CN112812994B - 一种能够防治丹参根腐病的铜绿假单胞菌及其应用 - Google Patents
一种能够防治丹参根腐病的铜绿假单胞菌及其应用 Download PDFInfo
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Abstract
一种能够防治丹参根腐病的铜绿假单胞菌,该菌株编号为铜绿假单胞菌(Pseudomonas aeruginosa)Pa2‑2,已保藏于已保藏于“中国典型培养物保藏中心”(简称CCTCC),保藏地址:湖北省武汉市,洪山区八一路,武汉大学山国典型培养物保藏中心,登记入册的保藏号为CCTCC M 2020928,保藏日期为2020年12月21日。本发明的铜绿假单胞菌对丹参根腐病病原菌均具有良好的抑制作用,对丹参根腐病病害的防效能达到40%以上,能够有效降低丹参根腐病的发病率,其效果优于现有的常用化学农药。本发明筛选的铜绿假单胞菌使用后不会产生抗药性、农药残留、环境污染等化学农药常见问题,对人畜健康、环境友好,符合人们对生态农业的需求。
Description
技术领域
本发明涉及一种能够防治丹参根腐病的铜绿假单胞菌及其应用,属于微生物技术领域。
背景技术
丹参Salvia miltiorrhizaBge为唇形科多年生草本,以干燥根和根茎入药,是治疗心脑血管疾病的首选药物之一。近年来,种植丹参已经成为河南省豫西贫困山区脱贫攻坚和乡村振兴的重要举措。为了保证丹参的产量,多数药材种植户在丹参的种植过程中广泛使用化学肥料和化学药剂,一定程度上降低了丹参的品质,严重情况下还会出现肥害和药害,甚至导致化学制剂残留超标,同时对种植区的生态环境也造成了污染。利用有益拮抗菌开展生物防治和土壤调控,是当前倍受关注的研究热点。目前,主要依靠微生物这种生物方式来防治植物病害,包括细菌、真菌和放线菌等。丹参根腐病是丹参生产中最重要的病害,严重降低了丹参的产量。研究发现,对丹参威胁最大的根腐病病原菌主要包括镰刀属的层出镰刀菌(Fusarium proliferatum)、腐皮镰刀菌(F.solani)、尖孢镰刀菌(F.oxysporum) 及细链格孢(Alternaria tenuissima)。生物防治在丹参根腐病病害控制方面已经受到学者的重视,有关丹参根腐病病原菌F.solani的生防菌筛选及其菌剂和应用已被广泛研究,已公开生防菌株主要有枯草芽孢杆菌、解淀粉芽孢杆菌、木霉等几类,而对病原菌F.proliferatum、F.oxysporum及链格孢属Alternaria sp有效的生防菌较少,因此筛选对丹参根腐病不同病原菌同时有效的生防菌株具有重要的理论和实践意义。
发明内容
针对现有技术的不足,本发明的目的是提供一种能够防治丹参根腐病的铜绿假单胞菌及其应用,以及该菌种的筛选方法和应用,该铜绿假单胞菌能够防治丹参根腐病。
本发明未解决上述技术问题所采用的技术方案是:
一种能够防治丹参根腐病的铜绿假单胞菌,该菌株编号为铜绿假单胞菌(Pseudomonas aeruginosa)Pa2-2,已保藏于已保藏于“中国典型培养物保藏中心”(简称CCTCC),保藏地址:湖北省武汉市,洪山区八一路,武汉大学山国典型培养物保藏中心,登记入册的保藏号为CCTCC M 2020928,保藏日期为2020年 12月21日。
所述的一种能够防治丹参根腐病的铜绿假单胞菌,其筛选方法为:在丹参田采集丹参根际土壤,称取1g土壤,加入10ml无菌水中充分振荡获得土壤悬浮液,然后将土壤悬浮液摇匀后进行稀释,取0.2ml稀释液涂布于LB固体平板上,培养,至菌落长出后分离纯化,然后利用平板拮抗试验筛选出对丹参根腐病具有抑制效果的菌株,并鉴定,得到铜绿假单胞菌Pa2-2。
所述的一种能够防治丹参根腐病的铜绿假单胞菌,其在防治丹参根腐病方面的应用。
所述的一种能够防治丹参根腐病的铜绿假单胞菌,其在制备防治丹参根腐病的菌剂中的应用。
所述的菌剂的制备方法为:将铜绿假单胞菌Pa2-2接种于LB培养基中,在 28℃、180rpm条件下振荡培养12h制备种子菌,种子菌以2%的接种量接种于LB 培养基中,在28℃、180rpm条件下发酵48h,形成发酵液,即为菌剂。
所述的发酵液中铜绿假单胞菌Pa2-2活菌的浓度为107~108CFU/mL。
所述的LB培养基为:胰蛋白胨10g,酵母提取物5g,氯化钠10g,添加蒸馏水定容至1000mL,混匀后分装于三角瓶中,121℃灭菌30min。
所述的菌剂的使用方法为:菌剂用无菌水稀释10倍,使稀释液活菌浓度为106~107CFU/ml,灌根于丹参根部周围;每7d灌根处理1次,共灌根2次。
本发明有益效果在于:本发明的目的是利用微生物防治丹参根腐病,因此本发明通过分离鉴定、平板拮抗、防病盆栽实验,从丹参根际土分离出能够防治丹参根病的Pa2-2,经鉴定为铜绿假单胞菌(Pseudomonas aeruginosa)。试验表明,本发明的铜绿假单胞菌对丹参根腐病病原菌均具有良好的抑制作用,对丹参根腐病病害的防效能达到55%以上,能够有效降低丹参根腐病的发病率,其效果优于现有的常用化学农药。本发明筛选的铜绿假单胞菌使用后不会产生抗药性、农药残留、环境污染等化学农药常见问题,对人畜健康、环境友好,符合人们对生态农业的需求。
附图说明
图1为菌株Pa2-2对层出镰孢菌、腐皮镰刀菌、尖孢镰刀菌、细极链格孢的平板拮抗试验结果对比图。
图2为菌株Pa2-2的菌落形态图。
图3为菌株Pa2-2对烟叶过敏性试验结果对比图,其中图左为菌株Pa2-2,图右为阳性对照。
具体实施方式
下面结合实施例和附图对本发明做进一步的说明。
实验材料:
LB培养基:胰蛋白胨10g,酵母提取物5g,氯化钠10g,琼脂粉17g(仅做固体培养基时添加),添加蒸馏水定容至1000ml,混匀后分装于三角瓶中, 121℃灭菌30min。
本发明所用病原菌及其它原料、试剂均可由市场购得。
(一)生防菌的分离
称取1g土壤,加入10ml无菌水中充分振荡获得土壤悬浮液,然后将土壤悬浮液摇匀后进行稀释,取200μL稀释液涂布于LB固体平板上,培养,至菌落长出,挑取大小、颜色或形态不同的菌落在LB固体培养基表面划线分离纯化,获得1株菌株,命名为Pa2-2。
(二)生防菌的筛选(平板拮抗试验)
利用平板对峙方法筛选生防菌:将丹参根腐病腐病原菌层出镰孢菌、腐皮镰刀菌、尖孢镰刀菌、细极链格孢分别接种在PDA培养基上,于28℃条件下培养 5d,使用灭好菌的直径为8mm的打孔器将培养好的丹参根腐病病原菌制成菌饼,接到新PDA平板(直径为90mm)的一侧,将直径为8mm的纸碟环灭好菌,放在PDA培养基另一侧,吸取10μl生防菌菌悬液缓慢滴加在纸碟环内,接种时保证生防菌与纸碟环处于同一条直线上。用封口膜封好后,28℃,倒置培养,待对照病原菌长至纸碟处后记录抑菌带的大小,并测量平板上相应病原菌的菌落半径。对照为丹参根腐病腐病原菌层出镰孢菌、腐皮镰刀菌、尖孢镰刀菌、细极链格孢与接无菌水纸碟在PDA上的培养,菌落处理最少3次重复,5d后检查对峙培养结果,结果见表1、图1,Pa2-2对4种病原菌均有效果。
表1:铜绿假单胞菌(Pa2-2)对丹参根腐病病原菌的抑菌效果
(三)生防菌鉴定
对筛选出拮抗效果好的菌株进行形态鉴定,经形态学、生理生化分析(见表 2)和分子生物学鉴定,确定为铜绿假单胞菌。贝莱斯芽抱杆菌在培养基上形成黄绿色菌落,不透明,中间略凸起,边缘不平整,挑起有劲性(见图2)。
表2:Pa2-2生理生化特征
注:“-”为阴性,“+”为阳性。
(四)生防菌测序
测序:对菌株Pa2-2进行测序,鉴定结果为铜绿假单胞菌(Pseudomonasaeruginosa)。
16SrDNA基因测序结果为:
(五)菌剂制备
将铜绿假单胞菌Pa2-2接种于LB培养基中,在28℃、180rpm条件下振荡培养 12h制备种子菌,种子菌以2%的接种量接种于LB培养基中,在28℃、180rpm条件下发酵48h,形成发酵液,即为菌剂。所述的发酵液中铜绿假单胞菌Pa2-2活菌的浓度为107~108CFU/mL。
(六)菌剂对烟叶过敏性试验
对烟草下表皮进行皮下接种,以10mM MgSO4为阴性对照,以烟草野火菌为阳性对照(YH)。在24℃,60%-80%的高湿度条件下,经24-48h后出现过敏性坏死枯斑的为阳性反应,3d后出现黄斑的为阴性反应。结果发现,如图3 所示,铜绿假单胞菌Pa2-2过敏性试验为阴性反应,因此铜绿假单胞菌Pa2-2无致病性,可作为防治丹参根腐病生防菌。
(七)生防菌的防病实验
将丹参种子在装有灭菌土的托盘上萌发,然后移入营养钵培养30天,采用灌根法接种生防菌,灌根前用少量清水湿润土壤,于根际接种铜绿假单胞菌Pa2-2 发酵液(活菌浓度为107~108CFU/mL),发酵液用无菌水稀释10倍,每株10mL,间隔7d再接种10mL;对照组同样方法接入等量LB培养基(CK),另外,设置药剂阳性对照组,对照药剂为50%多菌灵可湿性粉剂1000倍稀释液,每株丹参苗10mL,7d之后再灌根1次,共灌根2次。在各组最后一次灌根3d后接种丹参根腐病病原菌腐皮镰刀菌。接种方法:灌根接种,取10mL根腐病病原菌孢子悬浮液(浓度为1×106个/mL)对丹参幼苗进行灌根接种,3d浇水1次,置于28℃恒温培养箱中培养,定期观察丹参的发病情况。培养30d后调查、计算发病率、病情指数和防治效果。丹参根腐病病害分级标准参照相关文献进行,结果见表3。
表3铜绿假单胞菌Pa2-2对丹参根腐病盆栽防病效果
结果表明,铜绿假单胞菌Pa2-2对丹参根腐病的防效为40.4%,效果明显。
以上所述仅为本发明最佳的实施例,对于本领域的技术人员来说,本发明可以有各种更改和变化。凡在本发明的精神和原则之内,所作的任何修改、等同替换、改进等,均应包含在本发明的保护范围之内。
序列表
<110> 河南省农业科学院植物保护研究所
<120> 一种能够防治丹参根腐病的铜绿假单胞菌及其应用
<141> 2021-01-07
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1527
<212> DNA
<213> Pseudomonas aeruginosa
<400> 1
atntnvrsna gagtttgatc ctggctcagg acgaacgctg gcggcgtgcc taatacatgc 60
aagtcgagcg agattattta gaagcttgct tctaaataat ctagcggcgg acgggtgagt 120
aacacgtagg caacctgccc acaagacagg gataactacc ggaaacggta gctaataccc 180
gatacatctt tttcctgcat gggagaagga ggaaaggcgg agcaatctgt cacttgtgga 240
tgggcctgcg gcgcattagc tagttggtgg ggtaaaggcc taccaaggcg acgatgcgta 300
gccgacctga gagggtgatc ggccacactg ggactgagac acggcccaga ctcctacggg 360
aggcagcagt agggaatctt ccgcaatggg cgaaagcctg acggagcaac gccgcgtgag 420
tgatgaaggt tttcggatcg taaagctctg ttgccaggga agaacgtctt gtagagtaac 480
tgctacaaga gtgacggtac ctgagaagaa agccccggct aactacgtgc cagcagccgc 540
ggtaatacgt agggggcaag cgttgtccgg aattattggg cgtaaagcgc gcgcaggcgg 600
ctctttaagt ctggtgttta atcccgaggc tcaacttcgg gtcgcactgg aaactgggga 660
gcttgagtgc agaagaggag agtggaattc cacgtgtagc ggtgaaatgc gtagagatgt 720
ggaggaacac cagtggcgaa ggcgactctc tgggctgtaa ctgacgctga ggcgcgaaag 780
cgtggggagc aaacaggatt agataccctg gtagtccacg ccgtaaacga tgaatgctag 840
gtgttagggg tttcgatacc cttggtgccg aagttaacac attaagcatt ccgcctgggg 900
agtacggtcg caagactgaa actcaaagga attgacgggg acccgcacaa gcagtggagt 960
atgtggttta attcgaagca acgcgaagaa ccttaccagg tcttgacatc cctctgaccg 1020
ctgtagagat atggctttcc ttcgggacag aggagacagg tggtgcatgg ttgtcgtcag 1080
ctcgtgtcgt gagatgttgg gttaagtccc gcaacgagcg caacccttat gcttagttgc 1140
cagcaggtca agctgggcac tctaagcaga ctgccggtga caaaccggag gaaggtgggg 1200
atgacgtcaa atcatcatgc cccttatgac ctgggctaca cacgtactac aatggccggt 1260
acaacgggaa gcgaaggagc aatctggagc caatcctaga aaagccggtc tcagttcgga 1320
ttgtaggctg caactcgcct acatgaagtc ggaattgcta gtaatcgcgg atcagcatgc 1380
cgcggtgaat acgttcccgg gtcttgtaca caccgcccgt cacaccacga gagtttacaa 1440
cacccgaagt cggtgaggta accgcaaggg gccagccgcc gaaggtgggg tagatgattg 1500
gggtgaagtc gtaacaaggt agccgta 1527
Claims (6)
1.一种能够防治丹参根腐病的铜绿假单胞菌,该菌株编号为铜绿假单胞菌(Pseudomonas aeruginosa)Pa2-2,已保藏于“中国典型培养物保藏中心”,保藏地址:湖北省武汉市,洪山区八一路,武汉大学山国典型培养物保藏中心,登记入册的保藏号为CCTCCM 2020928,保藏日期为2020年12月21日。
2.根据权利要求1所述的铜绿假单胞菌在防治丹参根腐病方面的应用。
3.根据权利要求1所述的铜绿假单胞菌在制备防治丹参根腐病的菌剂中的应用。
4.根据权利要求3所述的应用,其特征在于,所述的菌剂的制备方法为:将铜绿假单胞菌Pa2-2接种于LB培养基中,在28℃、180rpm条件下振荡培养12h制备种子菌,种子菌以2%的接种量接种于LB培养基中,在28℃、180rpm条件下发酵48h,形成发酵液,即为菌剂。
5.根据权利要求4所述的菌剂的制备方法,其特征在于,所述的菌剂中铜绿假单胞菌Pa2-2活菌的浓度为107~108CFU/mL。
6.根据权利要求3所述的应用,其特征在于,菌剂的使用方法为:菌剂用无菌水稀释10倍,使稀释液活菌浓度为106~107CFU/ml,灌根于丹参根部周围;每7d灌根处理1次,共灌根2次。
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Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN104285962A (zh) * | 2010-08-05 | 2015-01-21 | 拜耳知识产权有限责任公司 | 包含丙硫菌唑和氟唑菌酰胺的活性化合物组合产品 |
CN104877943A (zh) * | 2015-06-11 | 2015-09-02 | 福建农林大学 | 一株防治地黄根腐病的拮抗菌及其应用 |
CN106676034A (zh) * | 2016-11-25 | 2017-05-17 | 浙江理工大学 | 具有抗三七根腐病作用的细菌及其用途 |
CN109234197A (zh) * | 2018-09-29 | 2019-01-18 | 河南省农业科学院植物保护研究所 | 一株能够同时防治烟草黑胫病和根黑腐病且具有促生作用的铜绿假单胞杆菌及其应用 |
CN111763643A (zh) * | 2020-07-09 | 2020-10-13 | 中国科学院南京土壤研究所 | 一种防治花生根腐病的复合菌群及其应用 |
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Patent Citations (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN104285962A (zh) * | 2010-08-05 | 2015-01-21 | 拜耳知识产权有限责任公司 | 包含丙硫菌唑和氟唑菌酰胺的活性化合物组合产品 |
CN104877943A (zh) * | 2015-06-11 | 2015-09-02 | 福建农林大学 | 一株防治地黄根腐病的拮抗菌及其应用 |
CN106676034A (zh) * | 2016-11-25 | 2017-05-17 | 浙江理工大学 | 具有抗三七根腐病作用的细菌及其用途 |
CN109234197A (zh) * | 2018-09-29 | 2019-01-18 | 河南省农业科学院植物保护研究所 | 一株能够同时防治烟草黑胫病和根黑腐病且具有促生作用的铜绿假单胞杆菌及其应用 |
CN111763643A (zh) * | 2020-07-09 | 2020-10-13 | 中国科学院南京土壤研究所 | 一种防治花生根腐病的复合菌群及其应用 |
Non-Patent Citations (3)
Title |
---|
Biodiscovery of antibacterial constituents from the endolichenic fungi isolated from Parmotrema rampoddense;Mario A. Tan et al.;《3 Biotech》;20200424;第10卷(第212期);第1-7页 * |
偏肿栓菌内生木霉菌株18-21抑菌效果研究;李强等;《微生物学杂志》;20160229;第 36 卷(第 1 期);第17-21页 * |
生防细菌PA2101和PG3402抑菌和促生特性的研究;千慧敏等;《中国生物防治学报》;20200202;第36卷(第1期);第940-948页 * |
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