ABSTRACT Modulator therapies have improved outcomes for people with cystic fibrosis (pwCF), and currently, more than 50% of pwCF are over the age of 18. This has resulted in an increased prevalence of atypical pathogens, including non-tuberculous mycobacteria (NTM). CF-isolation rates of NTM and Pseudomonas aeruginosa are high, and those co-colonized have worse clinical outcomes. We therefore investigated the behavior of these two organisms in a dual-species biofilm. We found that co-culture of Mycobacterium abscessus (MAB) promoted biofilm formation by P. aeruginosa . Confocal imaging revealed changes in biomass and structural organization of the P. aeruginosa biofilm during co-culture with MAB. DNase treatment slightly decreased dual-species biofilm, but biofilm formation was abrogated in a Pel-deficient mutant of P. aeruginosa . Moreover, MAB stimulated the overproduction of Pel polysaccharide in dual-species biofilms with P. aeruginosa . Furthermore, dual-species cultures promoted tolerance of P. aeruginosa to tobramycin treatment. Finally, transcriptomics analysis revealed that MAB alters the expression of genes that impact biofilm formation in P. aeruginosa . Overall, our findings highlight a Pel-dependent interaction between P. aeruginosa and M. abscessus that may result in bacterial persistence for pwCF during antibiotic therapy. IMPORTANCE Advances in treatments for cystic fibrosis have increased the incidence of “non-typical” infections, including those caused by non-tuberculous mycobacteria (NTM). NTM are often found in the lungs alongside the pathogen Pseudomonas aeruginosa , and patients infected with both tend to have worse clinical outcomes. In this study, we examined how these two organisms interact under biofilm conditions, an important lifestyle adaptation in chronic pulmonary infections. We found that when grown together, Mycobacterium abscessus enhanced biofilm formation by P. aeruginosa and reduced the effectiveness of antibiotic treatment. Our findings reveal a Pel-dependent interaction between two important lung pathogens that may contribute to more severe infections and poorer treatment outcomes in people with cystic fibrosis or other chronic lung diseases. Understanding these mechanisms opens new opportunities for developing strategies to combat these complex infections.
McDaniel et al. (2026) studied this question.