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Hybrid-pathogenic Escherichia coli represent an important group of strains associated with intestinal and extraintestinal infections. Recently, we described strain UPEC-46, a uropathogenic/enteroaggregative E. coli (UPEC/EAEC) strain presenting the aggregative adherence (AA) pattern on bladder and colorectal epithelial cells mediated by aggregate-forming pili (AFP). However, the role of AFP and other uninvestigated putative fimbriae operons in UPEC-46 pathogenesis remains unclear. Thus, this study evaluated the involvement of AFP and other adhesins in uropathogenicity and intestinal colonization using different in vitro and in vivo models. The strain UPEC-46 was able to adhere and invade intestinal and urinary cell lines. A library of transposon mutants also identified the involvement of type I fimbriae (TIF) in the adherence to HeLa cells, in addition to colorectal and bladder cell lines. The streptomycin-treated mouse in vivo model also showed an increased number of bacterial counts in the colon in the presence of AFP and TIF. In the mouse model of ascending urinary tract infection (UTI), AFP was more associated with kidney colonization, while TIF appears to mediate bladder colonization. Results observed in in vivo experiments were also confirmed by electron microscopy (EM) analyses. In summary, the in vitro and in vivo analyses show a synergistic role of AFP and TIF in the adherence and colonization of intestinal and urinary epithelia. Therefore, we propose that hybrid E. coli strains carrying AFP and TIF could potentially cause intestinal and urinary tract infections in the same patient.
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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.
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Background: During the COVID-19 pandemic, universities all over the world have had to reorganize their work for remote education to ensure continuity of learning. This situation has forced both teachers and students into an atypical, very specific situation, in which they need to cope with a number of psychological factors. Meanwhile, there is a research gap in academic knowledge of the social and psychological factors that determine attitudes toward forced remote learning. Objective: To analyze the psychological traits of students in relation to their attitudes toward forced remote learning. Methods: The study assessed self-regulation and communication skills, as well thinking and learning styles of 280 students in the situation of forced remote learning. We used the methods of surveying and psychological testing for data collection. The data were analyzed in R statistical software using the regression modeling methods. Results: We found that the number of students with positive (46%) and negative (54%) attitudes toward forced remote learning during the COVID-19 pandemic was approximately equal. Using regression analysis, we identified several statistically significant associations between the interpersonal communicative skills of students (self-regulation, shyness, alienation, manipulative and cooperative communication styles) and their thinking styles (right-hemispheric and integrated), on the one hand, and their attitude to remote learning, on the other. It was also illustrated that depending on the attitude to the forced remote learning, students differ by the percentage of assimilated learning materials while studying under the lockdown. Conclusion: The results identify that success of remote learning in many ways depends on the extent to which it accommodates psychological traits of students who are forced to acquire new knowledge without traditional classroom instruction.
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Escherichia coli EC121 is a multidrug-resistant (MDR) strain isolated from a bloodstream infection of an inpatient with persistent gastroenteritis and T-zone lymphoma that died due to septic shock. Despite causing an extraintestinal infection, previous studies showed that it did not have the usual characteristics of an extraintestinal pathogenic E. coli. Instead, it belonged to phylogenetic group B1 and harbored few known virulence genes. To evaluate the pathogenic potential of strain EC121, an extensive genome sequencing and in vitro characterization of various pathogenicity-associated properties were performed. The genomic analysis showed that strain EC121 harbors more than 50 complete virulence genetic clusters. It also displays the capacity to adhere to a variety of epithelial cell lineages and invade T24 bladder cells, as well as the ability to form biofilms on abiotic surfaces, and survive the bactericidal serum complement activity. Additionally, EC121 was shown to be virulent in the Galleria mellonella model. Furthermore, EC121 is an MDR strain harboring 14 antimicrobial resistance genes, including blaCTX-M-2. Completing the scenario, it belongs to serotype O154:H25 and to sequence type 101-B1, which has been epidemiologically linked to extraintestinal infections as well as to antimicrobial resistance spread. This study with E. coli strain EC121 shows that clinical isolates considered opportunistic might be true pathogens that go underestimated.
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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.
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Diarrhea is one of the main causes of infant mortality worldwide, mainly in the developing world. Among the various etiologic agents, Escherichia albertii is emerging as an important human enteropathogen. E. albertii promote attaching and effacing (AE) lesions due to the presence of the locus of enterocyte effacement (LEE) that encodes a type three secretion system (T3SS), the afimbrial adhesin intimin and its translocated receptor, Tir, and several effector proteins. We previously showed that E. albertii strain 1551-2 invades several epithelial cell lineages by a process that is dependent on the intimin-Tir interaction. To understand the contribution of T3SS-dependent effectors present in E. albertii 1551-2 during the invasion process, we performed a genetic analysis of the LEE and non-LEE genes and evaluated the expression of the LEE operons in various stages of bacterial interaction with differentiated intestinal Caco-2 cells. The kinetics of the ability of the 1551-2 strain to colonize and form AE lesions was also investigated in epithelial HeLa cells. We showed that the LEE expression was constant during the early stages of infection but increased at least 4-fold during bacterial persistence in the intracellular compartment. An in silico analysis indicated the presence of a new tccP/espFU subtype, named tccP3. We found that the encoded protein colocalizes with Tir and polymerized F-actin during the infection process in vitro. Moreover, assays performed with Nck null cells demonstrated that the 1551-2 strain can trigger F-actin polymerization in an Nck-independent pathway, despite the fact that TccP3 is not required for this phenotype. Our study highlights the importance of the T3SS during the invasion process and for the maintenance of E. albertii 1551-2 inside the cells. In addition, this work may help to elucidate the versatility of the T3SS for AE pathogens, which are usually considered extracellular and rarely reach the intracellular environment.
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Hybrid strains of Escherichia coli combine virulence traits of diarrheagenic (DEC) and extraintestinal pathogenic E. coli (ExPEC), but it is poorly understood whether these combined features improve the virulence potential of such strains. We have previously identified a uropathogenic E. coli (UPEC) strain (UPEC 252) harboring the eae gene that encodes the adhesin intimin and is located in the locus of enterocyte effacement (LEE) pathogenicity island. The LEE-encoded proteins allow enteropathogenic E. coli (EPEC) and enterohemorrhagic E. coli (EHEC) to form attaching and effacing (A/E) lesions in enterocytes. We sought to characterize UPEC 252 through whole-genome sequencing and phenotypic virulence assays. Genome analysis unveiled that this strain harbors a complete LEE region, with more than 97% of identity comparing to E2348/69 (EPEC) and O157:H7 Sakai (EHEC) prototype strains, which was functional, since UPEC 252 expressed the LEE-encoded proteins EspB and intimin and induced actin accumulation foci in HeLa cells. Phylogenetic analysis performed comparing 1,000 single-copy shared genes clustered UPEC 252 with atypical EPEC strains that belong to the sequence type 10, phylogroup A. Additionally, UPEC 252 was resistant to the bactericidal power of human serum and colonized cells of the urinary (T24 and HEK293-T) and intestinal (Caco-2 and LS174T) tracts. Our findings suggest that UPEC 252 is an atypical EPEC strain that emerges as a hybrid strain (aEPEC/UPEC), which could colonize new niches and potentially cause intestinal and extraintestinal infections.
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Escherichia coli EC121 is a multidrug-resistant (MDR) strain isolated from a bloodstream infection of an inpatient with persistent gastroenteritis and T-zone lymphoma that died due to septic shock. Despite causing an extraintestinal infection, previous studies showed that it did not have the usual characteristics of an extraintestinal pathogenic E. coli. Instead, it belonged to phylogenetic group B1 and harbored few known virulence genes. To evaluate the pathogenic potential of strain EC121, an extensive genome sequencing and in vitro characterization of various pathogenicity-associated properties were performed. The genomic analysis showed that strain EC121 harbors more than 50 complete virulence genetic clusters. It also displays the capacity to adhere to a variety of epithelial cell lineages and invade T24 bladder cells, as well as the ability to form biofilms on abiotic surfaces, and survive the bactericidal serum complement activity. Additionally, EC121 was shown to be virulent in the Galleria mellonella model. Furthermore, EC121 is an MDR strain harboring 14 antimicrobial resistance genes, including blaCTX-M-2. Completing the scenario, it belongs to serotype O154:H25 and to sequence type 101-B1, which has been epidemiologically linked to extraintestinal infections as well as to antimicrobial resistance spread. This study with E. coli strain EC121 shows that clinical isolates considered opportunistic might be true pathogens that go underestimated.
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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.
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The intimin protein is the major adhesin involved in the intimate adherence of atypicalenteropathogenicEscherichia coli(aEPEC) strains to epithelial cells, but little is knownabout the structures involved in their early colonization process. A previous studydemonstrated that the type III secretion system (T3SS) plays an additional role in theadherence of anEscherichia albertiistrain. Therefore, we assumed that the T3SS couldbe related to the adherence efficiency of aEPEC during the first stages of contactwith epithelial cells. To test this hypothesis, we examined the adherence of sevenaEPEC strains and theireae(intimin) isogenic mutants in the standard HeLa adherenceassay and observed that all wild-type strains were adherent while five isogeniceaemutants were not. The twoeaemutant strains that remained adherent were then usedto generate theeae/escNdouble mutants (encoding intimin and the T3SS ATPase,respectively) and after the adherence assay, we observed that one strain lost itsadherence capacity. This suggested a role for the T3SS in the initial adherence stepsof this strain. In addition, we demonstrated that this strain expressed the T3SS atsignificantly higher levels when compared to the other wild-type strains and that itproduced longer translocon-filaments. Our findings reveal that the T3SS-transloconcan play an additional role as an adhesin at the beginning of the colonization processof aEPEC.
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Typical enteropathogenic Escherichia coli strains (tEPEC) cause attaching/effacing lesions in eukaryotic cells and produce the bundle-forming pilus (BFP), which interweaves and aggregates bacteria, resulting in the localized adherence (LA) pattern on eukaryotic cells. Previously, we identified tEPEC strains (serotype O119:H6) that exhibited LA simultaneously with an aggregative adherence (AA)-like pattern (LA/AA-like+). Remarkably, AA is characteristically produced by strains of enteroaggregative E. coli (EAEC), another diarrheagenic E. coli pathovar. In one LA/AA-like?+?strain (Ec404/03), we identified a conjugative plasmid containing the pil operon, which encodes the Pil fimbriae. Moreover, a pil operon associated with an AA pattern and plasmid transfer had been previously described in the EAEC C1096 strain. In this study, we investigated the occurrence of the two pilS alleles (pilSEc404 and pilSC1096) in tEPEC strains of different serotypes, origins and years of isolation. We also examined the potential relationship of pilS with the AA-like phenotype, its ability to be transferred by conjugation, and occurrence among strains of the other E. coli pathovars. The pilS alleles were found in 90 (55.2%) of 163 tEPEC strains, with pilSEc404 occurring more often (30.7%) than pilSC1096 (25.1%). About 21 tEPEC serotypes carried pilS. The pilS alleles were found in tEPEC strains from Chile, Peru and different Brazilian cities, with the oldest strain being isolated in 1966. No absolute correlation was found between the presence of pilS and the AA-like pattern. Conjugative pilS transfer was detected in 26.2% of pilSEc404+ strains and in 65.1% of pilSC1096+ strains, but only pilSEc404+ transconjugants were AA-like+, thus suggesting that the latter allele might need a different genetic background to express this phenotype. pilS was found in all other E. coli pathovars, where it was most prevalent in enterotoxigenic E. coli. More studies are needed to understand the mechanisms involved in the regulation of Pil expression and production.
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The intimin protein is the major adhesin involved in the intimate adherence of atypicalenteropathogenicEscherichia coli(aEPEC) strains to epithelial cells, but little is knownabout the structures involved in their early colonization process. A previous studydemonstrated that the type III secretion system (T3SS) plays an additional role in theadherence of anEscherichia albertiistrain. Therefore, we assumed that the T3SS couldbe related to the adherence efficiency of aEPEC during the first stages of contactwith epithelial cells. To test this hypothesis, we examined the adherence of sevenaEPEC strains and theireae(intimin) isogenic mutants in the standard HeLa adherenceassay and observed that all wild-type strains were adherent while five isogeniceaemutants were not. The twoeaemutant strains that remained adherent were then usedto generate theeae/escNdouble mutants (encoding intimin and the T3SS ATPase,respectively) and after the adherence assay, we observed that one strain lost itsadherence capacity. This suggested a role for the T3SS in the initial adherence stepsof this strain. In addition, we demonstrated that this strain expressed the T3SS atsignificantly higher levels when compared to the other wild-type strains and that itproduced longer translocon-filaments. Our findings reveal that the T3SS-transloconcan play an additional role as an adhesin at the beginning of the colonization processof aEPEC.
RESUMO
Typical enteropathogenic Escherichia coli strains (tEPEC) cause attaching/effacing lesions in eukaryotic cells and produce the bundle-forming pilus (BFP), which interweaves and aggregates bacteria, resulting in the localized adherence (LA) pattern on eukaryotic cells. Previously, we identified tEPEC strains (serotype O119:H6) that exhibited LA simultaneously with an aggregative adherence (AA)-like pattern (LA/AA-like+). Remarkably, AA is characteristically produced by strains of enteroaggregative E. coli (EAEC), another diarrheagenic E. coli pathovar. In one LA/AA-like?+?strain (Ec404/03), we identified a conjugative plasmid containing the pil operon, which encodes the Pil fimbriae. Moreover, a pil operon associated with an AA pattern and plasmid transfer had been previously described in the EAEC C1096 strain. In this study, we investigated the occurrence of the two pilS alleles (pilSEc404 and pilSC1096) in tEPEC strains of different serotypes, origins and years of isolation. We also examined the potential relationship of pilS with the AA-like phenotype, its ability to be transferred by conjugation, and occurrence among strains of the other E. coli pathovars. The pilS alleles were found in 90 (55.2%) of 163 tEPEC strains, with pilSEc404 occurring more often (30.7%) than pilSC1096 (25.1%). About 21 tEPEC serotypes carried pilS. The pilS alleles were found in tEPEC strains from Chile, Peru and different Brazilian cities, with the oldest strain being isolated in 1966. No absolute correlation was found between the presence of pilS and the AA-like pattern. Conjugative pilS transfer was detected in 26.2% of pilSEc404+ strains and in 65.1% of pilSC1096+ strains, but only pilSEc404+ transconjugants were AA-like+, thus suggesting that the latter allele might need a different genetic background to express this phenotype. pilS was found in all other E. coli pathovars, where it was most prevalent in enterotoxigenic E. coli. More studies are needed to understand the mechanisms involved in the regulation of Pil expression and production.
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The number of diarrhea cases caused by atypical enteropathogenic Escherichia coli (aEPEC) has been increasing worldwide. Here, we report the draft whole-genome sequences of 10 aEPEC strains isolated in Brazil. These sequences will provide an important source for future studies concerning aEPEC pathogenicity and genetic markers of potentially virulent strains.
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The number of diarrhea cases caused by atypical enteropathogenic Escherichia coli (aEPEC) has been increasing worldwide. Here, we report the draft whole-genome sequences of 10 aEPEC strains isolated in Brazil. These sequences will provide an important source for future studies concerning aEPEC pathogenicity and genetic markers of potentially virulent strains.
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Enteroaggregative Escherichia coli (EAEC) has been recently associated with urinary tract infections (UTI). Since EAEC are found in feces of both diarrheic and asymptomatic individuals, their presence in the intestine may be a source of UTI. In this study, we detected in feces of diarrheic and healthy children a subset of EAEC strains with genetic markers of extraintestinal pathogenic E. coli (ExPEC). MLST grouped these EAEC with ExPEC markers in three main clusters along with prototypes strains of EAEC, uropathogenic E. coli and UTI-causing EAEC. Interestingly, the latter cluster was composed by EAEC with ExPEC markers belonging to phylogroup A and closely related to the uropathogenic EAEC O78:H10 strain. Such attributes suggest that these strains have uropathogenic abilities. Therefore, intestinal carriers of these strains are potentially in risk to develop UTIs.
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Diarrhea is the second leading cause of death of children up to five years old in the developing countries. Among the etiological diarrheal agents are atypical enteropathogenic Escherichia coli (aEPEC), one of the diarrheagenic E. coli pathotypes that affects children and adults, even in developed countries. Currently, genotypic and biochemical approaches have helped to demonstrate that some strains classified as aEPEC are actually E. albertii, a recently recognized human enteropathogen. Studies on particular strains are necessary to explore their virulence potential in order to further understand the underlying mechanisms of E. albertii infections. Here we demonstrated for the first time that infection of fragments of rat intestinal mucosa is a useful tool to study the initial steps of E. albertii colonization. We also observed that an E. albertii strain can translocate from the intestinal lumen to Mesenteric Lymph Nodes and liver in a rat model. Based on our finding of bacterial translocation, we investigated how E. albertii might cross the intestinal epithelium by performing infections of M-like cells in vitro to identify the potential in vivo translocation route. Altogether, our approaches allowed us to draft a general E. albertii infection route from the colonization till the bacterial spreading in vivo.
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Enteroaggregative Escherichia coli (EAEC) is an agent of acute and persistent diarrhea worldwide, categorized in typical or atypical subgroups. Some EAEC virulence factors are members of the serine protease autotransporters of Enterobacteriaceae (SPATE). The presence of SPATE-encoding genes of different E. coli pathotypes was searched in a large collection of EAEC strains, and a possible association between SPATES and E. coil phylogroups was investigated. Among 108 typical and 85 atypical EAEC, pic was the most prevalent gene, detected in 47.1% of the strains, followed by sat (24.3%), espl (21.2%), pet (19.2%), sepA (13.5%), sigA (4.1%), eatA (4.1%), vat (1.0%), espP and tsh, detected in one strain (0.5%) each; while epeA and espC were not detected. Phylogenetic analysis demonstrated that 39.9% of the strains belonged to group A, 233% to B1, 10.9% to B2, 7.8% to D, 8.8% to E and 1.5% to F. The majority of the SPATE genes were distributed in typical and atypical strains without association with any phylogroup. In addition, pic and pet were strongly associated with typical EAEC and sepA was detected in close association with atypical EAEC. Our data indicate that SPATEs may represent important virulence traits in both subgroups of EAEC.
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Most Escherichia coli strains live harmlessly in the intestines and rarely cause disease in healthy individuals. Nonetheless, a number of pathogenic strains can cause diarrhea or extraintestinal diseases both in healthy and immunocompromised individuals. Diarrheal illnesses are a severe public health problem and a major cause of morbidity and mortality in infants and young children, especially in developing countries. E. coli strains that cause diarrhea have evolved by acquiring, through horizontal gene transfer, a particular set of characteristics that have successfully persisted in the host. According to the group of virulence determinants acquired, specific combinations were formed determining the currently known E. coli pathotypes, which are collectively known as diarrheagenic E. coli. In this review, we have gathered information on current definitions, serotypes, lineages, virulence mechanisms, epidemiology, and diagnosis of the major diarrheagenic E. coli pathotypes.
Assuntos
Diarreia/diagnóstico , Diarreia/epidemiologia , Escherichia coli/patogenicidade , Infecções por Escherichia coli/epidemiologiaRESUMO
ABSTRACT Most Escherichia coli strains live harmlessly in the intestines and rarely cause disease in healthy individuals. Nonetheless, a number of pathogenic strains can cause diarrhea or extraintestinal diseases both in healthy and immunocompromised individuals. Diarrheal illnesses are a severe public health problem and a major cause of morbidity and mortality in infants and young children, especially in developing countries. E. coli strains that cause diarrhea have evolved by acquiring, through horizontal gene transfer, a particular set of characteristics that have successfully persisted in the host. According to the group of virulence determinants acquired, specific combinations were formed determining the currently known E. coli pathotypes, which are collectively known as diarrheagenic E. coli. In this review, we have gathered information on current definitions, serotypes, lineages, virulence mechanisms, epidemiology, and diagnosis of the major diarrheagenic E. coli pathotypes.