Molecular Detection of Mutations Associated with Antimicrobial Resistance

Detection of mutations associated with antimicrobial resistance in Mycoplasma genitalium by PCR.

• Non-gonococcal urethritis (NGU)

Urethral/Urogenital swab, vaginal swab, rectal swab, urine, Aptima tube remnant, ESwab, or DNA extracts. Other specimen types may be accepted with proper rationale for testing or by prior approval from the Streptococcus and STI Unit.

Specimens may be subject to rejection if they are not the appropriate sample type, have insufficient volume, or are not accompanied by relevant patient information.

Sterile cotton swab or Dacron polyester swab or flocked nylon swab.

Place swabs in at least 1 mL appropriate transport medium (such as Universal Transport Medium).   Store samples refrigerated or frozen until shipped for testing.  Ship frozen samples on dry ice and refrigerated samples on cold packs.

Shipping of specimens shall be done by a TDG certified individual in accordance with TDG regulations. For additional information regarding classification of specimens for the purposes of shipping, consult either Part 2 Appendix 3 of the TDG Regulations or section 3.6.2 of the IATA Dangerous Goods Regulations as applicable.

Infected with Mycoplasma genitalium.

Streptococcus and STI Unit requisition form, clearly specify organism testing request on the requisition form. Append all relevant clinical background information and testing performed.

Duplicate samples should not be submitted without prior consent as they may not be processed. Isolates collected from the same patient within 3 weeks of each other are considered duplicate.

DNA from all urogenital mycoplasma samples received at the NML for PCR testing is initially extracted using a commercially-available kit. Results are based upon PCR assays that detect mutations in 23S ribosomal RNA (associated with azithromycin resistance) and parC and gyrA (associated with moxifloxacin resistance).  The presence of the specific mutations predicts resistance to the associated antimicrobial. Sample is also tested for RNase-P to ensure DNA extraction was successful.

A negative PCR result indicates the absence of detectable bacterial DNA in the specimen, but does not rule-out infection as false-negative results may occur due to: inhibition of PCR reactions, sequence variability underlying the target primers and probes, or the presence of bacterial DNA in quantities less than the limit of detection of the respective assay.

As with any laboratory test, the results of the test should be interpreted with consideration of all of the laboratory and clinical findings available.

14 calendar days. Urgent specimens will be granted priority status, and completed as soon as possible.

1. Jensen JS. Protocol for the detection of Mycoplasma genitalium by PCR from clinical specimens and subsequent detection of macrolide resistance-mediating mutations in region V of the 23S rRNA gene. Methods in Molecular Biology. 2012; 903:129-139.
2. Shimada Y, Deguchi T, Nakane K, Masue T, et al. Emergence of clinical strains of Mycoplasma genitalium harbouring alterations in ParC associated with fluorouinolone resistance. 2010. Int J Antimicrob Agents. Sep;36(3):255-8.

Pathogen Safety Data Sheets: Infectious Substances – Mycoplasma genitalium

Canadian Guidelines on Sexually Transmitted Infections