Scrub typhus의 진단에 있어서 16s ribosomal RNA gene을 이용한 conventional PCR의 임상적 유용성

Abstract

(Background) A simple and more rapid laboratory diagnostic method for scrub typhus, a polymerase chain reaction(PCR) assay on the blood sample has become prevalent for the early diagnosis of scrub typhus. Also, Nested, real-time PCR methods have been introduced and yielded more sensitive diagnostic results. However, these newer methods need more experienced personnels for conducting molecular laboratory works, more expensive laboratory facilities. Various target genes used in PCR for the diagnosis of scrub typhus are introduced in several studies. Up to date, 47kDa, 56kDa and gro-EL STG gene have been used and investigated, frequently. Also, 16s ribosomal RNA gene could be used in PCR methods as target gene for the diagnosis of scrub typhus. 16s ribosomal RNA gene is highly conserved between different species of bacteria and archaea.(1) Moreover, 16S rRNA gene sequences contain hypervariable regions which can provide species-specific signature sequences useful for bacterial identification.(2) In this aspects, 16S rRNA gene sequencing could be used in medical microbiology as a rapid, accurate alternative to phenotypic methods of bacterial identification. But Studies for evaluating clinical values of these target genes used in conventional PCR for the diagnosis of scrub typhus has not been reported until recently.

(Method) In this study, we conducted a comparative study on 167 patient with acute febrile illness which is possible scrub typhus who were admitted to a single hospital(Chosun University Hospital, Korea). To assess the clinical usefulness of performing conventional PCR as a diagnostic modality, we designed the conventional PCR primers targetted to 16s ribosomal RNA and conducted comparative study with other conventional PCR results in which 47kD, 56kD, gro-EL STG gene were used for the diagnosis of scrub typhus. To estimate and compare the sensitivity and specificity of conventional, nested and real time PCR, the receiver operating characteristic(ROC) curve analysis was used.

(Result) The sensitivity and specificity of 16s ribosomal RNA conventional PCR for the diagnosis of scrub typhus were 86.8% and 100%, respectively. In case of 47 kDa , 56kDa, groEL-STG gene PCR, sensitivities of each gene are 3%, 8% and 61%. In this study, among conventional PCR, PCR using 16s ribosomal RNA gene is most sensitive for the diagnosis of scrub typhus. In pairwise comparison of ROC curves, AUC value of 16s ribosomal RNA conventional PCR(0.934(95% CI, 0.885 to 0.967)) was superior to that of 47kDa, 57kDa, groEL-STG conventional PCR.

(Conclusion) 16s ribosomal RNA gene could be used in conventional PCR for the diagnosis of scrub typhus, as the most sensitive and superior diagnostic target gene, compared to other conventional PCR results using 47kDa, 56kDa, groEL-STG gene. Even comparing with nested, realtime PCR methods, 16s ribosomal RNA conventional PCR showed superior diagnostic results in this study.