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Archives of Microbiology (v.174, #6)
No Title by Kathryn H. Engler; Raymond D. Coker; Ivor H. Evans (pp. 381-385).
Uptake of aflatoxin B1 (AFB1) and trichothecene T-2 toxin from growth medium by mycotoxin bioassay strains of Kluyveromyces marxianus and Bacillus megaterium was assessed by incubating, washing, and sonicating the cells, extracting samples with chloroform, and analysing the extracts by a combination of high-performance thin-layer chromatography (HPTLC) and fluorescence densitometry. Using cultures of K. marxianus, the entire AFB1 dose was recovered and no AFB1 metabolites were detected. Less than 1% of the AFB1 was recovered from the cells, and AFB1 did not inhibit growth. Methanol in the incubation medium had no significant effect on the levels of AFB1 associated with K. marxianus cells. The entire dose of T-2 toxin was also recovered from K. marxianus cultures, and no metabolites were detected; again, less than 1% of T-2 toxin was cell-associated, but growth was completely inhibited. AFB1 partially inhibited the growth of B. megaterium; approximately 12% of the dose could not be recovered, and no AFB1-related metabolites were detected. Methanol increased the levels of recoverable AFB1 associated with B. megaterium cells. In the case of T-2 toxin, around 8% of the dose was not recovered, and no metabolites were detected; growth of B. megaterium was stimulated. These results suggest irreversible binding of both toxins, or derivatives of them, to the cells of B. megaterium.
Keywords: Mycotoxin bioassay Uptake Aflatoxin B T-2 toxin Kluyveromyces marxianus Bacillus megaterium
No Title by Matthias Sipiczki; Aniko Bozsik (pp. 386-392).
Cytokinesis in the fission yeast Schizosaccharomyces begins by formation of a medially placed actomyosin ring, continues by progressive development of a septum, and is completed by cleavage of the mature septum to bring about cell separation. The cytological analysis of the Schizosaccharomyces pombe morphomutants sph2-3 and sep1-1 presented here demonstrates that the medial actomyosin ring colocalizes to the leading edge of the centripetally growing septum, and cleavage of the septum is triggered by rupture of the mother cell wall at the septal basis.
Keywords: Actin Cell wall Cytokinesis Fission yeast Morphogenesis Schizosaccharomyces septum
No Title by Ulrike Jonas; Elke Hammer; Erhard T.K. Haupt; Frieder Schauer (pp. 393-398).
Cells of the white rot fungus Pycnoporus cinnabarinus grown in glucose were able to hydroxylate biphenyl and diphenyl ether, although growth was inhibited by these substrates at concentrations above 250 µM. 2- and 4-Hydroxybiphenyl were detected as products of biphenyl metabolism and 2- and 4-hydroxydiphenyl ether as products of diphenyl ether metabolism in the culture media. After addition of 2-hydroxydiphenyl ether and 2-hydroxybiphenyl to cell-free supernatants containing laccase as the only ligninolytic enzyme, different coloured precipitates were formed. HPLC analysis revealed the formation of additional hydrophobic metabolites with one major product per transformation. Mass spectrometric analysis of the methyl derivatives of the polymer mixture indicated dimers and trimers with different binding types. The main products were identified as dimers with carbon-carbon bonds in para-position to the hydroxyl group of the monomers by mass spectroscopy and nuclear magnetic resonance spectroscopy.
Keywords: Diphenyl ether Biphenyl Laccase Pycnoporus cinnabarinus White rot fungi Coupling
No Title by Zhaomin Yang; Dongchuan Guo; M. Gabriela Bowden; Hong Sun; Leming Tong; Zhuo Li; Alfred E. Brown; Heidi B. Kaplan; Wenyuan Shi (pp. 399-405).
Myxococcus xanthus is a gram-negative soil bacterium that initiates a complex developmental program in response to starvation. A transposon insertion (Tn5-lac Ω109) mutant with developmental deficiencies was isolated and characterized in this study. A strain containing this insertion mutation in an otherwise wild-type background showed delayed developmental aggregation for about 12 h and sporulated at 1–2% of the wild-type level. Tn5-lac Ω109 was found to have disrupted the M. xanthus wbgB gene, which is located 2.1 kb downstream of the M. xanthus lipopolysacharide (LPS) O-antigen biosynthesis genes wzm wzt wbgA. The deduced polypeptide sequence of WbgB shares significant similarity with bacterial glycosyltransferases including M. xanthus WbgA. The wbgB::Tn5-lac Ω109 mutant was found to be defective in LPS O-antigen synthesis by immunochemical analysis. Further mutational analysis indicated that the defects of the wbgB::Tn5-lac Ω109 mutant were not the result of polar effects on downstream genes. Various motility assays demonstrated that the Tn5-lac Ω109 mutation affected both social (S) and adventurous (A) gliding motility of M. xanthus cells. The pleiotrophic effects of wbgB mutations indicate the importance of LPS O-antigen biosynthesis for various cellular functions in M. xanthus.
Keywords: Myxobacteria Myxococcus Lipopolysaccharide O-antigen Gliding motility Development Fruiting body
No Title by Biswarup Mukhopadhyay; Vipool J. Patel; Ralph S. Wolfe (pp. 406-414).
The pyruvate carboxylase (PYC) of the hyperthermophilic, strictly hydrogenotrophic, autotrophic and marine methanarchaeon Methanococcus jannaschii was purified to homogeneity. Optimal activity was at pH 8.5, ≥80 °C, and a KCl concentration of 0.175 M. This enzyme is the most thermophilic PYC so far studied. Unlike the Methanobacterium thermoautotrophicum enzyme, Mc. jannaschii PYC was expressed in cells grown without an external source of biotin and in the purified form was stable during storage at 4, –20 and –80 °C. However, it was rapidly inactivated at 80 °C. The enzyme was insensitive to aspartate and glutamate, mildly inhibited by α-ketoglutarate, and was strongly inhibited by ATP and ADP (apparent K m for ATP, 0.374±0.039 mM; apparent K i for ATP, 5.34±2.14 mM; K i for ADP, 0.89±0.18 mM). It was also strongly inhibited when the Mg2+ concentration in the assay exceeded that of ATP. Thus, this stable PYC could serve as a model for mechanistic studies on archaeal PYCs. It was apparently an α4β4-type PYC composed of a non-biotinylated 55.5-kDa subunit (PYCA) and a 64.2-kDa biotinylated subunit (PYCB). The determined NH2-terminal sequences for these subunits provided additional support for our earlier proposal to rename the ORFs MJ1229 and MJ1231 in the NCBI Mc. jannaschii genome sequence database as PYCA and PYCB, respectively; even very recently, these have been misidentified as a subunit of acetyl-CoA carbxoylase (AccC) and the α-subunit of ion-pumping oxaloacetate decarboxylase (OADα), respectively.
Keywords: Methanarchaea Methanococcus jannaschii Pyruvate carboxylase Extreme thermophile Storage stable Thermolability pyc genes
No Title by David L. Valentine; Douglas C. Blanton; William S. Reeburgh (pp. 415-421).
Hydrogen production was studied in four species of methanogens (Methanothermobacter marburgensis, Methanosaeta thermophila, Methanosarcina barkeri, and Methanosaeta concilii) under conditions of low (sub-nanomolar) ambient hydrogen concentration using a specially designed culture apparatus. Transient hydrogen production was observed and quantified for each species studied. Methane was excluded as the electron source, as was all organic material added during growth of the cultures (acetate, yeast extract, peptone). Hydrogen production showed a strong temperature dependence, and production ceased at temperatures below the growth range of the organisms. Addition of polysulfides to the cultures greatly decreased hydrogen production. The addition of bromoethanesulfonic acid had little influence on hydrogen production. These experiments demonstrate that some methanogens produce excess reducing equivalents during growth and convert them to hydrogen when the ambient hydrogen concentration becomes low. The lack of sustained hydrogen production by the cultures in the presence of methane provides evidence against "reverse methanogenesis" as the mechanism for anaerobic methane oxidation.
Keywords: Methanogens Hydrogen production Storage compounds Anaerobic methane oxidation
No Title by Hor-Gil Hur; Jackson O. Lay Jr.; Richard D. Beger; James P. Freeman; Fatemeh Rafii (pp. 422-428).
Fecal bacteria from a healthy individual were screened for the specific bacteria involved in the metabolism of dietary isoflavonoids. Two strains of bacteria capable of producing primary and secondary metabolites from the natural isoflavone glycosides daidzin and genistin were detected. The metabolites were identified by comparison of their HPLC/mass, 1H NMR and UV spectra with those of standard and synthetic compounds. Both Escherichia coli HGH21 and the gram-positive strain HGH6 converted daidzin and genistin to the their respective aglycones daidzein and genistein. Under anoxic conditions, strain HGH6 further metabolized the isoflavones daidzein and genistein to dihydrodaidzein and dihydrogenistein, respectively. The reduction of a double bond between C-2 and C-3 to a single bond was isoflavonoid-specific by strain HGH6, which did not reduce a similar bond in the flavonoids apigenin and chrysin. Strain HGH6 did not further metabolize dihydrodaidzein and dihydrogenistein. This is the first study in which specific colonic bacteria that are involved in the metabolism of daidzin and genistin have been detected.
Keywords: Isoflavonoid Phytoestrogen Intestinal bacteria Biotransformation Reduction
No Title by Arthur L. Koch (pp. 429-439).
The structure and conformation of the sacculus of bacteria at a scale much larger than just the component disaccharide penta-muropeptide is not well known and is crucially important for the understanding of bacterial growth and cell wall function. By computer simulations, the minimal energy conformations and the energy needed for stretching the component parts were found. The oligosaccharide chain, modeled as (GlcNAc-MurNAc)8 when under no tension, can assumed a variety of nearly iso-energetic conformations. These included a variety of bends and kinks, with the chain forming an irregular random coil. In the most relaxed and minimal energy state, the D-lactyl groups of the MurNAc (N-acetyl muramic acid) residues protruded at about an angle of 90° relative to the D-lactyl groups of their immediate MurNAc neighbors in the same chain. The cell wall penta-muropeptide precursor is identical for Escherichia coli and Bacillus subtilis; it also adopted many conformations, each of an energy almost equal to the global minimum. The cross-bridged structure of the tail-to-tail linkage of disaccharide nona-muropeptide has a second type of association, in addition to the covalent cross-bridge, which has not been considered before. This is the ionic interaction between the free D-Ala and the free amino group of the m-A2 pm. In vivo, when the cross-bridge is stretched (in the computer to simulate growth), this pairing dissociates. The possible biological significance of this is that it exposes the underlying 'tail-to-tail' peptide bond to autolysis and will expose both the ends of the m-A2 pm and the D-Ala-D-Ala groups that may then be able to react with nascent penta-muropeptides to form trimers. This suggests a new model for growth of the bacterial cell wall that depends on changes in the chemical conformation of the cross-bridge structure as it comes to bear stress.
Keywords: Oligoglycan Penta-muropeptide Nona-muropeptide Minimal energy conformation Tail-to-tail peptide bond
No Title by Maria V. Simankova; Oleg R. Kotsyurbenko; Erko Stackebrandt; Nadezhda A. Kostrikina; Anatoliy M. Lysenko; Georgiy A. Osipov; Alla N. Nozhevnikova (pp. 440-447).
A new psychrophilic, anaerobic, acetogenic bacterium from the tundra wetland soil of Polar Ural is described. The organism fermented H2/CO2, formate, methanol, and several sugars to acetate as the sole end-product. The temperature range for growth was 1–30 °C with an optimum at 20 °C. The bacterium showed no growth at 32 °C. Cells were gram-positive, oval-shaped, flagellated rods 0.7–1.1×1.1–4.0 µm in size when grown at 1–20 °C. At 25–30 °C, the cell size increased up to 2–3×10–15 µm due to a defect in cell division. The DNA G+C content of the organism was 39.2 mol%. Based upon 16S rDNA analysis and DNA-DNA reassociation studies, the organism was classified in the genus Acetobacterium as a new species, for which the name Acetobacterium tundrae sp. nov. is proposed. The type strain is Z-4493 (=DSM 9173T).
Keywords: Acetogenesis Anaerobe Psychrophile Psychrotroph Acetobacterium tundrae Tundra wetlands
No Title by Alexander V. Surkov; Michael E. Böttcher; Jan Kuever (pp. 448-451).
Stable sulfur isotope fractionation was investigated during reduction of thiosulfate by growing batch cultures of Dethiosulfovibrio russensis at a cell-specific reduction rate of 2.4±0.72 fmol cell–1 d–1 (28 °C). Citrate was used as carbon and energy source. The hydrogen sulfide produced by this sulfur- and thiosulfate-reducing bacterium was depleted in 34S by 11‰ compared to total thiosulfate sulfur, in agreement with previous results observed for sulfate-reducing bacteria. This indicates the operation of a similar pathway for thiosulfate reduction in these phylogenetically different bacteria.
Keywords: Thiosulfate reduction Stable sulfur isotopes Isotope fractionation
No Title by Henri L. Saenz; Vera Augsburger; Cuong Vuong; Ralph W. Jack; Friedrich Götz; Michael Otto (pp. 452-455).
AgrB has been suggested to be responsible for the posttranslational modification in staphylococci that leads to the production of the thiolactone-containing agr peptide pheromone. We demonstrate that AgrB is located in the cytoplasmic membrane. Vectors were constructed for the xylose-inducible overexpression of agrB, and of agrB and agrD together. A Staphylococcus epidermidis strain deleted for agr and containing these vectors was assayed for AgrB protein and pheromone production. The lack of adequate pheromone production suggests the involvement of additional factors in the production of the agr pheromone.
Keywords: agr Quorum-sensing Staphylococcus epidermidis Pheromone Posttranslational modification