The new case of Ebola Virus Disease in Beni is genetically linked to the 2018-2020 Nord Kivu/Ituri EVD outbreak and does not represent a new spillover event.**
Context
On Monday, August 15, 2022, the Institut National de Recherche Biomédicale (INRB) Beni mobile laboratory received an oropharyngeal secretion sample taken from a deceased patient who was being treated at the Beni General Reference Hospital (HGR-Beni). The sample tested positive for Ebola virus (EBOV) using the Cepheid Xpert Ebola assay with ct- values of 32.5 for viral glycoprotein (GP), and 28.2 for viral nucleoprotein (NP). For quality control and confirmation, samples were sent to the Rodolphe Mérieux INRB-Goma Laboratory on Tuesday August 16, 2022, where the analysis confirmed the EVD case.
The patient was admitted on July 23, 2022 to the Internal Medicine department of the HGR-Beni for cough, headache, polyarthralgia, intense physical asthenia dating back several days and for which she had self-medicated with non-specified drugs. She was HIV-positive on antiretroviral treatment, and she had also presented with tuberculosis for which she was receiving anti-tuberculosis treatment. On August 13, 2022, her health condition deteriorated with the onset of bronchial congestion and kidney failure, which prompted her transfer to the intensive care unit for better care. She died on August 15, 2022. In total her hospitalization lasted 23 days. The body was collected by the family the same day for burial.
On August 17, 2022, 131 contacts were listed, including 60 front-line healthcare workers and 71 co-patients. Of the 131 pre-listed contacts, 59 of the 60 HCWs are vaccinated. The vaccination status of the index case and 71 co-patients is not yet known.
Sequencing
Sequencing analysis was conducted at the Rodolphe Mérieux INRB-Goma Laboratory, thanks to genomic support provided by the Africa CDC Pathogen Genomic Initiative.
The swab sample, BEN1-22, was sequenced in triplicate (A/B/C) at the Pathogen Genomics Sequencing satellite Laboratory in Goma using the Oxford Nanopore Technologies (ONT) sequencing platform. Sequencing libraries were prepared using an amplicon sequencing approach (Mbala-Kingebeni, 2021; Grubaugh et al., 2019; Quick et al. 2017), generating identical genomes with 99.2%, 97% and 97.8% genome coverage, respectively.
The genome sequence generated from sample BEN1-22 group with other sequences from the Nord Kivu/Ituri Ebola virus disease (EVD) outbreak between 2018 and 2020. The new sequence is most closely related to a cluster of cases occurring in Beni around November/December 2018 and had six additional mutations from this cluster. If the new EVD case was the result of continued human-to-human transmission we would have expected many more mutations to have occurred during this time. Therefore, our initial findings indicate that this case likely represents a new flare-up of the 2018-2020 Nord Kivu/Ituri EVD outbreak, initiated by transmission of Ebola virus from a persistently infected survivor or a survivor who experienced a relapse. This case may be the index case, or there may be previous cases that were not detected. Epidemiological investigations are ongoing to determine the source.
Figure 1 | Time tree of BEN1-22 and 812 EBOV genome sequences from the Nord Kivu/Ituri outbreak. Tree built using the Nextstrain pipeline (Hadfield et al., 2018) BEN1-22 is linked to cases from November 2018.
Figure 2 | Maximum likelihood phylogenetic subtree of BEN1-22 and closest Nord Kivu/Ituri EBOV sequences . Six mutations separate the new case from the closest cluster of cases.
Data availability
The genome sequence is available [here](GitHub - inrb-drc/ebola-nord-kivu-aug-2022).
Partners and collaborators
Placide Mbala-Kingebeni (INRB, University of Kinshasa)
Daniel Mukadi (INRB, University of Kinshasa)
Michel Kasereka Tosalisana (MCZ/Beni)
Franck Muhindo Fikiri (HGR/Beni)
Guy Mutombo Ndongala (DPS/Nord-Kivu)
Serge Mbokani (HGR/Beni)
Fyfy Mbelu Matulu (INRB/Beni)
Janvier Kubuya Bonane (CD-DPS/Nord-Kivu)
Michel Ngimba (INRB-Goma)
Brigitte Modadra (INRB-Goma)
Elie Ishara (INRB-Goma)
Herve Viala (INRB-Goma)
Noëlla Mulopo Mukanya (INRB-Goma)
Hugo Kavunga (INRB)
Elisabeth Pukuta (INRB)
Eddy Kinganda Lusamaki (INRB, University of Kinshasa)
Amuri Aziza (INRB, University of Kinshasa)
Jean Claude Makangara (INRB, University of Kinshasa)
Emmanuel Lokilo (INRB)
Franck Edidi (INRB, University of Kinshasa)
Junior Bula Bula (INRB, University of Kinshasa)
John Otshudiema (WHO)
Raphael Lumembe (INRB, University of Kinshasa)
Gabriel Kabamba (INRB, University of Kinshasa)
Francis Chikuse (Africa CDC)
Dorcas Waruguru (Africa CDC)
Sofonias Tessema (Africa CDC)
Dieudonné Mwamba (DGLM)
Mike Wiley (University of Nebraska Medical Center [UNMC], PraesensBIO)
Catherine Pratt (UNMC)
Steve Ahuka-Mundeke (INRB, University of Kinshasa)
Jean-Jacques Muyembe (INRB, University of Kinshasa)
Statement on continuing work and analyses prior to publication
This genome is being shared pre-publication. Please note that this data is based on work in progress and should be considered preliminary. Our analyses of these data are ongoing and a publication communicating our findings on these and other published genomes is in preparation. If you intend to use these sequences prior to our publication, please communicate with Dr. Placide Mbala for coordination.
References
Hadfield[PCB2] J, Megill C, Bell SM, et al. Nextstrain: real-time tracking of pathogen evolution , Bioinformatics 2018 ; 34(23) :4121-4123. 10.1093/bioinformatics/bty407
Grubaugh ND, Gangavarapu K, Quick J, et al. An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar. Genome Biol 2019;20:8. 10.1186/s13059-018-1618-7 4 7
Mbala-Kingebeni P, Pratt C, Mutafali-Ruffin M, et al. Ebola Virus Transmission Initiated by Relapse of Systemic Ebola Virus Disease. N Engl J Med 2021; 384:1240-1247 10.1056/NEJMoa2024670 13
Quick, J., Grubaugh, N., Pullan, S. et al. Multiplex PCR method for MinION and Illumina sequencing of Zika and other virus genomes directly from clinical samples. Nat Protoc 2017;12:1261–1276. 10.1038/nprot.2017.066 2