Pan-Filovirus Serum Neutralizing Antibodies in a Subset of Congolese Ebolavirus Infection Survivors

Abstract

One year after a Zaire ebolavirus (EBOV) outbreak occurred in the Boende Health Zone of the Democratic Republic of the Congo during 2014, we sought to determine the breadth of immune response against diverse filoviruses including EBOV, Bundibugyo (BDBV), Sudan (SUDV), and Marburg (MARV) viruses. After assessing the 15 survivors, 5 individuals demonstrated some degree of reactivity to multiple ebolavirus species and, in some instances, Marburg virus. All 5 of these survivors had immunoreactivity to EBOV glycoprotein (GP) and EBOV VP40, and 4 had reactivity to EBOV nucleoprotein (NP). Three of these survivors showed serologic responses to the 3 species of ebolavirus GPs tested (EBOV, BDBV, SUDV). All 5 samples also exhibited ability to neutralize EBOV using live virus, in a plaque reduction neutralization test. Remarkably, 3 of these EBOV survivors had plasma antibody responses to MARV GP. In pseudovirus neutralization assays, serum antibodies from a subset of these survivors also neutralized EBOV, BDBV, SUDV, and Taï Forest virus as well as MARV. Collectively, these findings suggest that some survivors of naturally acquired ebolavirus infection mount not only a pan-ebolavirus response, but also in less frequent cases, a pan-filovirus neutralizing response.

Figure 1.

A, Serological assessment of donor reactivity against Zaire ebolavirus (EBOV) glycoprotein (GP) and nucleoprotein using a commercially available enzyme-linked immunosorbent assay and VP40 reactivity assessment using a luciferase immunoprecipitation assay. B, Reactivity of plasma antibodies to recombinant GP of ebolavirus (EBOV), Bundibugyo ebolavirus, Sudan ebolavirus, or Marburg ebolavirus, antibody titers for 4 assessed plasma specimens, which demonstrated multiple filovirus neutralization ability. C, Reactivity of polyclonal antibodies that were affinity purified from plasma of the subject with the most broadly reactive antibodies, subject ES 2. Data in B–D represent the mean ± standard deviation of technical triplicates. D, Representative binding curves for the 4 plasma specimens with the broadest GP reactivity profiles. Abbreviations: BDBV, Bundibugyo ebolavirus; EBOV, Ebola virus; ELISA, enzyme-linked immunosorbent assay; HA, hemagglutinin; LIPS, luciferase immunoprecipitation assay; MARV, Marburg virus; NP, nucleoprotein; OD₄₅₀, optical density 450 nanometers; pAb, polyclonal antibody; SUDV, Sudan ebolavirus; ZEBOV, Zaire ebolavirus.

Figure 2.

A, Ebolavirus species (ES) donor serum neutralizing activity against Zaire ebolavirus (EBOV), Taï Forest ebolavirus (TAFV), Sudan ebolavirus (SUDV), Bundibugyo ebolavirus (BDBV), or Marburg virus (MARV) or the arenavirus Lassa virus, using a vesicular stomatitis virus pseudovirus assay expressing a species-specific glycoprotein. Neutralization was assessed in technical triplicate using patient serum at a 1:50 dilution. B, Neutralizing activity in donor serum for EBOV or BDBV using the G-Luc entry inhibition assay. Neutralization was assessed in technical triplicate using patient serum at a 1:50 dilution. C, Plaque reduction neutralization antibody assay using live EBOV, measured in triplicate for each dilution. Results shown in colors indicate the ability to reduce EBOV plaque-forming units by at least 50% for those individuals that demonstrated a multiple filovirus neutralization pattern.

Figure 3.

Competition-binding assay with plasma antibodies and receptor binding site (RBS)–specific monoclonal antibodies (mAbs) (5 µg/mL) using cell surface–displayed Marburg virus (MARV) glycoprotein (GP). Subject ebolavirus species (ES) 2 plasma revealed the capacity to significantly reduce binding of RBS-specific mAbs to Jurkat-MARV GP at 1:25 and 1:50 dilutions, whereas normal plasma did not. Mean ± standard deviation of assay triplicates. Dashed line indicates MARV-specific mAb-only control and shows maximal binding to MARV GP. Background was determined from binding of MARV-specific mAb to an untransfected cell line that does not express GP. Results are expressed as the percentage of MARV-specific antibody binding in the presence of plasma relative to MARV-specific mAb-only control minus background.