AnIn VitroDeletion inribEEncoding Lumazine Synthase Contributes to Nitrofurantoin Resistance in Escherichia coli

ABSTRACTNitrofurantoin has been used for decades for the treatment of urinary tract infections (UTIs), but clinically significant resistance inEscherichia coliis uncommon. Nitrofurantoin concentrations in the gastrointestinal tract tend to be low, which might facilitate selection of nitrofurantoin-resistant (NIT-R) strains in the gut flora. We subjected two nitrofurantoin-susceptible intestinalE. colistrains (ST540-p and ST2747-p) to increasing nitrofurantoin concentrations under aerobic and anaerobic conditions. Whole-genome sequencing was performed for both susceptible isolates and selected mutants that exhibited the highest nitrofurantoin resistance levels aerobically (ST540-a and ST2747-a) and anaerobically (ST540-an and ST2747-an). ST540-a/ST540-an and ST2747-a (aerobic MICs of >64 μg/ml) harbored mutations in the known nitrofurantoin resistance determinantsnfsAand/ornfsB, which encode oxygen-insensitive nitroreductases. ST2747-an showed reduced nitrofurantoin susceptibility (aerobic MIC of 32 μg/ml) and exhibited remarkable growth deficits but did not harbornfsA/nfsBmutations. We identified a 12-nucleotide deletion inribE, encoding lumazine synthase, an essential enzyme involved in the biosynthesis of flavin mononucleotide (FMN), which is an important cofactor for NfsA and NfsB. Complementing ST2747-an with a functional wild-type lumazine synthase restored nitrofurantoin susceptibility. Six NIT-RE. coliisolates (NRCI-1 to NRCI-6) from stools of UTI patients treated with nitrofurantoin, cefuroxime, or a fluoroquinolone harbored mutations innfsAand/ornfsBbut notribE. Sequencing of theribEgene in six intestinal and three urinaryE. colistrains showing reduced nitrofurantoin susceptibility (MICs of 16 to 48 μg/ml) also did not identify any relevant mutations. NRCI-1, NRCI-2, and NRCI-5 exhibited up to 4-fold higher anaerobic MICs, compared to the mutants generatedin vitro, presumably because of additional mutations in oxygen-sensitive nitroreductases.

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Escherichia coli CFT073 Fitness Factors during Urinary Tract Infection: Identification Using an Ordered Transposon Library

Applied and Environmental Microbiology ◽

10.1128/aem.00691-20 ◽

2020 ◽

Vol 86 (13) ◽

Cited By ~ 1

Author(s):

Allyson E. Shea ◽

Juan Marzoa ◽

Stephanie D. Himpsl ◽

Sara N. Smith ◽

Lili Zhao ◽

...

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Urinary Tract Infections ◽

Human Urine ◽

Transposon Mutagenesis ◽

Content Type ◽

E Coli ◽

Tract Infections

ABSTRACT Urinary tract infections (UTI), the second most diagnosed infectious disease worldwide, are caused primarily by uropathogenic Escherichia coli (UPEC), placing a significant financial burden on the health care system. High-throughput transposon mutagenesis combined with genome-targeted sequencing is a powerful technique to interrogate genomes for fitness genes. Genome-wide analysis of E. coli requires random libraries of at least 50,000 mutants to achieve 99.99% saturation; however, the traditional murine model of ascending UTI does not permit testing of large mutant pools due to a bottleneck during infection. To address this, an E. coli CFT073 transposon mutant ordered library of 9,216 mutants was created and insertion sites were identified. A single transposon mutant was selected for each gene to assemble a condensed library consisting of 2,913 unique nonessential mutants. Using a modified UTI model in BALB/c mice, we identified 36 genes important for colonizing the bladder, including purB, yihE, and carB. Screening of the condensed library in vitro identified yigP and ubiG to be essential for growth in human urine. Additionally, we developed a novel quantitative PCR (qPCR) technique to identify genes with fitness defects within defined subgroups of related genes (e.g., genes encoding fimbriae, toxins, etc.) following UTI. The number of mutants within these subgroups circumvents bottleneck restriction and facilitates validation of multiple mutants to generate individual competitive indices. Collectively, this study investigates the bottleneck effects during UTI, provides two techniques for evading those effects that can be applied to other disease models, and contributes a genetic tool in prototype strain CFT073 to the field. IMPORTANCE Uropathogenic Escherichia coli strains cause most uncomplicated urinary tract infections (UTI), one of the most common infectious diseases worldwide. Random transposon mutagenesis techniques have been utilized to identify essential bacterial genes during infection; however, this has been met with limitations when applied to the murine UTI model. Conventional high-throughput transposon mutagenesis screens are not feasible because of inoculum size restrictions due to a bottleneck during infection. Our study utilizes a condensed ordered transposon library, limiting the number of mutants while maintaining the largest possible genome coverage. Screening of this library in vivo, and in human urine in vitro, identified numerous candidate fitness factors. Additionally, we have developed a novel technique using qPCR to quantify bacterial outputs following infection with small subgroups of transposon mutants. Molecular approaches developed in this study will serve as useful tools to probe in vivo models that are restricted by anatomical, physiological, or genetic bottleneck limitations.

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Cefoxitin as an Alternative to Carbapenems in a Murine Model of Urinary Tract Infection Due to Escherichia coli Harboring CTX-M-15-Type Extended-Spectrum β-Lactamase

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.06233-11 ◽

2012 ◽

Vol 56 (3) ◽

pp. 1376-1381 ◽

Cited By ~ 32

Author(s):

Raphaël Lepeule ◽

Etienne Ruppé ◽

Patrick Le ◽

Laurent Massias ◽

Françoise Chau ◽

...

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Murine Model ◽

Content Type ◽

E Coli ◽

Extended Spectrum ◽

Drug Concentrations ◽

Tract Infections

ABSTRACTWe investigated the efficiency of the cephamycin cefoxitin as an alternative to carbapenems for the treatment of urinary tract infections (UTIs) due toEscherichia coliproducing CTX-M-type extended-spectrum β-lactamases. The susceptible, UTI-inducingE. coliCFT073-RR strain and its transconjugant CFT073-RR Tc (pblaCTX-M-15), harboring ablaCTX-M-15carrying-plasmid, were used for all experiments. MICs of cefoxitin (FOX), ceftriaxone (CRO), imipenem (IMP), and ertapenem (ETP) for CFT073-RR and CFT073-RR Tc (pblaCTX-M-15) were 4 and 4, 0.125 and 512, 0.5 and 0.5, and 0.016 and 0.032 μg/ml, respectively. Bactericidal activity was similarly achievedin vitroagainst the two strains after 3 h of exposure to concentrations of FOX, IMI, and ETP that were 2 times the MIC, whereas CRO was not bactericidal against CFT073-RR Tc (pblaCTX-M-15). The frequencies of spontaneous mutants of the 2 strains were not higher for FOX than for IMP or ETP. In the murine model of UTIs, mice infected for 5 days were treated over 24 h. Therapeutic regimens in mice (200 mg/kg of body weight every 3 h or 4 h for FOX, 70 mg/kg every 6 h for CRO, 100 mg/kg every 2 h for IMP, and 100 mg/kg every 4 h for ETP) were chosen in order to reproduce the percentage of time that free-drug concentrations above the MIC are obtained in humans with standard regimens. All antibiotic regimens produced a significant reduction in bacterial counts (greater than 2 log10CFU) in kidneys and bladders for both strains (P< 0.001) without selecting resistant mutantsin vivo, but the reduction obtained with CRO against CFT073-RR Tc (pblaCTX-M-15) in kidneys was significantly lower than that obtained with FOX. In conclusion, FOX appears to be an effective therapeutic alternative to carbapenems for the treatment of UTIs due to CTX-M-producingE. coli.

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A Rhodobacter sphaeroides Protein Mechanistically Similar to Escherichia coli DksA Regulates Photosynthetic Growth

mBio ◽

10.1128/mbio.01105-14 ◽

2014 ◽

Vol 5 (3) ◽

Cited By ~ 7

Author(s):

Christopher W. Lennon ◽

Kimberly C. Lemmer ◽

Jessica L. Irons ◽

Max I. Sellman ◽

Timothy J. Donohue ◽

...

Keyword(s):

Escherichia Coli ◽

Structure Function ◽

Rhodobacter Sphaeroides ◽

Fatty Acid Content ◽

Regulatory Protein ◽

Growth Conditions ◽

Globular Domain ◽

Content Type ◽

E Coli

ABSTRACTDksA is a global regulatory protein that, together with the alarmone ppGpp, is required for the “stringent response” to nutrient starvation in the gammaproteobacteriumEscherichia coliand for more moderate shifts between growth conditions. DksA modulates the expression of hundreds of genes, directly or indirectly. Mutants lacking a DksA homolog exhibit pleiotropic phenotypes in other gammaproteobacteria as well. Here we analyzed the DksA homolog RSP2654 in the more distantly relatedRhodobacter sphaeroides, an alphaproteobacterium. RSP2654 is 42% identical and similar in length toE. coliDksA but lacks the Zn finger motif of theE. coliDksA globular domain. Deletion of the RSP2654 gene results in defects in photosynthetic growth, impaired utilization of amino acids, and an increase in fatty acid content. RSP2654 complements the growth and regulatory defects of anE. colistrain lacking thedksAgene and modulates transcriptionin vitrowithE. coliRNA polymerase (RNAP) similarly toE. coliDksA. RSP2654 reduces RNAP-promoter complex stabilityin vitrowith RNAPs fromE. coliorR. sphaeroides, alone and synergistically with ppGpp, suggesting that even though it has limited sequence identity toE. coliDksA (DksAEc), it functions in a mechanistically similar manner. We therefore designate the RSP2654 protein DksARsp. Our work suggests that DksARsphas distinct and important physiological roles in alphaproteobacteria and will be useful for understanding structure-function relationships in DksA and the mechanism of synergy between DksA and ppGpp.IMPORTANCEThe role of DksA has been analyzed primarily in the gammaproteobacteria, in which it is best understood for its role in control of the synthesis of the translation apparatus and amino acid biosynthesis. Our work suggests that DksA plays distinct and important physiological roles in alphaproteobacteria, including the control of photosynthesis inRhodobacter sphaeroides. The study of DksARsp, should be useful for understanding structure-function relationships in the protein, including those that play a role in the little-understood synergy between DksA and ppGpp.

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Occurrence of Antimicrobial-Resistant Escherichia coli and Salmonella enterica in the Beef Cattle Production and Processing Continuum

Applied and Environmental Microbiology ◽

10.1128/aem.03079-14 ◽

2014 ◽

Vol 81 (2) ◽

pp. 713-725 ◽

Cited By ~ 44

Author(s):

John W. Schmidt ◽

Getahun E. Agga ◽

Joseph M. Bosilevac ◽

Dayna M. Brichta-Harhay ◽

Steven D. Shackelford ◽

...

Keyword(s):

Escherichia Coli ◽

Salmonella Enterica ◽

Generation Cephalosporin ◽

Resistant Bacteria ◽

Processing Plant ◽

Cattle Production ◽

Content Type ◽

E Coli ◽

Beef Processing ◽

Tract Infections

ABSTRACTSpecific concerns have been raised that third-generation cephalosporin-resistant (3GCr)Escherichia coli, trimethoprim-sulfamethoxazole-resistant (COTr)E. coli, 3GCrSalmonella enterica, and nalidixic acid-resistant (NALr)S. entericamay be present in cattle production environments, persist through beef processing, and contaminate final products. The prevalences and concentrations of these organisms were determined in feces and hides (at feedlot and processing plant), pre-evisceration carcasses, and final carcasses from three lots of fed cattle (n= 184). The prevalences and concentrations were further determined for strip loins from 103 of the carcasses. 3GCrSalmonellawas detected on 7.6% of hides during processing and was not detected on the final carcasses or strip loins. NALrS. entericawas detected on only one hide. 3GCrE. coliand COTrE. coliwere detected on 100.0% of hides during processing. Concentrations of 3GCrE. coliand COTrE. colion hides were correlated with pre-evisceration carcass contamination. 3GCrE. coliand COTrE. coliwere each detected on only 0.5% of final carcasses and were not detected on strip loins. Five hundred and 42 isolates were screened for extraintestinal pathogenicE. coli(ExPEC) virulence-associated markers. Only two COTrE. coliisolates from hides were ExPEC, indicating that fed cattle products are not a significant source of ExPEC causing human urinary tract infections. The very low prevalences of these organisms on final carcasses and their absence on strip loins demonstrate that current sanitary dressing procedures and processing interventions are effective against antimicrobial-resistant bacteria.

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Single Clinical Isolates from Acute Uncomplicated Urinary Tract Infections Are Representative of Dominant In Situ Populations

mBio ◽

10.1128/mbio.01064-13 ◽

2014 ◽

Vol 5 (2) ◽

Cited By ~ 18

Author(s):

Dana Willner ◽

Serene Low ◽

Jason A. Steen ◽

Narelle George ◽

Graeme R. Nimmo ◽

...

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Urinary Tract Infections ◽

Clinical Isolates ◽

Content Type ◽

E Coli ◽

Strain Diversity ◽

Amplicon Pyrosequencing ◽

Culture Independent ◽

Tract Infections

ABSTRACTUrinary tract infections (UTIs) are one of the most commonly acquired bacterial infections in humans, and uropathogenicEscherichia colistrains are responsible for over 80% of all cases. The standard method for identification of uropathogens in clinical laboratories is cultivation, primarily using solid growth media under aerobic conditions, coupled with morphological and biochemical tests of typically a single isolate colony. However, these methods detect only culturable microorganisms, and characterization is phenotypic in nature. Here, we explored the genotypic identity of communities in acute uncomplicated UTIs from 50 individuals by using culture-independent amplicon pyrosequencing and whole-genome and metagenomic shotgun sequencing. Genus-level characterization of the UTI communities was achieved using the 16S rRNA gene (V8 region). Overall UTI community richness was very low in comparison to other human microbiomes. We strain-typedEscherichia-dominated UTIs using amplicon pyrosequencing of the fimbrial adhesin gene,fimH. There were nine highly abundantfimHtypes, and each UTI sample was dominated by a single type. Molecular analysis of the corresponding clinical isolates revealed that in the majority of cases the isolate was representative of the dominant taxon in the community at both the genus and the strain level. Shotgun sequencing was performed on a subset of eightE. coliurine UTI and isolate pairs. The majority of UTI microbial metagenomic sequences mapped to isolate genomes, confirming the results obtained using phylogenetic markers. We conclude that for the majority of acute uncomplicatedE. coli-mediated UTIs, single cultured isolates are diagnostic of the infection.IMPORTANCEIn clinical practice, the diagnosis and treatment of acute uncomplicated urinary tract infection (UTI) are based on analysis of a single bacterial isolate cultured from urine, and it is assumed that this isolate represents the dominant UTI pathogen. However, these methods detect only culturable bacteria, and the existence of multiple pathogens as well as strain diversity within a single infection is not examined. Here, we explored bacteria present in acute uncomplicated UTIs using culture-independent sequence-based methods.Escherichia coliwas the most common organism identified, and analysis ofE. colidominant UTI samples and their paired clinical isolates revealed that in the majority of infections the cultured isolate was representative of the dominant taxon at both the genus and the strain level. Our data demonstrate that in most cases single cultured isolates are diagnostic of UTI and are consistent with the notion of bottlenecks that limit strain diversity during UTI pathogenesis.

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Amdinocillin (Mecillinam) Resistance Mutations in Clinical Isolates and Laboratory-Selected Mutants of Escherichia coli

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.04819-14 ◽

2015 ◽

Vol 59 (3) ◽

pp. 1718-1727 ◽

Cited By ~ 58

Author(s):

Elisabeth Thulin ◽

Martin Sundqvist ◽

Dan I. Andersson

Keyword(s):

Escherichia Coli ◽

Clinical Isolates ◽

Cysteine Biosynthesis ◽

Resistance Mutations ◽

Content Type ◽

E Coli ◽

Major Mechanism ◽

Laboratory Selection ◽

Lactam Antibiotic ◽

Tract Infections

ABSTRACTAmdinocillin (mecillinam) is a β-lactam antibiotic that is used mainly for the treatment of uncomplicated urinary tract infections. The objectives of this study were to identify mutations that confer amdinocillin resistance on laboratory-isolated mutants and clinical isolates ofEscherichia coliand to determine why amdinocillin resistance remains rare clinically even though resistance is easily selected in the laboratory. Under laboratory selection, frequencies of mutation to amdinocillin resistance varied from 8 × 10−8to 2 × 10−5per cell, depending on the concentration of amdinocillin used during selection. Several genes have been demonstrated to give amdinocillin resistance, but here eight novel genes previously unknown to be involved in amdinocillin resistance were identified. These genes encode functions involved in the respiratory chain, the ribosome, cysteine biosynthesis, tRNA synthesis, and pyrophosphate metabolism. The clinical isolates exhibited significantly greater fitness than the laboratory-isolated mutants and a different mutation spectrum. ThecysBgene was mutated (inactivated) in all of the clinical isolates, in contrast to the laboratory-isolated mutants, where mainly other types of more costly mutations were found. Our results suggest that the frequency of mutation to amdinocillin resistance is high because of the large mutational target (at least 38 genes). However, the majority of these resistant mutants have a low growth rate, reducing the probability that they are stably maintained in the bladder. Inactivation of thecysBgene and a resulting loss of cysteine biosynthesis are the major mechanism of amdinocillin resistance in clinical isolates ofE. coli.

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Biological Cost of Single and Multiple Norfloxacin Resistance Mutations in Escherichia coli Implicated in Urinary Tract Infections

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.6.2343-2351.2005 ◽

2005 ◽

Vol 49 (6) ◽

pp. 2343-2351 ◽

Cited By ~ 91

Author(s):

Patricia Komp Lindgren ◽

Linda L. Marcusson ◽

Dorthe Sandvang ◽

Niels Frimodt-Møller ◽

Diarmaid Hughes

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Infection Model ◽

Resistance Mutations ◽

Topoisomerase Iv ◽

E Coli ◽

Tract Infections ◽

Subsequent Selection

ABSTRACT Resistance to fluoroquinolones in urinary tract infection (UTIs) caused by Escherichia coli is associated with multiple mutations, typically those that alter DNA gyrase and DNA topoisomerase IV and those that regulate AcrAB-TolC-mediated efflux. We asked whether a fitness cost is associated with the accumulation of these multiple mutations. Mutants of the susceptible E. coli UTI isolate Nu14 were selected through three to five successive steps with norfloxacin. Each selection was performed with the MIC of the selected strain. After each selection the MIC was measured; and the regions of gyrA, gyrB, parC, and parE, previously associated with resistance mutations, and all of marOR and acrR were sequenced. The first selection step yielded mutations in gyrA, gyrB, and marOR. Subsequent selection steps yielded mutations in gyrA, parE, and marOR but not in gyrB, parC, or acrR. Resistance-associated mutations were identified in almost all isolates after selection steps 1 and 2 but in less than 50% of isolates after subsequent selection steps. Selected strains were competed in vitro, in urine, and in a mouse UTI infection model against the starting strain, Nu14. First-step mutations were not associated with significant fitness costs. However, the accumulation of three or more resistance-associated mutations was usually associated with a large reduction in biological fitness, both in vitro and in vivo. Interestingly, in some lineages a partial restoration of fitness was associated with the accumulation of additional mutations in late selection steps. We suggest that the relative biological costs of multiple mutations may influence the evolution of E. coli strains that develop resistance to fluoroquinolones.

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The Repeat-In-Toxin Family Member TosA Mediates Adherence of Uropathogenic Escherichia coli and Survival during Bacteremia

Infection and Immunity ◽

10.1128/iai.05713-11 ◽

2011 ◽

Vol 80 (2) ◽

pp. 493-505 ◽

Cited By ~ 31

Author(s):

Patrick D. Vigil ◽

Travis J. Wiles ◽

Michael D. Engstrom ◽

Lev Prasov ◽

Matthew A. Mulvey ◽

...

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Virulence Factors ◽

Upper Urinary Tract ◽

Host Cells ◽

Content Type ◽

E Coli ◽

Wide Range ◽

Novel Target ◽

Tract Infections

ABSTRACTUropathogenicEscherichia coli(UPEC) is responsible for the majority of uncomplicated urinary tract infections (UTI) and represents the most common bacterial infection in adults. UPEC utilizes a wide range of virulence factors to colonize the host, including the novel repeat-in-toxin (RTX) protein TosA, which is specifically expressed in the host urinary tract and contributes significantly to the virulence and survival of UPEC.tosA, found in strains within the B2 phylogenetic subgroup ofE. coli, serves as a marker for strains that also contain a large number of well-characterized UPEC virulence factors. The presence oftosAin anE. coliisolate predicts successful colonization of the murine model of ascending UTI, regardless of the source of the isolate. Here, a detailed analysis of the function oftosArevealed that this gene is transcriptionally linked to genes encoding a conserved type 1 secretion system similar to other RTX family members. TosA localized to the cell surface and was found to mediate (i) adherence to host cells derived from the upper urinary tract and (ii) survival in disseminated infections and (iii) to enhance lethality during sepsis (as assessed in two different animal models of infection). An experimental vaccine, using purified TosA, protected vaccinated animals against urosepsis. From this work, it was concluded that TosA belongs to a novel group of RTX proteins that mediate adherence and host damage during UTI and urosepsis and could be a novel target for the development of therapeutics to treat ascending UTIs.

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Determination of nitrofurantoin and fosfomycin susceptibility among urinary Escherichia coli isolates

International Journal of Research in Medical Sciences ◽

10.18203/2320-6012.ijrms20203676 ◽

2020 ◽

Vol 8 (9) ◽

pp. 3274

Author(s):

Rachana Kanaujia ◽

Amit Kumar ◽

Malay Bajpai

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Urinary Tract Infections ◽

Urine Samples ◽

E Coli ◽

Tract Infections ◽

Therapeutic Alternatives ◽

Micro Organisms ◽

Staphylococcus Spp

Background: Urinary tract infections (UTIs) are one of the most common infections. For treatment of UTIs, there are limited antibiotics due to increased resistance among uropathogens. Two older antibiotics; Nitrofurantoin and Fosfomycin have become novel oral therapeutic options against uropathogens. Aim of the study was to identify UTI causing micro-organisms and evaluate in-vitro activity of nitrofurantoin and fosfomycin against most common isolated organism (E. coli).Methods: Results of urine samples culture and susceptibility testing over a period of 1 year were analysed and included in this study.Results: Micro-organisms were isolated from 568 urine samples. Most commonly isolated organism was Escherichia coli (40.50%), followed by Klebsiella spp. (20.07%) and Staphylococcus spp. (17.07%). Susceptibility of E. coli to nitrofurantoin and fosfomycin was 91.74% and 65.65% respectively. Conclusion: Good activity of nitrofurantoin and fosfomycin against E. coli indicates that these two drugs are potential therapeutic alternatives for urinary tract infections.

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The Globoseries Glycosphingolipid Sialosyl Galactosyl Globoside Is Found in Urinary Tract Tissues and Is a Preferred Binding Receptor In Vitro for Uropathogenic Escherichia coli Expressing pap-Encoded Adhesins

Infection and Immunity ◽

10.1128/iai.66.8.3856-3861.1998 ◽

1998 ◽

Vol 66 (8) ◽

pp. 3856-3861 ◽

Cited By ~ 65

Author(s):

A. E. Stapleton ◽

M. R. Stroud ◽

S. I. Hakomori ◽

W. E. Stamm

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Kidney Tissue ◽

Human Kidney ◽

Blood Group Antigens ◽

E Coli ◽

Vaginal Epithelial Cells ◽

History Of ◽

Tract Infections

ABSTRACT Women with a history of recurrent Escherichia coliurinary tract infections (UTIs) are significantly more likely to be nonsecretors of blood group antigens than are women without such a history, and vaginal epithelial cells (VEC) from women who are nonsecretors show enhanced adherence of uropathogenic E. coli isolates compared with cells from secretors. We previously extracted glycosphingolipids (GSLs) from native VEC and determined that nonsecretors (but not secretors) selectively express two extended globoseries GSLs, sialosyl galactosyl globoside (SGG) and disialosyl galactosyl globoside (DSGG), which specifically bound uropathogenicE. coli R45 expressing a P adhesin. In this study, we demonstrated, by purifying the compounds from this source, that SGG and DSGG are expressed in human kidney tissue. We also demonstrated that SGG and DSGG isolated from human kidneys bind uropathogenic E. coli isolates expressing each of the three classes ofpap-encoded adhesins, including cloned isolates expressing PapG from J96, PrsG from J96, and PapG from IA2, and the wild-type isolates IA2 and R45. We metabolically 35S labeled these five E. coli isolates and measured their relative binding affinities to serial dilutions of SGG and DSGG as well as to globotriaosylceramide (Gb3) and globotetraosylceramide (Gb4), two other globoseries GSLs present in urogenital tissues. Each of the five E. coli isolates bound to SGG with the highest apparent avidity compared with their binding to DSGG, Gb3, and Gb4, and each isolate had a unique pattern of GSL binding affinity. These studies further suggest that SGG likely plays an important role in the pathogenesis of UTI and that its presence may account for the increased binding of E. colito uroepithelial cells from nonsecretors and for the increased susceptibility of nonsecretors to recurrent UTI.

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