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Journal of Veterinary Science & Medical Diagnosis ISSN: 2325-9590

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Designing Polymerase Chain Reaction (PCR) Technique for the Detection of Specific Causes of Tuberculosis (TB) in Dairy Cattle and Human

Designing Polymerase Chain Reaction (PCR) Technique for the Detection of Specific Causes of Tuberculosis (TB) in Dairy Cattle and Human

Tuberculosis (TB) in cattle and human is caused by M. bovis, M. tuberculosis and M. avium subsp. var Paratuberculosis (M. paratuberculosis). Selected dairy cattle (N=700) and human (N=20) were tested using tuberculin tests and X-ray imaging. Aspiration biopsy of prescapular lymphnodes from tuberculin test positive cattle and urine, cough, pleural and peritoneal fluid from suspected human was further tested using Ziehl Neelsen staining. Genomic DNA from tuberculin test positive cattle (N=23) and suspected humans (N=20) were tested using multiplex polymerase chain reactions (PCR) targeting 16srRNA (1030bp, 180bp). Results showed that 23 cattle and 11 human were infected with TB. To detect infectivity due to M. bovis and M. tuberculosis selected cattle (N=11) and humans (N=11) samples were tested in uniplex PCR targeting MPB83 (600bp) and H37Rv Rv3479HP (667bp) genes respectively. Results of PCR showed that all of the bovine and seven human samples generated MPB83 gene specific 600bp amplicon. Two bovine and seven human DNA generated H37Rv Rv3479HP gene specific 667bp amplicons. Seven cattle were infected with M. bovis and two with M. tuberculosis. Two cattle were co-infected with M. bovis and M. paratuberculosis. Four humans were infected with M. bovis and seven with M. tuberculosis. The MPB83 gene is invariably shared by M. bovis and M. tuberculosis. The PCR protocol designed targeting fragment of H37Rv Rv3479HP (667bp) gene is selective for M. tuberculosis. Results of sequencing showed point mutation in 16srRNA, MPB83 and H37Rv Rv3479HP genes. Phylogenetic analyses of the selected genes of M. bovis and M. tuberculosis showed that the organisms were belonging to Lineage 1. Intradermal tuberculin test and smear microscopy unable to differentiate infectivity due to M. bovis or M. tuberculosis. The PCR technique designed appear specific for M. tuberculosis, can be used to detect causes of TB in mammals and designing future preventive strategies accordingly.

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