Bat-borne coronaviruses are of increasing interest due to their potential for zoonotic spillover, particularly within the Betacoronavirus genus. Partial genomic sequences related to MERS-like CoV was identified in multiple hosts, including humans, camelids, and bats from the families Vespertilionidae and Molossidae. In this study, we investigated the presence of coronaviruses in 65 Molossus molossus bats captured in southern Brazil using nested RT-PCR and high-throughput sequencing (HTS). One individual (1.5

Background: Infectious liver diseases are frequent in domestic cats. However, the etiological diagnosis remains challenging due to the nonspecific nature of clinical and laboratory findings. Domestic cat hepadnavirus (DCHV) (Orthohepadnavirus felisdomestici) is a circular DNA virus belonging to the genus Orthohepadnavirus within the family Hepadnaviridae, recently identified in domestic and wild felines. Similar to human hepatitis B virus (HBV), DCHV has been associated with chronic hepatitis and hepatocellular carcinoma in cats. The DCHV was firstly reported in Australia in 2018 and subsequently across multiple countries and continents countries, including Brazil. DCHV has been primarily detected in serum, whole blood, and liver tissue samples. However, viral DNA has also been identified in kidney, intestine, brain, heart, and lymph node tissues, suggesting the possibility of systemic. DCHV is currently classified in genotypes A and B, where genotype A is the most frequent reported worldwide, and genotype B was only reported in Japan and Brazil. The present study aimed to detect DCHV in serum and liver in a large domestic cat population from Porto Alegre and its metropolitan area, contributing to a better understanding of this virus molecular epidemiology. Materials, Methods & Results: In total, 528 individual clinical samples were analyzed, obtained from domestic cats of different age groups and clinical histories. Of these, 451 were serum samples collected in veterinary diagnostic laboratories, and 77 were liver tissue samples from cats necropsied at the Department of Veterinary Pathology of the Federal University of Rio Grande do Sul (UFRGS). Detection of DCHV was performed using two different PCR assays, both aiming to amplify the polymerase (P) gene, including internal controls in all reactions. The second DCHV PCR was designed to amplify both genotypes A and B. None of the samples analyzed yielded a positive result for DCHV genome. This finding suggests the absence or a low frequency of DCHV in the domestic cat population from Porto Alegre metropolitan area, South Brazil. Discussion: DCHV has been identified in cats from different regions worldwide, with prevalence varying among domestic cat populations. In the present study, a comprehensive investigation of DCHV occurrence was carried out in a large domestic cats sampling from Porto Alegre and its metropolitan area using two clinical specimens (serum and liver) as well as 2 different PCR assays. DCHV DNA was not detected in any of the animals evaluated, suggesting the absence or low frequency of this virus in the here evaluated domestic cat population. These findings are consistent with previous studies reporting low DCHV detection rates in some geographic regions, including Brazil. In contrast, higher frequencies have been described in locations such as Hong Kong, Thailand, and Malaysia, indicating that regional, population-related, specific epidemiological factors may influence viral transmission dynamics. Expanded surveillance and further investigations are essential to determine whether the currently low detection frequency reflects truly limited viral circulation or arises from sampling and methodological constraints. Considering the hepatotropic potential of DCHV, the results of this study highlight the importance of continuous monitoring of this agent in feline populations. Expanded investigations across diverse epidemiological contexts will be essential to determine whether the absence of DCHV here observed reflects truly limited viral circulation or arises from methodological constraints. Keywords: Orthohepadnavirus felisdomestici, Hepadnaviridae, molecular diagnosis, hepatotropic viruses, PCR.

Bovine viral diarrhea virus (BVDV) is a major pathogen in cattle, causing significant economic losses and frequently contaminating cell cultures through fetal calf serum, which can compromise biological products. Gene editing technologies, such as CRISPR/Cas9, offer promising tools for developing disease-resistant models. CD46, the primary cellular receptor for BVDV, has an incompletely understood role in viral infection. This study aimed to: (i) characterize and compare CD46 between bovine cell lines resistant (CRIB) and susceptible (MDBK); (ii) edit the BVDV-binding region of MDBK-CD46; and (iii) evaluate the susceptibility of CD46-edited MDBK cells to BVDV infection. CD46 was sequenced in BVDV-sensitive MDBK cells, BVDV-resistant CRIB cells, and bovine fibroblasts. CRISPR/Cas9 was used to delete exon 1 of CD46, which encodes the viral attachment platform. Two guide RNAs were cloned into px458 plasmids expressing GFP and co-transfected into MDBK cells. Genomic DNA was PCR-amplified, cloned, and sequenced. Results showed that CRIB cells express a wild-type CD46 protein, suggesting that CD46 is not responsible for their resistance to BVDV. This resistance is likely due to the truncated ADAM17 in CRIB cells, which may alter membrane composition and endocytic pathways, affecting viral entry. Three edited MDBK cell lines were generated: two with homozygous deletions (A and B), and one with a heteroallelic edit (C). The edited lines A and B exhibited over 90 % resistance to BVDV. These findings support the use of gene editing to generate BVDV-resistant models and to further explore alternative viral receptors and entry pathways.

Herpesviruses are among the most significant viral pathogens that affect captive and free-ranging testudines. Moreover, herpesviruses are known to establish latent infections. In this report, we describe 2 cases of Trachemys herpesvirus infection in captive adult black-bellied sliders (Trachemys dorbigni) submitted for necropsy after a 2-week history of respiratory signs and oral lesions. Gross examination revealed severe diphtheric stomatitis, rhinitis, tracheitis, esophagitis, gastritis, and reddened lungs. Histologically, there were multifocal areas of necrosis with syncytial cells and fibrin deposition. Intranuclear amphophilic to eosinophilic inclusions were frequent in epithelial and syncytial cells, especially in the respiratory tract. Oral cavity and esophageal swabs were collected during necropsy and subjected to a multiplex nested PCR assay targeting herpesvirus DNA. The amplified DNA was sequenced and analyzed phylogenetically, confirming the virus as Trachemys herpesvirus. This is the first detailed description of clinical disease and associated lesions caused by Trachemys herpesvirus infection.

The surveillance of wildlife viromes is essential for identifying zoonotic threats within the One Health framework. This study analyzed rectal and oral swabs from 88 individuals representing 13 species as felids, wild rodents, marsupials and non-human primates in Southern Brazil using metagenomic sequencing. Akodon montensis (n = 15 individuals) and Coendou spinosus (n = 4) harbored Chikungunya virus (ChikV, Togaviridae), marking its first detection in these hosts. Molossus molossus (n = 17) presented Coronaviridae and Orthoherpesviridae, while Eptesicus furinalis (n = 1) also carried Coronaviridae. A broad virome diversity, including Togaviridae and Adenoviridae members, was identified in Didelphis albiventris (n = 43), with significant relevance to human health. Additional species, such as Callithrix jacchus (n = 1), Leopardus guttulus (n = 1), Myocastor coypus (n = 1), Monodelphis iheringi (n = 1), Thaptomys nigrita (n = 1), Sooretamys angouya (n = 1), Brucepattersonius iheringi (n = 1), and Lasiurus blossevillii (n = 1), contributed to insights into viral reservoirs. These results underscore the importance of virome studies in regions harboring high biodiversity, emphasizing genomic surveillance as a vital tool for monitoring zoonotic viruses and safeguarding global health.

Hepacivirus is a genus of RNA viruses within the family Flaviviridae of which hepatitis C virus (HCV) is the prototype. Several hepaciviruses have been identified in mammals, including rodents of multiple families. Each rodent hepacivirus described so far has been found only in members of a single rodent species. Here, we report the discovery and characterization of a putative new genotype of an unclassified rodent hepacivirus in a wild cavy (Cavia aperea aperea) that was reported previously in Proechimys semispinosus. This virus was detected in one out of 14 (7.14%) wild cavy sera tested by RT-PCR. The complete genome sequence was obtained by high-throughput sequencing using an Illumina MiSeq platform. This is the first report of a hepacivirus in a member of the family Caviidae. Our findings show that members of different rodent species and even families can be infected by hepaciviruses of the same species. The identification and characterization of novel hepaciviruses might lead to the discovery of reservoirs of viruses that are genetically related to human pathogens, and this can help to elucidate the evolutionary origins of HCV and other hepaciviruses.

Domestic cat hepadnavirus (DCH) (Orthohepadnavirus felisdomestici) is an emerging virus related to the hepatitis B virus (HBV) already reported in many countries. The molecular prevalence of DCH varies widely in the regions investigated so far. In the present work, we reported the presence of DCH in Brazil. Sixty cat serum samples tested by DCH presence using PCR and 1.67% (1/60) were positive, similar to the low positive molecular rates reported in United States and Japan. The DCH full-length genome was classified in genotype B, which is uncommon since this genotype was only reported once in Japan. The DCH-positive sample was obtained in a stray cat female apparently healthy, presenting ALT, AST, and ALKP normal values, and negative for FIV and FeLV. Due the low positivity rate detected, some factors as alteration in hepatic enzymes and FIV/FeLV infection could not be evaluated. Other works are necessary to statistically validate these observations in Brazil.

Bat-borne viruses may affect public health and the global economy. These mammals have a wide geographical distribution and unique biological, physiological, and immunogenic characteristics, allowing the dissemination of many known and unknown viruses. Enteric viruses, such as adeno (AdV) and rotaviruses, are recognized as the main causative agents of disease and outbreaks. In the present study, the presence of viruses from Adenoviridae and Reoviridae families was evaluated in molossid, phyllostomid, and vespertilionid bats captured in Rio Grande do Sul, Southern Brazil, between September 2021 and July 2022. Sixty bat rectal swabs were analyzed by PCR. Eight (13.3%) samples were positive for adenovirus and classified as human mastadenovirus C (HAdV-C) (three samples) and HAdV-E (five samples) by sequencing followed by phylogenetic analysis. All samples were negative in rotavirus specific RT-PCR. This is the first study to describe the presence of HAdV in samples of Glossophaga soricina, Eptesicus brasiliensis, and Histiotus velatus. Furthermore, the presence of HAdV-E in bats was reported, which is unusual and may suggest that other HAdV genotypes, in addition to HAdV-C, may also be harbored by wild animals. The data generated in the present study reinforces the importance of eco-surveillance of viral agents related to diseases in humans and wild animals. In addition, it is essential to identify possible new hosts or reservoirs that increase the risk of spillover and dissemination of infectious pathogens, helping to prevent and control zoonotic diseases.

Bats are mammals with high biodiversity and wide geographical range. In Brazil, three haematophagous bat species are found. Desmodus rotundus is the most documented due to its role as a primary host of rabies virus in Latin America. Bats are known to harbor various emerging viruses causing severe human diseases. Beyond zoonotic viruses, these animals also harbor a diversity of non-zoonotic viruses. Papillomaviruses are circular double-stranded deoxyribonucleic acid (dsDNA) viruses that infect the epithelial and mucosal cells of many vertebrates, occasionally causing malignant lesions. High-throughput sequencing has enabled papillomaviruses discovery in different bat species. Here, 22 D. rotundus samples were collected through the rabies eradication program in Rio Grande do Sul. The DNA extracted from pooled intestines was amplified by the rolling-circle amplification (RCA) method and sequenced using the Illumina® MiSeq platform (San Diego, CA, USA).Analysis revealed three contigs corresponding to the Papillomaviridae family, representing three novel viruses named DrPV-1, DrPV-2, and DrPV-3. Phylogenetic analysis suggests DrPV-1 may constitute a new species within the Dyophipapillomavirus genus, while DrPV-2 and DrPV-3 may represent different types within the same species from a novel genus. This is the first description of a papillomavirus in the D. rotundus species, contributing to the characterization of PVs in the Chiropteran order.

The conjunctiva plays an essential role in eye health and immunity and acts as a barrier to the entry of microorganisms. Conjunctival infections are common in dogs and result from both the invasion of pathogenic microorganisms and the uncontrolled growth of the existing microbiota. Most of the existing data come from studies based on traditional culture methods. These reports indicate the predominance of gram-positive bacteria, especially Staphylococcus spp. In the present study, we analyzed the microbiota present on the conjunctival surface from a heterogeneous dog population without ophthalmological disorders using DNA sequencing. After a thorough ophthalmological examination, conjunctival swabs were collected from both eyes of 30 dogs. After processing and nucleic acid extraction, the sample pool was subjected to shotgun DNA sequencing through the Illumina platform and analyzed via the Metagenomic Rapid Annotations using Subsystems Technology (MGRAST) server. A predominance of the phylum Proteobacteria and the genera Ralstonia and Burkholderia were identified along with a minority of fungi, whereas viruses were not found. Microbial DNA sequencing has provided new data on this subject, revealing the presence of noncultivable organisms that were previously unknown as part of the ocular microbiome.

Resumo A conjuntiva desempenha papel fundamental na saúde e imunidade ocular e atua como uma barreira à entrada de microrganismos. As infecções conjuntivais são comuns em cães e resultam tanto da invasão de microrganismos patogênicos quanto do crescimento descontrolado da microbiota existente. A maior parte dos dados existentes provém de estudos baseados em métodos de cultura tradicionais. Esses relatos apontam para o predomínio de bactérias gram-positivas, principalmente Staphylococcus spp. O presente estudo analisou a comunidade microbiana presente na conjuntiva ocular de uma população heterogênea de cães sem distúrbios oftalmológicos por sequenciamento de DNA. Após exame oftálmico minucioso, foram coletados suabe conjuntivais de ambos os olhos de 30 cães. Após processamento e extração de ácidos nucleicos, o pool de amostras foi submetido ao sequenciamento shotgun de DNA por meio da plataforma Illumina e analisado no servidor Metagenomic Rapid Annotations using Subsystems Technology (MG-RAST). Foi identificada uma predominância do filo Proteobacteria e dos gêneros Ralstonia e Burkholderia juntamente com uma minoria de fungos, enquanto vírus não foram encontrados. O sequenciamento do DNA microbiano trouxe novos dados sobre o assunto, revelando a presença de organismos não cultiváveis até então desconhecidos como parte do microbioma ocular.

Animals are common hosts for many coronaviruses where bats and rodents are commonly regarded as primary reservoirs. The unquestionable emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a yet unknown animal host - in addition with reports of further anthropogenic spread and sustained transmission in mustelids, captive felids, and domestic dogs and cats owned by people previously tested as SARS-CoV-2-positive, rise some concerns about possible continuous maintenance of the virus in nature and domestic species. In this review, we discussed the current data about coronaviruses in domestic and farming animals, recombination events, animal species susceptibility, virus-cell receptor interactions, and clinical signs of most relevant Coronaviridae members of each genera. Also, we present what is known about SARS-CoV-2 in animals and what will be the potential role of those species in COVID-19 epidemiology. Apparently, the virus can infect pets on some occasions, where cats look to be more susceptible than dogs. Thus, pet infection by sick owners is not only likely but expected given the numerous opportunities for spill-over during a massive outbreak. Regarded to farm animals, attention should be focused on breeding species of the Mustelidae family since they are those that have been shown to be more susceptible in experimental infections and have also effectively exhibited animal-to-human transmission. Other intensively bred species such as poultry, swine, horses and ruminants seem to present little or no epidemiological risk so far. The continuous monitoring of SARS-CoV-2 in animals in close contact with people with COVID-19 may be a key in the understanding of this emergent disease and the animal’s role in epidemiology in the future. It is possible that some species will serve as important reservoirs and source of infection of COVID-19 for humans making it re-emergent in the future, as is theoretically proposed for the origin of SARS-CoV-2.

Pestiviruses are globally distributed and cause substantial economic losses to the cattle industry. In Brazil, the country with the world’s largest cattle population, pestivirus infections are well described in some regions, such as in the south, where a high frequency of BVDV-2 is described and contrasts with the high prevalence of HoBi-like pestivirus (HoBiPeV) in the northeast. However, there is a lack of information about pestiviruses in the Amazon Region, in northern Brazil, with a cattle population estimated at 55.7 million head, which has a significant impact on the international livestock market. Therefore, this study investigated the seroprevalence and genetic variability of ruminant pestiviruses in 944 bovine serum samples from four states in northern Brazil: Pará (PA), Amapá (AP), Roraima (RR), and Amazonas (AM). Our results showed that 45.4% of the samples were seropositive (19.8% for BVDV-1, 14.1% for BVDV-2, and 20.9% for HoBiPeV). All samples were tested by RT–qPCR, and three were positive and classified as HoBiPeV in a phylogenetic analysis. These serological and molecular results contrast with those from other regions of the world, suggesting that the northern Brazilian states have a high prevalence of all bovine pestiviruses including HoBiPeV.

Arboviral diseases comprise a group of important infectious diseases imposing a heavy burden to public health in many locations of the world. Orthoflaviviruses are viruses belonging to the genus Orthoflavivirus; this genus includes some of the most relevant arboviruses to human health. Orthoflaviviruses can infect several different hosts, with some species being transmitted in cycles involving birds and anthropophilic mosquitoes and others transmitted between mammals and mostly Aedes sp. mosquitoes. Some of the most important sylvatic reservoirs of orthoflaviviruses are non-human primates (NHPs). Many flaviviruses that infect NHPs in nature have the potential to cause epidemics in humans, as has been observed in the cases of Orthoflavivirus denguei (dengue virus - DENV), Orthoflavivirus flavi (yellow fever virus - YFV), and Orthoflavivirus zikaense (Zika virus - ZIKV). In this minireview, we discuss important aspects regarding history, ecology involving NHP, distribution, disease outcome, and pathogenesis of these three major orthoflaviviruses that affect humans and NHP and relate this information to the potential of using NHP as experimental models. In addition, we suggest some orthoflaviviruses that could be better investigated, both in nature and in experimental studies, in light of the recent revolution in molecular biology.

Chiroptera are one of the most diverse mammal orders. They are considered reservoirs of main human pathogens, where coronaviruses (CoVs) and paramyxoviruses (PMVs) may be highlighted. Moreover, the growing number of publications on CoVs and PMVs in wildlife reinforces the scientific community’s interest in eco-vigilance, especially because of the emergence of important human pathogens such as the SARS-CoV-2 and Nipha viruses. Considering that Brazil presents continental dimensions, is biologically rich containing one of the most diverse continental biotas and presents a rich biodiversity of animals classified in the order Chiroptera, the mapping of CoV and PMV genetics related to human pathogens is important and the aim of the present work. CoVs can be classified into four genera: Alphacoronavirus, Betacoronavirus, Deltacoronavirus and Gammacoronavirus. Delta- and gammacoronaviruses infect mainly birds, while alpha- and betacoronaviruses contain important animal and human pathogens. Almost 60% of alpha- and betacoronaviruses are related to bats, which are considered natural hosts of these viral genera members. The studies on CoV presence in bats from Brazil have mainly assayed phyllostomid, molossid and vespertilionid bats in the South, Southeast and North territories. Despite Brazil not hosting rhinophilid or pteropodid bats, which are natural reservoirs of SARS-related CoVs and henipaviruses, respectively, CoVs and PMVs reported in Brazilian bats are genetically closely related to some human pathogens. Most works performed with Brazilian bats reported alpha-CoVs that were closely related to other bat-CoVs, despite a few reports of beta-CoVs grouped in the Merbecovirus and Embecovirus subgenera. The family Paramyxoviridae includes four subfamilies (Avulavirinae, Metaparamyxovirinae, Orthoparamyxovirinae and Rubulavirinae), and bats are significant drivers of PMV cross-species viral transmission. Additionally, the studies that have evaluated PMV presence in Brazilian bats have mainly found sequences classified in the Jeilongvirus and Morbillivirus genera that belong to the Orthoparamyxovirinae subfamily. Despite the increasing amount of research on Brazilian bats, studies analyzing these samples are still scarce. When surveying the representativeness of the CoVs and PMVs found and the available genomic sequences, it can be perceived that there may be gaps in the knowledge. The continuous monitoring of viral sequences that are closely related to human pathogens may be helpful in mapping and predicting future hotspots in the emergence of zoonotic agents.

OBJECTIVES:The HIV-1 genetic diversity and the presence of transmitted drug resistance mutations (TDRMs) against integrase strand transfer inhibitors (INSTIs) were assessed sequencing samples of antiretroviral (ARV)-naive HIV-1-infected individuals from South Brazil.METHODS:Viral RNA from 42 ART-naive individuals was submitted to complete HIV-1 integrase gene amplification by RT-PCR and sequencing.RESULTS:Viral strains carrying TDRMs against INSTIs were not detected in the present study. However, the polymorphisms L74M and L74I were each observed in 4.8% of the individuals. These accessory mutations have been reported as putative causes of TDRMs in ART with raltegravir, but only when associated with additional major mutations. When submitted to HIV-1 subtyping, 50% were classified as subtype C, 21% as recombinant BC, 19% as subtype B, 4.8% as subtype F1 and 4.8% as recombinant CF1.CONCLUSIONS:All 42 ARV-naive individuals were apparently susceptible to INSTIs, included in the Brazilian therapeutic guideline since 2009. To the best of our knowledge, this is the first study to evaluate TDRMs against INSTIs in Brazil. The most prevalent HIV-1 subtypes were subtype C, followed by the recombinant BC and subtype B, which is in agreement with previous studies. However, the presence of subtype F1 and recombinant CF1 reported herein was not observed in previous studies.

Background: SARS-CoV-2, the virus that causes COVID-19, is constantly mutating, leading to new variants that culminate in a temporal lineages fluctuation. B.1.1.28 lineage has been evolving in Brazil since February 2020 and originated P.1 (VOC), P.2 (VOI) and other P.Xs proposed as new variants. Methods and results: In this study, through the Illumina platform, we performed the whole-genome sequencing of 26 positive samples of SARS-CoV-2. Employing variant calling analysis on FASTQ reads and phylogenetic inference, we report a brief dispersion of a potentially new B.1.1.28-derived variant detected between 2021 May and June in individuals crossing the border between Brazil and Argentina, and local spread to inpatients from hospitals at the Rio Grande do Sul state capital (Porto Alegre). Besides, the Rio Grande do Sul State SARS-CoV-2 genomic epidemiological data was analyzed and showed an important B.1.1.28 peak in RS at the same period (May-June), even in the presence of a major Gamma wave. Conclusions: The emergence of a putative B.1.1.28-derived lineage was identified in travelers crossing Brazil-Argentina border representing an important peak of B.1.1.28 in RS State with a decreased in Gamma variant frequency in the same period of time.

Zoonotic spillover is a phenomenon characterized by the transfer of pathogens between different animal species. Most human emerging infectious diseases originate from non-human animals, and human-related environmental disturbances are the driving forces of the emergence of new human pathogens. Synthesizing the sequence of basic events involved in the emergence of new human pathogens is important for guiding the understanding, identification, and description of key aspects of human activities that can be changed to prevent new outbreaks, epidemics, and pandemics. This review synthesizes the connections between environmental disturbances and increased risk of spillover events based on the One Health perspective. Anthropogenic disturbances in the environment (e.g., deforestation, habitat fragmentation, biodiversity loss, wildlife exploitation) lead to changes in ecological niches, reduction of the dilution effect, increased contact between humans and other animals, changes in the incidence and load of pathogens in animal populations, and alterations in the abiotic factors of landscapes. These phenomena can increase the risk of spillover events and, potentially, facilitate new infectious disease outbreaks. Using Brazil as a study model, this review brings a discussion concerning anthropogenic activities in the Amazon region and their potential impacts on spillover risk and spread of emerging diseases in this region.

EDITORIAL article Front. Vet. Sci., 18 October 2022Sec. Veterinary Epidemiology and Economics Volume 9 - 2022 | https://doi.org/10.3389/fvets.2022.1025314

The emergence of Variants of Concern (VOC) presenting an unusual number of new mutations is one of the most remarkable features of SARS-CoV-2. The Delta variant, since its appearance, replaced the VOC Gamma, which was responsible for the major COVID-19 wave in Brazil. In this study, we performed a Delta whole-genome sequencing of 183 samples as part of a major genomic surveillance study performed since the beginning of the pandemic. Here, we showed an emergence, widespread dispersion and consolidation of the Delta variant in Rio Grande do Sul State, completely replacing the Gamma variant in a four to five months period. Performing the phylogenetic and phylodynamic analysis, the majority of the sequences generated herein were classified as AY.99.2, AY.99.2-like and AY.101. AY.99.2 Delta-related lineage has been widely reported in Brazil and in the Americas as well. Altogether, our findings provided a mutational profile of the sequences and presented high substitutions per site in the root-to-tip phylogenetic tree, corroborating studies that show the high mutational rate of SARS-CoV-2 over time.