Rapid laboratory confirmation of the 2025 Sudan Virus Disease (Svd) index case in Uganda.

BACKGROUND: On January 30, 2025, the Uganda Ministry of Health declared an outbreak of Sudan virus disease (SVD) following laboratory confirmation from postmortem samples of a suspected case identified through routine mortality surveillance for viral hemorrhagic fevers (VHFs) at Mulago National Referral Hospital, Kampala. This report describes the laboratory procedures used to confirm the 2025 SVD index case, emphasizing the vital role of rapid diagnostics in containing VHFs.

METHODS: We leveraged existing surveillance infrastructure to collect, package, transport, test, and report results for the index-confirmed case. Testing was conducted within the framework of the established laboratory quality and bio-risk management system at the Central Emergency Response and Surveillance Laboratory. The workflow consisted of sample delivery, reception, preparation, nucleic acid extraction, master mix preparation, quantitative reverse transcriptase polymerase chain Reaction (RT-qPCR), differential diagnosis for VHFs, parallel PCR testing and typing for Sudan virus (SUDV) confirmation, result validation, and reporting.

RESULTS: The sample tested negative for Marburg, Yellow fever, Rift Valley fever, and Crimean-Congo Hemorrhagic Fever viruses. However, it tested positive for SUDV. Four parallel tests utilizing the Real Star® Filovirus Screen RT-PCR Kit 1.0 (Altona Diagnostics GmbH, Hamburg, Germany) on CFX96 Bio-Rad platforms, with varying sample volumes (140 µL, 90 µL, 50 µL, and 30 µL diluted with nuclease-free water), confirmed the presence of SUDV. The 50 µL sample had the lowest Ct values for the Orthoebolavirus target, while the 30 µL sample had the lowest Ct values for the internal control target. The turnaround time, from sample reception to results reporting to the Ministry of Health leadership, was less than 12 h.

CONCLUSION: Mortality surveillance of VHFs is vital for the swift identification of Filovirus outbreaks in high-risk areas. Comprehensive VHFs panel testing is crucial for detecting pathogens in suspected cases. Sample dilution impacts diagnostic sensitivity, making optimized testing protocols crucial for precise molecular diagnostics. A rapid turnaround time is critical to outbreaks, enabling timely public health decisions such as case isolation, contact tracing, and resource allocation.