【作者】 张喆；

【导师】 唐学玺；

【作者基本信息】 中国海洋大学 ， 生态学， 2008， 博士

【摘要】 本文采用分级稀释培养计数法和表面荧光显微镜计数法,分别于2006年8月(夏季)、2006年12月(冬季)、2007年4月(春季)和2007年10月(秋季),四次对山东近岸海域可培养异养细菌数量,海洋浮游细菌丰度、生物量及海洋浮游病毒丰度分布特征进行了调查研究,同时分析了夏、冬两季海洋细菌、病毒与环境生态因子之间的相关性,探讨了环境因子对浮游细菌及病毒分布的影响,以期为山东近岸海域微生物分布状况提供基础数据,为其后的海洋微生物生态学研究提供参考。同时,采用ARDRA方法分析了胶州湾内站位QD08及威海近岸站位QD13表层沉积物细菌多样性及其季节变化。1.可培养异养细菌的分布特征调查海区四个季节可培养异养细菌数量介于1.01×104~6.27×107 CFU/L,最大值出现在夏季小岛湾外站位QD15底层水中,最小值则出现在冬季QD03站位中层水,各季节可培养异养细菌分布为夏季>冬季>秋季>春季。四个季节均表现出沿岸可培养异养细菌数量大于远岸数量的趋势;夏、秋、冬季节北部海区可培养异养细菌数量要高于南部海区,而冬季则刚好相反。在垂直方向上,除冬季表现出各水层可培养异养细菌数量相似的规律外,其它三个季节则没有明显的垂直分布规律。夏、冬季节可培养异养细菌数量与温度及海水叶绿素a含量(Chla)没有相关性,而与总氮(N)、总磷(P)及磷酸盐(PO43-)呈显著相关,这说明总N、总P及PO43-可能是夏、冬季节可培养异养细菌数量分布的主要限制因子,而温度在可培养异养细菌数量分布上并没有起到限制性作用。2.浮游细菌丰度及生物量的分布特征调查海区四个季节浮游细菌丰度及生物量分别介于9.76×108~1.55×1010 cells/L和14.79~236.49μg/L。两者无论是在时间还是空间都表现出了相似的分布特征。浮游细菌丰度及生物量季节分布均为春季>夏季>秋季>冬季,冬季浮游细菌丰度及生物量明显低于其他三个季节,冬季的低温可能是造成这一现象的主要原因。各站位不同季节浮游细菌丰度及生物量有明显的垂直分布规律,但在夏季QD06及QD14站位可以观察到浮游细菌丰度的变化受到温度跃层的影响。调查海区春、秋、冬季三层水均呈现出近岸浮游细菌丰度及生物量大于远岸的趋势,且一般在胶州湾或小岛湾、丁字湾附近出现高值区;夏季表层与底层水浮游细菌丰度及生物量亦表现出近岸区域大于远岸区域的趋势。夏、冬季节各站位水柱浮游细菌丰度及生物量与温度、总有机碳及叶绿素a含量均呈现显著正相关关系,而与溶解氧、PO43-呈显著负相关关系,因此可以认为调查海区夏、冬季节浮游细菌分布同时受到上述几种环境生态因子的影响,而由于海区营养盐充足,减弱了营养盐对浮游细菌季节分布的限制作用。3.浮游病毒丰度的分布特征调查海区各季节浮游病毒丰度介于1.71×109~3.40×1011 VLPs/L,其季节分布规律较明显,为冬季>春季>夏季>秋季,海水盐度及溶解氧含量的季节变化是造成这一现象的主要原因。不同站位各季节浮游病毒丰度没有明显的垂直分布规律,四个季节各水层浮游病毒丰度基本随着由沿岸区域向外海延伸,呈现出下降趋势。夏、冬季节山东近岸浮游病毒丰度与海水中硝酸盐含量呈显著正相关关系,而与浮游细菌丰度及叶绿素a含量呈现弱的负相关性,说明海水中营养盐含量的变化对浮游病毒丰度有较大影响。冬季较高的营养盐含量有可能增加了细菌体内病毒的释放量,而病毒对其寄主——浮游细菌的裂解作用则可能是导致两者呈现出负的相关性的原因。4.QD08 / QD13站位表层沉积物细菌多样性分为夏、冬、春、秋四个季节对山东近岸海域表层沉积物细菌多样性进行了研究。利用提取纯化的沉积物样品总DNA为模版,扩增了沉积物样品中细菌的16S rDNA片段,并构建了相应的文库。从QD08站位四个季节沉积物样品中分别得到37、48、48及39个具有不同酶切图谱的阳性克隆子,而QD13站位四个季节表层沉积物样品中具有不同酶切图谱的阳性克隆子则分别为33、29、57及57个。通过对构建的8个文库克隆子的ARDRA分析,探讨了文库种群的多样性,结果表明:在所构建的8个16S rDNA克隆文库中,A8S、D8S、A13S和B13S 16S rDNA克隆文库具有较高的覆盖率,可以认为上述4个文库能够反映沉积物样品细菌多样性;B8S、C8S、C13S及D13S克隆文库文库覆盖率均低于80%,可能需要更多的克隆子来反映沉积物样品细菌多样性。从细菌多样性季节变化来看,两个站位各季节沉积物样品细菌主要属于变形杆菌(Proteobacteria)类群,且γ-Proteobacteria亚群属于优势种类,同时也检测到α-/γ-/δ-/ε-Proteobacteria亚群的存在,表明变形杆菌是海洋沉积物中的主要类群。同时在QD08站位表面沉积物样品发现的还有浮霉菌(Plancomycene)、放线菌( Actinobacteria )、芽单胞菌( Gemmatimonadetes )、酸杆菌(Acidobacteriaceae)、硝化螺旋菌(Nitrospirales)、疣微菌(Verrucomicrobial)、拟杆菌(Bacteroidetes)和绿弯菌(Chloroflexi)几大类群。QD13站位沉积物样品细菌类群与QD08站位略有不同,主要包含浮霉菌(Plancomycene)、芽单胞菌(Gemmatimonadetes)、酸杆菌(Acidobacteriaceae)、硝化螺旋菌(Nitrospirales)、拟杆菌(Bacteroidetes)、绿弯菌(Chloroflexi)、高G + C含量革兰氏阳性菌放线菌(Actinobacteria)和低G + C含量的革兰氏阳性菌梭菌(Clostridiales)几类。与QD13站位相比,QD08站位沉积物细菌多样性随季节变化不大,可能是由于QD08站位沉积物受陆地影响较小,沉积物环境季节变化不大。更多还原

【Abstract】 Based on four cruises from August, 2006 to October, 2007, the distribution patterns of cultivable heterotrophic bacteria biomass (CHBB), virioplankton and bacterioplankton abundance (BA) in coastal waters of Shandong were studied. Moreover, the relationships between them and the environmental ecological factors, temperature, dissolved oxygen (DO), chlorophyll a (Chl a), total N, total P, total organic carbon, NO3- and PO43- in summer and winter were also analyzed. At the same time, bacterial diversity in sediments of station QD08 and QD13 were also investigated used the method of ARDRA.1. Distribution of cultivable heterotrophic bacteria (CHBB)Cultivable heterotrophic bacteria biomass fluctuated between 1.01×104CFU/L and 6.27×107CFU/L in the four seasons. The highest number was detected in the substrate waters of station QD15 in summer, while the CHBB was lowest in middle layer water of station QD03 in winter. The highest CHBB in water columns in different stations was recorded during summer, the second highest CHBB occurred in winter and the lowest was in spring. The CHBB decreased with the distance far away from the coast on most of transects in summer, winter and august. There was no distinct spatial distribution pattern of the CHBB in vertical section.The CHBB has highly significant correlations (P < 0.01) with total nitrogen, phosphorus and PO43-, which indicated that the CHBB might be influenced by these environmental factors in summer and winter. No correlation was detected between CHBB and temperature and the Chl a.2. Distribution of bacterioplankton abundance and biomassThe abundance and biomass of bacterioplankton in coastal waters of Shandong was 9.76×108~1.55×1010cells/L and 14.79~236.49μg/L respectively. The distribution of bacterioplankton abundance and biomass almost demonstrated the same temporal and spatial pattern. The bacterioplankton abundance and biomass were much lower in winter than in the other three seasons. The highest bacterioplankton abundance and biomass in water columns were occurred in spring, while the second in summer. There was no distinct spatial distribution pattern of the bacterioplankton abundance and biomass in vertical section. The bacterioplankton abundance and biomass were higher in the coastal line area in spring, august and winter. Most of the highest bacterioplankton abundance areas were around Jiaozhou bay, Xiaodao bay and Dingzi bay.The correlations were highly significant between bacterioplankton abundance, biomass and temperature, total organic carbon and Chl a in summer and winter. At the mean time, bacterioplankton and biomass were negative correlated with DO and PO43-. The distribution of bacterioplankton abundance and biomass were affected by the environmental ecological factors above.3.Distribution of virioplankton abundanceThe virioplankton abundance was between 1.71×109 VLPs/L and 3.40×1011 VLPs/L in the four cruises. The highest virioplankton abundance was recorded during winter, the second highest virioplankton abundance occurred in spring and the lowest in august. Salinity and DO content in sea water were regarded as the main factors which leaded to the seasonal variation of virioplankton abundance. The distribution of virioplankton abundance in the four seasons almost demonstrated the same pattern, which showed higher abundance in the costal line.Virioplankton abundance was significant positive correlated with NO3- content in the sea water, and was weak negative correlated with bacterioplankton abundance and Chl a. NO3- content might be the primary factor which effected the distribution of virioplankton abundance in summer and winter.4.Bacterial diversity in the sediment of QD08 and QD13Bacterial diversity in the sediment of stations QD08 and QD13 were studied in summer, winter, and spring, august. 16S rDNA fragments of bacteria were amplified and libraries were constructed based on the purified DNA extracted from the sediments of station QD08 and QD13. In the station of QD08, the numbers of positive clones which have different enzyme digest maps were 37, 48, 48 and 39. The numbers of positive clones were 33, 29, 57 and 57 respectively in the station of QD13. The 16S rDNA sequences which these clones contented were than sequedced. The results indicated that the coverage C of A8S, D8S, A13S and B13S libraries are very high, while the coverage C of B8S, C8S, C13S and D13S libraries all below 80%.In the station QD08, the sequenced clones fell into nine major lineages of the domain bacteria: the alpha, gamma, delta and epsilon Proteobacteria, Plancomycene, Actinobacteria, Gemmatimonadetes, Acidobacteriaceae, Nitrospirales, Verrucomicrobial, Bacteroidetes and Chloroflexi. The sub class of Proteobacteria, especially gamma Proteobacteria predominated in this station. The results suggested that these clones were very diverse in phylogeny. In the station of QD13, the domain bacteria included the alpha, gamma, delta and epsilon Proteobacteria, Plancomycene, Gemmatimonadetes, Acidobacteriaceae, Nitrospirales, Bacteroidetes, Chloroflexi, Actinobacteria and Clostridiales.Compared to the station QD13, seasonal variation of bacteria diversity in the sediment of QD08 seemed to be very small. This phenomenon might indicate that the sediment environment did not change much during the year.更多还原