ABSTRACT Wild birds have great prominence in the transmission of diseases to humans, mainly due to their ease of access to human populations, raising concerns about the potential impact of that proximity on public health. The present study reports ARV and PBV occurrence in wild birds from an environmental protection area in the Amazon biome, in Pará state, Brazil. We obtained 155 fecal specimens from 47 species of wild birds for RNA extraction, ARV and PBV detection utilizing molecular methods, nucleotide sequencing, and phylogenetic analysis. ARV prevalence was 0.6% (1/155), being positive in an individual of Myrmotherula longipennis, and PBV prevalence was 1.29% (2/155), affecting two individuals of Guira guira. The PBV strains were assigned to genogroup I based on phylogenetic analysis, and they shared a strong phylogenetic link with strains isolated from different geographic locations. The ARV strain was more closely related to strains that had previously circulated in the same region. The presence of ARV and PBV in this habitat suggests that infection cycles of these agents occur naturally in the wild ecosystem, potentially triggering transmission events between bird species and humans. This is the first study on ARV detection in wild birds in Brazil and the first report on the occurrence of PBV in wild Guira guira. Additional studies are required to determine the epidemiology, origin, evolution, and emergence of new potentially pathogenic viruses in the Amazon.

RESUMO As aves silvestres têm grande destaque na transmissão de doenças ao homem, principalmente pela facilidade de acesso à população humana, levantando preocupações sobre o potencial impacto dessa proximidade no contexto da saúde pública. O presente estudo relata a ocorrência de ARV e PBV em aves silvestres de uma área de proteção ambiental no bioma Amazônico. Obtivemos 155 espécimes fecais de 47 espécies de aves silvestres, que foram submetidos a extração de RNA, detecção de ARV e PBV por técnicas moleculares, sequenciamento nucleotídico e análise filogenética. A prevalência de infecção por ARV foi 0,6% (1/155), sendo positivo em um indivíduo de Myrmotherula longipennis, e a prevalência de PBV foi 1,29% (2/155), afetando dois insdivíduos de Guira guira. A análise filogenética agrupou as cepas de PBV no genogrupo I, mostrando intensa relação filogenética com cepas isoladas de outras regiões geográficas. A cepa de ARV esteve mais relacionada filogeneticamente a cepas previamente circulantes na mesma região. Este é o primeiro estudo sobre detecção de ARV em aves silvestres no Brasil e o primeiro relato sobre a ocorrência de PBV em Guira guira. A circulação de ARV e PBV nesse ambiente demonstra que os ciclos desses agentes estão ocorrendo de forma natural no ecossistema silvestre, o que pode desencadear possíveis eventos de transmissão entre as espécies aviárias e humanos. Estudos adicionais são necessários para determinar a epidemiologia, origem, evolução e surgimento de novos vírus potencialmente patogênicos na Amazônia.

BACKGROUND In this study, we evaluated the role of free-living domestic pigeons (Columba livia) as a reservoir of arboviruses in the city of Belém, state of Pará, Brazil. We investigated the presence of antibodies against the most prevalent arboviruses. OBJECTIVES This study was aimed at evaluating some clinical and physical parameters of domestic pigeons, including the presence of antibodies to Amazon-endemic arboviruses. METHODS Eighty-five healthy pigeons were captured in Mangal das Garças Park, in Belém, and were bled. Upon capture, the birds were subjected to a clinical examination in search of alterations that could indicate the presence of arboviruses. Blood samples were converted to serum and tested using the haemagglutination inhibition (HI) technique with a panel of 19 antigens of arboviruses circulating in the Amazon. The confirmation assay for the positive reactions to the viral species tested by HI was a neutralisation test in new-born Swiss albino mice (Mus musculus) [mouse neutralisation test (MNT)]. FINDINGS A total of 10 (11.8%) serum samples tested positive for antiflavivirus antibodies by HI. All the samples positive for the HI test were subjected to MNT for detection of viruses and yielded negative results (logarithmic neutralisation index < 1.7). MAIN CONCLUSION The results represent the first serological detection of antiarbovirus antibodies in domestic pigeons as potential hosts of arboviruses in Brazil. The detection of haemagglutination-inhibiting antibodies against genus Flavivirus indicated that there was recent contact between the analysed domestic pigeons and these arboviruses. Further studies are needed to evaluate the role of free-living pigeons in the maintenance cycle and spread of arboviruses in the Amazon.

RESUMO: O estado do Pará corresponde a 26% da Amazônia brasileira, onde uma grande diversidade de arbovírus foi descrita. Este estudo procurou avaliar a prevalência e a distribuição de anticorpos inibidores da hemaglutinação (IH) contra antígenos de quatro alfavirus (Togaviridae: Alphavirus ), das espécies: Vírus da encefalite equina do leste (EEEV), Vírus da encefalite equina do oeste (WEEV), Vírus mayaro (MAYV) e Vírus mucambo (MUCV), de 753 amostras de soro de equinos no estado do Pará, Brasil. Todos os arbovirus pesquisados foram detectados, indicando que os equinos são suscetíveis a esses Alphavirus e mostrando evidências de sua circulação ativa em animais de fazenda na Amazônia brasileira.

ABSTRACT: 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 hemagglutination inhibition (HI) antibodies against antigens of four alphaviruses (Togaviridae: Alphavirus ) from the species: Eastern equine encephalitis (EEEV), Western equine encephalitis (WEEV), Mayaro virus (MAYV), and Mucambo virus (MUCV) in 753 serum samples of horses in Pará State, Brazil. All investigated arboviruses were detected and indicate that horses are susceptible to these alphaviruses, and show evidences of their active circulation in farm animals in the Brazilian Amazon.

<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>

<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>

Background The Amazon as a whole is the largest reservoir of arboviruses worldwide, while the Brazilian Amazon hosts the largest variety of arboviruses isolated to date. In this study, the results of an indirect sandwich IgG ELISA, standardized for 19 arbovirustypes circulating among horses in Brazilian Amazon, were compared to results of the hemagglutination inhibition test. A screening test assessed the conditional probability distribution and a Pearson linear correlation test determined the correlation strength among the absorbance values recorded for viruses from the same family.Findings Sensitivity varied between 40.85 and 100%; the specificity was low and ranged from 39.71 to 67.0%; and the accuracy varied between 41 and 65.2%. The test developed in this study yielded a large number of serological cross-reactions.Conclusions The test can be employed to detect IgG antibodies within one arbovirus family; however, the hemagglutination test or other more specific techniques, such as the serum neutralization test in mice or the plaque-reduction neutralization test, are essential complementary methods for positive cases.