## 3 Results
### Phylogenetic Analysis and Comparative Genomics
Regarding the VGs, significant differences (\(p<0.05\)) were found only for the human VG-content versus all other origins (Figure 4). In the next analysis, we aimed to juxtapose isolates of human and dairy origin with respect to their functional (CAZymes content) and safety (RGs-VGs) profile and elucidate whether the presence-absence patterns of these genes can distinguish isolates of different sources. The heatmap and hierarchical clustering showed an arrangement similar to that of  
Figure 4: Phylogenetic tree including the 24 _E. ficeum_ dairy isolates of this study and 161 isolates from various origins, parsed from NCBI. The outer bar graph denotes the abundance of RGs found in each strain. Alternating dark and light grey segments in the outer ring mark different hierBAPS clusters. The colored inner ring indicates the isolates’ origin according to the legend. Isolate labels with red font signify human clinical isolates form cases of bloodstream and urinary tract infections. The outlined, filled or absent orange circle at the tree’s leaves, shows whether an isolate carried vancomycin-resistance regulator gene(s) (_nurR_-like) only, regulator and vancomycin-resistance gene(s) (_nur_-like), or no vancomycin-related genes, respectively. The isolate labels of this study and of those retrieved from NCBI have a “C” and “GCA_” numerical prefix, respectively. #, number of the respective genes. the phylogenetic tree. Three major clusters were formed, which predominantly contained dairy and human isolates (clinical and GI) and a mixture of isolates, respectively (Figure 5). Statistical analysis for gene enrichment with Scoary showed no significant associations of dairy isolates with specific RGs or VGs, on the contrary, several genes, such as the _van_-like RGs and the _ecbA_ VG, showed a strong negative association (odds ratio (OR) \(<\) 1, _p_-value \(<\) 0.05). Conversely, several RGs (n = 19) and VGs (n = 4) were significantly enriched in the human-clinical (n = 13) and human-GI (n = 16) sources, while six genes were associated with both sources (OR \(>\) 1, _p_-value \(<\) 0.05). Important RGs enriched in the human sources were the vancomycin RGs (_van_-like) and regulator genes (_vanR_-like), the tetracycline RGs (_tet_-like), and the _lsaE_ MDR efflux pump. Noteworthy VGs were the _sgrA_ and _ecbA_ adhesins as well as the _scm_ adhesin. Several RGs and VGs, such as the _efmA_ efflux pump and _acm_ adhesin, were ubiquitous across all isolate origins (Figure 4). The majority of the CAZymes identified in the analyzed _E. facetium_ isolates belonged to the glycoside hydrolase (GH) families with the GH43 family being predominant, while  
Figure 5: Cluster heatmap generated using a resistance, virulence and CAZyme gene presence-absence data matrix of _E. facetium_ isolated from dairy, human-clinical and human-gastrointestinal origin. 62 different families were identified. The GH43 family as well as others from different groups, e.g., Glycosyltransferase (GT) 2, Carbohydrate Esterase (CE) 4 and Polysaccharide Lyase (PL) 9, were ubiquitous among isolates. Nevertheless, our comparative analysis showed that several CAZyme families were associated with either the human or dairy sources. The GH35, GH123, and the Carbohydrate-binding module (CBM) 70 families were significantly enriched in dairy-related isolates (OR \(>\) 1, \(p\)-value \(<\) 0.05). Within the human sources (clinical & GI), the CBM61, CE12, and GH154 CAZymes were significantly enriched and they showed strong negative association in the dairy-origin isolates. The AA10 CAZyme was strongly associated only with the human-clinical source. Lastly, analysis with Traitar for predicted phenotypic characteristics showed that, irrespective of their origin, all isolates can utilize sugars, such as lactose, galactose, maltose, sucrose, and D-mannitol (Figure 6). The clustered heatmap of predicted traits indicated an overlap of dairy and human origin isolates as well as a separate human-clinical cluster.