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Abstract Background The rapid spread and increase of chikungunya (CHIKV) and dengue (DENV) cases in Brazilian regions in 2023 has raised concerns about the impact of arboviruses on public health. Epidemiological and genomic surveillance was performed to estimate the introduction and spread of CHIKV and DENV in Brazil. Methods This study obtained results from the Hermes Pardini (HP), a private medical laboratory, and the Health Department of Minas Gerais state (SES-MG). We investigated the positivity rates of CHIKV and DENV by analyzing the results of 139,457 samples tested for CHIKV (44,029 in 2022 and 95,428 in 2023) and 491,528 samples tested for DENV (163,674 in 2022 and 327,854 in 2023) across the five representative geographical regions of Brazil. Genome sequencing was performed on 80 CHIKV and 153 DENV samples that had been positive for RT-PCR tests. Results In our sampling, the data from CHIKV tests indicated that the Northeast region had the highest regional positivity rate in 2022 (58.1%). However, in 2023, the Southeast region recorded the highest positivity rate (40.5%). With regard to DENV, the South region exhibited the highest regional positivity rate in both 2022 (40.8%) and 2023 (22.7%), followed by the Southeast region in both years (34.8% in 2022; 21.4% in 2023). During the first 30 epidemiological weeks of 2023 in the state of Minas Gerais (MG), there was a 5.8-fold increase in CHIKV cases and a 3.5-fold increase in DENV compared to the same period in 2022. Analysis of 151 new DENV-1 and 80 CHIKV genomes revealed the presence of three main clusters of CHIKV and circulation of several DENV lineages in MG. All CHIKV clades are closely related to genomes from previous Brazilian outbreaks in the Northeast, suggesting importation events from this region to MG. We detected the RNA of both viruses in approximately 12.75% of the confirmed positive cases, suggesting an increase of co-infection with DENV and CHIKV during the period of analysis. Conclusions These high rates of re-emergence and co-infection with both arboviruses provide useful data for implementing control measures of Aedes vectors and the urgent implementation of public health politics to reduce the numbers of CHIKV and DENV cases in the country.

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.

Since 2014, Brazil has experienced an unprecedented epidemic caused by chikungunya virus (CHIKV), with several waves of East-Central-South-African (ECSA) lineage transmission reported across the country. In 2018, Rio de Janeiro state, the third most populous state in Brazil, reported 41% of all chikungunya cases in the country. Here we use evolutionary and epidemiological analysis to estimate the timescale of CHIKV-ECSA-American lineage and its epidemiological patterns in Rio de Janeiro. We show that the CHIKV-ECSA outbreak in Rio de Janeiro derived from two distinct clades introduced from the Northeast region in mid-2015 (clade RJ1, n = 63/67 genomes from Rio de Janeiro) and mid-2017 (clade RJ2, n = 4/67). We detected evidence for positive selection in non-structural proteins linked with viral replication in the RJ1 clade (clade-defining: nsP4-A481D) and the RJ2 clade (nsP1-D531G). Finally, we estimate the CHIKV-ECSA's basic reproduction number (R0) to be between 1.2 to 1.6 and show that its instantaneous reproduction number (Rt) displays a strong seasonal pattern with peaks in transmission coinciding with periods of high Aedes aegypti transmission potential. Our results highlight the need for continued genomic and epidemiological surveillance of CHIKV in Brazil, particularly during periods of high ecological suitability, and show that selective pressures underline the emergence and evolution of the large urban CHIKV-ECSA outbreak in Rio de Janeiro.

Toxoplasmosis is an important zoonotic disease caused by the parasite Toxoplasma gondii and is especially fatal for neotropical primates. In Brazil, the Ministry of Health is responsible for national epizootic surveillance, but some diseases are still neglected. Here, we present an integrated investigation of an outbreak that occurred during the first year of the COVID-19 pandemic among eleven neotropical primates housed at a primatology center in Brazil. After presenting non-specific clinical signs, all animals died within four days. A wide range of pathogens were evaluated, and we successfully identified T. gondii as the causative agent within four days after necropsies. The liver was the most affected organ, presenting hemorrhage and hepatocellular necrosis. Tachyzoites and bradyzoite cysts were observed in histological examinations and immunohistochemistry in different organs; in addition, parasitic DNA was detected through PCR in blood samples from all specimens evaluated. A high prevalence of Escherichia coli was also observed, indicating sepsis. This case highlights some of the obstacles faced by the current Brazilian surveillance system. A diagnosis was obtained through the integrated action of researchers since investigation for toxoplasmosis is currently absent in national guidelines. An interdisciplinary investigation could be a possible model for future epizootic investigations in animals.

The development of high-throughput sequencing (HTS) technologies and metagenomics protocols deeply impacted the discovery of viral diversity. Moreover, the characterization of novel viruses in the Neotropical primates (NP) is central for the comprehension of viral evolution dynamics in those hosts, due to their evolutionary proximity to Old World primates, including humans. In the present work, novel anelloviruses were detected and characterized through HTS protocols in the NP Callithrix penicillata, the common black-tufted marmoset. De novo assembly of generated sequences was carried out, and a total of 15 contigs were identified with complete Anelloviridae ORF1 gene, two of them including a flanking GC-rich region, confirming the presence of two whole novel genomes of ~3 kb. The identified viruses were monophyletic within the Epsilontorquevirus genus, a lineage harboring previously reported anelloviruses infecting hosts from the Cebidae family. The genetic divergence found in the new viruses characterized two novel species, named Epsilontorquevirus callithrichensis I and II. The phylogenetic pattern inferred for the Epsilontorquevirus genus was consistent with the topology of their host species tree, echoing a virus-host diversification model observed in other viral groups. This study expands the host span of Anelloviridae and provides insights into their diversification dynamics, highlighting the importance of sampling animal viral genomes to obtain a clearer depiction of their long-term evolutionary processes.

Severe acute respiratory syndrome-related coronavirus (SARS-CoV-2) transmission occurs even among fully vaccinated individuals; thus, prompt identification of infected patients is central to control viral circulation. Antigen rapid diagnostic tests (Ag-RDTs) are highly specific, but sensitivity is variable. Discordant RT-qPCR vs. Ag-RDT results are reported, raising the question of whether negative Ag-RDT in positive RT-qPCR samples could imply the absence of infectious viruses. To study the relationship between negative Ag-RDT results with virological, molecular, and serological parameters, we selected a cross-sectional and a follow-up dataset and analyzed virus culture, subgenomic RNA quantification, and sequencing to determine infectious viruses and mutations. We demonstrated that RT-qPCR positive while SARS-CoV-2 Ag-RDT negative discordant results correlate with the absence of infectious virus in nasopharyngeal samples. A decrease in sgRNA detection together with an expected increase in detectable anti-S and anti-N IgGs was also verified in these samples. The data clearly demonstrate that a negative Ag-RDT sample is less likely to harbor infectious SARS-CoV-2 and, consequently, has a lower transmissible potential.

BackgroundThe coronavirus disease 2019 (COVID-19) pandemic had quite an impact on dental health care. Concerns about the risk of SARS-CoV-2 transmission through contaminant fluids and droplet formation during several dental procedures highly impacted dental health care, drastically reducing the number of dental practices worldwide. To monitor SARS-CoV-2 contamination in dental clinics, a longitudinal study was carried out during the return of dental practice at university.MethodsDental health care professionals [(DHCPs); teachers, undergraduate dental students, and dental assistants] and patients were screened for SARS-CoV-2 RNA in a dental school clinic environment from 11th January to 12th March 2021 (9 weeks). Serological testing was performed on DHCPs in two-time points. Additionally, samples with low Ct values were sequenced to identify the circulating SARS-CoV-2 variant and possible transmission clusters.ResultsWe found a low number of dental staff (5.8%), patients (0.9%), and environment sites (0.8%) positive for SARS-CoV-2. Most positive cases had asymptomatic to mild symptoms, and two asymptomatic DHCPs presented prolonged infection. In the first week after previous exposure to COVID-19, 16.2% of DHCPs had IgM or IgG antibodies against SARS-CoV-2, and 1/3 of them had undetected antibodies in the last weeks. The variant zeta (P.2) could be detected. No cross-infection was observed between participants.ConclusionOur study suggests that dental practice can be safely executed when adequate control measures and biosafety protocols are applied. DHCP and patient testing, patient telemonitoring, proper use of personal protection equipment, and sanitization of surfaces are essential to avoid SARS-CoV-2 cross-infection in dental practice.

The COVID-19 pandemic has created an unprecedented need for epidemiological monitoring using diverse strategies. We conducted a project combining prevalence, seroprevalence, and genomic surveillance approaches to describe the initial pandemic stages in Betim City, Brazil. We collected 3239 subjects in a population-based age-, sex- and neighborhood-stratified, household, prospective; cross-sectional study divided into three surveys 21 days apart sampling the same geographical area. In the first survey, overall prevalence (participants positive in serological or molecular tests) reached 0.46% (90% CI 0.12–0.80%), followed by 2.69% (90% CI 1.88–3.49%) in the second survey and 6.67% (90% CI 5.42–7.92%) in the third. The underreporting reached 11, 19.6, and 20.4 times in each survey. We observed increased odds to test positive in females compared to males (OR 1.88 95% CI 1.25–2.82), while the single best predictor for positivity was ageusia/anosmia (OR 8.12, 95% CI 4.72–13.98). Thirty-five SARS-CoV-2 genomes were sequenced, of which 18 were classified as lineage B.1.1.28, while 17 were B.1.1.33. Multiple independent viral introductions were observed. Integration of multiple epidemiological strategies was able to adequately describe COVID-19 dispersion in the city. Presented results have helped local government authorities to guide pandemic management.

BACKGROUND During routine Coronavirus disease 2019 (COVID-19) diagnosis, an unusually high viral load was detected by reverse transcription real-time polymerase chain reaction (RT-qPCR) in a nasopharyngeal swab sample collected from a patient with respiratory and neurological symptoms who rapidly succumbed to the disease. Therefore we sought to characterise the infection. OBJECTIVES We aimed to determine and characterise the etiological agent responsible for the poor outcome. METHODS Classical virological methods, such as plaque assay and plaque reduction neutralisation test combined with amplicon-based sequencing, as well as a viral metagenomic approach, were performed to characterise the etiological agents of the infection. FINDINGS Plaque assay revealed two distinct plaque phenotypes, suggesting either the presence of two severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strains or a productive coinfection of two different species of virus. Amplicon-based sequencing did not support the presence of any SARS-CoV-2 genetic variants that would explain the high viral load and suggested the presence of a single SARS-CoV-2 strain. Nonetheless, the viral metagenomic analysis revealed that Coronaviridae and Herpesviridae were the predominant virus families within the sample. This finding was confirmed by a plaque reduction neutralisation test and PCR. MAIN CONCLUSIONS We characterised a productive coinfection of SARS-CoV-2 and Herpes simplex virus 1 (HSV-1) in a patient with severe symptoms that succumbed to the disease. Although we cannot establish the causal relationship between the coinfection and the severity of the clinical case, this work serves as a warning for future studies focused on the interplay between SARS-CoV-2 and HSV-1 coinfection and COVID-19 severity.

Brazil currently ranks second in absolute deaths by COVID-19, even though most of its population has completed the vaccination protocol. With the introduction of Omicron in late 2021, the number of COVID-19 cases soared once again in the country. We investigated in this work how lineages BA.1 and BA.2 entered and spread in the country by sequencing 2173 new SARS-CoV-2 genomes collected between October 2021 and April 2022 and analyzing them in addition to more than 18,000 publicly available sequences with phylodynamic methods. We registered that Omicron was present in Brazil as early as 16 November 2021 and by January 2022 was already more than 99% of samples. More importantly, we detected that Omicron has been mostly imported through the state of São Paulo, which in turn dispersed the lineages to other states and regions of Brazil. This knowledge can be used to implement more efficient non-pharmaceutical interventions against the introduction of new SARS-CoV variants focused on surveillance of airports and ground transportation.

Brazil is one of the nations most affected by Coronavirus disease 2019 (COVID-19). The introduction and establishment of new virus variants can be related to an increase in cases and fatalities. The emergence of Omicron, the most modified SARS-CoV-2 variant, caused alarm for the public health of Brazil. In this study, we examined the effects of the Omicron introduction in Minas Gerais (MG), the second-most populous state of Brazil. A total of 430 Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) samples from November 2021 to June 2022 from Belo Horizonte (BH) city were sequenced. These newly sequenced genomes comprise 72% of all previously available SARS-CoV-2 genomes for the city. Evolutionary analysis of novel viral genomes reveals that a great diversity of Omicron sublineages have circulated in BH, a pattern in-keeping with observations across Brazil more generally. Bayesian phylogeographic reconstructions indicate that this diversity is a product of a large number of international and national importations. As observed previously, São Paulo state is shown as a significant hub for viral spread throughout the country, contributing to around 70% of all viral Omicron introductions detected in MG.

ABSTRACT Current guidelines for patient isolation in COVID-19 cases recommend a symptom-based approach, averting the use of control real-time reverse transcription PCR (rRT-PCR) testing. However, we hypothesized that patients with persistently positive results by RT-PCR for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) could be potentially infectious for a prolonged time, even if immunocompetent and asymptomatic, which would demand a longer social isolation period than presently recommended. To test this hypothesis, 72 samples from 51 mildly symptomatic immunocompetent patients with long-lasting positive rRT-PCR results for SARS-CoV-2 were tested for their infectiousness in cell culture. The serological response of samples from those patients and virus genomic integrity were also analyzed. Infectious viruses were successfully isolated from 34.38% (22/64) of nasopharynx samples obtained 14 days or longer after symptom onset. Indeed, we observed successful virus isolation up to 128 days. Complete SARS-COV-2 genome integrity was demonstrated, suggesting the presence of replication-competent viruses. No correlation was found between the isolation of infectious viruses and rRT-PCR cycle threshold values or the humoral immune response. These findings call attention to the need to review current isolation guidelines, particularly in scenarios involving high-risk individuals. IMPORTANCE In this study, we evaluated mildly symptomatic immunocompetent patients with long-lasting positive rRT-PCR results for SARS-CoV-2. Infectious viruses were successfully isolated in cell cultures from nasopharynx samples obtained 14 days or longer after symptom onset. Indeed, we observed successful virus isolation for up to 128 days. Moreover, SARS-CoV-2 genome integrity was demonstrated by sequencing, suggesting the presence of replication-competent viruses. These data point out the risk of continuous SARS-CoV-2 transmission from patients with prolonged detection of SARS-CoV-2 in the upper respiratory tract, which has important implications for current precaution guidelines, particularly in settings where vulnerable individuals may be exposed (e.g., nursing homes and hospitals).

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.

Arboviruses pose a major threat throughout the world and represent a great burden in tropical countries of South America. Although generally associated with moderate febrile illness, in more severe cases they can lead to neurological outcomes, such as encephalitis, Guillain-Barré syndrome, and Congenital Syndromes. In this context astrocytes play a central role in production of inflammatory cytokines, regulation of extracellular matrix, and control of glutamate driven neurotoxicity in the central nervous system. Here, we presented a comprehensive genome-wide transcriptome analysis of human primary astrocytes infected with Chikungunya, Mayaro, Oropouche, or Zika viruses. Analyses of differentially expressed genes (DEGs), pathway enrichment, and interactomes have shown that Alphaviruses up-regulated genes related to elastic fiber formation and N-glycosylation of glycoproteins, with down-regulation of cell cycle and DNA stability and chromosome maintenance genes. In contrast, Oropouche virus up-regulated cell cycle and DNA maintenance and condensation pathways while down-regulated extracellular matrix, collagen metabolism, glutamate and ion transporters pathways. Zika virus infection only up-regulated eukaryotic translation machinery while down-regulated interferon pathways. Reactome and integration analysis revealed a common signature in down-regulation of innate immune response, antiviral response, and inflammatory cytokines associated to interferon pathway for all arboviruses tested. Validation of interferon stimulated genes by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) corroborated our transcriptome findings. Altogether, our results showed a co-evolution in the mechanisms involved in the escape of arboviruses to antiviral immune response mediated by the interferon (IFN) pathway.

In an enclosure with nine collared peccaries (Pecari tajacu) from the Rio de Janeiro city Zoo, Brazil, one specimen was found dead and two others developed prostration, apathy and dehydration, resulting on its death. Necropsy of two animals pointed to pulmonary and renal damage. Histological examination revealed vasculitis in spleen from both P. tajacu, suggesting a systemic viral infection. Lungs from one specimen showed fibrinoid vasculitis, alveolar damage with hyaline membrane, and interstitial lymphocytes infiltration. Virome analysis in anal wash samples from the latter two animals revealed a new type of Betacoronavirus, lineage A, provisionally named Ptajacu-CoV.