The consequences of Lactobacillus vini and Dekkera bruxellensis as contaminants of the sugarcane-based ethanol fermentation

This work describes the effects of the presence of the yeast Dekkera bruxellensis and the bacterium Lactobacillus vini on the industrial production of ethanol from sugarcane fermentation. Both contaminants were quantified in industrial samples, and their presence was correlated to a decrease in ethanol concentration and accumulation of sugar. Then, laboratory mixed-cell fermentations were carried out to evaluate the effects of these presumed contaminants on the viability of Saccharomyces cerevisiae and the overall ethanol yield. The results showed that high residual sugar seemed the most significant factor arising from the presence of D. bruxellensis in the industrial process when compared to pure S. cerevisiae cultures. Moreover, when L. vini was added to S. cerevisiae cultures it did not appear to affect the yeast cells by any kind of antagonistic effect under stable fermentations. In addition, when L. vini was added to D. bruxellensis cultures, it showed signs of being able to stimulate the fermentative activity of the yeast cells in a way that led to an increase in the ethanol yield.     

The effect of iodoacetate and other inhibitors on phage production and lysis in three phage systems

The minimal bacteriostatic concentration of iodoacetate, azide, or proflavine was added at intervals during the latent periods of virus in three different bacterium-bacteriophage systems (M. aureus, B. mycoides, E. coli). For each interval at which inhibitor was added, the occurrence of lysis and the final yield of phage were determined. In the B. mycoides and E. coli systems, when added during the first part of the latent period, inhibitor prevented lysis and no phage was released. Introduction of inhibitor during the last part of the latent period resulted in normal lysis and in a linear increase in phage that progressively approached the yield obtained in the absence of inhibitor (the later the introduction, the higher the yield). In the M. aureus system, phage production and lysis in the presence of inhibitor followed the same general pattern, except that release of phage and normal lysis occurred in infected cells to which inhibitor had been added quite early in the latent period. Our results, when compared with those of Foster (1948) with proflavine and Bozeman et al. (1954) with chloramphenicol, suggest that (1) the final phage yields represent the amount of mature intracellular virus present at the time of addition of inhibitor and (2) the reactions leading to lysis proceed independently of those leading to the formation of mature virus once phage infection has reached a critical point in time.     

Influence of ammonium on the biosynthesis of the macrolide antibiotic tylosin

The effect of ammonium ions on growth and tylosin biosynthesis in Streptomyces fradiae NRRL 2702 cultured on a chemically defined medium was studied. Mycelial growth and tylosin production were not affected when ammonium sulphate was added to idiophase cultures to a final concentration of 10 mm or 20 mm; however, when ammonium sulphate was added to tylosin cultures to a final concentration of 20 mm before the onset of antibiotic biosynthesis (trophophase), tylosin production was severely suppressed while mycelial growth was stimulated. The activities of propionyl-coenzyme A carboxylase (EC 6.4.1.3) and methylmalonyl-coenzyme A carboxyltransferase (EC 2.1.3.1), enzymes involved in the synthesis of tylonolide precursors, were depressed in high ammonium cultures. The activity of macrocin 3′-o-methyltransferase, which catalyses the methylation of macrocin to form tylosin, was also affected by high concentrations of ammonium ions added in the trophophase.     

Formate Oxidation-Driven Calcium Carbonate Precipitation by Methylocystis parvus OBBP

Microbially induced carbonate precipitation (MICP) applied in the construction industry poses several disadvantages such as ammonia release to the air and nitric acid production. An alternative MICP from calcium formate by Methylocystis parvus OBBP is presented here to overcome these disadvantages. To induce calcium carbonate precipitation, M. parvus was incubated at different calcium formate concentrations and starting culture densities. Up to 91.4% ± 1.6% of the initial calcium was precipitated in the methane-amended cultures compared to 35.1% ± 11.9% when methane was not added. Because the bacteria could only utilize methane for growth, higher culture densities and subsequently calcium removals were exhibited in the cultures when methane was added. A higher calcium carbonate precipitate yield was obtained when higher culture densities were used but not necessarily when more calcium formate was added. This was mainly due to salt inhibition of the bacterial activity at a high calcium formate concentration. A maximum 0.67 ± 0.03 g of CaCO₃ g of Ca(CHOOH)₂⁻¹ calcium carbonate precipitate yield was obtained when a culture of 10⁹ cells ml⁻¹ and 5 g of calcium formate liter⁻¹ were used. Compared to the current strategy employing biogenic urea degradation as the basis for MICP, our approach presents significant improvements in the environmental sustainability of the application in the construction industry.     

Effect of various factors on the biosynthesis of piscarinines,secondary metabolites of the fungus <Emphasis Type="Italic">Penicillium piscarium</Emphasis> Westling

Piscarinines A and B were synthesized most actively during the surface cultivation of the fungus Penicillium piscarium in a complex medium (5.5 mg/l). Under conditions of submerged cultivation in a mineral medium, the yield of piscarinines was two times lower. An increase in the inoculum quantity of conidia treated with Tween-80 increased the culture productivity. The biosynthesis of the alkaloid was completely suppressed when mannitol was replaced with glucose or when zinc, iron, or copper ions were added to the culture medium. The metabolites were active against the prostate cancer cell line LNCAP (IC₅₀ were 2.195 and 1.914 μg/ml for piscarinines A and B, respectively).     

A two-stage glycine supplementation strategy enhances the extracellular expression of sortase A in Escherichia coli

Sortase A (SrtA) is a transpeptidase widely used in protein engineering. In this study, the enhancement of the extracellular expression of SrtA in Escherichia coli was investigated using a combined strategy based on the PelB signal peptide and chemical additives. First, glycine was identified to be the best additive for promoting the release of SrtA from the periplasm to the medium. Then, the effect of glycine concentration on cell growth and SrtA production was investigated, and a two-stage supplementation strategy was developed in order to control the impairment of cell growth and to achieve the maximum production of secretory SrtA. The results showed that when 0.5% glycine was added to the medium at the beginning of cell growth and 1% glycine was added at the end of the exponential phase, the extracellular yield of SrtA was 228.0 mg/L and the enzyme activity was 100.4 U/mL at the end of fermentation; these values were 5.3- and 8.6-fold higher, respectively, than those attained in the control culture without any additives. This result represents the highest yield of extracellular SrtA ever reported and demonstrates a promising process for the production of SrtA for large-scale industrial application.     

Does the combined application of organic and mineral nutrient sources influence maize productivity? A meta-analysis

The combined application of organic resources (ORs) and mineral fertilizers is increasingly gaining recognition as a viable approach to address soil fertility decline in sub-Saharan Africa (SSA). We conducted a meta-analysis to provide a comprehensive and quantitative synthesis of conditions under which ORs, N fertilizers, and combined ORs with N fertilizers positively or negatively influence Zea mays (maize) yields, agronomic N use efficiency and soil organic C (SOC) in SSA. Four OR quality classes were assessed; classes I (high quality) and II (intermediate quality) had >2.5% N while classes III (intermediate quality) and IV (low quality) had <2.5% N and classes I and III had <4% polyphenol and <15% lignin. On the average, yield responses over the control were 60%, 84% and 114% following the addition of ORs, N fertilizers and ORs + N fertilizers, respectively. There was a general increase in yield responses with increasing OR quality and OR-N quantity, both when ORs were added alone or with N fertilizers. Surprisingly, greater OR residual effects were observed with high quality ORs and declined with decreasing OR quality. The greater yield responses with ORs + N fertilizers than either resource alone were mostly due to extra N added and not improved N utilization efficiency because negative interactive effects were, most often, observed when combining ORs with N fertilizers. Additionally, their agronomic N use efficiency was not different from sole added ORs but lower than N fertilizers added alone. Nevertheless, positive interactive effects were observed in sandy soils with low quality ORs whereas agronomic use efficiency was greater when smaller quantities of N were added in all soils. Compared to sole added ORs, yield responses for the combined treatment increased with decreasing OR quality and greater yield increases were observed in sandy (68%) than clayey soils (25%). While ORs and ORs + N fertilizer additions increased SOC by at least 12% compared to the control, N fertilizer additions were not different from control suggesting that ORs are needed to increase SOC. Thus, the addition of ORs will likely improve nutrient storage while crop yields are increased and more so for high quality ORs. Furthermore, interactive effects are seldom occurring, but agronomic N use efficiency of ORs + N fertilizers were greater with low quantities of N added, offering potential for increasing crop productivity.     

Substrate gradient formation in the large-scale bioreactor lowers cell yield and increases by-product formation

A heterogeneous micro-environment was identified in a 12 m³ bioreactor with a height-to-diameter ratio of 2.5. The reactor was aerated by a ring sparger and stirred by three Rushton turbines. E. coli cells were cultivated in minimal medium to a cell density in the order of 30?g/l. Samples of glucose, the growth limiting component fed to the process, were taken at three levels in the bioreactor (top/middle/bottom). These showed that glucose concentration declined away from the feedpoint. The gradients depended on the mixing characteristics of the feedpoint, and concentrations of up to 400 times the mean value were found when feed was added to a relatively stagnant mixing zone. This resulted in up to 20% lower biomass yield compared to the bench scale. Gradients also affected the by-product formation, resulting in acetate formation in the large-scale bioreactor. IPTG induction of a recombinant protein was shown to influence important cell parameters and considerably increased the yield of carbon dioxide per glucose added, indicating an increased maintenance. The product formation rate was, however, not notably affected by the scale-up.     

Engineering of the glycerol decomposition pathway and cofactor regulation in an industrial yeast improves ethanol production

Glycerol is a major by-product of industrial ethanol production and its formation consumes up to 4 % of the sugar substrate. This study modified the glycerol decomposition pathway of an industrial strain of Saccharomyces cerevisiae to optimize the consumption of substrate and yield of ethanol. This study is the first to couple glycerol degradation with ethanol formation, to the best of our knowledge. The recombinant strain overexpressing GCY1 and DAK1, encoding glycerol dehydrogenase and dihydroxyacetone kinase, respectively, in glycerol degradation pathway, exhibited a moderate increase in ethanol yield (2.9 %) and decrease in glycerol yield (24.9 %) compared to the wild type with the initial glucose concentration of 15 % under anaerobic conditions. However, when the mhpF gene, encoding acetylating NAD⁺-dependent acetaldehyde dehydrogenase from Escherichia coli, was co-expressed in the aforementioned recombinant strain, a further increase in ethanol yield by 5.5 % and decrease in glycerol yield by 48 % were observed for the resultant recombinant strain GDMS1 when acetic acid was added into the medium prior to inoculation compared to the wild type. The process outlined in this study which enhances glycerol consumption and cofactor regulation in an industrial yeast is a promising metabolic engineering strategy to increase ethanol production by reducing the formation of glycerol.     

Interactions of Taxol-producing endophytic fungus with its host (Taxus spp.) during Taxol accumulation

Endophytic fungi (Fusarium mairei) culture broth (EFCB) was added to cell suspension cultures of Taxus cuspidata. After 5 days, cultures of T. cuspidata given 4 ml of EFCB produced a maximal yield of 6.11 mg/l paclitaxel, with a release ratio of 75%, 2- and 6.8-fold, respectively, greater than the controls. The active element in EFCB is an exopolysaccharide of ∼79 kD. Endophytic fungi produced 0.19 mg/l of paclitaxel in its producing medium. However, when the supernatant of Taxus cell suspension cultures from day 20 was added to the paclitaxel-producing medium, the biomass of fungi decreased by 24% and the yield of paclitaxel by 45%. In a co-culture system of plant and fungus, the yield of paclitaxel (12.8 mg/l) was >2-fold higher than that in the EFCB-treatment system.     

Constituents of human urine alter siderophore production ofEscherichia coli

Four uroisolates and four fæcal isolates ofEscherichia coli were randomly selected for studying the siderophore production in an iron-deficient, chemically defined, medium. Glucose, lactose, urea and creatinine were added individually, as well as in combination, to estimate their influence on siderophore production. No difference in siderophore production was observed between stool and urinary isolates ofE. coli. Alterations in phenolate production were observed to be constituent-dependent while a uniformly significant increase in hydroxamate production (p<0.05) was recorded after addition of the constituents, either each individually or in combination.     

Effects of selenium, iron and cobalt addition to growth and yessotoxin production of the toxic marine dinoflagellate Protoceratium reticulatum in culture

The marine dinoflagellate Protoceratium reticulatum has been recently identified as a source for the disulfated polyether toxin, yessotoxin (YTX), and may pose a risk to human health, aquaculture development and coastal environments. The requirements of P. reticulatum for selenium, iron and cobalt were assessed in culture. P. reticulatum was grown in nutrient enriched seawater (1/10 GP medium) without selenium or with 0.003 and 0.0003 μM selenium added; without iron or with 0.076 and 0.0076 μM iron added; and without cobalt or with 0.008 μM cobalt added. Test flasks were monitored for growth rate, cell yield and YTX production. P. reticulatum was found to exhibit a strong requirement for both selenium and iron. Growth rate and cell yield in treatments without added selenium were significantly (P<0.05) reduced to 60.2% (μ=0.15 day⁻¹) and 20.2% (4942 cell ml⁻¹), respectively, of those with selenium added (μ=0.23 day⁻¹ and 24, 387 cell ml⁻¹). YTX production was significantly increased by addition of selenium in two of three transfers tested. Cells of P. reticulatum subjected to medium without selenium added showed morphological changes observable at the light microscope level which included enlarged cell size. The diameter of cells in medium without selenium added were significantly (P<0.05) enlarged to 36.7±0.90 μm compared to cells in the medium with selenium added, 27.5±1.25 μm. Growth rate and cell yield in treatments without added iron were also significantly reduced to 70.1% (μ=0.16 day⁻¹) and 34.2% (8003 cells ml⁻¹), respectively, of those with iron added (μ=0.23 day⁻¹ and 23,416 cells ml⁻¹). No significant effect on YTX production was measured. In contrast to selenium and iron, no limitation of growth or cell yield or differences in YTX production were observed for flasks without cobalt as compared to those with cobalt added. The possibility that harmful algal events of P. reticulatum may be influenced by selenium or iron in neritic waters is discussed.     

Phage formation in Staphylococcus muscae cultures. VII. Partial purification of the protein factor necessary for virus synthesis

1. A substance is present in autolyzed pepsin solutions which stimulates the release of phage by some strains of S. muscae when added to Fildes' synthetic medium. 2. The substance is assayed by determining the quantity necessary to increase the phage yield to one-half the maximum value, using the one-step growth curve technique. 3. The substance has been concentrated and partially purified (500-fold) by heavy metal precipitation, butyl alcohol extraction, and absorption on norit. 4. No known amino acid or accessory growth substance tested could replace this substance.     

Two rhizobacterial strains,individually and in interactions with Rhizobium sp., enhance fusarial wilt control,growth, and yield in pigeon pea

A Pseudomonas aeruginosa strain, RRLJ 04, and a Bacillus cereus strain, BS 03, were tested both individually and in combination with a Rhizobium strain, RH 2, for their ability to enhance plant growth and nodulation in pigeon pea (Cajanus cajan L.) under gnotobiotic, greenhouse and field conditions. Both of the rhizobacterial strains exhibited a positive effect on growth in terms of shoot height, root length, fresh and dry weight, nodulation and yield over the non-treated control. Co-inoculation of seeds with these strains and Rhizobium RH 2 also reduced the number of wilted plants, when grown in soil infested with Fusarium udum. Gnotobiotic studies confirmed that the suppression of wilt disease was due to the presence of the respective PGPR strains. Seed bacterization with drug-marked mutants of RRLJ 04 and BS 03 confirmed their ability to colonize and multiply along the roots. The results suggest that co-inoculation of these strains with Rhizobium strain RH 2 can be further exploited for enhanced growth, nodulation and yield in addition to control of fusarial wilt in pigeon pea.     

Effects of Bicarbonate on Growth of Neisseria gonorrhoeae: Replacement of Gaseous CO2 Atmosphere

The effect of NaHCO₃ on the growth of Neisseria gonorrhoeae cultures was studied in a liquid and a semisolid growth medium. With a broth culture, NaHCO₃ (0.009 M) greatly reduced the lag phase and also increased the total growth. The same concentration of bicarbonate supported rapid growth when added to the semisolid medium if the plates were individually incubated in sealed plastic bags. In a container with a large air space, a higher concentration of NaHCO₃ was necessary to support growth. The assimilation of ¹⁴C-labeled NaHo₃ by growing cultures was also investigated.     

Development of diagnostics in the search for an explanation of aerotoxic syndrome

Aerotoxic syndrome is assumed to be caused by exposure to tricresyl phosphate, an additive in engine lubricants and hydraulic fluids that is activated to the toxic 2-(ortho-cresyl)-4H-1,3,2-benzodioxaphosphoran-2-one (CBDP). Currently, there is no laboratory evidence to support intoxication of airline crew members by CBDP. Our goal was to develop methods for testing in vivo exposure by identifying and characterizing biomarkers. Mass spectrometry was used to study the reaction of CBDP with human albumin, free tyrosine, and human butyrylcholinesterase. Human albumin made a covalent bond with CBDP, adding a mass of 170 amu to Tyr411 to yield the o-cresyl phosphotyrosine derivative. Human butyrylcholinesterase made a covalent bond with CBDP on Ser198 to yield five adducts with added masses of 80, 108, 156, 170, and 186 amu. The most abundant adduct had an added mass of 80 amu from phosphate (HPO₃), a surprising result given that no pesticide or nerve agent is known to yield phosphorylated serine with an added mass of 80 amu. The next most abundant adduct had an added mass of 170 amu to form o-cresyl phosphoserine. It is concluded that toxic gases or oil mists in cabin air may form adducts on plasma butyrylcholinesterase and albumin, detectable by mass spectrometry.     

Protoplast isolation for species in the Chamelaucium group and the effect of antioxidant enzymes (superoxide dismutase and catalase) on protoplast viability

An effective protocol for protoplast isolation from young leaves and somatic embryogenic cells of species in the Chamelaucium group and the use of superoxide dismutase (SOD) and catalase (CAT) to enhance protoplast viability are described. Mesophyll protoplasts were isolated from young leaves of a white Geraldton waxflower (Chamelaucium uncinatum) line 583, using a mixture of 1% (w/v) cellulase R10, 0.5% (w/v) macerozyme R10, and 0.1% (w/v) pectolyase. Viability of isolated mesophyll protoplasts increased dramatically when SOD and CAT were added. The highest increase of 7.61-fold in viability and 4.34-fold of viable protoplast yield were achieved when a combination of SOD at 500 units mL^?1 and CAT at 2,000 units mL^?1 was added to the enzyme mixture. Somatic embryogenic cell-derived protoplasts were isolated from embryogenic suspension cells of C. uncinatum line 583 when 1% (w/v) hemicellulase was added to a combination of 2% (w/v) cellulase R10, and 1% (w/v) macerozyme R10. Addition of SOD at 500 units mL^?1 and CAT at 2,000 units mL^?1 to the enzyme mixture improved viability only slightly, to above 90%, but improved yield significantly (6.6-fold). This combination of enzymes was also used to isolate protoplasts from embryogenic suspension cells of Chamelaucium repens and from young leaves of C. uncinatum, Actinodium calocephalum, Verticordia etheliana, Verticordia grandis, Verticordia hughanii, and Verticordia mitchelliana successfully with viability >80% and viable yield >7?×?10⁵ cells g^?1 fresh weight (or per milliliter packed cell volume in the case of suspension cells).     

Conversion of 5′-inosinic acid to inosine by Streptomyces aureus

The production of inosine by microbial conversion of 5′-inosinic acid (IMP) was investigated. Among the various strains of Streptomyces and Bacillus tested, Streptomyces aureus NCIB 9803 was selected for the microbial conversion process due to its high IMP-degrading activity. A maximum conversion yield of 0.43 (86% of the theoretical value) was obtained when IMP was added to the culture medium at 24 h. Kinetic studies with [8-¹⁴C] IMP showed that the difference from the theoretical values mainly attributable to the uptake of inosine by S. aureus.     

Influence of sugars on enantioselective reduction using Saccharomyces cerevisiae in organic solvent

Haploid Saccharomyces cerevisiae W303-1A cells grown on different carbon sources were employed as the biocatalyst for ethyl acetoacetate reduction in n-hexane. The effects of cell immobilization on montmorillonite, as well as the addition of trehalose or sucrose solutions, were also tested. Best conversions (∼50%) to the chiral alcohol ethyl (S)-(+)-3-hydroxybutanoate (ee > 99%) were obtained with cells grown under respiratory metabolism with glycerol–ethanol, and higher yields were observed when trehalose was added to the reaction media. Although cells with fermentative metabolism grown on glucose were able to reduce the substrate when sucrose was added, the disaccharide was consumed by the cells during the course of the reaction, and no enantioselective product was obtained. Immobilized cells also required the addition of trehalose in order to reduce the substrate with high yield. Thus, our results indicate that trehalose may be an efficient protector of immobilized or free yeast cells during enantioselective reductions in organic solvent.     

Growth Responses to Sodium by Bryophyllum tubiflorum under Conditions Inducing Crassulacean Acid Metabolism

The dry weight yield of plants of Bryophyllum tubiflorum Harvey, a species with Crassulacean acid metabolism characteristics, increased significantly (P < 1%) in response to added sodium (0.1 milliequivalents per liter NaCl was supplied to the culture solution initially containing less than 0.08 microequivalents per liter of Na) when grown under short day (8 hours) conditions but not when grown under long day conditions (16 hours).