| Soil microbial diversity,interaction and the effect of microbial functional diversity change on ecological function is one of the scientific problems in ecological research.With global climate change and the rapid development of animal husbandry,the shortage of natural grassland productivity is becoming more and more serious.The establishment of artificial grassland is beneficial to the development of animal husbandry and relieve the pressure of natural grassland.Monoculture and grass-legume mixtures are two common planting methods of artificial grassland.It is very important to utilize different planting methods,apply microbial ecology theory to optimize soil microbial community,improve soil microbial interaction,resist pathogen microorganism invasion and improve grassland ecological function.In this study,artificial grassland was established in the alpine grassland grazing area of Qinghai-Tibet Plateau as the test area.The local reference forages Elymus sibiricus L.cv.’chuancao No.2’,Elynus nutans Griseb.cv.’Aba’and Medicago sativa cv.’Beilin201’were used as the material to construct different artificial grassland communities.Monoculture and grass-legume mixtures plots were set up,and rhizobium seeding and organic fertilizer were set up.Conventional methods and Illumina high-throughput sequencing technology were used to study the short-term relationship between soil physical and chemical properties and ecological functions of different artificial grasslands and different treatments,such as forage biomass,soil microbial community diversity and function,molecular ecological network and microbial assembly mechanism.The purpose is to provide constructive scientific basis for the agronomic measures of artificial grassland in alpine region,agricultural sustainable development and ecological protection.The main research results are:1.Compared with monoculture,grass-legume mixtures significantly increased forage biomass（plant height,plant fresh weight,plant dry weight）,but had no significant effects on soil total nitrogen,total organic carbon,soil water content and soil conductivity.Theαdiversity of rhizosphere soil bacterial community was consistent.Based on the Bray-Curtis dissimilarity test,theβdiversity of the rhizosphere soil bacterial community of the two grasses was significantly different,and the Source Tracker analysis showed that the change was dominated by the mixed legume rhizosphere soil bacterial community.Proteobacteria,Actinomycetes and Firmicutes were the dominant bacteria in the rhizosphere.The abundance of Firmicutes in grass-legume mixtures was significantly lower than that in monoculture.At the genus level,Exiguobacterium,Arthrobacter,Pseudomonas and Acinetobacter were the dominant genera.Grass-legume mixutres increased the relative abundance of plant growth-promoting bacteria（Exiguobacterium）and decreased the relative abundance of pathogenic bacteria（Firmicutes）.Compared with monoculture,the bacterial community network scale of grass-legume mixtures was larger and more complex.The absolute values of positive Coheison and negative Coheison were significantly higher than those of monoculture,and the vulnerability of the network structure was significantly lower than that of monoculture.The complexity and stability of rhizosphere bacterial network were improved by grass-legume mixtures.The interaction,stability and complexity of bacterial network structure in rhizosphere of grass-legume mixtures were significantly positively correlated with the increase of forage biomass,which played a crucial role in the increase of forage biomass.2.The contents of NO3-N and TOC in soil of grass-legume mixtures were increased by rhizobium seeding,and the biomass of forage was increased by rhizobium seeding in short term.The diversity and richness of soil prokaryotes were decreased and the community structure was changed by rhizobium seeding.It significantly increased the relative abundance of beneficial bacteria such as Actinobacteria,Proteobacteria and Nitrososphaera in soil prokaryotes.The nitrate＿reduction effect was increased by 1.5%～3.3%,and the ecological function of nitrogen cycle was promoted.The abundance of nif H gene in rhizosphere soil was increased,and the nitrogen cycle of microbial-soil-forage growth was effectively improved.3.The contents of soil organic carbon（TOC）and soil total nitrogen（TN）were significantly increased by the application of organic fertilizer,but the aboveground and underground biomass of forage were not significantly increased.Theαdiversity of bulk soil bacterial community was decreased,and theαdiversity of bulk soil fungi was increased.The dominant fungal communities were Ascomycota,Basidiomycota,Zygomycota,Spheromycota,Chytridiomycota.Application of organic fertilizer increased the relative abundance of Ascomycota and decreased the relative abundance of Basidiomycota.The relative abundance of Exiguobacterium in rhizosphere soil was the highest,and that of Exiguobacterium in bulk soil was significantly lower than that in rhizosphere soil（P<0.05）.The application of organic fertilizer reduced the community size of grass and legume fungi.Reduced the stability（robustness and vulnerability）of fungal network structure;The interaction between soil fungi was reduced.The structural equation model showed that the application of organic fertilizer was negatively correlated with soil physicochemical properties and the stability of bacterial community network（-0.357）,and the stability of fungal community network was negatively correlated with herbage biomass（-0.702）.There was no significant path between the application of organic fertilizer and forage biomass.4.Based on the Molecular Ecological Network（MEN）and Taxonomic NST analysis,the core species of bacterial and fungal community network structure in the grass-legume mixtures were changed by organic fertilizer application.The network nodes in the network structure are reduced.Zero model analysis showed that the rhizosphere bacterial and fungal communities were dominated by random processes,which were increased by the application of organic fertilizer.The diversity and composition of the rhizosphere bacterial and fungal communities of grass and legume were changed by the application of organic fertilizer in the grass-legume mixtures in the alpine region,and the core species in the network structure and the assembly process of the community were also changed,which may be the main factor leading to the change of rhizosphere fungal community.5.By comparing the effects of rhizobium and organic fertilizer on soil microorganisms and forage biomass in grass-legume mixtures,it was proved that rhizobium seed dressing can increase soil beneficial microorganisms and improve forage biomass in the short term.The application of organic fertilizer increased the relative abundance of pathogenic bacteria,decreased the stability and complexity of fungal network structure,and increased the random process of bacterial and fungal communities.In a short period of time,the application of organic fertilizer in grass-legume mixtures was not conducive to improving the forage biomass.These results showed that grass-legume mixtures could significantly improve the productivity of forage,increased the relative abundance of growth-promoting bacteria in soil plants,effectively resist the invasion of pathogenic bacteria,improve the diversity and functional diversity of microbial community,increase the scale of microbial network,and improve the stability and complexity of the network.The abundance of nifH functional genes in nitrogen cycling was increased by rhizobium seeding,which was helpful to increase grass biomass.The application of organic fertilizer reduced the soil microbial diversity,reduced the stability of microbial network,and inhibited the growth and biomass of forage.This study provides scientific basis and data support for the establishment of artificial grassland in alpine region,and has important significance for promoting the systematic study of soil-microbial-forage interaction. |
