Time of Flight-Secondary Ion Mass Spectrometry on isolated extracellular fractions and intact biofilms of three species of benthic diatoms.
ABSTRACT Time of Flight-Secondary Ion Mass Spectrometry (ToF-SIMS) was used to study compositional characteristics of Extracellular Polymeric Substances (EPS) and compared these to characteristics of the EPS-matrix of intact diatom biofilms. Three benthic diatoms species were investigated, Cylindrotheca closterium, Navicula mutica and Nitzschia cf. brevissima. Comparison of the ToF-SIMS spectra of sequentially extracted EPS-fractions by cluster analysis and multidimensional scaling analysis (MDS) indicated that soluble and bound EPS were not distinguishable based on their ion spectra. On the contrary the water insoluble bicarbonate soluble (WIBS)-EPS-fraction formed a distinct cluster showing that this material was compositionally different from the other EPS-fractions. Ion spectra of the EPS-fractions were dissimilar to results obtained from intact biofilms. This suggested that during the extraction procedure, the structure of the EPS irreversibly changed, which alters the fragmentation patterns of the extracellular surface layer. Furthermore, from the examination of the positive ion spectra it was shown that the overall composition of EPS in the intact biofilms was different between diatom species. In spite of these differences, several common peak patterns were shared between different species. This suggests the presence of common structural components in the EPS of these diatoms that may play a role in building the surface EPS-layer.
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ABSTRACT: Atomic Force Microscopy (AFM) resolved the topography and mechanical properties of two distinct adhesive mucilages secreted by the marine, fouling diatom Craspedostauros australis. Tapping mode images of live cells revealed a soft and cohesive outer mucilage layer that encased most of the diatom's siliceous wall, and force curves revealed an adhesive force of 3.58 nN. High loading force, contact mode imaging resulted in cantilever 'cleaned' cell walls, which enabled the first direct observation of the active secretion of soft mucilage via pore openings. A second adhesive mucilage consisted of strands secreted at the raphe, a distinct slit in the silica wall involved in cell-substratum attachment and motility. Force measurements revealed a raphe adhesive strand(s) resistant to breaking forces up to 60 nN, and these strands could only be detached from the AFM cantilever probe using the manual stepper motor.Protist 04/2002; 153(1):25-38. · 4.14 Impact Factor
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ABSTRACT: Marine benthic diatoms excrete large quantities of extracellular polymeric substances (EPS), both as a function of their motility system and as a response to environmental conditions. Diatom EPS consists predominantly of carbohydrate-rich polymers and is important in the ecology of cells living on marine sediments. Production rates, production pathways, and monosaccharide composition of water-soluble (colloidal) carbohydrates, EPS, and intracellular storage carbohydrate (glucans) were investigated in the epipelic (mud-inhabiting) diatoms Cylindrotheca closterium (Ehrenburg), Navicula perminta (Grün.) in Van Heurck, and Amphora exigua Greg. under a range of experimental conditions simulating aspects of the natural environment. Cellular rates of colloidal carbohydrate, EPS, and glucan production were significantly higher during nutrient-replete compared with nutrient-limited growth for all three species. The proportion of EPS in the extracellular carbohydrate pool increased significantly (to 44%–69%) as cells became nutrient limited. Cylindrotheca closterium produced two types of EPS differing in sugar composition and production patterns. Nutrient-replete cells produced a complex EPS containing rhamnose, fucose, xylose, mannose, galactose, glucose, and uronic acids. Nutrient-limited cells produced an additional EPS containing mannose, galactose, glucose, and uronic acids. Both EPS types were produced under illuminated and darkened conditions. 14C-labeling revealed immediate production of 14C-glucan and significant increases in 14C-EPS between 3 and 4 h after addition of label. The glucan synthesis inhibitor 2,6-dichlorobenzonitrile significantly reduced 14C-colloidal carbohydrate and 14C-EPS. The glucanase inhibitor P-nitrophenyl β-d-glucopyranoside resulted in accumulation of glucan within cells and lowered rates of 14C-colloidal and 14C-EPS production. Cycloheximide prevented glucan catabolism, but glucan production and EPS synthesis were unaffected.Journal of Phycology 03/2004; 40(2):293 - 304. · 2.24 Impact Factor
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ABSTRACT: Biofilms dominated by pennate diatoms are important in fields as diverse as ship biofouling and marine littoral sediment stabilization. The architecture of a biofilm depends on the fact that much of its mass consists of extracellular polymers. Although most illuminated biofilms in nature are dominated by phototrophs, they also contain heterotrophic bacteria. Given the close spatial association of the two types of organisms, cell-cell interaction is likely. Fluorophore-conjugated lectins were used to demonstrate the sites of the various extracellular polymers in three species of diatoms. Based on their lectin staining properties, the polymers in different species appeared to be similar, but their involvement in the process of attachment to a surface differed. In a coculture Pseudoalteromonas sp. strain 4 or its sterilized spent medium reduced the ability of Amphora coffeaeformis and Navicula sp. strains 1 and D to adhere, inhibited motility, and caused agglutination and eventually diatom cell lysis. Diatoms could be protected from the negative effects of the bacterial spent medium if D-galactose or mannan was included in the incubation medium. The active principle of the spent medium is probably a lectin/agglutinin that is able to bind to the extracellular polymers of the diatoms that are involved in adhesion and motility. Awareness of interactions of this type is important in the study of natural biofilms.Applied and Environmental Microbiology 02/2005; 71(1):428-35. · 3.68 Impact Factor