Effect Of Quorum Sensing On The Cell Surface Properties,Phenanthrene Removal And Copper Tolerance In Novosphingobium Pentaromativorans US6-16-1

Quorum sensing?QS?is an important mechanism to regulate the physiological activities of bacteria,such as biofilm formation,extracellular polymeric substance?EPS?production.These physiological activities will further affect the cell surface characteristics and the interaction between bacteria and environmental pollutants,so QS is likely to participate in bioremediation in the way of affecting the cell surface properties.At present,there are few studies on the influential mechanism of QS on the removal of polycyclic aromatic hydrocarbons?PAHs?and the tolerance of Cu²⁺.Novosphingobium pentarotavorans US6-1 is a bacteria that can not only degrade a variety of PAHs,but also produce acyl homoserine lactones?AHLs?for QS.Therefore,this paper uses US6-1 to explore the regulatory characteristics of QS on cell surface properties and whether QS deletion will further affect the ability of bacteria to remove phenanthrene?phe?and tolerate Cu²⁺.Firstly,?nov I and?novR from US6-1,which are mutants with nov I and novR deletion,were confirmed.And their complementary strains,?nov I?p CM62-nov I?and?novR?p CM62-novR?,were constructed and confirmed,respectively.The cell surface properties of wild type strains?WT?US6-1?,mutants??nov I and?novR?,and complementary strains??nov I?p CM62-nov I?and?novR?p CM62-novR??were measured in P5Y3 liquid,in MM2 liquid with phe as the main carbon source,and in P5Y3 liquid with different concentration of Cu²⁺,respectively.The effects of QS on phe removal efficiency?PRE?were explored under different culture conditions in US6-1.And the effect of QS deletion on cell surface properties and PRE was further confired by q RT-PCR analysis.The main conclusions are as follows:?1?The results of AHLs assay indicate that mutants and the complementary strains were eligible for further experiments.In P5Y3 liquid,the growth trends were not significantly affected in mutants.Cell surface hydrophobicity?CSH?of mutants increased,EPS production of mutants decreased,the extension of C-O-C functional groups on the cell membrane surface and the cell membrane permeability changed in mutants.According to q RT-PCR analysis,the relative expression of spt in?nov I was up-regulated,which is the key gene response to synthesize glycosphingolipid in US6-1.And epsd in?novR,the gene responsible for synthesizing glycosyltransferase Eps D,was down-regulated.The results support the detection of CSH and EPS production in mutants.?2?The growth trends of WT?US6-1 and mutants were basically same in MM2liquid with phe as the main carbon source.The CSH of mutants was about three times of that of WT?US6-1 and complementary strains,and EPS production of them was similar.Phe makes the vibration of 600 cm^-1C-O-C functional groups on the membrane surface of mutants and the membrane permeability higher than that of WT?US6-1.The results show that the cell surface properties of mutants more affected by phe.For phe removal assay in MM2 liquid and on MM2 agar plates,PRE of mutants was higher than that of WT?US6-1,and the relative expression of most enzyme genes in PAHs degradation pathway was up-regulated in mutants,which was consistent with the results of PRE.The results show that PRE was negatively regulated by QS in US6-1.?3?US6-1 can tolerate 1 mmol/L Cu²⁺.Cu²⁺had no significant effect on CSH and functional groups of US6-1,however which increased EPS production and membrane permeability of US6-1,the growth and the surface properties of?nov I were more affected by Cu²⁺.Cu²⁺decreased the membrane electronegativity of WT?US6-1 and?nov I?p CM62-nov I?,but increased the membrane electronegativity of?nov I.The results show that QS can protect the surface properties of US6-1.On the third day of cultivation of copper-phe pollution,PRE of US6-1 was higher with the increase of Cu²⁺,and PRE of?nov I was higher than that of WT?US6-1 and?nov I?p CM62-nov I?with Cu²⁺.On the fifth day of cultivation of copper-phe pollution,PRE of US6-1 cultured with 0.5 mmol/L Cu²⁺was highest.And PRE of?nov I was significantly lower than that of WT?US6-1 and?nov I?p CM62-nov I?with 1 mmol/L Cu²⁺.The results show that 0.5 mmol/L Cu²⁺can promote PRE of US6-1,PRE of?nov I without QS more affected by Cu²⁺,but the specific mechanism of the effect needs to be further explored.