Application Of Metabolic Uncoupler TCS Control MBR Membrane Biofouling And Mechanism Analysis

Membrane fouling is recognized to inhibit the development of membrane filtration technology.Biological control of biofouling has recently been proposed as a novel anti-biofouling method,which aims to interfere with the biological attachment and biofilm formation process on filtration membrane.The application of metabolic uncoupler can effectively inhibit biofouling formation in MBR system.However,the applied concentration of uncoupler is relatively high which may affect the normal growth of sludge and therefore,it would be difficult to analyze the biofouling inhibitory mechanism of uncoupler.Based on biological control theory of biofouling,this study investigated the inhibitory effect of uncoupler on single bacteria biofilm and mixed bacteria biofilm in MBR to understand the biofouling control mechanism of uncoupler in MBR system.The typical metabolic uncoupler,3,3?,4?,5-Tetrachlorosalicyanilide(TCS),was chose to study the biofilm formation mechanisms of single bacteria and mixed bacteria in MBR.The gram-negative bacteria(Pseudomonas aeruginosa PAO1)and the gram-positive bacteria(Bacillus subtilis)were chose as typical bacteria to study single bacteria biofilm formation.Apply the real-time quantitative PCR(RT-q PCR)technology to analyze the effect of TCS on quorum sensing and biofilm related genes of P.aeruginosa PAO1 and B.subtilis.Three lab-scale parallel membrane bioreactor(MBR)systems were continuously operated for 140 days to investigate the effect of TCS on biofouling formation and the mechanism of biofouling control.The inhibitory effect and mechanism of TCS at different concentrations on biofilm formation of P.aeruginosa PAO1 was investigated.It was observed that when the added concentration of TCS was over 10 μg/L,the biofilm formation of P.aeruginosa PAO1 on filtration membrane was significantly inhibited.In the 2 h biological attachment test,when the concentrations of TCS in medium were 1 00 μg/L and 500 μg/L,the number of attached cells were reduced 27.78%和 19.19%.In the bacterial motility test,TCS concentrations at 100 μg/L and 500 μg/L both inhibited the swimming motility of P.aeruginosa PAO1 while 100 μg/L was found more effective.In the 30 h biofilm test,the biofilm in 100 μg/L and 500 μg/L TCS samples were reduced 28.02% and 50.15%.It was suggested that TCS at 500 μg/L greater suppressed bacterial biofilm development.In the analysis of RT-q PCR,it was observed that TCS at 500 μg/L repressed the expressions of las I and las R quorum sensing(QS)genes which involve in the regulation of biofilm formation but not 100 μg/L.This found suggested that 500 μg/L TCS could effectively control biofilm formation of P.aeruginosa PAO1 via regulating quorum sensing system.The inhibitory effect and mechanism of TCS on the biofilm formation of B.subtilis was investigated.It was found that the biofilm at the bottom of 12-well plate,and filtration membrane were reduced 53.08 % and 44.67% by 100 μg/L TCS without affecting the growth of B.subtilis.The analysis of bacterial motility,flocculability and surface thermodynamics found poorer bacterial motility,smaller biofloc,more negative surface,higher hydrophilicity and higher primary energy barrier were observed in the test samples after treated with 100 μg/L TCS,indicating that B.subtilis need to consume more energy to aggregate and develop biofilm.The effects of TCS on B.subtilis QS system and biofilm related gens were evaluated.It was found that the gene sin R which was modulated by CSF QS system,was upregulated by 100 μg/L TCS.The up-regulation of sin R repressed the expressions of extracellular polymeric substances(EPS)related genes eps E,tas A and yqx M.The biofilm inhibitory mechanism of TCS on B.subtilis was because the expressions of biofilm and EPS related genes were regulated.During 140-day continuous operation,we investigated the effect of 100 μg/L TCS on biological attachment and biofouling formation in MBR system.Compared to the control system MBR 1,the average fouling cycle of MBR 2 system with 100 μg/L TCS increased from 10.46 d to 55.5 d,indicating 100 μg/L TCS addition could effectively mitigate biofouling formation in MBR.However,the beneficial effect of TCS on biofouling alleviation was vanished when no longer dosing TCS.During 140-day operation,it was notable that the secretion of interspecies QS signal,autoinducer-2(AI-2),and intraspecific QS signals,N-acylhomoserine lactones(AHL),were all substantially inhibited by 100 μg/L TCS.Since bacterial EPS secretion has a close relationship with QS system,the concentrations of extracellular proteins and polysaccharides were reduced,suggesting the EPS secretion and QS system in MBR should be regulated by TCS.In addition,the measurements of observed sludge growth yield in system and COD and NH4-N concentrations in effluent indicated the addition of 100 μg/L TCS had no significant effect on sludge growth and effluent quality.This study indicated that the utilization of metabolic uncoupler could effectively inhibit biofouling in MBR system via controlling QS signals and EPS secretion without affecting sludge growth and effluent quality.