Nipah virus molecular detection from whole blood and respiratory swabs in a rapid field-ready protocol

Nipah virus (NiV) is a highly pathogenic, zoonotic paramyxovirus with significant public health implications due to high associated mortality and potential for human-to-human transmission. Current diagnostic testing options for NiV are limited and require extensive laboratory infrastructure.

Develop a field-deployable testing workflow for timely NiV detection.

A NiV real-time RT-PCR (rRT-PCR) was designed for a highly conserved region of the nucleocapsid gene and tested with RNA from Bangladesh and Malaysia NiV strains. The NiV rRT-PCR was evaluated on Rotor-Gene Q and Palm PCR S1e thermocyclers following instrument free RNA extraction (Extract & Store).

Initial analytical evaluation, on a Rotor-Gene Q, demonstrated dynamic amplification and a limit of detection (LoD) of 3.7-4.2 copies/µL without amplification of related paramyxoviruses. The assay was adapted for the portable, battery-powered, self-contained Palm PCR S1e thermocycler, and exhibited linear detection with a LoD of 30.7 copies/µL. RNA extraction from contrived whole blood and pharyngeal swabs using the Extract & Store workflow yielded comparable results to automated extraction on a KingFisher Apex instrument. The entire assay, including extracted and stabilized RNA controls from BSL-1 strains, was successfully transferred to Aga Khan University with ambient temperature shipping and yielded similar performance.

The combination of Extract & Store and the Palm PCR S1e device offers a viable solution for field-based molecular detection of NiV. While limitations were noted for reaction setup on the Palm PCR, this presents a flexible and accessible workflow for rapid, portable detection of high-consequence pathogens in resource-constrained settings.