Feline infectious peritonitis (FIP) is a worldwide fatal disease caused by a mutant feline coronavirus (FCoV). Simple and efficient molecular detection methods are needed. Here, sensitive, specific, rapid, and reliable colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) was developed to detect the ORF1a/1b gene of FCoV from cats with suspected FIP using neutral red as an indicator. Novel LAMP primers were specifically designed based on the gene of interest. The isothermal assay could visually detect FCoV at 58 °C for 50 min. The RT-LAMP assay was highly specific and had no cross-reactivity with other related feline viruses. The detection limit of FCoV detection by RT-LAMP was 20 fg/µL. A blind clinical test (n = 81) of the developed RT-LAMP procedure was in good agreement with the conventional PCR method. In the light of its performance specificity, sensitivity, and easy visualization, this neutral-red-based RT-LAMP approach would be a fruitful alternative molecular diagnostic tool for veterinary inspection of FCoV when combined with nucleotide sequencing or specific PCR to affirm the highly virulent FIP-associated FCoV. 

https://www.mdpi.com/2076-2615/12/16/2075/pdf?version=1661247396

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

Background and Aim: Feline infectious peritonitis (FIP) is an infectious, immune-mediated, and fatal disease in cats caused by a mutant feline coronavirus (FCoV) infection. Feline immunodeficiency virus (FIV) and feline leukemia virus (FeLV) are two common retroviruses that play a role in reducing feline immune function with opportunistic retrovirus infection being a predisposing factor for the development of FIP. This study aimed to evaluate the clinicopathological parameters of FIP in cats with and without retrovirus coinfection. Materials and Methods: In total, 62 cats presenting with pleural and/or peritoneal effusion at the Kasetsart University Veterinary Teaching Hospital, Bangkok, Thailand, were selected for the study. Effusion samples were collected and a reverse transcriptase-polymerase chain reaction (RT-PCR) assay was performed on all samples using the 3’ untranslated region primer. All FCoV-positive cats were tested for retrovirus infection using a commercial kit (Witness FeLV-FIV [Zoetis]; United States). Clinical signs, hematological, and biochemical parameters of these cats were investigated and grouped. Results: Of the 62 cats with pleural and/or peritoneal effusion, FCoV was detected in 32, of which 21 were highly suspicious for FIP. The cats suspected of FIP were divided into three subgroups following viral detection. A total of 14 had only FCoV infection (Group A), four had FCoV and FeLV infection (Group B), and three had FCoV, FeLV, and FIV infection (Group C). Of the rest, 11 had definitive diagnoses, which included three being FCoV and FeLV-positive (Group D), and eight were retrovirus-negative (Group E). Mild anemia and lymphopenia were found in cats infected with these three viruses. An albumin-to-globulin ratio lower than 0.5 was found in FIP cats with only FCoV infection. Conclusion: Typically, cats with clinical effusion and FIP, with and without retrovirus coinfection, had similar hematological findings. Clinical signs, blood parameters, fluid analysis with cytological assessment, and RT-PCR assays could identify better criteria to diagnose FIP with and without retrovirus coinfection.

https://www.veterinaryworld.org/Vol.16/April-2023/20.pdf

The first study on clinicopathological changes in cats with feline infectious peritonitis with and without retrovirus coinfection

Feline infectious peritonitis (FIP) is a lethal infectious disease of domestic cats caused by feline coronavirus (FCoV) infection. Feline infectious peritonitis virus (FIPV) is a mutant type of FCoV that is characterized by causing fibrinous serositis with effusions in the pleural and abdominal cavities (wet form) and/or granulomatous-necrotizing inflammatory lesions in several organs (dry form). There have been numerous studies on FIP worldwide, whereas information about this disease in Thailand is still limited. Most studies involving molecular surveillance and evaluation of FCoV field strains have examined the genetic diversity of the spike and accessory ORF3c coding regions. Of these, the S gene is more divergent and is responsible for the two FCoV serotypes, while ORF3c harbors mutations that result either in early termination or destruction of the protein. In this study, we investigated the genetic diversity and genetic relationships among the current Thai and global FCoV strains in the accessory and nucleocapsid genes using a virus-specific PCR method. Comparative sequence analysis suggested that the Thai FCoV isolates were most closely related to strains reported in the Netherlands, the USA, and China. In the ORF3ab sequences, some Thai strains were more than 99% identical to the DF-2 prototype strain. Truncation of the 3a gene product was found in Thai FCoV strains of group 2. Amino acid deletions were observed in the N, ORF3c, and ORF7b proteins of Thai FCoV sequences. The accessory gene sequence divergence may be responsible for driving the periodic emergence and continued persistence of FCoVs in Thai domestic cat populations. Our findings provide updated information about the molecular characteristics of the accessory and nucleocapsid genes of FCoV strains in circulation that were not previously documented in this country.

/pdf/comparative-sequence-analysis-of-the-accessory-and-132amp0x0b.pdf

Comparative sequence analysis of the accessory and nucleocapsid genes of feline coronavirus strains isolated from cats diagnosed with effusive feline infectious peritonitis.

Supansa Tuanthap

Papers

Comparative sequence analysis of the accessory and nucleocapsid genes of feline coronavirus strains isolated from cats diagnosed with effusive feline infectious peritonitis.