RESUMO
We have developed GmPcides from a peptidomimetic dihydrothiazolo ring-fused 2-pyridone scaffold that has antimicrobial activities against a broad spectrum of Gram-positive pathogens. Here, we examine the treatment efficacy of GmPcides using skin and soft tissue infection (SSTI) and biofilm formation models by Streptococcus pyogenes. Screening our compound library for minimal inhibitory (MIC) and minimal bactericidal (MBC) concentrations identified GmPcide PS757 as highly active against S. pyogenes. Treatment of S. pyogenes biofilm with PS757 revealed robust efficacy against all phases of biofilm formation by preventing initial biofilm development, ceasing biofilm maturation and eradicating mature biofilm. In a murine model of S. pyogenes SSTI, subcutaneous delivery of PS757 resulted in reduced levels of tissue damage, decreased bacterial burdens, and accelerated rates of wound healing, which were associated with down-regulation of key virulence factors, including M protein and the SpeB cysteine protease. These data demonstrate that GmPcides show considerable promise for treating S. pyogenes infections.
Assuntos
Biofilmes , Testes de Sensibilidade Microbiana , Piridonas , Infecções dos Tecidos Moles , Infecções Estreptocócicas , Streptococcus pyogenes , Streptococcus pyogenes/efeitos dos fármacos , Animais , Infecções dos Tecidos Moles/tratamento farmacológico , Infecções dos Tecidos Moles/microbiologia , Biofilmes/efeitos dos fármacos , Infecções Estreptocócicas/tratamento farmacológico , Infecções Estreptocócicas/microbiologia , Camundongos , Piridonas/farmacologia , Piridonas/química , Antibacterianos/farmacologia , Antibacterianos/química , Modelos Animais de Doenças , Tiazóis/farmacologia , Tiazóis/química , Dermatopatias Bacterianas/tratamento farmacológico , Dermatopatias Bacterianas/microbiologia , Feminino , Cicatrização/efeitos dos fármacos , HumanosRESUMO
Low-abundance members of microbial communities are difficult to study in their native habitats. This includes Escherichia coli, a minor, but common inhabitant of the gastrointestinal tract and opportunistic pathogen, including of the urinary tract, where it is the primary pathogen. While multi-omic analyses have detailed critical interactions between uropathogenic Escherichia coli (UPEC) and the bladder that mediate UTI outcome, comparatively little is known about UPEC in its pre-infection reservoir, partly due to its low abundance there (<1% relative abundance). To accurately and sensitively explore the genomes and transcriptomes of diverse E. coli in gastrointestinal communities, we developed E. coli PanSelect which uses a set of probes designed to specifically recognize and capture E. coli's broad pangenome from sequencing libraries. We demonstrated the ability of E. coli PanSelect to enrich, by orders of magnitude, sequencing data from diverse E. coli using a mock community and a set of human stool samples collected as part of a cohort study investigating drivers of recurrent urinary tract infections (rUTI). Comparisons of genomes and transcriptomes between E. coli residing in the gastrointestinal tracts of women with and without a history of rUTI suggest that rUTI gut E. coli are responding to increased levels of oxygen and nitrate, suggestive of mucosal inflammation, which may have implications for recurrent disease. E. coli PanSelect is well suited for investigations of native in vivo biology of E. coli in other environments where it is at low relative abundance, and the framework described here has broad applicability to other highly diverse, low abundance organisms.
RESUMO
Catheter-associated urinary tract infections (CAUTIs), a common cause of healthcare-associated infections, are caused by a diverse array of pathogens that are increasingly becoming antibiotic resistant. We analyze the microbial occurrences in catheter and urine samples from 55 human long-term catheterized patients collected over one year. Although most of these patients were prescribed antibiotics over several collection periods, their catheter samples remain colonized by one or more bacterial species. Examination of a total of 366 catheter and urine samples identify 13 positive and 13 negative genus co-occurrences over 12 collection periods, representing associations that occur more or less frequently than expected by chance. We find that for many patients, the microbial species composition between collection periods is similar. In a subset of patients, we find that the most frequently sampled bacteria, Escherichia coli and Enterococcus faecalis, co-localize on catheter samples. Further, co-culture of paired isolates recovered from the same patients reveals that E. coli significantly augments E. faecalis growth in an artificial urine medium, where E. faecalis monoculture grows poorly. These findings suggest novel strategies to collapse polymicrobial CAUTI in long-term catheterized patients by targeting mechanisms that promote positive co-associations.
Assuntos
Infecções Relacionadas a Cateter , Infecções Urinárias , Humanos , Escherichia coli , Infecções Relacionadas a Cateter/microbiologia , Catéteres , Infecções Urinárias/microbiologia , Enterococcus faecalis , BactériasRESUMO
We have developed GmPcides from a peptidomimetic dihydrothiazolo ring-fused 2-pyridone scaffold that have antimicrobial activities against a broad-spectrum of Gram-positive pathogens. Here we examine the treatment efficacy of GmPcides using skin and soft tissue infection (SSTI) and biofilm formation models by Streptococcus pyogenes. Screening our compound library for minimal inhibitory (MIC) and minimal bactericidal (MBC) concentrations identified GmPcide PS757 as highly active against S. pyogenes. Treatment of S. pyogenes biofilm with PS757 revealed robust efficacy against all phases of biofilm formation by preventing initial biofilm development, ceasing biofilm maturation and eradicating mature biofilm. In a murine model of S. pyogenes SSTI, subcutaneous delivery of PS757 resulted in reduced levels of tissue damage, decreased bacterial burdens and accelerated rates of wound-healing, which were associated with down-regulation of key virulence factors, including M protein and the SpeB cysteine protease. These data demonstrate that GmPcides show considerable promise for treating S. pyogenes infections.
RESUMO
Catheter-associated urinary tract infections (CAUTIs) contribute greatly to the burden of healthcare associated infections. Acinetobacter baumannii is a Gram-negative bacterium with high levels of antibiotic resistance that is of increasing concern as a CAUTI pathogen. A. baumannii expresses fibrinogen-binding adhesins (Abp1D and Abp2D) that mediate colonization and biofilm formation on catheters, which become coated with fibrinogen upon insertion. We developed a protein subunit vaccine against Abp1DRBD and Abp2DRBD and showed that vaccination significantly reduced bladder bacterial titers in a mouse model of CAUTI. We then determined that immunity to Abp2DRBD alone was sufficient for protection. Mechanistically, we defined the B cell response to Abp2DRBD vaccination and demonstrated that immunity was transferrable to naïve mice through passive immunization with Abp2DRBD-immune sera. This work represents a novel strategy in the prevention of A. baumannii CAUTI and has an important role to play in the global fight against antimicrobial resistance.
RESUMO
The ability to detect and quantify microbiota over time has a plethora of clinical, basic science, and public health applications. One of the primary means of tracking microbiota is through sequencing technologies. When the microorganism of interest is well characterized or known a priori, targeted sequencing is often used. In many applications, however, untargeted bulk (shotgun) sequencing is more appropriate; for instance, the tracking of infection transmission events and nucleotide variants across multiple genomic loci, or studying the role of multiple genes in a particular phenotype. Given these applications, and the observation that pathogens (e.g. Clostridioides difficile, Escherichia coli, Salmonella enterica) and other taxa of interest can reside at low relative abundance in the gastrointestinal tract, there is a critical need for algorithms that accurately track low-abundance taxa with strain level resolution. Here we present a sequence quality- and time-aware model, ChronoStrain, that introduces uncertainty quantification to gauge low-abundance species and significantly outperforms the current state-of-the-art on both real and synthetic data. ChronoStrain leverages sequences' quality scores and the samples' temporal information to produce a probability distribution over abundance trajectories for each strain tracked in the model. We demonstrate Chronostrain's improved performance in capturing post-antibiotic E. coli strain blooms among women with recurrent urinary tract infections (UTIs) from the UTI Microbiome (UMB) Project. Other strain tracking models on the same data either show inconsistent temporal colonization or can only track consistently using very coarse groupings. In contrast, our probabilistic outputs can reveal the relationship between low-confidence strains present in the sample that cannot be reliably assigned a single reference label (either due to poor coverage or novelty) while simultaneously calling high-confidence strains that can be unambiguously assigned a label. We also include and analyze newly sequenced cultured samples from the UMB Project.
RESUMO
Multidrug-resistant Acinetobacter baumannii infections are an urgent clinical problem and can cause difficult-to-treat nosocomial infections. During such infections, like catheter-associated urinary tract infections (CAUTI), A. baumannii rely on adhesive, extracellular fibers, called chaperone-usher pathway (CUP) pili for critical binding interactions. The A. baumannii uropathogenic strain, UPAB1, and the pan-European subclone II isolate, ACICU, use the CUP pili Abp1 and Abp2 (previously termed Cup and Prp, respectively) in tandem to establish CAUTIs, specifically to facilitate bacterial adherence and biofilm formation on the implanted catheter. Abp1 and Abp2 pili are tipped with two domain tip adhesins, Abp1D and Abp2D, respectively. We discovered that both adhesins bind fibrinogen, a critical host wound response protein that is released into the bladder upon catheterization and is subsequently deposited on the catheter. The crystal structures of the Abp1D and Abp2D receptor-binding domains were determined and revealed that they both contain a large, distally oriented pocket, which mediates binding to fibrinogen and other glycoproteins. Genetic, biochemical, and biophysical studies revealed that interactions with host proteins are governed by several critical residues in and along the edge of the binding pocket, one of which regulates the structural stability of an anterior loop motif. K34, located outside of the pocket but interacting with the anterior loop, also regulates the binding affinity of the protein. This study illuminates the mechanistic basis of the critical fibrinogen-coated catheter colonization step in A. baumannii CAUTI pathogenesis.
Assuntos
Acinetobacter baumannii , Infecções Urinárias , Humanos , Adesinas Bacterianas/genética , Adesinas Bacterianas/metabolismo , Infecções Urinárias/microbiologia , Catéteres , Acinetobacter baumannii/genética , Fibrinogênio/metabolismoRESUMO
The alarming rise of multidrug-resistant Gram-positive bacteria has precipitated a healthcare crisis, necessitating the development of new antimicrobial therapies. Here we describe a new class of antibiotics based on a ring-fused 2-pyridone backbone, which are active against vancomycin-resistant enterococci (VRE), a serious threat as classified by the Centers for Disease Control and Prevention, and other multidrug-resistant Gram-positive bacteria. Ring-fused 2-pyridone antibiotics have bacteriostatic activity against actively dividing exponential phase enterococcal cells and bactericidal activity against nondividing stationary phase enterococcal cells. The molecular mechanism of drug-induced killing of stationary phase cells mimics aspects of fratricide observed in enterococcal biofilms, where both are mediated by the Atn autolysin and the GelE protease. In addition, combinations of sublethal concentrations of ring-fused 2-pyridones and standard-of-care antibiotics, such as vancomycin, were found to synergize to kill clinical strains of VRE. Furthermore, a broad range of antibiotic resistant Gram-positive pathogens, including those responsible for the increasing incidence of antibiotic resistant healthcare-associated infections, are susceptible to this new class of 2-pyridone antibiotics. Given the broad antibacterial activities of ring-fused 2-pyridone compounds against Gram-positive (GmP) bacteria we term these compounds GmPcides, which hold promise in combating the rising tide of antibiotic resistant Gram-positive pathogens.
Assuntos
Bactérias Gram-Positivas , Piridonas , Enterococos Resistentes à Vancomicina , Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla , Bactérias Gram-Positivas/efeitos dos fármacos , Testes de Sensibilidade Microbiana , N-Acetil-Muramil-L-Alanina Amidase/farmacologia , Piridonas/farmacologia , Vancomicina/farmacologia , Enterococos Resistentes à Vancomicina/efeitos dos fármacosRESUMO
Recurrent urinary tract infections (rUTIs) are a major health burden worldwide, with history of infection being a significant risk factor. While the gut is a known reservoir for uropathogenic bacteria, the role of the microbiota in rUTI remains unclear. We conducted a year-long study of women with (n = 15) and without (n = 16) history of rUTI, from whom we collected urine, blood and monthly faecal samples for metagenomic and transcriptomic interrogation. During the study 24 UTIs were reported, with additional samples collected during and after infection. The gut microbiome of individuals with a history of rUTI was significantly depleted in microbial richness and butyrate-producing bacteria compared with controls, reminiscent of other inflammatory conditions. However, Escherichia coli gut and bladder populations were comparable between cohorts in both relative abundance and phylogroup. Transcriptional analysis of peripheral blood mononuclear cells revealed expression profiles indicative of differential systemic immunity between cohorts. Altogether, these results suggest that rUTI susceptibility is in part mediated through the gut-bladder axis, comprising gut dysbiosis and differential immune response to bacterial bladder colonization, manifesting in symptoms.
Assuntos
Infecções por Escherichia coli , Microbioma Gastrointestinal , Infecções Urinárias , Disbiose , Escherichia coli , Infecções por Escherichia coli/microbiologia , Feminino , Humanos , Leucócitos Mononucleares , Masculino , Infecções Urinárias/microbiologiaAssuntos
Microbioma Gastrointestinal , Microbiota , Infecções Urinárias , Feminino , Humanos , MasculinoRESUMO
BACKGROUND: Urinary tract infections (UTIs) affect 15 million women each year in the United States, with > 20% experiencing frequent recurrent UTIs. A recent placebo-controlled clinical trial found a 39% reduction in UTI symptoms among recurrent UTI sufferers who consumed a daily cranberry beverage for 24 weeks. Using metagenomic sequencing of stool from a subset of these trial participants, we assessed the impact of cranberry consumption on the gut microbiota, a reservoir for UTI-causing pathogens such as Escherichia coli, which causes > 80% of UTIs. RESULTS: The overall taxonomic composition, community diversity, carriage of functional pathways and gene families, and relative abundances of the vast majority of observed bacterial taxa, including E. coli, were not changed significantly by cranberry consumption. However, one unnamed Flavonifractor species (OTU41), which represented ≤1% of the overall metagenome, was significantly less abundant in cranberry consumers compared to placebo at trial completion. Given Flavonifractor's association with negative human health effects, we sought to determine OTU41 characteristic genes that may explain its differential abundance and/or relationship to key host functions. Using comparative genomic and metagenomic techniques, we identified genes in OTU41 related to transport and metabolism of various compounds, including tryptophan and cobalamin, which have been shown to play roles in host-microbe interactions. CONCLUSION: While our results indicated that cranberry juice consumption had little impact on global measures of the microbiome, we found one unnamed Flavonifractor species differed significantly between study arms. This suggests further studies are needed to assess the role of cranberry consumption and Flavonifractor in health and wellbeing in the context of recurrent UTI. TRIAL REGISTRATION: Clinical trial registration number: ClinicalTrials.gov NCT01776021 .
Assuntos
Bactérias/efeitos dos fármacos , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/genética , Extratos Vegetais/administração & dosagem , Vaccinium macrocarpon/química , Adulto , Bactérias/classificação , Bactérias/genética , Bebidas , Método Duplo-Cego , Fezes/microbiologia , Feminino , Microbioma Gastrointestinal/fisiologia , Humanos , Metagenoma , Metagenômica/métodos , Pessoa de Meia-Idade , Reinfecção/microbiologia , Reinfecção/prevenção & controle , Infecções Urinárias/microbiologia , Infecções Urinárias/prevenção & controleRESUMO
Urinary tract infections (UTIs) are predominantly caused by uropathogenic Escherichia coli (UPEC). UPEC pathogenesis requires passage through a severe population bottleneck involving intracellular bacterial communities (IBCs) that are clonal expansions of a single invading UPEC bacterium in a urothelial superficial facet cell. IBCs occur only during acute pathogenesis. The bacteria in IBCs form the founder population that develops into persistent extracellular infections. Only a small fraction of UPEC organisms proceed through the IBC cycle, regardless of the inoculum size. This dramatic reduction in population size precludes the utility of genomic mutagenesis technologies for identifying genes important for persistence. To circumvent this bottleneck, we previously identified 29 positively selected genes (PSGs) within UPEC and hypothesized that they contribute to virulence. Here, we show that 8 of these 29 PSGs are required for fitness during persistent bacteriuria. Conversely, 7/8 of these PSG mutants showed essentially no phenotype in acute UTI. Deletion of the PSG argI leads to arginine auxotrophy. Relative to the other arg genes, argI in the B2 clade (which comprises most UPEC strains) of E. coli has diverged from argI in other E. coli clades. Replacement of argI in a UPEC strain with a non-UPEC argI allele complemented the arginine auxotrophy but not the persistent bacteriuria defect, showing that the UPEC argI allele contributes to persistent infection. These results highlight the complex roles of metabolic pathways during infection and demonstrate that evolutionary approaches can identify infection-specific gene functions downstream of population bottlenecks, shedding light on virulence and the genetic evolution of pathogenesis.IMPORTANCE Uropathogenic Escherichia coli (UPEC) is the most common cause of human urinary tract infection (UTI). Population bottlenecks during early stages of UTI make high-throughput screens impractical for understanding clinically important later stages of UTI, such as persistence and recurrence. As UPEC is hypothesized to be adapted to these later pathogenic stages, we previously identified 29 genes evolving under positive selection in UPEC. Here, we found that 8 of these genes, including argI (which is involved in arginine biosynthesis), are important for persistence in a mouse model of UTI. Deletion of argI and other arginine synthesis genes resulted in (i) arginine auxotrophy and (ii) defects in persistent UTI. Replacement of a B2 clade argI with a non-B2 clade argI complemented arginine auxotrophy, but the resulting strain remained attenuated in its ability to cause persistent bacteriuria. Thus, argI may have a second function during UTI that is not related to simple arginine synthesis. This study demonstrates how variation in metabolic genes can impact virulence and provides insight into the mechanisms and evolution of bacterial virulence.
Assuntos
Adaptação Fisiológica , Arginina/biossíntese , Evolução Molecular , Filogenia , Sistema Urinário/microbiologia , Escherichia coli Uropatogênica/metabolismo , Animais , Infecções por Escherichia coli/microbiologia , Proteínas de Escherichia coli/metabolismo , Feminino , Aptidão Genética , Redes e Vias Metabólicas/genética , Camundongos , Camundongos Endogâmicos C3H , Escherichia coli Uropatogênica/genética , Virulência/genéticaRESUMO
Orthopaedic surgery has a rich history of publication of the science that supports the practice of our specialty, which dates from 1887. Orthopaedic publishing has evolved since that time, expanding from print to online access, with increasing variation in publication models, including open-access journals and article repositories, and methods of information delivery that include video, data archives, and commentary. This symposium provides an overview of the changes and challenges in the publication of orthopaedic science.
Assuntos
Ortopedia/tendências , Publicações Periódicas como Assunto/tendências , Ética em Pesquisa , Humanos , Publicação de Acesso Aberto/tendências , Revisão por Pares , Editoração/ética , Editoração/tendências , Projetos de Pesquisa , Má Conduta Científica/ética , Má Conduta Científica/tendênciasRESUMO
Chaperone-usher pathway pili are extracellular proteinaceous fibres ubiquitously found on Gram-negative bacteria, and mediate host-pathogen interactions and biofilm formation critical in pathogenesis in numerous human diseases1. During pilus assembly, an outer membrane macromolecular machine called the usher catalyses pilus biogenesis from the individual subunits that are delivered as chaperone-subunit complexes in the periplasm. The usher orchestrates pilus assembly using all five functional domains: a 24-stranded transmembrane ß-barrel translocation domain, a ß-sandwich plug domain, an amino-terminal periplasmic domain and two carboxy-terminal periplasmic domains (CTD1 and CTD2)2-6. Despite extensive structural and functional characterization, the mechanism by which the usher is activated to initiate pilus biogenesis is unknown. Here, we present the crystal structure of the full-length PapC usher from Escherichia coli in complex with its cognate PapDG chaperone-subunit complex in a pre-activation state, elucidating molecular details of how the usher is specifically engaged by allosteric interactions with its substrate preceding activation and how the usher facilitates the transfer of subunits from the amino-terminal periplasmic domain to the CTDs during pilus assembly. This work elucidates the intricate workings of a molecular machine that catalyses chaperone-usher pathway pilus assembly and opens the door for the development of potent inhibitors to block pilus biogenesis.
Assuntos
Proteínas de Escherichia coli/química , Escherichia coli/metabolismo , Fímbrias Bacterianas/química , Porinas/química , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Fímbrias Bacterianas/genética , Modelos Moleculares , Chaperonas Moleculares/química , Chaperonas Moleculares/genética , Proteínas Periplásmicas/genética , Ligação Proteica , Conformação Proteica , Domínios ProteicosRESUMO
Uropathogenic E. coli (UPEC), which cause urinary tract infections (UTI), utilize type 1 pili, a chaperone usher pathway (CUP) pilus, to cause UTI and colonize the gut. The pilus rod, comprised of repeating FimA subunits, provides a structural scaffold for displaying the tip adhesin, FimH. We solved the 4.2 Å resolution structure of the type 1 pilus rod using cryo-electron microscopy. Residues forming the interactive surfaces that determine the mechanical properties of the rod were maintained by selection based on a global alignment of fimA sequences. We identified mutations that did not alter pilus production in vitro but reduced the force required to unwind the rod. UPEC expressing these mutant pili were significantly attenuated in bladder infection and intestinal colonization in mice. This study elucidates an unappreciated functional role for the molecular spring-like property of type 1 pilus rods in host-pathogen interactions and carries important implications for other pilus-mediated diseases.
Assuntos
Adesinas Bacterianas/metabolismo , Adesinas Bacterianas/ultraestrutura , Aderência Bacteriana , Infecções por Escherichia coli/microbiologia , Fímbrias Bacterianas/metabolismo , Interações Hospedeiro-Patógeno , Escherichia coli Uropatogênica/ultraestrutura , Animais , Microscopia Crioeletrônica , Modelos Animais de Doenças , Proteínas de Fímbrias/genética , Proteínas de Fímbrias/metabolismo , Fímbrias Bacterianas/ultraestrutura , Camundongos , Infecções Urinárias/microbiologia , Escherichia coli Uropatogênica/fisiologiaRESUMO
Antibiotics have become the standard of care for bacterial infections. However, rising rates of antibiotic-resistant infections are outpacing the development of new antimicrobials. Broad-spectrum antibiotics also harm beneficial microbial communities inhabiting humans. To combat antibiotic resistance and protect these communities, new precision antimicrobials must be engineered to target specific pathogens.
Assuntos
Anti-Infecciosos/uso terapêutico , Microbioma Gastrointestinal/efeitos dos fármacos , Medicina de Precisão/métodos , Animais , Anti-Infecciosos/administração & dosagem , Anti-Infecciosos/efeitos adversos , Infecções Bacterianas/tratamento farmacológico , HumanosRESUMO
Urinary tract infections (UTIs) caused by uropathogenic Escherichia coli (UPEC) affect 150 million people annually. Despite effective antibiotic therapy, 30-50% of patients experience recurrent UTIs. In addition, the growing prevalence of UPEC that are resistant to last-line antibiotic treatments, and more recently to carbapenems and colistin, make UTI a prime example of the antibiotic-resistance crisis and emphasize the need for new approaches to treat and prevent bacterial infections. UPEC strains establish reservoirs in the gut from which they are shed in the faeces, and can colonize the periurethral area or vagina and subsequently ascend through the urethra to the urinary tract, where they cause UTIs. UPEC isolates encode up to 16 distinct chaperone-usher pathway pili, and each pilus type may enable colonization of a habitat in the host or environment. For example, the type 1 pilus adhesin FimH binds mannose on the bladder surface, and mediates colonization of the bladder. However, little is known about the mechanisms underlying UPEC persistence in the gut. Here, using a mouse model, we show that F17-like and type 1 pili promote intestinal colonization and show distinct binding to epithelial cells distributed along colonic crypts. Phylogenomic and structural analyses reveal that F17-like pili are closely related to pilus types carried by intestinal pathogens, but are restricted to extra-intestinal pathogenic E. coli. Moreover, we show that targeting FimH with M4284, a high-affinity inhibitory mannoside, reduces intestinal colonization of genetically diverse UPEC isolates, while simultaneously treating UTI, without notably disrupting the structural configuration of the gut microbiota. By selectively depleting intestinal UPEC reservoirs, mannosides could markedly reduce the rate of UTIs and recurrent UTIs.
Assuntos
Proteínas de Fímbrias/antagonistas & inibidores , Intestinos/efeitos dos fármacos , Intestinos/microbiologia , Manosídeos/farmacologia , Ácidos Ftálicos/farmacologia , Infecções Urinárias/prevenção & controle , Escherichia coli Uropatogênica/efeitos dos fármacos , Escherichia coli Uropatogênica/isolamento & purificação , Adesinas de Escherichia coli/metabolismo , Sequência de Aminoácidos , Animais , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/microbiologia , Fezes/microbiologia , Feminino , Proteínas de Fímbrias/metabolismo , Fímbrias Bacterianas/classificação , Fímbrias Bacterianas/efeitos dos fármacos , Fímbrias Bacterianas/genética , Fímbrias Bacterianas/metabolismo , Humanos , Intestinos/citologia , Manosídeos/uso terapêutico , Camundongos , Modelos Moleculares , Ácidos Ftálicos/uso terapêutico , Bexiga Urinária/efeitos dos fármacos , Bexiga Urinária/microbiologia , Infecções Urinárias/tratamento farmacológico , Infecções Urinárias/microbiologia , Escherichia coli Uropatogênica/classificação , Escherichia coli Uropatogênica/genéticaRESUMO
Positive selection in the two-domain type 1 pilus adhesin FimH enhances Escherichia coli fitness in urinary tract infection (UTI). We report a comprehensive atomic-level view of FimH in two-state conformational ensembles in solution, composed of one low-affinity tense (T) and multiple high-affinity relaxed (R) conformations. Positively selected residues allosterically modulate the equilibrium between these two conformational states, each of which engages mannose through distinct binding orientations. A FimH variant that only adopts the R state is severely attenuated early in a mouse model of uncomplicated UTI but is proficient at colonizing catheterized bladders in vivo or bladder transitional-like epithelial cells in vitro. Thus, the bladder habitat has barrier(s) to R state-mediated colonization possibly conferred by the terminally differentiated bladder epithelium and/or decoy receptors in urine. Together, our studies reveal the conformational landscape in solution, binding mechanisms, and adhesive strength of an allosteric two-domain adhesin that evolved "moderate" affinity to optimize persistence in the bladder during UTI.
Assuntos
Adesinas de Escherichia coli , Infecções por Escherichia coli , Escherichia coli , Proteínas de Fímbrias , Interações Hospedeiro-Parasita/fisiologia , Bexiga Urinária , Infecções Urinárias , Adesinas de Escherichia coli/química , Adesinas de Escherichia coli/genética , Adesinas de Escherichia coli/metabolismo , Animais , Escherichia coli/química , Escherichia coli/patogenicidade , Escherichia coli/fisiologia , Infecções por Escherichia coli/genética , Infecções por Escherichia coli/metabolismo , Infecções por Escherichia coli/patologia , Feminino , Proteínas de Fímbrias/química , Proteínas de Fímbrias/genética , Proteínas de Fímbrias/metabolismo , Camundongos , Domínios Proteicos , Bexiga Urinária/metabolismo , Bexiga Urinária/microbiologia , Bexiga Urinária/fisiologia , Infecções Urinárias/genética , Infecções Urinárias/metabolismo , Infecções Urinárias/microbiologia , Infecções Urinárias/patologiaRESUMO
Urinary tract infections (UTIs) are caused by uropathogenic Escherichia coli (UPEC) strains. In contrast to many enteric E. coli pathogroups, no genetic signature has been identified for UPEC strains. We conducted a high-resolution comparative genomic study using E. coli isolates collected from the urine of women suffering from frequent recurrent UTIs. These isolates were genetically diverse and varied in their urovirulence, that is, their ability to infect the bladder in a mouse model of cystitis. We found no set of genes, including previously defined putative urovirulence factors (PUFs), that were predictive of urovirulence. In addition, in some patients, the E. coli strain causing a recurrent UTI had fewer PUFs than the supplanted strain. In competitive experimental infections in mice, the supplanting strain was more efficient at colonizing the mouse bladder than the supplanted strain. Despite the lack of a clear genomic signature for urovirulence, comparative transcriptomic and phenotypic analyses revealed that the expression of key conserved functions during culture, such as motility and metabolism, could be used to predict subsequent colonization of the mouse bladder. Together, our findings suggest that UTI risk and outcome may be determined by complex interactions between host susceptibility and the urovirulence potential of diverse bacterial strains.