Identificador: TDX:2369
Autors: Soler Falgàs, Lara
Resum:
The genus Aeromonas comprises Gram-negative bacteria frequently isolated from water. They are important because they are found in food and can provoke diarrhoea and extraintestinal infections in humans. The present thesis demonstrates that they can be aetiological agents in up to 2% of traveller's diarrhoea.<br/>Aeromonas taxonomy is complex. The genus comprises 15 species difficult to identify with classical biochemical methods. Because of this we have developed a technique based on the 16S rRNA restriction patterns that allows the identification of all the accepted species of Aeromonas. <br/>Most clinical laboratories use miniaturized semiautomatic systems or other biochemical tests to identify Aeromonas species. As we demonstrate they can give erroneous identifications as Vibrio at both species and genus level. In an attempt to avoid this confusion, we designed a molecular probe, based on the gene that codifies for glycerophospholipid-cholesterol acyltransferase, which can detect Aeromonas colonies in primary isolation media in a fast and reliable manner.<br/>The correct delimitation and identification of the species that integrate the phenotypic 'A. hydrophila' complex (A. hydrohila, A. bestiarum, A. salmonicida and A. popoffii) constitutes one of the problems in the taxonomy of the genus. For that reason we considered it important to investigate the biochemical and genetic characteristics of this group of species. We have observed that the species A. bestiarum and A. salmonicida cannot be differentiated either with biochemical or with genetic methodology, suggesting that they may be unique species.<br/>Another objective was the study of the resistance patterns, in genetically identified Aeromonas species, to the antimicrobial agents. We demonstrate that clinical isolates of A. hydrophila, A. caviae and A. veronii species are susceptible to cephalosporins, aminoglycosides and fluoroquinolones. Moreover, there was evidence that quinolones and third generation cephalosporines should be considered as the treatment of choice of traveller's diarrhoea.<br/>In order to improve our knowledge of the epidemiology of Aeromonas, it is essential to have reliable molecular typing methods. We evaluated three techniques (intergenic spacer region 16S-23S RFLP, ERIC-PCR and REP-PCR) for the molecular typing of Aeromonas, and we found ERIC-PCR to be more discriminative. However, we also observed that the simultaneous combination of two of those techniques (ERIC and REP) further improved the typing.<br/>The virulence of Aeromonas is complex and possibly multifactorial. Numerous genes codifying for different enzymes (hemolysins, lipases, DNases, proteases.) have been described and their implication in virulence has been demonstrated, but there have been no studies to investigate simultaneously the incidence of all those genes in clinical and environmental isolates of all the species of the genus. In the present thesis we investigate the distribution of the genes codifying for those virulence factors, and their associated enzymatic activity on clinical and environmental isolates of all the species of the genus. We found a large presence of the genes in all the species, regardless of the origin. However, we also found a statistically significant association between the presence of genes codifying for aerolysin/hemolysin and serine protease and ß-hemolitic activity, which suggests that serine protease acts as aerolysin/hemolysin activator.<br/>Finally, we considered it of interest to determine whether the Aeromonas species with major clinical relevance possess the type III secretion system, described in other pathogenic micoorganisms, such as Escherichia coli, Yersinia spp. and Salmonella spp. The principal function of this system is to inject toxins directly into the host cell cytoplasm. We demonstrate that this system is present in most of the clinical A. veronii and A. hydrophila isolates, which indicates that those strains possess a virulence potential comparable to that of the other mentioned pathogens.
Repositori URV