长期施磷对石灰性土壤中编码碱性磷酸酶基因的细菌群落的影响

Effects of long-term phosphorus application on phoD harboring bacterial community in calcareous soil

摘要:

【目的】通过研究集约化玉米体系中长期不同供磷水平对土壤中编码碱性磷酸酶基因的细菌群落多样性、结构、组成和网络特征的影响及其与土壤有机磷形态之间的关系，为探究有机磷循环功能在微生物的群落特征及提高土壤中有机磷矿化和利用提供科学依据。【方法】本研究依托华北平原小麦玉米轮作体系石灰性土壤长期施磷的定位试验（始于2008年），试验处理包括6个供磷水平：0、12.5、25.0、50.0、100.0、200.0 kg P/hm²（分别表示为：P0、P12.5、P25、P50、P100和P200），采用核磁共振技术测定土壤中有机磷的分子形态，采用高通量测序技术分析编码碱性磷酸酶的phoD基因（alkaline phosphatase-encoding gene）的细菌群落，探究含phoD基因的细菌群落特征及其与土壤有机磷形态之间的关系。【结果】随着供磷水平的增加，土壤中磷酸单酯的含量无显著变化，磷酸二酯的含量显著增加；含phoD基因的细菌群落α-多样性整体呈先不变（P0-P50）后降低趋势（P50-P200），含phoD基因细菌的群落结构发生显著变化。优势类群中属水平的假单胞菌属（Pseudomona）和马赛菌属（Massilia）随供磷水平的增加显著降低，而壳聚糖酶产生菌属（Mitsuaria）和Kribbella随供磷水平的增加显著升高。其中，Mitsuaria与土壤中速效磷的浓度呈显著的正相关，同时与磷酸酶活性呈显著的负相关；Pseudomonas的相对丰度与土壤中总有机磷的浓度呈显著的负相关，溶杆菌属（Lysobacter）的相对丰度与土壤中总有机磷的浓度呈显著的正相关。网络分析中正相关连接线的比例在低磷条件下最高，P50处理次之，P100处理最低。其中在P0、P12.5、P25、P50处理中，特定的关键类群，如慢生根瘤菌属（Bradyrhizobium）、Stackebrandtia、伯克氏菌属（Burkholderia）、Lysobacter与有机磷（包括总有机磷、磷酸单酯和磷酸二酯）的浓度呈显著相关，而P100和P200处理下的关键类群与有机磷形态无显著相关。【结论】供磷水平通过显著影响土壤中pH、有机磷形态和数量等理化性质，进而影响含phoD基因细菌群落的α-多样性、群落结构、群落组成、网络特征及其关键类群的变化。

关键词:核磁共振;有机磷分子形态;α-多样性;群落组成;关键类群;phoD基因

Abstract:

[Objective] This study was aimed to investigate the effects of long-term phosphorus (P) supply on the diversity, structure, composition and network characteristics of bacterial community encoding alkaline phosphatase gene and their relationship with the forms of soil organic P, so as to provide scientific basis for exploring the community characteristics of organic P cycling functional microorganisms and improving the mineralization and utilization of soil organic P.[Methods] Based on the long-term P application experiment in calcareous soil of wheat-maize rotation system in North China Plain (started in 2008), the experiment included six P levels:0, 12.5, 25, 50, 100, 200 kg P/hm² (P0, P12.5, P25, P50, P100 and P200 respectively) and the molecular forms of organic P in soil were determined by nuclear magnetic resonance (NMR), high-throughput sequencing technology was used to analyze the bacterial community of phoD gene (encoding alkaline phosphatase), and explore the bacterial community characteristics of phoD gene and its relationship with the forms of organic P in soil.[Results] With the increase of P supply level, the concentration of phosphate monoester did not change significantly, but the concentration of phosphate diester increased significantly; the α-diversity of the phoD harboring bacterial community was first unchanged (P0 to P50) and then decreased (P50 to P200), and the community structure of phoD harboring bacteria changed significantly. The dominant taxa named Pseudomonas and Masslis at genus level decreased significantly, while Mitsuaria and Kribbella increased significantly with the increase of P supply level. Mitsuaria was positively correlated with the concentration of available P in soil, and negatively correlated with the activity of phosphatase. There was a significant negative correlation between the relative abundance of Pseudomonas and the concentration of total organic P in soil, and a significant positive correlation between the relative abundance of Lysobacter and the concentration of total organic P in soil. In the network analysis, the proportion of positive edges was the highest in P0 treatment, followed by P50 treatment, and the lowest in P100 treatment. In the treatments of P0, P12.5, P25 and P50, specific keystone taxa such as Bradyrhizobium, Stackbrandtia, Burkholderia, Bradyrhizobium and Lysobacter were significantly correlated with the concentration of organic P (including total organic P, phosphate monoester and phosphate diester), however, the keystone taxa under P100 and P200 treatments has no significant correlation with the forms of organic P.[Conclusion] The levels of P supply significantly affected the physicochemical properties of soil, such as pH, the forms and quantities of organic P, and then affected the changes of α-diversity, community structure, community composition, network characteristics and keystone taxa of phoD harboring bacterial community.

Key words:nuclear magnetic resonance;molecular speciation of organic P;α-diversity;community composition;keystone taxa;phoD gene