## 3 Discussion  
The detection of MVCs could be of great interest in medical diagnosis. The extraction of metabolites during the growth phase allows higher concentrations to be obtained although a bacterial culture step is necessary. Notably, the presence of the culture medium constituting a complex matrix often of indefinite chemical composition, is an obstacle to the detection of volatile bacterial metabolites. In addition, the volatile metabolic profile of bacteria includes compounds present at very low concentrations. The detection and identification of these molecules represent a real challenge. This work focused on statistically separating the different molecules identified in order to preserve only those produced by _S. aureus_. A methodology based on the screening without a priori by GC-MS following the extraction and concentration of compounds by SPME or SBSE/HSSE was developed. The SPME extraction using PDMS/DVB and CAR/PDMS fibers allowed to extract four volatile metabolites (all already described in the literature), while the SBSE/HSSE extraction using EGS and PDMS twisters(r) allowed the extraction of 17 volatile and semi-volatile metabolites from a bacterial culture medium. SBSE/HSSE has therefore a better extraction capacity than SPME. We also demonstrated the robustness of the method by analyzing 12 independent replicates and developing a process for the treatment of obtained GC data that has shown its effectiveness in statistically determining the compounds produced by bacteria. The diversity of results between the two techniques clearly shows the need for standardization of analytical methods and the use of replicates to obtain reliable results. Indeed, since the 1980s, many articles have described advances in the metabolic profile of _S. aureus_. In 2013, [PERSON] et al. [24] compiled a review of 15 studies on volatile bacterial metabolites produced by this bacterium. They identified 111 compounds and classified them into three main groups: produced compounds, not produced compounds, and compounds with conflicting results or showing little evidence. Despite the number of studies, only 10 metabolites were considered as produced by the bacterium, 86 metabolites were still awaiting confirmation and 15 have been specifically researched in _S. aureus_ culture and never recovered. Comparison between the literature and our results is summarized in Figure 3 on the basis of the three identical groups. Ten compounds tentatively identified by our method have already been described in the literature. Seven of them are in agreement with previous works (3-methylbutanoic acid, ethanol and acetic acid as produced compounds, (methyldisulfanyl)methane as not produced, and benzaldehyde, 3-methylbutanal and (methyltrisulfanyl)methane as potentially produced or consumed). The three other compounds did not match with the literature. Regarding acetaldehyde, its highly volatile nature makes it difficult to detect and quantify under these conditions [29]. So, this compound is found in our study as consumed by _S. aureus_. In addition, propan-2-one and 1H-indole, that have been  
Figure 3: SBSE/HSSE contribution to the detection of volatile/semi-volatile metabolites produced by _S. aureus_. In green: produced compounds; in orange: medium compounds, neither produced nor consumed during growth; and in red: consumed compounds. \({}^{\text{a}}\) detected by SPME, \({}^{\text{b}}\) detected by SBSE/HSSE, and \({}^{\text{c}}\) detected by both methods. described controversially [30; 31; 32; 33; 34; 35; 36], have been found as produced compounds. Contradictory results between studies might be explained by several variables like the different strains of _S. aureus_ used, the growth medium and the parameters of cultures, the measurements performed at different time points of growth, and the type of method used for the volatile extraction. Furthermore, the SBSE/HSSE method proposed in our study detected nine new molecules from _S.