趋磁细菌AMB-1培养条件优化及磁小体变化过程研究

趋磁细菌(Magnetotactic bacteria)的研究是国际微生物学研究热点之一。趋磁细菌体内含有纳米单磁畴的氧化铁/硫化铁(Fe3O4或Fe3S4)晶体，称为磁小体。由于趋磁细菌营养条件要求苛刻，在环境中需要微好氧条件，且营养类型属于化能自养，使得培养趋磁细菌时常遇到问题。 本研究首先通过正交试验优化趋磁细菌AMB-1菌株培养条件，在培养条件铁源为奎尼酸铁0.02 mmol/L，装瓶量75% ，pH值6.7，温度25 ℃时，AMB-1 OD600达到0.440(1.166×109 cells/ml)。同时运用磁收集传代法，使带有磁小体的AMB-1细胞比例占95％以上(Cmag值稳定在1.9-2.0)。 在AMB-1具有较好的生物量，同时又具有较好的含磁小体细胞比例后，研究磁小体的变化过程。通过透射电镜观察磁小体变化过程，发现培养24 h细菌体内已有较小晶体形成(平均27 nm，n=188)且沿长轴分布；48 h晶体长大(平均43 nm，n=203)且形成分段链沿长轴排列；72 h晶体进一步成熟(平均50 nm，n=191)仍以分段链沿长轴排列；随后细菌逐渐衰亡磁小体变小，168 h可见部分自溶细菌中仍有磁小体链(平均37 nm，n=186)；192 h细菌自溶磁小体链(平均33 nm，n=184)分散到环境中。 通过透射电镜在细胞水平上研究趋磁细菌细胞分裂时发现，磁小体在细菌分裂时采用两种分离方式：一种为磁小体分配到两个子细胞；另一种为磁小体只分配到一个子细胞。无磁小体的子细胞，在随后的生长过程又分为两种情况：一种为细胞逐渐产生磁小体，另一种为不再产生磁小体。这种现象的发现，解释了随着传代次数的增多，细菌磁性有所下降的原因(Cmag值降低)。 在对趋磁细菌磁小体合成机制的研究中，常使用基因敲除的办法获得缺陷型，并与野生型对比进行研究。但是，利用基因敲除获得缺陷型不仅操作繁琐并且所得缺陷型不稳定。本研究利用特殊的磁富集传代法，先将带有磁小体的菌体收集并连续传代，筛选获得了高磁菌株；利用这种方法，收集不含磁小体的菌体并连续传代，筛选获得了无磁菌株。 趋磁细菌磁小体在医疗、环保等领域具有广阔应用价值，但是目前由于趋磁细菌难以大规模培养，并且磁小体纯化存在成本高等原因，将磁小体真正实际应用尚有一段距离。通过研究磁小体在趋磁细菌中的变化过程发现，AMB-1菌株在培养192 h后自溶，并且磁小体随着细胞的破碎释放到环境中去。

Characterization of a homogeneous taxonomic group of marine magnetotactic cocci within a low tide zone in the China Sea

Magnetotactic bacteria are a heterologous group of motile prokaryotes, ubiquitous in aquatic habitats and cosmopolitan in distribution. Here, we studied the diversity of magnetotactic bacteria in a seawater pond within an intertidal zone at Huiquan Bay in the China Sea. The pond is composed of a permanently submerged part and a low tide subregion. The magnetotactic bacteria collected from the permanently submerged part display diversity in morphology and taxonomy. In contrast, we found a virtually homogenous population of ovoid-coccoid magnetotactic bacteria in the low tide subregion of the pond. They were bilophotrichously flagellated and exhibited polar magnetotactic behaviour. Almost all cells contained two chains of magnetosomes composed of magnetite crystals. Intriguingly, the combination of restriction fragment length polymorphism analysis (RFLP) and sequencing of cloned 16S rDNA genes from the low tide subregion samples as well as fluorescence in situ hybridization (FISH) revealed the presence of a homogenous population. Moreover, phylogenetic analysis indicated that the Qingdao Huiquan low tide magnetotactic bacteria belong to a new genus affiliated with the alpha-subclass of Proteobacteria. This finding suggests the adaptation of the magnetotactic bacterial population to the marine tide.

Isolation and characterization of a marine magnetotactic spirillum axenic culture QH-2 from an intertidal zone of the China Sea

Magnetotactic bacteria (MTB) are ubiquitous in aquatic habitats. Because of their fastidious requirements for growth conditions, only very few axenic MTB cultures have been obtained worldwide. In this study, we report a novel marine magnetotactic spirillum axenic culture, designated as QH-2, isolated from the China Sea. It was able to grow in semi-solid or liquid chemically defined medium. The cells were amphitrichously flagellated and contained one single magnetosome chain with an average number of 16 magnetosomes per cell. Phosphate and lipid granules were also observed in the cells. Both rock magnetism and energy-dispersive X-ray spectroscopy characterizations indicated that the magnetosomes in QH-2 were single-domain magnetites (Fe3O4). QH-2 cells swam mostly in a straight line at a velocity of 20-50 mu m/s and occasionally changed to a helical motion. Unlike other magnetotactic spirilla. QH-2 cells responded to light illumination. As a consequence of illumination, the cells changed the direction in which they swam from parallel to the magnetic field to antiparallel. This response appears to be similar to the effect of an increase in [O-2]. Analysis of the QH-2 16S rRNA sequence showed that it had greater than 11% sequence divergence from freshwater magnetotactic spirilla. Thus, the marine QH-2 strain seems to be both phylogenetically and magnetotactically distinct from the freshwater Magnetospirillum spp. studied previously. (C) 2010 Elsevier Masson SAS. All rights reserved.