Table_4_Evolution of monkeypox virus from 2017 to 2022: In the light of point mutations.XLSX

Monkeypox virus (MPXV) causing multi-country outbreak-2022 is related to viruses caused outbreak-2017–2018 in West Africa. Still not fully understood which proteins of the MPXV discovered in Nigeria in 2017 have mutated through different lineages to the extent that it could cause a multi-country outbreak in 2022; similarly, codon usage bias, host adaptation indices, and the role of selection or mutation pressure in the mutated genes are also not fully studied. Here we report that according to the available sequence data this monkeypox virus acquires point mutations in multiple proteins in each period, and these point mutations accumulate and become a virus that can root outbreak-2022. Viruses exported from Nigeria to Singapore, Israel, and the United Kingdom in 2018–2019 were developed as evolutionary ancestors to B.1 viruses (MPXVs causing multi-country outbreak-2022) through MPXV/United States/2021/MD virus. Although these exported viruses have different amino acid mutations in different proteins, amino acid mutations in 10 proteins are common among them. The MPXV-United Kingdom-P2 virus evolved with only mutations in these 10 proteins and further evolved into MPXV/United States/2021/MD with amino acid mutations in 26 (including amino acid mutations in 10 proteins of the MPXV-United States-P2) proteins. It is noteworthy that specific amino acid mutations in these 22/26 (presence in MPXV/United States/2021/MD) proteins are present in B.1 viruses. Further, analysis of Relative Synonymous Codon Usage (RSCU), Synonymous Codon Usage Fraction (SCUF), and Effective Number of Codons (ENc) revealed codon usage bias in genes that exhibited nucleotide mutations in lineage B.1. Also, host adaptation indices analyzes such as Codon Adaptation Index (CAI), Expected-CAI (eCAI), Relative Codon Deoptimization Index (RCDI) and Expected value for the RCDI (eRCDI) analyzes reveal that the genes that demonstrated nucleotide mutations in lineage B.1 are favorable for human adaptation. Similarly, ENc-GC3s plot, Neutrality plot, and Parity Rule 2 (PR2)-bias plot analyzes suggest a major role of selection pressure than mutation pressure in the evolution of genes displaying nucleotide mutations in lineage B.1. Overall, from 2017 to 2022, MPXV’s mutation and spread suggests that this virus continues to evolve through point mutation in the genes according to the available sequence data.

2022年引发多国暴发的猴痘病毒（Monkeypox virus, MPXV），与2017-2018年西非暴发的相关病毒存在关联。目前尚未完全明确，2017年于尼日利亚发现的MPXV的哪些蛋白通过不同谱系发生突变，以至于能够引发2022年的多国暴发；类似地，密码子使用偏好性、宿主适应指数，以及突变基因中选择压力或突变压的作用，也未得到充分研究。
本研究基于现有序列数据发现，该猴痘病毒在各阶段的多个蛋白中均获得点突变，此类点突变不断累积，最终形成可引发2022年暴发的病毒毒株。
2018-2019年从尼日利亚出口至新加坡、以色列及英国的病毒，以MPXV/United States/2021/MD毒株为进化中间体，演化成为B.1谱系毒株（即引发2022年多国暴发的MPXV）。尽管这些出口毒株在不同蛋白中存在各异的氨基酸突变，但其中10种蛋白的氨基酸突变具有共通性。英国P2株MPXV仅通过这10种蛋白的氨基酸突变完成演化，后续进一步突变为MPXV/United States/2021/MD毒株，该毒株在26种蛋白中出现氨基酸突变（包含前述10种蛋白的突变）。
值得注意的是，MPXV/United States/2021/MD毒株中22/26种蛋白的特异性氨基酸突变，同样存在于B.1谱系毒株中。
进一步对相对同义密码子使用度（Relative Synonymous Codon Usage, RSCU）、同义密码子使用分数（Synonymous Codon Usage Fraction, SCUF）以及有效密码子数（Effective Number of Codons, ENc）开展分析，结果显示B.1谱系中发生核苷酸突变的基因存在密码子使用偏好性。
宿主适应指数相关分析，包括密码子适应指数（Codon Adaptation Index, CAI）、预期密码子适应指数（Expected-CAI, eCAI）、相对密码子去优化指数（Relative Codon Deoptimization Index, RCDI）及其预期值（Expected RCDI, eRCDI），均表明B.1谱系中发生核苷酸突变的基因更适配人类宿主。
同理，ENc-GC3s绘图、中性绘图以及奇偶规则2（Parity Rule 2, PR2）偏倚绘图分析结果显示，在B.1谱系发生核苷酸突变的基因演化过程中，选择压力相较于突变压发挥了更为主要的作用。
综上，基于现有序列数据，2017年至2022年间MPXV的突变与传播过程表明，该病毒持续通过基因层面的点突变完成演化。