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Title

Development and Validation of Heminested RT-PCR and qRT-PCR Assays for Comprehensive Detection of Rabies Virus in the Suspected Rabid Brain and Saliva Samples

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Abstract

 Background: Rabies Virus (RV) is one of the most dangerous zoonotic disease and major public health problems in most of the world, especially underdeveloped countries. Rabies is preventable by proper vaccination, even shortly after exposure. Today, it seems a fast, sensitive, and reliable rabies diagnostic method is required, which might reduce the fi nancial burden of inappropriate diagnosis. Objectives: The aim of this study was to develop and validate two molecular techniques, including heminested RT-PCR and qRT-PCR assays, for comprehensive detection of Rabies Virus in the suspected rabid brain and saliva samples. Methods: In this study, we developed qRT-PCR as a fast, sensitive, and specifi c method for rapid detection of Rabies Virus in brain and saliva samples. Also, the sensitivity and specifi city of the method were compared with heminested RT-PCR test and Direct fl uorescent antibody (dFA) as a serologic gold standard method of World Health Organization (WHO) and MIT (mouse inoculation test) as a confi rmatory test. Results: A combination of compatible primers based on RNA-dependent RNA polymerase (L) gene of the Pasteur virus fi xed strain (PV) (accession number. M13215) was used for developing the qRT-PCR assay. Primer and probes were designed according to other Iran circulating viruses genomes that were available in public databases (GenBank). The clinical sensitivities of qRT-PCR and heminested RT-PCR methods were determined 97. 14% and 94. 3%, respectively. In addition, the clinical specifi cities of qRT-PCR and heminested RT-PCR methods were determined 93. 75% and 88. 24%, respectively. Also, the analytical sensitivities of qRT-PCR and heminested RT-PCR methods were about 5  10 FFU/mL, respectively. Conclusions: In this study, qRT-PCR assay as a diagnostic molecular method with high sensitivity and specifi city was developed for the detection of the Rabies Virus genome in both brain and saliva samples. Therefore, this rapid, accurate, and cost-eff ective detection and quantifi cation method may be used as an investigative tool, which can be valid for detection of target viral genome in the research and diagnosis fi eld.

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    APA: Copy

    AMIRI, FERESHTEH, Fadajan, Zohreh, Rasooli, Azadeh, SALAHSHOURIFAR, IMAN, BASHAR, ROUZBEH, ARAB BAFERANI, MASOUMEH, & FAZELI, MARYAM. (2019). Development and Validation of Heminested RT-PCR and qRT-PCR Assays for Comprehensive Detection of Rabies Virus in the Suspected Rabid Brain and Saliva Samples. ARCHIVES OF CLINICAL INFECTIOUS DISEASES, 14(2), 0-0. SID. https://sid.ir/paper/756172/en

    Vancouver: Copy

    AMIRI FERESHTEH, Fadajan Zohreh, Rasooli Azadeh, SALAHSHOURIFAR IMAN, BASHAR ROUZBEH, ARAB BAFERANI MASOUMEH, FAZELI MARYAM. Development and Validation of Heminested RT-PCR and qRT-PCR Assays for Comprehensive Detection of Rabies Virus in the Suspected Rabid Brain and Saliva Samples. ARCHIVES OF CLINICAL INFECTIOUS DISEASES[Internet]. 2019;14(2):0-0. Available from: https://sid.ir/paper/756172/en

    IEEE: Copy

    FERESHTEH AMIRI, Zohreh Fadajan, Azadeh Rasooli, IMAN SALAHSHOURIFAR, ROUZBEH BASHAR, MASOUMEH ARAB BAFERANI, and MARYAM FAZELI, “Development and Validation of Heminested RT-PCR and qRT-PCR Assays for Comprehensive Detection of Rabies Virus in the Suspected Rabid Brain and Saliva Samples,” ARCHIVES OF CLINICAL INFECTIOUS DISEASES, vol. 14, no. 2, pp. 0–0, 2019, [Online]. Available: https://sid.ir/paper/756172/en

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