This work investigates the efficacy of plasma-activated water (PAW) and plasma-activated acidified buffer (PAAB) on Enterobacter aerogenes in aqueous system and fruit systems. Reactive oxygen and nitrogen species in PAW have been suggested to provide antimicrobial and acidifying effects, causing the pH of treated water to drop. To isolate the effect of pH in microbial inactivation and to study the interactive effects of pH and reactive species on microbial inactivation, a citrate-phosphate buffer (pH 3.1) and PAAB (citrate-phosphate) were studied. A 1.92 ± 0.70 log CFU/mL reduction in E. aerogenes was observed in PAW, while no reduction was achieved in the buffer, suggesting that the inactivation was due to the reactive species in PAW and not the acidic pH. PAAB achieved a 5.11 ± 0.63 log CFU/mL reduction, suggesting an interactive effect of reactive species and low pH. Electrical conductivity and oxidation-reduction potential measurements suggest potential mechanisms for the greater antimicrobial efficacy of PAAB over PAW. Four surfaces of increasing roughness (glass slides, grape tomatoes, limes, and spiny gourds) were spot inoculated and washed with distilled water, PAW, buffer, and PAAB for 3 min. The smoothest surface (glass) showed the highest reduction (6.32 ± 0.43 log CFU per surface), while the roughest surface (spiny gourd) showed a significantly lower reduction (2.52 ± 0.46 log CFU per surface) when treated with PAAB. For treatment with PAW, no significant differences were observed between glass slides, limes, and spiny gourds. With PAW treatment, significantly lower reduction was observed on spiny gourds (1.70 ± 0.21 log CFU per surface) than on grape tomatoes (4.65 ± 1.34 log CFU per surface). PAW and PAAB both showed potential for their use in fresh produce sanitation.