A group B streptococcal type VII secreted LXG toxin mediates interbacterial competition and colonization of the female genital tract.

Group B Streptococcus (GBS) asymptomatically colonizes the vagina but can opportunistically ascend to the uterus and be transmitted vertically during pregnancy, resulting in neonatal pneumonia, bacteremia and meningitis. GBS is a leading etiologic agent of neonatal infection and understanding the mechanisms by which GBS persists within the polymicrobial female genital mucosa has potential to mitigate subsequent transmission and disease. Type VIIb secretion systems (T7SSb) are encoded by Firmicutes and often mediate interbacterial competition using LXG toxins that contain conserved N-termini important for secretion and variable C-terminal toxin domains that confer diverse biochemical activities. Our recent work characterized a role for the GBS T7SSb in vaginal colonization and ascending infection but the mechanisms by which the T7SSb promotes GBS persistence in this polymicrobial niche remain unknown. Herein, we investigate the GBS T7SS in interbacterial competition and GBS niche establishment in the female genital tract. We demonstrate GBS T7SS-dependent inhibition of mucosal pathobiont Enterococcus faecalis both in vitro using predator-prey assays and in vivo in the murine genital tract and found that a GBS LXG protein encoded within the T7SS locus (herein named group B streptococcal LXG Toxin A) that contributes to these phenotypes. We identify BltA as a T7SS substrate that is toxic to E. coli and S. aureus upon induction of expression along with associated chaperones. Finally, we show that BltA and its chaperones contribute to GBS vaginal colonization. Altogether, these data reveal a role for a novel T7b-secreted toxin in GBS mucosal persistence and competition.

Intrauterine Detection of Ureaplasma Species after Vaginal Colonization in Pregnancy and Neonatal Outcome.

INTRODUCTION: Intrauterine infection with Ureaplasma species (U.spp.) is mostly a result of vaginal colonization with subsequent ascending infection and is associated with adverse pregnancy outcome. Little is known about rates and risk factors for ascending infection. Aim of the current study was to analyse the frequency of ascending U.spp. infection in vaginally colonized pregnant women delivering preterm and subsequent short- and long-term outcome of infants. METHODS: Women delivering ≤32 weeks of gestation with available data on vaginal U.spp. colonization in early pregnancy as well as amniotic and placental colonization screening during caesarean section were included. Neonatal short- and long-term outcome was analysed depending on vaginal and intrauterine colonization. RESULTS: Seventy-two women giving birth to 104 preterm infants were included. Intrauterine microbial invasion was found in 23/72 (31.9%) pregnancies. The most commonly detected organisms were U.spp. (52.2%), followed by E. coli (21.7%) and Enterococcus faecalis (17.4%). Intrauterine growth of U.spp. occurred exclusively after previous vaginal colonization in early pregnancy (42/72; 58.3%) and was found in 12/42 (28.6%) cases. Ascending U.spp. infection mainly occurred in pregnancies delivering <28 weeks after preterm rupture of membranes or preterm labour (9/17, 52.3%). Intrauterine detection of U.spp., but not vaginal colonization, was associated with a significantly higher rate of severe intraventricular haemorrhage, retinopathy of prematurity, bronchopulmonary dysplasia, and unfavourable psychomotor outcome. CONCLUSION: Ascending U.spp. infection after previous vaginal colonization occurred in almost one-third of pregnancies delivering ≤32 weeks, with particularly high rates in those <28 weeks, and was associated with adverse outcome of preterm infants.

Identification of a DNA-cytosine methyltransferase that impacts global transcription to promote group B streptococcal vaginal colonization.

Group B Streptococcus (GBS) colonizes the female reproductive tract (FRT) and causes adverse pregnancy outcomes and invasive disease following vertical transmission to the fetus or newborn. Despite this major public health burden, the mechanisms of GBS FRT colonization are understudied. A recent transposon sequencing screen identified GBS factors contributing to vaginal colonization and ascending spread, including a putative DNA-cytosine methyltransferase (Dcm). We constructed a Δdcm deletion strain and confirmed that dcm contributes to murine FRT colonization. Investigation of the evolutionary origin of the dcm gene reveals that it is widely distributed across GBS and is encoded as part of a prophage genome that displays evidence of horizontal transfer between GBS strains. We further show that Dcm contributes to 5mC methylation and global regulation of genes involved in carbohydrate metabolism, transcription regulation, and known adhesins and metabolic factors involved in GBS colonization. Interestingly, GBS genes that are induced in the presence of the highly glycosylated vaginal mucin MUC5B were significantly downregulated in the ∆dcm mutant. Furthermore, the ∆dcm mutant exhibited reduced binding to immobilized mucin and was attenuated in its ability to grow on numerous carbon sources including the carbohydrates found on mucins. While the ∆dcm mutant displayed enhanced clearance from the FRT in wild-type mice, there was no significant difference in MUC5B -/- mice, indicating that Dcm-mediated regulation requires MUC5B to promote GBS colonization. This is the first report to characterize the impact of a DNA methyltransferase on GBS gene regulation and FRT colonization. IMPORTANCE Group B Streptococcus (GBS) colonizes the female reproductive tract (FRT) in one-third of women, and carriage leads to numerous adverse pregnancy outcomes including the preterm premature rupture of membranes, chorioamnionitis, and stillbirth. The presence of GBS in the FRT during pregnancy is also the largest predisposing factor for the transmission of GBS and invasive neonatal diseases, including pneumonia, sepsis, and meningitis. The factors contributing to GBS colonization are still being elucidated. Here, we show for the first time that GBS transcription is regulated by an orphan DNA cytosine methyltransferase (Dcm). Many GBS factors are regulated by Dcm, especially those involved in carbohydrate transport and metabolism. We show that GBS persistence in the FRT is dependent on the catabolism of sugars found on the vaginal mucin MUC5B. Collectively, this work highlights the regulatory importance of a DNA methyltransferase and identifies both host and bacterial factors required for GBS colonization.

Immunization with Multiple Virulence Factors Provides Maternal and Neonatal Protection against Group B Streptococcus Serotypes.

Group B streptococcus (GBS) commonly colonizes the vaginal tract and is a leading cause of life-threatening neonatal infections and adverse pregnancy outcomes. No effective vaccine is clinically available. Conserved bacterial virulence factors, including those of GBS, have been employed as vaccine components. We investigated serotype-independent protection against GBS by intranasal immunization with six conserved GBS virulence factors (GBSV6). GBSV6 induced systemic and vaginal antibodies and T cell responses in mice. The immunity reduced mouse mortality and vaginal colonization by various GBS serotypes and protected newborn mice of immunized dams against GBS challenge. Intranasal GBSV6 immunization also provided long-lasting protective immunity and had advantages over intramuscular GBSV6 immunization regarding restricting vaginal GBS colonization. Our findings indicate that intranasal immunization targeting multiple conserved GBS virulence factors induces serotype-independent immunity, which protects against GBS infection systemically and vaginally in dams and prevents newborn death. The study presents valuable strategies for GBS vaccine development.

In Vitro and In Vivo Antibiofilm Activity of Red Onion Scales: An Agro-Food Waste.

Red onion wastes (ROW) are valuable sources of bioactive metabolites with promising antimicrobial effects. Methicillin-resistant Staphylococcus aureus (MRSA) infections are a growing risk in hospitals and communities. This study aims to investigate the in vitro and in vivo antibiofilm activities of the acidified ethanolic extract of red onion scales (RO-T) and its fractions against an MRSA vaginal colonization model. The RO-T extract, as well as its anthocyanin-rich fraction (RO-P) and flavonoid-rich fraction (RO-S), recorded a promising antibacterial activity against highly virulent strains of bacteria (MRSA, Acinetobacter baumannii, Escherichia coli and Pseudomonas aeruginosa). RO-S showed the highest antibacterial activity (MBC of 0.33 ± 0.11 mg/mL) against MRSA USA300 and significantly eradicated its biofilm formation with an IC50 of 0.003. Using a rat model, in vivo assessment on all samples, which were formulated as a hydrogel, revealed a significant reduction of MRSA bacterial load recovered from an infected vagina compared to that of the negative control group (NCG). RO-T extract and vancomycin groups recorded the highest antibacterial activity with a bacterial load 2.998 and 3.358 logs lower than the NCG, respectively. The histopathological investigation confirmed our findings. RO-T and RO-S were standardized for their quercetin content. Finally, ROW offers a new potent antibiofilm agent mostly due to its high quercetin content.

Staphylococcus aureus Fibronectin-Binding Proteins Contribute to Colonization of the Female Reproductive Tract.

Methicillin-resistant Staphylococcus aureus (MRSA) is an opportunistic pathogen and frequent colonizer of human skin and mucosal membranes, including the vagina, with vaginal colonization reaching nearly 25% in some pregnant populations. MRSA vaginal colonization can lead to aerobic vaginitis (AV), and during pregnancy, bacterial ascension into the upper reproductive tract can lead to adverse birth outcomes. USA300, the most prominent MRSA lineage to colonize pregnant individuals, is a robust biofilm former and causative agent of invasive infections; however, little is known about how it colonizes and ascends in the female reproductive tract (FRT). Our previous studies showed that a MRSA mutant of seven fibrinogen-binding adhesins was deficient in FRT epithelial attachment and colonization. Using both monolayer and multilayer air-liquid interface cell culture models, we determine that one class of these adhesins, the fibronectin binding proteins (FnBPA and FnBPB), are critical for association with human vaginal epithelial cells (hVECs) and hVEC invasion through interactions with α5ß1 integrin. We observe that both FnBPs are important for biofilm formation as single and double fnbAB mutants exhibit reduced biofilm formation on hVECs. Using heterologous expression of fnbA and fnbB in Staphylococcus carnosus, FnBPs are also found to be sufficient for hVEC cellular association, invasion, and biofilm formation. In addition, we found that an ΔfnbAB mutant displays attenuated ascension in our murine vaginal colonization model. Better understanding of MRSA FRT colonization and ascension can ultimately inform treatment strategies to limit MRSA vaginal burden or prevent ascension, especially during pregnancy and in those prone to AV.

Group B Streptococcus: Virulence Factors and Pathogenic Mechanism.

Group B Streptococcus (GBS) or Streptococcus agalactiae is a major cause of neonatal mortality. When colonizing the lower genital tract of pregnant women, GBS may cause premature birth and stillbirth. If transmitted to the newborn, it may result in life-threatening illnesses, including sepsis, meningitis, and pneumonia. Moreover, through continuous evolution, GBS can use its original structure and unique factors to greatly improve its survival rate in the human body. This review discusses the key virulence factors that facilitate GBS invasion and colonization and their action mechanisms. A comprehensive understanding of the role of virulence factors in GBS infection is crucial to develop better treatment options and screen potential candidate molecules for the development of the vaccine.

Early-Onset Neonatal Pneumococcal Sepsis: An Old but Sometimes Forgotten Pathogen.

Streptococcus pneumoniae (SP) is an uncommon but potentially serious neonatal pathogen. SP is perceived as a significant cause of mortality and morbidity in infancy; however, there are relatively few cases of neonatal sepsis recorded, with an incidence between 1% and 11%. We aim to report the spectrum of morbidity associated with SP infections in the neonatal period. Two cases of neonatal SP infection are reported. The first neonate presented with a very early onset of severe clinical disease with bacteremia and pneumonia. She developed severe pulmonary hypertension and needed intensive ventilatory support, including nitric oxide, and vasoactive drugs. An SP serotype 23B was isolated from blood cultures and bronchial secretions as well as from the mother's vaginal secretions. In the second case, the baby presented with bacteremia and meningitis. He remained hemodynamically stable and did not need respiratory support. Blood and cerebrospinal fluid cultures revealed an SP serotype 8. In both cases, the neonates were treated with vancomycin and cefotaxime. Both mothers remained well and asymptomatic during the perinatal period. These reported cases emphasize the importance of considering a wide range of microorganisms in the differential diagnosis of early-onset neonatal sepsis. Although uncommon, SP can have different clinical manifestations and cause significant diseases in newborns. Specific preventive measures against early-onset sepsis for this pathogen are yet to be implemented due to the absence of sufficient scientific evidence. For this reason, prompt and aggressive treatment remains the best therapeutic approach.

The Group B Streptococcal Adhesin BspC Interacts with Host Cytokeratin 19 To Promote Colonization of the Female Reproductive Tract.

Streptococcus agalactiae, otherwise known as Group B Streptococcus (GBS), is an opportunistic pathogen that vaginally colonizes approximately one third of healthy women. During pregnancy, this can lead to in utero infection, resulting in premature rupture of membranes, chorioamnionitis, and stillbirths. Furthermore, GBS causes serious infection in newborns, including sepsis, pneumonia, and meningitis. Previous studies have indicated that GBS antigen (Ag) I/II family proteins promote interaction with vaginal epithelial cells; thus, we hypothesized that the Ag I/II Group B streptococcal surface protein C (BspC) contributes to GBS colonization of the female reproductive tract (FRT). Here, we show that a ΔbspC mutant has decreased bacterial adherence to vaginal, ecto-, and endocervical cells, as well as decreased auto-aggregation and biofilm-like formation on cell monolayers. Using a murine model of vaginal colonization, we observed that the ΔbspC mutant strain exhibited a significant fitness defect compared to wild-type (WT) GBS and was less able to ascend to the cervix and uterus in vivo, resulting in reduced neutrophil chemokine signaling. Furthermore, we determined that BspC interacts directly with the host intermediate filament protein cytokeratin 19 (K19). Surface localization of K19 was increased during GBS infection, and interaction was mediated by the BspC variable (V) domain. Finally, mice treated with a drug that targets the BspC V-domain exhibited reduced bacterial loads in the vaginal lumen and reproductive tissues. These results demonstrate the importance of BspC in promoting GBS colonization of the FRT and that it may be targeted therapeutically to reduce GBS vaginal persistence and ascending infection. IMPORTANCE Group B Streptococcus (GBS) asymptomatically colonizes the female reproductive tract (FRT) of up to one third of women, but GBS carriage can lead to adverse pregnancy outcomes, including premature rupture of membranes, preterm labor, and chorioamnionitis. GBS colonization during pregnancy is also the largest predisposing factor for neonatal GBS disease, including pneumonia, sepsis, and meningitis. The molecular interactions between bacterial surface proteins and the host cell receptors that promote GBS colonization are vastly understudied, and a better understanding would facilitate development of novel therapeutics to prevent GBS colonization and disease. Here, we characterize the role of the GBS surface protein BspC in colonization of the FRT. We show for the first time that GBS infection induces cytokeratin 19 (K19) surface localization on vaginal epithelial cells; GBS then uses the BspC V-domain to interact with K19 to promote colonization and ascending infection. Furthermore, this interaction can be targeted therapeutically to reduce GBS carriage.

Interrelated Effects of Zinc Deficiency and the Microbiome on Group B Streptococcal Vaginal Colonization.

Group B Streptococcus (GBS) in the vaginal tract is a risk factor for preterm birth and adverse pregnancy outcomes. GBS colonization is also transient in nature, which likely reflects the contributions of pathogen determinants, interactions with commensal flora, and host factors, making this environment particularly challenging to understand. Additionally, dietary zinc deficiency is a health concern on the global scale that is known to be associated with recurrent bacterial infection and increased rate of preterm birth or stillbirth. However, the impact of zinc deficiency on vaginal health has not yet been studied. Here we use a murine model to assess the role of dietary zinc on GBS burden and the impact of GBS colonization on the vaginal microbiome. We show that GBS vaginal colonization is increased in a zinc-deficient host and that the presence of GBS significantly alters the microbial community structure of the vagina. Using machine learning approaches, we show that vaginal community turnover during GBS colonization is driven by computationally predictable changes in key taxa, including several organisms not previously described in the context of the vaginal microbiota, such as Akkermansia muciniphila. We observed that A. muciniphila increases GBS vaginal persistence and, in a cohort of human vaginal microbiome samples collected throughout pregnancy, we observed an increased prevalence of codetection of GBS and A. muciniphila in patients who delivered preterm compared to those who delivered at full term. These findings reveal the importance and complexity of both host zinc availability and native microbiome to GBS vaginal persistence. IMPORTANCE The presence of group B Streptococcus (GBS) in the vaginal tract, perturbations in the vaginal microbiota, and dietary zinc deficiency are three factors that are independently known to be associated with increased risk of adverse pregnancy outcomes. Here, we developed an experimental mouse model to assess the impact of dietary zinc deficiency on GBS vaginal burden and persistence and to determine how changes in GBS colonization impact vaginal microbial structure. We have employed unique animal, in silica metabolic, and machine learning models, paired with analyses of human cohort data, to identify taxonomic biomarkers that contribute to host susceptibility to GBS vaginal persistence. Collectively, the data reported here identify that both dietary zinc deficiency and the presence of A. muciniphila could perpetuate an increased GBS burden and prolonged exposure in the vaginal tract, which potentiate the risk of invasive infection in utero and in the newborn.

Genomic Analyses Identify Manganese Homeostasis as a Driver of Group B Streptococcal Vaginal Colonization.

Group B Streptococcus (GBS) is associated with severe infections in utero and in newborn populations, including pneumonia, sepsis, and meningitis. GBS vaginal colonization of the pregnant mother is an important prerequisite for transmission to the newborn and the development of neonatal invasive disease; however, our understanding of the factors required for GBS persistence and ascension in the female reproductive tract (FRT) remains limited. Here, we utilized a GBS mariner transposon (Krmit) mutant library previously developed by our group and identified underrepresented mutations in 535 genes that contribute to survival within the vaginal lumen and colonization of vaginal, cervical, and uterine tissues. From these mutants, we identified 47 genes that were underrepresented in all samples collected, including mtsA, a component of the mtsABC locus, encoding a putative manganese (Mn2+)-dependent ATP-binding cassette transporter. RNA sequencing analysis of GBS recovered from the vaginal tract also revealed a robust increase of mtsA expression during vaginal colonization. We engineered an ΔmtsA mutant strain and found by using inductively coupled plasma mass spectrometry that it exhibited decreased concentrations of intracellular Mn2+, confirming its involvement in Mn2+ acquisition. The ΔmtsA mutant was significantly more susceptible to the metal chelator calprotectin and to oxidative stressors, including both H2O2 and paraquat, than wild-type (WT) GBS. We further observed that the ΔmtsA mutant strain exhibited a significant fitness defect in comparison to WT GBS in vivo by using a murine model of vaginal colonization. Taken together, these data suggest that Mn2+ homeostasis is an important process contributing to GBS survival in the FRT. IMPORTANCE Morbidity and mortality associated with GBS begin with colonization of the female reproductive tract (FRT). To date, our understanding of the factors required for GBS persistence in this environment remain limited. We identified several necessary systems for initial colonization of the vaginal lumen and penetration into the reproductive tissues via transposon mutagenesis sequencing. We determined that mutations in mtsA, the gene encoding a protein putatively involved in manganese (Mn2+) transport, were significantly underrepresented in all in vivo samples collected. We also show that mtsA contributes to Mn2+ acquisition and GBS survival during metal limitation by calprotectin, a metal-chelating protein complex. We further demonstrate that a mutant lacking mtsA is hypersusceptible to oxidative stress induced by both H2O2 and paraquat and has a severe fitness defect compared to WT GBS in the murine vaginal tract. This work reveals the importance of Mn2+ homeostasis at the host-pathogen interface in the FRT.

Role of MUC5B during Group B Streptococcal Vaginal Colonization.

The female reproductive tract (FRT) is a complex environment, rich in mucin glycoproteins that form a dense network on the surface of the underlying epithelia. Group B Streptococcus (GBS) asymptomatically colonizes 25-30% of healthy women, but during pregnancy can cause ascending infection in utero or be transmitted to the newborn during birth to cause invasive disease. Though the cervicovaginal mucosa is a natural site for GBS colonization, the specific interactions between GBS and mucins remain unknown. Here we demonstrate for the first time that MUC5B interacts directly with GBS and promotes barrier function by inhibiting both bacterial attachment to human epithelial cells and ascension from the vagina to the uterus in a murine model of GBS colonization. RNA sequencing analysis of GBS exposed to MUC5B identified 128 differentially expressed GBS genes, including upregulation of the pilus island-2b (PI-2b) locus. We subsequently show that PI-2b is important for GBS attachment to reproductive cells, binding to immobilized mucins, and vaginal colonization in vivo. Our results suggest that while MUC5B plays an important role in host defense, GBS upregulates pili in response to mucins to help promote persistence within the vaginal tract, illustrating the dynamic interplay between pathogen and host. IMPORTANCE Mucin glycoproteins are a major component that contributes to the complexity of the female reproductive tract (FRT). Group B Streptococcus (GBS) is present in the FRT of 25-30% of healthy women, but during pregnancy can ascend to the uterus to cause preterm birth and fetal infection in utero. Here we show that a prominent mucin found in the FRT, MUC5B, promotes host defense by inhibiting GBS interaction with epithelial cells found in the FRT and ascension from the vagina to the uterus in vivo. In response to MUC5B, GBS induces the expression of surface expressed pili, which in turn contributes to GBS persistence within the vaginal lumen. These observations highlight the importance and complexity of GBS-mucin interactions that warrant further investigation.

Human Milk Oligosaccharides versus Streptococcus: How a Human-Made Natural Product Protects Us from Pathogens.

Group B Streptococcus (GBS) is a Gram-positive bacterium that colonizes the lower gastrointestinal tract, and in females, the urogenital tract, in up to 30% of healthy adults. However, GBS is a leading cause of mortality and morbidity in newborns due to ascending infection of the womb or by neonatal acquisition during vaginal passage. GBS neonatal disease manifests as pneumonia, sepsis, or meningitis, and an estimated 4 million newborns die each year globally. This commentary reflects on recent work by Mejia and colleagues (M. E. Mejia, S. Ottinger, A. Vrbanac, P. Babu, et al., mSphere 6:e00885-21, 2022, https://doi.org/10.1128/msphere.00885-21) that has examined human milk oligosaccharides (HMOs) as a natural product with anti-GBS activity. They show that HMOs reduce GBS vaginal colonization without impacting the normal vaginal microbiota. This study advances the possibility of using novel therapeutics to limit GBS maternal colonization and subsequent neonatal disease.

Human Milk Oligosaccharides Reduce Murine Group B Streptococcus Vaginal Colonization with Minimal Impact on the Vaginal Microbiota.

Group B Streptococcus (GBS) colonizes the vaginal mucosa of a significant percentage of healthy women and is a leading cause of neonatal bacterial infections. Currently, pregnant women are screened in the last month of pregnancy, and GBS-positive women are given antibiotics during parturition to prevent bacterial transmission to the neonate. Recently, human milk oligosaccharides (HMOs) isolated from breastmilk were found to inhibit GBS growth and biofilm formation in vitro, and women that make certain HMOs are less likely to be vaginally colonized with GBS. Using in vitro human vaginal epithelial cells and a murine vaginal colonization model, we tested the impact of HMO treatment on GBS burdens and the composition of the endogenous microbiota by 16S rRNA amplicon sequencing. HMO treatment reduced GBS vaginal burdens in vivo with minimal alterations to the vaginal microbiota. HMOs displayed potent inhibitory activity against GBS in vitro, but HMO pretreatment did not alter adherence of GBS or the probiotic Lactobacillus rhamnosus to human vaginal epithelial cells. In addition, disruption of a putative GBS glycosyltransferase (Δsan_0913) rendered the bacterium largely resistant to HMO inhibition in vitro and in vivo but did not compromise its adherence, colonization, or biofilm formation in the absence of HMOs. We conclude that HMOs are a promising therapeutic bioactive to limit GBS vaginal colonization with minimal impacts on the vaginal microenvironment. IMPORTANCE During pregnancy, GBS ascension into the uterus can cause fetal infection or preterm birth. In addition, GBS exposure during labor creates a risk of serious disease in the vulnerable newborn and mother postpartum. Current recommended prophylaxis consists of administering broad-spectrum antibiotics to GBS-positive mothers during labor. Although antibiotics have significantly reduced GBS neonatal disease, there are several unintended consequences, including altered neonatal gut bacteria and increased risk for other types of infection. Innovative preventions displaying more targeted antimicrobial activity, while leaving the maternal microbiota intact, are thus appealing. Using a mouse model, we found that human milk oligosaccharides (HMOs) reduce GBS burdens without perturbing the vaginal microbiota. We conclude that HMOs are a promising alternative to antibiotics to reduce GBS neonatal disease.

Differential impact of abnormal vaginal colonization on perinatal outcome and association with early-onset neonatal sepsis: preterm labor vs. preterm premature rupture of membrane.

OBJECTIVE: The purpose of this study was to check whether the impact of abnormal vaginal colonization on perinatal outcomes would be different in patients with preterm labor (PTL) and premature membrane rupture (PPROM). We also sought to determine the concordance rate of microorganisms isolated from the maternal vagina and neonatal blood in cases of early-onset neonatal sepsis (EONS) in PTL and PPROM. METHODS: This retrospective study included 996 singleton pregnancies who were admitted to the high risk care unit of our institution due to PTL (n = 519) or PPROM (n = 477) and underwent vaginal culture examination at admission between January 2005 and April 2019. Abnormal vaginal colonization was defined upon isolation of aerobic microorganisms. The maternal baseline characteristics, delivery, and neonatal outcomes were compared according to the presence or absence of abnormal vaginal flora, both in PTL and PPROM. RESULTS: The rate of abnormal vaginal colonization in PTL and PPROM was 17.0 and 21.4%, respectively. Both in PTL and PPROM, the gestational age at admission was lower in the abnormal vaginal colonization group (PTL, 27.2 ± 3.5 vs. 28.2 ± 3.5 weeks, p = .024; PPROM, 26.1 ± 5.3 vs. 27.5 ± 4.5 weeks, p = .007). Multivariable analysis demonstrated that the group with abnormal bacteria in PPROM but not in PTL had a significantly higher rate of EONS than the group without abnormal bacteria after adjustment for confounders including gestational age at admission (PPROM, odds ratio, OR [95% confidence interval, CI]: 4.172 [1.426-12.206]; PTL, OR [95% CI]: 0.661 [0.079-5.505]). Concordance analysis showed that the maternal vaginal bacteria colonization by Escherichia coli (5.9 vs. 0.5%, p = .033) and Staphylococcus aureus (14.3 vs. 0.2%, p = .032) in PPROM was significantly correlated with the microorganisms from the neonatal blood culture EONS cases. In PTL, no specific microorganisms showed concordance between maternal vaginal bacteria and microorganisms causing EONS. CONCLUSION: Our data showed that maternal vaginal colonization in PPROM, but not in PTL, is an independent risk factor for EONS.

Group B Streptococcus Capsular Serotype Alters Vaginal Colonization Fitness.

BACKGROUND: Group B Streptococcus (GBS) remains a leading cause of infant morbidity and mortality. A candidate vaccine targets 6 GBS serotypes, offering a potential alternative to intrapartum antibiotic prophylaxis to reduce disease burden. However, our understanding of the contributions of specific capsule types to GBS colonization and disease remains limited. METHODS: Using allelic exchange, we generated isogenic GBS strains differing only in the serotype-determining region in 2 genetic backgrounds, including the hypervirulent clonal complex (CC) 17. Using a murine model of vaginal cocolonization, we evaluated the roles of the presence of capsule and of expression of specific capsular types in GBS vaginal colonization fitness independent of other genetic factors. RESULTS: Encapsulated wild-type strains COH1 (CC17, serotype III) and A909 (non-CC17, serotype Ia) outcompeted isogenic acapsular mutants in murine vaginal cocolonization. COH1 wild type outcompeted A909. Notably, expression of type Ia capsule conferred an advantage over type III capsule in both genetic backgrounds. CONCLUSIONS: Specific capsule types may provide an advantage in GBS vaginal colonization in vivo. However, success of certain GBS lineages, including CC17, likely involves both capsule and noncapsule genetic elements. Capsule switching in GBS, a potential outcome of conjugate vaccine programs, may alter colonization fitness or pathogenesis.

Expression of virulence genes in Group B Streptococcus isolated from symptomatic pregnant women with term and preterm delivery

Aims@#Maternal vaginal Group B Streptococcus (GBS) colonization is considered a risk factor for preterm delivery and, consequently, neonatal infections. Previous studies have portrayed the important roles of these virulence factors, including hemolytic pigment, hyaluronidase (HylB), serine-rich protein (Srr) and bacterial surface adhesion of GBS (BsaB) in mediating GBS colonization and intrauterine ascending infection, causing preterm delivery. This study aimed to investigate the association between mRNA expression of virulence genes in GBS isolates obtained from symptomatic pregnant women and preterm delivery.@*Methodology and results@#GBS isolates were obtained from high vaginal swabs of 40 symptomatic pregnant women of gestational age of less than 37 weeks. RNA was extracted from these GBS isolates and RT-qPCR was performed to determine the relative mRNA expression of GBS virulence genes, including CylE (encode enzyme required for the biosynthesis of the hemolytic pigment), HylB, Srr-1 and BsaB. Socio-demographic details and obstetric history were not found to be associated with the delivery outcomes of these women. The GBS isolates from symptomatic pregnant women who delivered prematurely showed a higher expression of CylE gene and a trend towards an elevated expression of HylB gene compared to women with term delivery. Meanwhile the expression of both Srr-1 and BsaB genes was similar between symptomatic pregnant women who had term or preterm delivery.@*Conclusion, significance and impact of study@#The results suggest that following vaginal colonization, both CylE and HylB genes are likely to contribute to intrauterine ascending infection and inflammation, leading to preterm delivery in humans. These virulence factors may be targeted for the pre-clinical stages of vaccine development or therapeutic intervention.

Group B Streptococcus CAMP Factor Does Not Contribute to Interactions with the Vaginal Epithelium and Is Dispensable for Vaginal Colonization in Mice.

The Gram-positive pathogen group B Streptococcus (GBS) is a leading cause of neonatal bacterial infections, preterm birth, and stillbirth. Although maternal GBS vaginal colonization is a risk factor for GBS-associated adverse birth outcomes, mechanisms promoting GBS vaginal persistence are not fully defined. GBS possesses a broadly conserved small molecule, CAMP factor, that is co-hemolytic in the presence of Staphylococcus aureus sphingomyelinase C. While this co-hemolytic reaction is commonly used by clinical laboratories to identify GBS, the contribution of CAMP factor to GBS vaginal persistence is unknown. Using in vitro biofilm, adherence and invasion assays with immortalized human vaginal epithelial VK2 cells, and a mouse model of GBS vaginal colonization, we tested the contribution of CAMP factor using GBS strain COH1 and its isogenic CAMP-deficient mutant (Δcfb). We found no evidence for CAMP factor involvement in GBS biofilm formation, or adherence, invasion, or cytotoxicity toward VK2 cells in the presence or absence of S. aureus. Additionally, there was no difference in vaginal burdens or persistence between COH1 and Δcfb strains in a murine colonization model. In summary, our results using in vitro human cell lines and murine models do not support a critical role for CAMP factor in promoting GBS vaginal colonization. IMPORTANCE Group B Streptococcus (GBS) remains a pervasive pathogen for pregnant women and their newborns. Maternal screening and intrapartum antibiotic prophylaxis to GBS-positive mothers have reduced, but not eliminated GBS neonatal disease, and have not impacted GBS-associated preterm birth or stillbirth. Additionally, this antibiotic exposure is associated with adverse effects on the maternal and neonatal microbiota. Identifying key GBS factors important for maternal vaginal colonization will foster development of more targeted, alternative therapies to antibiotic treatment. Here, we investigate the contribution of a broadly conserved GBS determinant, CAMP factor, to GBS vaginal colonization and find that CAMP factor is unlikely to be a biological target to control maternal GBS colonization.

Group B streptococcal infection of the genitourinary tract in pregnant and non-pregnant patients with diabetes mellitus: An immunocompromised host or something more?

Group B Streptococcus (GBS), also known as Streptococcus agalactiae is a Gram-positive bacterium commonly encountered as part of the microbiota within the human gastrointestinal tract. A common cause of infections during pregnancy, GBS is responsible for invasive diseases ranging from urinary tract infections to chorioamnionitis and neonatal sepsis. Diabetes mellitus (DM) is a chronic disease resulting from impaired regulation of blood glucose levels. The incidence of DM has steadily increased worldwide to affecting over 450 million people. Poorly controlled DM is associated with multiple health comorbidities including an increased risk for infection. Epidemiologic studies have clearly demonstrated that DM correlates with an increased risk for invasive GBS infections, including skin and soft tissue infections and sepsis in non-pregnant adults. However, the impact of DM on risk for invasive GBS urogenital infections, particularly during the already vulnerable time of pregnancy, is less clear. We review the evolving epidemiology, immunology, and pathophysiology of GBS urogenital infections including rectovaginal colonization during pregnancy, neonatal infections of infants exposed to DM in utero, and urinary tract infections in pregnant and non-pregnant adults in the context of DM and highlight in vitro studies examining why DM might increase risk for GBS urogenital infection.

Colonization Rate of Potential Neonatal Disease-Causing Bacteria, Associated Factors, and Antimicrobial Susceptibility Profile Among Pregnant Women Attending Government Hospitals in Hawassa, Ethiopia.

INTRODUCTION: Vaginal colonization with some species of bacteria during the last term of pregnancy can affect the health of fetuses and newborns resulting in high morbidity and mortality among newborns. OBJECTIVE: The aim of this study was to determine the colonization rate of potential neonatal disease-causing bacteria, factors associated with colonization rate, and the antimicrobial susceptibility profile of bacteria among pregnant women. METHODS: Institution-based cross-sectional study was conducted on pregnant women from October 13 to December 28, 2020, at government hospitals located in Hawassa, Ethiopia. Background data were captured using a structured questionnaire. Vaginal swabs were collected to isolate bacteria using the standard method. Antimicrobial susceptibility test was performed using the modified Kirby-Bauer disc diffusion method. Data were analyzed using SPSS. Factors that could predict vaginal colonization with potential neonatal disease-causing bacteria were determined using logistic regression. RESULTS: Overall bacterial colonization rate among pregnant women was 271 (98.9%) 95 CI (97.4‒100.1). The prevalence of potential neonatal disease-causing bacteria was 95 (34.7%) 95 CI (28.8‒40.1). The proportion of potential neonatal disease-causing bacteria were as follows: Escherichia coli (n=82, 29.9%), Acinetobacter species (n=9, 3.3%), Staphylococcus aureus (n=7. 2.6%), and Klebsiella pneumoniae (n=4, 1.5%). Pregnant women with a gestational age of 38‒40 weeks were 1.9 times (AOR= 1.9, 95% CI= 1.0-3.4, p=0.04) were more likely to be colonized by potential neonatal disease-causing bacteria. All E. coli, Klebsiella species, and Acinetobacter species were susceptible to gentamicin and imipenem. All S. aureus were susceptible to penicillin, tetracycline, clindamycin, and erythromycin. CONCLUSION: High proportion of pregnant women in this study were colonized with potential neonatal disease-causing bacteria. E. coli was the predominant bacteria. Most bacteria isolated in this study were susceptible to antimicrobial agents tested. Gestational age was significantly associated with the colonization rate of potential neonatal disease-causing bacteria.