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Applied Microbiology and Biotechnology (v.86, #3)

Cofactor-independent oxidases and oxygenases by Susanne Fetzner; Roberto A. Steiner (pp. 791-804).

Whereas the majority of O₂-metabolizing enzymes depend on transition metal ions or organic cofactors for catalysis, a significant number of oxygenases and oxidases neither contain nor require any cofactor. Among the cofactor-independent oxidases, urate oxidase, coproporphyrinogen oxidase, and formylglycine-generating enzyme are of mechanistic as well as medical interest. Formylglycine-generating enzyme is also a promising tool for protein engineering as it can be used to equip proteins with a reactive aldehyde function. PqqC, an oxidase in the biosynthesis of the bacterial cofactor pyrroloquinoline quinone, catalyzes an eight-electron ring-closure oxidation reaction. Among bacterial oxygenases, quinone-forming monooxygenases involved in the tailoring of polyketides, the dioxygenase DpgC found in the biosynthesis of a building block of vancomycin and teicoplanin antibiotics, luciferase monooxygenase from Renilla sp., and bacterial ring-cleaving 2,4-dioxygenases active towards 3-hydroxy-4(1H)-quinolones have been identified as cofactor-independent enzymes. Interestingly, the 3-hydroxy-4(1H)-quinolone 2,4-dioxygenases as well as Renilla luciferase use an α/β-hydrolase architecture for oxygenation reactions. Cofactor-independent oxygenases and oxidases catalyze very different reactions and belong to several different protein families, reflecting their diverse origin. Nevertheless, they all may share the common mechanistic concept of initial base-catalyzed activation of their organic substrate and “substrate-assisted catalysis.”

Keywords: Oxygen; Oxygenase; Oxidase; Cofactor-independent enzymes; α/β-hydrolase fold

The antiviral protein cyanovirin-N: the current state of its production and applications by Sheng Xiong; Jun Fan; Kaio Kitazato (pp. 805-812).

Human immunodeficiency virus (HIV)/AIDS continues to spread worldwide, and most of the HIV-infected people living in developing countries have little or no access to highly active antiretroviral therapy. The development of efficient and low-cost microbicides to prevent sexual transmission of HIV should be given high priority because there is no vaccine available yet. Cyanovirin-N (CVN) is an entry inhibitor of HIV and many other viruses, and it represents a new generation of microbicide that has specific and potent activity, a different mechanism of action, and unusual chemicophysical stability. In vitro and in vivo antiviral tests suggested that the anti-HIV effect of CVN is stronger than a well-known gp120-targeted antibody (2G12) and another microbicide candidate, PRO2000. CVN is a cyanobacteria-derived protein that has special structural features, making the artificial production of this protein very difficult. In order to develop an efficient and relatively low-cost approach for large-scale production of recombinant CVN to satisfy medical use, this protein has been expressed in many systems by trial and error. Here, to summarize the potential and remaining challenges for the development of this protein into an HIV prevention agent, the progress in the structural mechanism determination, heterologous production and pharmacological evaluation of CVN is reviewed.

Keywords: Human immunodeficiency virus; Microbicide; Cyanovirin-N; gp120 binding; Pharmacological evaluation; Heterologous production

Molecular mechanisms of compounds affecting bacterial biofilm formation and dispersal by Paolo Landini; Davide Antoniani; J. Grant Burgess; Reindert Nijland (pp. 813-823).

Bacteria can switch between planktonic forms (single cells) and biofilms, i.e., bacterial communities growing on solid surfaces and embedded in a matrix of extracellular polymeric substance. Biofilm formation by pathogenic bacteria often results in lower susceptibility to antibiotic treatments and in the development of chronic infections; thus, biofilm formation can be considered an important virulence factor. In recent years, much attention has been directed towards understanding the biology of biofilms and towards searching for inhibitors of biofilm development and of biofilm-related cellular processes. In this report, we review selected examples of target-based screening for anti-biofilm agents: We focus on inhibitors of quorum sensing, possibly the most characterized target for molecules with anti-biofilm activity, and on compounds interfering with the metabolism of the signal molecule cyclic di-GMP metabolism and on inhibitors of DNA and nucleotide biosynthesis, which represent a novel and promising class of biofilm inhibitors. Finally, we discuss the activation of biofilm dispersal as a novel mode of action for anti-biofilm compounds.

Keywords: Biofilm formation and dispersal; Quorum sensing; c-di-GMP; Target-directed screening; Structure-directed screening; Antimicrobial drugs

Biological control of microbial attachment: a promising alternative for mitigating membrane biofouling by Yanghui Xiong; Yu Liu (pp. 825-837).

Microbial attachment to a solid surface is a universal phenomenon occurring in both natural and engineering systems and is responsible for various types of biofouling. Membrane systems have been widely applied in drinking water production, wastewater reuse, and seawater desalination. However, membrane biofouling is the bottleneck that limits the development of membrane systems. In this review, some biological control strategies of microbial attachment which would have great potential in alleviating membrane biofouling are discussed, including inhibition of quorum sensing system, nitric oxide-induced biofilm dispersal, enzymatic disruption of extracellular polysaccharides, proteins, and DNA, inhibition of microbial attachment by energy uncoupling, use of cell wall hydrolases, and disruption of biofilm by bacteriophage. It appears that biological control of microbial attachment would be a novel and promising alternative for mitigating membrane biofouling and would be a new research niche that deserves further study.

Keywords: Microbial attachment; Membrane fouling; Quorum sensing; Autoinducers; Nitric oxide; EPS; eDNA; Energy uncoupling; Cell wall hydrolases; Bacteriophage

Current approaches for the assessment of in situ biodegradation by Petra Bombach; Hans H. Richnow; Matthias Kästner; Anko Fischer (pp. 839-852).

Considering the high costs and technical difficulties associated with conventional remediation strategies, in situ biodegradation has become a promising approach for cleaning up contaminated aquifers. To verify if in situ biodegradation of organic contaminants is taking place at a contaminated site and to determine if these processes are efficient enough to replace conventional cleanup technologies, a comprehensive characterization of site-specific biodegradation processes is essential. In recent years, several strategies including geochemical analyses, microbial and molecular methods, tracer tests, metabolite analysis, compound-specific isotope analysis, and in situ microcosms have been developed to investigate the relevance of biodegradation processes for cleaning up contaminated aquifers. In this review, we outline current approaches for the assessment of in situ biodegradation and discuss their potential and limitations. We also discuss the benefits of research strategies combining complementary methods to gain a more comprehensive understanding of the complex hydrogeological and microbial interactions governing contaminant biodegradation in the field.

Keywords: Microbial in situ degradation; Contaminated aquifer; In situ microcosm; Compound-specific isotope analysis; Metabolites; Functional genes

Effects of plastic composite support and pH profiles on pullulan production in a biofilm reactor by Kuan-Chen Cheng; Ali Demirci; Jeffrey M. Catchmark (pp. 853-861).

Pullulan is a linear homopolysaccharide which is composed of glucose units and often described as α-1, 6-linked maltotriose. The applications of pullulan range from usage as blood plasma substitutes to environmental pollution control agents. In this study, a biofilm reactor with plastic composite support (PCS) was evaluated for pullulan production using Aureobasidium pullulans. In test tube fermentations, PCS with soybean hulls, defatted soy bean flour, yeast extract, dried bovine red blood cells, and mineral salts was selected for biofilm reactor fermentation (due to its high nitrogen content, moderate nitrogen leaching rate, and high biomass attachment). Three pH profiles were later applied to evaluate their effects on pullulan production in a PCS biofilm reactor. The results demonstrated that when a constant pH at 5.0 was applied, the time course of pullulan production was advanced and the concentration of pullulan reached 32.9 g/L after 7-day cultivation, which is 1.8-fold higher than its respective suspension culture. The quality analysis demonstrated that the purity of produced pullulan was 95.8% and its viscosity was 2.4 centipoise. Fourier transform infrared spectroscopy spectra also supported the supposition that the produced exopolysaccharide was mostly pullulan. Overall, this study demonstrated that a biofilm reactor can be successfully implemented to enhance pullulan production and maintain its high purity.

Keywords: Pullulan; Aureobasidium pullulans ; PCS biofilm reactor; FTIR

Permeabilization of Microbacterium oxylans shifts the conversion of puerarin from puerarin-7-O-glucoside to puerarin-7-O-fructoside by Cigang Yu; Haidong Xu; Guodong Huang; Ting Chen; Guiyou Liu; Nan Chai; Yin Ji; Siyuan Wang; Yijun Dai; Sheng Yuan (pp. 863-870).

The main product of the conversion of puerarin by unpermeabilized cells of bacterium Microbacterium oxydans CGMCC 1788 was puerarin-7-O-glucoside (241 ± 31.9 µM). Permeabilization with 40% ethanol could not increase conversion yield, whereas it resulted in change of main product; a previous trace product became a main product (213 ± 48.0 µM) which was identified as a novel puerarin-7-O-fructoside by electrospray ionization time-of-flight MS, ¹³C NMR, ¹H NMR, and GC-MS analysis of sugar composition, and puerarin-7-O-glucoside became a trace product (14.8 ± 5.4 µM). However, the extract from cells of M. oxydans CGMCC 1788 permeabilized with ethanol converted puerarin to form 113.9 ± 27.7 µM puerarin-7-O-glucoside and 187.8 ± 29.5 µM puerarin-7-O-fructoside under the same conditions. When unpermeabilized intact cells were recovered and used repeatedly for the conversion of puerarin, with increase of reuse times, the yield of puerarin-7-O-glucoside gradually decreased, whereas the yield of puerarin-7-O-fructoside increased gradually in the conversion mixture. The main product of the conversion of puerarin by the tenth recycled unpremerbilized cells was puerarin-7-O-fructoside (288.4 ± 24.0 µM). Therefore, the change of permeability of cell membrane of bacterium M. oxydans CGMCC 1788 contributed to the change of conversion of the product’s composition.

Keywords: Puerarin; Puerarin-7-O-glucoside; Puerarin-7-O-fructoside; Conversion; Permeabilization; Microbacterium oxydans

Secretion of Kluyveromyces lactis Cu/Zn SOD: strategies for enhanced production by Stefano Raimondi; Daniela Uccelletti; Alberto Amaretti; Alan Leonardi; Claudio Palleschi; Maddalena Rossi (pp. 871-878).

The Kluyveromyces lactis Cu/Zn SOD gene (SOD1) was fused with the toxin K1 signal sequence to obtain extracellular production of superoxide dismutase. Kluyveromyces marxianus L3 and K. lactis MW98-8C strains were transformed and compared as hosts for the secretion. The effects of the media composition were evaluated: In K. lactis, the highest volumetric activity was obtained in YKK synthetic medium in the presence of Cu²⁺/Zn²⁺ cofactors (9.6 kU l⁻¹). In K. marxianus, active SOD was produced only in YPD medium supplemented with Cu²⁺ and Zn²⁺ (8.8 kU l⁻¹). In order to improve the production of secreted active SOD in K. lactis, the SOD1 copper carrier (CCS1) was overexpressed and targeted to the secretory apparatus. A positive effect was observed only when K. lactis was grown in a medium without Cu²⁺/Zn²⁺ supplement. The best performing culture conditions for K. lactis and K. marxianus recombinant strains were successfully applied to two laboratory-scale fed-batch processes, and volumetric SOD activities increased up to 19.4 and 24.1 kU l⁻¹, respectively.

Keywords: Kuyveromyces marxianus ; Kluyveromyces lactis ; Superoxide dismutase; Proteins secretion

Cloning, overexpression, and characterization of recombinant heparinase III from Bacteroides stercoris HJ-15 by Yang-Jin Hyun; Jeong Hoon Lee; Dong-Hyun Kim (pp. 879-890).

Recombinant heparinase III (rHepIII) from Bacteroides stercoris HJ-15 was cloned, expressed, and characterized. The full-length heparinase III gene from B. stercoris HJ-15 was identified by Southern blotting, and the sequence was deposited in GenBank. The heparinase III gene, which is 2,001-bp long, was cloned and overexpressed in Escherichia coli; highly active rHepIII was easily purified using only one step of immobilized Ni²⁺ affinity column chromatography. Enzymatic properties and substrate specificities of rHepIII were assessed, and its kinetic constants were calculated. rHepIII was most active in 50 mM sodium phosphate buffer with 350 mM NaCl (pH 6.6) at 45°C. Through amino acid modification studies and site-directed mutagenesis assay, cysteines and histidines were identified as crucial residues for enzymatic activity. Moreover, this enzyme digested not only heparan sulfate but also heparin and hyaluronic acid, and their degradation products were verified by strong anion exchange/high-performance liquid chromatography. These characteristics, including active residues and substrate specificities were interesting compared with those of existing heparinase III from other species. We anticipate that the convenience of purification and the characteristics of this enzyme will make it a powerful tool for studies of glycosaminoglycans and their lyases.

Keywords: Recombinant heparinase III; Bacteroides stercoris HJ-15; Cloning and overexpression; Characterization; SAX–HPLC

Characterization of modification enzyme NukM and engineering of a novel thioether bridge in lantibiotic nukacin ISK-1 by Kouki Shioya; Yoshitaka Harada; Jun-ichi Nagao; Jiro Nakayama; Kenji Sonomoto (pp. 891-899).

The lantibiotic nukacin ISK-1 is an antimicrobial peptide containing unusual amino acids such as lanthionine and dehydrobutyrine. The nukacin ISK-1 prepeptide (NukA) undergoes posttranslational modifications, such as the dehydration and cyclization reactions required to form the unusual amino acids by the modification enzyme NukM. We have previously constructed a system for the introduction of unusual amino acids into NukA by coexpression of NukM in Escherichia coli. Using this system, we describe the substrate specificity of NukM by the coexpression of a series of NukA mutants. Our results revealed the following characteristics of NukM: (1) its dehydration activity is not coupled to its cyclization activity; (2) its dehydration activity is site-specific; (3) the length of the substrate is important for its dehydration activity. Furthermore, we succeeded in introducing a novel thioether bridge in NukA by replacing an unmodified Ser at position 27 with a Cys residue.

Keywords: Lantibiotic; Unusual amino acids; Modification enzyme; Peptide engineering; Posttranslational modification

Identification in Agrobacterium tumefaciens of the d-galacturonic acid dehydrogenase gene by Harry Boer; Hannu Maaheimo; Anu Koivula; Merja Penttilä; Peter Richard (pp. 901-909).

There are at least three different pathways for the catabolism of d-galacturonate in microorganisms. In the oxidative pathway, which was described in some prokaryotic species, d-galacturonate is first oxidised to meso-galactarate (mucate) by a nicotinamide adenine dinucleotide (NAD)-dependent dehydrogenase (EC 1.1.1.203). In the following steps of the pathway mucate is converted to 2-keto-glutarate. The enzyme activities of this catabolic pathway have been described while the corresponding gene sequences are still unidentified. The d-galacturonate dehydrogenase was purified from Agrobacterium tumefaciens, and the mass of its tryptic peptides was determined using MALDI-TOF mass spectrometry. This enabled the identification of the corresponding gene udh. It codes for a protein with 267 amino acids having homology to the protein family of NAD(P)-binding Rossmann-fold proteins. The open reading frame was functionally expressed in Saccharomyces cerevisiae. The N-terminally tagged protein was not compromised in its activity and was used after purification for a kinetic characterization. The enzyme was specific for NAD and accepted d-galacturonic acid and d-glucuronic acid as substrates with similar affinities. NMR analysis showed that in water solution the substrate d-galacturonic acid is predominantly in pyranosic form which is converted by the enzyme to 1,4 lactone of galactaric acid. This lactone seems stable under intracellular conditions and does not spontaneously open to the linear meso-galactaric acid.

Keywords: meso-galactaric acid; Mucic acid; Lactone; Oxidative pathway; EC 1.1.1.203

Requirement of de novo synthesis of the OdhI protein in penicillin-induced glutamate production by Corynebacterium glutamicum by Jongpill Kim; Hirohisa Fukuda; Takashi Hirasawa; Keisuke Nagahisa; Kazuo Nagai; Masaaki Wachi; Hiroshi Shimizu (pp. 911-920).

We found that penicillin-induced glutamate production by Corynebacterium glutamicum is inhibited when a de novo protein synthesis inhibitor, chloramphenicol, is added simultaneously with penicillin. When chloramphenicol was added 4 h after penicillin addition, glutamate production was essentially unaffected. ³H-Leucine incorporation experiments revealed that protein synthesis continued for 1 h after penicillin addition and then gradually decreased. These results suggest that de novo protein synthesis within 4 h of penicillin treatment is required for the induction of glutamate production. To identify the protein(s) necessary for penicillin-induced glutamate production, proteome analysis of penicillin-treated C. glutamicum cells was performed with two-dimensional gel electrophoresis. Of more than 500 proteins detected, the amount of 13 proteins, including OdhI (an inhibitory protein for 2-oxoglutarate dehydrogenase complex), significantly increased upon penicillin treatment. Artificial overexpression of the odhI gene resulted in the decreased specific activity of the 2-oxoglutarate dehydrogenase complex and increased glutamate production without any triggers. These results suggest that the de novo synthesis of OdhI is the necessary factor for penicillin-induced glutamate overproduction by C. glutamicum. Moreover, continuous glutamate production was achieved by overexpression of odhI without any triggers. Thus, the odhI-overexpressing strain of C. glutamicum can be useful for efficient glutamate production.

Keywords: Corynebacterium glutamicum ; Glutamate; Penicillin; OdhI; De novo protein synthesis

Regulation of genes in Streptomyces bacteria required for catabolism of lignin-derived aromatic compounds by Jennifer R. Davis; Jason K. Sello (pp. 921-929).

The major utilization pathway for lignin-derived aromatic compounds in microorganisms is the β-ketoadipate pathway. Through this pathway, the aromatic compounds protocatechuate and catechol are converted to acetyl coenzyme A and succinyl coenzyme A. The enzymes of the protocatechuate branch of this pathway are encoded by the pca genes. Here, we describe a gene cluster in Streptomyces coelicolor containing the pca structural genes and a regulatory gene required for the catabolism of protocatechuate. We found that transcription of the structural genes in S. coelicolor is induced by protocatechuate and p-hydroxybenzoate. We also observed inducible transcription of pca structural genes in the ligninolytic strain Streptomyces viridosporus ATCC 39115. Disruption of a gene encoding a putative MarR family transcription factor that is divergently transcribed from the pca structural genes resulted in constitutive transcription of the structural genes. Thus, the transcription factor encoded by this gene is an apparent negative regulator of pca gene transcription in S. coelicolor. Our findings suggest how Streptomyces bacteria could be engineered for and used in biotechnology for the utilization and degradation of lignin and lignin-derived aromatic compounds.

Keywords: Streptomyces ; Lignin; β-Ketoadipate; Protocatechuate; Aromatic catabolism

FLO gene-dependent phenotypes in industrial wine yeast strains by Patrick Govender; Michael Bester; Florian F. Bauer (pp. 931-945).

Most commercial yeast strains are nonflocculent. However, controlled flocculation phenotypes could provide significant benefits to many fermentation-based industries. In nonflocculent laboratory strains, it has been demonstrated that it is possible to adjust flocculation and adhesion phenotypes to desired specifications by altering expression of the otherwise silent but dominant flocculation (FLO) genes. However, FLO genes are characterized by high allele heterogeneity and are subjected to epigenetic regulation. Extrapolation of data obtained in laboratory strains to industrial strains may therefore not always be applicable. Here, we assess the adhesion phenotypes that are associated with the expression of a chromosomal copy of the FLO1, FLO5, or FLO11 open reading frame in two nonflocculent commercial wine yeast strains, BM45 and VIN13. The chromosomal promoters of these genes were replaced with stationary phase-inducible promoters of the HSP30 and ADH2 genes. Under standard laboratory and wine making conditions, the strategy resulted in expected and stable expression patterns of these genes in both strains. However, the specific impact of the expression of individual FLO genes showed significant differences between the two wine strains and with corresponding phenotypes in laboratory strains. The data suggest that optimization of the flocculation pattern of individual commercial strains will have to be based on a strain-by-strain approach.

Keywords: Wine yeast; Flocculation; FLO gene expression; Cell wall proteins; Adhesion; Fermentation

Rapid solubilization of insoluble phosphate by a novel environmental stress-tolerant Burkholderia vietnamiensis M6 isolated from ginseng rhizospheric soil by Ki-Hyun Park; O-Mi Lee; Ho-Il Jung; Jin-Ha Jeong; Young-Dong Jeon; Dae-Youn Hwang; Chung-Yeol Lee; Hong-Joo Son (pp. 947-955).

We isolated and characterized novel insoluble phosphate (P)-solubilizing bacteria tolerant to environmental factors like high salt, low and high pHs, and low temperature. A bacterium M6 was isolated from a ginseng rhizospheric soil and confirmed to belong to Burkholderia vietnamiensis by BIOLOG system and 16S rRNA gene analysis. The optimal cultural conditions for the solubilization of P were 2.5% (w/v) glucose, 0.015% (w/v) urea, and 0.4% (w/v) MgCl₂·6H₂O along with initial pH 7.0 at 35°C. High-performance liquid chromatography analysis showed that B. vietnamiensis M6 produced gluconic and 2-ketogluconic acids. During the culture, the pH was reduced with increase in gluconic acid concentration and was inversely correlated with P solubilization. Insoluble P solubilization in the optimal medium was about 902 mg l⁻¹, which was approximately 1.6-fold higher than the yield in NBRIP medium (580 mg l⁻¹). B. vietnamiensis M6 showed resistance against different environmental stresses like 10–45°C, 1–5% (w/v) salt, and 2–11 pH range. The maximal concentration of soluble P produced by B. vietnamiensis M6 from Ca₃(PO₄)₂, CaHPO₄, and hydroxyapatite was 1,039, 2,132, and 1,754 mg l⁻¹, respectively. However, the strain M6 produced soluble P with 20 mg l⁻¹ from FePO₄ after 2 days and 100 mg l⁻¹ from AlPO₄ after 6 days, respectively. Our results indicate that B. vietnamiensis M6 could be a potential candidate for the development of biofertilizer applicable to environmentally stressed soil.

Keywords: Biofertilizer; Burkholderia vietnamiensis ; Insoluble phosphate; Solubilization; Stress

Light/electricity conversion by a self-organized photosynthetic biofilm in a single-chamber reactor by Koichi Nishio; Kazuhito Hashimoto; Kazuya Watanabe (pp. 957-964).

Biological energy-conversion systems are attractive in terms of their self-sustaining and self-organizing nature and are expected to be applied to low-cost and environment-friendly processes. Here we show a biofilm-based light/electricity-conversion system that was self-organized from a natural microbial community. A bioreactor equipped with an air cathode and graphite-felt anode was inoculated with a green hot-spring microbial mat. When the reactor was irradiated with light, electric current was generated between the anode and cathode in accordance with the formation of green biofilm on the anode. Fluorescence microscopy of the green biofilm revealed the presence of chlorophyll-containing microbes of ∼10 µm in size, and these cells were abundant close to the surface of the biofilm. The biofilm community was also analyzed by sequencing of polymerase chain reaction-amplified small-subunit rRNA gene fragments, showing that sequence types affiliated with Chlorophyta, Betaproteobacteria, and Bacteroidetes were abundantly detected. These results suggest that green algae and heterotrophic bacteria cooperatively converted light energy into electricity.

Keywords: Energy conversion; Microbial solar cell; Microalgae; Heterotrophs; Biofilm; Phylogeny

Saccharomyces cerevisiae CCMI 885 secretes peptides that inhibit the growth of some non-Saccharomyces wine-related strains by Helena Albergaria; Diana Francisco; Klaus Gori; Nils Arneborg; Francisco Gírio (pp. 965-972).

The nature of the toxic compounds produced by Saccharomyces cerevisiae CCMI 885 that induce the early death of Hanseniaspora guilliermondii during mixed fermentations, as well as their ability to inhibit the growth of other non-Saccharomyces wine-related strains, was investigated. The killing effect of mixed supernatants towards H. guilliermondii was inactivated by protease treatments, thus revealing the proteinaceous nature of the toxic compounds. Analysis of the protein pattern of mixed supernatants on Tricine SDS-PAGE showed that this S. cerevisiae strain secretes peptides (<10 kDa), which were detected only when death of H. guilliermondii was already established. Death-inducing supernatants were ultrafiltrated by 10 and 2 kDa membranes, respectively, and the inhibitory effect of those permeates were tested in H. guilliermondii cultures. Results indicated that the (2–10) kDa protein fraction of those supernatants seemed to contain antimicrobial peptides active against H. guilliermondii. Thus, the (2–10) kDa protein fraction was concentrated and its inhibitory effect tested against strains of Kluyveromyces marxianus, Kluyveromyces thermotolerans, Torulaspora delbrueckii and H. guilliermondii. Under the growth conditions used for these tests, the (2–10) kDa protein fraction of S. cerevisiae CCMI 885 supernatants exhibited a fungistatic effect against all the strains and a fungicidal effect against K. marxianus.

Keywords: Antimicrobial peptides; Yeast–yeast interactions; Wine fermentations; Saccharomyces cerevisiae ; Non-Saccharomyces ; Yeast population dynamic

Long-term performance of a plant microbial fuel cell with Spartina anglica by Ruud A. Timmers; David P. B. T. B. Strik; Hubertus V. M. Hamelers; Cees J. N. Buisman (pp. 973-981).

The plant microbial fuel cell is a sustainable and renewable way of electricity production. The plant is integrated in the anode of the microbial fuel cell which consists of a bed of graphite granules. In the anode, organic compounds deposited by plant roots are oxidized by electrochemically active bacteria. In this research, salt marsh species Spartina anglica generated current for up to 119 days in a plant microbial fuel cell. Maximum power production was 100 mW m⁻² geometric anode area, highest reported power output for a plant microbial fuel cell. Cathode overpotential was the main potential loss in the period of oxygen reduction due to slow oxygen reduction kinetics at the cathode. Ferricyanide reduction improved the kinetics at the cathode and increased current generation with a maximum of 254%. In the period of ferricyanide reduction, the main potential loss was transport loss. This research shows potential application of microbial fuel cell technology in salt marshes for bio-energy production with the plant microbial fuel cell.

Keywords: Plant microbial fuel cell; Rhizodeposition; Bio-energy; Spartina anglica ; Potential losses; Green electricity

Rapid and reliable detection of 11 food-borne pathogens using thin-film biosensor chips by Sulan Bai; Jinyi Zhao; Yaochuan Zhang; Wensheng Huang; Shi Xu; Haodong Chen; Liu-Min Fan; Ying Chen; Xing Wang Deng (pp. 983-990).

Traditional methods for identifying food-borne pathogens are time-consuming and laborious, so it is necessary to develop innovative methods for the rapid identification of food-borne pathogens. Here, we report the development of silicon-based optical thin-film biosensor chips for sensitive detection of 11 food-borne pathogens. Briefly, aldehyde-labeled probes were arrayed and covalently attached to a hydrazine-derivatized chip surface, and then, biotinylated polymerase chain reaction (PCR) amplicons were hybridized with the probes. After washing and brief incubation with an antibiotin immunoglobulin G–horseradish peroxidase conjugate and a precipitable horseradish peroxidase substrate, biotinylated chains bound to the probes were visualized as a color change on the chip surface (gold to blue/purple). Highly sensitive and accurate examination of PCR fragment targets can be completed within 30 min. This assay is extremely robust, sensitive, specific, and economical and can be adapted to different throughputs. Thus, a rapid, sensitive, and reliable technique for detecting 11 food-borne pathogens was successfully developed.

Keywords: Food-borne pathogen; Optical thin-film biosensor chip; Microarray; Detection

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Applied Microbiology and Biotechnology (v.86, #3)

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Applied Microbiology and Biotechnology (v.86, #3)