Effectiveness of the Fungal Metabolite 3-O-Methylfunicone towards Canine Coronavirus in a Canine Fibrosarcoma Cell Line (A72)

Abstract

Canine coronavirus (CCoV), an alphacoronavirus, may cause self-limiting enteric disease in dogs, especially in puppies. The noteworthy plasticity of coronaviruses (CoVs) occurs through mutation and recombination processes, which sometimes generate new dangerous variants. The ongoing SARS-CoV-2 pandemic and the isolation of a novel canine–feline recombinant alphacoronavirus from humans emphasizes the cross-species transmission ability of CoVs. In this context, exploring antiviral compounds is essential to find new tools for fighting against CoVs infections. Fungi produce secondary metabolites, which are often developed as antibiotics, fungicides, hormones, and plant growth regulators. Previous examinations of benzo-γ-pyrone 3-O-methylfunicone (OMF), obtained from Talaromyces pinophilus, showed that it reduces the infectivity of hepatitis C virus and bovine herpesvirus 1. Based on this evidence, this study evaluated the antiviral ability of OMF against CCoV infection in a canine fibrosarcoma (A72) cell line. During CCoV infection, a non-toxic dose of OMF markedly increased features of cell viability. Moreover, OMF induced a significant reduction in virus yield in the presence of an intense downregulation of the viral nucleocapsid protein (NP). These findings occurred in the presence of a marked reduction in the aryl hydrocarbon receptor (AhR) expression. Taken together, preliminary findings suggest that OMF inhibiting AhR shows promising activity against CCoV infection.

Fungal metabolite 6-pentyl-α-pyrone reduces canine coronavirus infection

“…We have performed Giemsa staining and acridine orange staining to study cell morphology [ 14 , 32 ] at 24 h and 48 h of infection on A72 and CRFK, treated with 6 PP, and infected with CCoV at MOI of 5 and at MOI of 1, respectively.…”

“…Monolayers of A72 and CRFK cells, treated or not with 6 PP, were infected or not with CCoV, at MOI 5 for A72 and at MOI 10 for CRFK, and incubated at 37 °C. Real-time PCR was used for CCoV quantification in A72 cells [ 14 , 22 ]. In CRFK cells, the virus titer was tested by 50% tissue culture infective dose (TCID 50 ) titration [ 29 , 37 ].…”

“…RT-qPCR was used for quantifying CCoV during infection in A72 cells [ 39 ]. Then, the quantification of CCoV was carried by the mean of a standard curve, making serial dilutions (from 3.5 × 10 9 to 3.5 × 10 2 TCID 50 /mL) of the quantified canine coronavirus viral stock and plotting threshold cycle number (Ct) against the TCID 50 /mL on a log scale [ 14 , 16 , 17 , 22 , 35 ].…”

“…In addition, some fungal molecules revealed antiviral properties [ [11] , [12] , [13] ]. As an example, studies about 3- O -methylfunicone (OMF), which is a benzo-γ-pyrone obtained by Talaromyces pinophilus , revealed that this compound is able to induce a decline in the infectivity of CCoV [ 14 ], hepatitis C virus [ 15 ], and bovine herpesvirus 1 [ 16 ]. Recently, other funicone-like substances, vermistatin (VER) as well as penisimplicissin (PS) have been evaluated for their in vitro inhibitory activity towards CCoV infection [ 17 ].…”

“…The Trichoderma fungal metabolite 6-pentyl-α-pyrone (6 PP) shows a significant role against plant pathogens [ 18 , 19 ], and in anti-biofilm-producing bacteria activity [ 20 , 21 ], but the antiviral activity of 6 PP has never been explored up to now. Hence, herein the potential efficacy of 6 PP anti-CCoV was tested using A72, which are canine fibrosarcoma cells, and Crandell-Rees Feline Kidney Cell (CRFK), a cell line resulting from the feline kidney, both are suitable for studying CCoV [ 14 , [22] , [23] , [24] ].…”