Effects And Mechanisms Of Polyamine-producing Bacteria On Reducing Cd And Pb Accumulation In Water Spinach

In recent years,due to sewage irrigation,decomposition of agricultural plastic products and unreasonable use of chemical fertilizers and pesticides,vegetable fields contaminated with Cd has seriously exceeded the standard,posing a serious threat to people's health.Based on the necessity of using heavy metal contaminated vegetable soil,microbial immobilization can achieve the purpose of passivating soil metals and preventing vegetables from accumulating heavy metals.Polyamine-producing bacteria,as a functional strain that can promote growth,immobilize heavy metals and erase polyamines,provides technical means for the safe utilization of heavy metal polluted vegetable fields,alleviating the toxic effects of heavy metals on crops,and providing scientific data and theoretical basis for maintaining crop yield and improving food quality and safety.In this study,109 polyamine-producing strains with strong tolerance to Cd and Pb were isolated from the abandoned vegetable field soil around Yuguang Gold and Lead Factory in Jiyuan City,Henan Province by using polyamine-production screening medium and LB solid plate with high concentration of Cd(100 mg/L)and Pb(1500 mg/L).Thirteen polyamine-producing bacteria with high removal effeciency of Cd and Pb were isolated by liquid shaking flask test using high-pressure liquid chromatography and inductively coupled plasma spectrometer.The adsorption efficiency of Cd and Pb in the solution was 27%-90%and 28%-94%,respectively.Meanwhile,the p H of fermentation broth was increased.Finally,the indole acetic acid and iron carrier capacity of the strains were analyzed,and two polyamine-producing strains were screened out by hydroponic experiments with water spinach,which prevented water spinach from accumulating Cd and Pb,namely Enterobacteria bugandensis(XY1)and Serratia marcescens(X43).XY1produced spermine(24.2 mg/L)and spermidine(19.5 mg/L),and X43 produced putrescine(53.5 mg/L).The indole acetic acid yields of the two strains were 78.65 mg/L and 55.26mg/L,respectively.Hydroponic experiment showed that XY1 and X43 increased biomass of water spinach by 1.1?2.35 times and 4%?98%,respectively,compared with the control without inoculation with strains.At the same time,XY1 and X43 significantly reduced the contents of Cd and Pb in tissues,the contents of Cd was reduced by 49%?59%and29%?38%,and the contents of Pb was reduced by 63%?72%and 30%?47%,indicating that the two strains had good growth promotion effect and the ability to reduce the content of heavy metals in edible parts of vegetables.The biological characteristics of XY1 and X43 and the mechanisms of heavy metal immobilization showed that XY1 and X43 were resistant to Cd(600 mg/L and 500 mg/L)and Pb(2300 mg/L and 1700 mg/L).The adsorption test of exogenous polyamines at 0.1m M illuminated that the adsorption rates of exogenous putrescine on Cd and Pb were 1.35%and 3.7%,respectively.The removal rates were 4.25%and 84.5%,respectively,under the combined treatment of exogenous spermine and spermidine,indicating that polyamines chelated heavy metals.The adsorption mechanism of polyamine-producing bacteria was characterized by scanning electron microscopy,infrared spectroscopy and X-ray diffraction.The results showed that bacteria adsorbed Cd²⁺and Pb²⁺by cell surface,mainly through the-OH,-NH,C-N and other functional groups on the cell wall,and generated Cd(OH)₂,Cd CO₃ and Pb O_x precipitates to passivate the heavy metals.Based on the efficiency in solution,the effect of functional strains to fix Cd and Pb in soil was studied by pure soil culture experiment(Cd²⁺concentration was 1 mg/kg,Pb²⁺concentration was 5 mg/kg).The results showed that p H was increased in the presence of XY1 and X43 by 0.47 units(7.02)and 0.39 units(6.94),respectively,which were beneficial to the fixation of heavy metal ions in the soil.Meanwhile,the content of extractable Cd and Pb(18%?39%,31%?37%)was droped,and differed from that,the content of residual increased by40%?54%and 33%?57%,respectively.Based on the conventional planting mode of vegetables,pot experiments were conducted to study the effects and mechanisms of polyamine-producing bacteria on the biomass,quality and metals content of water spinach under different concentrations of Cd and Pb pollution.The results signified that XY1 and X43 increased biomass of edible tissue(40.6%?80%)and contents of soluble protein(3%?24%),and helped to lowered the contents of Cd(23%?60%)and Pb(15%?46%)in vegetables and DTPA-Cd(23%?48%)and DTPA-Pb(17%?35%)in soil under heavy metal stress,compared with the control.The antioxidant system of water spinach also changed.The content of non-enzymatic antioxidant vitamin C was increased by 26%?39%,and the catalase activity in enzymatic antioxidant substances was up to 1.7 times higher than that of the control.While the activity of superoxide dismutase had a slightly increase(3%?7%).High-throughput sequencing results showed that the bacterial community diversity decreased inoculation with XY1 in the rhizosphere soil contaminated by low concentration of metals.Compared with the control group,the relative abundance of Proteobacteria,Firmicutes,Gemmatimonadetes and Bacteroidetes in the treatment group XY1 was significantly increased,and the relative abundance of Actinobacteria,Acidobacteria and Chloroflexi was significantly decreased.