Abstract
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. 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. 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. 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. The developed method for viral RNA quantification is a promising tool for the specific activity control of the HFRS vaccine.
Highlights
Hemorrhagic fever with renal syndrome (HFRS) holds a leading place among natural focal human diseases in Russian Federation
The immunogenicity of experimental vaccines was determined by the induction of neutralizing antibodies in BALB/c mice
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
Summary
Егорова М.С.1, Курашова С.С.1, Ишмухаметов А.А.1,2, Баловнева М.В.1, Девяткин А.А.3, Сафонова М.В.4, Ожерелков С.В.1, Хапчаев Ю.Х.1, Балкина А.С.1, Белякова А.В.1, Дзагурова Т.К.1, Ткаченко Е.А.1. Цель исследования – разработка чувствительной и специфичной методики проведения ПЦР в реальном времени с целью количественного определения вирусной РНК в инактивированной вакцине, а также установление корреляции между содержанием генетического материала вируса и выраженностью иммуногенности. Разработанная система оценки количественного содержания вирусной РНК делает перспективным её применение для контроля специфической активности вакцины против ГЛПС. Для цитирования: Егорова М.С., Курашова С.С., Ишмухаметов А.А., Баловнева М.В., Девяткин А.А., Сафонова М.В., Ожерелков С.В., Хапчаев Ю.Х., Балкина А.С., Белякова А.В., Дзагурова Т.К., Ткаченко Е.А. Разработка метода количественного определения вирусной РНК для контроля специфической активности вакцины против геморрагической лихорадки с почечным синдромом. – разработка концепции и дизайна исследования, анализ данных, написание текста; Ткаченко Е.А.
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