Fecal Pharmacokinetics and Gut Microbiome Effects of Oral Omadacycline Versus Vancomycin in Healthy Volunteers.

BACKGROUND: Clostridioides difficile infection (CDI) is a common healthcare-associated infection with limited treatment options. Omadacycline, an aminomethylcycline tetracycline, has potent in vitro activity against C difficile and a low propensity to cause CDI in clinical trials. We aimed to assess fecal pharmacokinetics and gut microbiome effects of oral omadacycline compared to oral vancomycin in healthy adults. METHODS: This was a phase 1, nonblinded, randomized clinical trial conducted in healthy volunteers aged 18-40 years. Subjects received a 10-day course of omadacycline or vancomycin. Stool samples were collected at baseline, daily during therapy, and at follow-up visits. Omadacycline and vancomycin stool concentrations were assessed, and microbiome changes were compared. RESULTS: Sixteen healthy volunteers with a mean age of 26 (standard deviation [SD], 5) years were enrolled; 62.5% were male, and participants' mean body mass index was 23.5 (SD, 4.0) kg/m2. Omadacycline was well tolerated with no safety signal differences between the 2 antibiotics. A rapid initial increase in fecal concentrations of omadacycline was observed compared to vancomycin, with maximum concentrations achieved within 48 hours. A significant difference in alpha diversity was observed following therapy in both the omadacycline and vancomycin groups (P < .05). Bacterial abundance and beta diversity analysis showed differing microbiome changes in subjects who received omadacycline versus vancomycin. CONCLUSIONS: Subjects given omadacycline had high fecal concentrations with a distinct microbiome profile compared to vancomycin. CLINICAL TRIALS REGISTRATION: NCT06030219.

Antibacterial activity of ibezapolstat against antimicrobial-resistant clinical strains of Clostridioides difficile.

Antimicrobial resistance is emerging in clinical strains of Clostridioides difficile. Ibezapolstat (IBZ) is a DNA polymerase IIIC inhibitor that has completed phase II clinical trials. IBZ has potent in vitro activity against wild-type, susceptible strains but its effect on C. difficile strains with reduced susceptibility to metronidazole (MTZ), vancomycin (VAN), or fidaxomicin (FDX) has not been tested. The primary objective of this study was to test the antibacterial properties of IBZ against multidrug-resistant C. difficile strains. The in vitro activity, bactericidal, and time-kill activity of IBZ versus comparators were evaluated against 100 clinical strains of which 59 had reduced susceptibility to other C. difficile antibiotics. Morphologic changes against a multidrug resistance strain were visualized by light and scanning electron microscopy. The overall IBZ MIC50/90 values (µg/mL) for evaluated C. difficile strains were 4/8, compared with 2/4 for VAN, 0.5/1 for FDX, and 0.25/4 for MTZ. IBZ MIC50/90 values did not differ based on non-susceptibility to antibiotic class or number of classes to which strains were non-susceptible. IBZ bactericidal activity was similar to the minimum inhibitory concentration (MIC) and maintained in wild-type and non-susceptible strains. Time-kill assays against two laboratory wild-type and two clinical non-susceptible strains demonstrated sustained IBZ activity despite reduced killing by comparator antibiotics for IBZ and VAN non-susceptible strains. Microscopy visualized increased cell lengthening and cellular damage in multidrug-resistant strains exposed to IBZ sub-MIC concentrations. This study demonstrated the potent antibacterial activity of IBZ against a large collection of C. difficile strains including multidrug-resistant strains. This study highlights the therapeutic potential of IBZ against multidrug-resistant strains of C. difficile.

The Novel DNA Binding Mechanism of Ridinilazole, a Precision Clostridiodes difficile Antibiotic.

Clostridioides difficile infection (CDI) causes substantial morbidity and mortality worldwide with limited antibiotic treatment options. Ridinilazole is a precision bisbenzimidazole antibiotic being developed to treat CDI and reduce unacceptably high rates of infection recurrence in patients. Although in late clinical development, the precise mechanism of action by which ridinilazole elicits its bactericidal activity has remained elusive. Here, we present conclusive biochemical and structural data to demonstrate that ridinilazole has a primary DNA binding mechanism, with a co-complex structure confirming binding to the DNA minor groove. Additional RNA-seq data indicated early pleiotropic changes to transcription, with broad effects on multiple C. difficile compartments and significant effects on energy generation pathways particularly. DNA binding and genomic localization was confirmed through confocal microscopy utilizing the intrinsic fluorescence of ridinilazole upon DNA binding. As such, ridinilazole has the potential to be the first antibiotic approved with a DNA minor groove binding mechanism of action.

Epigenetic regulation of 5α reductase-1 underlies adaptive plasticity of reproductive function and pubertal timing.

BACKGROUND: Women facing increased energetic demands in childhood commonly have altered adult ovarian activity and shorter reproductive lifespan, possibly comprising a strategy to optimize reproductive success. Here, we sought to understand the mechanisms of early-life programming of reproductive function, by integrating analysis of reproductive tissues in an appropriate mouse model with methylation analysis of proxy tissue DNA in a well-characterized population of Bangladeshi migrants in the UK. Bangladeshi women whose childhood was in Bangladesh were found to have later pubertal onset and lower age-matched ovarian reserve than Bangladeshi women who grew-up in England. Subsequently, we aimed to explore the potential relevance to the altered reproductive phenotype of one of the genes that emerged from the screens. RESULTS: Of the genes associated with differential methylation in the Bangladeshi women whose childhood was in Bangladesh as compared to Bangladeshi women who grew up in the UK, 13 correlated with altered expression of the orthologous gene in the mouse model ovaries. These mice had delayed pubertal onset and a smaller ovarian reserve compared to controls. The most relevant of these genes for reproductive function appeared to be SRD5A1, which encodes the steroidogenic enzyme 5α reductase-1. SRD5A1 was more methylated at the same transcriptional enhancer in mice ovaries as in the women's buccal DNA, and its expression was lower in the hypothalamus of the mice as well, suggesting a possible role in the central control of reproduction. The expression of Kiss1 and Gnrh was also lower in these mice compared to controls, and inhibition of 5α reductase-1 reduced Kiss1 and Gnrh mRNA levels and blocked GnRH release in GnRH neuronal cell cultures. Crucially, we show that inhibition of this enzyme in female mice in vivo delayed pubertal onset. CONCLUSIONS: SRD5A1/5α reductase-1 responds epigenetically to the environment and its downregulation appears to alter the reproductive phenotype. These findings help to explain diversity in reproductive characteristics and how they are shaped by early-life environment and reveal novel pathways that might be targeted to mitigate health issues caused by life-history trade-offs.

Fusobacteria behaving badly: Masquerading strains of strictly anaerobic Escherichiacoli misidentified due to the deletion of the hemB gene.

Three strictly anaerobic strains of Escherichia coli were misidentified as Fusobacterium mortiferum, due to a deletion of the hemB gene which is involved in anaerobic respiration. An unusual antimicrobial susceptibility pattern sparked the further diagnostic strategies that eventually identified these strains as true anaerobic E. coli This phenomenon is more common than appreciated and can have an impact on clinical practice including persistent and relapsing infections.

Efficacy, Safety, Pharmacokinetics, and Microbiome Changes of Ibezapolstat in Adults with Clostridioides difficile Infection: A Phase 2a Multicenter Clinical Trial.

BACKGROUND: This study was the first human validation of the gram-positive bacterial DNA polymerase IIIC target in patients with Clostridioides difficile infection. The primary objectives were to assess clinical cure rates and adverse events (AEs). Secondary objectives were to evaluate plasma/fecal pharmacokinetics, microbiologic eradication, microbiome and bile acid effects, and sustained clinical cure (SCC) with ibezapolstat. METHODS: This single-arm, open-label, phase 2a study enrolled adults with C. difficile infection at 4 US centers. Patients received ibezapolstat 450 mg orally every 12 hours for 10 days and followed for an additional 28 days to assess study objectives. RESULTS: Ten patients with a mean (standard deviation [SD]) age of 49 [15] years were enrolled. Seven AEs were reported classified as mild-moderate. Plasma levels of ibezapolstat ranged from 233 to 578 ng/mL while mean (SD) fecal levels were 416 (494) µg/g stool by treatment day 3 and >1000 µg/g stool by days 8-10. A rapid increase in alpha diversity in the fecal microbiome was noted after starting ibezapolstat therapy, which was maintained after completion of therapy. A proportional decrease in Bacteroidetes phylum was observed (mean change [SD], -10.0% [4.8%]; P = .04) with a concomitantly increased proportion of Firmicutes phylum (+14.7% [5.4%]; P = .009). Compared with baseline, total primary bile acids decreased by a mean (SD) of 40.1 (9.6) ng/mg stool during therapy (P < .001) and 40.5 (14.1) ng/mg stool after completion of therapy (P = .007). Rates of both initial clinical cure and SCC at 28 days were 100% (10 of 10 patients). CONCLUSIONS: In this phase 2a study, 10 of 10 patients achieved SCC, demonstrated favorable pharmacokinetics, minimal AEs, and beneficial microbiome and bile acids results. These results support continued clinical development.

Functional and Metagenomic Evaluation of Ibezapolstat for Early Evaluation of Anti-Recurrence Effects in Clostridioides difficile Infection.

Reduction of Clostridioides difficile infection (CDI) recurrence is an essential endpoint for CDI-directed antibiotic development that is often not evaluated until Phase III trials. The purpose of this project was to use a functional and metagenomic approach to predict the potential anti-CDI recurrence effect of ibezapolstat, a DNA polymerase IIIC inhibitor, in clinical development for CDI. As part of the Phase I ibezapolstat clinical study, stool samples were collected from 22 healthy volunteers, who were given either ibezapolstat or vancomycin. Stool samples were evaluated for microbiome changes and bile acid concentrations. Ibezapolstat 450 mg and vancomycin, but not ibezapolstat 300 mg, showed statistically significant changes in alpha diversity over time compared to that of a placebo. Beta diversity changes confirmed that microbiota were significantly different between study groups. Vancomycin had a more wide-ranging effect on the microbiome, characterized by an increased proportion of Gammaproteobacteria. Ibezapolstat demonstrated an increased proportion of Actinobacteria, including the Bifidobacteriaceae family. Using a linear regression analysis, vancomycin was associated with significant increases in primary bile acids as well as primary:secondary bile acid ratios. An overabundance of Enterobacteriaceae was most highly correlated with primary bile acid concentrations (r = 0.63; P < 0.0001). Using Phase I healthy volunteer samples, beneficial changes suggestive of a lower risk of CDI recurrence were associated with ibezapolstat compared to vancomycin. This novel omics approach may allow for better and earlier prediction of anti-CDI recurrence effects for antibiotics in the clinical development pipeline.

No impact of developmental conditions on serum estradiol levels among Bangladeshi women in the UK and Bangladesh.

INTRODUCTION: While many aspects of female ovarian function respond to environmental stressors, estradiol (E2) appears less sensitive to stressors than progesterone, except under extreme ecological conditions. However, earlier studies relied on saliva samples, considered less sensitive than blood. Here, we investigated E2 variation among 177 Bangladeshi and UK white women, aged 35-59, using single serum samples. Bangladeshi women either grew up in Sylhet, Bangladesh (exposed to poor sanitation, limited health care, and higher pathogen loads but not poor energy availability), or in the UK. METHODS: We collected samples on days 4-6 of the menstrual cycle in menstruating women and on any day for post-menopausal women. Participants included: (i) Bangladeshi sedentees (n = 36), (ii) Bangladeshis who migrated to the UK as adults (n = 52), (iii) Bangladeshis who migrated as children (n = 40), and (iv) UK white women matched for neighborhood residence to the migrants (n = 49). Serum was obtained by venipuncture and analyzed using electrochemiluminescence. We collected anthropometrics and supplementary sociodemographic and reproductive data through questionnaires. We analyzed the data using multivariate regression. RESULTS: E2 levels did not differ between migrant groups after controlling for age, BMI, physical activity, psychosocial stress, parity, and time since last birth (parous women). Paralleling results from salivary E2, serum E2 did not differ among women who experienced varying developmental conditions. CONCLUSION: Our results reinforce the hypothesis that E2 levels are stable under challenging environmental conditions. Interpopulation variation may only arise under chronic conditions of extreme nutritional scarcity, energy expenditure, and/or high disease burdens.

Multi-country surveillance of Clostridioides difficile demonstrates high prevalence of spores in non-healthcare environmental settings.

BACKGROUND: C. difficile spores are frequently isolated from hospital and non-healthcare settings but a worldwide analysis has not been done. The study objectives were to assess C. difficile spore contamination in the hospital and non-healthcare environments across a variety of countries. METHODS: Field studies assessed hospital vs. non-healthcare C. difficile spore contamination in hospitals, non-healthcare buildings, outdoor environments, and shoes. Swabs were cultured anaerobically for C. difficile and typed using PCR-fluorescent ribotyping. C. difficile contamination by swabbing area and geographic locations were compared. FINDINGS: A total of 7,857 unique samples were collected primarily from the USA (89%) in addition to 9 other countries. The global prevalence of C difficile from environmental samples was 25.3% and did not differ between countries. In USA based studies, C. difficile contamination rates were similar for healthcare buildings (23.2%), non-healthcare buildings (23.4%), and outdoor spaces (24.7%). Floor samples had significantly higher (p < 0.001) C. difficile contamination rate (46.5%) followed by non-floor samples (21.1%), and bathrooms (15.3%). In a comparison of USA to other country samples, C. difficile contamination rates were similar for USA samples (21.5%) compared to rest of world samples (22.3%; p = 0.61). The most common ribotypes included F014-020 (15.7%), F106 (12.6%), F010 (8.9%), F027 (8.8%), and F002 (8.1%) and did not differ significantly between USA and non-USA samples. Finally, 546 of 1,218 (44.8%) shoe soles swabbed from the USA were contaminated with C. difficile spores. INTERPRETATION: This large surveillance study of several countries demonstrated high prevalence of toxigenic C. difficile in non-healthcare environments with high contamination rates from floors and shoe soles.

Comparative clinical outcomes evaluation of hospitalized patients infected with Clostridioides difficile ribotype 106 vs. other toxigenic strains.

BACKGROUND: Although Clostridioides difficile surveillance often identifies emerging strains, clinical outcome evaluations are rarely performed. Ribotype (RT) 106 is a commonly isolated C. difficile strain worldwide; however, studies investigating RT 106 clinical outcomes are limited. The purpose of this study was to investigate clinical outcomes of RT 106 infections compared with two other endemic strains of varying virulence. METHODS: This multicenter study evaluated adults hospitalized with C. difficile infection (CDI). C. difficile samples underwent PCR ribotyping and patients infected with RT 106 were compared to patients infected with a known hypervirulent strain (RT 027) and a strain associated with less virulence (RT 014-020). Electronic medical records were reviewed by blinded investigators to assess the primary outcome of poor clinical outcome (composite of initial clinical failure, discharge to a higher level of care, 90-day CDI recurrence, and CDI-contributable mortality). RESULTS: A total of 396 patients with CDI were identified (RT 106, 32.3%; RT 027, 29.3%; RT 014-020, 38.3%). Patients infected with RT 014-020 less often experienced a poor clinical outcome (40%) compared with RT 106 (56%) and RT 027 (65%) infection (P < 0.0001). After controlling for covariates and using RT 014-020 as a comparator, patients infected with RT 106 (OR, 2.25; 95% CI, 1.36-3.73) or RT 027 (OR, 2.56; 95% CI, 1.52-4.31) had higher odds of poor clinical outcome. Using RT 027 as the comparator, only RT 014-020 was associated with lower odds of poor clinical outcome (OR, 0.42; 95% CI, 0.27-0.65). CONCLUSION: This study demonstrated that the emergent C. difficile RT 106 was associated with increased rates of poor clinical outcomes compared to RT 014-020 and comparable poor clinical outcomes to RT 027. These findings can help to better understand the clinical significance of this and future emerging ribotypes.

Visualization of fidaxomicin association with the exosporium layer of Clostridioides difficile spores.

BACKGROUND: Fidaxomicin has novel pharmacologic effects on C. difficile spore formation including outgrowth inhibition and persistent spore attachment. However, the mechanism of fidaxomicin attachment on spores has not undergone rigorous microscopic studies. MATERIALS & METHODS: Fidaxomicin attachment to C. difficile spores of three distinct ribotypes and C. difficile mutant spores with inactivation of exosporium or spore-coat protein-coding genes were visualized using confocal microscopy with a fidaxomicin-bodipy compound (green fluorescence). The pharmacologic effect of the fidaxomicin-bodipy compound was determined. Confocal microscopy experiments included direct effect on C. difficile wild-type and mutant spores, effect of exosporium removal, and direct attachment to a comparator spore forming organism, Bacillus subtilis. RESULTS: The fidaxomicin-bodipy compound MIC was 1 mg/L compared to 0.06 mg/L for unlabeled fidaxomicin, a 16-fold increase. Using confocal microscopy, the intracellular localization of fidaxomicin into vegetative C. difficile cells was observed consistent with its RNA polymerase mechanism of action and inhibited spore outgrowth. The fidaxomicin-bodipy compound was visualized outside of the core of C. difficile spores with no co-localization with the membrane staining dye FM4-64. Exosporium removal reduced fidaxomicin-bodipy association with C. difficile spores. Reduced fidaxomicin-bodipy was observed in C. difficile mutant spores for the spore surface proteins CdeC and CotE. CONCLUSION: This study visualized a direct attachment of fidaxomicin to C. difficile spores that was diminished with mutants of specific exosporium and spore coat proteins. These data provide advanced insight regarding the anti-spore properties of fidaxomicin.

Systems biology evaluation of refractory Clostridioides difficile infection including multiple failures of fecal microbiota transplantation.

BACKGROUND: Fecal microbiota transplantation (FMT) aims to cure Clostridioides difficile infection (CDI) through reestablishing a healthy microbiome and restoring colonization resistance. Although often effective after one infusion, patients with continued microbiome disruptions may require multiple FMTs. In this N-of-1 study, we use a systems biology approach to evaluate CDI in a patient receiving chronic suppressive antibiotics with four failed FMTs over two years. METHODS: Seven stool samples were obtained between 2016-18 while the patient underwent five FMTs. Stool samples were cultured for C. difficile and underwent microbial characterization and functional gene analysis using shotgun metagenomics. C. difficile isolates were characterized through ribotyping, whole genome sequencing, metabolic pathway analysis, and minimum inhibitory concentration (MIC) determinations. RESULTS: Growing ten strains from each sample, the index and first four recurrent cultures were single strain ribotype F078-126, the fifth was a mixed culture of ribotypes F002 and F054, and the final culture was ribotype F002. One single nucleotide polymorphism (SNP) variant was identified in the RNA polymerase (RNAP) ß-subunit RpoB in the final isolated F078-126 strain when compared to previous F078-126 isolates. This SNV was associated with metabolic shifts but phenotypic differences in fidaxomicin MIC were not observed. Microbiome differences were observed over time during vancomycin therapy and after failed FMTs. CONCLUSION: This study highlights the importance of antimicrobial stewardship in patients receiving FMT. Continued antibiotics play a destructive role on a transplanted microbiome and applies selection pressure for resistance to the few antibiotics available to treat CDI.

In Vitro Activity of Omadacycline, a New Tetracycline Analog, and Comparators against Clostridioides difficile.

Omadacycline is a potent aminomethylcycline with in vitro activity against Gram-positive, Gram-negative, and anaerobic bacteria. Preliminary data demonstrated that omadacycline has in vitro activity against Clostridioides difficile; however, large-scale in vitro studies have not been done. The purpose of this study was to assess the in vitro susceptibility of omadacycline and comparators on a large biobank of clinical C. difficile isolates. In vitroC. difficile susceptibility to omadacycline and comparators (fidaxomicin, metronidazole, and vancomycin) was assessed using the broth microdilution method. Minimum bactericidal concentrations (MBCs) and time-kill assays were assessed for pharmacodynamics analysis, and whole-genome sequencing was performed in a subset of isolates to assess distribution of MICs and resistance determinants. Two hundred fifty clinical C. difficile isolates collected between 2015 and 2018 were tested for in vitro susceptibility of omadacycline and comparators. Ribotypes included F001 (n = 5), F002 (n = 56), F014-020 (n = 66), F017 (n = 8), F027 (n = 53), F106 (n = 45), and F255 (n = 17). Omadacycline demonstrated potent in vitro activity with an MIC range of 0.016 to 0.13 µg/ml, an MIC50 of 0.031 µg/ml, and an MIC90 of 0.031 µg/ml. No difference was observed for omadacycline MIC50 and MIC90 values stratified by ribotype, disease severity, or vancomycin susceptibility. Bactericidal activity was confirmed in time-kill studies. No difference was observed in MIC based on C. difficile phylogeny. Further development of omadacycline as an intravenous and oral antibiotic directed toward C. difficile infection is warranted.

In vitro activity of eravacycline against common ribotypes of Clostridioides difficile.

BACKGROUND: Eravacycline is a novel synthetic fluorocycline antibacterial approved for complicated intra-abdominal infections. OBJECTIVES: The purpose of this study was to assess the in vitro activities of eravacycline and comparator antibiotics against contemporary clinical isolates of Clostridioides difficile representing common ribotypes, including isolates with decreased susceptibility to metronidazole and vancomycin. METHODS: Clinical C. difficile strains from six common or emerging ribotypes were used to test the in vitro activities of eravacycline and comparator antibiotics (fidaxomicin, vancomycin and metronidazole) by broth microdilution. In addition, MBC experiments, time-kill kinetic studies and WGS experiments were performed. RESULTS: A total of 234 isolates were tested, including ribotypes RT001 (n = 37), RT002 (n = 41), RT014-020 (n = 39), RT027 (n = 42), RT106 (n = 38) and RT255 (n = 37). MIC50/90 values were lowest for eravacycline (≤0.0078/0.016 mg/L), followed by fidaxomicin (0.016/0.063 mg/L), metronidazole (0.25/1.0 mg/L) and vancomycin (2.0/4.0 mg/L). MBCs were lower for eravacycline compared with vancomycin for all ribotypes tested. Both vancomycin and eravacycline demonstrated bactericidal killing, including for epidemic RT027. The presence of the tetM or tetW resistance genes did not affect the MIC of eravacycline. CONCLUSIONS: This study demonstrated potent in vitro activity of eravacycline against a large collection of clinical C. difficile strains that was not affected by ribotype, susceptibility to vancomycin or the presence of certain tet resistance genes. Further development of eravacycline as an antibiotic to be used in patients with Clostridioides difficile infection is warranted.

A randomized, double-blind, placebo-controlled, single and multiple ascending dose Phase 1 study to determine the safety, pharmacokinetics and food and faecal microbiome effects of ibezapolstat administered orally to healthy subjects.

BACKGROUND: Clostridioides difficile infection is the most common cause of healthcare-associated infections in the USA, with limited treatment options. Ibezapolstat is a novel DNA polymerase IIIC inhibitor with in vitro activity against C. difficile. OBJECTIVES AND METHODS: Randomized, double-blind, placebo-controlled study to assess the safety, tolerability and pharmacokinetics of ibezapolstat in healthy volunteers. Microbiome changes associated with ibezapolstat were compared with vancomycin over a 10 day course using shotgun metagenomics. RESULTS: A total of 62 subjects aged 31 ± 7 years (45% female; average BMI: 25 ± 3 kg/m2) were randomized. Ibezapolstat was well tolerated with a safety signal similar to placebo. Ibezapolstat had minimal systemic absorption with the majority of plasma concentrations less than 1 µg/mL. In the multiday, ascending dose study, ibezapolstat concentrations of 2000 µg/g of stool were observed by Day 2 and for the remainder of the dosing time period. In the multiday, multiple-dose arm, baseline microbiota was comparable between subjects that received ibezapolstat compared with vancomycin. At Day 10 of dosing, differential abundance analysis and ß-diversity demonstrated a distinct difference between the microbiome in subjects given vancomycin compared with either dose of ibezapolstat (P = 0.006). α-Diversity changes were characterized as an increase in the Actinobacteria phylum in subjects that received ibezapolstat and an increase in Proteobacteria in subjects given vancomycin. CONCLUSIONS: Ibezapolstat was shown to be safe and well tolerated, with minimal systemic exposure, high stool concentrations and a distinct microbiome profile compared with oral vancomycin. These results support further clinical development of ibezapolstat for patients with C. difficile infection.

Molecular epidemiology of toxigenic Clostridioides difficile isolates from hospitalized patients and the hospital environment in Dhaka, Bangladesh.

Epidemiology of Clostridioides difficile (syn. Clostridium difficile) infection (CDI) in Bangladesh is poorly understood. This study assessed the epidemiology of CDI in hospitalized patients and hospital environmental contamination of toxigenic C. difficile at two large urban Bangladesh hospitals. This 12-month prospective observational cohort study collected stool samples from adults with diarrhea and recent antimicrobial exposure during 2017. Environmental samples were collected by swabbing surfaces of hospital common areas. Samples underwent toxigenic culture. C. difficile isolates were tested for toxins A and B and PCR-ribotyped. Of 208 stool samples, 18 (8.7%) were positive for toxigenic C. difficile. Of 400 environmental samples, 45 (11%) were positive for toxigenic C. difficile. Ribotypes present in ≥10% of stool isolates were 017 (38%), 053-163 (13%), and a novel ribotype (FP435 [13%]). Common ribotypes in environmental isolates were 017 (22%), 053-163 (11%), 106 (24%). This is the first report describing current epidemiology of CDI in at risk hospitalized adult patients in Bangladesh.

Molecular epidemiology of Clostridioides difficile in domestic dogs and zoo animals.

Animals such as domestic dogs and zoo animals reside in close proximity to humans and could contribute to the dissemination of Clostridioides difficile spores which are common in the community environment. The purpose of this study was to assess C. difficile colonization in domestic dogs attending a day boarding facility and zoo animals receiving systemic antibiotics. Stool samples and paw swabs were collected from dogs who attended a day boarding facility. Stool samples were also collected from zoo animals starting systemic antibiotics. Finally, environmental samples were collected from nearby public parks. Stool samples and swabs were incubated anaerobically in enrichment broth for C. difficile growth, PCR was done to confirm presence of toxin genes, and PCR ribotyping was performed for strain characterization. During the study period, 136 dog stool samples were obtained, the paws of 16 dogs were swabbed, and 250 environmental swabs from surrounding public parks were obtained. Twenty-three of 136 dog stool samples (17%) and 9 of 16 dog paws sampled (56%) grew toxigenic C. difficile. One hundred and four stool samples from 49 zoo animals were collected of which 19 (18%) grew toxigenic C. difficile. Rates of toxigenic C. difficile colonization increased significantly during antibiotic therapy (33%) and then returned to baseline during the follow-up (11%) period (p = 0.019). Fifty-five of 250 environmental swabs from public parks (22%) grew toxigenic C. difficile. Ribotypes associated with human disease including 106 and 014-020 were isolated from all sources. This study demonstrated a high rate of toxigenic C. difficile colonization in domestic dogs and zoo animals with ribotypes similar to those causing human disease. These results demonstrate the relationship between humans, animals, and the environment in the dissemination of spores.

Clostridioides difficile ribotypes isolated from domestic environment and from patients in Bangladesh.

Clostridioides difficile infection (CDI) is an emerging but often understudied infectious disease in developing countries. This study was aimed to isolate and characterize C. difficile from shoe sole swabs and diarrheal patient's stool samples in Bangladesh. We collected 94 shoe sole swabs samples from urban communities in Dhaka and 208 diarrheal stool samples from hospitalized patients over a period of 4 months. Samples were incubated anaerobically for C. difficile growth, confirmed toxigenic, and PCR-ribotyped. Eleven of 94 (11.7%) shoe sole swabs and 4 of 208 (1.9%) stool samples were culture positive of which 9 shoe sole isolates were toxigenic. Six PCR ribotypes from the 9 toxigenic isolates were identified with ribotype F014-020 being the most common (n = 4; 44%). The recently identified ribotype 106 strain was also identified. To the best of our knowledge, this is the first report of C. difficile culture, isolation and characterization from environmental sources in Bangladesh.

The Dual Estrogen Receptor α Inhibitory Effects of the Tissue-Selective Estrogen Complex for Endometrial and Breast Safety.

The conjugated estrogen /: bazedoxifene tissue-selective estrogen complex (TSEC) is designed to minimize the undesirable effects of estrogen in the uterus and breast tissues and to allow the beneficial effects of estrogen in other estrogen-target tissues, such as the bone and brain. However, the molecular mechanism underlying endometrial and breast safety during TSEC use is not fully understood. Estrogen receptor α (ERα)-estrogen response element (ERE)-DNA pull-down assays using HeLa nuclear extracts followed by mass spectrometry-immunoblotting analyses revealed that, upon TSEC treatment, ERα interacted with transcriptional repressors rather than coactivators. Therefore, the TSEC-mediated recruitment of transcriptional repressors suppresses ERα-mediated transcription in the breast and uterus. In addition, TSEC treatment also degraded ERα protein in uterine tissue and breast cancer cells, but not in bone cells. Interestingly, ERα-ERE-DNA pull-down assays also revealed that, upon TSEC treatment, ERα interacted with the F-box protein 45 (FBXO45) E3 ubiquitin ligase. The loss-of- and gain-of-FBXO45 function analyses indicated that FBXO45 is involved in TSEC-mediated degradation of the ERα protein in endometrial and breast cells. In preclinical studies, these synergistic effects of TSEC on ERα inhibition also suppressed the estrogen-dependent progression of endometriosis. Therefore, the endometrial and breast safety effects of TSEC are associated with synergy between the selective recruitment of transcriptional repressors to ERα and FBXO45-mediated degradation of the ERα protein.

Life course effects on age at menopause among Bangladeshi sedentees and migrants to the UK.

OBJECTIVES: To assess how different variables experienced across the life course, but particularly during early life, might affect age at menopause among 174 Bangladeshi migrants to London by comparing them to 157 nonmigrant sedentees and 154 women of European descent in London. METHODS: Participants were aged 35-59 years, with no exogenous hormone use in the past three months, not pregnant or lactating, with no history of hysterectomy or oophorectomy. Face-to-face interviews and anthropometric measures were carried out. In addition to mean recalled age at natural menopause, median age was computed by probit analysis. Ages at menopause were examined by bivariate and Cox regression analyses in relation to demographic, reproductive, and lifestyle variables, and in relation to potential exposure to cyclones in early childhood. RESULTS: Ages at menopause were significantly earlier among Bangladeshi sedentees and immigrants compared to Londoners of European origin. Ages at menopause were earlier among sedentees compared to immigrants. Urban birthplace, more infectious diseases during childhood, and lower levels of education increased the risk of an earlier menopause. CONCLUSIONS: Changes in environmental conditions during adulthood appeared to modify age at menopause among Bangladeshi immigrants in London compared to women living in Bangladesh; however, Bangladeshi immigrants still experienced an earlier age at menopause compared with their London neighbors of European descent.