Metabolic Comparison and Molecular Networking of Antimicrobials in Species.

are well-known for producing bioactive secondary metabolites, with numerous antimicrobials essential to fight against infectious diseases. Globally, multidrug-resistant (MDR) microorganisms significantly challenge human and veterinary diseases. To tackle this issue, there is an urgent need for alternative antimicrobials. In the search for potent agents, we have isolated four species PC1, BT1, BT2, and BT3 from soils collected from various geographical regions of the Himalayan country Nepal, which were then identified based on morphology and 16S rRNA gene sequencing. The relationship of soil microbes with different species has been shown in phylogenetic trees. Antimicrobial potency of isolates was carried out against American Type Culture Collection (ATCC) 43300, ATCC 25931, ATCC 14028, ATCC 700603, and ATCC 25922. Among them, species PC1 showed the highest zone of inhibition against tested pathogens. Furthermore, ethyl acetate extracts of shake flask fermentation of these strains were subjected to liquid chromatography-tandem mass spectrometric (LC-MS/MS) analysis for their metabolic comparison and Global Natural Products Social Molecular Networking (GNPS) web-based molecular networking. We found very similar metabolite composition in four strains, despite their geographical variation. In addition, we have identified thirty-seven metabolites using LC-MS/MS analysis, with the majority belonging to the diketopiperazine class. Among these, to the best of our knowledge, four metabolites, namely cyclo-(Ile-Ser), 2-n-hexyl-5-n-propylresorcinol, 3-[(6-methylpyrazin-2-yl) methyl]-1H-indole, and cyclo-(d-Leu-l-Trp), were detected for the first time in species. Besides these, other 23 metabolites including surfactin B, surfactin C, surfactin D, and valinomycin were identified with the help of GNPS-based molecular networking.