Dot-ELISA Rapid Test Using Recombinant 56-kDa Protein Antigens for Serodiagnosis of Scrub Typhus

We developed a rapid dot-enzyme-linked immunosorbent assay (dot-ELISA) using the combination of recombinant 56-kDa protein antigens that exhibited broad reactivity with serum antibodies against the four most prevalent strains (Karp, Kato, Gilliam, and TA763) of Orientia tsutsugamushi. The assay is rapid (30 minutes), and can be done at room temperature, and results can be read by the naked eye. Only a simple shaker is required to wash the membrane. Sera from 338 patients suspected of being ill with scrub typhus from rural hospitals around Thailand were tested using this dot-ELISA. Seventy-five (22.2%) patients were found to be positive. The sensitivity and specificity of dot-ELISA were determined using the indirect immunofluorescent assay (IFA) test as the gold standard, with the cutoff titer of immunoglobulin peroxidase conjugate M (IgM)/G (IgG) greater than 1:400/1:400. The dot-ELISA had a sensitivity of 98.5%, a specificity of 96.3%, a positive predictive value of 86.7%, and a negative predictive value of 99.6% for the acute-phase specimens. The results indicate that dot-ELISA rapid test using recombinant 56-kDa protein antigen was comparable with the IFA test and may be very useful for the diagnosis of scrub typhus in rural hospitals, where IFA is not available.