Forgot your password?
New user?
New Window Opens on the Secret Life of Microbes: Scientists Develop First Microbial Profiles of Ecosystems
CAS announces the release of SciFinder Scholar for Mac OS X
Press Releases
Press Releases
| Check out our New Publishers' Select for Free Articles |
Applied Microbiology and Biotechnology (v.66, #5)
Archaea in protozoa and metazoa by Marianne Lange; Peter Westermann; Birgitte Kiær Ahring (pp. 465-474).
The presence of Archaea is currently being explored in various environments, including extreme geographic positions and eukaryotic habitats. Methanogens are the dominating archaeal organisms found in most animals, from unicellular protozoa to humans. Many methanogens can contribute to the removal of hydrogen, thereby improving the efficiency of fermentation or the reductive capacity of energy-yielding reactions. They may also be involved in tissue damage in periodontal patients. Recent molecular studies demonstrated the presence of Archaea other than methanogens in some animals—but so far, not in humans. The roles of these microorganisms have not yet been established. In the present review, we present the state of the art regarding the archaeal microflora in animals.
Cyclodextrin glucanotransferase: from gene to applications by Qingsheng Qi; Wolfgang Zimmermann (pp. 475-485).
Cyclodextrin glucanotransferase (CGTase) is an important industrial enzyme which is used to produce cyclodextrins. CGTase genes from more than 30 bacteria have been isolated and several of the enzymes have been identified and biochemically characterized. For a better understanding of the reaction mechanism and function of CGTase, the enzyme has been analyzed at gene level and protein level with regard to its structure and the similarity of different CGTase subgroups. The biological role of the enzyme is proposed based on the genetic and enzymatic analyses. Methods to enhance the production of active CGTase by bacteria are compared. The enzyme can be applied in biotechnology for the production of cyclodextrins and oligosaccharides with novel properties.
Botryococcus braunii: a rich source for hydrocarbons and related ether lipids by P. Metzger; C. Largeau (pp. 486-496).
This paper presents a review on Botryococcus braunii, a cosmopolitan green colonial microalga characterised by a considerable production of lipids, notably hydrocarbons. Strains like wild populations of this alga differ in the type of hydrocarbons they synthesise and accumulate: (1) n-alkadienes and trienes, (2) triterpenoid botryococcenes and methylated squalenes, or (3) a tetraterpenoid, lycopadiene. In addition to hydrocarbons and some classic lipids, these algae produce numerous series of characteristic ether lipids closely related to hydrocarbons. This review covers the algal biodiversity, the chemical structures and biosynthesis of hydrocarbons and ether lipids and the biotechnological studies related to hydrocarbon production.
Studies on the production and purification of an antimicrobial compound and taxonomy of the producer isolated from the marine environment of the Sundarbans by M. Saha; D. Ghosh Jr.; D. Ghosh; D. Garai; P. Jaisankar; K. K. Sarkar; P. K. Dutta; S. Das; T. Jha; J. Mukherjee (pp. 497-505).
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.
UV-A mediated induction of carotenoid accumulation in Dunaliella bardawil with retention of cell viability by Alonso Salguero; Rosa León; Annalisa Mariotti; Benito de la Morena; José M. Vega; Carlos Vílchez (pp. 506-511).
The effect of adding UV-A radiation (320–400 nm) to photosynthetically active radiation (PAR, 400–700 nm) during growth of the photosynthetic marine microalga Dunaliella bardawil was investigated in this work in terms of cell growth and carotenoid production. Although signs of slow cell growth (slight reduction of chlorophyll and protein content) were observed after 24 h of cell exposure to UV-A (40 μmol photons m−2 s−1 and 70 μmol photons m−2 s−1) plus 140 μmol photons m−2 s−1 PAR , 84 h exposure to these UV-A conditions slightly stimulated cell growth and increased the photosynthetic efficiency of the exposed cultures. The enhanced cell growth was coupled with an increase in total carotenoid content. Besides β-carotene as the major pigment, increases in the well-known antioxidants lutein and zeaxanthin of about 3-fold and 5-fold, respectively, were determined in cultures exposed to UV-A radiation of 70 μmol photons m−2 s−1for 84 h. As a consequence, far from being negative to cell growth, low and medium UV-A radiation are stress factors that could be successfully applied to long-term processes for large scale carotenoid production using D. bardawil cultures with retention of cell viability. UV-A exposure has the advantage of being a factor either easily applied or removed as required, in contrast to other nutrient stresses, which require medium replacement for their application.
Chemo-enzymatic synthesis of 2′-O-methoxyethyl ribonucleosides using a phosphodiesterase from Serratia marcescens by Guy Marais; Oreste Ghisalba (pp. 512-519).
An enzyme able to cleave the 3′,5′-phosphate ring of 2′-methoxyethyl cyclic nucleotides (3′,5′-cyclic nucleotide phosphodiesterase, EC 3.1.4.17) from Serratia marcescens DSM 30121 was used to deprotect the cyclic phosphate nucleotides after chemical alkylation. The process yielded 2′-O-alkylated nucleosides used as building blocks of antisense oligonucleotides for subsequent potential applications in therapeutics (antisense oligonucleotide synthesis) and diagnostics. The phosphodiesterase from the Gram-negative enteric bacterium S. marcescens was selected on account of the broad substrate range and high activity of the enzyme. The protein was purified by heat-treatment of the crude cell-free extract, followed by column chromatography (gel filtration). It was characterised and showed optimal activity at a broad pH range (pH 6.8–9.4, with a peak at ca. pH 8.5) and at a temperature of 60–65°C. No metal ions were required for activity, although Ba2+ was an activator. Conversion of 2′-O-methoxyethyl cAMP into the corresponding nucleoside derivative on a multi-gram scale was successfully performed in two steps, using the S. marcescens enzyme in conjunction with a commercially available alkaline phosphatase from Escherichia coli.
Expression of the Fusarium oxysporum lactonase gene in Aspergillus oryzae: molecular properties of the recombinant enzyme and its application by Kohsuke Honda; Hirokazu Tsuboi; Toshitaka Minetoki; Hanae Nose; Keiji Sakamoto; Michihiko Kataoka; Sakayu Shimizu (pp. 520-526).
The lactonase gene of Fusarium oxysporum was expressed in Aspergillus oryzae for optical resolution of dl-pantoyl lactone. When the chromosomal gene encoding the full-length form of the lactonase, which has its own NH2-terminal signal peptide, was introduced in the host cells, the resulting transformant produced an enzyme of 46,600 Da, which corresponded to the wild-type enzyme. In contrast, A. oryzae transformed with the cDNA coding the mature enzyme produced a protein of 41,300 Da. Deglycosylation analysis with an endoglycosidase revealed that the difference in molecular mass arose from the different sugar contents of the recombinant enzymes. The mycelia of the transformant were used as a catalyst for asymmetric hydrolysis of dl-pantoyl lactone. The initial velocity of the asymmetric hydrolysis reaction catalyzed by the transformant was estimated to be 30 times higher than that by F. oxysporum. When the mycelia of the transformant were incubated with a 20% dl-pantoyl lactone solution for 4 h, 49.9% of the racemic mixture was converted to d-pantoic acid (>95% ee).
Purification and characterization of recombinant Escherichia coli-expressed Pichia etchellsii β-glucosidase II with high hydrolytic activity on sophorose by Yukti Bhatia; Saroj Mishra; Virendra S. Bisaria (pp. 527-535).
β-Glucosidase II (Bgl II), encoded by the βglu2 gene of the thermo-tolerant yeast Pichia etchellsii, was purified from recombinant Escherichia coli pBG22:JM109. The enzyme had a molecular mass of 176 kDa and was a dimer with an apparent subunit mass of 83 kDa. It exhibited broad substrate specificity and hydrolyzed β-linked gluco-disaccharides and oligosaccharides, salicin, and cyanogenic glucoside amygladin. The unusually high hydrolytic activity of 7,680 units min−1 g−1 protein was obtained on sophorose. Competition experiments performed using differently linked β-disaccharides indicated these to be hydrolyzed at the same active site. Transglycosylation activity leading to the biosynthesis of several disaccharides and oligosaccharides was observed. The enzyme was placed in glycosyl hydrolase family 3, based on a statistical approach using amino acid composition data. The involvement of His as a catalytically important residue was confirmed by diethylpyrocarbonate modification. Pre-incubation of the purified enzyme with 5 mM p-nitrophenyl-β-d-glucoside offered 2.5-fold higher residual activity compared with unbound enzyme, indicating protection at the active site. The feasibility of this enzyme as a biocatalyst of choice for the synthesis of glyco-conjugates is discussed.
The AlnB protein of the bioemulsan alasan is a peroxiredoxin by R. Bekerman; G. Segal; E. Z. Ron; E. Rosenberg (pp. 536-541).
The bioemulsifier of Acinetobacter radioresistens KA53, referred to as alasan, is a high molecular weight complex of a polysaccharide and three proteins (AlnA, AlnB and AlnC). AlnA has previously been shown to be an OmpA-like protein that is largely responsible for the emulsifying activity of alasan. To further elucidate the nature of alasan, the gene coding for AlnB was cloned, sequenced and overexpressed in Escherichia coli. The overall 561 bp sequence of the hypothetical AlnB showed strong homology, including all conserved regions and residues known to be essential for enzymatic activity, to the ubiquitous family of thiol-specific antioxidant enzymes known as peroxiredoxins. Transgenic E. coli overexpressing AlnB exhibited increased resistance to cumene hydroperoxide both in liquid culture and on agar medium. Recombinant AlnB had no emulsifying activity but stabilized oil-in-water emulsion generated by AlnA.
Identification and characterization of a novel d-amidase gene from Variovorax paradoxus and its expression in Escherichia coli by L. Krieg; H. Slusarczyk; S. Verseck; M.-R. Kula (pp. 542-550).
The gene for the newly described d-amidase from Variovorax paradoxus (Krieg et al. 2002) was cloned and functionally expressed in Escherichia coli. Since native enzyme was available in minute amounts only, we determined the N-terminal sequence of the enzyme and utilized the Universal GenomeWalker Approach to make use of the common internal sequence of the amidase signature family. The high GC content of the gene made it necessary to employ an appropriate DNA polymerase in the amplification reactions. Thus, the sequence of the complete gene and the flanking regions was established. In independent experiments, the gene was then amplified from genomic DNA of V. paradoxus, expressed in E. coli, and characterized. The recombinant enzyme has a specific activity of 1.7 units/mg with racemic tert-leucine amide as substrate and is a homodimer of 49.6-kDa monomers.
Formation and analysis of mannosylerythritol lipids secreted by Pseudozyma aphidis by U. Rau; L. A. Nguyen; S. Schulz; V. Wray; M. Nimtz; H. Roeper; H. Koch; S. Lang (pp. 551-559).
Pseudozyma aphidis DSM 70725 was found to be a novel producer of mannosylerythritol lipids (MELs). The MELs were quantified by HPLC. Glucose as carbon source for precultivation supported growth well. By contrast, at concentrations >30 g l−1 in preculture, subsequent MEL formation in the main culture with soybean oil as sole carbon source was reduced. The type of substrate supply considerably influenced MEL formation. High concentrations of soybean oil (80 ml l−1) at init favored the production process when compared to a stepwise (20 ml l−1) addition. Mannose or erythritol were suitable second carbon sources that enhanced the MEL yield with soybean oil as preferred primary substrate. After 10 days, a maximum yield of 75 g l−1 was attained during shake-flask cultivation. Biofuel (rapeseed oil methyl ester) also resulted in high yields of MEL, but glucose reduced the MEL yield. Analysis by GC-MS showed that all fatty acids contained in MEL and derived from soybean oil or related methyl ester were degraded by C2-units to differing extents. The surface (water/air) and interfacial (water/hexadecane) tension of the MELs produced from different carbon sources were reduced to a minimum of 26.2 mN m−1 and 1 mN m−1, respectively.
Effect of nitrogen limitation and surplus upon trehalose metabolism in wine yeast by Maria Teresa Novo; Gemma Beltran; Nicolas Rozès; José Manuel Guillamón; Alberto Mas (pp. 560-566).
Trehalose metabolism in yeast has been related to stress and could be used as a stress indicator. Winemaking conditions are stressful for yeast and understanding trehalose metabolism under these conditions could be useful for controlling alcoholic fermentation. In this study, we analysed trehalose metabolism of a commercial wine yeast strain during alcoholic fermentation by varying the nitrogen levels from low (below adequate) to high (excess). We determined trehalose, nitrogen, sugar consumption and expression of NTH1, NTH2 and TPS1. Our results show that trehalose metabolism is slightly affected by nitrogen availability and that the main consumption of nitrogen occurs in the first 24 h. After this period, nitrogen is hardly taken up by the yeast cells. Although nitrogen and sugar are still available, no further growth is observed in high concentrations of nitrogen. Increased expression of genes involved in trehalose metabolism occurs mainly at the end of the growth period. This could be related to an adaptive mechanism for fine tuning of glycolysis during alcoholic tumultuous fermentation, as both anabolic and catabolic pathways are affected by such expression.
Continuous acetonitrile degradation in a packed-bed bioreactor by Taras Manolov; Håkansson Kristina; Guieysse Benoit (pp. 567-574).
A 20-l packed-bed reactor filled with foamed glass beads was tested for the treatment of acetonitrile HPLC wastes. Aeration was provided by recirculating a portion of the reactor liquid phase through an aeration tank, where the dissolved oxygen concentration was kept at 6 mg/l. At a feeding rate of 0.77 g acetonitrile l−1 reactor day−1, 99% of the acetonitrile was removed; and 86% of the nitrogen present in acetonitrile was released as NH3, confirming that acetonitrile volatilization was not significant. Increasing the acetonitrile loading resulted in lower removal efficiencies, but a maximum removal capacity of 1.0 g acetonitrile l−1 reactor day−1 was achieved at a feeding rate of 1.6 g acetonitrile l−1 reactor day−1. The removal capacity of the system was well correlated with the oxygenation capacity, showing that acetonitrile removal was likely to be limited by oxygen supply. Microbial characterization of the biofilm resulted in the isolation of a Comamonas sp. able to mineralize acetonitrile as sole carbon, nitrogen and energy source. This organism was closely related to C. testosteroni (91.2%) and might represent a new species in the Comamonas genus. This study confirms the potential of packed-bed reactors for the treatment of a concentrated mixture of volatile pollutants.
Monitoring the impact of bioaugmentation on the start up of biological phosphorus removal in a laboratory scale activated sludge ecosystem by Patrick Dabert; Jean-Philippe Delgenès; Jean-Jacques Godon (pp. 575-588).
The acclimatisation of activated sludge to enhanced biological phosphorus removal (EBPR) conditions requires a period of about 40–100 days but its output remains hazardous. The impact of bioaugmentation on the start-up of a laboratory scale EBPR sequencing batch reactor was evaluated by process parameters measurement and microbial community dynamics monitoring using 16S rDNA targeted polymerase chain reaction-single strand conformation polymorphism electrophoresis (PCR-SSCP). Bioaugmentation: (1) speeded up the installation of good and stable EBPR in the bioaugmented reactor by about 15 days; (2) correlated with the transient enrichment of the sludge in the added microbial populations; and (3) favoured the long-term enrichment of the sludge in the phosphorus-accumulating organism (PAO) Candidatus Accumulibacter phosphatis. However, despite a lag time period, the control non-bioaugmented reactor ended up with comparable reactor parameters and microbial community evolution, suggesting that the same PAO populations were already present from the beginning in the original non-P-accumulating seed sludge. The potential of a true installation of the added microbial populations within the bioaugmented reactor compared to their substitution by indigenous similar populations is discussed. Competition between PAOs and the antagonistic glycogen accumulating organism Candidatus Competibacter phosphatis is also highlighted during EBPR start-up.
Klebsiella planticola strain DSZ mineralizes simazine: physiological adaptations involved in the process by Mariela Sánchez; Carlos Garbi; Roberto Martínez-Álvarez; Luis T. Ortiz; José Luis Allende; Margarita Martín (pp. 589-596).
We examined the ability of a soil bacterium, Klebsiella planticola strain DSZ, to degrade the herbicide simazine (SZ). Strain DSZ is metabolically diverse and grows on a wide range of s-triazine and aromatic compounds. DSZ cells grown in liquid medium with SZ (in 10 mM ethanol) as carbon source mineralized 71.6±1.3% of 0.025 mM SZ with a yield of 4.6±0.3 μg cell dry weight mmol−1 carbon. The metabolites produced by DSZ during SZ degradation included ammeline, cyanuric acid, N-formylurea and urea. We studied the physiological adaptations which allow strain DSZ to metabolize SZ. Using scanning electron microscopy, we detected DSZ cells covering the surfaces of SZ crystals when the herbicide was used at high concentrations (0.1 mM). The membrane order observed by FTIR spectroscopy showed membrane activity at low temperature (4°C) to assimilate the herbicide. Membrane fatty acid analysis demonstrated that strain DSZ adapted to grow on SZ by increasing the degree of saturation of membrane lipid fatty acid; and the opposite effect was detected when both SZ and ethanol were used as carbon sources. This confirms the modulator effect of ethanol on membrane fluidity.