Real-time PCR assay development for the control of vaccine against hemorrhagic fever with renal syndrome

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Abstract

Introduction. Hemorrhagic fever with renal syndrome (HFRS) holds a leading place among natural focal human diseases in Russian Federation. There is no etiotropic therapy for the disease now. The vaccine prophylaxis is the most effective method to control this infection. The main criteria for inactivated vaccines evaluation are its immunogenicity and specific activity.

The study purposes were to develop a sensitive and specific real-time PCR method for viral RNA quantification in the inactivated vaccine and to study the correlation between the viral RNA amount and vaccine immunogenicity.

Material and methods. L-segment fragments of the Puumala, Hantaan, and Sochi vaccine strains were selected as diagnostic targets for oligonucleotides and fluorescent probes. The immunogenicity of experimental vaccines was determined by the induction of neutralizing antibodies in BALB/c mice.

Results. A highly specific, sensitive and reproducible real-time PCR method has been developed. The analytical sensitivity was 1.24 ± 1.5 x 102 copies/ml for Puumala virus; 1.16 ± 1.4 * 102 copies/ml for Hantaan; 1.32 ± 1.8 * 102 copies/ ml for Sochi, with a virus content of 1.5 ± 0.5 lg FFU/ml; 1.8 ± 0.5 lg FFU/ml and 2.2 ± 0.5 lg FFU/ml, respectively. The viral RNA amount in experimental vaccine preparations inactivated with β-propiolactone was proportional to the neutralizing antibodies titer observed in mice following the immunization.

Discussion. It was found that different virus inactivators differently affects the detected viral RNA amount, but not the vaccine immunogenicity, which indicates the same degree of the immunogenic proteins damage. The direct relationship between the viral RNA copy number and vaccine immunogenicity makes it possible to use this criterion for vaccine dosage preparation.

Conclusion. The developed method for viral RNA quantification is a promising tool for the specific activity control of the HFRS vaccine.

About the authors

M. S. Egorova

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Author for correspondence.
Email: masha_0787@mail.ru
ORCID iD: 0000-0002-6416-2573

Maria S. Egorova - Ph.D. (Biol.), Senior Researcher of the Laboratory of Hemorrhagic Fevers.

108819, Moscow

Russian Federation

S. S. Kurashova

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Email: svetlanak886@yandex.ru
ORCID iD: 0000-0001-9934-699X

Kurashova Svetlana Sergeevna - Researcher, Laboratory of Hemorrhagic Fevers.

108819, Moscow

Russian Federation

A. A. Ishmukhametov

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences; Institute for Translational Medicine and Biotechnology, FSAEI HE I.M. Sechenov First Moscow State Medical University of the Ministry of the Health of Russia

Email: aishmukhametov@gmail.com
ORCID iD: 0000-0001-6130-4145

Ishmukhametov Aidar Airatovich - PhD, MD,  Professor, Corresponding Member of the Russian Academy of Sciences.

108819, Moscow, 119991, Moscow

 

Russian Federation

M. V. Balovneva

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Email: mashasm@yandex.ru
ORCID iD: 0000-0003-2198-7521

Balovneva Maria Vladimirovna - PhD, Leading Researcher, Laboratory of Hemorrhagic Fevers.

108819, Moscow


Russian Federation

A. A. Deviatkin

Institute of Molecular Medicine, FSAEI HE I.M. Sechenov First Moscow State Medical University of the Ministry of the Health of Russia

Email: andreideviatkin@gmail.com
ORCID iD: 0000-0003-0789-4601

Devyatkin Andrey Andreevich - PhD, Leading Researcher, Laboratory of Molecular Biology and Biochemistry, Institute of Molecular Medicine.

119991, Moscow

Russian Federation

M. V. Safonova

FSHI Plague Control Center» of the Federal Service for Surveillance of Consumer Rights Protection and Human Wellbeing (Rospotrebnadzor)

Email: Marina-iris@mail.ru
ORCID iD: 0000-0002-5579-5694

Safonova Marina Viktorovna - Researcher, laboratory for viral infections I-II pathogenicity groups diagnostics of the Federal Public Health Institution "Anti-Plague Center".

119121, Moscow

Russian Federation

S. V. Ozherelkov

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Email: ozherelkov@yandex.ru
ORCID iD: 0000-0001-9778-0485

Ozherelkov Sergey Viktorovich – PhD, DSc, Leading Researcher, tick-borne encephalitis and other viral encephalitis laboratory.

108819, Moscow

Russian Federation

Yu. Kh. Khapchaev

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Email: hapchaev_uh@chumakovs.su
ORCID iD: 0000-0003-1613-5228

Khapchaev Yusuf Khadzhibekovich - PhD, DSc, Head of the Polio Vaccine Department.

108819, Moscow

Russian Federation

A. S. Balkina

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Email: balkina_as@chumakovs.su
ORCID iD: 0000-0002-9704-7774

Balkina Alexandra Sergeevna - Researcher, Laboratory of Hemorrhagic Fevers.

108819, Moscow

Russian Federation

A. V. Belyakova

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Email: belyakova_av@chumakovs.su

Belyakova Alla Vladimirovna - PhD, Scientific Secretary.

108819, Moscow

Russian Federation

T. K. Dzagurova

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Email: centrglps@yandex.ru
ORCID iD: 0000-0002-6656-1682

Dzagurova Tamara Kazbekovna - PhD, MD,  Leading Researcher, Head of the Laboratory of Hemorrhagic Fevers.

108819, Moscow

Russian Federation

E A. Tkachenko

FSBSI M.I. Chumakov Federal Scientific Center for Research and Development of Immune-and-Biological Products of Russian Academy of Sciences

Email: evgeniytkach@mail.ru
ORCID iD: 0000-0002-6829-1241

Tkachenko Evgeniy Aleksandrovich - PhD, MD, Professor, Scientific Superviser.

108819, Moscow

Russian Federation

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Copyright (c) 2021 Egorova M.S., Kurashova S.S., Ishmukhametov A.A., Balovneva M.V., Deviatkin A.A., Safonova M.V., Ozherelkov S.V., Khapchaev Y.K., Balkina A.S., Belyakova A.V., Dzagurova T.K., Tkachenko E.A.

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