Biology subjects

Darch, S. E.

Publications and source records attributed to Darch, S. E..

6 recordsLinked to original sources

bioRxiv2026

Microbial Crosstalk in the Neonatal Gut: Candida albicans Impacts Group B Streptococcus Pathogenicity

Background Group B Streptococcus (GBS) is a leading cause of bacterial sepsis globally. Late-onset (LO) GBS disease is preceded by colonization within the gastrointestinal tract in vulnerable neonates. Previous studies demonstrate that Candida albicans (C. albicans) promotes GBS colonization of host tissues including the bladder and vagina. However, its impact on GBS colonization of the intestinal tract is unknown. Methods We established a new animal model to study GBS and C. albicans co-colonization model in the context of the newborn host to examine C. albicans impact on GBS and further evaluated fluconazole prophylactic treatment to these same outcomes. Physical interactions between organisms in vivo were imaged and GBS gene expression was evaluated. Results C. albicans increased GBS burden in the neonatal gut and GBS was physically colocalized with C. albicans hyphae as evidenced by the colocalization analysis. Fluconazole treatment of co-colonized mice decreased extraintestinal GBS dissemination. C. albicans presence upregulated genes encoding proteins responsible for GBS adhesion and invasion of intestinal epithelial cells. Conclusion C. albicans enhances GBS burden and pathogenicity in the neonatal gut. These findings highlight the need for further investigation into C. albicans as a potential risk factor for GBS colonization and LO disease in human infants.

microbiology↗

bioRxiv2026

Host immune stress reveals a role for potassium homeostasis in Pseudomonas aeruginosa aggregate stability

Pseudomonas aeruginosa (Pa) forms multicellular aggregates during chronic airway infection, yet the physiological processes that regulate aggregate organization and dispersal remain incompletely understood. Here, we investigated how Pa aggregates respond to human neutrophil elastase (HNE), a host-derived protease abundant in the cystic fibrosis airway. Transcriptomic analysis of HNE-exposed aggregates identified strong induction of the kdpFABC high-affinity potassium transport system, with greater induction in aggregate than planktonic populations. Functional analysis of kdpA revealed a broader role in aggregate physiology: loss of kdpA increased total biomass while reducing average aggregate volume and accelerating dispersal, including in the absence of HNE. Genetic complementation restored key kdpA-dependent phenotypes, whereas co-culture with wild-type cells did not rescue the mutant. Increasing extracellular potassium modified aggregate volume, biomass, and dispersal in the kdpA mutant but did not uniformly restore these phenotypes to wild-type levels, demonstrating that kdpA-dependent aggregate behavior is potassium-responsive. Together, these findings identify KdpA as a link between potassium homeostasis and Pa aggregate behavior and suggest that host-derived stress increases engagement of a homeostatic system that contributes to the coordination of population growth, aggregate organization, and dispersal.

microbiology↗

bioRxiv2025

AFM characterization of early P. aeruginosa aggregates highlights emergent mechanical properties

Pseudomonas aeruginosa (Pa) is a leading cause of chronic lung infections in people with cystic fibrosis (pwCF), where its ability to form resilient, multicellular communities contributes to antibiotic tolerance and long-term persistence. While much of our understanding of Pa biofilms comes from surface-attached models, recent studies have emphasized the clinical relevance of suspended bacterial aggregates - dense, three-dimensional clusters that form early during infection and exhibit key biofilm-like properties. However, the physical characteristics of these aggregates remain poorly defined. Here, we apply atomic force microscopy (AFM) to visualize and quantify the structural and mechanical properties of Pa aggregates formed in synthetic cystic fibrosis sputum medium (SCFM2). Compared to planktonic cultures grown without mucin, aggregates formed in SCFM2 exhibited complex architecture and increased resistance to deformation, as measured by force spectroscopy. These differences emerged despite the absence of mature extracellular matrix components, suggesting that environmental cues and spatial organization alone may be sufficient to enhance aggregate mechanical resilience. Our results demonstrate that AFM provides a powerful, high-resolution approach for studying early-stage bacterial aggregates under physiologically relevant conditions. By resolving structural features and quantifying localized mechanical strength, this method offers new insight into how aggregate architecture contributes to persistence during chronic infection. These findings lay the groundwork for future studies targeting the physical robustness of bacterial communities as an early vulnerability in the pathogenesis of Pa both in CF and in other infection settings.

microbiology↗

bioRxiv2024

Exploring aggregation genes in a P. aeruginosa chronic infection model

Bacterial aggregates are observed in both natural and artificial environments. In the context of disease, aggregates have been isolated from both chronic and acute infections. Pseudomonas aeruginosa (Pa) aggregates contribute significantly to chronic infections, particularly in the lungs of people with cystic fibrosis (CF). Unlike the large biofilm structures observed in vitro, Pa in CF sputum forms smaller aggregates ([~]10-1000 cells), and the mechanisms behind their formation remain underexplored. This study aims to identify genes essential and unique to Pa aggregate formation in a synthetic CF sputum media (SCFM2). We cultured Pa strain PAO1 in SCFM2 and LB, both with and without mucin, and used RNA sequencing (RNA-seq) to identify differentially expressed genes. The presence of mucin revealed hundreds of significantly differentially expressed (DE) genes, predominantly downregulated, with 40% encoding hypothetical proteins unique to aggregates. Using high-resolution microscopy, we assessed the ability of mutants to form aggregates and identified 13 that were unable to form WT aggregates. Notably, no mutant exhibited a completely planktonic phenotype. Instead, we identified multiple spatial phenotypes described as normal, entropic, or impaired. Entropic mutants displayed tightly packed, raft-like structures, while impaired mutants had loosely packed cells. Predictive modeling linked the prioritized genes to metabolic shifts, iron acquisition, surface modification, and quorum sensing. Co-culture experiments with wild-type PAO1 revealed further spatial heterogeneity and the ability to rescue some mutant phenotypes, suggesting cooperative interactions during growth. This study enhances our understanding of Pa aggregate biology, specifically the genes and pathways unique to aggregation in CF-like environments. Importantly, it provides insights for developing therapeutic strategies targeting aggregate-specific pathways. ImportanceThis study identifies genes essential for the formation of Pseudomonas aeruginosa (Pa) aggregates in cystic fibrosis (CF) sputum, filling a critical gap in understanding their specific biology. Using a synthetic CF sputum model (SCFM2) and RNA sequencing, 13 key genes were identified, whose disruption led to distinct spatial phenotypes observed through high-resolution microscopy. The addition of wild-type cells either rescued the mutant phenotype or increased spatial heterogeneity, suggesting cooperative interactions are involved in aggregate formation. This research advances our knowledge of Pa aggregate biology, particularly the unique genes and pathways involved in CF-like environments, offering valuable insights for developing targeted therapeutic strategies against aggregate-specific pathways.

microbiology↗

bioRxiv2023

Group B Streptococcus drives major transcriptomic changes in the colonic epithelium.

Group B Streptococcus (GBS) is a leading cause of infant sepsis worldwide. Colonization of the gastrointestinal tract is a critical precursor to late-onset disease in exposed newborns. Neonatal susceptibility to GBS intestinal translocation stems from intestinal immaturity; however, the mechanisms by which GBS exploits the immature host remain unclear. {beta}-hemolysin/cytolysin ({beta}H/C) is a highly conserved toxin produced by GBS capable of disrupting epithelial barriers. However, its role in the pathogenesis of late-onset GBS disease is unknown. Our aim was to determine the contribution of {beta}H/C to intestinal colonization and translocation to extraintestinal tissues. Using our established mouse model of late-onset GBS disease, we exposed animals to GBS COH-1 (WT), a {beta}H/C-deficient mutant (KO), or vehicle control (PBS) via gavage. Blood, spleen, brain, and intestines were harvested 4 days post-exposure for determination of bacterial burden and isolation of intestinal epithelial cells. RNA-sequencing was used to examine the transcriptomes of host cells followed by gene ontology enrichment and KEGG pathway analysis. A separate cohort of animals was followed longitudinally to compare colonization kinetics and mortality between WT and KO groups. We demonstrate that dissemination to extraintestinal tissues occurred only in the WT exposed animals. We observed major transcriptomic changes in the colons of colonized animals, but not in the small intestines. We noted differential expression of genes that indicated the role of {beta}H/C in altering epithelial barrier structure and immune response signaling. Overall, our results demonstrate an important role of {beta}H/C in the pathogenesis of late-onset GBS disease.

microbiology↗

bioRxiv2022

Volatile hydrogen cyanide released by Pseudomonas aeruginosa provides a competitive advantage over Staphylococcus aureus in biofilm and in vivo lung environments

Diverse bacterial volatile compounds alter bacterial stress responses and physiology, but their contribution to population dynamics in polymicrobial communities is not well known. In this study, we showed that airborne volatile hydrogen cyanide (HCN) produced by a wide range of Pseudomonas aeruginosa clinical strains leads to at-a-distance inhibition of the growth of a wide array of Staphylococcus aureus strains. We determined that low oxygen level environments not only enhance P. aeruginosa HCN production but also increase S. aureus sensitivity to HCN, which impacts P. aeruginosa-S. aureus competition in microaerobic in vitro mixed biofilms as well as in an in vitro cystic fibrosis lung sputum medium. Consistently, we demonstrated that production of HCN provides a competitive advantage to P. aeruginosa in a mouse model of airways co-infected by P. aeruginosa and S. aureus. Our study therefore demonstrates that P. aeruginosa HCN contributes to local and distant airborne competition against S. aureus and potentially other HCN-sensitive bacteria in contexts relevant to cystic fibrosis and other polymicrobial infectious diseases. IMPORTANCEAirborne volatile compounds produced by bacteria are often only considered as attractive or repulsive scents, but they also directly contribute to bacterial physiology. Here we showed that volatile hydrogen cyanide (HCN) released by a wide range of Pseudomonas aeruginosa clinical strains inhibits Staphylococcus aureus growth in low oxygen in vitro biofilms or aggregates and in vivo lung environments. These results are of pathophysiological relevance, since lungs of cystic fibrosis patients are known to present microaerophilic areas and to be commonly associated with the presence of S. aureus and P. aeruginosa in polymicrobial communities. Our study therefore provides insights into how a bacterial volatile compound can contribute to the exclusion of S. aureus and other HCN-sensitive competitors from P. aeruginosa ecological niches. It opens new perspectives for the management or monitoring of P. aeruginosa infections in lower lung airway infections and other polymicrobial disease contexts.

microbiology↗