Characterization of PHB storage in activated sludge extended filamentous bacteria by automated colour image analysis

The storage of poly-β-hydroxybutyrate (PHB) in extended filamentous bacteria from activated sludge was monitored by Sudan Black staining: PHB granules were blue in the reddish filaments counterstained by safranin. By quantitative image analysis of colour images grabbed on an optical microscope, the distribution of the PHB loading of the extended filaments was estimated by determination of the proportion of blue pixels of their skeleton. The method was applied for different feed compositions to demonstrate its ability to monitor the PHB synthesis and storage capacity of filamentous bacteria in mixed cultures. Fast PHB storage, within a few hours, could be observed with acetate-based feeding solutions but the storage rate decreased with more complex feeds (meat extract based feed, wastewater).     

Morphogenesis in a community of filamentous cyanobacteria

Reversible differentiation was experimentally discovered in a community of modern filamentous cyanobacteria Oscillatoria terebriformis. Splitting of the initially uniform community into differentiated parts (strands, multiradiate aggregates, networks, etc.) occurs only for the duration of a function facilitating the activity of this community as an integral unit. The structures are formed as a result of regrouping of the filaments, without their specialization. A morphologically regulatory system (polygonal network) was found to develop under the impact of extreme factors. The levels of structural organization of filamentous cyanobacteria and multicellular eukaryotes were compared (individual cells in a filament—cell organelles; filaments—individual cells; community—organism), and the similarities and differences in morphogenesis of these groups were analyzed using the data on the embryonic regulation in multicellular eukaryotes. Spatial information in morphogenesis was shown to result not from direct realization of an inherited program but is created by the elements of integral organisms (cells and filaments) in the course of development.     

Filamentous Hydrous Ferric Oxide Biosignatures in a Pipeline Carrying Acid Mine Drainage at Iron Mountain Mine,California

A pipeline carrying acidic mine effluent at Iron Mountain, CA, developed Fe(III)-rich precipitate caused by oxidation of Fe(II)_aq. The native microbial community in the pipe included filamentous microbes. The pipe scale consisted of microbial filaments, and schwertmannite (ferric oxyhydroxysulfate, FOHS) mineral spheres and filaments. FOHS filaments contained central lumina with diameters similar to those of microbial filaments. FOHS filament geometry, the geochemical environment, and the presence of filamentous microbes suggest that FOHS filaments are mineralized microbial filaments. This formation of textural biosignatures provides the basis for a conceptual model for the development and preservation of biosignatures in other environments.     

Carbonate fabrics in the modern microbialites of Pavilion Lake: two suites of microfabrics that reflect variation in microbial community morphology,growth habit,and lithification

Modern microbialites in Pavilion Lake, BC, provide an analog for ancient non‐stromatolitic microbialites that formed from in situ mineralization. Because Pavilion microbialites are mineralizing under the influence of microbial communities, they provide insights into how biological processes influence microbialite microfabrics and mesostructures. Hemispherical nodules and micrite–microbial crusts are two mesostructures within Pavilion microbialites that are directly associated with photosynthetic communities. Both filamentous cyanobacteria in hemispherical nodules and branching filamentous green algae in micrite–microbial crusts were associated with calcite precipitation at microbialite surfaces and with characteristic microfabrics in the lithified microbialite. Hemispherical nodules formed at microbialite surfaces when calcite precipitated around filamentous cyanobacteria with a radial growth habit. The radial filament pattern was preserved within the microbialite to varying degrees. Some subsurface nodules contained well‐defined filaments, whereas others contained only dispersed organic inclusions. Variation in filament preservation is interpreted to reflect differences in timing and amount of carbonate precipitation relative to heterotrophic decay, with more defined filaments reflecting greater lithification prior to degradation than more diffuse filaments. Micrite–microbial crusts produce the second suite of microfabrics and form in association with filamentous green algae oriented perpendicular to the microbialite surface. Some crusts include calcified filaments, whereas others contained voids that reflect the filamentous community in shape, size, and distribution. Pavilion microbialites demonstrate that microfabric variation can reflect differences in lithification processes and microbial metabolisms as well as microbial community morphology and organization. Even when the morphology of individual filaments or cells is not well preserved, the microbial growth habit can be captured in mesoscale microbialite structures. These results suggest that when petrographic preservation is extremely good, ancient microbialite growth structures and microfabrics can be interpreted in the context of variation in community organization, community composition, and lithification history. Even in the absence of distinct microbial microfabrics, mesostructures can capture microbial community morphology.     

Photocontrol of growth pattern in a tissue isolated from the gametophytes of bracken fern

A tissue isolated from the gametophytes of bracken fern, Pteridiumaquilinum grew as unbranched filaments in red or green light,or in complete darkness. Irradiation with blue or white lightled to a shift from filamentous to two-dimensional pattern ofgrowth in the tissue. (Received February 13, 1969; )     

Enrichment and identification of large filamentous sulfur bacteria related to Beggiatoa species from brackishwater ecosystems of Tamil Nadu along the southeast coast of India

Beggiatoa species are filamentous sulfide-oxidizing bacteria belonging to the family Beggiatoaceae that contains several largest bacteria known today. These large sulfur bacteria occur in diverse ecosystems and play an important role in the global sulfur, nitrogen and phosphorus cycle. In this study, sediment samples from brackishwater shrimp culture ponds and other brackishwater ecosystems from Tamil Nadu, southeast coast of India, were enriched for Beggiatoa species. Extracted hay medium supplemented with catalase was used and were incubated for two weeks at 28°C. Out of seven set-ups, four yielded positive growth of filamentous sulfide-oxidizing bacteria. The filaments were several millimeters long, ranged in width between 2 and 15μm and exhibited typical gliding motility. The 16S rRNA gene of four single filaments representing the four positive enrichments was subjected to PCR-DGGE followed by sequencing. All four filaments were affiliated to the Beggiatoaceae, but showed less than 89% identity with the Beggiatoa type strain Beggiatoa alba and less than 93% identity with any other sequence of the family. One of the four filaments revealed a nearly full-length 16S rDNA sequence (1411bp) and it formed a monophyletic cluster with two of the partial DGGE-16S rRNA gene sequences (99-100% identity) within the Beggiatoa species cluster. These organisms could possibly represent a novel genus within the family Beggiatoaceae. The fourth partial sequence affiliated with less than 93% sequence identity to the genera Parabeggiatoa, Thioploca and Thiopilula, and was likewise strongly delineated from any sequence published in the family.     

Microscopic examination and fatty acid characterization of filamentous bacteria colonizing substrata around subtidal hydrothermal vents

Microscopic examination of the whitish mat that covered the substrata around subtidal hydrothermal vents at White Point in southern California revealed a Thiothrix-like bacterium containing sulfur inclusions as the dominant filamentous form in this microbial community. The matlike appearance developed as a result of the closely-packed manner inwhich the basal ends of the filaments were anchored to the substrate. The dominant phospholipid fatty acids of these filaments (16:0, 16:1w7c, 18:0, 18:1w7c) were similar to those recovered from a sample of Beggiatoa isolated from a spring in Florida. Filaments from both sources contained small quantities of C₁₈ and C₂₀ polyunsaturated fatty acids, as well. A larger but less abundant sheathless, filamentous form, which also contained sulfur inclusions and displayed a cell wall structure similar to a previously described Thioploca strain, also colonized the substrata around the subtidal mat. The preservation methods used in the preparation of thin-sections of the subtidal mat material were found to be inadequate for defining some key cellular structures of the large filaments. Nevertheless, the results demonstrate that the filamentous bacteria comprising the microbial mat in the vicinity of the subtidal vents exhibit some of the features of the free-living filamentous microorganisms found in deep-water hydrothermal areas.Published as Technical Report of the Southern California Ocean Studies Consortium, Long Beach, CA, USA     

Ultrastructural architecture of proteoglycans in the glomerular basement membrane. A cytochemical approach

Rat kidneys were perfused with fixative solutions containing either a) a polycationic dye (Alcian blue 8 GX, Astra blue 6 GLL, cuprolinic blue, ruthenium red), b) a monocationic dye (safranine 0), or c) Alcian blue in the presence of a 0.3 M MgCl2 concentration. Whereas solutions of a revealed the glomerular basement membrane proteoglycans as particles or threads 60 nm apart and arranged in a reticular pattern, solutions of b and c demonstrated new morphological aspects of these molecules. They appeared as tiny filamentous structures, about 100 to 160 nm long, ordered in a network-like pattern with a mesh of about 60-nm width. The filaments displayed lateral branches about 20 nm apart and about 25 nm long, projecting within the meshes. We suggest that the filamentous structures are the protein core, and the branches are the glycosaminoglycans of proteoglycan molecules. Because of this arrangement the negatively charged sites of the glomerular basement membrane would lie closer to each other than previously assumed.     

Proteoglycans in arterial smooth muscle cell cultures: an ultrastructural histochemical analysis

The extracellular matrix in cultures of arterial smooth muscle cells has been examined by ultrastructural histochemistry using each of the following cationic dyes: ruthenium red, Alcian blue, acridine orange, and safranin O. All dyes exhibited an affinity for a structural component that was either preserved as a granule with ruthenium red or Alcian blue, or as an extended filament or bottlebrush structure with acridine orange or safranin O. Both granules and filaments were removed when the cultures were pretreated with chondroitinase ABC, an enzyme that degrades the glycosaminoglycan moiety of some proteoglycans. These structural components of the extracellular matrix were not observed when cultures were prepared in the absence of the cationic dyes. Labeling experiments (35S-sulfate) revealed that approximately 40% of the total labeled proteoglycans were lost during routine processing for electron microscopy (i.e., fixation through dehydration). Inclusion of any one of the cationic dyes during fixation reduced the losses to less than 1%. The extended filamentous structure preserved by safranin O and acridine orange resembled the structure of purified proteoglycans prepared from the same cultures and spread on cytochrome c monolayer films. These observations suggest that proteoglycans exist as extended bottlebrush structures within the extracellular matrix, and support the interpretation that the granular deposits observed in the ruthenium red and Alcian blue preparations most likely represent individual proteoglycan monomers that have undergone molecular collapse during processing. In addition, the dyes also exhibited an affinity for chords of fine fibrils that contained small granules and/or filaments. Both the fibrillar material and the associated granular and filamentous structures enmeshed in the fibrils resisted digestion with chondroitinase ABC.     

A filamentous cytoskeleton in vertebrate smooth muscle fibers.

There are three classes of myofilaments in vertebrate smooth muscle fibers. The thin filaments correspond to actin and the thick filaments are identified with myosin. The third class of myofilaments (100 A diam) is distinguished from both the actin and the myosin on the basis of fine structure, solubility, and pattern of localization in the muscle fibers. Direct structural evidence is presented to show that the 100A filament constitute an integrated filamentous network with the dense bodies in the sarcoplasm, and that they are not connected to either the actin or myosin filaments. Examination of (a) isolated dense bodies, (b) series of consecutive sections through the dense bodies, and (c) redistributed dense bodies in stretched muscle fibers supports this conclusion. It follows that the 100-A filaments complexes constitute a structrally distinct filamentous network. Analysis of polyacrylamide gels after electrophoresis of cell fractions that are enriched with respect to the 100-A filaments shows the presence of a new muscle protein with a molecular weight of 55,000. This protein can form filamentous segments that closely resemble in structure the native, isolated 100-A filaments. The results indicate that the filamentous network has a structure and composition that distinguish it from the actin and myosin in vertebrate smooth muscle.     

Microscopic examination of distribution and phenotypic properties of phylogenetically diverse Chloroflexaceae-related bacteria in hot spring microbial mats

We investigated the diversity, distribution, and phenotypes of uncultivated Chloroflexaceae-related bacteria in photosynthetic microbial mats of an alkaline hot spring (Mushroom Spring, Yellowstone National Park). By applying a directed PCR approach, molecular cloning, and sequence analysis of 16S rRNA genes, an unexpectedly large phylogenetic diversity among these bacteria was detected. Oligonucleotide probes were designed to target 16S rRNAs from organisms affiliated with the genus Chloroflexus or with the type C cluster, a group of previously discovered Chloroflexaceae relatives of this mat community. The application of peroxidase-labeled probes in conjunction with tyramide signal amplification enabled the identification of these organisms within the microbial mats by fluorescence in situ hybridization (FISH) and the investigation of their morphology, abundance, and small-scale distribution. FISH was combined with oxygen microelectrode measurements, microscope spectrometry, and microautoradiography to examine their microenvironment, pigmentation, and carbon source usage. Abundant type C-related, filamentous bacteria were found to flourish within the cyanobacterium-dominated, highly oxygenated top layers and to predominate numerically in deeper orange-colored zones of the investigated microbial mats, correlating with the distribution of bacteriochlorophyll a. Chloroflexus sp. filaments were rare at 60 degrees C but were more abundant at 70 degrees C, where they were confined to the upper millimeter of the mat. Both type C organisms and Chloroflexus spp. were observed to assimilate radiolabeled acetate under in situ conditions.     

Ultrastructure of the secretory cells of the submucosal glands in the human maxillary sinus

Tissue samples obtained from the lateral wall of the maxillary sinuses of five patients were examined by light microscopical, histochemical, and ultrastructural techniques. Submucosal glands were tubulo-alveolar mixed glands. The acini consisted of either all serous or all mucous cells, or a mixture of both. Serous granules were stained by toluidine blue, or by hematoxylin and eosin (H and E), but showed little or no reaction with periodic acid-Schiff (PAS) or Alcian blue. Mucous granules were pale in toluidine blue or H and E preparations, and consisted primarily of acid mucosubstances, as demonstrated by their staining reaction with PAS and Alcian blue. At the electron microscope level, the serous granules were either homogeneously dense, or showed a substructure consisting of at least two layers of distinctly different electron-opacity. Typical mucous droplets consisted of a fibrillar network dispersed in a translucent matrix. A second secretory product was present in the mucous cells in the form of elongated, membrane-bounded structures containing numerous parallel filaments, which measured about 55 Å in diameter. The mucous droplets and the filamentous bodies appear to arise from the opposite faces of the Golgi complex in the mucous cells. The filamentous bodies showed a pronounced tendency to fuse with the mucous droplets. All acini were surrounded by a well-defined myoepithelial layer and contained intercellular nerve terminals.     

Protein-protein interactions of proteolytic fragments of actin.

Proteolytic fragments of actin, prepared by removal of up to sixty-eight residues from the N-terminal end of the molecule, can form filamentous structures after denaturation in urea solution. The filaments have a diameter similar to F-actin filaments and interact with myosin and tropomyosin. A fragment comprising residues 1 to 207 of the actin sequence did not form filaments or interact with myosin after the urea treatment.     

Composition and dynamics of epilithic algae in a forest stream at Shillong (India)

The seasonal dynamics of epilithic algae in a third order pristine forest stream were analyzed over a period of 2 years. Stream water was slightly acidic and nutrient poor. Encrusting, filamentous flocs, and filaments were found. Algal standing crop was high (mean concentration of Chl a 16–43 mg m^–2) in spring. Filamentous algae contributed most to standing crop. Diatoms made up over 85% by number of the epilithon. Blue-greens were abundant upstream, and chlorophytes downstream. This shift was ascribed to greater light availability downstream. The community was more diverse during spring. Water current was the most important variable regulating epilithon structure. Total phosphorus (TP), orthophosphate (O-PO _inf4 ^su3– ), silica (Si⁴⁺), nitrate (NO _inf3 ^su– ) and conductance correlated negatively with flow rate. Green algae showed a positive correlation to phosphorus during low and stable flow. During rapid runoff, diatoms were the most resistant forms. Seasonal change in the epilithic community was mainly regulated by fluctuations in flow rate.     

COLONIZATION AND COMMUNITY STRUCTURE OF TWO PERIPHYTON ASSEMBLAGES,WITH EMPHASIS ON THE DIATOMS (BACILLARIOPHYCEAE)

Examination of the three-dimensional structure of periphyton communities through time indicated that their microsuccession is analogous to higher plant succession. The development of attached diatom communities in two reservoirs was studied using artificial substrates, and the morphology of dominant organisms, patterns of spatial heterogeneity, and community interactions were documented with scanning electron microscopy. Of 93 taxa found, Gomphonema parvulum, G. olivaceum, Navicula graciloides, Nitzschia palea, and N. dissipata were dominant, depending upon season and reservoir. Comparisons of community diversity (SIMI) between reservoirs within seasons ranged from 0.023–0.843 (median = 0.254), indicating that the reservoirs were quite different with respect to diatoms present and their apportionments. Colonization was slow the first 2 wk in spring and fall, and throughout winter, but rapid during summer. Shifts in numerical dominance between certain species occurred in fall, spring, and summer. These inverse correlations of abundances and the functional dominance of overgrowth suggested competition for substrate surface area in the periphyton. The colonization sequence was often predictable—a presumably organic coating and a variety of bacteria, followed by low profile diatoms, and finally an upperstory of long-stalked and large-rosette diatoms and filamentous green algae. Periphyton microsuccession is similar to higher plant succession in the consistent change in vertical community structure from low to high physical stature, in the association of numerical dominance with large stature (via cell size or long mucilaginous stalks), and in the progressive slow-down in the rate of succession. Diatom mucilage also contributed to community structure by binding particulates and entrapping other algae and serving as the mechanism for substrate attachment.     

Recent increase of filamentous algae in shallow Swedish bays: Effects on the community structure of epibenthic fauna and fish

A summary is presented of estimates of distribution and growth of filamentous algae and its effect on the structure and functioning of epibenthic fauna and fish communities in shallow bays on the Swedish west coast. As a consequence of coastal eutrophication vegetation cover has gradually increased during the last decade, and during 1990's most bays in the Skagerrak-Kattegat area were variously covered with filamentous algae during spring and summer (May–July). In some areas filamentous algae (mainlyCladophora andEnteromorpha) completely covered the bottom. In field studies it was demonstrated that increased cover and dominance of filamentous algae result in structural changes of the epibenthic fauna community. Field studies showed that species richness and biomass of epibenthic fauna increased in a sandy bay with a moderate increase (30 to 50%) of filamentous algae cover. At higher cover (90%), biomass of epibenthic fauna was reduced, however, to the same level as for the sandy habitat, although the dominant epibenthic species were different. Heavy growth of epiphytic filamentous algae on eelgrass resulted in reduced biomass and a shift in the species composition of the epibenthic fauna community. Fish assemblage structure was also related to changes in vegetation. In eelgrass beds, fish species numbers were reduced with increasing cover of epiphytic filamentous algae, and at rocky bottoms with kelp algae (dominated byFucus), fish biomass decreased with increasing cover of attached filamentous algae. Further, foraging efficiency of juvenile cod and settling success of plaice were reduced as a response to increasing dominance of filamentous algae.     

Immunocytochemical studies of intermediate filament aggregates and their relationship to microtubules in cultured skin fibroblasts from patients with giant axonal neuropathy

Giant axonal neuropathy (GAN) is a severe autosomal recessive disease affecting both the peripheral and central nervous systems. It is characterized by segmental axonal ballooning due to large neurofilamentous masses and abnormal aggregation of filaments in other cell types including glial cells. Coomassie blue staining of the detergent-resistant cytoskeleton of cultured skin fibroblasts from three patients with GAN revealed the presence of large cytoplasmic filamentous aggregates in the great majority of cells. The aggregates were birefringent when viewed under polarization microscopy and electron microscopy showed that they were composed of aggregates of 8 to 10 nm intermediate filaments. The aggregates stained with antisera specific for vimentin but did not stain with antibodies to actin, tubulin, or the high molecular weight (HMW) microtubule associated protein. Examination of the fibroblasts containing the vimentin aggregates with antibodies to tubulin and the HMW protein showed that they had a normal distribution of microtubules and that the microtubules present were normally associated with the HMW protein. The results suggest that giant axonal neuropathy is a generalized inborn error of organization of intermediate filaments and that a defect in microtubules or their association with HMW protein is not responsible for the observed aggregation of intermediate filaments in this disease. Further study of GAN may be useful in understanding the function of intermediate filaments.     

Connections of intermediate filaments with the nuclear lamina and the cell periphery

We investigated the relationship between intermediate filaments (IFs) and other detergent- and nuclease-resistant filamentous structures of cultured liver epithelial cells (T51B cell line) using whole mount unembedded preparations which were sequentially extracted with Triton X-100 and nucleases. Immunogold labelling and stereoscopic observation facilitated the examination of each filamentous structure and their three-dimensional relationships to each other. After solubilizing phospholipid, nucleic acid and soluble cellular protein, the resulting cytoskeleton preparation consisted of a network of cytokeratin and vimentin IFs linked by 3 nm filaments. The IFs were anchored to and determined the position of the nuclear lamina filaments (NLF) network and the centrioles. The NLF was composed of the nuclear lamina filaments measuring 3-6 nm in diameter which radiated from and anchored to the skeleton of the nuclear pores. The IFs located in the nuclear region appeared to be interwoven with the NLF. At the cell surface, the IFs seemed to be attached to the putative actin filament network. They formed a focally interrupted plexus-like structure at the cell periphery. Fragments of vimentin filaments were found among the filamentous network located at the cell surface, and some filaments terminated blindly there.