Development of a tandem repeat-based multilocus typing system distinguishing Babesia bovis geographic isolates.

AGUSTINA PEREZ-LLANEZA; MARINA CABALLERO; EUGENIA BARAVALLE; MARIA MESPLET; JUAN MOSQUEDA; CARLOS E. SUAREZ; IGNACIO ECHAIDE; FRANK KATZER; GABRIELA M. PACHECO; MONICA FLORIN-CHRISTENSEN; LEONHARD SCHNITTGER

VETERINARY PARASITOLOGY

ELSEVIER SCIENCE BV

Lugar: Amsterdam; Año: 2010 vol. 167 p. 196 - 204

0304-4017

Mini- and microsatellite sequences have proven to be excellent tools for the differentiation of strains and populations in several protozoan parasites due to their high variability. In the present work we have searched the genome of the ticktransmitted bovine hemoprotozoon Babesia bovis for tandem repeats (TRs) that could be useful for amultilocus typing system. Hundred and nineteen sequenceswere shortlisted and tested in five common B. bovis reference isolates originating from distinct geographic locations of North and South America: Texas, USA (T2Bo), Mexico (RAD and Mo7), and Santa Fe and Salta, Argentina (R1A and S2P, respectively). Satellite sequences were PCRamplified using specific primers, separated by polyacrylamide gel electrophoresis,visualized by silver staining and sized. Fourteen TR sequences could be reliably amplified in all isolates and displayed length polymorphism. All primers used were specific for B.bovis and did not amplify genomic DNA from the bovine host or from Babesia bigemina,the principal co-infecting bovine parasite in the Americas, allowing their future use infield surveys. The 14 satellite markers identified are distributed throughout the fourchromosomes of B. bovis as follows: chromosome 1 (n = 3), chromosome 2 (n = 2),chromosome 3 (n = 5), and chromosome 4 (n = 4). Within the five B. bovis isolates weidentified nine satellite marker loci with two alleles, three with three alleles, one withfour and another with five alleles. In comparison to Theileria parva, a bovinehemoprotozoan that pertains to the same piroplasmida order and owns a genome ofsimilar size, the number of polymorphic TRs and the average number of alleles per TRlocus seem to be significantly reduced in the B. bovis genome. Furthermore, the ratio ofmicro- to minisatellites in both B. bovis and T. parva is considerably lower than in othereukaryotes, as confirmed by bioinformatic analysis. The multilocus genotype of the five B.bovis isolates was assessed and the genetic distance between each other determinedfollowed by cluster analysis based on neighbor joining. The resulting phenogramshowedthat B. bovis isolates segregated into three clusters according to their geographic origin.The presented marker system is suitable to explore various parameters of B. bovispopulations such as genetic diversity, infection dynamics and their structure under different epidemiological situations, which are of crucial importance for improved control strategies.