Research On Lead-resistant Endophytic Bacterium Isolated From Phytolacca Acinosa And Solanum Nigrum For Promotion Of Pb Accumulation In Pinus Massoniana.

Mineral resources are rich in China.Mineral development and industrial development exist side by side and play a part together,both which have played an important role in promoting the development of our national economy.With rapid development of high-energy-consuming industries,a large number of environmental problems have arisen due to the large-scale development of minerals.Especially the problem of heavy metal pollution,it has a serious impact on human life.It is an urgent problem for human beings to handle with heavy metal pollution.Pinus massoniana is a native tree species that grows in subtropical regions of China,which is the most widely distributed among Chinese pines.It has many special characters of strong adaptability to soil and climatic environment,easy breeding,rapid growth and wide application,therefore it is used as a pioneer species for greening barren hills.In this study,a variety of lead-tolerant endophytic bacteria are isolated from the lead-enriched plants Solanum nigrum and Phytolacca acinosa plants.Physiological and biochemical identification,molecular identification and quantitative analysis of lead tolerance are carried out in these strains.The extracted various strains are inoculated in the lead-treated Pinus massoniana,and physiological and biochemical analysis of Pinusmassoniana before and after inoculation was carried out.The results showed that 105 endophytic bacteria are isolated from the leaves of Solanum nigrum by lead stress culture,and 24 endophytic bacteria are obtained from Phytolacca acinosa.Through Gram staining and observed by a microscope,the shape of endophytic bacteria is mostly rod-shaped.The results of Gram staining is positive in bacteria.Further experiments such as methyl red and VP are carried out,and the physiological and biochemical properties of the isolated endophytes are identified.Then test for enriching lead capacity is carried out in lead-tolerant endophytic bacteria.It is found that after culture of endophytes in lead-containing medium for 16 hours,the average enrichment of lead ions in some bacterial cells reaches 200.31 ug/ml.The endophytic bacterial DNA is further extracted and the DNA is subjected to PCR amplification.The sequence of strains is analyzed by BLAST.The NJ and Bayesian phylogenetic tree are constructed by MEGA(6.0)and Mrbayes software.The endophytic bacteria isolated from Solanum nigrum are initially identified as Bacillus,Acinetobacter,and Enterobacter sakazaki.The endophytic bacteria of Phytolacca acinosa are Acinetobacter and Bacillus.Physiological and biochemical analysis under lead stress culture in the masson pine inoculated with lead-resistant endophytes show that the chlorophyll content and SOD activity of the Pinus massoniana pine needles are higher than that of the uninfectedPinus massoniana.However,the content of soluble protein,Pro and MDA in the pine needles of the inoculated masson pine is lower than the pine needle of the uninfected Pinus massoniana.The content of lead in the roots,stems and pine needles of Pinus massoniana was determined by the method of nitration-ICP: The average lead content of roots,stems and pine needles of Pinus massoniana was 472.210 times,279.841 times and48.214 times of the average lead content of the control group.This study provides a set of effective technical methods for bioremediation technologies in lead-contaminated areas.It also provides new ideas for the production of sustainable and stable economic benefits on the marginal soil contaminated by lead.