The ACE rs1799752 Variant Is Associated with COVID-19 Severity but Is Independent of Serum ACE Activity in Hospitalized and Recovered Patients.

This paper assesses the association of the insertion/deletion ACE (angiotensin-converting enzyme) variant (rs1799752 I/D) and the serum ACE activity with the severity of COVID-19 as well as its impact on post-COVID-19, and we compare these associations with those for patients with non-COVID-19 respiratory disorders. We studied 1252 patients with COVID-19, 104 subjects recovered from COVID-19, and 74 patients hospitalized with a respiratory disease different from COVID-19. The rs1799752 ACE variant was assessed using TaqMan® Assays. The serum ACE activity was determined using a colorimetric assay. The DD genotype was related to risk for invasive mechanical ventilation (IMV) requirement as an indicator of COVID-19 severity when compared to the frequencies of II + ID genotypes (p = 0.025, OR = 1.428, 95% CI = 1.046-1.949). In addition, this genotype was significantly higher in COVID-19 and post-COVID-19 groups than in the non-COVID-19 subjects. The serum ACE activity levels were lower in the COVID-19 group (22.30 U/L (13.84-32.23 U/L)), which was followed by the non-COVID-19 (27.94 U/L (20.32-53.36 U/L)) and post-COVID-19 subjects (50.00 U/L (42.16-62.25 U/L)). The DD genotype of the rs1799752 ACE variant was associated with the IMV requirement in patients with COVID-19, and low serum ACE activity levels could be related to patients with severe disease.

Rise and fall of SARS-CoV-2 variants in Rotterdam: Comparison of wastewater and clinical surveillance.

Monitoring of SARS-CoV-2 in wastewater (WW) is a promising tool for epidemiological surveillance, correlating not only viral RNA levels with the infection dynamics within the population, but also to viral diversity. However, the complex mixture of viral lineages in WW samples makes tracking of specific variants or lineages circulating in the population a challenging task. We sequenced sewage samples of 9 WW-catchment areas within the city of Rotterdam, used specific signature mutations from individual SARS-CoV-2 lineages to estimate their relative abundances in WW and compared them against those observed in clinical genomic surveillance of infected individuals between September 2020 and December 2021. We showed that especially for dominant lineages, the median of the frequencies of signature mutations coincides with the occurrence of those lineages in Rotterdam's clinical genomic surveillance. This, along with digital droplet RT-PCR targeting signature mutations of specific variants of concern (VOCs), showed that several VOCs emerged, became dominant and were replaced by the next VOC in Rotterdam at different time points during the study. In addition, single nucleotide variant (SNV) analysis provided evidence that spatio-temporal clusters can also be discerned from WW samples. We were able to detect specific SNVs in sewage, including one resulting in the Q183H amino acid change in the Spike gene, that was not captured by clinical genomic surveillance. Our results highlight the potential use of WW samples for genomic surveillance, increasing the set of epidemiological tools to monitor SARS-CoV-2 diversity.

Tracking the molecular evolution and transmission patterns of SARS-CoV-2 lineage B.1.466.2 in Indonesia based on genomic surveillance data.

BACKGROUND: As a new epi-center of COVID-19 in Asia and a densely populated developing country, Indonesia is facing unprecedented challenges in public health. SARS-CoV-2 lineage B.1.466.2 was reported to be an indigenous dominant strain in Indonesia (once second only to the Delta variant). However, it remains unclear how this variant evolved and spread within such an archipelagic nation. METHODS: For statistical description, the spatiotemporal distributions of the B.1.466.2 variant were plotted using the publicly accessible metadata in GISAID. A total of 1302 complete genome sequences of Indonesian B.1.466.2 strains with high coverage were downloaded from the GISAID's EpiCoV database on 28 August 2021. To determine the molecular evolutionary characteristics, we performed a time-scaled phylogenetic analysis using the maximum likelihood algorithm and called the single nucleotide variants taking the Wuhan-Hu-1 sequence as reference. To investigate the spatiotemporal transmission patterns, we estimated two dynamic parameters (effective population size and effective reproduction number) and reconstructed the phylogeography among different islands. RESULTS: As of the end of August 2021, nearly 85% of the global SARS-CoV-2 lineage B.1.466.2 sequences (including the first one) were obtained from Indonesia. This variant was estimated to account for over 50% of Indonesia's daily infections during the period of March-May 2021. The time-scaled phylogeny suggested that SARS-CoV-2 lineage B.1.466.2 circulating in Indonesia might have originated from Java Island in mid-June 2020 and had evolved into two disproportional and distinct sub-lineages. High-frequency non-synonymous mutations were mostly found in the spike and NSP3; the S-D614G/N439K/P681R co-mutations were identified in its larger sub-lineage. The demographic history was inferred to have experienced four phases, with an exponential growth from October 2020 to February 2021. The effective reproduction number was estimated to have reached its peak (11.18) in late December 2020 and dropped to be less than one after early May 2021. The relevant phylogeography showed that Java and Sumatra might successively act as epi-centers and form a stable transmission loop. Additionally, several long-distance transmission links across seas were revealed. CONCLUSIONS: SARS-CoV-2 variants circulating in the tropical archipelago may follow unique patterns of evolution and transmission. Continuous, extensive and targeted genomic surveillance is essential.

Genomic chronicle of SARS-CoV-2: a mutational analysis with over 1 million genome sequences.

Use of information technologies to analyse big data on SARS-CoV-2 genome provides an insight for tracking variations and examining the evolution of the virus. Nevertheless, storing, processing, alignment and analyses of these numerous genomes are still a challenge. In this study, over 1 million SARS-CoV-2 genomes have been analysed to show distribution and relationship of variations that could enlighten development and evolution of the virus. In all genomes analysed in this study, a total of over 215M SNVs have been detected and average number of SNV per isolate was found to be 21.83. Single nucleotide variant (SNV) average is observed to reach 31.25 just in March 2021. The average variation number of isolates is increasing and compromising with total case numbers around the world. Remarkably, cytosine deamination, which is one of the most important biochemical processes in the evolutionary development of coronaviruses, accounts for 46% of all SNVs seen in SARS-CoV-2 genomes within 16 months. This study is one of the most comprehensive SARS-CoV-2 genomic analysis study in terms of number of genomes analysed in an academic publication so far, and reported results could be useful in monitoring the development of SARS-CoV-2.

Prevalence of a Single-Nucleotide Variant of SARS-CoV-2 in Korea and Its Impact on the Diagnostic Sensitivity of the Xpert Xpress SARS-CoV-2 Assay.

The sensitivity of molecular diagnostics could be affected by nucleotide variants in pathogen genes, and the sites affected by such variants should be monitored. We report a single-nucleotide variant (SNV) in the nucleocapsid (N) gene of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), i.e., G29179T, which impairs the diagnostic sensitivity of the Xpert Xpress SARS-CoV-2 assay (Cepheid, Sunnyvale, CA, USA). We observed significant differences between the threshold cycle (Ct) values for envelope (E) and N genes and confirmed the SNV as the cause of the differences using Sanger sequencing. This SNV, G29179T, is the most prevalent in Korea and is associated with the B.1.497 virus lineage, which is dominant in Korea. Clinical laboratories should be aware of the various SNVs in the SARS-CoV-2 genome and consider their potential effects on the diagnosis of coronavirus disease 2019.

Host- and Species-Dependent Quasispecies Divergence of Severe Acute Respiratory Syndrome Coronavirus-2 in Non-human Primate Models.

Recently, newly emerging variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been continuously reported worldwide. However, the precise evaluation of SARS-CoV-2 microevolution in host is very limited because the exact genetic information of infected virus could not be acquired in human researches. In this report, we performed deep sequencing for seed virus and SARS-CoV-2 isolated in eight cynomolgus and rhesus macaques at 3 days postinoculation and evaluated single-nucleotide polymorphisms (SNPs) in SARS-CoV-2 by variant analysis. A total of 69 single-nucleotide variants (SNVs) were present in the 5'-untranslated region (UTR), 3'-UTR, ORF1ab, S, ORF3a, ORF8, and N genes of the seed virus passaged in VERO cells. Between those present on the seed virus and those on each SARS-CoV-2 isolated from the lungs of the macaques, a total of 29 variants was identified in 4 coding proteins (ORF1ab, S, ORF3a, and N) and non-coding regions (5'- and 3'-UTR). Variant number was significantly different according to individuals and ranged from 2 to 11. Moreover, the average major frequency variation was identified in six sites between the cynomolgus monkeys and rhesus macaques. As with diverse SNPs in SARS-CoV-2, the values of viral titers in lungs were significantly different according to individuals and species. Our study first revealed that the genomes of SARS-CoV-2 differ according to individuals and species despite infection of the identical virus in non-human primates (NHPs). These results are important for the interpretation of longitudinal studies evaluating the evolution of the SARS-CoV-2 in human beings and development of new diagnostics, vaccine, and therapeutics targeting SARS-CoV-2.

SARS-CoV-2 genomic analyses in cancer patients reveal elevated intrahost genetic diversity.

Numerous factors have been identified to influence susceptibility to Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection and disease severity. Cancer patients are more prone to clinically evolve to more severe COVID-19 conditions, but the determinants of such a more severe outcome remain largely unknown. We have determined the full-length SARS-CoV-2 genomic sequences of cancer patients and healthcare workers (non-cancer controls) by deep sequencing and investigated the within-host viral population of each infection, quantifying intrahost genetic diversity. Naso- and oropharyngeal SARS-CoV-2+ swabs from 57 cancer patients and 14 healthcare workers from the Brazilian National Cancer Institute were collected in April to May 2020. Complete genome amplification using ARTIC network V3 multiplex primers was performed followed by next-generation sequencing. Assemblies were conducted in Geneious R11, where consensus sequences were extracted and intrahost single nucleotide variants were identified. Maximum likelihood phylogenetic analysis was performed using PhyMLv.3.0 and lineages were classified using Pangolin and CoV-GLUE. Phylogenetic analysis showed that all but one strain belonged to clade B1.1. Four genetically linked mutations known as the globally dominant SARS-CoV-2 haplotype (C241T, C3037T, C14408T and A23403G) were found in the majority of consensus sequences. SNV signatures of previously characterized Brazilian genomes were also observed in most samples. Another 85 SNVs were found at a lower frequency (1.4%-19.7%) among the consensus sequences. Cancer patients displayed a significantly higher intrahost viral genetic diversity compared to healthcare workers. This difference was independent of SARS-CoV-2 Ct values obtained at the diagnostic tests, which did not differ between the two groups. The most common nucleotide changes of intrahost SNVs in both groups were consistent with APOBEC and ADAR activities. Intrahost genetic diversity in cancer patients was not associated with disease severity, use of corticosteroids, or use of antivirals, characteristics that could influence viral diversity. Moreover, the presence of metastasis, either in general or specifically in the lung, was not associated with intrahost diversity among cancer patients. Cancer patients carried significantly higher numbers of minor variants compared to non-cancer counterparts. Further studies on SARS-CoV-2 diversity in especially vulnerable patients will shed light onto the understanding of the basis of COVID-19 different outcomes in humans.