Characterization Of Cadmium Resistance Mechanism And Plant Growth Promoting Potential Of Burkholderia Contaminans Strain ZCC

Cadmium is a heavy metal that is biologically not necessary and very toxic.The accumulation of cadmium in the soil will cause damage to organisms,affect the growth and development of organisms,and can also accumulate in the human body through the food chain,causing serious harm to human health.Anti-cadmium microorganisms have great application value in the bioremediation of cadmium-contaminated soil,and revealing the mechanism of microbial cadmium resistance is the basis and premise of applying microbial remediation technology.In this study,the interaction of extracellular polymer（EPS）with cadmium,whole genome analysis and plant experiments revealed the cadmium resistance of Burkholderia contaminans ZCC isolated from the compound contaminated soil around the Zijinshan gold-copper mine in Fujian.The effects on plant growth under cadmium stress by strain ZCC was also estimated In this study.The main conclusions are as follows:Physiological characterization of heavy metal resistant strain ZCC found that the strain ZCC is a Gram-negative bacterium,facultative anaerobic and mobile.Strain ZCC displayed a temperature growth range of 4-37℃,and the optimal growth temperature was 28℃;growth was possible in the pH range of 4.13-9.39,the optimum growth pH was 4.13;the NaCl tolerance range was 0-5%,and the optimum NaCl growth concentration was 1%.Strain ZCC was shown to have a high tolerance to a variety of heavy metals.The minimal inhibitory concentration to cadmium was 7 m M,and strain ZCC had a high capability to remove Cd（Ⅱ）,with a removal rate of 48.23%.Strain ZCC was also highly tolerant to various antibiotics such as streptomycin,tetracycline,and gentamicin,and their tolerance levels were 400,400,and 200μg·m L^-1,respectively.Strain ZCC was able to use glucose,mannose,glycerol,sucrose and other carbon sources and ammonium sulfate,ammonium chloride,L-leucine and other nitrogen sources.Using 16S rRNA phylogenetic tree analysis,the strain ZCC was identified as Burkholderia contaminans.By studying the interaction between EPS secreted by strain ZCC and Cd（Ⅱ）,it was found that when the concentration of Cd（Ⅱ）is less than 0.1m M,Cd（Ⅱ）was able to stimulate ZCC to secrete more EPS,and EPS was able to lower cadmium toxicity compared to the control.The inhibitory of strain ZCC biomass and cell activity.When the concentration of Cd（Ⅱ）was greater than 0.1 m M,the amount of EPS secreted by strain ZCC dropped rapidly.When using three-dimensional fluorescence excitation emission spectroscopy,Fourier infrared spectroscopy,X-ray photoelectron spectroscopy to explore the mechanism of EPS and Cd（Ⅱ）complex,the results showed that EPS and Cd（Ⅱ）complex carboxyl,C-OH,P=O,C-O and phosphorus-containing groups played a key role,mainly due to the complexation of O atoms with Cd（Ⅱ）,in formation of C-O-Cd⁺.The above results indicated that Cd（Ⅱ）is physically adsorbed by EPS and undergoes a complexation reaction,which prevents Cd（Ⅱ）from entering the interior of the cell,so as to lower the toxic effect of Cd（Ⅱ）on strain ZCC.Genomics analysis of strain ZCC revealed that the strain ZCC has a total genome size of 9,002,241 bp,a G+C content of 66.74%,and containing 8,897 coding genes,There are 3 prophages and 3 CRISPRs on the genome of strain ZCC.Annotation of the genome of strain ZCC identified 5,185,6,171 and 4,704 genes in GO,GOG and KEGG databases,respectively,accounting for 62.4%、74.2%and 56.6%of the total number of genes.These genes are mainly related to such as carbohydrate metabolism,amino acid metabolism,signal transduction and energy generation and conversion.Annotation of strain ZCC genome identified a total of 5550 genes in RAST,among them,including 2 P_1B-type-ATPase,2czcCBA operons and 1 czcD related to cadmium resistance.Annotation of the genome of strain ZCC identified 147 genes encoding carbohydrate active enzymes in the carbohydrate active enzyme（CAZy）database,of which the genes related to glycoside hydrolase and glycosyltransferase accounted for the vast majority of the total number of genes In part,these two enzymes were related to the secretion of EPS by strain ZCC.These genotypic characteristics were consistent with the phenotypic characteristics of ZCC.By studying the effect of strain ZCC on plant growth under cadmium stress,it was discovered that strain ZCC has the ability to secrete ACC deaminase,indole-3-acetic acid（IAA）,siderophores,and dissolve phosphorus,and it is preliminarily judged that strain ZCC has plant growth promotion potential.Plant experiments showed that compared to the control group,the stress group Cd（Ⅱ）treatment inhibited the growth of soybean roots,reducing the accumulation of soybean dry weight,increasing the content of both cadmium in the aerial and aerial parts of the soybean and the content of malondialdehyde（MDA）in soybean roots.Compared to the stress group,the inoculation strain ZCC in the remission group could significantly increase the root length,dry weight and N content of soybean,as well as significantly reduce the content of both cadmium in the aerial and aerial parts of the soybean and the content of malondialdehyde in soybean roots.The above results indicate that the strain ZCC was able to relieve the toxicity of cadmium to soybeans and promote the growth of soybeans.In conclusion,under conditions of cadmium stress,strain ZCC may activate its own efflux system to expel intracellular Cd（Ⅱ）,and also secrete EPS to complex extracellular Cd（Ⅱ）,to alleviate the poisoning by Cd（Ⅱ）.These characteristics of strain ZCC can not only protect themselves from Cd（Ⅱ）poisoning,but also complex Cd（Ⅱ）by secreting EPS to relieve Cd（Ⅱ）stress to soybean.In addition,the strain ZCC was shown to also secrete IAA,iron siderophores,ACC deaminase and other substances to promote plant growth and make the plant more resistant to cadmium.