厚壳贻贝肠道细菌的生物被膜对其幼虫和稚贝附着的影响

徐嘉康; 王劲松; 方怡涵; 杨金龙; 梁箫

doi:10.12284/hyxb2021112

厚壳贻贝肠道细菌的生物被膜对其幼虫和稚贝附着的影响

doi: 10.12284/hyxb2021112

徐嘉康^{1, 2,},
王劲松^{1, 2},
方怡涵^{1, 2},
杨金龙^{1, 2, 3},
梁箫^{1, 2, ,}

1.
上海海洋大学国家海洋生物科学国际联合研究中心，上海 201306
2.
上海海洋大学水产种质资源发掘与利用教育部重点实验室，上海 201306
3.
南方海洋科学与工程广东省实验室（广州），广东广州 511458

基金项目: 上海市优秀学术带头人计划（20XD1421800）；国家自然科学基金（41876159）；南方海洋科学与工程广东省实验室（广州）人才团队引进重大专项（GML2019ZD0402）；国家重点研发计划项目（2020YFD0900804）。

详细信息

作者简介:
徐嘉康（1996-），男，辽宁省沈阳市人，从事海洋贝类分子生物学的研究。E-mail：m180100004@st.shou.edu.cn

通讯作者:
梁箫（1983-），女，博士，从事海洋贝类分子生物学的研究。E-mail：x-liang@shou.edu.cn

中图分类号: Q178.53
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- 被引次数: 0
出版历程
- 收稿日期: 2020-12-30
- 修回日期: 2021-03-31
- 网络出版日期: 2021-05-19
- 刊出日期: 2021-09-25

Effects of intestinal bacterial biofilms on settlement process of larvae and plantigrades in Mytilus coruscus

Xu Jiakang^{1, 2
,},
Wang Jinsong^{1, 2},
Fang Yihan^{1, 2},
Yang Jinlong^{1, 2, 3},
Liang Xiao^{1, 2
, ,}

1.
International Research Center for Marine Biosciences, Ministry of Science and Technology, Shanghai Ocean University, Shanghai 201306, China
2.
Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources, Ministry of Education, Shanghai Ocean University, Shanghai 201306, China
3.
Southern Marine Science and Engineering Guangdong Laboratory (Guangzhou), Guangzhou 511458, China

摘要

摘要: 为研究肠道细菌在厚壳贻贝(Mytilus coruscus)幼虫和稚贝生长发育过程中的作用，本研究从成体厚壳贻贝肠道中分离出了10株细菌，通过分别形成单一细菌生物被膜，检验其对厚壳贻贝幼虫和稚贝附着的影响和生物被膜特性。实验结果发现，10株肠道细菌所形成的生物被膜均能诱导厚壳贻贝幼虫和稚贝的附着，但不同种类肠道细菌的诱导能力不同，其中Bacillus sp.4对厚壳贻贝幼虫具有高诱导活性，Phaeobacter sp.1具有低诱导活性；Phaeobacter sp.1对厚壳贻贝稚贝具有高诱导活性，Bacillus sp.4具有低诱导活性。通过比较分析 Bacillus sp.4和Phaeobacter sp.1生物被膜的生物量及胞外产物发现，肠道细菌被膜细菌密度、膜厚和胞外脂类对厚壳贻贝幼虫的附着变态无影响，而胞外蛋白和胞外多糖可以影响幼虫的附着变态；对于厚壳贻贝稚贝的附着，肠道细菌被膜细菌密度、膜厚和胞外α-多糖均能影响其诱导活性，而胞外脂类和胞外蛋白无影响。本研究成果可为提高厚壳贻贝的健康生态养殖相关技术提供相关的指导，为解析生物被膜调控厚壳贻贝附着机制和该物种生态健康养殖提供理论依据。
- 肠道细菌 /
- 厚壳贻贝 /
- 生物被膜 /
- 诱导活性；附着
Abstract: Ten strains of bacteria were isolated from the gut of Mytilus coruscus to study its role in the settlement of larvae and plantigrade by forming biofilms. Results showed that the inducing ability of the biofilms formed by ten bacteria were different, although all of them could induce the settlement of larvae and plantigrade. In larvae, Bacillus sp.4 showed high inducing activity, while Phaeobacter sp.1 had low inducing activity. In plantigrades, Phaeobacter sp.1 showed high inducing activity, while Bacillus sp.4 showed low inducing activity. The polysaccharides and proteins from biofilms formed by Bacillus sp.4 and Phaeobacter sp.1 impacted larval settlement and metamorphosis, and the cell density, thickness and extracellular lipids of biofilm showed no effect on inducing activity of larval settlement and metamorphosis. For plantigrades, biofilm’s bacterial density, thickness and extracellular α-polysaccharide could induce plantigrades to the settlement, and the extracellular lipids and proteins of biofilms did not affect the settlement. This study is helpful for improving the healthy ecological culture of M. coruscus and to understand the settlement mechanism of M. coruscus.
- intestinal bacteria /
- Mytilus coruscus /
- biofilm /
- inducing activity; settlement

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图 1 实验所用贻贝肠道细菌的表型

Fig. 1 Phenotypes of the different bacterial strains from mussel intestinal in tested

下载: 全尺寸图片幻灯片

图 2 不同肠道细菌对厚壳贻贝的诱导作用

A. 对幼虫变态附着的诱导活性；B. 对稚贝附着的诱导活性；不同字母表示差异显著（p<0.05）

Fig. 2 Induction of settlement of Mytilus coruscus on the different intestinal bacterial biofilms

A. Inducing activity of larval settlement and metamorphosis; B. inducing activity of plantigrade settlement; values that are significantly different between each other at p<0.05 are indicated by different letters

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图 3 不同肠道细菌生物被膜的细菌密度

不同字母表示差异显著（p<0.05）

Fig. 3 The bacterial density of different intestinal bacterial biofilms

Values that are significantly different between each other at p<0.05 are indicated by different letters

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图 4 本实验中肠道细菌的系统进化树

Fig. 4 Phylogenetic tree of intestinal bacterial in tested

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图 5 Bacillus sp.4和Phaeobacter sp.1的生物被膜图像（A）及膜厚（B）

不同字母表示差异显著（p<0.05）

Fig. 5 Biofilm images (A) and biofilm thickness (B) of Bacillus sp.4 and Phaeobacter sp.1

Values that are significantly different between each other at p<0.05 are indicated by different letters

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图 6 生物被膜的CLSM图像分析

脂类、蛋白质、α-多糖和β-多糖在生物被膜上的分布（A）和生物量（B）；不同字母表示差异显著（p<0.05）

Fig. 6 The analysis of CLSM images of biofilms

The distribution (A) and biovolume (B) of lipids, proteins, α-polysaccharide and β-polysaccharide on BFs; values that are significantly different between each other at p < 0.05 are indicated by different letters

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表 1 肠道细菌16S rRNA基因序列分析

Tab. 1 16S rRNA gene sequence analysis of the intestinal bacterial strains

菌株来源	比对菌株	比对序列号	测试菌株	上传序列号	相似度/%
厚壳贻贝肠道	Bacillus sp.4	KF933662	ECSMC2	KU845379	98
厚壳贻贝肠道	Paracoccus sp.2	KJ648494	ECSMC12	KU845388	97
厚壳贻贝肠道	Flavobacterium sp.1	KF933689	ECSMC8	KU845384	99
厚壳贻贝肠道	Arenibacter sp.1	JQ898120	ECSMC15	KU845393	99
厚壳贻贝肠道	Mesoflavibacter sp.1	NR134082	ECSMC14	KU845390	99
厚壳贻贝肠道	Pseudoalteromonas sp.30	NR114190	ECSMB30	KX099925	100
厚壳贻贝肠道	Tenacibaculum sp.3	JN128275	ECSMC3	KU845380	99
厚壳贻贝肠道	Ruegeria sp.2	MF359423	ECSMC5	KU845381	99
厚壳贻贝肠道	Ahrensia sp.1	KJ700633	ECSMC1	KU845378	99
厚壳贻贝肠道	Phaeobacter sp.1	HE584770	ECSMC6	KU845382	100

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表 2 细菌密度与诱导活性的相关性分析

Tab. 2 Correlation analyses between the bacterial density and inducing activity

测试菌株	细菌密度
	幼虫诱导活性		稚贝诱导活性
	r	p	r	p
Bacillus sp.4	0.912 9	0.000 1*	−0.642 0	0.118 7
Paracoccus sp.2	0.804 1	0.000 1*	0.835 2	0.000 1*
Flavobacterium sp.1	0.227 2	0.182 7	0.874 2	0.000 1*
Arenibacter sp.1	−0.010 8	0.950 0	0.747 4	0.002 7*
Mesoflavibacter sp.1	0.247 0	0.146 5	0.426 1	0.196 3
Pseudoalteromonas sp.30	−0.672 7	0.000 1*	0.638 7	0.002 2*
Tenacibaculum sp.3	0.260 4	0.125 0	0.174 7	0.237 4
Ruegeria sp.2	−0.461 9	0.004 6*	0.791 6	0.000 1*
Ahrensia sp.1	0.344 1	0.039 9*	0.277 4	0.208 5
Phaeobacter sp.1	0.630 7	0.172 5	0.965 4	0.000 1*
注: *表示差异显著（p<0.05）。

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表 3 本实验中肠道细菌的遗传距离

Tab. 3 Genetic distances of intestinal bacterial in tested

测试菌株	2	12	8	15	14	30	3	5	1	6
2
12	0.267
8	0.254	0.011
15	0.319	0.286	0.308
14	0.317	0.307	0.294	0.097
30	0.265	0.194	0.221	0.303	0.290
3	0.336	0.285	0.305	0.118	0.099	0.274
5	0.253	0.063	0.076	0.118	0.314	0.230	0.311
1	0.255	0.144	0.117	0.303	0.295	0.217	0.292	0.139
6	0.303	0.102	0.084	0.310	0.315	0.237	0.317	0.039	0.138
注: 1, 2, ···, 30 分别表示菌株ECSMC1, ECSMC2, ···, ECSMC30。

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表 4 膜厚与诱导活性的相关性分析

Tab. 4 Correlation analyses between the biofilm thickness and inducing activity

测试菌株	生物被膜膜厚
	幼虫诱导活性		稚贝诱导活性
	r	p	r	p
Bacillus sp.4	0.376 4	0.108 7	0.264 2	0.075 6
Phaeobacter sp.1	0.375 3	0.115 4	0.886 4	0.000 1*
注: *表示差异显著（p<0.05）。

下载: 导出CSV

表 5 胞外产物与幼虫诱导活性的相关性分析

Tab. 5 Correlation analyses between extracellular product and inducing activity of larvae

测试菌株	生物被膜胞外产物生物量
	脂类		蛋白质		α-多糖		β-多糖
	r	p	r	p	r	p	r	p
Bacillus sp.4	0.363 4	0.094 2	0.486 5	0.000 1*	0.886 5	0.000 1*	0.753 2	0.000 1*
Phaeobacter sp.1	0.484 5	0.265 8	−0.864 3	0.028 7*	−0.642 4	0.147 5	−0.448 6	0.084 5
注: *表示差异显著（p < 0.05）。

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表 6 胞外产物与稚贝诱导活性的相关性分析

Tab. 6 Correlation analyses between the extracellular product and inducing activity of plantigrade

测试菌株	生物被膜胞外产物生物量
	脂类		蛋白质		α-多糖		β-多糖
	r	p	r	p	r	p	r	p
Bacillus sp.4	0.765 3	0.219 5	0.592 5	0.1209	−0.764 3	0.037 1*	−0.343 6	0.129 1
Phaeobacter sp.1	0.464 2	0.107 5	0.556 3	0.0753	0.669 8	0.044 3*	0.562 1	0.085 9
注: *表示差异显著（p < 0.05）。

下载: 导出CSV

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