Tamm-Horsfall protein augments neutrophil NETosis during urinary tract infection.

Urinary neutrophils are a hallmark of urinary tract infection (UTI), yet the mechanisms governing their activation, function, and efficacy in controlling infection remain incompletely understood. Tamm-Horsfall glycoprotein (THP), the most abundant protein in urine, uses terminal sialic acids to bind an inhibitory receptor and dampen neutrophil inflammatory responses. We hypothesized that neutrophil modulation is an integral part of THP-mediated host protection. In a UTI model, THP-deficient mice showed elevated urinary tract bacterial burdens, increased neutrophil recruitment, and more severe tissue histopathological changes compared to WT mice. Furthermore, THP-deficient mice displayed impaired urinary NETosis during UTI. To investigate the impact of THP on NETosis, we coupled in vitro fluorescence-based NET assays, proteomic analyses, and standard and imaging flow cytometry with peripheral human neutrophils. We found that THP increases proteins involved in respiratory chain, neutrophil granules, and chromatin remodeling pathways, enhances NETosis in an ROS-dependent manner, and drives NET-associated morphologic features including nuclear decondensation. These effects were observed only in the presence of a NETosis stimulus and could not be solely replicated with equivalent levels of sialic acid alone. We conclude that THP is a critical regulator of NETosis in the urinary tract, playing a key role in host defense against UTI.

Gestational diabetes augments group B Streptococcus infection by disrupting maternal immunity and the vaginal 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. Using a murine GDM model of GBS colonization and perinatal transmission, we find that GDM mice display greater GBS in utero dissemination and subsequently worse neonatal outcomes. Dual-RNA sequencing reveals differential GBS adaptation to the GDM reproductive tract, including a putative glycosyltransferase (yfhO), and altered host responses. GDM immune disruptions include reduced uterine natural killer cell activation, impaired recruitment to placentae, and altered maternofetal cytokines. Lastly, we observe distinct vaginal microbial taxa associated with GDM status and GBS invasive disease status. Here, we show a model of GBS dissemination in GDM hosts that recapitulates several clinical aspects and identifies multiple host and bacterial drivers of GBS perinatal disease.

Vaginal microbial dynamics and pathogen colonization in a humanized microbiota mouse model.

Vaginal microbial composition is associated with differential risk of urogenital infection. Although Lactobacillus spp. are thought to confer protection against infection, the lack of in vivo models resembling the human vaginal microbiota remains a prominent barrier to mechanistic discovery. Using 16S rRNA amplicon sequencing of C57BL/6J female mice, we found that vaginal microbial composition varies within and between colonies across three vivaria. Noting vaginal microbial plasticity in conventional mice, we assessed the vaginal microbiome of humanized microbiota mice (HMbmice). Like the community structure in conventional mice, HMbmice vaginal microbiota clustered into community state types but, uniquely, HMbmice communities were frequently dominated by Lactobacillus or Enterobacteriaceae. Compared to conventional mice, HMbmice were less susceptible to uterine ascension by urogenital pathobionts group B Streptococcus (GBS) and Prevotella bivia. Although Escherichia and Lactobacillus both correlated with the absence of uterine GBS, vaginal pre-inoculation with exogenous HMbmouse-derived E. coli, but not Ligilactobacillus murinus, reduced vaginal GBS burden. Overall, HMbmice serve as a useful model to elucidate the role of endogenous microbes in conferring protection against urogenital pathogens.

Breastfeeding Practices of Women with HIV in Sub-Saharan Africa.

OBJECTIVE: To identify sociodemographic factors that could enhance breastfeeding uptake among women with HIV in sub-Saharan Africa. DESIGN: This was a secondary analysis from a retrospective cohort study using the Demographic Health Surveys (DHS) on women and HIV data from 14 sub-Saharan African countries during the period from 2010 to 2018. Our study sample encompassed women aged 15 to 49 years with HIV, with childbearing history within the 3 to 5 years preceding the survey, living in any of the countries in sub-Saharan Africa for whom breastfeeding information was available. We used an adjusted survey log binomial regression model to examine factors associated with breastfeeding among participants. SETTING: Sub-Saharan Africa. PARTICIPANTS: Breastfeeding women with HIV from 15 sub-Saharan African countries. RESULTS: Of 138,920 women with HIV in sub-Saharan Africa, 49,479 (35.6%) breastfed their infants. Young women, aged 15 to 19 years (90.5%, n = 2,422) were more likely to breastfeed than those aged 25 to 29 years (13.7%, n = 5,266). Breastfeeding was more common among women who lived in rural areas (38.1%, n = 26,000) than among those in urban areas (33.2%, n = 23,479) (p < .01). The wealthiest women (43.3%, n = 13,710) breastfed more than those with the lowest economic resources (30.8%, n = 4,750) (p < .01). CONCLUSION: Recognizing the breastfeeding issues in this individualized sub-Saharan population of women with HIV can influence the creation of more accessibility to education, resources, supplies, counseling, and support throughout the region and beyond through policy changes in health care.

Vaginal microbial dynamics and pathogen colonization in a humanized microbiota mouse model.

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 ( HMb mice) would model a more human-like vaginal microbiota. Similar to humans and conventional mice, HMb mice vaginal microbiota clustered into five community state types ( h mCST). Uniquely, HMb mice vaginal communities were frequently dominated by Lactobacilli or Enterobacteriaceae . Compared to genetically-matched conventional mice, HMb mice 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 HMb mice 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 HMb mouse-derived E. coli , but not L. murinus , conferred protection against vaginal GBS burden. Overall, the HMb mice are an improved model to elucidate the role of endogenous microbes in conferring protection against urogenital pathogens. IMPORTANCE: An altered vaginal microbiota, typically with little to no levels of Lactobacillus , is associated with increased susceptibility to urogenital infections, although mechanisms driving this vulnerability are not fully understood. Despite known inhibitory properties of Lactobacillus against urogenital pathogens, clinical studies with Lactobacillus probiotics have shown mixed success. In this study, we characterize the impact of the vaginal microbiota on urogenital pathogen colonization using a humanized microbiota mouse model that more closely mimics the human vaginal microbiota. We found several vaginal bacterial taxa that correlated with reduced pathogen levels but showed discordant effects in pathogen inhibition between in vitro and in vivo assays. We propose that this humanized microbiota mouse platform is an improved model to describe the role of the vaginal microbiota in protection against urogenital pathogens. Furthermore, this model will be useful in testing efficacy of new probiotic strategies in the complex vaginal environment.

Phage Resistance Accompanies Reduced Fitness of Uropathogenic Escherichia coli in the Urinary Environment.

Urinary tract infection (UTI) is among the most common infections treated worldwide each year and is caused primarily by uropathogenic Escherichia coli (UPEC). Rising rates of antibiotic resistance among uropathogens have spurred a consideration of alternative treatment 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, namely, 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 to 8 h of coincubation. Whole-genome sequencing revealed that UPEC strains resistant to HP3 and ES17 harbored mutations in genes involved in lipopolysaccharide (LPS) biosynthesis. Phage-resistant strains displayed several in vitro phenotypes, including alterations to adherence to and invasion of human bladder epithelial HTB-9 cells and increased biofilm formation in some isolates. Interestingly, these phage-resistant UPEC isolates 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, phage-resistant UPEC isolates were attenuated in bladder colonization in a murine UTI model. In total, our findings suggest that while resistance to phages, such as HP3 and ES17, may arise readily in the urinary environment, phage resistance is accompanied by fitness costs which may render UPEC more susceptible to host immunity or antibiotics. IMPORTANCE 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. For successful UTI treatment, we must predict how bacteria will evade killing by phage and identify the downstream consequences of phage resistance during bacterial infection. In our current study, we found that while phage-resistant bacteria quickly emerged in vitro, these bacteria were 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, of which each is evaded only at the cost of bacterial fitness.

Pregnancy-Associated Diabetes Mellitus and Stillbirths by Race and Ethnicity among Hospitalized Pregnant Women in the United States.

OBJECTIVES: Racial disparities in preexisting diabetes mellitus (PDM) and gestational diabetes mellitus (GDM) remain largely unexplored. We examined national PDM and GDM prevalence trends by race/ethnicity and the association between these conditions and fetal death. METHODS: This was a retrospective cross-sectional analysis of 69,539,875 pregnancy-related hospitalizations from 2002 to 2017 including 674,040 women with PDM (1.0%) and 2,960,797 (4.3%) with GDM from the US Nationwide Inpatient Sample Survey. Joinpoint regression was used to evaluate trends in prevalence. Survey logistic regression was used to evaluate the association between exposures (PDM and GDM) and outcome. RESULTS: Overall, the average annual increase in prevalence was 5.2% (95% confidence interval [CI] 4.2-6.2) for GDM and 1.0% (95% CI -0.1 to 2.0) for PDM, during the study period. Hispanic (average annual percentage change 5.3, 95% CI 3.6 - 7.1) and non-Hispanic Black (average annual percentage change 0.9, 95% CI 0.1 - 1.7) women had the highest average annual percentage increase in the prevalence of GDM and PDM, respectively. After adjustment, the odds of stillbirth were highest for Hispanic women with PDM (odds ratio 2.41, 95% CI 2.23-2.60) and decreased for women with GDM (odds ratio 0.51, 95% CI 0.50-0.53), irrespective of race/ethnicity. CONCLUSIONS: PDM and GDM prevalence is increasing in the United States, with the highest average annual percentage changes seen among minority women. Furthermore, the reasons for the variation in the occurrence of stillbirths among mothers with PDM and GDM by race/ethnicity are not clear and warrant additional research.

Racial/ethnic disparities in infertility treatment utilization in the US, 2011-2019.

With delayed child-bearing age, there has been an increase in infertility rates globally and in the United States (US). Unsurprisingly, there has been a concomitant substantial increase in the number of individuals seeking infertility treatments over the last decade. This study aimed to examine the relationship between race/ethnicity and the utilization of different infertility treatments over the previous decade. We conducted this retrospective cohort study using the United States (US) Birth data files 2011-2019. We calculated the rates of infertility treatment and its subtypes over the study period. Descriptive statistics were utilized to examine the sociodemographic and birth characteristics for overall births and those associated with any infertility treatment and each of its subtypes. We calculated the level of association between race/ethnicity and utilization of infertility treatment and the subtypes using adjusted logistic regression models. We found that the rate of infertility treatments for all subtypes considered, had steadily increased by 63.7% within the past decade. In contrast, fertility enhancing drugs or Intrauterine Insemination (IUI) increased by 134%, and in vitro fertilization (IVF), gamete intrafallopian transfer (GIFT), and zygote intrafallopian transfer (ZIFT) treatments increased by 40% over the 9-year study period. Non-Hispanic (NH) Asian women had the highest rate of any infertility treatment with a rate of 25 per 1000 births whereas Hispanic women had the lowest rate of any infertility treatment at 5.8 per 1000 births. When compared with NH-White women, NH-Asian women had a modest 7% lower likelihood (OR = 0.93, 95% CI = 0.92-0.94) of receiving any infertility treatment while NH-Black and Hispanic women had about 70% lower likelihood of receiving any infertility treatment. Our report of increased assisted reproductive technology (ART) utilization rates, and marked racial/ethnic differences in ART utilization highlight the importance of expanding knowledge of inequities that continue to impact marginalized groups, a critical step for informing actionable strategy formulations (i.e., advocacy, policy change, patient education, provider training) to address these inequities.

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.