Please use this identifier to cite or link to this item: http://hdl.handle.net/2307/40413
Title: Shiga-toxin-producing Escherichia coli (STEC) : evolution of the different pathogenic clones and study of their interactions with the microbiota in the intestine and the environment
Authors: Gigliucci, Federica
Advisor: MORABITO, STEFANO
Keywords: HUMAN GUT
MICROBIOTA
METAGENOMICS
STEC
Issue Date: 14-Dec-2018
Publisher: Università degli studi Roma Tre
Abstract: Shiga toxin-producing E. coli are zoonotic pathogens causing a wide spectrum of diseases in humans, ranging from mild diarrhoea to haemorrhagic colitis (HC) and the haemolytic uremic syndrome (HUS). STEC pathogenicity relies on the production of potent cytotoxins, Shiga toxins (Stx), acquired upon infection with bacteriophages carrying stx genes. Given the tremendous genomic variability characterising the STEC strains and the many STEC types described so far, the aim of the present work was to investigate those STEC populations whose impact on public health was not completely understood, as well as to examine the hidden epidemiology of certain STEC types, how they can reach the final host and their effect on the host gut. In particular, we investigated novel or neglected reservoirs and transmission pathways for STEC infections and studied the interactions between STEC and the microbial communities, once they establish in the human intestine upon infection. In the first part of this work whole genome sequencing was used to characterize STEC strains producing the Stx2f, first identified in pigeons and for long time considered a host-adapted Stx subtype. These STEC were considered as having a limited impact on public health, but recently this picture started changing as isolates from cases of human diarrhoea and even HUS have been reported. The study included Stx2f-STEC strains isolated from human patients with diarrhoea or HUS and from healthy pigeons. The Virulotyping analysis highlighted the existence of at least three distinct subpopulations of Stx2f-producing E. coli, all sharing a minimum core of the virulence genes content. Strains from pigeons and from diarrhoea displayed a certain degree of similarity in the virulome, while the Stx2f-producing strains from HUS showed a virulence genes asset overlapping that of the other STEC commonly isolated from HUS and including mobile genetic elements conveying virulence genes associated with the colonization of host gut not observed in the other Stx2f-STEC strains studied. The results of our work suggested that the Stx2f-STEC may be pathogenic to humans in their own right and that the Stx2f subtype has the potential to cause HUS when produced by an isolate able to efficiently colonize the human intestinal tract. We formulated the hypothesis that pigeons can disseminate in the environment pathogenic Stx2f-STEC strains, or the stx2f-phages, which can in turn lysogenize E. coli strains with a virulence genes background associated with HUS, representing a serious risk for public health. The environment represents indeed an important source of pathogenic E. coli, given their ability to adapt and survive in a wide spectrum of ecological niches. Nowadays, the need, to high crop with a reduced usage of water and expensive fertilizers to amend soils, led to the use of Biosolids (BSO) as soil improvers, such as biomass resulting from sewage sludge derived from biological or chemical treatment of industrial, municipal and zootechnical wastewater, manure from livestock, and compost from green wastes. Given the increasing use of this organic matter in agricultural settings and in order to evaluate the possible presence of zoonotic agents, a set of BSO samples intended for soil fertilization and grazing activities was subjected to a molecular screening through Real Time PCR. The analysis gave positive results for virulence genes of human adenovirus, human norovirus and different diarrhoeagenic E. coli (DEC), including STEC. Additionally, the enrichment culture from BSO showed the presence of viable microbial cells of pathogenic E. coli. These findings indicated that the use of such organic matter to amend agricultural land may present the risk of spreading known or emerging hazards of anthropogenic or animal origin in the environment devoted to food production, suggesting the need to perform a deeper analysis of such BSO. To this purpose, we investigated the BSO samples with a metagenomic approach that allowed us to identify in the samples virulence genes characteristic of different DEC pathogroups, including STEC, and antimicrobial resistance determinants, as well as to obtain a complete taxonomic profile of the microbial communities present in the different BSO analysed. These studies provided evidences that BSO may act as a new environmental vehicle of important biological threats for human health, strengthening the need for a full risk assessment of the use of soil improvers in agricultural settings devoted to growing crops for human consumption or animal pasture. The advent of next generation sequencing technologies changed the paradigm of the investigation of complex systems. To complete the investigation of the whole axis of human exposure to STEC infections, a study was performed based on the analysis of faecal samples from STEC-infected patients and from coupled healed and healthy subjected, collected during an outbreak of STEC O26:H11 infections, using an enhanced metagenomic approach. The aim of the study was to investigate whether STEC may alter the composition of the human intestinal microbiota upon infection, eventually influencing the severity of disease. Briefly, the distribution of the microbial populations in the specimens from diseased patients, showed a lesser abundance of commensal Bifidobacteriales and Clostridiales spp. with respect to those from STEC negative samples, where those microorganisms predominated. This observation was confirmed by including in the analysis samples from patients with Crohn’s disease in order to assess whether the changes observed were due to a general status of dysbiosis rather than to the colonization by the STEC strain. Our study provided the first evidence of the dynamics occurring in the human gut upon infection with STEC and led to the hypothesis of a possible competition between beneficial microorganisms and STEC strains in human intestine. Such studies should be further encouraged as they may open the way to the deployment of measures to mitigate the impact of STEC infections and their progression towards the most severe forms.
URI: http://hdl.handle.net/2307/40413
Access Rights: info:eu-repo/semantics/openAccess
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