This study was carried out to determine the prevalence of the bacterial agents Chlamydia trachomatis (C. trachomatis), Neisseria gonorrhoeae (N. gonorrhoeae), Mycoplasma hominis (M. hominis) and Ureaplasma urealyticum (U. urealyticum) and the conditions which may play a role in the development of female infertility, in the county of Iaşi in North-Eastern Romania. Cervical and blood samples were collected from 176 infertile women and 45 pregnant women in the third trimester. Classical methods and real time PCR were applied to each cervical sample to detect the presence of these sexually transmitted microorganisms; the ELISA method was applied to blood samples to detect C. trachomatis antibodies (IgA, IgM and IgG). The proportion of C. trachomatis IgG was significantly higher in the infertile group (23.8%) than in the pregnant group (4.4%), p < 0.05. For C. trachomatis antigen (Ag) and N. gonorrhoeae Ag no differences were observed between the two groups. The prevalence of mycoplasma genital infections was higher in the pregnant group (U. urealyticum - 53.3% and M. hominis - 20%) than in the infertile group (U. urealyticum - 39.7% and M. hominis - 7.3%). Higher rate of co-infection with C. trachomatis and mycoplasma were observed among the infertile women (25.7%) than among the pregnant women (7.7%). This combination could be involved in the appearance of pelvic inflammatory disease (PID) and its sequela, including infertility. C. trachomatis IgG determination still remains the gold standard for the diagnosis of PID and should be used as a screening test for the prediction of tubal damage in infertile women. In view of the large number of cases involving the co-existence of genital infections with C. trachomatis, M. hominis and U. urealyticum, it is clearly necessary to perform screening for all three microorganisms among all women of reproductive age but especially those who are infertile.