Análise comparativa das propriedades fenotípicas e genotípicas de amostras de origem humana e animal de Streptococcus pseudoporcinus

• Autor(a) principal: Carvalho, Guilherme da Silva Lourenço
• Data de Publicação: 2017
• Outros Autores: [e-mail]
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Biblioteca Digital de Teses e Dissertações da UERJ
• Texto Completo: http://www.bdtd.uerj.br/handle/1/20472
• topic: Streptococcus pseudoporcinus; Streptococcus porcinus; Virulence; MALDI-TOF; Biofilms; Genomics; Proteomics; Streptococcus pseudoporcinus; Streptococcus porcinus; Virulência; MALDI-TOF MS; Biofilmes; Genômica; Proteômica; CIENCIAS BIOLOGICAS::MICROBIOLOGIA::MICROBIOLOGIA APLICADA::MICROBIOLOGIA MEDICA

The genus Streptococcus is constantly suffering reclassifications. The aim of this study was to evaluate the characteristics presented by 24 isolates of Streptococcus pseudoporcinus, including a derivative of the type strain, being 19 obtained from human clinical especimens and five from animal origin (milk samples). Six Streptococcus porcinus reference strains were included for comparative purposes. These isolates were characterized by physiological tests, protein profiles by SDS-PAGE, antimicrobial susceptibilities and virulence-related characteristics. Additionally, MALDI-TOF MS technique was used in order to characterize these isolates, allied with phylogenetic analysis and whole genome sequencing. Proteomic assays were carried out by mass spectrometry (ESI-Q-TOF). Physiological tests results indicated that hippurate hydrolysis was the only one able to differentiate the species. The results obtained by MALDI-TOF MS showed distinct scores values, depending on the protocol employed to the samples preparation. Protein extracts obtained by the treatment with formic acid and acetonitrile, have shown effective results in discriminate between S. pseudoporcinus and S. porcinus species. Additionally, high correlation degrees were identified among samples of S. pseudoporcinus obtained from milk. The data obtained by MALDI-TOF MS were also assessed to identify biomarkers, demonstrating the presence of intra and interspecies spectra. The analysis of the protein profiles obtained by SDS-PAGE revealed a great similarity between the species and it was not sufficiently discriminatory. The analysis by the rrs gene sequences that S. pseudoporcinus isolates belonging to human sources were less related to the isolates obtained from milk samples. Antimicrobial susceptibility testing demonstrated 66.0% of the S. pseudoporcinus of human origin were resistant to tetracycline. The isolates were able to form biofilms, and supplementation of the culture medium with sucrose or glucose considerably increases the amount of biomass adhered. Biofilms of type strains of both species showed inhibitory concentration values up to 40,000 times greater than planktonic cells. Several virulence-related proteins have been identified by proteomics and genomics analyses. Both species were able to adhere to human cells.