The Microbiome Characteristics Of Coptis Chinensis Planting Soil And Its Application Of Continuous Cropping Obstacles

Background:Coptis ranunculus is one of the most commonly used traditional Chinese medicinal materials in my country.High-frequency fungal diseases caused by continuous cropping,quality deterioration of medicinal materials,and yield reduction are important issues that need to be solved urgently.Traditional Coptis continuous cropping restoration methods include rest,quicklime treatment,pesticide spraying or fumigation incineration,etc.The restoration period is long and causes a large amount of waste of land resources and soil toxic substance residues.At present,there is no set of parameters for evaluating the effect of Coptidis soil restoration.The model can only wait for 5-7 years to test the yield and quality of Coptis after maturity,which greatly increases the time cost of repairing strategy research and development.Restoring the ecological balance of microorganisms is an important means to solve continuous obstacles and improve plant disease resistance and stress resistance.The study of the soil microbiome characteristics of Coptis chinensis has a guiding role in the development of continuous obstacle restoration strategies,and is an important research topic for the healthy development of the Coptis chinensis industry in Shizhu,Chongqing.Objective:From the perspective of micro-ecological balance,conduct microbiological research on Coptis chinensis planting soil.By analyzing microbial diversity and functions,analyzing soil physicochemical and enzyme activities and other factors,screening the key factors that affect the growth of Coptis chinensis,as a strategy for repairing the continuous cropping obstacles.This subject provides scientific support and new ideas for alleviating the continuous cropping obstacles of Coptis,developing new compound biological inoculants,and constructing the soil quality monitoring system of Coptis.Method:1.Use the method of absolute quantitative fluorescence quantitative PCR to detect5 samples in the cropping area and continuous cropping area.Draw a standard curve of absolute abundance of fungi and bacteria based on the copy numbers of 18 S and 16 S gene fragments and calculate the absolute abundance.2.Construct Coptis chinensis soil microbial gene library by amplicon sequencing and perform classification annotations,calculate the relative abundance and diversity index of Coptis chinensis soil microbial community,and analyze the cropping area with absolute abundance(PC group: Coptis chinensis planting;ZERO group: the original soil without Coptis chinensis)the structure of the true and bacterial communities.3.Using AMOVA,PCoA,and NMDS and other dimensionality reduction statistical analysis methods,the differences in the microbial communities of the Coptidis planting soil were studied,and the microbial communities with large differences in abundance and diversity were screened as biomarkers.4.Use the Tax4 Fun function to predict and obtain functional annotation information,and compare the shared or unique functional gene information between the PC group and the ZERO group in the cropping area.Use LEf Se to analyze and screen the Biomarker of fine and fungal communities,and combine the soil physicochemical and microbial functional characteristics to isolate representative communities with predictive functions and belong to biomarkers from the soil as a continuous cropping biological treatment strategy.5.Compare the two common restoration methods of quicklime(SSH)and WBM(WBM),compare the restoration effects of biological treatment(YSJ)from the three aspects of microbial community,soil physical and chemical and enzyme activity,and growth status of Coptis,and use the main component The analysis method constructs the SQI system of the Coptis soil evaluation system including biological and environmental factors,and calculates the soil quality index of the YSJ strategy.6.Using correlation analysis and combining environmental factors,an optimization method for YSJ to repair continuous cropping obstacles is proposed.Result:1.Calculation of soil microbial abundance of Coptis.Coptis planting will increase the overall abundance of bacteria and decrease the abundance of fungi in the soil.The bacterial abundance of the three treatment groups in the continuous cropping area was also significantly different.Compared with the normal cropping area(PC group and ZERO group),the bacterial abundance in the continuous cropping area was lower,while the abundance of fungi was lower.Has not changed much.2.Analysis of soil microbial structure and diversity of Coptidis.The PC group clustered 4285 bacterial OTUs,the ZERO group only had 2778,and the fungal OTU number was lower,only 1276-1707.The difference in the number of OTUs between the two groups was significant(p<0.05).The Chao1 and ACE diversity indexes of the PC group were higher than those of the ZERO group.Firmicutes(Firmicutes)is a type of bacteria whose abundance was significantly increased in the PC group.The abundance of some antagonistic bacteria such as Streptomyces,Bacillus,Paenibacillus,and Pseudomonas increased significantly in the PC group.At the same time,the abundance of plant pathogenic fungi such as Calophoma and Trichocladium also increased.3.Functional predictive analysis and screening of biomarkers.Six secondary metabolic pathways with relative abundance higher than 10% were found in the soil of Coptis chinensis.Among them,three pathways are related to carbon,nitrogen,and energy metabolism,and the abundance of pathways related to membrane transport has increased significantly(t-test,p<0.05).LEf Se analysis screened the bacteria Bacillales,Clostridiales,Methylophilaceae,Calophoma and Trichocladium as biomarkers.4.Isolation of bioremediation strains.Combined with soil enzyme activity analysis,a bacterium HL-1 that has both the ability to produce urease and belongs to the classification of biomarkers was finally screened from the soil.It was identified as Bacillus subtilis and used as a soil bioremediation strategy for Coptis chinensis.And named the YSJ processing group.5.YSJ continuous cropping repair effect evaluation.The repair effects of YSJ strategy were compared from the three aspects of microorganisms,environmental factors and the growth of Coptis chinensis.The YSJ group soil has higher OM,TN,AP and TP content and higher soil urease and cellulase activities.The Coptis SQI including soil physics and chemistry,soil microbial abundance and biomarkers was constructed,and the SQI evaluation model of the smallest data set with S-UE,Bacillus abundance and fungal abundance as evaluation factors was further optimized.The evaluation results of the two methods are consistent: the index of YSJ is higher than the other two repair methods.6.YSJ repair strategy optimization.The soil quality after YSJ restoration did not reach the level of Coptis chinensis soil.There is a very significant positive correlation between some antagonistic bacteria and biomarkers,and an optimization strategy to improve the continuous cropping soil of Coptis by “OM & TP & HL-1” is proposed.Conclusion:Coptis soil has higher bacterial abundance and diversity,and many antagonistic antibacterial and functional biomarkers are enriched in it.At the same time,Its soil microbiome also has higher carbon and nitrogen metabolism-related activities.The isolated HL-1 not only has carbon and nitrogen related enzyme production activities,but also belongs to important biomarkers.The bioremediation strategy with HL-1 as the continuous cropping area has a better restoration effect.The constructed soil quality evaluation system of Coptis chinensis can intuitively describe the difference in soil quality of different treatment groups.The soil quality after YSJ restoration is better than that of SSH and WBM.Combined with the related research,the optimization strategy of organic improvement + phosphorus enrichment + HL-1 to improve the continuous cropping soil of Coptis chinensis was further proposed.This research provides scientific support and new ideas for alleviating the continuous cropping obstacles of Coptis and the development of new composite biological inoculants.