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P489 Chlamydia trachomatis -specific gene transcripts as a more accurate marker for infection status
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  1. Kevin Janssen1,
  2. Christian Hoebe2,
  3. Romy Nijs1,
  4. Nicole Dukers-Muijrers2,
  5. Petra Wolffs3
  1. 1Maastricht University Medical Centre (MUMC), Department of Medical Microbiology, Care and Public Health Research Institute (CAPHRI), Maastricht, Netherlands
  2. 2Public Health Service South Limburg, Maastricht University Medical Center (MUMC), Sexual Health, Infectious Diseases and Environmental Health, Medical Microbiology, Care and Public Health Research Institute (CAPHRI), Heerlen, Netherlands
  3. 3Maastricht University Medical Center (MUMC), Medical Microbiology, Care and Public Health Research Institute (CAPHRI), Maastricht, Netherlands

Abstract

Background Current routine diagnostic methods for the detection of Chlamydia trachomatis (CT) do not provide information on CT viability. Previously, detection of messenger-RNA (mRNA) has been utilized as a marker for bacterial viability, as mRNA molecules are generally short-lived (half-life of minutes). However, only one study evaluated CT mRNA half-life times [t1/2] of two clinical isolates (serovar L2b and E) which ranged from 1 to ≥5000 minutes. Here we assess and confirm mRNA half-life times of serovar D to further facilitate evaluation of CT viability.

Methods CT serovar D was propagated in HeLa cells until 30 h post infection and treated with rifampicin to arrest gene transcription. Total RNA was isolated at 0 min (before treatment), and 10 min, 30 min and 60 min after treatment. RNA was converted to cDNA using random hexamers. RT-qPCR was used to amplify fragments of the unprocessed 16S (intermediate molecules in 16S rRNA synthesis), 16S (early), rpoD (early), omcB (mid), and hctA (late) gene transcripts. Half-life time was based on the fit of an exponential decay between values obtained at before and after transcriptional arrest.

Results In this study showed that the obtained t1/2 values for CT serovar D mRNA (median t1/218 min) were similar as previously reported for the CT serovars L2b and E (median t1/2 15 and 17 min, respectively). The observed half-lifes of rpoD (9 min), hctA (12 min), omcB (18 min), and unprocessed 16S (18 min) transcripts were relatively short, while 16S gene transcripts were more stable over time (t1/254 min).

Conclusion The detection of rpoD, hctA, omcB, and unprocessed 16S gene transcripts showed the most promising results as a potential marker for an active CT infection. Currently we are evaluating the time to clearance of mRNA molecules in patients after being treated.

Disclosure No significant relationships.

  • chlamydia
  • diagnosis

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