Molecular diversity and antimicrobial properties of extracts from different organs of Tunisian extremophilic plants

Plants are a rich source of bioactive molecules, including anti-infective compounds. Antimicrobial peptides have been found in seeds, leaves, roots, stems, and flowers. In a previous study, we reported antimicrobial activity in extracts from the leaves and roots of Astragalus armatus and Anthyllis sericea. In this study, we characterized the molecular diversity of several plant extracts using MALDI mass spectrometry and off-gel bottom-up proteomics approaches. In addition, the biological properties of the acidic extracts of the roots, leaves, and seeds of A. armatus, A. sericea, Genista microcephala, and Rhanterium suaveolens, 4 extremophilic plants from southern Tunisia, were evaluated. Their MALDI profiles showed substantial variability between plant species and organs. For the 4 plants, the seed profiles showed a higher molecular content compared to the leaf and root profiles. Furthermore, most of the masses determined for the different organs of the 4 plants were between 1 kDa and 3 kDa. This MALDI profiling revealed a large variety of molecules (401 in A. sericea, 564 in A. armatus, 398 in G. microcephala, and 300 in R. suaveolens) that remain to be further explored for their antimicrobial potential. The resulting extracts were tested against Gram-positive and Gram-negative bacteria, the filamentous fungus Aspergillus fumigatus, and the yeast Candida albicans, and showed variable antimicrobial activity. LC-MS analysis enabled the identification of 89 proteins in the plants, and the analysis of a fraction of A. artmatus roots, which successfully inhibited A. fumigatus, led to the identification and partial characterization of 3 proteins: Endochitinase 2, Thaumatin-Like protein and Peroxidase. SIGNIFICANCE: Antimicrobial resistance is one of the most pressing health challenges currently. In this context, extremophile plants represent an insufficiently explored source of bioactive molecules with potential therapeutic value. By applying proteomic approaches (MALDI-MS and LC-MS), our study provides new insights into the molecular diversity of acidic extracts from 4 Tunisian extremophile species and highlights their antimicrobial activities against resistant pathogenic bacteria and fungi. The importance of this work resides in its dual contribution: (i) expanding proteomic knowledge of Tunisian extremophile plants, which remain still poorly characterized, and (ii) opening new perspectives for the discovery of plant peptides and proteins as promising alternatives to conventional antimicrobials. This integrative approach bridges the gap between molecular characterization and functional validation, thereby contributing to the ongoing search for innovative strategies to combat multidrug resistance.