Nicole M. Gilbert

Immune Modulation by Group B Streptococcus Influences Host Susceptibility to Urinary Tract Infection by Uropathogenic Escherichia coli

ABSTRACT Urinary tract infection (UTI) is most often caused by uropathogenic Escherichia coli (UPEC). UPEC inoculation into the female urinary tract (UT) can occur through physical activities that expose the UT to an inherently polymicrobial periurethral, vaginal, or gastrointestinal flora. We report that a common urogenital inhabitant and opportunistic pathogen, group B Streptococcus (GBS), when present at the time of UPEC exposure, undergoes rapid UPEC-dependent exclusion from the murine urinary tract, yet it influences acute UPEC-host interactions and alters host susceptibility to persistent outcomes of bladder and kidney infection. GBS presence results in increased UPEC titers in the bladder lumen during acute infection and reduced inflammatory responses of murine macrophages to live UPEC or purified lipopolysaccharide (LPS), phenotypes that require GBS mimicry of host sialic acid residues. Taken together, these studies suggest that despite low titers, the presence of GBS at the time of polymicrobial UT exposure may be an overlooked risk factor for chronic pyelonephritis and recurrent UTI in susceptible groups, even if it is outcompeted and thus absent by the time of diagnosis.

Clinical Features of Bacterial Vaginosis in a Murine Model of Vaginal Infection with Gardnerella vaginalis

Bacterial vaginosis (BV) is a dysbiosis of the vaginal flora characterized by a shift from a Lactobacillus-dominant environment to a polymicrobial mixture including Actinobacteria and Gram-negative bacilli. BV is a common vaginal condition in women and is associated with increased risk of sexually transmitted infection and adverse pregnancy outcomes such as preterm birth. Gardnerella vaginalis is one of the most frequently isolated bacterial species in BV. However, there has been much debate in the literature concerning the contribution of G. vaginalis to the etiology of BV, since it is also present in a significant proportion of healthy women. Here we present a new murine vaginal infection model with a clinical isolate of G. vaginalis. Our data demonstrate that this model displays key features used clinically to diagnose BV, including the presence of sialidase activity and exfoliated epithelial cells with adherent bacteria (reminiscent of clue cells). G. vaginalis was capable of ascending uterine infection, which correlated with the degree of vaginal infection and level of vaginal sialidase activity. The host response to G. vaginalis infection was characterized by robust vaginal epithelial cell exfoliation in the absence of histological inflammation. Our analyses of clinical specimens from women with BV revealed a measureable epithelial exfoliation response compared to women with normal flora, a phenotype that, to our knowledge, is measured here for the first time. The results of this study demonstrate that G. vaginalis is sufficient to cause BV phenotypes and suggest that this organism may contribute to BV etiology and associated complications. This is the first time vaginal infection by a BV associated bacterium in an animal has been shown to parallel the human disease with regard to clinical diagnostic features. Future studies with this model should facilitate investigation of important questions regarding BV etiology, pathogenesis and associated complications.

Degradation, foraging and depletion of mucus sialoglycans by the vagina-adapted Actinobacterium Gardnerella vaginalis

Background: Mucus degradation is hypothesized to be important in bacterial vaginosis (BV), but mechanisms require investigation. Results: We characterize a Gardnerella vaginalis pathway that performs digestion/catabolism of mucus sialoglycans. Conclusion: G. vaginalis participates in mucosal sialic acid depletion in BV. Significance: G. vaginalis is the first example of a BV-associated organism that recapitulates a measurable clinical phenotype of mucus degradation in an animal model. Bacterial vaginosis (BV) is a polymicrobial imbalance of the vaginal microbiota associated with reproductive infections, preterm birth, and other adverse health outcomes. Sialidase activity in vaginal fluids is diagnostic of BV and sialic acid-rich components of mucus have protective and immunological roles. However, whereas mucus degradation is believed to be important in the etiology and complications associated with BV, the role(s) of sialidases and the participation of individual bacterial species in the degradation of mucus barriers in BV have not been investigated. Here we demonstrate that the BV-associated bacterium Gardnerella vaginalis uses sialidase to break down and deplete sialic acid-containing mucus components in the vagina. Biochemical evidence using purified sialoglycan substrates supports a model in which 1) G. vaginalis extracellular sialidase hydrolyzes mucosal sialoglycans, 2) liberated sialic acid (N-acetylneuraminic acid) is transported into the bacterium, a process inhibited by excess N-glycolylneuraminic acid, and 3) sialic acid catabolism is initiated by an intracellular aldolase/lyase mechanism. G. vaginalis engaged in sialoglycan foraging in vitro, in the presence of human vaginal mucus, and in vivo, in a murine vaginal model, in each case leading to depletion of sialic acids. Comparison of sialic acid levels in human vaginal specimens also demonstrated significant depletion of mucus sialic acids in women with BV compared with women with a “normal” lactobacilli-dominated microbiota. Taken together, these studies show that G. vaginalis utilizes sialidase to support the degradation, foraging, and depletion of protective host mucus barriers, and that this process of mucus barrier degradation and depletion also occurs in the clinical setting of BV.

Urinary Tract Infection as a Preventable Cause of Pregnancy Complications: Opportunities, Challenges, and a Global Call to Action

The urinary tract is a common site of infection in humans. During pregnancy, urinary tract infection (UTI) is associated with increased risks of maternal and neonatal morbidity and mortality, even when the infection is asymptomatic. By mapping available rates of UTI in pregnancy across different populations, we emphasize this as a problem of global significance. Many countries with high rates of preterm birth and neonatal mortality also have rates of UTI in pregnancy that exceed rates seen in more developed countries. A global analysis of the etiologies of UTI revealed familiar culprits as well as emerging threats. Screening and treatment of UTI have improved birth outcomes in several more developed countries and would likely improve maternal and neonatal health worldwide. However, challenges of implementation in resource-poor settings must be overcome. We review the nature of the barriers occurring at each step of the screening and treatment pipeline and highlight steps necessary to overcome these obstacles. It is our hope that the information compiled here will increase awareness of the global significance of UTI in maternal and neonatal health and embolden governments, nongovernmental organizations, and researchers to do their part to make urine screening and UTI treatment a reality for all pregnant women.

Impact of host age and parity on susceptibility to severe urinary tract infection in a murine model

The epidemiology and bacteriology of urinary tract infection (UTI) varies across the human lifespan, but the reasons for these differences are poorly understood. Using established monomicrobial and polymicrobial murine UTI models caused by uropathogenic Escherichia coli (UPEC) and/or Group B Streptococcus (GBS), we demonstrate age and parity as inter-related factors contributing to UTI susceptibility. Young nulliparous animals exhibited 10–100-fold higher bacterial titers compared to older animals. In contrast, multiparity was associated with more severe acute cystitis in older animals compared to age-matched nulliparous controls, particularly in the context of polymicrobial infection where UPEC titers were ∼1000-fold higher in the multiparous compared to the nulliparous host. Multiparity was also associated with significantly increased risk of chronic high titer UPEC cystitis and ascending pyelonephritis. Further evidence is provided that the increased UPEC load in multiparous animals required TLR4-signaling. Together, these data strongly suggest that the experience of childbearing fundamentally and permanently changes the urinary tract and its response to pathogens in a manner that increases susceptibility to severe UTI. Moreover, this murine model provides a system for dissecting these and other lifespan-associated risk factors contributing to severe UTI in at-risk groups.

Host-Like Carbohydrates Promote Bloodstream Survival of Vibrio vulnificus In Vivo

ABSTRACT Sialic acids are found on all vertebrate cell surfaces and are part of a larger class of molecules known as nonulosonic acids. Many bacterial pathogens synthesize related nine-carbon backbone sugars; however, the role(s) of these non-sialic acid molecules in host-pathogen interactions is poorly understood. Vibrio vulnificus is the leading cause of seafood-related death in the United States due to its ability to quickly access the host bloodstream, which it can accomplish through gastrointestinal or wound infection. However, little is known about how this organism persists systemically. Here we demonstrate that sialic acid-like molecules are present on the lipopolysaccharide of V. vulnificus, are required for full motility and biofilm formation, and also contribute to the organisms natural resistance to polymyxin B. Further experiments in a murine model of intravenous V. vulnificus infection demonstrated that expression of nonulosonic acids had a striking benefit for bacterial survival during bloodstream infection and dissemination to other tissues in vivo. In fact, levels of bacterial persistence in the blood corresponded to the overall levels of these molecules expressed by V. vulnificus isolates. Taken together, these results suggest that molecules similar to sialic acids evolved to facilitate the aquatic lifestyle of V. vulnificus but that their emergence also resulted in a gain of function with life-threatening potential in the human host.

Transient microbiota exposures activate dormant Escherichia coli infection in the bladder and drive severe outcomes of recurrent disease

Pathogens often inhabit the body asymptomatically, emerging to cause disease in response to unknown triggers. In the bladder, latent intracellular Escherichia coli reservoirs are regarded as likely origins of recurrent urinary tract infection (rUTI), a problem affecting millions of women worldwide. However, clinically plausible triggers that activate these reservoirs are unknown. Clinical studies suggest that the composition of a woman’s vaginal microbiota influences her susceptibility to rUTI, but the mechanisms behind these associations are unclear. Several lines of evidence suggest that the urinary tract is routinely exposed to vaginal bacteria, including Gardnerella vaginalis, a dominant member of the vaginal microbiota in some women. Using a mouse model, we show that bladder exposure to G. vaginalis triggers E. coli egress from latent bladder reservoirs and enhances the potential for life-threatening outcomes of the resulting E. coli rUTI. Transient G. vaginalis exposures were sufficient to cause bladder epithelial apoptosis and exfoliation and interleukin-1-receptor-mediated kidney injury, which persisted after G. vaginalis clearance from the urinary tract. These results support a broader view of UTI pathogenesis in which disease can be driven by short-lived but powerful urinary tract exposures to vaginal bacteria that are themselves not “uropathogenic” in the classic sense. This “covert pathogenesis” paradigm may apply to other latent infections, (e.g., tuberculosis), or for diseases currently defined as noninfectious because routine culture fails to detect microbes of recognized significance.

Relationship between nugent score and vaginal epithelial exfoliation

Objective Clue cells characteristic of bacterial vaginosis (BV) are thought to arise due to exfoliation of the vaginal epithelium; however, there is little published data connecting total numbers of epithelial cells to markers of BV. The purpose of this study was to enumerate exfoliated epithelial cells (independent of clue cells) and examine the relationship to Nugent score. Study design We conducted a cross-sectional sub-study of the Contraceptive CHOICE Project cohort. Vaginal swabs were used to create vaginal smears for Gram staining and these smears were later scored using the Nugent method, and then two blinded observers used microscopy to enumerate exfoliated epithelial cells. The degree of epithelial cell exfoliation was compared between women diagnosed as BV-negative (Nugent score 0–3), BV-intermediate (Nugent score 4–6), and BV-positive (Nugent score 7–10). BV specimens (Nugent 7–10) were randomly matched to specimens in the two other groups (Nugent low and Nugent-intermediate), in order to avoid comparing groups of women with potentially confounding baseline demographics. Results Exfoliated epithelial cell counts were higher in the vaginal smears from BV-positive women compared with BV-negative women. Higher levels of epithelial exfoliation were also evident in BV-intermediate women compared to those with low Nugent scores. After adjustment for clustering introduced by matching, the incidence ratio of increased epithelial cell counts was 2.09 (95% CI 1.50–2.90) for the BV-intermediate women and 1.71 (95% CI 1.23–2.38) for the BV positive women. Conclusion A vaginal epithelial exfoliation phenotype was measured in both Nugent-defined BV-positive and BV-intermediate women. Bacterial vaginosis and intermediate status (Nugent score >3) was associated with significantly more vaginal epithelial exfoliation compared to women with Lactobacillus-dominated microbiotas (Nugent 0–3).