Genetic characterization and strain differentiation of feline Calicivirus among cat population in Thailand by high resolution melting analysis

Abstract

Feline calicivirus (FCV) is wildly known as one of the significant causative viral pathogens causing self-limited upper respiratory tract and oral disease in cats. In the last decade, severe FCV form, as named a virulent systemic feline calicivirus (VS-FCV), has distinctly emerged worldwide even though commercially available vaccines for FCV are typically used in Thai cats. However, the data of either prevalence or molecular characterization has never been officially noticed in Thailand. Thus, this study aimed to provide molecular epidemiology and characterization of the FCV circulating in Thai cats during 2016-2021. Moreover, the high-resolution melting analysis (HRM) is utilized for simultaneous detection and strain typing. Immunohistochemical staining (IHC) and in situ hybridization (ISH) were also performed for viral localization in tissues. Our results demonstrated FCV incidence circulating in Thai cats as 46.7% and associated with gingivostomatitis lesion with an odds ratio of 9.02 (95% CI: 2.61-30.46; p<0.001). The FCV Thai strains (FCV-THs) were clustered in genogroup I without any viral recombination evidence. One of our cases had shown clinical signs, likely VS-FCV, and became sudden death, and we could complete the whole genome coding sequence of these samples. Moreover, our study successfully validated the HRM technique that could segregate between two commercially available FCV vaccine strains and five wild-type FCV Thai strains within a single PCR reaction. Furthermore, the reverse-transcription polymerase chain reactions (RT-PCRs) disclosed positive results in various organ tissues in one VS-FCV dead cat, which represented related clinical signs of virulent strain. Afterward, IHC, ISH, and transmission electron microscopy (TEM) were performed to confirm the localization, distribution, and existence of viral pathogens. As known that target cells of FCV are epithelial and endothelial cells, interestingly, our findings represented FCV antigen either protein or genomic signal in the neuronal cells. This was confirmed the existing intranuclear virion by TEM. In conclusion, we suggested that FCV circulation in Thai cats is still on a big scale, even though the vaccination is wildly used. So, monitoring emerging vaccine breakdown strains and possibly influencing the evolution undergone positive selection is still needed. Lastly, the insight study about FCV-associated neuropathy should be more investigated in further research