Microbial glycoside hydrolases as antibiofilm agents with cross-kingdom activity.

Galactosaminogalactan and Pel are cationic heteropolysaccharides produced by the opportunistic pathogens and , respectively. These exopolysaccharides both contain 1,4-linked -acetyl-d-galactosamine and play an important role in biofilm formation by these organisms. Proteins containing glycoside hydrolase domains have recently been identified within the biosynthetic pathway of each exopolysaccharide. Recombinant hydrolase domains from these proteins (Sph3 from and PelA from ) were found to degrade their respective polysaccharides in vitro. We therefore hypothesized that these glycoside hydrolases could exhibit antibiofilm activity and, further, given the chemical similarity between galactosaminogalactan and Pel, that they might display cross-species activity. Treatment of with Sph3 disrupted biofilms with an EC of 0.4 nM. PelA treatment also disrupted preformed biofilms with EC values similar to those obtained for Sph3 In contrast, Sph3 was unable to disrupt Pel-based biofilms, despite being able to bind to the exopolysaccharide. Treatment of hyphae with either Sph3 or PelA significantly enhanced the activity of the antifungals posaconazole, amphotericin B, and caspofungin, likely through increasing antifungal penetration of hyphae. Both enzymes were noncytotoxic and protected A549 pulmonary epithelial cells from -induced cell damage for up to 24 h. Intratracheal administration of Sph3 was well tolerated and reduced pulmonary fungal burden in a neutropenic mouse model of invasive aspergillosis. These findings suggest that glycoside hydrolases can exhibit activity against diverse microorganisms and may be useful as therapeutic agents by degrading biofilms and attenuating virulence.