Characterization of Epstein–Barr virus genotypes in multiple sclerosis through next generation sequencing approaches

Abstract

regarded as an important pathogenetic factor that contributes to chronic neuroinflammation.We determined a relationship between EBV, HHV-6 and HHV-7 viral load in the saliva and serum levels of pro-inflammatory cytokines (IFNalpha, IFNgamma, IL-1, IL-2) in a group of ME patients. Materials: 32 ME patients (11 female, mean age 32 ± 5) were enrolled into the study. The patients were divided into groups based on viral load and on viruses detected. 30 volunteers (12 female, mean age 32.3 ± 4.7) were enrolled as controls. Real-time quantitative PCR was used to assess the viral load, and ELISA was used to assess serum cytokine levels. Mann–Whitney U-test was used for statistical analysis. Results: Mean cytokine levels were higher in ME patients than in controls. To study the impact of EBV on cytokine levels, EBV-positiveME patients (n = 19) were divided into group 1 (EBV b 4 lg copies/ml) and group 2 (EBV N 4 lg copies/ml). Mean IFNgamma level differed significantly (p b 0.01) between groups (121.26 ± 41.76 pg/ml and 13.15 ± 5.76 pg/ml), and mean IL-2 level differed (p= 0.05) between groups (33.3 ± 15.66 pg/ml and 17 ± 16.8 pg/ml). Also, in group 2 EBV load correlatedwell with IFNgamma (r =−0.5, p b 0.05) andwith IL-2 levels (r =−0.56, p b 0.025). Mean IFNgamma and IL-2 levels were lower in patientswith EBV/HHV-6 coinfection then in patientswith EBV and without HHV-6, but the difference was not significant. To study the role of HHV-7 inME, we determined HHV-7 load in 22 random patients from ME group. In these patients, HHV-7 load N 6 lg copies/ml was a good marker for EBV load N 4 lg copies/ml (sensitivity 70%, specificity 100%), and HHV-7 load correlated well with EBV load (r = 0.8524, p b 0.001). 2 groups were formed based on HHV-7 load: group 1b (HHV-7 b 6 lg c/ml), and group 2b (HHV-7 N 6 lg c/ml). Mean EBV load differed significantly (p b 0.05) between groups (3.3 ± 0.6 lg copies/ml and 5.49 ± 0.76 lg copies/ml). Conclusions: ME patients differ in Th1-cell cytokine levels (IFNgamma and IL-2) depending on EBV viral load in the saliva. This result supports the suppressive impact of EBV on IFNgamma secretion described in vitro. HHV-7 viral load has predictive value for EBV viral load in ME patients. We are planning to assess cytokines in saliva as well as in blood serum of ME patients.