Question: What is the result of phylogenetic analysis of SARS-CoV-2 genome sequence?

Title: A snapshot of SARS-CoV-2 genome availability up to 30th March, 2020 and its implications Content: The SARS-CoV-2 pandemic has been growing exponentially, affecting nearly 900 thousand people and causing enormous distress to economies and societies worldwide. A plethora of analyses based on viral sequences has already been published, in scientific journals as well as through non-peer reviewed channels, to investigate SARS-CoV-2 genetic heterogeneity and spatiotemporal dissemination. We examined full genome sequences currently available to assess the presence of sufficient information for reliable phylogenetic and phylogeographic studies in countries with the highest toll of confirmed cases. Although number of-available full-genomes is growing daily, and the full dataset contains sufficient phylogenetic information that would allow reliable inference of phylogenetic relationships, country-specific SARS-CoV-2 datasets still present severe limitations. Studies assessing within country spread or transmission clusters should be considered preliminary at best, or hypothesis generating. Hence the need for continuing concerted efforts to increase number and quality of the sequences required for robust tracing of the epidemic. Significance Statement Although genome sequences of SARS-CoV-2 are growing daily and contain sufficient phylogenetic information, country-specific data still present severe limitations and should be interpreted with caution.

Title: A Snapshot of SARS-CoV-2 Genome Availability up to April 2020 and its Implications: Data Analysis Content: BACKGROUND: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has been growing exponentially, affecting over 4 million people and causing enormous distress to economies and societies worldwide. A plethora of analyses based on viral sequences has already been published both in scientific journals and through non-peer-reviewed channels to investigate the genetic heterogeneity and spatiotemporal dissemination of SARS-CoV-2. However, a systematic investigation of phylogenetic information and sampling bias in the available data is lacking. Although the number of available genome sequences of SARS-CoV-2 is growing daily and the sequences show increasing phylogenetic information, country-specific data still present severe limitations and should be interpreted with caution. OBJECTIVE: The objective of this study was to determine the quality of the currently available SARS-CoV-2 full genome data in terms of sampling bias as well as phylogenetic and temporal signals to inform and guide the scientific community. METHODS: We used maximum likelihood-based methods to assess the presence of sufficient information for robust phylogenetic and phylogeographic studies in several SARS-CoV-2 sequence alignments assembled from GISAID (Global Initiative on Sharing All Influenza Data) data released between March and April 2020. RESULTS: Although the number of high-quality full genomes is growing daily, and sequence data released in April 2020 contain sufficient phylogenetic information to allow reliable inference of phylogenetic relationships, country-specific SARS-CoV-2 data sets still present severe limitations. CONCLUSIONS: At the present time, studies assessing within-country spread or transmission clusters should be considered preliminary or hypothesis-generating at best. Hence, current reports should be interpreted with caution, and concerted efforts should continue to increase the number and quality of sequences required for robust tracing of the epidemic.

Title: A Snapshot of SARS-CoV-2 Genome Availability up to April 2020 and its Implications: Data Analysis Content: BACKGROUND: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has been growing exponentially, affecting over 4 million people and causing enormous distress to economies and societies worldwide. A plethora of analyses based on viral sequences has already been published both in scientific journals and through non–peer-reviewed channels to investigate the genetic heterogeneity and spatiotemporal dissemination of SARS-CoV-2. However, a systematic investigation of phylogenetic information and sampling bias in the available data is lacking. Although the number of available genome sequences of SARS-CoV-2 is growing daily and the sequences show increasing phylogenetic information, country-specific data still present severe limitations and should be interpreted with caution. OBJECTIVE: The objective of this study was to determine the quality of the currently available SARS-CoV-2 full genome data in terms of sampling bias as well as phylogenetic and temporal signals to inform and guide the scientific community. METHODS: We used maximum likelihood–based methods to assess the presence of sufficient information for robust phylogenetic and phylogeographic studies in several SARS-CoV-2 sequence alignments assembled from GISAID (Global Initiative on Sharing All Influenza Data) data released between March and April 2020. RESULTS: Although the number of high-quality full genomes is growing daily, and sequence data released in April 2020 contain sufficient phylogenetic information to allow reliable inference of phylogenetic relationships, country-specific SARS-CoV-2 data sets still present severe limitations. CONCLUSIONS: At the present time, studies assessing within-country spread or transmission clusters should be considered preliminary or hypothesis-generating at best. Hence, current reports should be interpreted with caution, and concerted efforts should continue to increase the number and quality of sequences required for robust tracing of the epidemic.

Title: Whole genome sequencing of SARS-CoV-2 isolated in Guangdong Province and factors influencing the sequencing/ 广东省新型冠状病毒全基因组序列测定及其影响因素分析 Content: Objective: To obtain the genome sequences of SARS-CoV-2 in respiratory specimens in Guangdong Province with next-generation sequencing (NGS) and analyze the factors influencing sequencing. Methods: Eight upper and lower respiratory tract specimens were collected from patients with SARS-CoV-2 infection in Guangdong Province in January 2020. RNA library construction was used to obtain the genome sequences of SARS-CoV-2. A bio-informatics software package (CLC Genomics Workbench 12.0) was used to analyze and compare the genomic sequences. Results: Five SARS-CoV-2 genome sequences were obtained from the eight specimens and two were obtained from lower respiratory tract specimens. The nucleotide homology to SARS-CoV-2 was 97.74%-99.90%. The Ct values were lower, while the sequencing depth, coverage, relative abundance and genome integrity were higher in sequencing the SARS-CoV-2 in lower respiratory tract specimens. Conclusions: The low Ct value of SARS-CoV-2 in the samples was good for sequencing.

Title: Early Phylogenetic Diversification of SARS-CoV-2: Determination of Variants and the Effect on Epidemiology, Immunology, and Diagnostics. Content: The phylogenetic clustering of 95 SARS-CoV-2 sequences from the first 3 months of the pandemic reveals insights into the early evolution of the virus and gives first indications of how the variants are globally distributed. Variants might become a challenge in terms of diagnostics, immunology, and effectiveness of drugs. All available whole genome sequence data from the NCBI database (March 16, 2020) were phylogenetically analyzed, and gene prediction as well as analysis of selected variants were performed. Antigenic regions and the secondary protein structure were predicted for selected variants. While some clusters are presenting the same variant with 100% identical bases, other SARS-CoV-2 lineages show a beginning diversification and phylogenetic clustering due to base substitutions and deletions in the genomes. First molecular epidemiological investigations are possible with the results by adding metadata as travelling history to the presented data. The advantage of variants in source tracing can be a challenge in terms of virulence, immune response, and immunological memory. Variants of viruses often show differences in virulence or antigenicity. This must also be considered in decisions like herd immunity. Diagnostic methods might not work if the variations or deletions are in target regions for the detection of the pathogen. One base substitution was detected in a primer binding site.

Title: Unsupervised cluster analysis of SARS-CoV-2 genomes reflects its geographic progression and identifies distinct genetic subgroups of SARS-CoV-2 virus Content: Over 10,000 viral genome sequences of the SARS-CoV-2 virus have been made readily available during the ongoing coronavirus pandemic since the initial genome sequence of the virus was released on the open access Virological website (http://virological.org/) early on January 11. We utilize the published data on the single stranded RNAs of 11, 132 SARS-CoV-2 patients in the GISAID (Elbe and Buckland-Merrett, 2017; Shu and McCauley, 2017) database, which contains fully or partially sequenced SARS-CoV-2 samples from laboratories around the world. Among many important research questions which are currently being investigated, one aspect pertains to the genetic characterization/classification of the virus. We analyze data on the nucleotide sequencing of the virus and geographic information of a subset of 7, 640 SARS-CoV-2 patients without missing entries that are available in the GISAID database. Instead of modelling the mutation rate, applying phylogenetic tree approaches, etc., we here utilize a model-free clustering approach that compares the viruses at a genome-wide level. We apply principal component analysis to a similarity matrix that compares all pairs of these SARS-CoV-2 nucleotide sequences at all loci simultaneously, using the Jaccard index (Jaccard, 1901; Tan et al., 2005; Prokopenko et al., 2016; Schlauch et al., 2017). Our analysis results of the SARS-CoV-2 genome data illustrates the geographic and chronological progression of the virus, starting from the first cases that were observed in China to the current wave of cases in Europe and North America. We also observe that, based on their sequence data, the SARS-CoV-2 viruses cluster in distinct genetic subgroups. It is the subject of ongoing research to examine whether the genetic subgroup could be related to diseases outcome and its potential implications for vaccine development.

Title: Regaining perspective on SARS-CoV-2 molecular tracing and its implications Content: During the past three months, a new coronavirus (SARS-CoV-2) epidemic has been growing exponentially, affecting over 100 thousand people worldwide, and causing enormous distress to economies and societies of affected countries. A plethora of analyses based on viral sequences has already been published, in scientific journals as well as through non-peer reviewed channels, to investigate SARS-CoV-2 genetic heterogeneity and spatiotemporal dissemination. We examined all full genome sequences currently available to assess the presence of sufficient information for reliable phylogenetic and phylogeographic studies. Our analysis clearly shows severe limitations in the present data, in light of which any finding should be considered, at the very best, preliminary and hypothesis-generating. Hence the need for avoiding stigmatization based on partial information, and for continuing concerted efforts to increase number and quality of the sequences required for robust tracing of the epidemic.

Title: Comparative genomics provides an operational classification system and reveals early emergence and biased spatio-temporal distribution of SARS-CoV-2 Content: Effective systems for the analysis of molecular data are of fundamental importance for real-time monitoring of the spread of infectious diseases and the study of pathogen evolution. While the Nextstrain and GISAID portals offer widely used systems for the classification of SARS-CoV-2 genomes, both present relevant limitations. Here we propose a highly reproducible method for the systematic classification of SARS-CoV-2 viral types. To demonstrate the validity of our approach, we conduct an extensive comparative genomic analysis of more than 20,000 SARS-CoV-2 genomes. Our classification system delineates 12 clusters and 4 super-clusters in SARS-CoV-2, with a highly biased spatio-temporal distribution worldwide, and provides important observations concerning the evolutionary processes associated with the emergence of novel viral types. Based on the estimates of SARS-CoV-2 evolutionary rate and genetic distances of genomes of the early pandemic phase, we infer that SARS-CoV-2 could have been circulating in humans since August-November 2019. The observed pattern of genomic variability is remarkably similar between all clusters and super-clusters, being UTRs and the s2m element, a highly conserved secondary structure element, the most variable genomic regions. While several polymorphic sites that are specific to one or more clusters were predicted to be under positive or negative selection, overall, our analyses also suggest that the emergence of novel genome types is unlikely to be driven by widespread convergent evolution and independent fixation of advantageous substitutions. While, in the absence of rigorous experimental validation, several questions concerning the evolutionary processes and the phenotypic characteristics (increased/decreased virulence) remain open, we believe that the approach outlined in this study can be of relevance for the tracking and functional characterization of different types of SARS-CoV-2 genomes.

Title: A High-Coverage SARS-CoV-2 Genome Sequence Acquired by Target Capture Sequencing Content: This manuscript is based on the method we developed urgently to deal with the research requirement in the conflict between achieving a complete genome sequence for the evolutionary history of SARS-CoV-2 study and the low viral RNA concentration. Here, in this manuscript, we developed a set of SARS-CoV-2 enrichment probes to increase the sensitivity of sequence-based virus detection and characterization via obtaining the comprehensive genome sequence. Following the CDC health and safety guidelines, we test the concept using the culturing supernatant contain SARS-CoV-2 particles, and its full-length sequence was used for further analysis. The fraction of SARS-CoV-2 endogenous DNA was 93.47% with Cluster Factor about 1.1, which demonstrate that the numbers of mapped reads to SARS-CoV-2 reference sequence significantly increased, compared to metagenomic sequencing technology, following SARS-CoV-2 probe enrichment. Moreover, based on the high-quality sequence, we discussed the heterozygosity and viral expression during replication of coronavirus, and its phylogenetic relationship with other selected high-quality samples from The Genome Variation Map (GVM) (on 2020/03/22). We believe this manuscript is valuable for all the researchers who are interested in using clinical warp samples to obtain the high coverage of SARS-CoV-2 genome sequence with a relatively low concentration of viral particles. This would allow the clinician to correlate the diagnostic data with molecular monitoring in viral evolutional, the most importantly, to track the functional mutation of SARS-CoV-2.

Title: Genomic characterization of a novel SARS-CoV-2 Content: A new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) associated with human to human transmission and extreme human sickness has been as of late announced from the city of Wuhan in China. Our objectives were to mutation analysis between recently reported genomes at various times and locations and to characterize the genomic structure of SARS-CoV-2 using bioinformatics programs. Information on the variation of viruses is of considerable medical and biological impacts on the prevention, diagnosis, and therapy of infectious diseases. To understand the genomic structure and variations of the SARS-CoV-2. The study analyzed 95 SARS-CoV-2 complete genome sequences available in GenBank, National MicrobiologyData Center (NMDC) and NGDC Genome Warehouse from December-2019 until 05 of April-2020. The genomic signature analysis demonstrates that a strong association between the time of sample collection, location of sample and accumulation of genetic diversity. We found 116 mutations, the three most common mutations were 8782C>T in ORF1ab gene, 28144T>C in ORF8 gene and 29095C>T in the N gene. The mutations might affect the severity and spread of the SARS-CoV-2. The finding heavily supports an intense requirement for additional prompt, inclusive investigations that combine genomic detail, epidemiological information and graph records of the clinical features of patients with COVID-19.

Title: Stability of SARS-CoV-2 Phylogenies Content: The SARS-CoV-2 pandemic has led to unprecedented, nearly real-time genetic tracing due to the rapid community sequencing response. Researchers immediately leveraged these data to infer the evolutionary relationships among viral samples and to study key biological questions, including whether host viral genome editing and recombination are features of SARS-CoV-2 evolution. This global sequencing effort is inherently decentralized and must rely on data collected by many labs using a wide variety of molecular and bioinformatic techniques. There is thus a strong possibility that systematic errors associated with lab-specific practices affect some sequences in the repositories. We find that some recurrent mutations in reported SARS-CoV-2 genome sequences have been observed predominantly or exclusively by single labs, co-localize with commonly used primer binding sites and are more likely to affect the protein coding sequences than other similarly recurrent mutations. We show that their inclusion can affect phylogenetic inference on scales relevant to local lineage tracing, and make it appear as though there has been an excess of recurrent mutation and/or recombination among viral lineages. We suggest how samples can be screened and problematic mutations removed. We also develop tools for comparing and visualizing differences among phylogenies and we show that consistent clade- and tree-based comparisons can be made between phylogenies produced by different groups. These will facilitate evolutionary inferences and comparisons among phylogenies produced for a wide array of purposes. Building on the SARS-CoV-2 Genome Browser at UCSC, we present a toolkit to compare, analyze and combine SARS-CoV-2 phylogenies, find and remove potential sequencing errors and establish a widely shared, stable clade structure for a more accurate scientific inference and discourse. Foreword We wish to thank all groups that responded rapidly by producing these invaluable and essential sequence data. Their contributions have enabled an unprecedented, lightning-fast process of scientific discovery---truly an incredible benefit for humanity and for the scientific community. We emphasize that most lab groups with whom we associate specific suspicious alleles are also those who have produced the most sequence data at a time when it was urgently needed. We commend their efforts. We have already contacted each group and many have updated their sequences. Our goal with this work is not to highlight potential errors, but to understand the impacts of these and other kinds of highly recurrent mutations so as to identify commonalities among the suspicious examples that can improve sequence quality and analysis going forward.

Title: Comparative genomic signature representations of the emerging COVID-19 coronavirus and other coronaviruses: High identity and possible recombination between Bat and Pangolin coronaviruses Content: Coronaviruses are responsible on respiratory diseases in animal and human. The combination of numerical encoding techniques and digital signal processing methods are becoming increasingly important in handling large genomic data. In this paper, we propose to analyze the SARS-CoV-2 genomic signature using the combination of different nucleotide representations and signal processing tools in the aim to identify its genetic origin. The sequence of SARS-CoV-2 was compared with 21 relevant sequences including bat, yak and pangolin coronavirus sequences. In addition, we developed a new algorithm to locate the nucleotide modifications. The results show that the Bat and Pangolin coronaviruses were the most related to SARS-CoV-2 with 96% and 86% of identity all along the genome. Within the S gene sequence, the Pangolin sequence presents local highest nucleotide identity. Those findings suggest genesis of SARS-Cov-2 through evolution from bat and pangolin strains. This study offers new ways to automatically characterize viruses.

Title: Genomic characterization of a novel SARS-CoV-2 Content: Abstract A new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) associated with human to human transmission and extreme human sickness has been as of late announced from the city of Wuhan in China. Our objectives were to mutation analysis between recently reported genomes at various times and locations and to characterize the genomic structure of SARS-CoV-2 using bioinformatics programs. Information on the variation of viruses is of considerable medical and biological impacts on the prevention, diagnosis, and therapy of infectious diseases. To understand the genomic structure and variations of the SARS-CoV-2. The study analyzed 95 SARS-CoV-2 complete genome sequences available in GenBank, National MicrobiologyData Center (NMDC) and NGDC Genome Warehouse from December-2019 until 05 of April-2020. The genomic signature analysis demonstrates that a strong association between the time of sample collection, location of sample and accumulation of genetic diversity. We found 116 mutations, the three most common mutations were 8782C>T in ORF1ab gene, 28144T>C in ORF8 gene and 29095C>T in the N gene. The mutations might affect the severity and spread of the SARS-CoV-2. The finding heavily supports an intense requirement for additional prompt, inclusive investigations that combine genomic detail, epidemiological information and graph records of the clinical features of patients with COVID-19.

Title: SARS-CoV Infection Was from at Least Two Origins in the Taiwan Area Content: OBJECTIVE: Severe acute respiratory syndrome (SARS) is caused by a new coronavirus. Genomic sequence analysis will provide the molecular epidemiology and help to develop vaccines. METHODS: We developed a rapid method to amplify and sequence the whole SARS-CoV genome from clinical specimens. The technique employed one-step multiplex RT-PCR to amplify the whole SARS-CoV genome, and then nested PCR was performed to amplify a 2-kb region separately. The PCR products were sequenced. RESULTS: We sequenced the genomes of SARS-CoV from 3 clinical specimens obtained in Taiwan. The sequences were similar to those reported by other groups, except that 17 single nucleotide variations and two 2-nucleotide deletions, and a 1-nucleotide deletion were found. All the variations in the clinical specimens did not alter the amino acid sequence. Of these 17 sequenced variants, two loci (positions 26203 and 27812) were segregated together as a specific genotype-T:T or C:C. Phylogenetic analysis showed two major clusters of SARS patients in Taiwan. CONCLUSION: We developed a very economical and rapid method to sequence the whole genome of SARS-CoV, which can avoid cultural influence. From our results, SARS patients in Taiwan may be infected from two different origins.

Title: CoV-Seq: SARS-CoV-2 Genome Analysis and Visualization Content: Summary COVID-19 has become a global pandemic not long after its inception in late 2019. SARS-CoV-2 genomes are being sequenced and shared on public repositories at a fast pace. To keep up with these updates, scientists need to frequently refresh and reclean datasets, which is ad hoc and labor-intensive. Further, scientists with limited bioinformatics or programming knowledge may find it difficult to analyze SARS-CoV-2 genomes. In order to address these challenges, we developed CoV-Seq, a webserver to enable simple and rapid analysis of SARS-CoV-2 genomes. Given a new sequence, CoV-Seq automatically predicts gene boundaries and identifies genetic variants, which are presented in an interactive genome visualizer and are downloadable for further analysis. A command-line interface is also available for high-throughput processing. Availability and Implementation CoV-Seq is implemented in Python and Javascript. The webserver is available at http://covseq.baidu.com/ and the source code is available from https://github.com/boxiangliu/covseq. Contact jollier.liu@gmail.com Supplementary information Supplementary information are available at bioRxiv online.

Title: Genome analysis of SARS-CoV-2 isolates occurring in India: Present scenario. Content: Background The origin of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is still a debatable topic. The association of the virus spread from the market is supported by the close relation of genome sequences of environmental surface samples with virus samples from earliest patients by phylogenetic analysis. Objectives To have an insight into the SARS-CoV-2 genome sequences reported from India for better understanding on their epidemiology and virulence. Methods Genome sequences of Indian isolates of SARS-CoV-2 were analyzed to understand their phylogeny and divergence with respect to other isolates reported from other countries. Amino acid sequences of individual open reading frames (ORFs) from SARS-CoV-2 Indian isolates were aligned with sequences of isolates reported from other countries to identify the mutations occurred in Indian isolates. Results Our analysis suggests that Indian SARS-CoV-2 isolates are closely related to isolates reported from other parts of the world. Most ORFs are highly conserved; mutations were also detected in some ORFs. We found that most isolates reported from India have key mutations at 614th position of the S protein and 84th position of the ORF 8, which has been reported to be associated with high virulence and high transmission rate. Conclusion An attempt was made to understand the SARS-CoV-2 virus reported from India. SARS-CoV-2 reported from India was closely similar to other SARS-CoV-2 reported from other parts of the world, which suggests that vaccines and other therapeutic methods generated from other countries might work well in India. In addition, available sequence data suggest that majority of Indian isolates are capable of high transmission and virulence.

Title: An Analysis of SARS-CoV-2 Using ViReport Content: The ongoing outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has resulted in millions of cases and hundreds of thousands of deaths. Given the current lack of treatments or vaccines available, it may be useful to trace the evolu-tion and spread of the virus to better develop methods of preventative intervention. In this study, we analyzed over 4,000 full genome sequences of human SARS-CoV-2 using novel tool ViReport [13], an automated workflow for performing phylogenetic analyses on viral sequences and generating comprehensive molecular epidemiologi-cal reports. The complete ViReport output can be found at https://github.com/mirandajsong/ViReport-SARS-CoV-2.

Title: Comparative genomic signature representations of the emerging COVID-19 coronavirus and other coronaviruses: High identity and possible recombination between Bat and Pangolin coronaviruses Content: Abstract Coronaviruses are responsible on respiratory diseases in animal and human. The combination of numerical encoding techniques and digital signal processing methods are becoming increasingly important in handling large genomic data. In this paper, we propose to analyze the SARS-CoV-2 genomic signature using the combination of different nucleotide representations and signal processing tools in the aim to identify its genetic origin. The sequence of SARS-CoV-2 was compared with 21 relevant sequences including bat, yak and pangolin coronavirus sequences. In addition, we developed a new algorithm to locate the nucleotide modifications. The results show that the Bat and Pangolin coronaviruses were the most related to SARS-CoV-2 with 96% and 86% of identity all along the genome. Within the S gene sequence, the Pangolin sequence presents local highest nucleotide identity. Those findings suggest genesis of SARS-Cov-2 through evolution from bat and pangolin strains. This study offers new ways to automatically characterize viruses.

Title: SARS-CoV-2 sequence typing, evolution and signatures of selection using CoVa, a Python-based command-line utility Content: The current global pandemic COVID-19, caused by SARS-CoV-2, has resulted in millions of infections worldwide in a few months. Global efforts to tackle this situation have produced a tremendous body of genomic data, which can be used for tracing transmission routes, characterization of isolates, and monitoring variants with potential for unusual virulence. Several groups have analyzed these genomes using different approaches. However, as new data become available, the research community needs a pipeline to perform a set of routine analyses, that can quickly incorporate new genome sequences and update the analysis reports. We developed a programmatic tool, CoVa, with this objective. It is a fast, accurate and user-friendly utility to perform a variety of genome analyses on hundreds of SARS-CoV-2 sequences. Using CoVa, we define a modified sequence typing nomenclature and identify sites under positive selection. Further analysis identified some peptides and sites showing geographical patterns of selection. Specifically, we show differences in sequence type distribution between sequences from India and those from the rest of the world. We also show that several sites show signatures of positive selection uniquely in sequences from India. Preliminary evolutionary analysis, using features that will be incorporated into CoVa in the near future, show a mutation rate of 7.4 × 10−4 substitutions/site/year, confirm a temporal signal with a November 2019 origin of SARS-CoV-2, and a heterogeneity in the geographical distribution of Indian samples.

Title: Genome analysis of SARS-CoV-2 isolates occurring in India: Present scenario Content: Background: The origin of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is still a debatable topic. The association of the virus spread from the market is supported by the close relation of genome sequences of environmental surface samples with virus samples from earliest patients by phylogenetic analysis. Objectives: To have an insight into the SARS-CoV-2 genome sequences reported from India for better understanding on their epidemiology and virulence. Methods: Genome sequences of Indian isolates of SARS-CoV-2 were analyzed to understand their phylogeny and divergence with respect to other isolates reported from other countries. Amino acid sequences of individual open reading frames (ORFs) from SARS-CoV-2 Indian isolates were aligned with sequences of isolates reported from other countries to identify the mutations occurred in Indian isolates. Results: Our analysis suggests that Indian SARS-CoV-2 isolates are closely related to isolates reported from other parts of the world. Most ORFs are highly conserved; mutations were also detected in some ORFs. We found that most isolates reported from India have key mutations at 614th position of the S protein and 84th position of the ORF 8, which has been reported to be associated with high virulence and high transmission rate. Conclusion: An attempt was made to understand the SARS-CoV-2 virus reported from India. SARS-CoV-2 reported from India was closely similar to other SARS-CoV-2 reported from other parts of the world, which suggests that vaccines and other therapeutic methods generated from other countries might work well in India. In addition, available sequence data suggest that majority of Indian isolates are capable of high transmission and virulence.

Title: Molecular characterization of SARS-CoV-2 in the first COVID-19 cluster in France reveals an amino-acid deletion in nsp2 (Asp268Del) Content: We present the first genetic characterization of a COVID-19 cluster in Europe using metagenomic next-generation sequencing (mNGS). Despite low viral loads, the mNGS workflow used herein allowed to characterize the whole genome sequences of SARS-CoV2 isolated from an asymptomatic patient, in 2 clinical samples collected 1 day apart. Comparison of these sequences suggests viral evolution with development of quasispecies. In addition, the present workflow identified a new deletion in nsp2 (Asp268Del) which was found in all 3 samples originating from this cluster as well as in 37 other viruses collected in England and in Netherlands, suggesting the spread of this deletion in Europe. The impact of Asp268Del on SARS-CoV-2 transmission and pathogenicity, as well as on PCR performances and anti-viral strategy should be rapidly evaluated in further studies.

Title: Investigating the genomic landscape of novel coronavirus (2019-nCoV) to identify non-synonymous mutations for use in diagnosis and drug design Content: This study presents a comprehensive phylogenetic analysis of SARS-CoV2 isolates to understand discrete mutations that are occurring between patient samples. The analysis unravel various amino acid mutations in the viral proteins which may provide an explanation for varying treatment efficacies of different inhibitory drugs and a future direction towards a combinatorial treatment therapies based on the kind of mutation in the viral genome.

Title: Quantification of Intra-Host Genomic Diversity of SARS-CoV-2 Allows a High-Resolution Characterization of Viral Evolution and Reveals Functionally Convergent Variants Content: A global cross-discipline effort is ongoing to characterize the evolution of SARS-CoV-2 virus and generate reliable epidemiological models of its diffusion. To this end, phylogenomic approaches leverage accumulating genomic mutations as barcodes to track the evolutionary history of the virus and can benefit from the surge of sequences deposited in public databases. Yet, such methods typically rely on consensus sequences representing the dominant virus lineage, whereas a complex sublineage architecture is often observed within single hosts. Furthermore, most approaches do not account for variants accumulation processes and might produce inaccurate results in condition of limited sampling, as witnessed in most countries affected by the epidemics. We introduce VERSO (Viral Evolution ReconStructiOn), a new comprehensive framework for the characterization of viral evolution and transmission from sequencing data of viral genomes. our approach accounts for accumulation of clonal mutations and uncertainty in the data, by taking advantage of the achievements of research in cancer evolution, to deliver robust phylogenomic lineage models, and exploits intra-host variant frequency profiles to characterize the sublineage similarity among samples, which may derive from uncovered infection events. The application of our approach to RNA-sequencing data of 162 SARS-CoV-2 samples generates a high-resolution model of evolution and spread, which improves recent findings on viral types and highlights the existence of patterns of co-occurrence of minor variants, revealing likely infection paths among hosts harboring the same viral lineage. The in-depth analysis of the mutational landscape of SARS-CoV-2 confirms a statistically significant increase of genomic diversity in time and identifies a number of variants that are transiting from minor to clonal state in the population. We also show that standard phylogenetic methods can produce unreliable results when handling datasets with noise and sampling limitations, as proven by the further application of VERSO to 12419 consensus sequences included in GISAID database. Notably, VERSO allows to pinpoint minor variants that might be positively or negatively selected across distinct lineages, thus driving the design of treatments and vaccines. In particular, minor variant g.29039A>U, detected in multiple viral lineages and validated on independent samples, shows that SARS-CoV-2 can lose its main Nucleocapsid immunogenic epitopes, raising concerns about the effectiveness of vaccines targeting the C-terminus of this protein. Finally, we here release the likely SARS-CoV-2 ancestral genome, obtained by resolving ambiguous SNPs that distinguish two widely-used reference genomes from human samples, by employing the Pangolin-CoV and the Bat-CoV-RaTG13 genomes. Our results show that the joint application of our framework and data-driven epidemiological models might allow to deliver a high-resolution platform for pathogen surveillance and analysis.

Title: SARS-CoV-2 Whole Genome Amplification and Sequencing for Effective Population-Based Surveillance and Control of Viral Transmission Content: Background With the gradual reopening of economies and resumption of social life, robust surveillance mechanisms should be implemented to control the ongoing COVID-19 pandemic. Unlike RT-qPCR, SARS-CoV-2 Whole Genome Sequencing (cWGS) has the added advantage of identifying cryptic origins of the virus, and the extent of community-based transmissions versus new viral introductions, which can in turn influence public health policy decisions. However, practical and cost considerations of cWGS should be addressed before it can be widely implemented. Methods We performed shotgun transcriptome sequencing using RNA extracted from nasopharyngeal swabs of patients with COVID-19, and compared it to targeted SARS-CoV-2 full genome amplification and sequencing with respect to virus detection, scalability, and cost-effectiveness. To track virus origin, we used open-source multiple sequence alignment and phylogenetic tools to compare the assembled SARS-CoV-2 genomes to publicly available sequences. Results We show a significant improvement in whole genome sequencing data quality and viral detection using amplicon-based target enrichment of SARS-CoV-2. With enrichment, more than 99% of the sequencing reads mapped to the viral genome compared to an average of 0.63% without enrichment. Consequently, a dramatic increase in genome coverage was obtained using significantly less sequencing data, enabling higher scalability and significant cost reductions. We also demonstrate how SARS-CoV-2 genome sequences can be used to determine their possible origin through phylogenetic analysis including other viral strains. Conclusions SARS-CoV-2 whole genome sequencing is a practical, cost-effective, and powerful approach for population-based surveillance and control of viral transmission in the next phase of the COVID-19 pandemic.

Title: On spatial molecular arrangements of SARS-CoV2 genomes of Indian patients Content: A pandemic caused by the SARS-CoV2 is being experienced by the whole world since December, 2019. A thorough understanding beyond just sequential similarities among the protein coding genes of SARS-CoV2 is important in order to differentiate or relate to the other known CoVs of the same genus. In this study, we compare three genomes namely MT012098 (India-Kerala), MT050493 (India-Kerala), MT358637 (India-Gujrat) from India with NC_045512 (China-Wuhan) to view the spatial as well as molecular arrangements of nucleotide bases of all the genes embedded in these four genomes. Based on different features extracted for each gene embedded in these genomes, corresponding phylogenetic relationships have been built up. Differences in phylogenetic tree arrangement with individual gene suggest that three genomes of Indian origin have come from three different origins or the evolution of viral genome is very fast process. This study would also help to understand the virulence factors, disease pathogenicity, origin and transmission of the SARS-CoV2.

Title: Genomic epidemiology of a densely sampled COVID19 outbreak in China Content: Analysis of genetic sequence data from the pandemic SARS Coronavirus 2 can provide insights into epidemic origins, worldwide dispersal, and epidemiological history. With few exceptions, genomic epidemiological analysis has focused on geographically distributed data sets with few isolates in any given location. Here we report an analysis of 20 whole SARS-CoV 2 genomes from a single relatively small and geographically constrained outbreak in Weifang, People's Republic of China. Using Bayesian model-based phylodynamic methods, we estimate the reproduction number for the outbreak to be 1.99(95% CI:1.48-3.14). We further estimate the number of infections through time and compare these estimates to confirmed diagnoses by the Weifang Centers for Disease Control. We find that these estimates are consistent with reported cases and there is unlikely to be a large undiagnosed burden of infection over the period we studied.

Title: Polymorphism of SARS-CoV Genomes Content: Abstract In this work, severe acute respiratory syndrome associated coronavirus (SARS-CoV) genome BJ202 (AY864806) was completely sequenced. The genome was directly accessed from the stool sample of a patient in Beijing. Comparative genomics methods were used to analyze the sequence variations of 116 SARS-CoV genomes (including BJ202) available in the NCBI Gen-Bank. With the genome sequence of GZ02 as the reference, there were 41 polymorphic sites identified in BJ202 and a total of 278 polymorphic sites present in at least two of the 116 genomes. The distribution of the polymorphic sites was biased over the whole genome. Nearly half of the variations (50.4%, 140/278) clustered in the one third of the whole genome at the 3′ end (19.0 kb-29.7 kb). Regions encoding Orf10–11, Orf3/4, E, M and S protein had the highest mutation rates. A total of 15 PCR products (about 6.0 kb of the genome) including 11 fragments containing 12 known polymorphic sites and 4 fragments without identified polymorphic sites were cloned and sequenced. Results showed that 3 unique polymorphic sites of BJ202 (positions 13 804, 15 031 and 20 792) along with 3 other polymorphic sites (26 428, 26 477 and 27 243) all contained 2 kinds of nucleotides. It is interesting to find that position 18379 which has not been identified to be polymorphic in any of the other 115 published SARS-CoV genomes is actually a polymorphic site. The nucleotide composition of this site is A (8) to G (6). Among 116 SARS-CoV genomes, 18 types of deletions and 2 insertions were identified. Most of them were related to a 300 bp region (27 700–28 000) which encodes parts of the putative ORF9 and ORF10–11. A phylogenetic tree illustrating the divergence of whole BJ202 genome from 115 other completely sequenced SARS-CoVs was also constructed. BJ202 was phylogeneticly closer to BJ01 and LLJ-2004.

Title: Insights into SARS-CoV-2 genome, structure, evolution, pathogenesis and therapies: Structural genomics approach Content: The sudden emergence of severe respiratory disease, caused by a novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has recently become a public health emergency. Genome sequence analysis of SARS-CoV-2 revealed its close resemblance to the earlier reported SARS-CoV and Middle East respiratory syndrome coronavirus (MERS-CoV). However, initial testing of the drugs used against SARS-CoV and MERS-CoV has been ineffective in controlling SARS-CoV-2. The present study highlights the genomic, proteomic, pathogenesis, and therapeutic strategies in SARS-CoV-2 infection. We have carried out sequence analysis of potential drug target proteins in SARS-CoV-2 and, compared them with SARS-CoV and MERS viruses. Analysis of mutations in the coding and non-coding regions, genetic diversity, and pathogenicity of SARS-CoV-2 has also been done. A detailed structural analysis of drug target proteins has been performed to gain insights into the mechanism of pathogenesis, structure-function relationships, and the development of structure-guided therapeutic approaches. The cytokine profiling and inflammatory signalling are different in the case of SARS-CoV-2 infection. We also highlighted possible therapies and their mechanism of action followed by clinical manifestation. Our analysis suggests a minimal variation in the genome sequence of SARS-CoV-2, may be responsible for a drastic change in the structures of target proteins, which makes available drugs ineffective.

Title: Chasing the origin of SARS-CoV-2 in Canada’s COVID-19 cases: A genomics study Content: The emergence and global spread of SARS-CoV-2 has had profound social and economic consequences and has shed light on the importance of continued and additional investment in global health and infectious disease surveillance. Identifying changes in viral genomes provides key insights into viral diversity, how viruses spread within populations, and viral strategies for evasion of host immune systems. Here we report twenty-five SARS-CoV-2 genome sequences collected from some of the first COVID-19 cases in eastern Ontario, Canada (March 18-30, 2020). The reported genomes belong to the S-clade (n=2) and G-clade (n=23) of SARS-CoV-2 and contain 45 polymorphic sites including one shared missense and three unique synonymous variants in the gene encoding the spike protein. A phylogenetic analysis enabled the tracing of viral origin and potential transmission into and within Canada. There may be as many as sixteen unique infection events represented in these samples, including at least three that were likely introduced from Europe and seven from the USA. In addition, four separate genomes are each shared by multiple patients, suggesting a common origin or community spread even during this early stage of infection. These results demonstrate how molecular epidemiology and evolutionary phylogenetics can help local health units track origins and vectors of spread for emerging diseases like SARS-CoV-2. Earlier detection and screening in this way could improve the effectiveness of regional public health interventions to prevent future pandemics.

Title: Insights into SARS-CoV-2 genome, structure, evolution, pathogenesis and therapies: Structural genomics approach Content: The sudden emergence of severe respiratory disease, caused by a novel severe acute respiratory syndrome coronavirus (SARS-CoV-2), has recently become a public health emergency. Genome sequence analysis of SARS-CoV-2 revealed its close resemblance to the earlier reported SARS-CoV and Middle East respiratory syndrome coronavirus (MERS-CoV). However, initial testing of the drugs used against SARS-CoV and MERS-CoV has been ineffective in controlling SARS-CoV-2. The present review looks to highlight the differences in genomic, proteomic, pathogenesis, and therapeutic strategies of SARS-CoV-2. We have carried out sequence analysis of potential drug target proteins in SARS-CoV-2 and, compared them with SARS-CoV-1 and MERS viruses. Analysis of mutations in the coding and non-coding regions, genetic diversity, and pathogenicity of SARS-CoV-2 has also been done. A detailed structural analysis of drug target proteins was performed to gain insights into the mechanism of pathogenesis, structure-function relationships, and the development of structure-guided therapeutic approaches. The cytokine profiling and inflammatory signalling are different in the case of SARS-CoV-2 infection. We also highlighted possible therapies and their mechanism of action followed by clinical manifestation. Our analysis suggests a minimal variation in the genome sequence of SARS-CoV-2, may be responsible for a drastic change in the structures of target proteins, makes available drugs ineffective.

Title: Sequence analysis of travel-related SARS-CoV-2 cases in the Greater Geelong region, Australia Content: This study reports the sequence analysis of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) from infected individuals within the Greater Geelong region, Victoria, Australia. All but one individual had recently returned from travelling abroad, and all had clinical signs consistent with SARS-CoV-2 infection. SARS-CoV-2 belonging to three lineages were detected and represent separate introductions of the virus into the region. Sequence data were consistent with the recent travel history of each case. Full virus genome sequencing can play an important role in supporting local epidemiological tracing and monitoring for community transmission. Quality of the SARS-CoV-2 sequences obtained was highly dependent on appropriate sample collection and handling.

Title: MicroGMT: A Mutation Tracker for SARS-CoV-2 and Other Microbial Genome Sequences Content: With the continued spread of SARS-CoV-2 virus around the world, researchers often need to quickly identify novel mutations in newly sequenced SARS-CoV-2 genomes for studying the molecular evolution and epidemiology of the virus. We have developed a Python package, MicroGMT, which takes either raw sequence reads or assembled genome sequences as input and compares against database sequences to identify and characterize indels and point mutations. Although our default setting is optimized for SARS-CoV-2 virus, the package can be also applied to any other microbial genomes. The software is freely available at Github URL https://github.com/qunfengdong/MicroGMT.

Title: Genome-Wide Identification and Characterization of Point Mutations in the SARS-CoV-2 Genome Content: OBJECTIVES: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged in Wuhan, China, in December 2019 and has been rapidly spreading worldwide. Although the causal relationship among mutations and the features of SARS-CoV-2 such as rapid transmission, pathogenicity, and tropism, remains unclear, our results of genomic mutations in SARS-CoV-2 may help to interpret the interaction between genomic characterization in SARS-CoV-2 and infectivity with the host. METHODS: A total of 4,254 genomic sequences of SARS-CoV-2 were collected from the Global Initiative on Sharing all Influenza Data (GISAID). Multiple sequence alignment for phylogenetic analysis and comparative genomic approach for mutation analysis were conducted using Molecular Evolutionary Genetics Analysis (MEGA), and an in-house program based on Perl language, respectively. RESULTS: Phylogenetic analysis of SARS-CoV-2 strains indicated that there were 3 major clades including S, V, and G, and 2 subclades (G.1 and G.2). There were 767 types of synonymous and 1,352 types of non-synonymous mutation. ORF1a, ORF1b, S, and N genes were detected at high frequency, whereas ORF7b and E genes exhibited low frequency. In the receptor-binding domain (RBD) of the S gene, 11 non-synonymous mutations were observed in the region adjacent to the angiotensin converting enzyme 2 (ACE2) binding site. CONCLUSION: It has been reported that the rapid infectivity and transmission of SARS-CoV-2 associated with host receptor affinity are derived from several mutations in its genes. Without these genetic mutations to enhance evolutionary adaptation, species recognition, host receptor affinity, and pathogenicity, it would not survive. It is expected that our results could provide an important clue in understanding the genomic characteristics of SARS-CoV-2.

Title: Companion vaccine Bioinformatic design tool reveals limited functional genomic variability of SARS-Cov-2 Spike Receptor Binding Domain Content: Tracking the genetic variability of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is crucial for several reasons, such as to identify target sequences to generate robust vaccines and neutralizing monoclonal antibodies; to track viral genetic temporal and geographic evolution; and to mine for viral variants associated with reduced or increased severity. Several online tools and bioinformatic analyses have been released for this purpose; four main viral clades have been described via phylogenetic analyses. Here, we present an open-source bioinformatic protocol focused on SARS-CoV-2 single mutations and minimal consensus sequence building as a companion vaccine design tool. Results on the whole GISAID sequence dataset at the time of the writing (April–June 2020) confirms no high variability pressure on the viral genome in terms of quality and quantity of mutations. We then focused our analysis on the receptor-binding domain region of the spike protein and only found a few variants associated with particular geographic locations that should be tracked over time. Finally, additional immunogenomic analyses revealed some variation in mutated epitope MHC compatibility and T-cell recognition for most frequent human HLAs.

Title: Genomic analysis of early SARS-CoV-2 strains introduced in Mexico Content: The COVID-19 pandemic has affected most countries in the world. Studying the evolution and transmission patterns in different countries is crucial to implement effective strategies for disease control and prevention. In this work, we present the full genome sequence for 17 SARS-CoV-2 isolates corresponding to the earliest sampled cases in Mexico. Global and local phylogenomics, coupled with mutational analysis, consistently revealed that these viral sequences are distributed within 2 known lineages, the SARS-CoV-2 lineage A/G, containing mostly sequences from North America, and the lineage B/S containing mainly sequences from Europe. Based on the exposure history of the cases and on the phylogenomic analysis, we characterized fourteen independent introduction events. Additionally, three cases with no travel history were identified. We found evidence that two of these cases represent local transmission cases occurring in Mexico during mid-March 2020, denoting the earliest events described in the country. Within this Mexican cluster, we also identified an H49Y amino acid change in the spike protein. This mutation is a homoplasy occurring independently through time and space, and may function as a molecular marker to follow on any further spread of these viral variants throughout the country. Our results depict the general picture of the SARS-CoV-2 variants introduced at the beginning of the outbreak in Mexico, setting the foundation for future surveillance efforts. This work is the result of the collaboration of five institutions into one research consortium: three public health institutes and two universities. From the beginning of this work, it was agreed that the experimental leader of each institution would share the first authorship. Those were the criteria followed to assign first co-first authorship in this manuscript. The order of the other authors was randomly assigned. IMPORTANCE Understanding the introduction, spread and establishment of SARS-CoV-2 within distinct human populations is crucial to implement effective control strategies as well as the evolution of the pandemics. In this work, we describe that the initial virus strains introduced in Mexico came from Europe and the United States and the virus was circulating locally in the country as early as mid-March. We also found evidence for early local transmission of strains having the mutation H49Y in the Spike protein, that could be further used as a molecular marker to follow viral spread within the country and the region.

Title: Genomic analysis and comparative multiple sequences of SARS-CoV2 Content: BACKGROUND: China announced an outbreak of new coronavirus in the city of Wuhan on December 31, 2019; lash to now, the virus transmission has become pandemic worldwide. Severe cases from the Huanan Seafood Wholesale market in Wuhan were confirmed pneumonia with a novel coronavirus (2019-nCoV). Understanding the molecular mechanisms of genome selection and packaging is critical for developing antiviral strategies. Thus, we defined the correlation in 10 severe acute respiratory syndrome coronavirus (SARS-CoV2) sequences from different countries to analyze the genomic patterns of disease origin and evolution aiming for developing new control pandemic processes. METHODS: We apply genomic analysis to observe SARS-CoV2 sequences from GenBank (http://www.ncbi.nim.nih.gov/genebank/): MN 908947 (China, C1), MN985325 (USA: WA, UW), MN996527 (China, C2), MT007544 (Australia: Victoria, A1), MT027064 (USA: CA, UC), MT039890 (South Korea, K1), MT066175 (Taiwan, T1), MT066176 (Taiwan, T2), LC528232 (Japan, J1), and LC528233 (Japan, J2) for genomic sequence alignment analysis. Multiple Sequence Alignment by Clustalw (https://www.genome.jp/tools-bin/clustalw) web service is applied as our alignment tool. RESULTS: We analyzed 10 sequences from the National Center for Biotechnology Information (NCBI) database by genome alignment and found no difference in amino acid sequences within M and N proteins. There are two amino acid variances in the spike (S) protein region. One mutation found from the South Korea sequence is verified. Two possible "L" and "S" SNPs found in ORF1ab and ORF8 regions are detected. CONCLUSION: We performed genomic analysis and comparative multiple sequences of SARS-CoV2. Studies about the biological symptoms of SARS-CoV2 in clinic animals and humans will manipulate an understanding on the origin of pandemic crisis.

Title: Identification of unique mutations in SARS-CoV-2 strains isolated from India suggests its attenuated pathotype Content: Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), which was first reported in Wuhan, China in November 2019 has developed into a pandemic since March 2020, causing substantial human casualties and economic losses. Studies on SARS-CoV-2 are being carried out at an unprecedented rate to tackle this threat. Genomics studies, in particular, are indispensable to elucidate the dynamic nature of the RNA genome of SARS-CoV-2. RNA viruses are marked by their unique ability to undergo high rates of mutation in their genome, much more frequently than their hosts, which diversifies their strengths qualifying them to elude host immune response and amplify drug resistance. In this study, we sequenced and analyzed the genomic information of the SARS-CoV-2 isolates from two infected Indian patients and explored the possible implications of point mutations in its biology. In addition to multiple point mutations, we found a remarkable similarity between relatively common mutations of 36-nucleotide deletion in ORF8 of SARS-CoV-2. Our results corroborate with the earlier reported 29-nucleotide deletion in SARS, which was frequent during the early stage of human-to-human transmission. The results will be useful to understand the biology of SARS-CoV-2 and itsattenuation for vaccine development.

Title: Molecular phylogeny of coronaviruses including human SARS-CoV Content: Phylogenetic tree of coronaviruses (CoVs) including the human SARS-associated virus is reconstructed from complete genomes by using our newly developed K-string composition approach. The relation of the human SARS-CoV to other coronaviruses, i.e. the rooting of the tree is suggested by choosing an appropriate outgroup. SARS-CoV makes a separate group closer but still distant from G2 (CoVs in mammalian host). The relation between different isolates of the human SARS virus is inferred by first constructing an ultrametric distance matrix from counting sequence variations in the genomes. The resulting tree is consistent with clinic relations between the SARS-CoV isolates. In addition to a larger variety of coronavirus genomes these results provide phylogenetic knowledge based on independent novel methodology as compared to recent phylogenetic studies on SARS-CoV.

Title: Unsupervised cluster analysis of SARS-CoV-2 genomes indicates that recent (June 2020) cases in Beijing are from a genetic subgroup that consists of mostly European and South(east) Asian samples, of which the latter are the most recent Content: Research efforts of the ongoing SARS-CoV-2 pandemic have focused on viral genome sequence analysis to understand how the virus spread across the globe. Here, we assess three recently identified SARS-CoV-2 genomes in Beijing from June 2020 and attempt to determine the origin of these genomes, made available in the GISAID database. The database contains fully or partially sequenced SARS-CoV-2 samples from laboratories around the world. Including the three new samples and excluding samples with missing annotations, we analyzed 7, 643 SARS-CoV-2 genomes. Using principal component analysis computed on a similarity matrix that compares all pairs of the SARS-CoV-2 nucleotide sequences at all loci simultaneously, using the Jaccard index, we find that the newly discovered virus genomes from Beijing are in a genetic cluster that consists mostly of cases from Europe and South(east) Asia. The sequences of the new cases are most related to virus genomes from a small number of cases from China (March 2020), cases from Europe (February to early May 2020), and cases from South(east) Asia (May to June 2020). These findings could suggest that the original cases of this genetic cluster originated from China in March 2020 and were re-introduced to China by transmissions from samples from South(east) Asia between April and June 2020.

Title: Monophyletic Relationship between Severe Acute Respiratory Syndrome Coronavirus and Group 2 Coronaviruses Content: Although primary genomic analysis has revealed that severe acute respiratory syndrome coronavirus (SARS CoV) is a new type of coronavirus, the different protein trees published in previous reports have provided no conclusive evidence indicating the phylogenetic position of SARS CoV. To clarify the phylogenetic relationship between SARS CoV and other coronaviruses, we compiled a large data set composed of 7 concatenated protein sequences and performed comprehensive analyses, using the maximum-likelihood, Bayesian-inference, and maximum-parsimony methods. All resulting phylogenetic trees displayed an identical topology and supported the hypothesis that the relationship between SARS CoV and group 2 CoVs is monophyletic. Relationships among all major groups were well resolved and were supported by all statistical analyses.

Title: WHOLE-GENOME SEQUENCING AND DE NOVO ASSEMBLY OF A 2019 NOVEL CORONAVIRUS (SARS-COV-2) STRAIN ISOLATED IN VIETNAM Content: The pandemic COVID-19 caused by the zoonotic virus SARS-CoV-2 has devastated countries worldwide, infecting more than 4.5 million people and leading to more than 300,000 deaths. Whole genome sequencing (WGS) is an effective tool to monitor emerging strains and provide information for intervention, thus help to inform outbreak control decisions. Here, we reported the first effort to sequence and de novo assemble the whole genome of SARS-CoV-2 using PacBio’s SMRT sequencing technology in Vietnam. We also presented the annotation results and a brief analysis of the variants found in our SARS-CoV-2 strain, which was isolated from a Vietnamese patient. The sequencing was successfully completed and de novo assembled in less than 30 hours, resulting in one contig with no gap and a length of 29,766 bp. All detected variants as compared to the NCBI reference were highly accurate as confirmed by Sanger sequencing. The results have shown the potential of long read sequencing to provide high quality WGS data to support public health responses, and advance understanding of this and future pandemics.

Title: Metagenomic Analysis Reveals Clinical SARS-CoV-2 Infection and Bacterial or Viral Superinfection and Colonization Content: BACKGROUND: More than 2 months separated the initial description of SARS-CoV-2 and discovery of its widespread dissemination in the United States. Despite this lengthy interval, implementation of specific quantitative reverse transcription (qRT)-PCR-based SARS-CoV-2 tests in the US has been slow, and testing is still not widely available. Metagenomic sequencing offers the promise of unbiased detection of emerging pathogens, without requiring prior knowledge of the identity of the responsible agent or its genomic sequence. METHODS: To evaluate metagenomic approaches in the context of the current SARS-CoV-2 epidemic, laboratory-confirmed positive and negative samples from Seattle, WA were evaluated by metagenomic sequencing, with comparison to a 2019 reference genomic database created before the emergence of SARS-CoV-2. RESULTS: Within 36 h our results showed clear identification of a novel human Betacoronavirus, closely related to known Betacoronaviruses of bats, in laboratory-proven cases of SARS-CoV-2. A subset of samples also showed superinfection or colonization with human parainfluenza virus 3 or Moraxella species, highlighting the need to test directly for SARS-CoV-2 as opposed to ruling out an infection using a viral respiratory panel. Samples negative for SARS-CoV-2 by RT-PCR were also negative by metagenomic analysis, and positive for Rhinovirus A and C. Unlike targeted SARS-CoV-2 qRT-PCR testing, metagenomic analysis of these SARS-CoV-2 negative samples identified candidate etiological agents for the patients' respiratory symptoms. CONCLUSION: Taken together, these results demonstrate the value of metagenomic analysis in the monitoring and response to this and future viral pandemics.

Title: Children’s Hospital Los Angeles COVID-19 Analysis Research Database (CARD) - A Resource for Rapid SARS-CoV-2 Genome Identification Using Interactive Online Phylogenetic Tools Content: Effective response to the Coronavirus Disease 2019 (COVID-19) pandemic requires genomic resources and bioinformatics tools for genomic epidemiology and surveillance studies that involve characterizing full-length viral genomes, identifying origins of infections, determining the relatedness of viral infections, performing phylogenetic analyses, and monitoring the continuous evolution of the SARS-CoV-2 viral genomes. The Children’s Hospital, Los Angeles (CHLA) COVID-19 Analysis Research Database (CARD) (https://covid19.cpmbiodev.net/) is a comprehensive genomic resource that provides access to full-length SARS-CoV-2 viral genomes and associated meta-data for over 30,000 (as of May 20, 2020) isolates collected from global sequencing repositories and the sequencing performed at the Center for Personalized Medicine (CPM) at CHLA. Reference phylogenetic trees of global and USA viral isolates were constructed and are periodically updated using selected high quality SARS-CoV-2 genome sequences. These provide the baseline and analytical context for identifying the origin of a viral infection, as well as the relatedness of SARS-CoV-2 genomes of interest. A web-based and interactive Phylogenetic Tree Browser supports flexible tree manipulation and advanced analysis based on keyword search while highlighting time series animation, as well as subtree export for graphical representation or offline exploration. A Virus Genome Tracker accepts complete or partial SARS-CoV-2 genome sequence, compares it against all available sequences in the database (>30,000 at time of writing), detects and annotates the variants, and places the new viral isolate within the global or USA phylogenetic contexts based upon variant profiles and haplotype comparisons, in a few seconds. The generated analysis can potentially aid in genomic surveillance to trace the transmission of any new infection. Using CHLA CARD, we demonstrate the identification of a candidate outbreak point where 13 of 31 CHLA internal isolates may have originated. We also discovered multiple indels of unknown clinical significance in the orf3a gene, and revealed a number of USA-specific variants and haplotypes.

Title: Genetic Diversity and Genomic Epidemiology of SARS-COV-2 in Morocco Content: COVID-A9 is an infection disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), declared as a pandemic due to its rapid expansion worldwide. In this study we investigate the genetic diversity and genomic epidemiology of SARS-CoV-2 using 22 virus genome sequences reported by three different laboratories in Morocco till the date 07/06/2020 as well as (40366) virus genomes from all around the world. The SARS-CoV-2 genomes from Moroccan patients revealed 62 mutations of which 30 were missense mutations. The mutations Spike_D614G and NSP12_P323L were present in all the 22 analyzed sequences, followed by N_G204R and N_R203K which occurred in 9 among the 22 sequences. The mutations NSP10_R134S, NSP15_D335N, NSP16_I169L, NSP3_L431H, NSP3_P1292L and Spike_V6F occurred one time in our sequences with no record in other sequence worldwide. These mutations should be investigated to figure out their potential effects on all around the world virulence. Phylogenetic analyses revealed that Moroccan SARS-CoV-2 genomes included 9 viruses pertaining to clade 20A, 9 to clade 20B and 2 to clade 20C. This finding suggest that the epidemic spread in Morocco did not show a predominant SARS-CoV-2 route. For multiple and unrelated introductions of SARS-CoV-2 into Morocco via different routes have occurred, giving rise to the diversity of virus genomes in the country. Furthermore, very likely, the SARS-CoV-2 virus circulated in cryptic way in Morocco starting from the fifteen January before the discovering of the first case the second of March.

Title: Controlling the SARS-CoV-2 outbreak, insights from large scale whole genome sequences generated across the world Content: Background SARS-CoV-2 most likely evolved from a bat beta-coronavirus and started infecting humans in December 2019. Since then it has rapidly infected people around the world, with more than 4.5 million confirmed cases by the middle of May 2020. Early genome sequencing of the virus has enabled the development of molecular diagnostics and the commencement of therapy and vaccine development. The analysis of the early sequences showed relatively few evolutionary selection pressures. However, with the rapid worldwide expansion into diverse human populations, significant genetic variations are becoming increasingly likely. The current limitations on social movement between countries also offers the opportunity for these viral variants to become distinct strains with potential implications for diagnostics, therapies and vaccines. Methods We used the current sequencing archives (NCBI and GISAID) to investigate 15,487 whole genomes, looking for evidence of strain diversification and selective pressure. Results We used 6,294 SNPs to build a phylogenetic tree of SARS-CoV-2 diversity and noted strong evidence for the existence of two major clades and six sub-clades, unevenly distributed across the world. We also noted that convergent evolution has potentially occurred across several locations in the genome, showing selection pressures, including on the spike glycoprotein where we noted a potentially critical mutation that could affect its binding to the ACE2 receptor. We also report on mutations that could prevent current molecular diagnostics from detecting some of the sub-clades. Conclusion The worldwide whole genome sequencing effort is revealing the challenge of developing SARS-CoV-2 containment tools suitable for everyone and the need for data to be continually evaluated to ensure accuracy in outbreak estimations.

Title: Metagenomic analysis reveals clinical SARS-CoV-2 infection and bacterial or viral superinfection and colonization Content: BACKGROUND: More than two months separated the initial description of SARS-CoV-2 and discovery of its widespread dissemination in the United States. Despite this lengthy interval, implementation of specific quantitative reverse transcription (qRT)-PCR-based SARS-CoV-2 tests in the US has been slow, and testing is still not widely available. Metagenomic sequencing offers the promise of unbiased detection of emerging pathogens, without requiring prior knowledge of the identity of the responsible agent or its genomic sequence. METHODS: To evaluate metagenomic approaches in the context of the current SARS-CoV-2 epidemic, laboratory-confirmed positive and negative samples from Seattle, Washington were evaluated by metagenomic sequencing, with comparison to a 2019 reference genomic database created before the emergence of SARS-CoV-2. RESULTS: Within 36 hours our results showed clear identification of a novel human Betacoronavirus, closely related to known Betacoronaviruses of bats, in laboratory-proven cases of SARS-CoV-2. A subset of samples also showed superinfection or colonization with human parainfluenza virus 3 or Moraxella species, highlighting the need to test directly for SARS-CoV-2 as opposed to ruling out an infection using a viral respiratory panel. Samples negative for SARS-CoV-2 by RT-PCR were also negative by metagenomic analysis, and positive for Rhinovirus A and C. Unlike targeted SARS-CoV-2 qRT-PCR testing, metagenomic analysis of these SARS-CoV-2 negative samples identified candidate etiological agents for the patients’ respiratory symptoms. CONCLUSION: Taken together, these results demonstrate the value of metagenomic analysis in the monitoring and response to this and future viral pandemics.

Title: Emergence of multiple variants of SARS-CoV-2 with signature structural changes Content: This study explores the divergence pattern of SARS-CoV-2 using whole genome sequences of the isolates from various COVID-19 affected countries. The phylogenomic analysis indicates the presence of at least four distinct groups of the SARS-CoV-2 genomes. The emergent groups have been found to be associated with signature structural changes in specific proteins. Also, this study reveals the differential levels of divergence patterns for the protein coding regions. Moreover, we have predicted the impact of structural changes on a couple of important viral proteins via structural modelling techniques. This study further advocates for more viral genetic studies with associated clinical outcomes and hosts’ response for better understanding of SARS-CoV-2 pathogenesis enabling better mitigation of this pandemic situation.

Title: Phylogenomic proximity and comparative proteomic analysis of SARS-CoV-2 Content: The coronavirus disease (COVID-19) belongs to the family Severe Acute Respiratory Syndrome (SARS-CoV). It can be more severe for some persons and can lead to pneumonia or breathing difficulties resulting in the death of immune-compromised patients. We performed a phylogenomic and phylogeographic tree from the collected datasets. Phylogenomic analysis or sequence-based phylogeny showed an evolutionary relationship between the geographical strains. The phylogenomic tree grouped into two major clades consists of various isolates of SARS-CoV-2 and Bat SARS-like coronavirus, Bat coronavirus, and Pangolin coronavirus. The phylogenetic neighbor of newly sequenced Indian strains (Accession: MT012098.1, MT050493.1) was revealed to identify the variations between the nCoV-19 strains. The results showed keen evidence that SARS-CoV-2 has evolved from Bat SARS-like coronavirus. The evolutionary history and comparative proteomic analysis provide a new avenue for the current scientific research related to the coronavirus.

Title: Genomic Analysis of Early SARS-CoV-2 Variants Introduced in Mexico Content: The COVID-19 pandemic has affected most countries in the world. Studying the evolution and transmission patterns in different countries is crucial to implement effective strategies for disease control and prevention. In this work, we present the full genome sequence for 17 SARS-CoV-2 isolates corresponding to the earliest sampled cases in Mexico. Global and local phylogenomics, coupled with mutational analysis, consistently revealed that these viral sequences are distributed within 2 known lineages, the SARS-CoV-2 lineage A/G, containing mostly sequences from North America, and the lineage B/S containing mainly sequences from Europe. Based on the exposure history of the cases and on the phylogenomic analysis, we characterized fourteen independent introduction events. Additionally, three cases with no travel history were identified. We found evidence that two of these cases represent local transmission cases occurring in Mexico during mid-March 2020, denoting the earliest events described for the country. Within this local transmission cluster, we also identified the H49Y amino acid change in the Spike protein. This mutation is a homoplasy occurring independently through time and space, and may function as a molecular marker to follow on any further spread of these viral variants throughout the country. Our results depict the general picture of the SARS-CoV-2 variants introduced at the beginning of the outbreak in Mexico, setting the foundation for future surveillance efforts.IMPORTANCE Understanding the introduction, spread and establishment of SARS-CoV-2 within distinct human populations is crucial to implement effective control strategies as well as the evolution of the pandemics. In this work, we describe that the initial virus strains introduced in Mexico came from Europe and the United States and the virus was circulating locally in the country as early as mid-March. We also found evidence for early local transmission of strains having the mutation H49Y in the Spike protein, that could be further used as a molecular marker to follow viral spread within the country and the region.

Title: Genomic Analysis of Early SARS-CoV-2 Variants Introduced in Mexico. Content: The COVID-19 pandemic has affected most countries in the world. Studying the evolution and transmission patterns in different countries is crucial to implement effective strategies for disease control and prevention. In this work, we present the full genome sequence for 17 SARS-CoV-2 isolates corresponding to the earliest sampled cases in Mexico. Global and local phylogenomics, coupled with mutational analysis, consistently revealed that these viral sequences are distributed within 2 known lineages, the SARS-CoV-2 lineage A/G, containing mostly sequences from North America, and the lineage B/S containing mainly sequences from Europe. Based on the exposure history of the cases and on the phylogenomic analysis, we characterized fourteen independent introduction events. Additionally, three cases with no travel history were identified. We found evidence that two of these cases represent local transmission cases occurring in Mexico during mid-March 2020, denoting the earliest events described for the country. Within this local transmission cluster, we also identified the H49Y amino acid change in the Spike protein. This mutation is a homoplasy occurring independently through time and space, and may function as a molecular marker to follow on any further spread of these viral variants throughout the country. Our results depict the general picture of the SARS-CoV-2 variants introduced at the beginning of the outbreak in Mexico, setting the foundation for future surveillance efforts.IMPORTANCE Understanding the introduction, spread and establishment of SARS-CoV-2 within distinct human populations is crucial to implement effective control strategies as well as the evolution of the pandemics. In this work, we describe that the initial virus strains introduced in Mexico came from Europe and the United States and the virus was circulating locally in the country as early as mid-March. We also found evidence for early local transmission of strains having the mutation H49Y in the Spike protein, that could be further used as a molecular marker to follow viral spread within the country and the region.

Title: CoV Genome Tracker: tracing genomic footprints of Covid-19 pandemic Content: Summary Genome sequences constitute the primary evidence on the origin and spread of the 2019-2020 Covid-19 pandemic. Rapid comparative analysis of coronavirus SARS-CoV-2 genomes is critical for disease control, outbreak forecasting, and developing clinical interventions. CoV Genome Tracker is a web portal dedicated to trace Covid-19 outbreaks in real time using a haplotype network, an accurate and scalable representation of genomic changes in a rapidly evolving population. We resolve the direction of mutations by using a bat-associated genome as outgroup. At a broader evolutionary time scale, a companion browser provides gene-by-gene and codon-by-codon evolutionary rates to facilitate the search for molecular targets of clinical interventions. Availability and Implementation CoV Genome Tracker is publicly available at http://cov.genometracker.org and updated weekly with the data downloaded from GISAID (http://gisaid.org). The website is implemented with a custom JavaScript script based on jQuery (https://jquery.com) and D3-force (https://github.com/d3/d3-force). Contact weigang@genectr.hunter.cuny.edu, City University of New York, Hunter College Supplementary Information All supporting scripts developed in JavaScript, Python, BASH, and PERL programming languages are available as Open Source at the GitHub repository https://github.com/weigangq/cov-browser.

Title: Insights into SARS-CoV-2, the Coronavirus Underlying COVID-19: Recent Genomic Data and the Development of Reverse Genetics Systems Content: The emergence and rapid worldwide spread of a novel pandemic of acute respiratory disease - eventually named coronavirus disease 2019 (COVID-19) by the World Health Organization (WHO) - across the human population has raised great concerns. It prompted a mobilization around the globe to study the underlying pathogen, a close relative of severe acute respiratory syndrome coronavirus (SARS-CoV) called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Numerous genome sequences of SARS-CoV-2 are now available and in-depth analyses are advancing. These will allow detailed characterization of sequence and protein functions, including comparative studies. Care should be taken when inferring function from sequence information alone, and reverse genetics systems can be used to unequivocally identify key features. For example, the molecular markers of virulence, host range and transmissibility of SARS-CoV-2 can be compared to those of related viruses in order to shed light on the biology of this emerging pathogen. Here, we summarize some recent insights from genomic studies and strategies for reverse genetics systems to generate recombinant viruses, which will be useful to investigate viral genome properties and evolution.

Title: Insights into SARS-CoV-2, the Coronavirus Underlying COVID-19: Recent Genomic Data and the Development of Reverse Genetics Systems. Content: The emergence and rapid worldwide spread of a novel pandemic of acute respiratory disease - eventually named coronavirus disease 2019 (COVID-19) by the World Health Organization (WHO) - across the human population has raised great concerns. It prompted a mobilization around the globe to study the underlying pathogen, a close relative of severe acute respiratory syndrome coronavirus (SARS-CoV) called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Numerous genome sequences of SARS-CoV-2 are now available and in-depth analyses are advancing. These will allow detailed characterization of sequence and protein functions, including comparative studies. Care should be taken when inferring function from sequence information alone, and reverse genetics systems can be used to unequivocally identify key features. For example, the molecular markers of virulence, host range and transmissibility of SARS-CoV-2 can be compared to those of related viruses in order to shed light on the biology of this emerging pathogen. Here, we summarize some recent insights from genomic studies and strategies for reverse genetics systems to generate recombinant viruses, which will be useful to investigate viral genome properties and evolution.

Title: Assessing uncertainty in the rooting of the SARS-CoV-2 phylogeny Content: The rooting of the SARS-CoV-2 phylogeny is important for understanding the origin and early spread of the virus. Previously published phylogenies have used different rootings that do not always provide consistent results. We use several different strategies for rooting the SARS-CoV-2 tree and provide measures of statistical uncertainty for all methods. We show that methods based on the molecular clock tend to place the root in the B clade, while methods based on outgroup rooting tend to place the root in the A clade. The results from the two approaches are statistically incompatible, possibly as a consequence of deviations from a molecular clock or excess back-mutations. We also show that none of the methods provide strong statistical support for the placement of the root in any particular edge of the tree. Our results suggest that inferences on the origin and early spread of SARS-CoV-2 based on rooted trees should be interpreted with caution.

Title: Evolutionary analysis of SARS-CoV-2: how mutation of Non-Structural Protein 6 (NSP6) could affect viral autophagy Content: BACKGROUND: SARS-CoV-2 is a new coronavirus that has spread globally, infecting more than 150000 people, and being declared pandemic by the WHO. We provide here bio-informatic, evolutionary analysis of 351 available sequences of its genome with the aim of mapping genome structural variations and the patterns of selection. METHODS: A Maximum likelihood tree has been built and selective pressure has been investigated in order to find any mutation developed during the SARS-CoV-2 epidemic that could potentially affect clinical evolution of the infection. FINDING: We have found in more recent isolates the presence of two mutations affecting the Non-Structural Protein 6 (NSP6) and the Open Reding Frame10 (ORF 10) adjacent regions. Amino acidic change stability analysis suggests both mutations could confer lower stability of the protein structures. INTERPRETATION: One of the two mutations, likely developed within the genome during virus spread, could affect virus intracellular survival. Genome follow-up of SARS-CoV-2 spread is urgently needed in order to identify mutations that could significantly modify virus pathogenicity.

Title: Whole genome and phylogenetic analysis of two SARS-CoV-2 strains isolated in Italy in January and February 2020: additional clues on multiple introductions and further circulation in Europe Content: Whole genome sequences of SARS-CoV-2 obtained from two patients, a Chinese tourist visiting Rome and an Italian, were compared with sequences from Europe and elsewhere. In a phylogenetic tree, the Italian patient’s sequence clustered with sequences from Germany while the tourist’s sequence clustered with other European sequences. Some additional European sequences in the tree segregated outside the two clusters containing the patients’ sequences. This suggests multiple SARS-CoV-2 introductions in Europe or virus evolution during circulation.

Title: Molecular analysis of several in-house rRT-PCR protocols for SARS-CoV-2 detection in the context of genetic variability of the virus in Colombia Content: The COVID-19 pandemic caused by SARS-CoV-2 is a public health problem unprecedented in the recent history of humanity. Different in-house real-time RT-PCR (rRT-PCR) methods for SARS-CoV-2 diagnosis and the appearance of genomes with mutations in primer regions have been reported. Hence, whole-genome data from locally-circulating SARS-CoV-2 strains contribute to the knowledge of its global variability and the development and fine tuning of diagnostic protocols. To describe the genetic variability of Colombian SARS-CoV-2 genomes in hybridization regions of oligonucleotides of the main in-house methods for SARS-CoV-2 detection, RNA samples with confirmed SARS-CoV-2 molecular diagnosis were processed through next-generation sequencing. Primers/probes sequences from 13 target regions for SARS-CoV-2 detection suggested by 7 institutions and consolidated by WHO during the early stage of the pandemic were aligned with Muscle tool to assess the genetic variability potentially affecting their performance. Finally, the corresponding codon positions at the 3' end of each primer, the open reading frame inspection was identified for each gene/protein product. Complete SARS-CoV-2 genomes were obtained from 30 COVID-19 cases, representative of the current epidemiology in the country. Mismatches between at least one Colombian sequence and five oligonucleotides targeting the RdRP and N genes were observed. The 3' end of 4 primers aligned to the third codon position, showed high risk of nucleotide substitution and potential mismatches at this critical position. Genetic variability was detected in Colombian SARS-CoV-2 sequences in some of the primer/probe regions for in-house rRT-PCR diagnostic tests available at WHO COVID-19 technical guidelines; its impact on the performance and rates of false-negative results should be experimentally evaluated. The genomic surveillance of SARS-CoV-2 is highly recommended for the early identification of mutations in critical regions and to issue recommendations on specific diagnostic tests to ensure the coverage of locally-circulating genetic variants.

Title: Whole genome and phylogenetic analysis of two SARS-CoV-2 strains isolated in Italy in January and February 2020: additional clues on multiple introductions and further circulation in Europe Content: Whole genome sequences of SARS-CoV-2 obtained from two patients, a Chinese tourist visiting Rome and an Italian, were compared with sequences from Europe and elsewhere. In a phylogenetic tree, the Italian patient's sequence clustered with sequences from Germany while the tourist's sequence clustered with other European sequences. Some additional European sequences in the tree segregated outside the two clusters containing the patients' sequences. This suggests multiple SARS-CoV-2 introductions in Europe or virus evolution during circulation.

Title: Genomic Analysis and Comparative Multiple Sequence of SARS-CoV2. Content: BACKGROUND China announced an outbreak of new coronavirus in the city of Wuhan on December 31, 2019; lash to now, the virus transmission has become pandemic worldwide. Severe cases from Huanan Seafood Wholesale market in Wuhan were confirmed pneumonia with a novel coronavirus. (2019-nCoV). Understanding the molecular mechanisms of genome selection and packaging is critical for developing antiviral strategies. Thus, we defined the correlation in ten SARS-CoV2 sequences from different countries to analyze the genomic patterns of disease origin and evolution aiming for development new control pandemic processes. METHODS We apply genomic analysis to observe SARS-Co-V2 sequences from GenBank (http:// www.ncbi.nim.nih.gov/genebank/): MN 908947 China, C1), MN985325 (USA:WA, UW), MN996527 (China, C2), MT007544 (Australia:Victoria, A1), MT027064 (USA:CA, UC), MT039890 (South Korea, K1), MT066175 (Taiwan,T1), MT066176 (Taiwan, T2), LC528232 (Japan, J1), and LC528233 (Japan, J2) for genomic sequence alignment an analysis. Multiple Sequence Alignment by Clustalw (https://www.genome.jp/tools-bin/clustalw) web is applied as our alignment tool. RESULTS We analyze ten sequences from NCBI database by genome alignment and find no difference in amino acid sequences within M and N proteins. There are two amino acid variances in Spike protein region. One mutation found from south Korea sequence is verified. Two possible "L" and "S" SNP found in ORF1ab and ORF8 regions are detected CONCLUSION:: We perform genomic analysis and comparative multiple sequence of SARS-CoV-2. Studies about the biological symptoms of SARS-CoV-2 in clinic animal and humans will manipulate an understanding the origin of pandemic crisis.

Title: Molecular analysis of several in-house rRT-PCR protocols for SARS-CoV-2 detection in the context of genetic variability of the virus in Colombia. Content: The COVID-19 pandemic caused by SARS-CoV-2 is a public health problem unprecedented in the recent history of humanity. Different in-house real-time RT-PCR (rRT-PCR) methods for SARS-CoV-2 diagnosis and the appearance of genomes with mutations in primer regions have been reported. Hence, whole-genome data from locally-circulating SARS-CoV-2 strains contribute to the knowledge of its global variability and the development and fine-tuning of diagnostic protocols. To describe the genetic variability of Colombian SARS-CoV-2 genomes in hybridization regions of oligonucleotides of the main in-house methods for SARS-CoV-2 detection, RNA samples with confirmed SARS-CoV-2 molecular diagnosis were processed through next-generation sequencing. Primers/probes sequences from 13 target regions for SARS-CoV-2 detection suggested by 7 institutions and consolidated by WHO during the early stage of the pandemic were aligned with Muscle tool to assess the genetic variability potentially affecting their performance. Finally, the corresponding codon positions at the 3' end of each primer, the open reading frame inspection was identified for each gene/protein product. Complete SARS-CoV-2 genomes were obtained from 30 COVID-19 cases, representative of the current epidemiology in the country. Mismatches between at least one Colombian sequence and five oligonucleotides targeting the RdRP and N genes were observed. The 3' end of 4 primers aligned to the third codon position, showed high risk of nucleotide substitution and potential mismatches at this critical position. Genetic variability was detected in Colombian SARS-CoV-2 sequences in some of the primer/probe regions for in-house rRT-PCR diagnostic tests available at WHO COVID-19 technical guidelines; its impact on the performance and rates of false-negative results should be experimentally evaluated. The genomic surveillance of SARS-CoV-2 is highly recommended for the early identification of mutations in critical regions and to issue recommendations on specific diagnostic tests to ensure the coverage of locally-circulating genetic variants.

Title: Molecular epidemiology of the coronavirus associated with severe acute respiratory syndrome: a review of data from the Chinese University of Hong Kong. Content: The epidemic of the severe acute respiratory syndrome (SARS) has swept through the globe with more than 8000 reported probable cases. In Hong Kong, the hardest hit areas included our teaching hospital and the Amoy Gardens apartment complex. A novel coronavirus, SARS-coronavirus (SARS-CoV), with a single-stranded plus sense RNA genome, was promptly implicated as the causative agent and subsequently fulfilled Koch's postulates. To aid the understanding of SARS-CoV, groups of investigators rapidly sequenced viral isolates around the world. We were the third group in the world to release the complete SARS-CoV genome sequence (isolate CUHK-W1) on the world-wide web. With other isolates from patients of distinct epidemiological backgrounds, we additionally sequenced four complete (CUHK-Su10, CUHK-AG01, CUHK-AG02, CUHK-AG03) and two partial SARS-CoV genomes. The reviewed data obtained from representative patients from the hospital and community outbreaks has documented the evolution of the virus in this epidemic. Their sequence variations also revealed a remarkable epidemiological correlation. We demonstrate that sequence variations in the SARS-CoV genome can be applied as a powerful molecular tool in tracing the route of transmission, when used adjunctively with standard epidemiology.

Title: Molecular analysis of several in-house rRT-PCR protocols for SARS-CoV-2 detection in the context of genetic variability of the virus in Colombia Content: The COVID-19 pandemic caused by SARS-CoV-2 is a public health problem unprecedented in the recent history of humanity. Different in-house real-time RT-PCR (rRT-PCR) methods for SARS-CoV-2 diagnosis and the appearance of genomes with mutations in primer regions have been reported. Hence, whole-genome data from locally-circulating SARS-CoV-2 strains contribute to the knowledge of its global variability and the development and fine tuning of diagnostic protocols. To describe the genetic variability of Colombian SARS-CoV-2 genomes in hybridization regions of oligonucleotides of the main in-house methods for SARS-CoV-2 detection, RNA samples with confirmed SARS-CoV-2 molecular diagnosis were processed through next-generation sequencing. Primers/probes sequences from 13 target regions for SARS-CoV-2 detection suggested by 7 institutions and consolidated by WHO during the early stage of the pandemic were aligned with Muscle tool to assess the genetic variability potentially affecting their performance. Finally, the corresponding codon positions at the 3′ end of each primer, the open reading frame inspection was identified for each gene/protein product. Complete SARS-CoV-2 genomes were obtained from 30 COVID-19 cases, representative of the current epidemiology in the country. Mismatches between at least one Colombian sequence and five oligonucleotides targeting the RdRP and N genes were observed. The 3′ end of 4 primers aligned to the third codon position, showed high risk of nucleotide substitution and potential mismatches at this critical position. Genetic variability was detected in Colombian SARS-CoV-2 sequences in some of the primer/probe regions for in-house rRT-PCR diagnostic tests available at WHO COVID-19 technical guidelines; its impact on the performance and rates of false-negative results should be experimentally evaluated. The genomic surveillance of SARS-CoV-2 is highly recommended for the early identification of mutations in critical regions and to issue recommendations on specific diagnostic tests to ensure the coverage of locally-circulating genetic variants.

Title: Comparative genomics suggests limited variability and similar evolutionary patterns between major clades of SARS-CoV-2 Content: Phylogenomic analysis of SARS-CoV-2 as available from publicly available repositories suggests the presence of 3 prevalent groups of viral episomes (super-clades), which are mostly associated with outbreaks in distinct geographic locations (China, USA and Europe). While levels of genomic variability between SARS-CoV-2 isolates are limited, to our knowledge, it is not clear whether the observed patterns of variability in viral super-clades reflect ongoing adaptation of SARS-CoV-2, or merely genetic drift and founder effects. Here, we analyze more than 1100 complete, high quality SARS-CoV-2 genome sequences, and provide evidence for the absence of distinct evolutionary patterns/signatures in the genomes of the currently known major clades of SARS-CoV-2. Our analyses suggest that the presence of distinct viral episomes at different geographic locations are consistent with founder effects, coupled with the rapid spread of this novel virus. We observe that while cross species adaptation of the virus is associated with hypervariability of specific protein coding regions (including the RDB domain of the spike protein), the more variable genomic regions between extant SARS-CoV-2 episomes correspond with the 3’ and 5’ UTRs, suggesting that at present viral protein coding genes should not be subjected to different adaptive evolutionary pressures in different viral strains. Although this study can not be conclusive, we believe that the evidence presented here is strongly consistent with the notion that the biased geographic distribution of SARS-CoV-2 isolates should not be associated with adaptive evolution of this novel pathogen.

Title: Whole Genome Sequencing of SARS-CoV-2: Adapting Illumina Protocols for Quick and Accurate Outbreak Investigation During a Pandemic Content: The COVID-19 pandemic spread very fast around the world. A few days after the first detected case in South Africa, an infection started a large hospital outbreak in Durban, KwaZulu-Natal. Phylogenetic analysis of SARS-CoV-2 genomes can be used to trace the path of transmission within a hospital. It can also identify the source of the outbreak and provide lessons to improve infection prevention and control strategies. In this manuscript, we outline the obstacles we encountered in order to genotype SARS-CoV-2 in real-time during an urgent outbreak investigation. In this process, we encountered problems with the length of the original genotyping protocol, reagent stockout and sample degradation and storage. However, we managed to set up three different library preparation methods for sequencing in Illumina. We also managed to decrease the hands on library preparation time from twelve to three hours, which allowed us to complete the outbreak investigation in just a few weeks. We also fine-tuned a simple bioinformatics workflow for the assembly of high-quality genomes in real-time. In order to allow other laboratories to learn from our experience, we released all of the library preparation and bioinformatics protocols publicly and distributed them to other laboratories of the South African Network for Genomics Surveillance (SANGS) consortium.

Title: Genomics of Indian SARS-CoV-2: Implications in genetic diversity, possible origin and spread of virus Content: World Health Organization (WHO) declared COVID-19 as a pandemic disease on March 11, 2020. Comparison of genome sequences from diverse locations allows us to identify the genetic diversity among viruses which would help in ascertaining viral virulence, disease pathogenicity, origin and spread of the SARS-CoV-2 between countries. The aim of this study is to ascertain the genetic diversity among Indian SARS-CoV-2 isolates. Initial examination of the phylogenetic data of SARS-CoV-2 genomes (n=3123) from different continents deposited at GISAID (Global Initiative on Sharing All Influenza Data) revealed multiple origin for Indian isolates. An in-depth analysis of 449 viral genomes derived from samples representing countries from USA, Europe, China, East Asia, South Asia, Oceania, Middle East regions and India revealed that most Indian samples are divided into two clusters (A and B) with cluster A showing more similarity to samples from Oceania and Kuwait and the cluster B grouping with countries from Europe, Middle East and South Asia. Diversity analysis of viral clades, which are characterized by specific non-synonymous mutations in viral proteins, discovered that the cluster A Indian samples belong to I clade (V378I in ORF1ab), which is an Oceania clade with samples having Iran connections and the cluster B Indian samples belong to G clade (D614G in Spike protein), which is an European clade. Thus our study identifies that the Indian SARS-CoV-2 viruses belong to I and G clades with potential origin to be countries mainly from Oceania, Europe, Middle East and South Asia regions, which strongly implying the spread of virus through most travelled countries. The study also emphasizes the importance of pathogen genomics through phylogenetic analysis to discover viral genetic diversity and understand the viral transmission dynamics with eventual grasp on viral virulence and disease pathogenesis.

Title: COVID-19 Variants Database: A repository for Human SARS-CoV-2 Polymorphism Data Content: COVID-19 is a newly communicable disease with a catastrophe outbreak that affects all over the world. We retrieved about 8,781 nucleotide fragments and complete genomes of SARS-CoV-2 reported from sixty-four countries. The CoV-2 reference genome was obtained from the National Genomics Data Center (NGDC), GISAID, and NCBI Genbank. All the sequences were aligned against reference genomes using Clustal Omega and variants were called using in-house built Python script. We intend to establish a user-friendly online resource to visualize the variants in the viral genome along with the Primer Infopedia. After analyzing and filtering the data globally, it was made available to the public. The detail of data available to the public includes mutations from 5688 SARS-CoV-2 sequences curated from 91 regions. This database incorporated 39920 mutations over 3990 unique positions. According to the translational impact, these mutations include 11829 synonymous mutations including 681 synonymous frameshifts and 21701 nonsynonymous mutations including 10 nonsynonymous frameshifts. Development of SARS-CoV-2 mutation genome browsers is a fundamental step obliging towards the virus surveillance, viral detection, and development of vaccine and therapeutic drugs. The SARS-COV-2 mutation browser is available at http://covid-19.dnageography.com.

Title: Large scale genomic analysis of 3067 SARS-CoV-2 genomes reveals a clonal geo-distribution and a rich genetic variations of hotspots mutations Content: In late December 2019, an emerging viral infection COVID-19 was identified in Wuhan, China, and became a global pandemic. Characterization of the genetic variants of SARS-CoV-2 is crucial in following and evaluating it spread across countries. In this study, we collected and analyzed 3,067 SARS-CoV-2 genomes isolated from 55 countries during the first three months after the onset of this virus. Using comparative genomics analysis, we traced the profiles of the whole-genome mutations and compared the frequency of each mutation in the studied population. The accumulation of mutations during the epidemic period with their geographic locations was also monitored. The results showed 782 variant sites, of which 512 (65.47%) had a non-synonymous effect. Frequencies of mutated alleles revealed the presence of 38 recurrent non-synonymous mutations, including ten hotspot mutations with a prevalence higher than 0.10 in this population and distributed in six SARS-CoV-2 genes. The distribution of these recurrent mutations on the world map revealed certain genotypes specific to the geographic location. We also found co-occurring mutations resulting in the presence of several haplotypes. Moreover, evolution over time has shown a mechanism of mutation co-accumulation which might affect the severity and spread of the SARS-CoV-2. On the other hand, analysis of the selective pressure revealed the presence of negatively selected residues that could be taken into considerations as therapeutic targets We have also created an inclusive unified database (http://genoma.ma/covid-19/) that lists all of the genetic variants of the SARS-CoV-2 genomes found in this study with phylogeographic analysis around the world.

Title: Temporal evolution and adaptation of SARS-COV 2 codon usage Content: The outbreak of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) has caused an unprecedented pandemic. Since the first sequenced whole-genome of SARS-CoV-2 on January 2020, the identification of its genetic variants has become crucial in tracking and evaluating their spread across the globe. In this study, we compared 15,259 SARS-CoV-2 genomes isolated from 60 countries since the outbreak of this novel coronavirus with the first sequenced genome in Wuhan to quantify the evolutionary divergence of SARS-CoV-2. Thus, we compared the codon usage patterns, every two weeks, of 13 of SARS-CoV-2 genes encoding for the membrane protein (M), envelope (E), spike surface glycoprotein (S), nucleoprotein (N), non-structural 3C-like proteinase (3CLpro), ssRNA-binding protein (RBP), 2’-O-ribose methyltransferase (OMT), endoRNase (RNase), helicase, RNA-dependent RNA polymerase (RdRp), Nsp7, Nsp8, and exonuclease ExoN. As a general rule, we find that SARS-CoV-2 genome tends to diverge over time by accumulating mutations on its genome and, specifically, on the coding sequences for proteins N and S. Interestingly, different patterns of codon usage were observed among these genes. Genes S, Nsp7, NSp8, tend to use a norrower set of synonymous codons that are better optimized to the human host. Conversely, genes E and M consistently use a broader set of synonymous codons, which does not vary with respect to the reference genome. We identified key SARS-CoV-2 genes (S, N, ExoN, RNase, RdRp, Nsp7 and Nsp8) suggested to be causally implicated in the virus adaptation to the human host.

Title: Emergence of genomic diversity and recurrent mutations in SARS-CoV-2 Content: SARS-CoV-2 is a SARS-like coronavirus of likely zoonotic origin first identified in December 2019 in Wuhan, the capital of China's Hubei province. The virus has since spread globally, resulting in the currently ongoing COVID-19 pandemic. The first whole genome sequence was published on January 5 2020, and thousands of genomes have been sequenced since this date. This resource allows unprecedented insights into the past demography of SARS-CoV-2 but also monitoring of how the virus is adapting to its novel human host, providing information to direct drug and vaccine design. We curated a dataset of 7666 public genome assemblies and analysed the emergence of genomic diversity over time. Our results are in line with previous estimates and point to all sequences sharing a common ancestor towards the end of 2019, supporting this as the period when SARS-CoV-2 jumped into its human host. Due to extensive transmission, the genetic diversity of the virus in several countries recapitulates a large fraction of its worldwide genetic diversity. We identify regions of the SARS-CoV-2 genome that have remained largely invariant to date, and others that have already accumulated diversity. By focusing on mutations which have emerged independently multiple times (homoplasies), we identify 198 filtered recurrent mutations in the SARS-CoV-2 genome. Nearly 80% of the recurrent mutations produced non-synonymous changes at the protein level, suggesting possible ongoing adaptation of SARS-CoV-2. Three sites in Orf1ab in the regions encoding Nsp6, Nsp11, Nsp13, and one in the Spike protein are characterised by a particularly large number of recurrent mutations (>15 events) which may signpost convergent evolution and are of particular interest in the context of adaptation of SARS-CoV-2 to the human host. We additionally provide an interactive user-friendly web-application to query the alignment of the 7666 SARS-CoV-2 genomes.

Title: SARS-CoV-2 detection in nasopharyngeal throat swabs by metagenomics Content: Metagenomics could detect SARS-CoV-2 in all eight nasopharyngeal/throat swabs with high/low viral loads, and rhinovirus in a co-infected patient. The sequenced viruses belonged to lineage B1. Because metagenomics could detect novel pathogen and co-infection, and generate sequence data for epidemiological investigation, it is an attractive approach for infectious-disease diagnosis.

Title: Tracking the genomic footprints of SARS-CoV-2 transmission Content: Abstract There is considerable public and scientific interest on the origin, spread and evolution of SARS-CoV-2. A recent study by Lu et al. [1], conducted genomic sequencing and analysis of SARS-CoV-2 in Guangdong, revealing its early transmission out of Hubei and shedding light on the effectiveness of controlling local transmission chains.

Title: Genome Analysis of SARS-CoV-2 Isolate from Bangladesh Content: Recently the first genome sequence for a Severe acute respiratory syndrome coronavirus 2 or SARS-CoV-2 isolate from Bangladesh became available. The sequencing was carried out by the Child Health Research Foundation and provided the first insight into the genetic details of the viral strain responsible for the SARS-CoV-2 infections in Bangladesh. Here we carried out a comparative study were we explored the phylogenetic relationship between the Bangladeshi isolate with other isolates from different parts of the world. Afterwards we identified single nucleotide variants in the Bangladeshi isolate, using the Wuhan virus reference sequence. We found a total of 9 variants in the Bangladeshi isolate using 2 separate tools. Barring 2, the rest of these variants were also observed in other isolates from different countries. Most of the variants occurred in the ORF1ab gen. Another noteworthy finding was a sequence of three consecutive variants in the N protein gene that were observed in other isolates as well. Lastly the phylogenetic analysis revealed a close relationship between the Bangladeshi isolate and those from Taiwan, Kazakhstan, Greece, California, Spain, Israel, and Sri Lanka.

Title: Analysis of SARS-CoV-2 RNA-Sequences by Interpretable Machine Learning Models Content: We present an approach to investigate SARS-CoV-2 virus sequences based on alignment-free methods for RNA sequence comparison. In particular, we verify a given clustering result for the GISAID data set, which was obtained analyzing the molecular differences in coronavirus populations by phylogenetic trees. For this purpose, we use alignment-free dissimilarity measures for sequences and combine them with learning vector quantization classifiers for virus type discriminant analysis and classification. Those vector quantizers belong to the class of interpretable machine learning methods, which, on the one hand side provide additional knowledge about the classification decisions like discriminant feature correlations, and on the other hand can be equipped with a reject option. This option gives the model the property of self controlled evidence if applied to new data, i.e. the models refuses to make a classification decision, if the model evidence for the presented data is not given. After training such a classifier for the GISAID data set, we apply the obtained classifier model to another but unlabeled SARS-CoV-2 virus data set. On the one hand side, this allows us to assign new sequences to already known virus types and, on the other hand, the rejected sequences allow speculations about new virus types with respect to nucleotide base mutations in the viral sequences. Author summary The currently emerging global disease COVID-19 caused by novel SARS-CoV-2 viruses requires all scientific effort to investigate the development of the viral epidemy, the properties of the virus and its types. Investigations of the virus sequence are of special interest. Frequently, those are based on mathematical/statistical analysis. However, machine learning methods represent a promising alternative, if one focuses on interpretable models, i.e. those that do not act as black-boxes. Doing so, we apply variants of Learning Vector Quantizers to analyze the SARS-CoV-2 sequences. We encoded the sequences and compared them in their numerical representations to avoid the computationally costly comparison based on sequence alignments. Our resulting model is interpretable, robust, efficient, and has a self-controlling mechanism regarding the applicability to data. This framework was applied to two data sets concerning SARS-CoV-2. We were able to verify previously published virus type findings for one of the data sets by training our model to accurately identify the virus type of sequences. For sequences without virus type information (second data set), our trained model can predict them. Thereby, we observe a new scattered spreading of the sequences in the data space which probably is caused by mutations in the viral sequences.

Title: CoV2ID: Detection and Therapeutics Oligo Database for SARS-CoV-2 Content: The ability to detect the SARS-CoV-2 in a widespread epidemic is crucial for screening of carriers and for the success of quarantine efforts. Methods based on real-time reverse transcription polymerase chain reaction (RT-qPCR) and sequencing are being used for virus detection and characterization. However, RNA viruses are known for their high genetic diversity which poses a challenge for the design of efficient nucleic acid-based assays. The first SARS-CoV-2 genomic sequences already showed novel mutations, which may affect the efficiency of available screening tests leading to false-negative diagnosis or inefficient therapeutics. Here we describe the CoV2ID (http://covid.portugene.com/), a free database built to facilitate the evaluation of molecular methods for detection of SARS-CoV-2 and treatment of COVID-19. The database evaluates the available oligonucleotide sequences (PCR primers, RT-qPCR probes, etc.) considering the genetic diversity of the virus. Updated sequences alignments are used to constantly verify the theoretical efficiency of available testing methods. Detailed information on available detection protocols are also available to help laboratories implementing SARS-CoV-2 testing.

Title: Evolutionary analysis of SARS-CoV-2: how mutation of Non-Structural Protein 6 (NSP6) could affect viral autophagy Content: Abstract Background SARS-CoV-2 is a new coronavirus that has spread globally, infecting more than 150000 people, and being declared pandemic by the WHO. We provide here bio-informatic, evolutionary analysis of 351 available sequences of its genome with the aim of mapping genome structural variations and the patterns of selection. Methods A Maximum likelihood tree has been built and selective pressure has been investigated in order to find any mutation developed during the SARS-CoV-2 epidemic that could potentially affect clinical evolution of the infection. Finding We have found in more recent isolates the presence of two mutations affecting the Non-Structural Protein 6 (NSP6) and the Open Reding Frame10 (ORF 10) adjacent regions. Amino acidic change stability analysis suggests both mutations could confer lower stability of the protein structures. Interpretation One of the two mutations, likely developed within the genome during virus spread, could affect virus intracellular survival. Genome follow-up of SARS-CoV-2 spread is urgently needed in order to identify mutations that could significantly modify virus pathogenicity.

Title: The novel coronavirus SARS-CoV-2: From a zoonotic infection to coronavirus disease 2019 Content: The novel coronavirus (CoV), severe acute respiratory syndrome (SARS)-CoV-2 is an international public health emergency. Until now, the intermediate host and mechanisms of the interspecies jump of this virus are unknown. Phylogenetic analysis of all available bat CoV complete genomes was performed to analyze the relationships between bat CoV and SARS-CoV-2. To suggest a possible intermediate host, another phylogenetic reconstruction of CoV genomes obtained from animals that were hypothetically commercialized in the Chinese markets was also carried out. Moreover, mutation analysis was executed to suggest genomic regions that may have permitted the adaptation of SARS-CoV-2 to the human host. The phylogenetic analysis demonstrated that SARS-CoV-2 formed a cluster with the bat CoV isolate RaTG13. Possible CoV interspecies jumps among bat isolates were also observed. The phylogenetic tree reconstructed from CoV strains belonging to different animals demonstrated that SARS-CoV-2, bat RaTG13, and pangolin CoV genomes formed a monophyletic cluster, demonstrating that pangolins may be suggested as SARS-CoV-2 intermediate hosts. Three AA substitutions localized in the S1 portion of the S gene were observed, some of which have been correlated to structural modifications of the S protein which may facilitate SARS-CoV-2 tropism to human cells. Our analysis shows the tight relationship between SARS-CoV-2 and bat SARS-like strains. It also hypothesizes that pangolins might have been possible intermediate hosts of the infection. Some of the observed AA substitutions in the S-binding protein may serve as possible adaptation mutations in humans but more studies are needed to elucidate their function.

Title: Analyzing hCov genome sequences: Applying Machine Intelligence and beyond Content: Covid-19 pandemic, caused by the sars-cov-2 strain of coronavirus, has affected millions of people all over the world and taken thousands of lives. It is of utmost importance that the character of this deadly virus be studied and its nature be analysed. We present here an analysis pipeline comprising phylogenetic analysis on strains of this novel virus to track its evolutionary history among the countries uncovering several interesting relationships, followed by a classification exercise to identify the virulence of the strains and extraction of important features from its genetic material that are used subsequently to predict mutation at those interesting sites using deep learning techniques. In a nutshell, we have prepared an analysis pipeline for hCov genome sequences leveraging the power of machine intelligence and uncovered what remained apparently shrouded by raw data.

Title: The phylogeny of SARS coronavirus Content: Different tree-building methods consistently place the SARS corona-virus (SARS-CoV) as a basal Group 2 coronavirus rather than as an ungrouped species as concluded by others. Detailed comparisons of the SARS-CoV genomic sequence with those of six other coronaviruses failed to find evidence of recombination or genomic rearrangement using computational methods designed for that purpose.

Title: The origin and underlying driving forces of the SARS-CoV-2 outbreak Content: The spread of SARS-CoV-2 since December 2019 has become a pandemic and impacted many aspects of human society. Here, we analyzed genetic variation of SARS-CoV-2 and its related coronavirus and found the evidence of intergenomic recombination. After correction for mutational bias, analysis of 137 SARS-CoV-2 genomes as of 2/23/2020 revealed the excess of low frequency mutations on both synonymous and nonsynonymous sites which is consistent with recent origin of the virus. In contrast to adaptive evolution previously reported for SARS-CoV in its brief epidemic in 2003, our analysis of SARS-CoV-2 genomes shows signs of relaxation of selection. The sequence similarity of the spike receptor binding domain between SARS-CoV-2 and a sequence from pangolin is probably due to an ancient intergenomic introgression. Therefore, SARS-CoV-2 might have cryptically circulated within humans for years before being recently noticed. Data from the early outbreak and hospital archives are needed to trace its evolutionary path and reveal critical steps required for effective spreading. Two mutations, 84S in orf8 protein and 251V in orf3 protein, occurred coincidentally with human intervention. The 84S first appeared on 1/5/2020 and reached a plateau around 1/23/2020, the lockdown of Wuhan. 251V emerged on 1/21/2020 and rapidly increased its frequency. Thus, the roles of these mutations on infectivity need to be elucidated. Genetic diversity of SARS-CoV-2 collected from China was two time higher than those derived from the rest of the world. In addition, in network analysis, haplotypes collected from Wuhan city were at interior and have more mutational connections, both of which are consistent with the observation that the outbreak of cov-19 was originated from China. SUMMARY In contrast to adaptive evolution previously reported for SARS-CoV in its brief epidemic, our analysis of SARS-CoV-2 genomes shows signs of relaxation of selection. The sequence similarity of the spike receptor binding domain between SARS-CoV-2 and a sequence from pangolin is probably due to an ancient intergenomic introgression. Therefore, SARS-CoV-2 might have cryptically circulated within humans for years before being recently noticed. Data from the early outbreak and hospital archives are needed to trace its evolutionary path and reveal critical steps required for effective spreading. Two mutations, 84S in orf8 protein and 251V in orf3 protein, occurred coincidentally with human intervention. The 84S first appeared on 1/5/2020 and reached a plateau around 1/23/2020, the lockdown of Wuhan. 251V emerged on 1/21/2020 and rapidly increased its frequency. Thus, the roles of these mutations on infectivity need to be elucidated.

Title: Phylogenetic analysis of the full‐length SARS‐CoV sequences: Evidence for phylogenetic discordance in three genomic regions Content: The origin of the severe acute respiratory syndrome‐coronavirus (SARS‐CoV) remains unclear. Evidence based on Bayesian scanning plots and phylogenetic analysis using maximum likelihood (ML) and Bayesian methods indicates that SARS‐CoV, for the largest part of the genome (∼80%), is more closely related to Group II coronaviruses sequences, whereas in three regions in the ORF1ab gene it shows no apparent similarity to any of the previously characterized groups of coronaviruses. There is discordant phylogenetic clustering of SARS‐CoV and coronaviruses sequences, throughout the genome, compatible with either ancient recombination events or altered evolutionary rates in different lineages, or a combination of both. J. Med. Virol. 74:369–372, 2004. © 2004 Wiley‐Liss, Inc.

Title: Close relationship between SARS-coronavirus and group 2 coronavirus. Content: The sudden appearance and potential lethality of severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV) in humans has resulted in a focusing of new attention on the determination of both its origins and evolution. The relationship existing between SARS-CoV and other groups of coronaviruses was determined via analyses of phylogenetic trees and comparative genomic analyses of the coronavirus genes: polymerase (Orf1ab), spike (S), envelope (E), membrane (M) and nucleocapsid (N). Although the coronaviruses are traditionally classed into 3 groups, with SARS-CoV forming a 4th group, the phylogenetic position and origins of SARS-CoV remain a matter of some controversy. Thus, we conducted extensive phylogenetic analyses of the genes common to all coronavirus groups, using the Neighbor-joining, Maximum-likelihood, and Bayesian methods. Our data evidenced largely identical topology for all of the obtained phylogenetic trees, thus supporting the hypothesis that the relationship existing between SARS-CoV and group 2 coronavirus is a monophyletic one. Additional comparative genomic studies, including sequence similarity and protein secondary structure analyses, suggested that SARS-CoV may bear a closer relationship with group 2 than with the other coronavirus groups. Although our data strongly suggest that group 2 coronaviruses are most closely related with SARS-CoV, further and more detailed analyses may provide us with an increased amount of information regarding the origins and evolution of the coronaviruses, most notably SARS-CoV.

Title: Insights into The Codon Usage Bias of 13 Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Isolates from Different Geo-locations Content: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of Coronavirus disease 2019 (COVID-19) which is an infectious disease that spread throughout the world and was declared as a pandemic by the World Health Organization (WHO). In the present study, we analyzed genome-wide codon usage patterns in 13 SARS-CoV-2 isolates from different geo-locations (countries) by utilizing different CUB measurements. Nucleotide and di-nucleotide compositions displayed bias toward A/U content in all codon positions and CpU-ended codons preference, respectively. Relative Synonymous Codon Usage (RSCU) analysis revealed 8 common putative preferred codons among all the 13 isolates. Interestingly, all of the latter codons are A/U-ended (U-ended: 7, A-ended: 1). Cluster analysis (based on RSCU values) was performed and showed comparable results to the phylogenetic analysis (based on their whole genome sequences) indicating that the CUB pattern may reflect the evolutionary relationship between the tested isolates. To investigate the force (mutation and/or selection) influencing the pattern of CUB in SARS-CoV-2 coding sequences, we employed the following; (i). Effective number of codons (ENc), (ii). ENc-GC3 plot, (iii). Neutrality plot, and (iv) Codon adaptation index (CAI). According to their results, natural selection and/or other factors (not investigated in this study) may be the dominant force driving SARS-CoV-2 CUB. It is also worth mentioning that, by using the most expressed genes in human lung tissues as a reference set, some viral genes such as Nucleocapsid phosphoprotein, ORF7a protein, and surface glycoprotein had high CAI values which may indicate for selection force acting on their codon usage, as they play important roles in viral assembly and may help viruses avoid the host immune system. The outcome of our study may help in understanding the underlying factors involved in the evolution of SARS-CoV-2 viruses, and the interactions with their host. Also, it may aid in vaccine design strategies.

Title: Comprehensive evolution and molecular characteristics of a large number of SARS-CoV-2 genomes revealed its epidemic trend and possible origins Content: Objectives To reveal epidemic trend and possible origins of SARS-CoV-2 by exploring its evolution and molecular characteristics based on a large number of genomes since it has infected millions of people and spread quickly all over the world. Methods Various evolution analysis methods were employed. Results The estimated Ka/Ks ratio of SARS-CoV-2 is 1.008 or 1.094 based on 622 or 3624 SARS-CoV-2 genomes, and the time to the most recent common ancestor (tMRCA) was inferred in late September 2019. Further 9 key specific sites of highly linkage and four major haplotypes H1, H2, H3 and H4 were found. The Ka/Ks, detected population size and development trends of each major haplotype showed H3 and H4 subgroups were going through a purify evolution and almost disappeared after detection, indicating H3 and H4 might have existed for a long time, while H1 and H2 subgroups were going through a near neutral or neutral evolution and globally increased with time. Notably the frequency of H1 was generally high in Europe and correlated to death rate (r>0.37). Conclusions In this study, the evolution and molecular characteristics of more than 16000 genomic sequences provided a new perspective for revealing epidemiology of SARS-CoV-2.

Title: Has India met this enemy before? From an eternal optimist’s perspective: SARS-CoV-2 Content: OBJECTIVE: We are presently going through a historic and unprecedented crisis for humanity with SARS-CoV-2 causing immense damage to life and world economics. It has been 3 months, since we had the first cluster in China and we felt the need to look into certain regional patterns of transmission of the virus with respect to some distinctive living conditions, incidence of malaria, the genomics of different strains, and its impact on severity. MATERIAL AND METHODS: Data for 107 countries was compiled and correlation analysis was done between incidence of malaria and number of SARS-CoV-2 cases. Possibility of genetic similarity between SARS-CoV-2 and reported zoonotic RNA viruses found associated previously with some Plasmodium species was explored by utilizing NCBI database. RESULTS: We found a significant inverse correlation between SARS-CoV-2 disease burden and incidence of Malaria. Our analysis also showed that a 12 base pair region encoding a part of surface glycoprotein of SARS- CoV-2 aligned with the Plasmodium associated zoonotic viral genome. CONCLUSION: Our analysis suggests a significantly lower SARS-CoV-2 disease burden in Malaria endemic regions and an inverse correlation with incidence of Malaria. The possibility of a pre-existing immunological memory for SARS-CoV-2 in Indians is possible and needs to be explored further

Title: Dinucleotide repeats in coronavirus SARS-CoV-2 genome: evolutionary implications Content: The ongoing global pandemic of infection disease COVID-19 caused by the 2019 novel coronavirus (SARS-COV-2, formerly 2019-nCoV) presents critical threats to public health and the economy since it was identified in China, December 2019. The genome of SARS-CoV-2 had been sequenced and structurally annotated, yet little is known of the intrinsic organization and evolution of the genome. To this end, we present a mathematical method for the genomic spectrum, a kind of barcode, of SARS-CoV-2 and common human coronaviruses. The genomic spectrum is constructed according to the periodic distributions of nucleotides, and therefore reflects the unique characteristics of the genome. The results demonstrate that coronavirus SARS-CoV-2 exhibits dinucleotide TT islands in the non-structural proteins 3, 4, 5, and 6. Further analysis of the dinucleotide regions suggests that the dinucleotide repeats are increased during evolution and may confer the evolutionary fitness of the virus. The special dinucleotide regions in the SARS-CoV-2 genome identified in this study may become diagnostic and pharmaceutical targets in monitoring and curing the COVID-19 disease.

Title: Population genomics insights into the recent evolution of SARS-CoV-2 Content: The current coronavirus disease 2019 (COVID-19) pandemic is caused by the SARS-CoV-2 virus and is still spreading rapidly worldwide. Full-genome-sequence computational analysis of the SARS-CoV-2 genome will allow us to understand the recent evolutionary events and adaptability mechanisms more accurately, as there is still neither effective therapeutic nor prophylactic strategy. In this study, we used population genetics analysis to infer the mutation rate and plausible recombination events that may have contributed to the evolution of the SARS-CoV-2 virus. Furthermore, we localized targets of recent and strong positive selection. The genomic regions that appear to be under positive selection are largely co-localized with regions in which recombination from non-human hosts appeared to have taken place in the past. Our results suggest that the pangolin coronavirus genome may have contributed to the SARS-CoV-2 genome by recombination with the bat coronavirus genome. However, we find evidence for additional recombination events that involve coronavirus genomes from other hosts, i.e., Hedgehog and Sparrow. Even though recombination events within human hosts cannot be directly assessed, due to the high similarity of SARS-CoV-2 genomes, we infer that recombinations may have recently occurred within human hosts using a linkage disequilibrium analysis. In addition, we employed an Approximate Bayesian Computation approach to estimate the parameters of a demographic scenario involving an exponential growth of the size of the SARS-CoV-2 populations that have infected European, Asian and Northern American cohorts, and we demonstrated that a rapid exponential growth in population size can support the observed polymorphism patterns in SARS-CoV-2 genomes.

Title: Coronavirus phylogeny based on base-base correlation. Content: With more and more complete genome sequences having been released, phylogenetic analysis is entering a new era--that of phylogenomics. In this paper, a novel phylogenomic method, named as Base-Base Correlation (BBC), has been proposed to infer phylogenetic relationships from complete genomes, with particular emphasis on coronavirus phylogeny. Following the high-profile publicity of SARS outbreaks, a renewed interest in coronavirus has been promoted and two novel human coronaviruses (NL63 and HKU1) have been identified. Coronavirus phylogenomics based on BBC is well consistent with that of previous studies. BBC, to study genome information structure based on information theory, provides a novel alignment-free phylogenomic methodology in postgenome informatics.

Title: Evidence of Increasing Diversification of Emerging SARS-CoV-2 Strains Content: BACKGROUND: On January 30th, 2020, an outbreak of atypical pneumonia caused by a novel Betacoronavirus (ßCoV), named SARS-CoV-2, was declared a public health emergency of international concern by the World Health Organization. For this reason, a detailed evolutionary analysis of SARS-CoV-2 strains currently circulating in different geographic regions of the world was performed. METHODS: A compositional analysis as well as a Bayesian coalescent analysis of complete genome sequences of SARS-CoV-2 strains recently isolated in Europe, North America, South America and Asia was performed. RESULTS: The results of these studies revealed a diversification of SARS-CoV-2 strains in three different genetic clades. Co-circulation of different clades in different countries, as well as different genetic lineages within different clades were observed. The time of the most recent common ancestor (tMRCA) was established to be around November 1, 2019. A mean rate of evolution of 6.57 x 10-4 substitutions per site per year was found. A significant migration rate per genetic lineage per year from Europe to South America was also observed. CONCLUSION: The results of these studies revealed an increasing diversification of SARS-CoV-2 strains. High evolutionary rates and fast population growth characterizes the population dynamics of SARS-CoV-2 strains. This article is protected by copyright. All rights reserved.

Title: SARS-CoV-2: virus mutations in specific European populations Content: Abstract The SARS-CoV-2 virus is being intensively studied, particularly, its evolution in the increasingly available sequences between countries/continents with classical phylogenetic tree representation. More recently, certain protein mutations are correlated with specific functional impacts. Our clinical data from patients suggest different clinical symptoms between European countries. Among others, SARS-CoV-2 mutations could explain these disparities. Our analyses point out an association of diverse mutations, including co-evolving ones, in a few SARS-CoV-2 proteins, with specific countries. We therefore suggest combining clinical information from patients and the determination of the associated SARS-CoV-2 genome to better understand the specific symptoms.

Title: [Analysis of variation and evolution of SARS-CoV-2 genome] Content: OBJECTIVE: To analyze the evolution and variation of SARS-CoV-2 during the epidemic starting at the end of 2019. METHODS: We downloaded the full-length genome sequence of SARS-CoV-2 from the databases of GISAID and NCBI. Using the software for bioinformatics including MEGA-X, BEAST, and TempEst, we constructed the genomic evolution tree, inferred the time evolution signal of the virus, calculated the tMRCA time of the virus and analyzed the selection pressure of the virus during evolution. RESULTS: The phylogenetic tree showed that SARS-CoV-2 belonged to the Sarbecovirus subgenus of ß Coronavirus genus together with bat coronavirus BetaCoV/bat/Yunnan/RaTG13/2013, bat-SL-CoVZC45, bat-SL-CoVZXC21 and SARS-CoV. The genomic sequences of SARS-CoV-2 isolated from the ongoing epidemic showed a weak time evolution signal with an average tMRCA time of 73 days (95% CI: 38.9-119.3 days). No positive time evolution signal was found between SARS-CoV-2 and BetaCoV/bat/Yunnan/RaTG13/2013, but the former virus had a strong positive temporal evolution relationship with bat-SL-CoVZC45 and SARS-CoV. The major cause for mutations of SARS-CoV-2 was the pressure of purification selection during the epidemic. CONCLUSIONS: SARS-CoV-2 may have emerged as early as November, 2019, originating most likely from bat-associated coronavirus. This finding may provide evidence for tracing the sources and evolution of the virus.

Title: Coronavirus phylogeny based on 2D graphical representation of DNA sequence Content: A novel coronavirus has been identified as the cause of the outbreak of severe acute respiratory syndrome (SARS). Previous phylogenetic analyses based on sequence alignments show that SARS‐CoVs form a new group distantly related to the other three groups of previously characterized coronaviruses. In this aritcle, a new approach based on the 2D graphical representation of the whole genome sequence is proposed to analyze the phylogenetic relationships of coronaviruses. The evolutionary distances are obtained through measuring the differences among the two‐dimensional curves. © 2006 Wiley Periodicals, Inc. J Comput Chem 27: 1196–1202, 2006

Title: Evidence of Increasing Diversification of Emerging SARS‐CoV‐2 Strains Content: BACKGROUND: On January 30th, 2020, an outbreak of atypical pneumonia caused by a novel Betacoronavirus (βCoV), named SARS‐CoV‐2, was declared a public health emergency of international concern by the World Health Organization. For this reason, a detailed evolutionary analysis of SARS‐CoV‐2 strains currently circulating in different geographic regions of the world was performed. METHODS: A compositional analysis as well as a Bayesian coalescent analysis of complete genome sequences of SARS‐CoV‐2 strains recently isolated in Europe, North America, South America and Asia was performed. RESULTS: The results of these studies revealed a diversification of SARS‐CoV‐2 strains in three different genetic clades. Co‐circulation of different clades in different countries, as well as different genetic lineages within different clades were observed. The time of the most recent common ancestor (tMRCA) was established to be around November 1, 2019. A mean rate of evolution of 6.57 x 10(‐4) substitutions per site per year was found. A significant migration rate per genetic lineage per year from Europe to South America was also observed. CONCLUSION: The results of these studies revealed an increasing diversification of SARS‐CoV‐2 strains. High evolutionary rates and fast population growth characterizes the population dynamics of SARS‐CoV‐2 strains. This article is protected by copyright. All rights reserved.

Title: Phyloevolutionary analysis of SARS-CoV-2 in Nigeria Content: Abstract Background Phyloepidemiologic approaches have given specific insight to understanding emergence and evolution of infection. Knowledge on the outbreak and spread of SARS-CoV-2 in Nigeria would assist in providing preventive measures to reduce transmission among populations at risk. Therefore, this study aimed at investigating the evolution of SARS-CoV-2 in Nigeria. Materials and Method A total of 39 complete genomes of SARS-CoV-2 were retrieved from the GISAID EpiFluTM database on March 29th 2020 to investigate its evolution in Nigeria. Sequences were selected based on the travel history of the patient and the collection date. Other sequences were not selected because they were short, contained artefacts, not from original source or had insufficient information. Evolutionary history was inferred using Maximum Likelihood method based on the General Time Reversible model. Phylogenetic tree was constructed to determine the common ancestor of each strain. Results The phylogenetic analysis showed the strain in Nigeria clustered in a monophyletic clade with a Wuhan sublineage. Nucleotide alignment also showed a 100% similarity indicating a common origin of evolution. Comparative analysis showed 27,972 (93.6%) identical sites and 97.6% pairwise identity with the consensus. Conclusion The study evidently showed the entire outbreak of COVID-19 infection in Nigeria stemmed from a single introduction sharing consensus similarity with the reference SARS-CoV-2 human genome from Wuhan. Preventive measures that can limit the spread of the infection among populations at risk should be implemented.

Title: Epidemiological and Genomic Analysis of SARS-CoV-2 in Ten Patients from a Mid-sized City outside of Hubei, China Content: A novel coronavirus known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of the ongoing COVID-19 pandemic. In this study, we performed a comprehensive epidemiological and genomic analysis of SARS-CoV-2 genomes from ten patients in Shaoxing, a mid-sized city outside of the epicenter Hubei province, China, during the early stage of the outbreak (late January to early February, 2020). We obtained viral genomes with > 99% coverage and a mean depth of 296X demonstrating that viral genomic analysis is feasible via metagenomics sequencing directly on nasopharyngeal samples with SARS-CoV-2 Real-time PCR Ct values less than 28. We found that a cluster of 4 patients with travel history to Hubei shared the exact same virus with patients from Wuhan, Taiwan, Belgium and Australia, highlighting how quickly this virus spread to the globe. The virus from another cluster of husband and wife without travel history but with a sick contact of a confirmed case from another city outside of Hubei accumulated significantly more mutations (9 SNPs vs average 4 SNPs), suggesting a complex and dynamic nature of this outbreak. We also found 70% patients in this study had the S genotype, consistent with an early study showing a higher prevalence of S genotype out of Hubei than that inside Hubei. We calculated an average mutation rate of 1.37x10-3 nucleotide substitution per site per year, which is similar to that of other coronaviruses. Our findings add to the growing knowledge of the epidemiological and genomic characteristics of SARS-CoV-2 that are important for guiding outbreak containment and vaccine development. The moderate mutation rate of this virus also lends hope that development of an effective, long-lasting vaccine may be possible.

Title: Genome-wide analysis of Indian SARS-CoV-2 genomes for the identification of genetic mutation and SNP Content: The wave of COVID-19 is a big threat to the human population. Presently, the world is going through different phases of lock down in order to stop this wave of pandemic; India being no exception. We have also started the lock down on 23rd March 2020. In this current situation, apart from social distancing only a vaccine can be the proper solution to serve the population of human being. Thus it is important for all the nations to perform the genome-wide analysis in order to identify the genetic variation in Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) so that proper vaccine can be designed. This fast motivated us to analyze publicly available 566 Indian complete or near complete SARS-CoV-2 genomes to find the mutation points as substitution, deletion and insertion. In this regard, we have performed the multiple sequence alignment in presence of reference sequence from NCBI. After the alignment, a consensus sequence is build to analyze each genome in order to identify the mutation points. As a consequence, we have found 933 substitutions, 2449 deletions and 2 insertions, in total 3384 unique mutation points, in 566 genomes across 29.9 K bp. Further, it has been classified into three groups as 100 clusters of mutations (mostly deletions), 1609 point mutations as substitution, deletion and insertion and 64 SNPs. These outcomes are visualized using BioCircos and bar plots as well as plotting entropy value of each genomic location. Moreover, phylogenetic analysis has also been performed to see the evolution of SARS-CoV-2 virus in India. It also shows the wide variation in tree which indeed vivid in genomic analysis. Finally, these SNPs can be the useful target for virus classification, designing and defining the effective dose of vaccine for the heterogeneous population.

Title: Origin of imported SARS-CoV-2 strains in The Gambia identified from Whole Genome Sequences Content: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is a positive-sense single stranded RNA virus with high human transmissibility. This study generated Whole Genome data to determine the origin and pattern of transmission of SARS-CoV-2 from the first six cases tested in The Gambia. Total RNA from SARS-CoV-2 was extracted from inactivated nasopharyngeal-oropharyngeal swabs of six cases and converted to cDNA following the ARTIC COVID-19 sequencing protocol. Libraries were constructed with the NEBNext ultra II DNA library prep kit for Illumina and Oxford Nanopore Ligation sequencing kit and sequenced on Illumina MiSeq and Nanopore GridION, respectively. Sequencing reads were mapped to the Wuhan reference genome and compared to eleven other SARS-CoV-2 strains of Asian, European and American origins. A phylogenetic tree was constructed with the consensus genomes for local and non-African strains. Three of the Gambian strains had a European origin (UK and Spain), two strains were of Asian origin (Japan). In The Gambia, Nanopore and Illumina sequencers were successfully used to identify the sources of SARS-CoV-2 infection in COVID-19 cases.

Title: Tracking the Genomic Footprints of SARS-CoV-2 Transmission Content: There is considerable public and scientific interest in the origin, spread, and evolution of SARS-CoV-2. Lu et al. recently conducted genomic sequencing and analysis of SARS-CoV-2 in Guangdong, revealing its early transmission out of Hubei and shedding light on the effectiveness of controlling local transmission chains.

Title: Emergence of genomic diversity and recurrent mutations in SARS-CoV-2 Content: Abstract SARS-CoV-2 is a SARS-like coronavirus of likely zoonotic origin first identified in December 2019 in Wuhan, the capital of China's Hubei province. The virus has since spread globally, resulting in the currently ongoing COVID-19 pandemic. The first whole genome sequence was published on January 52,020, and thousands of genomes have been sequenced since this date. This resource allows unprecedented insights into the past demography of SARS-CoV-2 but also monitoring of how the virus is adapting to its novel human host, providing information to direct drug and vaccine design. We curated a dataset of 7666 public genome assemblies and analysed the emergence of genomic diversity over time. Our results are in line with previous estimates and point to all sequences sharing a common ancestor towards the end of 2019, supporting this as the period when SARS-CoV-2 jumped into its human host. Due to extensive transmission, the genetic diversity of the virus in several countries recapitulates a large fraction of its worldwide genetic diversity. We identify regions of the SARS-CoV-2 genome that have remained largely invariant to date, and others that have already accumulated diversity. By focusing on mutations which have emerged independently multiple times (homoplasies), we identify 198 filtered recurrent mutations in the SARS-CoV-2 genome. Nearly 80% of the recurrent mutations produced non-synonymous changes at the protein level, suggesting possible ongoing adaptation of SARS-CoV-2. Three sites in Orf1ab in the regions encoding Nsp6, Nsp11, Nsp13, and one in the Spike protein are characterised by a particularly large number of recurrent mutations (>15 events) which may signpost convergent evolution and are of particular interest in the context of adaptation of SARS-CoV-2 to the human host. We additionally provide an interactive user-friendly web-application to query the alignment of the 7666 SARS-CoV-2 genomes.

Title: The establishment of reference sequence for SARS‐CoV‐2 and variation analysis Content: Starting around December 2019, an epidemic of pneumonia, which was named COVID‐19 by the World Health Organization, broke out in Wuhan, China, and is spreading throughout the world. A new coronavirus, named severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) by the Coronavirus Study Group of the International Committee on Taxonomy of Viruses was soon found to be the cause. At present, the sensitivity of clinical nucleic acid detection is limited, and it is still unclear whether it is related to genetic variation. In this study, we retrieved 95 full‐length genomic sequences of SARAS‐CoV‐2 strains from the National Center for Biotechnology Information and GISAID databases, established the reference sequence by conducting multiple sequence alignment and phylogenetic analyses, and analyzed sequence variations along the SARS‐CoV‐2 genome. The homology among all viral strains was generally high, among them, 99.99% (99.91%‐100%) at the nucleotide level and 99.99% (99.79%‐100%) at the amino acid level. Although overall variation in open‐reading frame (ORF) regions is low, 13 variation sites in 1a, 1b, S, 3a, M, 8, and N regions were identified, among which positions nt28144 in ORF 8 and nt8782 in ORF 1a showed mutation rate of 30.53% (29/95) and 29.47% (28/95), respectively. These findings suggested that there may be selective mutations in SARS‐COV‐2, and it is necessary to avoid certain regions when designing primers and probes. Establishment of the reference sequence for SARS‐CoV‐2 could benefit not only biological study of this virus but also diagnosis, clinical monitoring and intervention of SARS‐CoV‐2 infection in the future.

Title: The complete genome sequence of severe acute respiratory syndrome coronavirus strain HKU-39849 (HK-39). Content: The complete genomic nucleotide sequence (29.7kb) of a Hong Kong severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) strain HK-39 is determined. Phylogenetic analysis of the genomic sequence reveals it to be a distinct member of the Coronaviridae family. 5' RACE assay confirms the presence of at least six subgenomic transcripts all containing the predicted intergenic sequences. Five open reading frames (ORFs), namely ORF1a, 1b, S, M, and N, are found to be homologues to other CoV members, and three more unknown ORFs (X1, X2, and X3) are unparalleled in all other known CoV species. Optimal alignment and computer analysis of the homologous ORFs has predicted the characteristic structural and functional domains on the putative genes. The overall nucleotides conservation of the homologous ORFs is low (<5%) compared with other known CoVs, implying that HK-39 is a newly emergent SARS-CoV phylogenetically distant from other known members. SimPlot analysis supports this finding, and also suggests that this novel virus is not a product of a recent recombinant from any of the known characterized CoVs. Together, these results confirm that HK-39 is a novel and distinct member of the Coronaviridae family, with unknown origin. The completion of the genomic sequence of the virus will assist in tracing its origin.