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.

Phage Display Detection of Mimotopes that Are Shared Epitopes of Clinically and Epidemiologically Relevant Enterobacteria.

BACKGROUND: Escherichia coli and Salmonella are etiologic agents of intestinal infections. A previous study showed the presence of shared epitopes between lipopolysaccharides (LPSs) of E. coli O157 and Salmonella. AIM: Using phage display, the aim of this study is to identify mimotopes of shared epitopes in different enterobacterial LPSs. METHODS: We use anti-LPS IgG from E. coli O157 and Salmonella to select peptide mimotopes of the M13 phage. The amino acid sequence of the mimotopes is used to synthesize peptides, which are in turn used to immunize rabbits. The antibody response of the resulting sera against the LPSs and synthetic peptides (SPs) is analyzed by ELISA and by Western blot assays, indicating that LPS sites are recognized by the same antibody. In a complementary test, the reactions of human serum samples obtained from the general population against the SPs and LPSs are also analyzed. RESULTS: From the last biopanning phase, sixty phagotopes are selected. The analysis of the peptide mimotope amino acid sequences shows that in 4 of them the S/N/A/PF motif is a common sequence. Antibodies from the sera of immunized rabbits with SP287/3, SP459/1, SP308/3, and SP073/14 react against both their own peptide and the different LPSs. The Western blot test shows a sera reaction against both the lateral chains and the cores of the LPSs. The analysis of the human sera shows a response against the SPs and LPSs. CONCLUSION: The designed synthetic peptides are mimotopes of LPS epitopes of Salmonella and E. coli that possess immunogenic capacity. These mimotopes could be considered for use in the design of vaccines against both enterobacteria.

Persistencia, internalización y translocación de Escherichia coli O157:H7, O157:H16 y O105ab en plantas y frutos de tomate (Solanum lycopersicum L.) / Persistence, internalization and translocation of Escherichia coli O157:H7, O157:H16 and O105ab in plants and tomato fruits (Solanum lycopersicum L.)

La presencia de bacterias patógenas, como Escherichia coli, afecta la calidad e inocuidad de las hortalizas que se consumen en fresco y se relaciona con graves problemas de salud. El objetivo de este trabajo fue determinar si 3 cepas diferentes de E. coli tienen la capacidad de penetrar y permanecer en plantas y frutos de tomate. Se siguió un diseño experimental completamente al azar, para lo cual se estableció un cultivo de tomate (variedad «Cid¼) en condiciones de invernadero y se evaluaron 3 tratamientos, T1 (E. coli O157: H7), T2 (E. coli de cultivo de tomate -#91;EcT-#93; O157: H16), T3 (E. coli de cultivo de espinaca -#91;EcH-#93; EcH O105ab) y un testigo T4, con 100 plantas cada uno y 4 formas de inoculación: en el sustrato, en el tallo, en el pecíolo y en el pedúnculo. Se realizaron muestreos en etapa vegetativa, floración, fructificación y madurez fisiológica para cuantificar en placa las UFC/g y saber si las bacterias lograban moverse y recuperarse en la raíz, el tallo, la flor y el fruto. Los grupos filogenéticos a los que correspondieron las bacterias recuperadas fueron confirmados mediante pruebas bioquímicas, serotipificación y PCR. A los 120 días la recuperación de bacterias en la planta fue del 23% (E. coli O157: H7), 28% (EcT O157: H16) y 55% (EcH O105ab) con la inoculación al sustrato, mientras que con la inoculación por punción la recuperación fue (en igual orden) del 5%, 3% y 4% a los 30 días; del 37%, 35% y 30% a los 90 días; y del 42%, 39% y 13% a los 65 días. Las cepas utilizadas mostraron la capacidad de entrar en la planta de tomate y de permanecer en ella y transportarse hasta llegar al fruto, sin producir síntomas que indiquen su presencia.

The presence of pathogenic bacteria, such as Escherichia coli affects the quality and safety of vegetables that are consumed fresh and is associated with serious health problems. The objective of this study was to determine if three different strains of E. coli can penetrate and remain in plants and tomato fruits. A completely randomized experimental design was followed for which a tomato crop ("Cid" variety) was established under greenhouse conditions and three treatments were evaluated, T1 (E. coli O157: H7), T2 (E. coli from tomato cultivation -#91;EcT-#93; O157: H16), T3 (E. coli from spinach cultivation -#91;EcH-#93; O105ab) and a T4 control, with 100 plants each and four forms of inoculation: in the substrate, steam, petiole and the peduncle. Samples were carried out in vegetative stage, flowering, fruiting and physiological maturity to quantify in petri dish CFU/g and know if the bacteria managed to move around and recover in root, stem, flower and fruit. The phylogenetic groups that corresponded to the bacteria recovered were confirmed by biochemical tests, serotyping and PCR. At 120 days the recovery of bacteria in the plant was 23% (E. coli O157: H7), 28% (EcT O157: H16) and 55% (EcH O105ab) whit inoculation to the substrate while the inoculation by puncture the recovery was (in the same order) of 5%, 3%, and 4% at 30 days; 37%, 35% and 30% at 90 days; and 42%, 39% and 13% at 65 days. The strains submit the ability to enter the tomato plant and to stay in it and transported to the fruit, without producing that indicate their presence.

[Persistence, internalization and translocation of Escherichia coli O157:H7, O157:H16 and O105ab in plants and tomato fruits (Solanum lycopersicum L.)]. / Persistencia, internalización y translocación de Escherichia coli O157:H7, O157:H16 y O105ab en plantas y frutos de tomate (Solanum lycopersicum L.).

The presence of pathogenic bacteria, such as Escherichia coli affects the quality and safety of vegetables that are consumed fresh and is associated with serious health problems. The objective of this study was to determine if three different strains of E. coli can penetrate and remain in plants and tomato fruits. A completely randomized experimental design was followed for which a tomato crop ("Cid" variety) was established under greenhouse conditions and three treatments were evaluated, T1 (E. coli O157:H7), T2 (E. coli from tomato cultivation [EcT] O157:H16), T3 (E. coli from spinach cultivation [EcH] O105ab) and a T4 control, with 100 plants each and four forms of inoculation: in the substrate, steam, petiole and the peduncle. Samples were carried out in vegetative stage, flowering, fruiting and physiological maturity to quantify in petri dish CFU/g and know if the bacteria managed to move around and recover in root, stem, flower and fruit. The phylogenetic groups that corresponded to the bacteria recovered were confirmed by biochemical tests, serotyping and PCR. At 120 days the recovery of bacteria in the plant was 23% (E. coli O157:H7), 28% (EcT O157:H16) and 55% (EcH O105ab) whit inoculation to the substrate while the inoculation by puncture the recovery was (in the same order) of 5%, 3%, and 4% at 30 days; 37%, 35% and 30% at 90 days; and 42%, 39% and 13% at 65 days. The strains submit the ability to enter the tomato plant and to stay in it and transported to the fruit, without producing that indicate their presence.

Corrigendum: Effects of lng Mutations on LngA Expression, Processing, and CS21 Assembly in Enterotoxigenic Escherichia coli E9034A.

[This corrects the article on p. 1201 in vol. 7, PMID: 27536289.].

Data on enterobacteria activity on biofilm formation at surface mango fruit (Mangifera indica L.) cv Ataulfo.

Abiotic factors influenced the capacity of the strains to form biofilms. Classification of the adhesion type is related with the optical density measured on the biofilm formation of tested strains. The relationship between the biofilm formation in real values with theoretical values of the strains was used to determine the mechanism involved during mixed cultures.

Immunogenic peptide mimotopes from an epitope of Escherichia coli O157 LPS.

Escherichia coli O157:H7 is a subtype of Shiga toxin-producing E. coli that is associated with haemorrhagic colitis and haemolytic uremic syndrome (HUS). Studies of populations in endemic areas have reported that the presence of specific antibodies against the O157 lipopolysaccharide (LPS) is associated with a lower incidence of diarrhoea and HUS. Phage display and IgG anti-O157 LPS antibodies were used in the present study to select peptide mimotopes of O157 LPS expressed in protein III of the M13 phage. Synthetic peptides (SP) were designed using the derived amino acid sequences obtained from DNA nucleotides of 63 selected phagotopes. The LxP/YP/SxL motif was identified in five of the phagotope amino acid sequences. Antibody responses against the phagotopes and their corresponding SPs were evaluated. SP12, one of the designed SP, induced the production of antibodies against the homologous peptide (1:800) and O157 LPS (1:200). The specificity of anti-SP12 antiserum was confirmed by analyzing its response to SP3, an SP with a different amino acid sequence than that of SP12, as well as against an E. coli LPS different from O157. Competitive studies with SP12 and O157 LPS showed a significant decrease in anti-SP12 and anti-LPS O157 antiserum responses against SP12 and O157 LPS, respectively. Eighteen (82%) of the 22 human serum samples with positive reactivity against E coli O157 LPS reacted with SP12 SP (cut-off >0.4). These results support the idea that SP12 is an immunogenic mimotope of O157 LPS.

Effects of lng Mutations on LngA Expression, Processing, and CS21 Assembly in Enterotoxigenic Escherichia coli E9034A.

Enterotoxigenic Escherichia coli (ETEC) is a major cause of morbidity in children under 5 years of age in low- and middle-income countries and a leading cause of traveler's diarrhea worldwide. The ability of ETEC to colonize the intestinal epithelium is mediated by fimbrial adhesins, such as CS21 (Longus). This adhesin is a type IVb pilus involved in adherence to intestinal cells in vitro and bacterial self-aggregation. Fourteen open reading frames have been proposed to be involved in CS21 assembly, hitherto only the lngA and lngB genes, coding for the major (LngA) and minor (LngB) structural subunit, have been characterized. In this study, we investigated the role of the LngA, LngB, LngC, LngD, LngH, and LngP proteins in the assembly of CS21 in ETEC strain E9034A. The deletion of the lngA, lngB, lngC, lngD, lngH, or lngP genes, abolished CS21 assembly in ETEC strain E9034A and the adherence to HT-29 cells was reduced 90%, compared to wild-type strain. Subcellular localization prediction of CS21 proteins was similar to other well-known type IV pili homologs. We showed that LngP is the prepilin peptidase of LngA, and that ETEC strain E9034A has another peptidase capable of processing LngA, although with less efficiency. Additionally, we present immuno-electron microscopy images to show that the LngB protein could be localized at the tip of CS21. In conclusion, our results demonstrate that the LngA, LngB, LngC, LngD, LngH, and LngP proteins are essential for CS21 assembly, as well as for bacterial aggregation and adherence to HT-29 cells.

X-ray crystal structure of the passenger domain of plasmid encoded toxin(Pet), an autotransporter enterotoxin from enteroaggregative Escherichia coli (EAEC).

Autotransporters (ATs) represent a superfamily of proteins produced by a variety of pathogenic bacteria, which include the pathogenic groups of Escherichia coli (E. coli) associated with gastrointestinal and urinary tract infections. We present the first X-ray structure of the passenger domain from the Plasmid-encoded toxin (Pet) a 100 kDa protein at 2.3 Å resolution which is a cause of acute diarrhea in both developing and industrialized countries. Pet is a cytoskeleton-altering toxin that induces loss of actin stress fibers. While Pet (pdb code: 4OM9) shows only a sequence identity of 50% compared to the closest related protein sequence, extracellular serine protease plasmid (EspP) the structural features of both proteins are conserved. A closer structural look reveals that Pet contains a ß-pleaded sheet at the sequence region of residues 181-190, the corresponding structural domain in EspP consists of a coiled loop. Secondary, the Pet passenger domain features a more pronounced beta sheet between residues 135 and 143 compared to the structure of EspP.

CS21 positive multidrug-resistant ETEC clinical isolates from children with diarrhea are associated with self-aggregation, and adherence.

BACKGROUND: Enterotoxigenic Escherichia coli (ETEC) colonize the human intestinal mucosa using pili and non-pili colonization factors (CFs). CS21 (also designated Longus) is one of the most prevalent CFs encoded by a 14 kb lng DNA cluster located in a virulence plasmid of ETEC; yet limited information is available on the prevalence of CS21 positive ETEC isolates in different countries. The aim of this study was to evaluate the prevalence of CS21 among ETEC clinical isolates from Mexican and Bangladeshi children under 5 years old with diarrhea and to determine the phenotypic and genotypic features of these isolates. METHODS: ETEC clinical isolates positive to lngA gene were characterized by genotype, multidrug-resistance, self-aggregation, biofilm formation, and adherence to HT-29 cell line. RESULTS: A collection of 303 E. coli clinical isolates were analyzed, the 81.51% (247/303) were identified as ETEC, 30.76% (76/247) were st (+)/lt (+), and 25.10% (62/247) were positive for the lngA gene. Among the lngA (+) ETECs identified, 50% of isolates (31/62) were positive for LngA protein. The most frequent serotype was O128ac:H12 found in 19.35% (12/62) of lngA (+) ETEC studied. Multidrug-resistance (MDR) lngA (+) ETEC isolates was identified in 65% (39/60), self-aggregation in 48.38% (30/62), and biofilm formation in 83.87% (52/62). ETEC lngA (+) isolates were able to adhere to HT-29 cells at different levels. Two lngA isogenic mutants were constructed in the ETEC E9034A and ETEC73332 clinical isolate, showing a 77% and 98% reduction in adherence, respectively with respect to the wild type. CONCLUSION: ETEC isolates that have the lngA gene showed features associated with self-aggregation, and adherence to HT-29 cells, important characteristics in the human gut colonization process and pathogenesis.

Susceptibilidad antimicrobiana de aislamientos de Escherichia coli procedentes de ecosistemas dulceacuícolas / Antimicrobial susceptibility of Escherichia coli isolates from river water ecosystems

Introducción: la resistencia antimicrobiana constituye uno de los mayores problemas que afronta la salud pública mundial. La aparición de cepas resistentes no solo de origen clínico sino también ambiental agrava la situación. Entre los microorganismos que presentan esta característica se destaca la especie bacteriana Escherichia coli debido a su doble papel como indicador de contaminación fecal y como patógeno. Objetivos: aislar e identificar hasta especie aislamientos de Escherichia coli a partir de muestras de agua procedentes de ríos contaminados de La Habana y determinar la susceptibilidad antimicrobiana in vitro de estos aislados. Métodos: se estudiaron 113 aislamientos de bacterias coliformes aislados de 10 estaciones de muestreo ubicadas en la zona urbana de los ríos capitalinos Almendares, Quibú y Luyanó en el período comprendido de febrero de 2008 hasta junio de 2010. La identificación de los aislados, la determinación de la susceptibilidad antimicrobiana y la búsqueda de b-lactamasas de espectro extendido se realizó mediante el método automatizado VITEK. Resultados: se identificaron 113 cepas ambientales de Escherichia coli. Se demostró que 23 % de los aislamientos resultaron resistentes al menos a uno de los antimicrobianos evaluados. Los mayores porcentajes de resistencia se observaron frente a ampicilina, sulfametoxazol-trimetropin y ciprofloxacina. Conclusiones: la presencia de aislados de E. coli con multirresistencia antimicrobiana en estos ríos indica claramente el riesgo biológico que implica el uso de sus aguas.

Introduction: antimicrobial resistance is one of the biggest problems facing global public health. The emergence of resistant clinical and environmental strains worsens the situation. Among the microorganisms with antimicrobial resistance, Escherichia coli species stands out due to its dual role as fecal contamination indicator and pathogen. Objectives: to isolate and identify Escherichia coli isolates from water samples from polluted rivers located in La Habana, and to determine their antimicrobial in vitro susceptibility. Methods: one hundred thirteen isolates of coliform bacteria isolated from 10 sampling stations in the capital´s urban areas near Almendares, Quibú and Luyanó rivers were studied in the period of February 2008 to June 2010. The identification of isolates, the determination of antimicrobial susceptibility and the search for extended-spectrum b-lactamase were all performed using VITEK automated method. Results: one hundred thirteen environmental strains of Escherichia coli were identified. It showed that 23 % of the isolates were resistant to at least one of the tested antimicrobials. The highest percentages of resistance were observed to ampicilline, sulfamethoxazole-trimethoprim and ciprofloxacin. Conclusions: the presence of E. coli isolates with multiple antimicrobial resistances in these rivers clearly indicates the biological risk involving the use of their waters.

[Antimicrobial susceptibility of Escherichia coli isolates from river water ecosystems]. / Susceptibilidad antimicrobiana de aislamientos de Escherichia coli procedentes de ecosistemas dulceacuícolas.

INTRODUCTION: Antimicrobial resistance is one of the biggest problems facing global public health. The emergence of resistant clinical and environmental strains worsens the situation. Among the microorganisms with antimicrobial resistance, Escherichia coil species stands out due to its dual role as fecal contamination indicator and pathogen. OBJECTIVES: To isolate and identify Escherichia coil isolates from water samples from polluted rivers located in La Habana, and to determine their antimicrobial in vitro susceptibility. METHODS: One hundred thirteen isolates of coliform bacteria isolated from 10 sampling stations in the capital's urban areas near Almendares, Quibú and Luyanó rivers were studied in the period of February 2008 to June 2010. The identification of isolates, the determination of antimicrobial susceptibility and the search for extended-spectrum beta-lactamase were all performed using VITEK automated method. RESULTS: One hundred thirteen environmental strains of Escherichia coli were identified. It showed that 23% of the isolates were resistant to at least one of the tested antimicrobials. The highest percentages of resistance were observed to ampicilline, sulfamethoxazole-trimethoprim and ciprofloxacin. CONCLUSIONS: The presence of E. coil isolates with multiple antimicrobial resistances in these rivers clearly indicates the biological risk involving the use of their waters.