Effects of biogas slurry combined with chemical fertilizer on Allium fistulosum yield, soil nutrient, microorganism, and enzymatic activity
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摘要: 明确沼液替代化肥的合适比例以及沼液对大葱产量和土壤养分、微生物含量以及酶活性的影响, 可为沼液的合理施用提供理论依据。试验设计了不施肥(CK)、化肥(CF)、沼液替代25%化肥氮(25BS)、沼液替代50%化肥氮(50BS)、沼液替代75%化肥氮(75BS)、沼液替代100%化肥氮(100BS) 6个处理, 分析了大葱产量、土壤养分含量、土壤磷脂脂肪酸含量、土壤碳氮磷循环相关酶的活性, 并通过偏最小二乘法路径模型(PLS-PM)探究上述指标的因果关系。结果表明, 与CK相比, CF和各沼液处理(25BS、50BS、75BS和100BS)均能显著提高大葱产量(P<0.05), 分别提高37.2%、75.9%、118.9%、99.8%和59.3%, 大葱产量随着沼液替代化肥比例的增加呈现先增加后降低的趋势, 其中50BS处理的大葱产量最高达59.9 t·km−2。施用沼液可有效改善土壤养分状况, 与CK相比, 施用沼液显著提高土壤有机碳(SOC, 19.5%~65.8%)、全氮(TN, 40.5%~69.6%)、氨态氮(NH4+, 26.8%~77.4%)、硝态氮(NO3−, 30.1%~41.9%)、速效磷(AP, 10.5%~40.6%)、速效钾(5.4%~8.5%)含量。施用沼液可有效提高土壤微生物含量以及土壤酶活性, 与CK相比, 施用沼液显著提高细菌、真菌、放线菌等微生物的磷脂脂肪酸含量(P<0.05), 降低了革兰氏阳性细菌∶革兰氏阴性细菌的比例(P<0.05), 有助于提高土壤碳、氮、磷相关循环酶活性(P<0.05); 但是, 随着沼液替代比例的增加, 细菌、革兰氏阳性细菌、真菌、总磷脂脂肪酸含量以及碳、氮、磷相关循环酶活性呈先增加后降低的趋势(P<0.05)。PLS-PM分析表明, 沼液通过增加SOC、TN、NH4+、NO3−、AP养分含量, 进而提高土壤微生物含量以及碳、氮循环酶活性, 并提升大葱产量, 但是过量的沼液可导致土壤电导率升高, 并对土壤微生物活性和大葱生长产生抑制效果。本研究表明, 短期施用沼液可显著提高大葱产量, 有效改善土壤养分状况, 并有利于土壤微生物含量以及酶活性提高, 其中以沼液替代50%化肥氮的处理效果最优, 但是沼液并不能完全替代化肥, 施用过量的沼液容易造成土壤盐分累积, 不利于大葱和土壤微生物的生长。Abstract: In this study, we conducted an investigation to determine the optimal ratio of biogas slurry and chemical fertilizer, and the effect of biogas slurry combined with chemical fertilizer on Allium fistulosum yield, soil nutrient levels, microorganism contents, and enzymatic activity. The objective of this research was to establish a methodological and theoretical foundation for the rational utilization of biogas slurry. The field experiment consisted of six treatments: no fertilization (CK), chemical fertilizer (CF), 25% substitution inorganic N by biogas slurry N (25BS), 50% substitution inorganic N by biogas slurry N (50BS), 75% substitution inorganic N by biogas slurry N (75BS), and 100% substitution inorganic N by biogas slurry N (100BS). We measured the Allium fistulosum yield, soil nutrient content, phospholipid fatty acid (PLFA) content, and extracellular enzymes activities involved in C, N, P cycling. Through Partial Least Square Path Model (PLS-PM), we explored the variations in these parameters to elucidate their internal correlations. The results of the study indicated that CF and biogas slurry (25BS, 50BS, 75BS, 100BS) treatments significantly increased the Allium fistulosum yield compared to the CK (P<0.05), with increases of 37.2%, 75.9%, 118.9%, 99.8% and 59.3% respectively. Furthermore, as the substitution percentage of inorganic N by biogas slurry N increased, the yield showed a tendency of initial increase and then decrease, with the highest yield of 59.9 t·km-2 observed for the 50BS. The application of biogas slurry is effective for improving soil nutrient contents. Specifically, compared to CK, the biogas slurry significantly increased the contents of soil organic carbon (SOC), total nitrogen (TN), ammoniacal nitrogen (NH+ 4), nitrate nitrogen (NO- 3), available phosphorus (AP), available potassium by 19.5%~65.8%, 40.5%~69.6%, 26.8%~77.4%, 30.1%~41.9%, 10.5%~40.6%, and 5.4%~8.5%. The application of biogas slurry demonstrated a notable enhancement in both soil microbial contents and enzymatic activity. Compared to CK, the biogas slurry significantly increased the PLFAs contents of bacteria, fungi and actinomycete (P<0.05), while concurrently reducing the ratio of gram-positive to gram-negative bacteria. This shift was advantageous in improving the activity of extracellular enzymes involved in C, N, P cycling. However, with the increasing ratio of inorganic N substitution by biogas slurry, it is worth noting that the bacteria, gram-negative bacteria, fungi, total PLFAs contents and the extracellular enzymes activities involved in C, N, P cycling exhibited a tendency of initial increase followed by a subsequent decrease. The results of PLS-PM indicated that the observed increase of Allium fistulosum yield after biogas slurry application can be attributed to improvements of SOC, TN, NH+ 4, NO- 3, AP contents, microbial contents, and the enhanced activity of extracellular enzymes involved the N cycling. Nevertheless, the excessive application of biogas slurry led to the elevated soil electrical conductivity (EC), which inhibited the microbial activities and ultimately reduced the Allium fistulosum yield. In conclusion, this research illustrated that the temporary utilization of biogas slurry contributed to enhancements of the Allium fistulosum yield, effective improvement of soil nutrients level, and promotion of soil microbial contents and enzymes activities. Notably, optimal substitution percentage of inorganic N by biogas slurry was 50% to achieve the highest improvement. However, it is important to note that the biogas slurry cannot completely substitute the chemical fertilizer, for that excessive use may lead to the accumulation of soil salinity, adversely affecting the growth of Allium fistulosum and microorganisms.
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Key words:
- Biogas slurry /
- Soil fertility /
- Soil microorganism /
- Soil extracellular enzyme
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图 1 各施肥处理大葱产量
不同小写字母表示处理间显著差异(P<0.05)。Different lowercase letters on the bars indicate significant difference among treatment (P<0.05).
Figure 1. Allium fistulosum L. yields in the different fertilization treatments
图 2 各施肥处理磷脂脂肪酸(PLFA)含量
不同小写字母表示处理间显著差异(P<0.05); 最右侧图中的点线图分别为革兰氏阳性细菌与阴性细菌的比值和真菌与细菌的比值。Different lowercase letters on the bars indicate significant difference among treatment (P<0.05). The point plots in the right side of figure are ratio of gram-positive to gram-negative bacteria, and ratio of fungi to bacteria, respectively.
Figure 2. Phospholipid fatty acid (PLFA) contents in the different fertilization treatments
图 3 土壤微生物群落结构与土壤理化性质的冗余分析
Total PLFA: 总磷脂脂肪酸; Fungi: 真菌; Bacteria: 细菌; Act: 放线菌; G+/G-: 革兰氏阳性细菌/革兰氏阴性细菌; F/B: 真菌/细菌; C: 土壤有机碳; N: 土壤全氮; P: 土壤速效磷; K: 土壤速效钾。Act: Actinomycetes; G+/G-: ratio of gram-positive to gram-negative bacteria; F/B: ratio of fungi to bacteria; C: soil organic carbon; N: soil total nitrogen; P: soil available phosphorus; K: soil available potassium.
Figure 3. Redundancy analyses (RDA) between the soil microbial community composition and soil properties
图 4 聚类热图分析不同施肥处理对土壤酶活性的影响
图中数据为酶活性标准化后的结果; 颜色梯度表示酶活性的差异, 从浅灰到黑色表示酶活性从低到高; 小写字母代表不同施肥处理间酶活性的差异显著性(P<0.05); AG: α-葡糖苷酶; BG: β-葡糖苷酶; CB: β-纤维二糖苷酶; XYL: β-木糖苷酶; NAG: 乙酰氨基葡萄糖苷酶; LAP: 亮氨酸氨基肽酶; APP: 丙氨酸氨基肽酶; PHOS: 酸性磷酸酶。The data in the figure is the standardized enzyme activity. The color gradients indicate the differences in enzyme activities. The color from light gray to black indicate low to high for enzyme activities. Lowercase letters in each column represent significant differences among the fertilization treatments (P<0.05). AG: α-glucosidase; BG: β-glucosidase; CB: β-cellobiosidase; XYL: β-xylosidase; NAG: N-Acetyl-glucosaminidase; LAP: L-leucine aminopeptidase; APP: Alanine aminopeptidase; PHOS: Acid phosphatase.
Figure 4. The effect of different fertilization treatments on soil enzyme activities analyzed by Cluster heat map
图 5 土壤酶活性与土壤理化性质的冗余分析
AG: α-葡糖苷酶; BG: β-葡糖苷酶; CB: β-纤维二糖苷酶; XYL: β-木糖苷酶; NAG: 乙酰氨基葡萄糖苷酶; LAP: 亮氨酸氨基肽酶; APP: 丙氨酸氨基肽酶; PHOS: 酸性磷酸酶; C: 土壤有机碳; N: 土壤全氮; P: 土壤速效磷; K: 土壤速效钾。AG: α-glucosidase; BG: β-glucosidase; CB: β-cellobiosidase; XYL: β-xylosidase; NAG: N-Acetyl-glucosaminidase; LAP: L-leucine aminopeptidase; APP: alanine aminopeptidase; PHOS: acid phosphatase; C: soil organic carbon; N: soil total nitrogen; P: soil available phosphorus; K: soil available potassium.
Figure 5. The redundancy analyses (RDA) between the soil enzyme activities and soil properties
图 6 基于偏最小二乘回归法的土壤微生物对土壤酶活性影响的分析
AG: α-葡糖苷酶; BG: β-葡糖苷酶; CB: β-纤维二糖苷酶; XYL: β-木糖苷酶; NAG: 乙酰氨基葡萄糖苷酶; LAP: 亮氨酸氨基肽酶; APP: 丙氨酸氨基肽酶; PHOS: 酸性磷酸酶; B: 细菌; F: 真菌; Act: 放线菌; G+/G-: 革兰氏阳性细菌/革兰氏阴性细菌; F/B: 真菌/细菌。*和**分别表示P<0.05和P<0.01水平上的差异显著。AG: α-glucosidase; BG: β-glucosidase; CB: β-cellobiosidase; XYL: β-xylosidase; NAG: N-Acetyl-glucosaminidase; LAP: L-leucine aminopeptidase; APP: alanine aminopeptidase; PHOS: acid phosphatase; B: bacteria; F: Fungi; Act: actinomycetes; G+/G−: ratio of Gram-positive to Gram-negative bacteria; F/B: ratio of fungi to bacteria. * and ** indicate statistical significance at P<0.05 and P<0.01, respectively.
Figure 6. The influence of soil microbial groups on different soil enzyme activities using the partial least squares regression
图 7 偏最小二乘路径模型分析土壤生物化学性质对大葱产量的直接和间接影响
SOC: 土壤有机碳; TN: 土壤全氮; AP: 土壤速效磷; AG: α-葡糖苷酶; BG: β-葡糖苷酶; CB: β-纤维二糖苷酶; XYL: β-木糖苷酶; NAG: 乙酰氨基葡萄糖苷酶; LAP: 亮氨酸氨基肽酶; APP: 丙氨酸氨基肽酶; Enzy-C: 碳循环相关酶; Enzy-N: 氮循环相关酶; PLFA: 总磷脂脂肪酸; yield: 大葱产量。箭头旁数值为标准化路径系数, 实线表示影响显著, 虚线表示影响不显著; 模型拟合优度值(Goodness of fit, GOF)评价路径模型的拟合度; R2值表示大葱产量被其他变量解释的程度。SOC: soil organic carbon; TN: total nitrogen; AP: available phosphorus; AG: α-glucosidase; BG: β-glucosidase; CB: β-cellobiosidase; XYL: β-xylosidase; NAG: N-Acetyl-glucosaminidase; LAP: L-leucine aminopeptidase; APP: Alanine aminopeptidase; Enzy-C: enzymes activities involved in cabon cycling; Enzy-N: enzymes activities involved in nitrogen cycling; yield: Allium fistulosum yield. Number on the arrows indicate standardized path coefficients. Solid-line path indicates the significant impact, and dashed-line path indicates the non-significant impact. The path model is assessed using the Goodness of fit (GOF) index. R2 indicates the proportion of the Allium fistulosum L. yield explained by the other variables.
Figure 7. The direct and indirect effects of soil biological and chemical properties on the Allium fistulosum L. yield using partial least squares path modeling (PLS-PM)
表 1 各试验处理施肥量
Table 1. Amounts of fertilizers for each fertilization treatment
处理
Treatment全氮Total N(kg·km−2) P2O5 (kg·km−2) K2O (kg·km−2) 沼液
Biogas slurry化学氮肥
Chemical N fertilizer沼液
Biogas slurry化学磷肥
Chemical P fertilizer沼液
Biogas slurry化学钾肥
Chemical K fertilizerCK 0 0 0 0 0 0 CF 0 160 0 80 0 100 25BS 40 120 18.3 61.7 19.4 80.6 50BS 80 80 36.6 33.4 38.8 61.2 75BS 120 40 54.9 25.1 58.2 41.8 100BS 160 0 73.2 6.8 77.6 22.4 
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表 2 各施肥处理土壤养分含量
Table 2. The soil nutrition content in different fertilization treatments
处理
Treatment有机碳
Organic carbon
(g·kg−1)全氮
Total nitrogen
(g·kg−1)pH EC
(mS·cm−1)铵态氮
Ammoniumu nitrogen
(mg·kg−1)硝态氮
Nitrate nitrogen
(mg·kg−1)有效磷
Available
Phosphorus
(mg·kg−1)速效钾
Available potassium
(mg·kg−1)CK 8.89±0.45d 0.79±0.11d 7.99±0.11ab 0.44±0.05d 15.86±1.22d 92.7±9.3c 27.6±1.7c 129.0±11.3c CF 9.16±0.42d 1.05±0.02c 8.04±0.05a 0.45±0.05d 18.16±1.85cd 108.4±6.6cd 31.3±1.2b 137.6±4.1b 25BS 10.63±1.31c 1.11±0.03bc 7.97±0.05ab 0.48±0.02d 20.11±1.34c 120.6±12.7ab 30.5±3.9b 136.3±10.7b 50BS 12.01±0.92c 1.21±0.04ab 7.96±0.09ab 0.66±0.07c 24.67±1.45b 114.6±11.4ab 31.5±1.9b 131.3±4.7b 75BS 13.78±0.58b 1.27±0.06a 7.87±0.08b 1.01±0.10b 26.12±1.98ab 130.1±15.9ab 37.1±1.59a 144.0±6.1a 100BS 14.74±0.65a 1.34±0.04a 7.88±0.06b 1.44±0.12a 28.14±1.74a 131.5±10.6a 38.8±2.2a 147.3±5.1a 表中数据为平均值±标准差(n=3), 同列数据后不同小写字母表示处理间显著差异(P<0.05)。The data are means ± standard error (n=3). Values followed by different lowercase letters in the same column indicate significant difference among treatment (P<0.05).
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