Biology subjects

Barrack, K.

Publications and source records attributed to Barrack, K..

7 recordsLinked to original sources

bioRxiv2025

The Regional Landscape of the Human Colon Culturome in Health and Cystic Fibrosis

Cystic fibrosis (CF) alters gut physiology, yet its impact on microbial communities across colonic regions (ascending, transverse, descending colon) and microhabitats (lumen, mucosa) remains incompletely understood. Here, we applied culturomics to characterize gut microbiota in 32 individuals (22 nonCF, 10 CF). Persons with CF (pwCF) exhibited significantly higher viable bacterial loads than nonCF individuals, particularly in mucosal samples. Anaerobes predominated overall, with relative enrichment of aerobes in the mucosa of pwCF. Alpha diversity was reduced in mucosal samples and aerobic cultures for pwCF, whereas beta diversity was influenced by all the tested variables except the colonic region. Phylum-level analyses revealed enrichment of Proteobacteria and depletion of Actinobacteria, Bacteroidota, and Firmicutes in samples from pwCF, consistent with stool analysis. Random forest models identified selected oral-associated microbes as key predictive taxa and accurately classified polyp status with very high accuracy. Whole-genome sequencing of Bacteroides fragilis (n=21) and Escherichia coli (n=15) isolates, representing a subset of 109 gut bacterial genomes sequenced from this cohort, revealed minimal genomic variation across colonic regions and sample types, indicating intra-individual strain stability. The understandings from this pilot culturome study may help in developing targeted microbial therapeutic approaches to address the gut dysbiosis of CF. ImportanceThis pilot study represents the first culturome analysis of the cystic fibrosis colon. Our preliminary findings demonstrate that CF-associated gut dysbiosis is spatially specific, with mucosal bacterial communities showing pronounced alterations while luminal communities remain unchanged. This spatial specificity suggests the mucosal microenvironment as a potential therapeutic target and indicates that interventions focused solely on luminal bacteria may be insufficient. The promising predictive accuracy of culturome-based machine learning models in this small cohort suggests these viable bacterial signatures could serve as biomarkers for CF management pending larger validation studies. Additionally, our initial observations of complex CFTR modulator effects on gut microbial communities provide insights for future studies optimizing combination therapies.

microbiology↗

bioRxiv2025

A genotoxin associated with colorectal cancer linked to gut dysbiosis in children with cystic fibrosis

Cystic fibrosis (CF) substantially alters the gastrointestinal microbiome from an early age, leading to significant changes in microbial composition and functionality. This study explores the physiological and microbiological factors contributing to dysbiosis in children with cystic fibrosis (cwCF), characterized by an increase in potentially pathogenic Escherichia coli and a decrease in beneficial anaerobes such as Bacteroides. In this study, we employed an in vitro medium representative of the nutritional environment of the CF colon to test the role of factors including mucin, fat, bile, pH, antibiotics and features associated with inflammation (e.g., nitrate, sulfate, formate, reactive oxygen species) on growth of clinical isolates of E. coli and Bacteroides spp. We further examined interactions between these two microbes under CF-like conditions to understand modulators of microbial competition, and identified glycerol, a surrogate of increased fat, as a significant driver of altered microbial competition. Finally, we investigated genetic determinants influencing these microbial interactions, with the focus on glycerol metabolism, by performing a transposon mutagenesis screen in E. coli. Results of this screen pointed to the role of colibactin production in mediating this microbial competition; colibactin is a DNA-damaging genotoxin associated with the increased risk of colorectal cancer (CRC) in CF populations. This work enhances our understanding of mechanisms of microbial competition in the CF gut, while potentially enhancing our understanding of colorectal cancer risk in persons with CF through the identification of early-life microbial biomarkers. Significance StatementThe risk of CRC development in CF populations is significantly increased. This study examines the interplay of altered intestinal physiology in the microbial dysbiosis common in the CF gut, implicating the high fat environment in a competition-mediated depletion of immune-modulating Bacteroides. This work identifies candidate features of the young CF intestine and gut microbiome that may contribute to advanced development of CRC in these populations, informing potential therapeutic approaches.

microbiology↗

bioRxiv2023

An In Vitro Medium for Modeling Gut Dysbiosis Associated with Cystic Fibrosis

The gut physiology of pediatric and adult persons with cystic fibrosis (pwCF) is altered relative to healthy persons. The CF gut is characterized, in part, as having excess mucus, increased fat content, acidic pH, increased inflammation, increased antibiotic perturbation and the potential for increased oxygen availability. These physiological differences shift nutritional availability and the local environment for intestinal microbes, thus likely driving significant changes in microbial metabolism, colonization and competition with other microbes. The impact of any specific change in this physiological landscape is difficult to parse using human or animal studies. Thus, we have developed a novel culture medium representative of the CF gut environment, inclusive of all the aforementioned features. This medium, called CF-MiPro, maintains CF gut microbiome communities, while significantly shifting non-CF gut microbiome communities toward a CF-like microbial profile, characterized by low Bacteroidetes and high Proteobacteria abundance. This medium is able to maintain this culture composition for up to 5 days of passage. Additionally, microbial communities passaged in CF-MiPro produce significantly less immunomodulatory short chain fatty acids (SCFA), including propionate and butyrate, than communities passaged in MiPro, a culture medium representative of healthy gut physiology, confirming not only a shift in microbial composition but altered community function. Our results support the potential for this in vitro culture medium as a new tool for the study of gut dysbiosis in CF. ImportanceCystic fibrosis is an autosomal recessive disease that disrupts ion transport at mucosal surfaces, leading to mucus accumulation and altered physiology of both the lungs and the intestines, among other organs, with the resulting altered environment contributing to an imbalance of microbial communities. Culture media representative of the CF airway have been developed and validated; however, no such medium exists for modeling the CF intestine. Here, we develop and validate a first-generation culture medium inclusive of features that are altered in the CF colon. Our findings suggest this novel medium, called CF-MiPro, as a maintenance medium for CF gut microbiome samples and a flexible tool for studying key drivers of CF-associated gut dysbiosis.

microbiology↗

bioRxiv2022

Predicting Clinical Outcomes in Infants With Cystic Fibrosis From Stool Microbiota using Random Forest Algorithms

The gut-lung axis describes the interaction between microbiota in the gut and health status of the airway, whereby there is a bidirectional relationship facilitated by systemic transport of microbially-derived metabolites and immune factors. Cystic fibrosis (CF) is a genetic disease that is associated with dysbiosis of the gut microbiota. Recent literature has shown that the microbial dysbiosis in the CF gut can alter the hosts inflammatory status and that there are distinct microbial compositions in children with CF who have low versus high intestinal inflammation. These distinct microbial profiles can be used as signatures in children with CF (cwCF) to predict health outcomes. Here, we use supervised machine learning to train a random forest model on the distinct microbial composition of cwCF to predict: age (as a validation of the method), frequency of upper respiratory infection (URIfreq), and neutrophil to lymphocyte ratio (NLR), a clinical marker for systemic inflammation that negatively correlates with lung function. We find that the out of bag error, a measure of model accuracy, is lower when predicting age for cwCF compared to children without CF, consistent with previous data. We are able to predict high URIfreq with only 16% error and high NLR with 27% error. This machine learning pipeline may allow physicians and microbiome researchers to use the stool microbiota of cwCF as a tool for identifying individuals with the more negative airway clinical outcomes from this population, and potentially allow for early intervention. ImportanceChildren with CF (cwCF) often experience chronic respiratory infections, leading to progressive, irreversible lung function decline and significant morbidity and premature mortality. Modulator therapy has revolutionized the treatment of eligible adults with CF. cwCF as young as age 6 are now eligible to receive modulator therapy, although by early childhood many children have already experienced pulmonary exacerbations. Here we show that for cwCF, stool microbiota composition is associated with higher upper respiratory infection frequency and increased systemic inflammation. Our findings may aid in developing diagnostic tools that can allow physicians further understanding of which intestinal microbiota profiles are associated with health outcomes and to identify targets for preventative treatment for cwCF.

microbiology↗

bioRxiv2022

Development of the Intestinal Microbiome in Cystic Fibrosis in Early Life

Cystic Fibrosis is a heritable disease that causes altered physiology at mucosal sites; these changes result in chronic infection in the lung, significant gastrointestinal complications as well as dysbiosis of the gut microbiome, although the latter has been less well explored. Here, we describe the longitudinal development of the gut microbiome in a cohort of children with cystic fibrosis (cwCF) from birth through early childhood (0-4 years of age) using 16S rRNA gene amplicon sequencing of stool samples as a surrogate for the gut microbiota. Similar to healthy populations, alpha diversity of the gut microbiome increases significantly with age, but diversity plateaus [~]2 years of age for this CF cohort. Several taxa that have been associated with dysbiosis in CF change with age towards a more healthy-like composition; notable exceptions include Akkermansia, which decreases with age, and Blautia, which increases with age. We also examined the relative abundance and prevalence of nine taxa associated with CF lung disease, several of which persist across early life, highlighting the possibility of the lung being seeded directly from the gut early in life. Finally, we applied the Crohns dysbiosis index to each sample, and found that high Crohns-associated dysbiosis early in life (<2 years) was associated with significantly lower Bacteroides in samples collected from 2-4 years of age. Together, these data indicate a persisting dysbiosis in the gut microbiota as well as markers associated with inflammatory bowel disease early in life for cwCF. IMPORTANCECystic Fibrosis is a heritable disease that disrupts ion transport at mucosal surfaces, causing a buildup of mucus and dysregulation of microbial communities in both the lungs and the intestines. Persons with CF are known to have dysbiotic gut microbial communities, but the development of these communities over time beginning at birth have not been thoroughly studied. Here, we describe the development of the gut microbiome of cwCF throughout the first four years of life, during the critical window of both gut microbiome and immune development. Our findings indicate a persisting dysbiosis, the possibility of the gut microbiota as a reservoir of airway pathogens and a surprisingly early indication of a microbiota associated with inflammatory bowel disease.

microbiology↗

bioRxiv2022

Intestinal Bacteroides Modulates Systemic Inflammation and the Microbial Ecology in a Mouse Model of CF: Evidence for Propionate and other Short Chain Fatty Acids Reducing Systemic Inflammatory Cytokines

Persons with cystic fibrosis, starting in early life, have intestinal microbiome dysbiosis characterized in part by a decreased relative abundance of the genus Bacteroides. Bacteroides is a major producer of the intestinal short chain fatty acid (SCFA) propionate. We demonstrate here that CFTR-/- Caco-2 intestinal epithelial cells are responsive to the anti-inflammatory effects of propionate. Furthermore, Bacteroides isolates inhibit the IL-1{beta}-induced inflammatory response of CFTR-/- Caco-2 intestinal epithelial cells and do so in a propionate-dependent manner. Bacteroides isolates also produce low levels of butyrate; this SCFA is positively correlated with inhibition of the inflammatory response. Finally, the introduction of Bacteroides-supplemented stool from infants with CF into the gut of CftrF508del mice results in an increase in propionate in the stool as well as the reduction in several systemic pro-inflammatory cytokines. Bacteroides supplementation also reduced the fecal relative abundance of E. coli, indicating a potential interaction between these two microbes, consistent with previous clinical studies. Together, our data indicate the important role of Bacteroides and Bacteroides-derived propionate in the context of the developing microbiome in infants and children with CF, which could help explain the observed gut-lung axis in CF.

microbiology↗

bioRxiv2021

The Diguanylate Cyclase YfiN of Pseudomonas aeruginosa Regulates Biofilm Maintenance in Response to Peroxide

Pseudomonas aeruginosa forms surface-attached communities that persist in the face of antimicrobial agents and environmental perturbation. Published work has found extracellular polysaccharide (EPS) production, regulation of motility and induction of stress response pathways as contributing to biofilm tolerance during such insults. However, little is known regarding the mechanism(s) whereby biofilm maintenance is regulated when exposed to such environmental challenges. Here, we provide evidence that the diguanylate cyclase YfiN is important for the regulation of biofilm maintenance when exposed to peroxide. We find that, compared to the wild type (WT), static biofilms of the {Delta}yfiN mutant exhibit a maintenance defect, which can be further exacerbated by exposure to peroxide (H2O2); this defect can be rescued through genetic complementation. Additionally, we found that the {Delta}yfiN mutant biofilms produce less c-di-GMP than WT, and that H2O2 treatment enhanced motility of surface-associated bacteria and increased cell death for the {Delta}yfiN mutant grown as a biofilm compared to WT biofilms. These data provide evidence that YfiN is required for biofilm maintenance by P. aeruginosa, via c-di-GMP signaling, to limit motility and protect viability in response to peroxide stress. These findings add to the growing recognition that biofilm maintenance by P. aeruginosa is an actively regulated process that is controlled, at least in part, by the wide array of c-di-GMP metabolizing enzymes found in this microbe. ImportanceWe build on previous findings that suggest that P. aeruginosa utilizes c-di-GMP metabolizing enzymes to actively maintain a mature biofilm. Here, we explore how the diguanylate cyclase YfiN contributes to the regulation of biofilm maintenance during peroxide exposure. We find that mature P. aeruginosa biofilms require YfiN to synthesize c-di-GMP, regulate motility and to insure viability during peroxide stress. These findings provide further evidence that the modulation of c-di-GMP in response to environmental signals is an important mechanism by which biofilms are maintained.

microbiology↗