Immunological impact of cytokines on the chikungunya virus pathophysiology: A literature narrative review.

The chikungunya virus (CHIKV) is a member of the genus Alphavirus, family Togaviridae. CHIKV causes an acute systemic febrile condition, accompanied by severe polyarthralgia, intense muscle pain, and maculopapular exanthema, which may still occur in many patients. In rare cases, unusual symptoms may occur, eventually worsening the condition and resulting in a fatal outcome. It is a single-stranded, non-segmented RNA virus with a genome of approximately 11,805 nucleotides that organises a genetic and molecular chain that encodes non-structural proteins (nsP1, nsP2, nsP3, nsP4) and structural proteins (E3, E2, 6K, and E1). The fundamental role of immune response in the evolution of the disease is known. Understanding the role of immune response in the pathogenesis of CHIKV infection is challenging. In this context, innate and adaptive immune responses establish a connective interface that induces the production of various mediators that modulate the strategy of inhibiting viral replication. However, the immune escape articulated by the virus indicates that the action of pro-and anti-inflammatory cytokines contributes to the worsening of the disease and potentiates tissue damage with joint involvement. In this review, we discuss the role of the primary pro-and anti-inflammatory cytokines in the immunopathological processes of chikungunya fever.

Characterization of Triniti virus supports its reclassification in the family Peribunyaviridae.

Triniti virus (TNTV) has been isolated in Trinidad and Tobago and in Brazil. To date little is known about this virus, which is classified as an ungrouped virus within the family Togaviridae. Here, three isolates of TNTV were characterized both genetically and antigenically. The genome was shown to contain three RNA segments: small (S), medium (M) and large (L). Genome organization, protein sizes and protein motifs were similar to those of viruses in the genus Orthobunyavirus, family Peribunyaviridae. Antigenic reactivity revealed the three TNTV isolates to be closely related, but no serologic cross-reaction with other orthobunyaviruses. Morphological observation by transmission electron microscopy indicated that virus size and symmetry were compatible with those of viruses in the family Peribunyaviridae. Our serological, morphological and molecular results support the taxonomic reclassification of TNTV as a member of the genus Orthobunyavirus, family Peribunyaviridae.

Experimental infection of golden hamsters with Guama virus (Peribunyaviridae, Orthobunyavirus).

The Guama virus (GMAV) is a member of Peribunyaviridae family, Orthobunyavirus genus. Several strains of the virus were isolated in South and Central Americas from several hosts, such as humans, wild animals, including nonhuman primates, wild rodents and mosquitoes as well as mice used as sentinels. The virus is able to cause febrile disease in humans. Here we describe for the first time pathologic and biochemical findings in golden hamsters (Mesocricetus auratus) infected with the prototype GMAV. Blood and organs of infected and control animals were collected every 24 h after infection from the 1st to the 7th day post infection (dpi) and at 21 dpi when experiment was ended. The tissues were processed for histopathology and immunohistochemistry. The blood and serum were used to determine viremia and biochemical markers plus to detect anti-GMAV antibodies. The viremia was early detected already on the 1st dpi and it was no longer detected on the 3rd dpi. Total anti-GMAV antibodies were detected from the 6th dpi. Hepatic markers as ALT of infected animals were increased and showed statistically significant difference in comparison with control animals, indicating damage of the liver; indeed the liver was the most affected organ, but other organs presented lesions and positive GMAV immunostaining as brain, lung, liver, spleen, and kidney. Our findings indicate that golden hamsters are a good animal model for experimental infection of the GMAV.

First isolation of West Nile virus in Brazil.

BACKGROUND: Serological evidence of West Nile virus (WNV) infection has been reported in different regions of Brazil from equine and human hosts but the virus had never been isolated in the country. OBJECTIVES: We sought to identify the viral etiology of equine encephalitis in Espírito Santo state. METHODS: We performed viral culture in C6/36 cells, molecular detection of WNV genome, histopathology and immunohistochemistry from horse cerebral tissue. We also carried out sequencing, phylogenetic analysis and molecular clock. FINDINGS: Histopathologic analysis from horse cerebral tissue showed injury related to encephalitis and WNV infection was confirmed by immunohistochemistry. The virus was detected by reverse transcription quantitative polymerase chain reaction (RT-qPCR) from brain tissue and subsequently isolated in C6/36 cells. WNV full-length genome was sequenced showing the isolated strain belongs to lineage 1a. The molecular clock indicated that Brazilian WNV strain share the same common ancestor that were circulating in US during 2002-2005. MAIN CONCLUSIONS: Here we report the first isolation of WNV in Brazil from a horse with neurologic disease, which was clustered into lineage 1a with others US WNV strains isolated in beginning of 2000's decade.

In situ inflammasome activation results in severe damage to the central nervous system in fatal Zika virus microcephaly cases.

Zika virus (ZIKV) has caused substantial concern worldwide owing to its association with severe birth defects, such as microcephaly and other congenital malformations. Inflammasomes, i.e., multi-protein complexes that induce inflammation and pyroptosis, are predicted to contribute to the immune response to this flavivirus. Accordingly, in this study, the in situ inflammasome response was evaluated in fatal cases of ZIKV-linked microcephaly. Brain tissue samples were collected from eight babies, including four ZIKV-positive microcephalic neonates who died after birth and four flavivirus-negative neonatal controls who died of other causes and whose central nervous system (CNS) architecture was preserved. In the ZIKV-positive newborn/stillbirth babies, the major histopathological alterations included atrophy of the cortical layer, a predominance of mononuclear cell infiltration in the Virchow-Robin space, neuronal necrosis, vacuolization and neuronal degeneration, neuronophagy, and gliosis. An immunohistochemical analysis of tissues in the neural parenchyma showed significantly higher expression of the receptors NLRP1, NLRP3, and AIM2, cytokines IL-1ß, IL-18, and IL-33, and enzymes caspase 1, iNOS, and arginase 1 in ZIKV-positive microcephaly cases than in flavivirus-negative controls. These results suggest that inflammasome activation can aggravate the neuroinflammatory response and consequently increase CNS damage in neonates with fetal neural ZIKV infection and microcephaly.

Clinical and serological tests for arboviruses in free-living domestic pigeons (Columba livia).

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.

Antibody cross-reactivity and evidence of susceptibility to emerging Flaviviruses in the dengue-endemic Brazilian Amazon.

OBJECTIVES: Several Flaviviruses can co-circulate. Pre-existing immunity to one virus can modulate the response to a heterologous virus; however, the serological cross-reaction between these emerging viruses in dengue virus (DENV)-endemic regions are poorly understood. METHODS: A cross-sectional study was performed among the residents of Manaus city in the state of Amazonas, Brazil. The serological response was assessed by hemagglutination inhibition assay (HIA), enzyme-linked immunosorbent assay, and neutralization assay. RESULTS: A total of 74.52% of the participants were immunoglobulin G-positive (310/416), as estimated by lateral flow tests. Overall, 93.7% of the participants were seropositive (419/447) for at least one DENV serotype, and the DENV seropositivity ranged between 84.8% and 91.0%, as determined by HIA. About 93% had antiyellow fever virus 17D-reactive antibodies, whereas 80.5% reacted to wild-type yellow fever virus. Zika virus (ZIKV) had the lowest seropositivity percentage (52.6%) compared with other Flaviviruses. Individuals who were DENV-positive with high antibody titers by HIA or envelope protein domain III enzyme-linked immunosorbent assay reacted strongly with ZIKV, whereas individuals with low anti-DENV antibody titers reacted poorly toward ZIKV. Live virus neutralization assay with ZIKV confirmed that dengue serogroup and ZIKV-spondweni serogroup are far apart; hence, individuals who are DENV-positive do not cross-neutralize ZIKV efficiently. CONCLUSION: Taken together, we observed a high prevalence of DENV in the Manaus-Amazon region and a varying degree of cross-reactivity against emerging and endemic Flaviviruses. Epidemiological and exposure conditions in Manaus make its population susceptible to emerging and endemic arboviruses.

Isolation of Flaviviruses and Alphaviruses with Encephalitogenic Potential Diagnosed by Evandro Chagas Institute (Pará, Brazil) in the Period of 1954-2022: Six Decades of Discoveries.

Viruses with encephalitogenic potential can cause neurological conditions of clinical and epidemiological importance, such as Saint Louis encephalitis virus, Venezuelan equine encephalitis virus, Eastern equine encephalitis virus, Western equine encephalitis virus, Dengue virus, Zika virus, Chikungunya virus, Mayaro virus and West Nile virus. The objective of the present study was to determine the number of arboviruses with neuroinvasive potential isolated in Brazil that corresponds to the collection of viral samples belonging to the Department of Arbovirology and Hemorrhagic Fevers, Evandro Chagas Institute (SAARB/IEC) of the Laboratory Network of National Reference for Arbovirus Diagnosis from 1954 to 2022. In the analyzed period, a total of 1,347 arbovirus samples with encephalitogenic potential were isolated from mice; 5,065 human samples were isolated exclusively by cell culture; and 676 viruses were isolated from mosquitoes. The emergence of new arboviruses may be responsible for diseases still unknown to humans, making the Amazon region a hotspot for infectious diseases due to its fauna and flora species characteristics. The detection of circulating arboviruses with the potential to cause neuroinvasive diseases is constant, which justifies the continuation of active epidemiological surveillance work that offers adequate support to the public health system regarding the virological diagnosis of circulating arboviruses in Brazil.

Persistence of experimental Rocio virus infection in the golden hamster (Mesocricetus auratus).

Rocio virus (ROCV) is an encephalitic flavivirus endemic to Brazil. Experimental flavivirus infections have previously demonstrated a persistent infection and, in this study, we investigated the persistence of ROCV infection in golden hamsters (Mesocricetus auratus). The hamsters were infected intraperitoneally with 9.8 LD50/0.02 mL of ROCV and later anaesthetised and sacrificed at various time points over a 120-day period to collect of blood, urine and organ samples. The viral titres were quantified by real-time-polymerase chain reaction (qRT-PCR). The specimens were used to infect Vero cells and ROCV antigens in the cells were detected by immunefluorescence assay. The levels of antibodies were determined by the haemagglutination inhibition technique. A histopathological examination was performed on the tissues by staining with haematoxylin-eosin and detecting viral antigens by immunohistochemistry (IHC). ROCV induced a strong immune response and was pathogenic in hamsters through neuroinvasion. ROCV was recovered from Vero cells exposed to samples from the viscera, brain, blood, serum and urine and was detected by qRT-PCR in the brain, liver and blood for three months after infection. ROCV induced histopathological changes and the expression of viral antigens, which were detected by IHC in the liver, kidney, lung and brain up to four months after infection. These findings show that ROCV is pathogenic to golden hamsters and has the capacity to cause persistent infection in animals after intraperitoneal infection.

Emergence of New Immunopathogenic Factors in Human Yellow Fever: Polarisation of the M1/M2 Macrophage Response in the Renal Parenchyma.

Macrophages in the kidney play a pathogenic role in inflammation and fibrosis. Our study aimed to understand the polarisation of the M1 and M2 phenotypic profiles of macrophages in injured kidney tissue retrieved from fatal cases of yellow fever virus (YFV). A total of 11 renal tissue biopsies obtained from patients who died of yellow fever (YF) were analysed. To detect antibodies that promote the classical and alternative pathways of macrophage activation, immunohistochemical analysis was performed to detect CD163, CD68, inducible nitric oxide synthase (iNOS), arginase 1, interleukin (IL)-4, IL-10, interferon (IFN)-γ, IFN-ß, tumour necrosis factor (TNF)-α, IL-13, and transforming growth factor (TGF)-ß. There was a difference in the marker expression between fatal cases of YFV and control samples, with increased expression in the cortical region of the renal parenchyma. The immunoexpression of CD68 and CD163 receptors suggests the presence of activated macrophages migrating to infectious foci. The rise in IL-10, IL-4, and IL-13 indicated their potential role in the inactivation of the inflammatory macrophage response and phenotypic modulation of M2 macrophages. The altered expression of IFN-γ and IFN-ß demonstrates the importance of the innate immune response in combating microorganisms. Our findings indicate that the polarisation of M1 and M2 macrophages plays a vital role in the renal immune response to YFV.

Co-infection of Peruvian horse sickness virus and West Nile virus associated with neurological diseases in horses from Brazil.

In 2018, during the surveillance for West Nile virus (WNV) in horses with neurological clinical signs in the state of Espírito Santo (Brazil), 19 animals were investigated, and 52 biological samples were collected for WNV diagnostic. One brain sample was positive for WNV by RT-qPCR and the virus was isolated in C6/36 cell culture and sequenced. We obtained a nearly complete genome of WNV co-infected with Peruvian horse sickness virus (PHSV) in the cell culture. After confirmation of PHSV by next-generation sequencing, a new PHSV RT-qPCR protocol was developed, which was used to detect another horse positive only for PHSV. This assay provides a simple and direct method for easy identification of PHSV from biological samples from horses and may become a useful tool in the epidemiological surveillance of this virus. It is the first case of PHSV in Brazil, and only the third country overall to report, 23 years after the first confirmed notification in Peru. Moreover, it is the first reported co-infection of PHSV and WNV in a horse with neurological signs, confirmed by RT-qPCR.

Factors Involved in the Apoptotic Cell Death Mechanism in Yellow Fever Hepatitis.

Yellow fever (YF), a non-contagious infectious disease, is endemic or enzootic to the tropical regions of the Americas and Africa. Periodic outbreaks or epidemics have a significant impact on public health. Programmed cell death, or apoptosis, is generally characterised by distinct morphological changes and energy-dependent biochemical pathways. In this study, we performed immunohistochemistry analysis to identify and quantify proteases and protein targets involved in the cascade that triggers apoptosis in YF virus (YFV)-infected human hepatocytes. Liver tissue samples were collected from 26 individuals, among whom 21 were diagnosed as YF-positive, and five were flavivirus-negative and died due to other causes. The histopathological alterations in YFV-positive cases were characterised by the presence of apoptotic bodies, steatosis, cellular swelling, and extensive necrosis and haemorrhage in the hepatic lobules. Additionally, we observed an abundance of inflammatory infiltrates in the portal tract. The expression of various apoptotic markers in the hepatic parenchyma, including CASPASE 3, CASPASE 8, BAX, FAS, FASL, GRANZYME B, and SURVIVIN, differed between YFV-positive cases and controls. Collectively, this study confirmed the complexity of YFV infection-induced apoptosis in situ. However, our data suggest that apoptosis in liver parenchyma lesions may significantly contribute to the pathogenesis of fatal YF in humans.

Arboviruses in Free-Ranging Birds and Hematophagous Arthropods (Diptera, Nematocera) from Forest Remnants and Urbanized Areas of an Environmental Protection Area in the Amazon Biome.

The rapid and disorderly urbanization in the Amazon has resulted in the insertion of forest fragments into cities, causing the circulation of arboviruses, which can involve hematophagous arthropods and free-ranging birds in the transmission cycles in urban environments. This study aimed to evaluate the circulation of arboviruses in free-ranging birds and hematophagous arthropods captured in an Environmental Protection Area in the Belem metropolitan area, Brazil. Birds were captured using mist nets, and hematophagous arthropods were collected using a human protected attraction technique and light traps. The birds' sera were subjected to a hemagglutination inhibition test to detect antibodies against 29 arbovirus antigens. Arthropod macerates were inoculated into C6/36 and VERO cell cultures to attempt viral isolation and were tested using indirect immunofluorescence, subsequent genetic sequencing and submitted for phylogenetic analysis. Four bird sera were positive for arbovirus, and one batch of Psorophora ferox was positive for Flavivirus on viral isolation and indirect immunofluorescence. In addition, the Ilheus virus was detected in the sequencing and phylogenetic analysis. The presence of antibodies in sera from free-ranging birds and the isolation of Ilheus virus in Psorophora ferox indicate the circulation of arboviruses in forest remnants in the urban center of Belem.

Prevalence of arbovirus antibodies in young healthy adult population in Brazil.

BACKGROUND: The emergence and re-emergence of infectious diseases are a cause for worldwide concern. The introduction of Zika and Chikungunya diseases in the Americas has exposed unforeseen medical and logistical challenges for public health systems. Moreover, the lack of preventive measures and vaccination against known and emerging mosquito-transmitted pathogens, and the occurrence of unanticipated clinical complications, has had an enormous social and economic impact on the affected populations. In this study, we aimed to measure the seroprevalence of endemic and emerging viral pathogens in military personnel stationed in Manaus, Amazonas state. METHODS: We measured the seropositivity of antibodies against 19 endemic and emerging viruses in a healthy military personnel group using a hemagglutination inhibition assay (HIA). RESULTS: Overall, DENV positivity was 60.4%, and 30.9% of the individuals reacted against ZIKV. Also, 46.6%, 54.7%, 51.3% and 48.7% individuals reacted against West Nile virus (WNV), Saint Louis encephalitis virus (SLEV), Ilheus virus (ILHV) and Rocio virus (ROCV), respectively. Individuals with high DENV HIA titer reacted more frequently with ZIKV or WNV compared to those with low HIA titers. Observed cross-reactivity between Flaviviruses varied depending on the virus serogroup. Additionally, 0.6% and 0.3% individuals were seropositive for Oropouche virus (OROV) and Catu virus (CATUV) from the family Peribunyaviridae, respectively. All samples were negative for Eastern Equine Encephalitis virus (EEEV), Western Equine Encephalomyelitis virus (WEEV), Mayaro virus (MAYV), Mucambo virus (MUCV) and CHIKV from the family Togaviridae. CONCLUSIONS: A high proportion of individuals in our high-risk population (~ 60%) lacked antibodies against major endemic and emerging viruses, which makes them susceptible for further infections. Military personnel serving in the Amazon region could serve as sentinels to strengthen global infectious disease surveillance, particularly in remote areas.

Serological evidence of Eastern equine encephalitis circulation in equids in Pará state, Brazil.

Serum samples from 89 equids were analyzed (75 horses, 9 donkeys, and 5 mules) from the municipality of Viseu, Pará state, Brazil. Samples were collected in November 2014 and August 2015. The antibody prevalence against the following alphaviruses was estimated: Eastern equine encephalitis virus, Western equine encephalitis virus, Mucambo virus, and Mayaro virus. Seroprevalence was determined by the hemagglutination inhibition (HI) technique. Sera that exhibited HI antibodies with heterotypic reactions for the analyzed viruses were subjected to the 90% plaque reduction neutralization test (PRNT90). The HI prevalence of monotypic reactions to EEEV was 7.9%, and that of WEEV was 1.1%, as confirmed by PRNT90. Viral isolation attempts were negative for all tested blood samples. Our results suggest the circulation of equine encephalitis complex viruses. Future studies should evaluate the possible involvement of arthropod hosts and residents in the viral transmission in the study area.

Foram analisadas amostras de soros de 89 equídeos (75 equinos, 9 asininos e 5 muares) oriundos do município de Viseu, estado do Pará. As amostras foram coletadas no período de novembro de 2014 e agosto de 2015. Foram estimadas as prevalências de anticorpos contra os seguintes Alfavírus: vírus da Encefalite equina leste (EEEV), vírus da Encefalite equina oeste (WEEV), vírus Mucambo (MUCV), and vírus Mayaro (MAYV). A soroprevalência foi determinada por técnicas de Inibição da Hemaglutinação (IH), porém os soros que apresentaram anticorpos IH com reações heterotípicas para os vírus analisados foram submetidos ao PRNT90. A prevalência de reações monotípicas por IH para EEEV foi de 7, 9% e para WEEV foi de 1,1%, confirmadas por PRNT90. A tentativa de isolamento viral foi negativa em todas as amostras de sangue testadas. Nossos resultados sugerem a circulação de vírus do complexo das encefalites equinas. Estudos futuros devem ser realizados para avaliar possível envolvimento de hospedeiros artrópodes e residentes na área de abrangência do estudo.

First Description of the Mitogenome and Phylogeny of Culicinae Species from the Amazon Region.

The Culicidae family is distributed worldwide and comprises about 3587 species subdivided into the subfamilies Anophelinae and Culicinae. This is the first description of complete mitochondrial DNA sequences from Aedes fluviatilis, Aedeomyia squamipennis, Coquillettidia nigricans, Psorophora albipes, and Psorophora ferox. The mitogenomes showed an average length of 15,046 pb and 78.02% AT content, comprising 37 functional subunits (13 protein coding genes, 22 tRNAs, and two rRNAs). The most common start codons were ATT/ATG, and TAA was the stop codon for all PCGs. The tRNAs had the typical leaf clover structure, except tRNASer1. Phylogeny was inferred by analyzing the 13 PCGs concatenated nucleotide sequences of 48 mitogenomes. Maximum likelihood and Bayesian inference analysis placed Ps. albipes and Ps. ferox in the Janthinosoma group, like the accepted classification of Psorophora genus. Ae. fluviatilis was placed in the Aedini tribe, but was revealed to be more related to the Haemagogus genus, a result that may have been hampered by the poor sampling of Aedes sequences. Cq. nigricans clustered with Cq. chrysonotum, both related to Mansonia. Ae. squamipennis was placed as the most external lineage of the Culicinae subfamily. The yielded topology supports the concept of monophyly of all groups and ratifies the current taxonomic classification.

Prenatal disorders and congenital Zika syndrome in squirrel monkeys.

During the Zika virus (ZIKV) outbreak in Brazil (2015-2016), the clinical manifestations associated with its infection were complex and included miscarriage and congenital malformations, not previously described. In this study, we evaluated the prenatal conditions of pregnant female squirrel monkeys (Saimiri collinsi) infected during different gestational thirds (GTs) and assessed all clinical aspects, diagnostic imaging, viremia and the immune response. In our study, 75% of the infected animals in the 1st GT group had significant clinical manifestations, such as miscarriage and prolonged viremia associated with a late immune response. Consequently, their neonates showed fetal neuropathology, such as cerebral hemorrhage, lissencephaly or malformations of the brain grooves, ventriculomegaly, and craniofacial malformations. Thus, our study demonstrated the relevance of pregnant squirrel monkeys as a model for the study of ZIKV infection in neonates due to the broad clinical manifestations presented, including the typical congenital Zika syndrome manifestations described in humans.

Histopathological lesions of congenital Zika syndrome in newborn squirrel monkeys.

The absence of an adequate animal model for studies has limited the understanding of congenital Zika syndrome (CZS) in humans during the outbreak in America. In this study, we used squirrel monkeys (Saimiri collinsi), a neotropical primate (which mimics the stages of human pregnancy), as a model of Zika virus (ZIKV) infection. Seven pregnant female squirrel monkeys were experimentally infected at three different gestational stages, and we were able reproduce a broad range of clinical manifestations of ZIKV lesions observed in newborn humans. Histopathological and immunohistochemical analyses of early-infected newborns (2/4) revealed damage to various areas of the brain and ZIKV antigens in the cytoplasm of neurons and glial cells, indicative of CZS. The changes caused by ZIKV infection were intrauterine developmental delay, ventriculomegaly, simplified brain gyri, vascular impairment and neuroprogenitor cell dysfunction. Our data show that the ZIKV infection outcome in squirrel monkeys is similar to that in humans, indicating that this model can be used to help answer questions about the effect of ZIKV infection on neuroembryonic development and the morphological changes induced by CZS.

Mitochondrial genome sequencing and phylogeny of Haemagogus albomaculatus, Haemagogus leucocelaenus, Haemagogus spegazzinii, and Haemagogus tropicalis (Diptera: Culicidae).

The genus Haemagogus (Diptera: Culicidae) comprises species of great epidemiological relevance, involved in transmission cycles of the Yellow fever virus and other arboviruses in South America. So far, only Haemagogus janthinomys has complete mitochondrial sequences available. Given the unavailability of information related to aspects of the evolutionary biology and molecular taxonomy of this genus, we report here, the first sequencing of the mitogenomes of Haemagogus albomaculatus, Haemagogus leucocelaenus, Haemagogus spegazzinii, and Haemagogus tropicalis. The mitogenomes showed an average length of 15,038 bp, average AT content of 79.3%, positive AT-skews, negative GC-skews, and comprised 37 functional subunits (13 PCGs, 22 tRNA, and 02 rRNA). The PCGs showed ATN as start codon, TAA as stop codon, and signs of purifying selection. The tRNAs had the typical leaf clover structure, except tRNASer1. Phylogenetic analyzes of Bayesian inference and Maximum Likelihood, based on concatenated sequences from all 13 PCGs, produced identical topologies and strongly supported the monophyletic relationship between the Haemagogus and Conopostegus subgenera, and corroborated with the known taxonomic classification of the evaluated taxa, based on external morphological aspects. The information produced on the mitogenomes of the Haemagogus species evaluated here may be useful in carrying out future taxonomic and evolutionary studies of the genus.

Investigation about the Occurrence of Transmission Cycles of Arbovirus in the Tropical Forest, Amazon Region.

Because of its ecological characteristics, the Caxiuanã National Forest (FLONA) is a potential area of arbovirus circulation. The present study aimed to investigate the occurrence of arbovirus transmission cycles at FLONA de Caxiuanã. Five field trips were performed to capture mosquitoes and sylvatic vertebrates. For these vertebrates, we attempted viral isolation by cell monolayer inoculation from blood, and hemagglutination inhibition and further seroneutralization assays from sera. For mosquitoes, we performed tests of viral genome detection. A total of 338 vertebrates were captured, and the greatest representative was birds (251/74.26%). A total of 16,725 mosquitoes were captured, distributed among 56 species. There were no viruses isolated by newborn mouse inoculation. Among birds, antibodies against Ilheus virus were the most prevalent. Catu virus, Caraparu virus, and Mucambo virus were the most prevalent among mammals and reptiles. Fragments of Mucambo virus, Ilheus virus, Bussuquara virus, and Rocio virus genome were detected in a pool of mosquito samples. These results of the study suggest the occurrence of arbovirus transmission cycles in the FLONA of Caxiuanã. The proximity of human populations with elements, involved in transmission cycles, makes surveillance necessary in this population to avoid dispersion of arboviruses to naïve locations.