小麦和豌豆填闲对白菜幼苗生长和土壤微生物群落结构的影响

doi:10.3864/j.issn.0578-1752.2024.03.010

摘要/Abstract

摘要：

【目的】明确小麦和豌豆填闲对白菜幼苗生长和土壤微生物群落结构的影响，为麦、豆填闲减轻十字花科蔬菜的连作障碍提供理论依据和技术支撑。【方法】分别以白菜和白菜连作土为供试材料和供试土壤，设置小麦填闲（W）、箭筈豌豆填闲（P）及小麦和豌豆混合填闲（WP）3个填闲处理，以不填闲作物为对照（CK），分析不同填闲处理对白菜幼苗生长的影响，并利用实时荧光定量PCR（qPCR）和Illumina MiSeq技术，探究小麦和豌豆填闲对白菜根际土壤微生物群落的影响，通过Spearman相关性分析，找出与白菜幼苗生长相关的关键微生物类群，再通过环境因子关联性分析，探明土壤化学性质的变化对土壤微生物群落结构的影响。【结果】与对照相比，3种填闲处理均促进了白菜的生长，降低了土壤EC值，混合填闲处理显著降低了土壤速效钾含量，小麦填闲处理提高了土壤pH。qPCR结果表明，填闲处理对土壤总细菌丰度没有产生显著影响，但显著增加了土壤总真菌的丰度，豌豆填闲处理及小麦和豌豆混合填闲处理显著降低了芽孢杆菌和假单胞菌的丰度。Illumina MiSeq结果发现，在属水平上，填闲处理均显著增加了TM7a的相对丰度，但降低了Leptolyngbya_EcFYyyy-00、瓶毛壳属（Lophotrichus）、无茎真菌属（Acaulium）、Sodiomyces的相对丰度；小麦和豌豆混合填闲处理显著增加了鞘氨醇单胞菌属（Sphingomonas）和马赛菌属（Massilia）的相对丰度，显著降低了镰刀菌属（Fusarium）的相对丰度。基于Spearman的相关性分析表明，鞘氨醇单胞菌属（Sphingomonas）、TM7a、马赛菌属（Massilia）和芽单胞菌属（Gemmatimonas）与白菜生长呈显著正相关，Leptolyngbya_EcFYyyy-00、无茎真菌属（Acaulium）、瓶毛壳属（Lophotrichus）、Sodiomyces和镰刀菌属（Fusarium）与白菜生长呈显著负相关。填闲也改变了细菌和真菌的多样性指数，豌豆填闲处理降低了土壤细菌香农指数和逆辛普森指数，降低了真菌香农指数，增加了逆辛普森指数；小麦填闲处理和混合填闲处理增加了真菌香农指数，降低了逆辛普森指数。PCoA分析表明土壤微生物群落结构差异显著。土壤EC值是显著影响土壤微生物群落结构的土壤环境因子。【结论】填闲处理均能促进白菜幼苗生长，其中以小麦和豌豆混合填闲对白菜幼苗的促生效果最好。填闲处理增加了潜在促生菌鞘氨醇单胞菌属、TM7a、马赛菌等属的相对丰度，混合填闲处理显著降低了Leptolyngbya_EcFYyyy-00和镰刀菌属的相对丰度，增加了潜在生防菌毛壳菌的相对丰度。

关键词: 填闲, 白菜, 促生, 土壤化学性质, 微生物群落结构

Abstract:

【Objective】 This study aimed to investigate the impact of cover crops (wheat and common vetch) on the growth of Chinese cabbage seedlings and the structure of the soil microbial community. The findings could provide the theoretical and technical support for using wheat and common vetch cover crops to alleviate continuous cropping obstacles in Chinese cabbage production. 【Method】In this experiment, there were four treatments: wheat cover crop treatment (W), common vetch cover crop treatment (P), mixture of wheat and common vetch treatment (WP), and no cover crop treatment (CK). The effects of different cover crop treatments on the growth of Chinese cabbage seedlings were studied. Moreover, the effects of wheat and common vetch cover crops on the microbial community of Chinese cabbage rhizosphere were also investigated through qPCR and Illumina MiSeq techniques. In addition, Spearman correlation analysis was conducted to identify the key soil microbial taxa related to Chinese cabbage growth. Then, the changes in soil chemical properties on soil microbial community structure were explored by environmental factor correlation analysis. 【Result】Compared with CK, the cover crop treatments had positive effects on Chinese cabbage growth and decreased soil electrical conductivity (EC) value. The mixed cover crop treatment significantly decreased soil available potassium content, whereas wheat cover crop treatment increased soil pH. The qPCR results showed that the abundance of soil bacterial community was not significantly affected by the cover crop treatment, but increased the abundance of soil fungal community. Both common vetch cover crop treatment and mixed wheat and common vetch cover crop treatment significantly reduced the abundances of Bacillus spp. and Pseudomonas spp. communities. The Illumina MiSeq analysis showed that the relative abundance of genus TM7a was significantly increased by cover treatments, while the relative abundances of Leptolyngbya_EcFYyyy-00, Lophotrichus, Acaulium, and Sodiomyces were decreased. The mixed cover crop treatment significantly increased the relative abundance of Sphingomonas and Massilia and significantly decreased the relative abundance of Fusarium. Spearman correlation analysis showed that Sphingomonas, TM7a, Massilia, and Gemmatimonas were positively correlated with growth. Leptolyngbya_EcFYyyy-00, Acaulium, Lophotrichus, Sodiomyces, and Fusarium were significantly negatively correlated with the growth of Chinese cabbage. Moreover, these cover crop treatments influenced bacterial and fungal diversity indices. The Shannon index and inverse Simpson index for soil bacterial community and Shannon index of soil fungal community significantly decreased in cover common vetch treatment. In contrast, the inverse Simpson index of soil fungal community was increased. The mixed cover crop treatment increased the fungal Shannon index, while the inverse Simpson index of soil fungal community decreased. Principal Coordinates Analysis (PCoA) showed significant differences in soil microbial community structure, with soil EC value as a major environmental factor affecting the structure.【Conclusion】The cover crop treatments exhibited growth-promoting effects on Chinese cabbage seedlings, and the best effect was found in the mixed cover crop treatment. The relative abundances of some Sphingomonas, TM7a, Massilia, other potential growth-promoting bacteria were increased in the cover crop treatments. The relative abundances of some potential plant pathogens Leptolyngbya_EcFYyyy-00 and Fusarium were decreased and the relative abundance of potential biocontrol agent Chaetomium was increased in the mixed cover crop treatment.

Key words: cover, Chinese cabbage, promote growth, soil chemical properties, microbial community structure

王清慧, 李乃荟, 张一平, 狄成乾, 吴凤芝. 小麦和豌豆填闲对白菜幼苗生长和土壤微生物群落结构的影响[J]. 中国农业科学, 2024, 57(3): 555-569.

WANG QingHui, LI NaiHui, ZHANG YiPing, DI ChengQian, WU FengZhi. Effects of Wheat and Common Vetch Cover Crops on Chinese Cabbage Seedling Growth and Soil Microbial Community Structure[J]. Scientia Agricultura Sinica, 2024, 57(3): 555-569.

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链接本文: https://www.chinaagrisci.com/CN/10.3864/j.issn.0578-1752.2024.03.010

https://www.chinaagrisci.com/CN/Y2024/V57/I3/555

图/表 12

表1

qPCR反应体系和程序"

菌名 Bacterium name	引物 Primer	qPCR反应体系 qPCR reaction system (20 μL)	qPCR反应程序 qPCR reaction procedure
细菌 Bacteria	338F/518R^[19]	9 μL 2×SYBR Green，0.4 μL引物，2 μL样品DNA，8.6 μL ddH₂O 9 μL 2×SYBR Green, 0.4 μL primer, 2 μL sample DNA, 8.6 μL ddH₂O	95 ℃预变性5 min，95 ℃变性50 s，62 ℃退火30 s，72 ℃延伸1 min，72 ℃终延伸10 min，30个循环 Pre-denaturation at 95 ℃ for 5 min, denaturation at 95 ℃ for 50 s, annealing at 62 ℃ for 30 s, extension at 72 ℃ for 1 min, final extension at 72 ℃ for 10 min, 30 cycles
真菌 Fungus	ITS1F/ITS4F^[20]	10 μL 2×SYBR Green，1 μL引物，1 μL样品DNA，8 μL ddH₂O 10 μL 2×SYBR Green, 1 μL primer, 2 μL sample DNA, 8 μL ddH₂O	94 ℃预变性3 min，94℃变性5 s，58 ℃退火20 s，72 ℃延伸20 s，72 ℃终延伸10 min，43个循环 Pre-denaturation at 94 ℃ for 3 min, denaturation at 94 ℃ for 5 s, annealing at 58 ℃ for 20 s, extension at 72 ℃ for 20 s, final extension at 72 ℃ for 10 min, 43 cycles
芽孢杆菌 Bacillus spp.	BacF/BacR^[21]	9 μL 2×SYBR Green, 0.6 μL引物，2 μL样品DNA，8.4 μL ddH₂O 9 μL 2×SYBR Green, 0.6 μL primer, 2 μL sample DNA, 8.4 μL ddH₂O	95 ℃预变性3 min，95 ℃变性10 s，50 ℃退火30 s，72 ℃延伸32 s，72℃终延伸10 min，40个循环 Pre-denaturation at 95 ℃ for 3 min, denaturation at 95 ℃ for 10 s, annealing at 50 ℃ for 30 s, extension at 72 ℃ for 32 s, final extension at 72 ℃ for 10 min, 40 cycles
假单胞菌 Pseudomonas spp.	PsF/PsR^[22]	10 μL 2×SYBR Green，2 μL引物，2 μL样品DNA，6 μL ddH₂O 10 μL 2×SYBR Green, 2 μL primer, 2 μL sample DNA, 6 μL ddH₂O	95 ℃预变性10 min，95 ℃变性10 s，50 ℃退火30 s，72℃延伸32 s，72 ℃终延伸10 min，35个循环 Pre-denaturation at 95 ℃ for 10 min, denaturation at 95 ℃ for 10 s, annealing at 50 ℃ for 30 s, extension at 72 ℃ for 32 s, final extension at 72 ℃ for 10 min, 35 cycles

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处理 Treatment	EC值 EC Value (mS·cm^-1)	pH	速效磷 Available P (mg·kg^-1)	速效钾 Available K (mg·kg^-1)	有机碳 Organic carbon (g·kg^-1)	全碳 Total C (g·kg^-1)	全氮 Total N (g·kg^-1)
对照CK	0.44±0.01a	7.04±0.10b	120.81±2.28a	308.15±16.03a	19.67±0.90a	20.29±0.86a	2.46±0.16a
小麦W	0.34±0.01b	7.15±0.01a	120.44±2.86a	283.63±17.15ab	20.47±1.18a	21.96±1.80a	2.50±0.18a
箭筈豌豆P	0.36±0.02b	7.13±0.02ab	121.34±2.01a	273.13±6.52ab	19.46±0.10a	22.13±1.21a	2.50±0.10a
小麦+箭筈豌豆WP	0.35±0.03b	7.05±0.01ab	119.12±1.98a	247.52±35.40b	18.87±0.96a	20.12±1.87a	2.40±0.10a

属 Genus	CK	W	P	WP
鞘氨醇单胞菌属Sphingomonas	4.386±0.474c	5.192±0.205bc	7.292±1.884a	6.771±0.431ab
芽孢杆菌属Bacillus	2.784±0.403b	4.130±0.110a	3.964±0.640a	3.511±0.332ab
RB41	3.840±0.425a	3.258±0.051a	2.895±1.601a	2.711±0.631a
芽单胞菌属Gemmatimonas	1.508±0.154b	1.750±0.123ab	2.043±0.419a	1.927±0.108ab
TM7a	0.574±0.023c	0.917±0.098b	1.220±0.084a	1.104±0.122a
Skermanella	0.927±0.109b	1.394±0.210a	1.205±0.310ab	1.140±0.072ab
Leptolyngbya_EcFYyyy-00	3.117±0.311a	2.282±0.006b	0.805±0.487c	1.222±0.454c
Gaiella	0.727±0.088b	0.843±0.037ab	1.030±0.239a	0.833±0.079ab
Bryobacter	0.755±0.078c	1.046±0.093a	0.893±0.053bc	0.966±0.068ab
溶杆菌属Lysobacter	0.763±0.031b	0.795±0.062ab	0.869±0.226ab	1.051±0.126a
嗜盐囊菌属Haliangium	0.789±0.110ab	0.675±0.063b	0.901±0.152a	0.883±0.049a
慢生根瘤菌属Bradyrhizobium	0.607±0.083b	0.741±0.071ab	0.832±0.185ab	0.883±0.145a
马赛菌属Massilia	0.518±0.049c	0.611±0.019bc	1.163±0.280a	0.818±0.099b
芽球菌属Blastococcus	0.591±0.089b	0.746±0.136ab	0.914±0.175a	0.755±0.156ab
Ellin6055	0.648±0.054a	0.717±0.073a	0.845±0.210a	0.724±0.085a
Flavisolibacter	0.716±0.143a	0.846±0.064a	0.859±0.195a	0.939±0.100a
酸杆菌属Acidibacter	0.604±0.106a	0.717±0.015a	0.654±0.196a	0.699±0.215a
嗜酸菌属Acidovorax	0.948±0.086bc	1.104±0.042ab	0.813±0.151c	1.173±0.083a
Ellin6067	0.596±0.059a	0.651±0.110a	0.713±0.153a	0.683±0.109a
Actinoplanes	0.506±0.070a	0.543±0.019a	0.661±0.141a	0.593±0.029a

属Genus	CK	W	P	WP
油壶菌属Olpidium	17.448±3.632a	6.158±0.361b	19.032±0.892a	5.131±0.661b
腐质霉属Humicola	11.457±0.907a	9.157±0.101c	9.518±0.727bc	10.678±0.565ab
被孢霉属Mortierella	1.950±0.299c	12.448±0.413a	8.334±1.222b	9.261±0.986b
镰刀菌属Fusarium	8.535±0.668a	8.470±0.345a	6.131±0.345b	6.757±0.336b
光黑壳属Preussia	5.504±0.511a	4.029±0.026bc	3.246±0.064c	4.195±0.745b
枝鼻菌属Cladorrhinum	5.288±0.186a	3.379±0.121b	5.057±0.459a	5.361±0.202a
瓶毛壳属Lophotrichus	4.342±0.203a	3.070±0.172b	2.502±0.099c	2.576±0.321c
赤霉属Gibberella	0.155±0.031d	4.494±0.175a	0.821±0.068c	2.390±0.140b
被毛枝葡萄孢属Botryotrichum	0.734±0.031d	3.664±0.157a	1.422±0.136c	2.590±0.192b
Plectosphaerella	0.247±0.031c	7.769±0.149a	1.512±0.098b	0.388±0.058c
酵母属Remersonia	1.772±0.222a	0.849±0.109c	1.546±0.132ab	1.387±0.153b
毛壳菌属Chaetomium	0.822±0.126b	0.538±0.092b	0.729±0.046b	2.581±0.463a
闭小囊菌属Kernia	0.869±0.087ab	0.741±0.142bc	1.015±0.089ab	0.667±0.037c
无茎真菌属Acaulium	1.127±0.115a	0.792±0.054b	0.754±0.088b	0.639±0.064b
产油菌属Solicoccozyma	0.675±0.063ab	0.760±0.081a	0.626±0.065b	0.670±0.013ab
Sodiomyces	0.890±0.095a	0.549±0.055bc	0.666±0.118b	0.475±0.048c
曲霉属Aspergillus	0.626±0.123a	0.398±0.048b	0.458±0.066bc	0.577±0.052ab
枝顶孢属Acremonium	0.786±0.027a	0.193±0.032c	0.128±0.023d	0.297±0.047b
Tausonia	0.274±0.034bc	0.846±0.199a	0.331±0.088b	0.090±0.026c
头束霉属Cephalotrichum	0.216±0.061bc	1.008±0.186a	0.102±0.033c	0.323±0.028b

细菌Bacteria	r	真菌Fungi	r
鞘氨醇单胞菌属Sphingomonas	0.818**	无茎真菌属Acaulium	-0.867***
Leptolyngbya_EcFYyyy-00	-0.79**	瓶毛壳属Lophotrichus	-0.811**
TM7a	0.769**	Sodiomyces	-0.741**
马赛菌属Massilia	0.762**	镰刀菌属Fusarium	-0.706*
芽单胞菌属Gemmatimonas	0.706*	油壶菌属Olpidium	-0.552
溶杆菌属Lysobacter	0.622*	光黑壳属Preussia	-0.503
慢生根瘤菌属Bradyrhizobium	0.613*	毛壳菌属Chaetomium	0.441
嗜盐囊菌属Haliangium	0.559	Tausonia	-0.441
Actinoplanes	0.553	被毛枝葡萄孢属Botryotrichum	0.434
芽球菌属Blastococcus	0.483	赤霉属Gibberella	0.378
RB41	-0.483	枝顶孢属Acremonium	-0.357
Flavisolibacter	0.445	被孢霉属Mortierella	0.343
Bryobacter	0.434	酵母属Remersonia	-0.252
Ellin6055	0.378	枝鼻菌属Cladorrhinum	0.224
Gaiella	0.371	腐质霉属Humicola	-0.196
嗜酸菌属Acidovorax	0.336	Plectosphaerella	0.182
Ellin6067	0.336	产油菌属Solicoccozyma	-0.154
Skermanella	0.231	闭小囊菌属Kernia	-0.147
芽孢杆菌属Bacillus	0.196	曲霉属Aspergillus	0.042
酸杆菌属Acidibacter	0.032	头束霉属Cephalotrichum	0.021

化学性质 Soil property	细菌 Bacteria (R)	真菌 Fungi (R)
EC	0.309*	0.464**
pH	-0.118	0.164
有效磷 Available P (AP)	-0.068	-0.164
有效钾 Available K (AK)	0.141	0.239
有机碳 SOC	-0.109	0.018
全碳 TC	0.07	-0.099
全氮 TN	-0.19	-0.162