GBS Vaginal Colonization Research Articles

The Streptococcus agalactiae LytSR two-component regulatory system promotes vaginal colonization and virulence in vivo.

Streptococcus agalactiae (or group B Streptococcus, GBS) is a leading cause of neonatal sepsis and meningitis globally. To sense and respond to variations in its environment, GBS possesses multiple two-component regulatory systems (TCSs), such as LytSR. Here, we aimed to investigate the role of LytSR in GBS pathogenicity. We generated an isogenic lytS knockout mutant in a clinical GBS isolate and used a combination of phenotypic in vitro assays and in vivo murine models to investigate the contribution of lytS to the colonization and invasive properties of GBS. Deletion of the lytS gene in the GBS chromosome resulted in significantly higher survival rates in mice during sepsis, accompanied by reduced bacterial loads in blood, lung, spleen, kidney, and brain tissues compared to infection with the wild-type strain. In a mouse model of GBS vaginal colonization, we also observed that the lytS knockout mutant was cleared more readily from the vaginal tract compared to its wild-type counterpart. Interestingly, lower levels of proinflammatory cytokines were found in the serum of mice infected with the lytS mutant. Our results demonstrate that the LytSR TCS plays a key role in GBS tissue invasion and pathogenesis, and persistence of mucosal colonization.IMPORTANCEStreptococcus agalactiae (group B Streptococcus, or GBS) is a common commensal of the female urogenital tract and one of WHO's priority pathogens. The bacterium has evolved mechanisms to adapt and survive in its host, many of which are regulated via two-component signal transduction systems (TCSs); however, the exact contributions of TCSs toward GBS pathogenicity remain largely obscure. We have constructed a TCS lytS-deficient mutant in a CC-17 hypervirulent GBS clinical isolate. Using murine models, we showed that LytSR regulatory system is essential for vaginal colonization via promoting biofilm production. We also observed that lytS deficiency led to significantly attenuated virulence properties and lower levels of proinflammatory cytokines in blood. Our findings are of significant importance in that they unveil a previously unreported role for LytSR in GBS and pave the way toward a better understanding of its ability to transition from an innocuous commensal to a deadly pathogen.

Performances of two rapid LAMP-based techniques for the intrapartum detection of Group B Streptococcus vaginal colonization

PurposeGroup B Streptococcus (GBS) is the leading cause of invasive infections in newborns. The prevention of GBS neonatal disease relies on the administration of an intrapartum antibiotic prophylaxis to GBS-colonized women. In recent years, rapid intrapartum detection of GBS vaginal colonization using real-time nucleic acid amplification tests (NAATs) emerged as an alternative to antenatal culture screening methods.MethodsWe compared the performances of two loop-mediated isothermal amplification (LAMP) tests, the Ampliflash® GBS and the PlusLife® GBS tests, to standard culture for GBS detection in vaginal specimens from pregnant women. The study was conducted from April to July 2023 in a French hospital of the Paris area.ResultsA total of 303 samples were analyzed, including 85 culture-positive samples (28.1%). The Ampliflash® GBS test and the PlusLife® GBS tests gave a result for 100% and 96.3% tests, respectively. The performances of the tests were as follows: sensitivity 87.1% (95% confidence interval (CI) 78.3–92.6) and 98.7% (95% CI 93.0-99.8), specificity 99.1% (95% CI 96.7–99.8), and 91.9% (95% CI 87.3–95.0), respectively. False negative results of the Ampliflash® GBS test correlated with low-density GBS cultures. Time-to-results correlated with GBS culture density only for the PlusLife® GBS test (p < 0.001).ConclusionBoth techniques provide excellent analytical performances with high sensitivity and specificity together with a short turnaround time and results available in 10 to 35 min. Their potential to further reduce the burden of GBS neonatal disease compared with antenatal culture screening needs to be assessed in future clinical studies.

Correction for Thomas et al., "In silico and experimental analysis of the repeated domains in BvaP, a protein important for GBS vaginal colonization".

Correction for Thomas et al., "In silico and experimental analysis of the repeated domains in BvaP, a protein important for GBS vaginal colonization".

In silico and experimental analysis of the repeated domains in BvaP, a protein important for GBS vaginal colonization.

Streptococcus agalactiae (group B strep, GBS) infections in neonates are often fatal and strongly associated with maternal GBS vaginal colonization. Previously, we highlighted the importance of a formerly uncharacterized protein, BvaP, in GBS vaginal colonization. BvaP is highly conserved across GBS and is made up of repeated domains, with a variable number of repeats between strains. Here, we evaluate the prevalence of BvaP repeated domains and their relevance in phenotypes previously associated with vaginal colonization. Using in silico analysis, we found that the number of repeats in the BvaP protein does not generally appear to be associated with serotype, isolation site, or host. Using BvaP truncations in GBS strain A909, we determined that a smaller number of repeats was correlated with decreased bacterial chain length, but adherence to vaginal epithelial cells was complemented using BvaP containing one, two, three, or five repeats. Future research will be geared toward understanding the host immune response to BvaP in vivo and whether vaginal carriage or host response is dependent on the BvaP repeated domains.

Applicability of an in-house extraction protocol in a Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry system for the identification of Streptococcus agalactiae from broth-enriched vaginal/rectal swab specimens.

Early laboratory identification of group B Streptococcus (GBS, Streptococcus agalactiae) in the birth canal of pregnant women is critical for prompt administration of antimicrobial therapy and may further reduce the mortality rate due to GBS neonatal infection. A total of 164 vaginal/rectal swab specimens collected from pregnant women at 35-37 weeks of gestation were screened for GBS vaginal colonization. The matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS, Bruker Biotyper, Bruker Daltonik GmbH, Bremen, Germany) system was used to detect GBS from Carrot broth and LIM broth enrichment using an in-house extraction protocol. The results were compared to those by conventional broth-enriched culture/identification methods as the gold standard. BD MAX™ GBS assay (Becton Dickinson, Sparks, MD, USA) was also performed for Carrot broth-enriched specimen. Discordant results were investigated using the GeneXpert® GBS PCR assay (Cepheid Inc., Sunnyvale, CA, USA). Using the extraction protocol, 33 (20.1%) of the 164 specimens were positive in Carrot broth, and 19 (11.6%) were positive in LIM broth. Using the culture protocol, 38 (23.2%) samples in Carrot broth and 35 (21.3%) in LIM broth were positive. The sensitivity, specificity, and positive and negative predictive values using the extraction protocol in Carrot broth and LIM broth compared to the gold standard conventional culture/identification method were 86.8% and 50.0%, 100% and 100%, 100% and 100%, and 96.2% and 86.9%, respectively. The extraction protocol with MALDI-TOF MS from Carrot broth-enriched samples provides a more rapid turnaround time, lower cost, and acceptable sensitivity and specificity to correctly identify pathogens when compared to conventional culture/identification methods.

A Novel Conserved Protein in Streptococcus agalactiae, BvaP, Is Important for Vaginal Colonization and Biofilm Formation.

Streptococcus agalactiae (group B streptococcus [GBS]) infections in neonates are often fatal and strongly associated with maternal GBS vaginal colonization. Here, we investigated the role of an uncharacterized protein, BvaP, in GBS vaginal colonization. bvaP was previously identified as the most highly upregulated gene in the GBS A909 transcriptome when comparing vaginal colonization to growth in liquid culture. We found that the absence of BvaP affects the ability of GBS to adhere to extracellular matrix components and human vaginal epithelial cells, and the ability of a ΔbvaP mutant to colonize the murine vaginal tract was significantly decreased. Cellular morphological alterations such as changes in cell shape, chain length, and clumping were also observed in a knockout mutant strain. Given its high expression level in vivo, high degree of conservation among GBS strains, and role in vaginal colonization, BvaP may be an eligible target for GBS vaccination and/or drug therapy. IMPORTANCE Neonatal GBS disease is a major cause of morbidity and mortality, and maternal vaginal colonization is the leading risk factor for the disease. Colonization prevention would greatly impact the rates of disease transmission, but vaccine development has stalled as capsular polysaccharide vaccines have low immunogenicity in vivo. While these vaccines are still in development, the addition of a protein conjugate may prove fruitful in increasing immunogenicity and strain coverage across GBS serotypes. Previous research identified sak_1753 as a highly upregulated gene during murine vaginal colonization. This study reveals that Sak_1753 is required to maintain proper GBS cellular morphology and colonization phenotypes and is required for full in vivo vaginal colonization in a murine model. We have renamed Sak_1753 group B streptococcus vaginal adherence protein (BvaP). The findings of this study indicate that BvaP is important for GBS colonization of the vaginal tract and, given its high expression level in vivo and strain conservation, may be a candidate for vaccine development.

Group B Streptococcus Capsular Serotype Alters Vaginal Colonization Fitness.

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. 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. 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. 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.

Oral probiotics to reduce vaginal group B streptococcal colonization in late pregnancy

This study aimed to evaluate the potential of oral probiotics to eradicate vaginal GBS colonization during the third trimester of pregnancy. We screened 1058 women for GBS colonization at 33–37 gestational weeks using a combination of vaginal-to-rectal swab and culture-based methods. Women who tested GBS positive were randomized to either the verum group, receiving a dietary probiotic supplement of four viable strains of Lactobacillus twice-daily for 14 days, or to the placebo group. Women underwent follow-up smears, whereat GBS colonization upon follow-up was considered the primary endpoint. We found that 215 women (20.3%) were positive for GBS upon screening, of which 82 (38.1%) were eligible for study inclusion; 41 (50%) of these were randomized to the verum and placebo groups each. After treatment, 21/33 (63.6%) members of the verum group, and 21/27 (77.8%) of the placebo group were still GBS positive (p = 0.24). Four (9.8%) women in the verum group and one (2.4%) in the placebo group experienced preterm birth (p = 0.20); smokers showed significantly higher rates of preterm birth (p = 0.03). Hence, the findings did not support the hypothesis that oral probiotics can eradicate GBS during pregnancy, although we observed a trend toward reduced GBS persistence after probiotic intake.

Prevalence and capsular type distribution of group B Streptococcus isolated from vagina of pregnant women in Nghe An province, Vietnam

Identification of GBS serotypes provides helpful information for appropriate the development of suitable vaccines; however, no reports from Vietnam have been published. This study has been performed to find the prevalence and serotypes of group B Streptococcus isolated from vagina of pregnant women in Nghe An province, Vietnam. Vaginal swabs were collected from pregnant women at 35-37 weeks of gestation at the Nghe An Obstetrics and Pediatrics Hospital, Vietnam between May 2018 and July 2019. The swabs were cultured on 5% sheep blood agar for isolation of GBS. All isolates were identified using the Gram staining, CAMP test and specific PCR. GBS strains were serotyped using the multiplex PCR assays. The prevalence of vaginal GBS colonization was 9.20% of 750 participants. Among the isolates, serotypes III (39.13%) and V (31.89%) were the most frequent, followed by serotypes Ia (11.59%), VI (11.59%), Ib (2.90%), II (1.45%) and VII (1.45%), respectively. Serotypes IV, VIII and IX were not found. The prevalence of GBS in the Nghe An province of central Vietnam was similar to reports from other parts of the world. The predominat GBS serotypes (III, V, Ia and VI) were slightly different from those previously described from other regions around the world. The high frequency of serotype VI was a notable feature of the strains from pregnant women in Vietnam.

43: Effect of obesity on maternal and neonatal penicillin levels in women with Group B Streptococcus

Early onset Group B Streptococcal (EOGBS) disease is the leading cause of early onset neonatal sepsis in the United States. Despite intrapartum penicillin (PCN) prophylaxis, infants of obese women are at 70-80% increased risk of developing EOGBS disease compared to infants of non-obese women. Our objective was to determine whether the recommended PCN dosing achieves adequate PCN levels in obese women and their neonates. We performed a prospective cohort study from Feb 2018-Jan 2019 of term GBS positive women (by rectovaginal swab at 35-37 weeks) receiving PCN. The following outcomes were compared between obese women (BMI ≥35 kg/m2) and non-obese women (BMI ≤30 kg/m2): vaginal GBS colonization at admission and after 2 completed PCN doses, serum PCN concentration in maternal blood (after 2 completed PCN doses), umbilical cord blood PCN levels, and neonatal GBS colonization (via postnatal ear swab). Fifty-six women were needed to detect a 0.75 standard deviation difference in cord blood PCN levels. 56 women were enrolled and had all specimens collected; 51 had complete data for analysis (non-obese n=26, obese n=25). Approximately 40% of women were GBS negative on admission with no difference between groups (61% vs. 62%, p=0.96). There was no difference in the median maternal serum PCN levels between groups (Table 1). There was, however, a significant difference in median cord blood PCN levels between groups (6.7 mcg/mL vs. 2.7 mcg/mL, p< 0.001) with no difference in time from PCN to delivery. Postnatally, 8% of neonates born to women in both groups were GBS positive by ear swab with no difference between groups. There were no differences in infectious outcomes. Whether the 2.5 times lower median cord blood PCN levels in obese women affects rates of EOGBS disease requires future study. Importantly, our study demonstrates that current practices for EOGBS disease prevention result in unnecessary exposure in up to 40% of patients. These findings demand improved strategies to mitigate the risks of excess antibiotic usage while optimizing prevention of EOGBS infection.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

In-vitro effect of vaginal lactobacilli against group B Streptococcus

Streptococcus agalactiae(GBS) is a leading cause of infection during pregnancy, preterm birth and neonatal infection, with a significant clinical and socio-economic impact. To prevent maternal GBS vaginal colonization, new antibiotic-free approaches, based on lactobacilli probiotics, are advisable.The aim of this study was to assess the anti-GBS activity of 14 vaginal Lactobacillus strains, belonging to different species (L. crispatus, L. gasseri, L. vaginalis), isolated from healthy pre-menopausal women. In particular, we performed ‘inhibition’ experiments, evaluating the ability of both Lactobacillus cells and culture supernatants in reducing Streptococcus viability, after 60 min contact time.First, we demonstrated that the acidic milieu, produced by vaginal lactobacilli metabolism, is crucial in counteracting GBS growth in a pH-dependent manner. Experiments with organic/inorganic acid solutions confirmed the strict correlation between pH levels and the anti-GBS activity. GBS was more sensitive to lactic acid than to hydrochloric acid, indicating that the presence of H+ ions is necessary but not sufficient for the inhibitory activity. Moreover, experiments with Lactobacillus pH-adjusted supernatants led to exclude a direct role in the anti-GBS activity by other bioactive molecules.Second, we found that only a few Lactobacillus strains were able to reduce Streptococcus viability by means of cell pellets. The anti-GBS effect displayed by Lactobacillus cells was related to the their ability to interact and aggregate with Streptococcus cells. We found that the anti-GBS activity was retained after methanol/proteinase K treatment, but lost after lysozyme exposure of Lactobacillus cells. Therefore, we supposed that non-proteinaceous components of Lactobacillus cell wall could be responsible for the anti-GBS activity.In conclusion, we identified specific Lactobacillus strains able to interfere with GBS viability by multiple strategies and we elucidated some of the mechanisms of action. These strains could serve as probiotic formulations for the prevention of GBS vaginal colonization.

Maternal group B Streptococcus recto vaginal colonization increases the odds of stillbirth: evidence from Eastern Ethiopia

BackgroundGroup B Streptococcus (GBS) causes a significant number of stillbirths. Despite this, there is little documented information on the association between stillbirth and pregnant women’s GBS recto vaginal colonization in Sub Saharan Africa. As such, this study was aimed at identifying the association between stillbirth and pregnant women’s GBS recto vaginal colonization in Eastern Ethiopia.MethodsA health facility-based cross-sectional study was conducted among 1688 pregnant women who came for delivery service in Harar town, Eastern Ethiopia between June to October in 2016. Data were collected using a pre-tested structured questionnaire and checklist (which utilize clinical record). Group B streptococcus positivity of the pregnant women was confirmed by culture of recto vaginal swab using selective media. The association between GBS colonization and stillbirth was examined using multivariable logistic regression analysis. A statistical significance was declared at p-value ≤0.05.ResultsOf the 1688 pregnant women who participated in the study, 144 had stillbirths, representing a prevalence of 8.53% [(95% CI: (7.19, 9.86)]. Group B Streptococcus colonization at birth was detected in 231 women (13.68%; 95% CI 12.04, 15.32). Of these 144 stillbirths 59 (40.97%) were from colonized mothers and 72(59.03%) were from non-colonized mothers. Of these 59 stillbirth from colonized mothers, 32(54.23%) were intrapartum stillbirth, 27(45.77%) were antepartum stillbirth occur before exposed to intrapartum antibiotic prophylaxis (IAP). After controlling for potential confounders, the odds of having a stillbirth were 8.93 times higher among recto vaginal GBS colonized pregnant women [AOR = 8.93; 95% CI; (5.47, 14.56)].ConclusionsThis study demonstrated a significant association between maternal recto vaginal GBS colonization and stillbirth. Efforts to reduce stillbirth need to consider prevention of GBS colonization among pregnant women. Maternal vaccination may provide a feasible strategy to reduce stillbirth due to GBS.

Characterization of a Two-Component System Transcriptional Regulator, LtdR, That Impacts Group B Streptococcal Colonization and Disease

Streptococcus agalactiae (group B Streptococcus [GBS]) is often a commensal bacterium that colonizes healthy adults asymptomatically and is a frequent inhabitant of the vaginal tract in women. However, in immunocompromised individuals, particularly the newborn, GBS may transition to an invasive pathogen and cause serious disease. Despite the use of the currently recommended intrapartum antibiotic prophylaxis for GBS-positive mothers, GBS remains a leading cause of neonatal septicemia and meningitis. To adapt to the various host environments encountered during its disease cycle, GBS possesses multiple two-component regulatory systems (TCSs). Here we investigated the contribution of a transcriptional regulator containing a LytTR domain, LtdR, to GBS pathogenesis. Disruption of the ltdR gene in the GBS chromosome resulted in a significant increase in bacterial invasion into human cerebral microvascular endothelial cells (hCMEC) in vitro as well as the greater penetration of the blood-brain barrier (BBB) and the development of meningitis in vivo Correspondingly, infection of hCMEC with the ΔltdR mutant resulted in increased secretion of the proinflammatory cytokines interleukin-8 (IL-8), CXCL-1, and IL-6. Further, using a mouse model of GBS vaginal colonization, we observed that the ΔltdR mutant was cleared more readily from the vaginal tract and also that infection with the ΔltdR mutant resulted in increased cytokine production from human vaginal epithelial cells. RNA sequencing revealed global transcriptional differences between the ΔltdR mutant and the parental wild-type GBS strain. These results suggest that LtdR regulates many bacterial processes that can influence GBS-host interactions to promote both bacterial persistence and disease progression.

Number of colony forming units in urine at 35–37 weeks’ gestation as predictor of the vaginal load of Group B Streptococci at birth

ObjectiveTo evaluate GBS colony numbers in the urine at 35–37 weeks’ gestation to predict the load of GBS-colonization of the vagina at birth. Study designIn this prospective observational study, we included 902 unselected pregnant women. Exposure was GBS colony forming units (CFU) per mL urine at 35–37 weeks’ gestation. Outcome was vaginal GBS colonization at birth as assessed by a semi-quantitative culture of a vaginal swab sample (negative, +1, +2, +3). ResultsBacteriuria with GBS at 35–37 weeks’ gestation performed with a sensitivity of 30% concerning any degree of vaginal GBS colonization at birth (31 of 104 cases); 19% for light (+1), 17% for medium (+2), and 52% for high load (+3) vaginal GBS colonization. The colony count in case of GBS bacteriuria at 35–37 weeks’ gestation performed with positive predictive values of 35% for <104 CFU/mL, 70% for 104 CFU/mL, and 67% for >104 CFU/mL. ConclusionEven though the urinary GBS CFU at 35–37 weeks’ gestation is strongly associated with a high load of vaginal GBS colonization intrapartum, it may not perform satisfactorily as a standalone-screening marker for risk of early-onset GBS disease.

Risk-based screening combined with a PCR-based test for group B streptococci diminishes the use of antibiotics in laboring women

ObjectiveTo assess the performance of a polymerase chain reaction – group B streptococci test (PCR-GBS test) – in deciding antibiotic prophylaxis in term laboring women. Study designIn this observational study, we enrolled 902 unselected Danish term pregnant women. During labor, midwives obtained vaginal swabs that were used for both GBS cultures (reference standard) and for the PCR-GBS test. Furthermore, we recorded the presence of risk factors for EOGBS (Early Onset Group B Streptococcal disease): (1) Bacteriuria during current pregnancy, (2) Prior infant with EOGBS (3) Temperature above 38.0°C during labor, and (4) Rupture of membranes ≥18h. ResultsThe prevalence of GBS carriers was 12% (104 of 902), the sensitivity of the PCR-GBS test 83% (86 of 104), and the specificity 97% (774 of 798). Among the 108 with one or more EOGBS-risk factors, GBS was present in 23% (25 of 108), the sensitivity 92% (23 of 25), and the specificity 89% (74 of 83). ConclusionIn programs that aim to treat all laboring women with vaginal GBS-colonization (12% in the present study) with penicillin, the PCR-GBS will perform well (sensitivity 83% and specificity 97%). In programs aiming to treat only GBS-carriers among those with risk factors of EOGBS, a reduction of penicillin usage by two-thirds from 12% to 4% may be possible.

A Murine Model of Group B Streptococcus Vaginal Colonization.

Streptococcus agalactiae (group B Streptococcus, GBS), is a Gram-positive, asymptomatic colonizer of the human gastrointestinal tract and vaginal tract of 10 - 30% of adults. In immune-compromised individuals, including neonates, pregnant women, and the elderly, GBS may switch to an invasive pathogen causing sepsis, arthritis, pneumonia, and meningitis. Because GBS is a leading bacterial pathogen of neonates, current prophylaxis is comprised of late gestation screening for GBS vaginal colonization and subsequent peripartum antibiotic treatment of GBS-positive mothers. Heavy GBS vaginal burden is a risk factor for both neonatal disease and colonization. Unfortunately, little is known about the host and bacterial factors that promote or permit GBS vaginal colonization. This protocol describes a technique for establishing persistent GBS vaginal colonization using a single β-estradiol pre-treatment and daily sampling to determine bacterial load. It further details methods to administer additional therapies or reagents of interest and to collect vaginal lavage fluid and reproductive tract tissues. This mouse model will further the understanding of the GBS-host interaction within the vaginal environment, which will lead to potential therapeutic targets to control maternal vaginal colonization during pregnancy and to prevent transmission to the vulnerable newborn. It will also be of interest to increase our understanding of general bacterial-host interactions in the female vaginal tract.

Streptococcus salivarius K12 Limits Group B Streptococcus Vaginal Colonization.

Streptococcus agalactiae (group B streptococcus [GBS]) colonizes the rectovaginal tract in 20% to 30% of women and during pregnancy can be transmitted to the newborn, causing severe invasive disease. Current routine screening and antibiotic prophylaxis have fallen short of complete prevention of GBS transmission, and GBS remains a leading cause of neonatal infection. We have investigated the ability of Streptococcus salivarius, a predominant member of the native human oral microbiota, to control GBS colonization. Comparison of the antibacterial activities of multiple S. salivarius strains by use of a deferred-antagonism test showed that S. salivarius strain K12 exhibited the broadest spectrum of activity against GBS. K12 effectively inhibited all GBS strains tested, including disease-implicated isolates from newborns and colonizing isolates from the vaginal tract of pregnant women. Inhibition was dependent on the presence of megaplasmid pSsal-K12, which encodes the bacteriocins salivaricin A and salivaricin B; however, in coculture experiments, GBS growth was impeded by K12 independently of the megaplasmid. We also demonstrated that K12 adheres to and invades human vaginal epithelial cells at levels comparable to GBS. Inhibitory activity of K12 was examined in vivo using a mouse model of GBS vaginal colonization. Mice colonized with GBS were treated vaginally with K12. K12 administration significantly reduced GBS vaginal colonization in comparison to nontreated controls, and this effect was partially dependent on the K12 megaplasmid. Our results suggest that K12 may have potential as a preventative therapy to control GBS vaginal colonization and thereby prevent its transmission to the neonate during pregnancy.

Characterization of host immunity during persistent vaginal colonization by Group B Streptococcus.

Streptococcus agalactiae (Group B Streptococcus, GBS) is a Gram-positive bacterium, which colonizes the vaginal tract in 10–30% of women. Colonization is transient in nature, and little is known about the host and bacterial factors controlling GBS persistence. Gaining insight into these factors is essential for developing therapeutics to limit maternal GBS carriage and prevent transmission to the susceptible newborn. In this work, we have used human cervical and vaginal epithelial cells, and our established mouse model of GBS vaginal colonization, to characterize key host factors that respond during GBS colonization. We identify a GBS strain that persists beyond a month in the murine vagina, whereas other strains are more readily cleared. Correspondingly, we have detected differential cytokine production in human cell lines after challenge with the persistent strain versus other GBS strains. We also demonstrate that the persistent strain more readily invades cervical cells compared to vaginal cells, suggesting that GBS may potentially use the cervix as a reservoir to establish long-term colonization. Furthermore, we have identified IL-17 production in response to long-term colonization, which is associated with eventual clearance of GBS. We conclude that both GBS strain differences and concurrent host immune responses are crucial in modulating vaginal colonization.

A novel C5a-derived immunobiotic peptide reduces Streptococcus agalactiae colonization through targeted bacterial killing.

Streptococcus agalactiae (group B Streptococcus [GBS]) is a Gram-positive bacterium that colonizes the cervicovaginal tract in approximately 25% of healthy women. Although colonization is asymptomatic, GBS can be vertically transmitted to newborns peripartum, causing severe disease such as pneumonia and meningitis. Current prophylaxis, consisting of late gestation screening and intrapartum antibiotics, has failed to completely prevent transmission, and GBS remains a leading cause of neonatal sepsis and meningitis in the United States. Lack of an effective vaccine and emerging antibiotic resistance necessitate exploring novel therapeutic strategies. We have employed a host-directed immunomodulatory therapy using a novel peptide, known as EP67, derived from the C-terminal region of human complement component C5a. Previously, we have demonstrated in vivo that EP67 engagement of the C5a receptor (CD88) effectively limits staphylococcal infection by promoting cytokine release and neutrophil infiltration. Here, using our established mouse model of GBS vaginal colonization, we observed that EP67 treatment results in rapid clearance of GBS from the murine vagina. However, this was not dependent on functional neutrophil recruitment or CD88 signaling, as EP67 treatment reduced the vaginal bacterial load in mice lacking CD88 or the major neutrophil receptor CXCr2. Interestingly, we found that EP67 inhibits GBS growth in vitro and in vivo and that antibacterial activity was specific to Streptococcus species. Our work establishes that EP67-mediated clearance of GBS is likely due to direct bacterial killing rather than to enhanced immune stimulation. We conclude that EP67 may have potential as a therapeutic to control GBS vaginal colonization.

Group B Streptococcus CovR regulation modulates host immune signalling pathways to promote vaginal colonization

Streptococcus agalactiae (Group B Streptococcus, GBS) is a frequent commensal organism of the vaginal tract of healthy women. However, GBS can transition to a pathogen in susceptible hosts, but host and microbial factors that contribute to this conversion are not well understood. GBS CovR/S (CsrR/S) is a two component regulatory system that regulates key virulence elements including adherence and toxin production. We performed global transcription profiling of human vaginal epithelial cells exposed to WT, CovR deficient, and toxin deficient strains, and observed that insufficient regulation by CovR and subsequent increased toxin production results in a drastic increase in host inflammatory responses, particularly in cytokine signalling pathways promoted by IL-8 and CXCL2. Additionally, we observed that CovR regulation impacts epithelial cell attachment and intracellular invasion. In our mouse model of GBS vaginal colonization, we further demonstrated that CovR regulation promotes vaginal persistence, as infection with a CovR deficient strainresulted in a heightened host immune response as measured by cytokine production and neutrophil activation. Using CXCr2 KO mice, we determined that this immune alteration occurs, at least in part, via signalling through the CXCL2 receptor. Taken together, we conclude that CovR is an important regulator of GBS vaginal colonization and loss of this regulatory function may contribute to the inflammatory havoc seen during the course of infection.