Biology subjects

Kent, L. A.

Publications and source records attributed to Kent, L. A..

4 recordsLinked to original sources

bioRxiv2025

The acquisition of rmpADC can increase virulence of classical Klebsiella pneumoniae in the absence of other hypervirulence-associated genes

Klebsiella pneumoniae is one of the most common causes of nosocomial infections, and the rise of drug-resistant K. pneumoniae strains is complicating treatment and contributing to a mounting global health crisis. K. pneumoniae has two pathotypes, classical (cKp) and hypervirulent (hvKp). CKp typically cause opportunistic infections in immunocompromised individuals in healthcare settings and often are multi-drug resistant. HvKp can be community acquired and cause high-mortality infections in immunocompetent individuals. Concerningly, antibiotic-resistant cKp strains with hypervirulence-associated genes and traits have recently emerged. Determining if and how hv-associated genes contribute to increased virulence of cKp strains is essential to addressing this growing threat. The rmpADC operon is an hv-associated locus that confers hypermucoviscosity (HMV), a key virulence phenotype, and rmp genes are often found in convergent strains. In this study, we aimed to determine if the rmp genes alone could increase the virulence of classical strains in the absence of other hv-associated genes. We introduced genetically distinct rmp loci from different lineages into a broad array of cKp isolates and found that, while many isolates became HMV positive, only a subset of these strains showed an increase in virulence in a mouse model of pneumonia. Sequence type and capsule type were not predictive of how rmp acquisition impacted the clinical isolates. Our results indicate that HMV is likely necessary but not sufficient for hypervirulence and that rmp sequence can influence virulence potential in classical strains. IMPORTANCEKlebsiella pneumoniae is a global pathogen, and gene exchange between hypervirulent (hvKp) and classical (cKp) strains is a rising threat. It is essential to understand how hvKp genes impact virulence phenotypes and identify the cKp strain backgrounds most amenable to enhanced virulence. Hypermucoviscosity (HMV) is a critical virulence factor in hypervirulent K. pneumoniae, conferred by the rmpADC locus. The rmp genes are encoded on mobile genetic elements and have been detected in convergent antibiotic-resistant K. pneumoniae strains of concern. In this study, we explored the impact of rmp acquisition in a broad set of classical clinical isolates. We observed that HMV appears necessary, but not sufficient, for increased virulence. Sequence type, capsule type, and HMV capacity could not predict which classical isolates gain an rmp-dependent colonization benefit. These insights increase our understanding of the distinctions between cKp and hvKp and further our ability to identify and treat new strains of concern.

microbiology↗

bioRxiv2025

Anaerobic microbiota facilitate pathogen access to the airway epithelium in a novel co-culture model of colonization

Chronic rhinosinusitis (CRS) is a prevalent condition characterized by mucus stasis, persistent inflammation, and infection of the paranasal sinuses. It often involves the bacterium Pseudomonas aeruginosa, especially in individuals with cystic fibrosis or a history of antibiotic use. While P. aeruginosa is a known opportunistic pathogen that employs a diverse array of virulence factors to cause airway infections, its ability to thrive in the sinonasal environment is also likely influenced by its local microbial ecology. For instance, anaerobic bacterial genera, such as Streptococcus, Veillonella, and Prevotella, are also commonly found in CRS and may contribute to P. aeruginosa persistence. Here we sought to test the hypothesis that anaerobes promote P. aeruginosa colonization of the upper airways, specifically through degradation of mucin glycoproteins that decorate the epithelial surface. Using a novel dual oxic-anoxic culture platform termed DOAC, we co-cultured Calu-3 epithelial cells with a CRS-derived anaerobic microbial community. We observed increased expression of inflammatory marker genes and degradation of mucin glycoproteins, along with enhanced P. aeruginosa colonization of the epithelial surface after anaerobe pre-treatment. Furthermore, mucins isolated from anaerobe-treated Calu-3 cells promoted greater P. aeruginosa attachment to microtiter plates in vitro compared to intact mucins. These results suggest that anaerobic microbiota may shape the sinonasal environment in a way that favors P. aeruginosa persistence, offering new insights into CRS pathogenesis and potential therapeutic targets to disrupt detrimental bacterial interactions in chronic airway disease. IMPORTANCEThe prevalence and abundance of strict and facultative anaerobic bacteria in chronic sinusitis as detected by culture-independent sequencing has renewed interest in their potential role(s) in disease onset, progression, and treatment. However, reductionist study of interactions between anaerobic microbiota and the host has been limited by the lack of laboratory models compatible with their conflicting oxygen demands. The significance of this work lies in the use of a novel co-culture platform, termed DOAC, to interrogate anaerobe interactions with the airway epithelium. We use this platform to show that anaerobic CRS microbiota elicit a pro-inflammatory response, degrade mucin glycoproteins, and promote mucosal colonization by the canonical CRS pathogen, Pseudomonas aeruginosa. This work highlights highlight a potential role of anaerobic microbiota in conditioning the sinonasal environment to favor pathogen colonization.

microbiology↗

bioRxiv2022

An IS-mediated, RecA-dependent, bet-hedging strategy in Burkholderia thailandensis

Adaptation to fluctuating environmental conditions is difficult to achieve. Phase variation mechanisms can overcome this difficulty by altering genomic architecture in a subset of individuals, creating a phenotypically heterogeneous population with subpopulations optimized to persist when conditions change, or are encountered, suddenly. We have identified a phase variation system in Burkholderia thailandensis that generates a genotypically and phenotypically heterogeneous population. Genetic analyses revealed that RecA-mediated homologous recombination between a pair of insertion sequence (IS) 2-like elements duplicates a 208.6 kb region that contains 157 coding sequences. RecA-mediated homologous recombination also resolves merodiploids, and hence copy number of the region is varied and dynamic within populations. We showed that the presence of two or more copies of the region is advantageous for growth in a biofilm, and a single copy is advantageous during planktonic growth. While IS elements are well-known to contribute to evolution through gene inactivation, polar effects on downstream genes, and altering genomic architecture, we believe that this system represents a rare example of IS element-mediated evolution in which the IS elements provide homologous sequences for amplification of a chromosomal region that provides a selective advantage under specific growth conditions, thereby expanding the lifestyle repertoire of the species.

microbiology↗

bioRxiv2021

Anaerobic microbiota facilitate P. aeruginosa access to airway epithelia in a novel co-culture model of colonization

Strict and facultative anaerobic bacteria are widely associated with both acute and chronic airway disease. However, their potential role(s) in disease pathophysiology remain poorly understood due to inherent limitations of existing models and conflicting oxygen demands between anaerobes and host cells. To overcome these limitations, we optimized a dual oxic-anoxic culture (DOAC) platform that maintains an oxygen-limited microenvironment at the epithelial interface while host cells are oxygenated basolaterally. This approach enables host-bacterial co-culture for [~]24h, and here we demonstrate its utility via two applications. First, we show that anaerobe challenge results in epithelial mucus degradation, inflammatory marker gene expression, and enhanced pathogen colonization. Second, we combine DOAC with single cell RNA sequencing (scRNAseq) to reveal a cell type-specific transcriptional response of the airway epithelium to anaerobe infection. Together, these data illustrate the versatility of DOAC while revealing new insights into anaerobe-host interactions and their mechanistic contributions to airway disease pathophysiology.

microbiology↗