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Orientia tsutsugamushi  (Ots) frequently causes severe scrub typhus infections in the Asia-Pacific region. Korean investigators have demonstrated that Ots encodes five different autotransporter domain (ATD) proteins (ScaA-ScaE). ScaA functions as an adhesin and confers protective immunity in a lethal mouse model of Ots infection. Specific antibodies are detected against ScaA and ScaC in Korean scrub typhus patients. However, there is limited data on the distribution of the Sca protein genes in diverse isolates of Ots. By BLAST analysis with the conserved beta barrel autotransporter domain (ATD) regions of the  sca  proteins, we discovered a sixth gene  sca F among 3 of 10 new partial Ots genome sequences available at NCBI GenBank (Sido, Karp, AFSC7). We designed two to seven specific TaqMan assays to detect the ATD for each of the six  sca  genes. The TaqMan assays among those for each  sca  gene which gave the greatest sensitivity and linearity with DNA log dilutions were then used for screening DNAs from Ots isolates grown in L929 mouse cells for  sca  genes. The  sca  prevalence survey was performed for all six  sca  genes with 178 DNAs from isolates from 12 countries. The survey results were confirmed by conventional PCR with primers from conserved regions of the passenger domains (PD) and ATD of the  sca  proteins. The ATD was highly conserved between the DNAs of different genotypes compared to the  sca  PD but each TaqMan assay was  sca  specific. The percentage positivity for 56 kDa and  sca A genes in the 178 DNAs using Ha primers was 59.6% and 62.4%, respectively. Our  sca A conventional ATD PCR assay was positive in 98.3% but  sca A was present in all 178 DNAs (100%) by ATD TaqMan.  sca B,  sca C,  sca D,  sca E and  sca F were detected in 33.7%, 97.8%, 93.8%, 97.2% and 43.3% isolates by ATD TaqMan, respectively. The ATDs of Ots  sca  genes are thus sufficiently conserved between different genotypes for molecular assay design. Four  sca  genes are widely distributed among diverse Ots isolates from diverse geographical areas.  sca B and  sca F were detected in fewer Ots isolates and absent from some available genome sequences. Whether the utility of the ScaA, ScaC, ScaD, and ScaE antigenic passenger protein domains exceeds that of the mixed 56 kDa type surface antigens of Ots now used in combination diagnostic assays needs to be determined before they can be considered as suitable alternative serological antigens for diagnosis of scrub typhus.

Detection and distribution of Sca autotransporter protein antigens in diverse isolates of Orientia tsutsugamushi