Molecular epidemiology of SARS-CoV-2 isolated from COVID-19 family clusters

Abstract

Background: Transmission within families and multiple spike protein mutations have been associated with the rapid transmission of SARS-CoV-2. We aimed to: (1) describe full genome characterization of SARS-CoV-2 and correlate the sequences with epidemiological data within family clusters, and (2) conduct phylogenetic analysis of all samples from Yogyakarta and Central Java, Indonesia and other countries.

Methods: The study involved 17 patients with COVID-19, including two family clusters. We determined the full-genome sequences of SARS-CoV-2 using the Illumina MiSeq next-generation sequencer. Phylogenetic analysis was performed using a dataset of 142 full-genomes of SARS-CoV-2 from different regions.

Results: Ninety-four SNPs were detected throughout the open reading frame (ORF) of SARS-CoV-2 samples with 58% (54/94) of the nucleic acid changes resulting in amino acid mutations. About 94% (16/17) of the virus samples showed D614G on spike protein and 56% of these (9/16) showed other various amino acid mutations on this protein, including L5F, V83L, V213A, W258R, Q677H, and N811I. The virus samples from family cluster-1 (n = 3) belong to the same clade GH, in which two were collected from deceased patients, and the other from the survived patient. All samples from this family cluster revealed a combination of spike protein mutations of D614G and V213A. Virus samples from family cluster-2 (n = 3) also belonged to the clade GH and showed other spike protein mutations of L5F alongside the D614G mutation.

Conclusions: Our study is the first comprehensive report associating the full-genome sequences of SARS-CoV-2 with the epidemiological data within family clusters. Phylogenetic analysis revealed that the three viruses from family cluster-1 formed a monophyletic group, whereas viruses from family cluster-2 formed a polyphyletic group indicating there is the possibility of different sources of infection. This study highlights how the same spike protein mutations among members of the same family might show different disease outcomes.

Keywords: COVID-19 severity; Family cluster; Multiple spike protein mutations; Phylogenetic analysis; SARS-CoV-2 transmission; Whole genome sequencing.

Fig. 1

Phylogenetic analysis of SARS-CoV-2 genomes from Indonesia and different countries. A phylogenetic tree was constructed from 29.409 nt length of the open reading frame (ORF) of 142 SARS-CoV-2 virus sequences using Neighbor Joining statistical method with 2,000 bootstrap replications. The evolutionary distances were computed using the Kimura 2-parameter method and the rate variation among sites was modelled with a gamma distribution (estimated α = 0.14566). SARS-CoV-2 virus sequences from Indonesia (N = 89) followed by the date of collection are indicated in closed circles, while viruses for the study are indicated in red and viruses from the family clusters are color-shaded in yellow. The tree is rooted to Wuhan/Hu-1/2019 with the bootstrap percentage values less than 70% hidden and it is drawn to scale (0.0001) with branch lengths measured in the number of substitutions per site

Fig. 2

Timeline of COVID-19 symptoms and diagnosis from family clusters in Yogyakarta and Central Java

Fig. 3

Phylogenetic analysis and nucleic acid differences of family clusters’ virus sequences compared to the other SARS-CoV-2 virus sequences from Yogyakarta and Central Java. a The tree was constructed from 29.409 nt length of the open reading frame (ORF) of 142 SARS-CoV-2 virus sequences using Neighbor Joining statistical method and computed using the Kimura 2-parameter method with 2,000 bootstrap replications. The tree is rooted to Wuhan/Hu-1/2019 (NC_045512.2) with the bootstrap percentage values less than 70% hidden and it is drawn to scale (0.0001) with branch lengths measured in the number of substitutions per site. b The number of base differences per sequence from between sequences are shown. Codon positions included were 1st + 2nd + 3rd + Noncoding. All ambiguous positions were removed for each sequence pair. There was a total of 29,409 positions in the final dataset. Family clusters are indicated in asterisks: Case-1 (*) and Case-2 (**)