Effects of nitrogen and phosphorus additions on arbuscular mycorrhizal fungi community in the alpine meadow in Qinghai-Tibetan plateau
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摘要:目的
探究青藏高原高寒嵩草草甸生态系统中丛枝菌根真菌(AMF)群落对氮、磷添加的响应及其驱动因子，补充目前高寒草甸AMF对施肥响应研究的不足。
方法通过常规分析和高通量(Illumina-Miseq)测序，分析氮(0、7.5、15.0 g·m−2)、磷(0、7.5、15.0、30.0 g·m−2 P2O5)添加3年对青藏高原高寒小嵩草草甸土壤化学性质、AMF侵染率、OTU丰度、 Shannon多样性指数和AMF群落组成的影响。
结果测序共发现36个AMF的OTU，归属于7个科。氮、磷的添加及其交互作用对AMF侵染率、OTU丰度和 Shannon多样性指数均无显著影响。相对于低施氮处理，高施氮处理显著降低球囊霉门的相对丰度。土壤有机碳、硝态氮、有效磷和全磷含量均是影响AMF群落的土壤因子。
结论青藏高原高寒嵩草草甸根系AMF群落不受氮、磷添加的影响，群落分布与土壤因子有显著相关性。
Abstract:ObjectiveTo investigate the responses of arbuscular mycorrhizal fungi (AMF) community to nitrogen and phosphorus additions and the driven factors in the Qinghai-Tibetan alpine meadow, and have a better knowledge of the effects of fertilization on alpine meadow AMF community.
MethodThrough regular analyses and high-throughput sequencing (Illumina Miseq), we studied the effects of nitrogen addition (0, 7.5, 15.0 g·m−2) and phosphorus addition (0, 7.5, 15.0 and 30.0 g·m−2 P2O5) for three year on soil chemical property, AMF colonization rate, OTU richness, Shannon diversity index and AMF community composition in Qinghai-Tibetan alpine meadow.
ResultThe 36 AMF OTUs from seven families were detected. The addition of nitrogen, phosphorus and their interaction had no influence on AMF colonization rate, OTU richness and Shannon diversity index. Compared with low nitrogen addition treatment, high nitrogen input treatment significantly decreased the relative abundance of Glomeraceae. Soil organic carbon, nitrate nitrogen, available phosphorus and total phosphorus contents were the factors which impacted AMF community.
ConclusionRoot AMF community in Qinghai-Tibetan alpine Kobresia meadow wasn’ t affected by nitrogen or phosphorus addition. However, the distribution of community was significantly correlated with edaphic factors.
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图 1 不同施肥处理下丛枝菌根真菌侵染率
N0、N1、N2指氮施用量为0、7.5、15.0 g·m−2的处理；P0、P1、P2和P3指磷施用量为0、7.5、15.0、30.0 g·m−2的处理
Figure 1. Arbuscular mycorrhizal fungal colonization rates in different fertilization treatments
N0, N1, N2 indicated nitrogen application amount 0, 7.5, 15.0 g·m−2; P0, P1, P2, P3 indicated phosphorus application amount 0, 7.5, 15.0, 30.0 g·m−2
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图 2 不同施肥处理下分子测序的稀释曲线
N0、N1、N2指氮施用量为0、7.5、15.0 g·m−2的处理；P0、P1、P2和P3指磷施用量为0、7.5、15.0、30.0 g·m−2的处理
Figure 2. Rarefaction curve of sequencing samples in different fertilization treatments
N0, N1, N2 indicated nitrogen application amount 0, 7.5, 15.0 g·m−2; P0, P1, P2, P3 indicated phosphorus application amount 0, 7.5, 15.0, 30.0 g·m−2
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图 3 不同施肥处理下丛枝菌根真菌各科的相对丰度
N0、N1、N2指氮施用量为0、7.5、15.0 g·m−2的处理；P0、P1、P2和P3指磷施用量为0、7.5、15.0、30.0 g·m−2的处理
Figure 3. Relative abundance of different arbuscular mycorrhizal fungal families in different fertilization treatments
N0, N1, N2 indicated nitrogen application amount 0, 7.5, 15.0 g·m−2; P0, P1, P2, P3 indicated phosphorus application amount 0, 7.5, 15.0, 30.0 g·m−2
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图 4 36个丛枝菌根真菌OTU代表序列及其参考序列构建的邻接树
模型：p-distance；Boostrap值：1 000；DQ846895作为outgroup
Figure 4. Neighbor-joining tree constructed based on representative sequences of 36 arbuscular mycorrhizal fungal OTUs and their reference sequences
Model: p-distance;Boostrap value: 1 000; DQ846895 was used as an outgroup
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图 5 不同施肥处理丛枝菌根真菌OTU丰度、Shannon多样性指数及球囊霉科相对丰度
不同柱子上不同大写字母表示在P<0.001水平差异显著(Tukey’ s HSD检验)
Figure 5. OTU richness, Shannon diversity index of arbuscular mycorrhizal fungi and relative abundance of Glomeraceae in different fertilization treatments
Different capital letters on different columns indicated significant differences at P<0.001 level (Tukey’ s HSD test)
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图 6 不同施肥处理丛枝菌根真菌群落以及显著环境变量的典范对应分析图
Figure 6. Canonical correspondence analysis plot of arbuscular mycorrhizal fungal community distribution and significant environmental variables among different fertilization treatments
表 1 不同施肥处理土壤化学成分含量差异显著性分析
Table 1 Significance analyzes of soil chemical component content differences in different fertilization treatments
指标 Index 氮添加
Nitrogen addition磷添加
Phosphorus addition氮、磷添加交互作用
Interaction between nitrogen and phosphorus additionsF P F P F P 有机碳 Organic carbon 0.610 0.549 0.940 0.431 1.849 0.117 全氮 Total nitrogen 0.737 0.486 0.343 0.794 0.674 0.672 铵态氮 Ammonium nitrogen 15.662 < 0.001 1.094 0.367 1.939 0.105 硝态氮 Nitrate nitrogen 21.780 < 0.001 2.711 0.061 2.964 0.021 全磷全磷 Total phosphorus 0.304 0.740 17.135 < 0.001 0.389 0.881 有效磷 Available phosphorus 0.502 0.610 33.642 < 0.001 2.509 0.040 pH 1.336 0.276 0.329 0.804 0.948 0.473 
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表 2 不同施肥处理试验样地的土壤化学成分含量1)
Table 2 Soil chemical component contents of the experiment field in different fertilization treatments
处理 Treatment w/% w/(mg·kg−1) pH N P 有机碳
Organic carbon全氮
Total nitrogen全磷
Total phosphorus铵态氮
Ammonium nitrogen硝态氮
Nitrate nitrogen有效磷
Available phosphorusN0 P0 4.79±0.45a 0.26±0.01a 0.040±0.001cd 2.53±0.77b 7.49±1.43bc 2.72±0.49d 6.74±0.08a P1 3.61±0.79a 0.21±0.06a 0.040±0.006cd 3.91±2.16b 5.23±1.25bc 6.39±2.34cd 6.77±0.11a P2 3.25±0.58a 0.23±0.04a 0.044±0.007abcd 2.06±0.27b 6.26±1.90bc 13.64±7.19bcd 6.66±0.06a P3 3.69±0.65a 0.27±0.09a 0.057±0.012abc 2.15±1.37b 4.67±1.14c 21.78±4.00abc 6.65±0.12a N1 P0 3.59±0.42a 0.22±0.01a 0.038±0.001d 1.83±0.73b 15.32±5.27bc 2.17±1.35d 6.69±0.08a P1 3.35±0.40a 0.24±0.01a 0.046±0.003abcd 2.25±1.07b 6.96±0.95bc 9.73±0.83bcd 6.47±0.15a P2 3.77±0.47a 0.22±0.04a 0.042±0.004bcd 6.07±4.38b 9.00±0.49bc 5.49±2.61d 6.73±0.19a P3 3.83±0.73a 0.24±0.04a 0.060±0.010ab 6.28±5.37b 5.79±0.91bc 26.02±13.21ab 6.67±0.23a N2 P0 2.98±0.32a 0.19±0.01a 0.038±0.003d 27.07±11.00a 35.66±15.70ab 2.28±0.12d 6.64±0.11a P1 3.57±0.94a 0.23±0.08a 0.044±0.004abcd 13.72±9.81ab 24.76±22.05bc 4.54±1.96d 6.61±0.15a P2 3.29±0.39a 0.24±0.05a 0.047±0.003abcd 7.13±0.85b 14.94±3.58bc 11.30±5.63bcd 6.61±0.05a P3 4.24±1.50a 0.23±0.03a 0.060±0.010a 15.41±11.90ab 59.55±23.70a 35.21±5.05a 6.60±0.07a 1)表中数据为平均值±标准差；同列数据后不同小写字母表示差异显著(P < 0.05，Tukey′s HSD法)；N0、N1、N2指氮施用量为0、7.5、15.0 g·m −2的处理；P0、P1、P2和P3指磷施用量为0、7.5、15.0、30.0 g·m−2的处理
1) Data in the table were mean value ± standard deviation; Different lowercase letters in the same column indicated significant differences (P<0.05, Tukey′s HSD test); N0, N1, N2 indicated nitrogen application amount 0, 7.5, 15.0 g·m−2; P0, P1, P2, P3 indicated phosphorus application amount 0, 7.5, 15.0, 30.0 g·m−2
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