Detection of Chlamydia trachomatis mRNA using digital PCR as a more accurate marker of viable organism View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2018-11

AUTHORS

Samuel Phillips, Lenka A. Vodstrcil, Wilhelmina M. Huston, Amba Lawerence, Peter Timms, Marcus Y. Chen, Karen Worthington, Ruthy McIver, Catriona S. Bradshaw, Suzanne M. Garland, Sepehr N. Tabrizi, Jane S. Hocking

ABSTRACT

Spontaneous resolution of urogenital Chlamydia trachomatis (CT) without treatment has previously been described, but a limitation of these reports is that DNA or RNA-based amplification tests used do not differentiate between viable infection and non-viable DNA. We modified a previously published CT mRNA detection (omp2) method to differentiate between viable infection and non-viable DNA in a sample of CT DNA PCR positive women. We modified a CT mRNA detection (omp2) method from reverse transcriptase qPCR (RTqPCR) to digital PCR (dPCR) and evaluated it in samples from CT DNA positive women. Firstly, CT infected McCoy B cells treated with azithromycin in vitro identified detectable mRNA levels disappeared <2 days, while DNA persisted up to 6 days. We used 55 self-collected vaginal swabs from a cohort of women diagnosed as DNA positive for chlamydia obtained pre- and 7 days of post-azithromycin treatment. Concordance with DNA results was higher for dPCR than RTqPCR (74.5% versus 65.5%). At visit 1, there was a strong linear relationship between DNA and mRNA (r = 0.9, p < 0.000); 24 samples had both mRNA and DNA detected (82.8%) and 5 had only DNA detected with a potential false positive proportion of 17.2% (95%CI: 5.8, 35.8). At visit 2, there was poor correlation between DNA and mRNA (r = 0.14, p = 0.55); eight specimens had only DNA detected (42.1%; 95%CI: 20.25, 66.50) and one had mRNA detected. DNA detection methods alone may detect non-viable DNA. Consideration should be given to further develop mRNA assays as ancillary tests to improve detection of viable chlamydia. More... »

PAGES

2117-2122

References to SciGraph publications

Identifiers

URI

http://scigraph.springernature.com/pub.10.1007/s10096-018-3347-y

DOI

http://dx.doi.org/10.1007/s10096-018-3347-y

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1106150203

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/30109584


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25 schema:description Spontaneous resolution of urogenital Chlamydia trachomatis (CT) without treatment has previously been described, but a limitation of these reports is that DNA or RNA-based amplification tests used do not differentiate between viable infection and non-viable DNA. We modified a previously published CT mRNA detection (omp2) method to differentiate between viable infection and non-viable DNA in a sample of CT DNA PCR positive women. We modified a CT mRNA detection (omp2) method from reverse transcriptase qPCR (RTqPCR) to digital PCR (dPCR) and evaluated it in samples from CT DNA positive women. Firstly, CT infected McCoy B cells treated with azithromycin in vitro identified detectable mRNA levels disappeared <2 days, while DNA persisted up to 6 days. We used 55 self-collected vaginal swabs from a cohort of women diagnosed as DNA positive for chlamydia obtained pre- and 7 days of post-azithromycin treatment. Concordance with DNA results was higher for dPCR than RTqPCR (74.5% versus 65.5%). At visit 1, there was a strong linear relationship between DNA and mRNA (r = 0.9, p < 0.000); 24 samples had both mRNA and DNA detected (82.8%) and 5 had only DNA detected with a potential false positive proportion of 17.2% (95%CI: 5.8, 35.8). At visit 2, there was poor correlation between DNA and mRNA (r = 0.14, p = 0.55); eight specimens had only DNA detected (42.1%; 95%CI: 20.25, 66.50) and one had mRNA detected. DNA detection methods alone may detect non-viable DNA. Consideration should be given to further develop mRNA assays as ancillary tests to improve detection of viable chlamydia.
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