糙皮侧耳覆土栽培对土壤中抗生素抗性基因的影响

doi:10.13560/j.cnki.biotech.bull.1985.2024-1126

摘要/Abstract

摘要：

目的探究利用食用真菌覆土栽培模式降低土壤中抗生素抗性基因（antibiotic resistance genes, ARGs）丰度的可行性。方法以糙皮侧耳（Pleurotus ostreatus）为研究对象，采用高通量荧光定量PCR技术研究其覆土栽培模式下土壤细菌中ARGs和可移动遗传元件（mobile genetic elements, MGEs）丰度，利用16S rRNA测序技术分析出菇前后土壤细菌群落差异，并通过网络分析揭示ARGs、MGEs和细菌群落之间的共现模式。结果糙皮侧耳覆土栽培后，土壤中ARGs总相对丰度降低34.62%（P<0.01），总绝对丰度降低48.56%（P<0.01），其中氨基糖苷类、磺胺类、β-内酰胺类、大环内酯类-林肯酰胺类-链阳性菌素类和四环素类ARGs削减效果显著。MGEs总相对丰度下降20.63%，总绝对丰度降低32.99%（P<0.01）。细菌群落结构变化显著（P<0.01），解释了31.50%的ARGs变异，细菌群落结构与MGEs共同解释了8.01%的ARGs变异。结论糙皮侧耳菌丝在土壤中的增殖和发育改变了土壤微生物群落结构，达到了削减ARGs丰度的效果。利用糙皮侧耳覆土栽培的方法无需高温和发酵处理，实施操作简便，不仅能够降低土壤中ARGs丰度，而且能够正常收获食用的子实体，产生经济效益，为受ARGs污染的农业土壤的生物修复提供新思路。

关键词: 抗性污染, 农业土壤, 食用真菌, 微生物群落结构, 生物修复

Abstract:

Objective To investigate the feasibility of reducing the abundance of antibiotic resistance genes (ARGs) in soil using edible fungi soil-casing cultivation mode. Method The abundance of ARGs and mobile genetic elements (MGEs) in soil under soil-casing cultivation mode of Pleurotus ostreatus was investigated by high-throughput fluorescence quantitative PCR. The differences in soil bacterial communities before and after the formation of fruiting bodies were analyzed by using 16S rRNA sequencing. Co-occurrence patterns among ARGs, MGEs and bacterial communities were analyzed by using network analysis. Result After P. ostreatus cultivation, the total relative abundance of ARGs decreased by 34.62% (P<0.01), and the total absolute abundance decreased by 48.56% (P<0.01), with significant reductions observed in aminoglycoside, sulfonamide, β-lactamase, macrolide-lincosamide-streptogramin B, and tetracycline ARGs. The total relative abundance of MGEs decreased by 20.63%, and the total absolute abundance decreased by 32.99% (P<0.01). Bacterial community structure changed significantly (P<0.01), explaining 31.50% of the variation in ARGs, while the combined effect of bacterial community structure and MGEs explained 8.01% of the variation. Conclusion The proliferation and development of P. ostreatus mycelium in the soil changed the structure of microbial community and achieved the effect of reducing the abundance of ARGs in the soil. The method of using edible fungi soil-casing cultivation is easy to operate and requires no high temperature or fermentation treatment, which not only reduces the abundance of ARGs in the soil, but also enables the harvesting of edible mushroom, providing a new perspective for bioremediation of agricultural soils contaminated with ARGs.

Key words: resistance contamination, agricultural soil, edible fungi, microbial community structure, bioremediation

弥春霞, 许澍, 王守现, 刘宇, 宋庆港, 宋爽. 糙皮侧耳覆土栽培对土壤中抗生素抗性基因的影响[J]. 生物技术通报, 2025, 41(6): 335-343.

MI Chun-xia, Shu XU, WANG Shou-xian, LIU Yu, SONG Qing-gang, SONG Shuang. Effect of Soil-casing Cultivation of Pleurotus ostreatus on Antibiotic Resistance Genes in Soil[J]. Biotechnology Bulletin, 2025, 41(6): 335-343.

图/表 6

表1 土壤样品中ARGs和MGEs检出数量

Table 1 Number of ARGs and MGEs detected in soil samples

检测项目 Detected item	抗性分类 Resistance classification	检出数量 Number of detections
检测项目 Detected item	抗性分类 Resistance classification	S-CS	S-CG
ARGs	Aminoglycoside	24±1	26±1
	β-Lactamase	24±4	20±1
	MLSB	28±2	26±1
	Multidrug	22±2	18±0
	Phenicols	7±1	7±0
	Sulfonamide	6±4	6±1
	Tetracycline	26±2	27±0
	Vancomycin	9±2	7±3
MGEs	MGE	10±0	9±1
合计 Total		156±7	146±5

图1 糙皮侧耳覆土栽培对土壤ARGs和MGEs相对丰度和绝对丰度的影响A：ARGs和MGEs总相对丰度；B：不同抗性类别ARGs相对丰度；C：ARGs、MGEs和16S rRNA总绝对丰度；D：不同抗性类别ARGs绝对丰度。*、**和***分别表示P<0.05、P<0.01和P<0.001水平下存在显著差异

Fig. 1 Effects of soil-casing cultivation of P. ostreatus on the relative and absolute abundances of ARGs and MGEs in soilA: Total relative abundance of ARGs and MGEs. B: Relative abundance of ARGs in different resistance classifications. C: Total absolute abundance of ARGs, MGEs and 16S rRNAs. D: Absolute abundance of ARGs in different resistance classifications. *, ** and *** indicate significant differences at the P<0.05, P<0.01 and P<0.001 levels, respectively

图2 糙皮侧耳覆土栽培对土壤中整合子和转座子基因相对丰度的影响

Fig. 2 Effects of soil-casing cultivation of P. ostreatus on the relative abundance of integron and transposon genes in soil

图3 糙皮侧耳覆土栽培出菇前后土壤细菌群落变化A：香农指数；B：Chao 1指数；C：出菇前后土壤属水平细菌群落Venn图；D：土壤细菌群落在属水平上的丰度；E ：基于Bray-Curtis距离的细菌群落PCoA分析，椭圆边界代表相应组别的95%置信区间

Fig. 3 Changes in soil bacterial communities before and after the formation of fruiting bodies of P. ostreatus undersoil-casing cultivationA: Shannon index. B: Chao 1 index. C: Venn diagram of genus level before and after the formation of fruiting bodies. D: Abundance of bacterial communities at genus level. E: PCoA analysis of bacterial communities based on Bray-Curtis distance, ellipse boundaries indicate 95% confidence intervals of the corresponding groups

图4 糙皮侧耳覆土栽培出菇前后土壤理化性质变化

Fig. 4 Changes in soil physicochemical properties before and after the formation of fruiting bodies of P. ostreatus undersoil-casing cultivation

图5 糙皮侧耳覆土栽培对土壤ARGs变化的影响因素A：基于Pearson相关性（ρ>0.8，P<0.01）的ARGs、MGEs和属水平细菌群落的共现性分析。红色和绿色线条分别表示正相关和负相关，节点的大小与连接数成正比。红色节点代表细菌属，黄色节点代表MGEs，绿色节点代表ARGs；B：基于细菌群落和MGEs对ARGs变化的方差分解分析（VPA）

Fig. 5 Factors affecting the changes in ARGs of soil by soil-casing cultivation of P. ostreatusA: Co-occurrence analysis of ARGs, MGEs and genus-level bacterial communities based on Pearson correlation (ρ>0.8, P<0.01). Red and green lines indicate positive and negative correlations, respectively. The size of nodes is proportional to the number of connections. Red nodes indicate bacterial genera, yellow nodes indicate MGEs, and green nodes indicate ARGs. B: VPA analysis of changes in ARGs based on bacterial communities and MGEs

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