Evidence of aberrant anti-epstein-barr virus antibody response, though no viral reactivation, in people with post-stroke fatigue

Abstract

Background: Fatigue is a common complication of stroke that has a significant impact on quality of life. The biological mechanisms that underly post-stroke fatigue are currently unclear, however, reactivation of latent viruses and their impact on systemic immune function have been increasingly reported in other conditions where fatigue is a predominant symptom. Epstein-Barr virus (EBV) in particular has been associated with fatigue, including in long-COVID and myalgic encephalomyelitis/chronic fatigue syndrome, but has not yet been explored within the context of stroke.

Aims: We performed an exploratory analysis to determine if there is evidence of a relationship between EBV reactivation and post-stroke fatigue.

Methods: In a chronic ischemic stroke cohort (> 5 months post-stroke), we assayed circulating EBV by qPCR and measured the titres of anti-EBV antibodies by ELISA in patients with high fatigue (FACIT-F < 40) and low fatigue (FACIT-F > 41). Statistical analysis between two-groups were performed by t-test when normally distributed according to the Shapiro-Wilk test, by Mann-Whitney test when the data was not normally distributed, and by Fisher's exact test for categorical data.

Results: We observed a similar incidence of viral reactivation between people with low versus high levels of post-stroke fatigue (5 of 22 participants (24%) versus 6 of 22 participants (27%)). Although the amount of circulating EBV was similar, we observed an altered circulating anti-EBV antibody profile in participants with high fatigue, with reduced IgM against the Viral Capsid Antigen (2.244 ± 0.926 vs. 3.334 ± 2.68; P = 0.031). Total IgM levels were not different between groups indicating this effect was specific to anti-EBV antibodies (3.23 × 10⁵ ± 4.44 × 10⁴ high fatigue versus 4.60 × 10⁵ ± 9.28 × 10⁴ low fatigue; P = 0.288).

Conclusions: These data indicate that EBV is not more prone to reactivation during chronic stroke recovery in those with post-stroke fatigue. However, the dysregulated antibody response to EBV may be suggestive of viral reactivation at an earlier stage after stroke.

Keywords: Epstein-Barr virus; Fatigue; Ischaemic stroke.

Fig. 1

Fatigue and mood of cohort. (A) Fatigue, measured by the Functional Assessment of Chronic Illness Therapy – Fatigue (FACIT-F) tool, with threshold for high versus low fatigue demarcated by dotted lines (B) mood, as measured by the Stroke Impact Scale (SIS), and correlation between fatigue (FACIT-F) and mood (SIS3). Each data point represents an individual participant, n = 22 per group. Data show individual participants plus mean; analysed by (A, B) Mann-Whitney test and (C) linear regression, with best-fit line and 95% confidence band plotted; ****P < 0.0001

Fig. 2

Similar frequencies of EBV viraemia in people with and without post-stroke fatigue. (A) Frequency of circulating EBV measured by qPCR. (B) EBV load assessed in the subset of participants positive for EBV quantified with the delta-delta Ct method, using the housekeeping gene provided by the Norgen EBV PCR kit. (C) Stroke severity, measured by NIHSS, between participants with and without detectable circulating EBV by qPCR. Low fatigue participants are plotted as green squares and high fatigue as orange circles. Open shapes indicate participants with EBV viremia, as measured by qPCR. Data show individual participants plus mean, (A) n = 22, (B) n = 5–6, (C) n = 9–27 per group; (B) unpaired t-test; (C) Mann-Whitney test

Fig. 3

Anti-EBV seroresponse in people with post-stroke fatigue. (A) Schematic of expected circulating anti-EBV antibody titres following primary infection, and during latency, and reactivation. Created with BioRender. (B-G) Plasma antibodies to EBV were measured by ELISA. Low fatigue participants are plotted as green squares and high fatigue as orange circles. Open shapes indicate participants with EBV viremia, as measured by qPCR, and the dotted line (B, E, G) indicates the ELISA cut off control value. (B) Titre of anti-Epstein-Barr nuclear antigen 1 (EBNA1) IgG between participants with low versus high post-stroke fatigue. (C) Percent of participants within each group that are seropositive for anti-EBNA1 IgG. Seropositivity is considered any value > 10% over the cut-off control value. (D) Association between self-reported fatigue (FACIT-F) and IgG anti-EBNA1 titre. (E) Titre of IgG anti-viral capsid antigen (VCA) between low and high fatigue groups. (F) Titre of IgG anti-VCA between participants with and without EBV viremia, measured by qPCR. (G) Titre of IgM anti-viral capsid antigen (VCA) between low and high fatigue groups. Data show individual data points plus mean, (B, C, E, G) n = 20, (D) n = 12–30 per group.; *p < 0.05; (E) unpaired t-test (B, F, G) Mann-Whitney test; (D) linear regression, with best-fit line and 95% confidence band plotted

Fig. 4

IgM anti-VCA titre correlates with fatigue but not selected clinical characteristics. Titre of IgM against VCA were plotted against various characteristics, including (A) FACIT-F score, (B) age, (C) biological sex, (D) days from stroke to baseline, (E) mood as measured by SIS3, and (G) infarct volume. Each data point represents an individual participant, (A, B, D, E, F) n = 40, (C) n = 14–26. Analysed by linear regression with best-fit line and 95% confidence band plotted (A-B, D-F)

Fig. 5

Total circulating antibodies unchanged between low and high post-stroke fatigue groups. Amount of circulating IgM and total IgG antibodies, as well as C-reactive protein (CRP). Data show individual participants plus mean, n = 20 per group; analysed by Mann-Whitney test