Reporter genes and fluorescent probes for studying the colonisation of biofilms in a drinking water supply line by enteric bacteria

Abstract

Biofilms containing diverse microflora were developed on bitumen-painted steel and glass tiles suspended in a chemostat model of a water distribution system. Escherichia coli, taken from a naturally occurring biofilm, was transformed with a plasmid containing the anaerobically induced nirB promoter fused to the lacZ reporter gene. The resulting transformant, PRB1, was introduced into the chemostat. After 7 and 13 days, an E. coli strain with an anaerobically induced Lac+ phenotype was present in the biofilm. Development of an episcopic differential interference contrast technique combined with UV fluorescence microscopy enabled the simultaneous visualization of E. coli in the biofilm using a fluorescent probe to detect expression of the gusA reporter gene and a lacZ fluorescent probe to monitor anaerobic expression of beta-galactosidase from pnirB.

Detection of Escherichia coli in Biofilms from Pipe Samples and Coupons in Drinking Water Distribution Networks

“…Several studies at the lab scale have demonstrated that even in the presence of detectable levels of chlorine, E. coli can be incorporated in biofilms (34), survive in the biofilm for up to 40 days (1,17), and persist there in a metabolically active form (13,40). Some authors suggest that E. coli can even grow in drinking water distribution systems (7) because these bacteria can adapt to the oligotrophic environment.…”

“…Although some studies have shown that E. coli may grow in controlled laboratory-scale experiments (17,34,40), in general it is assumed that E. coli is not multiplying in the drinking water distribution network. The scarce distribution of individual cells observed in the present study might support the general assumption that the bacteria are most likely not multiplying since no microcolonies were detected.…”

“…Biofilms in distribution systems may provide a favorable condition for some bacteria, such as opportunistic pathogens (e.g., Legionella spp., Pseudomonas aeruginosa, and Mycobacterium avium), to colonize it and may harbor pathogens, such as Salmonella enterica serovar Typhimurium, which have entered the distribution system (1,5,18,28). It has been shown in lab-scale experiments that E. coli can survive in biofilters (24) and even multiply in the biofilm (17,34,40), which again raises the question of its suitability as a drinking water quality indicator. The aim of the present study was to investigate the presence of E. coli in the biofilm of actual water distribution networks in Europe by using a direct detection method.…”

Persistence of Helicobacter pylori in Heterotrophic Drinking-Water Biofilms

“…In fact, other microorganisms, such as Legionella pneumophila, Campylobacter jejuni, and even Escherichia coli, have been shown to prefer the microaerophilic environments demonstrated to be present in biofilm stacks (21,22,31) due to their intricate structure (Fig. 2d).…”

“…Even though the systems used to generate biofilms were different (the system used in the previous study was operated in batch mode, with planktonic H. pylori cells during the entire experiment), the altered behavior might be more logically explained by the heterotrophic nature of the biofilms in the current work. Heterotrophic DWDS-associated biofilms are known to create a safe haven that protects microorganisms from external stresses, such as temperature, shear, oxygen, and nutrient concentration stresses (24,31,41), and might help retain H. pylori attached to surfaces.…”