ABSTRACT
Infectious bronchitis virus (IBV) is prevented primarily by the use of live attenuated vaccines, which are known to have a limited strain range of protection. Alternative vaccines against the emerging new virus strains can improve control of the disease. The aim of this study was to evaluate the immunogenic potential of two recombinant viral proteins, when administered by eyedrop, without the assistance of a vector. The recombinant S1 (rS1) and N (rN) proteins of the M41 strain expressed in E. coli were tested, and the live attenuated vaccine H120 was used as a positive control. Protection was evaluated by re-isolation of virus from tracheas of vaccinated chickens after challenge with strain M41. After three immunizations, rS1 glycoprotein induced 40% protection, while vaccination with rN provided no protection. Vaccination with rS1, rN, or H120 induced a cellular immune response as demonstrated by in vitro ChIFN-γ production by splenocytes of vaccinated birds. Vaccination with H120, and to a lesser extent rS1, induced HI and virus-specific IgG antibody production. These findings indicate that recombinant viral proteins administered through the mucosal route can evoke an immune response without the assistance of a vector.
Subject(s)
Infectious bronchitis virus/immunology , Membrane Glycoproteins/immunology , Nucleocapsid Proteins/immunology , Poultry Diseases/immunology , Recombinant Proteins/immunology , Viral Envelope Proteins/immunology , Viral Vaccines/immunology , Administration, Mucosal , Animals , Antibodies, Viral/blood , Chickens , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Coronavirus Infections/veterinary , Coronavirus Nucleocapsid Proteins , Infectious bronchitis virus/metabolism , Interferon-gamma/biosynthesis , Membrane Glycoproteins/administration & dosage , Membrane Glycoproteins/genetics , Nucleocapsid Proteins/administration & dosage , Nucleocapsid Proteins/genetics , Poultry Diseases/prevention & control , Poultry Diseases/virology , Recombinant Proteins/administration & dosage , Recombinant Proteins/genetics , Spike Glycoprotein, Coronavirus , Vaccination , Viral Envelope Proteins/administration & dosage , Viral Envelope Proteins/genetics , Viral Vaccines/administration & dosage , Viral Vaccines/geneticsABSTRACT
A sensitive and specific method for the diagnosis of infectious bronchitis virus (IBV) is of great importance. In this study the development of a real-time TaqMan RT-PCR targeting the highly conserved nucleocapsid (N) gene of IBV and including an internal PCR control is described. The assay was specific for IBV and did not detect other avian pathogens, including turkey coronaviruses. A comparative limit of detection was determined for M41, an embryo-adapted strain, and IS/885/00, a poorly embryo-adapted variant. For M41 real-time RT-PCR and virus isolation were one or two times more sensitive than RT-PCR targeting the N or spike glycoprotein (S1) genes, respectively. For IS/885/00, real-time RT-PCR was more sensitive by tenfold than virus isolation and 30- or 40-fold than by N gene or S1 gene RT-PCR, respectively. Real-time RT-PCR and virus isolation were 17-75% more sensitive than RT-PCR targeting the S1 gene for testing tracheal swabs directly from experimentally infected chicks. When tracheal and cloacal swabs from clinical specimens were tested directly, 50% more samples were positive by real-time RT-PCR than by the S1 gene RT-PCR. Real-time RT-PCR targeting the N gene is more sensitive than common diagnostic assays, allowing rapid and accurate IBV detection directly from clinical specimens, facilitating differential diagnosis.
