Natural product biosynthesis capacities and anti-Candida activities in marine host-associated bacteria

Tese de mestrado, Biologia Molecular e Genética, Universidade de Lisboa, Faculdade de Ciências, 2021Numbers of drug-resistant fungal pathogens are rising, and new antifungal molecules are in great demand. The microbiomes of marine sponges and corals are prolific sources of novel natural products with diverse bioactivities of pharmaceutical value. The objective of this thesis was to identify isolates with promising secondary metabolism and antifungal activities among a collection of 70 marine animal associated bacteria. Their genomes were mined for secondary metabolite biosynthetic gene clusters (SM-BGCs) and isolates screened for antifungal activity against human-pathogenic Candida (C. albicans, C. glabrata wildtype and Δpdr1 mutant, C. auris, C. parapsilosis). Genomes presented diverse SM-BGC profiles, comprising 463 gene clusters. Among them were high numbers of antimicrobial peptide- and polyketide synthase-related SM-BGCs. Only 38 clusters showed similarity above 70% to SM-BGCs of known compounds, underlining the potential structural novelty encoded. Cross-streak assays showed that 39 isolates were active against C. glabrata, 13 of which were also active against C. albicans, C. parapsilosis and C. auris. Extracellular extracts, prepared from five highly active isolates (Aquimarina Aq135, unclassified Flavobacteriaceae RHTr2, Pseudoalteromonas Cy Y, Shimia Alg238-R152, Sulfitobacter EL44) grown at different temperatures were further investigated in disk diffusion and MIC assays. Extracts of Aquimarina sp. Aq135 grown at 20ºC, and Pseudoalteromonas sp. Cy Y grown at 10ºC showed most anti-Candida activity and were analysed by liquid chromatography-mass spectrometry. The metabolic profiles of Pseudoalteromonas Cy Y and Aquimarina Aq135 were highly distinct, and most compounds remained unidentified, highlighting this untapped metabolic reservoir. Some compounds were tentatively associated to known compound classes, among them depsipeptide-, ochratoxin- and depside-like molecule(s) in Aquimarina Aq135, and macrolide- like molecule(s) in Pseudoalteromonas Cy Y. In conclusion, this thesis identified bacterial isolates with potent anti-Candida activities and promising, novel secondary metabolism, paving the way for future studies on the structure elucidation and sustainable production of new antifungal compounds