## 4 Case Studies
### Case Study 2--Onion  
This case study considers \(a_{\mathrm{w}}=0.974\), \(0.982\) and \(0.990\), resulting in relative humidity scenarios of \(\varphi=97.4\)%, \(98.2\)% and \(99\)%, respectively. The growth of _Aeromonas hydroplilia_, _Bacillus cereus_, _Salmonella_ and _Shigella flexneri_ was considered. The analysis of this vegetable starts at \(T=2\)\({}^{\circ}\)C, the temperature at which _Aeromonas hydrophilia_ starts replicating, ending at \(T=18\)\({}^{\circ}\)C, for the reasons already mentioned. This is the temperature value at which the relative humidity scenario is more conducive to microbial growth, and the food safety threshold is reached in less than 24 h, for the full range of combined pH values, by reaching the infective dose of _Hydroplilia aeromonas_. Regarding the acidity levels in the study, pH values inherent to the onion itself were considered, comprised between \(5.3\leq\mathrm{pH}\leq 5.9\). The sampling of the results of remaining shelf life for onion is shown in Table 7.  
When, at the lowest analysed temperature (\(T=2\)\({}^{\circ}\)C), equal pH values in different water activities are compared, a large disparity caused by the different relative humidity scenarios is verified. For example, when \(T=2\)\({}^{\circ}\)C and pH = 5.4, for \(a_{\mathrm{w}}=0.974\), \(0.982\) and \(0.990\), the remaining shelf lives are, respectively, \(t=1580.4\), \(728.8\) and \(354.6\) h. Alternating between minimum and maximum relative humidity scenarios implies a decrease of \(\Delta t=1225.8\) h in the estimated remaining shelf life. However, this differential decreases significantly as storage temperatures increase. When \(T=10\)\({}^{\circ}\)C and pH = 5.6, shelf life is limited to \(t=173.4\), \(90.2\) and \(44.8\) h for \(a_{\mathrm{w}}=0.974\), \(0.982\) and \(0.990\), respectively, a difference of \(\Delta t=128.6\) h between the most extreme scenarios. At the highest temperature among those observed (\(T=18\)\({}^{\circ}\)C), for the same pH value, contamination by different bacteria is registered, depending on the water activity value in question. At a pH = 5.8  
\begin{table}
\begin{tabular}{c c c c c} \hline \hline \multirow{2}{*}{\(T\) (\({}^{\circ}\)C)} & \multirow{2}{*}{pH} & \multirow{2}{*}{\(a_{\mathrm{w}}\)} & **Remaining Shelf** & \multirow{2}{*}{**Contamination by**} \\  & & & **Life (\(t\), in Hours)** & \\ \hline \multirow{2}{*}{1} & 5.8 & 0.996 & 155.20 & _Listeria monocytogenes_ \\ \cline{2-5}  & 5.9 & 0.996 & 146.00 & _Listeria monocytogenes_ \\ \hline \multirow{2}{*}{4} & 5.8 & 0.996 & 79.60 & _Listeria monocytogenes_ \\ \cline{2-5}  & 5.9 & 0.996 & 73.60 & _Aeromonas hydrophilia_ \\ \hline \multirow{2}{*}{10} & 5.9 & 0.996 & 22.10 & _Aeromonas hydrophilia_ \\ \cline{2-5}  & 6.0 & 0.996 & 20.50 & _Aeromonas hydrophilia_ \\ \hline \hline \end{tabular}
\end{table}
Table 6: Sampling of results of DSS—Remaining shelf life of lettuce.  
when \(a_{\rm w}=0.974\), there is contamination by _Salmonella_. If, in turn, \(a_{\rm w}=0.982\) or \(0.990\), contamination is set by reaching the infective dose of _Aeromonas hydrophilia_.  
At the most unfavourable scenario for bacterial growth, that is, the minimum water activity, temperature, and pH values (\(a_{\rm w}=0.974\); \(T=2\)\({}^{\circ}\)C; pH = 5.3), the shelf life is reached \(t=1580.4\) h after the horticultural batch entrance into storage, at the threshold of contamination by _Aeromonas hydrophilia_, as it is the only bacterium capable of replicating under the selected conditions. On the other hand, when the selected water activity, temperature and pH values are maximum for the range under study (\(a_{\rm w}=0.990\); \(T=18\)\({}^{\circ}\)C; pH = 5.9) and, therefore, provide the most favourable scenario for bacterial replication, shelf life is dictated when the contamination threshold is reached, and also by _Aeromonas hydrophilia_, after \(t=11\) h of storage. In this sample, no cases of contamination by _Shigella flexneri_ were predicted.