Evaluation of adhesin genes and risk factors associated with urinary tract infections by drug-resistant uropathogenic Escherichia coli in North of Iran.

BACKGROUND: Uropathogenic Escherichia coli (UPEC) isolates, have a wide variety of virulence factors to promote colonization and survival in the urinary tract. This study aimed to evaluate adhesin genes, biofilm formation ability, antibiotic resistance profiles of UPEC strains, and the related risk factors in patients with UTIs caused by drug-resistant UPEC. METHODOLOGY: A total of 105 UPEC isolates were evaluated for biofilm formation using 96-well microtiter plates, the presence of adhesin genes by PCR assay and the antimicrobial susceptibility pattern using the disk diffusion method. Demographic and clinical characteristics of patients were investigated to identify predisposing factors for drug-resistant isolates. RESULTS: Out of 105 UPEC isolates, 84.8% were positive for biofilm formation. Biofilm-producing isolates exhibited a significantly higher prevalence of fimH, kpsMTII, csgA, afa/draBC, and pap adhesin genes compared to non-biofilm-producing strains (p < 0.05). The results also revealed that 52.4% of the isolates were ESBL-producing, and 84.8% were multidrug-resistant (MDR). Further analysis of antibiotic susceptibility among ESBL-producing strains showed the highest resistance rates to ampicillin, ciprofloxacin, and trimethoprim-sulfamethoxazole. Conversely, the highest susceptibility, in addition to carbapenems, was observed for fosfomycin, amikacin, cefoxitin, and nitrofurantoin. We identified hypertension as a potential risk factor for infection with ESBL-producing UPEC strains. CONCLUSIONS: Our results revealed a significant rate of drug resistance among UPEC isolates obtained from UTIs in our region. This underscores the importance of monitoring the empirical use of antibiotics and identifying specific risk factors in our geographical area to guide the selection of appropriate empirical treatment for UTIs.

Characterization of bacteriophages infecting multidrug-resistant uropathogenic Escherichia coli strains.

Uropathogenic Escherichia coli (UPEC) is the most common causative agent of urinary tract infections, and strains that are resistant to antibiotics are a major problem in treating these infections. Phage therapy is a promising alternative approach that can be used to treat infections caused by polyresistant bacterial strains. In the present study, 16 bacteriophages isolated from sewage and surface water were investigated. Phage host specificity was tested on a collection of 77 UPEC strains. The phages infected 2-44 strains, and 80% of the strains were infected by at least one phage. The susceptible E. coli strains belonged predominantly to the B2 phylogenetic group, including strains of two clones, CC131 and CC73, that have a worldwide distribution. All of the phages belonged to class Caudoviricetes and were identified as members of the families Straboviridae, Autographiviridae, and Drexlerviridae and the genera Kagunavirus, Justusliebigvirus, and Murrayvirus. A phage cocktail composed of six phages - four members of the family Straboviridae and two members of the family Autographiviridae - was prepared, and its antibacterial activity was tested in liquid medium. Complete suppression of bacterial growth was observed after 5-22 hours of cultivation, followed by partial regrowth. At 24 hours postinfection, the cocktail suppressed bacterial growth to 43-92% of control values. Similar results were obtained when testing the activity of the phage cocktail in LB and in artificial urine medium. The results indicate that our phage cocktail has potential to inhibit bacterial growth during infection, and they will therefore be preserved in the national phage bank, serving as valuable resources for therapeutic applications.

Multidrug-resistant Escherichia coli causing canine pyometra and urinary tract infections are genetically related but distinct from those causing prostatic abscesses.

Despite extensive characterisation of uropathogenic Escherichia coli (UPEC) causing urinary tract infections (UTIs), the genetic background of non-urinary extraintestinal pathogenic E. coli (ExPEC) in companion animals remains inadequately understood. In this study, we characterised virulence traits of 104 E. coli isolated from canine pyometra (n = 61) and prostatic abscesses (PAs) (n = 38), and bloodstream infections (BSIs) in dogs (n = 2), and cats (n = 3). A stronger association with UPEC of pyometra strains in comparison to PA strains was revealed. Notably, 44 isolates exhibited resistance to third-generation cephalosporins and/or fluoroquinolones, 15 were extended-spectrum ß-lactamase-producers. Twelve multidrug-resistant (MDR) strains, isolated from pyometra (n = 4), PAs (n = 5), and BSIs (n = 3), along with 7 previously characterised UPEC strains from dogs and cats, were sequenced. Genomic characteristics revealed that MDR E. coli associated with UTIs, pyometra, and BSIs belonged to international high-risk E. coli clones, including sequence type (ST) 38, ST131, ST617, ST648, and ST1193. However, PA strains belonged to distinct lineages, including ST12, ST44, ST457, ST744, and ST13037. The coreSNPs, cgMLST, and pan-genome illustrated intra-clonal variations within the same ST from different sources. The high-risk ST131 and ST1193 (phylogroup B2) contained high numbers of ExPEC virulence genes on pathogenicity islands, predominating in pyometra and UTI. Hybrid MDR/virulence IncF multi-replicon plasmids, containing aerobactin genes, were commonly found in non-B2 phylogroups from all sources. These findings offer genomic insights into non-urinary ExPEC, highlighting its potential for invasive infections in pets beyond UTIs, particularly with regards to high-risk global clones.

Peptidoglycan endopeptidase MepM of uropathogenic Escherichia coli contributes to competitive fitness during urinary tract infections.

BACKGROUND: Urinary tract infections (UTIs) are common bacterial infections, primarily caused by uropathogenic Escherichia coli (UPEC), leading to significant health issues and economic burden. Although antibiotics have been effective in treating UPEC infections, the rise of antibiotic-resistant strains hinders their efficacy. Hence, identifying novel bacterial targets for new antimicrobial approaches is crucial. Bacterial factors required for maintaining the full virulence of UPEC are the potential target. MepM, an endopeptidase in E. coli, is involved in the biogenesis of peptidoglycan, a major structure of bacterial envelope. Given that the bacterial envelope confronts the hostile host environment during infections, MepM's function could be crucial for UPEC's virulence. This study aims to explore the role of MepM in UPEC pathogenesis. RESULTS: MepM deficiency significantly impacted UPEC's survival in urine and within macrophages. Moreover, the deficiency hindered the bacillary-to-filamentous shape switch which is known for aiding UPEC in evading phagocytosis during infections. Additionally, UPEC motility was downregulated due to MepM deficiency. As a result, the mepM mutant displayed notably reduced fitness in causing UTIs in the mouse model compared to wild-type UPEC. CONCLUSIONS: This study provides the first evidence of the vital role of peptidoglycan endopeptidase MepM in UPEC's full virulence for causing UTIs. MepM's contribution to UPEC pathogenesis may stem from its critical role in maintaining the ability to resist urine- and immune cell-mediated killing, facilitating the morphological switch, and sustaining motility. Thus, MepM is a promising candidate target for novel antimicrobial strategies.

Whole genome sequencing of uropathogenic E. coli from Ireland reveals diverse resistance mechanisms and strong correlation with phenotypic (EUCAST) susceptibility testing.

Urinary tract infections (UTI) caused by uropathogenic Escherichia coli (UPEC) pose a global health concern. Resistance mechanisms, including genetic mutations in antimicrobial target genes, efflux pumps, and drug deactivating enzymes, hinder clinical treatment. These resistance factors often spread through mobile genetic elements. Molecular techniques like whole genome sequencing (WGS), multilocus sequence typing (MLST), and phylotyping help decode bacterial genomes and categorise resistance genes. In this study, we analysed 57 UPEC isolates from different UTI patients following EUCAST guidelines. A selection of 17 representative strains underwent WGS, phylotyping, MLST, and comparative analysis to connect laboratory susceptibility data with predictive genomics based on key resistance genes and chromosomal mutations in antimicrobial targets. Trimethoprim resistance consistently correlated with dfr genes, with six different alleles detected among the isolates. These dfr genes often coexisted with class 1 integrons, with the most common gene cassette combining dfr and aadA. Furthermore, 52.9% of isolates harboured the blaTem-1 gene, rendering resistance to ampicillin and amoxicillin. Ciprofloxacin-resistant strains exhibited mutations in GyrA, GyrB and ParC, plasmid-mediated quinolone resistance genes (qnrb10), and aac(6')-Ib-cr5. Nitrofurantoin resistance in one isolate stemmed from a four amino acid deletion in NfsB. These findings illustrate the varied strategies employed by UPEC to resist antibiotics and the correlation between clinical susceptibility testing and molecular determinants. As molecular testing gains prominence in clinical applications, understanding key resistance determinants becomes crucial for accurate susceptibility testing and guiding effective antimicrobial therapy.

Competitive fitness of asymptomatic bacteriuria E. coli strain 83972 against uropathogens in human urine.

Urinary tract infection (UTI) is one of the most common bacterial infections worldwide. The main causative agent of UTI is uropathogenic Escherichia coli (UPEC). There is an immediate need for novel prophylactic and treatment strategies against UTI because of the increasing incidence of antimicrobial resistance among uropathogens. ABU 83972, an asymptomatic bacteriuria-causing E. coli strain, prevents UTI by suppressing the colonization of UPEC. However, the nature of competition and growth repression of UPEC by ABU 83972 is unclear and is the subject of our investigation. Here, we characterized the growth kinetics of ABU 83972 and uropathogens in human urine and laboratory media. Next, we performed a series of competitive co-culture experiments where ABU 83972 and uropathogens were inoculated at a 1:1 ratio in human urine and in various media, and their relative abundance was determined. In human urine, ABU 83972 outcompeted UPEC and additional uropathogens, reaching up to 90% of the total population after 24 hours of incubation. In contrast, UPEC outcompeted ABU 83972 in LB and M9 minimal media and exhibited superior colonization than ABU 83972 in the mouse urinary bladder. Since engineered living materials (ELMs) can be used to retain an organism of interest in a particular location, we developed ABU 83972-containing ELMs that effectively outcompeted UPEC in human urine. In summary, our work establishes that ABU 83972 outcompetes UPEC in a milieu- and cell-density-dependent manner, highlighting the importance of the metabolites and nutrients found in the human urine as determinants of the competitive fitness of ABU 83972.

Turbidimetric bioassays: A solution to antimicrobial activity detection in asymptomatic bacteriuria isolates against uropathogenic Escherichia coli.

Traditional bacteriocin screening methods often face limitations due to diffusion-related challenges in agar matrices, which can prevent the peptides from reaching their target organism. Turbidimetric techniques offer a solution to these issues, eliminating diffusion-related problems and providing an initial quantification of bacteriocin efficacy in producer organisms. This study involved screening the cell-free supernatant (CFS) from eight uncharacterized asymptomatic bacteriuria (ABU) isolates and Escherichia coli 83972 for antimicrobial activity against clinical uropathogenic E. coli (UPEC) strains using turbidimetric growth methods. ABU isolates exhibiting activity against five or more UPEC strains were further characterized (PUTS 37, PUTS 58, PUTS 59, S-07-4, and SK-106-1). The inhibition of the CFS by proteinase K suggested that the antimicrobial activity was proteinaceous in nature, potentially bacteriocins. The activity of E. coli PUTS 58 and SK-106-1 was enhanced in an artificial urine medium, with both inhibiting all eight UPECs. A putative microcin H47 operon was identified in E. coli SK-106-1, along with a previously identified microcin V and colicin E7 in E. coli PUTS 37 and PUTS 58, respectively. These findings indicate that ABU bacteriocin-producers could serve as viable prophylactics and therapeutics in the face of increasing antibiotic resistance among uropathogens.

Gentamicin loaded niosomes against intracellular uropathogenic Escherichia coli strains.

Urinary tract infections (UTIs) are the most common bacterial infections and uropathogenic Escherichia coli (UPEC) is the main etiological agent of UTIs. UPEC can persist in bladder cells protected by immunological defenses and antibiotics and intracellular behavior leads to difficulty in eradicating the infection. The aim of this paper is to design, prepare and characterize surfactant-based nanocarriers (niosomes) able to entrap antimicrobial drug and potentially to delivery and release antibiotics into UPEC-infected cells. In order to validate the proposed drug delivery system, gentamicin, was chosen as "active model drug" due to its poor cellular penetration. The niosomes physical-chemical characterization was performed combining different techniques: Dynamic Light Scattering Fluorescence Spectroscopy, Transmission Electron Microscopy. Empty and loaded niosomes were characterized in terms of size, ζ-potential, bilayer features and stability. Moreover, Gentamicin entrapped amount was evaluated, and the release study was also carried out. In addition, the effect of empty and loaded niosomes was studied on the invasion ability of UPEC strains in T24 bladder cell monolayers by Gentamicin Protection Assay and Confocal Microscopy. The observed decrease in UPEC invasion rate leads us to hypothesize a release of antibiotic from niosomes inside the cells. The optimization of the proposed drug delivery system could represent a promising strategy to significatively enhance the internalization of antimicrobial drugs.

Cranberry Polyphenols and Prevention against Urinary Tract Infections: New Findings Related to the Integrity and Functionality of Intestinal and Urinary Barriers.

This work seeks to generate new knowledge about the mechanisms underlying the protective effects of cranberry against urinary tract infections (UTI). Using Caco-2 cells grown in Transwell inserts as an intestinal barrier model, we found that a cranberry-derived digestive fluid (containing 135 ± 5 mg of phenolic compounds/L) increased transepithelial electrical resistance with respect to control (ΔTEER = 54.5 Ω cm2) and decreased FITC-dextran paracellular transport by about 30%, which was related to the upregulation of the gene expression of tight junction (TJ) proteins (i.e., occludin, zonula occludens-1 [ZO-1], and claudin-2) (∼3-4-fold change with respect to control for claudin-2 and ∼2-3-fold for occludin and ZO-1). Similar protective effects, albeit to a lesser extent, were observed when Caco-2 cells were previously infected with uropathogenic Escherichia coli (UPEC). In a urinary barrier model comprising T24 cells grown in Transwell inserts and either noninfected or UPEC-infected, treatments with the cranberry-derived phenolic metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and phenylacetic acid (PAA) (250 µM) also promoted favorable changes in barrier integrity and permeability. In this line, incubation of noninfected T24 cells with these metabolites induced positive regulatory effects on claudin-2 and ZO-1 expression (∼3.5- and ∼2-fold change with respect to control for DOPAC and ∼1.5- and >2-fold change with respect to control for PAA, respectively). Overall, these results suggest that the protective action of cranberry polyphenols against UTI might involve molecular mechanisms related to the integrity and functionality of the urothelium and intestinal epithelium.

Plant phenolics inhibit focal adhesion kinase and suppress host cell invasion by uropathogenic Escherichia coli.

Traditional folk treatments for the prevention and management of urinary tract infections (UTIs) and other infectious diseases often include plants and plant extracts that are rich in phenolic compounds. These have been ascribed a variety of activities, including inhibition of bacterial interactions with host cells. Here, we tested a panel of four well-studied phenolic compounds-caffeic acid phenethyl ester (CAPE), resveratrol, catechin, and epigallocatechin gallate-for the effects on host cell adherence and invasion by uropathogenic Escherichia coli (UPEC). These bacteria, which are the leading cause of UTIs, can bind and subsequently invade bladder epithelial cells via an actin-dependent process. Intracellular UPEC reservoirs within the bladder are often protected from antibiotics and host defenses and likely contribute to the development of chronic and recurrent infections. In cell culture-based assays, only resveratrol had a notable negative effect on UPEC adherence to bladder cells. However, both CAPE and resveratrol significantly inhibited UPEC entry into the host cells, coordinate with attenuated phosphorylation of the host actin regulator Focal Adhesion Kinase (FAK or PTK2) and marked increases in the numbers of focal adhesion structures. We further show that the intravesical delivery of resveratrol inhibits UPEC infiltration of the bladder mucosa in a murine UTI model and that resveratrol and CAPE can disrupt the ability of other invasive pathogens to enter host cells. Together, these results highlight the therapeutic potential of molecules like CAPE and resveratrol, which could be used to augment antibiotic treatments by restricting pathogen access to protective intracellular niches.IMPORTANCEUrinary tract infections (UTIs) are exceptionally common and increasingly difficult to treat due to the ongoing rise and spread of antibiotic-resistant pathogens. Furthermore, the primary cause of UTIs, uropathogenic Escherichia coli (UPEC), can avoid antibiotic exposure and many host defenses by invading the epithelial cells that line the bladder surface. Here, we identified two plant-derived phenolic compounds that disrupt activation of the host machinery needed for UPEC entry into bladder cells. One of these compounds, resveratrol, effectively inhibited UPEC invasion of the bladder mucosa in a mouse UTI model, and both phenolic compounds significantly reduced host cell entry by other invasive pathogens. These findings suggest that select phenolic compounds could be used to supplement existing antibacterial therapeutics by denying uropathogens shelter within host cells and tissues and help explain some of the benefits attributed to traditional plant-based medicines.

Multi-step strategies for synergistic treatment of urinary tract infections based on D-xylose-decorated antimicrobial peptide carbon dots.

Urinary tract infections (UTIs) caused by Uropathogenic Escherichia coli (UPEC), often reoccur due to the formation of intracellular bacterial colonies (IBCs) and antibiotic resistance. Given the significance of YadC for UPEC infection in our previous study, we developed D-xylose-decorated ɛ-poly-L-lysine (εPL)-based carbon dots (D-xyl@εPLCDs) that can be traced, and employed multi-step approaches to elucidate the functional roles of D-xyl@εPLCDs in UPEC infection. Compared to undecorated particles, D-xyl@εPLCDs demonstrate YadC-dependent bacterial targeting and exhibit enhanced bactericidal activities both intracellularly and extracellularly. Moreover, pre-treatment of D-xyl@εPLCDs before infection blocked the subsequent adhesion and invasion of UPEC to bladder epithelial cells 5637. Increase of ROS production and innate immune responses were observed in bladder epithelial cells 5637 treated with D-xyl@εPLCDs. In addition, treatment of D-xyl@εPLCDs post-infection facilitated clearance of UPEC in the bladders of the UTI mouse model, and reduced ultimate number of neutrophils, macrophages and inflammatory responses raised by invaded bacteria. Collectively, we presented a comprehensive evaluating system to show that D-xyl@εPLCDs exhibits superior bactericidal effects against UPEC, making them a promising candidate for drug development in clinical UTI therapeutics.

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.

Bacteriological and molecular study of fosfomycin resistance in uropathogenic Escherichia coli.

The identification of genes associated with resistance has the potential to facilitate the development of novel diagnostic tests and treatment methods. The objective of this study was to examine the antibiotic resistance and Fosfomycin resistance genes in uropathogenic Escherichia coli (UPEC) in patients in Baghdad, Iraq. After analyzing 250 urine samples using various identification methods, including the examination of morphological characteristics, biochemical tests, and genetic detection, it was determined that E. coli was the most common bacteria present, accounting for 63.6% of the samples. Antibiotic susceptibility testing showed a significant prevalence of resistance to various antibiotics, with 99.3% of E. coli isolates exhibiting multiple drug resistance (MDR). Fosfomycin showed antibacterial properties against UPEC. The minimum inhibitory concentration (MIC) ranged from 512 to 1024 µg/mL, while the minimum bactericidal concentration (MBC) was 2048 µg/mL. In the time-kill assay, fosfomycin was effective against fosfomycin-resistant isolates within 8-12 h. The genetic determinants associated with fosfomycin resistance were examined through the utilization of polymerase chain reaction (PCR). The findings indicated that the genes murA, glpT, and cyaA were detected in all the isolates when genomic DNA was used as a template. However, all the tests yielded negative results when plasmid was used as a template. The genes fosA3 and fosA4 were detected in 8.6% and 5% of the isolates when genomic DNA was used as a template. When plasmid was used as a template, the genes fosA3 and fosA4 were found in 5.7% and 2.9% of the isolates, respectively. In conclusion, there is an increasing problem with antibiotic resistance in UPEC, with elevated rates of resistance to several antibiotics. The study also offers novel insights into the genetic foundation of fosfomycin resistance in UPEC.

Anti-biofilm activity of caffeine against uropathogenic E. coli is mediated by curli biogenesis.

Biofilms are assemblages of sessile microorganisms that form an extracellular matrix around themselves and mediate attachment to surfaces. The major component of the extracellular matrix of Uropathogenic E. coli and other Enterobacteriaceae are curli fibers, making biofilms robust and resistant to antimicrobials. It is therefore imperative to screen antibiofilm compounds that can impair biofilm formation. In the present study, we investigated the curli-dependent antibiofilm activity of caffeine against UPEC strain CFT073 and commensal strain E. coli K-12MG1655.Caffeine significantly reduced the biofilm formation of both UPEC and E. coli K-12 by 86.58% and 91.80% respectively at 48 mM caffeine as determined by Crystal Violet assay. These results were further confirmed by fluorescence microscopy and Scanning Electron Microscope (SEM). Caffeine significantly reduced the cytotoxicity and survivability of UPEC. Molecular docking analysis revealed a strong interaction between caffeine and curli regulator protein (Csg D) of E. coli. The qRT-PCR data also showed significant downregulation in the expression of CsgBA and the CsgDEFG operon at both 24 mM and 48 mM caffeine. The findings revealed that caffeine could inhibit E. coli biofilm formation by regulating curli assembly and thus may be used as an alternative therapeutic strategy for the treatment of chronic E. coli biofilm-related infections.

Comprehensive analysis of PNA-based antisense antibiotics targeting various essential genes in uropathogenic Escherichia coli.

Antisense peptide nucleic acids (PNAs) that target mRNAs of essential bacterial genes exhibit specific bactericidal effects in several microbial species, but our mechanistic understanding of PNA activity and their target gene spectrum is limited. Here, we present a systematic analysis of PNAs targeting 11 essential genes with varying expression levels in uropathogenic Escherichia coli (UPEC). We demonstrate that UPEC is susceptible to killing by peptide-conjugated PNAs, especially when targeting the widely-used essential gene acpP. Our evaluation yields three additional promising target mRNAs for effective growth inhibition, i.e.dnaB, ftsZ and rpsH. The analysis also shows that transcript abundance does not predict target vulnerability and that PNA-mediated growth inhibition is not universally associated with target mRNA depletion. Global transcriptomic analyses further reveal PNA sequence-dependent but also -independent responses, including the induction of envelope stress response pathways. Importantly, we show that 9mer PNAs are generally as effective in inhibiting bacterial growth as their 10mer counterparts. Overall, our systematic comparison of a range of PNAs targeting mRNAs of different essential genes in UPEC suggests important features for PNA design, reveals a general bacterial response to PNA conjugates and establishes the feasibility of using PNA antibacterials to combat UPEC.

Phenotypic and Genotypic Characteristics of Uropathogenic Escherichia coli Isolates from Kenya.

Introduction: Uropathogenic Escherichia coli (UPECs) are a significant cause of urinary tract infections (UTIs). In Kenya, UTIs are typically treated with ß-lactam antibiotics without antibiotic susceptibility testing, which could accelerate antibiotic resistance among UPEC strains. Aim: This study determined the occurrence of UPEC producing extended-spectrum ß-lactamases (ESBLs), the genes conferring resistance to ß-lactams, and the phylogenetic groups associated with ESBLs in Kenyan UPECs. Methodology: Ninety-five UPEC isolates from six Kenyan hospitals were tested for ESBL and plasmid-mediated AmpC ß-lactamase (pAmpC) production by combined disk diffusion and disk approximation tests, respectively. Real-time and conventional polymerase chain reactions (PCRs) were used to detect three ESBL and six pAmpC genes, respectively, and phylogenetic groups were assigned by a quadruplex PCR method. Results: Twenty-four percent UPEC isolates were ESBL producers with blaCTX-M (95.6%), blaTEM (95.6%), and blaSHV (21.7%) genes detected. Sixteen isolates had blaCTX-M/TEM, whereas five had blaTEM/CTX-M/SHV. A total of 5/23 ESBLs were cefoxitin resistant, but no AmpC genes were detected. The UPECs belonged predominantly to phylogenetic groups B2 (31/95; 32.6%) and D (30/95; 31.6%), while groups B2 and A had the most ESBL producers. Conclusions: ß-Lactam antibiotics have reduced utility for treating UTIs as a quarter of UPECs were ESBL producing. Single or multiple ESBL genes were present in UPECs, belonging primarily to phylogenetic groups B2 and A.

Differences in recipient ability of uropathogenic Escherichia coli strains in relation with their pathogenic potential.

Conjugation is recognized as a mechanism driving dissemination of antibacterial resistances and virulence factors among bacteria. In the presented work conjugative transfer frequency into clinical uropathogenic Escherichia coli strains (UPEC) isolated from patients with symptomatic urinary tract infections was investigated. From 93 obtained UPEC strains only 29 were suitable for conjugation experiments with the plasmid pOX38, a well-known F-plasmid derivative. The study was focused on comparison of conjugation frequencies in plankton and biofilm, including comparison of conjugation frequencies in high and low biofilm biomass with their virulence potential. It was shown that the conjugation frequency depended on the biofilm biomass and was significantly higher in thin (OD580 < 0.3) than in thick biofilm (OD580 ≥ 0.3). Nonmetric multidimensional scaling analysis revealed that higher conjugation frequencies in plankton and biofilm were directly positively correlated with the sum of virulence-associated genes of the recipient strain and presence of multidrug antibiotic resistances. On the other hand, the sum of insensitivities to different bacteriocins was negatively correlated with an increase in the conjugative transfer level. Our results obtained hence indicate that the evolution of potentially more pathogenic strains via conjugation is depended on the strains' ability to be a "good" recipient in the conjugative transfer, possibly due to the ability to form thinner biofilms.

Are Uropathogenic Bacteria Living in Multispecies Biofilm Susceptible to Active Plant Ingredient-Asiatic Acid?

Urinary tract infections (UTIs) are a serious health problem in the human population due to their chronic and recurrent nature. Bacteria causing UTIs form multispecies biofilms being resistant to the activity of the conventionally used antibiotics. Therefore, compounds of plant origin are currently being searched for, which could constitute an alternative strategy to antibiotic therapy. Our study aimed to determine the activity of asiatic acid (AA) against biofilms formed by uropathogenic Escherichia coli, Enterobacter cloacae, and Pseudomonas aeruginosa. The influence of AA on the survival, biofilm mass formation by bacteria living in mono-, dual-, and triple-species consortia as well as the metabolic activity and bacterial cell morphology were determined. The spectrophotometric methods were used for biofilm mass synthesis and metabolic activity determination. The survival of bacteria was established using the serial dilution assay. The decrease in survival and a weakening of the ability to create biofilms, both single and multi-species, as well as changes in the morphology of bacterial cells were noticed. As AA works best against young biofilms, the use of AA-containing formulations, especially during the initial stages of infection, seems to be reasonable. However, there is a need for further research concerning AA especially regarding its antibacterial mechanisms of action.

Resistencia antimicrobiana de uropatógenos en adultos mayores / Antimicrobial resistance of uropathogens in older adults

Introducción: En la actualidad, la resistencia antimicrobiana ha sido declarada por la Organización Mundial de la Salud como un problema de salud pública. Objetivo: Determinar el perfil de resistencia antimicrobiana de uropatógenos en adultos mayores. Métodos: Estudio descriptivo transversal de 567 urocultivos positivos de adultos mayores atendidos durante el año 2017 en una clínica privada en Lima, Perú. El análisis univariado se realizó por distribución de frecuencias, promedio y desviación estándar. Se estimó la asociación entre la producción de BLEE con respecto a las características epidemiológicas y el tipo de atención mediante Chi cuadrado con un nivel de significación de 0,05. Resultados: La edad promedio de la población fue de 74,1 años (DE:10,7). El 71,8 por ciento de los urocultivos positivos pertenecieron al sexo femenino. Los principales uropatógenos aislados en todos los niveles de atención fueron: E. coli, E. coli BLEE y K. pneumoniae BLEE. La E. coli presentó 69,3 por ciento de resistencia a ampicilina; y la E. coli BLEE tuvo el 100 por ciento de resistencia a ampicilina, ceftriaxona y ceftazidima. El 62 por ciento de microorganismos BLEE se encontraron en la atención ambulatoria. Se evidenció asociación estadísticamente significativa entre los agentes productores de BLEE y el sexo (p=0,004), mas no respecto al tipo de atención (p=0,144) ni subgrupos de edad (p=0,669). Conclusiones: La resistencia antimicrobiana es altamente prevalente en los adultos mayores. El sexo femenino fue el más afectado y el uropatógeno más frecuente la E. coli, este presenta una alta resistencia a ampicilina y mayor sensibilidad a nitrofurantoína. Se determinó un alto porcentaje de agentes productores de BLEE en la atención ambulatoria(AU)

Introduction: At present, the World Health Organization as a public health problem has declared antimicrobial resistance. Objective: To determine the antimicrobial resistance profile of uropathogens in older adults. Methods: Cross-sectional descriptive study of 567 positive urine cultures from older adults treated during 2017 in a private clinic in Lima, Peru. Univariate analysis was performed by frequency distribution, mean and standard deviation. The association between ESBL production with respect to epidemiological characteristics and type of care is estimated using Chi square with a significance level of 0.05. Results: The average age of the population was 74.1 years (SD: 10.7). 71.8percent of the positive urine cultures were from female sex. The main uropathogens isolated at all levels of care were E. coli, E. coli ESBL and K. pneumoniae ESBL. E. coli showed 69.3percent resistance to ampicillin; and E. coli ESBL had 100percent resistance to ampicillin, ceftriaxone, and ceftazidime. 62percent of ESBL microorganisms were found in outpatient care. There was a statistically significant association between ESBL-producing agents and gender (p = 0.004), but not with regard to type of care (p = 0.144) or age subgroups (p = 0.669). Conclusions: Antimicrobial resistance is highly prevalent in older adults. The female sex was the most affected and the most frequent uropathogen was E. coli, highly resistant to ampicillin and greater sensitivity to nitrofurantoin. High percentage of ESBL-producing agents was determined in outpatient care(AU)

Differences of virulence factors, and antimicrobial susceptibility according to phylogenetic group in uropathogenic Escherichia coli strains isolated from Korean patients.

BACKGROUND: Escherichia coli is among the most common uropathogens. Increased antibiotic resistance in Gram negative bacilli is global concern. Alternative therapeutic options including vaccines against uropathogenic E. coli (UPEC) have been developed. In this study, we compared the genotypic characteristics and antimicrobial susceptibility of UPEC according to phylogenetic groups. METHODS: We retrospectively reviewed the medical records of pyelonephritis patients with UPEC between February 2015 and June 2018. The study was conducted at a medical center in Korea. We compared the clinical and genotypic characteristics of UPEC according to phylogenetic groups. The phylogenetic groups and 29 virulence factors were identified using multiplex polymerase chain reaction. RESULTS: Phylogenetic group analysis revealed that most uropathogenic E. coli belonged to groups B2 and D: B2 (276, 77.7%), D (62, 17.5%), B1 (12, 3.4%), and A (5, 1.4%). Among the virulence factors, fyuA, fimH, traT, iutA, papG allele II, and papC were the most frequently observed. Phylogenetic group B2 was more closely related to virulence factors, including fimH, sfa/focED, focG, hlyA, cnf1, fyuA, and PAI, than group D. Groups B2 and D showed similar clinical presentations and complications. Group B2 had mostly healthcare-associated infections and antimicrobial resistance. Group D mostly had community-acquired infections. The K1 serotype was prevalent in group B2, and K5 was the most prevalent in group D. CONCLUSIONS: Phylogenetic group B2 had more proportions and types of virulence factors than group D. Group B2 showed a high presentation of virulence factors related to adhesions and toxins. An increased presentation of antimicrobial resistance and healthcare-associated infections was also noted. Considering the genetic characteristics of UPEC, alternative therapeutic options targeting frequent virulence factors might be considered in addition to antibiotics.