Secondary metabolites produced during Aspergillus fumigatus and Pseudomonas aeruginosa biofilm formation

Abstract In Cystic Fibrosis (CF), mucus plaques are formed in the patient’s lung, creating a hypoxic condition and a propitious environment for colonization and persistence of many microorganisms. There is clinical evidence showing that Aspergillus fumigatus can co-colonize CF patients with Pseudomonas aeruginosa , which has been associated with lung function decline. P. aeruginosa produces several compounds with inhibitory and anti-biofilm effects against A. fumigatus in vitro; however, little is known about the fungal compounds produced in counterattack. Here, we annotated fungal and bacterial secondary metabolites (SM) produced in mixed biofilms in normoxia and hypoxia conditions. We detected nine SMs produced by P. aeruginosa . Phenazines and different analogs of pyoverdin were the main compounds produced by P. aeruginosa , and their secretion were increased by the fungal presence. The roles of the two operons responsible for phenazines production ( phzA1 and phzA2 ) were also investigated showing both mutants are able to produce partial sets of phenazines. We detected a total of 20 SMs secreted by A. fumigatus either in monoculture or in co-culture with P. aeruginosa . All these compounds are secreted during biofilm formation either in normoxia or hypoxia. However, only eight compounds (demethoxyfumitremorgin C, fumitremorgin, ferrichrome, ferricrocin, tricetylfusigen, gliotoxin, gliotoxin E, and pyripyropene A) were detected during the biofilm formation by the co-culture of A. fumigatus and P. aeruginosa upon both normoxia and hypoxia conditions. Overall, we showed how diverse is SM secretion during A. fumigatus and P. aeruginosa mixed culture and how this can affect biofilm formation both in normoxia and hypoxia. Author Summary The interaction between Pseudomonas aeruginosa and Aspergillus fumigatus has been well-characterized in vitro . In this scenario, the bacterium exerts a strong inhibitory effect against the fungus. However, little is known about the metabolites produced by the fungus to counterattack the bacteria. Our work aimed to annotate secondary metabolites (SM) secreted during co-culture between P. aeruginosa and A. fumigatus during biofilm formation in both normoxia and hypoxia. The bacterium produces several different types of phenazines and pyoverdins, in response to the fungus presence. In contrast, we were able to annotate 29 metabolites produced during A. fumigatus biofilm formation but only eight compounds were detected during biofilm formation by the co-culture of A. fumigatus and P. aeruginosa upon both normoxia and hypoxia. In conclusion, we have detected many SMs secreted during A. fumigatus and P. aeruginosa biofilm formation. This analysis can provide several opportunities to understand the interaction between these two species.