三叶鬼针草内生细菌群体多样性及重金属耐受和吲哚乙酸产生潜力

Diversity,heavy-metal tolerance and indoleacetic acid production of bacterial endophytes in Bidens pilosa

【背景】定殖于植物的多样内生细菌与宿主生长和抗逆能力密切相关。三叶鬼针草(Bidenspilosa L.)极具入侵性且抗逆性强,但目前该植物内生细菌相关研究报道极少。【目的】探究三叶鬼针草内生细菌群体多样性,筛选获得兼具重金属耐受性及吲哚乙酸(Indoleacetic acid,IAA)产生潜力的内生细菌菌株。【方法】采用MiSeq高通量方法分析三叶鬼针草内生细菌群体多样性,可培养方法测定内生细菌菌株对重金属Pb、Cd、Ni、Hg的耐受能力及产生IAA的潜力。【结果】从三叶鬼针草内生细菌群体中共测得4 031个操作分类单元(OTU),可归属于25个门51个纲76个目182个科及536个属。属级水平上,三叶鬼针草根、茎、叶和种子中分别以肠杆菌属(Enterobacter)、不动杆菌属(Acinetobacter)、鞘脂单胞菌属(Sphingomonas)及假单胞菌属(Pseudomonas)丰度最高,伯克氏菌属(Burkholderia)、甲基杆菌属(Methylobacterium)、假单胞菌属、泛菌属(Pantoea)次之;从三叶鬼针草分离得到34株内生细菌,所有菌株均至少对1种测试的重金属具有耐受性。其中,GF-1、GF-8、YF-1、YF-2、JF-1、GF-2和JF-8这7个菌株能够产生IAA,其产率为57.48-312.22μg/mL;基于16S rRNA基因序列分析,初步鉴定菌株GF-1、GF-8、YF-1、YF-2、JF-1为芽孢杆菌(Bacillus spp.),菌株GF-2为假单胞菌(Pseudomonas sp.),菌株JF-8为伯克氏菌(Burkholderia sp.)。【结论】三叶鬼针草内生细菌群体具有丰富的种群多样性。三叶鬼针草内生细菌菌株GF-1、GF-8、YF-1、YF-2、JF-1、GF-2和JF-8兼具多重金属耐受性及高产IAA潜力,是重金属复合污染土壤生物修复的优良候选菌株。

[Background]Plants are inhabited by diverse bacterial endophytes that are closely related to the growth and stress tolerance of their hosts.Bidens pilisa L.is a highly invasive plant species with strong stress tolerance.However,up to date,there is limited literature reporting the research involving the bacterial endophytes of the plant species.[Objective]To investigate and characterize the diversity of bacterial endophytes community of B.pilisa,and to obtain the bacterial isolates from the plant species with both potential of heavy metal-tolerance and indoleacetic acid(IAA)production.[Methods]The diversity of bacterial endophytes community was analyzed by using MiSeq high-throughput sequencing method.The tolerance ability to heavy metals Pb,Cd,Ni,and Hg,as well as the IAA-producing ability of bacterial endophytes were evaluated by using culture-dependent method.[Results]We recovered a total of 4 031 distinct operational taxonomic units from the total bacterial endophytes community of B.pilosa,which could be affiliated with 25 distinct bacterial phyla,51 distinct bacterial classes,76 distinct bacterial orders,182 distinct bacterial families,and 536 distinct bacterial genera.At the genus level,the most dominant bacterial genera detected from the root,stem,leaf,and seed of B.pilosam were Enterobacter,Acinetobacter,Sphingomonas,and Pseudomonas,respectively,followed by Burkholderia,Methylobacterium,Pseudomonas,and Pantoea.Using culture-dependent method,we obtained 34 bacterial isolates from the internal tissues of B.pilisa and all obtained strains showed the tolerance at least to one tested heavy metal.Seven strains(numbered GF-1,GF-8,YF-1,YF-2,JF-1,GF-2,JF-8,respectively)could produce IAA with IAA yields varying in the ranging from 57.48-312.22μg/mL.Based on 16 S rRNA gene sequence analysis,the 7 IAA-producing strains were identified,and strains GF-1,GF-8,YF-1,YF-2,JF-1 as Bacillus spp.,strain GF-2 as Pseudomonas sp.,strain JF-8 as Burkholderia sp.[Conclusion]The bacterial endophytes community of B.pilosa has high population diversity.Bacterial endophytes strains GF-1,GF-8,YF-1,YF-2,JF-1,GF-2,and JF-8 obtained from B.pilosa show not only multi-heavy metals tolerance also high IAA production,being good candidates used as bio-inoculation agent for bioremediation of heavy metal-contaminated soil.