Y380Q novel mutation in receptor-binding domain of SARS-CoV-2 spike protein together with C379W interfere in the neutralizing antibodies interaction (preprint)

BackgroundThe emergence of SARS-CoV-2 variants is a current public health concern possibly impacting COVID-19 disease diagnosis, transmission patterns and vaccine effectiveness. ObjectivesTo describe the SARS-CoV-2 lineages circulating early pandemic among samples with S gene dropout and characterize a novel mutation in receptor-binding domain (RBD) of viral spike protein. Study designAdults and children older than 2 months with signs and symptoms of COVID-19 were prospectively enrolled from May to October 2020 in Porto Alegre, Brazil. All participants performed RT-PCR assays for diagnosing SARS-CoV-2, samples with S gene dropout and Ct < 30 (cycle threshold) were submitted to whole genome sequencing (WGS), and homology modeling and physicochemical properties analysis were performed. Results484/1,557 participants tested positive for SARS-CoV-2. The S gene dropout was detected in 7.4% (36/484) as early as May, and a peak was observed in early August. WGS was performed in 8 samples. The B.1.1.28, B.1.91 and B.1.1.33 lineages were circulating in early pandemic. The RBD novel mutation (Y380Q) was found in one sample occurring simultaneously with C379W and V395A, and the B.1.91 lineage in the spike protein. ConclusionMutations in the SARS-CoV-2 spike region were detected early in the COVID-19 pandemic in Southern Brazil, regarding the B.1.1.28, B.1.91 and B.1.1.33 lineages identified. The novel mutation (Y380Q) with C379W, modifies important RBD properties, which may interfere with the binding of neutralizing antibodies (CR3022, EY6A, H014, S304). HighlightsO_LICharacterization of novel mutation (Y380Q) in RBD of SARS-CoV-2 spike protein C_LIO_LIThe Y380Q and C379W modify important properties in the SARS-CoV-2 RBD region C_LIO_LIThe RBD mutations may interfere with the binding of neutralizing antibodies C_LIO_LIThe B.1.1.28, B.1.91 and B.1.1.33 lineages were circulating in early pandemic C_LI

Predominance of SARS-CoV-2 P.1 (Gamma) lineage inducing the recent wave in southern Brazil and the finding of an additional S: D614A mutation (preprint)

Since the first SARS-CoV-2 case in China in late 2019, this virus continues to impact many countries worldwide. Recently, the highest wave of SARS-CoV-2 epidemic was registered in Rio Grande do Sul (RS) State, Southern Brazil, considering the number of cases, deaths and hospitalization per day caused by COVID-19. In this study we described which lineages were circulating in the beginning of 2021 in southern Brazil to understand the cause of the highest health crisis caused by SARS-CoV-2 in the region, searching also possible additional SARS-CoV-2 sequence mutation. A total of 71 positive SARS-CoV-2 samples collected between December 15th, 2020, until April 23rd, 2021, were selected to sequencing. The samples were from 16 cities distributed in three RS regions. The library preparation was performed according to QIAseq® SARS-CoV-2 Primer Panel Handbook kit, and phylogenetic analysis was realized. Whole genome sequencing of 71 SARS-CoV-2 samples shows a predominance of Gamma lineage (67.14%, 47/71), followed by Zeta lineage (27.14%, 20/71) and B.1.1.28 (5,71%, 4/71). Two Gamma lineage consensus sequences presented a D614A mutation. Newly mutations could be emerging due the quick SARS-CoV-2 spreading. Thus, the greater understanding about immune protection and variants vigilance is essential to the better management of the health SARS-CoV-2 crisis.

Emergence of a putative novel SARS-CoV-2 P.X lineage harboring N234P and E471Q spike protein mutations in Southern Brazil (preprint)

Novel SARS-CoV-2 lineages are constantly reported worldwide, raising concerns about transmissibility, virulence, immune response and vaccine/antigenic escape. Variants of concern (VOCs), as B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma) and B.1.617.2 (Delta), caused epidemic outbreaks due their higher potential of transmissibility when compared with earlier waves of SARS-CoV-2 in 2019. B.1.1.28 lineage has been evolving in Brazil since February 2020 and originated P.1 (VOC), P.2 (VOI) and other P.Xs proposed as new variants. This lineage harbors specific defining mutations including two non-synonymous substitutions in the Spike (S) protein (D614G and V1176F). In this study, employing variant calling analysis on FASTQ reads and phylogenetic inference, we report a potentially new SARS-CoV-2 P.X variant. Variant calling mutational profile was investigated and presented additionally non-synonymous mutations when compared to B.1.1.28, including N234P and E471Q in S protein. Further studies are required to understand the spread of P.X variant and its potential effects on transmissibility and immune escape.

Early detection of SARS-CoV-2 P.1 variant in Southern Brazil and reinfection of the same patient by P.2 (preprint)

Multiple variants of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are constantly reported across the world. B.1.1.28 lineage has been evolving in Brazil since February 2020 and originated P.1 (VOC) and P.2 (VOI). Here we describe an early case of P.1 primary infection in Southern Brazil in late November 2020, soon after the emergence of the variant in Manaus, Northern Brazil. The same patient suffered a reinfection case by another B1.1.28 variant, namely P.2, in March, 2021. Genomic analysis showed genetically significant differences between the viruses recovered in both infections. P.1 lineage at the first episode and P.2 at the reinfection. We also have investigated the circulation of P.1 in the same region by differential RT-qPCR, showing that this was an isolated case of P.1 at the time of detection, whereas the variant disseminated and became prominent from late January 2021 to the end of March 2021.

Genomic Epidemiology of SARS-CoV-2 in Esteio, Rio Grande do Sul, Brazil (preprint)

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.

Pervasive transmission of E484K and emergence of VUI-NP13L with evidence of SARS-CoV-2 co-infection events by two different lineages in Rio Grande do Sul, Brazil (preprint)

Emergence of novel SARS-CoV-2 lineages are under the spotlight of the media, scientific community and governments. Recent reports of novel variants in the United Kingdom, South Africa and Brazil (B.1.1.28-E484K) have raised intense interest because of a possible higher transmission rate or resistance to the novel vaccines. Nevertheless, the spread of B.1.1.28 (E484K) and other variants in Brazil is still unknown. In this work, we investigated the population structure and genomic complexity of SARS-CoV-2 in Rio Grande do Sul, the southernmost state in Brazil. Most samples sequenced belonged to the B.1.1.28 (E484K) lineage, demonstrating its widespread dispersion. We were the first to identify two independent events of co-infection caused by the occurrence of B.1.1.28 (E484K) with either B.1.1.248 or B.1.91 lineages. Also, clustering analysis revealed the occurrence of a novel cluster of samples circulating in the state (named VUI-NP13L) characterized by 12 lineage-defining mutations. In light of the evidence for E484K dispersion, co-infection and emergence of VUI-NP13L in Rio Grande do Sul, we reaffirm the importance of establishing strict and effective social distancing measures to counter the spread of potentially more hazardous SARS-CoV-2 strains. HighlightsO_LIThe novel variant B.1.1.28 (E484K) previously described in Rio de Janeiro is currently spread across the southernmost state of Brazil; C_LIO_LIThe novel variant VUI-NP13L was also identified by causing a local outbreak in Rio Grande do Sul; C_LIO_LIB.1.1.28 (E484K) is able to establish successful coinfection events co-occurring simultaneously with different lineages of SARS-CoV-2. C_LI

Comparison Of Different Kits For SARS-CoV-2 RNA Extraction Marketed In Brazil (preprint)

December 2019 marked the begining of the greatest pandemic since Spanish Flu, the disease named Covid-19 that cause severe pneumonia. Until May 19, 2020 more than 4 million and 700 thousand cases were oficially notified with about 316 thousand deaths. Etiological agent of the disease was identified as being a new coronavirus, Severe acute respiratory syndrome-related coronavirus (SARS-CoV-2). In this study we compared four different manual methods for RNA isolation and purification for detection of SARS-CoV-2 through qRT-PCR, as well as the extraction quality itself through detection of RNAse P. Magnetic beads-based (MagMax) and silica column-based (Biopur(R)) methods presented the better performances. Concerning to the mean delay in CT values when compared to MagMax, TRIzol, Biopur(R) and EasyExtract presented 0,39, 0,95 and 5,23 respectively. Agreement between positive and negative results of different methods when compared with the one with better performance MagMax was 94,44% for silica column-based method (Biopur(R)), 88,89% for phenol-chroloform-based method (TRIzol) and 77,78% for EasyExtract. We aimed to evaluate how reliable each method is for diagnostic purposes and to propose alternatives when usual methods are not available. In this regard, magnectic beads and silica column-based methods are convenient and reliable choices and phenol-chloroform-based method could also be chosen as an alternative.