Hormetic Promotion of Biofilm Growth by Polyvalent Bacteriophages at Low Concentrations

Interactions between bacteriophages (phages) and biofilms are poorly understood despite their broad ecological and water quality implications. Here, we report that biofilm exposure to lytic polyvalent phages at low concentrations (i.e., 10²-10⁴ phages/mL) can counterintuitively promote biofilm growth and densification (corroborated by confocal laser scanning microscopy (CLSM)). Such exposure hormetically upregulated quorum sensing genes (by 4.1- to 24.9-fold), polysaccharide production genes (by 3.7- to 9.3-fold), and curli synthesis genes (by 4.5- to 6.5-fold) in the biofilm-dwelling bacterial hosts (i.e., Escherichia coli and Pseudomonas aeruginosa) relative to unexposed controls. Accordingly, the biofilm matrix increased its polysaccharide and extracellular DNA content relative to unexposed controls (by 41.8 ± 2.3 and 81.4 ± 2.2%, respectively), which decreased biofilm permeability and increased structural integrity. This contributed to enhanced resistance to disinfection with chlorine (bacteria half-lives were 6.08 ± 0.05 vs 3.91 ± 0.03 min for unexposed controls) and to subsequent phage infection (biomass removal was 18.2 ± 1.2 vs 32.3 ± 1.2% for unexposed controls), apparently by mitigating diffusion of these antibacterial agents through the biofilm. Overall, low concentrations of phages reaching a biofilm may result in unintended biofilm stimulation, which might accelerate biofouling, biocorrosion, or other biofilm-related water quality problems.