In silico analysis of non-structural protein 12 sequences from SARS-COV-2 found in Manaus, Amazonas, Brazil, reveals mutations linked to higher transmissibility.

The disease coronavirus COVID-19 has been the cause of millions of deaths worldwide. Among the proteins of SARS-CoV-2, non-structural protein 12 (NSP12) plays a key role during COVID infection and is part of the RNA-dependent RNA polymerase complex. The monitoring of NSP12 polymorphisms is extremely important for the design of new antiviral drugs and monitoring of viral evolution. This study analyzed the NSP12 mutations detected in circulating SARS-CoV-2 during the years 2020 to 2022 in the population of the city of Manaus, Amazonas, Brazil. The most frequent mutations found were P323L and G671S. Reports in the literature indicate that these mutations are related to transmissibility efficiency, which may have contributed to the extremely high numbers of cases in this location. In addition, two mutations described here (E796D and R914K) are close and have RMSD that is similar to the mutations M794V and N911K, which have been described in the literature as influential on the performance of the NSP12 enzyme. These data demonstrate the need to monitor the emergence of new mutations in NSP12 in order to better understand their consequences for the treatments currently used and in the design of new drugs.

The emergence of Omicron VOC and its rapid spread and persistence in the Western Amazon.

Genomic surveillance represents a strategy to understanding the evolutionary mechanisms, transmission, and infectivity of different SARS-CoV-2 variants. We evaluated 603 individuals positive for SARS-CoV-2 from 34 municipalities of Rondônia between December 2021 to December 2022. Nasopharyngeal samples were collected, RNA was extracted and screened using RT-qPCR for VOCs. RNA of the samples were sequenced and further analyzed for phylogeny, mutations, and lineages, totaling 96.19% of samples positive for Omicron VOC in this cohort. We observed that most individuals had at least two doses, however 18.97% were not vaccinated with any dose. 554 sequences were amenable to analysis for alignment and phylogenetic characterization; this group corresponded to the 27 subvariants of the Omicron VOC; a total of 100 mutations were identified, 48% of which were found in the S gene. In conclusion, the data demonstrated the rapid spread and persistence of Omicron VOC in Rondônia during the 12-month study period. Although high frequency of mutations was found in the analyzed samples, there were no individuals with a severe clinical profile, demonstrating that vaccination had a positive effect in those cases.

Case Report: SARS-CoV-2 Gamma Isolation From Placenta of a Miscarriage in Midwest, Brazil.

The present study investigated a SARS-CoV-2 infection in placenta and fetal samples from an early pregnancy miscarriage in Midwest Brazil. The Gamma variant was isolated and fully sequenced from the placenta sample, but not from fetal samples. Our findings highlight potential adverse perinatal outcomes caused by SARS-CoV-2 Gamma infection during pregnancy.

Active surveillance and early detection of community transmission of SARS-CoV-2 Mu variant (B.1.621) in the Brazilian Amazon.

Through active surveillance and contact tracing from outpatients, we aimed to identify and characterize SARS-CoV-2 variants circulating in Porto Velho-Rondônia, a city in the Brazilian Amazon. As part of a prospective cohort, we gathered information from 2,506 individuals among COVID-19 patients and household contacts. Epidemiological data, nasopharyngeal swabs, and blood samples were collected from all participants. Nasopharyngeal swabs were tested for antigen rapid diagnostic test and reverse transcription-polymerase chain reaction (RT-PCR) followed by genomic sequencing. Blood samples underwent ELISA testing for IgA, IgG, and IgM antibody levels. From 757 specimens sequenced, three were identified as Mu variant, none of the individuals carrying this variant had a travel history in the previous 15 days before diagnosis. One case was asymptomatic and two presented mild symptoms. Two infected individuals from different households caring viruses with additional amino acid substitutions ORF7a P45L and ORF1a T1055A compared to the Mu virus reference sequence. One patient presented IgG levels. Our results highlight that genomic surveillance for SARS-CoV-2 variants can assist in detecting the emergency of SARS-CoV-2 variants in the community, before its identification in other parts of the country.

Using amino acids co-occurrence matrices and explainability model to investigate patterns in dengue virus proteins.

BACKGROUND: Dengue is a common vector-borne disease in tropical countries caused by the Dengue virus. This virus may trigger a disease with several symptoms like fever, headache, nausea, vomiting, and muscle pain. Indeed, dengue illness may also present more severe and life-threatening conditions like hemorrhagic fever and dengue shock syndrome. The causes that lead hosts to develop severe infections are multifactorial and not fully understood. However, it is hypothesized that different viral genome signatures may partially contribute to the disease outcome. Therefore, it is plausible to suggest that deeper DENV genetic information analysis may bring new clues about genetic markers linked to severe illness. METHOD: Pattern recognition in very long protein sequences is a challenge. To overcome this difficulty, we map protein chains onto matrix data structures that reveal patterns and allow us to classify dengue proteins associated with severe illness outcomes in human hosts. Our analysis uses co-occurrence of amino acids to build the matrices and Random Forests to classify them. We then interpret the classification model using SHAP Values to identify which amino acid co-occurrences increase the likelihood of severe outcomes. RESULTS: We trained ten binary classifiers, one for each dengue virus protein sequence. We assessed the classifier performance through five metrics: PR-AUC, ROC-AUC, F1-score, Precision and Recall. The highest score on all metrics corresponds to the protein E with a 95% confidence interval. We also compared the means of the classification metrics using the Tukey HSD statistical test. In four of five metrics, protein E was statistically different from proteins M, NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5, showing that E markers has a greater chance to be associated with severe dengue. Furthermore, the amino acid co-occurrence matrix highlight pairs of amino acids within Domain 1 of E protein that may be associated with the classification result. CONCLUSION: We show the co-occurrence patterns of amino acids present in the protein sequences that most correlate with severe dengue. This evidence, used by the classification model and verified by statistical tests, mainly associates the E protein with the severe outcome of dengue in human hosts. In addition, we present information suggesting that patterns associated with such severe cases can be found mostly in Domain 1, inside protein E. Altogether, our results may aid in developing new treatments and being the target of debate on new theories regarding the infection caused by dengue in human hosts.

The ongoing evolution of variants of concern and interest of SARS-CoV-2 in Brazil revealed by convergent indels in the amino (N)-terminal domain of the spike protein.

Mutations at both the receptor-binding domain (RBD) and the amino (N)-terminal domain (NTD) of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Spike (S) glycoprotein can alter its antigenicity and promote immune escape. We identified that SARS-CoV-2 lineages circulating in Brazil with mutations of concern in the RBD independently acquired convergent deletions and insertions in the NTD of the S protein, which altered the NTD antigenic-supersite and other predicted epitopes at this region. Importantly, we detected the community transmission of different P.1 lineages bearing NTD indels ∆69-70 (which can impact several SARS-CoV-2 diagnostic protocols), ∆144 and ins214ANRN, and a new VOI N.10 derived from the B.1.1.33 lineage carrying three NTD deletions (∆141-144, ∆211, and ∆256-258). These findings support that the ongoing widespread transmission of SARS-CoV-2 in Brazil generates new viral lineages that might be more resistant to antibody neutralization than parental variants of concern.

A Potential SARS-CoV-2 Variant of Interest (VOI) Harboring Mutation E484K in the Spike Protein Was Identified within Lineage B.1.1.33 Circulating in Brazil.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) epidemic in Brazil was dominated by two lineages designated as B.1.1.28 and B.1.1.33. The two SARS-CoV-2 variants harboring mutations at the receptor-binding domain of the Spike (S) protein, designated as lineages P.1 and P.2, evolved from lineage B.1.1.28 and are rapidly spreading in Brazil. Lineage P.1 is considered a Variant of Concern (VOC) because of the presence of multiple mutations in the S protein (including K417T, E484K, N501Y), while lineage P.2 only harbors mutation S:E484K and is considered a Variant of Interest (VOI). On the other hand, epidemiologically relevant B.1.1.33 deriving lineages have not been described so far. Here we report the identification of a new SARS-CoV-2 VOI within lineage B.1.1.33 that also harbors mutation S:E484K and was detected in Brazil between November 2020 and February 2021. This VOI displayed four non-synonymous lineage-defining mutations (NSP3:A1711V, NSP6:F36L, S:E484K, and NS7b:E33A) and was designated as lineage N.9. The VOI N.9 probably emerged in August 2020 and has spread across different Brazilian states from the Southeast, South, North, and Northeast regions.

A Potential SARS-CoV-2 Variant of Interest (VOI) Harboring Mutation E484K in the Spike Protein Was Identified within Lineage B 1 1 33 Circulating in Brazil

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) epidemic in Brazil was dominated by two lineages designated as B.1.1.28 and B.1.1.33. The two SARS-CoV-2 variants harboring mutations at the receptor-binding domain of the Spike (S) protein, designated as lineages P.1 and P.2, evolved from lineage B.1.1.28 and are rapidly spreading in Brazil. Lineage P.1 is considered a Variant of Concern (VOC) because of the presence of multiple mutations in the S protein (including K417T, E484K, N501Y), while lineage P.2 only harbors mutation S:E484K and is considered a Variant of Interest (VOI). On the other hand, epidemiologically relevant B.1.1.33 deriving lineages have not been described so far. Here we report the identification of a new SARS-CoV-2 VOI within lineage B.1.1.33 that also harbors mutation S:E484K and was detected in Brazil between November 2020 and February 2021. This VOI displayed four non-synonymous lineage-defining mutations (NSP3:A1711V, NSP6:F36L, S:E484K, and NS7b:E33A) and was designated as lineage N.9. The VOI N.9 probably emerged in August 2020 and has spread across different Brazilian states from the Southeast, South, North, and Northeast regions.

A Single Haplotype of IFNG Correlating With Low Circulating Levels of Interferon-γ Is Associated With Susceptibility to Cutaneous Leishmaniasis Caused by Leishmania guyanensis.

BACKGROUND: Interferon-γ (IFN-γ) plays an important role in the control of Leishmania infection. Blockade of IFN-γ signaling in mice increases lesion size and parasite load. In endemic areas of Leishmaniasis, only a fraction of the population develop the disease. This suggest that host genetics may play a role in this response. We investigated whether single nucleotide polymorphisms (SNPs) in IFNG may be associated with elevated or decrease risk in the development of cutaneous leishmaniasis (CL). METHODS: We assessed 9 SNP and cytosine-adenine (CA) repeats in IFNG by nucleotide sequencing in 647 patients with CL caused by Leishmania guyanensis and 629 controls. Circulating plasma IFN-γ levels were also assayed in 400 patients with CL and 400 controls. RESULTS: The rs2069705TT genotype is associated with elevated risk of developing CL compared with the rs2069705CC genotype (OR, 1.7; 95% CI, 1.3-2.4; P = .0008). There is a 70% chance that this genotype raises the risk of developing CL. In a dominant model, carriers of the rs2069705T allele compared with the rs2069705CC genotype showed a 50% (range, 20-100%) increased risk of developing CL (OR, 1.5; 95% CI, 1.2-2.0; P = .0004). Haplotype analysis showed 1 haplotype (H1) associated with low levels of IFN-γ presented an increased risk of 60% of developing CL (OR, 1.6; 95% CI, 1.3-1.9; P = 5 × 10-5) compared with non-H1. CONCLUSIONS: IFNG variant rs2069705 seems to be a genetic modifier of clinical outcome of Leishmania infection; individuals with the H1 haplotype, associated with low levels of IFN-γ, have a 60% risk of developing CL.

Neisseria meningitidis disease-associated clones in Amazonas State, Brazil.

BACKGROUND: The aim of this study is to describe the molecular epidemiology of Neisseria meningitidis invasive disease before the introduction of serogroup C conjugate vaccine in Amazonas State in 2010. METHODS: Meningococcal disease reported cases were investigated in Amazonas State during the period 2000-2010. N. meningitidis isolates (n = 196) recovered from patients were genotyped by multilocus sequence typing (MLST) and sequencing of porB, porA, fetA, fHbp and penA. Antimicrobial susceptibility was determined using E-test. RESULTS: In the study period, 948 cases were reported; the incidence was 2.8 for the entire state and 4.8 per 100,000 in the capital of Manaus. Most meningococcal disease was caused by N. meningitidis belonging to ST-32 (72%; 141/196) or ST-103 (21%; 41/196) clonal complexes. Capsular switching (B→C) was suggested within clonal complex (cc) 32. There were 6 (3%; 6/196) strains with intermediate susceptibility to penicillin and a single strain was resistant to rifampicin. Since 2007, serogroup C strains belonging to the cc103 have predominated and case-fatality has increased. CONCLUSION: We demonstrate a high rate of meningococcal disease in Amazonas State, where, like other parts of Brazil, serogroup C replaced serogroup B during 2000s. These data serve as a baseline to measure impact of serogroup C conjugate vaccine introduction in 2010. This study emphasizes the need for enhanced surveillance to monitor changes in meningococcal disease trends following the introduction of meningococcal vaccines.

Detection of Herpesvirus, Enterovirus, and Arbovirus infection in patients with suspected central nervous system viral infection in the Western Brazilian Amazon.

Acute infections of the central nervous system (CNS) can be caused by various pathogens. In this study, the presence of herpesviruses (HHV), enteroviruses (EVs), and arboviruses were investigated in CSF samples from 165 patients with suspected CNS viral infection through polymerase chain reaction (PCR) and reverse transcriptase PCR. The genomes of one or more viral agents were detected in 29.7% (49/165) of the CSF samples. EVs were predominant (16/49; 32.6%) followed by Epstein-Barr virus (EBV) (22.4%), Varicella-Zoster virus (VZV) (20.4%), Cytomegalovirus (CMV) (18.4%), herpes simplex virus (HSV-1) (4.1%), (HSV-2) (4.1%), and the arboviruses (14.3%). Four of the arboviruses were of dengue virus (DENV) and three of oropouche virus (OROV). The detection of different viruses in the CNS of patients with meningitis or encephalitis highlight the importance of maintaining an active laboratory monitoring diagnostics with rapid methodology of high sensitivity in areas of viral hyperendemicity that may assist in clinical decisions and in the choice of antiviral therapy.

Clinical profile of concurrent dengue fever and Plasmodium vivax malaria in the Brazilian Amazon: case series of 11 hospitalized patients.

Malaria and dengue fever are the most prevalent vector-borne diseases worldwide. This study aims to describe the clinical profile of patients with molecular diagnosis of concurrent malaria and dengue fever in a tropical-endemic area. Eleven patients with concurrent dengue virus (DENV) and Plasmodium vivax infection are reported. Similar frequencies of DENV-2, DENV-3, and DENV-4 were found, including DENV-3/DENV-4 co-infection. In eight patients, the World Health Organization (WHO) criteria for severe malaria could be fulfilled (jaundice being the most common). Only one patient met severe dengue criteria, but warning signs were present in 10. Syndromic surveillance systems must be ready to identify this condition to avoid misinterpretation of severity attributed to a single disease.

Complete genome sequence of a Dengue virus serotype 4 strain isolated in Roraima, Brazil.

Dengue is the most important arboviral disease worldwide. We report the complete genome sequence of a dengue virus serotype 4, genotype II strain isolated in 2010 from a patient with classical dengue fever in Boa Vista, Roraima, Brazil.

Prevalência da infecção por Chlamydia Trachomatis e fatores associados em diferentes populações de ambos os sexos na cidade de Manaus / Prevalence of Chlamydia Trachomatis infections and associated risk factors in different populations of both gender in Manaus city

Introdução: a infecção por Chlamydia trachomatis (CT) é uma DST de alta prevalência no mundo e quando não diagnosticada pode, principalmente nas mulheres, progredir com seqüelas graves. No Brasil não se conhece com precisão o padrão de comportamento epidemiológico da infecção por CT. Objetivo: estimar prevalência e fatores associados à infecção por Chlamydia trachomatis em segmentos de população sexualmente ativa da cidade de Manaus. Métodos: durante 2004 e 2005 foi realizado estudo de corte seccional com 1732 pessoas (1007 mulheres e 755 homens) divididos em grupos de baixo risco ( 598 gestantes e 600 homens trabalhadores de indústrias) e de maior vulnerabilidade (409 mulheres e 155 homens em clínica de DST). Para o diagnóstico, empregou-se captura híbrida/DIGENE nas mulheres e PCR "Cobas Amplicor CT/NG/Roche" nos homens. Resultados: a prevalência global foi de 7,5%, em mulheres de 11,1% e nos homens de 2,8% (p = 0,000000). As taxas para gestantes e mulheres com DST foram de 11,9% e 10,0% (p = 0,36) e 3,0% e 1,9% para trabalhadores de indústrias e homens com DST (p = 0,65). Prevalências elevadas foram observadas nos adolescentes e nas gestantes (14,8%). Mulheres apresentaram risco de infecção quatro vezes maior do que os homens [ORprev. = 4,38 (IC 95% 2,66 - 7,26); p = 0,000000], a razão de prevalência (RP) foi 4 e a prevalência atribuível para mulheres foi de 8,3. Prevalência em mulheres com mais de um parceiro foi de 16,6% (42/253), superior àquelas que só tiveram um parceiro 9,3% (70/753 p = 0,001). O risco de infectar-se foi o dobro nas com mais de um parceiro [ORPrev.= 1,74 (OR 95% 1,26-2,99); p = 0,002]. Mulheres com parceiro portador de corrimento uretral representaram agravo [OR = 4,4 (IC 95% 2,15-9,21); p = 0,0000104]. A co-infecção com Neisseria gonorrhoeae ocorreu em 17,3%. Conclusão: a prevalência nas mulheres é intermediária (aproximadamente 10%) quando comparada com as taxas publicadas na literatura internacional.

Changing epidemiology of rotavirus-related hospitalizations in rio de janeiro, Brazil, from 2002 to 2006.

A prospective hospital-based sentinel study conducted in Rio de Janeiro identified a shift in the pattern (long to short electropherotype and P(8) to P(4) genotype) of rotavirus strains recovered from children with severe diarrhea a few months after the far-reaching Brazilian rotavirus immunization program was launched, posing new interesting challenges.