RESUMO
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.
Assuntos
Infecções por Escherichia coli/microbiologia , Escherichia coli O104/metabolismo , Proteínas de Escherichia coli/metabolismo , Microbioma Gastrointestinal , Animais , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Modelos Animais de Doenças , Escherichia coli O104/genética , Proteínas de Escherichia coli/genética , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Fatores de Virulência/genética , Fatores de Virulência/metabolismoRESUMO
Los serogrupos O26, O45, O103, O104, O111, O121, O145 y O157 de STEC se relacionan con un elevado número de casos de SUH a nivel mundial, por lo que están incluidos dentro de las categorías de mayor riesgo para los humanos, según los criterios de autoridades alimentarias de Estados Unidos y Europa. El método convencional de identificación de antígenos O y H se realiza por aglutinación con antisueros de conejo. Este método además de ser muy costoso y laborioso, no se encuentra disponible en el país para empleo masivo. En este contexto, el objetivo de este estudio observacional descriptivo de corte transverso ha sido la estandarización de una técnica de PCR múltiple para la detección de estos 8 serogrupos, a fin de contar con un sistema de detección eficiente, sensible y con potencial de aplicación en la industria alimentaria. Se estandarizaron reacciones de PCR empleando como controles positivos cepas E. coli de referencia correspondientes a la totalidad de los serogrupos citados. Se obtuvieron productos de tamaños esperados para cada serogrupo, no se observaron amplificaciones cruzadas o falsos positivos. Esta técnica estandarizada podría representar una herramienta rápida y menos costosa que la técnica serológica, con la capacidad de ser aplicada a diferentes matrices, permitiendo la detección de estos serogrupos en aislados STEC de ganado en pie, fuentes de agua de consumo, alimentos e incluso en aislamientos clínicos asociados a enfermedades humanas(AU)
STEC serogroups O26, O45, O103, O104, O111, O121, O145, and O157, are related to a high number of cases of HUS worldwide, so they are included in the categories of greatest risk for humans, according to the food administration criteria of the United States and Europe. The conventional method of identifying antigens O and H is carried out by agglutination with rabbit antisera. This method is very expensive and laborious and is not available in the country for massive-scale use. In this context, the objective of this cross-sectional descriptive observational study has been the standardization of a multiplex PCR technique for the detection of these 8 serogroups, in order to have an efficient and sensitive detection system with the potential for application in the food industry. PCR reactions were standardized using as positive controls reference E. coli strains to correspond to all the mentioned serogroups. Products of expected sizes were obtained for each serogroup; no cross-amplification or false positives were observed. This standardized technique could represent a quick and less expensive tool than the serological technique, with the possibility to be applied to different kind of samples, allowing the detection of these serogroups in STEC isolates of live cattle, sources of drinking water, food and even in clinical isolates associated with human diseases(AU)
Assuntos
Escherichia coli Shiga Toxigênica/isolamento & purificação , Reação em Cadeia da Polimerase Multiplex , Estudos Transversais , Escherichia coli O157/isolamento & purificação , Escherichia coli O157/genética , Escherichia coli Shiga Toxigênica/genética , Escherichia coli O104/isolamento & purificação , Escherichia coli O104/genéticaRESUMO
Enteroaggregative Escherichia coli (EAEC) from the O104:H4 specific serotype caused a large outbreak of bloody diarrhea with some complicated cases of hemolytic-uremic syndrome (HUS) in Europe in 2011. The outbreak strain consisted in an EAEC capable to produce the Shiga toxin (Stx) subtype 2a, a characteristic from enterohemorrhagic E. coli QseBC two-component system detects AI-3/Epi/NE and mediates the chemical signaling between pathogen and mammalian host. This system coordinates a cascade of virulence genes expression in important human enteropathogens. The blocking of QseC of EAEC C227-11 (Stx+) strain by N-phenyl-4-{[(phenylamino) thioxomethyl]amino}-benzenesulfonamide (also known as LED209) in vivo demonstrated a lower efficiency of colonization. The periplasmic protein VisP, which is related to survival mechanisms in a colitis model of infection, bacterial membrane maintenance, and stress resistance, here presented high levels of expression during the initial infection within the host. Under acid stress conditions, visP expression levels were differentiated in an Stx-dependent way. Together, these results emphasize the important role of VisP and the histidine kinase sensor QseC in the C227-11 (Stx+) outbreak strain for the establishment of the infectious niche process in the C57BL/6 mouse model and of LED209 as a promising antivirulence drug strategy against these enteric pathogens.IMPORTANCE EAEC is a remarkable etiologic agent of acute and persistent diarrhea worldwide. The isolates harbor specific subsets of virulence genes and their pathogenesis needs to be better understood. Chemical signaling via histidine kinase sensor QseC has been shown as a potential target to elucidate the orchestration of the regulatory cascade of virulence factors.