The current COVID-19 pandemic demands massive testing by Real-time RT-PCR (Reverse Transcription Polymerase Chain Reaction), which is considered the gold standard diagnostic test for the detection of the SARS-CoV-2 virus. However, the virus continues to evolve with mutations that lead to phenotypic alterations as higher transmissibility, pathogenicity or vaccine evasion. Another big issue are mutations in the annealing sites of primers and probes of RT-PCR diagnostic kits leading to false-negative results. Therefore, here we identify mutations in the N (Nucleocapsid) gene that affects the use of the GeneFinder COVID-19 Plus RealAmp Kit. We sequenced SARS-CoV-2 genomes from 17 positive samples with no N gene detection but with RDRP (RNA-dependent RNA polymerase) and E (Envelope) genes detection, and observed a set of three different mutations affecting the N detection: a deletion of 18 nucleotides (Del28877-28894), a substitution of GGG to AAC (28881-28883) and a frameshift mutation caused by deletion (Del28877-28878). The last one cause a deletion of six AAs (amino acids) located in the central intrinsic disorder region at protein level. We also found this mutation in 99 of the 14,346 sequenced samples by the Sao Paulo state Network for Pandemic Alert of Emerging SARS-CoV-2 variants, demonstrating the circulation of the mutation in Sao Paulo, Brazil. Continuous monitoring and characterization of mutations affecting the annealing sites of primers and probes by genomic surveillance programs are necessary to maintain the effectiveness of the diagnosis of COVID-19.
O Pará está entre os estados brasileiros com as mais elevadas taxas de prevalências da infecção pelo vírus T-linfotrópico humano (HTLV) nas populações onde este vírus já foi investigado. A infecção por HTLV geralmente apresenta prevalência mais elevada em mulheres com mais de 40 anos e com relacionamento estável. Além disso, as mulheres são mais susceptíveis à aquisição do vírus por via sexual do que os homens. Neste contexto, este estudo teve como objetivo detectar o genoma do provírus de HTLV em secreção cérvico-vaginal, com posterior confirmação em amostras de sangue, visando assim, propor uma nova metodologia de rastreamento desta infecção. Foram investigadas 400 mulheres de novembro de 2015 a dezembro de 2019, em Belém, Pará, Brasil. A coleta de secreção cérvico-vaginal se deu durante a realização do exame de Papanicolaou, e a de sangue periférico, durante contato posterior. O DNA das amostras foi extraído e realizada a análise molecular por Nested-PCR, seguida de digestão enzimática por Taq I, para pesquisa da infecção pelos tipos HTLV-1 e HTLV-2. Cinco (1,25%) das 400 mulheres tiveram resultado positivo para HTLV em secreção, sendo três para HTLV-1 e duas para HTLV-2. Por dificuldade de contato, só foi possível a confirmação da infecção no sangue em uma destas cinco mulheres. Nossos achados sugerem ser possível utilizar amostras de secreção cérvico-vaginal como mais uma forma de rastreamento da infecção por HTLV em mulheres.
São Paulo state has been the epicenter of the Coronavirus Disease 2019 (COVID-19) in Brazil, ranking first by state with over six million reported cases. In February 2021, the P.4 lineage was reported in 21 cities across the state by public health authorities due to the L452R mutation. Here, by analyzing 17,304 genome sequences of SARS-CoV-2 sampled between February and August of 2021 in 476 distinct cities in São Paulo, we assess the transmission dynamics of the P.4 lineage and other SARS-CoV-2 variants that were, at the time of the study, co-circulating in the state. Additionally, clinical parameters from the city of Araras, São Paulo (N = 251) were considered to estimate the potential risk and mortality rate associated with the P.4 lineage since its higher prevalence was observed in that city. Our data suggest a low frequency (0.55%) of the P.4 lineage across the state, with the gamma variant being the dominant form in all regions (90%) at that time. Furthermore, no evidence of increased transmissibility and disease severity related to the P.4 lineage was observed. The displacement through the time of different lineages in São Paulo highlights how challenging genomic surveillance appears to track the emergence of new SARS-CoV-2 lineages, which could better guide the implementation of control measures.
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