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Establishment of Recombinase Polymerase Amplification assay for rapid and sensitive detection of Orientia tsutsugamushi in Southeast Asia.
Acta Trop. 2020 Oct; 210:105541.AT

Abstract

Scrub typhus, caused by Orientia tsutsugamushi, is a common fever in parts of Southern and Southeast Asia. As delayed diagnosis of scrub typhus leads to inappropriate treatment and high mortality rates, of up to 70%, sensitive and rapid detection of O. tsutsugamushi is required for timely and appropriate treatment. Molecular assays, such as PCR and real-time PCR, have been shown to be more sensitive than conventional immunoassay, however, they are only available in centralized laboratories. In contrast to PCR assays, Recombinase Polymerase Amplification (RPA) is conducted under a constant temperature ranging from 24°C to 45°C. Therefore, this technology is very promising for nucleic acid testing in the field, and in resource-limited areas. An RPA assay for the detection of O. tsutsugamushi based on the target gene encoding for the 47 kDa outer membrane protein has been reported, but the primer and probe sequences of this assay are suboptimal for detection of the majority of recently published sequences of O. tsutsugamushi isolates from Southeast Asia. We have established a real-time RPA assay with primer and probe sequences that are optimized for most Southeast Asia's isolates of O. tsutsugamushi. As a result, the new RPA assay showed better performance than the previous assay in detecting O. tsutsugamushi in clinical samples of scrub typhus cases found in Vietnam. The specificity of RPA assay was also evaluated using genomic DNA from microorganisms commonly encountered in the differential diagnosis of scrub typhus, and blood samples from healthy controls and O. tsutsugamushi negative confirmed cases.

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Publisher Full Text (DOI)

Authors+Show Affiliations

Department of Infectious Disease, The 103 Military Hospital, Vietnam Military Medical University.

Department of Genomics and Cytogenetics, Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam.

Department of Genomics and Cytogenetics, Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam.

Department of Genomics and Cytogenetics, Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam.

Department of Genomics and Cytogenetics, Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam.

Department of Genomics and Cytogenetics, Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam.

Department of Genomics and Cytogenetics, Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam.

Department of Medical Microbiology, Military Institute of Preventive Medicine, Vietnam.

Department of Microbiology, The 108 Military Central Hospital.

Department of Hygiene, Vietnam Military Medical University, Vietnam.

Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam.

Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam.

The 103 Military Hospital, Vietnam Military Medical University, Vietnam.

Minerva Foundation Institute for Medical Research, Helsinki, Finland.

Department of Genomics and Cytogenetics, Institute of Biomedicine and Pharmacy (IBP), Vietnam Military Medical University, Vietnam; Department of Medical Microbiology, The 103 Military Hospital, Vietnam Medical University, Vietnam. Electronic address: hohuutho@vmmu.edu.vn.

MeSH

Asia, SoutheasternHumansNucleic Acid Amplification TechniquesOrientia tsutsugamushiReal-Time Polymerase Chain ReactionRecombinasesScrub TyphusSensitivity and Specificity

Pub Type(s)

Journal Article

Language

eng

PubMed ID

32492397

Citation

Nguyen, Chinh Trong, et al. "Establishment of Recombinase Polymerase Amplification Assay for Rapid and Sensitive Detection of Orientia Tsutsugamushi in Southeast Asia." Acta Tropica, vol. 210, 2020, p. 105541.

Nguyen CT, Nguyen UD, Le TT, et al. Establishment of Recombinase Polymerase Amplification assay for rapid and sensitive detection of Orientia tsutsugamushi in Southeast Asia. Acta Trop. 2020;210:105541.

Nguyen, C. T., Nguyen, U. D., Le, T. T., Bui, H. T., Nguyen, A. N. T., Thi Nguyen, A. N., Trieu, N. T., Trieu, L. P., Bui, S. T., Nguyen, C., Van Hoang, L., Ho, S. A., Van Nguyen, B., Stenman, J., & Ho, T. H. (2020). Establishment of Recombinase Polymerase Amplification assay for rapid and sensitive detection of Orientia tsutsugamushi in Southeast Asia. Acta Tropica, 210, 105541. https://doi.org/10.1016/j.actatropica.2020.105541

Nguyen CT, et al. Establishment of Recombinase Polymerase Amplification Assay for Rapid and Sensitive Detection of Orientia Tsutsugamushi in Southeast Asia. Acta Trop. 2020;210:105541. PubMed PMID: 32492397.

* Article titles in AMA citation format should be in sentence-case

TY - JOUR T1 - Establishment of Recombinase Polymerase Amplification assay for rapid and sensitive detection of Orientia tsutsugamushi in Southeast Asia. AU - Nguyen,Chinh Trong, AU - Nguyen,Ung Dinh, AU - Le,Thuy Thi, AU - Bui,Hoai Thi, AU - Nguyen,Anh Ngoc Thi, AU - Thi Nguyen,Anh Ngoc, AU - Trieu,Nguyet Thi, AU - Trieu,Long Phi, AU - Bui,Sy Tien, AU - Nguyen,Chuyen, AU - Van Hoang,Luong, AU - Ho,Son Anh, AU - Van Nguyen,Ba, AU - Stenman,Jakob, AU - Ho,Tho Huu, Y1 - 2020/05/31/ PY - 2019/10/30/received PY - 2020/05/05/revised PY - 2020/05/13/accepted PY - 2020/6/4/pubmed PY - 2020/12/29/medline PY - 2020/6/4/entrez KW - 47-kDa gene KW - Early detection KW - O. tsutsugamushi KW - Recombinase polymerase amplification KW - Scrub typhus KW - Southeast Asia SP - 105541 EP - 105541 JF - Acta tropica JO - Acta Trop VL - 210 N2 - Scrub typhus, caused by Orientia tsutsugamushi, is a common fever in parts of Southern and Southeast Asia. As delayed diagnosis of scrub typhus leads to inappropriate treatment and high mortality rates, of up to 70%, sensitive and rapid detection of O. tsutsugamushi is required for timely and appropriate treatment. Molecular assays, such as PCR and real-time PCR, have been shown to be more sensitive than conventional immunoassay, however, they are only available in centralized laboratories. In contrast to PCR assays, Recombinase Polymerase Amplification (RPA) is conducted under a constant temperature ranging from 24°C to 45°C. Therefore, this technology is very promising for nucleic acid testing in the field, and in resource-limited areas. An RPA assay for the detection of O. tsutsugamushi based on the target gene encoding for the 47 kDa outer membrane protein has been reported, but the primer and probe sequences of this assay are suboptimal for detection of the majority of recently published sequences of O. tsutsugamushi isolates from Southeast Asia. We have established a real-time RPA assay with primer and probe sequences that are optimized for most Southeast Asia's isolates of O. tsutsugamushi. As a result, the new RPA assay showed better performance than the previous assay in detecting O. tsutsugamushi in clinical samples of scrub typhus cases found in Vietnam. The specificity of RPA assay was also evaluated using genomic DNA from microorganisms commonly encountered in the differential diagnosis of scrub typhus, and blood samples from healthy controls and O. tsutsugamushi negative confirmed cases. SN - 1873-6254 UR - https://www.unboundmedicine.com/medline/citation/32492397/Establishment_of_Recombinase_Polymerase_Amplification_assay_for_rapid_and_sensitive_detection_of_Orientia_tsutsugamushi_in_Southeast_Asia_ DB - PRIME DP - Unbound Medicine ER -