ABSTRACT
Background P.1 lineage (Gamma) was first described in the State of Amazonas, northern Brazil, in the end of 2020, and has emerged as a very important variant of concern (VOC) of SARS-CoV-2 worldwide. P.1 has been linked to increased infectivity, higher mortality and immune evasion, leading to reinfections and potentially reduced efficacy of vaccines and neutralizing antibodies. Methods The samples of 276 patients from the State of Amazonas were sent to a central referral laboratory for sequencing by gold standard techniques, through Illumina MiSeq platform. Both global and regional phylogenetic analyses of the successfully sequenced genomes were conducted through maximum likelihood method. Multiple alignments were obtained including previously obtained unique human SARS-CoV-2 sequences. The evolutionary histories of spike and non-structural proteins from ORF1a of northern genomes were described and their molecular evolution was analyzed for detection of positive (FUBAR, FEL, and MEME) and negative (FEL and SLAC) selective pressures. To further evaluate the possible pathways of evolution leading to the emergence of P.1, we performed specific analysis for copy-choice recombination events. A global phylogenomic analysis with subsampled P.1 and B.1.1.28 genomes was applied to evaluate the relationship among samples. Results Forty-four samples from the State of Amazonas were successfully sequenced and confirmed as P.1 (Gamma) lineage. In addition to previously described P.1 characteristic mutations, we find evidence of continuous diversification of SARS-CoV-2, as rare and previously unseen P.1 mutations were detected in spike and non-structural protein from ORF1a. No evidence of recombination was found. Several sites were demonstrated to be under positive and negative selection, with various mutations identified mostly in P.1 lineage. According to the Pango assignment, phylogenomic analyses indicate all samples as belonging to the P.1 lineage. Conclusion P.1 has shown continuous evolution after its emergence. The lack of clear evidence for recombination and the positive selection demonstrated for several sites suggest that this lineage emergence resulted mainly from strong evolutionary forces and progressive accumulation of a favorable signature set of mutations.
ABSTRACT
Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infectivity is mediated by the androgen-promoted protease, transmembrane protease, serine 2 (TMPRSS2). Previously, we have shown that treatment with proxalutamide, a non-steroidal androgen receptor antagonist, accelerates viral clearance and clinical remission in outpatients with coronavirus disease 2019 (COVID-19) compared to placebo. The effects in hospitalized COVID-19 patients were unknown. Methods: Men and women hospitalized but not requiring mechanical ventilation were randomized (1:1 ratio) to receive 300 mg of proxalutamide per day or placebo for 14 days. The study was conducted at eight sites in the state of Amazonas, Brazil. The primary outcome measure was the clinical status (8-point ordinal scale) at 14-days post-randomization. The primary efficacy endpoint was the 14-day recovery ratio (alive hospital discharge [scores 1, 2]). Findings: A total of 645 patients were randomized (317 received proxalutamide, 328 placebo) and underwent intention-to-treat analysis. The 14-day median ordinal scale score in the proxalutamide group was 1 (interquartile range [IQR]=1–2) versus 7 (IQR=2–8) for placebo, P<0.001. The 14-day recovery rate was 81.4% for proxalutamide and 35.7% for placebo (recovery ratio, 2.28; 95% CI 1.95–2.66 [P<0.001]). The 28-day all-cause mortality rate was 11.0% for proxalutamide versus 49.4% for placebo (hazard ratio, 0.16; 95% CI 0.11–0.24). The median post-randomization time to recovery was 5 days (IQR=3–8) for proxalutamide versus 10 days (IQR=6–15) for placebo.Interpretation: Hospitalized COVID-19 patients not requiring mechanical ventilation receiving proxalutamide had a 128% higher recovery rate than those treated with placebo. Clinical Trial Registration Details: ClinicalTrials.gov number, NCT04728802Funding Information: Kintor Pharmaceuticals, Ltd.Declaration of Interests: Kintor Pharmaceuticals, Ltd. manufactures and plans to market proxalutamide, and has an investigational new drug (IND) application under United States Food and Drugs Administration to conduct a Phase 3 study for proxalutamide for COVID19. Applied Biology, Inc. has patents pending regarding antiandrogen therapy for COVID19. Dr. Goren, Dr. McCoy, and Dr. Li are employees of Applied Biology, Inc. Dr. Cadegiani has served as a clinical director for Applied Biology, Inc. Dr. Wambier has served as an advisor to Applied Biology, Inc. The other authors have no conflict of interest to declare.Ethics Approval Statement: The study was approved by an ethics committee and registered in clinicaltrials.gov (NCT04728802), and also approved by Brazilian National Ethics Committee, approval number 4.513.425; CAAE 41909121.0.0000.5553; Comitê de Ética em Pesquisa (CEP) of the Comitê Nacional de Ética em Pesquisa (CONEP) of the Ministry of Health (MS). (CEP/CONEP/MS).
Subject(s)
Alzheimer Disease , Porphyria, Erythropoietic , Severe Acute Respiratory Syndrome , Multiple Sclerosis , COVID-19ABSTRACT
Almost a year after the COVID-19 pandemic had begun, The United Kingdom, South Africa, and Brazil became the epicenter of new lineages, the Variant of Concern (VOCs), B.1.1.7, B.1.351, and P.1, respectively. These VOCs are increasingly associated with enhanced transmissibility, immunity evasion, and mortality. The previous most prevalent lineages in the state of Rio Grande do South (Brazil), B.1.1.28 and B.1.1.33 were rapidly replaced by P.1 and P.2, two B.1.1.28-derived lineages harboring the E484K mutation. To perform a genomic characterization of SARS-CoV-2 samples from COVID-19 patients from the metropolitan region of Porto Alegre (Rio Grande do Sul, Southern Brazil), in this second pandemic wave, we sequenced viral samples from patients of this region to: (i) identify the prevalence of SARS-CoV-2 lineages in the region, the state and bordering countries/states, (ii) characterize the mutation spectra, and (iii) hypothesize possible viral dispersal routes by using phylogenetic and phylogeographic approaches. As results, we not only confirmed that 96.4% of the samples belonged to the P.1 lineage but also that approximately 20% of which could be assigned as the newer P.1.2 (a P.1 derived new sublineage harboring new signature substitutions recently described and present in other Brazilian states and foreign countries). Moreover, P.1 sequences from this study were allocated in several distinct branches (four clades and five clusters) of the P.1 phylogeny, suggesting multiple introductions of P.1 in Rio Grande do Sul still in 2020 and placing this state as a potential core of diffusion and emergence of P.1-derived clades. It is still uncertain if the emergence of P.1.2 and other P.1 clades are related to further virological, clinical, or epidemiological consequences. However, the clear signs of viral molecular diversification from recently introduced P.1 warrant further genomic surveillance.
Subject(s)
COVID-19ABSTRACT
The COVID-19 pandemic has already reached approximately 110 million people and it is associated with 2.5 million deaths worldwide. Brazil is the third worst-hit country, with approximately 10.2 million cases and 250 thousand deaths. International efforts have been established to share information about SARS-CoV-2 epidemiology and evolution. However, sequencing facilities and research investments are very heterogeneous across different regions and countries. The understanding of the SARS-CoV-2 evolution plays a significant role in the development of effective strategies for public health and disease management. We aimed to analyze the available and high-quality genome sequences from Brazil between February 2020 and February 2021 to identify mutation hotspots, geographical and temporal distribution of SARS-CoV-2 lineages by using phylogenetics and phylodynamics analyses. We describe heterogeneous and episodic sequencing efforts, the progression of the different lineages along time, evaluating mutational spectra and frequency oscillations derived from the prevalence of novel and specific lineages across different Brazilian regions. We found at least seven major (1-7) and two minor clades (4.2 and 5.3) related to the six most prevalent lineages in the country and described its spatial distribution and dynamics. The emergence and recent frequency shift of lineages (P.1 and P.2) containing mutations of concern in the spike protein ( e. g ., E484K, N501Y) draws attention due to their association with immune evasion and enhanced receptor binding affinity. Improvements in genomic surveillance are of paramount importance and should be extended in Brazil to better inform policy makers and enable evidence-based decisions to fight the COVID-19 pandemic.
Subject(s)
COVID-19ABSTRACT
The COVID-19 pandemic caused by SARS-CoV-2 has affected millions of people since its beginning in 2019. The propagation of new lineages and the discovery of key mechanisms adopted by the virus to overlap the immune system are central topics for the entire public health policies, research and disease management. Since the second semester of 2020, the mutation E484K has been progressively found in the Brazilian territory, composing different lineages over time. It brought multiple concerns related to the risk of reinfection and the effectiveness of new preventive and treatment strategies due to the possibility of escaping from neutralizing antibodies. To better characterize the current scenario we performed genomic and phylogenetic analyses of the E484K mutated genomes sequenced from Brazilian samples in 2020. From October, 2020, more than 40% of the sequenced genomes present the E484K mutation, which was identified in three different lineages (P1, P2 and B.1.1.33) in four Brazilian regions. We also evaluated the presence of E484K associated mutations and identified selective pressures acting on the spike protein, leading us to some insights about adaptive and purifying selection driving the virus evolution.
Subject(s)
COVID-19ABSTRACT
Brazil is the third country most affected by Covid-19 pandemic. In spite of this, viral evolution in municipality resolution is poorly understood in Brazil and it is crucial to understand the epidemiology of viral spread. We identified four main circulating lineages in Esteio (Southern Brazil) and their relationship with global, national and regional lineages using phylogenetics and phylodynamics inferences from 21 SARS-CoV-2 genome sequences. We provided a comprehensive view of viral mutations from a time- and age-representative sampling from May to October 2020, in Esteio (RS, Brazil), highlighting two frequent mutations in Spike glycoprotein (D614G and V1176F), an emergent mutation (E484K) in Spike Receptor Binding Domain (RBD) characteristic of the South African lineage B.1.351, and the adjacent replacement of 2 amino acids in Nucleocapsid phosphoprotein (R203K and G204R). A significant viral diversity was evidenced with the identification of 80 different SNPs. The E484K replacement was found in two genomes (9.5%) from samples obtained in mid-October, which is to our best knowledge the earliest description of E484K harboring SARS-CoV-2 in South Brazil. This mutation identified in a small municipality from the RS state demonstrates that it was probably widely distributed in the Brazilian territory, but went unnoticed so far by the lack of genomic surveillance in Brazil. The introduction of E484K mutants shows temporal correlation with later increases in new cases in our state. Importantly, since it has been associated with immune evasion and enhanced interaction with hACE-2, lineages containing this substitution must be the subject of intense surveillance. Our date demonstrates multiple introductions of the most prevalent lineages (B.1.1.33 and B.1.1.248) and the major role of community transmission in viral spreading and the establishment of Brazilian lineages. This represents an important contribution to the epidemiology of SARS-CoV-2.
Subject(s)
COVID-19ABSTRACT
Background: Antiandrogens have demonstrated a protective effect for COVOD-19 patients in observational and interventional studies. The goal of this study was to determine if proxalutamide, an androgen receptor antagonist, could be an effective treatment for men with COVID-19 in an outpatient setting. Study design and methods: A randomized, double-blinded, placebo-controlled clinical trial was conducted at two outpatient centers (Brasilia, Brazil). Patients were recruited from October 21 to December 24, 2020 (ClinicalTrials.gov number, NCT04446429). Male patients with confirmed COVID-19 but not requiring hospitalization (COVID-19 8-point ordinal scale <3) were administered proxalutamide 200mg/day or placebo for up to 7 days. The primary endpoint was hospitalization rate at 30 days post-randomization.Results: A total of 268 men were randomized in a 1:1 ratio. 134 patients receiving proxalutamide and 134 receiving placebo were included in the intention-to-treat analysis. The 30-day hospitalization rate was 2.2% in men taking proxalutamide compared to 26% in placebo, P<0.001. The 30-day hospitalization risk ratio was 0.09; 95% confidence interval (CI) 0.03 to 0.27. Patients in the proxalutamide arm more frequently reported gastrointestinal adverse events, however, no patient discontinued treatment. In placebo group, 6 patients were lost to follow-up, and 2 patients died from acute respiratory distress syndrome.Conclusion: The hospitalization rate in proxalutamide treated men was reduced by 91% compared to usual care. Trial Registration: NCT04446429
Subject(s)
COVID-19 , Respiratory Distress SyndromeABSTRACT
Importance: A major barrier for successful therapeutic approaches to COVID-19 is the inability during the viral replication stage, when drugs with potential antiviral activity could demonstrate efficacy and preclude progression to more severe stages. Reasons that hamper an earlier diagnosis of COVID-19 include the unspecific and mild symptoms during the first stage, the delay in the diagnosis and specific management caused by the requirement of a rtPCR-SARS-CoV-2 for the diagnosis of COVID-19, and the insufficient sensitivity of the a rtPCR-SARS-CoV-2, oppositely to what is recommended for a screening test during an outbreak. More sensitive and earlier diagnostic tools for COVID-19 should be unraveled as a key strategy for an breakthrough change in the disease course and response to specific therapies, particularly those that target the blockage of viral shedding. We aimed to create an accurate, sensitive, easy-to-perform and intuitive clinical scoring for the diagnosis of COVID-19 without the need of a rtPCR-SARS-CoV-2 (termed as The AndroCoV Clinical Scoring for COVID-19 Diagnosis), resulted from a 1,757 population cohort, eo eventually encourage the management of patients with high pre-clinical likelihood of presenting COVID-19, independent of a rtPCR-SARS-COV-2 test, to avoid delays and loss of appropriate timing for potential therapies.Materials and methods: This is a post-hoc analysis of clinical data prospectively collected of the Pre-AndroCoV and AndroCov Trials, that resulted in scorings for clinical diagnosis of COVID-19, based on the likelihood of presenting actual COVID-19 according to the number of symptoms, presence of anosmia, and known positive household contact. Sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio, and accuracy were calculated for subjects screened in two different periods and both periods together, for females, males and both, in a total of nine different scenarios, according to combinations between one, two, or three or more symptoms, or presence of anosmia in subjects without known positive household contacts, and no symptoms, one, two, or three or more symptoms, or presence of anosmia or ageusia in subjects with known positive household contacts. Scorings that yielded the highest pre-test probability, sensitivity and accuracy were selected.Results: Of the 1,757 patients screened, 1,284 were diagnosed for COVID-19. The scoring that required: 1. Two or more symptoms, or anosmia or ageusia alone, for subjects without known contact; or 2. One or more symptoms, including anosmia or ageusia alone, when with known positive contacts presented the highest accuracy (80.4%) among all combinations attempted, and higher sensitivity (85.7%) than rtPCRSARS-CoV-2 commercially available kit tests.Conclusion: The AndroCoV Clinical Scoring for COVID-19 Diagnosis demonstrated to be a feasible, easy, costless, and sensitive diagnostic tool for clinical diagnosis of COVID-19. Because clinical diagnosis of COVID-19 avoids delays in specific treatments, particularly for high-risk populations, prevents false-negative diagnosis, and reduces diagnostic costs, this diagnostic tool should be considered as an option for COVID-19 diagnosis, at least while SARS-CoV-2 is the prevailing circulating virus and vaccination rate is below the required for herd immunity.