Persistent Immune Responses after Ebola Virus Infection

To the Editor:

Ebola virus is a highly virulent emerging pathogen and a causative agent of viral hemorrhagic fever.¹ Studies of the pathogenesis of Ebola virus infection in humans have indicated that recovery is largely dependent on the development of an immune response.^1-3 To investigate the persistence of immune response in humans, we examined levels of cytokine expression after in vitro whole-blood stimulation in persons 12 years after infection with the Gulu strain of Sudan Ebola virus (SUDV-gul).

The study was conducted in accordance with the Declaration of Helsinki and was approved by the Uganda Ministry of Health, the Uganda National Council for Science and Technology, and the ethics committee of the Uganda Virus Research Institute. All participants provided written informed consent.

We obtained whole-blood samples from six survivors of the Ebola virus outbreak that occurred in 2000 in the Gulu district of Uganda,⁴ as well as from four persons in the local community who were not infected. To obtain data on fresh whole-blood stimulation assays, despite the geographically remote nature of this cohort, reaction tubes were preloaded with antigen on site and incubated with whole blood. Plasma supernatants then were transported to the laboratory for analysis. Enzyme-linked immunosorbent assays (ELISAs) for detection of anti–SUDV-gul IgG antibody levels in serum and a viral plaque-reduction neutralization test with the lowest serum dilution inhibiting 80% or more of the plaques (PRNT₈₀) were performed.⁵

After samples obtained from survivors of Ebola virus infection underwent whole-blood stimulation with inactivated SUDV-gul and purified glycoprotein of SUDV-gul (GP_1-649), there was a significant increase in levels of cytokine expression associated with regulators of the proinflammatory and T-cell–derived immune response. Cytokine levels were measured in the plasma supernatants of samples after whole-blood stimulation in the field. As compared with the samples obtained from the control group, in the samples obtained from the survivors of Ebola virus infection there were significant increases in levels of interleukins 1α, 1β, 5, 8, 10, 12p70, and 15 and tumor necrosis factor α and interferon-γ. The levels of interleukins 2 and 17 also increased, but as compared with the samples from the control group, these increases were not significant, perhaps because of the sample size.

Survivors of Ebola virus infection and persons in the noninfected control group had similar baseline expression of cytokines and similar increased expression after stimulation with phytohemagglutinin leukoagglutinin. There were no differences in complete blood count, general medical history, or treatment during infection among persons who had been infected with Ebola virus.

Table 1. Table 1. Results of Enzyme-Linked Immunosorbent Assay IgG and PRNT₈₀ Assay Tests, According to Study Group.

Results of ELISA IgG against inactivated SUDV-gul and GP_1-649 revealed that all SUDV-gul survivors tested positive for both SUDV-gul and GP_1-649. No immunoreactivity was observed in the noninfected control group. PRNT₈₀ results showed that in the survivor group, four persons were positive for neutralization and two persons were negative. All serum samples obtained from noninfected persons that were not immunoreactive were negative in neutralization assays (Table 1).

Survivors of Ebola virus infection had persistent serum-neutralizing activity and IgG immunoreactivity against the viral protein GP_1-649 12 years after infection, and this protein stimulated high levels of cytokine expression after whole-blood stimulation.

Ariel Sobarzo, Ph.D.
David E. Ochayon, M.Sc.
Ben-Gurion University of the Negev, Beer-Sheva, Israel

Julius J. Lutwama, Ph.D.
Steven Balinandi, M.Sc.
Uganda Virus Research Institute, Entebbe, Uganda

Ofer Guttman, M.Sc.
Robert S. Marks, Ph.D.
Ben-Gurion University of the Negev, Beer-Sheva, Israel

Ana I. Kuehne, Ph.D.
John M. Dye, Ph.D.
U.S. Army Medical Research Institute of Infectious Diseases, Fort Detrick, MD

Victoria Yavelsky, Ph.D.
Eli C. Lewis, Ph.D.
Leslie Lobel, M.D., Ph.D.
Ben-Gurion University of the Negev, Beer-Sheva, Israel
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