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BRIEF REPORT
The interaction of EBOV with the human immune system re- EBOV GP–Specific T-Cell Responses
mains incompletely understood. We here report new insight To define EBOV-specific cellular immune responses, we investi-
into the long-lasting immune responses of an EVD survivor fol- gated the induction of 4 cytokines (IFN-γ, IL-2, TNF-α, and
lowing recovery. MIP-1β) in response to EBOV GP–specific OLP stimulation,
Ten days after first symptoms, the patient was transferred to via ICS (Figure 2). Antigen-specific T cells were predominantly
Hamburg, Germany, and treated in an isolation unit for indi- found in the CD8+ compartment and not in the CD4+ compart-
viduals with highly contagious diseases. The initial clinical ment. Whereas TNF-α/IFN-γ double producers were predomi-
course has been previously described [7]. He experienced a se- nantly observed upon SP and GP2 stimulation, GP1a
vere EBOV infection with the associated complications of sep- stimulation resulted in TNF-α induction (Figure 2A and 2B).
ticemia, encephalopathy, and respiratory failure [7]. After Only limited expression was observed for MIP-1β, and no induc-
recovery and EBOV-negative results of polymerase chain reac- tion was detectable for IL-2. Limited polyfunctionality (maximal-
tion analysis of plasma, he was transferred to the infectious dis- ly 3 functions) was observed in CD8+ T cells in response to GP2
ease unit (Supplementary Figure 1). Blood specimens were and SP peptide pools (Figure 2C), and none were observed in
collected in EDTA-lined tubes on days 37 and 46 after illness. CD4+ T cells (data not shown). Antigen-specific cytotoxic lym-
phocytes were characterized by staining with CD107a, with the
Lymphocyte Dynamics
Initial analysis of CD4+, CD8+, and CD19+ lymphocyte dynam- strongest induction observed in the GP1a pool (Figure 2B).
ics revealed minor differences in their respective frequencies as To increase sensitivity, EBOV-specific T-cell responses were
compared to findings for healthy controls (SF2). Subsequently, also assessed by IFN-γ ELISPOT (Figure 2D). Significant
the expression of CCR7 and CD45RA was evaluated (Figure 1A). responses were observed for peptide pools GP2.2 (376 spot-
While antigen-experienced effector memory (EM) T cells forming cells [SFCs] per million peripheral blood mononuclear
(CCR7−CD45RA−) were only slightly increased in the CD4+ cells [PBMCs] on day 37) and SP (202 SFCs per million PBMCs
compartment (21% on days 37 and 46, compared with a on day 37), confirming the ICS data described above. A higher
mean [±SD] of 13% ± 1.5% in controls), the proportion of response was observed for the master pool MP1.
EM CD8+ T cells expanded, compared with controls (39% on In summary, both ICS and ELISPOT revealed EBOV-specific
day 37 and 36% on day 46, compared with a mean [±SD] of antiviral T-cell activity, albeit with a low magnitude.
19% ± 7% in controls).
DISCUSSION
T-cell activation is a critical element of the antiviral immune
defense. We therefore assessed activation via HLA-DR and EBOV represents one of the most deadly human pathogens, and
CD38 staining (Figure 1B). Analysis of CD8+ T cells revealed to date little is understood about the human immunity to EBOV
a high frequency of activated cells, composing 68% (on day infection. Immunopathogenesis, factors that influence survival,
37) and 62% (on day 46) of the main CD8+ T-cell population, and correlates of protection have not been completely
compared with a mean (±SD) of 8% ± 4% among controls, and elucidated.
Figure 1. Lymphocyte dynamics during the convalescent phase of Ebola virus (EBOV) disease (EVD). A, CD4+ (left) and CD8+ (right) T-cell subsets were stained by
CCR7 and CD45RA to distinguish naive (CCR7+CD45RA+), effector memory (EM; CCR7−CD45RA−), central memory (CM; CCR7+CD45RA−), and effector memory RA
(CCR7−CD45RA+) T cells. In the upper row, data are represented in contour plots. The control plot depicts a representative experiment. The lower row shows the
distribution of T-cell subsets. The control pie chart represents the median value of measured results for 5 controls. B, T-cell activation was analyzed by HLA-DR and
CD38 staining to investigate CD4+, CD8+, and the respective EM T cells. At left, contour plots show the gating strategy. At right, the graph depicts the results (control
values are for 5 individuals). C, Cytotoxic T cells were assessed via GrzB staining. At left, contour plots show results for the patient with EVD on day 37 after symptom
onset for CD4+ (upper plot) and CD8+ (lower plot) T cells. Results are highlighted in the graph (red triangle, EVD survivor; circle, 2 healthy controls). D, Regulatory B
cells are enriched in transitional B cells (CD24+CD38high; blue gate; upper plot) and the CD19+CD24hiCD27+ population (lower plot). At left, contour plots depict rep-
resentative plots for the patient with EVD (on day 37) and 1 healthy control. At right, results are highlighted in the graph (data are for 5 controls). Values from controls
are shown as median values with SDs.
Figure 2. Ebola virus (EBOV) glycoprotein (GP)–specific T-cell responses found in CD8+ T cells. A, EBOV GP–specific interferon γ (IFN-γ)/tumor necrosis factor α
(TNF-α) T-cell responses. At left, graphs depict the observed IFN-γ/TNF-α responses in CD8+ (upper graph) and CD4+ (lower graph) compartments specific for the 4
EBOV GP peptide pools (control values are for 2 individuals). At right, representative dot plots show EBOV GP–specific IFN-γ/TNF-α T-cell responses for both CD8+
(upper panel) and CD4+ (lower panel) T cells (data are for patients with EVD on day 37 after first appearance of EVD symptoms). B, Cytokine and cytotoxic T-
lymphocyte responses upon stimulation with EBOV GP peptide pools (control values are for 2 individuals). C, The distribution of the functionality of CD8+ T-cell
responses to the 4 EBOV GP peptide pools on days 37 and 46 after first appearance of EVD symptoms. D, IFN-γ enzyme-linked immunospot assay of responses
against EBOV GP peptide pools. A CEF pool (consisting of peptides from cytomegalovirus, Epstein-Barr virus, and influenza virus) was used as positive control. The
assay was performed in duplicate. The graph shows median values and SDs (control values are for 2 individuals). Abbreviations: PBMC, peripheral blood mono-
nuclear cell; SFC, spot-forming cell.