Phthalate
esters (PAEs) are commonly released from plastic pipes
in some water distribution systems. Here, we show that exposure to
a low concentration (1–10 μg/L) of three PAEs (dimethyl
phthalate (DMP), di-n-hexyl phthalate (DnHP), and di-(2-ethylhexyl)
phthalate (DEHP)) promotes Pseudomonas biofilm formation and resistance to free chlorine. At PAE concentrations
ranging from 1 to 5 μg/L, genes coding for quorum sensing, extracellular
polymeric substances excretion, and oxidative stress resistance were
upregulated by 2.7- to 16.8-fold, 2.1- to 18.9-fold, and 1.6- to 9.9-fold,
respectively. Accordingly, more biofilm matrix was produced and the
polysaccharide and eDNA contents increased by 30.3–82.3 and
10.3–39.3%, respectively, relative to the unexposed controls.
Confocal laser scanning microscopy showed that PAE exposure stimulated
biofilm densification (volumetric fraction increased from 27.1 to
38.0–50.6%), which would hinder disinfectant diffusion. Biofilm
densification was verified by atomic force microscopy, which measured
an increase of elastic modulus by 2.0- to 3.2-fold. PAE exposure also
stimulated the antioxidative system, with cell-normalized superoxide
dismutase, catalase, and glutathione activities increasing by 1.8-
to 3.0-fold, 1.0- to 2.0-fold, and 1.2- to 1.6-fold, respectively.
This likely protected cells against oxidative damage by chlorine.
Overall, we demonstrate that biofilm exposure to environmentally relevant
levels of PAEs can upregulate molecular processes and physiologic
changes that promote biofilm densification and antioxidative system
expression, which enhance biofilm resistance to disinfectants.