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Inability to find consistent bacterial biocontrol agents of Pythium aphanidermatum in cucumber using screens based on ecophysiological traits.

A collection of 821 rhizobacteria from cucumber, originating from different root locations and stages of plant development, was screened for potential biocontrol agents of Pythium aphanidermatum (Edson) Fitzp. The screening procedure exploited carbon source utilization profiles and growth rates of bacteria as indicators of a partial niche overlap with the pathogen. The bacteria were tested for growth on nine carbon sources (glucose, fucose, sucrose, maltose, asparagine, alanine, galacturonic acid, succinic acid, and linoleic acid), most of which are reported to be used by the zoospores of P. aphanidermatum in the infection process. The isolates were classified as fast- or slow-growing, depending on their growth rate in 1/10 strength TSB. By nonhierarchical cluster analysis, 20 clusters were generated of bacteria with similar profiles of carbon source utilization. Redundancy analysis showed that the type of root sample explained 47% of the variance found in the relative abundance of bacteria from the clusters. Bacteria from clusters using none or few of the carbon sources, e.g., maltose and linoleic acid, with many slow-growing isolates, showed a preference for plants in the vegetative or generative stage, or for old root regions (root base). Bacteria from clusters with fast-growing isolates, using many carbon sources, were relatively abundant in the seedling stage. A selection of 127 bacteria from the different clusters was tested for disease suppressive capabilities in bioassays on young cucumber plants in nutrient solution, inoculated with zoospores of P. aphanidermatum. Nine of these bacteria produced biosurfactants, and 27 showed antibiosis against mycelial growth in plate assays. For 31 isolates, significant positive effects on plant biomass were shown, as analyzed with a general linear regression model. For most isolates, these effects occurred only in one of two replicate assays and no reductions in the degree of root and crown rot were found. Of the isolates that used many of the tested carbon sources, only four had positive effects on plant biomass. The majority of the isolates that positively affected plant biomass used few to moderate numbers of carbon sources and did not produce antibiotics or biosurfactants. In conclusion, competition for the tested carbon sources with the zoospores did not play a decisive role in disease suppression, and no clear relation was found between ecophysiological traits and disease suppression. Only isolate 3.1T8, isolated from root tips in the generative stage of plant growth, significantly increased plant biomass and suppressed root and crown rot symptoms in five out of six bioassays. The isolate produced an antifungal substance in plate assays and showed biosurfactant production in several (cucumber-derived) media.

Bacteria↗

Chelating agents in the treatment of acute vanadyl sulphate intoxication in mice.

Eighteen chelating or reducing agents were tested to determine their relative efficacy as antagonists in acute intramuscular vanadyl sulphate intoxication in mice. The chelating or reducing agents were administered intraperitoneally to male Swiss mice at doses equal to one-fourth of their respective LD50. Therapeutic effectiveness (TEF) was calculated. In a subsequent experiment, the effect of EDTA, glutathione, DFOA, ascorbic acid, succinic acid, monosodium phosphate, Tiron, DTPA, and 2-mercaptosuccinic acid on the excretion, and distribution of vanadium was determined. Of the compounds examined, Tiron followed by ascorbic acid, and 2-mercaptosuccinic acid were effective in increasing the urinary excretion of vanadium. Tiron, and 2-mercaptosuccinic acid were also effective in reducing the concentration of vanadium found in kidney, the main target organ of vanadium accumulation. Tiron appears to be the most effective agent of those tested in the prevention of acute vanadium (IV) intoxication in mice.

Animals↗

Decoloration of aqueous Brilliant Green by using glow discharge electrolysis.

This paper described a plasma degradation of Brilliant Green (BG) by glow discharge electrolysis. Various influencing factors such as the voltage, the distance between cathode and anode were examined. Ultraviolet (UV) absorption spectra, gas chromatogram-mass spectrum (GC-MS), and chemical oxygen demand (COD) were used to monitor the degradation process and to identify the major oxidation intermediates. It was confirmed that benzoic acid, 1,2,3,4,5,6-cyclohexanehexaol, and carboxylic acids (e.g., oxalic acid, succinic acid and hydroxyacetic acid) were produced in the degradation process. The results showed that BG rapidly underwent degradation and eventually mineralized into CO(2) and H(2)O.

Carbon Dioxide↗

Characterization of salicylate uptake across the basolateral membrane of the malpighian tubules of Drosophila melanogaster.

The organic anion salicylate is a plant secondary metabolite that can protect plants against herbivores. Transport of salicylate across the basolateral membrane of the Malpighian tubules of Drosophila melanogaster was studied using a radioisotope tracer technique. The uptake of [(14)C]salicylate by the Malpighian tubules was active, saturable and Na(+)-dependent; the maximum uptake rate (J(max)) and the half saturation concentration (K(t)) were 12.6 pmoltubule(-1)min(-1) and 30.7micromoll(-1), respectively. In contrast to organic anion transport by vertebrate renal tissues, salicylate uptake was not trans-stimulated by glutarate (0.01-1.0 mmoll(-1)) or cis-inhibited by high concentrations (5 mmoll(-1)) of various alpha-keto acids (glutaric acid, alpha-ketoglutaric acid, succinic acid, and citric acid). Changes in basolateral membrane potential or physiologically relevant changes in bathing saline pH did not affect the rate of [(14)C]salicylate uptake. Ring-structure monocarboxylic acids (benzoic acid, nicotinic acid, gentisic acid, unlabelled salicylic acid, alpha-cyano-4-hydroxycinnamic acid, probenecid, fluorescein, and P-aminohippuric acid) strongly inhibited [(14)C]salicylate uptake rate. In contrast, short-chain monocarboxylic acids had little (butyric acid) or no effect (lactic acid, pyruvic acid, and propionic acid). Our results suggest that salicylate uptake across the basolateral membrane of D. melanogaster Malpighian tubules is mediated by a non-electrogenic, alpha-cyano-4-hydroxycinnamic acid-sensitive, Na(+):salicylate cotransport system.

Animals↗

Directed evolution of Lactiplantibacillus plantarum for utilizing ethanol to produce postbiotics.

Alcohol is a recognized carcinogen worldwide. In this study, we aimed to utilize probiotics to metabolize ethanol and produce postbiotics. Initially, we identified a lactic acid bacteria community in kimchi with excellent probiotic activity. By employing our previously developed directed evolution techniques, a Lactiplantibacillus plantarum mutant with safe characteristics and an ethanol utilization capacity of 40 g/L and 0.15 g/L/OD was obtained. Genome sequencing and RT-qPCR analysis revealed the up-regulated expression of alcohol dehydrogenase and aldehyde dehydrogenase genes greatly contributed to ethanol utilization. Furthermore, the mutant strain demonstrated marked superiority in producing postbiotics, including antimicrobial peptides and beneficial organic acids such as lactic acid, phenyllactic acid, succinic acid, and indole-3-lactic acid. In the ethanol-fed fermentation process, the mutant strain achieved a lactic acid yield of 8.47 g/L and a carbon conversion rate of 21.8%. In vivo testing further validated its safety and ability to assist alcohol metabolism.

Adaptive laboratory evolution↗

Gas chromatographic-mass spectrometric analysis of acids and phenols in distilled alcohol beverages. Application of anion-exchange disk extraction combined with in-vial elution and silylation.

A GC-MS protocol for profiling spirits, based on 19 acids and phenolic compounds, has been proposed and evaluated. The method combined a simple preconcentration procedure based on solid-phase (anion-exchange) disk extraction, and in-vial elution and silylation of the analytes. The derivatized extract was directly injected into the GC-MS system. These analytes were: C6, C8, C10, C12 acids, pyruvic acid, 2-furoic acid, succinic acid, fumaric acid, glutaric acid, lactic acid, glycolic acid, malic acid, tartaric acid, citric acid, vanillin, syringaldehyde, coniferaldehyde, vanillic acid and gallic acid. The profiles of six different spirits were found reproducible from day-to-day with <20% RSD for measurements of most of the analytes at different concentrations. Recoveries of individual analytes appear to be affected by the level of tannins in the spirits, and they varied from sample to sample. The method of standard addition was used to quantify age-related analytes. Good linearity of response with correlation coefficients in the range of 0.992-0.999 was obtained. The results of the study indicate that for spirits of the same brand but of different ages, the amounts of these analytes appear to increase with the ageing period.

Acids↗

Influence of formulation on the physicochemical properties of casein microparticles.

Casein microparticles (CAS/MP) have a potential clinical use for targeting drugs. However, the use of organic solvents in their preparation is undesirable. This study was designed to investigate the influence of preparation procedures in aqueous media on the formulation and physicochemical properties of CAS/MP. The first stage involved the influence of the coacervating agents (lactic acid, succinic anhydride, succinic acid and tartaric acid). The second stage studied was the influence of the ionic strength and the third, the influence of adding a thickener, hydroxypropyl cellulose or hydroxypropyl methycellulose (HPC or HPMC), and a plasticizing agent (gelatin). Some physicochemical properties of CAS/MP were evaluated. While the infrared and the thermal analysis showed that all coacervating agents were appropriate for coacervation, the scanning electron microscopy studies showed that the external morphology of the particles was more homogeneous when lactic acid was used. Utilizing lactic acid as the coacervating agent, there was a trend effect of adding NaCl implying that the increasing of the ionic strength resulted in better stability. Finally, the addition of 0.1% HPC plus either 0.25 or 0.5% gelatin resulted in homogeneous formulations. In conclusion, the use of lactic acid plus 0.1% HPC and 0.25% gelatin results in biodegradable and homogeneous CAS/MP, presenting a potentially useful drug delivery system.

Animals↗

Characterization of cigar tobaccos by gas chromatographic/mass spectrometric analysis of nonvolatile organic acids: application to the authentication of Cuban cigars.

A reliable method based on gas chromatographic/mass spectrometric (GC/MS) profiling of nonvolatile organic acids is described for the characterization of cigars. The method involves an aqueous extraction of ground tobacco and selective isolation of the acids by simply stirring strong anion exchange (SAX) disks in the aqueous tobacco extract. The acids are then directly silylated on the disk with N-methyl-N-trimethylsilyl-trifluroacetamide (MSTFA) in acetonitrile in an autosampler vial. Elution of the derivatized acids in situ allows the sample to be directly analyzed by GC/MS without further sample handling. Compared to the conventional disk-extraction technique using a vacuum manifold, this method is much less labor intensive, and is desirable for multiple sample analysis. Nicotinic acid, succinic acid, glyceric acid, malic acid, pyroglutamic acid, threonic acid, citric acid, uracil, and an unidentified acid were reproducibly quantified in tobacco samples. Principal component analysis (PCA) of the acid profiles of the filler tobaccos of 18 Cuban cigars and 31 non-Cuban cigars shows separation of the two groups, indicating that the acid profiles are potentially useful in the authentication of Cuban cigars.

Carboxylic Acids↗

Micropore to macropore structure-designed silicas with regulated condensation of silicic acid nanoparticles.

A new preparation method for porous silica particles was developed using activated silica sols which are called nano-silica solutions in this paper. Several kinds of organic and inorganic acids are employed to neutralize diluted sodium silicate solutions to form the nano-silica solutions: formic acid, acetic acid, propionic acid, oxalic acid, succinic acid, dl-malic acid, citric acid, and tricarballylic acid as carboxylic acids, and sulfuric acid and hydrochloric acid as inorganic acids. The effect of salts in the nano-silica solution is also studied. The products were investigated using a field emission scanning electron microscope, an X-ray diffractometer, the nitrogen adsorption technique, and a mercury porosimeter. Microporous silicas were produced when carboxylic acids were applied; the formation of micropores was influenced by the pH of the nano-silica solutions and molecular sizes of the carboxylic acids. Addition of a salt in a citric acid solution increased the mesopore volume. Macropores were formed when inorganic acids including salts were applied; the salt nanoparticles which were crystallized in silica spheres acted as templates. The anion types and salt concentrations in the nano-silica solutions affected the aggregation condition of silica nanoparticles, following the Schulze-Hardy rule.

Journal Article↗

Identification of 3-hydroxy-3-methylglutaric acid (HMG) as a hypoglycemic principle of Spanish moss (Tillandsia usneoides).

Bioactivity-directed fractionation, using brine shrimp lethality and murine hypoglycemia, of an ethanol extract prepared from Tillandsia usneoides, led to the isolation of four apparently bioactive compounds from the water-soluble fraction. The compounds were identified as citric acid, succinic acid, 3-hydroxy-3-methylglutaric acid (HMG), and 3,6,3',5'-tetramethoxy-5,7,4'-trihydroxyflavone-7-O-beta-D-g lucoside. The brine shrimp lethality of the acids was simply due to acidity; however, HMG elicited significant hypoglycemic responses in fasting normal mice. Ethyl and methyl esters of citric acid were prepared and tested in the murine hypoglycemic assay. Five of the predominant sugars were identified by tlc. Free thymidine was also isolated. Further evaluation of HMG and other potential inhibitors of HMG CoA lyase, in the treatment of symptoms of diabetes mellitus, is suggested.

Animals↗

Evaluation of potential strontium chelators in an octanol-water system.

90Sr has a physical half-life of sufficient duration to make it a potentially dangerous contaminant from nuclear accidents and radioactive wastes. In the present study, the efficacy of 16 compounds as potential chelators of strontium was tested in vitro. Strontium solubilization from strontium carbonate and its distribution in an octanol-water system (Do/w) was determined in the absence and presence of alpha-ketoglutaric acid, Kryptofix 222, ethylenglycol-bis-(beta-amino-ethylether)-N,N-tetraacetic acid, diethylentriamine pentaacetic acid, Kryptofix 5, disodium chlodronate, disodium ethidronate, oxaloacetic acid, fumaric acid, D-gluconic acid, succinic acid, citric acid, D,L-2,3-diaminopropionic acid, 1,1-cyclohexanediacetic acid, tartaric acid, and trans-1,2-cy-clohexanediol. Kryptofix 222 and Kryptofix 5 significantly increased solubilized strontium, suggesting strontium chelation potential. Since in previous in vivo studies both compounds were also effective in the removal of strontium following internal contamination, it is concluded that the octanol-water system may be useful screening compounds with strontium chelation potential.

Bridged Bicyclo Compounds↗

Porphyromonas gulae sp. nov., an anaerobic, gram-negative coccobacillus from the gingival sulcus of various animal hosts.

A new species, Porphyromonas gulae sp. nov., is proposed to include strains isolated from the gingival sulcus of various animal hosts which are distinct from related strains of Porphyromonas gingivalis of human origin. This bacterium exhibits the following characteristics: black-pigmented colonies; asaccharolytic, obligate anaerobic growth; and Gram-negative, non-motile and non-spore-forming, rod-shaped cells. Colonies do not fluoresce under UV light. Vitamin K1 and haemin are required for growth. Cells haemagglutinate sheep erythrocytes. Major fatty acid end products are butyric acid, isovaleric acid, succinic acid and phenylacetic acid. Strains are catalase-positive and indole is produced. Alkaline phosphatase, trypsin-like and N-acetyl-beta-glucosaminidase activities are strong. A beta-galactosidase and a glutamylglutamic acid arylamidase are also present. The G+C content of the chromosomal DNA is 51 mol%. DNA-DNA homology data and 16S rRNA gene sequence analysis provide strong evidence that strains from the animal biotype of P. gingivalis represent a Porphyromonas species that is distinct from P. gingivalis. The type strain of P. gulae is Loup 1T (= ATCC 51700T = NCTC 13180T).

Animals↗

Production of alternatives to fuel oil from organic waste by the alkane-producing bacterium, Vibrio furnissii M1.

AIMS: We investigated the production of alternatives to fuel oil through the bacterial metabolism of organic waste. The availability for this purpose of various sources of organic waste for hydrocarbon production by the alkane-producing bacterium, Vibrio furnissii M1, was examined. METHODS AND RESULTS: We screened 17 authentic compounds which can generally be found in organic waste for their hydrocarbon production. Carbon (3 mmol) in a 50-ml culture with acetic acid, lactic acid, butyric acid, succinic acid, malic acid, pentanoic acid, hexanoic acid glucose, xylose, starch or sucrose yielded 10-27 mg of alkanes or alkenes. The chain length of these alkanes or alkenes varied according to the culture from C14 to C27. Varying the ratio of carbon to nitrogen in the culture had no effect on the hydrocarbon production. Crude blackstrap molasses were also converted into alkanes with a conversion ratio of 20% (half of that in an authentic sucrose medium) of the total carbon consumption. CONCLUSIONS: V. furnissii M1 could produce hydrocarbons corresponding to kerosene or light oil from volatile fatty acids and sugars. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on bacterial hydrocarbon production from organic waste.

Alkanes↗

Tetrachloroethene transformation to trichloroethene and cis-1,2-dichloroethene by sulfate-reducing enrichment cultures.

Tetrachloroethene, also known as perchloroethylene, was reductively dechlorinated to trichloroethene and cis-1,2-dichloroethene by laboratory sulfate-reducing enrichment cultures. The causative organism or group was not identified. However, tetrachloroethene was dechlorinated to trichloroethene in 50 mM bromoethane-sulfonate-inhibited enrichments and to trichloroethene and cis-1,2-dichloroethene in 3 mM fluoroacetate-inhibited enrichments. Overall transformation varied from 92% tetrachloroethene removal in 13 days to 22% removal in 65 days, depending on conditions of the inoculum, inhibitor used, and auxilliary substrate used. Neither lactate, acetate, methanol, isobutyric acid, valeric acid, isovaleric acid, hexanoic acid, succinic acid, nor hydrogen appeared directly to support tetrachloroethene dechlorination, although lactate-fed inocula demonstrated longer-term dechlorinating capability.

Bacteria↗

Influence of the nitrogen source on Saccharomyces cerevisiae anaerobic growth and product formation.

To prevent the loss of raw material in ethanol production by anaerobic yeast cultures, glycerol formation has to be reduced. In theory, this may be done by providing the yeast with amino acids, since the de novo cell synthesis of amino acids from glucose and ammonia gives rise to a surplus of NADH, which has to be reoxidized by the formation of glycerol. An industrial strain of Saccharomyces cerevisiae was cultivated in batch cultures with different nitrogen sources, i.e., ammonium salt, glutamic acid, and a mixture of amino acids, with 20 g of glucose per liter as the carbon and energy source. The effects of the nitrogen source on metabolite formation, growth, and cell composition were measured. The glycerol yields obtained with glutamic acid (0.17 mol/mol of glucose) or with the mixture of amino acids (0.10 mol/mol) as a nitrogen source were clearly lower than those for ammonium-grown cultures (0.21 mol/mol). In addition, the ethanol yield increased for growth on both glutamic acid (by 9%) and the mixture of amino acids (by 14%). Glutamic acid has a large influence on the formation of products; the production of, for example, alpha-ketoglutaric acid, succinic acid, and acetic acid, increased compared with their production with the other nitrogen sources. Cultures grown on amino acids have a higher specific growth rate (0.52 h-1) than cultures of both ammonium-grown (0.45 h-1) and glutamic acid-grown (0.33 h-1) cells. Although the product yields differed, similar compositions of the cells were attained. The NADH produced in the amino acid, RNA, and extracellular metabolite syntheses was calculated together with the corresponding glycerol formation. The lower-range values of the theoretically calculated yields of glycerol were in good agreement with the experimental yields, which may indicate that the regulation of metabolism succeeds in the most efficient balancing of the redox potential.

Amino Acids↗

L-pyroglutamate spontaneously formed from L-glutamate inhibits growth of the hyperthermophilic archaeon Sulfolobus solfataricus.

Identification of physiological and environmental factors that limit efficient growth of hyperthermophiles is important for practical application of these organisms to the production of useful enzymes or metabolites. During fed-batch cultivation of Sulfolobus solfataricus in medium containing L-glutamate, we observed formation of L-pyroglutamic acid (PGA). PGA formed spontaneously from L-glutamate under culture conditions (78 degrees C and pH 3.0), and the PGA formation rate was much higher at an acidic or alkaline pH than at neutral pH. It was also found that PGA is a potent inhibitor of S. solfataricus growth. The cell growth rate was reduced by one-half by the presence of 5.1 mM PGA, and no growth was observed in the presence of 15.5 mM PGA. On the other hand, the inhibitory effect of PGA on cell growth was alleviated by addition of L-glutamate or L-aspartate to the medium. PGA was also produced from the L-glutamate in yeast extract; the PGA content increased to 8.5% (wt/wt) after 80 h of incubation of a yeast extract solution at 78 degrees C and pH 3.0. In medium supplemented with yeast extract, cell growth was optimal in the presence of 3.0 g of yeast extract per liter, and higher yeast extract concentrations resulted in reduced cell yields. The extents of cell growth inhibition at yeast extract concentrations above the optimal concentration were correlated with the PGA concentration in the culture broth. Although other structural analogues of L-glutamate, such as L-methionine sulfoxide, glutaric acid, succinic acid, and L-glutamic acid gamma-methyl ester, also inhibited the growth of S. solfataricus, the greatest cell growth inhibition was observed with PGA. We also observed that unlike other glutamate analogues, N-acetyl-L-glutamate enhanced the growth of S. solfataricus. This compound was stable under cell culture conditions, and replacement of L-glutamate with N-acetyl-L-glutamate in the medium resulted in increased cell density.

Culture Media↗

CARBON DIOXIDE FIXATION IN BACILLUS ANTHRACIS.

Eastin, Jerry D. (U.S. Army Chemical Corps, Frederick, Md.) and Curtis B. Thorne. Carbon dioxide fixation in Bacillus anthracis J. Bacteriol. 85:410-417. 1963.-Virulent strains of Bacillus anthracis require a concentration of CO(2) greater than that of the normal atmosphere (air) for the production of capsular material (glutamyl polypeptide); avirulent strains may produce no polypeptide or may produce polypeptide in air. Fixation of C(14)O(2) by each of the three types tested resulted in labeling of aspartic acid, glycine, glutamic acid, succinic acid, and an unidentified organic acid. C(14) was detected in aspartic acid after less than 30 sec of exposure of cells to C(14)O(2). Subsequent flushing of the cells with C(12)O(2) displaced C(14) from aspartic acid but not from the other labeled intermediates. Aspartic acid appears to be closely associated with the primary CO(2)-fixation product, and the data suggest a fairly direct carbon pathway from CO(2) to aspartic acid (oxaloacetic acid) to glutamic acid to glutamyl polypeptide.

Journal Article↗

Growth and sporulation of Bacillus subtilis mutants blocked in the pyruvate dehydrogenase complex.

Two "ACE" mutants of Bacillus subtilis which require acetate for growth on glucose minimal medium have been isolated. They do not grow with acetoin, 2,3-butanediol, fatty acids, isoleucine, lipoic acid, malic acid, pyruvic acid, succinic acid, thiamine, or valine, but respond somewhat to glutamate or citrate. The mutants lack the activity of the pyruvate dehydrogenase complex; they excrete pyruvate and later acetoin. They grow in nutrient sporulation medium (NSMP) to one-half the normal turbidity and do not sporulate subsequently. When acetate is added to NSMP (at the optimal concentration of 0.07 m), the ACE mutants grow to the normal turbidity and then sporulate normally. Growth but not sporulation is restored in NSMP upon addition of 2,3-butanediol, citrate, glucose, glutamate, glycerol, or ribose, but not upon addition of acetoin, malate, oxaloacetate, pyruvate, and several other compounds. After growth in NSMP has stopped, the mutants incorporate uracil only at a very low rate, which can be increased by the addition of acetate, citrate, or glutamate. Furthermore, the metabolism of acetoin is prevented after growth has stopped but can be restored by the addition of acetate. All these results can be explained by a lack of reduced nicotinamide adenine dinucleotide (NADH) resulting from the deficiency in acetylcoenzyme A. In fact, after growth of the ACE mutants had stopped, the NADH concentration was at the borderline of measurability, whereas it increased significantly upon addition of glucose. The growing standard strain contains, at the same bacterial turbidity, at least 20 times more NADH (230 pmole/optical density unit at 600 nm) than the nongrowing ACE mutants. The isolated spores, obtained after growth in NSMP plus acetate, can be initiated to germinate in the presence of either l-alanine or the combination of l-asparagine, fructose, glucose, and potassium; addition of acetate is not required and has no effect.

Acetates↗