Detection of feline coronavirus spike gene mutations as a tool to diagnose feline infectious peritonitis

Abstract

Objectives Feline infectious peritonitis (FIP) is an important cause of death in the cat population worldwide. The ante-mortem diagnosis of FIP in clinical cases is still challenging. In cats without effusion, a definitive diagnosis can only be achieved post mortem or with invasive methods. The aim of this study was to evaluate the use of a combined reverse transcriptase nested polymerase chain reaction (RT-nPCR) and sequencing approach in the diagnosis of FIP, detecting mutations at two different nucleotide positions within the spike (S) gene. Methods The study population consisted of 64 cats with confirmed FIP and 63 cats in which FIP was initially suspected due to similar clinical or laboratory signs, but that were definitively diagnosed with another disease. Serum/plasma and/or effusion samples of these cats were examined for feline coronavirus (FCoV) RNA by RT-nPCR and, if positive, PCR products were sequenced for nucleotide transitions within the S gene. Results Specificity of RT-nPCR was 100% in all materials (95% confidence interval [CI] in serum/plasma 83.9-100.0; 95% CI in effusion 93.0-100.0). The specificity of the sequencing step could not be determined as none of the cats of the control group tested positive for FCoV RNA. Sensitivity of the 'combined RT-nPCR and sequencing approach' was 6.5% (95% CI 0.8-21.4) in serum/plasma and 65.3% (95% CI 50.4-78.3) in effusion. Conclusions and relevance A positive result is highly indicative of the presence of FIP, but as none of the control cats tested positive by RT-nPCR, it was not possible to confirm that the FCoV mutant described can only be found in cats with FIP. Further studies are necessary to evaluate the usefulness of the sequencing step including FCoV-RNA-positive cats with and without FIP. A negative result cannot be used to exclude the disease, especially when only serum/plasma samples are available.

Discrepancies between feline coronavirus antibody and nucleic acid detection in effusions of cats with suspected feline infectious peritonitis

“…Molecular methods have been used for detection of FCoV RNA in the effusions of cats with suspected FIP (Gut et al, 1999;Simons et al, 2004;Hornyák et al, 2012;Soma et al, 2013;Doenges et al, 2017;Felten et al, 2017;Longstaff et al, 2017). However, these methods display similar issues related to the diagnostic performances (lack of sensitivity and specificity).…”

Feline Infectious Peritonitis Virus Nsp5 Inhibits Type I Interferon Production by Cleaving NEMO at Multiple Sites

“…Spontaneous mutations of the S gene [6], ORF3abc [7,8], and ORF7ab [9,10] in the FCoV genome allow virions to efficiently replicate in macrophages and monocytes, ultimately resulting in FIPV [11]. Detection of FCoV S gene mutations is considered as a tool to diagnose FIPV [12,13]. FIP is typically characterized by a fibrinous and granulomatous serositis, protein-rich serous effusions, and pyogranulomatous lesions in several organs [14,15].…”

Limitations of using feline coronavirus spike protein gene mutations to diagnose feline infectious peritonitis

“…Two amino acid substitutions, M1058L and/or S1060A corresponding to nucleotide mutations 23531A>T/C and 23537T>G respectively in the S gene, together distinguished FCoVs found in the tissues of FIP cats from those found in the faeces of healthy cats without FIP in > 95% of cases. A subsequent study, which compared detection of FCoV by RT-qPCR alone to detection of FCoV by RT-qPCR combined with sequence analysis to confirm the presence of nucleotide mutations 23531A>T/C and 23537T>G, concluded that the addition of an assay for S gene mutation analysis did not alter the specificity of the FIP diagnosis, which was already 100%, but did decrease the sensitivity from 9.4 to 6.3% for serum/plasma samples and from 72 to 64% for effusion samples [19]. However, we have shown that the nucleotide mutations 23531A>T/C and 23537T>G are likely to be markers of systemic FCoV infection rather than FIP per se, being present in 91% of the FCoV-positive tissue samples from cats with FIP and 89% of the FCoV-positive tissue samples from cats without FIP [20].…”