| It has been aimed to investigate growth characteristics of a genetically engineered microorganism (GEM) Escherichia coli JM 109 (pGEX-AZR) and its ability to decolorize azo dyes. For the ecological safety, E. coli JM109 (pGEX-AZR) was fixed in the MBR to prevent the release of the GEM into the environment. Meanwhile, bio-augmentation of Escherichia coli JM109 (pGEX-AZR) to azo dyes decolorization was studied, and the kinetics and structure changes of microbial commnunity in augmented system were revealed by modern molecular techniques.The optimal growth conditions of E. coli JM109 (pGEX-AZR) are as follows: glucose 5 g·L-1, NH4C10.3 g·L-1, inorganic salt medium (KH2PO4 2.0 g·L-1, Na2HPO4 1.3 g·L-1, MgSO4 2.0 g·L-1, CaCl2 0.0364 g·L-1, FeCl3 0.00025 g·L-1), pH=7.5, inoculation amount 10 mL·L-1, temperature 35℃. When the optical density at 660 nm (OD660) of E. coli JM109 (pGEX-AZR) was up to 1.7～1.8, IPTG was added the culture concentration of 1 mmol L-1, and induction was conducted for another 8 h. Attempt to use lactose instead of IPTG inducing expression of azoductase by the recombinant strain E. coli JM109. Lactose was added so as to the final concentration to 40 mmol·L-1, recombinant protein expression concentration and azoreductase activity are 93.6% and 85.7% as much as that given by the strain induced by IPTG.The co-metabolism kinetics of acid red GR by the E. coli JM109 (pGEX-AZR) was studied using glucose as primary substrate. The process of glucose utilization by E. coli JM109 (pGEX-AZR) can be described by the Monod equation, where the maximum specific biodegradation rate coefficientμmax g is 0.07657 g·g-1·h-1, half-velocity constant Kg is 0.3324 g·L-1. Acid red GR exerted non-competitive inhibition on the glucose utilization, an inhibition coefficient Kig of 0.213 g·L-1. The kinetics of the acid red GR decolorization by the E. coli JM109 (pGEX-AZR) agrees with Andrews model, the kinetic parameters, rdye,max, Ks and Ki, have been found to be 42.45 mg L-1 h-1, 584.93 mg L1 and 556.89 mg L-1 respectively. The biomass yield coefficient of E. coli JM109 (pGEX-AZR) is 0.1 g·g-1, the biomass transformation capacity is 121.2mg·g-1, the endogenous decay constant is 0.0378 d-1.The decolorization of several kinds of azo dyes can be executed by the E. coli JM 109 (pGEX-AZR) . The optimal conditions of decolorization are anaerobic conditions and pH 7-8. The decolorization rate increases with the increase of temperature between 25～40℃. The process of decolorization of acid red GR is non-inhibit in the presence ot NaCl (1～5%), NaSO4 (1～5%). In contrast, the addition of NaNO3 (1～5%) had an adverse effect on the decolorization rate, this may be the competition in reduction reaction between the nitrate and the azo bonds, and the nitrate is obviously a better electron acceptor than the azo bond.Similar to acid red GR decolorization by suspended E. coli JM109 (pGEX-AZR), acid red GR decolorization by the immobilized cells on macro-porous foam and the sodium-alginate immobilized cells can be also described by the same Andrews model. The kinetic parameters rdey,max and Ks,. by the sodium-alginate immobilized cells decreased and Ki increased compared with suspended cell, which might be due to the low biological activity and slow mass transfer. The kinetic parameters rdye,max, Ks. and Ki by the immobilized cells on macro-porous foam carriers increased compared with suspended cell, which might be due to the improvement of stability.E. coli JM109 (pGEX-AZR) was used for the decolorization of acid red GR in the submerged anaerobic membrane bioreactor. The results from the study show that, the bioreactor with E. coli JM109 (pGEX-AZR) has a high capacity for decolorization, decolorization rate of the supernatant effluent is of 88%～93%, decolorization rate and COD removal rates of the membrane effluent are about 96% and 56%～72%, respectively. It is obvious that the membrane separation process can enhance the wastewater treatment effect. The concentration ofE. coli JM109 (pGEX-AZR) had grown from 1.97 g·L-1 to 2.12 g·L-1 at the beginning, and finally had kept at the concentration between 1.60 g·L-1 and 1.80 g·L-1.The E. coli JM109 (pGEX-AZR) was tested in anaerobic sequencing batch reactors (AnSBR) in order to enhance the acid red GR decolorization. The continuous operations of the four bioreactors with different E. coli JM109 (pGEX-AZR) immobilization supports showed that the E. coli JM109 (pGEX-AZR) showed bio-augmentation in AnSBRs with suspended and immobilized on macro-porous foam carriers. The tolerance to acid red GR concentration shock and the decolorization rate in these two bio-augmented AnSBRs were higher than those of the other two systems, control system and bio-augmented AnSBRs system with the sodium-alginate immobilized cells. Changes in microbial community were detected with ribosomal intergenic spacer analysis (RISA) and amplified ribosomal DNA restriction analysis (ARDRA), which revealed that the introduced E. coli JM109 (pGEX-AZR) is persistent in the augmented systems and maintained higher metabolic activity.
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