Biology subjects

Hoyland-Kroghsbo, N. M.

Publications and source records attributed to Hoyland-Kroghsbo, N. M..

6 recordsLinked to original sources

bioRxiv2026

Slow to Start, Free at Last: Dual Effects of Mucin on Escherichia coli Phage T4

Bacteriophages traversing the gastrointestinal tract are exposed to extreme physicochemical stresses that may rapidly compromise virion integrity and shape infection dynamics. While some phages bind to host-derived mucins at mucosal surfaces, the functional consequences of soluble mucin glycans for phage-host interactions remain incompletely understood. Here, we show that soluble mucin glycans exert dual effects on the Escherichia coli phage T4 by delaying infection initiation while simultaneously providing environmental virion stability. Mucin-coated T4 exhibits a lag in the onset of productive infection, consistent with transient steric occlusion from E. coli, yet without impairing overall phage progeny production once infection was established. We further show that E. coli can metabolize purified mucin, supporting a model in which dynamic remodeling of the mucin matrix gradually releases T4 and enables infection. Importantly, mucin coating substantially increases T4 survival under gastrointestinal-like stresses, including acidic pH and protease exposure. Moreover, we find that in a murine gut colonization model, a single oral dose of mucin-coated T4 displayed enhanced fecal persistence over a two-week period, which correlated with prolonged suppression of E. coli populations and delayed resolution of phage-associated functional shifts in the gut microbiome. Together, we find that that soluble mucin glycans actively shape T4 phage infection kinetics, virion stability, and ecological impact in the murine gut, and support mucin-based formulations as a strategy to extend the persistence and efficacy of orally delivered phages.

microbiology↗

bioRxiv2026

Anti-Quorum Sensing Phages Disarm Pseudomonas aeruginosa

By 2050, the death toll from previously preventable or easily curable bacterial infections is projected to surpass that caused by cancer, unless we prevent the spread of antibiotic resistance and develop new therapies. A promising approach is phage therapy, which exploits bacteriophages, natural predators of bacteria. However, bacteria fight back, which can limit its efficacy. Notably, many bacteria rely on cell-cell communication, known as quorum sensing, to orchestrate both virulence programs and phage defenses. To circumvent these, we have engineered anti-quorum sensing phages against the human pathogen Pseudomonas aeruginosa. Our engineered phages effectively degrade quorum-sensing molecules, reduce virulence factor production, and double the survival of P. aeruginosa-infected Galleria mellonella larvae. Moreover, we demonstrate that the anti-quorum sensing phages inhibit quorum sensing in mixed populations of phage-susceptible and phage-resistant cells, demonstrating the ability of the phages to disarm subpopulations phage-resistant P. aeruginosa, which often are selected for during phage treatment. Together, our findings highlight the future therapeutic promise of anti-quorum sensing phages as a dual-action strategy in killing susceptible cells while attenuating virulence across the bacterial population. This approach has the potential to enhance the robustness of phage therapy.

microbiology↗

bioRxiv2025

Overlooked signals: Highly stable quorum sensing molecule in phage lysates induces quorum sensing response

Phage-bacterial interaction studies routinely apply phage lysates at final concentrations of up to 10% of the culture. Consequently, bacterial metabolites such as quorum sensing (QS) signaling molecules are transferred along with the phage lysate to recipient bacterial cells. Here, we show that the Pseudomonas aeruginosa QS molecules 3OC12-HSL and C4-HSL are rapidly degraded in phage lysates. In contrast, the hydrophobic QS molecule PQS is remarkably stable, for at least one year, due to its binding within outer membrane vesicles. Strikingly, we find that PQS exceeds concentrations of 10 {micro}M in standard phage lysate preparations. We show that PQS carried over from phage lysates induces QS-controlled production of the virulence factor pyocyanin in P. aeruginosa. This PQS carryover does not oppose previous conclusions of phage infection-induced PQS production, as we show here, that this response is also triggered by PQS-free phage lysates. Since other bacterial species, including Paracoccus and Vibrio harveyi, also produce hydrophobic QS molecules that are bound within outer membrane vesicles, it is likely that phage lysates from these bacteria may similarly contain stable QS molecules. Collectively, we demonstrate that membrane-bound QS molecules may significantly confound QS-related physiological outcomes of phage-host interaction studies. This can be avoided by using QS synthase mutants for phage propagation or by purifying phage particles from lysates to eliminate QS molecule carryover.

microbiology↗

bioRxiv2023

The great divide: rhamnolipids mediate separation between P. aeruginosa and S. aureus

The coexistence of multiple bacterial species during infection can have significant impacts on pathogenesis. Pseudomonas aeruginosa and Staphylococcus aureus are opportunistic bacterial pathogens that can co-infect hosts and cause serious illness. The factors that dictate whether one species will outcompete the other or whether the two species can coexist are not fully understood. We investigated the role of surfactants in the interactions between these two species on a surface that enables P. aeruginosa to swarm. We found that P. aeruginosa swarms are repelled by colonies of clinical S. aureus isolates, creating physical separation between the two strains. This effect was abolished in mutants of S. aureus that were defective in the production of phenol-soluble modulins (PSMs), which form amyloid fibrils around wild-type colonies. We investigated the mechanism that establishes physical separation between the two species using the Imaging of Reflected Illuminated Structures (IRIS) method, which tracks the flow of the rhamnolipid surfactant layer produced by P. aeruginosa. We found that PSMs produced by S. aureus deflected the rhamnolipid surfactant layer flow, which in turn, altered the direction of P. aeruginosa swarms. These findings show that rhamnolipids mediate physical separation between P. aeruginosa and S. aureus, which enables these species to coexist in distinct microenvironments. Additionally, we found that a Bacillus subtilis surfactant and abiotic hydrophobic molecules repelled P. aeruginosa swarms through surfactant deflection. Our results suggest that surfactant interactions could have major impacts on bacteria-bacteria and bacteria-host relationships. In addition, our findings uncover a mechanism responsible for P. aeruginosa swarm development that does not rely on sensing but instead is guided largely by the flow of the surfactant layer and its boundaries.

microbiology↗

bioRxiv2023

Quorum sensing inhibits Type III-A CRISPR-Cas system activity through repressing positive regulators SarA and ArcR in Staphylococcus aureus

CRISPR-Cas is an adaptive immune system that protects prokaryotes from the invasion of foreign genetic elements. The components and immunity mechanisms of CRISPR-Cas have been extensively studied, but the regulation of this system in Staphylococci remains unclear. Here, we show that in the S. aureus Type III-A CRISPR-Cas system, the Pcas of 300 bp located in cas1 displays as a critical regulatory node that initiates the transcription of cas gene clusters. We discovered two transcriptional regulators, SarA and CRP-like ArcR, promote the expression and activity of the CRISPR-Cas system by directly binding to the novel promoter Pcas. Furthermore, we demonstrated that the cell-cell communication, known as quorum sensing (QS), inhibits the activity of CRISPR-Cas system by repressing the positive regulators SarA and ArcR. Bioinformatic analyses suggest that Pcas is conserved in many Type II and III CRISPR-Cas systems in Firmicutes. Our data reveal a new regulatory mechanism for QS-mediated repression of the Type III-A CRISPR-Cas system, which may allow S. aureus to acquire foreign genetic elements encoding antibiotic resistance or virulence factors specifically at high cell density.

microbiology↗

bioRxiv2021

Phage infection restores PQS signaling and enhances growth of a Pseudomonas aeruginosa lasI quorum-sensing mutant

Bacteriophage (phage) therapy is reemerging as a valuable tool to combat multidrug resistant bacteria. A major hurdle in developing efficacious bacteriophage therapies is that bacteria acquire resistance to phage killing. In this context, it is noteworthy that quorum sensing (QS), the bacterial cell-to-cell communication mechanism that promotes collective undertaking of group behaviors including anti-phage defenses, enhances bacterial survival in the face of phage attack. QS relies on the production, release, accumulation, and detection of signal molecules called autoinducers. In the opportunistic pathogen Pseudomonas aeruginosa, the LasI/R QS system induces the RhlI/R QS system, and these two systems control, in opposing manners, the PQS QS system that relies on the autoinducer called PQS. A {Delta}lasI mutant is impaired in PQS synthesis, leading to accumulation of the precursor molecule HHQ. HHQ suppresses growth of the P. aeruginosa {Delta}lasI strain. We uncover a phage infection-induced mechanism that restores expression of the pqsH gene in the P. aeruginosa {Delta}lasI QS mutant. PqsH converts HHQ into PQS, preventing HHQ-mediated growth inhibition. Thus, phage-infected P. aeruginosa {Delta}lasI cells exhibit superior growth compared to uninfected cells. Phage infection also restores expression of virulence factors and the CRISPR-cas anti-phage defense system in the P. aeruginosa {Delta}lasI strain. This study highlights a challenge for phage therapy, namely that phage infection may make particular bacterial strains faster growing, more virulent, and resistant to phage killing. ImportanceThe emergence of multidrug resistant bacteria necessitates development of new antimicrobial therapies. Phage therapy relies on exploiting phages, natural enemies of bacteria, in the fight against pathogenic bacteria. For successful phage therapy development, potent phages that exhibit low propensity for acquisition of bacterial resistance are desired. Here, we show that phage infection restores QS, a cell-to-cell communication mechanism in a P. aeruginosa QS mutant, which increases its virulence and resistance to phage killing. Importantly, clinical isolates of P. aeruginosa frequently harbor mutations in particular QS genes. Thus, phage therapies against such P. aeruginosa strains may inadvertently increase bacterial virulence. Our study underscores the importance of characterizing phage-host interactions in the context of bacterial mutants that are relevant in clinical settings prior to selecting phages for therapy.

microbiology↗