Queso Fresco (QF), a type of Hispanic-style soft and fresh cheese, is a popular food in Latin-American diet. Because of its high moisture content, near neutral pH, and moderate salt content, QF provides an optimal substrate for the growth of spoilage and pathogenic microorganisms. Currently, there are no effective commercial technologies to reduce microorganisms in soft cheeses such as QF. High voltage atmospheric cold plasma (HVACP) is a novel, non-thermal technology, which can be used to treat packaged food products and achieve significant reduction of pathogenic and spoilage microorganisms without compromising products’ qualities. In this study, various gases, such as dry air and MA50 (50% CO₂, 50% N₂), were evaluated for inactivation of Listeria innocua (LI), a non-pathogenic surrogate for Listeria monocytogenes, in QF by HVACP treatment. Survival LI after HVACP treatments were enumerated on Listeria selective agar and by thin agar layer method for injured cell enumeration. Quality effects were analyzed for lipid oxidation, pH, color, texture and moisture content. Plasma characterization was done using optical emission spectrometry. The results have shown that a maximal of 4.9 log₁₀ cfu/g LI reduction was achieved after dry air HVACP treatment. A slight increase in lipid oxidation was observed in QF samples after HVACP treatment in dry air. Negligible changes were observed in color, texture, moisture content and pH after HVACP treatments. Moreover, after direct HVACP treatments in dry air, LI population decreased with longer storage at 4°C. After 28 d, 1.4 log₁₀ cfu/g LI population was found on treated QF while 7.4 log₁₀ cfu/g was observed in control samples. Lastly, HVACP treatment in dry air reduced and inhibited the growth of background microorganisms. After 28 d of storage at 4°C, less than 2.0 log₁₀ cfu/g background microorganisms was observed in treated QF, while more than 6.0 log₁₀ cfu/g was recorded in control samples. The results demonstrate the efficacy of HVACP treatment for LI inactivation in QF and the potential of HVACP technology for non-thermal processing of delicate dairy products.