Project Summary Porcine Hemagglutinating Encephalomyelitis Virus (PHEV), a Betacoronavirus, has historically been associated with neurological diseases in swine and demonstrates wide seroprevalence in the US swine herds. However, recent shifts in clinical presentations suggest that PHEV may also be causing respiratory illnesses. This project aims to investigate interactions with Neural Cell Adhesion Molecule (NCAM-1). NCAM-1 is typically associated with neural cells but may also be present in the respiratory epithelium, suggesting a mechanism for PHEV's expanded tropism. Additionally, the high seroprevalence of PHEV demonstrates a good model to develop an in situ hybridization technique used in surveillance monitoring. Introduction PHEV belongs to the Betacoronavirus genus, sharing characteristics with other viruses like Bovine Coronavirus (BCoV) and SARS-CoV-2. PHEV primarily infects pigs, and while it has traditionally been neurotropic, recent findings indicate that it might also cause respiratory diseases. Serological studies have demonstrated a global dissemination of PHEV; including regions in Asia, Europe, the Americas, and Australia. Specifically, the United States has reports of seroconversion rates ranging from 11-99%. Additionally, the viral spike (S) glycoprotein of PHEV is critical in determining the cellular tropism, and facilitating entry into the host’s cells by interacting with NCAM-1. It has been demonstrated that NCAM-1 expression is present in neurons and microglia, where it mediates neurologic disease. However, there is a knowledge gap with whether or not NCAM-1 is expressed in respiratory epithelium. This receptor interaction could explain the dual tropism seen in PHEV, where it can cause both neurological and respiratory disease. Preliminary Data A retrospective investigation at Iowa State University's Veterinary Diagnostic Laboratory (ISU-VDL) analyzed 15 respiratory disease cases from swine between 2019 and 2021. These cases exhibited a necrotizing bronchitis and bronchointerstitial pneumonia of undetermined etiology. A qPCR targeting the spike gene confirmed the presence of PHEV in 72.2% of those cases. Phylogenetic analysis revealed that respiratory strains of PHEV clustered separately from neurological strains, suggesting genotypic differences. In situ hybridization (ISH) confirmed the localization of PHEV mRNA in bronchiolar epithelial cells, while immunohistochemistry (IHC) demonstrated the presence of NCAM-1 in the respiratory epithelium, supporting the hypothesis that this receptor may play a role in PHEV's respiratory infection.