CN110959562B - 牡蛎肽在促进海洋希瓦氏菌诱导厚壳贻贝稚贝附着上的应用 - Google Patents
牡蛎肽在促进海洋希瓦氏菌诱导厚壳贻贝稚贝附着上的应用 Download PDFInfo
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- 239000007788 liquid Substances 0.000 claims description 4
- 241000237536 Mytilus edulis Species 0.000 abstract description 6
- 235000020638 mussel Nutrition 0.000 abstract description 6
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- 244000241838 Lycium barbarum Species 0.000 description 1
- 235000015459 Lycium barbarum Nutrition 0.000 description 1
- 235000003140 Panax quinquefolius Nutrition 0.000 description 1
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Abstract
本发明公开了牡蛎肽在促进海洋希瓦氏菌诱导厚壳贻贝稚贝附着上的应用，海洋希瓦氏菌保藏于中国普通微生物菌种保藏管理中心，保藏号为CGMCC No:9798，牡蛎肽的浓度为0.01～1mg/mL，且为低聚肽质量百分含量不低于65％和分子量小于1kDa的牡蛎低聚肽，其对海洋希瓦氏菌诱导厚壳贻贝稚贝附着具有促进作用，有利于贻贝的水产养殖和海洋中贻贝种群的稳定，对探索厚壳贻贝稚贝附着机制和厚壳贻贝培育与养殖具有重要意义。
Description
技术领域
本发明涉及牡蛎肽的用途，具体涉及牡蛎肽在促进海洋希瓦氏菌(Shewanellaloihica)诱导厚壳贻贝稚贝附着上的应用。
背景技术
厚壳贻贝(Mytilus coruscus)，隶属于软体动物门(Mollusca)，双壳纲(Bivalvia)，贻贝目(Mytilodia)，贻贝科(Mytilidae)，分布于黄海、渤海和东海沿岸，是我国重要的海产贝类养殖品种。厚壳贻贝是从受精卵发育到眼点幼虫后，再经过附着变态这一过程，发育成稚贝，并最终发育为成贝。在由稚贝向成贝生长发育过程中，当环境发生变化时，厚壳贻贝稚贝能够自行切断足丝，开始爬行并重新选择适宜的附着基再次附着，稚贝附着对于贻贝的水产养殖和海洋中贻贝种群的稳定十分重要。牡蛎作为一种会极易相互附着集群的生物，由其制备的牡蛎肽能否同样吸引稚贝的附着，目前在还没有相关现有技术报道。在厚壳贻贝养殖过程中，稚贝附着率低是导致成贝存活率低的重要原因，探索牡蛎肽刺激希瓦氏菌生物被膜诱导厚壳贻贝附着具有重要的经济价值和生态学意义。
发明内容
本发明提供一种牡蛎肽在促进海洋希瓦氏菌诱导厚壳贻贝稚贝附着上的应用，其通过促进海洋希瓦细菌形成生物被膜和释放胞外产物诱导厚壳贻贝稚贝的附着。
进一步，所述牡蛎肽的浓度为0.01～1mg/mL，优选0.3mg/mL。
在某些实施方案中，所述牡蛎肽为牡蛎低聚肽，且低聚肽在总肽中的质量百分含量不低于65％，分子量小于1kDa，能溶于水且易被吸收。
在某些实施方案中，所述牡蛎低聚肽中，所述低聚肽在总肽中的质量百分含量不低于98％。
与现有技术相比，本发明发现加入一定浓度的牡蛎肽培养海洋希瓦氏菌形成生物被膜和释放胞外产物对诱导厚壳贻贝稚贝附着具有促进作用，相较于没有加入牡蛎肽时，其稚贝附着率约可提高35.72％，为贻贝的培育养殖提供理论基础，对探索厚壳贻贝稚贝附着机制和厚壳贻贝培育与养殖具有重要意义。
附图说明
图1是不同浓度的牡蛎肽培养海洋希瓦氏菌诱导厚壳贻贝稚贝附着的附着率。
具体实施方式
以下结合具体实施例进一步解释本发明的技术方案，但不限于此。
以下实施例中使用的海洋希瓦氏菌(Shewanella loihica)与中国专利ZL201410741373.4保藏的海洋希瓦氏细菌(Shewanella sp.ECSMB56)相同，来来自于浙江省嵊泗县枸杞岛(122°46′E 30°43′N)的自然生物被膜表面，在NCBI上的序列号为KM369861，其保藏信息为：该细菌菌株保藏于中国普通微生物菌种保藏管理中心，保藏日期为2014年10月20日，保藏号为CGMCC No：9798，保藏单位地址为北京市朝阳区北辰西路1号院3号，中国科学院微生物研究所。
以下实施例中使用的牡蛎肽是来自大连深蓝肽科技研发有限公司的牡蛎低聚肽粉，低聚肽在总肽中的质量百分含量不低于98％，分子量小于1kDa。
实施例1培养海洋希瓦氏菌
将海洋希瓦氏菌的纯种菌株先在2216E固体培养基进行涂板和划线得到单菌落，然后转接到Zobell 2216E液体培养基中扩大培养，离心洗涤并制成悬浮液。然后用吖啶橙染色并在荧光显微镜1000倍镜头下随机选取10个点进行计数，确定菌液总密度为4.3×109cells/mL。
实施例2海洋希瓦氏菌对厚壳贻贝稚贝附着的诱导实验
选用实施例1中初始菌液密度为1×108cells/mL的海洋希瓦氏菌液，在灭菌培养皿中放入一个灭菌的载玻片，加入一定量的含有不同浓度牡蛎肽的灭菌过滤海水和菌液使培养皿中液体总体积为20mL，牡蛎肽的浓度为0mg/mL、0.1mg/mL、0.3mg/mL、0.5mg/mL和1mg/mL，在18℃、黑暗条件下培养48h形成生物被膜。将一枚附有生物被膜的载玻片转移到新的含有灭菌过滤海水的培养皿中在载玻片左右均匀放入10只厚壳贻贝稚贝(壳长1.22±0.05mm，壳高0.75±0.02mm)，在24h时记录稚贝的附着率，即载玻片上附着的稚贝个数占该培养皿中稚贝总个数的百分比，结果如图1所示。
由图1可知，牡蛎肽对海洋希瓦氏菌的生物被膜诱导厚壳贻贝稚贝附着的诱导活性有增强作用，不同牡蛎肽浓度形成的生物被膜对稚贝的诱导活性也有所不同。在仅有海洋希瓦氏菌形成生物被膜的情况下稚贝附着的诱导率仅达30％，但在牡蛎肽浓度为0.3mg/mL的情况下诱导率高达46.67％，相比之下诱导率提高了35.72％。由此可见，该种牡蛎肽能促进海洋希瓦细菌形成生物被膜和释放胞外产物诱导厚壳贻贝稚贝的附着，且稚贝附着有利于贻贝的水产养殖和海洋中贻贝种群的稳定，对于阐明厚壳贻贝稚贝附着机制和厚壳贻贝培育与养殖均具有重要意义。
Claims (3)
1.牡蛎肽在促进海洋希瓦氏菌(Shewanella loihica)诱导厚壳贻贝稚贝附着上的应用；
所述牡蛎肽的浓度为0.01～1mg/mL，为分子量小于1kDa的牡蛎低聚肽，且低聚肽在总肽中的质量百分含量不低于65％；
所述牡蛎肽通过促进海洋希瓦细菌形成生物被膜和释放胞外产物诱导厚壳贻贝稚贝附着，其中：所述海洋希瓦氏菌保藏于中国普通微生物菌种保藏管理中心，保藏号为CGMCCNo：9798，其初始菌液密度为1×108～10×108cells/mL。
2.根据权利要求1所述的应用，其特征在于，所述牡蛎肽的浓度为0.3mg/mL。
3.根据权利要求1所述的应用，其特征在于，所述牡蛎肽中低聚肽在总肽中的质量百分含量不低于98％。
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