Controlling antibiotic-resistant (ART) bacteria in cheese fermentation is important for food safety and public health. A plant-maintained culture was found to be a potential source for ART bacterial contamination in cheese fermentation. Antibiotics had a detectable effect on the ART population from contamination in the finished product. The decrease in the prevalence of antibiotic resistance (AR) in retail cheese samples from 2010 compared to data from 2006 suggested the effectiveness of targeted AR mitigation in related products.The rapid emergence of antibiotic-resistant (ART) pathogens has been a major public health concern (11). The recent findings on the prevalence of antibiotic resistance (AR) genecontaining ART commensal bacteria in a broad spectrum of ready-to-consume items (3,12,18,20) suggested that the food chain likely has served as an important avenue for dissemination of ART bacteria to the general public. From 2005 to 2006, small-scale surveillance studies using retail samples purchased from national chain grocery stores in the Columbus, OH, area revealed the presence of high levels of ART bacteria and a representative AR gene pool in most cheese samples (multiple types and brands) examined (14, 18). Various commensal bacteria, such as Streptococcus thermophilus, Pseudomonas sp., Staphylococcus sp., Lactococcus lactis, and Lactococcus sp., were identified to be the isolates carrying AR genes (18). The AR genes from certain food-borne isolates were further transmitted to human-pathogenic and residential bacteria in laboratory settings by horizontal gene transfer (HGT) mechanisms, leading to acquired resistance in the recipient cells, which suggests the functionality and mobility of the food-borne AR genes (5,12,18). The high prevalence of ART bacteria and the AR gene pool in cheese products and the identification of certain isolates carrying AR genes as lactic acid bacteria raised concerns about the potential impact of the dairy fermentation process on the emergence and amplification of ART bacteria. Thus, the objective of this study was to reveal critical control points (CCP) in reducing the number of ART bacteria in dairy fermentation for targeted mitigation.CCP assessment in pilot-plant-scale cheese making. Cheddar-type cheeses were made at the Ohio State University (OSU) pilot plant to determine the impact of starter cultures, the fermentation process, and antibiotics on the development of ART bacteria. The mesophilic culture F-DVS R-604 (culture A, containing a mix of L. lactis subsp. lactis and L. lactis subsp. cremoris) and the thermophilic culture F-DVS RSF-621 (culture B, containing L. lactis, Lactobacillus helveticus, and S. thermophilus), both from the Chr. Hansen Company (Milwaukee, WI), were used in the study. Cultures were activated by inoculating 2 to 3 g of a frozen pellet into 940 ml shelf-stable ultrahigh-temperature (UHT) milk (Parmalat 2% reduced-fat milk; Farmland Dairies, Wallington, NJ) and incubated at 33°C (mesophilic) or 38°C (thermophilic) for 18 h. Raw milk was obtained ...