Caracterização de um novo fator de colonização (FC O25) de Escherichia coli isoladas de colibacilose neonatal bovina / Characterization of a new colonization (CF O25) of Escherichia coli isolated from neonatal bovine colibacillosis

AUTOR(ES)
DATA DE PUBLICAÇÃO

2006

RESUMO

Infections from Escherichia coli are frequent in domestic animals, affecting mainly newborn animals in the post weaning period, being thus responsible for important economic losses in the cattle, porcine and lamb herds. The establishment of the bacterium in the host is made by colonization and it is a process mediated by proteins called adhesins that are distributed along the bacterial surface. They present a great morphological variety and are called Colonization factors (CFs). Our objective in this work was to identify the new colonization factor (CF O25) in Escherichia coli strains isolated from bovine neonatal colibacillosis, this CF O25 presents antigenic similarities with the FCs F4(K88), CS31A and F41 without, however, present genetic identity to these factors in PCR assays. Based in these homologies, we investigated the hypothesis of the NFC be a member of this "family" of CFs. So we studied the sequence of the CF O25?s main subunit gene and its similarities with the genes of the homologous CFs (F4, F41 and CS31A) in polymerase chain reaction (PCR) assays. We found the CF O25 in 35 of the 41 studied strains. Through DNA sequencing, we verified that CF O25 protein presents 49.7% of homology with FaeG protein (F4), 35.4% with ClpG (CS31A) and 29.4% with FimF41. Using bioinformatics tools we determined the phylogenetic relationships between these proteins and the secondary structure of the CF O25. The genomic and plasmid DNA extraction assays indicates that the CF O25 gene is located in the bacterial chromosome. The CF O25 gene was cloned in the pET-151 expression vector, inserted in the E.coli BL21[DE3]-Star strain and the CF O25 protein was over expressed, purified and monospecific antiserum was obtained in rabbits. The purified CF O25 protein was submitted to Western Blotting assays with antissera against the FCs F4, F41 and CS31A and we verified recognition of the CF O25 in different degrees. The CG1905-B strain was used in the assays of electronic transmission microscopy using immunogold. The structure of the PCF is fibrillar resembling it the structure of the CS31A. This strain also was submitted the assays of adhesion to the eukaryotic culture cells and we observed adhesion to the Caco-2 cells (human colon carcinoma), this adhesion could be inhibited by anti-CFO25 antiserum. Our results suggest that the CF O25 can be important for the bovine pathogenicity of E. coli in Brazil, since we found CF O25 associated with other colonization factors and toxins, related with patogenicity in bovines, evidencing new relationships between virulence mechanisms that deserve new investigations, as well the elucidation of the complete structure of CF O25 operon

ASSUNTO(S)

calves escherichia coli f4 (k88) bovino fator de colonização diarrhea diarreia escherichia coli

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