2,4-D degradation in monoculture biofilm reactors

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Two strains of Alcaligenes eutrophus AEO106 were used to inoculate expanded bed biofilm reactors which were exposed to a range of concentrations and loading rates of 2,4-dichlorophenoxyacetic acid (2,4-D herbicide). Two bench-scale reactors were inoculated with bacterial strains containing different plasmids. Plasmid pRO101 expressed the 2,4-D degradative pathway in the presence of inducer (2,4-D or 3-chlorobenzoate), whereas plasmid pRO103 was engineered to allow constitutive expression of the pathway due to the deletion of a regulatory gene. Biofilm growth characteristics, loading rate and 2,4-D conversion efficiency relationships were compared in the two reactors. Actual loading rates tested ranged from 3.6 to 52.3 g 2,4-D per liter expanded bed volume per day (g/ld). The range of influent concentrations tested was between 166 and 728 mg/l (0.017 and 0.078%). Mineral nutrients were supplied in the feed solutions. At loadings up to 15 g/ld, reactors were aerated with filtered air. Pure oxygen was found to be necessary at higher loadings to sustain maximum bacterial activity. Removal of 2,4-D to below detectable concentrations (< 3 mg/l) was demonstrated at all but the highest loadings tested (at least up to 10 g/ld with air and > 25 g/ld with pure oxygen). However, minimum effluent chemical oxygen demand (COD) values were 25 mg/l even at lower loading conditions. Advantages noted with the AEO106 pRO103 (constitutive) biofilm included greater biomass development (up to 3 times higher volatile solids), 50–100% higher specific substrate conversion rates, quicker response to step increases in feed rate and concentration and more consistent effluent quality. Approximately one-third lower effluent COD concentrations, with one-half the standard deviation among individual analyses, were noted with pRO103 compared to pRO101. The genetically engineered biofilm reactor was shown to be a viable experimental treatment technology for one xenobiotic compound in these studies. However, pure culture conditions were not maintained throughout the experiments because of the low-level persistence of a sporadic contaminant bacterial species (identified as Xanthomonas maltophilia), which apparently incorporated plasmids from Alcaligenes eutrophus AEO106.