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In the present study, patients with acute OROV fever were classified as early seroconverters (IgM/IgG positive at baseline) or late seroconverters (IgM/IgG negative at baseline) and the timeline kinetics of the production of chemokines and cytokines were assessed at 1-3, 4-7, 8-10 and ≥11 days after patients have reported the first symptoms. Regardless immunoglobulin profile, all OROV fever patients presented higher levels of CXCL8, and IFN-α and lower levels of TNF and IL-10 at baseline as compared to healthy donors (HD). Lower levels of CCL2, CXCL10, and IFN-γ and higher levels of CCL2, CXCL10, IL-6, and IL-17A were detected in early and late seroconverters, respectively, as compared to HD. While early seroconverters presented the increasing levels of CCL2 along the timeline, late seroconverters displayed decreasing levels of CCL2, CXCL10, and IL-6 following days of disease onset. Noteworthy was that IFN-α was revealed as universal biomarker of human OROV fever, while CXCL8 & IL-5 and CXCL10 & IL-17 were consistently observed in early and late seroconverters, respectively. Thus, our results suggest that the production of IFN-α, CXCL10, and IL-17 precede the seroconversion bringing novel insights on the immunological events triggered by the OROV disease.

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The Yellow Fever (YF) vaccination is recommended for people living in endemic areas and represents the most effective strategy to reduce the risk of infection. Previous studies have warned that booster regimens should be considered to guarantee the long-term persistence of 17DD-YF-specific memory components in adults living in areas with YF-virus circulation. Considering the lower seroconversion rates observed in children (9-12 months of age) as compared to adults, this study was designed in order to access the duration of immunity in single-dose vaccinated children in a 10-years cross-sectional time-span. The levels of neutralizing antibodies (PRNT) and the phenotypic/functional memory status of T and B-cells were measured at a baseline, 30-45 days, 1, 2, 4, 7, and 10 years following primary vaccination. The results revealed that a single dose induced 85% of seropositivity at 30-45 days and a progressive time-dependent decrease was observed as early as 2 years and declines toward critical values (below 60%) at time-spans of ≥4-years. Moreover, short-lived YF-specific cellular immunity, mediated by memory T and B-cells was also observed after 4-years. Predicted probability and resultant memory analysis emphasize that correlates of protection (PRNT; effector memory CD8+ T-cells; non-classical memory B-cells) wane to critical values within ≥4-years after primary vaccination. Together, these results clearly demonstrate the decline of 17DD-YF-specific memory response along time in children primarily vaccinated at 9-12 months of age and support the need of booster regimen to guarantee the long-term persistence of memory components for children living in areas with high risk of YF transmission.

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Yellow fever virus (YFV) penetrates the skin through the bite of a vector mosquito and spreads to various organs, mainly the liver, where it causes lesions and induces necrosis and apoptosis. We evaluated the mRNA expression of various cytokines and the activation of caspases in HepG2 cells infected with YFV. We observed that interferon-α (IFN-α) expression decreased and IFN-ß, transforming growth factor (TGF)-ß IIIR, interleukin (IL)-6, and IL-8 expression increased in cells infected with genotype 1. In contrast, TNF-α expression increased in cells infected with genotype 2 but not with genotype 1. This provides insights into the role of cytokine regulation in yellow fever.

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Oropouche orthobunyavirus (OROV) has significant impact in public health in Amazon region. This arbovirus is one of the most common causes of febrile illness in Brazil, and is responsible for several epidemics since 1960's. In this study, we sequenced and characterized the complete coding sequences (S-, M- and L-RNA) of 35 OROV isolates from Brazil. Here, we classified 20 strains in genotype I from Pará and Maranhão states, nine as genotype II from Pará and Rondônia states confirmed, four classified into genotype III from Acre, Maranhão, Minas Gerais and Rondônia states and two genotype IV from Amazonas State. Also, we did not observe reassortment events involving the OROV isolates. In addition, we developed novel RT-PCR tools to identify reassortment events among OROV strains. These data will be useful to better understand the molecular epidemiology and diagnostic of OROV infections.

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Zika virus (ZIKV) is an arbovirus belonging to the genus Flavivirus (Flaviviridae). ZIKV infection is associated with alterations in various organs, including the liver, lungs, and kidneys. Studies on the influence of posttranscriptional control on viral infections have demonstrated that microRNAs (miRNAs) interfere with different stages of the replicative cycle of several viruses and may influence the disease outcome. To shed light on ZIKV-induced regulation of host miRNA-processing machinery in the above organs, we analyzed the expression of genes encoding key proteins of the miRNA pathway in different ZIKV-infected continuous primate cell lineages (HepG2, A549, and MA104) by reverse-transcription quantitative polymerase chain reaction (RT-qPCR). Expression of the genes encoding the miRNA-related proteins DGCR8, Ago1, and Ago3 in HepG2 cells and Drosha, Dicer, Ago2, and Ago3 in A549 and MA104 cells was significantly altered in the presence of ZIKV. Our results suggest that ZIKV modulates miRNA levels during infection in liver, lung, and kidney cells, which may be an additional mechanism of host cell subversion in these organs.

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In this investigation, machine-enhanced techniques were applied to bring about scientific insights to identify a minimum set of phenotypic/functional memory-related biomarkers for post-vaccination follow-up upon yellow fever (YF) vaccination. For this purpose, memory status of circulating T-cells (Naïve/early-effector/Central-Memory/Effector-Memory) and B-cells (Naïve/non-Classical-Memory/Classical-Memory) along with the cytokine profile (IFN/TNF/IL-5/IL-10) were monitored before-NV(day0) and at distinct time-points after 17DD-YF primary vaccination-PV(day30-45); PV(year1-9) and PV(year10-11). A set of biomarkers (eEfCD4; EMCD4; CMCD19; EMCD8; IFNCD4; IL-5CD8; TNFCD4; IFNCD8; TNFCD8; IL-5CD19; IL-5CD4) were observed in PV(day30-45), but not in NV(day0), with most of them still observed in PV(year1-9). Deficiencies of phenotypic/functional biomarkers were observed in NV(day0), while total lack of memory-related attributes was observed in PV(year10-11), regardless of the age at primary vaccination. Venn-diagram analysis pre-selected 10 attributes (eEfCD4, EMCD4, CMCD19, EMCD8, IFNCD4, IL-5CD8, TNFCD4, IFNCD8, TNFCD8 and IL-5CD4), of which the overall mean presented moderate accuracy to discriminate PV(day30-45)&PV(year1-9) from NV(day0)&PV(year10-11). Multi-parameter approaches and decision-tree algorithms defined the EMCD8 and IL-5CD4 attributes as the top-two predictors with moderated performance. Together with the PRNT titers, the top-two biomarkers led to a resultant memory status observed in 80% and 51% of volunteers in PV(day30-45) and PV(year1-9), contrasting with 0% and 29% found in NV(day0) and PV(year10-11), respectively. The deficiency of memory-related attributes observed at PV(year10-11) underscores the conspicuous time-dependent decrease of resultant memory following17DD-YF primary vaccination that could be useful to monitor potential correlates of protection in areas under risk of YF transmission.

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Yellow fever is a zoonotic disease caused by the yellow fever virus (YFV) and transmitted by mosquitoes of the family Culicidae. It is well known that cellular and viral microRNAs (miRNAs) are involved in modulation of viral and cellular gene expression, as well as immune response, and are considered by the scientific community as possible targets for an effective therapy against viral infections. This regulation may be involved in different levels of infection and clinical symptomatology. We used viral titration techniques, viral kinetics from 24 to 96 hours postinfection (hpi), and analyzed the expression of key proteins related to the miRNA pathway by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). The expression of Dicer was different when compared over the course of infection by the distinct YFV genotypes. Drosha expression was similar during infection by YFV genotype 1 or 2, with a decrease in their expression over time and a slight increase in 96 hpi. Ago1, Ago2, and Ago4 showed different levels of expression between the viral genotypes: for YFV genotype 1 infection, Ago1 presented a positive expression, while for YFV genotype 2, it showed a negative expression, when compared with negative controls. We conclude that YFV infection modulates the proteins involved in miRNA biogenesis, which can regulate both viral replication and cellular immune response.

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The State of Pará comprises 26% of Brazilian Amazon region where a large diversity of arboviruses has been described. This study sought to assess the prevalence and distribution of haemagglutination-inhibition antibodies against antigens of nine different types of arbovirus of the Bunyaviridae family, where eight were Orthobunyavirus: Guaroa virus, Maguari virus, Tacaiuma virus, Utinga virus, Belem virus, Caraparu virus, Oropouche virus and Catu virus, and one Phlebovirus: Icoaraci virus in sera samples of water buffaloes in Pará State, Brazil. For all Arboviruses investigated there were antibodies, with the exception of Belem virus. Antibodies to Maguari virus were more prevalent (7.33%). The water buffaloes of the present study showed variable levels of antibodies in monotypic and heterotypic reactions that may indicate there are movements from most bunyavirus studied in domestic buffaloes in the state of Pará, and the Maguari virus presents the largest circulation. Therefore, further studies are needed to investigate the role of water buffalo in the maintenance and dispersal of arboviruses, as well as whether these viruses can cause disease in that species, especially in cases of birth defects and abortions.(AU)

O Estado do Pará corresponde a 26% da Amazônica brasileira, onde uma grande quantidade de Arbovírus tem sido descrito. O presente trabalho teve como objetivo determinar a prevalência de anticorpos detectados pela técnica de inibição de hemaglutinação contra nove tipos diferentes de arbovírus da família Bunyaviridae, sendo oito do gênero Orthobunyavirus: vírus Guaroa, vírus Maguari, vírus Tacaiuma, vírus Utinga, vírus Belem, vírus Caraparu, vírus Oropouche e vírus Catu e um do gênero Phlebovirus: vírus Icoaraci, em soros de búfalos de água no Estado do Pará, Brasil. Para todos os Arbovírus investigados houve presença de anticorpos, com exceção do vírus Belém. Anticorpos para o vírus Maguari foram mais prevalentes (7,33%). O rebanho bubalino do presente estudo mostrou variáveis níveis de anticorpos em reações heterotípicas e monotípicas podendo indicar que há circulação da maioria dos bunyavírus estudados em búfalos domésticos no estado do Pará, e que o vírus Maguari é o de maior circulação. Por isso, são necessários outros estudos para investigar o papel dos búfalos de água na manutenção e dispersão de arbovírus, assim como se esses vírus podem causar enfermidades na referida espécie, principalmente, em casos de defeitos congênitos e abortamentos.(AU)

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title>Abstract /title> p>The State of Pará comprises 26% of Brazilian Amazon region where a large diversity of arboviruses has been described. This study sought to assess the prevalence and distribution of haemagglutination-inhibition antibodies against antigens of nine different types of arbovirus of the italic>Bunyaviridae /italic> family, where eight were italic>Orthobunyavirus: Guaroa virus, Maguari virus, Tacaiuma virus, Utinga virus, Belem virus, Caraparu virus, Oropouche virus /italic> and italic>Catu virus, /italic> and one italic>Phlebovirus: Icoaraci virus /italic> in sera samples of water buffaloes in Pará State, Brazil. For all Arboviruses investigated there were antibodies, with the exception of italic>Belem virus. /italic> Antibodies to italic>Maguari virus /italic>were more prevalent (7.33%). The water buffaloes of the present study showed variable levels of antibodies in monotypic and heterotypic reactions that may indicate there are movements from most bunyavirus studied in domestic buffaloes in the state of Pará, and the italic>Maguari virus /italic> presents the largest circulation. Therefore, further studies are needed to investigate the role of water buffalo in the maintenance and dispersal of arboviruses, as well as whether these viruses can cause disease in that species, especially in cases of birth defects and abortions. /p>

title>Resumo /title> p>O Estado do Pará corresponde a 26% da Amazônica brasileira, onde uma grande quantidade de Arbovírus tem sido descrito. O presente trabalho teve como objetivo determinar a prevalência de anticorpos detectados pela técnica de inibição de hemaglutinação contra nove tipos diferentes de arbovírus da família italic>Bunyaviridae, /italic> sendo oito do gênero italic>Orthobunyavirus: vírus Guaroa, vírus Maguari, vírus Tacaiuma, vírus Utinga, vírus Belem, vírus Caraparu, vírus Oropouche /italic> e italic>vírus Catu /italic>e um do gênero italic>Phlebovirus: vírus Icoaraci, /italic> em soros de búfalos de água no Estado do Pará, Brasil. Para todos os Arbovírus investigados houve presença de anticorpos, com exceção do italic>vírus Belém. /italic>Anticorpos para o italic>vírus Maguari /italic> foram mais prevalentes (7,33%). O rebanho bubalino do presente estudo mostrou variáveis níveis de anticorpos em reações heterotípicas e monotípicas podendo indicar que há circulação da maioria dos bunyavírus estudados em búfalos domésticos no estado do Pará, e que o vírus Maguari é o de maior circulação. Por isso, são necessários outros estudos para investigar o papel dos búfalos de água na manutenção e dispersão de arbovírus, assim como se esses vírus podem causar enfermidades na referida espécie, principalmente, em casos de defeitos congênitos e abortamentos. /p>

RESUMO

The State of Pará comprises 26% of Brazilian Amazon region where a large diversity of arboviruses has been described. This study sought to assess the prevalence and distribution of haemagglutination-inhibition antibodies against antigens of nine different types of arbovirus of the Bunyaviridae family, where eight were Orthobunyavirus: Guaroa virus, Maguari virus, Tacaiuma virus, Utinga virus, Belem virus, Caraparu virus, Oropouche virus and Catu virus, and one Phlebovirus: Icoaraci virus in sera samples of water buffaloes in Pará State, Brazil. For all Arboviruses investigated there were antibodies, with the exception of Belem virus. Antibodies to Maguari virus were more prevalent (7.33%). The water buffaloes of the present study showed variable levels of antibodies in monotypic and heterotypic reactions that may indicate there are movements from most bunyavirus studied in domestic buffaloes in the state of Pará, and the Maguari virus presents the largest circulation. Therefore, further studies are needed to investigate the role of water buffalo in the maintenance and dispersal of arboviruses, as well as whether these viruses can cause disease in that species, especially in cases of birth defects and abortions.

O Estado do Pará corresponde a 26% da Amazônica brasileira, onde uma grande quantidade de Arbovírus tem sido descrito. O presente trabalho teve como objetivo determinar a prevalência de anticorpos detectados pela técnica de inibição de hemaglutinação contra nove tipos diferentes de arbovírus da família Bunyaviridae, sendo oito do gênero Orthobunyavirus: vírus Guaroa, vírus Maguari, vírus Tacaiuma, vírus Utinga, vírus Belem, vírus Caraparu, vírus Oropouche e vírus Catu e um do gênero Phlebovirus: vírus Icoaraci, em soros de búfalos de água no Estado do Pará, Brasil. Para todos os Arbovírus investigados houve presença de anticorpos, com exceção do vírus Belém. Anticorpos para o vírus Maguari foram mais prevalentes (7,33%). O rebanho bubalino do presente estudo mostrou variáveis níveis de anticorpos em reações heterotípicas e monotípicas podendo indicar que há circulação da maioria dos bunyavírus estudados em búfalos domésticos no estado do Pará, e que o vírus Maguari é o de maior circulação. Por isso, são necessários outros estudos para investigar o papel dos búfalos de água na manutenção e dispersão de arbovírus, assim como se esses vírus podem causar enfermidades na referida espécie, principalmente, em casos de defeitos congênitos e abortamentos.

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