Abstract: Objectives Mycoplasma genitalium (MG) STIs represent a global concern, especially considering its rapid ability to acquire antimicrobial resistance. The objective of this study was to investigate the epidemiology of MG and antibiotic resistance among the general female and male populations in northern Spain. Methods Between April 2019 and May 2023, individuals attending healthcare facilities for STI screening were tested for MG and macrolide and fluoroquinolone resistance mutations. Phylogenetic analysis was conducted using single-locus sequence-based typing of the mgpB gene and complemented by dual-locus sequence-based typing combining the mgpB and MG309 genes. Results Of the 6350 people tested for MG during the study period, 5269 (83%) were women. Macrolide resistance mutations were identified in 25.4% of cases, with a higher prevalence in men (40% vs 17.14%) and coinfection with Chlamydia trachomatis was found predominantly in women (18.6% vs 7.5%). Phylogenetic analysis revealed two distinct epidemiological clades strongly correlated with gender. Clade A predominantly involved men with higher rates of HIV and syphilis history and detection of resistance mutations, while clade B exclusively comprised women, with only one case of macrolide resistance. Conclusions The predominance of women in STI screening requests reflects local trends in screening patterns and underscores the importance of understanding MG infection in women in our region. The clear separation of clades suggests two independent sexual networks, with clade A representing a high-risk population and dense connectivity. This research provides a foundation for future studies on the prevalence, transmission dynamics and impact of MG infection, particularly among the female population, where understanding remains limited.
