海绵共栖细菌抗菌活性及其群体感应信号的研究

利用平板涂布法从南海海绵生物样品中分离海绵共栖细菌,以枯草芽孢杆菌、金黄色葡萄球菌、大肠杆菌及酿酒酵母作为实验用指示菌对海绵共栖细菌进行了抗菌活性筛选。利用气相质谱联用仪(GC-MS)建立了群体感应信号分子酰基高丝氨酸内酯类化合物(AHLs)的检测方法,分析了活性菌株粗提液中信号分子的产生及种类。同时采用16S rDNA同源性和系统发育分析对活性菌株进行种属鉴定和系统发生学研究。结果表明,在分离到的94株海绵共栖细菌中18株(占19.1%)细菌具有抗菌活性。通过GC-MS检测确定大多数活性菌株都含有AHLs,因此提出了海绵共栖细菌抗菌活性与群体感应之间可能存在必然联系的推测。通过细菌分类鉴定,结果显示具有抗菌活性的细菌大部分属于芽孢杆菌属(Bacillus)。     

样品保藏方式与时间对海洋底栖细菌及原生生物荧光计数效能的影响

DAPI荧光染色技术已广泛应用于浮游细菌及原生生物的定量研究,但对底栖生物的定量效能尚缺必要的研究。比较了冷冻和冷藏两种保藏方式以及保存时间对DAPI荧光计数底栖细菌、蓝细菌、硅藻、不同粒级自养小鞭毛虫(PNF)和异养小鞭毛虫(HNF)的影响。对黄海冷水团三个站位表层2cm底栖样品进行的4℃冷藏与?20℃冷冻避光保存的比较研究表明,两种保藏方式下两个站位的所有研究对象均无显著差异,但另一站位的PNF(2~5μm和5~10μm)的冷藏保存显著优于冷冻保存。对选取的另外两个站位(0~2cm和2~5cm分层)的样品经1个月和4个月冷藏保存后的分析表明,对于底栖细菌、蓝细菌、PNF(5~10 μm)、PNF(10 μm)、HNF(10 μm)和硅藻在保存1个月和4个月后的计数没有显著差异,而对于PNF(2～5 μm)、HNF(2～5 μm)、HNF(5～10 μm)保存4个月的数量明显低于保存1个月的样品,如其中一个站位的0～2cm分层的PNF(2～5 μm)丰度减少了47.4%,2～5cm分层的丰度减少了59.6%,HNF(2～5 μm)和HNF(5～10 μm)经4个月后丰度降为0。本研究表明,对底栖细菌、蓝细菌以及原生生物的定量计数可因样品、保藏方式及保存时间的不同而产生差异,因此对于底栖样品短期内宜采用避光、冷藏保存,并在带回实验室后尽快分析。     

青岛海洋趋磁球菌QH-3 的鞭毛特征

趋磁细菌(magnetotactic bacteria,MTB)是一类能够沿着磁力线运动的特殊细菌。MTB在胞内磁小体的导向下借助自身鞭毛进行趋磁趋氧运动,以到达适宜的生境。本研究在青岛潮间带沉积物中发现大量的趋磁球菌,通过光镜与透射电镜对其运动方式与显微结构进行观察。结果显示优势趋磁细菌为卵球形,大小为2.5μm×2.1μm,宽长比为0.85,将其命名为QH-3。QH-3为趋北型(North-seeking)MTB,运动速度88μm/s,细胞内两侧各有一条由棱柱形颗粒组成的磁小体链。QH-3在菌体同一侧有两个由直径62 nm±9 nm的鞘包裹7根鞭毛组成的鞭毛束。这种特殊的鞭毛结构与已报道的致病菌的不同,而与法国地中海趋磁球菌MO-1的相似。     

一株深海耐嗜压菌的细胞毒活性次生代谢产物研究

通过CCK8法检测分离自深海一株深海耐压菌Shewanella piezotolerans WP3次生代谢产物的细胞毒活性,研究了温度、压力、噬菌体对WP3次生代谢产物细胞毒活性的影响.发现WP3次生代谢产物具有良好细胞毒活性.低温因素可提高其细胞毒活性,在100μg/cm^3的含量下,代谢产物对肿瘤细胞Bel7402、CNE2、A549、HT1080细胞毒活性均有提高,其中对A549细胞毒活性提高了1.3倍.在最适温度（20℃）情况下,高压可降低其细胞毒活性.20 MPa培养条件下,代谢产物对4株肿瘤细胞均未表现出明显活性,而常压培养条件下对SW 480具有93.3%的抑制率,对HT1080抑制率表现出87.5%,对CNE2抑制率是79.8%.在低温情况下,压力因素对其细胞毒活性无影响,噬菌体对细胞毒活性影响不大.HPLC指纹图谱分析发现不同温度条件下,其次生代谢产物指纹图谱有变化,结合细胞毒活性结果分析,为后期分离鉴定细胞毒活性化合物提供指导.     

拮抗香蕉枯萎病菌海洋细菌的筛选和鉴定

从93份中国南部海域近海海泥样品中分离出1800多株海洋细菌,以对峙培养法,测定海洋细菌对香蕉枯萎菌的拮抗作用,再以琼脂划线培养法(agar streak)测定菌株的抑菌活性,结果表明:约55.6%的菌株对香蕉枯萎菌有一定的拮抗作用,其中58株菌株抑制香蕉枯萎菌生长所形成的抑菌带宽度大于10.0mm,抑菌作用较好,复筛后其中13个菌株形成的抑菌带宽在15.0mm以上。经反复转接和对峙培养,筛选出拮抗效果好、抑菌活性稳定的菌株TC-1,初步鉴定为芽孢杆菌属的细菌,TC-1对14种病原微生物都有较好的抑菌效果。     

黄海冷水团海域微型异养鞭毛虫对异养细菌和蓝细菌摄食作用的初步研究

2006年10月在黄海冷水团海域的三个站点开展了微型异养鞭毛虫、异养细菌和蓝细菌的密度和生物量调查,进行了微型异养鞭毛虫的现场摄食实验,通过荧光标记细菌法和消化系数法获得该海区微型异养鞭毛虫对异养细菌和蓝细菌的摄食率,并估算了微型异养鞭毛虫对异养细菌和蓝细菌现存量及生产力的摄食压。结果显示,微型异养鞭毛虫、异养细菌和蓝细菌的密度分别为0.36×103~1.13×103,0.39×106~1.13×106和0.04×104~3.74×104cells/cm3,温跃层以上明显高于底层。微型异养鞭毛虫对异养细菌的摄食率为5.33~14.89个/(HF·h),对蓝细菌的摄食率为0.26×102~23.10×10-2cells/(HF·h),摄食率随深度而下降。微型异养鞭毛虫每天能消耗9.27%~33.08%的异养细菌现存量和8.12%~16.09%的蓝细菌现存量。同时,微型异养鞭毛虫对异养细菌和蓝细菌的日摄食量各占它们生产力的2.66%~13.10%和8.12%~16.09%。研究表明微型异养鞭毛虫的摄食可能不是秋季黄海冷水团海域浮游细菌及其生产力的主归宿。     

Growth of marine bacteria and ammonium regeneration from substrates in different C: N ratios

Natural assemblages of marine bacteria were chosen in a batch culture experiments. The impact of varying nitrogen substrate concentrations and the substrate C:N ratios (C:N_S) on the bacterial C:N ratio (C:N_B), the bacterial growth efficiency (BGE) and ammonium regeneration was mainly examined. The C:N_S ratios varied from 5:1 (carbon limitation) to 40:1 (nitrogen limitation) with varying combinations of glucose and NO₃^-. The C:N_B ratio had positive relationship with the C:N_S ratio (r=0.93, n=8), whose value was 3.77 when the C:N_S ratio was 5:1 but increased to 6.47 when the C:NS ratio was 40:1. These results indicate that the C:N_B ratio is a potential diagnostic tool for determining the bacterial growth in natural waters controlled by either, carbon or nitrogen. BGE decreased with the declining nitrate concentration and negatively related to C:N_s (r=-0.51, n=8). The average value of BGE was 0.20. This value was a little lower than other reports, which could be induced by the nitrogen source used in our experiments. Finally, regeneration time of ammonium delayed with the increasing C:N_S ratio, which indicates that there were different metabolism mechanisms when bacterial growth was limited by carbon source and nitrogen source.     

A culture-dependent survey of thermophilic bacteria from hot springs in Xiamen area in China

Microbes are believed to play important roles in ecosystem function in many environments. The hot springs of Xiamen Island are close to the Xiamen Sea, and may have some characteristics different from those of inland hot springs. Microbes living in the hot springs of Xiamen may have new characteristics. However, little is known about microbial communities of hot springs close to the Xiamen Sea. A culturedependent survey of microbial population in the Xiamen hot springs was performed by using an approach combining total cellular protein profile identification and 16S rRNA gene sequencing. A total of 328 isolates of bacteria were obtained from liquid and sediment samples from the Xiamen hot springs, including neutrophilic thermophilic bacteria and moderately thermophilic acidophiles. Neutrophilic thermophilic bacteria, which grow at a temperature range of 55—90℃ including Rhodothermus marinus (Strain 1), Thermus thermophilus (Strain 2), Thermus thiopara (Strain 3), Geobacillus stearothermophilus (Strain 4), Geobacillus thermoleovorans (Strain 5), and Pseudomonas pseudoalcaligenes (Strain 6), were recovered by 2216E plates. Moderately thermophilic acidophiles, which can grow at temperatures above 50℃ and a pH range of 1.8—3.5 such as Alicyclobacillus acidoterrestris (Strain 8), Sulfobacillus acidophilus (Strain 9), and Sulfobacillus thermosulfidooxidans (Strain 10), were isolated on selective solid medium containing sulfur and Fe2+. Among these strains, Rhodothermus marinus, Thermus thermophilus and Geobacillus stearothermophilus are not only thermophiles, but also halophiles. One bacterium strain (Strain 6) shared 99% nucleotide sequence homology with Pseudomonas pseudoalcaligenes on the 16S rRNA gene sequence, but was quite different from Pseudomonas pseudoalcaligenes in biological characteristics, suggesting that it may represent a novel thermophilic species. Results indicated that various species of neutrophilic thermophiles and moderately thermophilic acidophiles were widely distributed in the Xiamen hot springs and that Rhodothermus marinus and Thermus thermophilus dominated the cultivable microbial community.     

盐田藻垫细菌的分离及其胞外多糖的初步研究

从连云港台南盐场采集藻垫,在去除底泥等杂物的藻类层中分离出5株不同的细菌(编号为A,B,c,D和E).经VITEK-32鉴定,A和B可能是鲁氏不动杆菌(Acinetobacter lwoffii)或琼氏不动杆菌(Acinetobacter junii),C为溶血巴斯德菌(Pasteurella haemolytica),D为尿素放线杆菌(Actinobacillus ureae),E未鉴定出结果.在所试验的盐度范围内,细菌A、B和C的最适生长盐度都为25,细菌D的最适生长盐度为50～100,细菌E的最适生长盐度为25～50.细菌A,B,C,D和E细菌胞外多糖产率分别为0.628,0.362,0.177,0.269,0.719 g/g细胞干质量.细菌A,B,C,D和E胞外多糖的糖醛酸质量分数分别为5.4%,9.9%,7.4%,9.8%和23.4%,均不含蛋白质,均无乳化.絮凝及凝胶活性.实验结果显示细菌胞外多糖是构成盐田藻垫胶质的重要组成部分,但从其物化性质看,细菌胞外多糖对藻垫的胶结和黏附性没有直接的影响.     

Isolation and identification of bacteria associated with the surfaces of several algal species

We conducted this study to assess the diversity of bacteria associated with the surfaces of algae based on 16S rDNA sequence analyses.Twelve strains of bacteria were obtained from the surfaces of the following four species of algae:Gracilaria textorii,Ulva pertusa,Laminaria japonica,and Polysiphonia urceolata.The isolated strains of bacteria can be divided into two groups:Halomonas and Vibrio,in physiology,biochemical characteristics and 16S rDNA sequence analyses.The phylogenetic tree constructed based on ...