Molecular Epidemiology and Genetic Diversity of Human Respiratory Syncytial Virus in Sicily during Pre- and Post-COVID-19 Surveillance Seasons

Abstract

Human respiratory syncytial virus (hRSV) is an important pathogen of acute respiratory tract infection of global significance. In this study, we investigated the molecular epidemiology and the genetic variability of hRSV over seven surveillance seasons between 2015 and 2023 in Sicily, Italy. hRSV subgroups co-circulated through every season, although hRSV-B mostly prevailed. After the considerable reduction in the circulation of hRSV due to the widespread implementation of non-pharmaceutical preventive measures during the COVID-19 pandemic, hRSV rapidly re-emerged at a high intensity in 2022-2023. The G gene was sequenced for genotyping and analysis of deduced amino acids. A total of 128 hRSV-A and 179 hRSV-B G gene sequences were obtained. The phylogenetic analysis revealed that the GA2.3.5a (ON1) and GB5.0.5a (BA9) genotypes were responsible for the hRSV epidemics in Sicily.; only one strain belonged to the genotype GB5.0.4a. No differences were observed in the circulating genotypes during pre- and post-pandemic years. Amino acid sequence alignment revealed the continuous evolution of the G gene, with a combination of amino acid changes specifically appearing in 2022-2023. The predicted N-glycosylation sites were relatively conserved in ON1 and BA9 genotype strains. Our findings augment the understanding and prediction of the seasonal evolution of hRSV at the local level and its implication in the monitoring of novel variants worth considering in better design of candidate vaccines.

Keywords: BA9; GA2.3.5a; GB5.0.4a; GB5.0.5a; Italy; ON1; Sicily; genotype; hRSV-A; hRSV-B; molecular epidemiology.

Figure 1

Seasonal distribution of hRSV-A and hRSV-B positive samples identified in Sicily between 2015–2016 and 2022–2023.

Figure 2

Neighbor-joining (NJ) phylogenetic tree of nucleotide sequences from the second hypervariable region of the G gene of hRSV-A strains collected in Sicily between 2015 and 2023. The study sequences are indicated by solid circles, differently colored according to the surveillance season. Reference sequences are indicated in bold and are expressed in the following format: Genotype/GenBank accession number/country/year. The strain GA1 Y911262_USA_1956 was used as an outgroup. Genotypes are defined according to the classification proposed by Goya S et al. [10].

Figure 3

Neighbor-joining (NJ) phylogenetic tree of nucleotide sequences from the second hypervariable region of the G gene of hRSV-B strains collected in Sicily between 2015 and 2023. The study sequences are indicated by solid circles, differently colored according to the surveillance season. The unique strain belonging to GB5.0.4a is indicated red and highlighted in gray. Reference sequences are indicated in bold and are expressed in the following format: Genotype/GenBank accession number/country/year. The strain GB1 M73545_Sweden_1960 was used as an outgroup. Genotypes are defined according to the classification proposed by Goya S et al. [10].

Figure 4

Mutations of the deduced amino acid sequence of the second hypervariable region of the G gene of hRSV-A (A) and hRSV-B (B) Sicilian strains. * indicates a stop codon.

Figure 5

Amino acid variability of the second hypervariable region of the G gene represented by Shannon entropy plot. The threshold value was set at 0.2. Amino acid sites with entropy values < 0.2 are considered conserved, while values > 0.2 are considered variable. Horizontal blue and red bars indicate the analogous AA region followed by the duplicated AA region (23-amino acid for hRSV-A and 20-amino acid for hRSV-B, respectively).