Canine piroplasmids: Molecular detection and laboratory characterization in dogs from Brasilia, Brazil, with the first molecular evidence of dog exposure to a novel opossum-associated Babesia sp.

Canine piroplasmid infections can be caused by Babesia spp., Theileria spp. and Rangelia vitalii. In Brazil, canine babesiosis caused by Babesia vogeli is endemic and reported throughout the country. On the other hand, Rangeliosis caused by R. vitalii has only been described so far in the South and Southeast regions. Despite that, studies analyzing the laboratory and molecular characterization of these hemoprotozoa are still scarce. To investigate the occurrence, the laboratory features, the molecular characterization, and the diversity of piroplasmids from Midwestern Brazil, a survey was performed using blood samples obtained from 276 domestic dogs from Brasília, Federal District, Midwestern Brazil. A broad-range quantitative PCR (qPCR) targeting the mitochondrial large subunit ribosomal DNA (LSU4) was used to detect piroplasmid DNA. The overall molecular occurrence of piroplasmids was 11.2% (31/276), with 9.7% (27/276) of the sequences identified as Babesia vogeli (98-100% identity to B. vogeli isolate from the USA). Based on a partial 18S rRNA sequence pairwise alignment (-250 bp), 1.4% (4/276) of the sequences showed only 76.8% identity with B. vogeli but 100% identity with opossum-associated Babesia sp. (MW290046-53). These findings suggest the exposure of dogs from Brazil to a recently described Babesia sp. isolated from white-eared opossum. None of the analyzed dogs was positive for Theileria spp. or R. vitalii. Subsequently, all positive sequences were submitted to three additional PCR assays based on the 18S rRNA, cox-1, and cytb genes, aiming at performing a haplotype network analysis. Haplotype network using cox-1 sequences showed the presence of six different haplotypes of B. vogeli; one of them was shared with isolates from Brazil, the USA, and India. When including animals co-infected with other vector-borne diseases, piroplasmid-positive dogs had 2.3 times higher chance of having thrombocytopenia than the negative ones. The molecular results demonstrated that the compared Babesia vogeli sequences showed a low variability as well as evidence of exposure to a putative novel opossum-associated Babesia sp. in dogs from Midwestern Brazil.

Piroplasmid infection is not associated with clinicopathological and laboratory abnormalities in cats from Midwestern Brazil.

Feline piroplasmids include the genera Babesia spp., Cytauxzoon spp., and Theileria spp. In Brazil, there are few reports regarding these hemoprotozoans; however, clinicopathological and molecular data are scarce. This study aimed to characterize the clinical relevance of these parasites through hematological, biochemical, and molecular approaches. For this purpose, 166 cats from Brasilia, Federal District, Midwestern Brazil, were screened using a quantitative polymerase chain reaction (qPCR) for piroplasmids based on the LSU4 mitochondrial gene, which resulted in an overall prevalence of 36/166 (21.7%). Twelve of 166 samples (7.2%) were positive for C. felis, while 19/166 (11.4%) were positive for Babesia vogeli. No samples tested positive for Theileria spp. Babesia vogeli and Cytauxzoon spp. LSU4 sequences showed identities of 97-100% and 99.3%, respectively, to US isolates. The hematological and biochemical findings did not differ significantly between the cats that tested positive and negative for piroplasmids. Although the lack of abnormalities in clinical and laboratory parameters does not eliminate the possibility that these cats were sick and recovered, it may suggest that the Brazilian strain of Cytauxzoon spp. is not as pathogenic as that from the USA, despite the high molecular identity with North American isolates.

Molecular detection of piroplasmids in synanthropic rodents, marsupials, and associated ticks from Brazil, with phylogenetic inference of a putative novel Babesia sp. from white-eared opossum (Didelphis albiventris).

The order Piroplasmida encompasses tick-borne pathogens of veterinary and medical importance positioned in two main families: Babesiidae and Theileriidae. Even though previous studies carried out in Brazil recorded the occurrence of piroplasmid species circulating in small mammals, 18S RNA gene sequences were only partially sequenced, preventing the assessment of their phylogenetic positioning. The current study aimed to detect and characterize, using morphological, molecular, and bioinformatic approaches, piroplasmids from wild mammals and associated ticks sampled in Central-Western Brazil. Out of 67 Didelphis albiventris sampled, 22 (16.4%) were positive for piroplasmids by PCR. In contrast, none of the 48 small rodents and 14 capybaras (Hydrochoerus hydrochaeris) was PCR-positive. Four Amblyomma dubitatum ticks-one from Rattus rattus, one from H. hydrochaeris, and two from D. albiventris-out of 114 Amblyomma spp. DNA samples were positive for piroplasmids by PCR. The phylogenetic inference performed using the near-complete 18S rRNA gene positioned the putative novel piroplasmid species detected in D. albiventris and associated A. dubitatum ticks near to Babesia sensu lato clade (Western group-cluster III) and distant from the Australian marsupial-associated piroplasms. Phylogenetic inferences based on two additional molecular markers, namely hsp-70 and cox-1, supported the near-complete 18S rRNA gene phylogenetic inference. Finally, the partial 18S rRNA gene sequences detected in ticks from rodents (R. rattus and H. hydrochaeris) showed 97.2-99.4% identity with the Piroplasmida previously detected in a capybara from Brazil, raising evidence that a still uncharacterized piroplasmid species has been identified in the capybara, the largest rodent species from South America.