Effect Of Biocontrol Stain NEAU-Da4 On The Endophytic Actinobacteria Of The Soybean And Identification Of Specific Actinobacteria

Phytophthora stem and root rot, caused by Phytophthora sojae, is one of the most destructive diseases of soybean [Glycine max(L.) Merr.]. The incidence of this disease has been increasing in several soybean producing areas around the world. NEAU-Da4 producing Borrelidin was a strain with high inhibitory activity agaist Phytophthora sojae. In this study, we chose health soybean, the soybean infected by NEAU-Da4 and Phytophthora sojae(treatment 1), and the soybean infected by NEAU-Da4(treatment 2) as the separation source. The actinomyces from the roots, leaves,pods and beans of the soybean was separated by seven different culture media and two methods of grinding. The composition and difference of actinomyces community of differrnt treatment were figured out. The bioactivity of isolated actinobacteria was performed and the specific actinobacteria were identified. The results are as follows:(1) A total of 280 actinobacteria were successfully isolated from soybean specimens, with 102 from the positive control group, 89 from treatment 1 and 89 from treatment 2. According to the phenotypic and cultural characteristics, phylogenetic analysis based on 16 S rRNA, these strains were classified into five genera, and the dominant genus was Streptomyces and Micromonospora,while Baia, Plantactinospora and Glycomyces were also recovered. The Streptomyces is the advantage community of endogenous actinobacteria strains of soybean plants, which was 80% of total strains. The actinobacteria from different separation parts were comprehensive analysis, and we found that the maximum endophytic actinobacteria have been recovered from roots followed by pods, leaf and beans. In treatment 1, rare actinobacteria was accounted for 38.6%, treatment 2 was30%, and the two groups of treatment were significantly higher than that of health plant, which rare actinomyces accounted for 15.7%.(2) 32 strains with good biological activity were found from these actinobacteria, which were isolated from the soybean root. Active strains accounted for 7.1% of the total strain in health plant,22.8% in treatment 1 and 21.6% in treatment 2 respectively. The high rate of active actinobacteria in treatment 1 and treatment 2 as compared to health plant was found in our experiment.(3) Strain NEAU-gxj18, was characterized using a polyphasic approach. Phylogenetic analysis based on the 16 S r RNA gene sequences indicated that strain NEAU-gxj18 was phylogenetically related to members of the family Thermoactinomycetaceae with the highest sequence similarity to Geothermomicrobium terrae YIM 77562T(93.35 %). Growth was observed at 20-40 °C, optimaltemperature for growth was 37 °C. The diagnostic diamino acid was meso-diaminopimelic acid.The polar lipid profile consisted of diphosphatidylglycerol(DPG), phosphatidylethanolamine(PE),phosphatidylglycerol(PG), one unidentified aminophospholipid(APL) and one undientified phospholipid(PL). The whole-cell sugars were ribose, glucose and galactose. The only menaquinone was MK-7. The major fatty acids were iso-C15:0, C13:0and iso-C17:0. The DNA G+C content was 49.9 mol %. On the basis of the morphological and chemotaxonomic characteristics, phylogenetic analysis and characteristic patterns of 16 S rRNA gene signature nucleotides, strain NEAU-gxj18 represents a novel species of a new genus within the family Thermoactinomycetaceae, for which the name Baia soyae gen. nov., sp. nov. is proposed.