Background Macrolide-resistance in Mycoplasma genitalium (MG) exceeds 50% in many regions and quinolone-resistance is increasing. We recently reported that resistance-guided therapy (RGT) using doxycycline followed by sitafloxacin or 2.5g-azithromycin cured 92% and 95% of macrolide-resistant and macrolide-susceptible infections, respectively. We now present the data on RGT using doxycycline-moxifloxacin, the regimen recommended in international guidelines, and extend the data on the efficacy of doxyxycline-2.5g azithromycin and subsequent de novo macrolide-resistance. Methods Patients attending Melbourne Sexual Health Centre between 2017-2018 with STI-related syndromes were treated with doxycycline for 7 days and recalled if positive for MG. Macrolide-susceptible cases then received 2.5g azithromycin (1g, then 500mg daily for 3 days) and resistant cases received moxifloxacin (400 mg daily, 7 days). Test of cure (TOC) was recommended 14-28 days post-completion of antimicrobials. Adherence and adverse effects were recorded. Results A total of 383 patients (81 females/106 heterosexual males/196 men-who-have-sex-with-men) were included. Microbial cure following doxycycline-azithromycin was 95.4% (95% CI 89.7-98.0) and doxycycline-moxifloxacin was 92.0%(88.1-94.6). De novo macrolide-resistance was detected in 4.6% of cases. Combining doxycycline-azithromycin data with our prior RGT study (n=186) yielded a pooled cure of 95.7% (91.6-97.8). ParC mutations implicated in moxifloxacin failure were present in 15-22% of macrolide-resistant cases at baseline. Conclusion These findings support the inclusion of moxifloxacin in resistance-guided strategies and extend the evidence for use of 2.5g azithromycin, and presumptive use of doxycycline. These data provide an evidence-base for current UK, Australian and European guidelines for the treatment of MG, an STI which is increasingly challenging to cure.

Background The basis of fluoroquinolone treatment failure for Mycoplasma genitalium is poorly understood. Methods To identify mutations associated with failure we sequenced key regions of the M. genitalium parC and gyrA genes for patients undergoing sequential therapy with doxycycline-moxifloxacin (201 patients, including 21 failures) or doxycycline-sitafloxacin (126 patients, including 13 failures). Results The parC G248T/S83I mutation was more common among patients who failed sequential doxycycline-moxifloxacin (present in 76.2% of failures vs 7.8% cures, p<0.001) and doxycycline-sitafloxacin (50% failures vs 16.8% cures, p = 0.014). Doxycycline-sitafloxacin was more efficacious than doxycycline-moxifloxacin against infections carrying the ParC S83I mutation. Infections with a ParC S83I were more likely to fail treatment with a concurrent gyrA mutation (M95I or D99N) (for doxycycline- moxifloxacin group p = 0.067, for doxycycline-sitafloxacin group p = 0.0093), suggesting an additive effect. Conclusions This study indicates that parC G248T/S83I mutations contribute to failure of moxifloxacin and sitafloxacin, and will inform the development of quinolone resistance assays needed to ensure optimal selection of antimicrobials for M. genitalium.

Screening for Chlamydia trachomatis and Neisseria gonorrhoeae at the pharyngeal, urogenital, and anorectal sites is recommended for men who have sex with men (MSM). Combining the three individual-site samples into a single pooled sample could result in significant cost savings, provided there is no significant sensitivity reduction. The aim of this study was to examine the sensitivity of pooled samples for detecting chlamydia and gonorrhea in asymptomatic MSM using a nucleic acid amplification test. Asymptomatic MSM who tested positive for chlamydia or gonorrhoea were invited to participate. Paired samples were obtained from participants prior to administration of treatment. To form the pooled sample, the anorectal swab was agitated in the urine specimen transport tube and then discarded. The pharyngeal swab and 2 ml of urine sample were then added to the tube. The difference in sensitivity between testing of pooled samples and individual-site testing was calculated against an expanded gold standard, where an individual is considered positive if either pooled-sample or individual-site testing returns a positive result. All samples were tested using the Aptima Combo 2 assay. A total of 162 MSM were enrolled in the study. Sensitivities of pooled-sample testing were 86% (94/109; 95% confidence interval [CI], 79 to 92%]) for chlamydia and 91% (73/80; 95% CI, 83 to 96%) for gonorrhea. The sensitivity reduction was significant for chlamydia (P = 0.02) but not for gonorrhea (P = 0.34). Pooling caused 22 infections (15 chlamydia and 7 gonorrhoea) to be missed, and the majority were single-site infections (19/22). Pooling urogenital and extragenital samples from asymptomatic MSM reduced the sensitivity of detection by approximately 10% for chlamydia but not for gonorrhea.