Ecophysiology and Phylogeny of Vacuolate, Nitrate-Accumulating Sulfur Bacteria

液泡是细胞内充满液体的膜结合空间，具有多种功能，包括储存。直到最近，它已经明确地指出，液泡不发生在细菌。然而，在最近的研究中，包括在PI实验室进行的nsf支持的研究中，在属于Beggiatoa属，Thioploca属和Thiomargarita属的大型细菌中发现了液泡。这些结构约占这些细胞生物体积的80%，据信与硝酸盐的积累和储存有关，整个细胞体积的硝酸盐平均为0.1至0.5摩尔（高于环境水平10,000倍）。储存的硝酸盐可以作为氧化剂，从而转化为氨，同时允许由硫化氢厌氧氧化产生生物上有用的能量。这些积累硝酸盐的细菌形成了一个单一的进化集群，从其他野外研究来看，它们可能在海洋氮循环中起着重要的定量作用。目前的研究将研究这些细菌的生态生理学和代谢，重点是Beggiatoa，并将液泡细菌的研究扩展到附着的、丝状的、硫刺菌样菌株，这些菌株在华盛顿海岸和其他地方的深海热液喷口的各种表面上占主导地位。将对附着形式进行研究，以确定它们与先前鉴定的液泡细菌之间的进化关系，并确定液泡是否在不同物种中起相同的生理作用。为了更深入地研究液泡、硝酸盐还原、硫化物氧化细菌的生态生理，将对蒙特雷峡谷富含硫化物的渗漏（900米深）中容易获得的Beggiatoa种群进行详细研究。它们使用氧气和硝酸盐作为氧化剂的能力，以及它们使用有机和无机细胞碳源的能力将被确定。由于这些细菌在蒙特雷峡谷渗漏沉积物的上部15厘米处占主导地位，因此这些研究将通过分析细菌的丰度与深度，结合硫化物、硫酸盐和氨的沉积物孔隙水剖面以及某些代谢抑制剂对沉积物过程的影响，获得有关细菌原位代谢的信息。液泡细菌从未在纯培养中生长过，但所研究的群体可以获得足够的纯度，以便进行化学，酶学和分子表征。研究人员将尝试克隆Beggiatoa中编码中心代谢过程的基因，并将采用分子程序来确定克隆的基因是否实际上来自Beggiatoa。这种类型的分析可能最终使人们有可能了解液泡形成或细菌巨人症的遗传基础。这提供了将这些看似理想的特性植入其他培养细菌的长期可能性

Vacuoles, fluid-filled membrane-bound spaces within cells, may serve a variety of functions, including storage. Until recently, it has been categorically stated that vacuoles do not occur in bacteria. In recent studies, however, including NSF-supported studies undertaken in the PI's laboratory, vacuoles have been discovered in large bacteria belonging to the genera Beggiatoa, Thioploca and Thiomargarita. These structures, which comprise roughly 80% of the biovolume of these cells, are believed to be involved in the accumulation and storage of nitrate, which averages 0.1 to 0.5 molar (10,000-fold above ambient levels) computed over the entire cell volume. The stored nitrate can serve as an oxidant, thereby being converted to ammonia while allowing the generation of biologically useful energy driven by the anaerobic oxidation of hydrogen sulfide. These nitrate-accumulating bacteria form a single evolutionary cluster, and, judging from field studies of others, they may play a quantitatively important role in the marine nitrogen cycle. The current research will examine the ecophysiology and metabolism of these bacteria, focusing on Beggiatoa, and will extend the study of vacuolate bacteria to attached, filamentous, Thiothrix-like strains that dominate the biomass on a variety of surfaces at deep-sea hydrothermal vents off the coast of Washington and elsewhere. Studies of the attached forms will be undertaken to determine the evolutionary relationships between them and the previously identified vacuolate bacteria, and to determine whether the vacuole serves the same physiological role in the diverse species. To delve more deeply into the ecophysiology of vacuolate, nitrate reducing, sulfide-oxidizing bacteria, Beggiatoa populations readily available from sulfide-rich seeps (900 m depth) in Monterey Canyon will be studied in detail. Their ability to use both oxygen and nitrate as oxidants as well as their ability to use organic vs. inorganic sources of cell carbon will be determined. Because these bacteria dominate the upper 15 centimeters of the Monterey Canyon seep sediments, these studies will yield information on the bacteria's in situ metabolism through analyses of their abundance vs. depth in conjunction with a study of sediment porewater profiles of sulfide, sulfate and ammonia and the influence of certain metabolic inhibitors on sediment processes. No vacuolate bacterium has ever been grown in pure culture, but the populations under study can be harvested in sufficient purity to allow chemical, enzymological and molecular characterizations. The investigators will attempt to clone the genes encoding the central metabolic processes in Beggiatoa and will employ molecular procedures to determine whether the cloned genes were actually derive from Beggiatoa. Analyses of this type may eventually make it possible to understand the genetic basis for vacuole formation or gigantism in bacteria. This offers the long-term possibility of engineering these seemingly desirable traits into other cultivated bacteria