Advances and challenges in pretreatment strategies for nucleic acid test-based rapid detection of foodborne bacteria: a critical review

Foodborne bacteria are a significant contributor to foodborne diseases worldwide, and effective prevention requires accurate early detection and continuous monitoring. Nucleic acid test (NAT)-based detection methods have been extensively studied and applied due to their high specificity and rapid turnaround time. However, most NAT-based detection systems cannot achieve ideal sensitivity or short assay times without efficient sample pretreatment. Although reviews have individually addressed methods for nucleic acid (NA) extraction and methods for increasing the concentration of foodborne bacteria at low bacterial loads such as pre-enrichment and pre-concentration, a comprehensive review that compiles and critically analyzes all pretreatment steps as an integrated process has not yet been reported. This review summarizes key considerations and advances in all pretreatment stages, including sample preparation, enhancement of bacterial abundance, and NA extraction. Particular emphasis is placed on the compatibility and potential integration of each pretreatment steps and with NAT, while critically addressing practical challenges that are often overlooked. The evaluations presented in this review aim to enable the effective application of NAT for the detection of foodborne bacteria and to facilitate the development of fully automated workflows and field-deployable diagnostic systems.