## 3 Results and Discussion  
In this study, the strains _E. coli_ ATCC25922, _En. faecalis_ ATCC29212, and _P. aeruginosa_ ATCC27853 were used as indicator strains to test the efficacy of H\({}_{2}\)O\({}_{2}\) to process ice. The species _E. coli_, _En. faecalis_, and _P. aeruginosa_, generally transmitted by ice [31], represent the main pathogens used for the evaluation of the mandatory and accessory parameters provided by the Italian Legislative Decree No. 31/2001 [32] for the implementation of directive 98/83/EC. The results of survival tests, expressed as CFU/100 mL of thawed ice, are reported in Figure 2.  
After bacterial inoculation, the cell densities of control ice samples (0 OX) were 2.21, 2.10, and 2.26 Log CFU/100 mL for _E. coli_ ATCC25922, _En. faecalis_ ATCC29212, and _P. aeruginosa_ ATCC27853, respectively. In more detail, Figure 2 shows that the antibacterial activity of the food ice activated with different concentrations of H\({}_{2}\)O\({}_{2}\) was comparable among the different strains tested. No significant differences (\(p\) > 0.05) were found among the levels of microbial count for the trials 0 OX and 4 OX between the same bacteria strains, showing that the presence of 4 mg/L of H\({}_{2}\)O\({}_{2}\) in the food ice did not exert any disinfecting action. In contrast, the counts of the three bacteria decreased significantly (\(p\) < 0.001) with increasing H\({}_{2}\)O\({}_{2}\) concentration. In particular, _E. coli_ ATCC25922, _En. faecalis_ ATCC29212, and _P. aeruginosa_ ATCC27853 decreased consistently in the 8 OX trial and were not detected in the 12 OX samples, showing the bactericidal efficacy of H\({}_{2}\)O\({}_{2}\) at the concentration of 12 mg/L against both Gram-positive (_En. faecalis_) and Gram-negative (_E. coli_ and _P. aeruginosa_) bacteria commonly searched in water to establish its drinkability [33].  
[PERSON] et al. [34] demonstrated the effect of H\({}_{2}\)O\({}_{2}\) at 0.5% in combination with other products (benzalkonium chloride, cationic polymer, ethoxylated fatty alcohol, and ethyl alcohol) used in food industries against _E. coli_, _Staphylococcus aureus_, _Enterococcus hirae_, and _P. aeruginosa_ on stainless steel surfaces. However, the authors found differences in terms of bacterial sensitivity and evidenced a more pronounced effect of the mixtures used when the test organisms were _En. hirae_ and _P. aeruginosa_. [PERSON] et al. [35] demonstrated that H\({}_{2}\)O\({}_{2}\) at different concentrations were more than quaternary ammonium compounds effective against _S. aureus_ and _P. aeruginosa_ biofilms. Furthermore, [PERSON] et al. [36] demonstrated that a concentration of 0.5% H\({}_{2}\)O\({}_{2}\) represented a highly efficient antimicrobial strategy to disinfect medical devices. Regarding food applications of H\({}_{2}\)O\({}_{2}\), [PERSON] et al. [37] used several concentrations of this compound for raw cow's milk preservation, demonstrating that 0.14% of H\({}_{2}\)O\({}_{2}\) was suitable to extend the milk shelf-life where milk cooling facilities were not available. Hence, H\({}_{2}\)O\({}_{2}\) seems to be a proper compound to contain intestinal bacteria that can be found in water/food ice [5] at low cost.  
Figure 2: Counts of water-borne pathogenic bacteria in food ice activated with different concentrations of H\({}_{2}\)O\({}_{2}\). \(p\) value: \(\ast\ast\)\(p\) < 0.001; n.s.—not significant. The letters a, b, and c on the histograms indicate difference between the samples analyzed according to [PERSON]’s test.