Geovani de Oliveira Ribeiro

The Amapa State, Brazil, in the Amazon region and bordering French Guiana, is a strategic sentinel location for infectious disease surveillance. Ecological pressures such as deforestation, artisanal gold mining, and high human mobility contribute to complex transmission dynamics of arboviruses, bacterial infections, and other pathogens. This study aims to characterize the etiologies of acute febrile illness (AFI) reported in Amapa State in 2018-2020, focusing on arbovirus circulation, bacterial contributions, and emerging pathogens. A retrospective analysis was carried out with 727 AFI cases. Laboratory diagnoses were established using molecular and serological assays, with systematic testing for different pathogens. Definitive diagnoses were obtained in 185 individuals (25.4%), yielding 209 positive results and including 23 cases of co-infection. Dengue virus was the predominant etiological agent (n=109, 52%), followed by leptospirosis (n=20, 9.5%), chikungunya virus (n=16, 7.6%), parvovirus B19 (n=16, 7.6%), measles (n=15, 7.1%), zika virus (n=10, 4.8%), mayaro virus (n=1,0.5%), and other bacterial infections (n=5, 2.4%). Isolated cases of hepatitis A and B, malaria, SARS-CoV-2, enterovirus, and acute Chagas disease were also identified. No hepatitis E virus or oropouche cases were detected. Dengue virus was the leading cause of AFI. Leptospirosis, chikungunya virus, and parvovirus B19 were other prevalent and often underrecognized contributors. The detection of additional viruses highlights Amapa State as a potential zone of “silent expansion” for emerging arboviruses. Co-detection of multiple pathogens underscores diagnostic complexity in regions with overlapping transmission. Strengthening surveillance, expanding laboratory capacity, and prioritizing differential diagnostics are essential to capture epidemiological dynamics and guide public health interventions.

Several viruses belonging to the order Mononegavirales are recognized as highly pathogenic in humans and are often associated with lethal disease outcomes. Prominent examples include human respiratory syncytial virus (HRSV), Ebola virus (EBOV), rabies virus (RABV), and Marburg virus (MARV). In addition to their clinical severity, these viruses are considered potential biological threats due to their high transmissibility and virulence. In this study, we performed a metatranscriptomic analysis of Culex and Anopheles mosquitoes collected at the DBB (Diretoria de Biodiversidade e Biotecnologia), São Paulo, Brazil. Our analysis revealed viral sequences associated with two families within Mononegavirales : Lispiviridae and Rhabdoviridae . Phylogenetic analysis of the Lispiviridae sequences identified six variants, designated CxLispV-SP_03 , CxLispV-SP_09 , CxLispV-SP_12 , CxLispV-SP_13 , CxLispV-SP_14 , and CxLispV-SP_15 , that formed a strongly supported monophyletic clade with Canya virus, suggesting the discovery of a putative novel viral lineage. Examination of RNA‐dependent RNA polymerase (RdRp) domains in these sequences confirmed the presence of essential catalytic motifs. In contrast, the detected rhabdoviruses exhibited greater genomic structural heterogeneity, including the presence of accessory protein domains. Pairwise comparisons based on RdRp amino acid identity delineated four distinct groups among these sequences, pointing to substantial evolutionary divergence within the sampled rhabdovirus diversity. Together, these findings contribute to the growing characterization of mosquito‐associated mononegaviruses and provide a genomic foundation to support future surveillance efforts and public health risk assessments related to vector‐borne viruses.

The oxygen affinity of fish hemoglobins is regulated by organic phosphates, adjusting oxygen uptake and delivery. While adenosine triphosphate (ATP) and guanosine triphosphate (GTP) are common regulators, inositol pentaphosphate (IP ₅ ) is unique to certain fish and birds. The pirarucu ( Arapaima gigas (Schinz, 1822)), an air-breathing Amazonian fish, transitions from gill respiration to air-breathing, accompanied by changes in Hb-O2 affinity regulation. This study examined IP ₅ biosynthesis in pirarucu erythrocytes across development. Organic phosphate levels (ATP, GTP, IP ₅ ) and expression of genes involved in inositol phosphate metabolism ( ipmk, itpk-1, ip3k) were analyzed in aquatic (9.86 ± 0.38 cm) and air-breathing (60.2 ± 3.8 cm) specimens. Air-breathing pirarucu had higher erythrocyte IP ₅ and lower ATP and GTP. Gene expression analysis showed increased ipmk, itpk-1, and ip3k, suggesting regulatory rather than molecular changes. Phylogenetic analysis confirmed conservation of these genes. This ontogenetic increase in IP ₅ , along with the upregulation of genes involved in its synthesis, highlights a genetic and cellular adaptation that may contribute to improved oxygen regulation during aerial respiration in A. gigas.

Metagenomic studies of mosquito viromes demonstrated a more diverse composition than just an exclusive composition of pathogenic arboviruses transmitted to humans. In our study, the virome of 866 female mosquitoes collected throughout 2020 at the São Paulo Zoo, located in the city of São Paulo/SP—Brazil, was obtained. Specifically, in this paper, we describe a new virus found by viral RNA extraction and next-generation MiSeq sequencing of a group of 23 specimens of Anopheles (Nys.) strodei. The complete genome with a length of 9709 nucleotides was characterized by a positive orientation and a single strand, with a single large ORF, which encodes a polyprotein of 2987 amino acids. The phylogenetic analysis showed an association with the viral family Iflaviridae and the Riboviria realm. We carried out comparisons with translated sequences of the capsid regions of other iflavirus, and the identities in relation to our sequence were below the minimum limit of 90%, indicating that possibly it is a new species of iflavirus. Our findings contribute to expanding knowledge of virome composition among mosquito species in Brazil and globally. Moreover, we provide a viral genome reference specific to this geographic region and Culicidae family of mosquitoes. This resource facilitates future in silico recognition and assembly of viral genomes within metagenomic datasets.

There is a growing interest in phages as potential biotechnological tools in human health owing to the antibacterial activity of these viruses. In this study, we characterized a new member (named PhiV_005_BRA/2016) of the recently identified phage species Phietavirus Henu 2. PhiV_005_BRA/2016 was detected through metagenomic analysis of stool samples of individuals with acute gastroenteritis. PhiV_005_BRA/2016 contains double-stranded linear DNA (dsDNA), it has a genome of 43,513 base pairs (bp), with a high identity score (99%) with phage of the genus Phietavirus, species of Phietavirus Henu 2. Life style prediction indicated that PhiV_005_BRA/2016 is a lysogenic phage whose the main host is methicillin-resistant Staphylococcus aureus (MRSA). Indeed, we found PhiV_005_BRA/2016 partially integrated in the genome of distinct MRSA strains. Our findings highlights the importance of large-scale screening of bacteriophages to better understand the emergence of multi-drug resistant bacterial.

The genus totivirus in the family Totiviridae contains double-stranded RNA viruses. Their genome has two open reading frames (ORFs) that encode capsid protein (CP) and RNA-dependent RNA polymerase (RdRp). The toti-like viruses recently identified in Anopheles sp. and Aedes aegypti mosquitoes (AaTV) share the same genome organization as other totiviruses. The AaTVs that have been described in distinct geographical regions are monophyletic. In this study, we show that AaTV sequences can be grouped into at least three phylogenetic clades (named A, B, and C). Clades A and B are composed of AaTV sequences from mosquitoes collected in the Caribbean region (Guadeloupe), and clade C contains sequences from the USA. These clades may represent AaTV lineages that are locally adapted to their host populations. We also identified three recombinant AaTV strains circulating in mosquitoes in Guadeloupe. Although these strains have different chimeric patterns, the position of the recombination breakpoint was identical in all strains. Interestingly, this breakpoint is located in a hairpin-like structure in the intergenic region of the AaTV genome. This RNA structure may stall RNA polymerase processivity and consequently induce template switching. In vitro studies should be conducted to further investigate the biological significance of AaTV’s intergenic region as a recombination hotspot

The simultaneous transmission of two lineages of the chikungunya virus (CHIKV) was discovered after the pathogen’s initial arrival in Brazil. In Oiapoque (Amapá state, north Brazil), the Asian lineage (CHIKV-Asian) was discovered, while in Bahia state, the East-Central-South-African lineage (CHIKV-ECSA) was discovered (northeast Brazil). Since then, the CHIKV-Asian lineage has been restricted to the Amazon region (mostly in the state of Amapá), whereas the ECSA lineage has expanded across the country. Despite the fact that the Asian lineage was already present in the Amazon region, the ECSA lineage brought from the northeast caused a large outbreak in the Amazonian state of Roraima (north Brazil) in 2017. Here, CHIKV spread in the Amazon region was studied by a Zika–Dengue–Chikungunya PCR assay in 824 serum samples collected between 2013 and 2016 from individuals with symptoms of viral infection in the Amapá state. We found 11 samples positive for CHIKV-Asian, and, from these samples, we were able to retrieve 10 full-length viral genomes. A comprehensive phylogenetic study revealed that nine CHIKV sequences came from a local transmission cluster related to Caribbean strains, whereas one sequence was related to sequences from the Philippines. These findings imply that CHIKV spread in different ways in Roraima and Amapá, despite the fact that both states had similar climatic circumstances and mosquito vector frequencies.

Metagenomics based on the next-generation sequencing (NGS) technique is a target-independent assay that enables the simultaneous detection and genomic characterization of all viruses present in a sample. There is a limited amount of data about the virome of individuals with gastroenteritis (GI). In this study, the enteric virome of 250 individuals (92% were children under 5 years old) with GI living in the northeastern and northern regions of Brazil was characterized. Fecal samples were subjected to NGS, and the metagenomic analysis of virus-like particles (VLPs) identified 11 viral DNA families and 12 viral RNA families. As expected, the highest percentage of viral sequences detected were those commonly associated with GI, including rotavirus, adenovirus, norovirus (94.8%, 82% and 71.2%, respectively). The most common co-occurrences, in a single individual, were the combinations of rotavirus-adenovirus, rotavirus-norovirus, and norovirus-adenovirus (78%, 69%, and 62%, respectively). In the same way, common fecal-emerging human viruses were also detected, such as parechovirus, bocaporvirus, cosavirus, picobirnavirus, cardiovirus, salivirus, and Aichivirus. In addition, viruses that infect plants, nematodes, fungi, protists, animals, and arthropods could be identified. A large number of unclassified viral contigs were also identified. We show that the metagenomics approach is a powerful and promising tool for the detection and characterization of different viruses in clinical GI samples.

Dengue virus (DENV) is a mosquito-borne viral pathogen that plagues many tropical-climate nations around the world, including Brazil. Molecular epidemiology is a growing and increasingly invaluable tool for understanding the dispersal, persistence, and diversity of this impactful virus. In this study, plasma samples (n = 824) from individuals with symptoms consistent with an arboviral febrile illness were analyzed to identity the molecular epidemiological dynamics of DENV circulating in the Brazilian state of Amapá. Twelve DENV type 1 (DENV-1) genomes were identified, which were phylogenetically related to the BR4 lineage of genotype V. Phylodynamics analysis suggested that DENV-1 BR-4 was introduced into Amapá around early 2010, possibly from other states in northern Brazil. We also found unique amino acids substitutions in the DENV-1 envelope and NS5 protein sequences in the Amapá isolates. Characterization of the DENV-1 BR-4 sequences highlights the potential of this new lineage to drive outbreaks of dengue in the Amazon region.

Echoviruses (E) are a diverse group of viruses responsible for various pathological conditions in humans including aseptic meningitis, myocarditis, and acute flaccid paralysis. The detection and identification of echovirus genotypes in clinical samples is challenging due to its high genetic diversity. Here, we report the complete genome sequences of nine echoviruses, obtained by next-generation sequencing of 238 fecal samples from individuals with gastroenteritis in regions of Brazil. Detected viruses were classified into six genotypes: Three E1 sequences (BRA/TO-028, BRA/TO-069 and BRA/TO-236), one E3 (BRA/TO-018), one E11 (BRA/TO-086), one E20 (BRA/TO-016), two E29 (BRA/TO-030 and BRA/TO-193), and one E30 sequence (BRA/TO-032). Phylogenetic analysis indicated that the echoviruses E1 and E29 circulating in Brazil are divergent from strains circulating worldwide. The genotype diversity identified in our study may under-represent the total echovirus diversity in Brazil because of the small sample size and the restricted geographical distribution covered by the survey.