Molecular Characterization of the First African Swine Fever Virus Genotype II Strains Identified from Mainland Italy, 2022

Abstract

African swine fever (ASF) is responsible for important socio-economic effects in the global pig industry, especially for countries with large-scale piggery sectors. In January 2022, the African swine fever virus (ASFV) genotype II was identified in a wild boar population in mainland Italy (Piedmont region). This study describes the molecular characterization, by Sanger and next-generation sequencing (NGS), of the first index case 632/AL/2022 and of another isolate (2802/AL/2022) reported in the same month, in close proximity to the first, following multiple ASF outbreaks. Phylogenetic analysis based on the B646L gene and NGS clustered the isolates 632/AL/2022 and 2802/AL/2022 within the wide and most homogeneous p72 genotype II that includes viruses from European and Asian countries. The consensus sequence obtained from the ASFV 2802/AL/2022 isolate was 190,598 nucleotides in length and had a mean GC content of 38.38%. At the whole-genome level, ASF isolate 2802/AL/2022 showed a close genetic correlation with the other representative ASFV genotype II strains isolated between April 2007 and January 2022 from wild and domestic pigs in Eastern/Central European (EU) and Asian countries. CVR subtyping clustered the two Italian ASFV strains within the major CVR variant circulating since the first virus introduction in Georgia in 2007. Intergenic region I73R-I329L subtyping placed the Italian ASFV isolates within the variant identical to the strains frequently identified among wild boars and domestic pigs. Presently, given the high sequence similarity, it is impossible to trace the precise geographic origin of the virus at a country level. Moreover, the full-length sequences available in the NCBI are not completely representative of all affected territories.

Keywords: ASF outbreak; African swine fever virus (ASFV); genotype II; mainland Italy; next-generation sequencing (NGS); phylogenetic analysis; wild boar population.

Figure 1

Phylogenetic tree reconstruction based on the C-terminal region of the B646L gene (p72). The sample ON108572 632/AL/2022 characterized in this study is shown in bold. Bootstrap values > 70 are indicated at their respective nodes. Bars indicate the number of nucleotide substitutions per site.

Figure 2

Maximum likelihood (ML) phylogenetic tree of 36 ASFV complete genome sequences. The sample ON108571 2802/AL/2022 characterized in this study is shown in bold. Bootstrap values > 70 are indicated at their respective nodes. Bars indicate the number of nucleotide substitutions per site.

Figure 3

Amino acid sequence of the tetrameric repeats that constitute the central variable region (CVR) of the B602L gene identified in Italy. The single letters refer to the code of each tetrameric repeat: B = CADT; N = NVDT/NVGT; D = CASM; A = CAST; L = CTST; H = NEDT; P = NADT; S = SAST; O = NASI; F = NAST; Q = NADI; V = NANT; M = NANI; T = NVNT; C = GAST; K = CANT [17,22,29].

Figure 4

Nucleotide sequence alignment of the partial intergenic region between I73R and I329L from African swine fever genotype II viruses. Viruses characterized in this study are indicated in bold.