Effects of Fermentation Period on Metabolite Production and Bioactivity in Two Sponge-Associated Bacteria

Many marine bacteria are living in association with sponges and are known to produce secondary metabolites for the host's chemical defense. The secondary metabolite production in bacteria is influenced by the culture medium, growth conditions, and incubation time. In this study, the effect of fermentation period on secondary metabolite production was analyzed in two sponge-associated bacteria, Enterobacter sp. and Alcanivorax sp. The bacterial strains were subjected to a fermentation period of five days and then a 14-day period before centrifugation and solvent extraction from the supernatant. The extracts from the bacteria were then tested for antibacterial and antibiofilm activity against selected clinically significant bacterial strains. The composition of the extracts was confirmed using GC-MS. The results revealed variations in the activity of the extracts at different fermentation periods with differences in the composition of the metabolites observed in each extract. Enterobacter sp. cultured for 14 days of fermentation period showed higher antibacterial activity against Staphylococcus aureus ATCC 25923 and Escherichia coli ATCC 25922. While each extract demonstrated antibiofilm activity, in some cases the activity was higher after 5 days and in some cases after 14 days. GC-MS profiling clearly indicated a metabolic shift with the dominance of fatty alcohols and esters in the extracts obtained after 5 days and bioactive aromatic compounds and diketopiperazines after 14 days of fermentation. Overall, this study confirms that secondary metabolite enrichment in marine bacteria is fermentation time-dependent, and a longer fermentation period may enhance antibacterial activity. The changes in metabolites due to the fermentation period showed the significance of optimizing bacterial culture time to enhance the bioactive potentials of bacteria associated with marine organisms.