis for M. bovis, M. dispar and M. bovirhinis respectively by testing 44 BALF samples from calves. Conclusion: In conclusion, the RespoCheck PCR assay can be a valuable tool for timely and accurate detection of three Mycoplasma species associated with in bovine respiratory disease.

KW - Bovine Mycoplasma

KW - Bovine respiratory disease

KW - M. bovirhinis

KW - M. bovis

KW - M. dispar

KW - RespoCheck

KW - Triplex PCR

U2 - 10.1186/s12917-017-1023-6

DO - 10.1186/s12917-017-1023-6

M3 - Article

VL - 13

JO - BMC Veterinary Research

JF - BMC Veterinary Research

SN - 1746-6148

IS - 1

M1 - 97

ER -