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Clinical and Vaccine Immunology, September 2009, p. 1352-1359, Vol. 16, No. 9
1071-412X/09/$08.00+0     doi:10.1128/CVI.00211-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Identification of Novel Mycobacterium bovis Antigens by Dissection of Crude Protein Fractions{triangledown} ,{dagger}

V. Meikle,1* A. Alito,1 A. S. Llera,2 A. Gioffré,1 A. Peralta,1 B. M. Buddle,3 and A. Cataldi1

Institute of Biotechnology, INTA, Castelar, Argentina,1 Fundación Instituto Leloir, Buenos Aires, Argentina,2 AgResearch, Hopkirk Research Institute, Palmerston North, New Zealand3

Received 21 May 2009/ Returned for modification 16 June 2009/ Accepted 21 July 2009

Culture filtrate and cell extracts from Mycobacterium bovis cultures contain molecules which could promote protective immunity to tuberculosis in animals. Different protein fractions of M. bovis cultures were obtained by elution electrophoresis and were tested in experimentally infected cattle. The fractions that elicited gamma interferon (IFN-{gamma}) responses were resolved by two-dimensional gel electrophoresis, and individual proteins were identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry. The open reading frames were cloned, expressed as their recombinant forms, and retested with naturally and experimentally infected animals. Eleven protein fractions were highly reactive, from which the Rv1636, HspX, Rv0138, Rv2524, EsxI, and Rv3740 recombinant proteins were obtained. EsxI and HspX were the antigens most recognized by the IFN-{gamma} release assay. In summary, a proteomic approach allowed the identification of novel antigens useful for the diagnosis of bovine tuberculosis.

* Corresponding author. Mailing address: Institute of Biotechnology, INTA, Castelar, Argentina. Phone: 00 54 11 4621 1127. Fax: 00 54 11 4621 0199. E-mail: vmeikle{at}cnia.inta.gov.ar

{triangledown} Published ahead of print on 29 July 2009.

{dagger} Supplemental material for this article may be found at http://cvi.asm.org/.

Clinical and Vaccine Immunology, September 2009, p. 1352-1359, Vol. 16, No. 9
1071-412X/09/$08.00+0     doi:10.1128/CVI.00211-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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