Integrated genomic view of SARS-CoV-2 in India

Abstract

Background: India first detected SARS-CoV-2, causal agent of COVID-19 in late January 2020, imported from Wuhan, China. From March 2020 onwards, the importation of cases from countries in the rest of the world followed by seeding of local transmission triggered further outbreaks in India. Methods: We used ARTIC protocol-based tiling amplicon sequencing of SARS-CoV-2 (n=104) from different states of India using a combination of MinION and MinIT sequencing from Oxford Nanopore Technology to understand how introduction and local transmission occurred. Results: The analyses revealed multiple introductions of SARS-CoV-2 genomes, including the A2a cluster from Europe and the USA, A3 cluster from Middle East and A4 cluster (haplotype redefined) from Southeast Asia (Indonesia, Thailand and Malaysia) and Central Asia (Kyrgyzstan). The local transmission and persistence of genomes A4, A2a and A3 was also observed in the studied locations. The most prevalent genomes with patterns of variance (confined in a cluster) remain unclassified, and are here proposed as A4-clade based on its divergence within the A cluster. Conclusions: The viral haplotypes may link their persistence to geo-climatic conditions and host response. Multipronged strategies including molecular surveillance based on real-time viral genomic data is of paramount importance for a timely management of the pandemic.

Keywords: COVID; COVID-19; MinION; SARS-CoV-2; Whole Genome Sequencing; viral genomes.

Figure 1.. Haplotype network and phylogenetic analysis of SARS-CoV-2 sequences.

( A) The network analysis (integer neighbourhood joining network at reticulation tolerance value of 0.5, popART) of SARS-CoV-2 sequences from this study showing distinct clades with their geographical locations. A4 clade described in this study for the first time has widespread geographical affiliations. A3 being more confined to Ladakh and mostly these clusters represent introduction of the infection through travel history. Mutations are marked with hatch lines connecting the nodes. ( B) Phylogenetic tree (generated using MEGA) of SARS-CoV-2 genomes from sequences submitted from across India depicting major clade-based distribution of SARS-CoV-2 in India. ( C) Network analysis (Median spanning network) of A4 clade sequences submitted from Indian and neighbouring East Asians countries.

Figure 2.. Distribution of A4 clade within India and globally.

( A) Distribution of A4 clade variants across different geographical regions from the cohort. ( B) Distribution of A4 clade variants across different geographical regions across the globe. ( C) Comparison of A4 cluster sequences across the South-East Asian region showing sharing of variants and haplotype across South-East Asian region.

Figure 3.. Mapping of high frequency mutations on proteins displaying top-most mutations in SARS-CoV2.

( A) Nucleocapsid, the N- and C-terminal domains are coloured in red and purple, respectively, where the missing linker region is shown as dotted line. ( B) NSP12/RdRp, showing the five of its domains, NIRAN (olive), Palm (red), Thumb (green), Fingers(blue) and Interface(orange). ( C) Nucleic acid-binding domain (NAB) of Nsp3 protein shown in pink color. ( D) The trimeric spike protein showing 3 chain like structure. The right panel shows a single chain where the NTD, RBD and HR1 domains are coloured in orange, purple and green, respectively, whereas the rest of the structure is coloured grey. The mutations in all the proteins are marked within black boxes shown in yellow spheres.