Processed cheese is a dairy product with multiple end-use applications, where emulsifying salts play a fundamental role in physicochemical changes during production. Moreover, some of these salts may be a strategy to control spoilage and pathogenic microorganisms, contributing to safety and shelf life extension. This study aimed to evaluate the in vitro inhibitory activity of two emulsifying salts (ESSP = short polyP and BSLP = long polyP) against Bacillus thuringiensis CFBP 3476 and Clostridium perfringens ATCC 13124, and to compare the in situ effects of two emulsifying salts treatments (T1 = 1.5% ESSP and T2 = 1.0% ESSP + 0.5% BSLP) in processed cheeses obtained by two different methods (laboratory- and pilot-scales), during 45-day storage at 6 °C. C. perfringens ATCC 13124 growth was not affected in vitro or in situ (p > 0.05), but both of the treatments reduced B. thuringiensis CFBP 4376 counts in the tested condition. Counts of the treatments with B. thuringiensis CFBP 3476 presented a higher and faster reduction in cheeses produced by the laboratory-scale method (1.6 log cfu/g) when compared to the pilot-scale method (1.8 log cfu/g) (p < 0.05). For the first time, the inhibitory effect of emulsifying salts in processed cheeses obtained by two different methods was confirmed, and changes promoted by laboratory-scale equipment influenced important interactions between the processed cheese matrix and emulsifying salts, resulting in B. thuringiensis CFBP 4376 growth reduction.