外生菌根真菌卷边桩菇促进宿主灰杨氮吸收利用特征

Characteristics of Ectomycorrhizal Fungi Paxillus involutus Promoting Nitrogen Uptake and Utilization of Its Host Populus tremula×Populus alba

【目的】分析外生菌根真菌卷边桩菇对灰杨铵态氮和硝态氮吸收、同化和代谢的影响,揭示卷边桩菇促进灰杨氮吸收利用的生理特征。【方法】对砂培灰杨幼苗接种外生菌根真菌卷边桩菇,以正常根、菌根和叶片为研究对象,采用非损伤微测技术测定NH_(4)^(+)与NO_(3)^(-)吸收速率,利用qRT-PCR分析氮转运基因的转录水平,应用分光光度计法测定NH_(4)^(+)与NO_(3)^(-)含量以及氮同化酶活性,使用元素分析仪测定总碳和总氮。【结果】接种卷边桩菇16周后,灰杨根系形成明显的外生菌根结构,菌根定殖率达55.5%。菌根灰杨的净光合速率显著高于非菌根灰杨,生物量没有明显变化。菌根灰杨根尖的NH_(4)^(+)和NO_(3)^(-)吸收速率相较非菌根灰杨显著提高。接种卷边桩菇改变灰杨根尖吸收NO_(3)^(-)的空间特征,使最大吸收点从900μm前移至300μm。与氮的吸收速率一致,菌根灰杨根中NH_(4)^(+)转运蛋白编码基因(如AMT1;1、AMT2;1和AMT3;2等)和NO_(3)^(-)转运蛋白编码基因(如NPF1.2F、NPF2.11A和NPF6.3等)的转录表达显著高于非菌根灰杨,且菌根灰杨根中NH_(4)^(+)转运蛋白编码基因的上调程度强于NO_(3)^(-)转运蛋白编码基因,菌根灰杨根和叶中的NH_(4)^(+)以及根中的NO_(3)^(-)含量显著高于非菌根灰杨。菌根灰杨根和叶中的谷氨酰胺合成酶、谷氨酸合成酶和谷氨酸脱氢酶等NH_(4)^(+)氮同化酶活性显著升高。接种卷边桩菇引起灰杨根中的总氮含量显著升高,叶中的总碳含量显著降低,菌根灰杨根和叶中的碳/氮显著低于非菌根灰杨。【结论】外生菌根真菌卷边桩菇主要通过提高铵态氮的吸收、同化和代谢,增强宿主灰杨的氮营养。

【Objective】This study aims to analyze the effects of Paxillus involutus on the absorption,assimilation and metabolism of ammonium and nitrate in Populus tremula×Populus alba,and reveal the physiological characteristics of P.involutus in promoting the nitrogen absorption and utilization of P.tremula×P.alba.【Method】In this paper,the sandcultivated P.tremula×P.alba seedlings were inoculated with ectomycorrhizal fungi,P.involutus,to form ectomycorrhiza.Then,normal roots,ectomycorrhiza,and their leaves were used as research objects.The absorption rates of ammonium and nitrate were determined using a non-invasive micro-test technique.The contents of ammonium and nitrate and the activities of nitrogen assimilation enzymes were determined with a spectrophotometer.The transcription levels of nitrogen transporter encoding genes were analyzed by a method of qRT-PCR.【Result】After 16 weeks of inoculation with P.involutus,the ectomycorrhiza was emerged in the poplar roots,with the mycorrhizal colonization rate of 55.5%.The net photosynthetic rate of mycorrhizal(M)poplars was significantly higher than that of non-mycorrhizal(NM)poplars.However,there was no significant difference in the total biomass between M poplars and NM poplars.The net influxes of NH_(4)^(+)and NO_(3)^(-)in the root tips of M poplars were increased compared to those of NM poplars.It is worth noting that the inoculation of P.involutus led to change in spatial characteristics of NO_(3)^(-)fluxes on poplar roots.The maximum NO_(3)^(-)influx point on the poplar roots was shifted from 900μm to 300μm due to P.involutus colonization.In line with the results of the ion fluxes,the expression levels of genes involved in NH_(4)^(+)transport,such as AMT1;1,AMT2;1,and AMT3;2,and genes related to NO_(3)^(-)transport,such as NPF1.2F,NPF2.11A,and NPF6.3,were higher in the roots of M poplars than those in NM poplars.Meanwhile,the increase in transcript levels of genes involved in NH_(4)^(+)transport was stronger than that of genes related to NO_(3)^(-)transport in the roots of M poplars.Correspondingly,the contents of NH_(4)^(+)and NO_(3)^(-)in the roots and leaves of M poplars were higher than those of NM poplars.In addition,the activities of enzymes including glutamine synthase(GS),glutamate synthase(GOGAT)and glutamate dehydrogenase(GDH),which are involved in NH_(4)^(+)assimilation,were increased significantly in the roots and leaves of M poplars in comparison with NM poplars.Inoculation with P.involutus caused a significant increase in the total nitrogen in the roots and a significant decrease in the total carbon in the leaves of M poplars,resulting in significantly lower C/N levels in the roots and leaves of M poplars than those of NM poplars.【Conclusion】These results suggest that the ectomycorrhizal fungus,P.involutus,enhances the nitrogen nutrition of its host P.tremula×P.alba by mainly increasing the absorption,assimilation and metabolism of ammonium nitrogen.