Authors

Discussion

Background: There is an ever-increasing need to identify and produce novel antimicrobials and anti-parasitic compounds. Public Health England estimates 10 million deaths globally by the year 2050 if the current trend of rampant antimicrobial resistance is left unchecked. Co-culturing Streptomyces with competing microorganisms is a viable method to identify novel antimicrobial and anti-parasitic compounds.

Objective: This study aims to identify novel medically useful compounds by co-culturing S. coelicolor with A. flavus and A. parasiticus.

Methods: Wild-type (WT) and mutant (M1146, M1152, M1154) S. coelicolor strains were co-cultured with A. flavus and A. parasiticus under nutrient-deplete conditions to induce secretion of secondary metabolites by S. coelicolor. 

Results: Pronounced anti-fungal activity was observed in original minimal media developed for this study. Aspergillus radial growth was reduced, with the mean diameter of colonies decreased by ~12 mm when co-cultured with all strains of S. coelicolor. Zones of inhibition for Aspergillus sporulation around a seeded Streptomyces colony was 31 ± 1.5 mm and 22 ± 6.7 mm when co-cultured with the S. coelicolor strains M1152 and M1146 respectively, a significant increase compared to when co-cultured with WT S. coelicolor (4 ± 0.6 mm; P < 0.0001, n = 3). Quantitation by florescence microscopy indicate that S. coelicolor suppress Aspergillus growth by inhibiting fungal hyphae proliferation. SEM images show penetration of Aspergilli biofilm and damage to hyphal structures by Streptomyces strains. Extracts from co-culture conditions which suppress Aspergillus growth inhibit ESKAPE pathogens. Analyses of these extracts by HPLC-MS have identified known compounds with anti-microbial activity, along with novel compounds with predicted anti-microbial activity and anti-parasitic activity. More than 4000 novel secondary metabolites were observed in ethyl acetate extracts of co-cultured supernatants. RNA sequencing of co-cultured Streptomyces mutants is underway to study expression of known and novel bacterial cryptic gene clusters. 

Conclusions: The data argue that Streptomyces species competing with Aspergillus under specialized nutrient-deplete conditions secrete secondary metabolites with broad range anti-microbial, anti-parasitic and insecticidal activity. S. coelicolor mutant, M1152, exhibits significantly increased antimicrobial activity in co-cultured conditions as compared to WT or other mutants tested in this study.