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Lancefield group C and G streptococci in human infection : molecular typing, virulence and antimicrobial susceptibility

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Resumo:	Beta-hemolytic, large-colony-forming (diameter greater than 0.5 mm) Lancefield group C and group G streptococci (GCGS) is a group within the Streptococcus genus which includes several species recognized as either colonizers or pathogens in humans and animals. Streptococcus dysgalactiae subsp. equisimilis (SDE), which can express any of these two Lancefield group antigens, is the GCGS species most commonly reported in human infection worldwide and may cause a number of potentially life-threatening infections. SDE is increasingly regarded as an emerging global pathogen and is able to colonize and infect humans, while other GCGS species, such as Streptococcus canis, are mainly animal pathogens that occasionally infect the human host. The rising number of human infections reported to be caused by GCGS warrants a better study of their epidemiology, in order to establish the relevance of each species, and clarifying the clonal dynamics and the intra-specific factors influencing the virulence of the strains. This work aimed at determining the GCGS species responsible for human infection in Portugal and, by assessing the genetic diversity of the isolates recovered, to define the clonal structure of this population. Thus, a special emphasis on molecular typing techniques was given, including those more commonly used to type these streptococci: pulsed-field gel electrophoresis (PFGE), emm typing and multilocus sequence typing (MLST). The application of these techniques to characterize collections of SDE and S. canis isolates recovered from other geographic locations and, for the latter species, from animal hosts, allowed to elucidate their respective population structures and provided new insights into the biology and evolution of GCGS. Initially, the speciation and characterization of GCGS isolates recovered from human infections in Portugal identified the central role of SDE. Not only was there a weak correlation between emm typing and PFGE results, but the polyclonal origin of the SDE population in this region was revealed by each of the methods used that generated multiple partitions. Furthermore, a correlation between emm type and invasive disease potential was suggested. A more global snapshot of the clonal architecture of this pathogen population was achieved by developing a MLST scheme which was applied to an expanded collection of SDE isolates recovered from distinct continents. An association of Lancefield groups with distinct MLST partitions was found and the high prevalence of a small number of widely distributed MLST sequence types (STs) suggested that a few genetic lineages dominate among SDE causing human infection worldwide. The occurrence of intra-specific and inter-specific recombination with Streptococcus pyogenes (Lancefield group A streptococcus, [GAS]), involving the housekeeping genes used in MLST was detected. The poor correlation between emm typing and either PFGE or MLST defined groups was illustrated by lineages displaying distinct emm types and the presence of the same emm type in unrelated genetic backgrounds, as defined by both techniques. These observations suggested the existence of recombinational replacements involving the emm locus and question the value of emm typing to accurately ascertain the genetic relatedness of SDE strains. The characterization of the antimicrobial susceptibility patterns presented by SDE found a high proportion of levofloxacin-resistant isolates (12%) associated with multiple genetic lineages. Sequence analysis of the quinolone resistance-determining regions of the gyrA and parC genes of representative resistant and susceptible isolates showed that full resistance was associated with mutations in both GyrA and ParC. As observed for the housekeeping genes used in MLST, recombination with GAS DNA in some parC alleles was evident, though this phenomenon was not exclusively associated with resistance. The final part of the work in this thesis focused on S. canis, the second most frequent GCGS species isolated from human infections in Portugal. A collection of S. canis isolates recovered from infections in both humans and animals, collected in Portugal and abroad, were characterized by employing the same typing methods used for SDE. The S. canis population was polyclonal, and several genetic lineages were shown to possess the ability to infect the human host. The zoonotic nature of S. canis infection was demonstrated, as identical genetic lineages were found infecting house pets and humans, indicating that they constitute a single population. Phylogenetic analysis showed that S. canis was a divergent taxon of SDE and GAS and unveiled the acquisition of genetic material of SDE by S. canis. The presence of emm-like genes was restricted to a few S. canis isolates and correlated with some MLST-based genetic lineages. Globally, this thesis contributes to the current knowledge of the molecular epidemiology and evolutionary relationships among members of the two GCGS species studied. The clonal relationships among strains were elucidated and MLST schemes for SDE and S. canis were established, providing useful tools for future studies of their population dynamics. The use of emm typing was shown to be complemented by applying other typing methods and the role of the M protein in SDE virulence was reinforced. The evidence found for recombinational replacements between SDE and GAS in several loci and, to a smaller extent, between SDE and S. canis, indicates that horizontal gene transfer events are important mechanisms driving genetic variability in GCGS populations which may impact key bacterial functions such as virulence and antimicrobial resistance.
Autores principais:	Pinho, Marcos Daniel
Assunto:	S. dysgalactiae subsp. equisimilis S. canis População Recombinação Teses de doutoramento - 2014
Ano:	2014
País:	Portugal
Tipo de documento:	tese de doutoramento
Tipo de acesso:	acesso aberto
Instituição associada:	Universidade de Lisboa
Idioma:	inglês
Origem:	Repositório da Universidade de Lisboa