BACKGROUND: Bovine tuberculosis caused by Mycobacterium bovis is an important disease that has negative effects on public health and entails economic loss. Traditional controlling programs for cattle  focus on test and slaughter strategy, and false positive is one of the disadvantages associated with tuberculin skin test. To overcome this limitation, proteins with high specificity have to be utilized. OBJECTIVES: The objective of this study was to clone and express virulent protein CFP-10 from Mycobacterium bovis AN5. METHODS: Full-length genes of cfp-10 were amplified by PCR technique. In parallel, pET23a(+) and PCR products were double digested by EcoRI and HindIII. Ligation was performed at 16˚C followed by transformation into competence E. coli DH5α. After being identified with sequencing, the cloned vector was transformed into E. coli BL21. Induction was performed by isopropyl-β-D-thiogalactopyranoside (IPTG). Urea 8M was used to dissolve the expressed protein in the inclusion body form. Recombinant protein was purified by Nickle-Resin, and urea was eliminated by decreasing the gradient. RESULTS: The CFP-10 gene clone was proved by sequencing method. The CFP-10, as a 10 KDa protein, was confirmed by Western blotting using monoclonal antibodies. Based on the results, the recombinant protein was successfully cloned and expressed. CONCLUSIONS: The results showed that cfp-10 gene was successfully cloned and expressed in prokaryotic system, indicating that this recombinant protein could be utilized in diagnostic kits against bovine tuberculosis in the future.

In bovine tuberculosis caused by Mycobacterium bovis, cattle are known as the main host, and sheep are relatively resistant. Sheep become infected only when there is very close contact between cattle and sheep or when the mycobacterium is abundant in the environment. In the present study, a case of generalized tuberculosis was reported in the sheep of the Sanandaj slaughterhouse.The examined animal carcass was transferred from the slaughterhouse to the University of Kurdistan microbiology laboratory and was dissected under complete health safety conditions. In the carcass examination, necropsy lesions (generalized tubercular granulomas) were visible in different organs and mediastinal lymph nodes, bronchioles, mesentery, and liver portal system. These lesions were seen in the lymph nodes of different body parts, including the lung, liver, digestive system, reproductive system, bladder, and different muscles, including the peritoneal muscle, diaphragm muscle, and abdominal muscles.Diagnostic tests included clinical observations, Ziehl-Neelsen staining for Mycobacterium bacteria, and polymerase chain reaction (PCR) molecular technique. In Ziehl-Neelsen staining, Mycobacterium acid-fast bacilli were seen in red color on the pale blue background of the slide. Also, the product of 499 base pairs was amplified in the specific PCR reaction, which confirmed Mycobacterium bovis.The present report shows that Mycobacterium bovis strains present in the region can be transmitted to sheep and cause disease with severe clinical symptoms. This report is critical, particularly for mixed cattle and sheep farming. Since cattle and sheep are kept together in many breeding farms in Kurdistan province, special attention was paid to the possibility of transmission of bovine tuberculosis to sheep and humans.