Coronavirus Disease 2019 (COVID-19) is caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), a newly emerged coronavirus, and has been pandemic since March 2020 and led to many fatalities. Vaccines represent the most efficient means to control and stop the pandemic of COVID-19. However, currently there is no effective COVID-19 vaccine approved to use worldwide except for two human adenovirus vector vaccines, three inactivated vaccines, and one peptide vaccine for early or limited use in China and Russia. Safe and effective vaccines against COVID-19 are in urgent need. Researchers around the world are developing 213 COVID-19 candidate vaccines, among which 44 are in human trials. In this review, we summarize and analyze vaccine progress against SARS-CoV, Middle-East respiratory syndrome Coronavirus (MERS-CoV), and SARS-CoV-2, including inactivated vaccines, live attenuated vaccines, subunit vaccines, virus like particles, nucleic acid vaccines, and viral vector vaccines. As SARS-CoV-2, SARS-CoV, and MERS-CoV share the common genus, Betacoronavirus, this review of the major research progress will provide a reference and new insights into the COVID-19 vaccine design and development.

The current pandemic of COVID-19 is fueled by more infectious emergent Omicron variants. Ongoing concerns of emergent variants include possible recombinants, as genome recombination is an important evolutionary mechanism for the emergence and re-emergence of human viral pathogens. In this study, we identified diverse recombination events between two Omicron major subvariants (BA.1 and BA.2) and other variants of concern (VOCs) and variants of interest (VOIs), suggesting that co-infection and subsequent genome recombination play important roles in the ongoing evolution of SARS-CoV-2. Through scanning high-quality completed Omicron spike gene sequences, 18 core mutations of BA.1 (frequency >99%) and 27 core mutations of BA.2 (nine more than BA.1) were identified, of which 15 are specific to Omicron. BA.1 subvariants share nine common amino acid mutations (three more than BA.2) in the spike protein with most VOCs, suggesting a possible recombination origin of Omicron from these VOCs. There are three more Alpha-related mutations in BA.1 than BA.2, and BA.1 is phylogenetically closer to Alpha than other variants. Revertant mutations are found in some dominant mutations (frequency >95%) in the BA.1. Most notably, multiple characteristic amino acid mutations in the Delta spike protein have been also identified in the “Deltacron”-like Omicron Variants isolated since November 11, 2021 in South Africa, which implies the recombination events occurring between the Omicron and Delta variants. Monitoring the evolving SARS-CoV-2 genomes especially for recombination is critically important for recognition of abrupt changes to viral attributes including its epitopes which may call for vaccine modifications.

250 words 25 Importance: 150 words 26 Abstract 28 A novel coronavirus SARS-CoV-2 is associated with the current global pandemic of Coronavirus 29Disease 2019 . Spike protein receptor-binding domain (RBD) of SARS-CoV-2 is the 30 critical determinant of viral tropism and infectivity. To investigate whether the mutations in the RBD 31 have altered the receptor binding affinity and caused these strains more infectious, we performed 32 molecular dynamics simulations of the binding affinity between the mutant SARS-CoV-2 RBDs to 33 date and the human ACE2 receptor. Among 1609 genomes of global SARS-CoV-2 strains, 32 34 non-synonymous RBD mutants were identified and clustered into 9 mutant types under high positive 35 selection pressure. Three mutant types (V367F, W436R, and D364Y) emerging in Wuhan, Shenzhen, 36 Hong Kong, and France, displayed higher human ACE2 affinity, and probably higher infectivity. This 37 is due to the enhanced structural stabilization of the RBD beta-sheet scaffold. High frequencies of 38 RBD mutations were identified: V367F from five France and one Hong Kong mutants, 13 V483A and 39 7 G476S mutants from the U.S.A. This suggested they originated as novel sub-lineages. The 40 enhancement of the binding affinity of the mutant type (V367F) was further validated by the 41 receptor-ligand binding ELISA assay. The molecular dynamics simulations also indicated that it 42 would be difficult for bat SARS-like CoV to infect humans. However, the pangolin CoV is potentially 43 infectious to humans. The analysis of critical RBD mutations provides further insights into the 44 evolutionary history of SARS-CoV-2 under high selection pressure. An enhancement of the 45 SARS-CoV-2 binding affinity to human ACE2 receptor reveals higher infectivity of the mutant 46 strains. 47 48 Importance 49 A novel coronavirus SARS-CoV-2 has caused the pandemic of COVID-19. The origin of 50 SARS-CoV-2 was associated with zoonotic infections. The spike protein receptor-binding domain 51 (RBD) is identified as the critical determinant of viral tropism and infectivity. Thus, whether the 52 mutations in the RBD of the circulating SARS-CoV-2 strains have altered the receptor binding 53affinity and caused these strains more infectious, should be paid more attentions to. Here, 32 54 non-synonymous RBD mutants were identified and clustered into 9 mutant types under high positive 55 . CC-BY-NC-ND 4.0 International license (which was not certified by peer review) is the author/funder. It is made available under a selection pressure, suggesting they originated as novel sub-lineages. Three mutant types displayed 56 higher human ACE2 affinity, and probably higher infectivity, one of which (V367F) was validated 57 by wet bench. The RBD mutation analysis provides insights into SARS-CoV-2 evolution. The 58 emergence of RBD mutations with increased human ACE2 affinity reveals higher risk of severe 59 morbidity and mortality during a sustained COVID-19 pandemic, particularly if no effective 60 precautions are implemented. 61 62

The current pandemic of COVID-19 is caused by a novel coronavirus SARS-CoV-2. The SARS-CoV-2 spike protein receptor-binding domain (RBD) is the critical determinant of viral tropism and infectivity. To investigate whether naturally occurring RBD mutations during the early transmission phase have altered the receptor binding affinity and infectivity, firstly we analyzed in silico the binding dynamics between SARS-CoV-2 RBD mutants and the human ACE2 receptor. Among 32,123 genomes of SARS-CoV-2 isolates (January through March, 2020), 302 non-synonymous RBD mutants were identified and clustered into 96 mutant types. The six dominant mutations were analyzed applying molecular dynamics simulations (MDS). The mutant type V367F continuously circulating worldwide displayed higher binding affinity to human ACE2 due to the enhanced structural stabilization of the RBD beta-sheet scaffold. The MDS also indicated that it would be difficult for bat SARS-like CoV to infect humans. However, the pangolin CoV is potentially infectious to humans. The increased infectivity of V367 mutants was further validated by performing receptor-ligand binding ELISA, surface plasmon resonance, and pseudotyped virus assays. Phylogenetic analysis of the genomes of V367F mutants showed that during the early transmission phase, most V367F mutants clustered more closely with the SARS-CoV-2 prototype strain than the dual-mutation variants (V367F + D614G) which may derivate from recombination. The analysis of critical RBD mutations provides further insights into the evolutionary trajectory of early SARS-CoV-2 variants of zoonotic origin under negative selection pressure and supports the continuing surveillance of spike mutations to aid in the development of new COVID-19 drugs and vaccines. Importance A novel coronavirus SARS-CoV-2 has caused the pandemic of COVID-19. The origin of SARS-CoV-2 was associated with zoonotic infections. The spike protein receptor-binding domain (RBD) is identified as the critical determinant of viral tropism and infectivity. Thus, whether the mutations in the RBD of the circulating SARS-CoV-2 isolates have altered the receptor binding affinity and made them more infectious, has been the research hotspot. Given that SARS-CoV-2 is a novel coronavirus, the significance of our research is in identifying and validating the RBD mutant types emerging during the early transmission phase and increasing human ACE2 receptor binding affinity and infectivity. Our study provides insights into the evolutionary trajectory of early SARS-CoV-2 variants of zoonotic origin. The continuing surveillance of RBD mutations with increased human ACE2 affinity in human or other animals is critical to the development of new COVID-19 drugs and vaccines against these variants during the sustained COVID-19 pandemic.