Bt水稻对褐飞虱若虫肠道可培养细菌组成的影响

林胜; 李强; 夏晓峰; 尤民生

doi:10.19303/j.issn.1008-0384.2019.07.008

Bt水稻对褐飞虱若虫肠道可培养细菌组成的影响

doi: 10.19303/j.issn.1008-0384.2019.07.008

林胜^{1, 2, 3, 4,},
李强^{1, 2, 3, 4},
夏晓峰^{1, 2, 3, 4},
尤民生^{1, 2, 3, 4, ,}

1.
福建农林大学闽台作物有害生物生态防控国家重点实验室, 福建福州 350002
2.
福建农林大学应用生态研究所, 福建福州 350002
3.
教育部害虫生态防控国际合作联合实验室, 福建福州 350002
4.
农业部闽台作物有害生物综合治理重点实验室, 福建福州 350002

基金项目:

福建省自然科学基金项目 2017J01432

国家自然科学基金项目 31870520

福建农林大学科技创新专项基金 CXZX2017317

国家转基因生物新品种培育重大专项课题 2016ZX08011-001

详细信息

作者简介:
林胜(1979-), 男, 博士, 研究方向:昆虫生态与害虫综合治理(E-mail:linsn001@163.com)

通讯作者:
尤民生(1954-), 男, 教授, 研究方向:昆虫生态与害虫综合治理(E-mail:msyou@fafu.edu.cn)

中图分类号: S663.4
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- 被引次数: 0
出版历程
- 收稿日期: 2018-12-10
- 修回日期: 2019-06-09
- 刊出日期: 2019-07-20

Effects of Bt Rice on the Structure of Culturable Bacteria in the Gut of Nilaparvata lugens Nymphae

LIN Sheng^{1, 2, 3, 4
,},
LI Qiang^{1, 2, 3, 4},
XIA Xiao-feng^{1, 2, 3, 4},
YOU Min-sheng^{1, 2, 3, 4
, ,}

1.
State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China
2.
Institute of Applied Ecology, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China
3.
Joint International Research Laboratory of Ecological Pest Control, Ministry of Education, Fuzhou, Fujian 350002, China
4.
Key Laboratory of Integrated Pest Management of Fujian and Taiwan, China Ministry of Agriculture, Fuzhou, Fujian 350002, China

摘要

摘要: 目的探究Bt水稻对褐飞虱若虫肠道可培养细菌组成结构的影响，为转基因水稻的安全性评价提供科学依据。方法利用传统微生物纯培养法分别从取食非转基因（CK）水稻和转crylAb抗虫（Bt）水稻的褐飞虱肠道内分离细菌，利用16s rDNA对分离菌株进行鉴定，比较分析转基因水稻对褐飞虱若虫肠道微生物群落结构的影响。结果研究结果显示在取食CK水稻和Bt水稻的褐飞虱肠道内各分离获得7株细菌。取食CK水稻的褐飞虱肠道细菌分别属于3个门5个属；而取食Bt水稻的褐飞虱肠道细菌分别属于2个门4个属；分离获得频率最高的是变形菌门细菌Proteobateria，其中取食两种水稻均能分离获得变形菌门的泛菌属Pantoea，以及厚壁菌门Firmicutes的葡萄球菌属Staphylococcus等2个属的细菌。取食CK水稻的褐飞虱肠道内分离获得变形菌门的草螺菌属Herbaspirillum和不动杆菌属Acinetobacter，以及放线菌门的微杆菌属Microbacterium等3个特异性细菌属；取食Bt水稻的褐飞虱肠道内分离得到变形菌门的肠杆菌属Enterobacter和伯克氏菌属Burkholderia等2个特异性细菌属。结论取食非转基因水稻和转基因水稻的褐飞虱肠道可培养细菌组成存在一定的差异。本研究为后续深入研究褐飞虱肠道共生细菌的功能，以及转基因水稻对非靶标生物肠道细菌的影响提供了材料和基础。
- 褐飞虱 /
- 转基因水稻 /
- 肠道细菌 /
- 非靶标效应 /
- 生物安全
Abstract: Objective The present study aims to explore the effects of Bt rice on gut bacterial structure of the Nilaparvata lugens nymphae and to provide evidence for rational safety assessment of Bt rice. Method Gut bacteria from BPH feeding on non-transgenic rice (CK) and transgenic rice (Bt) were isolated using traditional culture methods, and were identified using 16s rDNA. The composition of isolates was compared between these two treatments. Results The results showed that seven bacterial strains were isolated from the gut of N. lugens feeding on either CK or Bt rice. Three phyla and five genera of the gut bacteria were identified from N. lugens feeding on CK rice, while two phyla and four genera of the gut bacteria from N. lugens feeding on Bt rice. The taxa of bacteria identified from the guts of N. lugens either feeding on CK rice or Bt rice were Proteobateria (including the genus of Pantoea) and Firmicutes (including the genus of Staphylococcus). Herbaspirillum Acinetobacter and Microbacterium were specific genera from the CK treatment, while Enterobacter, and Burkholderia from the Bt treatment. Conclusion The composition of culturable bacteria in the N. lugens gut was different beween the CK treatment and the Bt treatment. The findings provide an important foundation for further studies on the function of symbiotic bacteria in N. lugens, and the effects of transgenic rice on the gut bacteria of non-target species.
- Nilaparvata lugens /
- genetically modified rice /
- gut bacteria /
- non-target effect /
- biosafety

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图 1 取食非转基因水稻的褐飞虱肠道细菌菌落

Figure 1. Colony morphology of bacteria strains isolated from N. lugens feeding on the non-transgenic rice

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图 2 取食转基因水稻的褐飞虱肠道细菌菌落

Figure 2. Colony morphology of bacteria strains isolated from N. lugens feeding on the transgenic rice

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图 3 取食非转基因水稻的褐飞虱肠道细菌16S rDNA PCR产物电泳图

注：M泳道为100~5 000 bp DNA marker，1~14泳道分别为7株菌株CK1~CK7的2次重复，15泳道为阴性对照。

Figure 3. Gel electrophoresis of 16S rDNA PCR products of gut bacteria from Nilaparvata lugens feeding on the non-transgenic rice

Note: Lane M is DNA marker(100-5 000 bp), Lane 1-14 are PCR products of CK1 to CK7 with 1 repeat of each strain, respectively, Lane 15 is negative control.

下载: 全尺寸图片幻灯片

图 4 取食转基因水稻褐飞虱肠道细菌16S rDNA PCR产物电泳图

注：M泳道为100~5 000 bp DNA marker，1~14泳道分别为7株菌株Bt1~Bt7的2次重复，15泳道为阴性对照。

Figure 4. Gel electrophoresis of 16S rDNA PCR products of gut bacteria from N. lugens feeding on the transgenic rice

Note: Lane M is DNA marker (100-5 000 bp), Lane 1-14 are PCR products of Bt1 to Bt7 with 1 repeat of each strain, respectively, Lane 15 is negative control.

下载: 全尺寸图片幻灯片

图 5 取食非转基因水稻的褐飞虱肠道细菌16S rDNA系统进化分析

注：CK1~7序列分离自取食非转基因水稻褐飞虱肠道，分支间的数值是基于bootstrap验证的自展检验值(100次重复)，刻度标尺表示遗传距离为0.5。

Figure 5. Phylogenetic analysis based on 16S rDNA sequences from the gut bacteria of N. lugens feeding on the non-transgenic rice

Note: The sequences of CK1-7 were isolated from N. lugens gut bacteria feeding on the non-transgenic rice. The bootstrap test (100 replicates) was shown next to the branches. The scale bar represents genetic distance of 0.5.

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图 6 取食转基因水稻的褐飞虱肠道细菌16S rDNA系统进化分析

注：Bt1~7序列分离自取食转基因水稻褐飞虱肠道，分支间的数值是基于bootstrap验证的自展检验值(100次重复)，刻度标尺表示遗传距离为0.05。

Figure 6. Phylogenetic analysis based on 16S rDNA sequences from the gut bacteria of N. lugens feeding on the transgenic rice

Note: The sequences of Bt1-7 were isolated from N. lugens gut bacteria feeding on the transgenic rice. The bootstrap test (100 replicates) was shown next to the branches. The scale bar represents genetic distance of 0.05.

下载: 全尺寸图片幻灯片

表 1 褐飞虱肠道细菌的16S rDNA序列的Blast比对结果

Table 1. Blast-based sequence alignments of 16S rDNA from the gut bacteria from N. lugens

菌株 Strain ID	登录号 GenBank Accession No.	相似性最高菌株(GenBank登录号) Most closely related hit in GenBank (GenBank Accession No.)	相似性 Similarity/%	分类地位(门) Taxonomy (Phylum)
CK1	KU863620	Pantoea ananatis (KT380684)	99	Proteobacteria
CK2	KU863621	Staphylococcus sciuri (JX134627)	99	Firmicutes
CK3	KU863622	Acinetobacter baylyi (KJ372731)	100	Proteobacteria
CK4	KU863623	Microbacterium laevaniformans (JQ229794)	99	Actinobacteria
CK5	KU863624	Pantoea sp. (LC484790)	99	Proteobacteria
CK6	KU863625	Staphylococcus saprophyticus (MK208692)	100	Firmicutes
CK7	KU863626	Herbaspirillum sp. (KM253161)	99	Proteobacteria
Bt1	KU863627	Enterobacter sp. (LC484731)	99	Proteobacteria
Bt2	KU863628	Staphylococcus pasteuri (KT728837)	100	Firmicutes
Bt3	KU863629	Burkholderia gladioli (MG386180)	99	Proteobacteria
Bt4	KU863630	Staphylococcus epidermidis (MK883070)	99	Firmicutes
Bt5	KU867651	Staphylococcus sciuri (KX023250)	99	Firmicutes
Bt6	KU867652	Burkholderia gladioli (KX907740)	99	Proteobacteria
Bt7	KU867653	Pantoea dispersa (GQ200831)	99	Proteobacteria

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