Detection of anti-feline infectious peritonitis virus activity of a Chinese herb extract using geneLEAD VIII, a fully automated nucleic acid extraction/quantitative PCR testing system

Abstract

The geneLEAD VIII is a fully-automated nucleic acid extraction/quantitative PCR equipment developed by Precision System Science Co., Ltd., (PSS). To take advantage of its capability, we developed a quantitative assay system to measure growth of animal viruses. The system was used to assay one of the Chinese herbal extracts whose anti-malarial activities were previously reported and demonstrated its dose-dependent anti-viral activity against feline infectious peritonitis virus (FIPV), a feline coronavirus causing the fatal diseases in cats, and relatively low cell toxicity. The assay developed in this study is useful to screen antiviral drugs and the anti-FIPV activity of the herbal extract identified have a potential to lead to development of new drugs against FIPV and other coronaviruses, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

Keywords: Chinese herbal extract; assay; automated-qPCR; drug; feline infectious peritonitis virus.

Fig. 1.

A. Inhibition of the growth of feline infectious peritonitis virus (FIPV) 79–1146 strain with CsA and a control drug, FK506. After inoculation of the FIPV, fcwf-4 cells were incubated for 22 hr with the serial dilutions of the indicated drugs and culture supernatants were collected to quantitate the viral genome copies with geneLEAD VIII. Values and error bars in the figure indicate averages and standard errors of the percentages to the control of three independent measurements. B. Cytotoxicity of CsA and a control drug, FK506, to fcwf-4. Cell Counting Kit-8 was used to quantitate viability of fcwf-4 cells after incubation for 22 hr with the serial dilutions of the drugs. Values and error bars in the figure indicate averages and standard errors of the percentages to the control of three independent measurements.

Fig. 2.

A. Inhibition of the growth of feline infectious peritonitis virus (FIPV) 79–1146 strain with an herbal extract, D3. After inoculation of the FIPV, fcwf-4 cells were incubated for 22 hr with serial dilutions of D3, and culture supernatants were collected to quantitate extracellular viral copies and supernatants from the infected cells after three-times freeze/thaw at −80°C and a centrifuge at 10,000 × g for 10 min at 4°C to quantitate intracellular viral copies with geneLEAD VIII. Values and error bars in the figure indicate averages and standard errors of the percentages to the control of the three independent measurements. B. Cytotoxicity of an herbal extract, D3, to fcwf-4. Cell Counting Kit-8 was used to quantitate viability of fcwf-4 cells after incubation for 22 hr with the serial dilutions of D3. Values and error bars in the figure indicate averages and standard errors of the percentages to the control of three independent measurements.