Correlation Between Changes Of Microorganism Community And Enzyme Activities In Soil Of Astragalus Membranaceus And The Root Rot Disease Severity

In recent years,the serious occurrence of root rot has brought great challenges to the sustainable development of Astragalus membranaceus.The use of biocontrol agents for regulation is the most environmentally friendly and promising disease prevention and control means at present.In this paper,the differences of soil microbial flora,bacterial diversity,community structure and soil enzyme activity in the diseased and healthy soil samples of 1-6 years were studied in order to clarify the microecological mechanism of Astragalus root rot,and then to find the early warning biological indication factor of disease occurrence,so as to provide theoretical basis for the biological control and prevention of root rot.(1)According to the results of field investigation,representative soil samples(1-4 years of Astragalus membranaceus.root circumference disease/healthy soil)were collected,and the count of bacteria,fungi and actinomycetes was analyzed by dilution plate method.It was found that,in the samples of 1-4 years surrounding soils,the number of microbial groups and their proportion in total were bacteria > actinomycetes > fungi.The total amounts of microorganisms in diseased soils were greater than that in healthy ones in 1-4 years.And the comparison result of bacterial number in diseased and healthy soils was the same as the total amount of microorganisms,but it was different about fungi and actinomycetes.At the same time,the B/F and B/A in the diseased soils were significantly higher than those in the healthy ones,indicating that the proportion of bacteria in the diseased soil was greater than that in the healthy ones.In summary,the above results indicate that the dynamic changes in the number and proportion of bacteria are the most obvious features of the root rot of Astragalus membranaceus.(2)The soil samples in the onset period of Astragalus membranaceus root rot(diseased ang healthy soils of Astragalusmembranaceus in1-6 years)were exzamined through PCR-DGGE to probe into the correlation between the microflora,microbial diversity and community structure and the severity of the root rot disease.The results showed that the bacterial diversity index of the diseased soil was lower than that of the soil at 2-5 years,and higher than that of the soil at 6 years.In addition,the diversity index of diseased soils gradually decreases in2-5 years,and rebounds in 6 years.Combined with the field survey,the change in diversity between 2-6 years was basically negatively correlated with the degree of disease.At the same time,PCA analysis found that the second year was the critical point of diversity reduction and morbidity increase.In summary,the reduction of bacterial diversity is the main cause of the root rot disease.(3)Recover the advantages and specific bands after DGGE imaging,and then perform sequencing analysis to explore the change of bacterial community structure.The results indicated that the sequence similarity rates of No.8,No.9,No.10,No.11,No.12 and No.13 were less than 95%among the 17 bacterial band,which might be potential new species.The phylogenetic tree was constructed for the other 11 bands,and it was found that all,except the No.15,belonged to Proteobacteria,which mainly distributed in Pseudomonas and Stenotrophomonas of?-proteobacteria,and Achromobacter of ?-proteobacteria.From the perspective of diseased/healthy soil,it was found that the specific bands of No.1 and No.4 in the soil were uncultured Pseudomonas sp.and Pseudomonas fluorescens,respectively.From the perspective of years,it was found that No.6 and No.16 were the common bands of each sample,which were uncultured Pseudomonas sp.and A.xylosoxidans.Among them,the abundance of No.6 uncultured Pseudomonas was significantly negatively correlated with the incidence rate,and the correlation coefficient was 0.819(P<0.05).The abundance of No.16 A.xylosoxidans was not significantly negatively correlated with the incidence.Therefore,the above four bacteria can be further studied as potential indicators ofindicating soil condition.(4)The soil enzyme activity in the diseased and healthy soil samples,including urease,cellulase,sucrase and catalase,was measured to explain the effects of changes in soil enzyme activity on root rot.The results showed that the relative reduction of urease activity and the relative increase of cellulase activity and sucrase activity were positively correlated with incidence of root rot,indicating that the level of nitrogen metabolism was significantly reduced in the diseased soil,and the level of carbon metabolism is significantly increased.At 5 and 6 years,the reduction of urease activity,the increase activity of cellulase and sucrase in diseased soil were decreasing,indicating that since the 5th year,the soil enzyme activity in the disease soil began to gradually recover to the healthy state,but the degree of different enzyme recovery was different.