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Infection and Immunity, October 2009, p. 4356-4361, Vol. 77, No. 10
0019-9567/09/$08.00+0     doi:10.1128/IAI.00242-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

High-Throughput Identification of New Protective Antigens from a Yersinia pestis Live Vaccine by Enzyme-Linked Immunospot Assay{triangledown} ,{dagger}

Bei Li,1,2,{ddagger} Lei Zhou,1,{ddagger} JingYu Guo,1 Xiaoyi Wang,1 Bin Ni,1 Yuehua Ke,1 Ziwen Zhu,1 Zhaobiao Guo,1 and Ruifu Yang1*

Laboratory of Analytical Microbiology, State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Beijing 100071, China,1 Yunyang Medical College, Shiyan, Hubei Province, China2

Received 3 March 2009/ Returned for modification 9 April 2009/ Accepted 20 July 2009

Yersinia pestis, the plague pathogen, is a facultative intracellular bacterium. Cellular immunity plays important roles in defense against infections. The identification of T-cell targets is critical for the development of effective vaccines against intracellular bacteria; however, the function of cellular immunity in protection from plague was not clearly understood. In this study, 261 genes from Y. pestis were selected on the basis of bioinformatics analysis and previous research results for expression in Escherichia coli BL21(DE3). After purification, 101 proteins were qualified for examination of their abilities to induce the production of gamma interferon in mice immunized with live vaccine EV76 by enzyme-linked immunospot assay. Thirty-four proteins were found to stimulate strong T-cell responses. The protective efficiencies for 24 of them were preliminarily evaluated using a mouse plague model. In addition to LcrV, nine proteins (YPO0606, YPO1914, YPO0612, YPO3119, YPO3047, YPO1377, YPCD1.05c, YPO0420, and YPO3720) may provide partial protection against challenge with a low dose (20 times the 50% lethal dose [20x LD50]) of Y. pestis, but only YPO0606 could partially protect mice from infection with Y. pestis at a higher challenge dosage (200x LD50). These proteins would be the potential components for Y. pestis vaccine development.

* Corresponding author. Mailing address: Institute of Microbiology and Epidemiology, No. 20, Dongdajie, Fengtai, Beijing 100071, China. Phone: 0086-10-66948595. Fax: 0086-10-63815689. E-mail: ruifuyang{at}gmail.com

{triangledown} Published ahead of print on 3 August 2009.

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

Editor: J. B. Bliska

{ddagger} B.L. and L.Z. contributed equally to this work.

Infection and Immunity, October 2009, p. 4356-4361, Vol. 77, No. 10
0019-9567/09/$08.00+0     doi:10.1128/IAI.00242-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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