Molecular Cloning and Sequence Analysis of a Rickettsia Tsutsugamushi 22 KDa Antigen Containing B- and T-cell Epitopes

The identification of Rickettsia tsutsugamushi T-cell epitopes is necessary for the characterization of the protective immune response (of which the T-cell response is essential) against scrub typhus rickettsiae. A T-helper cell line derived from R. tsutsugamushi (Karp strain) immune mice reacted with rickettsial protein antigens eluted from the 18-35 kDa region of polyacrylamide gels. Within this region is a 22 kDa protein which is reactive with immune serum. The gene encoding the 22 kDa scrub typhus antigen (sta22) was cloned and expressed in Escherichia coli. Nucleotide sequence analysis of the sta22 gene revealed a potential open reading frame (ORF) in the sta22 sequence encoding a 22 kDa protein. A recombinant 22 kDa protein synthesized in E. coli maxicells was reactive with anti-rickettsial antibodies. The codon usage of the adenine and thymine rich sta22 sequence was similar to other previously sequenced R. tsutsugamushi genes. Computer analysis of the deduced amino acid sequence suggested that the Sta22 protein has several amphipathic regions which may be potential T-cell epitopes. The recombinant Sta22 protein eluted from polyacrylamide gels induced a strong proliferative response from the scrub typhus rickettsiae reactive T-cell line. Recognition of the R. tsutsugamushi Sta22 polypeptide by both cellular and humoral immune mechanisms implicates this antigen as one of potential importance in vaccine development.