Evolutionary dynamics of Erinaceus coronavirus in Italy: spike protein adaptation and variable CD200 ortholog insertion

Introduction In 2013, a novel coronavirus (CoV) species was identified in European hedgehogs ( Erinaceus europaeus ) in Germany. Since then, only a limited number of studies reported the circulation of this virus, recognized as Erinaceus coronavirus (EriCoV), and demonstrated its evolutionary relationship with Middle East respiratory syndrome coronavirus (MERS-CoV). Given that MERS-CoV is one of the three zoonotic coronaviruses known to cause severe disease in humans, these findings raise concerns about the zoonotic potential of EriCoV strains. In a previous study conducted in Italy, eight complete EriCoV genomes were obtained from hedgehogs sampled in Northern Italy, revealing substantial genetic heterogeneity among circulating strains. Genome analyses also identified the insertion of a gene encoding the CD200 ortholog, containing a short region potentially involved in host immune modulation. The present study aimed to investigate the circulation and genetic diversity of coronaviruses in hedgehogs from different areas of Italy and to assess the occurrence and distribution of the CD200 ortholog insertion. Methods Complete EriCoV genomes were obtained from 35 hedgehogs using Next Generation Sequencing (NGS) by Ion Torrent machine. For an additional 20 animals, full genome sequences could not be generated; therefore, sequences were obtained by nested RT-PCR amplifying partial sequences of the RNA-dependent RNA polymerase (RdRp) gene and, when present, on the CD200 insertion. Phylogenetic analysis of sequences obtained were performed. Results Phylogenetic analysis revealed no clear geographic clustering, with sequences from Northern Italy interspersed with those from Central and Southern Italy. Analysis of the spike (S) protein identified 25 sites evolving under positive selection, three of which were located within the receptor-binding domain (RBD). The CD200 ortholog was confirmed in 15 complete genomes and in 5 additional samples by RT-PCR, collected from both Northern and Southern Italy. Discussion Overall, the study confirms that EriCoV related strains are found across the country. A CD200 insertion appears in both Northern and Southern regions, but it is not consistently present, indicating that this insertion is not fixed within the viral population. Sites found under positive selection in spike protein, including three in the receptor-binding domain, that may be relevant for host–virus interactions and merit further investigation in the context of cross-species transmission risks.