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.

UTI is one of the most common causes of outpatient antibiotic use, and rising antibiotic resistance threatens the ability to control UTI unless alternative treatments are developed. Bacteriophage (phage) therapy is gaining renewed interest; however, much like with antibiotics, bacteria can readily become resistant to phages.

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. Additionally, 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.

Group B Streptococcus (GBS) is a pervasive neonatal pathogen accounting for a combined half a million deaths and stillbirths annually. The most common source of fetal or neonatal GBS exposure is the maternal microbiota.

Group B Streptococcus (GBS) is a pervasive perinatal pathogen, yet factors driving GBS dissemination in utero are poorly defined. Gestational diabetes mellitus (GDM), a complication marked by dysregulated immunity and maternal microbial dysbiosis, increases risk for GBS perinatal disease. We interrogated host-pathogen dynamics in a novel murine GDM model of GBS colonization and perinatal transmission. GDM mice had greater GBS in utero dissemination and subsequently worse neonatal outcomes. Dual-RNA sequencing revealed differential GBS adaptation to the GDM reproductive tract, including a putative glycosyltransferase (yfhO), and altered host responses. GDM disruption of immunity included reduced uterine natural killer cell activation, impaired recruitment to placentae, and altered vaginal cytokines. Lastly, we observed distinct vaginal microbial taxa associated with GDM status and GBS invasive disease status. Our translational model of GBS perinatal transmission in GDM hosts recapitulates several clinical aspects and enables discovery of host and bacterial drivers of GBS perinatal disease.

Vaginal microbiota composition is associated with differential risk of urogenital infection. Although vaginal Lactobacillus spp. are thought to confer protection through acidification, bacteriocin production, and immunomodulation, lack of an in vivo model system that closely resembles the human vaginal microbiota remains a prominent barrier to mechanistic discovery. We performed 16S rRNA amplicon sequencing of wildtype C57BL/6J mice, commonly used to study pathogen colonization, and found that the vaginal microbiome composition varies highly both within and between colonies from three distinct vivaria. Because of the strong influence of environmental exposure on vaginal microbiome composition, we assessed whether a humanized microbiota mouse (HMbmice) would model a more human-like vaginal microbiota. Similar to humans and conventional mice, HMbmice vaginal microbiota clustered into five community state types (hmCST). Uniquely, HMbmice vaginal communities were frequently dominated by Lactobacilli or Enterobacteriaceae. Compared to genetically-matched conventional mice, HMbmice were less susceptible to uterine ascension by urogenital pathobionts group B Streptococcus (GBS) and Prevotella bivia, but no differences were observed with uropathogenic E. coli. Specifically, vaginal Enterobacteriaceae and Lactobacillus were associated with the absence of uterine GBS. Anti-GBS activity of HMbmice vaginal E. coli and L. murinus isolates, representing Enterobacteriaceae and Lactobacillus respectively, were characterized in vitro and in vivo. Although L. murinus reduced GBS growth in vitro, vaginal pre-inoculation with HMbmouse-derived E. coli, but not L. murinus, conferred protection against vaginal GBS burden. Overall, the HMbmice are an improved model to elucidate the role of endogenous microbes in conferring protection against urogenital pathogens.

Urinary tract infections (UTIs) are among the most common infections treated worldwide each year and are primarily caused by uropathogenic E. coli (UPEC). Rising rates of antibiotic resistance among uropathogens have spurred consideration of alternative strategies such as bacteriophage (phage) therapy; however, phage-bacterial interactions within the urinary environment are poorly defined. Here, we assess the activity of two phages, HP3 and ES17, against clinical UPEC isolates using in vitro and in vivo models of UTI. In both bacteriologic medium and pooled human urine, we identified phage resistance arising within the first 6-8 hours of coincubation. Whole genome sequencing revealed that UPEC resistant to HP3 and ES17 harbored mutations in genes involved in lipopolysaccharide (LPS) biosynthesis. These mutations coincided with several in vitro phenotypes, including alterations to adherence to and invasion of human bladder epithelial HTB-9 cells, and increased biofilm formation. Interestingly, these phage-resistant UPEC demonstrated reduced growth in pooled human urine, which could be partially rescued by nutrient supplementation, and were more sensitive to several outer membrane targeting antibiotics than parental strains. Additionally, these phage-resistant UPEC were attenuated in a murine UTI model. In total, our findings suggest that while resistance to phages, such as LPS-targeted HP3 and ES17, may readily arise in the urinary environment, phage resistance is accompanied by fitness costs rendering UPEC more susceptible to host immunity or antibiotics.IMPORTANCEUTIs are one of the most common causes of outpatient antibiotic use, and rising antibiotic resistance threatens the ability to control these infections unless alternative treatments are developed. Bacteriophage (phage) therapy is gaining renewed interest, however, much like antibiotics, bacteria can readily become resistant to phage. For successful UTI treatment, we must predict how bacteria will evade killing by phage and identify the downstream consequences of phage-resistant bacterial infections. In our current study, we found that while phage-resistant mutant bacteria quickly emerged, these mutations left bacteria less capable of growing in human urine and colonizing the murine bladder. These results suggest that phage therapy poses a viable UTI treatment if phage resistance confers fitness costs for the uropathogen. These results have implications for developing cocktails of phage with multiple different bacterial targets, each of which is only evaded at the cost of bacterial fitness.

Microorganisms colonizing the human vaginal mucosa are associated with healthy states, as well as conditions such as bacterial vaginosis and infection-associated preterm birth. Here, we report complete genome sequences of 37 bacterial isolates from the human vaginal tract.