- Introduction
- Materials and Methods
- Chimeric proteins.
- Cloning of 56-kD protein genes from Karp, Kato, Gilliam, and TA763 strains of O. tsutsugamushi.
- Expression of recombinant 56-kD proteins from BL21(DE3) transformants.
- Extraction and solubilization of inclusion bodies containing recombinant 56-kD (r56) proteins.
- Chromatographic purification of recombinant 56-kD proteins in 6 M urea.
- Refolding of pure recombinant 56-kD proteins by dialysis.
- Analysis of purified recombinant 56-kD protein and verification of reactivity with antibody by Western blotting.
- Mouse immunization.
- Analysis of reactivity of various recombinant 56-kD proteins and serum samples from mice challenged by various strains of O. tsutsugamushi.
- Results
- Discussion
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Analysis of the Cross-Reactivity of Various 56 kDa Recombinant Protein Antigens with Serum Samples Collected after Orientia tsutsugamushi Infection by ELISA
Orientia tsutsugamushi, the etiologic agent of scrub typhus, has a highly expressed and immunodominant 56-kD outer membrane protein. This protein is one of the leading candidates for diagnosis and vaccine development for scrub typhus. Previous studies using recombinant 56-kD protein (r56s) derived from Karp strain (Kpr56) in a mouse model have shown good homologous protection but only moderate to poor heterologous protection. We evaluated the cross-reactivity of recombinant 56-kD proteins from Karp, Kato, Gilliam, TA763, and three chimeric 56-kD proteins. Not all r56s are equally reactive with strain-specific serum samples. These data provide a first glance of how reactive these r56s are toward the antiserum of different strains and which r56 exhibits the broadest reactivity. A formulation of this combination has the potential to provide broad protection against the heterologous challenge and to be used in a highly sensitive diagnostic assay.
Author Notes
Financial support: This study was supported by Work Unit Number (WUN) 6000.RAD1.J.A0310.
Disclosure: Chien-Chung Chao and Wei-Mei Ching are employees of the U.S. Government. This work was prepared as part of their official duties.
Authors' addresses: Chien-Chung Chao, Viral and Rickettsial Diseases Department, Infectious Diseases Directorate, Naval Medical Research Center, Silver Spring, MD, E-mail: chien-chung.chao@med.navy.mil. Erin S. Huber, Zhiwen Zhang, and Wei-Mei Ching, Viral and Rickettsial Diseases Department, Infectious Diseases Directorate, Naval Medical Research Center, Silver Spring, MD and Uniformed Services University of the Health Sciences, Bethesda, MD, E-mails: ehuber@jhsph.edu, zhiwen.zhang@med.navy.mil, and wei.ching@med.navy.mil.