Escherichia coli isolated from pyometra and cystitis in the same animal exhibit a wide phenotypic similarity.

AIMS: Pyometra and cystitis caused by Escherichia coli are common diseases identified in canine or feline females. The origin of pyometra infection remains uncertain, and effective prevention strategies for this disease are still unknown. This study aimed to provide a phenotypic characterization, including antimicrobial resistance and virulence profiles, of endometrial pathogenic (EnPEC) and uropathogenic (UPEC) E. coli strains isolated simultaneously from the same animal. METHODS AND RESULTS: Sixteen E. coli strains, from eight different animals, were analyzed in this study. The antimicrobial susceptibility profile of EnPEC and UPEC strains was determined using the disc diffusion method, which showed a similar susceptibility profile among strains (EnPEC and UPEC) from the same animal. The virulence profile of the strains was assessed through biofilm formation, as well as serum resistance abilities. EnPEC and UPEC strains from the same animal exhibited slight variations in their virulence and antimicrobial resistance capabilities. Overall, most of the strain pairs showed a high similarity in their ability to establish biofilms and survive in serum complement activity. CONCLUSIONS: Overall, strains of E. coli isolated from both pyometra and cystitis in the same animal, despite presenting distinct clinical diseases, exhibit a wide phenotypic similarity, suggesting a common origin for the strains.

Biofilm formation and antimicrobial resistance pattern of uropathogenic E. coli ST131 isolated from children with malignant tumors.

The multidrug-resistant clone identified as Escherichia coli sequence type 131 (E. coli ST131) has spread world-wide. This study sought to ascertain the frequency and biofilm formation of E. coli ST131 isolated from children with various malignancies. A total of 60 uropathogenic E. coli (UPEC) isolates from children without cancer and 30 UPEC isolates from children with cancer were assessed in this study. The microdilution method was used to investigate the sensitivity of bacteria to antibiotics. The microtiter plate (MTP) approach was used to phenotypically assess biofilm formation. The lasR, pelA, and lecA biofilm-encoding genes were detected by PCR in biofilm-producing isolates of E. coli. Thirty-seven out of 90 E. coli isolates were found to be ST131 (41.1%), with 17 (56.7%) from cancer-affected children and 20 (33.3%) from children without cancer, respectively (P-value = 0.036). The frequency of antimicrobial resistance was higher in ST131 strains were compared to non-ST131 strains and when they were isolated from healthy children vs. those who had cancer. In contrast to non-ST131 isolates, ST131 isolates were more biofilm-producers. There was a significant difference between the percentage of biofilm producers between the 22 (100%) ST131-O16 isolates and the 13 (86.7%) ST131-O25b isolates (P-value = 0.04). Children with cancer are more likely than children without cancer to develop biofilm forming E. coli ST131, the latter having a higher profile of antibiotic resistance. Interestingly, E. coli ST131 isolates from non-cancer patients had higher levels of overall antibiotic resistance and while more E. coli ST131isolates from cancer patients formed biofilms.

Studies on Virulence and Extended-Spectrum ß-Lactamase-Producing Uropathogenic Escherichia coli Isolates and Therapeutic Effect of Fosfomycin in Acute Pyelonephritis Mice.

The understanding about virulence factors (VFs) and the drug resistance of uropathogenic Escherichia coli (UPEC) helps us understand the pathogenesis of urinary tract infections (UTIs) and make better decisions for clinical treatment. This study examined the correlation between the extended-spectrum ß-lactamases (ESBLs) phenotype and VFs in UPEC strains. In addition, we validated the therapeutic potential of fosfomycin in acute pyelonephritis mice. From May 2017 to November 2018, 22 nonduplicate E coli. strains were isolated from UTI patients. PCR was utilized to detect the distribution of virulence genes. We also analyzed the ESBL phenotype in E coli. We further evaluated the therapeutic effect of intravenous fosfomycin treatment in the acute pyelonephritis (APN) model. All 22 UPEC strains expressed the type 1 fimbriae (FimH) gene and more than 50% (12/22) of strains produced ESBLs. The detection rates of the iron acquisition-associated genes ChuT and IutA were 77.3% (n = 17) and 50% (n = 11) and those of P fimbria papA and papC genes were 45% (n = 10) and 50% (n = 11), respectively. Though the VFs were closely related with pathologenicity, the relationship between VFs and ESBLs still needs further investigation. Furthermore, intravenous fosfomycin 800 mg/kg significantly reduced the bacterial load and the inflammatory infiltration in the bladder and kidney, maintaining the structural integrity of the kidney. Intravenous fosfomycin administration can be used for the treatment of acute pyelonephritis caused by highly pathogenic and drug-resistant UPEC strains.

Prevalence of common carbapenemase genes and multidrug resistance among uropathogenic Escherichia coli phylogroup B2 isolates from outpatients in Wasit Province/ Iraq.

Carbapenems are the last resort antimicrobials for the treatment of extended spectrum ß-lactamases (ESBLs) producing Enterobacteriaceae. Emergence of carbapenems resistant group B2 uropathogenic E. coli (UPEC) is a major concern because of their high virulence. Prevalence of these enzymes and multidrug resistance (MDR) among B2 UPEC isolates from Iraqi outpatients with acute urinary tract infection (UTI) was evaluated in this research. Urine cultures were performed and the isolates were identified biochemically. Escherichia coli isolates were tested for phylogroup reference by quadraplex PCR, then B2 isolates were detected for antimicrobial resistance by disc diffusion test and carbapenemase genes by PCR. Escherichia coli was the most prevalent among Gram-negative isolates (66.6%) and B2 was the most detected phylogroup among E. coli isolates (33.9%). Most of B2 isolates showed high resistance rates to tested antimicrobials, especially ß-lactams with MDR revealed in 100% of them. Whereas, low resistance rates were noted against carbapenems, aminoglycosides and nitrofurantoin. Carbapenemase genes were detected in 76.3% of B2 isolates. Of which, blaOXA-48 was the most frequent (57.8%), followed by blaPER (47.3%), blaKPC (15.7%), blaVEB and blaVIM (10.5%, for each). Whereas, blaGES and blaIMP genes were not found. Coproduction of these genes occurred among 17 isolates. The combination of blaOXA-48 and blaPER was the most frequent (41.1%). All carbapenemase producing isolates were MDR. These results revealed high prevalence of carbapenemase genes and MDR among B2 UPEC recovered in this study. In the study area. it is strongly advised to use aminoglycosides and nitrofurantoin for empirical treatment of UPEC.

Molecular epidemiology of blaCTX-M gene-producing uropathogenic Escherichia coli among Iranian kidney transplant patients: clonal dissemination of CC131 and CC10.

BACKGROUND: This study aimed to investigate the phylogenetic characterization and virulence traits of uropathogenic Escherichia coli (UPEC) isolated from kidney transplant patients (KTPs) as well as non-KTPs and analyze the clonal distribution of Extended spectrum ß-lactamases (ESBLs)-producing UPEC containing blaCTX-M gene. METHODS: To this end, we determined virulence marker and the phylogenetic characterization of UPEC in non-KTPs (n = 65) and KTPs (n = 46). The non-KTPs were considered the control group of the study. Also, according to the Achtman scheme, we performed multilocus sequence typing to assess the relationship between twenty-nine of ESBL-producing isolates containing blaCTX-M gene. RESULTS: According to the results of PCR assay, the prevalence of virulence factor genes ranged from 0% (cnf and papG III) to 93.7% (fimH). Also, KTP isolates significantly differed from non-KTP isolates only in terms of the prevalence of pap GI elements. Moreover, the most frequent UPEC isolates were in phylogenetic group B2, followed by group D (18.9%), and group A (13.5%). Furthermore, except for phylogenetic group C, there was no significant correlation between phylogenetic distribution in KTPs and non-KTPs. Additionally, MLST analysis of blaCTX-M carrying isolates identified 18 unique sequence types (ST) the most common of which was ST131 (24.1%), followed by ST1193 (10.3%), while fourteen STs were detected only once. CONCLUSIONS: The results further revealed significant differences between the UPEC isolates from KTPs and non-KTPs regarding the phylogroups C and PAI gene. Based on MLST analysis, we also observed a relatively high diversity in UPEC isolates obtained from KTPs and non-KTPs. Moreover, clonal complex (CC) 131 and ST131 were found to be the most prevalent clones and ST types, respectively. Besides, for the first time, ST8503 were reported in KTPs. These results suggested regular studies on characterization of UPEC isolates among KTPs.

Targeting of Uropathogenic Escherichia coli papG gene using CRISPR-dot nanocomplex reduced virulence of UPEC.

Urinary tract infections (UTI) are the most common infectious diseases in the world. It is becoming increasingly tough to treat because of emergence of antibiotic resistance. So, there is an exigency to develop novel anti-virulence therapeutics to combat multi-drug resistance pathogenic strains. Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) discovery has revolutionized the gene editing technology for targeted approach. The greatest obstacle for CRISPR/Cas9 is cargo delivery systems and both viral and plasmid methods have disadvantages. Here, we report a highly efficient novel CRISPR based gene editing strategy, CRISPR-dots for targeting virulence factor Fimbrial Adhesion (papG gene), the bacterial adhesion molecule. Carbon quantum dots (CQD) were used as a delivery vehicle for Cas9 and gRNA into CFT073, a UPEC strain. CQDs were covalently conjugated to cas9 and papG-targeted guide RNA (gRNA) forming a nanocomplex CRISPR-dots (Cri-dots) as confirmed by DLS and transmission electron microscopy. Cri-dots-papG significantly targeted papG as demonstrated by decrease in the expression of papG.Further papG deficient UPEC had significantly reduced adherence ability and biofilm forming ability as demonstrated by fluorescence microscopy and scanning electron microscopy. Also, papG deficient UPEC had reduced virulence as shown by significantly increased survival of Caenorhabditis elegans (C. elegans) worms compared to UPEC. Our findings suggest that targeting of papG gene using Cri-dots nanocomplexes significantly reduced the pathogenicity of UPEC. Thus, Cri-dots nanocomplex offer a novel anti-bacterial strategy against multi-drug resistant UPEC.

Characterization of Anti-Bacterial Effect of the Two New Phages against Uropathogenic Escherichia coli.

Urinary tract infections (UTIs) are among the events that most frequently need medical intervention. Uropathogenic Escherichia coli are frequently their causative agents and the infections are sometimes complicated by the presence of polyresistant nosocomial strains. Phage therapy is a tool that has good prospects for the treatment of these infections. In the present study, we isolated and characterized two bacteriophages with broad host specificity against a panel of local uropathogenic E. coli strains and combined them into a phage cocktail. According to genome sequencing, these phages were closely related and belonged to the Tequatrovirus genus. The newly isolated phages showed very good activity on a panel of local clinical E. coli strains from urinary tract infections. In the form of a two-phage cocktail, they were active on E. coli strains belonging to phylogroups B2 and D, with relatively lower activity in B1 and no response in phylogroup A. Our study is a preliminary step toward the establishment of a national phage bank containing local, well-characterized phages with therapeutic potential for patients in Slovakia.

Virulence characterization and clonal analysis of uropathogenic Escherichia coli metallo-beta-lactamase-producing isolates.

BACKGROUND: Uropathogenic Escherichia coli (UPEC) is a major cause of urinary tract infection (UTI); however, treatment of UTI has been challenging due to increased antimicrobial resistance (AMR). One of the most important types of AMR is carbapenem resistance (CR). CR bacteria are known as an important threat to global public health today. Class B metallo-beta-lactamases (MBLs) are one of the major factors for resistance against carbapenems. We aimed to investigate the characteristics of UPEC isolates producing MBL. METHODS: A cross-sectional study was conducted from October 2018 to December 2019 in Ahvaz; Iran. UPEC isolates were identified by biochemical and molecular methods. Metallo-beta-lactamase-producing isolates were detected using modified carbapenem inactivation method (mCIM) and EDTA-CIM (eCIM) tests. MBL genes, phylogenetic group, and virulence genes profile of carbapenem resistant isolates were determined. Conjugation assay and plasmid profiling were conducted to evaluate the ability of transferring of CR to other E. coli isolates. Clonal similarity of isolates were assessed using Enterobacterial intergenic repetitive element sequence (ERIC)-PCR. RESULTS: Among 406 UPEC isolates, 12 (2.95%) carbapenem-resistant were detected of which 11 were phenotypically MBL-producing strains. Four isolates were resistant to all investigated antimicrobial agents and were considered possible pandrug-resistant (PDR). blaNDM, blaOXA-48, blaIMP-1, and blaIMP-2 genes were found in 9, 5, 1, and 1 isolates, respectively. Among 30 virulence genes investigated, the traT, fyuA followed by fimH, and iutA with the frequency of 8 (66.7%), 8 (66.7%), 7 (58.3%), and 7 (58.3%) were the most identified genes, respectively. Siderophore production was the main virulence trait among carbapenem-resistant UPEC isolates. Except for two, all other isolates showed weak to moderate virulence index. In all recovered isolates, CR was readily transmitted via plasmids to other isolates during conjugation experiments. CONCLUSION: MBL and carbapenemase genes, especially blaNDM and blaOXA-48 are spreading rapidly among bacteria, which can be a threat to global public health. Therefore monitoring the emergence and dissemination of new AMR is necessary to continuously refine guidelines for empiric antimicrobial therapy. Understanding the mechanisms of resistance and virulence in this group of bacteria can play an effective role in providing new therapeutic methods.

Two Novel Mutations Associated with Polymyxin-B Resistance in a Pandemic Lineage of Uropathogenic Escherichia coli of the Sequence Type 69.

BACKGROUND: Uropathogenic Escherichia coli (UPEC) are frequent pathogens worldwide, impacting on the morbidity and economic costs associated with antimicrobial treatment. OBJECTIVES: We report two novel mutations associated with polymyxin-B resistance in an UPEC isolate collected in 2019. METHODS: Isolate was submitted to antimicrobial susceptibility testing including broth microdilution for polymyxin B. Whole genome was sequenced and analyzed. RESULTS: Polymyxin-B total inhibition occurred at 16 mg/L (resistant). UPEC isolate was assigned to the phylogroup D, serotype O117:H4, and Sequence Type 69. mcr genes were not detected, but two novel mutations in the pmrA/basS (A80S) and pmrB/basR (D149N) genes were identified. CONCLUSIONS: The occurrence of non-mcr polymyxin resistance in E. coli from extraintestinal infections underscores the need of a continuous surveillance of this evolving pathogen.

Waterborne Urinary Tract Infections: Have We Overlooked an Important Source of Exposure?

The presence of intestinal pathogenic Escherichia coli in drinking water is well recognized as a risk for diarrhea. The role of drinking water in extraintestinal infections caused by E. coli-such as urinary tract infections (UTIs)-remains poorly understood. Urinary tract infections are a leading cause of outpatient infections globally, with a lifetime incidence of 50-60% in adult women. We reviewed the scientific literature on the occurrence of uropathogenic E. coli (UPEC) in water supplies to determine whether the waterborne route may be an important, overlooked, source of UPEC. A limited number of studies have assessed whether UPEC isolates are present in drinking water supplies, but no studies have measured whether their presence in water may increase UPEC colonization or the risk of UTIs in humans. Given the prevalence of drinking water supplies contaminated with E. coli across the globe, efforts should be made to characterize UTI-related risks associated with drinking water, as well as other pathways of exposure.

Virulence genes and phylogenetic groups of uropathogenic Escherichia coli isolates from patients with urinary tract infection and uninfected control subjects: a case-control study.

BACKGROUND: Urinary Tract Infection (UTI) is one of the most common bacterial infectious diseases which causes considerable morbidity and costly health problems. Uropathogenic Escherichia coli (UPEC), the most common pathogen causing UTI, is a highly heterogeneous group of extraintestinal pathogenic E. coli (ExPEC) which may carry a variety of virulence factors and belonging to different phylogenetic backgrounds. The current study aimed to investigate the frequency and association between various virulence factors (VFs) and phylogenetic groups of UPEC and commensal isolates. METHODS: UPEC and commensal E. coli strains isolated from UTI and feces of healthy humans were compared for the presence of VFs and phylogenetic groups. Association between virulence genes was investigated and cluster analysis was employed. RESULTS: According to the results, among a 30 virulence markers tested, the pathogenicity-associated island (PAI), papAH, papEF, fimH, fyuA, and traT genes prevalence were statistically significant in UPEC isolates. A strong association was found between the B2 and D phylogenetic groups and clinical isolates of UPEC; while, commensal isolates were mostly associated with phylogenetic group A. The aggregated VFs scores were more than twice higher in the UPEC isolates in comparison with the commensal isolates. Interestingly, the B2 group in both UPEC and commensal isolates had the highest VF scores. A strong positive association was found between several virulence genes. The clustering results demonstrated that UPEC or commensal E. coli isolates were highly heterogeneous due to different composition of their virulence gene pool and pathogenicity islands. CONCLUSION: Genetic structure and VFs of UPEC strains vary from region to region; therefore, to control the UTI, the epidemiological aspects and characterization of the UPEC isolates need to be investigated in different regions. Since UPEC isolates are generally originate from the commensal strains, it may be feasible to reduce the UTI burden by interfering the intestinal colonization, particularly in the highly pathogenic clonal lineages such as B2.

Uropathogenic E. coli induces DNA damage in the bladder.

Urinary tract infections (UTIs) are among the most common outpatient infections, with a lifetime incidence of around 60% in women. We analysed urine samples from 223 patients with community-acquired UTIs and report the presence of the cleavage product released during the synthesis of colibactin, a bacterial genotoxin, in 55 of the samples examined. Uropathogenic Escherichia coli strains isolated from these patients, as well as the archetypal E. coli strain UTI89, were found to produce colibactin. In a murine model of UTI, the machinery producing colibactin was expressed during the early hours of the infection, when intracellular bacterial communities form. We observed extensive DNA damage both in umbrella and bladder progenitor cells. To the best of our knowledge this is the first report of colibactin production in UTIs in humans and its genotoxicity in bladder cells.

Non-lactose-fermenting uropathogenic Escherichia coli from dogs: virulence profile characterization.

Non-lactose-fermenting Escherichia coli (NLFEC) has a few descriptive studies restricted to human infections. In the present study, isolates of NLFEC obtained from urine samples of dogs with hyperadrenocorticism were characterized regarding their virulence ability, biofilm formation capacity and antimicrobial susceptibility profile. Escherichia coli lactose-fermenting strains from urinary infection in dogs with the same conditions were analysed to provide comparisons. The non-lactose-fermenting E. coli strains were classified as belonging to clade I E. coli, whereas the lactose-fermenting strains were classified in phylogroup B2. All strains presented virulence markers to adhesion, iron acquisition, toxins, colicin and cytotoxin production, and biofilm regulation. Components of the extracellular matrix in addition to the in vitro biofilm formation ability were observed in the strains. Multidrug resistance (MDR) profiles were observed by in vitro susceptibility tests to all NLFEC strains. In summary, non-lactose-fermenting uropathogenic E. coli from dogs behaves similar to lactose-fermenting E. coli, exhibiting MDR profile, and pathogenic potential of promote animal infections.

Characterization of Plasmid-Mediated Quinolone Resistance and Serogroup Distributions of Uropathogenic Escherichia coli among Iranian Kidney Transplant Patients.

INTRODUCTION: Urinary tract infection (UTI) is one of the most frequent infections in kidney transplant patients (KTPs). This infection is mainly caused by uropathogenic Escherichia coli (UPEC). Plasmid-mediated quinolone resistance (PMQR) was also increasingly identified in UPEC. This study proposed to investigate the frequency of quinolone-resistance plasmid genes and the O-antigen serogroup among UPEC isolated from KTPs and non-KTP with UTI. METHODS: Totally, 114 UPEC isolates from 49 KTPs and 65 non-KTPs patients diagnosed with an UPEC-associated UTI were obtained from June 2019 to December 2019 at three laboratory centers in Isfahan, Iran. The isolates were confirmed through phenotypic and genotypic methods. Moreover, the antimicrobial susceptibility test to nalidixic acid, ciprofloxacin, norfloxacin, and ofloxacin was performed using a disk diffusion method. The presence of the qnr gene as well as the serogroup distribution was identified using the PCR method. RESULT: According to data, the distribution of O1, O2, O4, O16, and O25 serogroups were 3.5%, 2.6, 3.5, 3.5, and 20.2%, respectively. Antibiotic susceptibility pattern revealed that the highest and lowest resistance rates were to nalidixic acid (69.3%) and norfloxacin (43.9%), respectively. Also, the frequency of qnrS and qnrB genes were 33.3% and 15.8%, respectively, while none of the isolates was found to be positive for the qnrA gene. There was no significant association between the presence of qnr genes and higher antibiotic resistance. CONCLUSION: This study recognized that the qnrS gene, O25 serotype, and resistance to nalidixic acid had the highest frequencies in UPEC strains isolated from UTI patients.

Isolation and characterization of Uropathogenic Escherichia coli (UPEC) from red panda (Ailurus fulgens).

BACKGROUND: Disease prevention and control is a significant part in the ex-situ conservation of the endangered red panda (Ailurus fulgens), being bacterial infection is one of the most important health threats to the captive population. To date, studies about the infection caused by Escherichia coli in the red panda are scarce. This study was conducted to determine the cause of death of a captive red panda through clinical symptoms, complete blood count, biochemical analysis, pathological diagnosis and bacterial whole genome sequencing. CASE PRESENTATION: The following report describes a case of a 1.5 year old captive red panda (Ailurus fulgens) that was found lethargic and anorectic. She was moved to the quarantine area for daily treatment with 50 mg of Cefpodoxime Proxetil. During the three-day treatment, she did not eat or defecate, and then died. Clinical hematology revealed the values of neutrophils, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and blood urea nitrogen (BUN) were significantly higher. Histological analysis demonstrated major pathological damage in the kidneys, liver and lungs, characterized by hyperemia, parenchymal cell degeneration and necrosis and inflammatory cell infiltration which were predominantly neutrophilic. A bacterial strain confirmed as Escherichia coli was isolated post mortem. Whole genome sequencing of the E. coli showed the complete genome size was 4.99 Mbp. PapA, PapC, OmpA, OmpU and other virulence factors which specific to Uropathogenic Escherichia coli (UPEC) were found in the isolate. Among the virulence factors, P pili, type I pili and related factors of the iron uptake system were associated with nephrotoxicity. CONCLUSION: The red panda died of bacterial infection caused by an uropathogenic strain of Escherichia coli. The pathogenic mechanisms of the strain are closely related to the expression of specific virulence genes.

Metabolic acidosis exacerbates pyelonephritis in mice prone to vesicoureteral reflux.

Acute pyelonephritis is a common, serious bacterial infection in children. The prevalence of acute pyelonephritis is due at least in part to vesicoureteral reflux (VUR). Although an association between abnormalities in electrolyte and acid-base balance and pyelonephritis is common in young children, the impact of metabolic acidosis (MA) on progression of acute pyelonephritis is not fully understood. In this study, the effect of MA on pyelonephritis was studied in C3H mouse strains prone to VUR. MA induced by ammonium chloride supplementation in food specifically impaired clearance of urinary tract infection with uropathogenic Escherichia. coli (UPEC-UTI) in innate immune competent C3H strains (HeOuJ, HeN), whereas kidney UPEC burden in Tlr-4-deficient HeJ mice was unaffected. Antibody-mediated depletion of myeloid cells (monocytes, neutrophil) markedly increased UPEC burden in the bladder and kidney confirming the pivotal role of neutrophils and tissue-resident macrophages in clearance of UPEC-UTI. MA concurrent with UPEC-UTI markedly increased expression of cytokine (TNFα, IL-1ß, IL-6) and chemokine (CXCL 1, 2, and 5) mRNA in isolated kidney CD cells and kidney neutrophil infiltrates were increased four- to fivefold compared to normal, UPEC-infected mice. Thus, MA intensified pyelonephritis and increased the risk of kidney injury by impairing clearance of UPEC-UTI and potentiating renal inflammation characterized by an elevated kidney neutrophil infiltrate.

Significant association between genes encoding virulence factors with antibiotic resistance and phylogenetic groups in community acquired uropathogenic Escherichia coli isolates.

BACKGROUND: Antibiotic resistance is an increasing phenomenon in many bacterial pathogens including uropathogenic Escherichia coli. Hypothetical anti-virulent agents could be a solution, but first clear virulence associated gene-pool of antibiotic resistant isolates have to be determined. The aim of this study is to investigate the significant associations between genes encoding VFs with antibiotic resistance and phylogenetic groups in UPEC isolates. RESULTS: The majority of 248 UPEC isolates belonged to phylogenetic group B2 (67.3%). The maximum and minimum resistance was attributed to amoxicillin (90.3%) and both fosfomycin and imipenem (1.6%) respectively. 11.3% of isolates were resistant to all antibiotic agents except that of imipenem, nitrofurantoin and fosfomycin. These highly resistant isolates were placed only in group B2 and D. The most prevalent virulence gene was ompA (93.5%). The hlyA was the only virulence gene that was significantly more prevalent in the highly resistant isolates. The ompA, malX and hlyA genes were obviously more abundant in the antibiotic resistant isolates in comparison to susceptible isolates. The papC gene was associated with amoxicillin resistance (p-value = 0.006, odds ratio: 26.00). CONCLUSIONS: Increased resistance to first line drugs prescribed for UTIs were detected in CA-UPEC isolates in our study.. Minimal resistance was observed against nitrofurantoin, fosfomycin and imipenem. Therefore, they are introduced for application in empirical therapy of UTIs. Fosfomycin may be the most effective antibiotic agent against highly resistant UPEC isolates. The presence of the ompA, malX and hlyA genes were significantly associated with resistance to different antibiotic agents. We assume that the ability of UPEC isolates to upgrade their antibiotic resistance capacity may occurs in compliance with the preliminary existence of specific virulence associated genes. But, more investigation with higher number of bacterial isolates, further virulence associated genes and comparison of gene pools from CA-UPEC isolates with HA-UPEC are proposed to confirm these finding and discovering new aspects of this association.

Prevalence of fluoroquinolone-resistant and broad-spectrum cephalosporin-resistant community-acquired urinary tract infections in Rio de Janeiro: Impact of Escherichia coli genotypes ST69 and ST131.

Uropathogenic Escherichia coli (UPEC) is the leading cause of community-acquired urinary tract infection (CA-UTI). The increasing prevalence of CA-UTI caused by UPEC strains resistant to broad-spectrum drugs complicates clinical management of these infections. Here we assessed the prevalence of antimicrobial drug resistance, genotypes and beta-lactamase genes among UPEC isolated from cases of CA-UTI in Rio de Janeiro, Brazil during November 2015 to determine if the prevalence of drug-resistant CA-UTI is determined by multiple genotypes of resistant UPEC or dissemination of key lineages of UPEC. Among 499 UPEC isolates, 98 (20%) were ciprofloxacin (CIP) resistant and 41 (8%) produced extended-spectrum beta-lactamase (ESBL). Sequence types (ST) 69 and 131 were the most common genotypes, representing 77 (15%) and 42 (8%) of all UPEC isolates, respectively. Of fluoroquinolone-resistant isolates, ST69 and ST131 together accounted for 57%, while of ESBL-producers, ST131 represented 21%. Only 5 (2%) of 255 susceptible isolates belonged to these STs (p < .001). blaCTX-M-15 was detected in 17 (42%) of the 41 ESBL-producing isolates. Comparison with a collection of UPEC isolates obtained a decade earlier from the same community showed that a large proportion (60% and 25%, respectively) of the increase in CA-UTI caused by fluoroquinolone-resistant and ESBL-producing UPEC appears to be due to just two pandemic lineages ST131 and ST69. These findings indicate that much of the prevalence of broad-spectrum drug-resistant CA-UTI in Rio de Janeiro is due to a limited set of pandemic lineages of UPEC circulating in the community instead of multiple genotypes selected by antimicrobial agents.

Biofilm formation, antimicrobial susceptibility and virulence genes of Uropathogenic Escherichia coli isolated from clinical isolates in Uganda.

INTRODUCTION: Uropathogenic E. coli is the leading cause of Urinary tract infections (UTIs), contributing to 80-90% of all community-acquired and 30-50% of all hospital-acquired UTIs. Biofilm forming Uropathogenic E. coli are associated with persistent and chronic inflammation leading to complicated and or recurrent UTIs. Biofilms provide an environment for poor antibiotic penetration and horizontal transfer of virulence genes which favors the development of Multidrug-resistant organisms (MDRO). Understanding biofilm formation and antimicrobial resistance determinants of Uropathogenic E. coli strains will provide insight into the development of treatment options for biofilm-associated UTIs. The aim of this study was to determine the biofilm forming capability, presence of virulence genes and antimicrobial susceptibility pattern of Uropathogenic E. coli isolates in Uganda. METHODS: This was a cross-sectional study carried in the Clinical Microbiology and Molecular biology laboratories at the Department of Medical Microbiology, Makerere University College of Health Sciences. We randomly selected 200 Uropathogenic E. coli clinical isolates among the stored isolates collected between January 2018 and December 2018 that had significant bacteriuria (> 105 CFU). All isolates were subjected to biofilm detection using the Congo Red Agar method and Antimicrobial susceptibility testing was performed using the Kirby disk diffusion method. The isolates were later subjected PCR for the detection of Urovirulence genes namely; Pap, Fim, Sfa, Afa, Hly and Cnf, using commercially designed primers. RESULTS: In this study, 62.5% (125/200) were positive biofilm formers and 78% (156/200) of these were multi-drug resistant (MDR). The isolates were most resistant to Trimethoprim sulphamethoxazole and Amoxicillin (93%) followed by gentamycin (87%) and the least was imipenem (0.5%). Fim was the most prevalent Urovirulence gene (53.5%) followed by Pap (21%), Sfa (13%), Afa (8%), Cnf (5.5%) and Hyl (0%). CONCLUSIONS: We demonstrate a high prevalence of biofilm-forming Uropathogenic E. coli strains that are highly associated with the MDR phenotype. We recommend routine surveillance of antimicrobial resistance and biofilm formation to understand the antibiotics suitable in the management of biofilm-associated UTIs.

PasT of Escherichia coli sustains antibiotic tolerance and aerobic respiration as a bacterial homolog of mitochondrial Coq10.

Antibiotic-tolerant persisters are often implicated in treatment failure of chronic and relapsing bacterial infections, but the underlying molecular mechanisms have remained elusive. Controversies revolve around the relative contribution of specific genetic switches called toxin-antitoxin (TA) modules and global modulation of cellular core functions such as slow growth. Previous studies on uropathogenic Escherichia coli observed impaired persister formation for mutants lacking the pasTI locus that had been proposed to encode a TA module. Here, we show that pasTI is not a TA module and that the supposed toxin PasT is instead the bacterial homolog of mitochondrial protein Coq10 that enables the functionality of the respiratory electron carrier ubiquinone as a "lipid chaperone." Consistently, pasTI mutants show pleiotropic phenotypes linked to defective electron transport such as decreased membrane potential and increased sensitivity to oxidative stress. We link impaired persister formation of pasTI mutants to a global distortion of cellular stress responses due to defective respiration. Remarkably, the ectopic expression of human coq10 largely complements the respiratory defects and decreased persister levels of pasTI mutants. Our work suggests that PasT/Coq10 has a central role in respiratory electron transport that is conserved from bacteria to humans and sustains bacterial tolerance to antibiotics.