Identificação, caracterização e análise da expressão de genes de resposta ao estresse oxidativo em Lactobacillus delbrueckii UFV H2b20

• Ano de defesa: 2008
• Autor(a) principal: Leite, Marta Cristina Teixeira
• Orientador(a): Moraes, Célia Alencar de
• Banca de defesa: Borges, Arnaldo Chaer , Mantovani, Hilário Cuquetto , Carvalho, Antônio Fernandes de , Salomão, Tânia Maria Fernandes
• Tipo de documento: Tese
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Viçosa
• Programa de Pós-Graduação: Doutorado em Microbiologia Agrícola
• Departamento: Associações micorrízicas; Bactérias láticas e probióticos; Biologia molecular de fungos de interesse
• País: BR
• Palavras-chave em Português: Lactobacillus delbrueckii; Probióticos; Estresse oxidativo
• Palavras-chave em Inglês: Lactobacillus delbrueckii; Probiotics; Oxidative stress
• Área do conhecimento CNPq: CNPQ::CIENCIAS AGRARIAS::AGRONOMIA::FITOSSANIDADE::MICROBIOLOGIA AGRICOLA
• Link de acesso: http://locus.ufv.br/handle/123456789/1537

Oxidative stress is an important cause of viability loss in lactic acid bacteria used in food industry and as probiotics. This work aimed at identifying and characterize genes involved in the oxidative stress response in Lactobacillus delbrueckii UFV H2b20, a potencial probiotic strain. Therefore, dps, which encodes a ferritin-like protein, and pox, which encodes the pyruvate oxidase enzyme, were identified in L. delbrueckii UFV H2b20 and sequenced. The operon bearing the spx gene, encoding a transcriptional factor, was also identified and partially characterized. The operon bearing spx is conserved in sequence and organization in L. delbrueckii UFV H2b20. A high sequence identity (98%) was found between the spx operon and its homologs of L. delbrueckii ATCC 11842 and L. delbrueckii ATCC BAA-365, as well as between pox of L. delbrueckii UFV H2b20 and its homologs of those bacteria. The dps gene is absent in the genome of L. delbrueckii ATCC 11842 and L. delbrueckii ATCC BAA-365, but it is present in L. delbrueckii UFV H2b20 bearing 99% identity with its homologs of Lactobacillus plantarum, Lactobacillus rhamnosus e Lactobacillus sakei, three probiotic bacteria that belong to a group different from L. delbrueckii. Phylogenetic analyses based on spx and pox demonstrate that L. delbrueckii UFV H2b20 is always related to L. delbrueckii ATCC 11842 and L. delbrueckii ATCC BAA-365, confirming that L. delbrueckii UFV H2b20 belongs to this species, although it has some unique characteristics. Phylogenetic analyses based on dps sequences demonstrate that L. delbrueckii UFV H2b20 is always related to L. plantarum, L. rhamnosus, L. reuteri e L. sakei, indicating the possible aquisition of this gene by horizontal gene transfer. Besides, these results indicate the possible involvement of dps with the probiotic features of this strain. In this work, the expression of those three genes in response to oxidative stress was also evaluated. Transcription of all genes was induced in response to oxygen and dps is induced by hydrogen peroxide in a dose-dependent way. The spx and dps genes were induced at the beggining of stationary phase demonstrating that some regulatory factors that control the expression of these genes are independent of oxygen, during this growth phase. The pox gene displays extreme sensibility to the presence or absence of oxygen, being fastly induced or repressed, respectively. This gene is also repressed by excess of glucose, since it is regulated by catabolite repression. These results were also observed when the enzyme activity was evaluated in excess of glucose or under restriction of this sugar and in the presence of galactose as well. The involvement of spx, dps and pox in the oxidative stress response of L. delbrueckii UFV H2b20 was clearly demonstrated. The demonstration of the role of these genes in the oxidative stress response is important for the development of the industrial process.