Phylogeny-aware linear B-cell epitope predictor detects targets associated with immune response to orthopoxviruses.

We introduce a phylogeny-aware framework for predicting linear B-cell epitope (LBCE)-containing regions within proteins. Our approach leverages evolutionary information by using a taxonomic scaffold to build models trained on hierarchically structured data. The resulting models present performance equivalent or superior to generalist methods, despite using simpler features and a fraction of the data volume required by current state-of-the-art predictors. This allows the utilization of available data for major pathogen lineages to facilitate the prediction of LBCEs for emerging infectious agents. We demonstrate the efficacy of our approach by predicting new LBCEs in the monkeypox (MPXV) and vaccinia viruses. Experimental validation of selected targets using sera from infected patients confirms the presence of LBCEs, including candidates for the differential serodiagnosis of recent MPXV infections. These results point to the use of phylogeny-aware predictors as a useful strategy to facilitate the targeted development of immunodiagnostic tools.

Differential diagnosis of exanthematous viruses during the 2022 Mpox outbreak in Minas Gerais, Brazil.

The monkeypox virus (MPXV) outbreak, primarily endemic to Africa, has spread globally, with Brazil reporting the second-highest number of cases. The emergence of MPXV in non-endemic areas has raised concerns, particularly due to the co-circulation of other exanthematous viruses such as varicella-zoster virus (VZV) and molluscum contagiosum virus (MOCV). To perform an accurate differential diagnosis of MPXV during the ongoing outbreak in Minas Gerais, Brazil, a 5PLEX qPCR assay targeting orthopoxviruses (OPV), VZV, and MOCV was used to retrospectively analyze all clinical samples that tested negative for MPXV in the initial screening conducted at Funed. In summary, our study analyzed 1,175 clinical samples received from patients suspected of MPXV infection and found a positivity rate of 33.8% (397 samples) for MPXV using the non-variola qPCR assay. Testing the 778 MPXV-negative clinical samples using the 5PLEX qPCR assay revealed that 174 clinical samples (22.36%) tested positive for VZV. MOCV DNA was detected in 13 and other OPV in 3 clinical samples. The sequencing of randomly selected amplified clinical samples confirmed the initial molecular diagnosis. Analysis of patient profiles revealed a significant difference in the median age between groups testing positive for MPXV and VZV and a male predominance in MPXV cases. The geographic distribution of positive cases was concentrated in the most populous mesoregions of Minas Gerais state. This study highlights the challenges posed by emerging infectious diseases. It emphasizes the importance of epidemiological surveillance and accurate diagnosis in enabling timely responses for public health policies and appropriate medical care. IMPORTANCE: Brazil ranks second in the number of cases during the global monkeypox epidemic. The study, conducted in Minas Gerais, the second most populous state in Brazil with over 20 million inhabitants, utilized differential diagnostics, revealing a significant number of positive cases for other exanthematous viruses and emphasizing the need for accurate diagnoses. During the study, we were able to assess the co-circulation of other viruses alongside monkeypox, including varicella-zoster virus, molluscum contagiosum virus, and other orthopoxviruses. The significance of the research is underscored by the concentration of positive cases in populous areas, highlighting the challenges posed by emerging infectious diseases. This demographic context further amplifies the importance of the research in guiding public health policies and medical interventions, given the substantial population at risk. The study not only addresses a global concern but also holds critical implications for a state with such a large population and geographic expanse within Brazil. Overall, the study emphasizes the pivotal role of surveillance and precise diagnosis in guiding effective public health responses and ensuring appropriate medical interventions.

Dengue diagnostics: serious inaccuracies are likely to occur if pre-analytical conditions are not strictly followed.

BACKGROUND: The heat-labile nature of Dengue virus (DENV) in serum samples must be considered when applying routine diagnostic tests to avoid issues that could impact the accuracy of test results with direct implications for case management and disease reporting. OBJECTIVES: To check if pre-analytical variables, such as storage time and temperature, have an impact on the accuracy of the main routine diagnostic tests for dengue. METHODS: Virus isolation, reverse transcription real-time polymerase chain reaction (RT-PCR) and NS1 enzyme-linked immunosorbent assay (ELISA) were evaluated using 84 samples submitted to different pre-analytical conditions. FINDINGS: Sensitivity and negative predictive value were directly affected by sample storage conditions. RT-PCR and virus isolation showed greater dependence on well-conserved samples for an accurate diagnosis. Interestingly, even storage at -30ºC for a relatively short time (15 days) was not adequate for accurate results using virus isolation and RT-PCR tests. On the other hand, NS1 ELISA showed no significant reduction in positivity for aliquots tested under the same conditions as in the previous tests. MAIN CONCLUSIONS: Our results support the stability of the NS1 marker in ELISA diagnosis and indicate that the accuracy of routine tests such as virus isolation and RT-PCR is significantly affected by inadequate transport and storage conditions of serum samples.

Needlestick-Associated Rocky Mountain Spotted Fever, Brazil.

We report a fatal case of Rocky Mountain spotted fever (RMSF) in a man in Brazil without recent history of tick bites or environmental exposure. He received an accidental needlestick while working as a nurse. The nurse and his patient died. Both cases were confirmed as RMSF by molecular methods.

Metagenomic Analysis for Diagnosis of Hemorrhagic Fever in Minas Gerais, Brazil.

Viral hemorrhagic fever poses a significant public health challenge due to its severe clinical presentation and high mortality rate. The diagnostic process is hindered by similarity of symptoms across different diseases and the broad spectrum of pathogens that can cause hemorrhagic fever. In this study, we applied viral metagenomic analysis to 43 serum samples collected by the Public Health Laboratory (Fundação Ezequiel Dias, FUNED) in Minas Gerais State, Brazil, from patients diagnosed with hemorrhagic fever who had tested negative for the standard local hemorrhagic disease testing panel. This panel includes tests for Dengue virus (DENV) IgM, Zika virus IgM, Chikungunya virus IgM, yellow fever IgM, Hantavirus IgM, Rickettsia rickettsii IgM/IgG, and Leptospira interrogans IgM, in addition to respective molecular tests for these infectious agents. The samples were grouped into 18 pools according to geographic origin and analyzed through next-generation sequencing on the NextSeq 2000 platform. Bioinformatic analysis revealed a prevalent occurrence of commensal viruses across all pools, but, notably, a significant number of reads corresponding to the DENV serotype 2 were identified in one specific pool. Further verification via real-time PCR confirmed the presence of DENV-2 RNA in an index case involving an oncology patient with hemorrhagic fever who had initially tested negative for anti-DENV IgM antibodies, thereby excluding this sample from initial molecular testing. The complete DENV-2 genome isolated from this patient was taxonomically classified within the cosmopolitan genotype that was recently introduced into Brazil. These findings highlight the critical role of considering the patient's clinical condition when deciding upon the most appropriate testing procedures. Additionally, this study showcases the potential of viral metagenomics in pinpointing the viral agents behind hemorrhagic diseases. Future research is needed to assess the practicality of incorporating metagenomics into standard viral diagnostic protocols.

Exploring Dengue Infection in a Vaccinated Individual: Preliminary Molecular Diagnosis and Sequencing Insights.

This study examines a case involving a 7-year-old child who developed dengue symptoms following Qdenga vaccination. Despite initial negative diagnostic results, molecular analysis confirmed an infection with DENV4. Next-generation sequencing detected viral RNA from both DENV2 and DENV4 serotypes, which were identified as vaccine-derived strains using specific primers. Phylogenetic analysis further confirmed that these sequences belonged to the Qdenga vaccine rather than circulating wild-type viruses. This case underscores the critical need for precise diagnostic interpretation in vaccinated individuals to avoid misdiagnosis and to strengthen public health surveillance. A comprehensive understanding of vaccine-induced viremia is essential for refining dengue surveillance, improving diagnostic accuracy, and informing public health strategies in endemic regions.

Retrospective Analysis of Omicron in Minas Gerais, Brazil: Emergence, Dissemination, and Diversification.

Brazil is one of the countries most affected by COVID-19, with the highest number of deaths recorded. Brazilian Health Institutions have reported four main peaks of positive COVID-19 cases. The last two waves were characterized by the emergence of the VOC Omicron and its sublineages. This study aimed to conduct a retrospective surveillance study illustrating the emergence, dissemination, and diversification of the VOC Omicron in 15 regional health units (RHUs) in MG, the second most populous state in Brazil, by combining epidemiological and genomic data. A total of 5643 confirmed positive COVID-19 samples were genotyped using the panels TaqMan SARS-CoV-2 Mutation and 4Plex SC2/VOC Bio-Manguinhos to define mutations classifying the BA.1, BA.2, BA.4, and BA.5 sublineages. While sublineages BA.1 and BA.2 were more prevalent during the third wave, BA.4 and BA.5 dominated the fourth wave in the state. Epidemiological and viral genome data suggest that age and vaccination with booster doses were the main factors related to clinical outcomes, reducing the number of deaths, irrespective of the Omicron sublineages. Complete genome sequencing of 253 positive samples confirmed the circulation of the BA.1, BA.2, BA.4, and BA.5 subvariants, and phylogenomic analysis demonstrated that the VOC Omicron was introduced through multiple international events, followed by transmission within the state of MG. In addition to the four subvariants, other lineages have been identified at low frequency, including BQ.1.1 and XAG. This integrative study reinforces that the evolution of Omicron sublineages was the most significant factor driving the highest peaks of positive COVID-19 cases without an increase in more severe cases, prevented by vaccination boosters.

Potential use of high-resolution melting analyses for SARS-CoV-2 genomic surveillance.

The pandemic caused by COVID-19 and the emergence of new variants of SARS-CoV-2 have generated clinical and epidemiological impacts on a global scale. The use of strategies for monitoring viral circulation and identifying mutations in genomic regions involved in host interaction are important measures to mitigate viral dissemination and reduce its likely complications on population health. In this context, the objective of this work was to explore the potential of high-resolution melting (HRM) analysis combined with one-step real-time reverse transcription PCR in a closed-tube system, as a fast and convenient method of screening for SARS-CoV-2 mutations with possible implications on host-pathogen interactions. The HRM analyses allowed the distinction of the Gamma, Zeta, Alpha, Delta, and Omicron variants against the predecessors (B.1.1.28, B.1.1.33) of occurrence in Brazil. It is concluded that the molecular tool standardized here has the potential to optimize the genomic surveillance of SARS-CoV-2, and could be adapted for genomic surveillance of other pathogens, due to its ability to detect, prior to sequencing, samples suggestive of new variants, selecting them more assertively and earlier for whole genome sequencing when compared to random screening.

Genomic epidemiology unveils the dynamics and spatial corridor behind the Yellow Fever virus outbreak in Southern Brazil.

Despite the considerable morbidity and mortality of yellow fever virus (YFV) infections in Brazil, our understanding of disease outbreaks is hampered by limited viral genomic data. Here, through a combination of phylogenetic and epidemiological models, we reconstructed the recent transmission history of YFV within different epidemic seasons in Brazil. A suitability index based on the highly domesticated Aedes aegypti was able to capture the seasonality of reported human infections. Spatial modeling revealed spatial hotspots with both past reporting and low vaccination coverage, which coincided with many of the largest urban centers in the Southeast. Phylodynamic analysis unraveled the circulation of three distinct lineages and provided proof of the directionality of a known spatial corridor that connects the endemic North with the extra-Amazonian basin. This study illustrates that genomics linked with eco-epidemiology can provide new insights into the landscape of YFV transmission, augmenting traditional approaches to infectious disease surveillance and control.

Seroprevalence estimate and risk factors for Coxiella burnetii infections among humans in a highly urbanised Brazilian state.

BACKGROUND: Q fever is among the top 13 global priority zoonoses, however, it is still neglected and under-reported in most of the world, including Brazil. Thus, we evaluated the seroprevalence of and the risk factors for Coxiella burnetii infections in humans from Minas Gerais, a highly urbanised Brazilian state. METHODS: Coxiella burnetii was searched for patient samples (n=437), which were suspected of then later confirmed as negative for dengue fever, by the indirect immunofluorescence technique and real-time PCR. Risk factors for infections and spatial clusters for both C. burnetii-seropositive individuals and livestock concentration were evaluated. RESULTS: We found that 21 samples (4.8%; 95% CI 3.0 to 7.2%) were reactive for at least one class of anti-C. burnetii antibodies (titer of ≥64), with rural residence (p=0.036) being a risk factor. Also, two spatial clusters of seropositivity were found within a significant area by Scan, and a probable relationship between the Scan result and the livestock concentration by area was found. CONCLUSIONS: Seropositive individuals were associated with rural residence, with a likely relationship with the livestock concentration. Thus, this study establishes baseline figures for C. burnetii seroprevalence in humans in a state of Brazil, allowing the monitoring of trends and setting of control targets, as well as more representative longitudinal and risk analysis studies.

A Retrospective Overview of Zika Virus Evolution in the Midwest of Brazil.

Since the introduction of the Zika virus (ZIKV) into Brazil in 2015, its transmission dynamics have been intensively studied in many parts of the country, although much is still unknown about its circulation in the midwestern states. Here, using nanopore technology, we obtained 23 novel partial and near-complete ZIKV genomes from the state of Goiás, located in the Midwest of Brazil. Genomic, phylogenetic, and epidemiological approaches were used to retrospectively explore the spatiotemporal evolution of the ZIKV-Asian genotype in this region. As a likely consequence of a gradual accumulation of herd immunity, epidemiological data revealed a decline in the number of reported cases over 2018 to 2021. Phylogenetic reconstructions revealed that multiple independent introductions of the Asian lineage have occurred in Goiás over time and revealed a complex transmission dynamic between epidemic seasons. Together, our results highlight the utility of genomic, epidemiological, and evolutionary methods to understand mosquito-borne epidemics. IMPORTANCE Despite the considerable morbidity and mortality of arboviral infections in Brazil, such as Zika, chikungunya, dengue fever, and yellow fever, our understanding of these outbreaks is hampered by the limited availability of genomic data to track and control the epidemic. In this study, we provide a retrospective reconstruction of the Zika virus transmission dynamics in the state of Goiás by analyzing genomic data from areas in Midwest Brazil not covered by other previous studies. Our study provides an understanding of how ZIKV initiates transmission in this region and reveals a complex transmission dynamic between epidemic seasons. Together, our results highlight the utility of genomic, epidemiological, and evolutionary methods to understand mosquito-borne epidemics, revealing how this toolkit can be used to help policymakers prioritize areas to be targeted, especially in the context of finite public health resources.

Retrospective Genomic Surveillance of Chikungunya Transmission in Minas Gerais State, Southeast Brazil.

Brazil accounted for a total number of 1,276,194 reported cases of chikungunya fever between 2014 and 2022. Additionally, since 2015, the country has experienced an increasing death toll, in which the Northeast and Southeast regions appear to report the worst scenarios. Although the CHIKV transmission dynamics have been studied in many parts of the country since its introduction in 2014, little is still known about chikungunya virus (CHIKV) transmission and genetic diversity in the state of Minas Gerais, located in southeast Brazil. Moreover, no studies have been published characterizing CHIKV genomic surveillance in this state. Thus, to retrospectively explore the CHIKV epidemic in Minas Gerais, we generated 40 genomes from clinical samples using Nanopore sequencing. Phylogenetic analysis indicated that multiple introductions of CHIKV occurred, likely from the northeastern Brazilian states, with the most recent common ancestral strain dating to early March 2016, which is in agreement with local epidemiological reports. Additionally, epidemiological data reveals a decline in the number of reported cases from 2017 to 2021, indicating that population immunity or changes in vector activity may have contributed to the decreasing waves of CHIKV infection. Together, our results shed light on the dispersion dynamics of CHIKV and show that infections decreased from March 2017 to January 2021 despite multiple introductions into Minas Gerais State. In conclusion, our study highlights the importance of combining genomic and epidemiological data in order to assist public health laboratories in monitoring and understanding the patterns and diversity of mosquito-borne viral epidemics. IMPORTANCE Arbovirus infections in Brazil, including chikungunya, dengue, yellow fever, and Zika, result in considerable morbidity and mortality and are pressing public health concerns. However, our understanding of these outbreaks is hampered by the limited availability of genomic data. In this study, we combine epidemiological analysis and portable genome sequencing to retrospectively describe the CHIKV epidemic in Minas Gerais between 2017 and 2021. Our results indicate that the East/Central/South African (ECSA) CHIKV lineage was introduced into Minas Gerais by three distinct events, likely from the North and Northeast regions of Brazil. Our study provides an understanding of how CHIKV initiates transmission in the region and illustrates that genomics in the field can augment traditional approaches to infectious disease surveillance and control.

Genomic epidemiology of the SARS-CoV-2 epidemic in Brazil.

The high numbers of COVID-19 cases and deaths in Brazil have made Latin America an epicentre of the pandemic. SARS-CoV-2 established sustained transmission in Brazil early in the pandemic, but important gaps remain in our understanding of virus transmission dynamics at a national scale. We use 17,135 near-complete genomes sampled from 27 Brazilian states and bordering country Paraguay. From March to November 2020, we detected co-circulation of multiple viral lineages that were linked to multiple importations (predominantly from Europe). After November 2020, we detected large, local transmission clusters within the country. In the absence of effective restriction measures, the epidemic progressed, and in January 2021 there was emergence and onward spread, both within and abroad, of variants of concern and variants under monitoring, including Gamma (P.1) and Zeta (P.2). We also characterized a genomic overview of the epidemic in Paraguay and detected evidence of importation of SARS-CoV-2 ancestor lineages and variants of concern from Brazil. Our findings show that genomic surveillance in Brazil enabled assessment of the real-time spread of emerging SARS-CoV-2 variants.

Monitoring the Establishment of VOC Gamma in Minas Gerais, Brazil: A Retrospective Epidemiological and Genomic Surveillance Study.

Since its first identification in Brazil, the variant of concern (VOC) Gamma has been associated with increased infection and transmission rates, hospitalizations, and deaths. Minas Gerais (MG), the second-largest populated Brazilian state with more than 20 million inhabitants, observed a peak of cases and deaths in March-April 2021. We conducted a surveillance study in 1240 COVID-19-positive samples from 305 municipalities distributed across MG's 28 Regional Health Units (RHU) between 1 March to 27 April 2021. The most common variant was the VOC Gamma (71.2%), followed by the variant of interest (VOI) zeta (12.4%) and VOC alpha (9.6%). Although the predominance of Gamma was found in most of the RHUs, clusters of Zeta and Alpha variants were observed. One Alpha-clustered RHU has a history of high human mobility from countries with Alpha predominance. Other less frequent lineages, such as P.4, P.5, and P.7, were also identified. With our genomic characterization approach, we estimated the introduction of Gamma on 7 January 2021, at RHU Belo Horizonte. Differences in mortality between the Zeta, Gamma and Alpha variants were not observed. We reinforce the importance of vaccination programs to prevent severe cases and deaths during transmission peaks.

Genomic epidemiology reveals the impact of national and international restrictions measures on the SARS-CoV-2 epidemic in Brazil.

Brazil has experienced some of the highest numbers of COVID-19 cases and deaths globally and from May 2021 made Latin America a pandemic epicenter. Although SARS-CoV-2 established sustained transmission in Brazil early in the pandemic, important gaps remain in our understanding of virus transmission dynamics at the national scale. Here, we describe the genomic epidemiology of SARS-CoV-2 using near-full genomes sampled from 27 Brazilian states and a bordering country - Paraguay. We show that the early stage of the pandemic in Brazil was characterised by the co-circulation of multiple viral lineages, linked to multiple importations predominantly from Europe, and subsequently characterized by large local transmission clusters. As the epidemic progressed under an absence of effective restriction measures, there was a local emergence and onward international spread of Variants of Concern (VOC) and Variants Under Monitoring (VUM), including Gamma (P.1) and Zeta (P.2). In addition, we provide a preliminary genomic overview of the epidemic in Paraguay, showing evidence of importation from Brazil. These data reinforce the usefulness and need for the implementation of widespread genomic surveillance in South America as a toolkit for pandemic monitoring that provides a means to follow the real-time spread of emerging SARS-CoV-2 variants with possible implications for public health and immunization strategies.

CCL3, CCL5, IL-15, IL-1Ra and VEGF compose a reliable algorithm to discriminate classes of adverse events following 17DD-YF primary vaccination according to cause-specific definitions.

In the present study, a range of serum biomarkers were quantified in suspected cases of adverse events following YF immunization (YEL-AEFI) to propose a reliable laboratorial algorithm to discriminate confirmed YEL-AEFI ("A1" class) from cases with other illnesses ("C" class). Our findings demonstrated that increased levels of CXCL8, CCL2, CXCL10, IL-1ß, IL-6 and TNF-α were observed in YEL-AEFI ("A1" and "C" classes) as compared to primary vaccines without YEL-AEFI [PV(day 3-28)] and reference range (RR) controls. Notably, increased levels of CCL3, CCL4, CCL2, CCL5, IL-1ß, IL-15, IL-1Ra and G-CSF were found in "A1" as compared to "C" class. Venn diagrams analysis allowed the pre-selection of biomarkers for further analysis of performance indices. Data demonstrated that CCL3, CCL5, IL-15 and IL-1Ra presented high global accuracy (AUC = 1.00) to discriminate "A1" from "C". Decision tree was proposed with a reliable algorithm to discriminate YEL-AEFI cases according to cause-specific definitions with outstanding overall accuracy (91%). CCL3, CCL5, IL-15 and IL-1Ra appears as root attributes to identify "A1" followed by VEGF as branch nodes to discriminate Wild Type YFV infection ("C(WT-YFV)") from cases with other illnesses ("C*"). Together, these results demonstrated the applicability of serum biomarker measurements as putative parameters towards the establishment of accurate laboratorial tools for complementary differential diagnosis of YEL-AEFI cases.

Field and classroom initiatives for portable sequence-based monitoring of dengue virus in Brazil.

Brazil experienced a large dengue virus (DENV) epidemic in 2019, highlighting a continuous struggle with effective control and public health preparedness. Using Oxford Nanopore sequencing, we led field and classroom initiatives for the monitoring of DENV in Brazil, generating 227 novel genome sequences of DENV1-2 from 85 municipalities (2015-2019). This equated to an over 50% increase in the number of DENV genomes from Brazil available in public databases. Using both phylogenetic and epidemiological models we retrospectively reconstructed the recent transmission history of DENV1-2. Phylogenetic analysis revealed complex patterns of transmission, with both lineage co-circulation and replacement. We identified two lineages within the DENV2 BR-4 clade, for which we estimated the effective reproduction number and pattern of seasonality. Overall, the surveillance outputs and training initiative described here serve as a proof-of-concept for the utility of real-time portable sequencing for research and local capacity building in the genomic surveillance of emerging viruses.

Pluripotency of Wolbachia against Arboviruses: the case of yellow fever.

Background: Yellow fever outbreaks have re-emerged in Brazil during 2016-18, with mortality rates up to 30%. Although urban transmission has not been reported since 1942, the risk of re-urbanization of yellow fever is significant, as Aedes aegypti is present in most tropical and sub-tropical cities in the World and still remains the main vector of urban YFV. Although the YFV vaccine is safe and effective, it does not always reach populations at greatest risk of infection and there is an acknowledged global shortage of vaccine supply. The introgression of Wolbachia bacteria into Ae. aegypti mosquito populations is being trialed in several countries ( www.worldmosquito.org) as a biocontrol method against dengue, Zika and chikungunya. Here, we studied the ability of Wolbachia to reduce the transmission potential of Ae. aegypti mosquitoes for Yellow fever virus (YFV). Methods: Two recently isolated YFV (primate and human) were used to challenge field-derived wild-type and Wolbachia-infected ( wMel +) Ae. aegypti mosquitoes. The YFV infection status was followed for 7, 14 and 21 days post-oral feeding (dpf). The YFV transmission potential of mosquitoes was evaluated via nano-injection of saliva into uninfected mosquitoes or by inoculation in mice. Results: We found that Wolbachia was able to significantly reduce the prevalence of mosquitoes with YFV infected heads and thoraces for both viral isolates. Furthermore, analyses of mosquito saliva, through indirect injection into naïve mosquitoes or via interferon-deficient mouse model, indicated Wolbachia was associated with profound reduction in the YFV transmission potential of mosquitoes (14dpf). Conclusions: Our results suggest that Wolbachia introgression could be used as a complementary strategy for prevention of urban yellow fever transmission, along with the human vaccination program.

Persistence of Yellow fever virus outside the Amazon Basin, causing epidemics in Southeast Brazil, from 2016 to 2018.

BACKGROUND: Yellow fever (YF) is endemic in the Brazilian Amazon Basin, and sporadic outbreaks take place outside the endemic area in Brazil. Since 2016, YF epidemics have been occurring in Southeast Brazil, with more than 1,900 human cases and more than 1,600 epizooties of non-human primates (NHPs) reported until April 2018. Previous studies have demonstrated that Yellow fever virus (YFV) causing outbreaks in 2017 formed a monophyletic group. METHODOLOGY/PRINCIPAL FINDINGS: Aiming to decipher the origin of the YFV responsible for the recent epidemics, we obtained nucleotide sequences of YFV detected in humans (n = 6) and NHPs (n = 10) from Minas Gerais state during 2017-2018. Next, we performed evolutionary analyses and discussed the results in the light of epidemiological records (official numbers of YFV cases at each Brazilian Federative unit, reported by the Brazilian Ministry of Health). Nucleotide sequences of YFV from Southeast Brazil from 2016 to 2018 were highly conserved and formed a monophyletic lineage (BR-YFV_2016/18) within the genotype South America I. Different clusters were observed within lineage BR-YFV_2016/18, one containing the majority of isolates (from humans and NHPs), indicating the sylvatic transmission of YFV. We also detected a cluster characterized by two synapomorphies (amino acid substitutions) that contained YFV only associated with NHP what should be further investigated. The topology of lineage BR-YFV_2016/18 was congruent with epidemiological and temporal patterns of the ongoing epidemic. YFV isolates detected in 2016, in São Paulo state were located in the most basal position of the lineage, followed by the isolates from Minas Gerais and Espírito Santo obtained in 2017 and 2018. The most recent common ancestor of the lineage BR-YFV_2016/18 dated to 2015 (95% credible intervals = 2014-2016), in a period that was coincident with the reemergence of YFV in the Midwest region of Brazil. CONCLUSIONS: The results demonstrated a single introduction of YFV in the Southeast region and the silent viral circulation before the onset of the outbreaks in 2016. Evolutionary analyses combined with epidemiological records supported the idea that BR-YFV_2016/18 was probably introduced from the Midwest into the Southeast region, possibly in São Paulo state. The persistence of YFV in the Southeast region, causing epidemics from 2016 to 2018, suggests that this region presents suitable ecological and climatic conditions for YFV maintenance during the epidemic and interepidemic seasons. This fact poses risks for the establishing of YF enzootic cycles and epidemics, outside the Amazon Basin in Brazil. YF surveillance and studies of viral dynamics deserve particular attention, especially in Midwest, Southeast and neighbor regions which are the main areas historically associated with YF outbreaks outside the Amazon Basin. YFV persistence in Southeast Brazil should be carefully considered in the context of public health, especially for public health decision-makers and researchers.

Genomic epidemiology of the SARS-CoV-2 epidemic in Brazil

The high numbers of COVID-19 cases and deaths in Brazil have made Latin America an epicentre of the pandemic. SARS-CoV-2 established sustained transmission in Brazil early in the pandemic, but important gaps remain in our understanding of virus transmission dynamics at a national scale. We use 17,135 near-complete genomes sampled from 27 Brazilian states and bordering country Paraguay. From March to November 2020, we detected co-circulation of multiple viral lineages that were linked to multiple importations (predominantly from Europe). After November 2020, we detected large, local transmission clusters within the country. In the absence of effective restriction measures, the epidemic progressed, and in January 2021 there was emergence and onward spread, both within and abroad, of variants of concern and variants under monitoring, including Gamma (P.1) and Zeta (P.2). We also characterized a genomic overview of the epidemic in Paraguay and detected evidence of importation of SARS-CoV-2 ancestor lineages and variants of concern from Brazil. Our findings show that genomic surveillance in Brazil enabled assessment of the real-time spread of emerging SARS-CoV-2 variants.