An algD-bioluminescent reporter plasmid to monitor alginate production in biofilms.

A broad-host range algD-lux bioluminescent reporter plasmid was developed to examine the role of exopolymer production in biofilm function. The algD-lux reporter plasmid will allow rapid on-line in situ detection of environmental factors that induce alginate biosynthesis. The algD promoter was stimulated by factors previously reported to induce alginate production, including ethanol and NaCl, and differences were observed with different nitrogen sources. With growth on minimal media with either glucose or succinate as a carbon source, succinate had a greater inductive effect on the algD promoter. An increase in light output of 1.3-fold and 1.7-fold was seen with cultures amended with 50 and 150 mM NaCI, respectively, compared to cultures with succinate alone. NaCl induction of the algD promoter was confirmed by algD RNA slot blots. Light output increased 2.0-fold and 1.7-fold with 0.25% and 0.5% ethanol, respectively, compared with controls grown with succinate only. While the rate of algD promoter response was initially similar when either NH4 or NO3 was used as a nitrogen source, NH4-grown cultures maintained a higher light output during late log phase compared to NO3-grown cultures.