Evaluation of Diagnostic Assay for Rickettsioses Using Duplex Real-Time PCR in Multiple Laboratories in Japan

Fumihiko Kawamori; Yukie Shimazu; Hiroko Sato; Naota Monma; Asaka Ikegaya; Seigo Yamamoto; Hiromi Fujita; Hiroshi Morita; Yukiko Tamaki; Naoya Takamoto; Hongru Su; Masahiko Shimada; Yuko Shimamura; Shuichi Masuda; Shuji Ando; Norio Ohashi

doi:10.7883/yoken.JJID.2017.447

Original Article

Evaluation of Diagnostic Assay for Rickettsioses Using Duplex Real-Time PCR in Multiple Laboratories in Japan

Fumihiko Kawamori, Yukie Shimazu, Hiroko Sato, Naota Monma, Asaka Ikegaya, Seigo Yamamoto, Hiromi Fujita, Hiroshi Morita, Yukiko Tamaki, Naoya Takamoto, Hongru Su, Masahiko Shimada, Yuko Shimamura, Shuichi Masuda, Shuji Ando, Norio Ohashi

Author information

Fumihiko Kawamori
Shizuoka Institute of Environment and Hygiene
Department of Food and Nutritional Sciences, Graduate School of Integrated Pharmaceutical and Nutritional Sciences, University of Shizuoka
Yukie Shimazu
Hiroshima Prefectural Technology Research Institute, Public Health and Environment Center
Hiroko Sato
Akita Research Center for Public Health and Environment
Naota Monma
Fukushima Institute for Public Health
Asaka Ikegaya
Shizuoka Institute of Environment and Hygiene
Seigo Yamamoto
Miyazaki Prefectural Institute for Public Health and Environment
Hiromi Fujita
Mahara Institute of Medical Acarology
Hiroshi Morita
Myojin Clinic
Yukiko Tamaki
Tamaki Hospital
Naoya Takamoto
Department of Food and Nutritional Sciences, Graduate School of Integrated Pharmaceutical and Nutritional Sciences, University of Shizuoka
Hongru Su
Department of Food and Nutritional Sciences, Graduate School of Integrated Pharmaceutical and Nutritional Sciences, University of Shizuoka
Masahiko Shimada
Department of Food and Nutritional Sciences, Graduate School of Integrated Pharmaceutical and Nutritional Sciences, University of Shizuoka
Yuko Shimamura
Department of Food and Nutritional Sciences, Graduate School of Integrated Pharmaceutical and Nutritional Sciences, University of Shizuoka
Shuichi Masuda
Department of Food and Nutritional Sciences, Graduate School of Integrated Pharmaceutical and Nutritional Sciences, University of Shizuoka
Shuji Ando
Department of Virology I, National Institute of Infectious Diseases
Norio Ohashi
Department of Food and Nutritional Sciences, Graduate School of Integrated Pharmaceutical and Nutritional Sciences, University of Shizuoka

Keywords: rickettsiosis, diagnosis, Orientia tsutsugamushi, Rickettsia japonica, duplex real-time PCR

JOURNAL FREE ACCESS

2018 Volume 71 Issue 4 Pages 267-273

DOI https://doi.org/10.7883/yoken.JJID.2017.447

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Abstract

Tsutsugamushi disease and Japanese spotted fever are representative rickettsioses in Japan, and are caused by infection with Orientia tsutsugamushi and Rickettsia japonica, respectively. For molecular-based diagnosis, conventional PCR assays, which independently amplify respective rickettsial DNA, are usually used; however, this approach is time-consuming. Here, we describe a new duplex real-time PCR assay for the simultaneous detection of O. tsutsugamushi and spotted fever group rickettsiae, and its evaluation using several PCR conditions in 6 public health laboratories. The detection limit of the assay was estimated to be 10² copies and the sensitivity was almost identical to that of 3 conventional PCR methods. A total of 317 febrile patients were selected as clinically suspected or confirmed cases of rickettsioses. The detection efficiency of this assay for O. tsutsugamushi from blood or skin (eschar) specimens appeared to be almost the same as that of the conventional PCR method, even when performed in different laboratories, whereas the efficiency for spotted fever group rickettsiae tended to be higher than that of the 2 traditional double PCR assays. Our duplex real-time PCR is thus a powerful tool for the rapid diagnosis of rickettsioses, especially at the acute stage of infection.

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