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Phycobilisomes in blue-green algae.

Fifteen species of freshwater blue-green algae, including unicellular, filamentous, and colonial forms, were subjected to a variety of fixatives, fixation conditions, and stains for comparison of the preservation of phycobilisomes. Absorption spectra of the corresponding in vivo and released photosynthetic pigments, in 10 of the species that were maintained in culture, demonstrated the presence of phycocyanin in all 10 species and phycoerythrin in only 2 of them. Spectroscope and electron microscope evidence was obtained for localization of phycobiliproteins in phycobilisomes of Nostoc muscorum. Phycobilisomes were observed in all species examined in situ, strenghening the hypothesis that phycobilisomes are common to all phycobiliprotein-containing photosynthetic blue-green algae.

Cell Fractionation↗

Carbon dioxide assimilation in blue-green algae: initial studies on the structure of ribulose 1,5-bisphosphate carboxylase.

D-Ribulose 1,5-bisphosphate carboxylase was purified from the blue-green alga Anabaena cylindrica (Lemm) by procedures involving acid precipitation, ammonium sulfate fractionation, and Sephadex G-200 gel filtration. The enzyme was homogeneous by the criterion of polyacrylamide disc gel electrophoresis and was a multimer of a single-size polypeptide chain of 54,000 daltons. The carboxylases from four species of blue-green algae (Anabaena, Nostoc strain MAC, Agmenellum quadruplicatum strain PR-6, and Anacystis nidulans strain TX20) were closely similar in molecular size, since enzyme activity was eluted at the same volume after sucrose gradient centrifugation. Further analysis by gel filtration indicated that the four blue-green algal carboxylases were nearly identical in molecular weight, ranging from 449 to 453,000. The amino acid composition of the Anabaena carboxylase was determined and was found to resemble closely the composition of the large subunit from eukaryotic photosynthetic organisms.

Amino Acids↗

Feedback regulation of arginine biosynthesis in blue-green algae and photosynthetic bacteria.

Hoare, D. S. (The University of Texas, Austin), and S. L. Hoare. Feedback regulation of arginine biosynthesis in blue-green algae and photosynthetic bacteria. J. Bacteriol. 92:375-379. 1966.-A number of blue-green algae and photosynthetic bacteria synthesize arginine from glutamate via acetylated intermediates. Cell-free extracts of these photosynthetic microorganisms contain an N-acetyl glutamate phosphokinase, which is specifically inhibited by arginine. They also contain a transacetylase which forms ornithine from alphaN-acetyl ornithine and glutamate. The transacetylase appears to be specific for l-glutamate. Arginine synthesis and its regulation by feedback inhibition in photosynthetic microorganisms differ from that in Escherichia coli and other Enterobacteriaceae.

Journal Article↗

Biochemical basis of obligate autotrophy in blue-green algae and thiobacilli.

Differential rates of incorporation of sugars, organic acids, and amino acids during autotrophic growth of several blue-green algae and thiobacilli have been determined. In obligate autotrophs (both blue-green algae and thiobacilli), exogenously furnished organic compounds make a very small contribution to cellular carbon; acetate, the most readily incorporated compound of those studied, contributes about 10% of newly synthesized cellular carbon. In Thiobacillus intermedius, a facultative chemoautotroph, acetate contributes over 40% of newly synthesized cellular carbon, and succinate and glutamate almost 90%. In the obligate autotrophs, carbon from pyruvate, acetate, and glutamate is incorporated into restricted groups of cellular amino acids, and the patterns of incorporation in all five organisms are essentially identical. These patterns suggest that the tricarboxylic acid cycle is blocked at the level of alpha-ketoglutarate oxidation. Enzymatic analyses confirmed the absence of alpha-ketoglutarate dehydrogenase in the obligate autotrophs, and also revealed that they lacked reduced nicotinamide adenine dinucleotide oxidase, and had extremely low levels of malic and succinic dehydrogenase. These enzymatic deficiencies were not manifested by the two facultative chemoautotrophs examined. On the basis of the data obtained, an interpretation of obligate autotrophy in both physiological and evolutionary terms has been developed.

Acetates↗

Measurement of steady-state growth rates of a thermophilic alga directly in nature.

Steady-state growth rates of thermophilic algae can be determined directly in nature in the flowing waters of a hot spring channel by measuring the rate of loss of algal cells when the channel is darkened. The half time of the loss rate in the steady state is identical to the generation time. We studied the unicellular blue-green alga Synechococcus in Yellowstone National Park. Temperature and flow rate remained relatively constant throughout the experiment. Quantitative cores were taken and homogenized; the algal cells were then counted by use of a Petroff-Hausser counting chamber. After ascertaining that the algal population was in a steady state, the channel was darkened in such a way that neither flow rate nor temperature was altered. The population began to decrease within 1 day; the loss rate was exponential with time for 2 to 3 decades. Half times were then calculated from these loss rates. The growth rates found were considerably lower than those for the same organism in laboratory culture. The results suggest that in nature the organism may be an obligate phototroph. In two cases, after the algal populations decreased to an undetectable level, the dark covers were removed and the rate of recolonization was measured. The kinetics of recolonization were different from the kinetics of washout.

Cell Division↗

Alga associated with diarrhea in patients with acquired immunodeficiency syndrome and in travelers.

Spherical bodies resembling coccidian oocysts and measuring 8.0 to 9.0 microns in diameter were seen in the stools of eight persons with explosive, watery diarrhea. Seven had recently traveled to tropical countries, mostly in the Caribbean, and four had acquired immunodeficiency syndrome. The structures were easily discernible in wet mounts by light microscopy and contained variable numbers of granular inclusions, but were refractory to, or stained partially with, 12 commonly used laboratory stains. Electron microscopy revealed an outer fibrillar coat, a thin cell wall, granules, and organelles which were not surrounded by membranes. One type of organelle was similar to the thylakoid photosynthesizing organelles of blue-green algae (cyanobacteria). These findings indicate that the bodies may be a species of blue-green algae.

Acquired Immunodeficiency Syndrome↗

Biochemical and pathogenic properties of Shewanella alga and Shewanella putrefaciens.

We characterized 49 strains of Shewanella spp. from clinical (n = 31) and nonhuman (n = 18) sources. Most Shewanella alga organisms (Gilardi biovar 2; Centers for Disease Control and Prevention [CDC] biotype 2) originated from clinical material (92%), failed to produce acid from carbohydrates other than D-ribose, and were biochemically and enzymatically fairly homogeneous. In contrast, Shewanella putrefaciens organisms (Gilardi biovars 1 and 3; CDC biotype 1) were more often associated with nonhuman sources (70%), were able to utilize a number of sugars (sucrose, L-arabinose, and maltose), and were found to exhibit wider variations in biochemical characteristics; three biotypes within S. putrefaciens were detected. Notable differences between the two species in enzymatic activity, determined with the API-ZYM system (bioMérieux, Hazelwood, Mo.), and cellular fatty acid profiles, determined by the MIDI system (Microbial ID Inc., Newark, Del.), were also detected. Pathogenicity studies of mice indicate that S. alga appears to be the more virulent species, possibly due to the production of a hemolytic substance.

Animals↗

First case of osteomyelitis due to Shewanella algae.

Shewanella spp. are infrequently recovered from clinical specimens. We report here on the first case of osteomyelitis due to Shewanella algae. This bacterium, at first misidentified by phenotypic tests as Shewanella putrefaciens, was subsequently identified correctly as S. algae by 16S rRNA gene sequence analysis.

Bacterial Typing Techniques↗

Viruses and viruslike particles of eukaryotic algae.

Until recently there was little interest or information on viruses and viruslike particles of eukaryotic algae. However, this situation is changing. In the past decade many large double-stranded DNA-containing viruses that infect two culturable, unicellular, eukaryotic green algae have been discovered. These viruses can be produced in large quantities, assayed by plaque formation, and analyzed by standard bacteriophage techniques. The viruses are structurally similar to animal iridoviruses, their genomes are similar to but larger (greater than 300 kbp) than that of poxviruses, and their infection process resembles that of bacteriophages. Some of the viruses have DNAs with low levels of methylated bases, whereas others have DNAs with high concentrations of 5-methylcytosine and N6-methyladenine. Virus-encoded DNA methyltransferases are associated with the methylation and are accompanied by virus-encoded DNA site-specific (restriction) endonucleases. Some of these enzymes have sequence specificities identical to those of known bacterial enzymes, and others have previously unrecognized specificities. A separate rod-shaped RNA-containing algal virus has structural and nucleotide sequence affinities to higher plant viruses. Quite recently, viruses have been associated with rapid changes in marine algal populations. In the next decade we envision the discovery of new algal viruses, clarification of their role in various ecosystems, discovery of commercially useful genes in these viruses, and exploitation of algal virus genetic elements in plant and algal biotechnology.

Chlorella↗

The non-light-dependent reduction of 2,6-dichlorophenolindophenol by cells of the blue-green alga Anacystis nidulans.

Whole cells of the blue-green alga Anacystis nidulans reduced, in the dark, the oxidation-reduction dye, 2,6-dichlorophenolindophenol at rates severalfold higher than those of the other algae tested. Under anaerobiosis, the endogenous reductant was depleted after up to 80 nmol of dye were reduced per microliter of cells. Cells held in darkness for several hours exhibited lowered dark reduction rates relative to cells held in light. Treatment with lysozyme and ethylenediaminetetraacetic acid yielded cells that would photoreduce the dye, whereas untreated cells would not. Comparisons of photoreduction and dark reduction revealed that the dark reduction proceeded independently of the photoreduction. It was concluded that the dark reduction represents a pool of endogenous reductant of sufficiently low oxidation-reduction potenital to reduce completely 2,6-dichlorophenolindophenol. Additionally, untreated cells were shown to be permeable to the dye although they did not photoreduce it; thus lysozyme/ethylenediaminetetraacetic acid treatment was considered to make the oxidant accessible to the photosynthetic machinery.

Anaerobiosis↗

Basis for the susceptibility of several algae to microbial decomposition.

Partially purified cellulase and a cellulase-containing polygalacturonase but notlysozyme extensively degraded the walls of Chlamydomonas reinhardtii and Ulothrix fimbrata and converted intact cells of the algae to spheroplasts. A streptomycete cellulase cochromatographed with the enzyme system releasing glucose from walls of these organisms, and this preparation also converted the algal cells to spheroplasts. The dominant constituent in the walls was carbohydrate, and glucose and small quantities of galacturonic acid but no amino sugars were present in acid hydrolysates of the walls. Glucose accounted for essentially all of the material solobilized by the cellulase preparation. Lysozyme acted on Cylindrospermum sp. walls, and it, but not the otherenzymes, converted some of the Cylindrospermum sp. cells to spheroplasts. Streptomycete enzymes lysing Micrococcus lysodeikticus cochromatographed with the proteins releasing reducing sugars from Cylindrospermum sp. walls, and components in the active fraction converted cells of this alga into spheroplasts. X-ray diffraction revealed that the walls of C. reinhardtii and U. fimbrata but not those of Cylindrospermum sp. contained cellulose. The data suggest that the susceptibility of the first twospecies to microbial degradation in natural ecosystems results from an attack on the cellulose in their walls, and the susceptibility of the third is linked with the microbial production of a lysozyme.

Carbohydrates↗

Assimilation of citric acid and adipic acid by the blue-green alga Anabaena variabilis.

The assimilation of [1,6-14C] citric acid and [1,6-14C] adipic acid by the blue-green alga Anabaena variabius was studied in the dark and in the light. Citric acid was assimilated in the dark and in the light but adipic acid showed only limited assimilation in the dark. In the light the assimilation of adipic acid did not enhance the growth of the alga at a concentration of 2.85 X 10(-8) M. Growth was inhibited at adipic acid concentrations greater than 10(-3) M. Analysis of the products of adipic acid metabolism showed the presence of aspartic acid, glutamic acid, leucine, proline, and threonine and threonine amongst other unidentified compounds. A mechanism of beta-oxidation is proposed.

Adipates↗

Speculations on a possible essential function of the gelatinous sheath of blue-green algae.

Voluminous and often fluffy sheaths surrounding blue-green algal cells are observed (a) in productive natural waters, (b) in bacteria-containing laboratory cultures growing in inorganic nutrient media with added bacteria-assimilable organic matter, and (c) in axenic cultures in the same inorganic media even without added organic matter. The sheaths of bacteria-associated species in inorganic media without added organic matter are, by comparison, thin, and growth is meager. Repeated observations show that voluminous sheaths and vigorous growth of algal species are associated. It is suggested that formation and retention of a voluminous shealth provide a microenvironment around the algal cell where essential nutrients, present at only submarginal levels in the surrounding water, are concentrated and become readily available to the cell. The increase in nutrient concentration above a critical level, in turn, leads to vigorous algal growth. The voluminous sheath produced by the alga is not attacked by alga-associated bacteria when other assimilable organic matter is available; but in the absence of a more suitalble food, the bacteria feed on the less desirable gelatinous sheath, markedly reducing its thickness and causing meager algal growth.

Bacteria↗

Removal of the nitro and phenyl groups from NPPB decreases its inhibitory effect on cytoplasmic streaming in the alga Nitella hookeri.

Structural analogues of the arylaminobenzoate 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB), prepared using a simple reductive amination sequence, were tested for their effects on cytoplasmic streaming rates in the alga Nitella hookeri. Cytoplasmic streaming was sensitive to NPPB, with an IC50 value of 24 micromol/L. Removal of the nitro group from the benzoate ring decreased the IC50 to 455 micromol/L. The introduction of an extra carbon or double bond into the aliphatic chain had no effect on activity. Loss of the phenyl group decreased potency, with an IC50 of 6.4 mmol/L. These data are the first documenting the relative inhibitory effects of structural changes to arylaminobenzoates in algae. Patch-clamp data and the effects of tetrapentyl ammonium chloride on streaming suggest that the nitro and phenyl groups may act by inhibiting both K+ and Cl- channels. This is likely, through changes in the membrane potential, to affect Ca2+ fluxes and action potentials, thereby slowing cytoplasmic streaming.

Aminobenzoates↗

Isolation and characterization of a new agglutinin from the red marine alga Hypnea cervicornis J. Agardh.

The biochemical characterization of a new lectin (Hypnea cervicornis agglutinin or HCA) isolated from the Brazilian red alga H. cervicornis is reported. The haemagglutinating activity of the lectin was only inhibited by the glycoprotein porcine stomach mucin at a minimum inhibitory concentration of 19 microg x mL(-1). No haemagglutination inhibition was detected after the addition of simple sugars. The MALDI-TOF molecular masses of native and reduced and carbamidomethylated HCA were, respectively, 9196.6 Da and 9988.2 Da, indicating that the primary structure of the protein is crosslinked by 7 disulfide bonds. This unusual structural feature among lectins, along with its N-terminal sequence and amino-acid composition, clearly shows that HCA belongs to a protein family distinct from the isolectins Hypnin A1 and A2 isolated from the related Japanese alga Hypnea japonica. On the other hand, HCA displayed a high degree of similarity to the agglutinin from the Brazilian species Hypnea musciformis. Our data indicate the occurrence of structural diversity among lectins of closely related species living in distant ecosystems, i.e., the Pacific coast of Japan and the Atlantic coast of Brazil, and support the hypothesis that the lectin content (lectinome) might serve as a biomarker for taxonomical purposes.

Agglutinins↗

Involvement of the hap gene (mucinase) in the survival of Vibrio cholerae O1 in association with the blue-green alga, Anabaena sp.

Mucinase is a soluble haemagglutinin protease, which may be important for the survival of Vibrio cholerae in association with mucilaginous blue-green algae (cyanobacteria). A comparative survival study was carried out with an Anabaena sp. and a wild-type V. cholerae O1 strain hap+ gene (haemagglutinin-protease), together with its isogenic mutant hap (hap-deleted gene). A simple spread plate technique was followed to count culturable V. cholerae O1 on taurocholate tellurite gelatin agar plate. The fluorescent antibody technique of Kogure et al. (1979) was used for the microscopical viable count of V. cholerae O1. Polymerase chain reaction (PCR) and Southern blot hybridization were carried out to detect a lower number of viable but nonculturable (VBNC) V. cholerae O1 from the laboratory-based experiments. The wild and mutant V. cholerae O1 strains survived in culturable form for 22 and 10 days. respectively, in association with the Anabaena sp., with the difference being statistically significant (P < 0.01). The fluorescent antibody technique, PCR, and hybridization results also showed that the wild strain survived better in the VBNC state than did the mutant VBNC strain in association with an Anabaena sp. These results indicate that the enzyme mucinase may play an important role in the association and long-term survival of V. cholerae O1 with a mucilaginous blue-green alga, Anabaena sp.

Anabaena↗

CO2 concentrating mechanisms in algae: mechanisms, environmental modulation, and evolution.

The evolution of organisms capable of oxygenic photosynthesis paralleled a long-term reduction in atmospheric CO2 and the increase in O2. Consequently, the competition between O2 and CO2 for the active sites of RUBISCO became more and more restrictive to the rate of photosynthesis. In coping with this situation, many algae and some higher plants acquired mechanisms that use energy to increase the CO2 concentrations (CO2 concentrating mechanisms, CCMs) in the proximity of RUBISCO. A number of CCM variants are now found among the different groups of algae. Modulating the CCMs may be crucial in the energetic and nutritional budgets of a cell, and a multitude of environmental factors can exert regulatory effects on the expression of the CCM components. We discuss the diversity of CCMs, their evolutionary origins, and the role of the environment in CCM modulation.

Biological Evolution↗

Giant viruses infecting algae.

Paramecium bursaria chlorella virus (PBCV-1) is the prototype of a family of large, icosahedral, plaque-forming, double-stranded-DNA-containing viruses that replicate in certain unicellular, eukaryotic chlorella-like green algae. DNA sequence analysis of its 330, 742-bp genome leads to the prediction that this phycodnavirus has 376 protein-encoding genes and 10 transfer RNA genes. The predicted gene products of approximately 40% of these genes resemble proteins of known function. The chlorella viruses have other features that distinguish them from most viruses, in addition to their large genome size. These features include the following: (a) The viruses encode multiple DNA methyltransferases and DNA site-specific endonucleases; (b) PBCV-1 encodes at least part, if not the entire machinery to glycosylate its proteins; (c) PBCV-1 has at least two types of introns--a self-splicing intron in a transcription factor-like gene and a splicesomal processed type of intron in its DNA polymerase gene. Unlike the chlorella viruses, large double-stranded-DNA-containing viruses that infect marine, filamentous brown algae have a circular genome and a lysogenic phase in their life cycle.

Biological Evolution↗