Mycoplasma genitalium molecular typing in men with non-gonococcal urethritis discriminates between phylogenetic clusters based on sexual preference and antibiotic resistance

<b>Introduction.</b> <i>Mycoplasma genitalium</i> is a sexually transmitted bacterium associated with non-gonococcal urethritis (NGU) in men. The rising macrolide and fluoroquinolone resistance in <i>M. genitalium</i> has become a public health concern, requiring close surveillance.<b>Gap statement.</b> <i>MgpB/MG309</i> typing is commonly used to study genotype distribution and resistance patterns of <i>M. genitalium</i> in men who have sex with men (MSM); however, data for men who have sex with women (MSW) are limited.<b>Aim.</b> The aim of this study was to explore the epidemiology of <i>M. genitalium</i> based on <i>mgpB</i>/<i>MG309</i> molecular typing in isolates from men diagnosed with NGU, comparing MSM and MSW. Additionally, antibiotic resistance was evaluated to assess associations between the <i>mgpB/MG309</i> genotypes, antimicrobial resistance profiles, and epidemiological determinants.<b>Methodology.</b> A subset of previously collected <i>M. genitalium</i> isolates from men diagnosed with NGU in Amsterdam, the Netherlands, between May 2018 and November 2019 was analysed. Molecular typing was performed by sequencing relevant regions of the <i>mgpB</i> and <i>MG309</i> loci. Macrolide resistance was assessed by detecting mutations in the 23S rRNA gene via quantitative polymerase chain reaction, while fluoroquinolone resistance was determined through sequencing <i>parC</i> and <i>gyrA</i>.<b>Results.</b> A total of 62 <i>M</i>. <i>genitalium</i> samples were analysed from 33 MSM and 29 MSW. The overall macrolide and fluoroquinolone resistance was 75.8% and 24.2 %, respectively. At the <i>mgpB</i> locus, 24 sequence types (STs) were identified, with ST4 most prevalent in MSM and ST2 in MSW. The <i>MG309</i> locus revealed 12 distinct short tandem repeat numbers, with repeat 10 being most common in both groups. Phylogenetic analysis based on <i>mgpB</i> sequences revealed two clusters: cluster A included significantly more MSW, whereas cluster B predominantly comprised MSM (<i>P</i><0.001). Macrolide and fluoroquinolone resistance was significantly higher in cluster B compared with cluster A (<i>P</i><0.01 and <i>P</i><0.05, respectively).<b>Conclusion.</b> Molecular typing of <i>M. genitalium</i> revealed two clusters that differed by sexual preference and antibiotic resistance, highlighting the importance of surveillance of resistance across genotypes. The findings suggest multiclonal spread of resistance through independent mutations. Future studies using next-generation sequencing are needed to further explore the links between sexual transmission and genetic diversity in <i>M. genitalium</i>.