Molecular Evolution of Human Coronavirus 229E in Russia

Human coronavirus 229E (hCoV-229E) is an endemic alphacoronavirus that causes mild to moderate respiratory infections in humans. Despite its long-term circulation in the human population, the genetic diversity and evolutionary dynamics of hCoV-229E remain insufficiently studied, especially in Russia. The aim of this study was to analyze the genetic diversity and molecular characteristics of hCoV-229E strains circulating in Russia from 2017 to 2025. A total of 69,053 nasopharyngeal swabs from hospitalized patients with symptoms of acute respiratory viral infections, mainly from Saint Petersburg, were analyzed. Respiratory pathogens were identified by multiplex RT-PCR. Whole-genome sequencing was performed on selected hCoV-229E-positive samples. Phylogenetic analysis was carried out using the maximum likelihood method, and amino acid substitutions in the S protein were analyzed. Potential N-linked glycosylation sites were predicted using NetNGlyc. hCoV-229E circulated during all eight epidemic seasons, with a pronounced spring peak; its proportion among acute respiratory viral infections ranged from 0.1% to 11%. The most common coinfections were combinations with SARS-CoV-2 (144 cases), human respiratory syncytial virus hRSV (62), and human rhinovirus hRV (48). Based on the whole-genome tree topology, we propose two novel genotypes, 7 and 8 (Bootstrap >70%), with genotype 8 further differentiating into sublineages 8a and 8b. Phylogenetic analysis demonstrated that all contemporary Russian isolates are assigned to these two genotypes. A total of 137 Russian whole-genome sequences were deposited in the international GenBank database, accounting for 23.22% of all available hCoV-229E genomes. In the receptor-binding domain (RBD), the amino acid substitutions E309D, R312S, and H314R (loop 1) were identified, which are characteristic of Russian genotype 8 isolates. The P358A substitution (loop 2) was found exclusively in three isolates from Saint Petersburg in 2023. Thirty-five potential N-glycosylation sites were predicted, of which 26 were conserved across all strains. At the same time, the N714 glycosylation site in the S2 subunit, present in the reference strain and in the 1989 isolate, was lost in all modern viruses. This study provides a comprehensive characterization of the circulation and evolution of hCoV-229E in Russia, demonstrating a shift in dominant genotypes, the formation of regional phylogenetic clades with unique molecular markers, and changes in the glycosylation profile. The results emphasize the need for continuous genomic monitoring of endemic coronaviruses for the timely detection of new genetic variants.