Detection and characterization of novel luchacoviruses, genus Alphacoronavirus, shed in saliva and feces of meso-carnivores in the northeastern United States

Small to mid-sized carnivores, or meso-carnivores, comprise a group of diverse mammals, many of which can adapt to anthropogenically disturbed environments. Wild meso-carnivores living in urban areas may get exposed to or spread pathogens to other species, including stray/feral domestic animals. Several coronaviruses (CoVs) have been detected in domesticated and farmed meso-carnivores, but knowledge of CoVs circulating in free-ranging wild meso-carnivores remains limited. In this study, we analyzed 321 samples collected between 2016 and 2022 from 9 species of free-ranging wild meso-carnivores and stray/feral domestic cats in the northeastern United States. Using a pan-CoV PCR, we screened tissues, feces, and saliva, nasal, and rectal swabs. We detected CoV RNA in fecal and saliva samples of animals in four species: fisher (Pekania pennanti), bobcat (Lynx rufus), red fox (Vulpes vulpes), and domestic cat (Felis catus). Next-generation sequencing revealed that all these viruses belonged to the Luchacovirus subgenus (Alphacoronavirus genus), previously reported only in rodents and lagomorphs (i.e., rabbits). Genetic comparison of the 3’-end of the genome (~12,000bp) revealed that although the viruses detected group with, and have a genetic organization similar to other luchacoviruses, they are genetically distinct from those from rodents and lagomorphs. Genetic characterization of the spike protein revealed that the meso-carnivore luchacoviruses do not have an S1/S2 cleavage motif but do have highly variable structural loops containing cleavage motifs similar to those identified in certain pathogenic CoVs. This study highlights the importance of characterizing the spike protein of CoVs in wild species for further targeted epidemiologic monitoring.