Comprehensive In Vitro and Technological Characterization of <i>Pediococcus acidilactici</i> <scp>NITDSCH2</scp> : Isolation, Probiotic Efficacy, and Safety for Fermented Food Production

ABSTRACT Escalating foodborne diseases and food spoilage have driven the overreliance on synthetic preservatives, antibiotics and fungicidal agents. This overreliance, in turn is associated with severe health risks such as antimicrobial resistance, allergic reactions, and other long‐term health issues. In light of these challenges, probiotics have emerged as a promising alternative by targeting pathogens and spoilage microorganisms while enhancing food safety and human health. This study reports the isolation and characterization of Pediococcus acidilactici NITDSCH2, a probiotic strain isolated from “Churpi”, an indigenous fermented food of Northeast India, for its potential applications in the food industry. P. acidilactici NITDSCH2 exhibited thermostable, broad‐spectrum antimicrobial activity, effectively inhibiting both bacterial and fungal pathogens. The strain also displayed excellent survivability under simulated gastrointestinal conditions, with only a minor reduction of 0.64 log units when subjected to pancreatic stress during oro‐gastrointestinal transit assay. Safety profiling revealed γ ‐hemolysis, no DNase activity, and Multiple Antibiotic Resistance Index below the 0.2 safety threshold, supporting its suitability for Generally Recognized as Safe status. In addition, the strain produced a substantial amount of lactic acid (26.37 ± 2 mg/mL). While previous studies on churpi‐derived probiotics have mainly isolated from Western Himalayan region, this study explore a largely unexplored Northeast Indian niche and provide the first evidence of antifungal activity of P. acidilactici against Sclerotinia sclerotiorum. Overall, Pediococcus acidilactici NITDSCH2 emerges as a promising candidate for functional food development and biocontrol agent, offering a sustainable solution to food spoilage and health challenges.