Title : Vaginal colonization by uropathogenic microorganisms: A key contributor to reproductive failure in mice
Abstract:
Infections of the urogenital tract are often convicted in 15% of the cases of infertility in females. The microorganisms present in vagina play an important role in determining several aspects of reproductive health. However, well designed studies on infections and infertility are still lacking. Hence, the present study was conducted with an aim to investigate the role of microorganism in infertility. This study utilized standard uropathogenic strains and on the basis of sperm-microbe interaction they were categorized into three groups: sperm agglutinating (SA) (Enterobacter aerogenes and Klebsiella pneumoniae), sperm immobilizing (SI) (Pseudomonas aeruginosa and Candida albicans), and non-sperm agglutinating/non-sperm immobilizing (NSA-NSI) (Proteus mirabilis and Streptococcus pyogenes). SA microorganisms impaired sperm motility via sperm agglutination, while SI microorganisms led to immobilization without agglutination. Conversely, NSA-NSI microorganisms had no adverse effect on sperm motility. To determine if decreased sperm motility in case of SA/SI was due to physical contact or by secretory factors, spermatozoa were incubated with washed cells or cell-free supernatant, showing that agglutination was caused by washed cells while immobilization resulted from secretory factors. SA and SI microorganisms could also reduce the viability, sperm Mg²?-ATPase activity and also led to premature acrosomal loss and induced morphological defects in spermatozoa. These effects were noticeably absent in case of NSA-NSI microorganisms. Further, to assess in-vivo relevance, female mice were intravaginally inoculated with 10?, 10?, or 10? cfu of SA, SI, or NSA-NSI microorganisms for ten consecutive days followed by mating with proven breeder males on day 12. 100% infertility in female mice inoculated with SA/SI microorganisms was observed in all groups, as evidenced by absence of pregnancy-related signs such as weight gain, abdominal distension, string of pearls, and delivery of pups. Histological analysis showed no corpus luteum in ovaries and decidual formation in uterus. In contrast, mice treated with NSA-NSI/PBS displayed normal fertility, with consistent weight gain, string of pearls, and delivery of pups. Histological analysis showed a well-defined corpus luteum, increased endometrial thickness, uterine gland proliferation, and decidual formation. To validate the hypothesis that infertility induced by SA/SI microorganisms was linked to their colonization, SA (E. aerogenes) and SI (P. aeruginosa) microorganisms were established in vagina followed by their eradication with oral administration of cefotaxime (10mg/kg). The complete eradication of E. aerogenes and P. aeruginosa from mouse vagina occurred by day 13 and 15, respectively. After clearance, mating with proven breeder male resulted in delivery of pups indicating restoration of fertility. Furthermore, no clinical or histopathological changes were observed in the reproductive organs (ovary, uterus and vagina), suggesting that colonization of the genital tract with sperm-impairing micro-organisms could be a feasible reason for female infertility. This highlighted that infertility in this case was not due to inflammation but rather the transformation of the vaginal environment into the one hostile to sperm. Protein profiling of vaginal lavages to study changes in proteins produced depicted distinct protein bands in SA/SI-inoculated mice whereas, PBS/NSA-NSI instilled group did not exhibit apparently any protein band, suggesting that microbial products might be responsible for infertility.
