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Development and validation of a real-time PCR for Chlamydia suis diagnosis in swine and humans.

De Puysseleyr K, De Puysseleyr L, Geldhof J, Cox E, Vanrompay D - PLoS ONE (2014)

Bottom Line: The PCR was successfully validated using conjunctival, pharyngeal and stool samples of slaughterhouse employees, as well as porcine samples from two farms with evidence of reproductive failure and one farm without clinical disease.PCR results were confirmed by culture in McCoy cells.The tet(C) gene was only present in porcine C. suis isolates.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biotechnology, Faculty of Bioscience Engineering, University of Ghent, Ghent, Belgium.

ABSTRACT
Pigs are the natural host for Chlamydia suis, a pathogen which is phylogenetically highly related to the human pathogen C. trachomatis. Chlamydia suis infections are generally treated with tetracyclines. In 1998, tetracyline resistant C. suis strains emerged on U.S. pig farms and they are currently present in the Belgian, Cypriote, German, Israeli, Italian and Swiss pig industry. Infections with tetracycline resistant C. suis strains are mainly associated with severe reproductive failure leading to marked economical loss. We developed a sensitive and specific TaqMan probe-based C. suis real-time PCR for examining clinical samples of both pigs and humans. The analytical sensitivity of the real-time PCR is 10 rDNA copies/reaction without cross-amplifying DNA of other Chlamydia species. The PCR was successfully validated using conjunctival, pharyngeal and stool samples of slaughterhouse employees, as well as porcine samples from two farms with evidence of reproductive failure and one farm without clinical disease. Chlamydia suis was only detected in diseased pigs and in the eyes of humans. Positive humans had no clinical complaints. PCR results were confirmed by culture in McCoy cells. In addition, Chlamydia suis isolates were also examined by the tet(C) PCR, designed for demonstrating the tetracycline resistance gene tet(C). The tet(C) gene was only present in porcine C. suis isolates.

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Related in: MedlinePlus

Real-time PCR quantitation of chlamydial S45 genomic DNA (A) and pGemT::CSIC control plasmid (B).The standard curve data points are the average of 3 replications, standard deviations are shown as error bars. The equations and R2 linearity values resulting from the linear regression analysis, are shown on the graphs.
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pone-0096704-g001: Real-time PCR quantitation of chlamydial S45 genomic DNA (A) and pGemT::CSIC control plasmid (B).The standard curve data points are the average of 3 replications, standard deviations are shown as error bars. The equations and R2 linearity values resulting from the linear regression analysis, are shown on the graphs.

Mentions: Standard curves made using 106 to 101 copies of the pGemT::CSIC control plasmid (Figure 1) showed slopes around −3.103, which correlates to an efficiency of 109%, with correlation coefficients > 98%.


Development and validation of a real-time PCR for Chlamydia suis diagnosis in swine and humans.

De Puysseleyr K, De Puysseleyr L, Geldhof J, Cox E, Vanrompay D - PLoS ONE (2014)

Real-time PCR quantitation of chlamydial S45 genomic DNA (A) and pGemT::CSIC control plasmid (B).The standard curve data points are the average of 3 replications, standard deviations are shown as error bars. The equations and R2 linearity values resulting from the linear regression analysis, are shown on the graphs.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4016100&req=5

pone-0096704-g001: Real-time PCR quantitation of chlamydial S45 genomic DNA (A) and pGemT::CSIC control plasmid (B).The standard curve data points are the average of 3 replications, standard deviations are shown as error bars. The equations and R2 linearity values resulting from the linear regression analysis, are shown on the graphs.
Mentions: Standard curves made using 106 to 101 copies of the pGemT::CSIC control plasmid (Figure 1) showed slopes around −3.103, which correlates to an efficiency of 109%, with correlation coefficients > 98%.

Bottom Line: The PCR was successfully validated using conjunctival, pharyngeal and stool samples of slaughterhouse employees, as well as porcine samples from two farms with evidence of reproductive failure and one farm without clinical disease.PCR results were confirmed by culture in McCoy cells.The tet(C) gene was only present in porcine C. suis isolates.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biotechnology, Faculty of Bioscience Engineering, University of Ghent, Ghent, Belgium.

ABSTRACT
Pigs are the natural host for Chlamydia suis, a pathogen which is phylogenetically highly related to the human pathogen C. trachomatis. Chlamydia suis infections are generally treated with tetracyclines. In 1998, tetracyline resistant C. suis strains emerged on U.S. pig farms and they are currently present in the Belgian, Cypriote, German, Israeli, Italian and Swiss pig industry. Infections with tetracycline resistant C. suis strains are mainly associated with severe reproductive failure leading to marked economical loss. We developed a sensitive and specific TaqMan probe-based C. suis real-time PCR for examining clinical samples of both pigs and humans. The analytical sensitivity of the real-time PCR is 10 rDNA copies/reaction without cross-amplifying DNA of other Chlamydia species. The PCR was successfully validated using conjunctival, pharyngeal and stool samples of slaughterhouse employees, as well as porcine samples from two farms with evidence of reproductive failure and one farm without clinical disease. Chlamydia suis was only detected in diseased pigs and in the eyes of humans. Positive humans had no clinical complaints. PCR results were confirmed by culture in McCoy cells. In addition, Chlamydia suis isolates were also examined by the tet(C) PCR, designed for demonstrating the tetracycline resistance gene tet(C). The tet(C) gene was only present in porcine C. suis isolates.

Show MeSH
Related in: MedlinePlus