Bacterial Hyaluronidase Promotes Ascending GBS Infection and Preterm Birth.

UNLABELLED: Preterm birth increases the risk of adverse birth outcomes and is the leading cause of neonatal mortality. A significant cause of preterm birth is in utero infection with vaginal microorganisms. These vaginal microorganisms are often recovered from the amniotic fluid of preterm birth cases. A vaginal microorganism frequently associated with preterm birth is group B streptococcus (GBS), or Streptococcus agalactiae However, the molecular mechanisms underlying GBS ascension are poorly understood. Here, we describe the role of the GBS hyaluronidase in ascending infection and preterm birth. We show that clinical GBS strains associated with preterm labor or neonatal infections have increased hyaluronidase activity compared to commensal strains obtained from rectovaginal swabs of healthy women. Using a murine model of ascending infection, we show that hyaluronidase activity was associated with increased ascending GBS infection, preterm birth, and fetal demise. Interestingly, hyaluronidase activity reduced uterine inflammation but did not impact placental or fetal inflammation. Our study shows that hyaluronidase activity enables GBS to subvert uterine immune responses, leading to increased rates of ascending infection and preterm birth. These findings have important implications for the development of therapies to prevent in utero infection and preterm birth. IMPORTANCE: GBS are a family of bacteria that frequently colonize the vagina of pregnant women. In some cases, GBS ascend from the vagina into the uterine space, leading to fetal injury and preterm birth. Unfortunately, little is known about the mechanisms underlying ascending GBS infection. In this study, we show that a GBS virulence factor, HylB, shows higher activity in strains isolated from cases of preterm birth than those isolates from rectovaginal swabs of healthy women. We discovered that GBS rely on HylB to avoid immune detection in uterine tissue, but not placental tissue, which leads to increased rates of fetal injury and preterm birth. These studies provide novel insight into the underlying mechanisms of ascending infection.

Use of nonhuman primate models to investigate mechanisms of infection-associated preterm birth.

Preterm birth is the most important direct cause of neonatal mortality and remains a major challenge for obstetrics and global health. Intrauterine infection causes approximately 50% of early preterm births. Animal models using pregnant mice, rabbits or sheep demonstrate the key link between infection and premature birth, but differ in the mechanisms of parturition and placental structure from humans. The nonhuman primate (NHP) is a powerful model which emulates many features of human placentation and parturition. The contributions of the NHP model to preterm birth research are reviewed, emphasising the role of infections and the potential development of preventative and therapeutic strategies.

Differential host response to LPS variants in amniochorion and the TLR4/MD-2 system in Macaca nemestrina.

OBJECTIVES: Microbial-specific factors are likely critical in determining whether bacteria trigger preterm labor. Structural variations in lipopolysaccharide (LPS), a component of gram-negative bacteria, can determine whether LPS has an inflammatory (agonist) or anti-inflammatory (antagonist) effect through Toll-like receptor 4 (TLR4). Our objective was to determine whether amniochorion could discriminate between LPS variants in a nonhuman primate model. We also cloned Macaca nemestrina TLR4 and MD-2 and compared this complex functionally to the human homologue to establish whether nonhuman primates could be used to study TLR4 signaling in preterm birth. STUDY DESIGN: Amniochorion explants from M. nemestrina were stimulated with a panel of LPS variants for 24 h. Supernatants were analyzed for IL-1beta, TNF-alpha, IL-6, IL-8 and prostaglandins E2 and F2alpha. Tissue expression of TLR1, 2, 4, 6, MyD88 and NF-kappaB was studied by RT-PCR. M. nemestrina TLR4 and MD-2 genes were cloned and compared with their human counterparts in a recombinant TLR4 signaling system to determine LPS sensitivity. RESULTS: LPS variants differentially stimulated cytokines and prostaglandins, which was not related to transcriptional changes of TLR4 or other TLRs. Nearly all elements of LPS binding and TLR4 leucine-rich repeats were conserved between humans and M. nemestrina. TLR4/MD-2 signaling complexes from both species were equally sensitive to LPS variants. CONCLUSIONS: LPS variants elicit a hierarchical inflammatory response within amniochorion that may contribute to preterm birth. LPS sensitivity is similar between M. nemestrina and humans, validating M. nemestrina as an appropriate model to study TLR4 signaling in preterm birth.

Dynamic changes in fetal microchimerism in maternal peripheral blood mononuclear cells, CD4+ and CD8+ cells in normal pregnancy.

OBJECTIVE: Cell trafficking during pregnancy results in persistence of small populations of fetal cells in the mother, known as fetal microchimerism (FMc). Changes in cell-free fetal DNA during gestation have been well described, however, less is known about dynamic changes in fetal immune cells in maternal blood. We have investigated FMc in maternal peripheral blood mononuclear cells (PBMC) longitudinally across gestation. STUDY DESIGN: Thirty-five women with normal pregnancies were studied. FMc was identified in PBMC, CD4+ and CD8+ subsets employing quantitative PCR assays targeting fetal-specific genetic polymorphisms. FMc quantities were reported as fetal genome equivalents (gEq) per 1,000,000 gEq mother's cells. Poisson regression modeled the rate of FMc detection. MAIN OUTCOME MEASURE: FMc in PBMC. RESULTS: The probability of detecting one fetal cell equivalent increased 6.2-fold each trimester [Incidence Rate Ratio (IRR) 95% CI: 1.73, 21.91; p = 0.005]. Although FMc in PBMC was not detected for the majority of time points, 7 of 35 women had detectable FMc during pregnancy at one or more time points, with the majority of positive samples being from the third trimester. There was a suggestion of greater HLA-sharing in families where women had FMc in PBMC. FMc was detected in 9% of CD4+ (2/23) and 18% of CD8+ (3/25) subsets. CONCLUSIONS: FMc in PBMC increased as gestation progressed and was found within CD4+ and CD8+ subsets in some women in the latter half of gestation. A number of factors could influence cellular FMc levels including sub-clinical fetal-maternal interface changes and events related to parturition. Whether FMc during pregnancy predicts persistent FMc and/or correlates with fetal-maternal HLA relationships also merits further study.