**5. Staphylococcal infection and female infertility**

Poland group associated *S. aureus* infection with abnormal semen parameters or other

Besides these prospective studies, several *in vitro* studies also support the effect of *Staphylo‐ coccus* on sperm activity and its relation with infertility. It was revealed that infection of the normal human ejaculated spermatozoa with *S. haemolyticus* profoundly impacted the archi‐ tecture and integrity of the sperm plasma membrane. Bacterial infection serves as a contribu‐ ting factor for severe injury of sperm membrane stability and mitochondrial activity with potential consequences of male fertility [29]. In addition, exposure of ejaculated spermatozoa to *S. haemolyticus* was found to trigger a simultaneous decrease in the percentage of sperm with normal ΔΨm and an increase in the proportion of sperm with Annexin V staining, indicative of apoptotic cells. The data suggested a determinant role of staphylococcal infection in sperm fate [30]. Despite the above findings, the role for staphylococci in male infertility has remained somewhat controversial. A previous study showed that, although *Staphylococcus* sp. was the most common bacteria isolated in 299 asymptomatic men undergoing fertility evaluation, the

In an effort to understand the mechanism whereby staphylococci modulate sperm activity, investigators currently identified some of the key molecules that have profound effect on sperm activity. Kaur and Prabha *et al.* firstly identified sperm agglutinating factor (SAF). Based on the observation that *S. aureus* can adhere to the sperm head as well as sperm tail and agglutinate mouse spermatozoa, the group finally isolated a protein with a molecular weight of approximately 57 kDa from *S. aureus* and implicated the protein in control of sperm motility and survival. Moreover, SAF potentially affects various sperm parameters such as Mg2+ dependent ATPase activity, acrosome status, and apoptosis. In support of this, a profound morphological alteration occurs in the spermatozoa upon binding with SAF, as detected with scanning electron microscopy. Also, SAF has a spermicidal effect at high concentrations and may have the potential to function as active ingredient of a vaginal contraceptive. Further studies indicate that the interaction of SAF with spermatozoa is receptor mediated, and the receptor has been isolated and purified from human spermatozoa. This sperm surface receptor component showed homology to glutamate decarboxylase and major histocompatibility complex (MHC) class I molecule [32]. Intriguingly, the receptor was shown to be able to counteract the detrimental effects of SAF on sperm parameters and alleviate SAF-induced infertility in mice [33]. In addition, Prabha *et al.* isolated the SAF from an *E. coli* strain. This kind of SAF also leads to sperm agglutination, the compromised Mg2+-dependent ATPase

In 2009, another protein with sperm regulatory effect, named sperm immobilization factor (SIF), was identified from *S. aureus*. SIF is a protein with the molecular weight of approximately 20 kDa. Similar to SAF, SIF causes multiple defects in the head, midpiece, neck, and tail region of human spermatozoa [34]. It can completely inhibit Mg2+ ATPase activity of spermatozoa at the concentration of 100 μg/ml and reduce calcium ionophore–induced acrosome reaction. The interaction between SIF and spermatozoa is also ligand-receptor dependent. The analysis by matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) showed that the receptor shares sequence similarity with MHC class II antigen. Interestingly, SIF has been

bacterial counts were not correlated with semen parameters [31].

surogenital tract infection [28].

164 Genital Infections and Infertility

activity, and spermatozoa apoptosis.

Bacterial infection in female reproductive system, such as staphylococcal infection, can profoundly affect all the phases of a woman's life in relation to the period of pre-pregnancy, fertilization, pregnancy, and reproduction. It has been demonstrated that BV is the most common lower genital tract disorder among women of reproductive age. In particular, staphylococcal infection is presumed to be a contributing factor for the adverse pregnancy outcomes and female infertility. Staphylococcal infection causes malodorous vaginal discharge and is causally associated with sexually transmitted infections. Also, it has been implicated in the development of endometritis (endometriosis), another crucial factor to female infertility.

#### **5.1. Inflammatory response and hypothalamic endocrine**

An immune/inflammatory challenge is considered as an important factor that impinges the reproduction process in animals and humans [38]. It can affect reproduction at the level of the hypothalamus, pituitary gland, or gonads. Nonetheless, the major impact is thought to occur within the brain or the pituitary gland [39]. Bacterial endotoxins can trigger the release of cytokines and other immune mediators in the hypothalamus, where the luteinizing hormone (LH)-releasing hormone and gonadotropin-releasing hormone (GnRH) neurons are located [40]. Numerous *in vitro* and *in vivo* studies showed that the immune stress and the subsequent action of proinflammatory cytokines have a profound impact on the secretory activity of GnRH and LH neurons in the hypothalamus [41]. These interconnections between the immune and the neuroendocrine systems are suggested to be based on the mutual sharing of receptors and mediators.

As described above, staphylococcal infection either in peripheral or directly in vagina can arouse strong immune/inflammatory reaction systematically or locally. The released cytokines and chemokines then act on the pituitary gland and reproductive organs, which may finally lead to menoxenia, irregular ovulation, and infertility. Evidences have shown that staphylo‐ coccal infection generates a large quantity of cytokines in female reproductive system. These mediators have an important role in the control of reproductive neuroendocrine, ovarian physiology, fetal implantation and development, and placenta function. Previous study revealed a correlation between BV, elevated IL-1β and IL-8, and idiopathic infertility, sug‐ gesting that abnormal vaginal flora and the vaginal inflammatory response may be responsible for the idiopathic infertility in women undergoing *in vitro* fertilization [42].

#### **5.2. Premature ovarian failure**

Female mammals are born with a finite number of oocytes that gradually decreases during prepubertal development and adult life [43]. Each oocyte is encircled by somatic granulosa cells (GCs) to form the basic functioning unit of the ovary—the follicle. The size of the oocytes at birth and the rate of endowment depletion dominate the ovarian functional lifespan. On the other hand, programmed cell death (apoptosis) has been considered one of the most prevalent mechanisms that contribute to the age-related exhaustion of oocytes. Therefore, a precise balance has to be achieved between prosurvival and proapoptotic molecules to maintain the final destiny of the follicle [44, 45].

It is well recognized that immune/inflammatory response participates in many aspects of reproductive physiology, such as ovulation, menstruation, and implantation. Recent studies suggest that the inflammatory stress caused by staphylococcal infection may also affect ovarian reserve and cyclicity in women. Proinflammatory reaction, such as its correspondent neuro‐ transmitter secretion, inflammatory gene transcription, and signaling pathway activation, was thought to play essential roles in the process. Moreover, the production of neurotransmitter, such as sphingolipid ceramide, further acts as a second messenger to promote age-related apoptosis of oocytes. Evidence showed that lower ceramide levels observed in acid sphingo‐ myelinase-deficient mice resulted in a larger postnatal pool of oocytes compared with their wild-type counterparts. Conversely, Bax-null female mice exhibited to extend the ovarian lifespan [46]. These data may provide novel perspectives on the regulation of oocyte dynamics by bacterial infection and link the critical biological processes such as infection, inflammation, cell survival, and female fertility.

#### **5.3. Bacterial vaginosis and endometritis**

BV is a polymicrobial syndrome mainly due to an imbalance of vaginal microbiota. Coloniza‐ tion and proliferation of staphylococci is supposed to be one of the reasons for the increase in pathogenic bacteria, anaerobic bacteria, or microaerophiles. During the pathogenesis of BV, the overgrowth of anaerobes promotes the production of noxious substances, such as polya‐ mines and other compounds. These metabolic products may further trigger the release of proinflammatory cytokines IL-1β and IL-8 and thus cause tissue damage and physiological imbalance [47]. BV can directly affect congenital formation and female fertility as an ascending dissemination of the related bacteria species proved to cause tubal factor infertility.

Another complication accompanied by vaginal staphylococcal infection is chronic endome‐ tritis (CE), a local inflammatory disease characterized by unusual plasmacytic infiltration in the endometrial stromal areas [48]. CE frequently happens in the later stage of the infection or under repeated infection. It tends to be neglected in gynecologic practice because of its less apparent symptom and the requirement of time-consuming histopathologic examinations. In most cases, the diagnosis is made based on gynecological indications, such as abnormal uterine bleeding (AUB) and infertility [49]. In a study on 64 CE patients and 28 healthy women, the biochemical analysis revealed that IL-6, IL-1β, and TNF-α levels were markedly higher in menstrual effluents of women with CE when compared with control subjects [50]. It is suggested that the infection and inflammation alter the endometrial cytokine profiles, which may further impair endometrial function and lead to menstrual abnormalities and reduced embryo receptivity [51]. Moreover, the proteomic analysis identified the key signaling pathways involved in inflammation and oxidative stress are closely related with carcinoge‐ netic processes. These studies associate the onset of CE with female infertility, obstetric and neonatal abnormity, and complications. The altered endometrial gene expression may explain the impaired endometrial receptivity and endometrial hyperplastic lesions observed in women affected by CE [52].

#### **5.4. Abnormal fetal implantation**

revealed a correlation between BV, elevated IL-1β and IL-8, and idiopathic infertility, sug‐ gesting that abnormal vaginal flora and the vaginal inflammatory response may be responsible

Female mammals are born with a finite number of oocytes that gradually decreases during prepubertal development and adult life [43]. Each oocyte is encircled by somatic granulosa cells (GCs) to form the basic functioning unit of the ovary—the follicle. The size of the oocytes at birth and the rate of endowment depletion dominate the ovarian functional lifespan. On the other hand, programmed cell death (apoptosis) has been considered one of the most prevalent mechanisms that contribute to the age-related exhaustion of oocytes. Therefore, a precise balance has to be achieved between prosurvival and proapoptotic molecules to maintain the

It is well recognized that immune/inflammatory response participates in many aspects of reproductive physiology, such as ovulation, menstruation, and implantation. Recent studies suggest that the inflammatory stress caused by staphylococcal infection may also affect ovarian reserve and cyclicity in women. Proinflammatory reaction, such as its correspondent neuro‐ transmitter secretion, inflammatory gene transcription, and signaling pathway activation, was thought to play essential roles in the process. Moreover, the production of neurotransmitter, such as sphingolipid ceramide, further acts as a second messenger to promote age-related apoptosis of oocytes. Evidence showed that lower ceramide levels observed in acid sphingo‐ myelinase-deficient mice resulted in a larger postnatal pool of oocytes compared with their wild-type counterparts. Conversely, Bax-null female mice exhibited to extend the ovarian lifespan [46]. These data may provide novel perspectives on the regulation of oocyte dynamics by bacterial infection and link the critical biological processes such as infection, inflammation,

BV is a polymicrobial syndrome mainly due to an imbalance of vaginal microbiota. Coloniza‐ tion and proliferation of staphylococci is supposed to be one of the reasons for the increase in pathogenic bacteria, anaerobic bacteria, or microaerophiles. During the pathogenesis of BV, the overgrowth of anaerobes promotes the production of noxious substances, such as polya‐ mines and other compounds. These metabolic products may further trigger the release of proinflammatory cytokines IL-1β and IL-8 and thus cause tissue damage and physiological imbalance [47]. BV can directly affect congenital formation and female fertility as an ascending

Another complication accompanied by vaginal staphylococcal infection is chronic endome‐ tritis (CE), a local inflammatory disease characterized by unusual plasmacytic infiltration in the endometrial stromal areas [48]. CE frequently happens in the later stage of the infection or under repeated infection. It tends to be neglected in gynecologic practice because of its less apparent symptom and the requirement of time-consuming histopathologic examinations. In most cases, the diagnosis is made based on gynecological indications, such as abnormal uterine bleeding (AUB) and infertility [49]. In a study on 64 CE patients and 28 healthy women, the

dissemination of the related bacteria species proved to cause tubal factor infertility.

for the idiopathic infertility in women undergoing *in vitro* fertilization [42].

**5.2. Premature ovarian failure**

166 Genital Infections and Infertility

final destiny of the follicle [44, 45].

cell survival, and female fertility.

**5.3. Bacterial vaginosis and endometritis**

In rodents and humans, implantation is the first coordinated encounter between mother and baby. The abnormal implantation and placentation may lead to various dysfunctions through‐ out the pregnancy. Pre-eclampsia (PE) is a pregnancy-induced disorder characterized by hypertension and proteinuria. It is estimated to affect about 8% of pregnancies and is thought to be unique to humans [53]. The etiology of PE still remains poorly understood, but abnormal placentation is proved to be a major reason for this disease. In addition, local infection and immune responses are critically involved in the process of implantation [54]. Increased placental secretion of proinflammatory cytokines as well as the angiogenic regulators has been implicated in the widespread maternal endothelial dysfunction and the development of PE. It has been shown that cytokines produced within the uterine microenvironment can alter trophoblast action [55], impair implantation, and placenta vascularization, which may account for the recurrent miscarriage in women [56]. Elevated circulating IL-15 levels, proportional to severity of diseases, have been detected in the serum of PE mothers when compared with healthy controls [57]. Also, IL-11 is also proved to be a contributing factor for the impaired trophoblast invasion, spiral artery remodeling, and altered placental labyrinth morphology. These functional abnormities further lead to the development of PE-like features, such as elevated systolic blood pressure (SBP), proteinuria, and kidney glomerular pathology. Another proinflammatory cytokine, interferon (IFN)-γ, was also elevated in plasma, circulating leukocytes, and decidua of patients with PE [58]. Besides this, decidual natural killer (dNK) cells, the predominant immune cell, coincide with the decidualization at the maternal-fetal interface in human and mice. All of these studies point to a critical role of immune/inflamma‐ tory response during the implanting and placental development. More importantly, vaginal *Staphylococcus* infection, an infectious agent frequently observed in the uterus, may participate in the development of implantation disorder presumably by the induction of local immune reaction.
