Obtaining recombinant capsid protein VP60 of rabbit haemorrhagic disease virus and its antigenic and immunogenic activity study

Viral hemorrhagic disease of rabbits is an acute, highly contagious disease characterized by the phenomena of hemorrhagic diathesis in all organs, especially in the lungs and liver. The causative agent of viral haemorrhagic disease of rabbits is a virus of haemorrhagic disease of rabbits, belonging to the family Caliciviridae, genus Lagovirus. Currently, there are 4 genogroups of lagoviruses, two pathogenic: GI1 (GI1a-GI1d) and GI2, and two non-pathogenic: GI3 and GI4. The greatest danger to rabbits in the Russian Federation is posed by viruses of the GI1 genotype. The virulence of these viruses for rabbits is extremely high, the incubation period is 48-72 hours. Clinically, the disease is almost not manifested. Mortality can reach 100%. For the prevention of HBV in the Russian Federation, inactivated tissue vaccines are used, which are a suspension of the liver of rabbits infected with virulent strains of the rabbit hemorrhagic disease virus. Currently, in veterinary practice, subunit recombinant vaccines based on virus proteins obtained in the baculovirus gene expression system are increasingly used. The authors obtained the recombinant VP60 virus of rabbit hemorrhagic disease of the GI1 genotype in the baculovirus gene expression system and studied its antigenic and immunogenic activity for rabbits. It was found that the recombinant capsid protein VP60 of the hemorrhagic disease virus, administered to rabbits at a dose of 50 pg, causes the synthesis of specific antibodies in animals, detected by enzyme immunoassay and in the hemagglutination inhibition reaction, and protects 80% of animals during control infection with the virulent strain «Voronezh-87» at a dose of 1000 LD 50. These data indicate the possibility of using this protein as a specific component of a subunit vaccine against rabbit hemorrhagic disease caused by strains of the GI1 gene group.

Recombinant Lactobacillus casei Expressing Capsid Protein VP60 can Serve as Vaccine Against Rabbit Hemorrhagic Disease Virus in Rabbits

Rabbit hemorrhagic disease virus (RHDV) is the causative agent of rabbit hemorrhagic disease (RHD). RHD, characterized by hemorrhaging, liver necrosis, and high morbidity and mortality in rabbits and hares, causes severe economic losses in the rabbit industry worldwide. Due to the lack of an efficient in-vitro propagation system for RHDV, the current vaccine is produced via chemical inactivation of crude RHDV preparation derived from the livers of infected rabbits. Inactivated vaccines are effective for controlling RHD, but the potential problems of biosafety and animal welfare have negative effects on the application of inactivated vaccines. In this study, an oral Lactobacillus casei (L. casei) vaccine was used as an antigen delivery system to express RHDV capsid protein VP60(VP1)-eGFP fusion protein. The expression of the recombinant protein was confirmed via western blotting and immunofluorescence (IFA). Our results indicate that oral administration of this probiotic vaccine can stimulate secretory immunoglobulin A (SIgA)-based mucosal and IgG-based humoral immune responses in rabbits. The immunized rabbits were completely protected against challenge with RHDV. Our findings indicate that the L. casei expression system is a new strategy for the development of a safe and efficient vaccine against RHDV.

Minor structural protein VP2 in rabbit hemorrhagic disease virus downregulates the expression of the viral capsid protein VP60

Rabbit hemorrhagic disease virus (RHDV) has two structural proteins: the major capsid protein VP60 and the minor capsid protein VP2. VP2 is speculated to play an important role in the virus life cycle. To investigate the effect of VP2 on VP60 expression, three types of experiment (baculovirus–insect cell system, mammalian–luciferase assay system and in vitro coupled transcription/translation system) were used to express VP60 alone or co-expressed with VP2. Both forms of VP60 were able to form virus-like particles in insect cells. Western blot analysis and dual-luciferase assays demonstrated that the presence of VP2 results in downregulation of the expression of VP60 in vivo. Real-time RT-PCR of mRNA levels showed that downregulation of VP60 occurs at the transcriptional level. The ability of the viral minor structural protein VP2 to regulate capsid protein levels may contribute to effective virus infection.

Horizontal Transmissible Protection against Myxomatosis and Rabbit Hemorrhagic Disease by Using a Recombinant Myxoma Virus

ABSTRACT We have developed a new strategy for immunization of wild rabbit populations against myxomatosis and rabbit hemorrhagic disease (RHD) that uses recombinant viruses based on a naturally attenuated field strain of myxoma virus (MV). The recombinant viruses expressed the RHDV major capsid protein (VP60) including a linear epitope tag from the transmissible gastroenteritis virus (TGEV) nucleoprotein. Following inoculation, the recombinant viruses induced specific antibody responses against MV, RHDV, and the TGEV tag. Immunization of wild rabbits by the subcutaneous and oral routes conferred protection against virulent RHDV and MV challenges. The recombinant viruses showed a limited horizontal transmission capacity, either by direct contact or in a flea-mediated process, promoting immunization of contact uninoculated animals.