[Study on gfp labeling of a 2,4-D degrading strain and its detection in a wastewater biotreatment system]

A 2,4-dichlorophenoxyacetic acid (2,4-D) degrading special bacteria Achromobacter sp. was chromosomally labeled with a green fluorescent protein gene (gfp) using a mini-Tn7 transposon delivery system. The growth status, fluorescence expression and degradation ability of the strain before and after labeling were compared. Methods to quantify the strain in different biotreatment systems (activated sludge or granular sludge system) after inoculation were also investigated. Results showed that the labeled Achromobacter sp. and its control strain demonstrated a similar growth pattern and 2,4-D degradation ability: both of them could completely remove 2, 4-D of about 100 mg/L within 103-112 h. The labeled strain could express fluorescence stably during the course of growth and degradation with fluorescence intensity/D600 stabilized at about 4500. For an activated sludge system bioaugmented with this labeled strain, its abundance could determined through direct measuring fluorescence emitted by the sludge mixture, for it was linearly associated to the percentage of the strain in the range of 0-75% (R2 = 0.9952). For a granular sludge system bioaugmented with this strain, fluorescence of the sludge mixture could be measured after homogenous pretreatment, and the percentage of the strain in the range of 0-42% was also linearly related to the fluorescence intensity emitted by the sludge mixture (R2 = 0.9801). Overall, this gfp labeling method based on Tn7 delivery system can be used to monitor specific bacteria in a biotreatment system.