Advancing public health preparedness: Establishment of Nipah virus molecular diagnostic test at the National Reference Laboratory, Sri Lanka

Abstract

Objective: To establish Nipah virus diagnostic capabilities at the National Reference Laboratory in Sri Lanka using the NIV Pune real-time PCR kit. Methods: Strict safety precautions were adhered during testing due to the high pathogenicity of the Nipah virus, with all diagnostics conducted in a BSL2+ laboratory at the Medical Research Institute in Sri Lanka. RNA extraction was performed using the QIAamp Viral RNA Mini kit. The NIV Pune in-house real-time PCR kit was employed, following established primer/probe sequences and controls. The assay was validated using the Rotor-Gene Q Series Real-time PCR platform. Results: The validation run of the Nipah virus real-time PCR test demonstrated robust performance, with positive controls consistently detecting Nipah RNA at a Ct value of 21.50±0.01. Negative controls confirmed assay specificity with an external negative control which was also used as an extraction control and showed no interference. The internal control exhibited stable behavior, enhancing confidence in PCR results. The qPCR analysis graph illustrated the successful detection of internal and positive controls, validating the reliability of the assay. Conclusions: Establishing Nipah virus diagnostic capabilities in Sri Lanka signifies a proactive and collaborative response to the persistent global health threat.

Keywords

• nipah virus
• real-time pcr
• validation
• molecular diagnostics