Application of Type Specific Spike Protein in the Differentiation of Type I and II Feline Coronavirus Infection
Date Issued
2011
Date
2011
Author(s)
Wang, Ying-Ting
Abstract
Feline infectious peritonitis (FIP), is a fatal disease caused by feline coronavirus (FCoV). According to the serum neutralizing capacity and the antigenicity relation to canine coronavirus, FCoVs can be divided into two serotypes, namely, Type I and Type II. Type I FCoV is predominant in the field. Despite the higher prevalence of Type I FCoV infection, infection of Type II FCoV appears to be highly significantly correlated to FIP. Coronavirus infection is determined by the interaction between the spike protein and its corresponding receptors on the target cells. S gene has been used in the differentiation of different type(s) of FCoV infection due to the low similarity between the two viruses. The seroprevalence of different types of FCoV infection in Taiwan is unknown yet. This study aim to investigate the seroprevalence of different types of FCoV infection in Taiwan through the expression of type specific spike proteins from Type I and II FCoV. Spike gene of two types of FCoV were aligned, and a low similarity region corresponding to the receptor binding domain of spike protein from Type II virus and the related region of Type I virus were selected, amplified and cloned into a baculovirus expression system. Two recombinant viruses with the titer up to 2 × 106 PFU/ml have been obtained. Cells were infected with these recombinant viruses and total protein was harvested, the type-specific recombinant proteins could be recognized by anti-serum with the expected size of 23-25kDa. While Sf9 cells were infected with these two recombinant viruses at a multiplicities of infection 0.1, the highest protein yield can be obtained at 72 hour and 96 hour post infection for type I and II RBD recombinant viruses, respectively. The specificity of type I recombinant protein could be evaluated by co-immunoprecipitation. Sera from cats infected with different types of FCoV could specifically identify corresponding recombinant proteins by immunofluorescence assay. The intensity of fluorescence was declined through serial dilution of anti-serum, which indicates the proteins is type-specific and could be used for the titration of the specific antibody. These type-specific recombinant proteins will be purified and used for the establishment of ELISA for further serological surveillance.
Subjects
feline coronavirus
spike protein
differentiation
Type
thesis
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