<p>Porcine hemagglutinating encephalomyelitis virus (PHEV), first identified in 1957, has been described as a neurotropic virus of pigs. However, more recent strains were mainly associated with respiratory problems. This study compared the replication kinetics of the historical neurotropic PHEV strain isolated in Belgium in 1972 (VW572) with the replication kinetics of a contemporary 2020 strain (PS412; isolated from nasal secretions during a monitoring study), using RPD (rein de porc diploid) cells, nasal and ethmoidal mucosa explants, and primary porcine respiratory epithelial cells (PoRECs). EGTA was used to disrupt intercellular junctions to facilitate the access of the virus to basolateral receptors. In RPD cells, VW572 showed an early advantage in initiating infection, whereas PS412 exhibited stronger replication and viral release at later stages, highlighting distinct replication dynamics between the two strains. The replication kinetics of both strains in nasal explants and PoRECs did not differ significantly. In ethmoidal explants, PHEV only replicated in sustentacular cells with VW572 infecting more cells than PS412. EGTA treatment had no effect on the PHEV infection in nasal explants but significantly increased in PoRECs and ethmoidal explants for VW572, suggesting a stronger reliance on basolateral receptors in sustentacular cells. Genomic analysis identified 2 spike protein mutations that led to amino acid changes at the receptor binding site: N512K, K576E. These changes switched the regional polarity on the surface of the spike protein that may affect the PHEV receptor use. In conclusion, the historical and contemporary PHEV isolates VW572 and PS412 replicate differently in the respiratory/neural tissues of the upper respiratory tract which may be associated with amino acid variations in the receptor binding domain of the spike protein.</p>