Diversidade genética da sequência 18S do DNA ribossomal de Theileria equi em equinos oriundos das cinco regiões geográficas do Brasil

Abstract

Equine piroplasmosis, caused by the hemoparasite Theileria equi, is endemic in Brazil and is considered one of the major equine diseases in the world. The objective of the present study was to investigate the genetic diversity of geographically distinct isolates of 18S rRNA gene sequences of T. equi in naturally infected horses from the five Brazilian regions, characterize the hypervariable regions of the T. equi 18S rRNA sequences from this study and to elucidate the taxonomic relationships between T. equi isolates from Brazil and of the world. Equine blood samples were collected in the five regions of the country. The genomic DNA was extracted and the samples were submitted to real-time PCR for the selection of positive samples. Subsequently, the selected samples were submitted to a conventional PCR for amplification of the 18S rRNA gene (~1600 bp) of T equi, the products being sequenced. The sequences were aligned using ClustalW, and the alignment was submitted to the Maximum Likelihood reconstruction method with molecular distances estimated by the General Time Reversible Model, using 1000 replicates. From this alignment a similarity matrix was developed and the hypervariable regions were observed. Phylogenetic analyzes were performed in three different ways: using T. equi sequences from Brazil, using sequences from different species of Theileria and Babesia, and using T. equi sequences from all over the world. Twenty-three partial sequences of the 18S rRNA gene of T. equi were obtained, of which thirteen were unique variants of the gene in Brazil. The hypervariable gene regions V2, V4 and V8 were identified in the study samples, with V4 being the most variable. These hypervariable regions were confirmed by entropy analysis. Two distinct genotypes circulating in the Brazilian regions were identified. Only a single genotype was observed in the North region, while in the other regions both were observed. Samples were grouped into two clades (A and C), most of the study sequences being clustered in clade C. In another tree, all T. equi sequences grouped into group 1, different from the other theilerias and babesias, present in groups 2 and 3. In relation to the T. equi samples of the world, three clades were observed: A, B and C. This study adds important information on the genetic diversity of T. equi in horses from all regions of Brazil based on the 18S rRNA sequence.