| Extracellular polymers substances(EPS)are the important components of membrane biofoulants,which is one of the critical issues in the successful application of membrane bioreactor(MBR)for water treatment.Acyl-homoserine lactone(AHL)mediated quorum sensing(QS)system has been identified as one of the most significant biological factors responsible for the process of biofouling formation.Thus,it is important to reveal the influences of AHL-QS on the characteristics of EPS and its membrane fouling behavior.In this work,5 dominant strains with AHL-QS were isolated from membrane fouling layer in a MBR.Among them,Aeromonas veronii was selected and taken as a model.Further AHL-lacking mutant stains were constructed and used to test how AHL mediated quorum sensing contributing to the characteristics of EPS and its membrane fouling behavior.The main research results were shown as follows:(1)The phylogenetically identified result of 16S rDNA sequencing suggested that 5 dominant strains with AHL-QS were Chromobacterium violaceum,Aeromonas veronii,Aeromonas caviae,Aeromonas hydrophila and Aeromonas aquariorum,respectively.The testing results of the report bacterium of Chromobacterium violaceum 026,thin layer chromatography(TLC)and high performance liquid chromatography-mass spectrometry(HPLC-MS/MS)indicated that 1 strain produced C6-HSL and the other 4 strains produced C4-HSL.(2)Upstream-and downstream-flanking sequences of LuxI and LuxR were amplified respectively.Then a chloramphenicol resistance cassette was linked between the two sequences by fusing PCR to get a recombinant suicide plasmid pCVD442.The transconjugants in which the Lux gene was replaced by a Cmr cassette were selected and then the Lux mutant strains,N8/?LuxI and N8/?LuxR,were generated.(3)There were obvious differences between the LuxIR mutant strains and the wild strain in the characteristics of EPS.Compared with the wild strain,the deletion of LuxI made the EPS produced by A.veronii were different in the compositions(protein and polysaccharide),but the LuxR mutant strain was produced fewer EPS.X-ray photoelectron spectroscopy(XPS)results showed that protein and polysaccharide were the main components of EPS and there were some differences in the compositions of functional groups among the three strains.Three-dimension excitation emission matrix fluorescence spectroscopy(EEM)results indicated that the main composition of protein in EPS was tryptophan-like protein.And loosely bound EPS(LB-EPS)had a little humic-like material,but tightly bound EPS(TB-EPS)had not.The tryptophan-like protein in the LB-EPS produced by the mutant strains was more complex than that of the wild strain,but the humic-like material had a simpler structure.In the TB-EPS produced by the mutant strains,both tryptophan-like protein and aromatic protein were simpler.Gel filtration chromatography(GFC)results showed that the EPS of the wild strain was smaller,but its molecular weigth distribution was wider,which indicate that the EPS produced by A.veronii has higher molecular weight and simpler structure after the AHL-QS system was interrupted.Zeta potential results suggested that EPS has ionizable functional groups rendering it negatively charged.Meanwhile,LB-EPS was more inclined to coagulate than TB-EPS.To compare with LuxR,LuxI had a greater influence in the stability of EPS produced by A.veronii.(4)The mutant strains had a different performance of the filtration with the wild strain.The results of filtration studies exhibited EPS had a greater fouling potential as its concentrations increasing.Atomic force microscopy(AFM)results showed the adhesion of EPS made the surfaces of PVDF membrane smoother and the value of Rq and Ra become smaller.There was less LB-EPS adsorb to the surfaces of PVDF membrane than TB-EPS,so the surfaces of PVDF membrane adsorbed TB-EPS was smoother.Infrared spectroscopy(IR)results suggested that the functional groups of adsorbed foulants after filtration were amino,hydroxyl and carboxyl group and so on.And that protein and polysaccharide are the main components of EPS.Confocal laser scanning microscope(CLSM)images showed there were a large amount of proteins and polysaccharides adsorbed on the membrane surface after filtration,and they were varied in distribution.The fouling layer of LB-EPS produced by the wild strain was thinnest and densest.The fouling layer of N8/? LuxI was thick but loose,and that of N8/? LuxR much more denser.For the fouling layer of TB-EPS,the wild strain's was thickest with most ?-polysaccharides and lest proteins,but that of N8/? LuxR was thinnest with most proteins. |
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