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Forward Genetic Approaches in Chlamydia trachomatis
Published on: October 23, 2013
Inhibition of PCR in genital and urine specimens submitted for Chlamydia trachomatis testing
1Department of Pathology and Laboratory Medicine, Ottawa General Hospital, University of Ottawa, Ontario, Canada. btoye@ogh.on.ca
Abstract:
We determined the frequency of PCR inhibition in genital and urine specimens submitted for Chlamydia trachomatis testing using the internal control DNA provided with the COBAS AMPLICOR C. trachomatis test and assessed methods to remove it. Inhibition occurred in 65 of 906 (7%) cervical swabs, 23 of 51 (45%) urethral swabs, and 2 of 175 (1.1%) urine samples. Overall, inhibition was eliminated in processed specimens after storage at 4 degrees C in 77 of 90 specimens (86%), freezing at -70 degrees C in 59 of 82 specimens (72%), storage at 4 degrees C followed by either 1:100 dilution in 37 of 43 specimens (86%) or 1:10 dilution in 42 of 47 specimens (89%), and phenol-chloroform extraction in 79 of 80 specimens (99%). No positive specimens were missed due to inhibition. We conclude that PCR inhibition is rare with urine specimens and infrequent with endocervical swabs but occurs frequently with urethral swabs. The frequency of PCR inhibition may be significantly reduced by methods which can be easily incorporated into the processing of specimens.
Insights
PCR inhibition is rare in urine and infrequent in cervical samples for Chlamydia trachomatis testing, but common in urethral swabs. Several methods effectively reduce PCR inhibition, ensuring accurate test results.
Area of Science:
- Clinical Microbiology
- Molecular Diagnostics
Background:
- Polymerase Chain Reaction (PCR) is crucial for detecting Chlamydia trachomatis.
- Specimen inhibition can lead to false-negative results in PCR assays.
Purpose of the Study:
- To determine the frequency of PCR inhibition in genital and urine specimens for Chlamydia trachomatis testing.
- To evaluate methods for removing PCR inhibition.
Main Methods:
- Utilized internal control DNA with the COBAS AMPLICOR C. trachomatis test.
- Assessed inhibition in cervical swabs, urethral swabs, and urine samples.
- Evaluated various inhibition removal methods: cold storage, freezing, dilution, and phenol-chloroform extraction.
Main Results:
- PCR inhibition occurred in 7% of cervical swabs, 45% of urethral swabs, and 1.1% of urine samples.
- Phenol-chloroform extraction was highly effective (99%) in eliminating inhibition.
- Cold storage (86%) and dilution (86-89%) also demonstrated significant effectiveness.
Conclusions:
- PCR inhibition is infrequent in urine and cervical samples but frequent in urethral samples for C. trachomatis detection.
- Simple methods like cold storage, dilution, and phenol-chloroform extraction can significantly reduce PCR inhibition.
- No positive specimens were missed due to inhibition in this study.
