The Diversity Analysis Of Legume Green Manure Symbiotic Rhizobia And Mining Of Dominant Species Resources In China

Green manure is a cultivated or wild green plant that can be directly or after being piled applied to the soil as an organic fertilizer.Among them,leguminous green manure can symbiosis with rhizobium to form nodules,so it can perform bio-nitrogen fixation.Common vetch,hairy vetch,Chinese milk vetch,white clover et al.were common leguminous green manures.It was also fitted with the national agricultural policy of "reducing fertilizer and reducing pesticide",to give full play to the biological nitrogen fixation effect of legumes and rhizobia,and effectively reduce the use of chemical fertilizers.But the legume must select the compatible rhizobia for effectively establishing the symbiotic interaction.However,there is no systematic research on the resources and genetic diversity of symbiotic rhizobia of leguminous green manure in China.Based upon the above,this study was the first study to systematically collect resources,analyze diversities and the distribution of biogeography of rhizobia nodulated with common vetch,hairy vetch and Chinese milk vetch in main leguminous green manure cultivate areas of China.Finally,explore the resources of dominant species and lay the foundation for the industrial production of fungicides.Symbiotic rhizobial resources of common vetch and hairy vetch were systematically collected at 7 sampling sites in the three provinces of northwestern China,including Gansu,Qinghai,and Shanxi provinces,and rhizobium resources of Chinese milk vetch at 13 samples sites in the four provinces of southwestern China,including Sichuan,Hubei,Hunan,and Guangxi Provinces in this study and analysis the diversity of the collected resources.The first work was to collect the soils and capture the symbiotic rhizobia using the related leguminous green manures.A total of 202 common vetch rhizobia,253 hairy vetch rhizobia and 451 Chinese milk vetch rhizobia were obtained after isolation and purification.The molecular identification of all the isolates was carried out based on the phylogenetic analysis of IGS-PCR RFLP,16 S rRNA gene,housekeeping genes(atpD,recA and glnII)and nodulation gene(nodC).The results showed that: 1)Rhizobium laguerreae,R.sophorae and three novel species(Rhizobium novel genospecies I-III)were the common rhizobia species that could nodulate with both of common vetch and hairy vetch,additionally,hairy vetch also could establish the effective symbiotic relationship with an another novel species(Rhizobium novel genospecies IV).However,all the test strains belong to the three known species:Mesorhizobium huakuii,M.jarvisii and M.qingshengii that could nodulate with Chinese milk vetch;2)The soil were weak alkaline(pH=7.1-7.7)in common vetch and hairy vetch sampling sites,while the pH in Chinese milk vetch sampling sites ranged from 4.9 to 8.0;3)Common vetch and hairy vetch are in the same leguminous cross inoculation group,it means that the rhizobia,which could nodulated with them,also could nodulat with legumes in this group such as plants of Vicia,Lens,Pisum and Phaseolus genera;while the rhizobia nodulated with Chinese milk vetch has strict host specificity and its symbiotic genes are highly conservative;4)Rhizobium novel genospecies III,R.sophorae and M.jarvisii were the dominant rhizobia species nodulating with common vetch,hairy vetch and Chinese milk vetch respectively.The Canonical Correlation Analysis for the physicochemical characteristics of soils and the genetic diversities of the rhizobia were carried out in the end of this study.The results showed that: the biogeographic distribution of the rhizobia was apparently affected by the pH,salinity,available phosphorus,for example,the distribution of Rhizobium novel genospecies III,which was the dominant population of common vetch,was positively correlated with available phosphorus and pH,but negatively correlated with salt content;the distribution of Rhizobium sophorae,which was the dominant population of hairy vetch,was positively correlated with available phosphorus and salt content,but negatively correlated with pH;the distribution of the dominant genotype T1 of Chinese milk vetch rhizobia was positively correlated with salt content and pH while negatively correlated with available phosphorus.Finally,the dominant species resources were excavated according to the distribution characteristics of the rhizobia population.The results showed that for the common vetch rhizobia,the strains of IGS type T3,T5 and T9 were the dominant rhizobial resources in Gansu,Qinghai and Shanxi provinces respectively;for the hairy vetch rhizobia,the strains of IGS type T21 were the dominant rhizobial resources of Shanxi Province,and the strains of IGS type T11 is the dominant rhizobial resources of Qinghai Province and Gansu Province;for the Chinese milk vetch rhizobia,the strains of IGS type T1 was the dominant rhizobial resources of four provinces in southwest China.