| Pinus sylvestris var.mongolica is one of the most important afforestation plant species in northern of China,which has strong ability of windbreak and sand fixation.Honghuaerji is the most precious and characteristic distribution area of natural P.sylvestris var.mongolica in China.There are a lot of planted forests in this area.In recent years,we found there was a large area of death phenomenon of P.sylvestris var.mongolica plantation with different ages in this area.It is well know that P.sylvestris var.mongolica is a typical mycorrhizal-dependent tree.Whether the death of large-scale plantation is related to the change of the ectomycorrhizal fungi(ECMF)community structure in the rhizosphere of them.Therefore,this study attempted to analyze the structural characteristics of ECMF community in the rhizosphere of P.sylvestris var.mongolica in different forest stands and age classes in order to investigate the causes of death of P.sylvestris var.mongolica plantation from mycorrhizal biological point of view.Illumina MiSeq sequencing was used to analyze the the ECMF community structure characteristics of P.sylvestris var.mongolica natural forest(NF),robust planted forest(PF)and dead planted forest(DF).And P.sylvestris var.mongolica trees were divided into 4 age classes,<10,11?20,21?30 and 31?40 years old,respectively graded as ?,?,? and ?.At the same time,ECMF strains were artificially inoculated 2-month-old seedings of P.sylvestris var.mongolica under the pot condition and 2-years-old seedings of P.sylvestris var.mongolica under the nursery condition.The mycorrhizal infection rate,growth,physiological and biochemical indexes,soil physical and chemical properties and soil enzyme activities were measured after drought stress,and the excellent fungi-tree for drought resistant was selected based on these paramters.In addition,the ECMF community structure of P.sylvestris var.mongolica in different treatments under the nursery condition was analyzed.Finally,it provides some reliable evidence for directional cultivation of mycorrhizal seedings of P.sylvestris var.mongolica.The results were as followed:(1)A total of 409384 ECMF sequences and 249 operational taxonomic units(OTUs)were obtained by Illumina MiSeq from 33 soil samples,belonging to 2 phlya,4 classes,12 orders,28 families and 54 genera.The genus with largest proportion is Inocybe,followed by Russula,Tricholoma and Cortinarius.(2)Diversity and richness indices were different in different forest stands and age classes.It was showed that Simpson diversity index in DF was significantly higher than that in NF in I age class.And the observed richness,Chaol and Ace richness index in NF and PF were signicantly higher than those in DF in ? and ? age classes.However,all indices in NF and PF had no significant difference in ? age class.(3)The main dominant genera(accounting for ?10%)of Inocybe,Cortinarius,Suillus,Tricholoma and Russula in NF ? and PF ?,Tricholoma,Suillus,Pseudotomentella,Laccaria and Hygrocybe in NF ? and PF ? and Cortinarius,Amphinema and Inocybe in NF ? and PF? all had small proportion in the corresponding age classes of DF and were not dominant genera.In addition,non-metric multidimensional scaling(NMDS)and analysis of similarity(ANOSIM)showed that there were significant differences in ECMF community composition among NF,PF and DF in ? and ? age classes.However,there was no significant difference in ECMF community composition in ? and ? age classes.In NF,there was no significant difference in ECMF community composition among four age classes.However,there were significant difference in ECMF community composition in PF and DF among different age classes.(4)Redundancy analysis(RDA)showed that soil pH,available N and available P were the main factors affecting the ECMF community composition.Different soil physical and chemical properties had different effects on different dominant genera.And Ceratobasidium and Russula genera ECMF had prefer growth in high soil organic matter environment,Cortinarius genera ECMF was prefer growth in high available K environment.Tricholoma genera ECMF was more suitable for habitat in high pH and available N environments.Inocybe genera ECMF was more suitable for habitat in high available P environment.Hygrophorus,Amphinema,Lactarius,Thelephora,Hygrocybe,Tomentella,Pseudotomentella,Wilcoxina and Suillus had no obvious environment bias.(5)Five strains of Rhizopogon luteolus,Leucocortinarius bulbiger,Suillus bovinus,Pisolithus tinctorius,Chroogomphidius viscidus and their mixed strains were selected to inoculate 2-month-old seedings of P.sylvestris var.mongolica under the pot condition.After 5 month,all treatments were treated with normal moisture and drought stress.All strains could successfully infect P.sylvestris var.mongolica and form typical ectomycorrhiza,but did not in CK.Under drought stress condition,ECMF enhanced the drought resistance of seedings by increasing the underground biomass,root-shoot rate,chlorophyll content,total N content,total P content,soluble sugar content,soluble protein content,starch content,SOD activity and POD activity.Under drought stress condition,the activities of soil catalase,urease,alkaline phosphatase and sucrase in all inoculation treatments were significant higher than those in CK.(6)Under the pot condition in laboratory,the combination of mixed strains and P.sylvestris var.mongolica had the best drought resistance,followed by L.bulbiger,C.viscidus,P.tinctorius,S.bovinus and R.luteolus.(7)Under the nursery condition,the above 5 strains and their mixed strains were selected to inoculate 2-years-old seedings of P.sylvestris var.mongolica.After 12 month,all treatments were treated with normal moisture and drought stress.All strains could successfully infect P.sylvestris var.mongolica and form typical ectomycorrhiza.Under drought stress condition,ECMF enhanced the drought resistance of seedings by increasing the ground diameter,underground biomass,root-shoot rate,chlorophyll content,soluble sugar content,soluble protein content,starch content,SOD activity and POD activity.Under drought stress condition,the activities of soil alkaline phosphatase and sucrase in all inoculation treatments were significant higher than those in CK.(8)Under the nursery condition,the combination of mixed strains and P.sylvestris var.mongolica had the best drought resistance,followed by C.viscidus,P.tinctorius,L.bulbiger,S.bovinus and R.luteolus.(9)A total of 121 OTUs were obtained by Illumina MiSeq from 42 soil samples,belonging to 2 phlya,4 classes,12 orders,21 families and 30 genera.The genus with largest proportion is Inocybe,which is consistent with field experiments.Under drought stress condition,the observed richness,Chao1 and Ace richness indices in all inoculation treatment were significantly higher than those in CK.NMDS and ANOSIM analysis showed that there were significant differences in ECMF community composition among all treatments under normal moisture and drought stress condition.RDA showed that soil pH was the primary factor affecting the ECMF community composition,followed by available P and available N. |
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