Methodological bases of differential detection of the EBV1/EBV2 and HHV6A/HHV6B

• Main Authors: M. Popkova, O. Utkin, E. Soboleva, N. Sakharnov, D. Bryzgalova, A. Senatskaia, E. Kulova
• Format: Article
• Language: Russian
• Published: Sankt-Peterburg : NIIÈM imeni Pastera 2019-06-01
• Series: Infekciâ i Immunitet
• Subjects: ebv1; ebv2; hhv6a; hhv6b; viral load; pcr; genotyping; differential detection.
• Online Access: https://www.iimmun.ru/iimm/article/view/1661

In Russia, of the whole variety of EBV- and HHV6-etiology diseases, only infectious mononucleosis is subject to official statistical reporting, which significantly complicates an objective assessment of their etiological structure, incidence rate, characteristics of the development of the epidemic process. Currently, information on the genetic heterogeneity of EBV, even at the level of the main types (EBV1 and EBV2), as well as HHV6A and HHV6B, their prevalence and clinical significance are limited mainly by foreign data. In Russia, there are isolated publications devoted to this issue. At the same time, the study of the circulation of genetic types (variants) and the use of this information in the implementation of epidemiological surveillance of some other infections have already become routine practice. One of the key issues is the level of development of laboratory support for molecular genetic monitoring. The purpose of this work was to improve the methodological base of differential detection of HHV6A/B and the main types of EBV. The material for the study was peripheral blood leukocytes of children aged 1-15 years with acute infectious mononucleosis (n = 50) and without clinical symptoms of this disease (n = 29). The detection and quantification of EBV DNA and HHV6 DNA was performed using real-time PCR. For differential determination of EBV1/EBV2 and HHV6A /HHV6B, an optimized one-round PCR with electrophoretic detection of amplification products in an agarose gel was used. According to the results of our own research, the frequency of detection of  EBV DNA and HHV6 DNA in acute infectious mononucleosis was 74% and 72%, and in the control group - 35% and 74%, respectively. It was found that among the examined children of the Nizhny Novgorod region, EBV1 and HHV6B prevail in the viral population, which is consistent with existing views about their geographical distribution in the adjacent territories. EBV2 was found in a single sample in the control group only. HHV6A was not detected in any of the studied groups. The methodological approach optimized in this work makes it possible to separately detect HHV6A/HHV6B and the main types of EBV according to a single laboratory protocol, and in combination with an additional stage of DNA concentration increases the diagnostic sensitivity of PCR analysis, minimizes the proportion of discordant and false negative results. Such an integrated approach can be applied for diagnostic, epidemiological and research purposes.