Pneumonia enzoótica suína: isolamento do agente e expressão de antígeno recombinante em pichia pastoris

Abstract

Mycoplasma hyopneumoniae is the causative agent of Enzootic Pneumonia (EP), one of the main respiratory diseases in pigs. The EP is a disease that affects pigs worldwide, causing major economic losses to the pig industry. Vaccination is the most effective method to control the EP. Commercial vaccines available on the market are bacterins, produced from whole-cell inactivated bacteria, and confer only partial protection for the pigs. In addition to the bacterins, recombinant M. hyopneumoniae proteins produced in E. coli have been tested as potential vaccine targets. This study had two main objectives: (1) obtaining isolates of M. hyopneumoniae from lung samples collected from an abattoir, aiming at assessing the genetic diversity of strains circulating in the RS, and (2) the expression of a chimera composed by three M. hyopneumoniae antigens (P42, R1 and NrdF) fused with a mucosal adjuvant, in the yeast Pichia pastoris in order to obtain the antigen in a large amount and with a different folding to the protein expressed in E. coli. For isolation, 62 lung samples were collected, processed and cultured. M. hyopneumoniae isolates obtained were contaminated with other Mycoplasma spp. It was not possible to obtain a pure isolated to proceed with the study of genetic diversity. For expression of a chimera in Pichia pastoris, the chimera synthetic gene was efficiently cloned in KM71H strain, and expression or the protein was observed by Western blot. Both, the insolation of the agent, as well as the expression of recombinant antigens, are important steps for the development of an effective vaccine for the control of EP.