Molecular Characterization of Uropathogenic Escherichia coli Reveals Emergence of Drug Resistant O15, O22 and O25 Serogroups.
ABSTRACT: Background and Objectives: Uropathogenic Escherichia coli (UPEC) are common pathogens causing urinary tract infections (UTIs). We aimed to investigate the relationship among clinical manifestation, serogroups, phylogenetic groups, and antimicrobial resistance among UPEC. Materials and Methods: One-hundred Escherichia coli isolates recovered from urine and ureteral scrapings were used for the study. The prevalence of antimicrobial resistance was determined by using European Committee on Antimicrobial Susceptibility Testing (EUCAST) recommendations. E. coli serogroups associated with UTI, as well as phylogenetic diversity were analyzed using multiplex PCR reactions. Results: Eighty-seven strains (87%) were isolated from females, while 13 (13%) from males. A high frequency of resistance to cephalosporins (43%) and fluoroquinolones (31%) was observed. Among UTI-associated serogroups O15 (32.8%), O22 (23.4%), and O25 (15.6%) were dominant and demonstrated elevated resistance rates. The E. coli phylogenetic group B2 was most common. These observations extended to pregnant patients with asymptomatic bacteriuria. Conclusions: Due to high rates of resistance, strategies using empirical therapy of second-generation cephalosporins and fluoroquinolones should be reconsidered in this population.
Project description:Urinary tract infections (UTIs) caused by Escherichia coli have been historically managed with oral antibiotics including the cephalosporins, fluoroquinolones and trimethoprim-sulfamethoxazole. The use of these agents is being compromised by the increase in extended spectrum ?-lactamase (ESBL)-producing organisms, mostly caused by the emergence and clonal expansion of E. coli multilocus sequence typing (ST) 131. In addition, ESBL isolates show co-resistance to many of oral agents. Management of UTIs caused by ESBL and fluoroquinolone-resistant organisms is becoming increasingly challenging to treat outside of the hospital setting with clinicians having to resort to intravenous agents. The aim of this study was to assess the prevalence of ESBL phenotypes and genotypes among UTI isolates of E. coli collected in the US during 2017 as well as the impact of co-resistance to oral agents such as the fluoroquinolones and trimethoprim-sulfamethoxazole. The national prevalence of ESBL phenotypes of E. coli was 15.7% and was geographically distributed across all nine Census regions. Levofloxacin and trimethoprim-sulfamethoxazole-resistance rates were ? 24% among all isolates and this co-resistance phenotype was considerably higher among isolates showing an ESBL phenotype (? 59.2%) and carrying blaCTX-M-15 (? 69.5%). The agents with the highest potency against UTI isolates of E. coli, including ESBL isolates showing cross-resistance across oral agents, were the intravenous carbapenems. The results of this study indicate that new oral options with the spectrum and potency similar to the intravenous carbapenems would address a significant unmet need for the treatment of UTIs in an era of emergence and clonal expansion of ESBL isolates resistant to several classes of antimicrobial agents, including oral options.
Project description:Urinary tract infection (UTI) is a prominent global health care burden. Although UTI is readily treated with antibiotics in healthy adults, complicated cases in immune-compromised individuals and the emerging antibiotic resistance of several uropathogens have accelerated the need for new treatment strategies. Here, we surveyed the composition of urinary exosomes in a mouse model of uropathgenic Escherichia coli (UPEC) UTI to identify specific urinary tract defense constituents for therapeutic development. We found an enrichment of the iron-binding glycoprotein lactoferrin in the urinary exosomes of infected mice. In subsequent in vitro studies, we identified human bladder epithelial cells as a source of lactoferrin during UPEC infection. We further established that exogenous treatment with human lactoferrin (hLf) reduces UPEC epithelial adherence and enhances neutrophil antimicrobial functions including bacterial killing and extracellular trap production. Notably, a single intravesicular dose of hLf drastically reduced bladder bacterial burden and neutrophil infiltration in our murine UTI model. We propose that lactoferrin is an important modulator of innate immune responses in the urinary tract and has potential application in novel therapeutic design for UTI.
Project description:Urinary tract infection (UTI) is the second most common infection in humans, making it a global health priority. Nearly half of all women will experience a symptomatic UTI, with uropathogenic Escherichia coli (UPEC) being the major causative agent of the infection. Although there has been extensive research on UPEC virulence determinants, the importance of host-specific metabolism remains understudied. We report here that UPEC upregulates the expression of ethanolamine utilization genes during uncomplicated UTIs in humans. We further show that UPEC ethanolamine metabolism is required for effective bladder colonization in the mouse model of ascending UTI and is dispensable for bladder colonization in an immunocompromised mouse model of UTI. We demonstrate that although ethanolamine metabolism mutants do not show increased susceptibility to antimicrobial responses of neutrophils, this metabolic pathway is important for surviving the innate immune system during UTI. This study reveals a novel aspect of UPEC metabolism in the host and provides evidence for an underappreciated link between bacterial metabolism and the host immune response.
Project description:To characterise the dissemination patterns of uropathogenic Escherichia coli (UPEC) in a community, we conducted a study utilising molecular and fundamental descriptive epidemiology. The subjects, consisted of women having community-acquired acute urinary tract infection (UTI), were enrolled in the study from 2011 to 2012. UPEC isolates were subjected to antibacterial-susceptibility testing, O serogrouping, phylotyping, multilocus-sequence typing with phylogenetic-tree analysis and pulsed-field-gel electrophoresis (PFGE). From the 209 unique positive urinary samples 166 UPEC were isolated, of which 129 were fully susceptible to the tested antibiotics. Of the 53 sequence types (STs), the four most prevalent STs (ST95, ST131, ST73 and ST357) accounted for 60% of all UPEC strains. Antimicrobial resistance was less frequently observed for ST95 and ST73 than for the others. A majority of rare STs and a few common STs constituted the diversity pattern within the population structure, which was composed of the two phylogenetically distinct clades. Eleven genetically closely related groups were determined by PFGE, which accounted for 42 of the 166 UPEC isolates, without overt geo-temporal clustering. Our results indicate that a few major lineages of UPEC, selected by unidentified factors, are disseminated in this community and contribute to a large fraction of acute UTIs.
Project description:Virulence genes from different E. coli pathotypes are blended in hybrid strains. E. coli strains with hybrid enteroaggregative/uropathogenic (EAEC/UPEC) genotypes have sporadically emerged causing outbreaks of extraintestinal infections, however their association with routine infections is yet underappreciated. We assessed 258 isolates of E. coli recovered from 86 consecutive cases of extraintestinal infections seeking EAEC and hybrid genotype (EAEC/UPEC) strains. Extensive virulence genotyping was carried out to detect 21 virulence genes, including molecular predictors of EAEC and UPEC strains. Phylogenetic groups and sequence types (STs) were identified, as well as it was performed phylogenetic analyses in order to evaluate whether hybrid EAEC/UPEC strains belonged to intestinal or extraintestinal lineages of E. coli. Adhesion assays were performed to evaluate the biofilm formation by hybrid strains in human urine and cell culture medium (DMEM). Molecular predictors of UPEC were detected in more than 70% of the strains (chuA in 85% and fyuA in 78%). Otherwise, molecular predictors of EAEC (aatA and aggR) were detected in only 3.4% (9/258) of the strains and always along with the UPEC predictor fyuA. Additionally, the pyelonephritis-associated pilus (pap) gene was also detected in all of the hybrid EAEC/UPEC strains. EAEC/UPEC strains were recovered from two cases of community-onset urinary tract infections (UTI) and from a case of bacteremia. Analyses revealed that hybrid EAEC/UPEC strains were phylogenetically positioned in two different clades. Two representative strains, each recovered from UTI and bacteremia, were positioned into a characteristic UPEC clade marked by strains belonging to phylogenetic group D and ST3 (Warwick ST 69). Another hybrid EAEC/UPEC strain was classified as phylogroup A-ST478 and positioned in a commensal clade. Hybrid EAEC/UPEC strains formed biofilms at modest, but perceptible levels either in DMEM or in urine samples. We showed that different lineages of E. coli, at least phylogenetic group A and D, can acquire and gather EAEC and UPEC virulence genes promoting the emergence of hybrid EAEC/UPEC strains.
Project description:Background/Purpose:Uropathogenic E. coli (UPEC) is the main cause of urinary tract infection (UTI) and it is known that pregnant women have a higher risk for UTI. UPEC has a variety of virulence and antibiotic resistance factors that facilitate its pathogenic success and it is crucial to know which are the susceptibility patterns, Extended-Spectrum-?-Lactamase (ESBL) production, virulence genes, pathogenicity islands (PAI), phylogenetic groups and serotypes among strains isolated from pregnant and non-pregnant women. Methods:One hundred fifty UPEC strains were isolated from pregnant and non-pregnant women from two different Mexican states (Sonora and Puebla). Strains were analyzed using the Kirby-Bauer method for the determination of antibiotic susceptibility and ESBL. Virulence genes, PAIs and phylogenetic groups were determined using a multiplex PCR. Strains were serotyped by an agglutination assay. Blood agar and CAS agar were used for phenotypic assays. Results:92.7% of UPEC strains showed multidrug-resistant (MDR), 6.7% extremely-resistant (XDR) and 0.6% pandrug-resistant (PDR). The highest resistance was determined to be for ?-lactam antibiotics (>72% in both states) and 44.5% of the UPEC strains were ESBL+. The predominant virulence genes found were fimH (100%), iucD (85%) and iha (60%). The strains isolated from pregnant women from Puebla presented a large percentage of genes associated with upper urinary tract infections. PAIs were found in 51% and 68% of the strains from Sonora and Puebla, respectively. All the strains were siderophores producers and 41.5% produced hemolysis. The serotypes found were diverse and belonged to phylogroups A, B2 and C. Conclusion:The UPEC strains from this study are MDR with tendency to XDR or PDR, they can cause upper UTIs and are serotypically and phylogenetically diverse, which supports the need to develop new strategies for UTI treatment in pregnant and non-pregnant Mexican women.
Project description:Objectives:To evaluate temporal trends in antimicrobial resistance, over 16 years, in bacteria isolated from dogs and cats with urinary tract infection (UTI) and the clonal lineages of bacteria harbouring critical antimicrobial resistance mechanisms. Methods:Antimicrobial susceptibility testing was conducted for 948 bacteria isolated from dogs and cats with UTI (1999-2014). Resistance mechanisms were detected by PCR, namely ESBL/AmpC in third-generation cephalosporin (3GC)-resistant Escherichia coli and Proteus mirabilis, mecA in methicillin-resistant staphylococci, and aac(6')-Ieaph(2?)-Ia and aph(2?)-1d in high-level gentamicin-resistant (HLGR) enterococci. Resistant bacteria were typed by MLST, and temporal trends in E. coli and Enterobacteriaceae antimicrobial resistance were determined by logistic regression. Results:Enterobacteriaceae had a significant temporal increase in resistance to amoxicillin/clavulanate, 3GCs, trimethoprim/sulfamethoxazole, fluoroquinolones, gentamicin and tetracycline (P?<?0.001). An increase in MDR was also detected (P?<?0.0001). 3GC resistance was mainly caused by the presence of blaCTX-M-15 and blaCMY-2 in E. coli and the presence of blaCMY-2 in P. mirabilis. Two major 3GC-resistant E. coli clonal lineages were detected: O25b:H4-B2-ST131 and ST648. The mecA gene was detected in 9.2% (n?=?11/119) of Staphylococcus spp., including MRSA clonal complex (CC) 5 (n?=?2) and methicillin-resistant Staphylococcus epidermidis CC5 (n?=?4). A temporal increase in MDR methicillin-resistant Staphylococcus pseudintermedius was detected (P?=?0.0069). Some ampicillin-resistant and/or HLGR Enterococcus spp. were found to belong to hospital-adapted CCs, namely Enterococcus faecalis ST6-CC6 (n?=?1) and Enterococcus faecium CC17 (n?=?8). Conclusions:The temporal increase in antimicrobial resistance and in MDR bacteria causing UTI in dogs and cats creates important therapeutic limitations in veterinary medicine. Furthermore, the detection of MDR high-risk clonal lineages raises public health concerns since companion animals with UTI may contribute to the spread of such bacteria.
Project description:Objectives: The aim of this study was to assess the clonal structure, virulence potential and antibiotic susceptibility of uropathogenic Escherichia coli (UPEC) isolates causing community acquired urinary tract infection (CAUTI) in unselected primary care patients in Switzerland. Methods: We performed multilocus sequence typing, virulence factor determination, and phenotypic and genotypic antimicrobial resistance testing on 44 non-duplicate UPEC isolates. Results: Twenty-seven different sequence types (STs) were identified. Major UPEC clones were represented by 19 (43.2%) of the isolates, including E. coli ST131, ST69 (both 13.6%), ST73 (6.8%), ST10 (4.5%), ST127, ST140, (both 2.3%). Five (11.4%) isolates belonged to ST141. Aggregate virulence factor (VF) scores were highest among isolates belonging to ST127 and ST141. Overall, 50% of the isolates were susceptible to all 12 antimicrobials tested, and all isolates remained susceptible to fosfomycin and nitrofurantoin. Resistance to sulfamethoxazole and ciprofloxacin were found in 31.8, and 15.9% of the isolates, respectively. Plasmid-mediated resistance genes were detected in ST69 and ST131 and included aac(6')-Ib-cr (2.3% of all isolates) blaCTX-M-14 and blaCTX-M-15 (9%), and mph(A) (13.6%). None of the isolates tested positive for mcr-1 or mcr-2. Conclusions: Our results show that CAUTI in Switzerland is caused by a wide variety of UPEC STs for which fosfomycin remains a good treatment option. We suggest that ST141 is an emerging clone associated with UTI in the community, and warrants closer attention. Moreover, the high rate of E. coli harboring mph(A) from patients without a history of antimicrobial therapy or hospitalization indicates that UPEC is an important reservoir for mph(A).
Project description:Uropathogenic Escherichia coli (UPEC) is the most common pathogen of urinary tract infections (UTIs). Antibiotic therapy is the conventional measure to manage such infections. However, the rapid emergence of antibiotic resistance has reduced the efficacy of antibiotic treatment. Given that the bacterial factors required for the full virulence of the pathogens are potential therapeutic targets, identifying such factors may facilitate the development of novel therapeutic strategies against UPEC UTIs. The peptidoglycan (PG) endopeptidase Spr (also named MepS) is required for PG biogenesis in E. coli. In the present study, we found that Spr deficiency attenuated the ability of UPEC to infect kidneys and induced a fitness defect during bladder colonization in a mouse model of UTI. Based on the liquid chromatography (LC)/mass spectrometry (MS)/MS analysis of the bacterial envelope, spr deletion changed the levels of some envelope-associated proteins, suggesting that Spr deficiency interfere with the components of the bacterial structure. Among the proteins, FliC was significantly downregulated in the spr mutant, which is resulted in reduced motility. Lack of Spr might hinder the function of the flagellar transcriptional factor FlhDC to decrease FliC expression. The motility downregulation contributed to the reduced fitness in urinary tract colonization. Additionally, spr deletion compromised the ability of UPEC to evade complement-mediated attack and to resist intracellular killing of phagocytes, consequently decreasing UPEC bloodstream survival. Spr deficiency also interfered with the UPEC morphological switch from bacillary to filamentous shapes during UTI. It is known that bacterial filamentation protects UPEC from phagocytosis by phagocytes. In conclusion, Spr deficiency was shown to compromise multiple virulence properties of UPEC, leading to attenuation of the pathogen in urinary tract colonization and bloodstream survival. These findings indicate that Spr is a potential antimicrobial target for further studies attempting to develop novel strategies in managing UPEC UTIs.
Project description:Urinary tracts infection (UTI) caused by uropathogenic Escherichia coli (UPEC) is a common infectious disease. With the shortage of new antimicrobial agents, the increase in UPEC resistance to commonly used drugs, such as fluoroquinolones and ?-lactams including carbapenems is a critical issue. UPEC invades urinary tract cells, where it aggregates, and subsequently, forms biofilm-like multicellular colonies termed intracellular bacterial communities (IBCs). This process allows the bacteria to establish infections and so may be a good potential target for new drugs to treat infections. Here, we show that deletion of the tolB gene, encoding a protein of the Tol-Pal system that was originally characterized as a protein complex for colicin uptake and maintenance of the outer membrane, decreases the level of bacterial internalization into and aggregation within cultured bladder epithelial cells and also inhibits the colonization of mice urinary tracts. The tolB mutant also exhibited defective motility because of impaired flagellum syntheses. The fliC and motA mutants, which are non-motile strains, also exhibited lower levels of bacterial internalization and aggregation than their wild-type parent. Additional deletion of tolB in the fliC mutant did not further decrease these, suggesting that the attenuated virulence of the tolB mutant is a result of defective motility. The tolA, tolQ, tolR, and pal mutants that lack other members of the Tol-Pal system also exhibited lower levels of motility and aggregation within bladder epithelial cells compared to their wild-type parent. These combined results suggest another role of the Tol-Pal system, i.e., that it is responsible for optimal internalization, aggregation followed by IBC formation within urinary tract cells, and bacterial motility.