Aplicação da glicoproteína D recombinante de Herpesvírus bovino 5 como antígeno em teste de imunodiagnóstico e vacina de subunidade

Abstract

The new strategies in vaccine design against Bovine herpesvirus 1 and 5 are focused in the viral envelope glycoprotein, such as the gD, which acts in attachment and fusion of viral envelope with the permissive cell membranes. This glycoprotein is present at large quantities in the viral envelope and stimulates strong humoral and cellular immune responses at the host. The serological diagnostic based on immunoenzymatic assays, as the Indirect ELISA, also applies the envelope glycoproteins, once they are expose in the viral structure. Thus, the aim of this study was to apply the gD from BoHV-5 expressed in Pichia pastoris, after its antigenic characterization, as antigen in a subunit vaccine and in Indirect ELISA, aiming the control and diagnosis of BoHV-5, which is the causative agent of herpetic meningoencephalitis outbreaks, resulting in economic losses for the Brazilian livestock. The subunit vaccine developed was able to stimulate humoral immune response in mice, with development of neutralizing antibodies when the rgD was administrated alone or in combination with immunological adjuvants. The cellular immune response was observed through the increase in the mRNA expression associated with this kind of responses, as IFN-!, IL-17 and GM-CSF, when the vaccine was formulated with the oil adjuvant Emulsigen-DDA. The Indirect ELISA developed shown high sensitivity (100%) and specificity (92.9%) when compared with the virus neutralization test, which is considered the gold standard in BoHV-1 and -5 serological diagnosis. The rgD from BoHV-5 expressed in P. pastoris possess similar tridimensional conformation from the native BoHV-5 gD. The rgD is immunogenic, capable of inducing neutralizing antibodies development and the subunit vaccine developed with the rgD as antigens induces a balanced Th1/Th2 immune response. The recombinant protein antigenicity was confirmed through the Indirect ELISA, which may be used to a fast screen of BoHV-5 infected herds.