# A Lab-Made E-Nose-MOS Device for Assessing the Bacterial Growth in a Solid Culture Medium
## 3 Results and Discussion
### Quantification of Bacteria CFUs Using the E-Nose-MOS Lab-Made Device  
As previously mentioned, few studies report the use of E-noses as semi-quantitative [23; 26] or quantitative [27; 28] tools to assess the levels of bacteria. In this sense, the present study also aimed to evaluate the quantitative performance of the lab-made E-nose-MOS for determining the number of each type of bacterium (i.e., CFUs) in the BHI agar medium. For that, MLRM were developed relating the decimal logarithmic of the CFUs (log\({}_{10}\)(CFU)) of each one of the four bacteria as a multiple linear function of the selected feature extracted parameters derived from the E-nose-MOS response towards the VOCs emitted by every single bacterium grown in solid medium. The predictive models' performances were evaluated based on two goodness of fitting parameters (\(R^{2}\) and RMSE) for the LOO-CV variant. Table 3 shows the goodness of fitting data as well as the information regarding the concentration range studied and the number of parameters included in each developed MLRM, which were selected by the SA algorithm. \begin{table}
\begin{tabular}{c c c c c c c} \hline \hline \multirow{3}{*}{**Actual Bacterium**} & \multicolumn{6}{c}{**Predicted Bacterium**} \\ \cline{2-7}  & \multicolumn{2}{c}{**Gram-Negative**} & \multicolumn{2}{c}{**Gram-Positive**} & \multirow{2}{*}{**Total**} & \multirow{2}{*}{**Sensitivity**} \\ \cline{3-3} \cline{5-6}  & _E. coli_ & & _P. aeruginosa_ & & _E. faecalis_ & & _S. aureus_ \\ \hline \multirow{2}{*}{Gram-negative} & _E. coli_ & 9 & 0 & 0 & 1 & 10 & 90\% \\  & _P. aeruginosa_ & 1 & 8 & 0 & 1 & 10 & 80\% \\ \hline \multirow{2}{*}{Gram-positive} & _E. faecalis_ & 0 & 1 & 9 & 0 & 10 & 90\% \\  & _S. aureus_ & 0 & 0 & 0 & 10 & 10 & 100\% \\ \hline \multirow{2}{*}{Total} & Total & 10 & 9 & 9 & 12 & 40 & 90\% \\  & Specificity & 90\% & 89\% & 100\% & 83\% & 91\% & — \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Confusion matrix for the internal validation classification (LOO-CV) performance of the E-nose-MOS-LDA-SA model concerning the discrimination of the four studied bacteria (_E. faecalis_, _S. aureus_, _E. coli_; and _P. aeruginosa_) grown in BHI agar medium. The satisfactory predictive (LOO-CV) values of \(R^{2}\) and RMSE (\(0.943\leq R^{2}\leq 0.994\) and \(0.158\leq\) RMSE \(\leq 0.602\) log\({}_{10}\)(CFU)), which are confirmed by the visualization of Figure 2, support the use of the lab-made E-nose-MOS for quantifying the number of CFU of each one of the four bacteria studied. Indeed, based on the RMSE values, the number of CFUs initially inoculated in BHI agar medium, could be estimated after 24 h of incubation (at 37 \({}^{\circ}\)C) with an accuracy as low as 1 to 4 CFUs, in-line with the accuracies previously reported in the literature for quantitative assessment of bacteria in food samples [27; 28]. The possibility of using the E-nose-MOS as an alternative routine tool for estimating the number of CFU grown in a solid medium was further checked following the methodology previously described by the research team [29], which is based on the XPT 90-210 French standard. For each bacterium, the decimal logarithmic of the CFU, predicted by the E-nose-MOS-MLRM were plotted versus the decimal logarithmic of the experimental number of CFU, determined by counting plate technique, and the respective parameters (slope and intercept values) of the single regression trend line were calculated. The regression analysis allowed us to verify if the slope and intercept values were statistically equal to one and zero, respectively, which would correspond to a perfect linear fit. Table 4 shows the parameters of the single linear regressions (\(R^{2}\), slope and intercept values and the respective 95% confidence intervals, CI) for the LOO-CV procedure. The results clearly demonstrate that, at 5% significance level, the slope and intercept values were statistically equal to the expected theoretic values (i.e., the slope CI included the value 1; and, the intercept CI contained the value zero).