• ISSN 1008-505X
  • CN 11-3996/S

蚯蚓联合生防细菌Bacillus velezensis改善连作百合土壤细菌群落结构及防治枯萎病的效果

鲁耀雄, 高鹏, 彭福元, 李卫东, 李静, 崔新卫, 黄国林, 潘素君, 王运生

鲁耀雄, 高鹏, 彭福元, 李卫东, 李静, 崔新卫, 黄国林, 潘素君, 王运生. 蚯蚓联合生防细菌Bacillus velezensis改善连作百合土壤细菌群落结构及防治枯萎病的效果[J]. 植物营养与肥料学报, 2024, 30(1): 147-159. DOI: 10.11674/zwyf.2023262
引用本文: 鲁耀雄, 高鹏, 彭福元, 李卫东, 李静, 崔新卫, 黄国林, 潘素君, 王运生. 蚯蚓联合生防细菌Bacillus velezensis改善连作百合土壤细菌群落结构及防治枯萎病的效果[J]. 植物营养与肥料学报, 2024, 30(1): 147-159. DOI: 10.11674/zwyf.2023262
LU Yao-xiong, GAO peng, PENG Fu-yuan, LI Wei-dong, LI Jing, CUI Xin-wei, HUANG Guo-lin, PAN Su-jun, WANG Yun-sheng. Earthworm combined with Bacillus velezensis improves bacterial community structure and the control effect of Fusarium wilt diseases of continuous cropping lily[J]. Journal of Plant Nutrition and Fertilizers, 2024, 30(1): 147-159. DOI: 10.11674/zwyf.2023262
Citation: LU Yao-xiong, GAO peng, PENG Fu-yuan, LI Wei-dong, LI Jing, CUI Xin-wei, HUANG Guo-lin, PAN Su-jun, WANG Yun-sheng. Earthworm combined with Bacillus velezensis improves bacterial community structure and the control effect of Fusarium wilt diseases of continuous cropping lily[J]. Journal of Plant Nutrition and Fertilizers, 2024, 30(1): 147-159. DOI: 10.11674/zwyf.2023262

蚯蚓联合生防细菌Bacillus velezensis改善连作百合土壤细菌群落结构及防治枯萎病的效果

基金项目: 湖南省重点研发计划项目(2021NK2008,2022NK2062);湖南省自然科学基金面上项目(2020JJ4393);长沙市自然科学基金项目(kq2208105)。
详细信息
    作者简介:

    鲁耀雄 E-mail: 37204824@qq.com

    通讯作者:

    潘素君 E-mail: sujunpan@126.com

    王运生 E-mail: wyunsheng@gmail.com

Earthworm combined with Bacillus velezensis improves bacterial community structure and the control effect of Fusarium wilt diseases of continuous cropping lily

  • 摘要:
    目的 

    从土壤理化性状和微生物群落结构及多样性角度,研究蚯蚓联合生防细菌改善百合连作障碍的机理和效果,为湖南百合产业化种植提供技术措施。

    方法 

    采用盆栽试验方法,在湖南长沙连续种植了两季百合 (Lilium lancifolium Thunb.)。供试土壤为红壤,生防菌为Bacillales velezensis YFB3-1菌液(菌体浓度为109 CFU/mL),蚯蚓为赤子爱胜蚓(Eisenia fetida)。试验设单施菌液(T1)、只接种蚯蚓(T2)、同时使用菌液和蚯蚓(T3) 3个处理,以不使用菌剂和蚯蚓为对照(CK)。调查了两茬百合产量和枯萎病病情指数,分析了第二茬百合根际土壤理化性质、细菌群落结构和多样性,以及连作百合产量与根际土壤理化性质、细菌群落和枯萎病的相关性。

    结果 

    与第一茬相比,同一处理的第二茬百合产量都显著下降。两年连作百合产量都以同时使用菌液和蚯蚓(T3)处理为最高,分别为16464、15674 kg/hm2,都显著高于同年份其他处理。两年连作百合枯萎病发病率和病情指数都以T3为最低,都显著低于同年份其他处理,T3处理百合枯萎病病情指数分别为18.15 (2019年)、25.00 (2020年),并且两年的防治效果 (T3) 都明显优于单施菌液 (T1) 和接种蚯蚓 (T2)。相比于对照 (CK), 2019、2020年T3处理百合枯萎病防治效果分别为44.55%、37.66%,分别增产15.05%、14.78%。蚯蚓联合生防菌YFB3-1增加了根际土壤Rhizobiales、Flavobacteriales、Pseudomonadales、Bacillales等有益微生物的种群丰度,蚯蚓与生防菌YFB3-1在抑制连作百合枯萎病方面具有显著的互作效应,因而缓解连作百合产量下降的效果最佳。

    结论 

    在施用芽孢杆菌生防菌剂防治连作百合枯萎病的同时,可以增施有机肥 (牛粪) 来提高土壤中蚯蚓数量,有效预防百合枯萎病发生,减少连作百合产量损失。

    Abstract:
    Objectives 

    We studied the effect and mechanism of earthworm and biocontrol bacteria strain on improving physiochemical properties, and microbial structure and diversity in rhizosphere soil of continuous cropping lily, to propose an effective measurement to alleviate the impaction of Fusarium wilt diseases on the industry of lily production in Hunan Province.

    Methods 

    A pot experiment was conducted in Changsha City, Hunan Province. The test crop lily (Lilium lancifolium Thunb.) was cultured in 2019 and 2020 continuously. The test bacteria strain was Bacillales velezensis YFB3-1 liquid (bacterial concentration ×109 CFU/mL), and the earthworm was Eisenia fetida. The four treatments included neither applying bacteria nor earthworm control (CK), merely applying bacteria liquid (T1), earthworm (T2), and the combination use of bacteria strain and earthworm (T3). The yield and Fusarium wilt diseases of lily were investigated. The rhizosphere soil were sampled after the harvest of the second lily crop for the determination of soil physiochemical properties, bacterial community structures and diversities.

    Results 

    Compared with the first crop, the yield of the second lily crop of the same treatment was significantly reduced. The highest yield of lily was all recorded in T3 treatment in 2019 and 2020 two years (16464 kg/hm2, and 15674 kg/hm2, respectively), which were significantly (P<0.05) higher than the other treatments in the same year. The diseases incidence and disease index was the lowest in T3 treatment (18.15 and 25.00, respectively) in the two years as well, which were significantly (P<0.05) lower than those in the other treatments in the same year. T3 treatment had stronger control effect on lily wilt diseases (P<0.05) than T1 and T2 treatment did in two years. The lily wilt diseases incidence in T3 treatment was 44.55% and 37.66% lower than in CK, and the lily yield was 15.05%, and 14.78% higher than in CK in 2019 and 2020. T3 treatment increased the population abundance of beneficial microorganisms such as Rhizobiales, Flavobacteriales, Pseudomonadales and Bacillales in the rhizosphere soil of lily, and the bacteria strain and earthworm showed significant interaction effects in inhibiting Fusarium wilt diseases of continuous cropping lily, thus alleviated the yield decline of continuous cropping lily.

    Conclusions 

    While using Bacillus biocontrol agent to control the continuous cropping lily wilt diseases, farmers should increase the number of earthworms by applying organic fertilizer (cow dung), for better preventing effect of the lily wilt diseases and less yield loss of continuous cropping lily.

  • 百合为单子叶植物亚纲百合科 (Liliaceae) 百合属 (Lilium) 的多年生草本植物[1],具有较高的药用、食用和观赏价值[23]。我国百合种植具有明显的区域特征,主要集中在湖南龙山和隆回、甘肃兰州、江西万载、江苏宜兴等地,这些地区的百合种植面积大,历史悠久,连作现象十分普遍[45],连作障碍问题严重,产量急剧下降。其主要原因是百合长期连作会改变土壤理化性质,存在化感物质自毒作用[6],定向选择根际微生物[78],导致土壤微生物群落结构失调[910],主要表现为细菌型土壤向真菌型土壤转变[11],特别是镰刀菌数量增多[1213]。多年连作百合的镰刀菌枯萎病发病率高达70%以上[14],是百合连作障碍中危害最大的一种真菌性土传病害[15],其中尖孢镰孢菌是其主要的病原微生物[16]。为了防止连作百合产量下降[17],种植户需要喷洒大量的农药进行防治,导致环境污染、生态失衡和农药残留超标,不利于产区百合种植及相关产业的可持续发展。

    蚯蚓是土壤动物的重要组成,种类多,数量大,分布广泛,每天能吞食大量土壤,有利于改善土壤理化性质和微生物群落结构,被称为“生态系统工程师”[18],在维持土壤生态系统的结构和功能方面占有重要地位[19]。蚯蚓偏好取食富含有机质的土壤,通过掘穴、取食、消化和排泄等活动,能够很好地将有机物和土壤混合[20],对土壤物理结构及养分循环都有显著的促进作用[19],影响了土壤微生物组成、丰富度和活性[21],激活整个土壤生态系统,抑制土传病害的发生与传播[22],为缓解作物的连作障碍问题提供一条新途径[23]。蚯蚓与微生物之间具有复杂相互关系[24],通过取食、肠道研磨和生物扰动等破坏真菌菌丝,减少真菌数量[25],从而减轻作物真菌病害的发生。同时,蚯蚓肠道的高湿、pH、C/N等被认为是最适宜细菌繁殖的生境,对细菌生长和繁殖有刺激作用,尤其是与蚯蚓存在互利共生的细菌会成为优势菌群[26]。代金君等[27]认为蚯蚓取食土壤消化后产生的蚓粪微生物群落结构发生变化,显著增加了细菌的群落多样性[28],细菌数量的增加比放线菌和真菌明显一些,并且微生物群落活性随着接种蚯蚓耕作年限的延长增加值提高[29]。蚯蚓粪中含有一些拮抗微生物,在一定程度上能够控制作物土传病害的发生[30],缓解土壤连作障碍的发生。蚯蚓还能够刺激微生物产生促生活性物质和抗生素等次生代谢物质,前者促进作物根系生长,提高了产量和品质,后者抑制真菌及病原菌的繁殖,减少植物土传病害的发生,提高了产量和品质[24]。然而,有关蚯蚓活动联合芽孢杆菌生防菌对百合连作土壤理化性质、根际微生物群落的影响和枯萎病的防治效果缺乏深入研究。

    本研究通过采集百合连作土壤进行连续两年的盆栽试验,研究喷洒生防菌菌液、接种蚯蚓、接种蚯蚓联合喷洒生防菌菌液等处理,对百合连作土壤理化性质、根际土壤细菌群落结构、百合枯萎病发生和产量的影响,分析百合产量与连作土壤理化性质、根际土壤细菌群落、枯萎病发生的相关关系,以期为连作百合高效生态栽培提供理论依据和技术支撑。

    供试百合品种为卷丹百合(L. lancifolium Thunb.),由湖南龙山绿叶百合农产品有限公司提供。蚯蚓品种为赤子爱胜蚓(Eisenia fetida)大平2号,来源于湖南省湘北蚯蚓养殖有限公司,每条重约0.40~0.45 g。Bacillales velezensis YFB3-1是湖南省农业环境生态研究所循环农业课题组,从蚯蚓肠道内含物中分离纯化的具有广谱抗百合病原真菌的一种生防细菌,保藏编号CCTCC NO:M20221140。YFB3-1生防菌经牛肉膏蛋白胨液体培养基30℃发酵48 h,菌体浓度为109 CFU/mL。

    供试土壤为红壤,取自湖南省农业科学院药用植物研究中心高桥基地百合连作两年的旱地土壤,连作百合表现出不同程度的枯萎变黄、死亡株数明显增多等连作障碍现象。土壤基本化学性质为:pH 5.42,有机质 20.6 g/kg,全氮1.72 g/kg,全磷0.91 g/kg,全钾21.2 g/kg,碱解氮89.7 mg/kg,有效磷77.7 mg/kg,速效钾92.5 mg/kg。

    牛粪来源于中国科学院亚热带农业生态研究所长沙农业环境观测研究站的牛场,其理化性质如下:pH 8.87,有机碳346 g/kg,全氮14.9 g/kg,P2O5 6.58 g/kg,K2O 9.76 g/kg,含水率76.4%,C/N 23.3。

    盆栽试验在湖南省农业环境生态研究所的网室进行,设4个处理:1) 不施菌剂对照(CK),百合种球覆盖土壤后,在两行之间表施牛粪22500 kg/hm2 (干基);2) 喷洒YFB3-1菌液 (T1),于9月28日百合种植后和翌年4月1日百合出苗后,分别喷洒YFB3-1菌液1次,每次菌液用量为30 L/hm2,兑水稀释10倍,喷洒在试验盆内土壤表面,其他施肥措施同CK;3) 接种蚯蚓 (T2),投放蚯蚓75 kg/hm2,约每株百合投放蚯蚓1条,其他施肥措施同CK;4) 接种蚯蚓联合喷洒YFB3-1菌液 (T3),菌液喷洒用量和方式同T1,投放蚯蚓用量和方式同T2,其他施肥措施同CK。每处理设3次重复,每重复有5盆,共计60盆,随机放置在平整好的试验地上。

    试验用盆为长方体,盆内长、宽、高分别为0.40、0.30 m、0.16 m,底部开有小孔,每盆装土量为20.88 kg,装土深度约为0.15 m,每盆栽种4株百合。盆中栽种百合的行距为0.25 m,间距为0.15 m,距盆边为0.075 m。每处理养分投入量按照N、P2O5、 K2O分别为450、375、600 kg/hm2计算,包括牛粪带入的养分量,折合N、P2O5、K2O量分别为10.0、8.3、13.3 g/盆,供试化肥包括尿素 (N 46.4%)、过磷酸钙 (P2O5 12%) 和硫酸钾 (K2O 50%),将化肥一次性撒施盆内,并与土壤混合均匀。百合种球消毒后播种,两行百合之间开宽×深为0.15 m×0.05 m的浅沟,将新鲜牛粪0.5 kg/盆表施于浅沟内。由于供试蚯蚓为表层蚯蚓,卷丹百合的茎根也生长在表层土壤,所以牛粪采用表施。为防止鲜牛粪过快干燥和杂草生长,在所有处理上面覆盖1层1~2 cm厚的稻草。

    卷丹百合为跨年生长采收的药食两用植物,第一茬于2018年9月26日种植,2019年3月中旬出苗,5月22日打花蕾,7月28日采收百合种球。2019年重复种植一茬,栽培和管理时间与2018年相同。

    在第二茬百合旺长期 (2020年5月10日),每重复随机抽取1盆,在两株百合间的中点位置,用环刀采集土壤样品,用于测定孔隙度、容重。

    然后,用小铲挖取每个盆中的百合4株,仔细挑出每盆的蚯蚓并统计好数量。轻微抖动百合茎根,轻取沾着其上的土作为根际土壤,四株百合茎根土壤制成一个混合根际土壤样品。

    每个根际土壤样品分成两份,一份自然风干后,常规法测定土壤理化性质[31]:土水比为1∶5浸提,PHS-3C型pH计测定pH,硫酸–重铬酸钾氧化法测定有机质,碱解扩散法测定碱解氮,碳酸氢钠提取—钼锑抗比色法测定有效磷,乙酸铵浸提−火焰光度法测定速效钾;另一份装入灭菌离心管中,−80℃超低温冰箱中速冻后,快递到上海美吉生物医药科技有限公司分析生物学性状。土壤总DNA采用E.Z.N.A.®Soil试剂盒 (Omega Bio-tek, Norcross, GA, U.S.) 提取,DNA浓度和纯度利用NanoDrop 2000检测,DNA提取质量利用1%琼脂糖凝胶电泳检测。采用序列为338F (5′-ACTCCTACGGGAGGCAGCAG-3′) 和806R (5′-GGACTACHVGGGTWTCTAAT-3′) 的引物对16S rRNA基因的V3~V4可变区片段进行PCR扩增,扩增程序为:95℃预变性3 min后,接着27个循环 (95℃变性30 s,55℃退火30 s,72℃延伸30 s),最后72℃延伸10 min (PCR仪:ABI GeneAmp® 9700型)。扩增体系总量为20 μL,含4 μL 5×FastPfu缓冲液,2 μL 2.5 mmol/L dNTPs,0.8 μL引物 (5 μmol/L),0.4 μL FastPfu聚合酶,10 ng DNA 模板。使用2%琼脂糖凝胶回收PCR产物,利用AxyPrep DNA Gel Extraction Kit (Axygen Biosciences, Union City, CA, USA) 进行纯化,Tris-HCl洗脱,2%琼脂糖电泳检测。利用QuantiFluor™-ST (Promega, USA) 进行检测定量。根据Illumina MiSeq平台 (Illumina, San Diego, USA) 标准操作规程用纯化后的扩增片段构建PE 2×300的文库。利用Illumina公司的Miseq PE300平台标准操作规程进行基因测序。

    在两茬百合成熟期(2019、2020年7月18日前后),调查百合枯萎病发病率和病情指数。枯萎病发病率和病情指数以株为单位,调查每个处理各级病株数。调查分级标准如下[32]:植株茎秆正常,全株无病叶,为0级;病株底部变黄或变紫叶片数不超过整株叶片数的25%,茎顶端变浅紫色,心叶向一侧轻度弯曲,为1级;病株底部叶片枯萎或枯萎叶片数占整株的25%~50%,茎上部变紫色且明显弯曲,为2级;病株枯萎叶片超过50%,茎中上部变紫色,且严重弯曲,为3级;全株叶片都枯萎或整株枯死,茎基部维管束变褐,为4级。

    发病率=(发病株数/总株数)×100%

    病情指数=[∑(各级病株数×相应病级数)]/(总株数×最高病级数)×100%

    相对防治效果=(对照病情指数−处理病情指数)/对照病情指数×100%

    于2019、2020年的7月28日前后收获百合并测定产量,同一处理每重复随机选择5株百合并称重。

    细菌群落结构采用测序后经过抽平处理的OTU数据描述。α多样性是指一个特定区域或者生态系统内的物种多样性,常用的度量标准有ace index、chao index、shannon index、coverage等,运用统计学检测每两组之间的指数值是否具有显著性差异。β多样性分析是对不同微生物群落间的物种多样性进行组间比较分析,主坐标分析 (principal co-ordinates analysis,PCoA) 是β多样性分析中的一种非约束性数据降维分析方法,可用来研究样本群落组成的相似性或差异性。菌群结构组成柱形图是根据分类学分析结果,得知不同样本在各分类水平上的物种组成情况,根据群落柱形图可以直观呈现各样本在某一分类学水平上含有哪些优势物种和各优势物种的相对丰度 (所占比重)。LEfSe (http://huttenhower.sph.harvard.edu/galaxy/root?tool_id=lefse_upload) 根据分类学组成对样本按照不同的分组条件进行线性判别分析 (linear discriminant analysis,LDA),找出对样本划分产生显著性差异影响的群落或物种。冗余分析(redundancy analysis,RDA) 基于线性模型在同一个二维排序图上反映样本菌群与环境因子之间关系。Mantel test在生态学上检验群落距离矩阵和环境变量距离矩阵之间的相关性。采用WPS Office 2020对试验数据进行记录、整理,采用SPSS 18.0对数据进行相关统计分析,利用最小显著差异法 (least significant difference,LSD) 进行多重比较。百合产量、枯萎病发病率和病情指数采用WPS Office 2020作图,细菌的菌群结构组成柱形图、PCoA、RDA采用R语言 (version 3.3.1) 统计分析和作图。利用R语言 (version 3.3.1),用“plspm”包进行偏最小二乘法路径回归分析 (PLS-PM)。

    图1可以看出,同一处理2020年的百合产量均低于2019年,说明随着连作年限的增加不同处理都具有减产的现象。与CK相比,T1、T2、T3处理2019和2020年百合产量均显著增加,且增加幅度均为T3>T2>T1 (P<0.05);在所有处理中,2019年T3的百合产量最高,为16464 kg/hm2,显著高于其他处理,其次是2020年的T3处理,为15674 kg/hm2,2019年的T3处理比同年的CK提高了15.05%,2020年的T3处理比同年的CK处理提高了14.78%;2020年T3处理的产量与2019年的T2处理差异不显著,但是两者的产量都显著低于2019年的T3处理,而显著高于其他处理;2020年T1处理的产量与2019年的CK差异不显著,都显著高于2020年的CK。表明菌剂和蚯蚓对百合产量都有显著提升效果,二者联合使用可进一步显著提升其增产效果。

    图  1  蚯蚓和菌剂对百合产量、枯萎病发病率和病情指数的影响
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。数据为平均值±标准误差 (n=3)。柱上不同小写字母表示处理间差异显著 (P<0.05)。
    Figure  1.  Yield, morbidity and disease index of Fusarium wilt of lilies as affected by earthworm and bacteria strain treatments
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. Values are means±SE (n=3). Different lowercase letters above the bars indicate significant differences among treatments (P<0.05).

    对比2019和2020年百合枯萎病发病率和病情指数可以看出,在同一处理中2020年的连作百合枯萎病发病率和病情指数都高于2019年,说明随着连作年限的增加不同处理的百合枯萎病发病率和病情指数都增大。2019年T3处理的百合枯萎病发病率和病情指数都是最低的,分别为37.5%、18.15,都显著低于其他处理;2020年T3处理的百合枯萎病发病率和病情指数分别为45.83%、25.00,其发病率与2019年的T2处理差异不显著,显著高于2019年的T3处理,显著低于其他处理,其病情指数与2019年的T1、T2处理差异不显著,显著高于2019年的T3处理,显著低于其他处理;与同年CK处理相比,T3处理的枯萎病防治效果分别为44.55% (2019年)和37.66% (2020年)。同一年份中,T3处理百合发病率和病情指数均最低,且显著低于其他处理;T2处理百合发病率和病情指数也显著低于T1和CK处理,而2020年T1处理与CK的发病率无显著差异。表明接种蚯蚓可以显著降低百合发病率和发病指数,蚯蚓与菌剂联合使用可进一步显著增加抗病效果。

    表1显示,T3处理的蚯蚓数量为127.78条/m2,T2处理为119.44条/m2,T3处理的蚯蚓数量与T2处理差异不显著。土壤pH、有机质含量和总孔隙度都以T3或T2处理最大,T3处理的土壤pH、有机质含量和总孔隙度与T2处理差异不显著,但是都显著高于T1和CK处理,而T3和T2处理的土壤容重都显著小于T1和CK处理,表明蚯蚓活动可以缓解连作百合土壤的酸化,显著增加土壤的有机质含量和总孔隙度,显著降低土壤容重。

    表  1  蚯蚓和菌剂对连作百合土壤理化性质的影响
    Table  1.  Soil physicochemical properties of continuous cropping lilies as affected by earthworm and bacteria strain treatments
    处理
    Treatment
    pH 有机质
    Organic matter
    (g/kg)
    总孔隙度
    Total porosity
    (%)
    容重
    Bulk density
    (g/cm3)
    碱解氮
    Available N
    (mg/kg)
    有效磷
    Available P
    (mg/kg)
    速效钾
    Available K
    (mg/kg)
    蚯蚓数量
    Earthworm number
    (PCS/m2)
    CK 5.66±0.08 b 23.03±0.24 b 51.44±0.57 b 1.13±0.02 a 113.56±6.57 c 92.82±2.86 c 121.36±6.57 c
    T1 5.69±0.09 b 22.93±0.45 b 50.92±1.44 b 1.13±0.01 a 117.85±6.21 c 105.42±5.95 bc 114.27±9.35 c
    T2 6.34±0.06 a 25.84±0.66 a 53.60±1.12 a 1.01±0.02 b 136.07±3.95 b 117.85±6.13 b 155.49±7.05 b 119.44±50.92 a
    T3 6.33±0.10 a 25.85±0.42 a 53.89±1.57 a 1.00±0.02 b 154.81±9.03 a 135.87±6.45 a 176.50±9.63 a 127.78±29.27 a
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。数据为平均值±标准误差 (n=3)。同列数据后不同小写字母表示处理间差异显著 (P<0.05)。
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. Values are means ± SE (n=3). Different lowercase letters after data in the same column indicate significant differences among treatments (P<0.05).
    下载: 导出CSV 
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    土壤碱解氮、有效磷和速效钾含量以T3处理最大,都显著高于其他处理,其次分别都是T2处理,T2处理的碱解氮和速效钾含量都显著高于T1和CK处理,表明蚯蚓活动可以增加土壤碱解氮、有效磷和速效钾等有效养分含量,蚯蚓与菌剂联合使用可进一步显著增加土壤有效养分含量。

    表2可知,4个处理的覆盖率没有显著差异,范围为98.22%~98.49%,说明各处理的样本OTU覆盖度已经饱和,未被检到基因序列的概率很低,本次测序结果能够代表不同处理方式下连作百合茎根土壤细菌群落的真实情况。

    表  2  不同处理的连作百合根际土壤微生物α多样性
    Table  2.  Microbial α diversity in rhizosphere soil of continuous cropping lilies in different treatments
    处理
    Treatment
    Ace指数
    Ace index
    (×102)
    Chao1指数
    Chao1 index
    (×102)
    Shannon指数
    Shannon
    index
    覆盖率
    Coverage
    (%)
    CK 16.40±1.86 a 16.46±1.63 a 5.65±0.20 b 98.49±0.15 a
    T1 18.89±1.92 a 18.89±2.00 a 5.84±0.19 ab 98.22±0.22 a
    T2 16.72±1.81 a 16.87±1.63 a 5.73±0.12 ab 98.42±0.21 a
    T3 19.27±0.63 a 19.17±0.39 a 6.03±0.08 a 98.23±0.09 a
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。数据为平均值±标准误差 (n=3)。同列数据后不同小写字母表示处理间差异显著 (P<0.05)。
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. Values are means ± SE (n = 3). Different lowercase letters after data in the same column indicate significant differences among treatments (P<0.05).
    下载: 导出CSV 
    | 显示表格

    T1、T2处理的Ace、Chao1和Shannon 指数与CK均无显著差异。T3处理的Ace、Chao1指数与其他处理没有显著差异,但Shannon指数显著高于CK。

    在OTU水平对不同处理连作百合茎根土壤细菌群落进行PCoA分析(图2),其中PC1的贡献率为42.31%,PC2的贡献率为18.58%,二者累积贡献率为60.89%。相同处理的样品明显聚集,不同处理的样品明显分离,说明相同处理样品的细菌群落结构差异较小,不同处理样品的细菌群落结构差异较大。

    图  2  不同处理的百合根际土壤细菌群落的PCoA分析
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。
    Figure  2.  PCoA of bacterial community in rhizosphere soil of continuous cropping lilies in different treatments
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution.

    在目水平上,共获得202个类群。将平均相对丰度<1%类群归类为其他,得到34个类群 (图3)。其中Rhizobiales (14.97%~21.12%)、Clostridiales (4.62%~7.09%)、Myxococcales (3.81%~9.44%)、Micrococcales (3.59%~5.85%)、Betaproteobacteriales (3.59%~5.24%)、Caulobacterales (2.46%~5.69%) 为5个不同处理条件下连作百合根际土壤的前6个优势菌目,他们相对丰度占总序列的39.37%~44.07%。T1处理在目水平丰度相对含量最高的类群有Rhizobiales (21.12%)、Chitinophagales (4.04%)、Microtrichales (4.15%)、Saccharimonadales (3.95%);T2处理在目水平的丰度相对含量最高的类群有Micrococcales (5.85%)、Caulobacterales (5.69%);T3处理在目水平的丰度相对含量最高的类群有Flavobacteriales (6.05%)。

    图  3  不同处理土壤细菌目水平相对丰度(前33个细菌目)
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。Others指相对丰度小于0.01者。
    Figure  3.  The relative abundances of the top 33 bacterial order present in different trentment soils
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. The relative abundance of others is less than 0.01.

    基于效应大小线性判别分析 (linear discriminant analysis effect size,LEfSe)方法,分析了不同处理的茎根土壤细菌在目水平有显著性差异的标志性物种 (biomarker,图4)。设置线性判别分析 (linear discriminant analysis,LDA,LDA≥4.0),4个处理共筛选出21个Biomarkers,T1处理在科水平Devosiaceae、Chitinophagaceae和属水平Devosia 3类物种相对丰度显著高于其它处理;T2处理在门水平Patescibacteria,科水平Hyhpomonadaceae、Flavobacteriaceae 3类物种相对丰度显著高于其它处理;T3处理在科水平Flavobacteriaceae、Pseudomonadaceae、Xanthobateraceae,目水平Flavobacteriales、Pseudomonadales,属水平Flavobacterium、Pseudomonas 7类物种相对丰度显著高于其它处理。并且选择B. velezensis YFB3-1所在的Bacillales进行对比分析,T3处理中Bacillales的丰度相对含量最大,其次分别为T2、T1处理,最低为CK处理,T3处理中Bacillales的丰度相对含量与T2、T1处理差异不显著,但是显著高于CK处理。

    图  4  基于LEfSe分析不同处理具有显著性差异的物种 (LDA≥4.0)
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。CK、T1、T2和T3分别以红色、蓝色、绿色、粉红色表示;线性判别分析的阈值设置为≥4.0。
    Figure  4.  LEfSe analysis on species with significant difference in different treatments (LDA≥4.0)
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. CK, T1, T2 and T3 treatments are shown in red, blue, green, and pink, respectively. The threshold of linear discriminant analysis (LDA) is set to ≥4.0.

    不同处理下连作百合根际土壤细菌目水平细菌相对丰度与环境因子的RDA见图5,RDA1的解释量为32.00%,RDA2的解释量为19.02%,两者累计解释量为51.02%。同一处理的样点相对聚集,不同处理的样点明显分开,该数据结果与基于PCoA的分析结果相似。此外,基于RDA分析发现不同的环境因子对连作百合土壤细菌群落特征具有不同影响程度(解释量),本研究中各环境因子的影响从大到小依次为:pH>有机质、有效磷>容重、总孔隙度>速效钾、碱解氮,其中pH、有机质、有效磷、容重、总孔隙度是影响连作百合土壤细菌群落特征的主要因子。基于Mantel test的不同处理方式的连作百合根际土壤细菌目水平上相对丰度与环境变量的斯皮尔曼相关性分析见表3。环境变量pH、有机质、容重、碱解氮、有效磷、速效钾与细菌目水平丰度存在极显著(P<0.01)相关关系,但环境变量总孔隙度与细菌目水平丰度变化无显著相关关系。由此可见,本研究中所选的环境变量中,pH、有机质、容重、碱解氮、有效磷、速效钾五个变量主要调控连作百合根际土壤细菌目水平的群落结构,这与RDA分析结果略有区别。因此,综合RDA和Mantel test的分析结果,pH、有机质、容重、有效磷为影响连作百合根际土壤细菌目水平群落结构的关键因子。

    图  5  不同处理目水平细菌相对丰度与环境因子的冗余分析
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。OM—有机质;TP—总孔隙度;BD—容重;AN—碱解氮;AP—有效磷;AK—速效钾。图中红色箭头表示数量型环境因子,箭头长短代表环境因子对细菌群落结构影响程度 (解释量) 的大小。
    Figure  5.  RDA analysis on the relative abundance of bacterial order and environmental factors in different treatments
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. OM—Organic matter; TP—Total porosity; BD—Bulk density; AN—Available N; AP—Available P; AK—Available K. The red arrows represent environmental factors, and the arrow length represent the degree of influence on the species data (the amount of interpretation).
    表  3  基于Mantel test的细菌目水平相对丰度与环境变量的相关性分析
    Table  3.  The Spearman’s correlations (r) between the relative abundances of bacterial order and the environmental variables determined by Mantel test
    环境因子 Environmental factor r P
    pH 0.5009 0.003
    土壤有机质 Soil organic matter 0.4324 0.006
    总孔隙度 Total porosity 0.0351 0.841
    容重 Bulk density 0.4117 0.006
    碱解氮 Available N 0.5155 0.005
    有效磷 Available P 0.7320 0.001
    速效钾 Available K 0.4004 0.008
    下载: 导出CSV 
    | 显示表格

    在蚯蚓与YFB3-1互作条件下,为了更好的理解蚯蚓、芽孢杆菌、土壤理化指标、细菌群落、镰刀菌枯萎病以及产量的相互关系,将上述归类的环境变量指标进行降维处理,选择每一类环境变量指标主成分分析 (principal component analysis,PCA) 的第一个成分表示来构建偏最小二乘路径模型 (PLS-PM,图6)。结果显示,蚯蚓 (earthworms)与芽孢杆菌丰度 (Bacillales)、细菌群落 (bacterial community) 正相关,与土壤理化性质 (soil properties,r=0.90,P<0.001) 具有极显著正相关;土壤理化性质与连作百合产量正相关,与细菌群落 (r=0.56,P<0.05) 显著正相关;细菌群落与连作百合镰刀菌枯萎病 (fusarium wilt diseases,r=−0.82,P<0.001) 呈极显著负相关;连作百合镰刀菌枯萎病与产量负相关。结果表明在蚯蚓和生防菌YFB3-1互作条件下,通过改善土壤理化性质,直接和间接作用于细菌群落结构来影响连作百合镰刀菌枯萎病发生,缓解产量下降,其中蚯蚓大于芽孢杆菌的作用效果,该模型的拟合度较好,不同环境变量对连作百合产量解释率为80% (R2=0.8)。

    图  6  蚯蚓与YFB3-1互作条件下连作百合产量与土壤理化性质、细菌群落和镰刀菌枯萎病的最小二乘路径模型分析
    注:OM—有机质;TP—总孔隙度;BD—容重;AN—碱解氮;AP—有效磷;AK—速效钾;IR—发病率;DI—病情指数. 图中箭头的宽度表示标准化路径系数的强度。实线表示正路径系数,虚线表示负路径系数,R2值表示每个内生变量解释量的比例。
    Figure  6.  Directed graph of the partial least squares path model (PLS-PM) of continuous cropping lilies yield, soil physical and chemical properties, bacterial community, and Fusarium wilt disease under interaction between earthworms and YFB3-1
    Note: OM—Organic matter; TP—Total porosity; BD—Bulk density; AN—Available N; AP—Available P; AK—Available K; IR—Incidence rate; DI—Disease index. The width of the arrows indicates the strength of the standardized path coefficient. The solid lines indicate positive path coefficients and dashed lines indicate negative path coefficients, R2 values represent the proportion of the variance explained for each endogenous variable.

    蚯蚓被称为“生态系统工程师”,蚯蚓活动改良土壤理化性质,促进有机质转化和团聚体形成,提高土壤养分含量和有效性[33],促进土壤养分循环[34],有利于作物养分吸收和生长发育。百合的根系分球根和茎根,球根生长在地表以下大约8~20 cm,茎根根层较浅,生长在地表以下大约0~8 cm,茎根根际土壤层刚好是接种的赤子爱胜蚓—大平2号(表层蚯蚓)活动区域,蚯蚓活动促进了有机质(表施牛粪转化的蚯蚓粪)的扩散,相比未接种蚯蚓的CK和T1处理,接种蚯蚓的T2和T3处理提高了土壤有机质含量,增加了土壤孔隙度,调节了土壤微生物群落结构,提高了土壤碱解氮、有效磷和速效钾的含量,有利于养分的吸收利用,促进连作百合的生长,增强其对病害侵染的抗逆性,提高产量。

    长期连作通常会引起土壤酸化,进而对土壤微生物群落结构产生很大影响。酸化土壤中细菌多样性的减少和真菌的增加会促进土传病原菌的增殖和作物病害的发生[35],Weyman-Kaczmarkowa等[36]报道,当土壤pH从4.5增加到7.0,真菌的数量和生物量分别减少了50%和42%。因此,缓解土壤酸化有利于减少连作百合枯萎病的发生。Acidobacteriales与枯萎病发病率显著正相关[37]Acidobacteriales会引起土壤的pH变化[38],大量研究[3941]表明,旱地连作土壤中酸杆菌处在优势菌群的前列,蚯蚓活动增强了百合根际土壤的通气性,是其前10优势细菌目没有厌氧菌Acidobacteriales的原因,从而调节土壤的pH。同时,蚯蚓活动促进有机质的转化和扩散,提高了土壤有机质含量,也起到缓解土壤酸化的作用,所以接种蚯蚓的T2和T3处理土壤pH高于未接种蚯蚓的对照和T1处理。此外,蚯蚓分泌物包括碳酸钙物质和氮物质,也可以提高酸性土壤的pH,缓解了土壤酸化[4243]。在施用牛粪有机质后接种蚯蚓提高了百合根际土壤有机质含量,有利于调节连作百合根际土壤的pH,降低土壤容重,增加孔隙度,改善土壤结构。相比单独接种蚯蚓(T2)或喷洒YFB3-1菌剂(T1),蚯蚓活动联合YFB3-1进一步优化微生物群落结构,增加了影响pH和有效磷相关微生物的丰度,提高土壤的有效养分含量,因此,在接种蚯蚓联合喷洒YFB3-1菌剂的条件下,pH、有机质、容重、有效磷成为影响连作百合根际土壤细菌目水平群落结构的关键因子。

    百合枯萎病是由尖孢镰孢菌(F. oxysporum)、串珠镰孢菌(F. moniliforme)和茄腐皮镰孢菌(F. solani)等复合侵染引起的一种真菌性病害,是百合栽培过程中危害最大的一种土传病害[15],随着种植年限增加,连作百合的枯萎病、病情指数都是逐年增大,而连作百合产量则逐年减少。

    Liu等[44]研究表明,非豆科类的百合与Rhizobiales存在共生关系,使Rhizobiales成为百合茎根和球根的优势细菌目,并且蚯蚓活动有利于Rhizobiales的传播和繁殖[45],增加了Rhizobiales丰度[4],百合根际土壤Rhizobiales相对丰度的增加起到了固氮作用,增加植物的氮素营养,促进百合的生长。接种蚯蚓联合喷洒生防菌YFB3-1菌剂(T3)的根际土壤,Flavobacteriales、Pseudomonadales相对丰度显著高于单独接种蚯蚓(T2)或喷洒生防菌YFB3-1菌剂(T1)等其它处理,Flavobacteriales、Pseudomonadales又有利于土壤的磷活化[46],说明蚯蚓活动与YFB3-1互作更有利于促进根际活磷微生物的繁殖,提高连作百合土壤的有效磷含量,增加土壤肥力,促进百合生长健壮增强抗逆性。林威鹏等[37]研究发现Pseudomonadales与枯萎病发病率呈显著负相关关系,主要是Pseudomonadales能够产生抗菌次级代谢产物,可以有效抑制土传病害的发生[47],从而减少连作百合枯萎病的发生。接种蚯蚓处理(T2和T3)的Bacillales数量高于对照(CK),说明蚯蚓活动增加了Bacillales的数量,而蚯蚓活动结合喷洒YFB3-1处理(T3)的Bacillales丰度最大,高于单独接种蚯蚓处理(T2)或喷洒YFB3-1菌剂处理(T1),说明蚯蚓活动联合喷洒B. velezensis YFB3-1更加促进了Bacillales的繁殖,二者存在互作关系。芽孢杆菌类群具有分布广泛、抗逆性强、对多种病原菌的拮抗效果好、易于在植物根际进行有效定殖等优势,可以预防多种作物的土传病害,而且还具有较强的促生作用[48],朱菲莹等[49]通过Spearman分析发现,在Bacillales存在下芽孢杆菌属与西瓜枯萎病发生率呈负相关。B. velezensisBacillales中的新种,能够产生抑菌次级代谢产物[5051],对百合多种土传病原真菌具有较强的抑制效果,从而降低连作百合枯萎病等土传病害的发病率。因此,相比对照处理,单独接种蚯蚓和喷洒B. velezensis YFB3-1菌剂或者两者联合施用,都可以提高连作百合产量,并且联合作用下的百合生长最好,枯萎病发病率和病情指数最低,产量最高,说明蚯蚓活动联合YFB3-1对防治连作百合枯萎病存在较好的互作关系。

    蚯蚓活动能够影响土壤微生物结构,调节土壤微生物群落丰富度和多样性[21,52]。Ace指数、Chao1指数用于表征物种丰富度,数值越大,物种的丰富度就越高;Shannon指数用来表征生物多样性,它的数值越大,生物多样性越高。相比单独接种蚯蚓或喷洒YFB3-1菌剂,接种蚯蚓联合喷洒生防菌YFB3-1(T3)处理的Ace指数、Chao1指数和Shannon指数最大,说明蚯蚓活动联合生防菌YFB3-1更有利于提高连作百合根际土壤细菌群落结构物种的丰富度和多样性,从而改善土壤的微生态环境,尤其在真菌性土壤向细菌性土壤转变过程中发挥较好的作用。同时,蚯蚓活动联合YFB3-1菌剂进一步优化连作百合的细菌群落结构,增加了土壤有益微生物的群落丰度(如Rhizobiales、Flavobacteriales、PseudomonadalesBacillales),相比单独接种蚯蚓或喷洒YFB3-1菌剂,两者的互作有利于进一步活化土壤的碱解氮、有效磷和速效钾等养分,从而促进百合的养分吸收利用,增强连作百合抗逆性,减少百合枯萎病的发生,减少连作百合的产量损失,达到缓解百合连作障碍的目的。然而蚯蚓的采食和挖掘等活动对生防菌YFB3-1的扩繁、定殖以及其抗病机理等,还需要进一步采用荧光标记和代谢组等手段进行定量分析和深入研究。

    接种蚯蚓显著改善了土壤结构,蚯蚓活动联合B. velezensis YFB3-1进一步优化了微生物群落结构,提高了土壤有效养分含量,因此,两者协作显著降低了连作百合枯萎病的发病率和病情指数,增加了百合产量。同时接种蚯蚓与YFB3-1菌剂对优化微生物群落结构丰富度和多样性,增加根际土壤中Flavobacteriales、Pseudomonadales、Bacillales等有益微生物的种群丰度,抑制连作百合枯萎病具有显著的互作效应。其中FlavobacterialesPseudomonadales有利于土壤磷活化,促进百合健壮增强抗逆性,PseudomonadalesBacillales产生的抗菌次级代谢产物,能够有效抑制镰刀菌等病原真菌的生长。因此,种植户在喷洒芽孢杆菌生防菌剂防治连作百合枯萎病的同时,可以增施有机肥(牛粪)来提高土壤蚯蚓数量,强化蚯蚓与芽孢杆菌生防菌的互作关系,有效地预防百合枯萎病等土传病害的发生,减少连作百合产量损失。

  • 图  1   蚯蚓和菌剂对百合产量、枯萎病发病率和病情指数的影响

    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。数据为平均值±标准误差 (n=3)。柱上不同小写字母表示处理间差异显著 (P<0.05)。

    Figure  1.   Yield, morbidity and disease index of Fusarium wilt of lilies as affected by earthworm and bacteria strain treatments

    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. Values are means±SE (n=3). Different lowercase letters above the bars indicate significant differences among treatments (P<0.05).

    图  2   不同处理的百合根际土壤细菌群落的PCoA分析

    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。

    Figure  2.   PCoA of bacterial community in rhizosphere soil of continuous cropping lilies in different treatments

    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution.

    图  3   不同处理土壤细菌目水平相对丰度(前33个细菌目)

    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。Others指相对丰度小于0.01者。

    Figure  3.   The relative abundances of the top 33 bacterial order present in different trentment soils

    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. The relative abundance of others is less than 0.01.

    图  4   基于LEfSe分析不同处理具有显著性差异的物种 (LDA≥4.0)

    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。CK、T1、T2和T3分别以红色、蓝色、绿色、粉红色表示;线性判别分析的阈值设置为≥4.0。

    Figure  4.   LEfSe analysis on species with significant difference in different treatments (LDA≥4.0)

    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. CK, T1, T2 and T3 treatments are shown in red, blue, green, and pink, respectively. The threshold of linear discriminant analysis (LDA) is set to ≥4.0.

    图  5   不同处理目水平细菌相对丰度与环境因子的冗余分析

    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。OM—有机质;TP—总孔隙度;BD—容重;AN—碱解氮;AP—有效磷;AK—速效钾。图中红色箭头表示数量型环境因子,箭头长短代表环境因子对细菌群落结构影响程度 (解释量) 的大小。

    Figure  5.   RDA analysis on the relative abundance of bacterial order and environmental factors in different treatments

    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. OM—Organic matter; TP—Total porosity; BD—Bulk density; AN—Available N; AP—Available P; AK—Available K. The red arrows represent environmental factors, and the arrow length represent the degree of influence on the species data (the amount of interpretation).

    图  6   蚯蚓与YFB3-1互作条件下连作百合产量与土壤理化性质、细菌群落和镰刀菌枯萎病的最小二乘路径模型分析

    注:OM—有机质;TP—总孔隙度;BD—容重;AN—碱解氮;AP—有效磷;AK—速效钾;IR—发病率;DI—病情指数. 图中箭头的宽度表示标准化路径系数的强度。实线表示正路径系数,虚线表示负路径系数,R2值表示每个内生变量解释量的比例。

    Figure  6.   Directed graph of the partial least squares path model (PLS-PM) of continuous cropping lilies yield, soil physical and chemical properties, bacterial community, and Fusarium wilt disease under interaction between earthworms and YFB3-1

    Note: OM—Organic matter; TP—Total porosity; BD—Bulk density; AN—Available N; AP—Available P; AK—Available K; IR—Incidence rate; DI—Disease index. The width of the arrows indicates the strength of the standardized path coefficient. The solid lines indicate positive path coefficients and dashed lines indicate negative path coefficients, R2 values represent the proportion of the variance explained for each endogenous variable.

    表  1   蚯蚓和菌剂对连作百合土壤理化性质的影响

    Table  1   Soil physicochemical properties of continuous cropping lilies as affected by earthworm and bacteria strain treatments

    处理
    Treatment
    pH 有机质
    Organic matter
    (g/kg)
    总孔隙度
    Total porosity
    (%)
    容重
    Bulk density
    (g/cm3)
    碱解氮
    Available N
    (mg/kg)
    有效磷
    Available P
    (mg/kg)
    速效钾
    Available K
    (mg/kg)
    蚯蚓数量
    Earthworm number
    (PCS/m2)
    CK 5.66±0.08 b 23.03±0.24 b 51.44±0.57 b 1.13±0.02 a 113.56±6.57 c 92.82±2.86 c 121.36±6.57 c
    T1 5.69±0.09 b 22.93±0.45 b 50.92±1.44 b 1.13±0.01 a 117.85±6.21 c 105.42±5.95 bc 114.27±9.35 c
    T2 6.34±0.06 a 25.84±0.66 a 53.60±1.12 a 1.01±0.02 b 136.07±3.95 b 117.85±6.13 b 155.49±7.05 b 119.44±50.92 a
    T3 6.33±0.10 a 25.85±0.42 a 53.89±1.57 a 1.00±0.02 b 154.81±9.03 a 135.87±6.45 a 176.50±9.63 a 127.78±29.27 a
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。数据为平均值±标准误差 (n=3)。同列数据后不同小写字母表示处理间差异显著 (P<0.05)。
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. Values are means ± SE (n=3). Different lowercase letters after data in the same column indicate significant differences among treatments (P<0.05).
    下载: 导出CSV

    表  2   不同处理的连作百合根际土壤微生物α多样性

    Table  2   Microbial α diversity in rhizosphere soil of continuous cropping lilies in different treatments

    处理
    Treatment
    Ace指数
    Ace index
    (×102)
    Chao1指数
    Chao1 index
    (×102)
    Shannon指数
    Shannon
    index
    覆盖率
    Coverage
    (%)
    CK 16.40±1.86 a 16.46±1.63 a 5.65±0.20 b 98.49±0.15 a
    T1 18.89±1.92 a 18.89±2.00 a 5.84±0.19 ab 98.22±0.22 a
    T2 16.72±1.81 a 16.87±1.63 a 5.73±0.12 ab 98.42±0.21 a
    T3 19.27±0.63 a 19.17±0.39 a 6.03±0.08 a 98.23±0.09 a
    注:CK—无菌剂和蚯蚓接种对照;T1—喷洒YFB3-1菌液;T2—接种蚯蚓;T3—接种蚯蚓联合喷洒YFB3-1菌液。数据为平均值±标准误差 (n=3)。同列数据后不同小写字母表示处理间差异显著 (P<0.05)。
    Note: CK—No bacteria and earthworm inoculation control; T1—Spray with YFB3-1 bacteria solution; T2—Inoculation with earthworms; T3—Earthworm inoculation and spray with YFB3-1 bacteria solution. Values are means ± SE (n = 3). Different lowercase letters after data in the same column indicate significant differences among treatments (P<0.05).
    下载: 导出CSV

    表  3   基于Mantel test的细菌目水平相对丰度与环境变量的相关性分析

    Table  3   The Spearman’s correlations (r) between the relative abundances of bacterial order and the environmental variables determined by Mantel test

    环境因子 Environmental factor r P
    pH 0.5009 0.003
    土壤有机质 Soil organic matter 0.4324 0.006
    总孔隙度 Total porosity 0.0351 0.841
    容重 Bulk density 0.4117 0.006
    碱解氮 Available N 0.5155 0.005
    有效磷 Available P 0.7320 0.001
    速效钾 Available K 0.4004 0.008
    下载: 导出CSV
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  • 收稿日期:  2023-06-13
  • 录用日期:  2023-08-28
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