Development of an early warning system for Nipah outbreak prevention: on-site inactivation, PCR surveillance and sequencing in Bangladesh

Summary Background Mobile laboratory diagnostic technologies for Nipah virus outbreak prevention, mitigation and response remain limited, despite the critical need for such capacities in remote, low-resource regions where most cases occur. We aim to address this gap by implementing a workflow that includes method development, laboratory validation, and field demonstration of a mobile Nipah virus complex diagnostic solution. Methods We developed a flexible mobile laboratory workflow incorporating PCR capacity, a novel amplicon-based sequencing protocol, and a validated Nipah virus inactivation procedure. Following development and validation, we demonstrated the feasibility of this workflow through repeated field sampling of bat colonies in Nipah virus endemic regions of Bangladesh across multiple field campaigns. Findings We demonstrated the feasibility of this system for early outbreak response and as a potential early warning tool prior to the emergence of human cases. We detected two urine samples from flying foxes that tested positive and performed full-scale on-site analysis, including qPCR diagnostics and NGS sequencing, within 24 hours. Interpretation As highlighted in the present study, active surveillance enables outbreak prevention by identifying bat colonies that are actively shedding viruses in real time, even in rural settings. Also, this method can provide rapid, on-site sequence data to track and better understand the genomic diversity of Nipah virus in natural reservoirs during both outbreak and non-outbreak periods. In this study we aimed to establish the foundations of a standard procedure for safe and rapid field testing of Nipah virus in remote areas. Funding This study was funded by the National Research, Development and Innovation Fund of Hungary – Grant details: Strengthening of Regional Research Infrastructure, Capacity Building and Pandemic Preparedness (2024-1.2.3-HU-RIZONT-2024-00041). Research in context Evidence before this study We searched the PubMed database with the following search term without fixed timeframe: (“Nipah virus” OR Nipah) AND (Bangladesh OR India OR Southeast-Asia) AND (“mobile laboratory” OR “field laboratory” OR “point-of-care” OR portable OR onsite) The search yielded 12 publications that address specific elements of mobile laboratory solutions and provide opinions on the necessity of mobile technologies (point-of-care testing, rapid diagnostics). One technological paper was identified in relation to a bus-based mobile laboratory solution in India, which is also referenced in our manuscript. Further manuscripts are related to the molecular and clinical diagnostics of Nipah virus, and a genome sequencing protocol validated on clinical samples was also identified. The health risk posed by Nipah virus can be reduced by investments in research, development and technology, as described in the literature. However, the available techniques can currently be interpreted as separate modules and a comprehensive diagnostic method validated in terms of safety, complex technical solutions and usability in real world conditions is still lacking. Added value of this study This complex study, which ranges from laboratory development to validation of methods and demonstration in real-world conditions, is the first of its kind. There is currently a limited number of methodologies in the literature and a significant need, which is also included in the WHO regional Nipah virus epidemic prevention plan. The development of rapid on-site surveillance techniques, starting from validated virus inactivation protocol, through the on-site real-time RT-PCR screening and amplicon-based sequencing method represents a significant advancement in early detection and monitoring of NiV. This approach facilitates swift responses in the early stages of human cases and contact tracing strategies or identification present of virus in bat colonies even before human cases emerge, bolstering outbreak prevention efforts. Implication of all the available evidence Our study highlights the need for a critically important technology integration. As the use of mobile laboratories expands and the demand for rapidly deployable, mobilizable technologies grows, there is an increasing need to move toward standardized operational approaches. The first step is to test different technologies in real-world situations beyond the development phase and provide alternatives for use during or in the prevention of epidemic situations.