Filovirus Seroreactivity Beyond Virus-Specific Outbreak Regions in the Democratic Republic of the Congo, 2015-2023

The ongoing Bundibugyo virus disease (BVD) outbreak in the Democratic Republic of the Congo (DRC) is the second-largest Ebola disease outbreak in recorded history, and only the second recorded BVD outbreak in the country. While the DRC has recorded 18 filovirus disease outbreaks, the historical frequency and geographic distribution of human filovirus exposures remain poorly understood. Using a multiplexed multi-filovirus immunoassay, we assessed antibody reactivity against antigens of filoviruses linked to human disease, Bundibugyo virus (BDBV), Ebola virus (EBOV), Marburg virus (MARV), and Sudan virus (SUDV), among eleven cohorts from seven provinces between 2015-2023 (n=5,852). Participants with median fluorescence intensity values exceeding established antigen-specific cutoffs were classified as seroreactive. Seroreactivity against glycoprotein (GP) targets of all four viruses were detected in every cohort, demonstrating evidence of filovirus or an antigenically related exposure well beyond known outbreak regions. Seroreactivity estimates varied by cohort site, with 60.8% of participants (n=175/288) in Mongala Province in April 2016 considered seroreactive against BDBV GP. In this cohort, 4.2%, 29.9%, and 34.7% of participants were seroreactive against EBOV, MARV and SUDV GP, respectively yet only 1.7% were seroreactive against all four GP antigens. In adjusted analyses, site was the primary predictor of seroreactivity, with odds for BDBV seroreactivity 9.1-fold (95%CI 7.0-11.9) and 9.5-fold (95%CI 6.9-13.2) higher in Mongala and Tshuapa provinces than in Kwilu. Similarly, the adjusted odds of MARV and SUDV GP seroreactivity were 4.5-fold and 7.1-fold higher in Mongala, and 3.7-fold and 6.1-fold higher in Tshuapa, than Kwilu. Among 280 individuals sampled longitudinally, BDBV GP detected seroconversion was comparable by site, with 12.8% in Kwilu and 10.4% in Mongala having detected seroreactivity three and five years after initial contact, respectively. Overall, these results indicate widespread, geographically heterogeneous filovirus seroreactivity across sampled cohorts. These findings may reflect zoonotic spillover events, leading to undetected human-to-human filovirus circulation, including paucisymptomatic or asymptomatic infections, or exposure to as-yet unidentified antigenically cross-reactive viruses. These data highlight gaps in understanding the historical and geographic distribution of filovirus exposure in the DRC and support further prospective serological, epidemiological, and molecular investigations, especially given the current outbreak.