Whole-Genome Sequencing and Genetic Diversity of Human Respiratory Syncytial Virus in Patients with Influenza-like Illness in Sicily (Italy) from 2017 to 2023

Abstract

Monitoring the genetic variability of human respiratory syncytial virus (hRSV) is of paramount importance, especially for the potential implication of key antigenic mutations on the emergence of immune escape variants. Thus, to describe the genetic diversity and evolutionary dynamics of hRSV circulating in Sicily (Italy), a total of 153 hRSV whole-genome sequences collected from 770 hRSV-positive subjects between 2017 and 2023, before the introduction of expanded immunization programs into the population, were investigated. The phylogenetic analyses indicated that the genotypes GA.2.3.5 (ON1) for hRSV-A and GB.5.0.5a (BA9) for hRSV-B co-circulated in our region. Amino acid (AA) substitutions in the surface and internal proteins were evaluated, including the F protein antigenic sites, as the major targets of immunoprophylactic monoclonal antibodies and vaccines. Overall, the proportion of AA changes ranged between 1.5% and 22.6% among hRSV-A, whereas hRSV-B varied in the range 0.8-16.9%; the latter was more polymorphic than hRSV-A within the key antigenic sites. No AA substitutions were found at site III of both subgroups. Although several non-synonymous mutations were found, none of the polymorphisms known to potentially affect the efficacy of current preventive measures were documented. These findings provide new insights into the global hRSV molecular epidemiology and highlight the importance of defining a baseline genomic picture to monitor for future changes that might be induced by the selective pressures of immunological preventive measures, which will soon become widely available.

Keywords: Italy; Sicily; amino acid change; community; mAb; molecular surveillance; respiratory syncytial virus; vaccine; whole-genome sequencing.

Figure 1

Clade assignment of Sicilian hRSV-A whole-genome sequences, as defined by Nextstrain.

Figure 2

Clade assignment of Sicilian hRSV-B whole-genome sequences, as defined by Nextstrain.

Figure 3

Neighbor-joining phylogenetic tree of whole-genome nucleotide sequences of hRSV-A strains collected in Sicily between October 2017 and April 2023. The study sequences are indicated in blue and by solid circles, differently colored according to the lineage. Reference sequences are indicated in black and are expressed in the following format: subgroup/country/strain/year–GISAID accession number. The strain A2 (GenBank: KT992094) was used as an outgroup (indicated as a black dot). Genotypes are defined according to the classification proposed by Goya S et al., 2020 [43].

Figure 4

Neighbor-joining (NJ) phylogenetic tree of whole-genome nucleotide sequences of hRSV-B strains collected in Sicily between October 2017 and April 2023. The study sequences are indicated in blue and by solid circles, differently colored according to the lineage. Reference sequences are indicated in black and are expressed in the following format: subgroup/country/strain/year–GISAID accession number. The strain 9320 (GenBank: AY353550) was used as an outgroup (indicated as a black dot). Genotypes are defined according to the classification proposed by Goya S et al., 2020 [43].

Figure 5

Amino acid variation frequencies detected in the F protein open reading frame of hRSV-A and hRSV-B Sicilian strains. Proportion of mutated AA positions: 6.3% for hRSV-A and 4.5% for hRSV-B.