Biology subjects

Hunter, R. C.

Publications and source records attributed to Hunter, R. C..

4 recordsLinked to original sources

bioRxiv2018

Agmatine accumulation by Pseudomonas aeruginosa clinical isolates confers antibiotic tolerance and dampens host inflammation.

In the cystic fibrosis (CF) airways, Pseudomonas aeruginosa undergoes diverse physiological changes in response to inflammation, antibiotic pressure, oxidative stress and a dynamic bioavailable nutrient pool. These include loss-of-function mutations that result in reduced virulence, altered metabolism and other phenotypes that are thought to confer a selective advantage for long-term persistence. Recently, clinical isolates of P. aeruginosa that hyperproduce agmatine (decarboxylated arginine) were cultured from individuals with CF. Sputum concentrations of this metabolite were also shown to correlate with disease severity. This raised the question of whether agmatine accumulation might also confer a selective advantage for P. aeruginosa in vivo during chronic colonization of the lung. Here, we screened a library of P. aeruginosa CF clinical isolates and found that ~5% of subjects harbored isolates with an agmatine hyperproducing phenotype. Agmatine accumulation was a direct result of mutations in aguA, encoding the arginine deiminase that catalyzes the conversion of agmatine into various polyamines. We also found that agmatine hyperproducing isolates (aguA-) had increased tolerance to the cationic antibiotics gentamicin, tobramycin and colistin relative to their chromosomally complemented strains (aguA+). Finally, we revealed that agmatine diminishes IL-8 production by airway epithelial cells in response to bacterial infection, with a consequent decrease in neutrophil recruitment to the murine airways in an acute pneumonia model. These data highlight a potential new role for bacterial-derived agmatine that may have important consequences for the long-term persistence of P. aeruginosa in the CF airways.

microbiology↗

bioRxiv2018

Cross-feeding modulates antibiotic tolerance in bacterial communities

Microbes frequently rely on metabolites excreted by other bacterial species, but little is known about how this cross-feeding influences the effect of antibiotics. We hypothesized that when species rely on each other for essential metabolites, the minimum inhibitory concentration (MIC) for all species will drop to that of the \"weakest link\" - the species least resistant in monoculture. We tested this hypothesis in an obligate cross-feeding system that was engineered between Escherichia coli, Salmonella enterica, and Methylobacterium extorquens. The effect of tetracycline and ampicillin were tested on both liquid and solid media. In all cases, resistant species were inhibited at significantly lower antibiotic concentrations in the cross-feeding community than in monoculture or a competitive community. However, deviation from the \"weakest link\" hypothesis was also observed in cross-feeding communities apparently as result of changes in the timing of growth and cross-protection. Comparable results were also observed in a clinically relevant system involving facultative cross-feeding between Pseudomonas aeruginosa and an anaerobic consortium found in the lungs of cystic fibrosis patients. P. aeruginosa was inhibited by lower concentrations of ampicillin when cross-feeding than when grown in isolation. These results suggest that cross-feeding significantly alters tolerance to antibiotics in a variety of systems.

microbiology↗

bioRxiv2017

16S rRNA Gene Sequencing Reveals Site-Specific Signatures Of The Upper And Lower Airways Of Cystic Fibrosis Patients

RationaleChronic rhinosinusitis (CRS) is an inflammatory disorder of the sinonasal mucosa associated with microbial colonization. Metastasis of sinus microbiota into the lower airways is thought have significant implications in the development of chronic respiratory disease. However, this dynamic has not been thoroughly investigated in cystic fibrosis (CF) patients, where lower airway infections are the primary driver of patient mortality. Given the high prevalence of CRS in CF patients and the proposed infection dynamic between the upper and lower airways, a better understanding of sinus-lung continuum is warranted.\n\nObjectiveTo compare the microbiome of matched sinus mucus and lung sputum samples from CF subjects undergoing functional endoscopic sinus surgery (FESS) for treatment of CRS.\n\nMethodsMucus was isolated from the sinuses and lungs of twelve CF patients undergoing FESS. 16S ribosomal RNA gene sequencing was then performed to compare bacterial communities of the CF lung and sinus niches. Finally, functional profiling was performed to predict bacterial metagenomes from the 16S dataset, and was used to compare pathogenic bacterial phenotypes between the upper and lower airways.\n\nMeasurements and Main ResultsBacterial richness was comparable between airway sites, though sinus and lung environments differed in community evenness, with the sinuses harboring a higher prevalence of dominant microorganisms. Beta diversity metrics also revealed that samples clustered more consistently by airway niche rather than by individual. Finally, predicted metagenomes showed that anaerobic metabolism was enriched in the lung environment, while genes associated with both biofilm formation and Gram identity were not variable between sites.\n\nConclusionsSinus and lung microbiomes are distinct with respect to richness and evenness, while sinus communities have a higher incidence of a dominant taxon. Additionally, ordination analyses point to sinus and lung environments as being stronger determinants of microbial community structure than the individual patient. Finally, BugBase-predicted metagenomes revealed anaerobic phenotypes to be in higher abundance in the lung relative to the sinuses. Our findings indicate that while the paranasal sinuses and lungs may still comprise a unified airway in which lower airways are seeded by sinus microbiota, these discrete airway microenvironments harbor distinct bacterial communities.

microbiology↗

bioRxiv2016

Genome-wide survey of Pseudomonas aeruginosa PA14 reveals a role for the glyoxylate pathway and extracellular proteases in the limited utilization of mucin

Chronic airway infections by the opportunistic pathogen Pseudomonas aeruginosa are major cause of mortality in cystic fibrosis (CF) patients. While this bacterium has been extensively studied for its virulence mechanisms, biofilm growth and immune evasion within the CF airways, comparatively little is known about the nutrient sources that sustain its growth in vivo. Respiratory mucins represent a potentially abundant bioavailable nutrient source, though we have recently shown that canonical pathogens inefficiently use these host glycoproteins as a growth substrate. Yet, given that P. aeruginosa, particularly in its biofilm mode of growth is thought to grow slowly in vivo, the inefficient use of mucin glycoproteins may have relevance to its persistence within the CF airways. To this end, here we use whole genome fitness analysis combining transposon mutagenesis with high throughput sequencing (TnSeq) to identify genetic determinants required for P. aeruginosa growth using intact purified mucins as a sole carbon source. Our analysis reveals a biphasic growth phenotype, during which the glyoxylate pathway and amino acid biosynthetic machinery are required for mucin utilization. Secondary analyses confirmed the simultaneous liberation and consumption of acetate during mucin degradation and revealed a central role for the extracellular proteases LasB and AprA. Together, these studies identified the P. aeruginosa genes required for mucin-based nutrient acquisition and reveal a host-pathogen dynamic that may contribute to its persistence within the CF airways.

microbiology↗