Detection of genetic diversity of Anaplasma marginale isolates in Minas Gerais, Brazil

Abstract

Bovine anaplasmosis, caused by the tick-borne rickettsia Anaplasma marginale, is endemic in tropical and subtropical regions of the world and results in economic losses in the cattle industry. Major surface proteins (MSPs) have been used as markers for the genetic characterization of A. marginale strains and demonstrate that many isolates may occur in a given geographic area. However, in Brazil, little is known about the genetic diversity of A. marginale isolates within individual herds. This study was designed to examine the genetic variation among A. marginale infecting calves in a farm in the south of Minas Gerais State, Brazil. Blood samples collected from 100 calves were used to prepare Giemsastained smears that were microscopically examined for the presence of A. marginale. From each blood sample, DNA was extracted and analyzed by a polymerase chain reaction (PCR), followed by sequencing to determine diversity among the isolates. Examination of blood smears showed that 48% of the calves were infected with A. marginale, while the real-time PCR detected 70.2% positivity. Congenital infections were found in four calves. The microsatellite and tandem repeat analyses showed high genetic diversity among the isolates.Keywords: Anaplasma marginale, MSP1a, DNA sequencing, microsatellites, tandem repeats, Brazil. ResumoA anaplasmose bovina, causada pela rickettsia Anaplasma marginale e transmitida por carrapatos, é endêmica em regiões tropicais e subtropicais no mundo e causa grandes perdas econômicas na industria de bovinos. Proteínas principais de superfície (MSPs) foram usados como marcadores para a caracterização genética de amostras de A. marginale, demonstrando que diferentes isolados podem ocorrer numa certa região geográfica. Porém, no Brasil pouco se sabe sobre a variedade genética de isolados de A. marginale em rebanhos individuais. Este estudo teve como objetivo investigar a ocorrência de variação genética entre bezerros infectados com A. marginale numa fazenda do sul de Minas Gerais, Brasil. Amostras de sangue coletadas de 100 bezerros foram utilizadas para o preparo de esfregaços sanguíneos, corados pelo Giemsa, para detecção da infecção por A. marginale. Amostras de DNA extraídas de cada amostra foram analisadas através de PCR seguido de sequenciamento. O exame dos esfregaços demonstrou que 48% dos bezerros estavam infectados com A. marginale, enquanto que o PCR detectou 70,2% de positividade. Infecção congênita foi detectada em quatro bezerros. As análises de microsatélites e 'tandem repeats' comprovaram uma grande diversidade genética entre os isolados.Palavras-chave: Anaplasma marginale, MSP1a, sequenciamento de DNA, microsatélites, tandem repeats, Brasil.

Outbreak of anaplasmosis associated with the presence of different Anaplasma marginale strains in dairy cattle in the states of São Paulo and Goiás, Brazil

“…The results found in Lins are similar to those reported by Pohl et al (2013) from cattle sampled in Belo Horizonte, state of Minas Gerais, southeastern Brazil, in which eight different strains of A. marginale and four different genotypes (B, D, E and G) were observed in the 13 samples sequenced. In our study, the high diversity of A. marginale in cattle from Lins, state of São Paulo, is most likely a consequence of different transmission mechanisms (biological and mechanical), each contributing to the genetic diversity of A. marginale in the herd.…”

“…Estrada-Peña et al (2009) evaluated the distribution of nine different genotypes in four distinct ecosystems worldwide and observed that the genotype E is the most common in South America, especially in Brazil and Argentina. However, the genotypes B, C, D and G have also been described previously in Brazil and Argentina VIDOTTO et al, 2006;POHL et al, 2013;SILVA et al, 2015a, b). The present study describes for the first time the presence of the H genotype in Brazil.…”

“…Brazil is considered an endemic area for anaplasmosis in cattle (VIDOTTO et al, 2006;POHL et al, 2013), and the infection of cattle by multiple strains of A. marginale has recently been reported in the country (POHL et al, 2013;SILVA et al, 2015a, b). The global distribution and variation in pathogenicity of A. marginale are associated to the genetic diversity and variability of this bacterium (FUENTE et al, 2007).…”

“…The α-β 3 -Γ and τ-10-15 strains have been previously described in outbreaks of bovine anaplasmosis in Mexico (ALMAZÁN et al, 2008) and Argentina (RUYBAL et al, 2009). In addition, the 72-62-61 strain has been reported as the most common in a cattle herd from the state of Minas Gerais, Brazil (POHL et al, 2013). In Brazil, significant variations in the MSP1a tandem repeats of strains of A. marginale have been found in the states of Paraná (VIDOTTO et al, 2006), Minas Gerais POHL et al, 2013), Rio de Janeiro (SILVA et al, 2015a) and São Paulo (SILVA et al, 2015b).…”

“…In addition, the 72-62-61 strain has been reported as the most common in a cattle herd from the state of Minas Gerais, Brazil (POHL et al, 2013). In Brazil, significant variations in the MSP1a tandem repeats of strains of A. marginale have been found in the states of Paraná (VIDOTTO et al, 2006), Minas Gerais POHL et al, 2013), Rio de Janeiro (SILVA et al, 2015a) and São Paulo (SILVA et al, 2015b). In MSP1a tandem repeats of strains of A. marginale described in Brazil, the most frequent sequences are 16, α, τ in Paraná (VIDOTTO et al, 2006), 72 and α in Minas Gerais (POHL et al, 2013), 4 and τ in Rio de Janeiro (SILVA et al, 2015a) and, α and β in São Paulo (SILVA et al, 2015b).…”