CN112608862B - 一株石油烃降解功能菌scsio 19801及其应用 - Google Patents
一株石油烃降解功能菌scsio 19801及其应用 Download PDFInfo
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Abstract
本发明公开了一株石油烃降解功能菌SCSIO 19801及其应用。本发明的戈登氏菌(Gordonia sp.)SCSIO 19801与Gordonia terrae相似性最高(100％)，该菌株于2020年12月07日保藏于中国微生物菌种保藏管理委员会普通微生物中心，保藏号为：CGMCC No.21142。所述菌株(Gordonia sp.)SCSIO 19801具有高效降解石油烃化合物的能力，可广泛地应用于降解石油烃以及对石油污染的修复，如对海洋环境石油污染的水体修复等。
Description
技术领域：
本发明属于生物技术领域，涉及石油污染生物处理，具体涉及一株石油烃降解戈登氏菌(Gordonia sp.)SCSIO 19801及其应用。
背景技术：
据统计，全球每年进入海洋的石油类在1.0×1010kg以上，海洋溢油已成为严重的世界性海洋污染。生态环境部近年来中国海洋生态环境状况公报表明，石油类已成为我国石油类已成为我国海洋的主要污染物之一。海洋运输(船舶溢油、船舶和码头作业排污等)、海洋石油开采、陆源油污输入等是海洋溢油污染的主要来源。大面积的溢油在海面扩散形成油膜减弱了太阳辐射透入海水的能量和减少水中的含氧量，影响水生动物呼吸和水生植物的光和作用，导致海洋生物的死亡。亚致死量的溢油污染也会影响生物的摄食、繁殖和生长等过程。此外，石油烃类化合物对生物具有化学毒性，难降解的组分如多环芳烃等具有致畸、致癌和致突变效应，毒性化合物通过食物链富集传递等途径也会对人体健康造成极大的危害。
自然界中能降解石油污染的微生物分布广泛，约有100余属、200多种，包括细菌、真菌、放线菌、霉菌、以及藻类等。目前已报道的海洋来源石油降解细菌，主要包括无色杆菌属(Achromobacter)、假单胞菌属(Pseudomonas)、不动杆菌属(Acinetobacter)、鞘氨醇单胞菌属(Sphingomonas)、产碱杆属(Alcaligenes)、分枝杆菌属(Mycobacterium)、节杆菌属(Archrobacter)和芽孢杆菌属(Bacillus)等。微生物通过新陈代谢将石油烃转化为自身的生物量和无毒的小分子化合物，是海洋石油烃污染物生物降解的主要途径。由于微生物修复具有高效、环境友好和低成本等优势，是石油污染修复的主要研究方向。目前石油烃降解菌主要分离自沿海和陆地污染区域，对于开放海域石油烃降解微生物的研究相对较少。研究表明，随着人类活动的影响，石油类污染物已扩散到偏远的大洋中，包括岛礁等。由于环境因子(温度、盐度和营养物质等)是影响石油烃微生物降解关键的因素，因此从原位环境中分离具有高效降解石油烃的土著菌株有利于石油烃污染的修复。
近年来，戈登氏菌因在环境修复中发挥重要的作用而备受关注。研究表明，戈登氏菌在废水处理、多环芳烃和邻苯二甲酸酯类等有机物降解、重金属的生物吸附、生物脱硫、石油降解等方面具有潜在的应用价值。但已报道的戈登氏菌主要来源于陆地，且对石油烃降解能力较低，因此开发海洋来源高效降解石油烃的菌株对于溢油污染的原位修复具有重要的意义。
发明内容：
本发明的第一个目的是提供一种具有石油烃降解功能的戈登氏菌(Gordoniasp.)SCSIO 19801。
本发明的戈登氏菌(Gordonia sp.)SCSIO 19801，已于2020年12月07日在中国微生物菌种保藏管理委员会普通微生物中心保藏，保藏地址：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，邮政编码：100101，保藏号为：CGMCC No.21142。
实验证明，戈登氏菌SCSIO 19801可高效降解石油烃类化合物。在无机盐液体培养基中分别加入多环芳烃、液态烷烃和柴油为唯一碳源，接种该菌株后在28℃条件下培养7天，对菲的降解率约为83％，对芘的降解率约为51％，正十二烷的降解率为26％，正十六烷降解率为12％，对柴油的降解率约为23％。
本发明的第二个目的是提供上述戈登氏菌SCSIO 19801在降解石油烃中应用。
优选，是用戈登氏菌SCSIO 19801对石油污染的海洋水体进行修复。
所述石油烃类化合物包括但不限于多环芳烃类、液态烷烃、柴油等烃类化合物。
所述的多环芳烃类包括但不限于菲或芘。
所述的液态烷烃包括但不限于正十二烷或正十六烷。
本发明的第三个目的是提供一种降解石油烃的制剂，含有所述的戈登氏菌SCSIO19801作为活性成分。
本发明从中国南海岛礁表层海水分离获得具有石油烃降解功能的戈登氏菌SCSIO19801，其对多环芳烃类、液态烷烃、柴油等烃类化合物具有较好的降解功能，由于本发明的戈登氏菌SCSIO 19801来源于海洋，可以用于海洋的溢油污染进行原位修复，具有重要的意义。
戈登氏菌(Gordonia sp.)SCSIO 19801，已于2020年12月07日在中国微生物菌种保藏管理委员会普通微生物中心(CGMCC)保藏，保藏地址：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，邮政编码：100101，保藏号为：CGMCC No.21142。
附图说明
图1为戈登氏菌(Gordonia sp.)SCSIO 19801在LB培养基上的菌落形态(A)和电镜图(B)。
图2为戈登氏菌(Gordonia sp.)SCSIO 19801对多环芳烃、液态烷烃和柴油的降解率。
具体实施方式：
以下实施例是对本发明的进一步说明，而不是对本发明的限制。
实施例1：戈登氏菌(Gordonia sp.)SCSIO 19801菌株的筛选鉴定
配制无机盐培养基MSM(g/L)：K2HPO4，0.27；KH2PO4，0.35；NH4Cl，2.7；MgCl2·6H2O，0.1；CaCl2·2H2O，0.1；FeCl2·4H2O，0.009；MnCl2·4H2O，0.004；ZnCl2，0.0014；CoCl4·6H2O，0.001；(NH4)6Mo7O24·4H2O，0.001；pH 7.0。在无机盐培养基中加入1％(V:V)石油和柴油配制成富集培养基。将采集于南海岛礁表层的海水样品加入到富集培养基中，28℃条件下培养富集培养15天。以体积比1％接种量将富集培养物转接到上述新鲜富集培养基中，相同条件下进行培养，并重复转接富集培养3次。取第三次转接培养后的培养物涂布到LB培养基中，培养2-3天后从平板中挑取形态不同的的菌落，分离培养，获得分离纯化的菌株SCSIO19801。
1、菌株SCSIO 19801菌株形态特征
将单菌落划线接种于LB平板培养基上，28℃条件下培养2-3天。菌株SCSIO 19801菌落在LB平板上呈橙色，菌落为圆形，边缘规则。革兰氏阳性，透射电镜观察结果显示，该菌株为棒状，长度0.6-1.1μm，参见图1。最适合生长条件为：pH 7-9，28-35℃。
2、分子生物学鉴定
提取菌株SCSIO 19801的基因组DNA，利用27F/1492R通用引物扩增16S rRNA基因片段并测序，其序列如SEQ ID No.1所示。对16S rRNA基因序列在NCBI网站上进行比对，利用序列比对软件Clustal X 2.1软件和***发育分析软件MEGA7构建进化树。结果表明，本发明的菌株SCSIO 19801属于Gordonia属，与Gordonia terrae strain W3S2-1菌株相似性最高，相似度为100％，将其命名为戈登氏(Gordonia sp.)SCSIO 19801，该菌于2020年12月07日保藏于中国微生物菌种保藏管理委员会普通微生物中心(CGMCC)，地址：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，保藏编号：CGMCC No.21142。
实施例2：戈登氏菌(Gordonia sp.)SCSIO 19801菌株降解烷烃和多环芳烃能力的测定
配制无机盐培养基MSM并高温灭菌，分别加入终浓度为100mg/L的多环芳烃(菲、芘)、1％(V:V)柴油和液态烷烃(正十二烷、正十六烷)，得到测试培养基。加入1％体积比OD600＝0.6的戈登氏菌(Gordonia sp.)SCSIO19801菌液接入到上述测试培养基中，28℃条件下培养7天，通过GC-MS等方法对培养物中残余的石油烃进行检测。结果显示，戈登氏菌(Gordonia sp.)SCSIO 19801菌株对菲的降解率为83％，对芘的降解率为51％，对正十二烷的降解率为26％，对正十六烷的降解率为12％，对柴油的降解率为23％，参见图2。上述结果表明，本发明保藏的戈登氏菌(Gordonia sp.)SCSIO 19801菌株对石油烃具有优良的降解能力，可广泛用于石油烃降解及石油污染的修复，如石油污染的海洋水体修复等。
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<110> 中国科学院南海海洋研究所
<120> 一株石油烃降解功能菌SCSIO19801及其应用
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<211> 1344
<212> DNA
<213> 戈登氏菌SCSIO19801(Gordonia sp.)
<400> 1
ggccaccggc ttcgggtgtt accgactttc atgacgtgac gggcggtgtg tacaaggccc 60
gggaacgtat tcaccgcagc gttgctgatc tgcgattact agcgactccg acttcatggg 120
gtcgagttgc agaccccaat ccgaactgag actggcttta agggattcgc tccacctcac 180
ggtatcgcag ccctctgtac cagccattgt agcatgtgtg aagccctgga cataaggggc 240
atgatgactt gacgtcatcc ccaccttcct ccgagttgac cccggcagtc tcctgcaagt 300
ccccggcata acccgctggc aatacaggac aagggttgcg ctcgttgcgg gacttaaccc 360
aacatctcac gacacgagct gacgacagcc atgcaccacc tgtacaccaa ccacaaggga 420
acgactatct ctagccgcgt ctggtgtatg tcaaacccag gtaaggttct tcgcgttgca 480
tcgaattaat ccacatgctc cgccgcttgt gcgggccccc gtcaattcct ttgagtttta 540
gccttgcggc cgtactcccc aggcggggta cttaatgcgt tagctacggc acggatcccg 600
tgaaaaggaa cccacaccta gtacccaccg tttacggcgt ggactaccag ggtatctaat 660
cctgttcgct acccacgctt tcgctcctca gcgtcagtta ctacccagag acccgccttc 720
gccaccggtg ttcctcctga tatctgcgca tttcaccgct acaccaggaa ttccagtctc 780
ccctgtagta ctcaagtctg cccgtatcgc ctgcacgcct gcaattgagt tgcagaattt 840
cacagacgac gcgacaaacc gcctacgagc tctttacgcc cagtaattcc ggacaacgct 900
cgcaccctac gtattaccgc ggctgctggc acgtagttgg ccggtgcttc ttctccaggt 960
accgtcactt ccgcttcgtc cctggtgaaa gaggtttaca acccgaaggc cgtcatccct 1020
cacgcggcgt cgctgcatca ggcttgcgcc cattgtgcaa tattccccac tgctgcctcc 1080
cgtaggagtc tgggccgtgt ctcagtccca gtgtggccga tcaccctctc aggtcggcta 1140
cccgtcgtcg ccttggtagg ccattacccc accaacaagc tgataggccg cgggcccatc 1200
ccacaccgca aaagctttcc accaaccacc atgcgacagt tggtcatatc cggtattaga 1260
cccagtttcc caggcttatc ccagagtgca gggcagatca cccacgtgtt actcacccgt 1320
tcgccactcg agtacccagc aagc 1344
Claims (4)
1.戈登氏菌(Gordonia sp.)SCSIO 19801，保藏号为：CGMCC No.21142。
2.权利要求1所述的戈登氏菌SCSIO 19801在降解菲、芘、正十二烷、正十六烷、柴油中应用。
3.根据权利要求2所述的应用，其特征在于，是用戈登氏菌SCSIO 19801对菲、芘、正十二烷、正十六烷、柴油污染的海洋水体进行修复。
4.一种降解石油烃的制剂，其特征在于，含有权利要求1所述的戈登氏菌SCSIO 19801作为活性成分。
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