Enhancing SARS-CoV-2 Lineage Surveillance through the Integration of a Simple and Direct qPCR-Based Protocol Adaptation with Established Machine Learning Algorithms.

Emerging and evolving Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) lineages, adapted to changing epidemiological conditions, present unprecedented challenges to global public health systems. Here, we introduce an adapted analytical approach that complements genomic sequencing, applying a cost-effective quantitative polymerase chain reaction (qPCR)-based assay. Viral RNA samples from SARS-CoV-2 positive cases detected by diagnostic laboratories or public health network units in Ceará, Brazil, were tracked for genomic surveillance and analyzed by using paired-end sequencing combined with integrative genomic analysis. Validation of a key structural variation was conducted with gel electrophoresis for the presence of a specific open reading frame 7a(ORF7a) gene deletion within the "BE.9" lineages tracked. The analytical innovation of our method is the optimization of a simple intercalating dye-based qPCR assay through repositioning primers from the ARTIC v4.1 amplicon panel to detect large molecular patterns. This assay distinguishes between "BE.9" and "non-BE.9" lineages, particularly BQ.1, without the need for expensive probes or sequencing. The protocol was validated against lineage predictions from next-generation sequencing (NGS) using 525 paired samples, achieving 93.3% sensitivity, 95.1% specificity, and 92.4% agreement, as measured by Cohen's Kappa coefficient. Machine learning (ML) models were trained using the melting curves from intercalating dye-based qPCR of 1724 samples, enabling highly accurate lineage assignment. Among them, the support vector machine (SVM) model had the best performance and after fine-tuning showed ∼96.52% (333/345) accuracy in comparison to the test data set. Our integrated approach provides an adapted analytical method that is both cost-effective and scalable, suitable for rapid assessment of emerging variants, especially in resource-limited settings. In this work, the protocol is applied to improve the monitoring of SARS-CoV-2 sublineages but can be extended to track any key molecular signature, including large insertions and deletions (indels) commonly observed in pathogenic agent subtypes. By offering a complement to traditional sequencing methods and utilizing easily trainable machine learning algorithms, our methodology contributes to enhanced molecular surveillance strategies and supports global efforts in pandemic control.

Predictive biomarkers of mortality in patients with severe COVID-19 hospitalized in intensive care unit.

Objectives: This study was performed to identify predictive markers of worse outcomes in patients with severe COVID-19 in an intensive care unit. Methods: Sixty patients with severe COVID-19, hospitalized in the Intensive Care Unit (ICU) between March and July 2021, were stratified into two groups according to the outcome survivors and non-survivors. After admission to the ICU, blood samples were collected directly for biomarker analysis. Routine hematological and biochemical biomarkers, as well as serum levels of cytokines, chemokines, and immunoglobulins, were investigated. Results: Lymphopenia, neutrophilia, and thrombocytopenia were more pronounced in non-surviving patients, while the levels of CRP, AST, creatinine, ferritin, AST, troponin I, urea, magnesium, and potassium were higher in the non-surviving group than the survival group. In addition, serum levels of IL-10, CCL2, CXCL9, and CXCL10 were significantly increased in patients who did not survive. These changes in the biomarkers evaluated were associated with increased mortality in patients with severe COVID-19. Conclusion: The present study confirmed and expanded the validity of laboratory biomarkers as indicators of mortality in severe COVID-19.

Gut Microbiota Composition Correlates with Disease Severity in Myelodysplastic Syndrome.

The myelodysplastic syndrome (MDS) is a heterogeneous group of clonal disorders of hematopoietic progenitor cells related to ineffective hematopoiesis and an increased risk of transformation to acute myelogenous leukemia. MDS is divided into categories, namely lineage dysplasia (MDS-SLD), MDS with ring sideroblasts (MDS-RS), MDS with multilineage dysplasia (MDS-MLD), MDS with excess blasts (MDS-EB). The International Prognostic Classification System (IPSS) ranks the patients as very low, low, intermediate, high, and very high based on disease evolution and survival rates. Evidence points to toll-like receptor (TLR) abnormal signaling as an underlying mechanism of this disease, providing a link between MDS and immune dysfunction. Microbial signals, such as lipopolysaccharides from gram-negative bacteria, can activate or suppress TLRs. Therefore, we hypothesized that MDS patients present gut microbiota alterations associated with disease subtypes and prognosis. To test this hypothesis, we sequenced the 16S rRNA gene from fecal samples of 30 MDS patients and 16 healthy elderly controls. We observed a negative correlation between Prevotella spp. and Akkermansia spp. in MDS patients compared with the control group. High-risk patients presented a significant increase in the genus Prevotella spp. compared to the other risk categories. There was a significant reduction in the abundance of the genus Akkermansia spp. in high-risk patients compared with low- and intermediate-risk. There was a significant decrease in the genus Ruminococcus spp. in MDS-EB patients compared with controls. Our findings show a new association between gut dysbiosis and higher-risk MDS, with a predominance of gram-negative bacteria.

Spatiotemporal transmission of SARS-CoV-2 lineages during 2020-2021 in Pernambuco-Brazil.

SARS-CoV-2 virus emerged as a new threat to humans and spread around the world, leaving a large death toll. As of January 2023, Brazil is among the countries with the highest number of registered deaths. Nonpharmacological and pharmacological interventions have been heterogeneously implemented in the country, which, associated with large socioeconomic differences between the country regions, has led to distinct virus spread dynamics. Here, we investigate the spatiotemporal dispersion of SARS-CoV-2 lineages in the Pernambuco state (Northeast Brazil) throughout the distinct epidemiological scenarios that unfolded in the first 2 years of the pandemic. We generated a total of 1,389 new SARS-CoV-2 genomes from June 2020 to August 2021. This sampling captured the arrival, communitary transmission, and the circulation of the B1.1, B.1.1.28, and B.1.1.33 lineages; the emergence of the former variant of interest P.2; and the emergence and fast replacement of all previous variants by the more transmissible variant of concern P.1 (Gamma). Based on the incidence and lineage spread pattern, we observed an East-to-West to inner state pattern of transmission, which is in agreement with the transmission of more populous metropolitan areas to medium- and small-size country-side cities in the state. Such transmission patterns may be partially explained by the main routes of traffic across municipalities in the state. Our results highlight that the fine-grained intrastate analysis of lineages and incidence spread can provide actionable insights for planning future nonpharmacological intervention for air-borne transmissible human pathogens.IMPORTANCEDuring the COVID-19 pandemic, Brazil was one of the most affected countries, mainly due its continental-size, socioeconomic differences among regions, and heterogeneous implementation of intervention methods. In order to investigate SARS-CoV-2 dynamics in the state of Pernambuco, we conducted a spatiotemporal dispersion study, covering the period from June 2020 to August 2021, to comprehend the dynamics of viral transmission during the first 2 years of the pandemic. Throughout this study, we were able to track three significant epidemiological waves of transmission caused by B1.1, B.1.1.28, B.1.1.33, P.2, and P.1 lineages. These analyses provided valuable insights into the evolution of the epidemiological landscape, contributing to a deeper understanding of the dynamics of virus transmission during the early years of the pandemic in the state of Pernambuco.

Spatiotemporal dynamics and epidemiological impact of SARS-CoV-2 XBB lineage dissemination in Brazil in 2023.

The SARS-CoV-2 XBB is a group of highly immune-evasive lineages of the Omicron variant of concern that emerged by recombining BA.2-descendent lineages and spread worldwide during 2023. In this study, we combine SARS-CoV-2 genomic data (n = 11,065 sequences) with epidemiological data of severe acute respiratory infection (SARI) cases collected in Brazil between October 2022 and July 2023 to reconstruct the space-time dynamics and epidemiologic impact of XBB dissemination in the country. Our analyses revealed that the introduction and local emergence of lineages carrying convergent mutations within the Spike protein, especially F486P, F456L, and L455F, propelled the spread of XBB* lineages in Brazil. The average relative instantaneous reproduction numbers of XBB* + F486P, XBB* + F486P + F456L, and XBB* + F486P + F456L + L455F lineages in Brazil were estimated to be 1.24, 1.33, and 1.48 higher than that of other co-circulating lineages (mainly BQ.1*/BE*), respectively. Despite such a growth advantage, the dissemination of these XBB* lineages had a reduced impact on Brazil's epidemiological scenario concerning previous Omicron subvariants. The peak number of SARI cases from SARS-CoV-2 during the XBB wave was approximately 90%, 80%, and 70% lower than that observed during the previous BA.1*, BA.5*, and BQ.1* waves, respectively. These findings revealed the emergence of multiple XBB lineages with progressively increasing growth advantage, yet with relatively limited epidemiological impact in Brazil throughout 2023. The XBB* + F486P + F456L + L455F lineages stand out for their heightened transmissibility, warranting close monitoring in the months ahead. IMPORTANCE: Brazil was one the most affected countries by the SARS-CoV-2 pandemic, with more than 700,000 deaths by mid-2023. This study reconstructs the dissemination of the virus in the country in the first half of 2023, a period characterized by the dissemination of descendants of XBB.1, a recombinant of Omicron BA.2 lineages evolved in late 2022. The analysis supports that XBB dissemination was marked by the continuous emergence of indigenous lineages bearing similar mutations in key sites of their Spike protein, a process followed by continuous increments in transmissibility, and without repercussions in the incidence of severe cases. Thus, the results suggest that the epidemiological impact of the spread of a SARS-CoV-2 variant is influenced by an intricate interplay of factors that extend beyond the virus's transmissibility alone. The study also underlines the need for SARS-CoV-2 genomic surveillance that allows the monitoring of its ever-shifting composition.

Expanding the phenotypic spectrum of CLCN2-related leucoencephalopathy and ataxia.

Mutations in CLCN2 are a rare cause of autosomal recessive leucoencephalopathy with ataxia and specific imaging abnormalities. Very few cases have been reported to date. Here, we describe the clinical and imaging phenotype of 12 additional CLCN2 patients and expand the known phenotypic spectrum of this disorder. Informed consent was obtained for all patients. Patients underwent either whole-exome sequencing or focused/panel-based sequencing to identify variants. Twelve patients with biallelic CLCN2 variants are described. This includes three novel likely pathogenic missense variants. All patients demonstrated typical MRI changes, including hyperintensity on T2-weighted images in the posterior limbs of the internal capsules, midbrain cerebral peduncles, middle cerebellar peduncles and cerebral white matter. Clinical features included a variable combination of ataxia, headache, spasticity, seizures and other symptoms with a broad range of age of onset. This report is now the largest case series of patients with CLCN2-related leucoencephalopathy and reinforces the finding that, although the imaging appearance is uniform, the phenotypic expression of this disorder is highly heterogeneous. Our findings expand the phenotypic spectrum of CLCN2-related leucoencephalopathy by adding prominent seizures, severe spastic paraplegia and developmental delay.

Evolutionary deletions within the SARS-CoV-2 genome as signature trends for virus fitness and adaptation.

Coronaviruses are large RNA viruses that can infect and spread among humans and animals. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), responsible for coronavirus disease 2019, has evolved since its first detection in December 2019. Deletions are a common occurrence in SARS-CoV-2 evolution, particularly in specific genomic sites, and may be associated with the emergence of highly competent lineages. While deletions typically have a negative impact on viral fitness, some persist and become fixed in viral populations, indicating that they may confer advantageous benefits for the virus's adaptive evolution. This work presents a literature review and data analysis on structural losses in the SARS-CoV-2 genome and the potential relevance of specific signatures for enhanced viral fitness and spread.

An assessment of the public health surveillance strategy based on molecular testing during three major pandemic waves of COVID-19 in Brazil.

A national laboratory-based surveillance system was adapted to monitor the situation of SARS-CoV-2 in Brazil. The objective of the study was to compare the challenges in implementing COVID-19 surveillance strategies based on the Ministry of Health's (MoH) distribution of RT-PCR tests to different types of laboratories. This retrospective study analyzed the MoH's testing policies and distribution of RT-PCR tests to laboratories during the first, second, and third waves. Recipient laboratories were divided into groups: public health laboratories that belonged to the national network of public health laboratories (Group 1); public laboratories granted authorization during the pandemic (Group 2); and High-Capacity Testing Centers (Group 3). We analyzed the timing and duration of COVID-19 testing policies and the allocation of tests to laboratories by group and wave. Using t-tests, we analyzed the difference in the weekly average of tests distributed to labs by group and using Pearson's correlation coefficient, analyzed the test distribution according to infection and death rates. Between epiweek 9, 2020, and epiweek 22, 2022, the MoH distributed an average of 263,004 RT-PCR tests per week. The weekly average of tests distributed was highest in the second wave (310,327 tests), followed by the first (218,005 tests) and third waves (201,226 tests). There was a significant increase in the mean weekly tests distributed in the second wave compared to the first and third waves (p = 0.047; IC 8.29-1110.71). We found a significant difference between the weekly average of tests distributed in the first and second wave (p < 0.001; IC -209.83-76.20) to Group 2. Group 3 received the second-highest number of tests from the MoH overall, with a reduction during the third wave to first-wave levels. The distribution of RT-PCR tests was not correlated with the case and death incidence.

Persistent SARS-COV-2 infection in vaccinated individual with three doses of COVID-19 vaccine.

We describe a case of a 24-year-old Brazilian woman previously vaccinated with CoronaVac and a booster dose of Pfizer-BioNTech, with mild-to-moderate COVID-19, with persistent viral shedding. We evaluated viral load, antibody dynamics for SARS-CoV-2 and performed genomic analysis to identify the viral variant. The female remained positive for 40 days following symptom onset (cycle quantification mean: 32.54 ± 2.29). The humoral response was characterized by absence of IgM for the viral spike protein, increased IgG for the viral spike (1800.60 to 19558.60 AU/mL) and for the nucleocapsid (from 0.03 to 8.9 index value) proteins, and high titers of neutralizing antibodies (>488.00 IU/mL). The variant identified was the sublineage BA. 5.1. of Omicron (B.1.1.529). Our results suggest that even though the female produced an antibody response against SARS-CoV-2, the persistent infection can be explained by antibody decline and/or the immune evasion by the Omicron variant, illustrating the need to revaccinate or update vaccines.

Molecular characterization of a new SARS-CoV-2 recombinant cluster XAG identified in Brazil.

Recombination events have been described in the Coronaviridae family. Since the beginning of the SARS-CoV-2 pandemic, a variable degree of selection pressure has acted upon the virus, generating new strains with increased fitness in terms of viral transmission and antibody scape. Most of the SC2 variants of concern (VOC) detected so far carry a combination of key amino acid changes and indels. Recombination may also reshuffle existing genetic profiles of distinct strains, potentially giving origin to recombinant strains with altered phenotypes. However, co-infection and recombination events are challenging to detect and require in-depth curation of assembled genomes and sequencing reds. Here, we present the molecular characterization of a new SARS-CoV-2 recombinant between BA.1.1 and BA.2.23 Omicron lineages identified in Brazil. We characterized four mutations that had not been previously described in any of the recombinants already identified worldwide and described the likely breaking points. Moreover, through phylogenetic analysis, we showed that the newly named XAG lineage groups in a highly supported monophyletic clade confirmed its common evolutionary history from parental Omicron lineages and other recombinants already described. These observations were only possible thanks to the joint effort of bioinformatics tools auxiliary in genomic surveillance and the manual curation of experienced personnel, demonstrating the importance of genetic, and bioinformatic knowledge in genomics.

SARS-CoV-2 antibody dynamics in blood donors and COVID-19 epidemiology in eight Brazilian state capitals: A serial cross-sectional study.

Background: The COVID-19 situation in Brazil is complex due to large differences in the shape and size of regional epidemics. Understanding these patterns is crucial to understand future outbreaks of SARS-CoV-2 or other respiratory pathogens in the country. Methods: We tested 97,950 blood donation samples for IgG antibodies from March 2020 to March 2021 in 8 of Brazil's most populous cities. Residential postal codes were used to obtain representative samples. Weekly age- and sex-specific seroprevalence were estimated by correcting the crude seroprevalence by test sensitivity, specificity, and antibody waning. Results: The inferred attack rate of SARS-CoV-2 in December 2020, before the Gamma variant of concern (VOC) was dominant, ranged from 19.3% (95% credible interval [CrI] 17.5-21.2%) in Curitiba to 75.0% (95% CrI 70.8-80.3%) in Manaus. Seroprevalence was consistently smaller in women and donors older than 55 years. The age-specific infection fatality rate (IFR) differed between cities and consistently increased with age. The infection hospitalisation rate increased significantly during the Gamma-dominated second wave in Manaus, suggesting increased morbidity of the Gamma VOC compared to previous variants circulating in Manaus. The higher disease penetrance associated with the health system's collapse increased the overall IFR by a minimum factor of 2.91 (95% CrI 2.43-3.53). Conclusions: These results highlight the utility of blood donor serosurveillance to track epidemic maturity and demonstrate demographic and spatial heterogeneity in SARS-CoV-2 spread. Funding: This work was supported by Itaú Unibanco 'Todos pela Saude' program; FAPESP (grants 18/14389-0, 2019/21585-0); Wellcome Trust and Royal Society Sir Henry Dale Fellowship 204311/Z/16/Z; the Gates Foundation (INV- 034540 and INV-034652); REDS-IV-P (grant HHSN268201100007I); the UK Medical Research Council (MR/S0195/1, MR/V038109/1); CAPES; CNPq (304714/2018-6); Fundação Faculdade de Medicina; Programa Inova Fiocruz-CE/Funcap - Edital 01/2020 Number: FIO-0167-00065.01.00/20 SPU N°06531047/2020; JBS - Fazer o bem faz bem.

Predicting SARS-CoV-2 Variant Spread in a Completely Seropositive Population Using Semi-Quantitative Antibody Measurements in Blood Donors.

SARS-CoV-2 serologic surveys estimate the proportion of the population with antibodies against historical variants, which nears 100% in many settings. New approaches are required to fully exploit serosurvey data. Using a SARS-CoV-2 anti-Spike (S) protein chemiluminescent microparticle assay, we attained a semi-quantitative measurement of population IgG titers in serial cross-sectional monthly samples of blood donations across seven Brazilian state capitals (March 2021−November 2021). Using an ecological analysis, we assessed the contributions of prior attack rate and vaccination to antibody titer. We compared anti-S titer across the seven cities during the growth phase of the Delta variant and used this to predict the resulting age-standardized incidence of severe COVID-19 cases. We tested ~780 samples per month, per location. Seroprevalence rose to >95% across all seven capitals by November 2021. Driven by vaccination, mean antibody titer increased 16-fold over the study, with the greatest increases occurring in cities with the highest prior attack rates. Mean anti-S IgG was strongly correlated (adjusted R2 = 0.89) with the number of severe cases caused by Delta. Semi-quantitative anti-S antibody titers are informative about prior exposure and vaccination coverage and may also indicate the potential impact of future SARS-CoV-2 variants.

Temporal trends and spatial clusters of gastric cancer mortality in Brazil.

Objective: To identify nationwide temporal trends and spatial patterns of gastric cancer-related mortality in Brazil. Methods: An ecological study was performed using death certificates registered from 2000 to 2019 in which gastric cancer was recorded as any cause of death (an underlying or associated cause). Trends over time were assessed using joinpoint regression models. Spatial and spatiotemporal clusters were identified by Kulldorff's space-time scan statistics to identify high-risk areas. Results: In 276 897/22 663 091 (1.22%) death certificates gastric cancer was recorded as any cause of death. Age-adjusted gastric cancer-related mortality increased significantly over time (annual percentage change [APC]: 0.7, 95% confidence interval [CI]: 0.5 to 0.8). The increase in mortality was more pronounced in the less-developed North and Northeast Regions (North Region, APC: 3.1, 95% CI: 2.7 to 3.5; Northeast Region, APC: 3.1, 95% CI: 2.5 to 3.7). Eight spatiotemporally associated high-risk clusters of gastric cancer-related mortality were identified in the North, South, Northeast and Central-West Regions, as well as a major cluster covering a wide geographical range in the South and Southeast Regions of Brazil during the first years of the study period (2000 to 2009). Conclusions: More recently, during 2010 to 2019, clusters of gastric cancer have been identified in the Northeast Region. The nationwide increase in mortality in this analysis of 20 years of data highlights the persistently high burden of gastric cancer in Brazil, especially in socioeconomically disadvantaged regions. The identification of these areas where the population is at high risk for gastric cancer-related mortality emphasizes the need to develop effective and intersectoral control measures.

Emergence and Spread of the SARS-CoV-2 Variant of Concern Delta across Different Brazilian Regions.

The SARS-CoV-2 variant of concern (VOC) Delta was first detected in India in October 2020. The first imported cases of the Delta variant in Brazil were identified in April 2021 in the southern region, followed by more cases in different regions during the following months. By early September 2021, Delta was already the dominant variant in the southeastern (87%), southern (73%), and northeastern (52%) Brazilian regions. This study aimed to understand the spatiotemporal dissemination dynamics of Delta in Brazil. To this end, we employed a combination of maximum likelihood (ML) and Bayesian methods to reconstruct the evolutionary relationship of 2,264 VOC Delta complete genomes (482 from this study) recovered across 21 of the 27 Brazilian federal units. Our phylogeographic analyses identified three major transmission clusters of Delta in Brazil. The clade BR-I (n = 1,560) arose in Rio de Janeiro in late April 2021 and was the major cluster behind the dissemination of the VOC Delta in the southeastern, northeastern, northern, and central-western regions. The AY.101 lineage (n = 207) that arose in the Paraná state in late April 2021 and aggregated the largest fraction of sampled genomes from the southern region. Lastly, the AY.46.3 lineage emerged in Brazil in the São Paulo state in early June 2021 and remained mostly restricted to this state. In the rapid turnover of viral variants characteristic of the SARS-CoV-2 pandemic, Brazilian regions seem to occupy different stages of an increasing prevalence of the VOC Delta in their epidemic profiles. This process demands continuous genomic and epidemiological surveillance toward identifying and mitigating new introductions, limiting their dissemination, and preventing the establishment of more significant outbreaks in a population already heavily affected by the COVID-19 pandemic. IMPORTANCE Amid the SARS-CoV-2 continuously changing epidemic profile, this study details the space-time dynamics of the emergence of the Delta lineage across Brazilian territories, pointing out its multiple introductions in the country and its most prevalent sublineages. Some of these sublineages have their emergence, alongside their genomic composition and geographic distribution, detailed here for the first time. A special focus is given to the emergence process of Delta outside the country's south and southeast regions, the most populated and subjects of most published SARS-CoV-2 studies in Brazil. In summary, the study allows a better comprehension of the evolution process of a SARS-CoV-2 lineage that would be associated with a significant recrudescence of the pandemic in Brazil.

Phylogenetic-based inference reveals distinct transmission dynamics of SARS-CoV-2 lineages Gamma and P.2 in Brazil.

The COVID-19 epidemic in Brazil experienced two major lineage replacements until mid-2021. The first was driven by lineage P.2, in late 2020, and the second by lineage Gamma, in early 2021. To understand how these SARS-CoV-2 lineages spread in Brazil, we analyzed 11,724 genomes collected throughout the country between September 2020 and April 2021. Our findings indicate that lineage P.2 probably emerged in July 2020 in the Rio de Janeiro state and Gamma in November 2020 in the Amazonas state. Both states were the main hubs of viral disseminations to other Brazilian locations. We estimate that Gamma was 1.56-3.06 times more transmissible than P.2 in Rio de Janeiro and that the median effective reproductive number (Re) of Gamma varied according to the geographic context (Re = 1.59-3.55). In summary, our findings support that lineage Gamma was more transmissible and spread faster than P.2 in Brazil.

Unusual SARS-CoV-2 intrahost diversity reveals lineage superinfection.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has infected almost 200 million people worldwide by July 2021 and the pandemic has been characterized by infection waves of viral lineages showing distinct fitness profiles. The simultaneous infection of a single individual by two distinct SARS-CoV-2 lineages may impact COVID-19 disease progression and provides a window of opportunity for viral recombination and the emergence of new lineages with differential phenotype. Several hundred SARS-CoV-2 lineages are currently well phylogenetically defined, but two main factors have precluded major coinfection/codetection and recombination analysis thus far: (i) the low diversity of SARS-CoV-2 lineages during the first year of the pandemic, which limited the identification of lineage defining mutations necessary to distinguish coinfecting/recombining viral lineages; and the (ii) limited availability of raw sequencing data where abundance and distribution of intrasample/intrahost variability can be accessed. Here, we assembled a large sequencing dataset from Brazilian samples covering a period of 18 May 2020 to 30 April 2021 and probed it for unexpected patterns of high intrasample/intrahost variability. This approach enabled us to detect nine cases of SARS-CoV-2 coinfection with well characterized lineage-defining mutations, representing 0.61 % of all samples investigated. In addition, we matched these SARS-CoV-2 coinfections with spatio-temporal epidemiological data confirming its plausibility with the cocirculating lineages at the timeframe investigated. Our data suggests that coinfection with distinct SARS-CoV-2 lineages is a rare phenomenon, although it is certainly a lower bound estimate considering the difficulty to detect coinfections with very similar SARS-CoV-2 lineages and the low number of samples sequenced from the total number of infections.

COVID-19-associated meningoencephalitis in a Brazilian patient: case report and literature review.

The global pandemic of COVID-19 is caused by the novel coronavirus SARS-CoV-2, which often causes flu-like symptoms and can progress to severe respiratory illness. Thus, as the disease spreads, COVID-19 cases have multiplied across the world, and manifestations involving multiple systems have been described. We report a case of COVID-19-associated meningoencephalitis in a Brazilian male patient who presented with seizures and altered mental status. To the best of our knowledge, this is the first reported case of COVID-19-associated meningoencephalitis in Brazil. COVID-19-associated meningoencephalitis is a rare manifestation of this viral infection and clinicians should be aware of this possible complication.

Spread of Gamma (P.1) Sub-Lineages Carrying Spike Mutations Close to the Furin Cleavage Site and Deletions in the N-Terminal Domain Drives Ongoing Transmission of SARS-CoV-2 in Amazonas, Brazil.

The Amazonas was one of the most heavily affected Brazilian states by the COVID-19 epidemic. Despite a large number of infected people, particularly during the second wave associated with the spread of the Variant of Concern (VOC) Gamma (lineage P.1), SARS-CoV-2 continues to circulate in the Amazonas. To understand how SARS-CoV-2 persisted in a human population with a high immunity barrier, we generated 1,188 SARS-CoV-2 whole-genome sequences from individuals diagnosed in the Amazonas state from 1st January to 6th July 2021, of which 38 were vaccine breakthrough infections. Our study reveals a sharp increase in the relative prevalence of Gamma plus (P.1+) variants, designated Pango Lineages P.1.3 to P.1.6, harboring two types of additional Spike changes: deletions in the N-terminal (NTD) domain (particularly Δ144 or Δ141-144) associated with resistance to anti-NTD neutralizing antibodies or mutations at the S1/S2 junction (N679K or P681H) that probably enhance the binding affinity to the furin cleavage site, as suggested by our molecular dynamics simulations. As lineages P.1.4 (S:N679K) and P.1.6 (S:P681H) expanded (Re > 1) from March to July 2021, the lineage P.1 declined (Re < 1) and the median Ct value of SARS-CoV-2 positive cases in Amazonas significantly decreases. Still, we did not find an increased incidence of P.1+ variants among breakthrough cases of fully vaccinated patients (71%) in comparison to unvaccinated individuals (93%). This evidence supports that the ongoing endemic transmission of SARS-CoV-2 in the Amazonas is driven by the spread of new local Gamma/P.1 sublineages that are more transmissible, although not more efficient to evade vaccine-elicited immunity than the parental VOC. Finally, as SARS-CoV-2 continues to spread in human populations with a declining density of susceptible hosts, the risk of selecting more infectious variants or antibody evasion mutations is expected to increase. IMPORTANCE The continuous evolution of SARS-CoV-2 is an expected phenomenon that will continue to happen due to the high number of cases worldwide. The present study analyzed how a Variant of Concern (VOC) could still circulate in a population hardly affected by two COVID-19 waves and with vaccination in progress. Our results showed that the answer behind that was a new generation of Gamma-like viruses, which emerged locally carrying mutations that made it more transmissible and more capable of spreading, partially evading prior immunity triggered by natural infections or vaccines. With thousands of new cases daily, the current pandemics scenario suggests that SARS-CoV-2 will continue to evolve and efforts to reduce the number of infected subjects, including global equitable access to COVID-19 vaccines, are mandatory. Thus, until the end of pandemics, the SARS-CoV-2 genomic surveillance will be an essential tool to better understand the drivers of the viral evolutionary process.

Temporal trends and spatial clusters of gastric cancer mortality in Brazil / Tendencias temporales y conglomerados espaciales de mortalidad por cáncer gástrico en Brasil / Tendências temporais e aglomerados espaciais de mortalidade por câncer gástrico no Brasil

ABSTRACT Objective. To identify nationwide temporal trends and spatial patterns of gastric cancer-related mortality in Brazil. Methods. An ecological study was performed using death certificates registered from 2000 to 2019 in which gastric cancer was recorded as any cause of death (an underlying or associated cause). Trends over time were assessed using joinpoint regression models. Spatial and spatiotemporal clusters were identified by Kulldorff's space-time scan statistics to identify high-risk areas. Results. In 276 897/22 663 091 (1.22%) death certificates gastric cancer was recorded as any cause of death. Age-adjusted gastric cancer-related mortality increased significantly over time (annual percentage change [APC]: 0.7, 95% confidence interval [CI]: 0.5 to 0.8). The increase in mortality was more pronounced in the less-developed North and Northeast Regions (North Region, APC: 3.1, 95% CI: 2.7 to 3.5; Northeast Region, APC: 3.1, 95% CI: 2.5 to 3.7). Eight spatiotemporally associated high-risk clusters of gastric cancer-related mortality were identified in the North, South, Northeast and Central-West Regions, as well as a major cluster covering a wide geographical range in the South and Southeast Regions of Brazil during the first years of the study period (2000 to 2009). Conclusions. More recently, during 2010 to 2019, clusters of gastric cancer have been identified in the Northeast Region. The nationwide increase in mortality in this analysis of 20 years of data highlights the persistently high burden of gastric cancer in Brazil, especially in socioeconomically disadvantaged regions. The identification of these areas where the population is at high risk for gastric cancer-related mortality emphasizes the need to develop effective and intersectoral control measures.

RESUMEN Objetivo. Identificar las tendencias temporales y los patrones espaciales de la mortalidad relacionada con el cáncer gástrico a nivel nacional en Brasil. Métodos. Se realizó un estudio ecológico, empleando certificados de defunción registrados entre los años 2000 y 2019 en los que se notificó cáncer gástrico como cualquier causa de muerte (subyacente o asociada). Se evaluaron las tendencias con el transcurso del tiempo mediante modelos de regresión de punto de inflexión (joinpoint). Se identificaron los conglomerados espaciales y espaciotemporales mediante la técnica estadística de exploración espaciotemporal de Kulldorff para determinar cuáles eran las áreas de alto riesgo. Resultados. En 276 897 de 22 663 091 certificados de defunción (1,22%), se registró cáncer gástrico como cualquier causa de muerte. La mortalidad relacionada con el cáncer gástrico ajustada por edad aumentó significativamente con el tiempo (cambio porcentual anual: 0,7; intervalo de confianza [IC] del 95%: 0,5 a 0,8). El aumento de la mortalidad fue más acusado en la regiones Norte y Noreste, menos desarrolladas, (región Norte, cambio porcentual anual: 3,1, IC del 95%: 2,7 a 3,5; región Noreste, cambio porcentual anual: 3,1, IC del 95%: 2,5 a 3,7). Durante los primeros años del período de estudio (del 2000 al 2009), se identificaron ocho conglomerados de alto riesgo de mortalidad relacionada con el cáncer gástrico y con asociación espacial y temporal en las regiones Norte, Sur, Noreste y Centro-Oeste, así como un conglomerado importante que cubría un amplio rango geográfico en las regiones Sur y Sureste de Brasil. Conclusiones. Más recientemente, del 2010 al 2019, se han identificado conglomerados de cáncer gástrico en la región noreste. El aumento nacional de la mortalidad en este análisis de veinte años de datos destaca la carga persistentemente alta del cáncer gástrico en Brasil, especialmente en las regiones socioeconómicamente desfavorecidas. La identificación de estas áreas en que la población presenta un alto riesgo de mortalidad relacionada con el cáncer gástrico subraya la necesidad de elaborar medidas de control intersectoriales y efectivas.

RESUMO Objetivo. Identificar tendências temporais e padrões espaciais de mortalidade relacionada ao câncer gástrico em todo o Brasil. Métodos. Realizou-se um estudo ecológico a partir de declarações de óbito registradas de 2000 a 2019 em que o câncer gástrico foi indicado como qualquer causa de morte (causa básica ou associada). As tendências ao longo do tempo foram avaliadas a partir de modelos de regressão por pontos de inflexão (joinpoint). Os aglomerados espaciais e espaço-temporais foram identificados por estatística de varredura espaço-temporal de Kulldorff para detectar áreas de alto risco. Resultados. O câncer gástrico foi registrado como qualquer causa de morte em 276.897/22.663.091 (1,22%) declarações de óbito. A mortalidade relacionada ao câncer gástrico ajustada por idade aumentou significativamente ao longo do tempo [variação percentual anual (VPA): 0,7, intervalo de confiança (IC) de 95%: 0,5 a 0,8]. O aumento da mortalidade foi mais acentuado no Norte e Nordeste, regiões menos desenvolvidas (região Norte, VPA: 3,1, IC 95%: 2,7 a 3,5; região Nordeste, VPA: 3,1, IC 95%: 2,5 a 3,7). Identificaram-se oito aglomerados de alto risco de mortalidade relacionada ao câncer gástrico em associação espaço-temporal nas regiões Norte, Sul, Nordeste e Centro-Oeste, além de um grande aglomerado que abrangia uma larga faixa geográfica nas regiões Sul e Sudeste do Brasil durante os primeiros anos do período de estudo (2000 a 2009). Conclusões. Mais recentemente, no período de 2010 a 2019, identificaram-se aglomerados de câncer gástrico na região Nordeste. O aumento da mortalidade em todo o país nesta análise de dados relativos a 20 anos evidencia a persistência da alta carga de câncer gástrico no Brasil, sobretudo em regiões desfavorecidas do ponto de vista socioeconômico. A identificação dessas áreas em que a população corre alto risco de morte relacionada ao câncer gástrico enfatiza a necessidade de desenvolver medidas de controle efetivas e intersetoriais.

COVID-19-associated meningoencephalitis in a Brazilian patient: case report and literature review

ABSTRACT The global pandemic of COVID-19 is caused by the novel coronavirus SARS-CoV-2, which often causes flu-like symptoms and can progress to severe respiratory illness. Thus, as the disease spreads, COVID-19 cases have multiplied across the world, and manifestations involving multiple systems have been described. We report a case of COVID-19-associated meningoencephalitis in a Brazilian male patient who presented with seizures and altered mental status. To the best of our knowledge, this is the first reported case of COVID-19-associated meningoencephalitis in Brazil. COVID-19-associated meningoencephalitis is a rare manifestation of this viral infection and clinicians should be aware of this possible complication.