Investigating the interaction of zno nanoparticles with flagellum and fimbriae in multi-drug resistant uropathogenic bacteria encoding fli and fim genes.

Due to the increasing occurrence of drug resistant urinary tract infections (UTI) among children, there is a need to investigate alternative effective treatment protocols such as nanoparticles. Flagella and fimbriae are primary factors contributing the virulence of urinary tract infecting bacteria. The aim of this study was to assess the antibacterial effects of zinc oxide nanoparticles which have been synthesized using both chemical and green methods on multi-drug resistant (MDR) uropathogenic bacteria encoding fli and fim genes and investigating their binding ability to bacterial appendage proteins. A total of 30 urine culture samples were collected from children under 2 years old diagnosed with urinary tract infection. The isolates underwent antibiotic suseptibility assessment and the isolates demonstrating MDR were subjected to molecular amplification of fimG (fimbrial) and fliD and fliT (flagellal) genes. The confirmation of cellular appendages was achieved through silver nitrate staining. The antibacterial efficacy of the synthetized nanoparticles was assessed using the micro and macrodilution methods. The successful binding of nanoparticles to bacterial appendage proteins was confirmed through mobility shift and membrane filter assays. The dimensions of chemically synthesized ZnO nanoparticles and green nanoparticles were measured at 30 nm and 85 nm, respectively, with the exhibition of hexagonal geometries. The nanoparticles synthesized through chemical and green methods exhibited minimum inhibitory concentrations (MIC) of 0.0062-0.025 g/L and 0.3 g/L, respectively. The ability of ZnO nanoparticles to bind bacterial appendage proteins and to combat MDR uropathogenic bacteria are promising for new treatment protocols against UTI in children in future.

Draft genome of three uropathogenic Escherichia coli strains.

Uropathogenic Escherichia coli (UPEC) remains the main etiological agent of urinary tract infections affecting females and males. The draft genome sequence of three strains of UPEC isolated from senior citizens and pregnant women in the state of Puebla, Mexico, is reported here.

Draft genome sequence of uropathogenic Escherichia coli U13824, a multidrug-resistant (MDR) and extended-spectrum-ß-lactamase (ESBL)-producing UPEC strain isolated from an adult woman with urinary tract infection.

Urinary tract infections (UTIs) caused by multidrug-resistant and extended-spectrum ß-lactamase-producing uropathogenic Escherichia coli are a worldwide concern. We report the draft genome of E. coli U13824 isolated from a female outpatient with UTI. This genome's availability strengthens the genomic surveillance of antimicrobial resistance and the spreading of these strains.

A novel chaperone-effector-immunity system identified in uropathogenic Escherichia coli UMN026.

Background: Urinary tract infections (UTIs) are very common worldwide. According to their symptomatology, these infections are classified as pyelonephritis, cystitis, or asymptomatic bacteriuria (AB). Approximately 75-95% of UTIs are caused by uropathogenic Escherichia coli (UPEC), which is an extraintestinal bacterium that possesses virulence factors for bacterial adherence and invasion in the urinary tract. In addition, UPEC possesses type 6 secretion systems (T6SS) as virulence mechanisms that can participate in bacterial competition and in bacterial pathogenicity. UPEC UMN026 carries three genes, namely, ECUMN_0231, ECUMN_0232, and ECUMN_0233, which encode three uncharacterized proteins related to the T6SS that are conserved in strains from phylogroups B2 and D and have been proposed as biomarkers of UTIs. Aim: To analyze the frequency of the ECUMN_0231, ECUMN_0232, ECUMN_0233, and vgrG genes in UTI isolates, as well as their expression in Luria Bertani (LB) medium and urine; to determine whether these genes are related to UTI symptoms or bacterial competence and to identify functional domains on the putative proteins. Methods: The frequency of the ECUMN and vgrG genes in 99 clinical isolates from UPEC was determined by endpoint PCR. The relationship between gene presence and UTI symptomatology was determined using the chi2 test, with p < 0.05 considered to indicate statistical significance. The expression of the three ECUMN genes and vgrG was analyzed by RT-PCR. The antibacterial activity of strain UMN026 was determined by bacterial competence assays. The identification of functional domains and the docking were performed using bioinformatic tools. Results: The ECUMN genes are conserved in 33.3% of clinical isolates from patients with symptomatic and asymptomatic UTIs and have no relationship with UTI symptomatology. Of the ECUMN+ isolates, only five (15.15%, 5/33) had the three ECUMN and vgrG genes. These genes were expressed in LB broth and urine in UPEC UMN026 but not in all the clinical isolates. Strain UMN026 had antibacterial activity against UPEC clinical isolate 4014 (ECUMN-) and E. faecalis but not against isolate 4012 (ECUMN+). Bioinformatics analysis suggested that the ECUMN genes encode a chaperone/effector/immunity system. Conclusions: The ECUMN genes are conserved in clinical isolates from symptomatic and asymptomatic patients and are not related to UTI symptoms. However, these genes encode a putative chaperone/effector/immunity system that seems to be involved in the antibacterial activity of strain UMN026.

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.

Chitosan-Coated Silver Nanoparticles Inhibit Adherence and Biofilm Formation of Uropathogenic Escherichia coli.

Urinary tract infections are commonly caused by uropathogenic Escherichia coli (UPEC), which usually presents multiple virulence and resistance mechanisms, making it difficult to treat. It has been demonstrated that silver and polymeric nanoparticles had potential against these pathogens. In this study, we synthesized thiol chitosan-coated silver nanoparticles (SH-Cs-AgNPs) and evaluated their antibacterial, antibiofilm and antiadherence activity against clinical isolates of UPEC. The SH-Cs-AgNPs showed a spherical shape with a size of 17.80 ± 2.67 nm and zeta potential of 18 ± 2 mV. We observed a potent antibacterial and antibiofilm activity as low as 12.5 µg/mL, as well as a reduction in the adherence of UPEC to mammalian cells at concentrations of 1.06 and 0.53 µg/mL. These findings demonstrate that SH-Cs-AgNPs have potential as a new therapeutic compound against infections caused by UPEC.

Detection and characterization of potentially hybrid enteroaggregative Escherichia coli (EAEC) strains isolated from urinary tract infection.

Uropathogenic Escherichia coli (UPEC) have the potential to receive the virulence markers of intestinal pathotypes and transform into various important hybrid pathotypes. This study aimed to investigate the frequency and characteristics of hybrid enteroaggregative E. coli (EAEC)/UPEC strains. Out of 202 UPEC strains, nine (4.5%) were detected as hybrid EAEC/UPEC. These strains carried one to four iron uptake systems. Among nine investigated pathogenicity islands (PAIs), PAI IV536, PAI II536, and PAI ICFT073 were found in 9 (100%), 3 (33.3%), and 1 (11.1%) strains, respectively. The chuA and sitA genes were detected in 5 (55.5%) and 3 (33.3%) hybrid strains, respectively. Six hybrid strains were found to be typical extraintestinal pathogenic E. coli (ExPEC) according to their virulence traits. Most of the hybrid strains belonged to the phylogenetic group E (6/9). Among the hybrid strains, seven (7/9) were able to form biofilm and adhere to cells; however, only two strains penetrated into the HeLa cells. Our findings reveal some of the virulence characteristics of hybrid strains that lead to fitness and infection in the urinary tract. These strains, with virulence factors of intestinal and non-intestinal pathotypes, may become emerging pathogens in clinical settings; therefore, further studies are needed to reveal their pathogenicity mechanisms and so that preventive measures can be taken.

Phenotypic and genotypic characteristics of Escherichia coli strains isolated during a longitudinal follow-up study of chronic urinary tract infections.

Worldwide, Urinary Tract Infections (UTIs) are an important health problem with many cases reported annually, women being the most affected. UTIs are relevant because they can become a recurrent condition, associated with different factors that contribute to the chronicity of the disease (cUTI). cUTI can be classified as persistent (peUTI) when the causative agent is the same each time the infection occurs or as reinfection (reUTI) when the associated microorganism is different. The purpose of this work was to characterize Escherichia coli isolates obtained in two prospective studies of patients with cUTI, to define which of them corresponded to peUTI and which to reUTI. A total of 394 isolates of E. coli were analyzed by agglutination with specific sera, antimicrobial susceptibility by diffusion disc test, and the phylogroups and presence of genes associated with virulence by PCR assays. Additionally, in some characterized strains adherence, invasiveness, and biofilm formation were analyzed by in vitro assays. The results showed that the peUTI strains belonged mainly to the classical UPEC serogroups (O25, O75, O6), were included in the B2 phylogroup, carried a great number of virulence genes, and were adherent, invasive, and biofilm-forming. Meanwhile, reUTI strains showed great diversity of serogroups, belonged mainly in the A phylogroup, and carried fewer virulence genes. Both peUTI and reUTI strains showed extensively drug-resistant (XDR) and multidrug-resistant (MDR) profiles in the antimicrobial susceptibility test. In conclusion, it appears that peUTIs are caused principally by classical UPEC strains, while reUTIs are caused by strains that appear to be a part of the common E. coli intestinal biota. Moreover, although both peUTI and reUTI strains presented different serotypes and phylogroups, their antimicrobial resistance profile (XDR and MDR) was similar, confirming the importance of regulating prophylactic treatments and seeking alternatives for the treatment and control of cUTI. Finally, it was possible to establish the features of the E. coli strains responsible for peUTI and reUTI which could be helpful to develop a fast diagnostic methodology.

Association Between Uropathogenic Escherichia coli Virulence Genes and Severity of Infection and Resistance to Antibiotics.

Purpose: Urinary tract infection (UTI) is the most frequent bacterial infection. Some uropathogenic Escherichia coli (UPEC) genes have been associated with disease severity and antibiotic resistance. The aim was to determine the association of nine UPEC virulence genes with UTI severity and antibiotic resistance of strains collected from adults with community-acquired UTI. Patients and Methods: A case-control study (1:3) (38 urosepsis/pyelonephritis and 114 cystitis/urethritis) was conducted. The fimH, sfa/foc, cvaC, hlyA, iroN, fyuA, ireA, iutA, and aer (the last five are siderophore genes) virulence genes were determined by PCR. The information of antibiotic susceptibility pattern of the strains was collected from medical records. This pattern was determined using an automated system for antimicrobial susceptibility testing. Multidrug-resistant (MDR) was defined as resistance to three or more antibiotic families. Results: fimH was the most frequently detected virulence gene (94.7%), and sfa/foc was the least frequently detected (9.2%); 55.3% (83/150) of the strains were MDR. The evaluated genes were not associated with UTI severity. Associations were found between the presence of hlyA and carbapenem resistance (Odds ratio [OR] = 7.58, 95% confidence interval [CI], 1.50-35.42), iutA and fluoroquinolone resistance (OR = 2.35, 95% CI, 1.15-4.84, and aer (OR = 2.8, 95% CI, 1.20-6.48) and iutA (OR = 2.95, 95% CI, 1.33-6.69) with penicillin resistance. In addition, iutA was the only gene associated with MDR (OR = 2.09, 95% CI,1.03-4.26). Conclusion: There was no association among virulence genes and UTI severity. Three of the five iron uptake genes were associated with resistance to at least one antibiotic family. Regarding the other four non-siderophore genes, only hlyA was associated with antibiotic resistance to carbapenems. It is essential to continue studying bacterial genetic characteristics that cause the generation of pathogenic and multidrug-resistant phenotypes of UPEC strains.

Prevalência e perfil de suscetibilidade aos antimicrobianos de Escherichia coli em uroculturas de pacientes atendidos em um hospital de ensino / Prevalence and antimicrobial susceptibility profile of Escherichia coli in urine cultures from patients treated at a teaching hospital

Introdução: As infecções do trato urinário (ITU) são geralmente causadas por bactérias da ordem Enterobacterales, principalmente por Escherichia coli uropatogênica (UPEC). Esta linhagem apresenta fatores de virulência que a torna capaz de colonizar e infectar o trato urinário. Apesar da maioria dos quadros de ITU ser solucionado com terapia antimicrobiana, linhagens de UPEC resistentes aos antimicrobianos representam uma séria ameaça à saúde pública. Objetivo: Avaliar a prevalência de Escherichia coli em uroculturas de pacientes atendidos em um hospital de ensino, bem como seu perfil de suscetibilidade aos antimicrobianos e os fenótipos de resistência. Material e Métodos: Trata-se de um estudo descritivo que analisou uroculturas de pacientes ambulatoriais e hospitalares atendidos em um hospital de ensino localizado no município de Juiz de Fora, Minas Gerais, Brasil, no período de janeiro de 2020 a dezembro de 2021. Resultados: Entre as uroculturas analisadas, 858 foram positivas para bactérias, sendo Escherichia coli a espécie predominante, com 27,2% (n= 233) dos isolados. Das 858 uroculturas: 608 foram de pacientes hospitalizados, com 124 (20,4%) isolados de UPEC; 250 foram de pacientes ambulatoriais, com 109 (43,6%) isolados de UPEC. O perfil de resistência aos antimicrobianos das linhagens isoladas nas amostras hospitalares e ambulatoriais, foi, respectivamente: 65% e 32% para ampicilina; 56% e 26% para amoxicilina + ácido clavulânico; 50% e 26% para ciprofloxacino; 42% e 33% para sulfazotrim; 38% e 20% para cefepime; 17% e 8% para gentamicina; 2,5% e 0,4% para ertapenem, meropenem e imipenem. Das linhagens de Escherichia coli resistentes aos beta-lactâmicos, 43 (18%) apresentaram fenótipos de resistência do tipo beta lactamase de espectro ampliado (ESBL) e 7 (3%) foram produtoras de carbapenemases. Conclusão: Escherichia coli foi a espécie mais isolada das uroculturas. UPEC apresentou taxas de resistência a todos os antimicrobianos testados, produzindo fenótipos do tipo ESBL e carbapenemase, principalmente em amostras de pacientes hospitalizados.

Introduction: Urinary tract infections (UTI) are usually caused by bacteria of the Enterobacterales order, mainly by uropathogenic Escherichia coli (UPEC). This strain has virulence factors that make it able to colonize and infect the urinary tract. Although most cases of UTI are resolved with antimicrobial therapy, antimicrobial-resistant UPEC strains pose a serious threat to public health. Objective: To assess the prevalence of Escherichia coli in urine cultures of patients treated at a teaching hospital, as well as their antimicrobial susceptibility profile and resistance phenotypes. Material and Methods: This is a descriptive study that analyzed urine cultures of outpatient and hospital patients treated at a teaching hospital located in the city of Juiz de Fora, Minas Gerais, Brazil from January 2020 to December 2021. Results: Among the analyzed urine cultures, 858 were positive for bacteria, with Escherichia coli being the predominant species, with 27.2% (n= 233) of the isolates. Of the 858 urine cultures: 608 were from hospitalized patients, with 124 (20.4%) UPEC isolates; 250 were from outpatients, with 109 (43.6%) UPEC isolates. The antimicrobial resistance profile of the strains isolated from hospital and outpatient samples was, respectively: 65% and 32% for Ampicillin; 56% and 26% for Amoxicillin+Clavulanic acid; 50% and 26% for Ciprofloxacin; 42% and 33% for Sulfazotrim; 38% and 20% for Cefepime; 17% and 8% for Gentamicin; 2.5% and 0.4% for Ertapenem, Meropenem and Imipenem. Of the Escherichia coli strains resistant to beta-lactams, 43 (18%) showed extended-spectrum beta-lactamase (ESBL) resistance phenotypes and 7 (3%) were carbapenemases producers.Conclusion: Escherichia coli was the most isolated species from urine cultures. UPEC showed rates of resistance to all tested antimicrobials, producing ESBL and carbapenemase-like phenotypes, mainly in samples from hospitalized patients.

Uropathogenic Escherichia coli in Mexico, an Overview of Virulence and Resistance Determinants: Systematic Review and Meta-analysis.

BACKGROUND: Urinary tract infections (UTI) are one of the most common pathologies in Mexico and the majority are caused by uropathogenic Escherichia coli (UPEC). UPEC possesses virulence and resistance determinants that promote UTI development and affect diagnosis and treatment. This study aims to systematically review published reports of virulence genes, antibiotic resistance, and phylogenetic groups prevalent in clinical isolates of UPEC in the Mexican population. METHODS: Systematic review with meta-analysis was performed following PRISMA guidelines. Articles in both English and Spanish were included. Total prevalence with a 95% confidence interval of each characteristic was calculated. Heterogeneity between studies and geographical areas was assessed by the Cochran Q test (Q), I-square (I2), and H-square (H2). Egger's test was used for risk of bias in publications and asymmetry evaluations. RESULTS: Forty-two articles were analyzed. The most prevalent virulence genes were ecp (97.25%; n = 364) and fimH (82.34%; n = 1,422), which are associated with lower UTI, followed by papGII (40.98%; n = 810), fliC (38.87%; n = 319), hlyA (23.55%; n = 1,521), responsible for with upper UTI. More than 78.13% (n = 1,893) of the isolates were classified as multidrug-resistant, with a higher prevalence of resistance to those antibiotics that are implemented in the basic regimen in Mexico. The most frequently reported Extended Spectrum ß-Lactamase (ESBL) was CTX-M-1 (55.61%; n = 392), and the predominant phylogroup was B2 (35.94%; n = 1,725). CONCLUSION: UPEC strains are responsible for a large portion of both lower and upper UTI in Mexico, and their multi-drug resistance drastically reduces the number of therapeutic options available.

Estratégias de clonagem de proteínas de alta massa molecular: α-hemolisina e CNF1 produzidas por Escherichia coli uropatogênica como modelos

Uropathogenic Escherichia coli (UPEC) is the main cause of urinary tract infection (UTI). Among the virulence factors produced by UPEC, alpha hemolysin (HlyA) and cytotoxic necrotizing factor 1 (CNF1) stand out. Keeping in mind that up to 60% of UPEC strains produce HlyA and about 40% express CNF1, these toxins become good targets for diagnosis and therapeutic interventions. The importance of HlyA and CNF1 toxins in the cases of UTI caused by UPEC and the enormous challenges in the production of recombinant proteins, led to this work proposal to obtain different structural forms of these toxins through the methodology of cloning and heterologous expression, of integral toxins, low molecular mass and intermediate immunogenic toxins. Different structural forms of these high molecular weight toxins have been proposed, from which the production integrates to low molecular weight proteins. Recombinant toxins were analyzed in silico, expressed and purified. Among the cloning and expression approaches, we highlight for HlyA the strategy to obtain the intermediate immunogen of this toxin. For CNF1, both the construction of the full protein and the recombinant protein using only its catalytic portion. In addition, a bacterial collection of UPEC was analyzed to define local epidemiological data on the prevalence of toxin genes and the ability to cause hemolysis and multinucleation. The hlyA and cnf1 genes were present in the isolates in 36% and 23%, respectively, and the hemolysis phenotype occurred in 33% and the multinucleation in 23% of the bacterial isolates.

Escherichia coli uropatogênica (UPEC) é a principal causa de infecção do trato urinário (ITU). Entre os fatores de virulência produzidos pela UPEC, destacam- se a alfa hemolisina (HlyA) e o fator necrosante citotóxico 1 (CNF1). Tendo em vista que até 60% das cepas de UPEC produzem HlyA e cerca de 40% expressam CNF1, estas toxinas se tornam bons alvos para o diagnóstico e intervenções terapêuticas. A importância das toxinas HlyA e CNF1 nos casos de ITU causados por UPEC e os enormes desafios na produção de proteínas recombinantes, levou a este trabalho que se propôs a obter diferentes formas estruturais destas toxinas pela metodologia de clonagem e expressão heteróloga. Diferentes formas estruturais destas toxinas de alta massa molecular foram propostas, deste a produção integra a proteínas de baixo peso molecular. As toxinas recombinantes foram analisadas in silico, expressas e purificadas. Dentre as abordagens de clonagem e expressão destacamos para HlyA a estratégia de obtenção do imunógeno intermediário dessa toxina. Já para CNF1, tanto a construção da proteína integra quanto a proteína recombinante utilizando apenas sua porção catalítica. Além disso, analisou-se uma coleção bacteriana de UPEC para definir dados epidemiológicos locais da prevalência dos genes das toxinas e capacidade de causar hemólise e multinucleação. A genes hlyA e cnf1 estavam presentes nos isolados em 36% e 23%, respectivamente e o fenótipo de hemólise ocorreu em 33% e a multinucleação em 23% dos isolados bacterianos.

Isolation of Staphylococcus aureus, Uropathogenic Escherichia coli, and Nontuberculous Mycobacteria Strains from Pasteurized Cheeses and Unpasteurized Cream Sold at Traditional Open Markets in Mexico City.

ABSTRACT: Fresh cheeses and cream are important garnishes of traditional Mexican food, often purchased at street or itinerant open markets or tianguis. However, there is scarce information regarding the microbiological quality of cheeses and cream sold in tianguis. For 2 years, three dairy stalls from three tianguis in Mexico City were visited once each season, trading practices were registered, and 96 dairy products were purchased. In total 72 fresh pasteurized cheeses that were hand-cut to order (24 Panela, 24 Canasto, and 24 Doble Crema) and 24 unpasteurized Crema de Rancho samples were collected. All dairy products remained without refrigeration for 8 h. Based on the National Guidelines limits, 87.5% of cheeses and 8% of Crema de Rancho samples were of low microbiological quality, and 1 sample of each type of cheese and 3 samples of Crema de Rancho exceeded the guidelines limits for Staphylococcus aureus. All dairy products were negative for Salmonella, Listeria monocytogenes, and all diarrheagenic Escherichia coli pathotypes, including Shiga toxin-producing E. coli. Among the 96 dairy samples, the prevalence of uropathogenic E. coli (UPEC) and of mycobacteria strains were determined because food items contaminated with these strains have been associated with urinary tract infections and mycobacteriosis, respectively. UPEC strains were isolated from 43% of cut-to-order cheeses and 29% of Crema de Rancho samples. Nontuberculous mycobacteria (NTM) strains were identified in 12.5% of Doble Crema cheese samples and 21% of Crema de Rancho samples. From the eight NTM-positive samples, 10 strains were identified (3 strains of Mycolicibacterium fortuitum, 2 of Mycobacteroides abscessus, 2 of Mycobacteroides chelonae, 2 of Mycolicibacterium porcinum, and 1 of Mycolicibacterium rhodesiae). All produced biofilms, and 70% had sliding motility (both virulence traits). Trading practices of cut-to-order pasteurized cheeses and unpasteurized Crema de Rancho in tianguis increase the risk of microbiological contamination of these products, including with human pathogens, and their consumption may cause human illness.

Efficacy of Fosfomycin against Planktonic and Biofilm-Associated MDR Uropathogenic Escherichia coli Clinical Isolates.

Urinary tract infections (UTI) are a severe public health problem and are caused mainly by the uropathogenic Escherichia coli (UPEC). Antimicrobial resistance and limited development of new antimicrobials have led to the reuse of old antibiotics such as fosfomycin. The aim of this study was to evaluate the in vitro efficacy of fosfomycin on a collection of multidrug-resistant (MDR) UPEC and the degradative activity on biofilm producers. A total of 100 MDR UPEC clinical isolates were collected from patients at Mexican second- and third-level hospitals. Microorganism identification was performed using an automated system, the evaluation of the susceptibility of clinical isolates to fosfomycin was performed using the resazurin microtiter assay, and the identification of biofilm producers and the effect of fosfomycin in biofilms were evaluated using the crystal violet method. Among planktonic MDR UPEC, 93% were susceptible to fosfomycin. Eighty-three MDR UPEC were categorized as weak (39.8%), moderate (45.2%), and strong (14.5%) biofilm producers. Fosfomycin exhibited degradative activity ranging from 164.4 µg/mL to 1045 µg/mL. Weak producers required statistically lower concentrations of fosfomycin to destroy the biofilm, contrary to moderate and strong producers. In conclusion, fosfomycin could be an option for the treatment of infections caused by MDR UPEC, for which the antimicrobial treatment is more often becoming limited.

Molecular typification of Escherichia coli from community-acquired urinary tract infections in Mexico.

One hundred and five uropathogenic Escherichia coli (UPEC) strains from patients with community-acquired urinary tract infections were characterized according to phylogenetic group, virulence factors, serogroup, antibiotic resistance, and genotype. The pathogenic phylogenetic groups (B2, D, and F) were found in 71.4% of the tested strains. Among them, the main uropathogenic serogroups were O8, O25, and O75, in which 97.1% of the strains had a multidrug-resistant profile. Sixteen virulence genes were analysed using a combination of polymerase chain reaction (PCR) assays, with the fimH, irp-2, iutA, aer, iucC, PAI, sat, iroN, usp, and cnf1 genes being mainly found in pathogenic phylogroups. The E. coli O25b-ST131 clone was identified in 32% of the strains assigned to the pathogenic phylogroup B2. These findings demonstrate that virulence genes encoding adhesin components, iron-acquisition systems, toxins, and pathogenicity-associated islands were highly prevalent among the pathogenic phylogroup of UPEC strains.

Community-Acquired Uropathogenic Escherichia coli, Antimicrobial Susceptibility, and Extended-Spectrum Beta-Lactamase Detection.

INTRODUCTION: Urinary tract infection is the second-leading reason for consults in primary health care. Bacterial urinary tract infections are the most common, of which Escherichia coli is the main etiologic agent. Antimicrobial resistance and multidrug resistance complicate effective community treatment, especially if resistance is caused by extended-spectrum beta-lactamase production. WHO recommends that antimicrobial susceptibility be evaluated in different regions of the world at different times. Community-acquired E. coli's susceptibility to colistin has not yet been studied in Cuba, and mcr-1 gene screening is necessary. OBJECTIVE: Evaluate community-acquired uropathogenic E. coli isolates' susceptibility to antibiotics, including colistin, and identify extended-spectrum beta-lactamase-producing bacteria. METHODS: We conducted a descriptive cross-sectional study that included 281 community-acquired uropathogenic E. coli isolates (153 from the Isle of Youth Special Municipality's Hygiene, Epidemiology, and Microbiology Center and 128 from Microbiology Laboratories of 7 institutions in Havana) from June 2016 through July 2018. We used the disk diffusion method to determine susceptibility to ampicillin, ampicillin/sulbactam, cefazolin, trimethoprim/sulfamethoxazole, ciprofloxacin, nitrofurantoin and fosfomycin. The disk elution method was used to determine susceptibility to colistin. The combined disk method was used to identify extended-spectrum beta-lactamases. Estimates were made regarding the frequency and percentages of antimicrobial susceptibility and resistance, as well as multidrug-resistance patterns. RESULTS: Of the 281 isolates, 68.3% (192/281) were resistant to ampicillin, 54.8% (154/281) were resistant to ciprofloxacin, and 49.5% (139/281) were resistant to trimethoprim/sulfamethoxazole. Resistance to colistin was not detected. On the other hand, 14.2% (40/281) were susceptible to the 8 antibiotics we evaluated, 22.1% (62/281) showed resistance to only 1 antibiotic, and 63.7% (179/281) were resistant to 2 or more antibiotics. In the extended-spectrum beta-lactamase determination, 34.5% (97/281) had inhibition zones ≤14 mm with cefazolin. Of those with inhibition zones, 64.9% (63/97) were positive in the phenotype test, and 35.1% (34/97) were negative. In extended-spectrum beta-lactamase-producing bacteria, 1.6% (1/63) were resistant to fosfomycin, and 3.2% (2/63) were resistant to nitrofurantoin. The most common multidrug-resistance pattern (22.9%; 30/131) was to ampicillin/sulbactam, ampicillin, cefazolin, ciprofloxacin, and trimethoprim/sulfamethoxazole. CONCLUSIONS: Uropathogenic E. coli resistance to the antibiotics most frequently used in community medical practice is quite common, and extended-spectrum beta-lactamase-producing bacteria is the mechanism for beta-lactam antibiotic resistance. Multidrug-resistance patterns include resistance to the antibiotics most used in community-acquired infections. Fosfomycin and nitrofurantoin are the most active in extended-spectrum beta-lactamase producing bacteria. All the isolates were susceptible to colistin.

Susceptibility to first choice antimicrobial treatment for urinary tract infections to Escherichia coli isolates from women urine samples in community South Brazil.

E. coli is the main pathogen of UTI. It is important to be aware the local epidemiological data for an appropriate initial treatment. Resistance to antimicrobial agents has increased, especially to first-choice antibiotics in the treatment of cystitis. There are few studies on the sensivity profile of community uropathogen in our region. OBJECTIVE: To characterize antimicrobials the sensitivity profile to E. coli isolated from urocultures of women treated at Basic Health Units and Emergency Care Units of Londrina- Paraná- Brazil during a period of 12 months (June 1, 2016 to June 1, 2017). METHODOLOGY: A cross-sectional study was carried out from June 2016 to June 2017. All urine samples collected in the Basic Health Units and Emergency Departments in the city of Londrina (Paraná State, Brazil) were sent to a Central Laboratory where the identification and antimicrobial susceptibility testing were performed. Clinical Laboratory Standards Institute (CLSI) breakpoints were used for the interpretation of susceptibility testing results. RESULTS: 56,555 urine cultures were performed in the period, of which 8,832 were positive, of which 5,377 were women. Of these samples, 4.7% were enterobacteria producing extended-spectrum beta-lactamases (ESBL) and 15.5% resistant to quinolones. TMP- SMX was resistant in more than 30% of the samples in all age groups. Among quinolone-resistant isolates, resistance to cephalothin, ampicillin and sulfamethoxazole-trimethoprim was greater than 60%. Nitrofurantoin was the only antimicrobial that showed 90% of sensitivity. CONCLUSION: The antimicrobials sensitivity profile was similar to that reported in the literature, with TMP- SMX resistance greater than 30% in the studied samples. Nitrofurantoin maintains high sensitivity rates greater than 90%. Resistance to quinolones increases proportionally with age, as well ESBL.

Saccharomyces boulardii exerts renoprotection by modulating oxidative stress, renin angiotensin system and uropathogenic microbiota in a murine model of diabetes.

AIMS: We aimed to investigate whether Saccharomyces boulardii strain might exert renoprotective effects by modulating renal renin angiotensin system, oxidative stress and intestinal microbiota in streptozotocin-diabetic mice. MAIN METHODS: Thirty-six C57BL/6 male mice were divided into four groups: control (C), control + probiotic (CP), diabetes (D), diabetes + probiotic (DP). Diabetes was induced by one intraperitoneal injection of streptozotocin and Saccharomyces boulardii was administered by oral gavage for 8 weeks. Blood glucose, albuminuria and urinary volume were measured. Renal levels of angiotensin peptides (angiotensin I, II and 1-7) and the activities of angiotensin-converting enzyme (ACE) and ACE2 were determined, besides that, renal morphology, serotonin and dopamine levels and also microbiota composition were analyzed. KEY FINDINGS: Probiotics significantly increased C-peptide secretion and reduced blood glucose of diabetic animals. Saccharomyces boulardii also improved renal antioxidant defense, restored serotonin and dopamine concentration, and activated the renin-angiotensin system (RAS) vasodilator and antifibrotic axis. The modulation of these markers was associated with a beneficial impact on glomerular structure and renal function of diabetic treated animals. The phenotypic changes induced by Saccharomyces boulardii were also related to modulation of intestinal microbiota, evidenced by the decreased abundance of Proteus and Escherichia-Shigella, considered diabetic nephropathy biomarkers. SIGNIFICANCE: Therefore, probiotic administration to streptozotocin-induced diabetic mice improves kidney structure and function in a murine model and might represent a reasonable strategy to counteract nephropathy-associated maladaptive responses in diabetes.

ß-lactamasas de espectro extendido y factores de virulencia en Escherichia coli uropatógenas en asilos de ancianos en Lima, Perú / ß-lactamases and virulence factors in uropathogenic Escherichia coli in nursing homes in Lima, Peru

RESUMEN Los asilos de ancianos son instituciones con una alta prevalencia de infecciones del tracto urinario ocasionado por Escherichia coli productoras de ß-lactamasas de espectro extendido (BLEE), con diversos factores de virulencia. El objetivo del estudio fue determinar la frecuencia del gen bla CTX-M y de ocho genes de virulencia en 35 E. coli uropatógenas productoras de BLEE provenientes de seis asilos en Perú, durante el 2018. El 57,1% (20/35) de las E. coli fueron portadores del gen bla CTX-M. Además, se obtuvo una frecuencia del 46% (15/35) y 37% (13/35) de hly-alfa y cnf-1, respectivamente; elevada presencia de los genes iucC (63%, 22/35), aer (94%, 33/35) y chuA (94%, 33/34) y una frecuencia del 46% (16/35) y del 91% (32/34) de los genes pap GII y nanA, respectivamente. Existe predominancia en la distribución del gen bla CTX-M, además de una alta frecuencia de exotoxinas que le confieren una ventaja competitiva para diseminarse hacia el torrente sanguíneo.

ABSTRACT Nursing homes are institutions with high prevalence of urinary tract infections caused by ESBL-producing E. coli with several virulence factors. The aim of this study was to determine the frequency of the bla CTX-M gene and eight virulence genes in 35 ESBL-producing uropathogenic E. coli from six nursing homes in Peru during 2018. Of the E. coli samples, 57.1% (20/35) were carriers of the bla CTX-M gene. Furthermore, we obtained frequencies of 46% (15/35) and 37% (13/35) for hly-alpha and cnf-1, respectively; we also found high presence of the iucC (63%, 22/35), aer (94%, 33/35) and chuA genes (94%, 33/34) as well as a frequency of 46% (16/35) and 91% (32/34) for the pap GII and nanA genes, respectively. The bla CTX-M gene is predominant and a high frequency of exotoxins gives it a competitive advantage for spreading into the bloodstream.

In Silico Prediction and Design of Uropathogenic Escherichia coli Alpha-Hemolysin Generate a Soluble and Hemolytic Recombinant Toxin.

The secretion of α-hemolysin by uropathogenic Escherichia coli (UPEC) is commonly associated with the severity of urinary tract infections, which makes it a predictor of poor prognosis among patients. Accordingly, this toxin has become a target for diagnostic tests and therapeutic interventions. However, there are several obstacles associated with the process of α-hemolysin purification, therefore limiting its utilization in scientific investigations. In order to overcome the problems associated with α-hemolysin expression, after in silico prediction, a 20.48 kDa soluble α-hemolysin recombinant denoted rHlyA was constructed. This recombinant is composed by a 182 amino acid sequence localized in the aa542-723 region of the toxin molecule. The antigenic determinants of the rHlyA were estimated by bioinformatics analysis taking into consideration the tertiary form of the toxin, epitope analysis tools, and solubility inference. The results indicated that rHlyA has three antigenic domains localized in the aa555-565, aa600-610, and aa674-717 regions. Functional investigation of rHlyA demonstrated that it has hemolytic activity against sheep red cells, but no cytotoxic effect against epithelial bladder cells. In summary, the results obtained in this study indicate that rHlyA is a soluble recombinant protein that can be used as a tool in studies that aim to understand the mechanisms involved in the hemolytic and cytotoxic activities of α-hemolysin produced by UPEC. In addition, rHlyA can be applied to generate monoclonal and/or polyclonal antibodies that can be utilized in the development of diagnostic tests and therapeutic interventions.