Production of the antibiotic cyanobacterin LU-1 by Nostoc linckia CALU 892 (cyanobacterium)

Cyanobacterin LU-1, produced by Nostoc linckia CALU 892, inhibits the growth of many cyanobacteria and eukaryotic algae. The minimum effective dose of a crude preparation to Synechococcus sp. R-2 is ca 1 µg ml^?1. The antibiotic hinders cell division and light-dependent oxygen evolution in Synechococcus sp. R-2 (PCC 7942) cells. It is not active against heterotrophic bacteria and fungi, and is non-toxic to mice. Purified cyanobacterin LU-1 contains a nitrous heterocycle with sugar and phenolic substituents. Cyanobacterin LU-1 accumulates in the medium during the course of growth, although not in direct proportion to cell density. Productivity of the culture depends on temperature.     

Three new D:A Friedo-oleanane triterpenes from the stem bark of Anacolosa poilanei and their cytotoxic activities

Three new D:A friedo-oleanane triterpenes, 3α-p-coumaroyl-D:A-friedo-oleanan-27-oic acid (1), 3α-(3,4-dihydroxycinnamoyl)-D:A-friedo-oleanan-27-oic acid (2), and 3α-(3,4-dihydroxycinnamoyl)-D:A-friedo-oleanan-27,15α-lactone (3) along with three known compounds, trichadenic acid A (4), trichadonic acid (5), and amentoflavone (6), were isolated from the stem barks of Anacolosa poilanei Gagnep. Their structures were established by spectral analysis, such as mass spectrometry, 1D-NMR, and 2D-NMR. Compound 1 exhibited cytotoxicity against LU-1, HepG2, MCF7, and KB cell lines. Compounds 2 and 3 were more active against KB and HepG2 compared to the LU-1 and MCF7 cells.     

Activity of the natural algicide, cyanobacterin, on angiosperms

Cyanobacterin is a secondary metabolite produced by the cyanobacterium (blue-green alga) Scytonema hofmanni. The compound had previously been isolated and chemically characterized. It was shown to inhibit the growth of algae at a concentration of approximately 5 micromolar. Cyanobacterin also inhibited the growth of angiosperms, including the aquatic, Lemna, and terrestrial species such as corn and peas. In isolated pea chloroplasts, cyanobacterin inhibited the Hill reaction when p-benzoquinone, K₃Fe(CN)₆, dichlorophenolindophenol, or silicomolybdate were used as electron acceptors. The concentration needed to inhibit the Hill reaction in photosystem II was generally lower than the concentration of the known photosystem II inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethyl urea. Cyanobacterin had no effect on electron transport in photosystem I. The data indicate that cyanobacterin inhibits O₂ evolving photosynthetic electron transport in all plants and that the most probable site of action is in photosystem II.     

Studies on Growth Inhibition of Hiochi-bacteria,Specific Saprophytes of Saké

The authors have succeeded in isolating an antibiotic, from the cultured filtrate of Asp. oryzae, responsible for, at least, the majority of the active substances against hiochi-bacteria, as pale yellow crystals of m.p. 152~153°C. It has been proved that this antibiotic is identical with hydroxyaspergillic acid. The minimum concentration for complete inhibition in diluted Saké-peptone medium against true hiochi-bacilli, Lactobacillus homohiochii H-42 and Lactobacillus heterohiochii H-l, was 10 γ/ml, and that against hiochi-lactobacilli H-7 and H-34 20 γ/ml.     

Studies on the production and purification of an antimicrobial compound and taxonomy of the producer isolated from the marine environment of the Sundarbans

A microorganism isolated from the Sundarbans region of the Bay of Bengal, India, showed potent antimicrobial activity against gram-positive and gram-negative bacteria, molds, yeast and several multiple-drug-resistant (MDR) bacteria, including methicillin-resistant Staphylococcus aureus (MRSA). The isolate grew in the presence of 20% (w/v) NaCl, antibiotic production being maximum with 5% (w/v) NaCl in the production medium. Natural seawater stimulated antibiotic biosynthesis. The absence of catabolite repression during the synthesis of the antimicrobial substance was demonstrated by the utilization of glucose by this isolate. The 16S rRNA gene of this aerobic, gram-positive, mycelium- and spore-forming microorganism was amplified, and molecular phylogenetic analysis of the DNA sequence showed less than 93% similarity with its closest relative, indicating differentiation at the genus level. The highly stable, active principle was purified by butyl acetate extraction and silica-gel chromatography and a single compound was found to posses the broad-spectrum activity. Molecular characterization showed that the active compound is a lipid. Bioreactor studies demonstrated that antibiotic production is strongly dependent on the scale of operation and there is a definite relation between the dissolved oxygen concentration, medium pH, glucose utilization, cell differentiation and antibiotic production. Maximum production in 30 h could be obtained by regulation of the medium pH in the alkaline range by a combination of controlled addition of NaOH, regulation of the air supply and changes in the reactor configuration. Considering all of the above evidences and based on comparison with the current literature, a novel antimicrobial appears to have been isolated.     

Directed biosynthesis of novel derivatives of echinomycin by Streptomyces echinatus. I. Effect of exogenous analogues of quinoxaline-2-carboxylic acid on the fermentation

Streptomyces echinatus A8331 cultured on a maltose minimal salts medium normally produces a single antibiotic, echinomycin (quinomycin A), containing two quinoxaline-2-carbonyl chromophores. Echinomycin is powerfully active against experimental tumours and can be assayed by its activity against Gram-positive bacteria. Grown in the presence of aromatic carboxylic acids related to quinoxaline, S. echinatus responds in favourable circumstances by incorporating the added material into analogues of the natural antibiotic having replacement chromophores. Both mono- and bis-substituted derivatives are formed. With quinoline-2-carboxylic acid as precursor, large quantities of analogues are produced, and the time course of synthesis, extraction, purification, assay, and characterization of the derivatives are described. Twenty-two other aromatic acids have been tested as potential substrates for antibiotic analogue biosynthesis. Half of them did not significantly affect growth and echinomycin production. Five appeared to stimulate antibiotic synthesis, while the remainder proved inhibitory. New biologically active antibiotics were detected in cultures supplemented with 7-chloroquinoxaline-2-carboxylic acid; 1,2,4-benzo-as-triazine-3-carboxylic acid; thieno[3,2-b]pyridine-5-carboxylic acid; and 6-methylquinoline-2-carboxylic acid.     

Characterization of a mutant of Anacystis nidulans r2 resistant to the natural herbicide, cyanobacterin

Cyanobacterin, a secondary metabolite produced by the cysnobacterium, Scytonema hofmanni, inhibits electron transport at a site in photosystem II. It was previously shown that a DCMU-resistant mutant of A. nidulans R2 was still susceptible to cyanobacterin (Gleason et al., Plant Science, 46 (1986) 5–10). Apparently, cyanobacterin acts at a site different from that of DCMU and similar PS II inhibitors. To confirm this conclusion, a cyanobacterin-resistant strain of A. nidulans R2 was produced by nitrosoguanidine mutagenesis and selected by growth in the presence of 4.7 μM cyanobacterin. Hill activity in mutant thylakoids was compared to that of the wild type membranes in the presence of ferricyanide and silicomolybdate as electron acceptors. Photosynthetic electron transport in the mutant membranes shows a high degree of resistance to cyanobacterin in both reactions. In contrast, the mutant exhibits the same susceptibility to DCMU inhibition as the wild type R2. Cyanobacterin acts at a unique site, inhibiting electron flow from quinone-A to quinone-B.     

Staining the nuclei in cells and protoplasts of living yeasts,moulds and green algae with the antibiotic lomofungin

Lomofungin, an antibiotic active against yeasts, mycelial fungi and bacteria imparts a red colour to living yeast, fungal and algal nuclei when it is present in the culture medium at concentration of 20–100 g/ml. Staining is equally distinct at all stages of the cell cycle, including mitosis and meiosis. The coloured nuclei are plainly visible in the light microscope and the presence in them of lomofungin has been confirmed by electron microscopy.Non-Common Abbreviations YNBG yeast nitrogen base glucose medium - DMSO dimethylsulfoxyde     

Glutamine synthetase from pig brain: binding of adenosine triphosphate.

Glutamine synthetase was purified to homogeneity from fresh brain homogenates by a procedure that makes use of the affinity of this ATP requiring ligase to Cibacron 3G-A Blue-Sephadex G-75. It is shown that the interaction of pig brain enzyme with the dye does not concern the active centre but a non-catalytic although specific ATP binding site. Both sides contain a cysteine residue which may react with N-ethylmaleimide. The unprotected succinimidated enzyme is inactive, and aggregates. ATP alone protects only against aggregation, not against inactivation; the ATP/Mg2 complex also hinders inactivation. A tryptic heptadecapeptide isolated from the derivatized enzyme representing the carboxy terminal seems to belong to the active centre; its amino acid sequence was partially determined.     

Production,by filamentous,nitrogen-fixing cyanobacteria,of a bacteriocin and of other antibiotics that kill related strains

Colonies of sixty-five filamentous cyanobacteria were screened for the production of temperate phages and/or antibiotics on solid medium. None of them was observed to release phages. However, seven N₂-fixing strains were found to produce antibiotics very active against other cyanobacteria. The antibiotic produced by Nostoc sp. 78-11 A-E represents a bacteriocin of low molecular weight. Nostoc sp. ATCC 29132 appears to secrete, together with an antibiotic, a protein that inhibits its action.     

Characterization and partial purification of a broad spectrum antibiotic AS-48 produced by Streptococcus faecalis

Streptococcus faecalis S-48 produces a broad spectrum antibiotic, active against Gram-positive and Gram-negative bacteria. This substance is produced in solid and liquid media and also in a defined basal medium. It is sensitive to protease, pronase, or trypsin, heating at 70 degrees C, and alkaline pH, but resistant to treatment with lipase, lysozyme, alkaline phosphatase, DNAase, RNAase, acidic or neutral pHs, and also lower temperatures (60 degrees C). Several organic solvents cause precipitation, but not inactivation. This antibiotic has been partially purified by gel filtration and further ion-exchange chromatography. Its molecular weight has been estimated close to 2000. The biological activity of this antagonistic substance against the selected indicator strains, Streptococcus faecalis S-47 and Escherichia coli U-9, is bactericidal. The characterization of this substance, initially classified as a bacteriocin, indicates that it is an antibiotic of peptidic nature. The significance of antibiotic occurrence in group D of the genus Streptococcus is also discussed.     

Isolation and Characterization of Antibacterial Compound from a Mangrove-Endophytic Fungus, Penicillium chrysogenum MTCC 5108

Microorganisms, especially endophytic fungi that reside in the tissue of living mangrove plants, seem to play a major role in meeting the general demand for new biologically active substances. During the course of screening for biologically active secondary metabolites from marine microorganisms, an antibiotic compound containing an indole and a diketopiperazine moiety was isolated from the culture medium of Penicilliumchrysogenum, (MTCC 5108), an endophytic fungus on the mangrove plant Porteresiacoarctata (Roxb.). The cell free culture medium of P. chrysogenum showed significant activity against Vibriocholerae, (MCM B-322), a pathogen causing cholera in humans. Bioassay guided chemical characterization of the crude extract led to the isolation of a secondary metabolite possessing a molecular formula C₁₉H₂₁O₂N₃. Its antibacterial activity was comparable with standard antibiotic, streptomycin. This compound (1) was found to be (3,1′-didehydro-3[2″(3′″,3′″-dimethyl-prop-2-enyl)-3″-indolylmethylene]-6-methyl pipera-zine-2,5-dione) on the basis of mass spectrometry, infrared spectroscopy and one and two-dimensional nuclear magnetic resonance analysis.     

Violet-Pigmented Pseudomonads With Antifungal Activity From the Rhizosphere of Beans

Bean rhizosphere bacteria antagonistic to four root-infecting fungi and an antibiotic produced by these bacteria were studied. The bacteria were violet-pigmented gram-negative rods, probably belonging to the genus Pseudomonas. The antibiotic, which was localized largely in the bacterial cell mass, was easily extracted with acetone. It was selectively active against a wide variety of plant-pathogenic and saprophytic fungi tested in vitro but was relatively inactive against bacteria. The compound, partially purified by chromatography, was soluble in all organic solvents tried, but nearly insoluble in water. It demonstrated no characteristic ultraviolet- or visible-absorption spectrum and was chemically unidentified. The antagonistic bacteria or crude antibiotic applied to buried buckwheat segments suppressed the colonization of this substrate by Rhizoctonia spp. The data suggested that the bacteria or the antibiotic may play a role in the suppression of root-infecting fungi in soil.     

Cloning of a DNA region of Actinoplanes teichomyceticus conferring teicoplanin resistance

Teicoplanin is a glycopeptide antibiotic, produced by Actinoplanes teichomyceticus, active against Gram positive bacteria and recently introduced into clinical practice. It blocks cell wall biosynthesis by inhibiting peptidoglycan polymerization. The mechanism(s) of resistance of the producer strains of this class of antibiotics have not yet been characterized. We have constructed a genomic bank of A. teichomyceticus in Streptomyces lividans. A clone from this bank, PTR168, was able to confer resistance to teicoplanin on its sensitive host. The restriction map of plasmid pTR168 and the hybridization pattern to A. teichomyceticus DNA were determined; we have also studied the mechanism of this resistance which seems correlated with a reduced binding of the antibiotic to the cell wall.     

MYCOBACTOCIDIN, A NEW ANTIBIOTIC ACTIVE AGAINST MYCOBACTERIA.

Fregnan, G. B. (University of Wisconsin, Madison) and D. W. Smith. Mycobactocidin, a new antiboitic active against mycobacteria. J. Bacteriol. 83:1069-1076. 1962.-A new antibiotic, produced by a strain of Staphylococcus epidermidis, is described. The name mycobactocidin is proposed because of the specificity of the antibiotic against mycobacteria. Methods for extraction and purification of the antibiotic are described. The water-soluble fraction is shown to be active in vitro. No toxicity could be detected in mice after intraperitoneal or subcutaneous injection. Enzymatic hydrolysis suggests a glyco-protein nature for the antibiotic. Mycobactocidin is soluble in distilled water at neutrality but is generally insoluble in organic solvents. The antibiotic is precipitated from aqueous solution by heavy metals. It is stable in the pH range of 2 to 8, but is precipitated at pH 3 to 4. An aqueous solution of the antibiotic is stable for several days at 5 C, and for longer periods if lyophilized and stored at -20 C. It does not pass through a dialysis membrane and its activity is retained after 15 to 20 hr of dialysis at 5 C. The activity was shown to be unrelated to the formation of hydrogen peroxide.     

Production of a potentially novel anti-microbial substance by Bacillus polymyxa

A bacterial strain, SCE2, identified as Bacillus polymyxa, produced an anti-microbial substance active against yeasts, fungi and different genera of Gram-positive and-negative bacteria, in liquid medium and in plate assays. This substance appeared to be an antibiotic different from the polymyxin group, mainly because of its action against the majority of Gram-positive bacteria tested and its lack of activity against Pseudomonas aeruginosa, a species usually killed by polymyxins. Preliminary characterization showed resistance to heat (65°C, 2 h), to proteases, trypsin, lysozyme, deoxyribonuclease I, ribonuclease A, phospholipase C, ethanol, acetone, chloroform, ether and to strong alkali treatment (2 M NaOH). The molecular weight was less than 3500. The B. polymyxa strain harboured a plasmid that did not correlate with antibiotic production; after curing experiments, a derivative strain, SCE2(46), was isolated that lacked the plasmid pES1, but showed the same inhibitory spectrum as the wild-type strain.     

Site of action of the natural algicide,cyanobacterin, in the blue-green alga,Synechococcus sp.

Cyanobacterin is a secondary metabolite produced by the cyanobacterium, Scytonema hofmanni. Highly purified cyanobacterin was found to inhibit the growth of many cyanobacteria at a minimum effective dose of 2 g/ml (4.6 M). The antibiotic had no effect on eubacteria including the photosynthetic Rhodospirillum rubrum. The site of action of cyanobacterin was further investigated in the unicellular cyanobacterium, Synechococcus sp. Electron micrographs of antibiotic-treated Synechococcus cells indicated that cyanobacterin affects thylakoid membrane structure. The antibiotic also inhibited light-dependent oxygen evolution in Synechococcus cells and in spheroplasts. These data support our conclusion that cyanobacterin specifically inhibits photosynthetic electron transport. This activity is similar to herbicides such as 3-(3,4-dichlorophenyl)-1,1-dimethyl urea (DCMU). The anhydro analog of cyanobacterin had no biological activity.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-dimethyl urea - DCPIP dichlorophenolindophenol     

Thermomonospora sp. T-SA-125 and its production of a growth promoting antibiotic

Thermomonospora sp. T-SA-125 is a true thermophilic actinomycete isolated from a soil sample collected from the Saudi Arabian desert. It is characterized by the formation of single spores at the tips of dichotomously branched aerial mycelium and differs from Thermomonospora curvata and T. viridis in certain aspects. It produces a basic water-soluble antibiotic which is active against Gram-positive bacteria, moderately active against Gram-negative bacteria and inactive against fungi. At high concentrations, this antibiotic, stimulated the growth of both Hordeum coleoptile and lettuce hypocotyl.     

Bacterial efflux systems and efflux pumps inhibitors

It is now well established that bacterial resistance to antibiotics has become a serious problem of public health that concerns almost all antibacterial agents and that manifests in all fields of their application. Among the three main mechanisms involved in bacterial resistance (target modification, antibiotic inactivation or default of its accumulation within the cell), efflux pumps, responsible for the extrusion of the antibiotic outside the cell, have recently received a particular attention. Actually, these systems, classified into five families, can confer resistance to a specific class of antibiotics or to a large number of drugs, thus conferring a multi-drug resistance (MDR) phenotype to bacteria. To face this issue, it is urgent to find new molecules active against resistant bacteria. Among the strategies employed, the search for inhibitors of resistance mechanisms seems to be attractive because such molecules could restore antibiotic activity. In the case of efflux systems, efflux pump inhibitors (EPIs) are expected to block the pumps and such EPIs, if active against MDR pumps, would be of great interest. This review will focus on the families of bacterial efflux systems conferring drug resistance, and on the EPIs that have been identified to restore antibiotic activity.     

The concentration dynamics of inorganic polyphosphates during the cephalosporin C synthesis by Acremonium chrysogenum

The contents of five fractions of energy-rich inorganic polyphosphates (polyPs), ATP, and H⁺-ATPase activity in the plasma membrane were determined in a low-activity cephalosporin C (cephC) producer Acremonium chrysogenum ATCC 11550 and selected highly efficient producer strain 26/8 grown on glucose or a synthetic medium providing for active synthesis of this antibiotic. It was shown that strain 26/8 on the synthetic medium produced 26-fold higher amount of cephC as compared with strain ATCC 11550. This was accompanied by a drastic decrease in the cell contents of ATP and the high-molecular-weight fractions polyP2, polyP3, and polyP5 with a concurrent increase in the low-molecular-weight fraction polyP1. These data suggest that polyPs are involved in the cephC synthesis as a source of energy. H⁺-ATPase activity insignificantly changed at both low and high levels of cephC production. This confirms the assumption that A. chrysogenum has other alternative antibiotic transporters in addition to cefT. The obtained results can be used for optimizing commercial-scale cephC biosynthesis.