POR structural domains important for the enzyme activity in R. capsulatus complementation system

NADPH:protochlorophyllide oxidoreductase (POR) catalyzes hydrogen transfer from NADPH to protochlorophyllide (PChlide) in the course of chlorophyll biosynthesis in photosynthetic organisms and is involved in the regulation of the development of photosynthetic apparatus in higher plants, algae and cyanobacteria. To approach molecular factors determining the enzyme activity in a living cell, several mutants of POR from pea (Pisum sativum) with site-directed modifications in different parts of the enzyme were generated. The mutant enzymes were expressed in a R. capsulatus mutant deficient in BChl biosynthesis, and their catalytic activity and ability to integrate in bacterial metabolism were analyzed. Our results demonstrate that in heterologous bacterial cell system, higher plant POR is integrated in the porphyrin biosynthesis network and its activity leads to the formation of photosynthetic chlorophyll-proteins (CPs). The study of POR mutants in R. capsulatus reveals several POR domains important for the association of the enzyme with other subcellular components and for its catalytic activity, including identification of putative enzyme reaction center and substrate binding site. The study also demonstrated that an unknown structural factor is important for the formation of the enzyme photoactive complex in etiolated plants. Moreover, our findings suggest that POR might be directly involved in the regulation of the metabolism of other porphyrins. This revised version was published online in June 2006 with corrections to the Cover Date.     

THE ROLE OF UNICELLS IN THE POLYMORPHIC SCENEDESMUS ARMATUS (CHLOROPHYCEAE)1

The UTEX 2193 strain of Scenedesmus armatus (Chod.) Chod, when cultured in any of several media (whether natural or artificial, concentrated or dilute) produced a variety of colonial morphologies as well as a unicell population. Morphological expression was related to culture ape. When the initial cell density was just a feu1 hundred cells per mL. the culture first produced a unicell population, then spiny colonies, and as stationary phase was approached, spine-less colonies. Two classes of spiny colonies were detected. Type I colonies had elongate cells with the terminal cells shorter than median cells. Spines were longer than cell length. The wider, oval, grainy cells of Type II colonies were uniform m length. Spines were shorter and thicker than those on Type I colonies. Only Type I colonies produced unicells: the latter appeared as two morphs. The smaller unicell was obovoid with four delicate spines: the larger had ovate cells bearing four thicker spines. Control of unicell development in all media was achieved by carefully monitoring colony type and cell number used for the inoculum. A unicellular population developed in batch culture in defined media, both concentrated and dilute, when the initial cell density (either Type I or Type II colonies) was low (below 1000 cells-mL^?1), as well as in synchronous cultures. With higher initial cell densities, e.g. 2 × 10⁴ cells·mL^?1, the inoculum had to contain Type I colonies to produce unicells. Unicells were also produced in water from Agronomy Pond, where the strain originated. We discuss the role of unicell populations in the distribution of Scenedesmus.     

Neosiphonia flavimarina gen. et sp. nov. with a taxonomic reassessment of the genus Polysiphonia (Rhodomelaceae, Rhodophyta)

Vegetative and reproductive development of Neosiphonia flavimarina gen. et sp. nov. (Rhodomelaceae, Ceramiales) from Bangpo on the western coast of Korea was investigated. This species is superficially similar to Polysiphonia, but differs distinctly from the latter in vegetative and reproductive structures. The plants attach by a solid disk composed of a dense cluster of rhizoids cut off from the pericentral cell wall, and bear erect indeterminate branches producing the lateral-branch initials from successive segments in a spiral arrangement. The procarps have a three-celled carpo-gonial branch. Spermatangial branches are formed on a primary branch of the trichoblasts, terminating in a single or occasionally two large, sterile cells. Tetra-sporangia are produced from the second pericentral cell adjacent to the trichoblast basal cell on indeterminate branches, and arranged spirally. Comparing several taxonomic characters among related genera, Neosiphonia occupies an independent phylogenetic position from Polysiphonia and leads to the conclusion that the genus may have a strong link with Fernandosiphonia which has a unilateral branching system. Relevant nomenclatural changes for several Polysiphonia species are also proposed.     

CHLAINOMONAS KOLII(HARDY ET CURL) COMB. NOV. (CHLOROPHYTA,VOLVOCALES), A REVISION OF THE SNOW ALGA,TRACHELOMONAS KOLII HARDY ET CURL (EUGLENOPHYTA,EUGLENALES)1,2

The snow alga Trachelomonas kolii is transferred from the Euglenophyta (Euglenales) to the Chlorophyta (Volvocales) as Chlainomonas kolii comb. nov. As a result of critical examination of both living and type material, this species was found to have 4 flagella per vegetative cell, true starch, and 1 axial plastid per cell with several peripheral lobes. Vegetative cells of Trachelomonas kolii were described originally as having 1 flagellum, lacking true starch but having paramylum, and as having several parietal plastids per cell. The reticulate markings on the outer envelope of vegetative cells were found to be different from those in the original illustrations. Vegetative cells and resting spores of Chlainomonas kolii and Chlainomonas rubra are compared. The similarities of resting spores of Chlainomonas kolii and Chlamydomonas nivalis are discussed. These are the first records of Chlainomonas kolii from snow in Washington State.     

Sporogenesis in Eusporangiate Ferns: I. Monoplastidic Meiosis in Angiopteris (Marattiales)

Angiopteris (Marattiales) undergoes the more primitive form of monoplastidic meiosis, while other ferns have evolved the polyplastidic type typical of seed plants. In monoplastidic cell division, the single plastid divides and serves as site of the microtubule organizing center (MTOC) for spindle formation resulting in coordinated division of plastid, nucleus, and cytoplasm. In plants with polyplastidic cell division, the MTOC is diffuse and generally perinuclear. Monoplastidic cell division is seen as a plesiomorphic feature that was inherited from algal ancestors containing a single plastid and modified through evolution. Monoplastidic meiosis occurs in all groups of bryophytes (although in only a few hepatics), Isoetes, Selaginella, certain generic segregates of Lycopodium, and in members of the Marattiales. It is not known to occur in psilophytes, Equisetum, leptosporangiate ferns, or seed plants. Received 30 January 2001/ Accepted in revised form 24 April 2001     

Selection for resistance to filtrates of Fusarium spp. in embryogenic cell suspension culture of Medicago sativa L.

A highly embryogenic cell suspension of alfalfa derived from a genotype sensitive to Fusarium oxysporum was successfully used for selection in vitro for resistance to culture filtrates of F. oxysporum, F. solani and F. avenaceum. Fifty two stable resistant cell lines were obtained and 500 plants regenerated from them. Among the 167 regenerants tested under glass there were 12–20% more plants with increased resistance to pathogens than in the group of plants regenerated from a control cell line. It was also found that the cell suspension cultures derived from genotypes of alfalfa with increased resistance to Fusarium spp. better tolerated filtrates of the pathogen. The results of a comparison of virulence of individual isolates of several species of Fusarium with toxicity of their filtrates to plants in vivo and in cell cultures were not unequivocal.     

NEW ENDOLITHIC CYANOPHYTES FROM THE NORTH ATLANTIC OCEAN. III. HYELLA PYXIS LUKAS & HOFFMAN SP. NOV.1

A new species of marine endolithic cyanophyte, Hyella pyxis Lukas and Hoffman (Order: Pleurocapsales), differs from other species of Hyella in its cell and filament dimensions, the manner in which its branches are initiated and the presence of gloeocapsin in the sheaths of colonies from the intertidal zone. Hyella pyxis colonies consist of a small cluster of coccoid cells located at the substrate surface and long, conspicuously branched filaments composed of cells that are longer than they are wide. Branches are initiated by the reorientation of the distal end of a filament cell or by the elongation of a filament cell, usually at one of its distal corners. Chromatic adaptation was not observed perhaps accounting for the relatively shallow depth limit of this species. Hyella pyxis was found within mollusk shells from the continental margin of eastern Florida to a depth of 50 m and carbonate rocks in the intertidal zone on Bermuda.     

EFFECTS OF LIGHT INTENSITY ON DIVISION RATE,STIMULABLE BIOLUMINESCENCE AND CELL SIZE OF THE OCEANIC DINOFLAGELLATES DISSODINIUM LUNULA,PYROCYSTIS FUSIFORMIS AND P. NOCTILUCA12

Preadapted cultures were grown in a 12:12 LD cycle at a series of light intensities under cool-white, fluorescent lamps. Pyrocystis fusiformis Murray maintained high division rates at low light intensities at the expense of cell size. In contrast, Dissodinium lunula (Schuett) Taylor had relatively lower division rates at low light intensities with little concomitant decrease in size. The response of P. noctiluca Murray was intermediate between these two species. For all three, cell numbers did not increase above an intensity of 5–10 μEin·m^?2·sec^?1 and division rate was saturated at ca. 30, 60, and 60μEin·m^?2·sec^?1 for P. fusiformis, P. noctiluca, and D. lunula, respectively. The capacity for stimulable bioluminescence was saturated at light intensities of 0.15 μEin·m^?2·day in short-term (2-day) experiments. In cultures of P. fusiformis and P. noctiluca, maintained for at least one month at lower intensities than needed to saturate division rate, a decrease in the capacity for stimulable bioluminescence was accompanied by a reduction in cell size. Our results suggest that cell size and bioluminescent capacity may prove to be a potentially useful indication of the history of exposure of natural populations of Pyrocystis spp. to ambient intensities.     

New ribosome-inactivating proteins with polynucleotide:adenosine glycosidase and antiviral activities from Basella rubra L. and Bougainvillea spectabilis Willd.

New single-chain (type 1) ribosome-inactivating proteins (RIPs) were isolated from the seeds of Basella rubra L. (two proteins) and from the leaves of Bougainvillea spectabilis Willd. (one protein). These RIPs inhibit protein synthesis both in a cell-free system, with an IC₅₀ (concentration causing 50% inhibition) in the 10⁻¹⁰ M range, and by various cell lines, with IC₅₀s in the 10⁻⁸–10⁻⁶ M range. All three RIPs released adenine not only from rat liver ribosomes but also from Escherichia coli rRNA, polyadenylic acid, herring sperm DNA, and artichoke mottled crinkle virus (AMCV) genomic RNA, thus being polynucleotide:adenosine glycosidases. The proteins from Basella rubra had toxicity to mice similar to that of most type 1 RIPs (Barbieri et al., 1993, Biochim Biophys Acta 1154: 237–282) with an LD₅₀ (concentration that is 50% lethal) ≤ 8 mg · kg⁻¹ body weight, whilst the RIP from Bougainvillea spectabilis had an LD₅₀ >32 mg · kg⁻¹. The N-terminal sequence of the two RIPs from Basella rubra had 80–93% identity, whereas it differed from the sequence of the RIP from Bougainvillea spectabilis. When tested with antibodies against various RIPs, the RIPs from Basella gave some cross-reactivity with sera against dianthin 32, and weak cross-reactivity with momordin I and momorcochin-S, whilst the RIP from Bougainvillea did not cross-react with any antiserum tested. An RIP from Basella rubra and one from Bougainvillea spectabilis were tested for antiviral activity, and both inhibited infection of Nicotiana benthamiana by AMCV. Received: 5 March 1997 / Accepted: 27 May 1997     

The characterization of Listeria spp. isolated from food products and the food‐processing environment

Aim: To enhance the information pertaining to the epidemiology of a collection of 378 Listeria spp. isolates obtained from several food‐processing plants in Ireland over a 3‐ year period (2004–2007). Methods and results: The collection was characterized by pulsed‐field gel electrophoresis (PFGE). The most prevalent pulse‐type was PFGE profile I (n = 14·5%) that consisted mainly of environmental Listeria spp. samples. Serotyping of 145 Listeria monocytogenes isolates was performed. The most common serovar was 1/2a and comprised 57·4% (n = 77) of the L. monocytogenes collection. The other serovars were as follows: 4b (14·1%, n = 19), 1/2b (9·7%, n = 13), 4c (4·4%, n = 6) and 1/2c (6·7%, n = 9), respectively. Eleven isolates were identified as non‐Listeria spp., the remaining ten L. monocytogenes isolates were nontypeable. The antimicrobial susceptibility testing revealed the antibiotic that isolates displayed the most resistance to was gentamicin (5%) followed by sulfamethoxazole‐trimethoprim (2%), tetracycline and ciprofloxacin (1·5%). Conclusions: The subtyping has indicated the diversity of the Listeria spp. The presence of serotype 1/2a, 1/2b and 4b in both raw and cooked ready‐to‐eat food products is a public health concern, as these serotypes are frequently associated with foodborne outbreaks and sporadic cases of human listeriosis. In addition, the emergence of antimicrobial‐resistant L. monocytogenes isolates could have serious therapeutic consequences. Significance and Impact of Study: The molecular subtyping and the further characterization of these isolates may be valuable particularly in the context of a suspected common source outbreak in the future.     

Cochlodinium fulvescens sp. nov. (Gymnodiniales, Dinophyceae), a new chain-forming unarmored dinoflagellate from Asian coasts

Cellular morphology and the phylogenetic position of a new unarmored photosynthetic dinoflagellate Cochlodinium fulvescens Iwataki, Kawami et Matsuoka sp. nov. were examined by light microscopy and molecular phylogenetic analyses based on partial large subunit ribosomal DNA (LSU rDNA) and small subunit ribosomal DNA (SSU rDNA) sequences. The cells of C. fulvescens closely resemble C. polykrikoides, one of the most harmful red tide forming dinoflagellates, due to it possessing a cingulum encircling the cell approximately twice, a spherical nucleus positioned in the anterior part of the cell and an eyespot‐like orange pigmented body located in the dorsal side of the epicone, as well as formation of cell‐chains. However, this species is clearly distinguished from C. polykrikoides based on several morphological characteristics, namely, cell size, shape of chloroplasts and the position of narrow sulcus situated in the cell surface. The sulcus of C. fulvescens is located at the intermediate position of the cingulum in the dorsal side, whereas that of C. polykrikoides is situated immediately beneath the cingulum. LSU rDNA phylogenies indicated that C. fulvescens is clearly distinct from, but closely related to C. polykrikoides among dinoflagellates.     

Dynamics of limiting cell wall porosity in plant suspension cultures

Changes in the limiting porosity of cell walls, i.e. the size limit for permeation of neutral molecules through the wall, were studied in several higher-plant cell-suspension cultures. For this purpose, samples of biomass fixed at different cultivation times were investigated using a method based on size-exclusion chromatography of polydisperse dextrans before and after equilibration with the extracted cell clusters. In suspension cultures of Chenopodium album L., Dioscorea deltoidea Wall. and Medicago sativa L., the mean size limit (MSL; critical Stokes' radius for exclusion of neutral polymers from half of the intracellular space) was found to vary between 2.4 and 3.8 nm. It decreased significantly during transition from the growth phase to the stationary phase. In the case of the C. album culture this change was found to be irrespective of whether sucrose in the medium was completely depleted at the end of the growth phase or not. The MSL was kept constant for long periods of the stationary phase if cell viability was maintained by repeated sucrose supplement. In a suspension strain of Triticum aestivum L., the MSL of cell wall permeation was comparatively small (1.75 nm) and remained constant during all cultivation phases. Relations between limiting porosity and cell wall growth, loss of pectic compounds to the medium, cross-linking activities and cell wall stiffening are discussed. Received: 19 December 1996 / Accepted: 23 April 1997     

INFLUENCE OF METHOD FOR REMOVAL OF SESTON ON THE DISSOLVED ORGANIC MATTER. II. COBALAMINS

Comparisons of the 3 previously used methods (continuous flow centrifugation, Millipore®filtration, and biodialysis) for making water samples seston-free before analysis for cobalamins point to several likely sources of error: (a) unpredictable occlusion of pores in Millipore®membranes during filtration; (b) irreversible adsorption of cobalamins by Millipore®membranes; (c) probable cell injury and leakage by centrifugation. Some water samples gave good agreement on soluble vitamin B₁₂ by all 3 methods, while other samples had appreciable discrepancies. The validity of total vs. sestonic cobalamin determinations therefore is questioned. One lake had time-dependent fluctuation in soluble cobalamin.     

Biotransformation of monoterpenes and sesquiterpenes by cell suspension cultures of Achillea millefolium L. ssp. millefolium

The transformation capacity of Achillea millefolium L. ssp. millefolium (yarrow) cell suspension cultures was investigated using geraniol (50mg/l) and borneol, menthol, thymol and farnesols (25mg/l) as substrates. Apart from converting these substrates into several biotransformation products, the cell suspension cultures were also able to glycosylate both the substrates and the biotransformation products. aa]Key Words bb]Achillea millefolium L. ssp. millefolium bb]Yarrow bb]Compositae bb]Biotransformation bb]Glycosylation bb]Geraniol bb]Borneol bb]Menthol bb]Thymol bb]Farnesols     

CHARACTERIZATION OF MORPHOSPECIES AND STRAINS OF MICROCYSTIS (CYANOBACTERIA) FROM NATURAL POPULATIONS AND LABORATORY CLONES USING CELL PROBES (LECTINS AND ANTIBODIES)1

The genus Microcystis (cyanobacteria) includes toxic and bloom-forming morphotypes which are usually arranged into species based on morphological features. Immunofluorescence assays using polyclonal and preadsorbed antibodies, as well as FITC-labebd lectins were used to characterize three morphospecies of Microcystis (M. viridis, M. wesenbergii, and M. aeruginosa) from natural populations (several lakes/reservoirs in Denmark and Spain) and laboratory clones. The cell probes used were unaffected by the different phases of the cell division cycle, growth phase, or environmental factors, such as culture medium, light, or temperature. Anhbody and lectin binding patterns were specific to each clone. In nature, the cell probes were useful tools to characterize Microcystis populations. Antibodies and lectins revealed geographic differentiation within the same morphospecies. Differentiation was moderate among nearby locales and intensified among areas distant from one another. Microcystis aeruginosa from Spain has very different cell surface antigens and lectin binding sites than M. aeruginosa from Denmark. A taxonomy of Microcystis based on cell probes reveals some discrepancies with classical morphospecies. The binding affinities were more closely related to the geographic origin of the tested material than to the morphospecies identification. Different morphospecies from the same lake in some cases were more similar than the same morphospecies from different lakes. Microcystis viridis and M. aeruginosa from Danish lakes appeared to be closely related species, whereas M. wesenbergii emerged as a different species.     

Russian Article pp. 309–318

It is demonstrated that aliphatic hydrocarbons (alkanes) penetrate into bacteria cells by the way of passive diffusion. The mechanism of this process is different for several bacteria species. A hydrophobic cell wall is essential for that process. In saprophytic Mycobacteria hydrocarbons are solubilized in the thick hydrophobic cell wall. During the process of absorption hydrocarbons pass through the whole cell wall up to the membrane. In the case of Arthrobacteria the hydrocarbons might pass not through the whole cell wall, but through special lipophilie canals. Mobile hydrocarbon-oxidizing bacteria g. Pseudomonas form peptidoglycolipid and excrete it into the medium. The peptidoglycolipid emulsifies hydrocarbon substrate.     

Prevention of aluminium toxicity with supplemental boron. I. Maintenance of root elongation and cellular structure

Aluminium toxicity is an important factor limiting plant growth mi acid soils. Symptoms of B deficiency and Al toxicity are very similar and generally associated with impaired membrane Function and root growth. Thus the objective of this study was to determine whether supplemental B prevents Al inhibition of root growth and development. Squash (Cucurbita pepo L. cv. Sunbar) was grown in hydroponic nutrient media with 44 mmol m^?3 free Al and B concentrations extending from 5 to 100 mmol m^?3. Our results establish that B protects against Al inhibition of root growth. Protection was apparent at all levels of organization examined: primary root and lateral root lengths; primary root cell elongation, cell production rate, tissue organization and cell structure; primary root morphology and maturation. Protection against Al inhibition was also apparent for shoot growth. These studies were undertaken in solution culture to limit the variables examined; however, the underlying motivation for this study is the problem of worldwide Al toxicity in soils. Therefore, the effect of adding additional B to a high-Al soil was also investigated and is the subject of the companion paper (Le Noble. Blevins & Miles 1996, Plant, Cell and Environment 19, 1143–1148).     

Immobilization of Burkholderia cepacia in polyurethane-based foams: embedding efficiency and effect on bacterial activity

Immobilization of the trichloroethylene-degrading bacterium Burkholderia cepacia was evaluated using hydrophilic polyurethane foam. The influence of several foam formulation parameters upon cell retention was examined. Surfactant type was a major determinant of retention; a lecithin-based compound retained more cells than pluronic- or silicone-based surfactants. Excessive amounts of surfactant led to increased washout of bacteria. Increasing the biomass concentration in the foam from 4.8 to 10.5% dry weight per wet weight of foam resulted in fewer cells being washed out. Embedding at reduced temperature did not significantly affect retention, while the use of a silane binding agent gave inconsistent results. The optimal formulation retained all but 0.2% of total embedded cells during passage of 2 L of water through columns containing 2 g of foam. All foam formulations tested reduced the culturability of embedded cells by several orders of magnitude, but O₂ consumption and CO₂ evolution rates of embedded cells were never less than 50% of those of free cells. Nutrient amendments stimulated an increase in cell volume and ribosomal activity in immobilized cells as indicated by hybridization studies using fluorescently labeled ribosomal probes. These results indicate that, although immobilized cells were mostly nonculturable, they were metabolically active and thus could be used for biodegradation of toxic compounds. Received 23 December 1996/ Accepted in revised form 13 March 1997