An adaptable microreactor to investigate the influence of interfaces on Pseudomonas aeruginosa biofilm growth

Biofilms are ubiquitous and notoriously difficult to eradicate and control, complicating human infections and industrial and agricultural biofouling. However, most of the study had used the biofilm model that attached to solid surface and developed in liquid submerged environments which generally have neglected the impact of interfaces. In our study, a reusable dual-chamber microreactor with interchangeable porous membranes was developed to establish multiple growth interfaces for biofilm culture and test. Protocol for culturing Pseudomonas aeruginosa (PAO1) on the air-liquid interface (ALI) and liquid-liquid interface (LLI) under static environmental conditions for 48 h was optimized using this novel device. This study shows that LLI model biofilms are more susceptible to physical disruption compared to ALI model biofilm. SEM images revealed a unique "dome-shaped" microcolonies morphological feature, which is more distinct on ALI biofilms than LLI. Furthermore, the study showed that ALI and LLI biofilms produced a similar amount of extracellular polymeric substances (EPS). As differences in biofilm structure and properties may lead to different outcomes when using the same eradication approaches, the antimicrobial effect of an antibiotic, ciprofloxacin (CIP), was chosen to test the susceptibility of a 48-h-old P. aeruginosa biofilms grown on ALI and LLI. Our results show that the minimum biofilm eradication concentration (MBEC) of 6-h CIP exposure for ALI and LLI biofilms is significantly different, which are 400 μg/mL and 200 μg/mL, respectively. These results highlight the importance of growth interface when developing more targeted biofilm management strategies, and our novel device provides a promising tool that enables manipulation of realistic biofilm growth. 

Key points: • A novel dual-chamber microreactor device that enables the establishment of different interfaces for biofilm culture has been developed. • ALI model biofilms and LLI model biofilms show differences in resistance to physical disruption and antibiotic susceptibility.