APPLICATION OF NUCLEIC ACID SEQUENCE-BASED AMPLIFICATION FOR THE DETECTION OF VIABLE FOODBORNE PATHOGENS: PROGRESS AND CHALLENGES DAVID RODRÍGUEZ-LÁZARO 1,4 , MARTA HERNÁNDEZ 2 , MARTIN D’AGOSTINO 3 and NIGEL COOK 3 1 Division of Veterinary Pathology, Infection and Immunity Faculty of Medical and Veterinary Sciences University of Bristol Langford, Bristol BS40 5DU, U.K. 2 Laboratorio de Biología Molecular Subdirección de Investigación y Tecnología Instituto Tecnológico Agrario de Castilla y León, Spain 3 Central Science Laboratory Sand Hutton, York, U.K. Accepted for Publication May 18, 2006 ABSTRACT Nucleic acid sequence-based amplification (NASBA) is a sensitive transcription-based amplification system that uses a battery of three enzymes (avian myeloblastosis virus reverse transcriptase, RNase H and T7 RNA polymerase) leading to a main amplification product of single-stranded RNA, and is specifically designed for the specific detection of RNA. NASBA is an established diagnostic tool in clinical use, with a theoretically bigger analyti- cal sensitivity than reverse transcription-polymerase chain reaction (RT-PCR) for pathogen detection, but is not progressing toward implementation in food analysis. This is unfortunate, because it has a potential for detection of viable cells through selective amplification of messenger RNA, even in a background of genomic DNA, which PCR does not possess. However, in some instances, an unexpected amplification of genomic DNA has been observed using the NASBA technique. The availability of methods for rapid, sensitive and selec- tive detection of viable microbial pathogens in foods is a goal worth pursuing, 4 Corresponding author. TEL: +44-0-117-331-9007; FAX: +44-0-117-928-9505; EMAIL: david.rodriguez@bris.ac.uk Journal of Rapid Methods & Automation in Microbiology 14 (2006) 218–236. All Rights Reserved. © 2006, The Author(s) Journal compilation © 2006, Blackwell Publishing 218