Background and common lesions in the female reproductive organs of giant anteaters (Myrmecophaga tridactyla).

The giant anteater (Myrmecophaga tridactyla) is a vulnerable species in South America and is considered endangered or near extinction in Central America. Therefore, studies describing the reproductive characteristics of this species are pivotal for its conservation. Thus, this study aimed to provide a morphological description of the female reproductive tissues of this species. We collected tissue samples from six female giant anteaters and performed gross, morphological, and histochemical analyses. Five adult subjects and one juvenile were included in the study. In the ovary, classifications were made according to the follicle and oocyte sizes: primordial, primary, secondary, early antral, or antral. Typical follicles with a single oocyte surrounded by a simple or stratified layer of cubic epithelium, atretic follicles, corpora lutea, corpora albicans, and ovarian cysts were also observed. No ovarian lesions were observed. By contrast, endometritis, metritis, mucometra, and endometrial cysts were identified in the uterus. Uterine alterations in these subjects were frequent and could affect reproduction.

Molecular Detection and Characterization of Mycoplasma spp. in Marine Mammals, Brazil.

Mycoplasma spp. are wall-less bacteria able to infect mammals and are classified as hemotropic (hemoplasma) and nonhemotropic. In aquatic mammals, hemoplasma have been reported in California sea lions (Zalophus californianus) and river dolphins (Inia spp.). We investigated Mycoplasma spp. in blood samples of West Indian manatees (Trichechus manatus), pinnipeds (5 species), and marine cetaceans (18 species) that stranded or were undergoing rehabilitation in Brazil during 2002-2022. We detected Mycoplasma in blood of 18/130 (14.8%) cetaceans and 3/18 (16.6%) pinnipeds. All tested manatees were PCR-negative for Mycoplasma. Our findings indicate that >2 different hemoplasma species are circulating in cetaceans. The sequences from pinnipeds were similar to previously described sequences. We also detected a nonhemotropic Mycoplasma in 2 Franciscana dolphins (Pontoporia blainvillei) that might be associated with microscopic lesions. Because certain hemoplasmas can cause disease and death in immunosuppressed mammals, the bacteria could have conservation implications for already endangered aquatic mammals.

Naturally Acquired Rabies in White-Eared Opossum, Brazil.

Opossums are considered resistant to rabies. Nonhematophagous bats are reservoirs of rabies in urban areas of South America. We analyzed bats and opossums tested for rabies during 2021 in a highly urbanized city in Brazil to understand spillover in an urban setting. Wildlife surveillance is necessary to prevent rabies in humans and domestic animals.

Detection of herpesviruses in neotropical primates from São Paulo, Brazil.

Transmission of herpesvirus between humans and non-human primates represents a serious potential threat to human health and endangered species conservation. This study aimed to identify herpesvirus genomes in samples of neotropical primates (NTPs) in the state of São Paulo, Brazil. A total of 242 NTPs, including Callithrix sp., Alouatta sp., Sapajus sp., and Callicebus sp., were evaluated by pan-herpesvirus polymerase chain reaction (PCR) and sequencing. Sixty-two (25.6%) samples containing genome segments representative of members of the family Herpesviridae, including 16.1% for Callitrichine gammaherpesvirus 3, 6.1% for Human alphaherpesvirus 1, 2.1% for Alouatta macconnelli cytomegalovirus, and 0.83% for Cebus albifrons lymphocryptovirus 1. No co-infections were detected. The detection of herpesvirus genomes was significantly higher among adult animals (p = 0.033) and those kept under human care (p = 0.008671). These findings confirm the importance of monitoring the occurrence of herpesviruses in NTP populations in epizootic events.

Herpesvirus and adenovirus surveillance in threatened wild West Indian (Trichechus manatus) and Amazonian manatees (Trichechus inunguis), Brazil.

THE FAMILY TRICHECHIDAE (ORDER SIRENIA) COMPRISES THREE SPECIES: African (Trichechus senegalenses), West Indian (T. manatus), WIM)], and the Amazonian manatees (T. inunguis, AMM). Whereas WIM inhabits both riverine and coastal systems in the western Atlantic, AMM is the only exclusively freshwater sirenian, endemic to the Amazon River Basin. The study of infectious agents is essential to species conservation, especially considering that both species are classified as Vulnerable by the IUCN Red List and as Endangered by the Brazilian Red List. The current knowledge about viral agents in sirenians is scarce. Herpesviruses and adenovirus are DNA viruses able to infect and cause disease in a wide range of hosts. Herein, we used panPCR protocols to survey herpesvirus and adenovirus in blood samples of wild WIM (n = 23) and AMM (n = 26) under human care in Brazil. Herpesvirus DNA was detected in one juvenile female WIM (1/23; 4.3%; 95% CI -4.7 - 13.3) from Ceará state and in four AMM (two juvenile females, a juvenile male, and an adult female; 4/26; 15.4%; 95% CI 0.5 - 30.3) from Amazonas state. The two different gammaherpesvirus DNA polymerase sequence types identified (one per species, a sequence type in a WIM and another one in three AMM) were highly similar (99% nucleotide identity) to Trichechid herpesvirus 1, reported in West Indian manatees of Florida (USA), and 100% identical when translated into amino acids. A herpesviral glycoprotein B sequence was identified in two AMM. None of the samples was positive to adenovirus. To the authors' knowledge, this is the first herpesvirus detection in manatees from South America, expanding the herpesvirus geographical range, and the first in WIM and AMM worldwide. Our findings suggest (i) that West Indian and Amazonian manatees are possibly the natural hosts of the detected herpesvirus, and (ii) coevolution of that gammaherpesvirus with Trichechus. Future studies are necessary to characterize the obtained virus and elucidate potential pathological effects (if any) in these species.

World Health Organization critical priority Escherichia coli clone ST648 in magnificent frigatebird (Fregata magnificens) of an uninhabited insular environment.

Antimicrobial resistance is an ancient natural phenomenon increasingly pressured by anthropogenic activities. Escherichia coli has been used as markers of environmental contamination and human-related activity. Seabirds may be bioindicators of clinically relevant bacterial pathogens and their antimicrobial resistance genes, including extended-spectrum-beta-lactamase (ESBL) and/or plasmid-encoded AmpC (pAmpC), in anthropized and remote areas. We evaluated cloacal swabs of 20 wild magnificent frigatebirds (Fregata magnificens) of the Alcatrazes Archipelago, the biggest breeding colony of magnificent frigatebirds in the southern Atlantic and a natural protected area with no history of human occupation, located in the anthropized southeastern Brazilian coast. We characterized a highly virulent multidrug-resistant ST648 (O153:H9) pandemic clone, harboring bla CTX-M-2, bla CMY-2, qnrB, tetB, sul1, sul2, aadA1, aac(3)-VIa and mdfA, and virulence genes characteristic of avian pathogenic (APEC) (hlyF, iroN, iss, iutA, and ompT) and other extraintestinal E. coli (ExPEC) (chuA, kpsMII, and papC). To our knowledge, this is the first report of ST648 E. coli co-producing ESBL and pAmpC in wild birds inhabiting insular environments. We suggest this potentially zoonotic and pathogenic lineage was likely acquired through indirect anthropogenic contamination of the marine environment, ingestion of contaminated seafood, or by intra and/or interspecific contact. Our findings reinforce the role of wild birds as anthropization sentinels in insular environments and the importance of wildlife surveillance studies on pathogens of critical priority classified by the World Health Organization.

Erysipelas in a stranded common bottlenose dolphin: first report in a South American odontocete.

Erysipelas is a zoonotic disease caused by Erysipelothrix rhusiopathiae. In cetaceans, this disease has two main clinical forms: a cutaneous one, grossly characterized by rhomboid lesions, and a septicemic and often fatal form. Erysipelas is considered an important cause of morbidity and mortality in captive cetaceans; however, information in free-ranging cetaceans is limited. An adult common bottlenose dolphin (Tursiops truncatus) was found dead and in advanced autolysis in Paraíba state, northeastern Brazil, on July 19th, 2020. Upon gross examination, 80% of the body surface presented disseminated rhomboid cutaneous lesions ranging from 4 to 6 cm-width, characterized by well-defined edges and occasional ulceration, consistent with erysipelas. Additionally, anthropic-made postmortem linear cuts and partial mechanical removal of the flank musculature were noted. Skin samples were collected for histopathologic and molecular analyses. Microscopically, it was possible to observe multifocal dermatitis with vasculitis. Erysipelothrix sp. was detected by PCR. Despite previous reports of human consumption of cetacean meat in northeastern Brazil, the observed marks and advanced carcass autolysis suggested that the animal was most likely used as bait for fishing instead of human intake. This case highlights the value of postmortem examination and PCR even in poorly preserved cadavers and contributes to the understanding of the epidemiology of cutaneous erysipelas in free-ranging cetaceans (first report in an odontocete from the Southern Hemisphere). Due to the zoonotic potential of certain Erysipelothrix species (i.e., E. rhusiopathiae), active public health policies are required to inform field professionals and the general public about the health threats associated with marine mammal manipulation and consumption.

Ovarian Filariasis in a Wild Southern Tamandua (Tamanduatetradactyla; Mammalia: Myrmecophagidae).

Knowledge of reproductive health in wild southern tamanduas (Tamandua tetradactyla; Mammalia: Myrmecophagidae) is fragmentary. During necropsies of roadkill xenarthran species in Brazil, a case of ovarian filariasis in an adult female southern tamandua was observed. Macroscopically, both ovaries were irregularly enlarged and had numerous smooth protuberances. Histologically, the affected ovarian parenchyma presented adult nematodes (including females with microfilaria) surrounded by pleocellular inflammatory infiltrates. The morphological characteristics of the nematodes were consistent with the superfamily Filarioidea (order Spirurida). The adjacent ovarian parenchyma had developing and atretic follicles at different stages of maturation. Filarial nematodes were not observed in other tissues. The cause of death of this tamandua was fatal acute polytrauma as a consequence of the motor vehicle collision. This case adds to a prior report of ovarian filariasis in two southern tamanduas in Nicaragua and Guatemala, dating back almost 100 years, and suggests filarial infections could potentially have an impact on reproductive success in southern tamanduas and possibly other xenarthrans. Several xenarthran species are under different levels of threat and knowledge of their basic reproductive health is crucial for conservation programs.

Toxoplasmosis in a free-ranging hairy dwarf porcupine (Sphiggurus spinosus) with a potential novel genotype.

Toxoplasmosis is a zoonotic disease caused by the ubiquitous coccidia Toxoplasma gondii. Rodents play an important role in maintaining its life cycle, as they are one of the main diet sources for felids (wild and domestic), the unique definitive hosts. However, reports of toxoplasmosis in porcupines (Order Rodentia) are uncommon, with gaps concerning its pathophysiology. South America is the continent with the greatest genetic diversity of rodents and T. gondii. A free-ranging hairy dwarf porcupine was admitted to a wildlife rescue centre with a history of trauma. During rehabilitation, the animal presented neurological symptoms (sporadic episodes of hind limbs paresis) and died 5 months later. The main findings during necropsy were brain congestion and severe incisor overgrowth associated with maxillary perforation. The histopathological exam showed moderate encephalitis, with variable-sized round cysts, positive for PAS stain and immunohistochemistry for T. gondii. Additionally, two cysts were observed in the medulla of the adrenal gland. Molecular techniques were performed to characterize the parasite load by qPCR (Cq = 30) and the genotype by PCR-RFLP with 11 markers, which revealed a potential new genotype. This case adds to the body of knowledge in comparative pathology of Neotropical Rodentia and reports a new potential genotype circulating in South America.

Adenovirus surveillance in wild carnivores from Brazil.

Landscape transformation favors the spread of new pathogens that can be shared between domestic and wild animals. Certain adenoviruses (e.g., canine adenovirus 1 and 2, family Adenoviridae) can infect domestic and wild carnivores. In domestic canids, these viruses are associated with hepatic and respiratory diseases (among others). Nevertheless, information regarding adenovirus pathogenicity and molecular features in wild carnivores is still limited. Herein we surveyed adenovirus in free-ranging carnivores from Brazil. Total DNA was extracted from and subsequently tested by a nested panPCR in spleen and/or lung of 52 carnivores, representing species of the following families: Canidae (n = 4), Felidae (n = 3), Mustelidae (n = 2) and Procyonidae (n = 2). The obtained sequences were compared to others available at GenBank. Available tissue samples from the positive cases were evaluated histopathologically. One out of 52 (1.9%, CI 95%, 0.0-5.7%) carnivores was positive; a roadkilled ocelot (Leopardus pardalis). The obtained sequence presented a low deduced amino acid (78.1%) similarity with the closest adenovirus, identified in a pinniped from the United States of America. This fact and its detection in a novel host suggest it may be representative of a novel species and denominated ocelot adenovirus 1. None of the gross and microscopic findings of the positive case were associated with adenovirus. To the authors' knowledge, this is the first report of adenovirus in wild felids of South America and the second worldwide. Further studies are necessary to assess the epidemiology and potential pathogenicity of this agent in wild carnivores.

Myeloid and histiocytic sarcomas in subantarctic fur seals Arctocephalus tropicalis, Brazil.

Histopathological and immunohistochemical methods were used to diagnose round cell tumors in 2 subantarctic fur seals Arctocephalus tropicalis with marked anemia. Although wild-born, both individuals were placed under human care while juveniles in a Brazilian aquarium. Both pinnipeds were PCR tested for herpesvirus, and 1 was infected with otariid gammaherpesvirus 5 (OtHV-5), previously described in a subantarctic fur seal stranded in Brazil. Although some gammaherpesviruses can cause sarcomas and other neoplasms, it was not possible to definitively associate OtHV-5 with the neoplasm. To our knowledge, these are the first neoplasm records in subantarctic fur seals.

Extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli survey in wild seabirds at a pristine atoll in the southern Atlantic Ocean, Brazil: First report of the O25b-ST131 clone harboring blaCTX-M-8.

Antimicrobial resistance is among the most serious public health threats of the 21st century, with great impact in terms of One Health. Among antimicrobial resistant bacteria (ARB), extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli (ESBL-EC) represent major challenges to human healthcare. Wild birds have been commonly used as environmental bioindicators of ESBL-EC. Remote locations represent a unique opportunity to evaluate the occurrence, dissemination and epidemiology of ARB in the environment. Herein we surveyed ESBL-EC in 204 cloacal swabs from six nonsynanthropic seabird species at the pristine Rocas Atoll, Brazil. We identified ESBL-EC isolates in 2.4% (5/204) of the tested seabirds, all in magnificent frigatebirds (Fregata magnificens). We isolated strains of O25b-ST131-fimH22 harboring gene blaCTX-M-8 (3 clones), ST117 harboring gene blaSHV-12, and a novel ST11350 (clonal complex 349) harboring genes blaCTX-M-55 and fosA3. All the isolates presented Extraintestinal pathogenic E. coli (ExPEC) virulence profiles. We suggest that magnificent frigatebirds may act as "flying bridges", transporting ESBL-EC and ARGs from an anthropogenically-impacted archipelago geographically close to our pristine and remote study site. The characteristics of our isolates suggest zoonotic potential and, despite the apparent good health of all the evaluated birds, may represent a hypothetical potential threat to the avian population using the atoll. To our knowledge, this is the first description of: (1) the pandemic and public health relevant ST131-O25b harboring blaCTX-M-8 worldwide; (2) ST131-fimH22 in wild birds; and (3); fosA3 in wildlife. Our findings expand the current epidemiological knowledge regarding host and geographical distribution of ESBL-EC and ARGs in wild birds, and emphasize the disseminating characteristics and adaptability of ST131 and ST117 strains within the human-animal-interface. Herein we discuss the involvement of nonsynanthropic wild birds in the epidemiology of antimicrobial resistance and their potential as sentinels of ESBL E. coli in insular environments.

Novel gammaherpesvirus associated with primary gastric T-cell lymphoma in a free-ranging giant armadillo in Brazil.

The number of viral-associated neoplasms reported in wildlife has increased over the last decades, likely because of growing research efforts and a potentially greater burden of carcinogenic pathogens. Herein, we describe a primary gastric T-cell lymphoma in one free-ranging giant armadillo (Priodontes maximus) from Brazilian Pantanal infected by a novel gammaherpesvirus, proposed as Cingulatid gammaherpesvirus 1 (CiHV-1). By chromogenic in situ hybridisation against Epstein-Barr virus some neoplastic cells were labeled. Subsequently, a molecular screening was carried out to detect the occurrence of this pathogen in other giant armadillos in the same region. Overall, this novel virus was detected in 14.3% (3/21) of the tested giant armadillos. We suggest this herpesvirus, the first in Xenarthra, as a plausible aetiology of the neoplasm. The implications of CiHV-1 for this species are uncertain; while no outbreaks of disease have been recorded, the present study raises concerns. Further research is warranted to assess the real significance of CiHV-1 and its potential oncogenic role in this species.

Novel alphaherpesvirus in a wild South American sea lion (Otaria byronia) with pulmonary tuberculosis.

In 2017, an adult male South American sea lion (Otaria byronia), presenting emaciation and a cervical abscess, stranded alive in Florianópolis, southern Brazil. The animal was directed to a rehabilitation center, dying a few days later. On necropsy, the main gross findings were necrotizing lymphadenitis of the right prescapular lymph node and nodular bronchopneumonia. A novel alphaherpesvirus, tentatively named Otariid alphaherpesvirus 1, was amplified in several tissue samples. No histopathologic findings associated with viral infection were observed. Additionally, pulmonary tuberculosis by Mycobacterium pinnipedii was diagnosed by histopathological, immunohistochemical, and molecular techniques. Several bacteria were cultured from antemortem and postmortem samples, including Proteus mirabilis from the cervical abscess and cardiac blood, and Escherichia coli from the cervical abscess and pericardial effusion. Flavivirus, morbillivirus, and Apicomplexa were not detected by molecular techniques. Herein, we report a novel alphaherpesvirus in a pinniped species of the family Otariidae. Although previously described in Southern Hemisphere pinniped species, including South American sea lions, there is limited information regarding M. pinnipedii impact over this group. Further research is required to determine the associated pathogenesis of this novel herpesvirus, and prevalence of Otariid alphaherpesvirus 1 and M. pinnipedii in the reproductive colonies.

Galectin-3 immunolabelling correlates with BCL2 expression in canine cutaneous mast cell tumours.

Mast cell tumour (MCT) is the most frequent skin neoplasm in dogs. These tumours are characterised by variable behaviour and clinical presentation that make prognosis an important and challenging task in the veterinary practice. Galectin-3 (Gal-3) is known to influence several biological processes that are important in the cancer context and has been described as a prognostic marker for several human cancers. The aim of the present work was to characterise Gal-3 immunolabelling in canine cutaneous MCTs and to investigate its value as a prognostic marker for the disease. Thirty-four random cases of canine cutaneous MCT that were surgically treated with wide margins were included in this study. Gal-3 expression was evaluated using immunohistochemistry and the results were compared with the expression of apoptosis-related proteins, Ki67 index, histopathological grades, mortality due to the disease and post-surgical survival. The majority of the MCTs (65.8%) were positive for Gal-3. Gal-3 immunolabelling was variable among the samples (2.7%-86.8% of the neoplastic cells). The protein was located in the cytoplasm or in the cytoplasm and the nucleus. Gal-3 positivity was correlated with BCL2 expression (P < 0.001; r = 0.604), but not with Ki67 and BAX. No significant differences were detected between histological grades or in the survival analysis. Gal-3 expression correlates with BCL2 expression in MCTs. Although an efficient marker for several human neoplasms, the results presented herein suggest that Gal-3 immunolabelling is not an independent prognostic indicator for this disease.

Yellow Fever as Cause of Death of Titi Monkeys (Callicebus Spp.).

From 2016 to 2018, an epidemic wave of yellow fever (YF) occurred in Brazil, affecting a large number of Platyrrhini monkeys. Titi monkeys (Callicebus spp.) were severely affected yet pathological characterizations are lacking. This study characterized epizootic YF in 43 titi monkeys (Callicebus spp.) with respect to the microscopic lesions in liver, kidney, spleen, heart, brain, and lung, as well as the distribution of immunolabeling for YF virus antigen, and the flaviviral load in the liver. Of 43 titi monkeys examined, 18 (42%) were positive for yellow fever virus (YFV) by immunohistochemistry or reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR). Affected livers had consistent marked panlobular necrotizing hepatitis, lipidosis, and mild inflammation, with intense immunolabeling for YFV mainly in centrilobular hepatocytes (zone 1; P = .05). In the spleen, consistent findings were variable lymphoid depletion (10/11), lymphoid necrosis (lymphocytolysis; 4/11), and immunolabeling for YFV in histiocytic cells (3/16). The main finding in the kidney was multifocal acute necrosis of tubular epithelium (5/7) that was occasionally associated with intracytoplasmic immunolabeling for YFV (6/15). These data indicate that titi monkeys are susceptible to YFV infection, developing severe hepatic lesions and high viral loads, comparable to humans and Alouatta spp. Thus, Callicebus spp. may be reliable sentinels for YF surveillance.

Occurrence and Quantification of Antimicrobial Resistance Genes in the Gastrointestinal Microbiome of Two Wild Seabird Species With Contrasting Behaviors.

Antimicrobial resistance genes (ARGs) are environmental pollutants and anthropization indicators. We evaluated human interference in the marine ecosystem through the ocurrence and quantification (real-time PCRs) of 21 plasmid-mediated ARGs in enema samples of 25 wild seabirds, upon admission into rehabilitation: kelp gull (Larus dominicanus, n = 14) and Magellanic penguin (Spheniscus magellanicus, n = 11). Overall, higher resistance values were observed in kelp gulls (non-migratory coastal synanthropic) in comparison with Magellanic penguins (migratory pelagic non-synanthropic). There were significant differences between species (respectively, kelp gull and Magellanic penguin): ARGs occurrence (bla TEM [p = 0.032]; tetM [p = 0.015]; tetA [p = 0.003]; and sulII [p = 0.007]), mean number of ARGs per sample (p = 0.031), ARGs mean load percentage (aadA [p = 0.045], tetA [p = 0.031], tetM [p = 0.016], bla TEM [p = 0.032], sulII [p = 0.008]), percentage of genes conferring resistance to an antimicrobial class (betalactams [p = 0.036] and sulfonamides [p = 0.033]), mean number of genes conferring resistance to one or more antimicrobial classes (p = 0.024]), percentage of multiresistant microbiomes (p = 0.032), and clustering (p = 0.006). These differences are likely due to these species' contrasting biology and ecology - key factors in the epidemiology of ARGs in seabirds. Additionally, this is the first report of mecA in seabirds in the Americas. Further studies are necessary to clarify the occurrence and diversity of ARGs in seabirds, and their role as potential sources of infection and dispersal within the One Health chain of ARGs.

Case report of respiratory aspergillosis and candidiasis in wild Magellanic penguins (Spheniscus magellanicus), Brazil.

Magellanic penguins (Spheniscus magellanicus) migrate to the continental shelf of southern-southeastern Brazil during austral winter. Stranded penguins are directed to rehabilitation centers, where they occasionally develop fungal diseases. Aspergillosis, a mycosis caused by Aspergillus spp., is one of the most important diseases of captive penguins, while Candida sp. has been detected in penguins undergoing rehabilitation. Nevertheless, their occurrence in the wild is poorly understood. This study surveyed the occurrence of mycoses in free-ranging Magellanic penguins wintering in southeastern Brazil. These penguins were either found dead or stranded alive and died during transport to a rehabilitation center. Overall, 61 fresh to moderate autolyzed carcasses were necropsied. Upon necropsy, three juvenile males (4.9%) presented mycotic-consistent gross lesions. Histopathology and panfungal PCRs confirmed the mycoses. Major microscopic findings were marked chronic necrotizing multifocal to coalescent pneumonia, airsacculitis, and esophageal/gastric serositis with two types of intralesional fungal structures: (a) septated acute-angled branching hyphae (n = 2) and (b) yeast structures (n = 1), both PAS- and Grocott-positive. Sequences identical to Aspergillus sp. were retrieved in two cases, while the third had sequences identical to Candida palmioleophila. This study describes two cases of aspergillosis and one of candidiasis in free-ranging Magellanic penguins, confirming the species' susceptibility in the wild. These mycoses could be associated with the animals' poor body condition, and/or impaired immunity, and natural and anthropogenic challenges related to migration. To the authors' knowledge, this is the first report of aspergillosis in free-ranging Magellanic penguins in the Atlantic Ocean and of candidiasis in penguins worldwide.

Pulmonary adiaspiromycosis in armadillos killed by motor vehicle collisions in Brazil.

Knowledge of infectious diseases in wildlife provides important information for preventing potential outbreaks of zoonotic diseases. Adiaspiromycosis is a neglected human disease caused by dimorphic Onygenales fungi. The disease is produced by the inflammatory response against growing adiaspores, leading to granulomatous pneumonia. In humans, adiaspiromycosis is relevant in immunosuppressed patients. In animals, it is associated with pneumonia in fossorial species. Given the potential role of armadillos in the epidemiology of adiaspiromycosis, in this study, we sought to investigate the occurrence and pathological features of adiaspiromycosis in roadkilled armadillos. In total, 54 armadillo carcasses were suitable for postmortem pathologic examinations between February 2017 and 2020. Adiaspores, associated with granulomatous lesions, were observed in ten six-banded (Euphractus sexcinctus) and two southern naked-tailed armadillos (Cabassous unicinctus). A previously uncharacterized Onygenales species was molecularly identified in two E. sexcinctus. In summary, herein we report 12 cases of pulmonary adiaspiromycosis (PA) in two species of free-living armadillos in Brazil. Both, the morphology of the fungus, as well as the histopathological findings (granulomatous inflammatory response to adiaspores) are consistent with PA; however, as the molecular identification differs from the reported species, the potential impact of this fungus for human PA is unknown, and we cannot rule out its impact on public health.