We investigated the antimicrobial susceptibilities of mycoplasmas in Gabonese men and women. A total of 1,332 men and women were included in the study. Sperm, urine, ureteral or vaginal swabs were collected from the subjects. Mycoplasmas identification and antimicrobial susceptibility to azithromycin, clarithromycin, erythromycin, josamycin, pristinamycin, doxycycline, tetracycline, ofloxacin and ciprofloxacin were tested using the Mycoplasma IST 2 kit. 794 subjects were positive for Mycoplasma. Respectively, 1.6 % and 82.24 % of subjects were singly infected with M. hominis and Ureaplasma urealyticum and 15.87 % had a mixed infection. M. hominis isolates were resistant to erythromycin and had an intermediate (I) to resistant (R) profile to azithromycin and clarithromycin. 84.6 % of M. hominis strains were sensitive (S) to josamycin and pristinamycin. 30.8 % and 92.3 % of M. hominis strains were sensitive to tetracycline and doxycycline, respectively. 76.9 and 84.6 % of M. hominis isolates were sensitive to ciprofloxacin and ofloxacin, respectively. The sensitivity rates of U. urealyticum strains were 45.23 %, 47.7 %, 63.84 %, 90.8 % and 92 % for azithromycin, erythromycin, clarithromycin, pristinamycin and josamycin, respectively. U. urealyticum strains showed 62.2 % and 79.7 % sensitivity to tetracycline and doxycycline, respectively. The resistance rates to azithromycin, clarithromycin and erythromycin for samples with mixed infection were 72.8 %, 84.7 % and 85.6 %, respectively. Josamycin and pristinamycin were 81.5 % effective on samples with mixed infection. The sensitivity rates of samples with mixed infection to tetracycline, doxycycline, ciprofloxacin and ofloxacin were 32 %, 69.6 %, 8.9 % and 18.5 %, respectively. Sub-Saharan Africa needs to use antibiotics rationally, as falling to do so would compromise the management of infectious diseases.