Coding-Complete Genome Sequences of Alpha and Delta SARS-CoV-2 Variants from Kamphaeng Phet Province, Thailand, from May to July 2021

ABSTRACT We report coding-complete genome sequences of 44 severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strains of the alpha and delta variants identified from patients in Kamphaeng Phet, Thailand. Two nonsense mutations in open reading frame 3a (ORF3a) (G254*) and ORF8 (K68*) were found in the alpha variant sequences. Two lineages of the delta variant, B.1.617.2 and AY.30, were found.

, GISAID clade nomenclature (10), and phylogenetic analysis (11)(12)(13) were used to determine SARS-CoV-2 lineages. Nextclade v1.6.0 (14) was used to identify variants. All tools were run with default parameters. b Nucleotide and amino acid substitutions and annotation were analyzed using an in-house bioinformatics pipeline (19). All alpha variant sequences were aligned with the first alpha variant sequence collected in Thailand (GISAID accession number EPI_ISL_1346626), which was collected on 21 December 2020. All delta variant sequences were aligned with the first delta variant sequence collected in Thailand (GISAID accession number EPI_ISL_2104743), which was collected on 2 May 2021.
Individual genome characteristics are summarized in Table 1. The reads obtained were 35 to 251 nucleotides in length, and the average length was 217 nucleotides. Consensus sequences of coding regions were 29,387 to 29,402 bp in length, with the mean DOC ranging from 3,636Â to 8,642Â. Of 44 sequences, 20 and 24 were identified as alpha and delta variants, respectively. The alpha variants were found from 3 May 2021 to 18 July 2021, whereas the delta variants were found from 24 June 2021 to 18 July 2021. The phylogenetic tree is shown in Fig. 1.
Amino acid substitutions found in the alpha and delta variants from KPP when aligned with the sequences of the first corresponding variants collected in Thailand are shown in Table 1. Two nonsense mutations, i.e., G254* and K68* in open reading frame 3a (ORF3a) and ORF8 genes, respectively, were not found in the first alpha variant virus in Thailand but were found in all alpha variant viruses in this study. K68* was reported previously (15). G254* in ORF3a resulted in the predicted absence of 18 amino acid residues (positions 254 to 271) at the C terminus of the protein, located in a region thought to carry several B cell epitopes (16). Mutations in ORF3a were previously described as potentially having an impact on viral infectivity and pathogenesis (16)(17)(18). Among the 24 delta variant viruses from KPP, 8 sequences were identified as B.1.617.2 lineage and 16 sequences were identified as AY.30 lineage.
In conclusion, the two variants of concern, alpha and delta, were identified from May to July 2021 in KPP. Nonsense mutations in ORF3a and ORF8 were found in the alpha variant sequences. Two lineages of the delta variant were found.
Data availability. The sequences from this study were deposited in GenBank (accession numbers MZ888515 to MZ888557 and MZ895505). Individual accession numbers are indicated in Table 1. The raw reads were deposited in the NCBI Sequence Read Archive (SRA) (accession numbers SRR15571382 to SRR15571425). The BioProject accession number is PRJNA757144. The BioSample accession numbers are SAMN20934606 to SAMN20934649. FIG 1 Maximum likelihood phylogenetic tree of 47 SARS-CoV-2 coding sequences, including 44 sequences from this study (black), the first collected alpha variant sequence from Thailand (GISAID accession number EPI_ISL_1346626), which was collected on 21 December 2020 (green), the first collected delta variant sequence from Thailand (GISAID accession number EPI_ISL_2104743), which was collected on 2 May 2021 (green), and the reference Wuhan-Hu-1 genome sequence (GenBank accession number NC_045512.2) (blue). Multiple sequence alignments were performed using MAFFT v7.475 with default settings (11). The tree was constructed by using IQ-TREE v2.1.2 (12) with substitution model TIM21F1I and 1,000 ultrafast bootstrap replicates and was visualized by using FigTree v1.4.2 (13).