RESUMO
The global spread of new SARS-CoV-2 variants of concern underscore an urgent need of simple deployed molecular tools that can differentiate these lineages. Several tools and protocols have been shared since the beginning of the COVID-19 pandemic, but they need to be timely adapted to cope with SARS-CoV-2 evolution. Although whole-genome sequencing (WGS) of the virus genetic material has been widely used, it still presents practical difficulties such as high cost, shortage of available reagents in the global market, need of a specialized laboratorial infrastructure and well-trained staff. These limitations result in SARS-CoV-2 surveillance blackouts across several countries. Here we propose a rapid and accessible protocol based on Sanger sequencing of a single PCR fragment that is able to identify and discriminate all SARS-CoV-2 variants of concern (VOCs) identified so far, according to each characteristic mutational profile at the Spike-RBD region (K417N/T, E484K, N501Y, A570D). Twelve COVID-19 samples from Brazilian patients were evaluated for both WGS and Sanger sequencing: three P.2, two P.1, six B.1.1 and one B.1.1.117 lineage. All results from the Sanger sequencing method perfectly matched the mutational profile of VOCs and non-VOCs RBD's characterized by WGS. In summary, this approach allows a much broader network of laboratories to perform molecular surveillance of SARS-CoV-2 VOCs and report results within a shorter time frame, which is of utmost importance in the context of rapid public health decisions in a fast evolving worldwide pandemic.
Assuntos
COVID-19/virologia , Variação Genética , SARS-CoV-2/genética , Proteínas Virais/metabolismo , Regulação Viral da Expressão Gênica , Humanos , Reprodutibilidade dos Testes , Proteínas Virais/genéticaRESUMO
Abstract Background: Acute coronary syndrome (ACS) is a cardiovascular disease caused by obstruction of coronary arteries by atheromatous plaque. Susceptibility to this disease may be related to genetic variations, such as single nucleotide polymorphisms (SNPs). Objective: In this study, we evaluated the relationship between SNPs in IL8 (rs4073; -251 A/T) and IL16 (rs11556218; T/G) genes and SCA in a Brazilian population. Materials and Methods: A sample of 200 patients with ACS and 50 non-ACS patients hospitalized at the Real Hospital Português, Recife - PE, Brazil, and 220 blood donors (donors) was used. Genotyping was carried out by polymerase chain reaction, and DNA sequencing. Statistical analyzes were performed using the Williams G, Chi-square and Kruskal Wallis tests, using the BioEstat 5.0 program, and the data with a value of p < 0.05 were considered significant. Results: In the IL8 gene, the AT genotype was the most frequent (p > 0.05) in all three groups. In the IL16 gene, genotypic distributions were different between patients with ACS and the donor group (p = 0.002), with the most frequent G allele in the second group (p = 0.0052). The IL-16 cytokine was higher in donors than in patients with ACS (p = 0.04) and the G (TG + GG) allele had higher values of this cytokine (p = 0.01). Conclusions: The results demonstrate the important role of the rs11556218 SNP in IL16 gene in SCA, evidencing that the G allele may be associated with a decreased risk of the disease.