Showing papers in "Journal of Industrial Microbiology & Biotechnology in 2002"

Diversity of halophilic microorganisms: environments, phylogeny, physiology, and applications.

Abstract: The phylogenetic diversity of microorganisms living at high salt concentrations is surprising. Halophiles are found in each of the three domains: Archaea, Bacteria, and Eucarya. The metabolic diversity of halophiles is great as well: they include oxygenic and anoxygenic phototrophs, aerobic heterotrophs, fermenters, denitrifiers, sulfate reducers, and methanogens. The diversity of metabolic types encountered decreases with salinity. The upper salinity limit at which each dissimilatory process takes place is correlated with the amount of energy generated and the energetic cost of osmotic adaptation. Our understanding of the biodiversity in salt-saturated environments has increased greatly in recent years. Using a combination of culture techniques, molecular biological methods, and chemotaxonomic studies, we have obtained information on the nature of the halophilic Archaea as well as the halophilic Bacteria that inhabit saltern crystallizer ponds. Several halophilic microorganisms are being exploited in biotechnology. In some cases, such as the production of ectoine, the product is directly related to the halophilic behavior of the producing microorganism. In other cases, such as the extraction of β-carotene from Dunaliella or the potential use of Haloferax species for the production of poly-β-hydroxyalkanoate or extracellular polysaccharides, similar products can be obtained from non-halophiles, but halophilic microorganisms may present advantages over the use of non-halophilic counterparts. Journal of Industrial Microbiology & Biotechnology (2002) 28, 56–63 DOI: 10.1038/sj/jim/7000176

Biofilm material properties as related to shear-induced deformation and detachment phenomena.

Abstract: Biofilms of various Pseudomonas aeruginosa strains were grown in glass flow cells under laminar and turbulent flows. By relating the physical deformation of biofilms to variations in fluid shear, we found that the biofilms were viscoelastic fluids which behaved like elastic solids over periods of a few seconds but like linear viscous fluids over longer times. These data can be explained using concepts of associated polymeric systems, suggesting that the extracellular polymeric slime matrix determines the cohesive strength. Biofilms grown under high shear tended to form filamentous streamers while those grown under low shear formed an isotropic pattern of mound-shaped microcolonies. In some cases, sustained creep and necking in response to elevated shear resulted in a time-dependent fracture failure of the "tail" of the streamer from the attached upstream "head." In addition to structural differences, our data suggest that biofilms grown under higher shear were more strongly attached and were cohesively stronger than those grown under lower shears.

Factors affecting the yield and properties of bacterial cellulose.

Abstract: Acetobacter xylinum E(25) has been applied in our studies in order to find optimal culture conditions for effective bacterial cellulose (BC) production The strain displays significantly higher stability in BC production under stationary culture conditions In contrast, intensive agitation and aeration appear to drastically reduce cellulose synthesis since such conditions induced formation of spontaneous cellulose nonproducing mutants (Cel-), which dominated in the culture Mutation frequency strictly depends on the medium composition in agitated cultures Enrichment of the standard SH and Yamanaka media with 1% ethanol significantly enhanced BC production in stationary cultures Horizontal fermentors equipped with rotating discs or rollers were successfully applied in order to improve culture conditions Relatively slow rotation velocity (4 rpm) and large surface area enabling effective cell attachment are optimal parameters for cellulose production Physical properties of BC samples synthesized either in stationary cultures or in a horizontal fermentor revealed that cellulose from stationary cultures demonstrated a much higher value of Young's modulus, but a much lower value of water-holding capacity

Succinate production in dual-phase Escherichia coli fermentations depends on the time of transition from aerobic to anaerobic conditions

Abstract: We examined succinic acid production in Escherichia coli AFP111 using dual-phase fermentations, which comprise an initial aerobic growth phase followed by an anaerobic production phase. AFP111 has mutations in the pfl, ldhA, and ptsG genes, and we additionally transformed this strain with the pyc gene (AFP111/pTrc99A-pyc) to provide metabolic flexibility at the pyruvate node. Aerobic fermentations with these two strains were completed to catalog physiological states during aerobic growth that might influence succinate generation in the anaerobic phase. Activities of six key enzymes were also determined for these aerobic fermentations. From these results, six transition times based on physiological states were selected for studying dual-phase fermentations. The final succinate yield and productivity depend greatly on the physiological state of the cells at the time of transition. Using the best transition time, fermentations achieved a final succinic acid concentration of 99.2 g/l with an overall yield of 110% and productivity of 1.3 g/l h. Journal of Industrial Microbiology & Biotechnology (2002) 28, 325–332 DOI: 10.1038/sj/jim/7000250

Lactic acid bacteria and yeasts in kefir grains and kefir made from them.

Abstract: In an investigation of the changes in the microflora along the pathway: kefir grains (A)→kefir made from kefir grains (B)→kefir made from kefir as inoculum (C), the following species of lactic acid bacteria (83–90%) of the microbial count in the grains) were identified: Lactococcus lactis subsp lactis, Streptococcus thermophilus, Lactobacillus delbrueckii subsp bulgaricus, Lactobacillus helveticus, Lactobacillus casei subsp pseudoplantarum and Lactobacillus brevis Yeasts (10–17%) identified were Kluyveromyces marxianus var lactis, Saccharomyces cerevisiae, Candida inconspicua and Candida maris In the microbial population of kefir grains and kefir made from them the homofermentative lactic streptococci (52–65% and 79–86%, respectively) predominated Within the group of lactobacilli, the homofermentative thermophilic species L delbrueckii subsp bulgaricus and L helveticus (70–87% of the isolated bacilli) predominated Along the pathway A→B→C, the streptococcal proportion in the total kefir microflora increased by 26–30% whereas the lactobacilli decreased by 13–23% K marxianus var lactis was permanently present in kefir grains and kefirs, whereas the dominant lactose-negative yeast in the total yeast flora of the kefir grains dramatically decreased in kefir C Journal of Industrial Microbiology & Biotechnology (2002) 28, 1–6 DOI: 101038/sj/jim/7000186

Antagonistic effects of Streptomyces violaceusniger strain G10 on Fusarium oxysporum f.sp. cubense race 4: indirect evidence for the role of antibiosis in the antagonistic process.

Abstract: f.sp. cubense is the causal pathogen of wilt disease of banana. A cost-effective measure of control for this disease is still not available. Streptomyces violaceusniger strain G10 acts as an antifungal agent antagonistic towards many different phytopathogenic fungi, including different pathogenic races of the Fusarium wilt pathogen. In an attempt to understand the mode of action of this antagonist in nature, the interaction between S. violaceusniger strain G10 and F. oxysporum f.sp. cubense was first studied by paired incubation on agar plates. Evidence for the in vitro antibiosis of strain G10 was demonstrated by inhibition zones in the “cross-plug” assay plates. Microscopic observations showed lysis of hyphal ends in the inhibited fungal colonies. Culture of strain G10 in liquid media produces antifungal metabolites, which showed in vitro antagonistic effects against F. oxysporum f.sp. cubense such as swelling, distortion and excessive branching of hyphae, and inhibition of spore germination. An indirect method was used to show that antibiosis is one of the mechanisms of antagonism by which strain G10 acts against F. oxysporun f.sp. cubense in soil. This study suggests the potential of developing strain G10 for the biological control of Fusarium wilt disease of banana. Journal of Industrial Microbiology & Biotechnology (2002) 28, 303–310 DOI: 10.1038/sj/jim/7000247

Use of Chlorella vulgaris for CO2 mitigation in a photobioreactor

Abstract: Carbon dioxide (CO2) is a colorless gas that exists at a concentration of approximately 330 ppm in the atmosphere and is released in great quantities when fossil fuels are burned. The current flux of carbon out of fossil fuels is about 600 times greater than that into fossil fuels. With increased concerns about global warming and greenhouse gas emissions, there have been several approaches proposed for managing the levels of CO2 emitted into the atmosphere. One of the most understudied methods for CO2 mitigation is the use of biological processes in engineered systems such as photobioreactors. This research project describes the effectiveness of Chlorella vulgaris, used in a photobioreactor with a very short gas residence time, in sequestering CO2 from an elevated CO2 airstream. We evaluated a flow-through photobioreactor's operational parameters, as well as the growth characteristics of the C. vulgaris inoculum when exposed to an airstream with over 1850 ppm CO2. When using dry weight, chlorophyll, and direct microscopic measurements, it was apparent that the photobioreactor's algal inoculum responded well to the elevated CO2 levels and there was no build-up of CO2 or carbonic acid in the photobioreactor. The photobioreactor, with a gas residence time of approximately 2 s, was able to remove up to 74% of the CO2 in the airstream to ambient levels. This corresponded to a 63.9-g/m3/h bulk removal for the experimental photobioreactor. Consequently, this photobioreactor shows that biological processes may have some promise for treating point source emissions of CO2 and deserve further study.

Production technology for entomopathogenic nematodes and their bacterial symbionts.

Abstract: Entomopathogenic nematodes (genera Steinernema and Heterorhabditis) kill insects with the aid of mutualistic bacteria. The nematode-bacteria complex is mass produced for use as biopesticides using in vivo or in vitro methods, i.e., solid or liquid fermentation. In vivo production (culture in live insect hosts) is low technology, has low startup costs, and resulting nematode quality is high, yet cost efficiency is low. In vitro solid culture, i.e., growing the nematodes and bacteria on crumbled polyurethane foam, offers an intermediate level of technology and costs. In vivo production and solid culture may be improved through innovations in mechanization and streamlining. In vitro liquid culture is the most cost-efficient production method but requires the largest startup capital and nematode quality may be reduced. Liquid culture may be improved through progress in media development, nematode recovery, and bioreactor design. A variety of formulations is available to facilitate nematode storage and application.

Microbial diversity and complexity in hypersaline environments: a preliminary assessment.

Abstract: The microbial communities in solar salterns and a soda lake have been characterized using two techniques: BIOLOG, to estimate the metabolic potential, and amplicon length heterogeneity analysis, to estimate the molecular diversity of these communities. Both techniques demonstrated that the halophilic Bacteria and halophilic Archaea populations in the Eilat, Israel saltern are dynamic communities with extensive metabolic potentials and changing community structures. Halophilic Bacteria were detected in Mono Lake and the lower salinity ponds at the Shark Bay saltern in Western Australia, except when the crystallizer samples were stressed by exposure to Acid Green Dye #9899. At Shark Bay, halophilic Archaea were found only in the crystallizer samples. These data confirm both the metabolic diversity and the phylogenetic complexity of the microbial communities and assert the need to develop more versatile media for the cultivation of the diversity of bacteria in hypersaline environments.

The effects of temperature on the aerobic stability of wheat and corn silages.

Abstract: The aim of this work was to study the effects of temperature on the aerobic stability of wheat and corn silages. Three silage samples from each crop were taken from the faces of six different commercial bunker silos immediately after unloading them. The samples were exposed to air for 3 or 6 days at 10, 20, 30 or 40 degrees C. The most intensive deterioration occurred at 30 degrees C. Samples incubated at 30 degrees C had the highest yeast counts, most prolific CO(2) production and greatest increases in pH. Silage samples exposed to 10 or 40 degrees C remained stable. The duration of exposure had a significant effect on aerobic stability, especially at 30 degrees C. Temperature has a significant effect on silage aerobic stability. In a warm climate, special care should be taken during unloading of silage in order to prevent intensive aerobic deterioration.

Halocins and sulfolobicins: the emerging story of archaeal protein and peptide antibiotics.

Abstract: Production of antibiotic peptides and proteins is a near-universal feature of living organisms regardless of phylogenetic classification. Bacteriocins (proteinaceous antimicrobials from the domain Bacteria) have been studied for over 75 years, and the eucaryocins (proteinaceous antimicrobials from the domain Eucarya) since the early 1960s. However, one domain of organisms, the Archaea, containing hyperthermophiles, extreme halophiles and the methanogens, is just beginning to be scrutinized for the production of peptide antibiotics. Production of archaeal proteinaceous antimicrobials (archaeocins) from extreme halophiles (halocins) is a nearly universal feature of the rod-shaped haloarchaea. Halocin activity is first detectable in culture supernatants at the beginning of the transition into stationary phase, concomitant with an induction of transcription of the structural gene. Halocins are diverse in size, consisting of proteins as large as 35 kDa and peptide “microhalocins” as small as 3.6 kDa. The 36 amino acids of microhalocin HalS8 are located in the interior of a 311-residue pro-protein from which they are liberated by an unknown mechanism. Microhalocins are hydrophobic and robust, withstanding heat, desalting and exposure to organic solvents. Unlike the peptide bacteriocins and the eucaryocins, microhalocins possess a large number of neutral residues and are not cationic, leaving their mechanism(s) of action mostly a mystery. While microhalocins affect a variety of haloarchaeal genera (kingdom Euryarchaeota), they also exhibit cross-kingdom toxicity, inhibiting or killing Sulfolobus species (kingdom Crenarchaeota). Finally, archaeocins also are produced by the hyperthermophile “Sulfolobus islandicus”. These 20-kDa protein antibiotics are not excreted into the environment, but are associated with small particles apparently derived from the cell's S-layer. Journal of Industrial Microbiology & Biotechnology (2002) 28, 23–31 DOI: 10.1038/sj/jim/7000190

Application of metabolic engineering to improve both the production and use of biotech indigo.

Abstract: A fermentation process was developed for production of indigo from glucose using recombinant Escherichia coli. This was achieved by modifying the tryptophan pathway to cause high-level indole production and adding the Pseudomonas putida genes encoding naphthalene dioxygenase (NDO). In comparison to a tryptophan-overproducing strain, the first indigo-producing strain made less than half of the expected amount of indigo. Severe inactivation of the first enzyme of aromatic biosynthesis, 3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthase (the aroG fbr gene product), was observed in cells collected from indigo fermentations. Subsequent in vitro experiments revealed that DAHP synthase was inactivated by exposure to the spontaneous chemical conversion of indoxyl to indigo. Indigo production was thereafter improved by increasing the gene dosage of aroG fbr or by increasing substrate availability to DAHP synthase in vivo by either amplifying the tktA (transketolase) gene or inactivating both isozymes of pyruvate kinase. By combining all three strategies for enhancing DAHP formation in the cell, a 60% increase in indigo production was achieved. Metabolic engineering was then further applied to eliminate a byproduct of the spontaneous conversion of indoxyl to indigo, thereby solving a serious problem with the use of bio-indigo in the final denim dyeing application. Journal of Industrial Microbiology & Biotechnology (2002) 28, 127–133 DOI: 10.1038/sj/jim/7000228

Production of butanol from starch-based waste packing peanuts and agricultural waste.

Abstract: We examined the fermentation of starch-based packing peanuts and agricultural wastes as a source of fermentable carbohydrates using Clostridium beijerinckii BA101. Using semidefined P2 medium containing packing peanuts and agricultural wastes, instead of glucose as a carbohydrate source, we measured characteristics of the fermentation including solvent production, productivity, and yield. With starch as substrate (control), the culture produced 24.7 g l−1 acetone–butanol–ethanol (ABE), while with packing peanuts it produced 21.7 g l−1 total ABE with a productivity of 0.20 g l−1 h−1 and a solvent (ABE) yield of 0.37. Cell growth in starch, packing peanuts, and agricultural wastes medium was different, possibly due to the different nature of these substrates. Using model agricultural waste, 20.3g l−1 ABE was produced; when using actual waste, 14.8 g l−1 ABE was produced. The use of inexpensive substrates will increase the economic viability of the conversion of biomass to butanol, and can provide new markets for these waste streams. Journal of Industrial Microbiology & Biotechnology (2002) 29, 117–123 doi: 10.1038/sj.jim.7000285

Deep-sea hydrothermal vents: a new source of innovative bacterial exopolysaccharides of biotechnological interest?

Abstract: Polysaccharides and, in particular, microbial polysaccharides represent a class of important products of growing interest for many sectors of industry. Although many known marine bacteria produce exopolysaccharides (EPS), continuation in looking for new polysaccharide-producing microorganisms is promising. Hydrothermal deep-sea vents could be a source of novel EPS as indicated by the screening of a number of mesophilic heterotrophic bacteria recovered from different locations. Although originating from such extreme environment, some bacteria were shown to biosynthesize innovative EPS under laboratory conditions. Their specific rheological properties either in the presence or absence of monovalent and divalent ions, biological activities, metal binding capabilities, and novel chemical composition mean that these EPS are expected to find many applications in the near future.

Substrate-dependent autoaggregation of Pseudomonas putida CP1 during the degradation of mono-chlorophenols and phenol.

Abstract: A bacterium, CP1, identified as Pseudomonas putida strain, was investigated for its ability to grow on and degrade mono-chlorophenols and phenols as sole carbon sources in aerobic shaking batch culture. The organism degraded up to 1.56 mM 2- and 3-chlorophenol, 2.34 mM 4-chlorophenol and 8.5 mM phenol using an ortho-cleavage pathway. P. putida CP1, acclimated to degrade 2-chlorophenol, was capable of 3-chlorocatechol degradation, while P. putida, acclimated to 4-chlorophenol degradation, degraded 4-chlorocatechol. Growth of P. putida CP1 on higher concentrations of the mono-chlorophenols, >or=1.56 mM 4-chlorophenol and >or=0.78 mM 2- and 3-chlorophenol, resulted in decreases in cell biomass despite metabolism of the substrates, and the formation of large aggregates of cells in the culture medium. Increases in cell biomass with no clumping of the cells resulted from growth of P. putida CP1 on phenol or on lower concentrations of mono-chlorophenol. Bacterial adherence to hydrocarbons (BATH) assays showed cells grown on the higher concentrations of mono-chlorophenol to be more hydrophobic than those grown on phenol and lower concentrations of mono-chlorophenol. The results suggested that increased hydrophobicity and autoaggregation of P. putida CP1 were a response to toxicity of the added substrates.

Fermentation of sugar mixtures using Escherichia coli catabolite repression mutants engineered for production of L-lactic acid.

Abstract: Conversion of lignocellulose to lactic acid requires strains capable of fermenting sugar mixtures of glucose and xylose. Recombinant Escherichia coli strains were engineered to selectively produce L-lactic acid and then used to ferment sugar mixtures. Three of these strains were catabolite repression mutants (ptsG −) that have the ability to simultaneously ferment glucose and xylose. The best results were obtained for ptsG − strain FBR19. FBR19 cultures had a yield of 0.77 (g lactic acid/g added sugar) when used to ferment a 100 g/l total equal mixture of glucose and xylose. The strain also consumed 75% of the xylose. In comparison, the ptsG + strains had yields of 0.47–0.48 g/g and consumed 18–22% of the xylose. FBR19 was subsequently used to ferment a variety of glucose (0–40 g/l) and xylose (40 g/l) mixtures. The lactic acid yields ranged from 0.74 to 1.00 g/g. Further experiments were conducted to discover the mechanism leading to the poor yields for ptsG + strains. Xylose isomerase (XI) activity, a marker for induction of xylose metabolism, was monitored for FBR19 and a ptsG + control during fermentations of a sugar mixture. Crude protein extracts prepared from FBR19 had 10–12 times the specific XI activity of comparable samples from ptsG + strains. Therefore, higher expression of xylose metabolic genes in the ptsG − strain may be responsible for superior conversion of xylose to product compared to the ptsG + fermentations.

Production, recovery and immunogenicity of the protective antigen from a recombinant strain of Bacillus anthracis.

Abstract: The protective antigen (PA) is one of the three components of the anthrax toxin. It is a secreted nontoxic protein with a molecular weight of 83 kDa and is the major component of the currently licensed human vaccine for anthrax. Due to limitations found in the existing vaccine formulation, it has been proposed that genetically modified PA may be more effective as a vaccine. The expression and the stability of two recombinant PA (rPA) variants, PA-SNKE-ΔFF-E308D and PA-N657A, were studied. These proteins were expressed in the nonsporogenic avirulent strain BH445. Initial results indicated that PA-SNKE-ΔFF-E308D, which lacks two proteolysis-sensitive sites, is more stable than PA-N657A. Process development was conducted to establish an efficient production and purification process for PA-SNKE-ΔFF-E308D. pH, media composition, growth strategy and protease inhibitors composition were analyzed. The production process chosen was based on batch growth of B. anthracis using tryptone and yeast extract as the only source of carbon, pH control at 7.5, and antifoam 289. Optimal harvest time was 14–18 h after inoculation, and EDTA (5 mM) was added upon harvest for proteolysis control. Recovery of the rPA was performed by expanded-bed adsorption (EBA) on a hydrophobic interaction chromatography (HIC) resin, eliminating the need for centrifugation, microfiltration and diafiltration. The EBA step was followed by ion exchange and gel filtration. rPA yields before and after purification were 130 and 90 mg/l, respectively. The purified rPA, without further treatment, treated with small amounts of formalin or adsorbed on alum, induced, high levels of IgG anti-PA with neutralization activities. Journal of Industrial Microbiology & Biotechnology (2002) 28, 232–238 DOI: 10.1038/sj/jim/7000239

Selection of Escherichia coli-inhibiting strains of Lactobacillus paracasei subsp. paracasei

Abstract: The aim of this study was to select Escherichia coli-inhibiting strains among lactic acid bacteria. On the basis of phenotypical and technological characteristics, 20 strains of lactic acid bacteria were screened from a total of 225 isolates, obtained from nine samples of artisanal Caprino d'Aspromonte cheese, made from raw goats' milk. The antagonistic activity of these 20 strains was detected in plates against three different strains of E. coli. Two strains of Lactobacillus paracasei subsp. paracasei showed a marked anti-E. coli activity against all three strains tested; the other lactic acid bacteria did not exhibit inhibiting activity. The E. coli inhibition can be ascribed to production of bacteriocin-like compounds. The use of L. paracasei subsp. paracasei strains to increase the safety of the cheeses made from raw milk is recommended because these cultures strongly inhibit E. coli, without foreseeable adverse sensory changes.

Biodegradation of a crude oil by three microbial consortia of different origins and metabolic capabilities.

Abstract: Microbial consortia were obtained three by sequential enrichment using different oil products. Consortium F1AA was obtained on a heavily saturated fraction of a degraded crude oil; consortium TD, by enrichment on diesel and consortium AM, on a mixture of five polycyclic aromatic hydrocarbons [PAHs]. The three consortia were incubated with a crude oil in order to elucidate their metabolic capabilities and to investigate possible differences in the biodegradation of these complex hydrocarbon mixtures in relation to their origin. The efficiency of the three consortia in removing the saturated fraction was 60% (F1AA), 48% (TD) and 34% (AM), depending on the carbon sources used in the enrichment procedures. Consortia F1AA and TD removed 100% of n-alkanes and branched alkanes, whereas with consortium AM, 91% of branched alkanes remained. Efficiency on the polyaromatic fraction was 19% (AM), 11% (TD) and 7% (F1AA). The increase in aromaticity of the polyaromatic fraction during degradation of the crude oil by consortium F1AA suggested that this consortium metabolized the aromatic compounds primarily by oxidation of the alkylic chains. The 500-fold amplification of the inocula from the consortia by subculturing in rich media, necessary for use of the consortia in bioremediation experiments, showed no significant decrease in their degradation capability.

Ensiling whole-crop wheat and corn in large containers with Lactobacillus plantarum and Lactobacillus buchneri

Abstract: The effect of applying Lactobacillus buchneri, alone or in combination with Lactobacillus plantarum, at ensiling, on the aerobic stability of wheat and corn silages was studied in 50-l plastic containers. Treatments comprised control (no additives), L. plantarum, L. buchneri and a combination of L. plantarum+L. buchneri. After 3 months of storage, the wheat silages treated with L. buchneri had higher acetic acid contents than the control or L. plantarum-treated silages, and were free of mold, whereas the top layers of the control or L. plantarum-treated silages were moldy. In an aerobic stability test the L. buchneri-treated silages were stable, whereas those treated with L. plantarum deteriorated. In the corn silages the effects of L. buchneri were not as clear and the top layer was moldy in all silages. However, L. buchneri also improved the aerobic stability of the corn silage, as indicated by lower yeast numbers, less CO2 production and stable pH. It is concluded that L. buchneri has a potential as a silage additive that protects the silage upon aerobic exposure. The 50-l plastic containers can serve as an appropriate model to test silage additives before conducting full-scale farm experiments. Journal of Industrial Microbiology & Biotechnology (2002) 28, 7–11 DOI: 10.1038/sj/jim/7000207

Gellan gum biosynthesis in Sphingomonas paucimobilis ATCC 31461: genes, enzymes and exopolysaccharide production engineering.

Abstract: The commercial gelling agent, gellan, is an extracellular polysaccharide (EPS) produced by Sphingomonas paucimobilis ATCC 31461. In recent years, significant progress in understanding the relationship between gellan structure and properties and elucidation of the biosynthesis and engineering of this recent product of biotechnology has been made. This review focuses on recent advances in this field. Emphasis is given to identification and characterization of genes and enzymes involved, or predicted to be involved, in the gellan biosynthetic pathway, at the level of synthesis of sugar-activated precursors, of the repeat unit assembly and of gellan polymerization and export. Identification of several genes, biochemical characterization of the encoded enzymes and elucidation of crucial steps of the gellan pathway indicate that possibilities now exist for exerting control over gellan production at any of the three levels of its biosynthesis. However, a better knowledge of the poorly understood steps and of the bottlenecks and regulation of the pathway, the characterization of the composition, structure and functional properties of gellan-like polymers produced either by the industrial strain under different culture conditions or by mutants are still required for eventual success of the metabolic engineering of gellan production.

Glucooligosaccharides from Leuconostoc mesenteroides B-742 (ATCC 13146): a potential prebiotic.

Abstract: There is an emerging market for functional oligosaccharides for use in foods. Currently, technology for the production of oligosaccharides is limited to extraction from plant sources, acid or enzymatic hydrolysis of polysaccharides or synthesis by transglycosylation reactions. Oligosaccharides can also be produced using a Leuconostoc fermentation and restricting the polymer size by addition of maltose. Maltose limits the dextransucrase reaction, yielding high concentrations of α-glucooligosaccharides. Branched oligomers produced by this process were readily catabolized by bifidobacteria and lactobacilli but were not readily utilized by either Salmonella sp. or Escherichia coli, pointing toward their use in intestinal microflora modification. Journal of Industrial Microbiology & Biotechnology (2002) 29, 196–199 doi:10.1038/sj.jim.7000269

The influence of fluid shear on the structure and material properties of sulphate-reducing bacterial biofilms.

Abstract: Biofilms of sulphate-reducing Desulfovibrio sp. EX265 were grown in square section glass capillary flow cells under a range of fluid flow velocities from 0.01 to 0.4 m/s (wall shear stress, τw, from 0.027 to 1.0 N/m2). In situ image analysis and confocal scanning laser microscopy revealed biofilm characteristics similar to those reported for aerobic biofilms. Biofilms in both flow cells were patchy and consisted of cell clusters separated by voids. Length-to-width ratio measurements (l c:w c) of biofilm clusters demonstrated the formation of more “streamlined” biofilm clusters (l c:w c=3.03) at high-flow velocity (Reynolds number, Re, 1200), whereas at low-flow velocity (Re 120), the l c:w c of the clusters was approximately 1 (l c:w c of 1 indicates no elongation in the flow direction). Cell clusters grown under high flow were more rigid and had a higher yield point (the point at which the biofilm began to flow like a fluid) than those established at low flow and some biofilm cell aggregates were able to relocate within a cluster, by travelling in the direction of flow, before attaching more firmly downstream.

Influence of the yeast strain on the changes of the amino acids, peptides and proteins during sparkling wine production by the traditional method.

Abstract: The influence of five yeast strains on the nitrogen fractions, amino acids, peptides and proteins, during 12 months of aging of sparkling wines produced by the traditional or Champenoise method, was studied. High-performance liquid chromatography (HPLC) techniques were used for analysis of the amino acid and peptide fractions. Proteins plus polypeptides were determined by the colorimetric Bradford method. Four main stages were detected in the aging of wines with yeast. In the first stage, a second fermentation took place; amino acids and proteins plus polypeptides diminished, and peptides were liberated. In the second stage, there was a release of amino acids and proteins, and peptides were degraded. In the third stage, the release of proteins and peptides predominated. In the fourth stage, the amino acid concentration diminished. The yeast strain used influenced the content of free amino acids and peptides and the aging time in all the nitrogen fractions.

Planktonic nitrate-reducing bacteria and sulfate-reducing bacteria in some western Canadian oil field waters.

Abstract: Oil fields that use water flooding to enhance oil recovery may become sour because of the production of H2S from the reduction of sulfate by sulfate-reducing bacteria (SRB) The addition of nitrate to produced waters can stimulate the activities of nitrate-reducing bacteria (NRB) and control sulfide production Many previous studies have focused on chemolithotrophic bacteria that can use thiosulfate or sulfide as energy sources while reducing nitrate Little attention has been given to heterotrophic NRB in oil field waters Three different media were used in this study to enumerate various types of planktonic NRB present in waters from five oil fields in western Canada The numbers of planktonic SRB and bacteria capable of growth under aerobic conditions were also determined In general, microbial numbers in the produced waters were very low (<10 ml−1) in samples taken near or at wellheads However, the numbers increased in the aboveground facilities No thiosulfate-oxidizing NRB were detected in the oil field waters, but other types of NRB were detected in 16 of 18 produced water samples The numbers of heterotrophic NRB were equal to or greater than the number of sulfide-oxidizing, chemolithotrophic NRB in 12 of 15 samples These results showed that each of the oil fields contained NRB, which might be stimulated by nitrate amendment to control H2S production by SRB Journal of Industrial Microbiology & Biotechnology (2002) 29, 83–92 doi:101038/sjjim7000274

Role of dose concentration in biocide efficacy against Pseudomonas aeruginosa biofilms.

Abstract: entrapped in alginate gel beads to form artificial biofilms resisted killing by chlorine, glutaraldehyde, 2,2-dibromo-3-nitrilopropionamide (DBNPA), and an alkyl dimethyl benzyl ammonium compound (ADBAC). The degree of resistance was quantified by a resistance factor that compared killing times for biofilm and planktonic cells in response to the same concentration of antimicrobial agent. Resistance factors averaged 120 for chlorine, 34 for glutaraldehyde, 29 for DBNPA, and 1900 for ADBAC. In every case, resistance factors decreased with increasing concentration of the antimicrobial agent. An independent analysis of the concentration dependence of the apparent rates of killing of planktonic and biofilm bacteria showed that elevating the treatment concentration increased bacterial killing more in the biofilm than it did in a suspension culture. Calculation of a transport modulus comparing the rates of biocide reaction and diffusion suggested that at least part of the biofilm resistance to chlorine, glutaraldehdye, and DBNPA could be attributed to incomplete or slow penetration of these agents into the biofilm. Time-kill curves were nonlinear for biofilm bacteria in some cases. The shapes of these curves implicated retarded antimicrobial penetration for chlorine and glutaraldehyde and the presence of a tolerant subpopulation for DBNPA and ADBAC. The results indicate that treating biofilms with a concentrated dose of biocide is more effective than using prolonged doses of a lower concentration. Journal of Industrial Microbiology & Biotechnology (2002) 29, 10–15 doi:10.1038/sj.jim.7000256

Production and properties of an alkaline, thermophilic lipase from Pseudomonas fluorescens NS2W.

Abstract: Eighteen bacterial strains were isolated from soil samples and screened for alkaline, thermophilic lipase production. Pseudomonas fluorescens NS2W was selected and its production of lipase was optimized in shake flasks using a statistical experimental design. Cell growth and lipase production were studied in shake flasks and in a 1-l fermenter in the optimized medium. Maximum lipase yields were 69.7 and 68.7 U ml−1, respectively. The optimized medium resulted in about a five-fold increase in the enzyme production, compared to that obtained in the basal medium. The lipase had an optimal activity at pH 9.0 and was stable over a wide pH range of 3–11 with more than 70% activity retention. The lipase had an optimal activity at 55°C and was stable up to 60°C with more than 70% activity retention for at least 2 h. Journal of Industrial Microbiology & Biotechnology (2002) 28, 344–348 DOI: 10.1038/sj/jim/7000254

Isolation and characterization of a feather-degrading bacterium from the poultry processing industry

Abstract: A Flavobacterium sp producing a high keratinolytic activity was isolated from a poultry industry after growth on selective feather meal agar This bacterium grew on feather meal broth, producing keratinase, and was also capable of complete degradation of raw feathers The proteolytic activity was assessed in the presence of specific protease inhibitors The crude enzyme showed mainly metalloprotease character This novel isolate would have potential biotechnological use in processes involving keratin hydrolysis

Extracellular production of L-ascorbic acid by Chlorella protothecoides, Prototheca species, and mutants of P. moriformis during aerobic culturing at low pH.

Abstract: Nine strains of Chlorella protothecoides and 43 strains representing the five species of Prototheca were screened in flask culture for their ability to synthesize L-ascorbic acid (AA). Ascorbic acid was detected in all strains, ranging from 4.8 to 0.38 mg AA g x (-1) of dry cells. Organisms selected for further study grew well and maintained their AA productivity above a pH of 3.5. They can produce AA using a variety of carbon and nitrogen sources. Aerobic fermentation of selected strains resulted in extracellular accumulation of AA up to 76 mg x l(-1). By classical mutagenesis and selection methods, we created mutants of Prototheca moriformis ATCC 75669 that produced greater quantities of AA than the wild-type strain (78.4 vs 21.9 mg AA g x (-1) of cells). A process based on extracellular production could greatly reduce the cost of AA manufacture by eliminating the need for extraction of the AA from the cells.

A mathematical model of quorum sensing in a growing bacterial biofilm.

Abstract: In a process called quorum sensing, bacteria monitor their population density via extracellular signaling molecules and modulate gene expression accordingly. This paper describes a one-dimensional model of a growing Pseudomonas aeruginosa biofilm. Quorum sensing has been included in the model by the addition of equations describing the production, degradation, and diffusion of acyl-homoserine lactones in the biofilm. In order for quorum sensing to initiate near the substratum, in accordance with experimental observations, model results suggest that cells in oxygen-deficient regions of the biofilm must still be synthesizing the signal compound. This result highlights the importance of careful study of the relationship between metabolic activity of the bacterium and signal synthesis.