Mapping niche-specific two-component system requirements in uropathogenic Escherichia coli.

Sensory systems allow pathogens to differentiate between different niches and respond to stimuli within them. A major mechanism through which bacteria sense and respond to stimuli in their surroundings is two-component systems (TCSs). TCSs allow for the detection of multiple stimuli to lead to a highly controlled and rapid change in gene expression. Here, we provide a comprehensive list of TCSs important for the pathogenesis of uropathogenic Escherichia coli (UPEC). UPEC accounts for >75% of urinary tract infections (UTIs) worldwide. UTIs are most prevalent among people assigned female at birth, with the vagina becoming colonized by UPEC in addition to the gut and the bladder. In the bladder, adherence to the urothelium triggers E. coli invasion of bladder cells and an intracellular pathogenic cascade. Intracellular E. coli are safely hidden from host neutrophils, competition from the microbiota, and antibiotics that kill extracellular E. coli. To survive in these intimately connected, yet physiologically diverse niches E. coli must rapidly coordinate metabolic and virulence systems in response to the distinct stimuli encountered in each environment. We hypothesized that specific TCSs allow UPEC to sense these diverse environments encountered during infection with built-in redundant safeguards. Here, we created a library of isogenic TCS deletion mutants that we leveraged to map distinct TCS contributions to infection. We identify-for the first time-a comprehensive panel of UPEC TCSs that are critical for infection of the genitourinary tract and report that the TCSs mediating colonization of the bladder, kidneys, or vagina are distinct.IMPORTANCEWhile two-component system (TCS) signaling has been investigated at depth in model strains of Escherichia coli, there have been no studies to elucidate-at a systems level-which TCSs are important during infection by pathogenic Escherichia coli. Here, we report the generation of a markerless TCS deletion library in a uropathogenic E. coli (UPEC) isolate that can be leveraged for dissecting the role of TCS signaling in different aspects of pathogenesis. We use this library to demonstrate, for the first time in UPEC, that niche-specific colonization is guided by distinct TCS groups.

Longitudinal profiling of the intestinal microbiome in children with cystic fibrosis treated with elexacaftor-tezacaftor-ivacaftor.

The intestinal microbiome influences growth and disease progression in children with cystic fibrosis (CF). Elexacaftor-tezacaftor-ivacaftor (ELX/TEZ/IVA), the newest pharmaceutical modulator for CF, restores the function of the pathogenic mutated CF transmembrane conductance regulator (CFTR) channel. We performed a single-center longitudinal analysis of the effect of ELX/TEZ/IVA on the intestinal microbiome, intestinal inflammation, and clinical parameters in children with CF. Following ELX/TEZ/IVA, children with CF had significant improvements in body mass index and percent predicted forced expiratory volume in one second, and required fewer antibiotics for respiratory infections. Intestinal microbiome diversity increased following ELX/TEZ/IVA coupled with a decrease in the intestinal carriage of Staphylococcus aureus, the predominant respiratory pathogen in children with CF. There was a reduced abundance of microbiome-encoded antibiotic resistance genes. Microbial pathways for aerobic respiration were reduced after ELX/TEZ/IVA. The abundance of microbial acid tolerance genes was reduced, indicating microbial adaptation to increased CFTR function. In all, this study represents the first comprehensive analysis of the intestinal microbiome in children with CF receiving ELX/TEZ/IVA.IMPORTANCECystic fibrosis (CF) is an autosomal recessive disease with significant gastrointestinal symptoms in addition to pulmonary complications. Recently approved treatments for CF, CF transmembrane conductance regulator (CFTR) modulators, are anticipated to substantially improve the care of people with CF and extend their lifespans. Prior work has shown that the intestinal microbiome correlates with health outcomes in CF, particularly in children. Here, we study the intestinal microbiome of children with CF before and after the CFTR modulator, ELX/TEZ/IVA. We identify promising improvements in microbiome diversity, reduced measures of intestinal inflammation, and reduced antibiotic resistance genes. We present specific bacterial taxa and protein groups which change following ELX/TEZ/IVA. These results will inform future mechanistic studies to understand the microbial improvements associated with CFTR modulator treatment. This study demonstrates how the microbiome can change in response to a targeted medication that corrects a genetic disease.

Raising the alarm: fosfomycin resistance associated with non-susceptible inner colonies imparts no fitness cost to the primary bacterial uropathogen.

IMPORTANCE: While fosfomycin resistance is rare, the observation of non-susceptible subpopulations among clinical Escherichia coli isolates is a common phenomenon during antimicrobial susceptibility testing (AST) in American and European clinical labs. Previous evidence suggests that mutations eliciting this phenotype are of high biological cost to the pathogen during infection, leading to current recommendations of neglecting non-susceptible colonies during AST. Here, we report that the most common route to fosfomycin resistance, as well as novel routes described in this work, does not impair virulence in uropathogenic E. coli, the major cause of urinary tract infections, suggesting a re-evaluation of current susceptibility guidelines is warranted.

Survey of the infant male urobiome and genomic analysis of Actinotignum spp.

The urinary bladder harbors a community of microbes termed the urobiome, which remains understudied. In this study, we present the urobiome of healthy infant males from samples collected by transurethral catheterization. Using a combination of enhanced culture and amplicon sequencing, we identify several common bacterial genera that can be further investigated for their effects on urinary health across the lifespan. Many genera were shared between all samples suggesting a consistent urobiome composition among this cohort. We note that, for this cohort, early life exposures including mode of birth (vaginal vs. Cesarean section), or prior antibiotic exposure did not influence urobiome composition. In addition, we report the isolation of culturable bacteria from the bladders of these infant males, including Actinotignum spp., a bacterial genus that has been associated with urinary tract infections in older male adults. Herein, we isolate and sequence 9 distinct strains of Actinotignum spp. enhancing the genomic knowledge surrounding this genus and opening avenues for delineating the microbiology of this urobiome constituent. Furthermore, we present a framework for using the combination of culture-dependent and sequencing methodologies for uncovering mechanisms in the urobiome.

The QseB response regulator imparts tolerance to positively charged antibiotics by controlling metabolism and minor changes to LPS.

The modification of lipopolysaccharide (LPS) in Escherichia coli and Salmonella spp. is primarily controlled by the two-component system PmrAB. LPS modification allows bacteria to avoid killing by positively charged antibiotics like polymyxin B (PMB). We previously demonstrated that in uropathogenic E. coli (UPEC), the sensor histidine kinase PmrB also activates a non-cognate transcription factor, QseB, and this activation somehow augments PMB tolerance in UPEC. Here, we demonstrate-for the first time-that in the absence of the canonical LPS transcriptional regulator, PmrA, QseB can direct some modifications on the LPS. In agreement with this observation, transcriptional profiling analyses demonstrate regulatory overlaps between PmrA and QseB in terms of regulating LPS modification genes. However, both PmrA and QseB must be present for UPEC to mount robust tolerance to PMB. Transcriptional and metabolomic analyses also reveal that QseB transcriptionally regulates the metabolism of glutamate and 2-oxoglutarate, which are consumed and produced during the modification of lipid A. We show that deletion of qseB alters glutamate levels in the bacterial cells. The qseB deletion mutant, which is susceptible to positively charged antibiotics, is rescued by exogenous addition of 2-oxoglutarate. These findings uncover a previously unknown mechanism of metabolic control of antibiotic tolerance that may be contributing to antibiotic treatment failure in the clinic. IMPORTANCE Although antibiotic prescriptions are guided by well-established susceptibility testing methods, antibiotic treatments oftentimes fail. The presented work is significant because it uncovers a mechanism by which bacteria transiently avoid killing by antibiotics. This mechanism involves two closely related transcription factors, PmrA and QseB, which are conserved across Enterobacterales. We demonstrate that PmrA and QseB share regulatory targets in lipid A modification pathway and prove that QseB can orchestrate modifications of lipid A in Escherichia coli in the absence of PmrA. Finally, we show that QseB controls glutamate metabolism during the antibiotic response. These results suggest that rewiring of QseB-mediated metabolic genes could lead to stable antibiotic resistance in subpopulations within the host, thereby contributing to antibiotic treatment failure.

Longitudinal Profiling of the Intestinal Microbiome in Children with Cystic Fibrosis Treated with Elexacaftor-Tezacaftor-Ivacaftor.

The intestinal microbiome influences growth and disease progression in children with cystic fibrosis (CF). Elexacaftor-tezacaftor-ivacaftor (ELX/TEZ/IVA), the newest pharmaceutical modulator for CF, restores function of the pathogenic mutated CFTR channel. We performed a single-center longitudinal analysis of the effect of ELX/TEZ/IVA on the intestinal microbiome, intestinal inflammation, and clinical parameters in children with CF. Following ELX/TEZ/IVA, children with CF had significant improvements in BMI, ppFEV1 and required fewer antibiotics for respiratory infections. Intestinal microbiome diversity increased following ELX/TEZ/IVA coupled with a decrease in the intestinal carriage of Staphylococcus aureus, the predominant respiratory pathogen in children with CF. There was a reduced abundance of microbiome-encoded antibiotic-resistance genes. Microbial pathways for aerobic respiration were reduced after ELX/TEZ/IVA. The abundance of microbial acid tolerance genes was reduced, indicating microbial adaptation to increased CFTR function. In all, this study represents the first comprehensive analysis of the intestinal microbiome in children with CF receiving ELX/TEZ/IVA.

Mapping Niche-specific Two-Component System Requirements in Uropathogenic Escherichia coli.

Sensory systems allow pathogens to differentiate between different niches and respond to stimuli within them. A major mechanism through which bacteria sense and respond to stimuli in their surroundings is two-component systems (TCSs). TCSs allow for the detection of multiple stimuli to lead to a highly controlled and rapid change in gene expression. Here, we provide a comprehensive list of TCSs important for the pathogenesis of uropathogenic Escherichia coli (UPEC). UPEC accounts for >75% of urinary tract infections (UTIs) worldwide. UTIs are most prevalent among people assigned female at birth, with the vagina becoming colonized by UPEC in addition to the gut and the bladder. In the bladder, adherence to the urothelium triggers E. coli invasion of bladder cells and an intracellular pathogenic cascade. Intracellular E. coli are safely hidden from host neutrophils, competition from the microbiota, and antibiotics that kill extracellular E. coli. To survive in these intimately connected, yet physiologically diverse niches E. coli must rapidly coordinate metabolic and virulence systems in response to the distinct stimuli encountered in each environment. We hypothesized that specific TCSs allow UPEC to sense these diverse environments encountered during infection with built-in redundant safeguards. Here, we created a library of isogenic TCS deletion mutants that we leveraged to map distinct TCS contributions to infection. We identify - for the first time - a comprehensive panel of UPEC TCSs that are critical for infection of the genitourinary tract and report that the TCSs mediating colonization of the bladder, kidneys, or vagina are distinct.

The Role of Mobile Genetic Elements in Virulence Factor Carriage from Symptomatic and Asymptomatic Cases of Escherichia coli Bacteriuria.

Uropathogenic Escherichia coli (UPEC) is extremely diverse genotypically and phenotypically. Individual strains can variably carry diverse virulence factors, making it challenging to define a molecular signature for this pathotype. For many bacterial pathogens, mobile genetic elements (MGEs) constitute a major mechanism of virulence factor acquisition. For urinary E. coli, the total distribution of MGEs and their role in the acquisition of virulence factors is not well defined, including in the context of symptomatic infection versus asymptomatic bacteriuria (ASB). In this work, we characterized 151 isolates of E. coli, derived from patients with either urinary tract infection (UTI) or ASB. For both sets of E. coli, we catalogued the presence of plasmids, prophage, and transposons. We analyzed MGE sequences for the presence of virulence factors and antimicrobial resistance genes. These MGEs were associated with only ~4% of total virulence associated genes, while plasmids contributed to ~15% of antimicrobial resistance genes under consideration. Our analyses suggests that, across strains of E. coli, MGEs are not a prominent driver of urinary tract pathogenesis and symptomatic infection. IMPORTANCE Escherichia coli is the most common etiological agent of urinary tract infections (UTIs), with UTI-associated strains designated "uropathogenic" E. coli or UPEC. Across urinary strains of E. coli, the global landscape of MGEs and its relationship to virulence factor carriage and clinical symptomatology require greater clarity. Here, we demonstrate that many of the putative virulence factors of UPEC are not associated with acquisition due to MGEs. The current work enhances our understanding of the strain-to-strain variability and pathogenic potential of urine-associated E. coli and points toward more subtle genomic differences distinguishing ASB from UTI isolates.

Defining the Infant Male Urobiome and Moving Towards Mechanisms in Urobiome Research.

The urinary bladder harbors a community of microbes termed the urobiome, which remains understudied. In this study, we present the urobiome of healthy infant males from samples collected by transurethral catheterization. Using a combination of extended culture and amplicon sequencing, we identify several common bacterial genera that can be further investigated for their effects on urinary health across the lifespan. Many genera were shared between all samples suggesting a consistent urobiome composition among this cohort. We note that, for this cohort, early life exposures including mode of birth (vaginal vs. Caesarean section), or prior antibiotic exposure did not influence urobiome composition. In addition, we report the isolation of culturable bacteria from the bladders of these infant males, including Actinotignum schaalii, a bacterial species that has been associated with urinary tract infection in older male adults. Herein, we isolate and sequence 9 distinct strains of A. schaalii enhancing the genomic knowledge surrounding this species and opening avenues for delineating the microbiology of this urobiome constituent. Furthermore, we present a framework for using the combination of culture-dependent and sequencing methodologies for uncovering mechanisms in the urobiome.

The QseB response regulator imparts tolerance to positively charged antibiotics by controlling metabolism and minor changes to LPS.

The modification of lipopolysaccharide (LPS) in Escherichia coli and Salmonella spp . is primarily controlled by the two-component system PmrAB. LPS modification allows bacteria to avoid killing by positively charged antibiotics like polymyxin B. We previously demonstrated that in uropathogenic E. coli (UPEC), the sensor histidine kinase PmrB also activates a non-cognate transcription factor, QseB, and this activation somehow augments polymyxin B tolerance in UPEC. Here, we demonstrate - for the first time - that in the absence of the canonical LPS transcriptional regulator, PmrA, QseB can direct some modifications on the LPS. In agreement with this observation, transcriptional profiling analyses demonstrate regulatory overlaps between PmrA and QseB in terms of regulating LPS modification genes. However, both PmrA and QseB must be present for UPEC to mount robust tolerance to polymyxin B. Transcriptional and metabolomic analyses also reveal that QseB transcriptionally regulates the metabolism of glutamate and 2-oxoglutarate, which are consumed and produced during the modification of lipid A. We show that deletion of qseB alters glutamate levels in the bacterial cells. The qseB deletion mutant, which is susceptible to positively charged antibiotics, is rescued by exogenous addition of 2-oxoglutarate. These findings uncover a previously unknown mechanism of metabolic control of antibiotic tolerance that may be contributing to antibiotic treatment failure in the clinic. IMPORTANCE: Although antibiotic prescriptions are guided by well-established susceptibility testing methods, antibiotic treatments oftentimes fail. The presented work is significant, because it uncovers a mechanism by which bacteria transiently avoid killing by antibiotics. This mechanism involves two closely related transcription factors, PmrA and QseB, which are conserved across Enterobacteriaceae. We demonstrate that PmrA and QseB share regulatory targets in lipid A modification pathway and prove that QseB can orchestrate modifications of lipid A in E. coli in the absence of PmrA. Finally, we show that QseB controls glutamate metabolism during the antibiotic response. These results suggest that rewiring of QseB-mediated metabolic genes can lead to stable antibiotic resistance in subpopulations within the host, thereby contributing to antibiotic treatment failure.

Functional HIV-1/HCV cross-reactive antibodies isolated from a chronically co-infected donor.

Despite prolific efforts to characterize the antibody response to human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) mono-infections, the response to chronic co-infection with these two ever-evolving viruses is poorly understood. Here, we investigate the antibody repertoire of a chronically HIV-1/HCV co-infected individual using linking B cell receptor to antigen specificity through sequencing (LIBRA-seq). We identify five HIV-1/HCV cross-reactive antibodies demonstrating binding and functional cross-reactivity between HIV-1 and HCV envelope glycoproteins. All five antibodies show exceptional HCV neutralization breadth and effector functions against both HIV-1 and HCV. One antibody, mAb688, also cross-reacts with influenza and coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We examine the development of these antibodies using next-generation sequencing analysis and lineage tracing and find that somatic hypermutation established and enhanced this reactivity. These antibodies provide a potential future direction for therapeutic and vaccine development against current and emerging infectious diseases. More broadly, chronic co-infection represents a complex immunological challenge that can provide insights into the fundamental rules that underly antibody-antigen specificity.

Serine Deamination Is a New Acid Tolerance Mechanism Observed in Uropathogenic Escherichia coli.

Escherichia coli associates with humans early in life and can occupy several body niches either as a commensal in the gut and vagina, or as a pathogen in the urinary tract. As such, E. coli has an arsenal of acid response mechanisms that allow it to withstand the different levels of acid stress encountered within and outside the host. Here, we report the discovery of an additional acid response mechanism that involves the deamination of l-serine to pyruvate by the conserved l-serine deaminases SdaA and SdaB. l-serine is the first amino acid to be imported in E. coli during growth in laboratory media. However, there remains a lack in knowledge as to how l-serine is utilized. Using a uropathogenic strain of E. coli, UTI89, we show that in acidified media, l-serine is brought into the cell via the SdaC transporter. We further demonstrate that deletion of the l-serine deaminases SdaA and SdaB renders E. coli susceptible to acid stress, similar to other acid stress deletion mutants. The pyruvate produced by l-serine deamination activates the pyruvate sensor BtsS, which in concert with the noncognate response regulator YpdB upregulates the putative transporter YhjX. Based on these observations, we propose that l-serine deamination constitutes another acid response mechanism in E. coli. IMPORTANCE The observation that l-serine uptake occurs as E. coli cultures grow is well established, yet the benefit E. coli garners from this uptake remains unclear. Here, we report a novel acid tolerance mechanism where l-serine is deaminated to pyruvate and ammonia, promoting survival of E. coli under acidic conditions. This study is important as it provides evidence of the use of l-serine as an acid response strategy, not previously reported for E. coli.

Redox-Mediated Inactivation of the Transcriptional Repressor RcrR is Responsible for Uropathogenic Escherichia coli's Increased Resistance to Reactive Chlorine Species.

The ability to overcome stressful environments is critical for pathogen survival in the host. One challenge for bacteria is the exposure to reactive chlorine species (RCS), which are generated by innate immune cells as a critical part of the oxidative burst. Hypochlorous acid (HOCl) is the most potent antimicrobial RCS and is associated with extensive macromolecular damage in the phagocytized pathogen. However, bacteria have evolved defense strategies to alleviate the effects of HOCl-mediated damage. Among these are RCS-sensing transcriptional regulators that control the expression of HOCl-protective genes under non-stress and HOCl stress. Uropathogenic Escherichia coli (UPEC), the major causative agent of urinary tract infections (UTIs), is particularly exposed to infiltrating neutrophils during pathogenesis; however, their responses to and defenses from HOCl are still completely unexplored. Here, we present evidence that UPEC strains tolerate higher levels of HOCl and are better protected from neutrophil-mediated killing compared with other E. coli. Transcriptomic analysis of HOCl-stressed UPEC revealed the upregulation of an operon consisting of three genes, one of which encodes the transcriptional regulator RcrR. We identified RcrR as a HOCl-responsive transcriptional repressor, which, under non-stress conditions, is bound to the operator and represses the expression of its target genes. During HOCl exposure, however, the repressor forms reversible intermolecular disulfide bonds and dissociates from the DNA resulting in the derepression of the operon. Deletion of one of the target genes renders UPEC significantly more susceptible to HOCl and phagocytosis indicating that the HOCl-mediated induction of the regulon plays a major role for UPEC's HOCl resistance. IMPORTANCE How do pathogens deal with antimicrobial oxidants produced by the innate immune system during infection? Uropathogenic Escherichia coli (UPEC), the most common etiological agent of urinary tract infections (UTIs), is particularly exposed to infiltrating neutrophils and, therefore, must counter elevated levels of the antimicrobial oxidant HOCl to establish infection. Our study provides fundamentally new insights into a defense mechanism that enables UPEC to fend off the toxic effects of HOCl stress. Intriguingly, the defense system is predominantly found in UPEC and absent in noninvasive enteropathogenic E. coli. Our data suggest expression of the target gene rcrB is exclusively responsible for UPEC's increased HOCl tolerance in culture and contributes to UPEC's survival during phagocytosis. Thus, this novel HOCl stress defense system could potentially serve as an attractive drug target to increase the body's own capacity to fight UTIs.

Uropathogenic Escherichia coli subverts mitochondrial metabolism to enable intracellular bacterial pathogenesis in urinary tract infection.

Urinary tract infections are among the most common human bacterial infections and place a significant burden on healthcare systems due to associated morbidity, cost and antibiotic use. Despite being a facultative anaerobe, uropathogenic Escherichia coli, the primary cause of urinary tract infections, requires aerobic respiration to establish infection in the bladder. Here, by combining bacterial genetics with cell culture and murine models of infection, we demonstrate that the widely conserved respiratory quinol oxidase cytochrome bd is required for intracellular infection of urothelial cells. Through a series of genetic, biochemical and functional assays, we show that intracellular oxygen scavenging by cytochrome bd alters mitochondrial physiology by reducing the efficiency of mitochondrial respiration, stabilizing the hypoxia-inducible transcription factor HIF-1 and promoting a shift towards aerobic glycolysis. This bacterially induced rewiring of host metabolism antagonizes apoptosis, thereby protecting intracellular bacteria from urothelial cell exfoliation and preserving their replicative niche. These results reveal the metabolic basis for intracellular bacterial pathogenesis during urinary tract infection and identify subversion of mitochondrial metabolism as a bacterial strategy to facilitate persistence within the urinary tract.

BioKIT: a versatile toolkit for processing and analyzing diverse types of sequence data.

Bioinformatic analysis-such as genome assembly quality assessment, alignment summary statistics, relative synonymous codon usage, file format conversion, and processing and analysis-is integrated into diverse disciplines in the biological sciences. Several command-line pieces of software have been developed to conduct some of these individual analyses, but unified toolkits that conduct all these analyses are lacking. To address this gap, we introduce BioKIT, a versatile command line toolkit that has, upon publication, 42 functions, several of which were community-sourced, that conduct routine and novel processing and analysis of genome assemblies, multiple sequence alignments, coding sequences, sequencing data, and more. To demonstrate the utility of BioKIT, we conducted a comprehensive examination of relative synonymous codon usage across 171 fungal genomes that use alternative genetic codes, showed that the novel metric of gene-wise relative synonymous codon usage can accurately estimate gene-wise codon optimization, evaluated the quality and characteristics of 901 eukaryotic genome assemblies, and calculated alignment summary statistics for 10 phylogenomic data matrices. BioKIT will be helpful in facilitating and streamlining sequence analysis workflows. BioKIT is freely available under the MIT license from GitHub (https://github.com/JLSteenwyk/BioKIT), PyPi (https://pypi.org/project/jlsteenwyk-biokit/), and the Anaconda Cloud (https://anaconda.org/jlsteenwyk/jlsteenwyk-biokit). Documentation, user tutorials, and instructions for requesting new features are available online (https://jlsteenwyk.com/BioKIT).

A tRNA modifying enzyme as a tunable regulatory nexus for bacterial stress responses and virulence.

Post-transcriptional modifications can impact the stability and functionality of many different classes of RNA molecules and are an especially important aspect of tRNA regulation. It is hypothesized that cells can orchestrate rapid responses to changing environmental conditions by adjusting the specific types and levels of tRNA modifications. We uncovered strong evidence in support of this tRNA global regulation hypothesis by examining effects of the well-conserved tRNA modifying enzyme MiaA in extraintestinal pathogenic Escherichia coli (ExPEC), a major cause of urinary tract and bloodstream infections. MiaA mediates the prenylation of adenosine-37 within tRNAs that decode UNN codons, and we found it to be crucial to the fitness and virulence of ExPEC. MiaA levels shifted in response to stress via a post-transcriptional mechanism, resulting in marked changes in the amounts of fully modified MiaA substrates. Both ablation and forced overproduction of MiaA stimulated translational frameshifting and profoundly altered the ExPEC proteome, with variable effects attributable to UNN content, changes in the catalytic activity of MiaA, or availability of metabolic precursors. Cumulatively, these data indicate that balanced input from MiaA is critical for optimizing cellular responses, with MiaA acting much like a rheostat that can be used to realign global protein expression patterns.

Urinary tract infections in cystic fibrosis patients.

Improved understanding of non-respiratory infections in cystic fibrosis (CF) patients will be vital to sustaining the increased life span of these patients. To date, there has not been a published report of urinary tract infections (UTIs) in CF patients. We performed a retrospective chart review at a major academic medical center during 2010-2020 to determine the features of UTIs in 826 CF patients. We identified 108 UTI episodes during this period. Diabetes, distal intestinal obstruction syndrome (DIOS), and nephrolithiasis were correlated with increased risk of UTIs. UTIs in CF patients were less likely to be caused by Gram-negative rods compared to non-CF patients and more likely to be caused by Enterococcus faecalis. The unique features of UTIs in CF patients highlight the importance of investigating non-respiratory infections to ensure appropriate treatment.

Cytochrome bd promotes Escherichia coli biofilm antibiotic tolerance by regulating accumulation of noxious chemicals.

Nutrient gradients in biofilms cause bacteria to organize into metabolically versatile communities capable of withstanding threats from external agents including bacteriophages, phagocytes, and antibiotics. We previously determined that oxygen availability spatially organizes respiration in uropathogenic Escherichia coli biofilms, and that the high-affinity respiratory quinol oxidase cytochrome bd is necessary for extracellular matrix production and biofilm development. In this study we investigate the physiologic consequences of cytochrome bd deficiency in biofilms and determine that loss of cytochrome bd induces a biofilm-specific increase in expression of general diffusion porins, leading to elevated outer membrane permeability. In addition, loss of cytochrome bd impedes the proton mediated efflux of noxious chemicals by diminishing respiratory flux. As a result, loss of cytochrome bd enhances cellular accumulation of noxious chemicals and increases biofilm susceptibility to antibiotics. These results identify an undescribed link between E. coli biofilm respiration and stress tolerance, while suggesting the possibility of inhibiting cytochrome bd as an antibiofilm therapeutic approach.