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Comparison of DRB3.2 alleles among false-negative and positive tuberculin skin test cattle with bovine tuberculosis

M Encinas(1) X Ferrara Muñiz(1) R A Sammarruco(2) V Ruiz Menna(2) C Garro(2) F Delgado(2) M Zumárraga(1) S Garbaccio(2) H A Carignano(3) M E Eirin(1)

1:Instituto de Agrobiotecnología y Biotecnología Molecular (IABIMO), UEDD INTA-CONICET; CICVyA, Instituto Nacional de Tecnología Agropecuaria; 2:Instituto de Patobiología Veterinaria (IPVeT), UEDD INTA-CONICET; CICVyA, Instituto Nacional de Tecnología Agropecuaria; 3:Instituto de Virología e Innovaciones tecnológicas (IVIT), UEDD CONICET-INTA, Instituto Nacional de Tecnología Agropecuaria (INTA), INTA-CONICET

Mycobacterium bovis is worldwide distributed mainly affecting livestock. Tuberculin skin test (TST), the primary screening tool, detect the infection in cattle. Some animals remain undetected (false-negatives) limiting sanitation programs success. Bovine lecukocyte antigen (BoLA) DRB3 gene is involved in antigenic presentation. BoLA-DRB3.2 polimorpysms were identified in false-negative TST cattle, comparing to TST+ bovines. Twenty-nine TST+ and 18 false-negative dairy bovines were studied. DNA extraction from blood (PuriPrep-S kit, InbioHigway®), PCR amplification of BoLA-DRB3.2 (319bp, primers DRB3FRW/DRB3REV) and genotyping by sequencing (BigDye ® v3.1, Applied Biosystems Inc.) were performed. Genotypes were assigned using Haplofinder and allelic frequency was compared using Epidat 3.1. Among TST+, *0101, *1501 and *1001 were the most frequent alleles (86.3%, accumulated frecuency) followed by *1101, *0902, *0601 and *1701. Among false-negatives, *1501, *1101, *0101, *4401 and *2703 were the most frequent alleles (71.5%, accumulated frequency), followed by *3701, *1001, *0902, *0601,*1701,*14011 and *1602. The *0101 allele was more prevalent in TST+ (p=0.01), while the frequency associated to the other allelles were similar among both, false-negative and TST+ groups (p>0.05). Alelles *1601, *3701, *4401, *14011 and *2703 were only detected in false-negatives, but in a frequency ≤ 8.3%. In this preliminary study, a higher diversity of BoLA-DRB3.2 alleles was observed in false-negative cattle comparing to TST+; with some alleles only identified among false-negatives bovines, although in low proportion. Further studies are needed to really comprenhend the implicancies of the DRB3 diversity on the false negative TSTphenotype.

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