Correction: Short Report: Early genomic detection of SARS-CoV-2 P.1 variant in Northeast Brazil.

[This corrects the article DOI: 10.1371/journal.pntd.0009591.].

Potential use of high-resolution melting analyses for SARS-CoV-2 genomic surveillance

The pandemic caused by COVID-19 and the emergence of new variants of SARS-CoV-2 have generated clinical and epidemiological impacts on a global scale. The use of strategies for monitoring viral circulation and identifying mutations in genomic regions involved in host interaction are important measures to mitigate viral dissemination and reduce its likely complications on population health. In this context, the objective of this work was to explore the potential of high-resolution melting (HRM) analysis combined with one-step real-time reverse transcription PCR in a closed-tube system, as a fast and convenient method of screening for SARS-CoV-2 mutations with possible implications on host-pathogen interactions. The HRM analyses allowed the distinction of the Gamma, Zeta, Alpha, Delta, and Omicron variants against the predecessors (B.1.1.28, B.1.1.33) of occurrence in Brazil. It is concluded that the molecular tool standardized here has the potential to optimize the genomic surveillance of SARS-CoV-2, and could be adapted for genomic surveillance of other pathogens, due to its ability to detect, prior to sequencing, samples suggestive of new variants, selecting them more assertively and earlier for whole genome sequencing when compared to random screening.

SARS-CoV-2 Spillback to Wild Coatis in Sylvatic-Urban Hotspot, Brazil.

We tested coatis (Nasua nasua) living in an urban park near a densely populated area of Brazil and found natural SARS-CoV-2 Zeta variant infections by using quantitative reverse transcription PCR, genomic sequencing, and serologic surveillance. We recommend a One Health strategy to improve surveillance of and response to COVID-19.

Global SARS-CoV-2 genomic surveillance: What we have learned (so far).

The COVID-19 pandemic has brought significant challenges for genomic surveillance strategies in public health systems worldwide. During the past thirty-four months, many countries faced several epidemic waves of SARS-CoV-2 infections, driven mainly by the emergence and spread of novel variants. In that line, genomic surveillance has been a crucial toolkit to study the real-time SARS-CoV-2 evolution, for the assessment and optimization of novel diagnostic assays, and to improve the efficacy of existing vaccines. During the pandemic, the identification of emerging lineages carrying lineage-specific mutations (particularly those in the Receptor Binding domain) showed how these mutations might significantly impact viral transmissibility, protection from reinfection and vaccination. So far, an unprecedented number of SARS-CoV-2 viral genomes has been released in public databases (i.e., GISAID, and NCBI), achieving 14 million genome sequences available as of early-November 2022. In the present review, we summarise the global landscape of SARS-CoV-2 during the first thirty-four months of viral circulation and evolution. It demonstrates the urgency and importance of sustained investment in genomic surveillance strategies to timely identify the emergence of any potential viral pathogen or associated variants, which in turn is key to epidemic and pandemic preparedness.

Retrospective Analysis of the SARS-CoV-2 Infection Profile in COVID-19 Positive Patients in Vitoria da Conquista, Northeast Brazil.

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is responsible for causing Coronavirus Disease-2019 (COVID-19), a heterogeneous clinical condition that manifests varying symptom severity according to the demographic profile of the studied population. While many studies have focused on the spread of COVID-19 in large urban centers in Brazil, few have evaluated medium or small cities in the Northeast region. The aims of this study were: (i) to identify risk factors for mortality from SARS-CoV-2 infection, (ii) to evaluate the gene expression patterns of key immune response pathways using nasopharyngeal swabs of COVID-19 patients, and (iii) to identify the circulating SARS-CoV-2 variants in the residents of a medium-sized city in Northeast Brazil. A total of 783 patients infected with SARS-CoV-2 between May 2020 and August 2021 were included in this study. Clinical-epidemiological data from patients who died and those who survived were compared. Patients were also retrospectively divided into three groups based on disease severity: asymptomatic, mild, and moderate/severe. Samples were added to a qPCR array for analyses of 84 genes involved with immune response pathways and sequenced using the Oxford Nanopore MinION technology. Having pre-existing comorbidity; being male; having cardiovascular disease, diabetes, and/or chronic obstructive pulmonary disease; and PCR cycle threshold (Ct) values under 22 were identified as risk factors for mortality. Analysis of the expression profiles of inflammatory pathway genes showed that the greater the infection severity, the greater the activation of inflammatory pathways, triggering the cytokine storm and downregulating anti-inflammatory pathways. Viral genome analysis revealed the circulation of multiple lineages, such as B.1, B.1.1.28, Alpha, and Gamma, suggesting that multiple introduction events had occurred over time. This study's findings help identify the specific strains and increase our understanding of the true state of local health. In addition, our data demonstrate that epidemiological and genomic surveillance together can help formulate public health strategies to guide governmental actions.

Dynamics of SARS-CoV-2 Variants of Concern in Vaccination Model City in the State of Sao Paulo, Brazil.

From a country with one of the highest SARS-CoV-2 morbidity and mortality rates, Brazil has implemented one of the most successful vaccination programs. Brazil's first model city vaccination program was performed by the CoronaVac vaccine (Sinovac Biotech) in the town of Serrana, São Paulo State. To evaluate the vaccination effect on the SARS-CoV-2 molecular dynamics and clinical outcomes, we performed SARS-CoV-2 molecular surveillance on 4375 complete genomes obtained between June 2020 and April 2022 in this location. This study included the period between the initial SARS-CoV-2 introduction and during the vaccination process. We observed that the SARS-CoV-2 substitution dynamics in Serrana followed the viral molecular epidemiology in Brazil, including the initial identification of the ancestral lineages (B.1.1.28 and B.1.1.33) and epidemic waves of variants of concern (VOC) including the Gamma, Delta, and, more recently, Omicron. Most probably, as a result of the immunization campaign, the mortality during the Gamma and Delta VOC was significantly reduced compared to the rest of Brazil, which was also related to lower morbidity. Our phylogenetic analysis revealed the evolutionary history of the SARS-CoV-2 in this location and showed that multiple introduction events have occurred over time. The evaluation of the COVID-19 clinical outcome revealed that most cases were mild (88.9%, 98.1%, 99.1% to Gamma, Delta, and Omicron, respectively) regardless of the infecting VOC. In conclusion, we observed that vaccination was responsible for reducing the death toll rate and related COVID-19 morbidity, especially during the gamma and Delta VOC; however, it does not prevent the rapid substitution rate and morbidity of the Omicron VOC.

Genomic epidemiology of the SARS-CoV-2 epidemic in Brazil.

The high numbers of COVID-19 cases and deaths in Brazil have made Latin America an epicentre of the pandemic. SARS-CoV-2 established sustained transmission in Brazil early in the pandemic, but important gaps remain in our understanding of virus transmission dynamics at a national scale. We use 17,135 near-complete genomes sampled from 27 Brazilian states and bordering country Paraguay. From March to November 2020, we detected co-circulation of multiple viral lineages that were linked to multiple importations (predominantly from Europe). After November 2020, we detected large, local transmission clusters within the country. In the absence of effective restriction measures, the epidemic progressed, and in January 2021 there was emergence and onward spread, both within and abroad, of variants of concern and variants under monitoring, including Gamma (P.1) and Zeta (P.2). We also characterized a genomic overview of the epidemic in Paraguay and detected evidence of importation of SARS-CoV-2 ancestor lineages and variants of concern from Brazil. Our findings show that genomic surveillance in Brazil enabled assessment of the real-time spread of emerging SARS-CoV-2 variants.

Molecular dynamics simulations of the SARS-CoV-2 Spike protein and variants of concern: structural evidence for convergent adaptive evolution.

The Spike protein's structure of the SARS-CoV-2 provides a unique opportunity to consider perturbations at the atomic level. We used the cryo-electron microscopy structure of the open conformation of the Spike protein to assess the impact of the mutations observed in the variants of concern at the molecular level. Molecular dynamics were subsequently performed with both the wt and the mutated forms to compare the flexibility and variation data for each residue of the three-dimensional fluctuations in the region associated with each alpha carbon. Additionally, protein-protein docking was used to investigate the interaction of each mutated profile with the ACE-2 receptor. After the molecular dynamics, the results show that the mutations increased the stability of the trimeric protein, with greater stability observed in the Gamma variant harboring the 10 characteristic mutations. The results of molecular dynamics, as shown by RMSF demonstrated for the residues that comprise the binding domain receptor (RBD), exhibited a reduction in flexibility, which was more pronounced in the Gamma variant. Finally, protein-protein docking experiments revealed an increase in the number of hydrophobic interactions and hydrogen bonds in the Gamma variant against the ACE-2 receptor, as opposed to the other variants. Taken together, these in silico experiments suggest that the evolution of the mutations favored the increased stability of Spike protein while potentially improving its interaction with the ACE-2 receptor, which in turn may indicate putative structural outcomes of the selection of these mutations in the convergent adaptive evolution as it has been observed for SARS-CoV-2.Communicated by Ramaswamy H. Sarma.

SARS-CoV-2 epidemic in Brazil: how the displacement of variants has driven distinct epidemic waves.

Brazil ranks as third in terms of total number of reported SARS-CoV-2 cases globally. The COVID-19 epidemic in Brazil was characterised by the co-circulation of multiple variants as a consequence of multiple independent introduction events occurring through time. Here, we describe the SARS-CoV-2 variants that are currently circulating and co-circulating in the country, with the aim to highlight which variants have driven the different epidemic waves. For this purpose, we retrieved metadata information of Coronavirus sequences collected in Brazil and available at the GISAID database. SARS-CoV-2 lineages have been identified along with eleven variants, labelled as VOCs (Alpha, Gamma, Beta, Delta and Omicron) VOIs (Lambda and Mu) VUMs (B.1.1.318) and FMVs (Zeta, Eta and B.1.1.519). Here we show that, in the Brazilian context, after 24 months of sustained transmission and evolution of SARS-CoV-2, local variants (among them the B.1.1.28 and B.1.1.33) were displaced by recently introduced VOCs firstly with the Gamma, followed by Delta and more recently Omicron. The rapid spread of some of those VOCs (such as Gamma and Omicron) was also mirror by a large increase in the number of cases and deaths in the country. This in turn reinforces that, due to the emergence of variants that appear to induce a substantial evasion against neutralizing antibody response, it is important to strengthen genomic effort within the country and how vaccination still remains a critical process to protect the vulnerable population, still at risk of infection and death.

Replacement of the Gamma by the Delta variant in Brazil: Impact of lineage displacement on the ongoing pandemic.

The coronavirus disease 2019 (COVID-19) epidemic in Brazil was driven mainly by the spread of Gamma (P.1), a locally emerged variant of concern (VOC) that was first detected in early January 2021. This variant was estimated to be responsible for more than 96 per cent of cases reported between January and June 2021, being associated with increased transmissibility and disease severity, a reduction in neutralization antibodies and effectiveness of treatments or vaccines, and diagnostic detection failure. Here we show that, following several importations predominantly from the USA, the Delta variant rapidly replaced Gamma after July 2021. However, in contrast to what was seen in other countries, the rapid spread of Delta did not lead to a large increase in the number of cases and deaths reported in Brazil. We suggest that this was likely due to the relatively successful early vaccination campaign coupled with natural immunity acquired following prior infection with Gamma. Our data reinforce reports of the increased transmissibility of the Delta variant and, considering the increasing concern due to the recently identified Omicron variant, argues for the necessity to strengthen genomic monitoring on a national level to quickly detect the emergence and spread of other VOCs that might threaten global health.

Introduction of SARS-CoV-2 C.37 (WHO VOI lambda) in the Sao Paulo State, Southeast Brazil.

The Lambda variants of interest (VOI) (C37/GR/452Q.V1/21G) was initially reported in Lima, Peru but has gained rapid dissemination through other Latin American countries. Nevertheless, the dissemination and molecular epidemiology of the Lambda VOI in Brazil is unknown apart from a single case report. In this respect, we characterized the circulation of the SARS-CoV-2 Lambda VOI (C37/GR/452Q.V1/21G) in Sao Paulo State, Brazil. From March to June 2021, we identified seven Lambda isolates in a set of approximately 8000 newly sequenced genomes of the Network for Pandemic Alert of Emerging SARS-CoV-2 variants from Sao Paulo State. Interestingly, in three of the positive patients, the Lambda VOI infection was probably related to a contact transmission. These individuals were fully vaccinated to COVID-19 and presented mild symptoms. The remaining positive for Lambda VOI individuals showed different levels of COVID-19 symptoms and one of them needed hospitalization (score 5, WHO). In our study, we present a low level of Lambda VOI circulation in the Sao Paulo State. This reinforces the essential role of molecular surveillance for the effective SARS-CoV-2 pandemic response, especially in regard to circulating variants.

Genomic monitoring unveil the early detection of the SARS-CoV-2 B.1.351 (beta) variant (20H/501Y.V2) in Brazil.

Sao Paulo State, currently experiences a second COVID-19 wave overwhelming the healthcare system. Due to the paucity of SARS-CoV-2 complete genome sequencing, we established a Network for Pandemic Alert of Emerging SARS-CoV-2 Variants to rapidly understand and monitor the spread of SARS-CoV-2 variants into the state. Through analysis of 210 SARS-CoV-2 complete genomes obtained from the largest regional health departments we identified cocirculation of multiple SARS-CoV-2 lineages such as B.1.1 (0.5%), B.1.1.28 (23.2%), B.1.1.7 (alpha variant, 6.2%), B.1.566 (1.4%), B.1.544 (0.5%), C.37 (0.5%) P.1 (gamma variant, 66.2%), and P.2 (zeta variant, 1.0%). Our analysis allowed also the detection, for the first time in Brazil, the South African B.1.351 (beta) variant of concern, B.1.351 (501Y.V2) (0.5%), characterized by the following mutations: ORF1ab: T265I, R724K, S1612L, K1655N, K3353R, SGF 3675_F3677del, P4715L, E5585D; spike: D80A, D215G, L242_L244del, A262D, K417N, E484K, N501Y, D614G, A701V, C1247F; ORF3a: Q57H, S171L, E: P71L; ORF7b: Y10F, N: T205I; ORF14: L52F. The most recent common ancestor of the identified strain was inferred to be mid-October to late December 2020. Our analysis demonstrated the P.1 lineage predominance and allowed the early detection of the South African strain for the first time in Brazil. We highlight the importance of SARS-CoV-2 active monitoring to ensure the rapid detection of potential variants for pandemic control and vaccination strategies. Highlights Identification of B.1.351 (beta) variant of concern in the Sao Paulo State. Dissemination of SARS-CoV-2 variants of concern and interest in the Sao Paulo State. Mutational Profile of the circulating variants of concern and interest.

SARS-CoV-2 shifting transmission dynamics and hidden reservoirs potentially limit efficacy of public health interventions in Italy.

We investigated SARS-CoV-2 transmission dynamics in Italy, one of the countries hit hardest by the pandemic, using phylodynamic analysis of viral genetic and epidemiological data. We observed the co-circulation of multiple SARS-CoV-2 lineages over time, which were linked to multiple importations and characterized by large transmission clusters concomitant with a high number of infections. Subsequent implementation of a three-phase nationwide lockdown strategy greatly reduced infection numbers and hospitalizations. Yet we present evidence of sustained viral spread among sporadic clusters acting as "hidden reservoirs" during summer 2020. Mathematical modelling shows that increased mobility among residents eventually catalyzed the coalescence of such clusters, thus driving up the number of infections and initiating a new epidemic wave. Our results suggest that the efficacy of public health interventions is, ultimately, limited by the size and structure of epidemic reservoirs, which may warrant prioritization during vaccine deployment.

Genomic surveillance activities unveil the introduction of the SARS-CoV-2 B.1.525 variant of interest in Brazil: Case report.

The appearance of new variants of SARS-CoV-2 has recently challenged public health authorities with respect to tracking transmission and mitigating the impact in the evolving pandemic across countries. B.1.525 is considered a variant under investigation since it carries specific genetic signatures present in P.1, B.1.1.7, and B.1.351. Here we report genomic evidence of the first likely imported case of the SARS-CoV-2 B.1.525 variant, isolated in a traveler returning from Nigeria.

Betacoronaviruses genome analysis reveals evolution toward specific codons usage: Implications for SARS-CoV-2 mitigation strategies.

Since the start of the coronavirus disease 2019 (COVID-19) pandemic, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly widespread worldwide becoming one of the major global public health issues of the last centuries. Currently, COVID-19 vaccine rollouts are finally upon us carrying the hope of herd immunity once a sufficient proportion of the population has been vaccinated or infected, as a new horizon. However, the emergence of SARS-CoV-2 variants brought concerns since, as the virus is exposed to environmental selection pressures, it can mutate and evolve, generating variants that may possess enhanced virulence. Codon usage analysis is a strategy to elucidate the evolutionary pressure of the viral genome suffered by different hosts, as possible cause of the emergence of new variants. Therefore, to get a better picture of the SARS-CoV-2 codon bias, we first identified the relative codon usage rate of all Betacoronaviruses lineages. Subsequently, we correlated putative cognate transfer ribonucleic acid (tRNAs) to reveal how those viruses adapt to hosts in relation to their preferred codon usage. Our analysis revealed seven preferred codons located in three different open reading frame which appear preferentially used by SARS-CoV-2. In addition, the tRNA adaptation analysis indicates a wide strategy of competition between the virus and mammalian as principal hosts highlighting the importance to reinforce the genomic monitoring to prompt identify any potential adaptation of the virus into new potential hosts which appear to be crucial to prevent and mitigate the pandemic.

Sixteen novel lineages of SARS-CoV-2 in South Africa.

The first severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in South Africa was identified on 5 March 2020, and by 26 March the country was in full lockdown (Oxford stringency index of 90)1. Despite the early response, by November 2020, over 785,000 people in South Africa were infected, which accounted for approximately 50% of all known African infections2. In this study, we analyzed 1,365 near whole genomes and report the identification of 16 new lineages of SARS-CoV-2 isolated between 6 March and 26 August 2020. Most of these lineages have unique mutations that have not been identified elsewhere. We also show that three lineages (B.1.1.54, B.1.1.56 and C.1) spread widely in South Africa during the first wave, comprising ~42% of all infections in the country at the time. The newly identified C lineage of SARS-CoV-2, C.1, which has 16 nucleotide mutations as compared with the original Wuhan sequence, including one amino acid change on the spike protein, D614G (ref. 3), was the most geographically widespread lineage in South Africa by the end of August 2020. An early South African-specific lineage, B.1.106, which was identified in April 2020 (ref. 4), became extinct after nosocomial outbreaks were controlled in KwaZulu-Natal Province. Our findings show that genomic surveillance can be implemented on a large scale in Africa to identify new lineages and inform measures to control the spread of SARS-CoV-2. Such genomic surveillance presented in this study has been shown to be crucial in the identification of the 501Y.V2 variant in South Africa in December 2020 (ref. 5).

Short Report: Early genomic detection of SARS-CoV-2 P.1 variant in Northeast Brazil.

Tracking the spread of SARS-CoV-2 variants of concern is crucial to inform public health efforts and control the ongoing pandemic. Here, we report genetic evidence for circulation of the P.1 variant in Northeast Brazil. We advocate for increased active surveillance to ensure adequate control of this variant throughout the country.

Molecular surveillance of the on-going SARS-COV-2 epidemic in Ribeirao Preto City, Brazil.

The Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of an unprecedented worldwide pandemic. Brazil demonstrates one of the highest numbers of confirmed SARS-CoV-2 cases, and São Paulo State is the epicenter of the pandemics in the country. Nevertheless, little is known about the SARS-CoV-2 circulation in other cities in the State than São Paulo city. The objective of this study was to analyze phylogenetically SARS-CoV-2 strains circulating in city of Ribeirão Preto at the beginning of the pandemic and during the actual second wave. Twenty-nine nasopharyngeal SARS-CoV-2 RNA positive samples were sequenced by nanopore technology (18 obtained at the initial period of the pandemic and 11 during the second wave) and analyzed them phylogenetically. The performed analysis demonstrated that the majority of the strains obtained in the initial period of the pandemic in Ribeirão Preto belonged mainly to the B1.1.33 lineage (61.1%), but B.1.1 (27.8%) and B.1.1.28 (11.1%) lineages were also identified. In contrast, the second wave strains were composed exclusively by the Brazilian variant of concern (VOC) P.1 (91%) and P.2 (9%) lineages. The obtained phylogenetic results were suggestive of successive SARS-CoV-2 lineage substitution in this Brazilian region by the P.1 VOC. The performed study examines the SARS-CoV-2 genotypes in Ribeirão Preto city via genomic surveillance data. The obtained findings can contribute for continuous long-term genomic surveillance of SARS-CoV-2 due to the accelerated dynamics of viral lineage substitution, predict further waves and examine lineage behavior during SARS-CoV-2 vaccination.