MIXED-SPECIES BIOFILMS AND RESISTANCE TO HEAVY METALS AND DISINFECTANTS: IMPLICATIONS FOR UPEC AND UTIS

Abstract

Intro: About 80% of urinary tract infections (UTIs) are caused by uropathogenic E. coli (UPEC) in young, sexually active women, and polymicrobial infections are common. It is established that mixed-species biofilms can cause persistent and recalcitrant infections, and complicate therapy. The aim of the study was to examine the effect of heavy metals (Cu2+, Zn2+, Ag+, and Hg+), peracetic acid (PAA), and hexadecyl trimethyl benzyl ammonium bromide (HDTMA-Br) on single and mixed-species biofilm containing UPEC strains. Methods: Cultures of four strains each of E. coli, K. pneumoniae, P. aeruginosa, and S. aureus were grown in microtiter plates at 37oC for 3 days with shaking, and biofilm production was quantified by the crystal violet assay. Bacteria, in triplicates, were examined singly and in duplex (1:1 ratio), triplex (1:1:1), and quadruplex (1:1:1:1) combinations, and data were analyzed using Microsoft Excel. Findings: All strains were characterized as high or medium biofilm producers, and most (96%) maintained this in duplex and triplex combinations in the absence of stress. However, in quadplex combinations, only one combination was classified as medium biofilm production: all others were low. UPEC strains were intolerant (no biofilm) of heavy metal and disinfectant stress. However, when combined with one other bacterial species, biofilm production was significantly enhanced (to high). With two other species, average biofilm production hovered about the high cut-point but was lower than when UPEC was co-cultured with one other species. In quadplex cultures, there was a significant reduction in biofilm production, however, medium levels of biofilm were observed in the presence of heavy metals (Zn2+, Ag+, Hg+). Conclusion: UPEC strains only benefitted from the presence of 1 or 2 other species in tolerating stresses, and are likely due to cooperation/synergism among the 2-3 species co-cultured and/or antagonism/competition when 4 species were co-cultured. Multispecies biofilms could pose a significant risk for treatment failures in UTIs.