Genomic Dissection of an Enteroaggregative Escherichia coli Strain Isolated from Bacteremia Reveals Insights into Its Hybrid Pathogenic Potential.

Escherichia coli is a frequent pathogen isolated from bloodstream infections. This study aimed to characterize the genetic features of EC092, an E. coli strain isolated from bacteremia that harbors enteroaggregative E. coli (EAEC) genetic markers, indicating its hybrid pathogenic potential. Whole-genome sequencing showed that EC092 belongs to phylogroup B1, ST278, and serotype O165:H4. Genes encoding virulence factors such as fimbriae, toxins, iron-uptake systems, autotransporter proteins (Pet, Pic, Sat, and SepA), and secretion systems were detected, as well as EAEC virulence genes (aggR, aatA, aaiC, and aap). EC092 was found to be closely related to the other EAEC prototype strains and highly similar in terms of virulence to three EAEC strains isolated from diarrhea. The genomic neighborhood of pet, pic, sat, sepA, and the EAEC virulence genes of EC092 and its three genetically related fecal EAEC strains showed an identical genomic organization and nucleotide sequences. Also, EC092 produced and secreted Pet, Pic, Sat, and SepA in the culture supernatant and resisted the bactericidal activity of normal human serum. Our results demonstrate that the strain EC092, isolated from bacteremia, is a hybrid pathogenic extraintestinal E. coli (ExPEC)/EAEC with virulence features that could mediate both extraintestinal and intestinal infections.

Enteroaggregative Escherichia coli (EAEC) isolates obtained from non-diarrheic children carry virulence factor-encoding genes from Extraintestinal Pathogenic E. Coli (ExPEC).

Enteroaggregative E. coli (EAEC) is one of the most frequent pathogens isolated from diarrheal patients as well as from healthy individuals in Brazil and has recently also been implicated as an extraintestinal pathogenic E. coli (ExPEC) associated with bloodstream and urinary tract infections. In this study, 37 EAEC isolates, obtained from fecal samples of non-diarrheic children, were molecularly and phenotypically characterized to access the pathogenic features of these isolates. The EAEC isolates were assigned into the phylogroups A (54.1%), D (29.7%), B1 (13.5%) and B2 (2.7%); and harbored genes responsible for encoding the major pilin subunit of the aggregative adherence fimbriae (AAFs) or aggregate-forming pili (AFP) adhesins as follows: aggA (24.3%), agg3A (5.4%), agg4A (27.0%), agg5A (32.4%) and afpA (10.8%). The most frequent O:H serotypes were O15:H2 (8.1%), O38:H25 (5.4%) and O86:H2 (5.4%). Twenty-one isolates (56.8%) produce the aggregative adherence (AA) pattern on HeLa cells, and biofilm formation was more efficient among EAEC isolates harboring the aggA and agg5A genes. PFGE analysis showed that 31 (83.8%) of the isolates were classified into 10 distinct clusters, which reinforces the high diversity found among the isolates studied. Of note, 40.5% (15/37) of the EAEC isolates have a genetic profile compatible with E. coli isolates with intrinsic potential to cause extraintestinal infections in healthy individuals, and therefore, classified as EAEC/ExPEC hybrids. In conclusion, we showed the presence of EAEC/ExPEC hybrids in the intestinal microbiota of non-diarrheic children, possibly representing the source of some endogenous extraintestinal infections.

Caracterización molecular de Escherichia coli diarreogénica proveniente de población pediátrica ambulatoria con diarrea, atendida en dos hospitales de Buenos Aires, Argentina / Molecular characterization of diarrheagenic Escherichia coli from an outpatient pediatric population with diarrhea attended in two hospitals from Buenos Aires, Argentina

Resumen Escherichia coli diarreogénica abarca un grupo heterogéneo de patotipos o variantes patogénicas que comparten características fenotípicas con marcadas diferencias en genes de virulencia, sitios de colonización, patogenia, presentación clínica y epidemiología de la infección. Los patotipos más estudiados son E.coli productora de toxina Shiga (STEC), E.coli enterotoxigénica (ETEC), E.coli enteropatogénica (EPEC), E.coli enteroagregativa (EAEC) y E.coli enteroinvasiva (EIEC). El objetivo del estudio fue caracterizar los aislamientos de E.coli diarreogénica provenientes de población pediátrica ambulatoria con diarrea, atendida en dos hospitales públicos de Buenos Aires, Argentina. Los patotipos de E.coli diarreogénica se investigaron mediante la amplificación de fragmentos de genes de virulencia característicos: intimina (eae), toxina termolábil (lt), toxinas termoestables (stp, sth), antígeno plasmídico de invasiónH (ipaH), activador transcripcional R (aggR) y toxinas Shiga (stx1, stx2). La subtipificación molecular de aislamientos se realizó mediante PFGE (XbaI). E.coli diarreogénica fue detectada en el 14% (84/601) de los casos. El patotipo EAEC fue prevalente, mientras que ETEC, STEC, EPEC y EIEC fueron hallados en menor proporción. Los aislamientos de EAEC presentaron un alto grado de diversidad genética. Todos los patotipos fueron hallados en niños menores de 5años, mientras que solamente EAEC, EIEC y ETEC fueron detectados en población de mayor edad. Futuros estudios que incluyan la caracterización de aislamientos a partir de un mayor número de genes y población de otras áreas geográficas serán necesarios para determinar la relevancia de E.coli diarreogénica en Argentina.

Abstract Diarrheagenic Escherichia coli comprises a heterogeneous group of pathotypes or pathogenic variants that share phenotypic characteristics with marked differences in virulence genes, colonization sites, pathogenesis, clinical presentation, and epidemiology of infection. The most studied pathotypes are Shiga toxin-producing E.coli (STEC), enterotoxigenic E.coli (ETEC), enteropathogenic E.coli (EPEC), enteroaggregative E.coli (EAEC), and enteroinvasive E.coli (EIEC). The objective of the study was to characterize the isolates of diarrheagenic E.coli from an outpatient pediatric population with diarrhea attended in two public hospitals from Buenos Aires, Argentina. Diarrheagenic E.coli pathotypes were investigated by amplifying characteristic virulence gene fragments: intimin (eae), heat-labile toxin (lt), heat-stable toxins (stp, sth), invasion plasmid antigen H (ipaH), transcriptional activator R (aggR) and Shiga toxins (stx1, stx2). Molecular subtyping of isolates was performed using PFGE (XbaI). Diarrheagenic E.coli was detected in 14% (84/601) of cases. The EAEC pathotype was prevalent, while ETEC, STEC, EPEC and EIEC were found in a lower proportion. EAEC isolates exhibited a high degree of genetic diversity. All pathotypes were found in children under 5years of age, while only EAEC, EIEC and ETEC were detected in the older population. Future studies that include the characterization of isolates from a greater number of genes and populations from other geographical areas will be necessary to determine the relevance of diarrheagenic E.coli in Argentina.

[Molecular characterization of diarrheagenic Escherichia coli from an outpatient pediatric population with diarrhea attended in two hospitals from Buenos Aires, Argentina]. / Caracterización molecular de Escherichia coli diarreogénica proveniente de población pediátrica ambulatoria con diarrea, atendida en dos hospitales de Buenos Aires, Argentina.

Diarrheagenic Escherichia coli comprises a heterogeneous group of pathotypes or pathogenic variants that share phenotypic characteristics with marked differences in virulence genes, colonization sites, pathogenesis, clinical presentation, and epidemiology of infection. The most studied pathotypes are Shiga toxin-producing E.coli (STEC), enterotoxigenic E.coli (ETEC), enteropathogenic E.coli (EPEC), enteroaggregative E.coli (EAEC), and enteroinvasive E.coli (EIEC). The objective of the study was to characterize the isolates of diarrheagenic E.coli from an outpatient pediatric population with diarrhea attended in two public hospitals from Buenos Aires, Argentina. Diarrheagenic E.coli pathotypes were investigated by amplifying characteristic virulence gene fragments: intimin (eae), heat-labile toxin (lt), heat-stable toxins (stp, sth), invasion plasmid antigen H (ipaH), transcriptional activator R (aggR) and Shiga toxins (stx1, stx2). Molecular subtyping of isolates was performed using PFGE (XbaI). Diarrheagenic E.coli was detected in 14% (84/601) of cases. The EAEC pathotype was prevalent, while ETEC, STEC, EPEC and EIEC were found in a lower proportion. EAEC isolates exhibited a high degree of genetic diversity. All pathotypes were found in children under 5years of age, while only EAEC, EIEC and ETEC were detected in the older population. Future studies that include the characterization of isolates from a greater number of genes and populations from other geographical areas will be necessary to determine the relevance of diarrheagenic E.coli in Argentina.

Detection of Escherichia coli and other Enterobacteriales members in seabirds sampled along the Brazilian coast.

Escherichia coli, an Enterobacterales member, is a normal representative of the microbiota of homeothermic animals. Most strains are commensal, but several pathotypes can cause disease, and numerous antimicrobial resistance factors have been identified. These bacteria have spread rapidly in recent years, highlighting the importance of screening the environment and non-human reservoirs for virulent strains and/or those presenting resistance factors, in addition to other microorganisms of public health importance. In this context, this study aimed to survey Enterobacteriales present in seabirds sampled along the Brazilian coast, comparing findings between migratory and resident birds, as well as between wrecked and non-wrecked animals. Escherichia coli pathotypes were also characterized through rapid seroagglutination and polymerase chain reaction techniques and antimicrobial resistance profiles were investigated through the disc agar diffusion method. Cloacal, ocular, oral, tracheal, and skin lesion swabs, as well as fresh feces, were collected from 122 seabirds. The findings indicate these animals as important hosts for opportunistic human pathogens. Escherichia coli strains were identified in 70 % of the analyzed seabirds, 62 % of which displaying resistant or intermediate profiles to at least one antimicrobial, while 7% were multiresistant. Resistance to tetracycline (22 %), nalidixic acid (15 %), trimethoprim-sulfamethozaxol (14 %) and ampicillin (12 %) were the most prevalent. Resistance to cefoxitin, a critically important antimicrobial for human medicine, was also detected. Virulence genes for one of the EAEC, ETEC or EPEC pathotypes were detected in 30 % of the identified strains, the first two described in seabirds for the first time. The EAEC gene was detected in 25 % of the sampled seabirds, all resident, 8 % of which exhibited a multidrug-resistant profile. Thus, seabirds comprise important reservoirs for this pathotype. Escherichia coli was proven an ubiquitous and well-distributed bacterium, present in all evaluated bird species and sampling sites (except Marajó Island). According to the chi-square test, no significant differences between E. coli prevalences or antimicrobial resistance profiles between migratory and resident and between wrecked and non-wrecked seabirds were observed. Thus, migratory birds do not seem to contribute significantly to E. coli frequencies, pathotypes or antimicrobial resistance rates on the Brazilian coast.

Transposon mutagenesis of atypical enteroaggregative Escherichia coli reveals a hemagglutinin-associated protein that mediates cell adhesion and contributes to the Galleria mellonella virulence.

Twenty-two atypical enteroaggregative Escherichia coli isolates from a previous epidemiological study harboring EAEC virulence genes were examined for their adhesion properties. Nine strains showed a typical aggregative adherence (AA) pattern, while 13 strains showed variant AA, such as AA with lined up cells characteristic of the chain-like adhesion (CLA) and AA mainly to HeLa cells characteristic of the diffuse adherence (DA). The aggregative forming pilus (AFP) genes afpA2 and afpR were detected only in strain Q015B, which exhibited an AA/DA pattern. Using Tn5-based transposon mutagenesis on Q015B strain, we identified a 5517-bp open reading frame (ORF) encoding a predicted 1838-amino-acid polypeptide that is genetically related to a putative filamentous hemagglutinin identified in E. coli strain 7-233-03_S3_C2. Therefore, the ORF was named orfHA. The regions flanking orfHA were sequenced and two ORFs were found; upstream, an ORF that encodes a 603-amino-acid polypeptide with 99% identity to hemolysin secretion/activation proteins of the ShlB/FhaC/HecB family, and downstream, another ORF, which encodes a 632-amino-acid polypeptide with 72% identity to the glycosyltransferase EtpC. An orfHA mutant (Q015BΔorfHA) was constructed from strain Q015B. Q015BΔorfHA strain did not adhere to HeLa cells, whereas Q015BΔ orfHA transformed with a pACYC184 plasmid carrying orfHA restored the AA/DA phenotype of strain Q015B. Furthermore, the Q015ΔorfHA mutant had a marked effect on the ability of strain Q015B to kill the larvae of Galleria mellonella. Our results suggest that the AA/DA pattern of strain Q015B is mediated by a hemagglutinin-associated protein which also contributes to its virulence in the G. mellonella model.

Boundaries That Prevent or May Lead Animals to be Reservoirs of Escherichia coli O104:H4.

Escherichia coli O104:H4, a hybrid serotype carrying virulence factors from enteroaggregative (EAEC) and Shiga toxin-producing (STEC) pathotypes, is the reported cause of a multicountry outbreak in 2011. Evaluation of potential routes of human contamination revealed that this strain is a foodborne pathogen. In contrast to STEC strains, whose main reservoir is cattle, serotype O104:H4 has not been commonly isolated from animals or related environments, suggesting an inability to naturally colonize the gut in hosts other than humans. However, contrary to this view, this strain has been shown to colonize the intestines of experimental animals in infectious studies. In this minireview, we provide a systematic summary of reports highlighting potential evolutionary changes that could facilitate the colonization of new reservoirs by these bacteria.

The aggregate-forming pili (AFP) mediates the aggregative adherence of a hybrid-pathogenic Escherichia coli (UPEC/EAEC) isolated from a urinary tract infection.

Enteroaggregative Escherichia coli (EAEC) comprises an important diarrheagenic pathotype, while uropathogenic E. coli (UPEC) is the most important agent of urinary tract infection (UTI). Recently, EAEC virulence factors have been detected in E. coli strains causing UTI, showing the importance of these hybrid-pathogenic strains. Previously, we detected an E. coli strain isolated from UTI (UPEC-46) presenting characteristics of EAEC, e.g., the aggregative adherence (AA) pattern and EAEC-associated genes (aatA, aap, and pet). In this current study, we analyzed the whole genomic sequence of UPEC-46 and characterized some phenotypic traits. The AA phenotype was observed in cell lineages of urinary and intestinal origin. The production of curli, cellulose, bacteriocins, and Pet toxin was detected. Additionally, UPEC-46 was not capable of forming biofilm using different culture media and human urine. The genome sequence analysis showed that this strain belongs to serotype O166:H12, ST10, and phylogroup A, harbors the tet, aadA, and dfrA/sul resistance genes, and is phylogenetically more related to EAEC strains isolated from human feces. UPEC-46 harbors three plasmids. Plasmid p46-1 (~135 kb) carries some EAEC marker genes and those encoding the aggregate-forming pili (AFP) and its regulator (afpR). A mutation in afpA (encoding the AFP major pilin) led to the loss of pilin production and assembly, and notably, a strongly reduced adhesion to epithelial cells. In summary, the genetic background and phenotypic traits analyzed suggest that UPEC-46 is a hybrid strain (UPEC/EAEC) and highlights the importance of AFP adhesin in the adherence to colorectal and bladder cell lines.

Identification of a hybrid atypical enteropathogenic and enteroaggregative Escherichia coli (aEPEC/EAEC) clone of serotype O3:H2 associated with a diarrheal outbreak in Brazil.

Enteropathogenic (EPEC) and enteroaggregative (EAEC) Escherichia coli are two of the major pathotypes of diarrheagenic E. coli causing disease worldwide. Here, we report a diarrheal outbreak caused by E. coli of serotype O3:H2, harboring virulence markers from EPEC (eae) and/or EAEC (aggR). This is likely the first E. coli diarrheal outbreak caused by a hybrid atypical-EPEC/EAEC clone reported in Brazil.

Human microbiota modulation via QseC sensor kinase mediated in the Escherichia coli O104:H4 outbreak strain infection in microbiome model.

BACKGROUND: The intestinal microbiota plays a crucial role in human health, adjusting its composition and the microbial metabolites protects the gut against invading microorganisms. Enteroaggregative E. coli (EAEC) is an important diarrheagenic pathogen, which may cause acute or persistent diarrhea (≥14 days). The outbreak strain has the potent Shiga toxin, forms a dense biofilm and communicate via QseBC two-component system regulating the expression of many important virulence factors. RESULTS: Herein, we investigated the QseC histidine sensor kinase role in the microbiota shift during O104:H4 C227-11 infection in the colonic model SHIME® (Simulator of the Human Intestinal Microbial Ecosystem) and in vivo mice model. The microbiota imbalance caused by C227-11 infection affected ỿ-Proteobacteria and Lactobacillus spp. predominance, with direct alteration in intestinal metabolites driven by microbiota change, such as Short-chain fatty acids (SCFA). However, in the absence of QseC sensor kinase, the microbiota recovery was delayed on day 3 p.i., with change in the intestinal production of SCFA, like an increase in acetate production. The higher predominance of Lactobacillus spp. in the microbiota and significant augmented qseC gene expression levels were also observed during C227-11 mice infection upon intestinal depletion. Novel insights during pathogenic bacteria infection with the intestinal microbiota were observed. The QseC kinase sensor seems to have a role in the microbiota shift during the infectious process by Shiga toxin-producing EAEC C227-11. CONCLUSIONS: The QseC role in C227-11 infection helps to unravel the intestine microbiota modulation and its metabolites during SHIME® and in vivo models, besides they contribute to elucidate bacterial intestinal pathogenesis and the microbiota relationships.

The aggregate-forming pili (AFP) mediates the aggregative adherence of a hybrid-pathogenic Escherichia coli (UPEC/EAEC) isolated from a urinary tract infection

Enteroaggregative Escherichia coli (EAEC) comprises an important diarrheagenic pathotype, while uropathogenic E. coli (UPEC) is the most important agent of urinary tract infection (UTI). Recently, EAEC virulence factors have been detected in E. coli strains causing UTI, showing the importance of these hybrid-pathogenic strains. Previously, we detected an E. coli strain isolated from UTI (UPEC-46) presenting characteristics of EAEC, e.g., the aggregative adherence (AA) pattern and EAEC-associated genes (aatA, aap, and pet). In this current study, we analyzed the whole genomic sequence of UPEC-46 and characterized some phenotypic traits. The AA phenotype was observed in cell lineages of urinary and intestinal origin. The production of curli, cellulose, bacteriocins, and Pet toxin was detected. Additionally, UPEC-46 was not capable of forming biofilm using different culture media and human urine. The genome sequence analysis showed that this strain belongs to serotype O166:H12, ST10, and phylogroup A, harbors the tet, aadA, and dfrA/sul resistance genes, and is phylogenetically more related to EAEC strains isolated from human feces. UPEC-46 harbors three plasmids. Plasmid p46-1 (~135 kb) carries some EAEC marker genes and those encoding the aggregate-forming pili (AFP) and its regulator (afpR). A mutation in afpA (encoding the AFP major pilin) led to the loss of pilin production and assembly, and notably, a strongly reduced adhesion to epithelial cells. In summary, the genetic background and phenotypic traits analyzed suggest that UPEC-46 is a hybrid strain (UPEC/EAEC) and highlights the importance of AFP adhesin in the adherence to colorectal and bladder cell lines.

Human microbiota modulation via QseC sensor kinase mediated in the Escherichia coli O104:H4 outbreak strain infection in microbiome model

Background: The intestinal microbiota plays a crucial role in human health, adjusting its composition and the microbial metabolites protects the gut against invading microorganisms. Enteroaggregative E. coli (EAEC) is an important diarrheagenic pathogen, which may cause acute or persistent diarrhea (≥14 days). The outbreak strain has the potent Shiga toxin, forms a dense biofilm and communicate via QseBC two-component system regulating the expression of many important virulence factors. Results: Here in, we investigated the QseC histidine sensor kinase role in the microbiota shift during O104:H4 C227–11 infection in the colonic model SHIME® (Simulator of the Human Intestinal Microbial Ecosystem) and in vivo mice model. The microbiota imbalance caused by C227–11 infection affected ỿ-Proteobacteria and Lactobacillus spp. predominance, with direct alteration in intestinal metabolites driven by microbiota change, such as Short-chain fatty acids (SCFA). However, in the absence of QseC sensor kinase, the microbiota recovery was delayed on day 3 p.i., with change in the intestinal production of SCFA, like an increase in acetate production. The higher predominance of Lactobacillus spp. in the microbiota and significant augmented qseC gene expression levels were also observed during C227–11 mice infection upon intestinal depletion. Novel insights during pathogenic bacteria infection with the intestinal microbiota were observed. The QseC kinase sensor seems to have a role in the microbiota shift during the infectious process by Shiga toxin-producing EAEC C227–11. Conclusions: The QseC role in C227–11 infection helps to unravel the intestine microbiota modulation and its metabolites during SHIME® and in vivo models, besides they contribute to elucidate bacterial intestinal pathogenesis and the microbiota relationships.

New Insights Into DAEC and EAEC Pathogenesis and Phylogeny.

Diarrheagenic E. coli can be separated into six distinct pathotypes, with enteroaggregative (EAEC) and diffusely-adherent E. coli (DAEC) among the least characterized. To gain additional insights into these two pathotypes we performed whole genome sequencing of ten DAEC, nine EAEC strains, isolated from Mexican children with diarrhea, and one EAEC plus one commensal E. coli strains isolated from an adult with diarrhea and a healthy child, respectively. These genome sequences were compared to 85 E. coli genomes available in public databases. The EAEC and DAEC strains segregated into multiple different clades; however, six clades were heavily or exclusively comprised of EAEC and DAEC strains, suggesting a phylogenetic relationship between these two pathotypes. EAEC strains harbored the typical virulence factors under control of the activator AggR, but also several toxins, bacteriocins, and other virulence factors. DAEC strains harbored several iron-scavenging systems, toxins, adhesins, and complement resistance or Immune system evasion factors that suggest a pathogenic paradigm for this poorly understood pathotype. Several virulence factors for both EAEC and DAEC were associated with clinical presentations, not only suggesting the importance of these factors, but also potentially indicating opportunities for intervention. Our studies provide new insights into two distinct but related diarrheagenic organisms.

Surface Protein Dispersin of Enteroaggregative Escherichia coli Binds Plasminogen That Is Converted Into Active Plasmin.

Dispersin is a 10.2 kDa-immunogenic protein secreted by enteroaggregative Escherichia coli (EAEC). In the prototypical EAEC strain 042, dispersin is non-covalently bound to the outer membrane, assisting dispersion across the intestinal mucosa by overcoming electrostatic attraction between the AAF/II fimbriae and the bacterial surface. Also, dispersin facilitates penetration of the intestinal mucus layer. Initially characterized in EAEC, dispersin has been detected in other E. coli pathotypes, including those isolated from extraintestinal sites. In this study we investigated the binding capacity of purified dispersin to extracellular matrix (ECM), since dispersin is exposed on the bacterial surface and is involved in intestinal colonization. Binding to plasminogen was also investigated due to the presence of conserved carboxy-terminal lysine residues in dispersin sequences, which are involved in plasminogen binding in several bacterial proteins. Moreover, some E. coli components can interact with this host protease, as well as with tissue plasminogen activator, leading to plasmin production. Recombinant dispersin was produced and used in binding assays with ECM molecules and coagulation cascade compounds. Purified dispersin bound specifically to laminin and plasminogen. Interaction with plasminogen occurred in a dose-dependent and saturable manner. In the presence of plasminogen activator, bound plasminogen was converted into plasmin, its active form, leading to fibrinogen and vitronectin cleavage. A collection of E. coli strains isolated from human bacteremia was screened for the presence of aap, the dispersin-encoding gene. Eight aap-positive strains were detected and dispersin production could be observed in four of them. Our data describe new attributes for dispersin and points out to possible roles in mechanisms of tissue adhesion and dissemination, considering the binding capacity to laminin, and the generation of dispersin-bound plasmin(ogen), which may facilitate E. coli spread from the colonization site to other tissues and organs. The cleavage of fibrinogen in the bloodstream, may also contribute to the pathogenesis of sepsis caused by dispersin-producing E. coli.

Analysis of the Virulence Profile and Phenotypic Features of Typical and Atypical Enteroaggregative Escherichia coli (EAEC) Isolated From Diarrheal Patients in Brazil.

Enteroaggregative Escherichia coli (EAEC) is an important agent of acute and persistent diarrhea in children and adults worldwide. Here we report a characterization of 220 EAEC isolates, 88.2% (194/220) of which were typical and 11.8% (26/220) were atypical, obtained from diarrheal patients during seven years (2010-2016) of epidemiological surveillance in Brazil. The majority of the isolates were assigned to phylogroups A (44.1%, 97/220) or B1 (21.4%, 47/220). The aggregative adherence (AA) pattern was detected in 92.7% (204/220) of the isolates, with six of them exhibiting AA concomitantly with a chain-like adherence pattern; and agg5A and agg4A were the most common adhesin-encoding genes, which were equally detected in 14.5% (32/220) of the isolates. Each of 12 virulence factor-encoding genes (agg4A, agg5A, pic, aap, aaiA, aaiC, aaiG, orf3, aar, air, capU, and shf) were statistically associated with typical EAEC (P < 0.05). The genes encoding the newly described aggregate-forming pili (AFP) searched (afpB, afpD, afpP, and afpA2), and/or its regulator (afpR), were exclusively detected in atypical EAEC (57.7%, 15/26), and showed a significant association with this subgroup of EAEC (P < 0.001). In conclusion, we presented an extensive characterization of the EAEC circulating in the Brazilian settings and identified the afp genes as putative markers for increasing the efficiency of atypical EAEC diagnosis.

An Stx-EAEC O59:NM[H19] strain isolated from a hemolytic uremic syndrome case in Argentina / Cepa EAEC-Stx O59:NM[H19] aislada de un caso de síndrome urémico hemolítico en Argentina

Abstract Shiga toxin-producing Escherichia coli (STEC) are a heterogeneous group of foodborne pathogens causing a broad spectrum of human disease, from uncomplicated diarrhea to hemolytic uremic syndrome (HUS). In this study, we report an HUS case associated with an O59:NM H19 mstrain, harboring stx2a, iha, lpfAO26, lpfAO113 genes associated with STEC, and aatA, aap, pic, sigA, agg4A genes associated with enteroaggregative E. coli (EAEC), named Stx-EAEC. The strain showed low toxicity on Vero cells, and was resistant to streptomycin and trimethoprim/sulfonamides. The child carried the bacteria for more than 100 days. Since the large outbreak associated with Stx-EAEC O104:H4, many strains with similar profiles have been described. In Germany, an O59:NM[H19] strain, with comparable characteristics to the Argentine strain, was isolated from a bloody diarrhea case. In Argentina, this is the first report of an HUS case associated with a Stx-EAEC infection, and represents a new challenge for the surveillance system. © 2019 Published by Elsevier Espana, S.L.U. on behalf of Asociacion Argentina de Microbiolog´a. This is an open access article under the CC BY-NC-ND license (https://creativecommons.org/ licenses/by-nc-nd/4.0/).

Resumen Escherichia coli productor de la toxina Shiga (STEC) es un grupo heterogéneo de patógenos transmitidos por alimentos que causan un amplio espectro de enfermedades humanas, desde diarrea no complicada hasta síndrome urémico hemolítico (SUH). Nosotros informamos de un caso de SUH por O59:NM[H19], que portaba los genes stx2a, iha, lpfAo26, lpfAoii3 asociados con STEC, y los genes aatA, aap, pic, sigA, agg4A de E. coli enteroagregativo (EAEC), llamado EAEC-Stx. La cepa mostró baja citotoxicidad en las células Vero, y fue resistente a estreptomicina y trimetoprima/sulfonamidas. El niño excretó la bacteria durante más de 100 días. Desde el brote asociado con EAEC-Stx O104:H4, se describieron muchas cepas con perfiles similares. En Alemania se aisló una cepa O59:NM[H19] de una diarrea sanguinolenta, con características comparables a la cepa argentina. Este es el primer informe de un caso de SUH asociado a una infección por EAEC-Stx, y representa un nuevo desafío para el sistema de vigilancia. © 2019 Publicado por Elsevier Espana, S.L.U. en nombre de Asociación Argentina de Microbiología. Este es un artículo Open Access bajo la licencia CC BY-NC-ND (http://creativecommons. org/licenses/by-nc-nd/4.0/).

An Stx-EAEC O59:NM[H19] strain isolated from a hemolytic uremic syndrome case in Argentina.

Shiga toxin-producing Escherichia coli (STEC) are a heterogeneous group of foodborne pathogens causing a broad spectrum of human disease, from uncomplicated diarrhea to hemolytic uremic syndrome (HUS). In this study, we report an HUS case associated with an O59:NM[H19] strain, harboring stx2a, iha, lpfAO26, lpfAO113 genes associated with STEC, and aatA, aap, pic, sigA, agg4A genes associated with enteroaggregative E. coli (EAEC), named Stx-EAEC. The strain showed low toxicity on Vero cells, and was resistant to streptomycin and trimethoprim/sulfonamides. The child carried the bacteria for more than 100 days. Since the large outbreak associated with Stx-EAEC O104:H4, many strains with similar profiles have been described. In Germany, an O59:NM[H19] strain, with comparable characteristics to the Argentine strain, was isolated from a bloody diarrhea case. In Argentina, this is the first report of an HUS case associated with a Stx-EAEC infection, and represents a new challenge for the surveillance system.

Surface protein dispersin of enteroaggregative escherichia coli binds plasminogen that Is converted Into active plasmin

Dispersin is a 10.2 kDa-immunogenic protein secreted by enteroaggregative Escherichia coli (EAEC). In the prototypical EAEC strain 042, dispersin is non-covalently bound to the outer membrane, assisting dispersion across the intestinal mucosa by overcoming electrostatic attraction between the AAF/II fimbriae and the bacterial surface. Also, dispersin facilitates penetration of the intestinal mucus layer. Initially characterized in EAEC, dispersin has been detected in other E. coli pathotypes, including those isolated from extraintestinal sites. In this study we investigated the binding capacity of purified dispersin to extracellular matrix (ECM), since dispersin is exposed on the bacterial surface and is involved in intestinal colonization. Binding to plasminogen was also investigated due to the presence of conserved carboxy-terminal lysine residues in dispersin sequences, which are involved in plasminogen binding in several bacterial proteins. Moreover, some E. coli components can interact with this host protease, as well as with tissue plasminogen activator, leading to plasmin production. Recombinant dispersin was produced and used in binding assays with ECM molecules and coagulation cascade compounds. Purified dispersin bound specifically to laminin and plasminogen. Interaction with plasminogen occurred in a dose-dependent and saturable manner. In the presence of plasminogen activator, bound plasminogen was converted into plasmin, its active form, leading to fibrinogen and vitronectin cleavage. A collection of E. coli strains isolated from human bacteremia was screened for the presence of aap, the dispersin-encoding gene. Eight aap-positive strains were detected and dispersin production could be observed in four of them. Our data describe new attributes for dispersin and points out to possible roles in mechanisms of tissue adhesion and dissemination, considering the binding capacity to laminin, and the generation of dispersin-bound plasmin(ogen), which may facilitate E. coli spread from the colonization site to other tissues and organs. The cleavage of fibrinogen in the bloodstream, may also contribute to the pathogenesis of sepsis caused by dispersin-producing E. coli.