No description available.

No description available.

No description available.

No description available.

No description available.

Symbiotic relationships of bacteria with higher organisms are commonly observed in nature; however, the functional role of these relationships is only rarely understood. This is particularly evident in epibiotic bacterial associations in the marine environment where the bacteria are often a diverse ensemble of microorganisms, thus complicating the identification of the functionally important members. Classical microbiological techniques, relying primarily on culturing these organisms, have provided an incomplete picture of these relationships. Molecular genetic techniques, focusing on the analyses of bacterial 16S rRNA sequences cloned directly from natural microbial populations, are now available which allow a more thorough examination of these associated bacterial populations. This study sought to characterize the epibiotic bacterial population associated with a very unique organism, Alvinella pompejana, using such a molecular approach. Alvinella pompejana is a polychaetous annelid that inhabits active deep-sea hydrothermal vent sites along the East Pacific Rise. This worm colonizes the walls of actively venting high temperature chimneys and is thought to be one of the most thermotolerant metazoans known. The chimney environment is characterized by high concentrations of sulfide and heavy metals in the vicinity of the worm colonies. A morphologically diverse epibiotic microflora is associated with the worm's dorsal integument, with a highly integrated filamentous morphotype clearly dominating the microbial biomass. It has been suggested that this bacterial population participates in either the nutrition of the worm or in detoxification of the worm's immediate environment; however, previous studies have been unable to confirm such a role. The primary goal of this study is to phylogenetically characterize the dominant epibionts through the analysis of 16S rRNA gene sequences. Nucleic acids were extracted from bacteria collected from the dorsal surface of Alvinella pompejana. 16S rRNA genes were amplified with universal bacterial primers by the polymerase chain reaction (PCR). These genes were subsequently cloned and the resulting clone library was screened by restriction fragment length polymorphism (RFLP) analysis to identify unique clone types. Thirty-two distinct clone families were found in the library. Four of these families were clearly dominant, representing over 65% of the library. The main assumption in this study is that the numerical dominance of the phylotypes in the starting population will be reflected in the clone library. Thus, representative clones from the four most abundant clone families were chosen for complete gene sequencing and phylogenetic analysis. These gene sequences were analyzed using a variety of phylogenetic inference methods and were found to be related to the newly established epsilon subdivision of the Proteobacteria. In future studies, these gene sequences will be used to construct specific oligodeoxynucleotide probes which can be used to confirm the morphology of the clone types in the epibiont population. / Graduation date: 1995

Thesis (M.S.)--Oregon State University, 2007. / Printout. Includes bibliographical references (leaves 70-73). Also available on the World Wide Web.

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Siderophores are secreted by marine bacteria to increase Fe uptake when Fe is limiting but are not produced when sufficient Fe is present to saturate growth. These results are well established in laboratory batch cultures of a number of isolates obtained from the open sea. Little is known, however, regarding the kinetics of siderophore secretion by heterotrophic bacteria in response to transients in Fe deprivation and resupply. We examined growth, hydroxamate siderophore concentration, and electron transport chain activity (a biochemical measure of Fe nutritional state) of Pseudoalteromonas haloplanktis, a representative gamma-proteobacterium from the Fe deficient region of the subarctic Pacific Ocean. Hydroxamate concentration was roughly 5-fold higher in batch cultures grown in low than in high Fe medium. Iron injection to the low Fe cultures repressed hydroxamic acid production and increased growth and ETC activity. Steady-state hydroxamate concentration in the chemostat increased 5-fold as Fe-limited growth rate declined from 9.8 to 2.8 d -1. This increase compounded to a 2.8-fold change in hydroxamates cell-1 reflecting the greater costs of growth at low Fe. Three types of Fe perturbation were made to Fe-limited chemostat cultures: 1) A switch perturbation that decreased the dilution rate of the chemostat-by ∼3-fold caused a transient increase in cell density that subsequently declined to a new steady state level. Hydroxamate concentration increased linearly over the same time. 2) A transient addition of dissolved Fe increased the total hydroxamate concentration in the chemostat within 1-3 hours which was followed by a decrease and then subsequent increase as the cells re-entered Fe-limitation. Dilution rate affected the response. Normalized to bacteria density, hydroxamate concentration remained constant for the first 2 hours after the Fe addition and then declined and returned to pre-infusion levels. Thus, Fe addition stimulated siderophore production by increasing the density of bacteria, which continued to secrete hydroxamates at a Fe-limited rate. 3) A continuous addition of low levels of dissolved Fe increased bacteria density and siderophore concentration. The net secretion rate of siderophores was proportional to the increase in Fe supply rate to the chemostat. At high Fe concentration, hydroxamate concentration declined to undetectable levels as the bacteria became Fe-sufficient and C-limited. Siderophore secretion by Fe-limited P. haloplanktis was repressed after 2 hours (corresponding roughly to 1-2 cell generations) following Fe re-supply.

The lipopolysaccharide (LPS) translocation process was studied by a pulse-chase experimental procedure designed to follow the fate of newly synthesized LPS through the cell wall fractions of the marine bacterium Alteromonas haloplanktis, strain 214, variant 3 (ATCC 19855). It was determined that newly synthesized LPS I (the prominent LPS species under the conditions employed) initially enters an LPS fraction which can be released from the cell wall along with periplasmic material. This is followed by rapid entry of the newly synthesized LPS into a loosely bound LPS fraction (released from the cell wall by washing with NaCl) before final insertion into the outer membrane. The inhibition of protein synthesis with chloramphenicol was observed to have little or no effect on the rate of translocation of newly synthesized LPS. In contrast, both the respiratory inhibitor NaCN (10 mM) and the proton ionophore 3,3',4',5-tetrachlorosalicylanilide (TCS, 30 (mu)M), if added during the chase period, resulted in an immediate and complete inhibition of further LPS translocation. The results suggest an independence of the LPS transloction process from continued protein synthesis, but a requirement for an energy source. It was observed that newly synthesized LPS sedimented to a lower density position than previously formed LPS on sucrose density gradients, and displayed a faster migration rate during electrophoresis on sodium dodecyl sulfate polyarylamide gels (SDS-PAGE). This indicated compositional differences between the two LPS populations. Extended electrophoresis on SDS-PAGE separated both newly synthesized and previously formed LPS I into a number of distinct radiolabelled peaks and shoulders, indicating compositional microheterogeneity within each species. The fate of added radiolabelled galactose was also investigated. A one minute pulse with {('14)C} galactose resulted in a preferential labelling of components associated with the cell wall such that 57.3% of the total w