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ARS Home » Southeast Area » Athens, Georgia » U.S. National Poultry Research Center » Exotic & Emerging Avian Viral Diseases Research » Research » Research Project #444708

Research Project: Development and Validation of Oxford Nanopore Technologies (ONT) Sequencing as a Molecular Point-of-Care Diagnostic (POCD) Assay

Location: Exotic & Emerging Avian Viral Diseases Research

Project Number: 6040-32000-081-033-R
Project Type: Reimbursable Cooperative Agreement

Start Date: Jun 1, 2023
End Date: May 31, 2025

The 2022 highly pathogenic avian influenza (HPAI) outbreak demonstrated that the speed of response is critical for the success of outbreak control. This includes the speed of identification and characterization of the infectious agent along with results sharing. The current diagnostic tools against foreign animal diseases (FAD), including Real-time PCR and sequencing, are sensitive, specific and accurate. However, the turnaround time between sample collection to generation and sharing of the results still needs days to be completed, which represents one of the major bottlenecks in any outbreak control process. Point-of-care diagnostics (POCD) is viewed as a potential solution to alleviate this bottleneck. The Overall objective of this project is to adopt Oxford Nanopore Technology (ONT), a third-generation sequencing platform, as a molecular POCD for avian FAD (IAV-A and vND) in the field or in resource-limited setups. Achieving this will provide accurate and rapid results to producers and decision makers to act effectively and timely. The successful adoption of ONT as a molecular POCD can be used not only for avian FAD, but has the potential to be used in a variety of applications, including at the borders to aid in controlling all FAD. This project explores new technology in an applied approach to strengthen the nation’s capacity to respond to any future outbreak of FAD.

By the end of this project, we aim to customize and validate Oxford Nanopore Technology sequencing as a Point-of-care diagnostics (POCD) that can directly provide accurate and reliable diagnosis of avian FAD (IAV-A and vND) in the field or resource limited setups with simple workflow that would allow easy use of it. Four specific objectives will serve as milestones towards achieving the goal of this project: Objective 1-Evaluate the suitability and reliability of different ONT workflow for field application (RNA extraction, library preparation and data analysis); Objective 2-Assembly of Mobile-Sequencing Kit (MSK) and test the newly validated protocols under field conditions (at farm level or resource limited lab); Objective 3-The development of a Portable-Bioinformatics Pipeline (PBP) for the real-time data analysis on local computing resources independent of internet connection; Objective 4-Head-to-head comparison of results generated from clinical samples submitted to one of NALHN labs and the newly validated POCD protocols.