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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.

Anaplasmataceae agents are obligatory intracellular Gram-negative α-proteobacteria that are transmitted mostly by arthropod vectors. Although mammals of the Superorder Xenarthra (sloths, anteaters, and armadillos) have been implicated as reservoirs for several zoonotic agents, only few studies have sought to detect Anaplasmataceae agents in this group of mammals. This study aimed to investigate the occurrence and genetic diversity of Anaplasma spp. and Ehrlichia spp. in blood and spleen samples of free-living Xenarthra from four different states in Brazil (São Paulo, Mato Grosso do Sul, Rondônia, and Pará). Nested and conventional PCR screening assays were performed to detect the rrs and dsb genes of Anaplasma spp. and Ehrlichia spp., respectively. The assays were positive in 27.57% (91/330) of the Anaplasma spp. and 24.54% (81/330) of the Ehrlichia spp. Of the 91 positive Anaplasma spp. samples, 56.04% were positive in a conventional PCR assay targeting the 23S-5S intergenic region. Phylogenetic and distance analyses based on the rrs gene allocated Anaplasma sequences from sloths captured in Rondônia and Pará states in a single clade, which was closely related to the A. marginale, A. ovis, and A. capra clades. The sequences detected in southern anteaters from São Paulo were allocated in a clade closely related to sequences of Anaplasma spp. detected in Nasua nasua, Leopardus pardalis, and Cerdocyon thous in Brazil. These sequences were positioned close to A. odocoilei sequences. Genotype analysis corroborated previous findings and demonstrated the circulation of two distinct Anaplasma genotypes in animals from north and southeast Brazil. The first genotype was new. The second was previously detected in N. nasua in Mato Grosso do Sul state. The intergenic region analyses also demonstrated two distinct genotypes of Anaplasma. The sequences detected in Xenarthra from Pará and Rondônia states were closely related to those in A. marginale, A. ovis, and A. capra. Anaplasma spp. sequences detected in Xenarthra from São Paulo and were allocated close to those in A. phagocytophilum. The analyses based on the dsb gene grouped the Ehrlichia spp. sequences with sequences of E. canis (São Paulo, Mato Grosso do Sul, and Pará) and E. minasensis (Rondônia and Pará). The data indicate the occurrence of E. canis and E. minasensis and two possible new Candidatus species of Anaplasma spp. in free-living mammals of the Superorder Xenarthra in Brazil.