Reverse Genetics for Type I Feline Coronavirus Field Isolate To Study the Molecular Pathogenesis of Feline Infectious Peritonitis

Abstract

Feline infectious peritonitis (FIP), one of the most important lethal infections of cats, is caused by feline infectious peritonitis virus (FIPV), the high-virulence biotype of feline coronaviruses (FCoVs). FIPVs are suggested to emerge from feline enteric coronaviruses (FECVs) by acquiring mutations in specific genes in the course of persistent infections. Although numerous studies identified mutations predicted to be responsible for the FECV-FIPV biotype switch, the presumed roles of specific genetic changes in FIP pathogenesis have not been confirmed experimentally. Reverse genetics systems established previously for serotype I and the less common serotype II FCoVs were based on cell culture-adapted FIPV strains which, however, were shown to be unsuitable for FIP pathogenesis studies in vivo. To date, systems to produce and manipulate recombinant serotype I field viruses have not been developed, mainly because these viruses cannot be grown in vitro. Here, we report the first reverse genetics system based on a serotype I FECV field isolate that is suitable to produce high-titer stocks of recombinant FECVs. We demonstrate that these recombinant viruses cause productive persistent infections in cats that are similar to what is observed in natural infections. The system provides an excellent tool for studying FCoVs that do not grow in standard cell culture systems and will greatly facilitate studies into the molecular pathogenesis of FIP. Importantly, the system could also be adapted for studies of other RNA viruses with large genomes whose production and characterization in vivo are currently hampered by the lack of in vitro propagation systems.

Pathogenesis of oral type I feline infectious peritonitis virus (FIPV) infection: Antibody-dependent enhancement infection of cats with type I FIPV via the oral route

“…It is unclear which of these regions is involved in the pathogenicity of FIPV-I KU-2. It is now possible to prepare recombinant type I FCoV by reverse genetics [4, 20]. It is desired to mutate the regions associated with pathogenicity in FIPV-I KU-2, inoculate cats with these mutants through various routes, and confirm whether the mutant causes FIP.…”

Development of Feline Ileum- and Colon-Derived Organoids and Their Potential Use to Support Feline Coronavirus Infection

“…Serotype I recFECV-GFP and recFECV-S 79 -GFP were produced in vitro using the reverse genetic system for FCoV field strains described previously [ 34 ]. Recombinant virus stocks of recFECV-S 79 -GFP were titrated by plaque assay on routinely used felis catus whole fetus (FCWF) cells [ 26 , 27 , 28 , 35 , 36 ].…”

“…Recombinant virus stocks of recFECV-S 79 -GFP were titrated by plaque assay on routinely used felis catus whole fetus (FCWF) cells [ 26 , 27 , 28 , 35 , 36 ]. Virus stocks of recFECV-GFP which cannot be cultivated in standard cell culture systems were quantified by comparative Western blot analysis of the FCoV M protein together with recFECV-S 79 -GFP [ 34 ]. The cell line FCWF was provided by the diagnostic laboratory at the Justus Liebig University Giessen and maintained in culture media (DMEM with 1× penicillin/streptomycin (Thermo, Waltham, MA, USA) and 10% FBS (Biochrom, Cambridge, UK)).…”

Development of Colorimetric Reverse Transcription Loop-Mediated Isothermal Amplification Assay for Detecting Feline Coronavirus

“…It is widely accepted that the highly virulent biotype, referred to as feline infectious peritonitis virus (FIPV) develops from less virulent FCoV in individual cat through mutations in certain genes (spike glycoproteins and probably some accessory genes, 3c and 7b); the hypothesis of which is called internal mutation theory [ 15 ]. It was estimated to occur in approximately 5% of persistently infected cats [ 16 , 17 , 18 , 19 ]. FIPV is responsible for monocytes and macrophages infection leading to fatal immune-mediated disease [ 15 , 18 , 20 , 21 ].…”