Clonagem e análise genética do gene iss, e avaliação da atividade do anticorpo IgY anti-proteína recombinante Iss de Escherichia coli patogênica para aves (APEC)

AUTOR(ES)
DATA DE PUBLICAÇÃO

2009

RESUMO

Escherichia coli strains designated as avian pathogenic E. coli (APEC) are responsible to avian colibacillosis, which is the principal cause of high economic losses in the modern poultry industry, due the increase of mortality, medication cost and condemnation of carcasses. The APEC pathogenicity is related to presence of several virulence factors, being the Iss (Increased survives Serum) protein a principal responsible characteristic of resistance to complement system, increasing its capacity to survive in the serum. The objectives of this work were both study the diversity sequence iss gene from APEC and characterize the recombinant Iss protein by its biological activity. The iss gene from APEC 9 strain of 309 pb was cloned in the pET SUMO vector and sub-cloned in the pET 30. The iss gene was classified as iss type 1 by differentiation of the three iss gene allele types. The recombinant protein (rIss) was expressed in E. coli BL21 (DE3) by induction of IPTG and purified in resin charged with the nickel ion. The molecular mass of rIss-SUMO was 22 kDa, corresponding 11 kDa of Iss protein and 11 kDa SUMO protein and the rIss from pET30 was14 kDa. The rIss was used to immunize hens and the results showed induction of immune response. Antibodies IgY anti rIss, purified from eggs of immunized hens, reacted by Western blotting with rIss showing specific molecular mass and they were utilized in the serum resistance inhibition tests and . The results demonstrated inhibition the bacterial grown in the serum and decrease of lesions score in the chicken one day old. In conclusion, the iss gene from APEC 9 strain is the genotype 1 that is highly prevalent among APEC and it could be used in diagnostic protocols, and these data increase the perspectives of new investigation about Iss protein.

ASSUNTO(S)

colibacilose ave - doenças colibacillosis birds diseases escherichia coli

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