Isolation, Characterization, And Antifungal Activity Of Endophytic Plant Growth Promoting Bacterial From Pigeon Pea [Cajanus Cajan (L.) Millsp.]

Nowadays,the yield of crops was enhanced mainly through the use of chemical fertilizers and chemical pesticides,but there are many adverse effects such as unsustainable effects and soil environmental pollution.The harm caused by chemical fertilizers and chemical pesticides can be improved and the sustainable development of agriculture can be achieved by applying microbial fertilizers and biocontrol agents to replace chemical fertilizers and pesticides.Endophytic bacteria are a type of microorganisms that live in various tissues and organs of healthy plants without causing any disease in the host plant.Endophytic bacteria,which can produce different kinds of antibiotics,are of great significance in controlling pests.The development of microbial fertilizers and biocontrol preparations by utilizing the e plant growth-promoting bacterial endophytes effects has great research significance in agriculture and pharmaceuticals.This study was the first to take the medicinal plant Pigeon pea as material.Isolate the endogenous bacteria of Pigeon pea using gradient dilution coating method and identify its species classification by combining morphological observation and 16S rRNA sequence analysis method.Using Salkowski colorimetric method,Mo Antimony colorimetry,CAS detection and other methods were used to study the growth-promoting characteristics of phytoalexins(IAA),phosphate solubilization,and iron-producing carriers by endogenous bacteria of peas.Study and use PCR method to further verify the antibacterial mechanism of the strain;According to the Voges-Proskauer method,B.subtilis BE-R4 was determined to contain 3-hydroxy-2-butanone,a substance that promotes plant growth.Verify the growth promotion effect of probiotic strains on wheat seedlings by in vitro pot experiments.The results of this study are as follows:1.Using pigeonpea roots,stems and leaves as materials,the tissues with different gradients were diluted and coated on three media of NA,LB and TSA for isolation and culture,passaged and purified,and 54 purified strains were finally obtained.Among them,20 strains were obtained from NA medium,18 strains were obtained from LB medium,and 16 strains were obtained from TSA medium;63%were isolated from roots,29%were isolated from leaves,and from stems.The isolates accounted for 16%.2.With reference to the "Bacterial Identification Manual" and molecular biological methods,the 54 endogenous bacteria isolated were mainly classified as:Pseudomonas sp.,Enterobacter sp.,Bacillus sp.according to the different morphology and 16S rRNA sequences of endogenous bacteria in pigeon peas.3.The 54 isolates of endophytic bacteria were isolated for proliferative research by colorimetric method and plate detection method,and 39 strains could secrete indoleacetic acid(IAA),and the secreted IAA concentration range was 0.86-80.8 ?g/mL.Among them,the concentration of the IAA produced by 8 strains were more than 10 ?g/mL;the 36 strains were able to dissolve phosphorus,and the amount of dissolved phosphorus was in the range of 2.8-25.8 ?g/mL;42 strains were capable of producing siderophores,and the R value was 1.2-8.8.There are 24 strains that can produce amylase,and R value is between 1.1 and 1.6;17 strains can produce protease,R value is between 1.2 and 1.5;28 strains can produce cellulase,R value is 1.1-6.1.4.The bacteriostatic effect of 5 pathogens was evaluated by the plate bacteriostatic method.B.subtilis BE-R4 has a better inhibitory effect on Fusarium oxysporum and Fusarium solani,and the bacteriostatic rates are 52.5%and 50%,respectively.The strain was found to contain the genes PKS-I and NRPS encoding polyketones and non-ribosomal peptide antibiotics biosynthesis through PCR detection.According to the Voges-Proskauer method,B.subtilis BE-R4 was determined to contain 3-hydroxy-2-butanone,a substance that promotes plant growth.5,B.subtilis BE-R4 showed strong life-promoting and bacteriostatic activities through screening and determination of the life-promoting strains.Pot experiments showed that the biomass of wheat seedlings could be significantly increased after treatment with B.subtilis BE-R4.The average root and leaf lengths of wheat increased by 52.1%and 44.4%,respectively;the fresh weight increase ratios of leaves and roots were 71.2%and 60.2%;the dry weight increase ratios of roots and leaves were 75.8%and 59.3%,respectively.