Science.gov

Sample records for anaerobe clostridium acetobutylicum

  1. Atmospheric vs. anaerobic processing of metabolome samples for the metabolite profiling of a strict anaerobic bacterium, Clostridium acetobutylicum.

    PubMed

    Lee, Sang-Hyun; Kim, Sooah; Kwon, Min-A; Jung, Young Hoon; Shin, Yong-An; Kim, Kyoung Heon

    2014-12-01

    Well-established metabolome sample preparation is a prerequisite for reliable metabolomic data. For metabolome sampling of a Gram-positive strict anaerobe, Clostridium acetobutylicum, fast filtration and metabolite extraction with acetonitrile/methanol/water (2:2:1, v/v) at -20°C under anaerobic conditions has been commonly used. This anaerobic metabolite processing method is laborious and time-consuming since it is conducted in an anaerobic chamber. Also, there have not been any systematic method evaluation and development of metabolome sample preparation for strict anaerobes and Gram-positive bacteria. In this study, metabolome sampling and extraction methods were rigorously evaluated and optimized for C. acetobutylicum by using gas chromatography/time-of-flight mass spectrometry-based metabolomics, in which a total of 116 metabolites were identified. When comparing the atmospheric (i.e., in air) and anaerobic (i.e., in an anaerobic chamber) processing of metabolome sample preparation, there was no significant difference in the quality and quantity of the metabolomic data. For metabolite extraction, pure methanol at -20°C was a better solvent than acetonitrile/methanol/water (2:2:1, v/v/v) at -20°C that is frequently used for C. acetobutylicum, and metabolite profiles were significantly different depending on extraction solvents. This is the first evaluation of metabolite sample preparation under aerobic processing conditions for an anaerobe. This method could be applied conveniently, efficiently, and reliably to metabolome analysis for strict anaerobes in air.

  2. Effect of iron limitation and fur gene inactivation on the transcriptional profile of the strict anaerobe Clostridium acetobutylicum.

    PubMed

    Vasileva, Delyana; Janssen, Holger; Hönicke, Daniel; Ehrenreich, Armin; Bahl, Hubert

    2012-07-01

    Iron is a nutrient of critical importance for the strict anaerobe Clostridium acetobutylicum, as it is involved in numerous basic cellular functions and metabolic pathways. A gene encoding a putative ferric uptake regulator (Fur) has been identified in the genome of C. acetobutylicum. In this work, we inactivated the fur gene by using insertional mutagenesis. The resultant mutant showed a slow-growing phenotype and enhanced sensitivity to oxidative stress, but essentially no dramatic change in its fermentation pattern. A unique feature of its physiology was the overflowing production of riboflavin. To gain further insights into the role of the Fur protein and the mechanisms for establishment of iron balance in C. acetobutylicum, we characterized and compared the gene-expression profile of the fur mutant and the iron-limitation stimulon of the parental strain. Not surprisingly, a repertoire of iron-transport systems was upregulated in both microarray datasets, suggesting that they are regulated by Fur according to the availability of iron. In addition, iron limitation and inactivation of fur affected the expression of several genes involved in energy metabolism. Among them, two genes, encoding a lactate dehydrogenase and a flavodoxin, were highly induced. In order to support the function of the latter, the ribDBAH operon responsible for riboflavin biosynthesis was also upregulated significantly. Furthermore, the iron-starvation response of C. acetobutylicum involved transcriptional modifications that were not detected in the fur mutant, suggesting that there exist additional mechanisms for adaptation to low-iron environments. Collectively, these results demonstrate that the strict anaerobe C. acetobutylicum senses and responds to availability of iron on multiple levels using a sophisticated system, and that Fur plays an important role in this process.

  3. Anaerobic biotransformation of 2,4-dinitrotoluene and 2,6-dinitrotoluene by Clostridium acetobutylicum: A pathway through dihydroxylamino intermediates

    SciTech Connect

    Hughes, J.B.; Wang, C.Y.; Zhang, C.

    1999-04-01

    Experiments were conducted to isolate and identify the intermediates and products of 2,4-dinitrotoluene and 2,6-dinitrotoluene metabolism by Clostridium acetobutylicum. Transformation of both dinitrotoluenes initially resulted in the formation of hydroxylaminonitrotoluenes. Subsequent transformation favored the formation of dihydroxylaminotoluenes, with a limited reduction to aminonitrotoluene isomers. In cell cultures, metabolism beyond the level of dihydroxylaminotoluene was not observed. In cell extracts, where activity could be maintained for periods in excess of those in cell cultures, further transformation yielded aminohydroxylaminotoluenes and eventually diaminotoluenes. These findings further demonstrate the potential for hydroxylamines to be significant intermediates of nitroaromatic transformation under anaerobic fermentative conditions. Interestingly, the rearrangement of dihydroxylaminotoluenes was not observed, as was the case in previous studies of 2,4-dihydroxylamino-6-nitrotoluene metabolism. Dihydroxylaminotoluenes were found to be quite unstable, decomposing rapidly upon exposure to oxygen, complicating the assessment of their fate in remediation processes.

  4. Cellulolytic Activity of Clostridium acetobutylicum.

    PubMed

    Lee, S F; Forsberg, C W; Gibbins, L N

    1985-08-01

    Clostridium acetobutylicum NRRL B527 and ATCC 824 exhibited extracellular and cell-bound endoglucanase and cellobiase activities during growth in a chemically defined medium with cellobiose as the sole source of carbohydrate. For both strains, the endoglucanase was found to be mainly extracellular (70 to 90%) during growth in continuous or batch cultures with the pH maintained at 5.2, whereas the cellobiase was mainly cell associated (60 to 90%). During continuous cultivation of strain B527 with cellobiose as the limiting nutrient, maximum production of the endoglucanase and cellobiase occurred at pH values of 5.2 and 4.8, respectively. In the carbon-limited continuous cultures, strain 824 produced similar levels of endoglucanase, cellobiosidase, and cellobiase activities regardless of the carbon source used. However, in ammonium- or phosphate-limited cultures, with an excess of glucose, only 1/10 of the endoglucanase was produced, and neither cellobiosidase nor cellobiase activities were detectable. A crude extracellular enzyme preparation from strain B527 hydrolyzed carboxymethylcellulose and phosphoric acid-swollen cellulose readily and microcrystalline cellulose (A vicel) to a lesser extent. Glucose accounted for more than 90% of the reducing sugar produced by the hydrolysis of acid-swollen cellulose and Avicel. Strain B527 did not grow in medium with acid-swollen cellulose as the sole source of carbohydrate, although it grew readily on the products obtained by hydrolyzing the cellulose in vitro with a preparation of extracellular cellulase derived from the same organism.

  5. Annotation of the Clostridium Acetobutylicum Genome

    SciTech Connect

    Daly, M. J.

    2004-06-09

    The genome sequence of the solvent producing bacterium Clostridium acetobutylicum ATCC824, has been determined by the shotgun approach. The genome consists of a 3.94 Mb chromosome and a 192 kb megaplasmid that contains the majority of genes responsible for solvent production. Comparison of C. acetobutylicum to Bacillus subtilis reveals significant local conservation of gene order, which has not been seen in comparisons of other genomes with similar, or, in some cases, closer, phylogenetic proximity. This conservation allows the prediction of many previously undetected operons in both bacteria.

  6. Fermentative hydrogen production in an up-flow anaerobic biofilm reactor inoculated with a co-culture of Clostridium acetobutylicum and Desulfovibrio vulgaris.

    PubMed

    Barca, Cristian; Ranava, David; Bauzan, Marielle; Ferrasse, Jean-Henry; Giudici-Orticoni, Marie-Thérèse; Soric, Audrey

    2016-12-01

    Dark fermentation systems often show low H2 yields and unstable H2 production, as the result of the variability of microbial dynamics and metabolic pathways. Recent batch investigations have demonstrated that an artificial consortium of two anaerobic bacteria, Clostridium acetobutylicum and Desulfovibrio vulgaris Hildenborough, may redirect metabolic fluxes and improve H2 yields. This study aimed at evaluating the scale-up from batch to continuous H2 production in an up-flow anaerobic packed-bed reactor (APBR) continuously fed with a glucose-medium. The effects of various parameters, including void hydraulic retention time (HRTv), pH, and alkalinity, on H2 production performances and metabolic pathways were investigated. The results demonstrated that a stable H2 production was reached after 3-4days of operation. H2 production rates increased significantly with decreasing HRTv from 4 to 2h. Instead, H2 yields remained almost stable despite the change in HRTv, indicating that the decrease in HRTv did not affect the global metabolism.

  7. Effects of butanol on Clostridium acetobutylicum.

    PubMed Central

    Bowles, L K; Ellefson, W L

    1985-01-01

    The internal pH of Clostridium acetobutylicum was determined at various stages during the growth of the organism. Even in the presence of significant quantities of acetic, butyric, and lactic acids, an internal pH of 6.2 was maintained. Experiments using N,N'-dicyclohexylcarbodiimide indicated that a functioning H+-ATPase is necessary for internal pH control. Butanol, one of the end products of the fermentation, had numerous harmful effects on C. acetobutylicum. At a concentration high enough to inhibit growth, butanol destroyed the ability of the cell to maintain internal pH, lowered the intracellular level of ATP, and inhibited glucose uptake. Experiments done at two different external pH values suggested that the butanol-mediated decrease in ATP concentration was independent of the drop in internal pH. Glucose uptake was not affected by arsenate, suggesting that uptake was not ATP dependent. The effects of butanol on C. acetobutylicum are complex, inhibiting several interrelated membrane processes. PMID:2868690

  8. Feasibility of installing and maintaining anaerobiosis using Escherichia coli HD701 as a facultative anaerobe for hydrogen production by Clostridium acetobutylicum ATCC 824 from various carbohydrates.

    PubMed

    Hassan, Sedky H A; Morsy, Fatthy Mohamed

    2015-12-01

    Using Escherichia coli for installing and maintaining anaerobiosis for hydrogen production by Clostridium acetobutylicum ATCC 824 is a cost-effective approach for industrial hydrogen production, as it does not require reducing agents or sparging with inert gases. This study was devoted for investigating the feasibility for installing and maintaining anaerobiosis of hydrogen production by C. acetobutylicum ATCC 824 when using E. coli HD701 utilizable versus non utilizable sugars as a-carbon source. Using E. coli HD701 for installing anaerobiosis showed a comparable hydrogen production yield and efficiency to the use of reducing agents and nitrogen sparging in case of hydrogen production from the E. coli HD701 non utilizable sugars. In contrast, using E. coli HD701 for installing anaerobiosis showed a lower hydrogen production yield and efficiency than the use of reducing agents and nitrogen sparging in case of using glucose as a substrate. This is possibly because E. coli HD701 when using glucose compensate for the substrate, and produce hydrogen with lower efficiency than C. acetobutylicum ATCC 824. These results indicated that the use of E. coli HD701 for installing anaerobiosis would not be economically feasible when using E. coli HD701 utilizable sugars as a carbon source. In contrast, the use of this approach for installing anaerobiosis for hydrogen production from sucrose and starch would have a high potency for industrial applications.

  9. Metabolite Analysis of Clostridium acetobutylicum: Fermentation in a Microbial Fuel Cell

    DTIC Science & Technology

    2010-01-01

    Metabolite analysis of Clostridium acetobutylicum : Fermentation in a microbial fuel cell Amethist S. Finch, Timothy D. Mackie, Christian J. Sund...Fermentation products Clostridium acetobutylicum Current generation a b s t r a c t Microbial fuel cells (MFCs) were used to monitor metabolism...changes in Clostridium acetobutylicum fer- mentations. When MFCs were inoculated with C. acetobutylicum , they generated a unique voltage output pattern

  10. Ribulokinase and transcriptional regulation of arabinose metabolism in Clostridium acetobutylicum.

    PubMed

    Zhang, Lei; Leyn, Semen A; Gu, Yang; Jiang, Weihong; Rodionov, Dmitry A; Yang, Chen

    2012-03-01

    The transcription factor AraR controls utilization of L-arabinose in Bacillus subtilis. In this study, we combined a comparative genomic reconstruction of AraR regulons in nine Clostridium species with detailed experimental characterization of AraR-mediated regulation in Clostridium acetobutylicum. Based on the reconstructed AraR regulons, a novel ribulokinase, AraK, present in all analyzed Clostridium species was identified, which was a nonorthologous replacement of previously characterized ribulokinases. The predicted function of the araK gene was confirmed by inactivation of the araK gene in C. acetobutylicum and biochemical assays using purified recombinant AraK. In addition to the genes involved in arabinose utilization and arabinoside degradation, extension of the AraR regulon to the pentose phosphate pathway genes in several Clostridium species was revealed. The predicted AraR-binding sites in the C. acetobutylicum genome and the negative effect of L-arabinose on DNA-regulator complex formation were verified by in vitro binding assays. The predicted AraR-controlled genes in C. acetobutylicum were experimentally validated by testing gene expression patterns in both wild-type and araR-inactivated mutant strains during growth in the absence or presence of L-arabinose.

  11. Fermentation of residual glycerol by Clostridium acetobutylicum ATCC 824 in pure and mixed cultures.

    PubMed

    Dams, Rosemeri I; Guilherme, Alexandre A; Vale, Maria S; Nunes, Vanja F; Leitão, Renato C; Santaella, Sandra T

    2016-12-01

    The aim of this research was to estimate the production of hydrogen, organic acids and alcohols by the strain of Clostridium acetobutylicum ATCC 824 using residual glycerol as a carbon source. The experiments were carried out in pure and mixed cultures in batch experiments. Three different sources of inocula for mixed culture were used. Ruminal liquid from goats and sludge collected from two upflow anaerobic sludge blanket reactors treating municipal wastewater and brewery effluent were tested for hydrogen, organic acids and alcohols production with or without C. acetobutylicum ATCC 824. The main detected end-products from the glycerol fermentation were hydrogen, organic acids (acetic, propionic, butyric and caproic) and alcohol (ethanol and 1,3-propanediol - 1,3PD). High hydrogen (0.44 mol H2/mol glycerol consumed) and 1,3PD (0.32 mol 1,3PD/mol glycerol consumed) yields were obtained when the strain C. acetobutylicum ATCC 824 was bioaugmented into the sludge from municipal wastewater using 5 g/L of glycerol. Significant concentrations of n-caproic acid were detected in the ruminal liquid when amended with C. acetobutylicum ATCC 824. The results suggest that glycerol can be used for the generation of H2, 1,3PD and n-caproic acid using C. acetobutylicum ATCC 824 as agent in pure or mixed cultures.

  12. Genetic and biochemical analysis of solvent formation in Clostridium acetobutylicum

    SciTech Connect

    Bennett, G.N.; Rudolph, F.B.

    1998-05-01

    The anaerobic organism Clostridium acetobutylicum has been used for commercial production of important organic solvents due to its ability to convert a wide variety of crude substrates to acids and alcohols. Current knowledge concerning the molecular genetics, cell regulation and metabolic engineering of this organism is still rather limited. The objectives are to improve the knowledge of the molecular genetics and enzymology of Clostridia in order to make genetic alterations which will more effectively channel cell metabolism toward production of desired products. Two factors that limit butanol production in continuous cultures are: (1) The degeneration of the culture, with an increase in the proportion of cells which are incapable of solvent production. Currently isolated degenerate strains are being evaluated to analyze the molecular mechanism of degeneration to determine if it is due to a genetic loss of solvent related genes, loss of a regulatory element, or an increase in general mutagenesis. Recent studies show two general types of degenerates, one which seems to have lost essential solvent pathway genes and another which has not completely lost all solvent production capability and retains the DNA bearing solvent pathway genes. (2) The production of hydrogen which uses up reducing equivalents in the cell. If the reducing power were more fully directed to the reduction reactions involved in butanol production, the process would be more efficient. The authors have studied oxidation reduction systems related to this process. These studies focus on ferredoxin and rubredoxin and their oxidoreductases.

  13. Stable Escherichia coli-Clostridium acetobutylicum shuttle vector for secretion of murine tumor necrosis factor alpha.

    PubMed

    Theys, J; Nuyts, S; Landuyt, W; Van Mellaert, L; Dillen, C; Böhringer, M; Dürre, P; Lambin, P; Anné, J

    1999-10-01

    Recombinant plasmids were constructed to secrete mouse tumor necrosis factor alpha (mTNF-alpha) from Clostridium acetobutylicum. The shuttle plasmids contained the clostridial endo-beta1, 4-glucanase (eglA) promoter and signal sequence that was fused in frame to the mTNF-alpha cDNA. The construction was first tested in Escherichia coli and then introduced in C. acetobutylicum DSM792 by electroporation. Controls confirmed the presence and stability of the recombinant plasmids in this organism. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an in vitro cytotoxic assay were used to monitor expression and secretion of mTNF-alpha during growth. Significant levels of biologically active mTNF-alpha were measured in both lysates and supernatants. The present report deals with investigations on the elaboration of a gene transfer system for cancer treatment using anaerobic bacteria.

  14. Purification and Characterization of an Autolysin from Clostridium acetobutylicum

    PubMed Central

    Webster, Jocelyn R.; Reid, Sharon J.; Jones, David T.; Woods, David R.

    1981-01-01

    A proteinaceous substance with antibiotic-like activity, resembling that of a bacteriocin, was isolated from an industrial-scale acetone-butanol fermentation of Clostridium acetobutylicum. The substance, purified by acetone precipitation, diethylaminoethyl cellulose chromatography, and polyacrylamide gel electrophoresis, was characterized as a glycoprotein with a molecular weight of 28,000. The glycoprotein was partially inactivated by certain protease enzymes. It had no effect on deoxyribonucleic acid, ribonucleic acid, or protein synthesis, and it did not result in the loss of intracellular adenosine triphosphate. The glycoprotein lysed sodium dodecyl sulfate-treated cells and cell wall preparations, and therefore it is referred to as an autolysin. The autolysin gene appeared to be chromosomal since plasmid deoxyribonucleic acid was not detected in the C. acetobutylicum strain. PMID:16345710

  15. Redox-switch regulatory mechanism of thiolase from Clostridium acetobutylicum

    PubMed Central

    Kim, Sangwoo; Jang, Yu-Sin; Ha, Sung-Chul; Ahn, Jae-Woo; Kim, Eun-Jung; Hong Lim, Jae; Cho, Changhee; Shin Ryu, Yong; Kuk Lee, Sung; Lee, Sang Yup; Kim, Kyung-Jin

    2015-01-01

    Thiolase is the first enzyme catalysing the condensation of two acetyl-coenzyme A (CoA) molecules to form acetoacetyl-CoA in a dedicated pathway towards the biosynthesis of n-butanol, an important solvent and biofuel. Here we elucidate the crystal structure of Clostridium acetobutylicum thiolase (CaTHL) in its reduced/oxidized states. CaTHL, unlike those from other aerobic bacteria such as Escherichia coli and Zoogloea ramegera, is regulated by the redox-switch modulation through reversible disulfide bond formation between two catalytic cysteine residues, Cys88 and Cys378. When CaTHL is overexpressed in wild-type C. acetobutylicum, butanol production is reduced due to the disturbance of acidogenic to solventogenic shift. The CaTHLV77Q/N153Y/A286K mutant, which is not able to form disulfide bonds, exhibits higher activity than wild-type CaTHL, and enhances butanol production upon overexpression. On the basis of these results, we suggest that CaTHL functions as a key enzyme in the regulation of the main metabolism of C. acetobutylicum through a redox-switch regulatory mechanism. PMID:26391388

  16. Metronidazole activation and isolation of Clostridium acetobutylicum electron transport genes.

    PubMed Central

    Santangelo, J D; Jones, D T; Woods, D R

    1991-01-01

    An Escherichia coli F19 recA, nitrate reductase-deficient mutant was constructed by transposon mutagenesis and shown to be resistant to metronidazole. This mutant was a most suitable host for the isolation of Clostridium acetobutylicum genes on recombinant plasmids, which activated metronidazole and rendered the E. coli F19 strain sensitive to metronidazole. Twenty-five E. coli F19 clones containing different recombinant plasmids were isolated and classified into five groups on the basis of their sensitivity to metronidazole. The clones were tested for nitrate reductase, pyruvate-ferredoxin oxidoreductase, and hydrogenase activities. DNA hybridization and restriction endonuclease mapping revealed that four of the C. acetobutylicum insert DNA fragments on recombinant plasmids were linked in an 11.1-kb chromosomal fragment. DNA sequencing and amino acid homology studies indicated that this DNA fragment contained a flavodoxin gene which encoded a protein of 160 amino acids that activated metronidazole and made the E. coli F19 mutant very sensitive to metronidazole. The flavodoxin and hydrogenase genes which are involved in electron transfer systems were linked on the 11.1-kb DNA fragment from C. acetobutylicum. Images PMID:1991710

  17. Cellulose fermentation by a coculture of a mesophilic cellulolytic Clostridium and Clostridium acetobutylicum

    SciTech Connect

    Fond, O.; Petitdemange, E.; Petitdemange, H.; Engasser, J.M.

    1983-01-01

    A coculture of a mesophilic cellulolytic Clostridium with Clostridium acetobutylicum can yield a direct conversion of cellulose into chemicals. In 13 days 30 g/l Solka Floc is degraded and fermented into 14 g/l butyric acid, 4 g/l acetic acid, 3 g/l ethanol, and 1 g/l butanol. A four times higher rate of cellulose hydrolysis than in pure culture of the cellulolytic Clostridium is thus obtained. Fed-batch fermentations of C. acetobutylicum at different glucose feeding rate show that solvents are only produced at a sufficient high rate of glucose supply to the medium. Acids are thus the main products of the coculture because of the limited rate of cellulolysis by the mesophilic strain. 7 references, 5 figures.

  18. Pervaporative butanol fermentation by Clostridium acetobutylicum B18

    SciTech Connect

    Geng, Q.; Park, C.H. . Dept. of Agricultural Engineering)

    1994-04-15

    Extractive acetone-butanol-ethanol (ABE) fermentation was carried out successfully using pervaporation and a low-acid-producing Clostridium acetobutylicum B18. A pervaporation module with 0.17 m[sup 2] of surface area was made of silicone membrane of 240 [mu]m thickness. Pervaporation experiments using make-up solutions showed that butanol and acetone fluxes increased linearly with their concentrations in the aqueous phase. Fickian diffusion coefficients were constants for fixed air flow rates, and increased at higher sweep air flow rates. During batch and fed-batch fermentation, pervaporation at an air flow rate of 8 L/min removed butanol and acetone efficiently. Butanol concentration was maintained below 4.5 g/L even though Clostridium acetobutylicum B18 produced butanol steadily. Pervaporation could not remove organic acids efficiently, but organic acids did not accumulate because strain B18 produced little organic acid and recycled added organic acids efficiently. With pervaporation, glucose consumption rate increased compared to without pervaporation, and up to 160 g/L of glucose was consumed during 80 h. Cell growth was not inhibited by possible salt accumulation or oxygen diffusion through the silicone tubing. The culture volume was maintained relatively constant during fed-batch operation because of an offsetting effect of water and product removal by pervaporation and addition of nutrient supplements.

  19. In situ hydrogen, acetone, butanol, ethanol and microdiesel production by Clostridium acetobutylicum ATCC 824 from oleaginous fungal biomass.

    PubMed

    Hassan, Elhagag Ahmed; Abd-Alla, Mohamed Hemida; Bagy, Magdy Mohamed Khalil; Morsy, Fatthy Mohamed

    2015-08-01

    An in situ batch fermentation technique was employed for biohydrogen, acetone, butanol, ethanol and microdiesel production from oleaginous fungal biomass using the anaerobic fermentative bacterium Clostridium acetobutylicum ATCC 824. Oleaginous fungal Cunninghamella echinulata biomass which has ability to accumulate up to 71% cellular lipid was used as the substrate carbon source. The maximum cumulative hydrogen by C. acetobutylicum ATCC 824 from crude C. echinulata biomass was 260 ml H2 l(-1), hydrogen production efficiency was 0.32 mol H2 mole(-1) glucose and the hydrogen production rate was 5.2 ml H2 h(-1). Subsequently, the produced acids (acetic and butyric acids) during acidogenesis phase are re-utilized by ABE-producing clostridia and converted into acetone, butanol, and ethanol. The total ABE produced by C. acetobutylicum ATCC 824 during batch fermentation was 3.6 g l(-1) from crude fungal biomass including acetone (1.05 g l(-1)), butanol (2.19 g l(-1)) and ethanol (0.36 g l(-1)). C. acetobutylicum ATCC 824 has ability to produce lipolytic enzymes with a specific activity 5.59 U/mg protein to hydrolyze ester containing substrates. The lipolytic potential of C. acetobutylicum ATCC 824 was used as a biocatalyst for a lipase transesterification process using the produced ethanol from ABE fermentation for microdiesel production. The fatty acid ethyl esters (microdiesel) generated from the lipase transesterification of crude C. echinulata dry mass was analyzed by GC/MS as 15.4% of total FAEEs. The gross energy content of biohydrogen, acetone, butanol, ethanol and biodiesel generated through C. acetobutylicum fermentation from crude C. echinulata dry mass was 3113.14 kJ mol(-1). These results suggest a possibility of integrating biohydrogen, acetone, butanol and ethanol production technology by C. acetobutylicum with microdiesel production from crude C. echinulata dry mass and therefore improve the feasibility and commercialization of bioenergy production.

  20. Systems-level metabolic flux profiling elucidates a complete, bifurcated tricarboxylic acid cycle in Clostridium acetobutylicum.

    PubMed

    Amador-Noguez, Daniel; Feng, Xiao-Jiang; Fan, Jing; Roquet, Nathaniel; Rabitz, Herschel; Rabinowitz, Joshua D

    2010-09-01

    Obligatory anaerobic bacteria are major contributors to the overall metabolism of soil and the human gut. The metabolic pathways of these bacteria remain, however, poorly understood. Using isotope tracers, mass spectrometry, and quantitative flux modeling, here we directly map the metabolic pathways of Clostridium acetobutylicum, a soil bacterium whose major fermentation products include the biofuels butanol and hydrogen. While genome annotation suggests the absence of most tricarboxylic acid (TCA) cycle enzymes, our results demonstrate that this bacterium has a complete, albeit bifurcated, TCA cycle; oxaloacetate flows to succinate both through citrate/alpha-ketoglutarate and via malate/fumarate. Our investigations also yielded insights into the pathways utilized for glucose catabolism and amino acid biosynthesis and revealed that the organism's one-carbon metabolism is distinct from that of model microbes, involving reversible pyruvate decarboxylation and the use of pyruvate as the one-carbon donor for biosynthetic reactions. This study represents the first in vivo characterization of the TCA cycle and central metabolism of C. acetobutylicum. Our results establish a role for the full TCA cycle in an obligatory anaerobic organism and demonstrate the importance of complementing genome annotation with isotope tracer studies for determining the metabolic pathways of diverse microbes.

  1. Spontaneous large-scale autolysis in Clostridium acetobutylicum contributes to generation of more spores

    PubMed Central

    Liu, Zhen; Qiao, Kai; Tian, Lei; Zhang, Quan; Liu, Zi-Yong; Li, Fu-Li

    2015-01-01

    Autolysis is a widespread phenomenon in bacteria. In batch fermentation of Clostridium acetobutylicum ATCC 824, there is a spontaneous large-scale autolysis phenomenon with significant decrease of cell density immediately after exponential phase. To unravel the role of autolysis, an autolysin-coding gene, CA_C0554, was disrupted by using ClosTron system to obtain the mutant C. acetobutylicum lyc::int(72). The lower final cell density and faster cell density decrease rate of C. acetobutylicum ATCC 824 than those of C. acetobutylicum lyc::int(72) indicates that CA_C0554 was an important but not the sole autolysin-coding gene responding for the large-scale autolysis. Similar glucose utilization and solvents production but obvious lower cell density of C. acetobutylicum ATCC 824 comparing to C. acetobutylicum lyc::int(72) suggests that lysed C. acetobutylicum ATCC 824 cells were metabolic inactive. On the contrary, the spore density of C. acetobutylicum ATCC 824 is 26.1% higher than that of C. acetobutylicum lyc::int(72) in the final culture broth of batch fermentation. We speculated that spontaneous autolysis of metabolic-inactive cells provided nutrients for the sporulating cells. The present study suggests that one important biological role of spontaneous large-scale autolysis in C. acetobutylicum ATCC 824 batch fermentation is contributing to generation of more spores during sporulation. PMID:26441884

  2. Systems Biology of Clostridium Acetobutylicum: Sugar Metabolism and TNT Reduction

    NASA Astrophysics Data System (ADS)

    Hurley, Margaret; Sund, Christian; Servinsky, Matthew

    2010-03-01

    Rapid advancements in biotechnology are expected to impact multiple areas of interest to the Army, including decontamination, degradation of toxic chemicals and biofuels. This project is a joint experimental/computational effort to map out the metabolic pathways in Clostridium acetobutylicum, and use this information to develop a systems biology model of this system. This organism has been chosen specifically due to the fact that it has potential application to both biofuel production and nitroaromatic degradation. It is hoped that a systems biology model may provide key information to enhance both of these processes. Details will be presented of a first-generation model of central carbon metabolism in C. Acet., developed upon gene expression data accumulated from bacteria grown on different carbohydrate sources. Additional work will discuss the effect of TNT exposure and potential relevant enhancements of the model.

  3. Metabolic flexibility of a butyrate pathway mutant of Clostridium acetobutylicum.

    PubMed

    Yoo, Minyeong; Croux, Christian; Meynial-Salles, Isabelle; Soucaille, Philippe

    2017-01-31

    Clostridium acetobutylicum possesses two homologous buk genes, buk (or buk1) and buk2, which encode butyrate kinases involved in the last step of butyrate formation. To investigate the contribution of buk in detail, an in-frame deletion mutant was constructed. However, in all the Δbuk mutants obtained, partial deletions of the upstream ptb gene were observed, and low phosphotransbutyrylase and butyrate kinase activities were measured. This demonstrates that i) buk (CA_C3075) is the key butyrate kinase-encoding gene and that buk2 (CA_C1660) that is poorly transcribed only plays a minor role; and ii) strongly suggests that a Δbuk mutant is not viable if the ptb gene is not also inactivated, probably due to the accumulation of butyryl-phosphate, which might be toxic for the cell. One of the ΔbukΔptb mutants was subjected to quantitative transcriptomic (mRNA molecules/cell) and fluxomic analyses in acidogenic, solventogenic and alcohologenic chemostat cultures. In addition to the low butyrate production, drastic changes in metabolic fluxes were also observed for the mutant: i) under acidogenic conditions, the primary metabolite was butanol and a new metabolite, 2-hydroxy-valerate, was produced ii) under solventogenesis, 58% increased butanol production was obtained compared to the control strain under the same conditions, and a very high yield of butanol formation (0.3gg(-1)) was reached; and iii) under alcohologenesis, the major product was lactate. Furthermore, at the transcriptional level, adhE2, which encodes an aldehyde/alcohol dehydrogenase and is known to be a gene specifically expressed in alcohologenesis, was surprisingly highly expressed in all metabolic states in the mutant. The results presented here not only support the key roles of buk and ptb in butyrate formation but also highlight the metabolic flexibility of C. acetobutylicum in response to genetic alteration of its primary metabolism.

  4. Capturing the response of Clostridium acetobutylicum to chemical stressors using a regulated genome-scale metabolic model

    DOE PAGES

    Dash, Satyakam; Mueller, Thomas J.; Venkataramanan, Keerthi P.; ...

    2014-10-14

    Clostridia are anaerobic Gram-positive Firmicutes containing broad and flexible systems for substrate utilization, which have been used successfully to produce a range of industrial compounds. Clostridium acetobutylicum has been used to produce butanol on an industrial scale through acetone-butanol-ethanol (ABE) fermentation. A genome-scale metabolic (GSM) model is a powerful tool for understanding the metabolic capacities of an organism and developing metabolic engineering strategies for strain development. The integration of stress related specific transcriptomics information with the GSM model provides opportunities for elucidating the focal points of regulation.

  5. Enhancing Butanol Production under the Stress Environments of Co-Culturing Clostridium acetobutylicum/Saccharomyces cerevisiae Integrated with Exogenous Butyrate Addition.

    PubMed

    Luo, Hongzhen; Ge, Laibing; Zhang, Jingshu; Zhao, Yanli; Ding, Jian; Li, Zhigang; He, Zhenni; Chen, Rui; Shi, Zhongping

    2015-01-01

    In this study, an efficient acetone-butanol-ethanol (ABE) fermentation strategy integrating Clostridium acetobutylicum/Saccharomyces cerevisiae co-culturing system with exogenous butyrate addition, was proposed and experimentally conducted. In solventogenic phase, by adding 0.2 g-DCW/L-broth viable S. cerevisiae cells and 4.0 g/L-broth concentrated butyrate solution into C. acetobutylicum culture broth, final butanol concentration and butanol/acetone ratio in a 7 L anaerobic fermentor reached the highest levels of 15.74 g/L and 2.83 respectively, with the increments of 35% and 43% as compared with those of control. Theoretical and experimental analysis revealed that, the proposed strategy could, 1) extensively induce secretion of amino acids particularly lysine, which are favorable for both C. acetobutylicum survival and butanol synthesis under high butanol concentration environment; 2) enhance the utilization ability of C. acetobutylicum on glucose and over-produce intracellular NADH for butanol synthesis in C. acetobutylicum metabolism simultaneously; 3) direct most of extra consumed glucose into butanol synthesis route. The synergetic actions of effective amino acids assimilation, high rates of substrate consumption and NADH regeneration yielded highest butanol concentration and butanol ratio in C. acetobutylicum under this stress environment. The proposed method supplies an alternative way to improve ABE fermentation performance by traditional fermentation technology.

  6. Enhancing Butanol Production under the Stress Environments of Co-Culturing Clostridium acetobutylicum/Saccharomyces cerevisiae Integrated with Exogenous Butyrate Addition

    PubMed Central

    Luo, Hongzhen; Ge, Laibing; Zhang, Jingshu; Zhao, Yanli; Ding, Jian; Li, Zhigang; He, Zhenni; Chen, Rui; Shi, Zhongping

    2015-01-01

    In this study, an efficient acetone-butanol-ethanol (ABE) fermentation strategy integrating Clostridium acetobutylicum/Saccharomyces cerevisiae co-culturing system with exogenous butyrate addition, was proposed and experimentally conducted. In solventogenic phase, by adding 0.2 g-DCW/L-broth viable S. cerevisiae cells and 4.0 g/L-broth concentrated butyrate solution into C. acetobutylicum culture broth, final butanol concentration and butanol/acetone ratio in a 7 L anaerobic fermentor reached the highest levels of 15.74 g/L and 2.83 respectively, with the increments of 35% and 43% as compared with those of control. Theoretical and experimental analysis revealed that, the proposed strategy could, 1) extensively induce secretion of amino acids particularly lysine, which are favorable for both C. acetobutylicum survival and butanol synthesis under high butanol concentration environment; 2) enhance the utilization ability of C. acetobutylicum on glucose and over-produce intracellular NADH for butanol synthesis in C. acetobutylicum metabolism simultaneously; 3) direct most of extra consumed glucose into butanol synthesis route. The synergetic actions of effective amino acids assimilation, high rates of substrate consumption and NADH regeneration yielded highest butanol concentration and butanol ratio in C. acetobutylicum under this stress environment. The proposed method supplies an alternative way to improve ABE fermentation performance by traditional fermentation technology. PMID:26489085

  7. Microbiological production of acetone-butanol by Clostridium acetobutylicum.

    PubMed

    Abou-Zeid, A A; Fouad, M; Yassein, M

    1978-01-01

    Trials succeeded in raising the efficiencies of the fermentation medium, used in the fermentative production of acetone-butanol by Clostridium acetobutylicum. Egyptian black strap molasses (50.0% sugars) was suitable as carbon source in the fermentation medium, and (NH4)2SO4 was utilized with great success as inorganic nitrogen source. 140.0 g/l black strap molasses (about 7.0% sugars) and 3.0 g/l (NH4)2SO4 were the optimum concentrations for obtaining good yields of acetone and butanol. Molasses and (NH4)2SO4 were preferred because they are cheaper than the other carbon and organic nitrogen sources, used in the fermentative production of acetone-butanol. The percentage increase of the total solvents produced in the fermentation (production medium) was increased by 64.0. The slop (by-product of the acetone-butanol fermentation after distillation) was re-used in the fermentation medium as organic nitrogen source and supported the microorganisms for a good production of acetone and butanol, while when stillage was used in the production medium, the total solvents output was less than that produced in the medium containing slop.

  8. Sequences affecting the regulation of solvent production in Clostridium acetobutylicum.

    PubMed

    Scotcher, Miles C; Huang, Ke-xue; Harrison, Mary L; Rudolph, Frederick B; Bennett, George N

    2003-07-01

    The high solvent phenotype of Clostridium acetobutylicum mutants B and H was complemented by the introduction of a plasmid that contains either an intact or partially-deleted copy of solR, restoring acetone and butanol production to wild-type levels. This demonstrates that the solR open reading frame on pSOLThi is not required to restore solvent levels. The promoter region upstream of alcohol dehydrogense E (adhE) was examined in efforts to identify sites that play major roles in the control of expression. A series of adhE promoter fragments was constructed and the expression of each in acid- and solvent-phases of growth was analyzed using a chloramphenicol acetyl-transferase reporter system. Our results show that a region beyond the 0A box is needed for full induction of the promoter. Additionally, we show that the presence of sequences around a possible processing site designated S2 may have a negative role in the regulation of adhE expression.

  9. Control of butanol formation in Clostridium acetobutylicum by transcriptional activation.

    PubMed

    Thormann, Kai; Feustel, Lothar; Lorenz, Karin; Nakotte, Stephan; Dürre, Peter

    2002-04-01

    The sol operon of Clostridium acetobutylicum is the essential transcription unit for formation of the solvents butanol and acetone. The recent proposal that transcriptional regulation of this operon is controlled by the repressor Orf5/SolR (R. V. Nair, E. M. Green, D. E. Watson, G. N. Bennett, and E. T. Papoutsakis, J. Bacteriol. 181:319-330, 1999) was found to be incorrect. Instead, regulation depends on activation, most probably by the multivalent transcription factor Spo0A. The operon is transcribed from a single promoter. A second signal identified in primer extension studies results from mRNA processing and can be observed only in the natural host, not in a heterologous host. The first structural gene in the operon (adhE, encoding a bifunctional butyraldehyde/butanol dehydrogenase) is translated into two different proteins, the mature AdhE enzyme and the separate butanol dehydrogenase domain. The promoter of the sol operon is preceded by three imperfect repeats and a putative Spo0A-binding motif, which partially overlaps with repeat 3 (R3). Reporter gene analysis performed with the lacZ gene of Thermoanaerobacterium thermosulfurigenes and targeted mutations of the regulatory region revealed that the putative Spo0A-binding motif, R3, and R1 are essential for control. The data obtained also indicate that an additional activator protein is involved.

  10. Orf5/SolR: a transcriptional repressor of the sol operon of Clostridium acetobutylicum?

    PubMed

    Thormann, K; Dürre, P

    2001-11-01

    The gene of Orf5 (SolR) of Clostridium acetobutylicum DSM 792 was subcloned and overexpressed in Escherichia coli. The protein was purified with Ni-NTA agarose and used for DNA binding assays. No DNA binding of Orf5 to regions upstream of the sol operon from C. acetobutylicum was observed. Overexpression of Orf5 in C. acetobutylicum led to a change in the organism's pattern of glycosylated exoproteins. The Orf5 protein was localized in the cell membrane fraction and to a small extent in the supernatant medium. Based on these results Orf5 (SolR) appears not to act as a transcriptional repressor in C. acetobutylicum, but instead may be an enzyme involved in glycosylation or deglycosylation.

  11. Predictive modeling in Clostridium acetobutylicum fermentations employing Raman spectroscopy and multivariate data analysis for real-time culture monitoring

    NASA Astrophysics Data System (ADS)

    Zu, Theresah N. K.; Liu, Sanchao; Germane, Katherine L.; Servinsky, Matthew D.; Gerlach, Elliot S.; Mackie, David M.; Sund, Christian J.

    2016-05-01

    The coupling of optical fibers with Raman instrumentation has proven to be effective for real-time monitoring of chemical reactions and fermentations when combined with multivariate statistical data analysis. Raman spectroscopy is relatively fast, with little interference from the water peak present in fermentation media. Medical research has explored this technique for analysis of mammalian cultures for potential diagnosis of some cancers. Other organisms studied via this route include Escherichia coli, Saccharomyces cerevisiae, and some Bacillus sp., though very little work has been performed on Clostridium acetobutylicum cultures. C. acetobutylicum is a gram-positive anaerobic bacterium, which is highly sought after due to its ability to use a broad spectrum of substrates and produce useful byproducts through the well-known Acetone-Butanol-Ethanol (ABE) fermentation. In this work, real-time Raman data was acquired from C. acetobutylicum cultures grown on glucose. Samples were collected concurrently for comparative off-line product analysis. Partial-least squares (PLS) models were built both for agitated cultures and for static cultures from both datasets. Media components and metabolites monitored include glucose, butyric acid, acetic acid, and butanol. Models were cross-validated with independent datasets. Experiments with agitation were more favorable for modeling with goodness of fit (QY) values of 0.99 and goodness of prediction (Q2Y) values of 0.98. Static experiments did not model as well as agitated experiments. Raman results showed the static experiments were chaotic, especially during and shortly after manual sampling.

  12. Direct selection of Clostridium acetobutylicum fermentation mutants by a proton suicide method

    SciTech Connect

    Cueto, P.H.; Mendez, B.S. )

    1990-02-01

    Clostridium acetobutylicum ATCC 10132 mutants altered in acetic acid synthesis or in the shift to solventogenesis were directly selected by a proton suicide method after mutagenic treatment, by using bromide and bromate as selective agents. The mutants were characterized according to their solvent and acid production. On the selection plates they differed in colony phenotype from the parent strain.

  13. Discovery of External Modulators of the Fe-Fe Hydrogenase Enzyme in Clostridium acetobutylicum

    DTIC Science & Technology

    2015-02-01

    ARL-TR-7189 ● FEB 2015 US Army Research Laboratory Discovery of External Modulators of the Fe-Fe Hydrogenase...ARL-TR-7189 ● FEB 2015 US Army Research Laboratory Discovery of External Modulators of the Fe-Fe Hydrogenase Enzyme in Clostridium...October 2014 4. TITLE AND SUBTITLE Discovery of External Modulators of the Fe-Fe Hydrogenase Enzyme in Clostridium acetobutylicum 5a. CONTRACT

  14. Production of 1,3-propanediol from glycerol by Clostridium acetobutylicum and other Clostridium species

    SciTech Connect

    Forsberg, C.W.

    1987-04-01

    Glycerol was fermented with the production of 1,3-propanediol as the major fermentation product by four strains of Clostridium acetobutylicum, six of C. butylicum, two of C. beijerinckii, one of C. kainantoi, and three of C. butylicum. 1,3-Propanediol was identified by its retention times in gas chromatography and high-pressure liquid chromatography and by its mass spectrum. During growth of C. butylicum B593 in a chemostat culture at pH 6.5, 61% of the glycerol fermented was converted to 1,3-propanediol. When the pH was decreased to 4.9, growth and 1,3-propanediol production were substantially reduced.

  15. Regulation and localization of amylolytic enzymes in Clostridium acetobutylicum ATCC 824

    SciTech Connect

    Annous, B.A.; Blaschek, H.P. )

    1990-08-01

    Amylolytic activity was primarily cell associated when Clostridium acetobutylicum was grown on glucose or maltose and primarily extracellular when grown on dextrin or starch. Total amylolytic activity decreased with increasing glucose concentration. When this microorganism was grown in P2 medium containing starch, the intracellular amylolytic activity was 90% membrane bound and 10% cytoplasmic in nature. The addition of 1% glucose to 2% starch-based P2 medium at different stages of growth indicated that this carbohydrate repressed the synthesis of C. acetobutylicum amylolytic enzymes at the level of transcription.

  16. Cap0037, a Novel Global Regulator of Clostridium acetobutylicum Metabolism

    PubMed Central

    Nguyen, Ngoc-Phuong-Thao; Linder, Sonja; Flitsch, Stefanie K.; Schiel-Bengelsdorf, Bettina; Dürre, Peter

    2016-01-01

    ABSTRACT An operon comprising two genes, CA_P0037 and CA_P0036, that encode proteins of unknown function that were previously shown to be highly expressed in acidogenic cells and repressed in solventogenic and alcohologenic cells is located on the pSOL1 megaplasmid of Clostridium acetobutylicum upstream of adhE2. A CA_P0037::int (189/190s) mutant in which an intron was inserted at position 189/190 in the sense strand of CA_P0037 was successfully generated by the Targetron technique. The resultant mutant showed significantly different metabolic flux patterns in acidogenic (producing mainly lactate, butyrate, and butanol) and alcohologenic (producing mainly butyrate, acetate, and lactate) chemostat cultures but not in solventogenic or batch cultures. Transcriptomic investigation of the CA_P0037::int (189/190s) mutant showed that inactivation of CA_P0037 significantly affected the expression of more than 258 genes under acidogenic conditions. Surprisingly, genes belonging to the Fur regulon, involved in iron transport (CA_C1029-CA_C1032), or coding for the main flavodoxin (CA_C0587) were the most significantly expressed genes under all conditions, whereas fur (coding for the ferric uptake regulator) gene expression remained unchanged. Furthermore, most of the genes of the Rex regulon, such as the adhE2 and ldhA genes, and of the PerR regulon, such as rbr3A-rbr3B and dfx, were overexpressed in the mutant. In addition, the whole CA_P0037-CA_P0036 operon was highly expressed under all conditions in the CA_P0037::int (189/190s) mutant, suggesting a self-regulated expression mechanism. Cap0037 was shown to bind to the CA_P0037-CA_P0036 operon, sol operon, and adc promoters, and the binding sites were determined by DNA footprinting. Finally, a putative Cap0037 regulon was generated using a bioinformatic approach. PMID:27703070

  17. 13C metabolic flux analysis in Clostridium acetobutylicum during growth on L-arabinose

    NASA Astrophysics Data System (ADS)

    Hurley, Margaret; Sund, Christian; Liu, Sanchao; Germane, Katherine; Servinsky, Matthew; Gerlach, Elliot

    2015-03-01

    Clostridium acetobutylicum's metabolic pathways have been studied for decades due to its metabolic diversity and industrial value, yet many details of its metabolism are continuing to emerge. To elucidate the role of xylulose-5-P/fructose-6-P phosphoketolase (XFP), and the recently discovered Pentose Phosphate Pathway (PKP) in C. acetobutylicum, experimental and computational metabolic isotope analysis was performed under growth on glucose, xylose, and arabinose. Results indicate that PKP utilization increased with increasing xylose concentration and this trend was further pronounced during growth on arabinose. This was confirmed by mutation of the gene encoding XFP, which almost completely abolished flux through the PKP during growth on arabinose and resulted in decreased acetate:butyrate ratios. We discuss these experimental and computational results here, and the implications for our understanding of sugar metabolism in C. acetobutylicum.

  18. Biological butanol production from microalgae-based biodiesel residues by Clostridium acetobutylicum.

    PubMed

    Cheng, Hai-Hsuan; Whang, Liang-Ming; Chan, Kun-Chi; Chung, Man-Chien; Wu, Shu-Hsien; Liu, Cheng-Pin; Tien, Shih-Yuan; Chen, Shan-Yuan; Chang, Jo-Shu; Lee, Wen-Jhy

    2015-05-01

    This study conducted batch experiments to evaluate the potential of butanol production from microalgae biodiesel residues by Clostridium acetobutylicum. The results indicated that with 90 g/L of glucose as the sole substrate the highest butanol yield of 0.2 g/g-glucose was found, but the addition of butyrate significantly enhanced the butanol yield. The highest butanol yield of 0.4 g/g-glucose was found with 60 g/L of glucose and 18 g/L of butyrate. Using microalgae biodiesel residues as substrate, C. acetobutylicum produced 3.86 g/L of butanol and achieved butanol yield of 0.13 g/g-carbohydrate via ABE fermentation, but the results indicated that approximately one third of carbohydrate was not utilized by C. acetobutylicum. Biological butanol production from microalgae biodiesel residues can be possible, but further research on fermentation strategies are required to improve production yield.

  19. Comparative shotgun proteomic analysis of Clostridium acetobutylicum from butanol fermentation using glucose and xylose

    PubMed Central

    2011-01-01

    Background Butanol is a second generation biofuel produced by Clostridium acetobutylicum through acetone-butanol-ethanol (ABE) fermentation process. Shotgun proteomics provides a direct approach to study the whole proteome of an organism in depth. This paper focuses on shotgun proteomic profiling of C. acetobutylicum from ABE fermentation using glucose and xylose to understand the functional mechanisms of C. acetobutylicum proteins involved in butanol production. Results We identified 894 different proteins in C. acetobutylicum from ABE fermentation process by two dimensional - liquid chromatography - tandem mass spectrometry (2D-LC-MS/MS) method. This includes 717 proteins from glucose and 826 proteins from the xylose substrate. A total of 649 proteins were found to be common and 22 significantly differentially expressed proteins were identified between glucose and xylose substrates. Conclusion Our results demonstrate that flagellar proteins are highly up-regulated with glucose compared to xylose substrate during ABE fermentation. Chemotactic activity was also found to be lost with the xylose substrate due to the absence of CheW and CheV proteins. This is the first report on the shotgun proteomic analysis of C. acetobutylicum ATCC 824 in ABE fermentation between glucose and xylose substrate from a single time data point and the number of proteins identified here is more than any other study performed on this organism up to this report. PMID:22008648

  20. Comparative shotgun proteomic analysis of Clostridium acetobutylicum from butanol fermentation using glucose and xylose

    SciTech Connect

    Sivagnanam, Kumaran; Raghavan, Vijaya G. S.; Shah, Manesh B; Hettich, Robert {Bob} L; Verberkmoes, Nathan C; Lefsrud, Mark G

    2011-01-01

    Background: Butanol is a second generation biofuel produced by Clostridium acetobutylicum through acetonebutanol- ethanol (ABE) fermentation process. Shotgun proteomics provides a direct approach to study the whole proteome of an organism in depth. This paper focuses on shotgun proteomic profiling of C. acetobutylicum from ABE fermentation using glucose and xylose to understand the functional mechanisms of C. acetobutylicum proteins involved in butanol production. Results: We identified 894 different proteins in C. acetobutylicum from ABE fermentation process by two dimensional - liquid chromatography - tandem mass spectrometry (2D-LC-MS/MS) method. This includes 717 proteins from glucose and 826 proteins from the xylose substrate. A total of 649 proteins were found to be common and 22 significantly differentially expressed proteins were identified between glucose and xylose substrates. Conclusion: Our results demonstrate that flagellar proteins are highly up-regulated with glucose compared to xylose substrate during ABE fermentation. Chemotactic activity was also found to be lost with the xylose substrate due to the absence of CheW and CheV proteins. This is the first report on the shotgun proteomic analysis of C. acetobutylicum ATCC 824 in ABE fermentation between glucose and xylose substrate from a single time data point and the number of proteins identified here is more than any other study performed on this organism up to this report.

  1. Control of Carbon and Electron Flow in Clostridium acetobutylicum Fermentations: Utilization of Carbon Monoxide to Inhibit Hydrogen Production and to Enhance Butanol Yields

    PubMed Central

    Kim, Byung Hong; Bellows, Para; Datta, Rathin; Zeikus, J. G.

    1984-01-01

    Extracts prepared from non-solvent-producing cells of Clostridium acetobutylicum contained methyl viologen-linked hydrogenase activity (20 U/mg of protein at 37°C) but did not display carbon monoxide dehydrogenase activity. CO addition readily inhibited the hydrogenase activity of cell extracts or of viable metabolizing cells. Increasing the partial pressure of CO (2 to 10%) in unshaken anaerobic culture tube headspaces significantly inhibited (90% inhibition at 10% CO) both growth and hydrogen production by C. acetobutylicum. Growth was not sensitive to low partial pressures of CO (i.e., up to 15%) in pH-controlled fermentors (pH 4.5) that were continuously gassed and mixed. CO addition dramatically altered the glucose fermentation balance of C. acetobutylicum by diverting carbon and electrons away from H2, CO2, acetate, and butyrate production and towards production of ethanol and butanol. The butanol concentration was increased from 65 to 106 mM and the butanol productivity (i.e., the ratio of butanol produced/total acids and solvents produced) was increased by 31% when glucose fermentations maintained at pH 4.5 were continuously gassed with 85% N2-15% CO versus N2 alone. The results are discussed in terms of metabolic regulation of C. acetobutylicum saccharide fermentations to achieve maximal butanol or solvent yield. PMID:16346643

  2. Capturing the response of Clostridium acetobutylicum to chemical stressors using a regulated genome-scale metabolic model

    SciTech Connect

    Dash, Satyakam; Mueller, Thomas J.; Venkataramanan, Keerthi P.; Papoutsakis, Eleftherios T.; Maranas, Costas D.

    2014-10-14

    Clostridia are anaerobic Gram-positive Firmicutes containing broad and flexible systems for substrate utilization, which have been used successfully to produce a range of industrial compounds. Clostridium acetobutylicum has been used to produce butanol on an industrial scale through acetone-butanol-ethanol (ABE) fermentation. A genome-scale metabolic (GSM) model is a powerful tool for understanding the metabolic capacities of an organism and developing metabolic engineering strategies for strain development. The integration of stress related specific transcriptomics information with the GSM model provides opportunities for elucidating the focal points of regulation.

  3. Phosphoketolase flux in Clostridium acetobutylicum during growth on L-arabinose.

    PubMed

    Sund, Christian J; Liu, Sanchao; Germane, Katherine L; Servinsky, Matthew D; Gerlach, Elliot S; Hurley, Margaret M

    2015-02-01

    Clostridium acetobutylicum's metabolic pathways have been studied for decades due to its metabolic diversity and industrial value, yet many details of its metabolism continue to emerge. The flux through the recently discovered pentose phosphoketolase pathway (PKP) in C. acetobutylicum has been determined for growth on xylose but transcriptional analysis indicated the pathway may have a greater contribution to arabinose metabolism. To elucidate the role of xylulose-5-phosphate/fructose-6-phosphate phosphoketolase (XFP), and the PKP in C. acetobutylicum, experimental and computational metabolic isotope analyses were performed under growth conditions of glucose or varying concentrations of xylose and arabinose. A positional bias in labelling between carbons 2 and 4 of butyrate was found and posited to be due to an enzyme isotope effect of the thiolase enzyme. A correction for the positional bias was applied, which resulted in reduction of residual error. Comparisons between model solutions with low residual error indicated flux through each of the two XFP reactions was variable, while the combined flux of the reactions remained relatively constant. PKP utilization increased with increasing xylose concentration and this trend was further pronounced during growth on arabinose. Mutation of the gene encoding XFP almost completely abolished flux through the PKP during growth on arabinose and resulted in decreased acetate/butyrate ratios. Greater flux through the PKP during growth on arabinose when compared with xylose indicated the pathway's primary role in C. acetobutylicum is arabinose metabolism.

  4. Effects of nutritional enrichment on the production of acetone-butanol-ethanol (ABE) by Clostridium acetobutylicum.

    PubMed

    Choi, Sung Jun; Lee, Joungmin; Jang, Yu-Sin; Park, Jin Hwan; Lee, Sang Yup; Kim, In Ho

    2012-12-01

    Clostridium acetobutylicum is an industrially important organism that produces acetone-butanol-ethanol (ABE). The main objective of this study was to characterize the effects of increased cell density on the production of ABE during the phase transition from acidogenesis to solventogenesis in C. acetobutylicum. The increased ABE productivity of C. acetobutylicum was obtained by increasing the cell density using a newly designed medium (designated C. a cetobutylicum medium 1; CAM1). The maximum OD(600) value of C. acetobutylicum ATCC 824 strain obtained with CAM1 was 19.7, which is 1.8 times higher than that obtained with clostridial growth medium (CGM). The overall ABE productivity obtained in the CAM1-fermetation of the ATCC 824 strain was 0.83 g/L/h, which is 1.5 times higher than that (0.55 g/L/h) obtained with CGM. However, the increased productivity obtained with CAM1 did not result in an increase in the final ABE titer, because phase transition occurred at a high titer of acids.

  5. Mechanisms of microbial oil recovery by Clostridium acetobutylicum and Bacillus strain JF-2

    SciTech Connect

    Marsh, T.L.; Zhang, X.; Knapp, R.M.; McInerney, M.J.; Sharma, P.K.; Jackson, B.E.

    1995-12-31

    Core displacement experiments at elevated pressures were conducted to determine whether microbial processes are effective under conditions that simulate those found in an actual oil reservoir. The in-situ growth of Clostridium acetobutylicum and Bacillus strain JF-2 resulted in the recovery of residual oil. About 21 and 23% of the residual oil was recovered by C. acetobutylicum and Bacillus strain JF-2, respectively. Flooding cores with cell-free culture fluids of C. acetobutylicum with and without the addition of 50 mM acetone and 100 mM butanol did not result in the recovery of residual oil. Mathematical simulations showed that the amount of gas produced by the clostridial fermentation was not showed that the amount of gas produced by the clostridial fermentation was not sufficient to recover residual oil. Oil recovery by Bacillus strain JF-2 was highly correlated to surfactant production. A biosurfactant-deficient mutant of strain JF-2 was not capable of recovering residual oil. These data show that surfactant production is an important mechanism for microbially enhanced oil recovery. The mechanism for oil recovery by C. acetobutylicum is not understood at this time, but the production of acids, solvents, or gases alone cannot explain the observed increases in oil recovery by this organism.

  6. Metabolic Engineering of Clostridium acetobutylicum ATCC 824 for Isopropanol-Butanol-Ethanol Fermentation

    PubMed Central

    Lee, Joungmin; Jang, Yu-Sin; Choi, Sung Jun; Im, Jung Ae; Song, Hyohak; Cho, Jung Hee; Seung, Do Young; Papoutsakis, E. Terry; Bennett, George N.

    2012-01-01

    Clostridium acetobutylicum naturally produces acetone as well as butanol and ethanol. Since acetone cannot be used as a biofuel, its production needs to be minimized or suppressed by cell or bioreactor engineering. Thus, there have been attempts to disrupt or inactivate the acetone formation pathway. Here we present another approach, namely, converting acetone to isopropanol by metabolic engineering. Since isopropanol can be used as a fuel additive, the mixture of isopropanol, butanol, and ethanol (IBE) produced by engineered C. acetobutylicum can be directly used as a biofuel. IBE production is achieved by the expression of a primary/secondary alcohol dehydrogenase gene from Clostridium beijerinckii NRRL B-593 (i.e., adhB-593) in C. acetobutylicum ATCC 824. To increase the total alcohol titer, a synthetic acetone operon (act operon; adc-ctfA-ctfB) was constructed and expressed to increase the flux toward isopropanol formation. When this engineering strategy was applied to the PJC4BK strain lacking in the buk gene (encoding butyrate kinase), a significantly higher titer and yield of IBE could be achieved. The resulting PJC4BK(pIPA3-Cm2) strain produced 20.4 g/liter of total alcohol. Fermentation could be prolonged by in situ removal of solvents by gas stripping, and 35.6 g/liter of the IBE mixture could be produced in 45 h. PMID:22210214

  7. Efficient gene knockdown in Clostridium acetobutylicum by synthetic small regulatory RNAs.

    PubMed

    Cho, Changhee; Lee, Sang Yup

    2017-02-01

    Clostridium is considered a promising microbial host for the production of valuable industrial chemicals. However, Clostridium is notorious for the difficulty of genetic manipulations, and consequently metabolic engineering. Thus, much effort has been exerted to develop novel tools for genetic and metabolic engineering of Clostridium strains. Here, we report the development of a synthetic small regulatory RNA (sRNA)-based system for controlled gene expression in Clostridium acetobutylicum, consisting of a target recognition site, MicC sRNA scaffold, and an RNA chaperone Hfq. To examine the functional operation of sRNA system in C. acetobutylicum, expression control was first examined with the Evoglow fluorescent protein as a model protein. Initially, a C. acetobutylicum protein annotated as Hfq was combined with the synthetic sRNA based on the Escherichia coli MicC scaffold to knockdown Evoglow expression. However, C. acetobutylicum Hfq did not bind to E. coli MicC, while MicC scaffold-based synthetic sRNA itself was able to knockdown the expression of Evoglow. When E. coli hfq gene was introduced, the knockdown efficiency assessed by measuring fluorescence intensity, could be much enhanced. Then, this E. coli MicC scaffold-Hfq system was used to knock down adhE1 gene expression in C. acetobutylicum. Knocking down the adhE1 gene expression using the synthetic sRNA led to a 40% decrease in butanol production (2.5 g/L), compared to that (4.5 g/L) produced by the wild-type strain harboring an empty vector. The sRNA system was further extended to knock down the pta gene expression in the buk mutant C. acetobutylicum strain PJC4BK for enhanced butanol production. The PJC4BK (pPta-Hfq(Eco) ) strain, which has the pta gene expression knocked down, was able to produce 16.9 g/L of butanol, which is higher than that (14.9 g/L) produced by the PJC4BK strain, mainly due to reduced acetic acid production. Fed-batch culture of PJC4BK (pPta-Hfq(Eco) ) strain coupled with

  8. Biobutanol production by a new local isolate of Clostridium acetobutylicum YM1

    NASA Astrophysics Data System (ADS)

    Al-Shorgani, Najeeb Kaid; Tibin, El Mubarak; Kalil, Mohd Sahaid; Yusoff, Wan Mohtar Wan; Hamid, Aidil Abdul

    2014-09-01

    Increasing demand of energy and awareness about environmental pollution has led to increase interest in alternative, clean and renewable energy sources. Biobutanol is considered as the candidate liquid biofuel to replace gasoline. In this study, the capability of a newly isolated strain of local Clostridium acetobutylicum YM1 was tested to produce biobutanol in batch fermentation. Various culture conditions including glucose concentration, initial pH, incubation temperature and inoculum size were investigated for their effects on production of biobutanol using strain YM1. The results showed that the optimal biobutanol production was obtained at glucose concentration 50 g/L, initial pH 6.2, temperature 30°C and inoculum size 10%. These results show that C. acetobutylicum YM1 as a mesophilic bacterium is a potential candidate for biobutanol production.

  9. Parallel labeling experiments validate Clostridium acetobutylicum metabolic network model for (13)C metabolic flux analysis.

    PubMed

    Au, Jennifer; Choi, Jungik; Jones, Shawn W; Venkataramanan, Keerthi P; Antoniewicz, Maciek R

    2014-11-01

    In this work, we provide new insights into the metabolism of Clostridium acetobutylicum ATCC 824 obtained using a systematic approach for quantifying fluxes based on parallel labeling experiments and (13)C-metabolic flux analysis ((13)C-MFA). Here, cells were grown in parallel cultures with [1-(13)C]glucose and [U-(13)C]glucose as tracers and (13)C-MFA was used to quantify intracellular metabolic fluxes. Several metabolic network models were compared: an initial model based on current knowledge, and extended network models that included additional reactions that improved the fits of experimental data. While the initial network model did not produce a statistically acceptable fit of (13)C-labeling data, an extended network model with five additional reactions was able to fit all data with 292 redundant measurements. The model was subsequently trimmed to produce a minimal network model of C. acetobutylicum for (13)C-MFA, which could still reproduce all of the experimental data. The flux results provided valuable new insights into the metabolism of C. acetobutylicum. First, we found that TCA cycle was effectively incomplete, as there was no measurable flux between α-ketoglutarate and succinyl-CoA, succinate and fumarate, and malate and oxaloacetate. Second, an active pathway was identified from pyruvate to fumarate via aspartate. Third, we found that isoleucine was produced exclusively through the citramalate synthase pathway in C. acetobutylicum and that CAC3174 was likely responsible for citramalate synthase activity. These model predictions were confirmed in several follow-up tracer experiments. The validated metabolic network model established in this study can be used in future investigations for unbiased (13)C-flux measurements in C. acetobutylicum.

  10. Genome analysis of a hyper acetone-butanol-ethanol (ABE) producing Clostridium acetobutylicum BKM19.

    PubMed

    Cho, Changhee; Choe, Donghui; Jang, Yu-Sin; Kim, Kyung-Jin; Kim, Won Jun; Cho, Byung-Kwan; Papoutsakis, E Terry; Bennett, George N; Seung, Do Young; Lee, Sang Yup

    2017-02-01

    Previously the development of a hyper acetone-butanol-ethanol (ABE) producing Clostridium acetobutylicum BKM19 strain capable of producing 30.5% more total solvent by random mutagenesis of its parental strain PJC4BK, which is a buk mutant C. acetobutylicum ATCC 824 strain is reported. Here, BKM19 and PJC4BK strains are re-sequenced by a high-throughput sequencing technique to understand the mutations responsible for enhanced solvent production. In comparison with the C. acetobutylicum PJC4BK, 13 single nucleotide variants (SNVs), one deletion and one back mutation SNV are identified in the C. acetobutylicum BKM19 genome. Except for one SNV found in the megaplasmid, all mutations are found in the chromosome of BKM19. Among them, a mutation in the thlA gene encoding thiolase is further studied with respect to enzyme activity and butanol production. The mutant thiolase (thlA(V5A) ) is showed a 32% higher activity than that of the wild-type thiolase (thlA(WT) ). In batch fermentation, butanol production is increased by 26% and 23% when the thlA(V5A) gene is overexpressed in the wild-type C. acetobutylicum ATCC 824 and in its derivative, the thlA-knockdown TKW-A strain, respectively. Based on structural analysis, the mutation in thiolase does not have a direct effect on the regulatory determinant region (RDR). However, the mutation at the 5(th) residue seems to influence the stability of the RDR, and thus, increases the enzymatic activity and enhances solvent production in the BKM19 strain.

  11. Transformation of heat-treated Clostridium acetobutylicum protoplasts with pUB110 plasmid DNA

    SciTech Connect

    Lin, Y.L.; Blaschek, H.P.

    1984-10-01

    Heat treatment of Clostridium acetobutylicum SA-1 protoplasts at 55/sup 0/C for 15 min before transformation resulted in expression in this microorganism of the kanamycin resistance determinant associated with plasmid pUB110. No heat treatment, or heat treatment at 65 or 44/sup 0/C for various time intervals, resulted in no kanamycin resistance transformants being recovered on selective kanamycin-containing regeneration medium. DNase plate assay indicated that treatment at 55/sup 0/C for 15 min completely inactivated the DNase activity associated with SA-1 protoplasts. Treatment of protoplasts at 65 or 55/sup 0/C for various periods under simulated transformation conditions had an inhibitory effect, although prolonged treatment at 55 or 44/sup 0/C appeared to stimulate DNase activity. Inactivation of protoplast-associated DNase activity by heat treatment at 55/sup 0/C for 15 min correlated with successful expression of kanamycin resistance and suggests that an extremely active, heat-sensitive, protoplast-associated DNase may be a factor in the polyethylene glycol-induced transformation of C. acetobutylicum SA-1 protoplasts. Plasmid pUB110 DNA was isolated from C. acetobutylicum SA-1 kanamycin-resistant (Km/sup r/) transformant cultures by a modification of the procedure used for C. perfringens plasmids. Detection of pUB110 DNA was possible only when diethyl pyrocarbonate was incorporated into isolation protocols to inactivate DNase activity. Restriction studies further verified the presence of pUB110 DNA in C. acetobutylicum SA-1 Km/sup r/ transformants. 36 references, 4 figures, 1 table.

  12. Butanol production by immobilised Clostridium acetobutylicum in repeated batch, fed-batch, and continuous modes of fermentation.

    PubMed

    Dolejš, Igor; Krasňan, Vladimír; Stloukal, Radek; Rosenberg, Michal; Rebroš, Martin

    2014-10-01

    Clostridium acetobutylicum immobilised in polyvinylalcohol, lens-shaped hydrogel capsules (LentiKats(®)) was studied for production of butanol and other products of acetone-butanol-ethanol fermentation. After optimising the immobilisation protocol for anaerobic bacteria, continuous, repeated batch, and fed-batch fermentations in repeated batch mode were performed. Using glucose as a substrate, butanol productivity of 0.41 g/L/h and solvent productivity of 0.63 g/L/h were observed at a dilution rate of 0.05 h(-1) during continuous fermentation with a concentrated substrate (60 g/L). Through the process of repeated batch fermentation, the duration of fermentation was reduced from 27.8h (free-cell fermentation) to 3.3h (immobilised cells) with a solvent productivity of 0.77 g/L/h (butanol 0.57 g/L/h). The highest butanol and solvent productivities of 1.21 and 1.91 g/L/h were observed during fed-batch fermentation operated in repeated batch mode with yields of butanol (0.15 g/g) and solvents (0.24 g/g), respectively, produced per gram of glucose.

  13. Enhanced butanol production by Clostridium acetobutylicum NCIMB 13357 grown on date fruit as carbon source in P2 medium.

    PubMed

    Khamaiseh, Emran I; Abdul Hamid, Aidil; Abdeshahian, Peyman; Wan Yusoff, Wan Mohtar; Kalil, Mohd Sahaid

    2014-01-01

    The production of biobutanol was studied by the cultivation of Clostridium acetobutylicum NCIMB 13557 in P2 medium including date fruit as the sole substrate. The effect of P2 medium and the effect of different concentrations of date fruit ranging from 10 to 100 g/L on biobutanol production were investigated. Anaerobic batch culture was carried out at 35 °C incubation temperature and pH 7.0 ± 0.2 for 72 h. Experimental results showed that the lowest yield of biobutanol and acetone-butanol-ethanol (ABE) was 0.32 and 0.35 gram per gram of carbohydrate consumed (g/g), respectively, when an initial date fruit concentration of 10 g/L was utilized. At this fruit date concentration a biobutanol production value of 1.56 g/L was obtained. On the other hand, the maximum yield of biobutanol (0.48 g/g) and ABE (0.63 g/g) was produced at 50 g/L date fruit concentration with a biobutanol production value as high as 11 g/L. However, when a higher initial date fruit concentration was used, biobutanol and ABE production decreased to reach the yield of 0.22 g/g and 0.35 g/g, respectively, where 100 g/L date fruit was used. Similar results also revealed that 10.03 g/L biobutanol was produced using 100 g/L date fruit.

  14. Molecular characterization of an aldehyde/alcohol dehydrogenase gene from Clostridium acetobutylicum ATCC 824.

    PubMed Central

    Nair, R V; Bennett, G N; Papoutsakis, E T

    1994-01-01

    A gene (aad) coding for an aldehyde/alcohol dehydrogenase (AAD) was identified immediately upstream of the previously cloned ctfA (J. W. Cary, D. J. Petersen, E. T. Papoutsakis, and G. N. Bennett, Appl. Environ. Microbiol. 56:1576-1583, 1990) of Clostridium acetobutylicum ATCC 824 and sequenced. The 2,619-bp aad codes for a 96,517-Da protein. Primer extension analysis identified two transcriptional start sites 83 and 243 bp upstream of the aad start codon. The N-terminal section of AAD shows homology to aldehyde dehydrogenases of bacterial, fungal, mammalian, and plant origin, while the C-terminal section shows homology to alcohol dehydrogenases of bacterial (which includes three clostridial alcohol dehydrogenases) and yeast origin. AAD exhibits considerable amino acid homology (56% identity) over its entire sequence to the trifunctional protein encoded by adhE from Escherichia coli. Expression of aad from a plasmid in C. acetobutylicum showed that AAD, which appears as a approximately 96-kDa band in denaturing protein gels, provides elevated activities of NADH-dependent butanol dehydrogenase, NAD-dependent acetaldehyde dehydrogenase and butyraldehyde dehydrogenase, and a small increase in NADH-dependent ethanol dehydrogenase. A 957-bp open reading frame that could potentially encode a 36,704-Da protein was identified upstream of aad. Images PMID:8300540

  15. Novel substrate specificity of glutathione synthesis enzymes from Streptococcus agalactiae and Clostridium acetobutylicum

    SciTech Connect

    Kino, Kuniki . E-mail: kkino@waseda.jp; Kuratsu, Shoko; Noguchi, Atsushi; Kokubo, Masahiro; Nakazawa, Yuji; Arai, Toshinobu; Yagasaki, Makoto; Kirimura, Kohtaro

    2007-01-12

    Glutathione (GSH) is synthesized by {gamma}-glutamylcysteine synthetase ({gamma}-GCS) and glutathione synthetase (GS) in living organisms. Recently, bifunctional fusion protein, termed {gamma}-GCS-GS catalyzing both {gamma}-GCS and GS reactions from gram-positive firmicutes Streptococcus agalactiae, has been reported. We revealed that in the {gamma}-GCS activity, S. agalactiae {gamma}-GCS-GS had different substrate specificities from those of Escherichia coli {gamma}-GCS. Furthermore, S. agalactiae {gamma}-GCS-GS synthesized several kinds of {gamma}-glutamyltripeptide, {gamma}-Glu-X{sub aa}-Gly, from free three amino acids. In Clostridium acetobutylicum, the genes encoding {gamma}-GCS and putative GS were found to be immediately adjacent by BLAST search, and had amino acid sequence homology with S. agalactiae {gamma}-GCS-GS, respectively. We confirmed that the proteins expressed from each gene showed {gamma}-GCS and GS activity, respectively. C. acetobutylicum GS had broad substrate specificities and synthesized several kinds of {gamma}-glutamyltripeptide, {gamma}-Glu-Cys-X{sub aa}. Whereas the substrate specificities of {gamma}-GCS domain protein and GS domain protein of S. agalactiae {gamma}-GCS-GS were the same as those of S. agalactiae {gamma}-GCS-GS.

  16. Cell growth behaviors of Clostridium acetobutylicum in a pervaporation membrane bioreactor for butanol fermentation.

    PubMed

    Yao, Peina; Xiao, Zeyi; Chen, Chunyan; Li, Weijia; Deng, Qing

    2016-01-01

    Acetone-butanol-ethanol fermentation using Clostridium acetobutylicum was studied in the continuous and closed-circulating fermentation (CCCF) system. The experiment lasting for 192 H was carried out by integrating fermentation with in situ pervaporation. In the entire process, the cell growth profile took place in the following two phases: the logarithmic phase during early 28 H and the linear phase from 130 to 150 H. This was a unique characteristic compared with the curve of traditional fermentation, and the fitting equations of two growth phases were obtained by Origin software according to the kinetic model of cell growth. Besides, the kinetic parameters that include the butanol yield, maximum specific growth rate, average specific formation rate, and volumetric productivity of butanol were measured as 0.19 g g(-1) , 0.345 H(-1) , 0.134 H(-1) and 0.23 g L(-1)  H(-1) , respectively. The C. acetobutylicum in the CCCF system showed good adaptability and fermentation performance, and the prolonged fermentation period and high production were also the main advantages of CCCF technology.

  17. Arabinose is metabolized via a phosphoketolase pathway in Clostridium acetobutylicum ATCC 824.

    PubMed

    Servinsky, M D; Germane, K L; Liu, S; Kiel, J T; Clark, A M; Shankar, J; Sund, C J

    2012-12-01

    In this report, a novel zymogram assay and coupled phosphoketolase assay were employed to demonstrate that Clostridium acetobutylicum gene CAC1343 encodes a bi-functional xylulose-5-P/fructose-6-P phosphoketolase (XFP). The specific activity of purified recombinant XFP was 6.9 U/mg on xylulose-5-P and 21 U/mg on fructose-6-P, while the specific activity of XFP in concentrated C. acetobutylicum whole-cell extract was 0.094 and 0.52 U/mg, respectively. Analysis of crude cell extracts indicated that XFP activity was present in cells grown on arabinose but not glucose and quantitative PCR was used to show that CAC1343 mRNA expression was induced 185-fold during growth on arabinose when compared to growth on glucose. HPLC analysis of metabolites revealed that during growth on xylose and glucose more butyrate than acetate was formed with final acetate:butyrate ratios of 0.72 and 0.83, respectively. Growth on arabinose caused a metabolic shift to more oxidized products with a final acetate:butyrate ratio of 1.95. The shift towards more oxidized products is consistent with the presence of an XFP, suggesting that arabinose is metabolized via a phosphoketolase pathway while xylose is probably metabolized via the pentose phosphate pathway.

  18. Integrated, systems metabolic picture of acetone-butanol-ethanol fermentation by Clostridium acetobutylicum.

    PubMed

    Liao, Chen; Seo, Seung-Oh; Celik, Venhar; Liu, Huaiwei; Kong, Wentao; Wang, Yi; Blaschek, Hans; Jin, Yong-Su; Lu, Ting

    2015-07-07

    Microbial metabolism involves complex, system-level processes implemented via the orchestration of metabolic reactions, gene regulation, and environmental cues. One canonical example of such processes is acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum, during which cells convert carbon sources to organic acids that are later reassimilated to produce solvents as a strategy for cellular survival. The complexity and systems nature of the process have been largely underappreciated, rendering challenges in understanding and optimizing solvent production. Here, we present a system-level computational framework for ABE fermentation that combines metabolic reactions, gene regulation, and environmental cues. We developed the framework by decomposing the entire system into three modules, building each module separately, and then assembling them back into an integrated system. During the model construction, a bottom-up approach was used to link molecular events at the single-cell level into the events at the population level. The integrated model was able to successfully reproduce ABE fermentations of the WT C. acetobutylicum (ATCC 824), as well as its mutants, using data obtained from our own experiments and from literature. Furthermore, the model confers successful predictions of the fermentations with various network perturbations across metabolic, genetic, and environmental aspects. From foundation to applications, the framework advances our understanding of complex clostridial metabolism and physiology and also facilitates the development of systems engineering strategies for the production of advanced biofuels.

  19. Clostridium acetobutylicum Mutants That Produce Butyraldehyde and Altered Quantities of Solvents.

    PubMed

    Rogers, P; Palosaari, N

    1987-12-01

    Spontaneous mutants of Clostridium acetobutylicum NRRL B643 that were resistant to allyl alcohol (AA) were selected and characterized. These mutants contained 10- to 100-fold reduced activities of butanol and ethanol alcohol dehydrogenase. The AA mutants formed two groups and produced no ethanol. Type 1 AA mutants produced significant amounts of a new solvent, butyraldehyde, and contained normal levels of the coenzyme A-dependent butyraldehyde dehydrogenase (BAD). Type 2 AA mutants produced no significant butyraldehyde and lower levels of all solvents, and they contained 45- to 100-fold lower activity levels of BAD. Following ethyl methanesulfonate mutagenesis, low-acid-producing (Acid) mutants were selected and characterized as superinduced solvent producers, yielding more than 99% of theoretical glucose carbon as solvents and only small amounts of acetate and butyrate. Following ethyl methanesulfonate mutagenesis, 13 sporulation-negative (Spo) mutants were characterized; and 3 were found to produce only butyrate and acetate, a minor amount of acetone, and no alcohols. These Spo mutants contained reduced butanol dehydrogenase activity and no BAD enzyme activity. The data support the view that the type 2 AA, the Acid, and the Spo mutants somehow alter normal regulated expression of the solvent pathway in C. acetobutylicum.

  20. A Quantitative System-Scale Characterization of the Metabolism of Clostridium acetobutylicum

    PubMed Central

    Yoo, Minyeong; Bestel-Corre, Gwenaelle; Croux, Christian; Riviere, Antoine; Meynial-Salles, Isabelle

    2015-01-01

    ABSTRACT Engineering industrial microorganisms for ambitious applications, for example, the production of second-generation biofuels such as butanol, is impeded by a lack of knowledge of primary metabolism and its regulation. A quantitative system-scale analysis was applied to the biofuel-producing bacterium Clostridium acetobutylicum, a microorganism used for the industrial production of solvent. An improved genome-scale model, iCac967, was first developed based on thorough biochemical characterizations of 15 key metabolic enzymes and on extensive literature analysis to acquire accurate fluxomic data. In parallel, quantitative transcriptomic and proteomic analyses were performed to assess the number of mRNA molecules per cell for all genes under acidogenic, solventogenic, and alcohologenic steady-state conditions as well as the number of cytosolic protein molecules per cell for approximately 700 genes under at least one of the three steady-state conditions. A complete fluxomic, transcriptomic, and proteomic analysis applied to different metabolic states allowed us to better understand the regulation of primary metabolism. Moreover, this analysis enabled the functional characterization of numerous enzymes involved in primary metabolism, including (i) the enzymes involved in the two different butanol pathways and their cofactor specificities, (ii) the primary hydrogenase and its redox partner, (iii) the major butyryl coenzyme A (butyryl-CoA) dehydrogenase, and (iv) the major glyceraldehyde-3-phosphate dehydrogenase. This study provides important information for further metabolic engineering of C. acetobutylicum to develop a commercial process for the production of n-butanol. PMID:26604256

  1. Sustainable biobutanol production from pineapple waste by using Clostridium acetobutylicum B 527: Drying kinetics study.

    PubMed

    Khedkar, Manisha A; Nimbalkar, Pranhita R; Gaikwad, Shashank G; Chavan, Prakash V; Bankar, Sandip B

    2017-02-01

    Present investigation explores the use of pineapple peel, a food industry waste, for acetone-butanol-ethanol (ABE) production using Clostridium acetobutylicum B 527. Proximate analysis of pineapple peel shows that it contains 35% cellulose, 19% hemicellulose, and 16% lignin on dry basis. Drying experiments on pineapple peel waste were carried out in the temperature range of 60-120°C and experimental drying data was modeled using moisture diffusion control model to study its effect on ABE production. The production of ABE was further accomplished via acid hydrolysis, detoxification, and fermentation process. Maximum total sugar release obtained by using acid hydrolysis was 97g/L with 95-97% and 10-50% removal of phenolics and acetic acid, respectively during detoxification process. The maximum ABE titer obtained was 5.23g/L with 55.6% substrate consumption when samples dried at 120°C were used as a substrate (after detoxification).

  2. The mechanism of switching from an acidogenic to butanol-acetone fermentation by Clostridium acetobutylicum

    SciTech Connect

    Rogers, P.

    1992-01-01

    The overall objective of this project is to elucidate the detailed mechanism by which solvent-forming bacteria such as Clostridium acetobutylicum regulate the well-known shift in fermentation pathway between alcohol-acetone and organic acid production. It is desired to eventually isolate and describe: (1) the regulatory genes and protein elements that determine induction of synthesis of the solvent-pathway enzymes; and (2) how this regulation system interacts with the sporulatin induction and development program and with related pathways such as granulse and exopolysaccharide formation in clostridia. A working model forhow clostridial control systems work can be derived from recent research on stress systems in E. coli and sporulation in Bacillus subtilis.

  3. Reduction of 2,4,6-trinitrotoluene by Clostridium acetobutylicum through hydroxylamino-nitrotoluene intermediates

    SciTech Connect

    Hughes, J.B.; Wang, C.Y.; Bhadra, R.; Richardson, A.; Bennett, G.N.; Rudolph, F.B.

    1998-03-01

    Studies were conducted to isolate and identify intermediates of 2,4,6-trinitrotoluene (TNT) transformation by Clostridium acetobutylicum and to quantify their concentrations in active whole cell cultures. Only two intermediates of TNT reduction were detected in cell cultures and were identified as 4-hydroxylamino-2,6-dinitrotoluene and 2,4-dihydroxylamino-6-nitrotoluene. Structures were confirmed with {sup 1}H-NMR, {sup 13}C-NMR, and desorption chemical ionization mass spectroscopy. When cells were suspended in a non-growth saline medium, both hydroxylamine forms accumulated. In media capable of supporting cell growth, the 2,4-dihydroxylamino-6-nitrotoluene accumulated with concentrations of 4-hydroxylamino-2,6-dinitrotoluene remaining near detection limits. Studies using purified 2,4-dihydroxylamino-6-nitrotoluene confirmed that its biotransformation rate in active cultures greatly exceeded abiotic decomposition in aqueous medium.

  4. Continuous lactose fermentation by Clostridium acetobutylicum--assessment of acidogenesis kinetics.

    PubMed

    Napoli, Fabio; Olivieri, Giuseppe; Russo, Maria Elena; Marzocchella, Antonio; Salatino, Piero

    2011-01-01

    An assessment of the growth kinetics of acidogenic cells of Clostridium acetobutylicum DSM 792 is reported in the paper. Tests were carried out in a continuous stirred tank reactor under controlled conditions adopting a complex medium supplemented with lactose as carbon source to mimic cheese whey. The effects of acids (acetic and butyric), solvents (acetone, ethanol and butanol) and pH on the growth rate of acidogenic cells were assessed. The conversion process was characterized under steady-state conditions in terms of concentration of lactose, cells, acids, total organic carbon and pH. The growth kinetics was expressed by means of a multiple product inhibition and interacting model including a novel formulation to account for the role of pH. The model has the potential to predict microorganism growth rate under a broad interval of operating conditions, even those typical of solvents production.

  5. Nutritional Factors Affecting the Ratio of Solvents Produced by Clostridium acetobutylicum

    PubMed Central

    Bahl, H.; Gottwald, M.; Kuhn, A.; Rale, V.; Andersch, W.; Gottschalk, G.

    1986-01-01

    Fermentation of whey by Clostridium acetobutylicum yielded butanol and acetone in a ratio of approximately 100:1. This ratio amounted to only 2:1 in synthetic media with glucose, lactose, or glucose plus galactose as substrates. Removal of citrate from whey and addition of minerals resulted in an increase in the amount of acetone produced. Experiments carried out in a chemostat with a low-phosphate synthetic medium revealed that the butanol/acetone ratio could be increased from 2:1 to 3.8:1 by cofermentation of l-lactate and from 2:1 to 8:1 by iron limitation. The performance of the fermentation in a low-iron glucose medium above pH 5.1 yielded l-lactate as the main product. PMID:16347104

  6. Nutritional factors affecting the ratio of solvents produced by Clostridium acetobutylicum

    SciTech Connect

    Bahl, H.; Gottwald, M.; Kuhn, A.; Rale, V.; Andersch, W.; Gottschalk, G.

    1986-07-01

    Fermentation of whey by Clostridium acetobutylicum yielded butanol and acetone in a ratio of approximately 100:1. This ratio amounted to only 2:1 in synthetic media with glucose, lactose, or glucose plus galactose as substrates. Removal of citrate from whey and addition of minerals resulted in an increase in the amount of acetone produced. Experiments carried out in a chemostat with a low-phosphate synthetic medium revealed that the butanol/acetone ratio could be increased from 2:1 to 3.8:1 by cofermentation of L-lactate and from 2:1 to 8:1 by iron limitation. The performance of the fermentation in a low-iron glucose medium above pH 5.1 yielded L-lactate as the main product. 42 references.

  7. Shotgun proteomic monitoring of Clostridium acetobutylicum during stationary phase of butanol fermentation using xylose and comparison with the exponential phase

    SciTech Connect

    Sivagnanam, Kumaran; Raghavan, Vijaya G. S.; Shah, Manesh B; Hettich, Robert {Bob} L; Verberkmoes, Nathan C; Lefsrud, Mark G

    2012-01-01

    Economically viable production of solvents through acetone butanol ethanol (ABE) fermentation requires a detailed understanding of Clostridium acetobutylicum. This study focuses on the proteomic profiling of C. acetobutylicum ATCC 824 from the stationary phase of ABE fermentation using xylose and compares with the exponential growth by shotgun proteomics approach. Comparative proteomic analysis revealed 22.9% of the C. acetobutylicum genome and 18.6% was found to be common in both exponential and stationary phases. The proteomic profile of C. acetobutylicum changed during the ABE fermentation such that 17 proteins were significantly differentially expressed between the two phases. Specifically, the expression of five proteins namely, CAC2873, CAP0164, CAP0165, CAC3298, and CAC1742 involved in the solvent production pathway were found to be significantly lower in the stationary phase compared to the exponential growth. Similarly, the expression of fucose isomerase (CAC2610), xylulose kinase (CAC2612), and a putative uncharacterized protein (CAC2611) involved in the xylose utilization pathway were also significantly lower in the stationary phase. These findings provide an insight into the metabolic behavior of C. acetobutylicum between different phases of ABE fermentation using xylose.

  8. Metabolic engineering of Clostridium acetobutylicum for the enhanced production of isopropanol-butanol-ethanol fuel mixture.

    PubMed

    Jang, Yu-Sin; Malaviya, Alok; Lee, Joungmin; Im, Jung Ae; Lee, Sang Yup; Lee, Julia; Eom, Moon-Ho; Cho, Jung-Hee; Seung, Do Young

    2013-01-01

    Butanol is considered as a superior biofuel, which is conventionally produced by clostridial acetone-butanol-ethanol (ABE) fermentation. Among ABE, only butanol and ethanol can be used as fuel alternatives. Coproduction of acetone thus causes lower yield of fuel alcohols. Thus, this study aimed at developing an improved Clostridium acetobutylicum strain possessing enhanced fuel alcohol production capability. For this, we previously developed a hyper ABE producing BKM19 strain was further engineered to convert acetone into isopropanol. The BKM19 strain was transformed with the plasmid pIPA100 containing the sadh (primary/secondary alcohol dehydrogenase) and hydG (putative electron transfer protein) genes from the Clostridium beijerinckii NRRL B593 cloned under the control of the thiolase promoter. The resulting BKM19 (pIPA100) strain produced 27.9 g/l isopropanol-butanol-ethanol (IBE) as a fuel alcohols with negligible amount of acetone (0.4 g/l) from 97.8 g/l glucose in lab-scale (2 l) batch fermentation. Thus, this metabolically engineered strain was able to produce 99% of total solvent produced as fuel alcohols. The scalability and stability of BKM19 (pIPA100) were evaluated at 200 l pilot-scale fermentation, which showed that the fuel alcohol yield could be improved to 0.37 g/g as compared to 0.29 g/g obtained at lab-scale fermentation, while attaining a similar titer. To the best of our knowledge, this is the highest titer of IBE achieved and the first report on the large scale fermentation of C. acetobutylicum for IBE production.

  9. Genome shuffling of Clostridium acetobutylicum CICC 8012 for improved production of acetone-butanol-ethanol (ABE).

    PubMed

    Gao, Xiaofeng; Zhao, Hai; Zhang, Guohua; He, Kaize; Jin, Yanling

    2012-08-01

    Genome shuffling was applied to increase ABE production of the strict anaerobe C. acetobutylicum CICC 8012. By using physical and chemical mutagenesis, strains with superior streptomycin sulfate, 2-deoxy-D-glucose and butanol tolerance levels were isolated. These strains were used for genome shuffling. The best performing strain F2-GA was screened after two rounds of genome shuffling. With 55 g glucose/l as carbon source, F2-GA produced 22.21 g ABE/l in 72 h and ABE yield reached 0.42 g/g which was about 34.53 % improvement compared to the wild type. Fermentation parameters and gene expression of several key enzymes in ABE metabolic pathways were varied significantly between F2-GA and the wild type. These results demonstrated the potential use of genome shuffling to microbial breeding which were difficult to deal with traditional methods.

  10. Enhanced butanol production obtained by reinforcing the direct butanol-forming route in Clostridium acetobutylicum.

    PubMed

    Jang, Yu-Sin; Lee, Jin Young; Lee, Joungmin; Park, Jin Hwan; Im, Jung Ae; Eom, Moon-Ho; Lee, Julia; Lee, Sang-Hyun; Song, Hyohak; Cho, Jung-Hee; Seung, Do Young; Lee, Sang Yup

    2012-10-23

    Butanol is an important industrial solvent and advanced biofuel that can be produced by biphasic fermentation by Clostridium acetobutylicum. It has been known that acetate and butyrate first formed during the acidogenic phase are reassimilated to form acetone-butanol-ethanol (cold channel). Butanol can also be formed directly from acetyl-coenzyme A (CoA) through butyryl-CoA (hot channel). However, little is known about the relative contributions of the two butanol-forming pathways. Here we report that the direct butanol-forming pathway is a better channel to optimize for butanol production through metabolic flux and mass balance analyses. Butanol production through the hot channel was maximized by simultaneous disruption of the pta and buk genes, encoding phosphotransacetylase and butyrate kinase, while the adhE1(D485G) gene, encoding a mutated aldehyde/alcohol dehydrogenase, was overexpressed. The ratio of butanol produced through the hot channel to that produced through the cold channel increased from 2.0 in the wild type to 18.8 in the engineered BEKW(pPthlAAD(**)) strain. By reinforcing the direct butanol-forming flux in C. acetobutylicum, 18.9 g/liter of butanol was produced, with a yield of 0.71 mol butanol/mol glucose by batch fermentation, levels which are 160% and 245% higher than those obtained with the wild type. By fed-batch culture of this engineered strain with in situ recovery, 585.3 g of butanol was produced from 1,861.9 g of glucose, with the yield of 0.76 mol butanol/mol glucose and productivity of 1.32 g/liter/h. Studies of two butanol-forming routes and their effects on butanol production in C. acetobutylicum described here will serve as a basis for further metabolic engineering of clostridia aimed toward developing a superior butanol producer. IMPORTANCE Renewable biofuel is one of the answers to solving the energy crisis and climate change problems. Butanol produced naturally by clostridia has superior liquid fuel characteristics and thus has

  11. High-efficient n-butanol production by co-culturing Clostridium acetobutylicum and Saccharomyces cerevisiae integrated with butyrate fermentative supernatant addition.

    PubMed

    Luo, Hongzhen; Zeng, Qingwei; Han, Shuo; Wang, Zhaoyu; Dong, Qing; Bi, Yanhong; Zhao, Yuping

    2017-04-01

    Butanol is not only an important chemical intermediate and solvent in pharmaceutical and cosmetics industries, but also considered as an advanced biofuel. Although species of the natural host Clostridium have been engineered, butanol titers in the anaerobe seem to be limited by its intolerance to butanol less than 13 g/L. Here we aimed to develop a technology for enhancing butanol production by a co-culture system with butyrate fermentative supernatant addition. First, when adding 4.0 g/L butyrate into the acetone-butanol-ethanol (ABE) fermentation broth with single-shot at 24 h, the "acid crash" phenomenon occurred and the ABE fermentation performance deteriorated. Subsequently, we found that adding certain amino acids could effectively enhance butyrate re-assimilation, butanol tolerance and titer (from 11.1 to 14.8 g/L). Additionally, in order to decrease the raw material cost, butyrate fermentative supernatant produced by Clostridium tyrobutyricum was applied to butanol production in the Clostridium acetobutylicum/Saccharomyces cerevisiae co-culture system, instead of adding synthetic butyrate. Final butanol and total ABE concentrations reached higher levels of 16.3 and 24.8 g/L with increments of 46.8 and 37.8%, respectively. These results show that the proposed fermentation strategy has great potential for efficiently butanol production with an economic approach.

  12. Structural Variation in Bacterial Glyoxalase I Enzymes: Investigation of the Metalloenzyme Glyoxalase I from Clostridium acetobutylicum

    SciTech Connect

    Suttisansanee U.; Swaminathan S.; Lau, K.; Lagishetty, S.; Rao, K. N.; Sauder, J. M.; Burley, S. K.; Honek, J. F.

    2011-11-04

    The glyoxalase system catalyzes the conversion of toxic, metabolically produced {alpha}-ketoaldehydes, such as methylglyoxal, into their corresponding nontoxic 2-hydroxycarboxylic acids, leading to detoxification of these cellular metabolites. Previous studies on the first enzyme in the glyoxalase system, glyoxalase I (GlxI), from yeast, protozoa, animals, humans, plants, and Gram-negative bacteria, have suggested two metal activation classes, Zn{sup 2+} and non-Zn{sup 2+} activation. Here, we report a biochemical and structural investigation of the GlxI from Clostridium acetobutylicum, which is the first GlxI enzyme from Gram-positive bacteria that has been fully characterized as to its three-dimensional structure and its detailed metal specificity. It is a Ni{sup 2+}/Co{sup 2+}-activated enzyme, in which the active site geometry forms an octahedral coordination with one metal atom, two water molecules, and four metal-binding ligands, although its inactive Zn{sup 2+}-bound form possesses a trigonal bipyramidal geometry with only one water molecule liganded to the metal center. This enzyme also possesses a unique dimeric molecular structure. Unlike other small homodimeric GlxI where two active sites are located at the dimeric interface, the C. acetobutylicum dimeric GlxI enzyme also forms two active sites but each within single subunits. Interestingly, even though this enzyme possesses a different dimeric structure from previously studied GlxI, its metal activation characteristics are consistent with properties of other GlxI. These findings indicate that metal activation profiles in this class of enzyme hold true across diverse quaternary structure arrangements.

  13. Acetone-butanol-ethanol production with high productivity using Clostridium acetobutylicum BKM19.

    PubMed

    Jang, Yu-Sin; Malaviya, Alok; Lee, Sang Yup

    2013-06-01

    Conventional acetone-butanol-ethanol (ABE) fermentation is severely limited by low solvent titer and productivities. Thus, this study aims at developing an improved Clostridium acetobutylicum strain possessing enhanced ABE production capability followed by process optimization for high ABE productivity. Random mutagenesis of C. acetobutylicum PJC4BK was performed by screening cells on fluoroacetate plates to isolate a mutant strain, BKM19, which exhibited the total solvent production capability 30.5% higher than the parent strain. The BKM19 produced 32.5 g L(-1) of ABE (17.6 g L(-1) butanol, 10.5 g L(-1) ethanol, and 4.4 g L(-1) acetone) from 85.2 g L(-1) glucose in batch fermentation. A high cell density continuous ABE fermentation of the BKM19 in membrane cell-recycle bioreactor was studied and optimized for improved solvent volumetric productivity. Different dilution rates were examined to find the optimal condition giving highest butanol and ABE productivities. The maximum butanol and ABE productivities of 9.6 and 20.0 g L(-1)  h(-1) , respectively, could be achieved at the dilution rate of 0.85 h(-1) . Further cell recycling experiments were carried out with controlled cell-bleeding at two different bleeding rates. The maximum solvent productivities were obtained when the fermenter was operated at a dilution rate of 0.86 h(-1) with the bleeding rate of 0.04 h(-1) . Under the optimal operational condition, butanol and ABE could be produced with the volumetric productivities of 10.7 and 21.1 g L(-1)  h(-1) , and the yields of 0.17 and 0.34 g g(-1) , respectively. The obtained butanol and ABE volumetric productivities are the highest reported productivities obtained from all known-processes.

  14. Expression of Clostridium acetobutylicum ATCC 824 Genes in Escherichia coli for Acetone Production and Acetate Detoxification

    PubMed Central

    Bermejo, Lourdes L.; Welker, Neil E.; Papoutsakis, Eleftherios T.

    1998-01-01

    A synthetic acetone operon (ace4) composed of four Clostridium acetobutylicum ATCC 824 genes (adc, ctfAB, and thl, coding for the acetoacetate decarboxylase, coenzyme A transferase, and thiolase, respectively) under the control of the thl promoter was constructed and was introduced into Escherichia coli on vector pACT. Acetone production demonstrated that ace4 is expressed in E. coli and resulted in the reduction of acetic acid levels in the fermentation broth. Since different E. coli strains vary significantly in their growth characteristics and acetate metabolism, ace4 was expressed in three E. coli strains: ER2275, ATCC 11303, and MC1060. Shake flask cultures of MC1060(pACT) produced ca. 2 mM acetone, while both strains ER2275(pACT) and ATCC 11303(pACT) produced ca. 40 mM acetone. Glucose-fed cultures of strain ATCC 11303(pACT) resulted in a 150% increase in acetone titers compared to those of batch shake flask cultures. External addition of sodium acetate to glucose-fed cultures of ATCC 11303(pACT) resulted in further increased acetone titers. In bioreactor studies, acidic conditions (pH 5.5 versus 6.5) improved acetone production. Despite the substantial acetone evaporation due to aeration and agitation in the bioreactor, 125 to 154 mM acetone accumulated in ATCC 11303(pACT) fermentations. These acetone titers are equal to or higher than those produced by wild-type C. acetobutylicum. This is the first study to demonstrate the ability to use clostridial genes in nonclostridial hosts for solvent production. In addition, acetone-producing E. coli strains may be useful hosts for recombinant protein production in that detrimental acetate accumulation can be avoided. PMID:9501448

  15. Purification and characterization of the extracellular. alpha. -amylase from Clostridium acetobutylicum ATCC 824

    SciTech Connect

    Paquet, V.; Croux, C.; Goma, G.; Soucaille, P. )

    1991-01-01

    The extracellular {alpha}-amylase (1,4-{alpha}-D-glucanglucanohydrolase; EC 3.2.1.1) from Clostridium acetobutylicum ATCC 824 was purified to homogeneity by anion-exchange chromatography (Mono Q) and gel filtration (Superose 12). The enzyme had an isoelectric point of 4.7 and a molecular weight of 84,000, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It was a monomeric protein, the 19-amino-acid N terminus of which displayed 42% homology with the Bacillus subtilis saccharifying {alpha}-amylase. The amino acid composition of the enzyme showed a high number of acidic and hydrophobic residues and only one cysteine residue per mole. The activity of the {alpha}-amylase was not stimulated by calcium ions (or other metal ions) or inhibited by EDTA, although the enzyme contained seven calcium atoms per molecule. {alpha}-Amylase activity on soluble starch was optimal at pH 5.6 and 45{degree}C. The {alpha}-amylase was stable at an acidic pH but very sensitive to thermal inactivation. It hydrolyzed soluble starch, with a K{sub m} of 3.6 g {center dot} liter{sup {minus}1} and a K{sub cat} of 122 mol of reducing sugars {center dot} s{sup {minus}1} {center dot} mol{sup {minus}1}. The {alpha}-amylase showed greater activity with high-molecular-weight substrates than with low-molecular-weight maltooligosaccharides, hydrolyzed glycogen and pullulan slowly, but did not hydrolyze dextran or cyclodextrins. The major end products of maltohexaose degradation were glucose, maltose, and maltotriose; maltotetraose and maltopentaose were formed as intermediate products. Twenty seven percent of the glucoamylase activity generally detected in the culture supernatant of C. acetobutylicum can be attributed to the {alpha}-amylase.

  16. Enhanced Butanol Production Obtained by Reinforcing the Direct Butanol-Forming Route in Clostridium acetobutylicum

    PubMed Central

    Jang, Yu-Sin; Lee, Jin Young; Lee, Joungmin; Park, Jin Hwan; Im, Jung Ae; Eom, Moon-Ho; Lee, Julia; Lee, Sang-Hyun; Song, Hyohak; Cho, Jung-Hee; Seung, Do Young; Lee, Sang Yup

    2012-01-01

    ABSTRACT Butanol is an important industrial solvent and advanced biofuel that can be produced by biphasic fermentation by Clostridium acetobutylicum. It has been known that acetate and butyrate first formed during the acidogenic phase are reassimilated to form acetone-butanol-ethanol (cold channel). Butanol can also be formed directly from acetyl-coenzyme A (CoA) through butyryl-CoA (hot channel). However, little is known about the relative contributions of the two butanol-forming pathways. Here we report that the direct butanol-forming pathway is a better channel to optimize for butanol production through metabolic flux and mass balance analyses. Butanol production through the hot channel was maximized by simultaneous disruption of the pta and buk genes, encoding phosphotransacetylase and butyrate kinase, while the adhE1D485G gene, encoding a mutated aldehyde/alcohol dehydrogenase, was overexpressed. The ratio of butanol produced through the hot channel to that produced through the cold channel increased from 2.0 in the wild type to 18.8 in the engineered BEKW(pPthlAAD**) strain. By reinforcing the direct butanol-forming flux in C. acetobutylicum, 18.9 g/liter of butanol was produced, with a yield of 0.71 mol butanol/mol glucose by batch fermentation, levels which are 160% and 245% higher than those obtained with the wild type. By fed-batch culture of this engineered strain with in situ recovery, 585.3 g of butanol was produced from 1,861.9 g of glucose, with the yield of 0.76 mol butanol/mol glucose and productivity of 1.32 g/liter/h. Studies of two butanol-forming routes and their effects on butanol production in C. acetobutylicum described here will serve as a basis for further metabolic engineering of clostridia aimed toward developing a superior butanol producer. PMID:23093384

  17. Solvents Production from a Mixture of Glucose and Xylose by Mixed Fermentation of Clostridium acetobutylicum and Saccharomyces cerevisiae.

    PubMed

    Qi, Gao-Xiang; Xiong, Lian; Huang, Chao; Chen, Xue-Fang; Lin, Xiao-Qing; Chen, Xin-De

    2015-10-01

    To overcome the xylose utilization defect in ethanol fermentation by wide-type Saccharomyces cerevisiae and alleviate the carbon catabolite repression (CCR) in acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum, a novel mixed fermentation of S. cerevisiae and C. acetobutylicum was developed. When S. cerevisiae was inoculated 24 h earlier than C. acetobutylicum CH02, a higher solvents yield was achieved with 0.41 g/g, compared to 0.38 g/g in ABE fermentation, and when S. cerevisiae and C. acetobutylicum CH02 were inoculated simultaneously, a higher productivity was achieved with 0.32 g/L/h, compared to 0.15 g/L/h in ABE fermentation. The total solvents yield was improved by the high ethanol yield from glucose. The CCR in mixed fermentation was alleviated when glucose was utilized quickly by S. cerevisiae, and therefore, the productivity was improved. This study suggests that mixed fermentation is an effective solvents production method from a mixture of glucose and xylose.

  18. Development of real-time PCR primer and probe sets for detecting degenerated and non-degenerated forms of the butanol-producing bacterium Clostridium acetobutylicum ATCC 824.

    PubMed

    Lee, Sun-Mi; Cho, Min Ok; Um, Youngsoon; Sang, Byoung-In

    2010-05-01

    Degeneration is one of the limiting factors in butanol fermentation, and it must be monitored and prevented for stable butanol production. In Clostridium acetobutylicum ATCC 824, the most well-known butanol-producing microorganism, degeneration is caused by the loss of the pSOL1 plasmid that carries essential genes involved in solvent production. In this study, we designed two specific primer and probe sets for real-time qPCR (RT-qPCR) detection of C. acetobutylicum ATCC 824 (the C. aceto set) and pSOL1-possessing C. acetobutylicum ATCC 824 (the DGS set). Specific primer and probe sets were designed on the basis of the 16S rDNA sequence and pSOL1 sequence. The number of degenerated C. acetobutylicum could be quantified by subtracting the number of C. acetobutylicum ATCC 824 containing pSOL1 from the total number of C. acetobutylicum ATCC 824. The primer and probe sets permitted the specific detection and quantification of degenerated C. acetobutylicum and total butanol-producing C. acetobutylicum by RT-qPCR.

  19. Confirmation and elimination of xylose metabolism bottlenecks in glucose phosphoenolpyruvate-dependent phosphotransferase system-deficient Clostridium acetobutylicum for simultaneous utilization of glucose, xylose, and arabinose.

    PubMed

    Xiao, Han; Gu, Yang; Ning, Yuanyuan; Yang, Yunliu; Mitchell, Wilfrid J; Jiang, Weihong; Yang, Sheng

    2011-11-01

    Efficient cofermentation of D-glucose, D-xylose, and L-arabinose, three major sugars present in lignocellulose, is a fundamental requirement for cost-effective utilization of lignocellulosic biomass. The Gram-positive anaerobic bacterium Clostridium acetobutylicum, known for its excellent capability of producing ABE (acetone, butanol, and ethanol) solvent, is limited in using lignocellulose because of inefficient pentose consumption when fermenting sugar mixtures. To overcome this substrate utilization defect, a predicted glcG gene, encoding enzyme II of the D-glucose phosphoenolpyruvate-dependent phosphotransferase system (PTS), was first disrupted in the ABE-producing model strain Clostridium acetobutylicum ATCC 824, resulting in greatly improved D-xylose and L-arabinose consumption in the presence of D-glucose. Interestingly, despite the loss of GlcG, the resulting mutant strain 824glcG fermented D-glucose as efficiently as did the parent strain. This could be attributed to residual glucose PTS activity, although an increased activity of glucose kinase suggested that non-PTS glucose uptake might also be elevated as a result of glcG disruption. Furthermore, the inherent rate-limiting steps of the D-xylose metabolic pathway were observed prior to the pentose phosphate pathway (PPP) in strain ATCC 824 and then overcome by co-overexpression of the D-xylose proton-symporter (cac1345), D-xylose isomerase (cac2610), and xylulokinase (cac2612). As a result, an engineered strain (824glcG-TBA), obtained by integrating glcG disruption and genetic overexpression of the xylose pathway, was able to efficiently coferment mixtures of D-glucose, D-xylose, and L-arabinose, reaching a 24% higher ABE solvent titer (16.06 g/liter) and a 5% higher yield (0.28 g/g) compared to those of the wild-type strain. This strain will be a promising platform host toward commercial exploitation of lignocellulose to produce solvents and biofuels.

  20. Carbon 13-Metabolic Flux Analysis derived constraint-based metabolic modelling of Clostridium acetobutylicum in stressed chemostat conditions.

    PubMed

    Wallenius, Janne; Maaheimo, Hannu; Eerikäinen, Tero

    2016-11-01

    The metabolism of butanol producing bacteria Clostridium acetobutylicum was studied in chemostat with glucose limited conditions, butanol stimulus, and as a reference cultivation. COnstraint-Based Reconstruction and Analysis (COBRA) was applied using additional constraints from (13)C Metabolic Flux Analysis ((13)C-MFA) and experimental measurement results. A model consisting of 451 metabolites and 604 reactions was utilized in flux balance analysis (FBA). The stringency of the flux spaces considering different optimization objectives, i.e. growth rate maximization, ATP maintenance, and NADH/NADPH formation, for flux variance analysis (FVA) was studied in the different modelled conditions. Also a previously uncharacterized exopolysaccharide (EPS) produced by C. acetobutylicum was characterized on monosaccharide level. The major monosaccharide components of the EPS were 40n-% rhamnose, 34n-% glucose, 13n-% mannose, 10n-% galactose, and 2n-% arabinose. The EPS was studied to have butanol adsorbing property, 70(butanol)mg(EPS)g(-1) at 37°C.

  1. Butanol production from hexoses and pentoses by fermentation of Clostridium acetobutylicum.

    PubMed

    Raganati, Francesca; Olivieri, Giuseppe; Götz, Peter; Marzocchella, Antonio; Salatino, Piero

    2015-08-01

    The present paper reports the characterization of ABE (acetone-butanol-ethanol) production by Clostridium acetobutylicum DSM 792 for sugars representative of hydrolysed lignocellulosic biomass (glucose, mannose, arabinose, xylose). The attention was focused on: the selection of an optimal medium for the simultaneous conversion of the investigated sugars; the assessment of interference-synergistic effects during the fermentation of mixtures of the investigated sugars. The synthetic medium was optimised in terms of nutritional factors: the KH2PO4-K2HPO4 concentration was increased up to 5 g/L; the MgSO4 concentration was increased up to 2 g/L; the MnSO4 concentration was increased up to 0.1 g/L; the FeSO4 concentration ranged between 0.002 and 0.01 g/L); the CaCO3 concentration was increased up to 10 g/L. The optimal concentration of the investigated factors was assessed and it varied from one sugar to another. The batch fermentations of a mixture of the four sugars highlighted their synergistic effects. Once set the initial concentration of the sugars (60 g/L), the butanol and solvent concentration increased up to 14.6 and 20.6 g/L, respectively, when the four sugars were present.

  2. Enhanced butanol fermentation using metabolically engineered Clostridium acetobutylicum with ex situ recovery of butanol.

    PubMed

    Lee, Sang-Hyun; Kim, Sooah; Kim, Jung Yeon; Cheong, Nam Yong; Kim, Kyoung Heon

    2016-10-01

    In this study, metabolic target reactions for strain engineering were searched via intracellular coenzyme A (CoA) metabolite analysis. The metabolic reactions catalyzed by thiolase (AtoB) and aldehyde-alcohol dehydrogenase (AdhE1) were considered potential rate-limiting steps. In addition, CoA transferase (CtfAB) was highlighted as being important for the assimilation of organic acids, in order to achieve high butanol production. Based on this quantitative analysis, the BEKW_E1AB-atoB strain was constructed by overexpressing the thl (atoB), adhE1, and ctfAB genes in Clostridium acetobutylicum strain BEKW, which has the phosphotransacetylase (pta) and butyrate kinase (buk) genes knocked out. After 100h of continuous fermentation coupled with adsorptive ex situ butanol recovery, the concentrations found after considering desorption, yield, and productivity for the BEKW_E1AB-atoB strain were 55.7g/L, 0.38g/g, and 2.64g/L/h, respectively. The level of butanol production achieved (2.64g/L/h) represents the highest reported value obtained after adsorptive, long-term fermentation.

  3. Immobilization of Clostridium acetobutylicum onto natural textiles and its fermentation properties.

    PubMed

    Zhuang, Wei; Liu, Xiaojing; Yang, Jing; Wu, Jinglan; Zhou, Jingwei; Chen, Yong; Liu, Dong; Ying, Hanjie

    2017-03-01

    Immobilized fermentation has several advantages over traditional suspended fermentation, including simple and continuous operation, improved fermentation performance and reduced cost. Carrier is the most adjustable element among three elements of immobilized fermentation, including carrier, bacteria and environment. In this study, we characterized carrier roughness and surface properties of four types of natural fibres, including linen, cotton, bamboo fibre and silk, to assess their effects on cell immobilization, fermentation performance and stability. Linen with higher specific surface area and roughness could adsorb more bacteria during immobilized fermentation, thereby improving fermentation performance; thus, linen was selected as a suitable carrier and was applied for acetone-butanol-ethanol (ABE) fermentation. To further improve fermentation performance, we also found that microbes of Clostridium acetobutylicum were negatively charged surfaces during fermentation. Therefore, we then modified linen with polyetherimide (PEI) and steric acid (SA) to increase surface positive charge and improve surface property. During ABE fermentation, the adhesion between modified linen and bacteria was increased, adsorption was increased about twofold compared with that of unmodified linen, and butanol productivity was increased 8.16% and 6.80% with PEI- and SA-modified linen as carriers respectively.

  4. Expression and nucleotide sequence of the Clostridium acetobutylicum beta-galactosidase gene cloned in Escherichia coli.

    PubMed Central

    Hancock, K R; Rockman, E; Young, C A; Pearce, L; Maddox, I S; Scott, D B

    1991-01-01

    A gene library for Clostridium acetobutylicum NCIB 2951 was constructed in the broad-host-range cosmid pLAFR1, and cosmids containing the beta-galactosidase gene were isolated by direct selection for enzyme activity on X-Gal (5-bromo-4-chloro-3-indolyl-beta-D-galactoside) plates after conjugal transfer of the library to a lac deletion derivative of Escherichia coli. Analysis of various pSUP202 subclones of the lac cosmids on X-Gal plates localized the beta-galactosidase gene to a 5.1-kb EcoRI fragment. Expression of the Clostridium beta-galactosidase gene in E. coli was not subject to glucose repression. By using transposon Tn5 mutagenesis, two gene loci, cbgA (locus I) and cbgR (locus II), were identified as necessary for beta-galactosidase expression in E. coli. DNA sequence analysis of the entire 5.1-kb fragment identified open reading frames of 2,691 and 303 bp, corresponding to locus I and locus II, respectively, and in addition a third truncated open reading frame of 825 bp. The predicted gene product of locus I, CbgA (molecular size, 105 kDa), showed extensive amino acid sequence homology with E. coli LacZ, E. coli EbgA, and Klebsiella pneumoniae LacZ and was in agreement with the size of a polypeptide synthesized in maxicells containing the cloned 5.1-kb fragment. The predicted gene product of locus II, CbgR (molecular size, 11 kDa) shares no significant homology with any other sequence in the current DNA and protein sequence data bases, but Tn5 insertions in this gene prevent the synthesis of CbgA. Complementation experiments indicate that the gene product of cbgR is required in cis with cbgA for expression of beta-galactosidase in E. coli. Images PMID:1850729

  5. Structural analysis of Clostridium acetobutylicum ATCC 824 glycoside hydrolase from CAZy family GH105

    SciTech Connect

    Germane, Katherine L.; Servinsky, Matthew D.; Gerlach, Elliot S.; Sund, Christian J.; Hurley, Margaret M.

    2015-07-29

    The crystal structure of the protein product of the C. acetobutylicum ATCC 824 gene CA-C0359 is structurally similar to YteR, an unsaturated rhamnogalacturonyl hydrolase from B. subtilis strain 168. Substrate modeling and electrostatic studies of the active site of the structure of CA-C0359 suggests that the protein can now be considered to be part of CAZy glycoside hydrolase family 105. Clostridium acetobutylicum ATCC 824 gene CA-C0359 encodes a putative unsaturated rhamnogalacturonyl hydrolase (URH) with distant amino-acid sequence homology to YteR of Bacillus subtilis strain 168. YteR, like other URHs, has core structural homology to unsaturated glucuronyl hydrolases, but hydrolyzes the unsaturated disaccharide derivative of rhamnogalacturonan I. The crystal structure of the recombinant CA-C0359 protein was solved to 1.6 Å resolution by molecular replacement using the phase information of the previously reported structure of YteR (PDB entry (http://scripts.iucr.org/cgi-bin/cr.cgi?rm)) from Bacillus subtilis strain 168. The YteR-like protein is a six-α-hairpin barrel with two β-sheet strands and a small helix overlaying the end of the hairpins next to the active site. The protein has low primary protein sequence identity to YteR but is structurally similar. The two tertiary structures align with a root-mean-square deviation of 1.4 Å and contain a highly conserved active pocket. There is a conserved aspartic acid residue in both structures, which has been shown to be important for hydration of the C=C bond during the release of unsaturated galacturonic acid by YteR. A surface electrostatic potential comparison of CA-C0359 and proteins from CAZy families GH88 and GH105 reveals the make-up of the active site to be a combination of the unsaturated rhamnogalacturonyl hydrolase and the unsaturated glucuronyl hydrolase from Bacillus subtilis strain 168. Structural and electrostatic comparisons suggests that the protein may have a slightly different substrate

  6. Targeted mutagenesis of the Clostridium acetobutylicum acetone-butanol-ethanol fermentation pathway.

    PubMed

    Cooksley, Clare M; Zhang, Ying; Wang, Hengzheng; Redl, Stephanie; Winzer, Klaus; Minton, Nigel P

    2012-11-01

    The production of the chemical solvents acetone and butanol by the bacterium Clostridium acetobutylicum was one of the first large-scale industrial processes to be developed, and in the first part of the last century ranked second in importance only to ethanol production. After a steep decline in its industrial use, there has been a recent resurgence of interest in the acetone-butanol-ethanol (ABE) fermentation process, with a particular emphasis on butanol production. In order to generate strains suitable for efficient use on an industrial scale, metabolic engineering is required to alter the AB ratio in favour of butanol, and eradicate the production of unwanted products of fermentation. Using ClosTron technology, a large-scale targeted mutagenesis in C. acetobutylicum ATCC 824 was carried out, generating a set of 10 mutants, defective in alcohol/aldehyde dehydrogenases 1 and 2 (adhE1, adhE2), butanol dehydrogenases A and B (bdhA, bdhB), phosphotransbutyrylase (ptb), acetate kinase (ack), acetoacetate decarboxylase (adc), CoA transferase (ctfA/ctfB), and a previously uncharacterised putative alcohol dehydrogenase (CAP0059). However, inactivation of the main hydrogenase (hydA) and thiolase (thl) could not be achieved. Constructing such a series of mutants is paramount for the acquisition of information on the mechanism of solvent production in this organism, and the subsequent development of industrial solvent producing strains. Unexpectedly, bdhA and bdhB mutants did not affect solvent production, whereas inactivation of the previously uncharacterised gene CAP0059 resulted in increased acetone, butanol, and ethanol formation. Other mutants showed predicted phenotypes, including a lack of acetone formation (adc, ctfA, and ctfB mutants), an inability to take up acids (ctfA and ctfB mutants), and a much reduced acetate formation (ack mutant). The adhE1 mutant in particular produced very little solvents, demonstrating that this gene was indeed the main contributor to

  7. Secretion and assembly of functional mini-cellulosomes from synthetic chromosomal operons in Clostridium acetobutylicum ATCC 824

    PubMed Central

    2013-01-01

    Background Consolidated bioprocessing (CBP) is reliant on the simultaneous enzyme production, saccharification of biomass, and fermentation of released sugars into valuable products such as butanol. Clostridial species that produce butanol are, however, unable to grow on crystalline cellulose. In contrast, those saccharolytic species that produce predominantly ethanol, such as Clostridium thermocellum and Clostridium cellulolyticum, degrade crystalline cellulose with high efficiency due to their possession of a multienzyme complex termed the cellulosome. This has led to studies directed at endowing butanol-producing species with the genetic potential to produce a cellulosome, albeit by localising the necessary transgenes to unstable autonomous plasmids. Here we have explored the potential of our previously described Allele-Coupled Exchange (ACE) technology for creating strains of the butanol producing species Clostridium acetobutylicum in which the genes encoding the various cellulosome components are stably integrated into the genome. Results We used BioBrick2 (BB2) standardised parts to assemble a range of synthetic genes encoding C. thermocellum cellulosomal scaffoldin proteins (CipA variants) and glycoside hydrolases (GHs, Cel8A, Cel9B, Cel48S and Cel9K) as well as synthetic cellulosomal operons that direct the synthesis of Cel8A, Cel9B and a truncated form of CipA. All synthetic genes and operons were integrated into the C. acetobutylicum genome using the recently developed ACE technology. Heterologous protein expression levels and mini-cellulosome self-assembly were assayed by western blot and native PAGE analysis. Conclusions We demonstrate the successful expression, secretion and self-assembly of cellulosomal subunits by the recombinant C. acetobutylicum strains, providing a platform for the construction of novel cellulosomes. PMID:23962085

  8. Increased productivity of Clostridium acetobutylicum fermentation of acetone, butanol, and ethanol by pervaporation through supported ionic liquid membrane.

    PubMed

    Izák, Pavel; Schwarz, Katrin; Ruth, Wolfgang; Bahl, Hubert; Kragl, Udo

    2008-03-01

    Pervaporation proved to be one of the best methods to remove solvents out of a solvent producing Clostridium acetobutylicum culture. By using an ionic liquid (IL)-polydimethylsiloxane (PDMS) ultrafiltration membrane (pore size 60 nm), we could guarantee high stability and selectivity during all measurements carried out at 37 degrees C. Overall solvent productivity of fermentation connected with continuous product removal by pervaporation was 2.34 g l(-1) h(-1). The supported ionic liquid membrane (SILM) was impregnated with 15 wt% of a novel ionic liquid (tetrapropylammonium tetracyano-borate) and 85 wt% of polydimethylsiloxane. Pervaporation, accomplished with the optimized SILM, led to stable and efficient removal of the solvents butan-1-ol and acetone out of a C. acetobutylicum culture. By pervaporation through SILM, we removed more butan-1-ol than C. acetobutylicum was able to produce. Therefore, we added an extra dose of butan-1-ol to run fermentation on limiting values where the bacteria would still be able to survive its lethal concentration (15.82 g/l). After pervaporation was switched off, the bacteria died from high concentration of butan-1-ol, which they produced.

  9. Regulation of nitrogen metabolism, starch utilisation and the beta-hbd-adh1 gene cluster in Clostridium acetobutylicum.

    PubMed

    Woods, D R; Reid, S J

    1995-10-01

    The successful genetic manipulation of Clostridium acetobutylicum for the increased production of solvents will depend on an understanding of gene structure and regulation in the bacterium. The glutamine synthetase (glnA) gene is regulated by antisense RNA, transcribed from a downstream promoter, in the opposite direction to the glnA gene. An open reading frame (ORF) was detected downstream of the glnA gene, which has sequence homology to response regulators with anti-termination activity and may be involved in sensing nitrogen conditions. The expression of the linked beta-hbd, adh1 and fixB genes was investigated throughout the bacterial growth cycle by RNA hybridisation techniques. The adh1 gene was independently expressed as a 2.4-kb transcript which peaked at 12 h, immediately prior to the solventogenic phase. The beta-hbd and fixB genes were transcribed throughout the acidogenic and solventogenic phases. A regulator gene, regA, which complements a Bacillus subtilis ccpA mutant, has been identified and sequenced from C. acetobutylicum P262. The regA gene repressed the degradation of starch by an uncharacterised C. acetobutylicum gene, and may therefore play a role in the utilisation of carbohydrate substrates in this organism.

  10. Structural analysis of Clostridium acetobutylicum ATCC 824 glycoside hydrolase from CAZy family GH105

    PubMed Central

    Germane, Katherine L.; Servinsky, Matthew D.; Gerlach, Elliot S.; Sund, Christian J.; Hurley, Margaret M.

    2015-01-01

    Clostridium acetobutylicum ATCC 824 gene CA_C0359 encodes a putative unsaturated rhamnogalacturonyl hydrolase (URH) with distant amino-acid sequence homology to YteR of Bacillus subtilis strain 168. YteR, like other URHs, has core structural homology to unsaturated glucuronyl hydrolases, but hydrolyzes the unsaturated disaccharide derivative of rhamnogalacturonan I. The crystal structure of the recombinant CA_C0359 protein was solved to 1.6 Å resolution by molecular replacement using the phase information of the previously reported structure of YteR (PDB entry 1nc5) from Bacillus subtilis strain 168. The YteR-like protein is a six-α-hairpin barrel with two β-sheet strands and a small helix overlaying the end of the hairpins next to the active site. The protein has low primary protein sequence identity to YteR but is structurally similar. The two tertiary structures align with a root-mean-square deviation of 1.4 Å and contain a highly conserved active pocket. There is a conserved aspartic acid residue in both structures, which has been shown to be important for hydration of the C=C bond during the release of unsaturated galacturonic acid by YteR. A surface electrostatic potential comparison of CA_C0359 and proteins from CAZy families GH88 and GH105 reveals the make-up of the active site to be a combination of the unsaturated rhamnogalacturonyl hydrolase and the unsaturated glucuronyl hydrolase from Bacillus subtilis strain 168. Structural and electrostatic comparisons suggests that the protein may have a slightly different substrate specificity from that of YteR. PMID:26249707

  11. Biobutanol production in a Clostridium acetobutylicum biofilm reactor integrated with simultaneous product recovery by adsorption

    PubMed Central

    2014-01-01

    Background Clostridium acetobutylicum can propagate on fibrous matrices and form biofilms that have improved butanol tolerance and a high fermentation rate and can be repeatedly used. Previously, a novel macroporous resin, KA-I, was synthesized in our laboratory and was demonstrated to be a good adsorbent with high selectivity and capacity for butanol recovery from a model solution. Based on these results, we aimed to develop a process integrating a biofilm reactor with simultaneous product recovery using the KA-I resin to maximize the production efficiency of biobutanol. Results KA-I showed great affinity for butanol and butyrate and could selectively enhance acetoin production at the expense of acetone during the fermentation. The biofilm reactor exhibited high productivity with considerably low broth turbidity during repeated batch fermentations. By maintaining the butanol level above 6.5 g/L in the biofilm reactor, butyrate adsorption by the KA-I resin was effectively reduced. Co-adsorption of acetone by the resin improved the fermentation performance. By redox modulation with methyl viologen (MV), the butanol-acetone ratio and the total product yield increased. An equivalent solvent titer of 96.5 to 130.7 g/L was achieved with a productivity of 1.0 to 1.5 g · L-1 · h-1. The solvent concentration and productivity increased by 4 to 6-fold and 3 to 5-fold, respectively, compared to traditional batch fermentation using planktonic culture. Conclusions Compared to the conventional process, the integrated process dramatically improved the productivity and reduced the energy consumption as well as water usage in biobutanol production. While genetic engineering focuses on strain improvement to enhance butanol production, process development can fully exploit the productivity of a strain and maximize the production efficiency. PMID:24401161

  12. Effect of zinc supplementation on acetone-butanol-ethanol fermentation by Clostridium acetobutylicum.

    PubMed

    Wu, You-Duo; Xue, Chuang; Chen, Li-Jie; Bai, Feng-Wu

    2013-05-10

    In this article, effect of zinc supplementation on acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum was studied. It was found that when 0.001 g/L ZnSO4·7H2O was supplemented into the medium, solventogenesis was initiated earlier, with 21.0 g/L ABE (12.6 g/L butanol, 6.7 g/L acetone and 1.7 g/L ethanol) produced with a fermentation time of 40 h, compared to 19.4 g/L ABE (11.7 g/L butanol, 6.4 g/L acetone and 1.3g/L ethanol) produced with a fermentation time of 64 h in the control without zinc supplementation, and correspondingly ABE and butanol productivities were increased to 0.53 and 0.32 g/L/h from 0.30 and 0.18 g/L/h, increases of 76.7% and 77.8%, respectively, but their yields were not compromised. The reason for this phenomenon was attributed to rapid acids re-assimilation for more efficient ABE production, which was in accordance with relatively high pH and ORP levels maintained during the fermentation process. The maximum cell density increased by 23.8%, indicating that zinc supplementation stimulated cell growth, and consequently facilitated glucose utilization. However, more zinc supplementation exhibited an inhibitory effect, indicating that zinc supplementation at very low levels such as 0.001 g/L ZnSO4·7H2O will be an economically competitive strategy for improving butanol production.

  13. Global transcriptional changes of Clostridium acetobutylicum cultures with increased butanol:acetone ratios.

    PubMed

    Hönicke, Daniel; Janssen, Holger; Grimmler, Christina; Ehrenreich, Armin; Lütke-Eversloh, Tina

    2012-05-15

    Artificial electron carriers have been widely used to shift the solvent ratio toward butanol in acetone-butanol-ethanol (ABE) fermentation of solventogenic clostridia according to decreased hydrogen production. In this study, first insights on the molecular level were gained to explore the effect of methyl viologen addition to cultures of Clostridium acetobutylicum. Employing batch fermentation in mineral salts medium, the butanol:acetone ratio was successively increased from 2.3 to 12.4 on a 100-ml scale in serum bottles and from 1.4 to 16.5 on a 1300-ml scale in bioreactors, respectively. The latter cultures were used for DNA microarray analyses to provide new information on the transcriptional changes referring to methyl viologen exposure and thus, exhibit gene expression patterns according to the manipulation of the cellular redox balance. Methyl viologen-exposed cultures revealed lower expression levels of the sol operon (CAP0162-0164) and the adjacent adc gene (CAP0165) responsible for solvent formation as well as iron and sulfate transporters and the CAC0105-encoded ferredoxin. On the contrary, genes for riboflavin biosynthesis, for the butyrate/butanol metabolic pathway and genes coding for sugar transport systems were induced. Interestingly, the adhE2-encoded bifunctional NADH-dependent aldhehyde/alcohol-dehydrogenase (CAP0035) was upregulated up to more than 100-fold expression levels as compared to the control culture without methyl viologen addition. The data presented here indicate a transcriptional regulation for decreased acetone biosynthesis and the redox-dependent substitution of adhE1 (CAP0162) by adhE2.

  14. The influence of the pentose's pathway of the Clostridium Acetobutylicum on the production of butanol: Insights from mathematical modeling

    NASA Astrophysics Data System (ADS)

    Soares, Rafael Ferreira; da Silva, Fabrício Alves Barbosa; Guimarães, Ana Carolina Ramos; Caffarena, Ernesto Raul

    2016-12-01

    This work is a pilot study for further analysis of the organism Trypanosoma cruzi (T. cruzi) and the influences of the Pentose's Pathway on the parasite Clostridium acetobutylicum, already cataloged in the database of OptFlux program. We used the approach parcimonius Flux Balance Analysis (pFBA) to simulate the wild type organism and the mutant with an inhibition of the R_01056 reaction in pentose's pathway. Results showed a reduction of approximately 1/3 of the biomass and 2/3 of the butanol production. This reduction shows the direct influence of the Pentose's Pathway on the primary production of metabolites and the biomass generation from the Clostridium metabolites. This information prompted us to build in the future an SBML parameter file to represent the flow of T.cruzi pathways, which will be essential for the development of new drugs against.

  15. Recent advances and state-of-the-art strategies in strain and process engineering for biobutanol production by Clostridium acetobutylicum.

    PubMed

    Xue, Chuang; Zhao, Jingbo; Chen, Lijie; Yang, Shang-Tian; Bai, Fengwu

    Butanol as an advanced biofuel has gained great attention due to its environmental benefits and superior properties compared to ethanol. However, the cost of biobutanol production via conventional acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum is not economically competitive, which has hampered its industrial application. The strain performance and downstream process greatly impact the economics of biobutanol production. Although various engineered strains with carefully orchestrated metabolic and sporulation-specific pathways have been developed, none of them is ideal for industrial biobutanol production. For further strain improvement, it is necessary to develop advanced genome editing tools and a deep understanding of cellular functioning of genes in metabolic and regulatory pathways. Processes with integrated product recovery can increase fermentation productivity by continuously removing inhibitory products while generating butanol (ABE) in a concentrated solution. In this review, we provide an overview of recent advances in C. acetobutylicum strain engineering and process development focusing on in situ product recovery. With deep understanding of systematic cellular bioinformatics, the exploration of state-of-the-art genome editing tools such as CRISPR-Cas for targeted gene knock-out and knock-in would play a vital role in Clostridium cell engineering for biobutanol production. Developing advanced hybrid separation processes for in situ butanol recovery, which will be discussed with a detailed comparison of advantages and disadvantages of various recovery techniques, is also imperative to the economical development of biobutanol.

  16. Production of an acetone-butanol-ethanol mixture from Clostridium acetobutylicum and its conversion to high-value biofuels.

    PubMed

    Sreekumar, Sanil; Baer, Zachary C; Pazhamalai, Anbarasan; Gunbas, Gorkem; Grippo, Adam; Blanch, Harvey W; Clark, Douglas S; Toste, F Dean

    2015-03-01

    Clostridium acetobutylicum is a bacterial species that ferments sugar to a mixture of organic solvents (acetone, butanol and ethanol). This protocol delineates a methodology to combine solventogenic clostridial fermentation and chemical catalysis via extractive fermentation for the production of biofuel blendstocks. Extractive fermentation of C. acetobutylicum is operated in fed-batch mode with a concentrated feed solution (500 grams per liter glucose and 50 grams per liter yeast extract) for 60 h, producing in excess of 40 g of solvents (acetone, butanol and ethanol) between the completely immiscible extractant and aqueous phases of the bioreactor. After distillation of the extractant phase, the acetone, butanol and ethanol mixture is upgraded to long-chain ketones over a palladium-hydrotalcite (Pd-HT) catalyst. This reaction is generally carried out in batch with a high-pressure Q-tube for 20 h at 250 °C. Following this protocol enables the production of ∼0.5 g of high-value biofuel precursors from a 1.7-g portion of fermentation solvents.

  17. Multiple orphan histidine kinases interact directly with Spo0A to control the initiation of endospore formation in Clostridium acetobutylicum

    PubMed Central

    Steiner, Elisabeth; Dago, Angel E.; Young, Danielle I.; Heap, John T.; Minton, Nigel P.; Hoch, James A.

    2011-01-01

    The phosphorylated Spo0A transcription factor controls the initiation of endospore formation in Clostridium acetobutylicum, but genes encoding key phosphorelay components, Spo0F and Spo0B, are missing in the genome. We hypothesized that the five orphan histidine kinases of C. acetobutylicum interact directly with Spo0A to control its phosphorylation state. Sequential targeted gene disruption and gene expression profiling provided evidence for two pathways for Spo0A activation, one dependent on a histidine kinase encoded by cac0323, the other on both histidine kinases encoded by cac0903 and cac3319. Purified Cac0903 and Cac3319 kinases autophosphorylated and transferred phosphoryl groups to Spo0A in vitro, confirming their role in Spo0A activation in vivo. A cac0437 mutant hyper-sporulated, suggesting that Cac0437 is a modulator that prevents sporulation and maintains cellular Spo0A~P homeostasis during growth. Accordingly, Cac0437 has apparently lost the ability to autophosphorylate in vitro; instead it catalyses the ATP-dependent dephosphorylation of Spo0A~P releasing inorganic phosphate. Direct phosphorylation of Spo0A by histidine kinases and dephosphorylation by kinase-like proteins may be a common feature of the clostridia that may represent the ancestral state before the great oxygen event some 2.4 billion years ago, after which additional phosphorelay proteins were recruited in the evolutionary lineage that led to the bacilli. PMID:21401736

  18. Chemostat cultivation and transcriptional analyses of Clostridium acetobutylicum mutants with defects in the acid and acetone biosynthetic pathways.

    PubMed

    Hönicke, Daniel; Lütke-Eversloh, Tina; Liu, Ziyong; Lehmann, Dörte; Liebl, Wolfgang; Ehrenreich, Armin

    2014-12-01

    Clostridium acetobutylicum is a model organism for the biotechnologically important acetone-butanol-ethanol (ABE) fermentation. With the objective to rationally develop strains with improved butanol production, detailed insights into the physiological and genetic mechanisms of solvent production are required. Therefore, pH-controlled phosphate-limited chemostat cultivation and DNA microarray technology were employed for an in-depth analysis of knockout mutants with defects in the central fermentative metabolism. The set of studied mutants included strains with inactivated phosphotransacetylase (pta), phosphotransbutyrylase (ptb), and acetoacetate decarboxylase (adc) encoding genes, as well as an adc/pta double knockout mutant. A comprehensive physiological characterization of the mutants was performed by continuous cultivation, allowing for a well-defined separation of acidogenic and solventogenic growth, combined with the advantage of the high reproducibility of steady-state conditions. The ptb-negative strain C. acetobutylicum ptb::int(87) exhibited the most striking metabolite profile: Sizable amounts of butanol (29 ± 1.3 mM) were already produced during acidogenic growth. The product patterns of the mutants as well as accompanying transcriptomic data are presented and discussed.

  19. Modifying the product pattern of Clostridium acetobutylicum: physiological effects of disrupting the acetate and acetone formation pathways.

    PubMed

    Lehmann, Dörte; Hönicke, Daniel; Ehrenreich, Armin; Schmidt, Michael; Weuster-Botz, Dirk; Bahl, Hubert; Lütke-Eversloh, Tina

    2012-05-01

    Clostridial acetone-butanol-ethanol (ABE) fermentation is a natural source for microbial n-butanol production and regained much interest in academia and industry in the past years. Due to the difficult genetic accessibility of Clostridium acetobutylicum and other solventogenic clostridia, successful metabolic engineering approaches are still rare. In this study, a set of five knock-out mutants with defects in the central fermentative metabolism were generated using the ClosTron technology, including the construction of targeted double knock-out mutants of C. acetobtuylicum ATCC 824. While disruption of the acetate biosynthetic pathway had no significant impact on the metabolite distribution, mutants with defects in the acetone pathway, including both acetoacetate decarboxylase (Adc)-negative and acetoacetyl-CoA:acyl-CoA transferase (CtfAB)-negative mutants, exhibited high amounts of acetate in the fermentation broth. Distinct butyrate increase and decrease patterns during the course of fermentations provided experimental evidence that butyrate, but not acetate, is re-assimilated via an Adc/CtfAB-independent pathway in C. acetobutylicum. Interestingly, combining the adc and ctfA mutations with a knock-out of the phosphotransacetylase (Pta)-encoding gene, acetate production was drastically reduced, resulting in an increased flux towards butyrate. Except for the Pta-negative single mutant, all mutants exhibited a significantly reduced solvent production.

  20. Transcriptional analysis of micronutrient zinc-associated response for enhanced carbohydrate utilization and earlier solventogenesis in Clostridium acetobutylicum

    PubMed Central

    Wu, You-Duo; Xue, Chuang; Chen, Li-Jie; Wan, Hui-Hui; Bai, Feng-Wu

    2015-01-01

    The micronutrient zinc plays vital roles in ABE fermentation by Clostridium acetobutylicum. In order to elucidate the zinc-associated response for enhanced glucose utilization and earlier solventogenesis, transcriptional analysis was performed on cells grown in glucose medium at the exponential growth phase of 16 h without/with supplementary zinc. Correspondingly, the gene glcG (CAC0570) encoding a glucose-specific PTS was significantly upregulated accompanied with the other two genes CAC1353 and CAC1354 for glucose transport in the presence of zinc. Additionally, genes involved in the metabolisms of six other carbohydrates (maltose, cellobiose, fructose, mannose, xylose and arabinose) were differentially expressed, indicating that the regulatory effect of micronutrient zinc is carbohydrate-specific with respects to the improved/inhibited carbohydrate utilization. More importantly, multiple genes responsible for glycolysis (glcK and pykA), acidogenesis (thlA, crt, etfA, etfB and bcd) and solventogenesis (ctfB and bdhA) of C. acetobutylicum prominently responded to the supplementary zinc at differential expression levels. Comparative analysis of intracellular metabolites revealed that the branch node intermediates such as acetyl-CoA, acetoacetyl-CoA, butyl-CoA, and reducing power NADH remained relatively lower whereas more ATP was generated due to enhanced glycolysis pathway and earlier initiation of solventogenesis, suggesting that the micronutrient zinc-associated response for the selected intracellular metabolisms is significantly pleiotropic. PMID:26586044

  1. Fermentation characterization and flux analysis of recombinant strains of Clostridium acetobutylicum with an inactivated solR gene.

    PubMed

    Harris, L M; Blank, L; Desai, R P; Welker, N E; Papoutsakis, E T

    2001-11-01

    The effect of solR inactivation on the metabolism of Clostridium acetobutylicum was examined using fermentation characterization and metabolic flux analysis. The solR-inactivated strain (SolRH) of this study had a higher rate of glucose utilization and produced higher solvent concentrations (by 25%, 14%, and 81%, respectively, for butanol, acetone, and ethanol) compared to the wild type. Strain SolRH(pTAAD), carrying a plasmid-encoded copy of the bifunctional alcohol/aldehyde dehydrogenase gene (aad) used in butanol production, produced even higher concentrations of solvents (by 21%, 45%, and 62%, respectively, for butanol, acetone, and ethanol) than strain SolRH. Clarithromycin used for strain SolRH maintenance during SolRH(pTAAD) fermentations did not alter product formation; however, tetracycline used for pTAAD maintenance resulted in 90% lower solvent production.

  2. Assessment of morphological changes of Clostridium acetobutylicum by flow cytometry during acetone/butanol/ethanol extractive fermentation.

    PubMed

    González-Peñas, Helena; Lu-Chau, Thelmo Alejandro; Moreira, Maria Teresa; Lema, Juan Manuel

    2015-03-01

    Acetone/butanol/ethanol (ABE) fermentation by Clostridium acetobutylicum was investigated in extractive fed-batch experiments. In conventional fermentations, metabolic activity ceases when a critical threshold products concentration is reached (~21.6 g solvents l(-1)). Solvents production was increased up to 36.6 and 37.2 g l(-1), respectively, using 2-butyl-1-octanol (aqueous to organic ratio: 1:0.25 v/v) and pomace olive oil (1:1 v/v) as extraction solvents. The morphological changes of different cell types were monitored and quantified using flow cytometry. Butanol production in extractive fermentations with pomace olive oil was achieved mainly by vegetative cells, whereas the percentage of sporulating cells was lower than 10%.

  3. Effect of pH and lactose concentration on solvent production from whey permeate using Clostridium acetobutylicum

    SciTech Connect

    Ennis, B.M.; Maddox, I.S.

    1987-02-20

    A study was performed to optimize the production of solvents from whey permeate in batch fermentation using Clostridium acetobutylicum P262. Fermentations performed at relatively low pH values resulted in high solvent yields and productivities, but lactose utilization was incomplete. At higher pH values, lactose-utilization was improved but acid production dominated over solvent production. When operating at the higher pH values, an increase in the initial lactose concentration of the whey permeate resulted in lower rates of lactose utilization, and this was accompanied by increased solvent production and decreased acid production. Analysis of data from several experiments revealed a strong inverse relationship between solvent yield and lactose utilization rate. Thus, conditions which minimize the lactose utilization rate such as low culture pH values or high initial lactose concentrations, favor solventogenesis at the expense of acid production. 12 references.

  4. Effect of cocultures on the production of butanol by Clostridium sp. [C. butyricum, C. pasteurianum, C. butylicum, and C. acetobutylicum

    SciTech Connect

    Bergstrom, S.L.; Foutch, G.L.

    1983-01-01

    One of the problems with the production of butanol by fermentation is that a mixture of other solvents, primarily acetone and ethanol, are also produced. The ratio of butanol to acetone to ethanol is approximately 6:3:1. If the metabolism of the Clostridium can be shifted away from acetone and ethanol, then a higher percentage of the sugar carbon can be converted to butanol. This study examines the effects that cocultures of Clostridium have on the yield of butanol from glucose. C. butyricum and C. pasteurianum produced high concentrations of butyric acid from these sugars. C. butylicum and C. acetobutylicum are capable of utilizing this butyric acid for energy, resulting in butanol production. By using combinations of these strains in coculture a higher initial conversion to butyric acid would be expected followed by a higher overall yield of butanol. This coculture would not be expected to have an increased tolerance for butanol in the fermentation broth. 6 references, 1 figure, 3 tables.

  5. Transcriptional Analysis of spo0A Overexpression in Clostridium acetobutylicum and Its Effect on the Cell's Response to Butanol Stress

    PubMed Central

    Alsaker, Keith V.; Spitzer, Thomas R.; Papoutsakis, Eleftherios T.

    2004-01-01

    Spo0A is the regulator of stationary-phase events and is required for transcription of solvent formation genes in Clostridium acetobutylicum. In order to elucidate the role of spo0A in differentiation, we performed transcriptional analysis of 824(pMSPOA) (a spo0A-overexpressing C. acetobutylicum strain with enhanced sporulation) against a plasmid control strain. DNA microarray data were contrasted to data from a spo0A knockout strain (SKO1) that neither sporulates nor produces solvents. Transcripts of fatty acid metabolism genes, motility and chemotaxis genes, heat shock protein genes, and genes encoding the Fts family of cell division proteins were differentially expressed in the two strains, suggesting that these genes play roles in sporulation and the solvent stress response. 824(pMSPOA) alone showed significant downregulation of many glycolytic genes in stationary phase, which is consistent with metabolic flux analysis data. Surprisingly, spo0A overexpression resulted in only nominal transcriptional changes of regulatory genes (abrB and sigF) whose expression was significantly altered in SKO1. Overexpression of spo0A imparted increased tolerance and prolonged metabolism in response to butanol stress. While most of the differentially expressed genes appear to be part of a general stress response (similar to patterns in two plasmid control strains and a groESL-overexpressing strain), several genes were expressed at higher levels at early time points after butanol challenge only in 824(pMSPOA). Most of these genes were related to butyryl coenzyme A and butyrate formation and/or assimilation, but they also included the cell division gene ftsX, the gyrase subunit-encoding genes gyrB and gyrA, DNA synthesis and repair genes, and fatty acid synthesis genes, all of which might play a role in the immediate butanol stress response, and thus in enhanced butanol tolerance. PMID:15028679

  6. Elucidating the contributions of multiple aldehyde/alcohol dehydrogenases to butanol and ethanol production in Clostridium acetobutylicum

    PubMed Central

    Dai, Zongjie; Dong, Hongjun; Zhang, Yanping; Li, Yin

    2016-01-01

    Ethanol and butanol biosynthesis in Clostridium acetobutylicum share common aldehyde/alcohol dehydrogenases. However, little is known about the relative contributions of these multiple dehydrogenases to ethanol and butanol production respectively. The contributions of six aldehyde/alcohol dehydrogenases of C. acetobutylicum on butanol and ethanol production were evaluated through inactivation of the corresponding genes respectively. For butanol production, the relative contributions from these enzymes were: AdhE1 > BdhB > BdhA ≈ YqhD > SMB_P058 > AdhE2. For ethanol production, the contributions were: AdhE1 > BdhB > YqhD > SMB_P058 > AdhE2 > BdhA. AdhE1 and BdhB are two essential enzymes for butanol and ethanol production. AdhE1 was relatively specific for butanol production over ethanol, while BdhB, YqhD, and SMB_P058 favor ethanol production over butanol. Butanol synthesis was increased in the adhE2 mutant, which had a higher butanol/ethanol ratio (8.15:1) compared with wild type strain (6.65:1). Both the SMB_P058 mutant and yqhD mutant produced less ethanol without loss of butanol formation, which led to higher butanol/ethanol ratio, 10.12:1 and 10.17:1, respectively. To engineer a more efficient butanol-producing strain, adhE1 could be overexpressed, furthermore, adhE2, SMB_P058, yqhD are promising gene inactivation targets. This work provides useful information guiding future strain improvement for butanol production. PMID:27321949

  7. Efficient acetone-butanol-ethanol production (ABE) by Clostridium acetobutylicum XY16 immobilized on chemically modified sugarcane bagasse.

    PubMed

    Kong, Xiangping; He, Aiyong; Zhao, Jie; Wu, Hao; Jiang, Min

    2015-07-01

    Sugarcane bagasse was chemically modified by polyethylenimine (PEI) and glutaraldehyde (GA) and then used as a support to immobilize Clostridium acetobutylicum XY16 in the process of butanol production. Compared with batch fermentation using unmodified sugarcane bagasse, 22.3 g/L total solvents were produced by cells immobilized on 4 g/L PEI treated sugarcane bagasse with high solvent productivity of 0.62 g/(L h) and glucose consumption rate of 1.67 g/(L h). Improvement of 14, 43, and 37 % in total solvent titer, solvent productivity and glucose consumption rate was observed, respectively. Enhanced solvent production of 25.14 g/L was obtained when using a high concentration of glucose of 80 g/L. Continuous fermentation was studied using PEI/GA modified sugarcane bagasse as immobilization support with a range of dilution which rates from 0.2 to 2.5 to find an optimal condition. The maximum solvent productivity of 11.32 g/(L h) was obtained at a high dilution rate of 2.0 h(-1).

  8. A Standard Operating Procedure (SOP) for the preparation of intra- and extracellular proteins of Clostridium acetobutylicum for proteome analysis.

    PubMed

    Schwarz, Katrin; Fiedler, Tomas; Fischer, Ralf-Jörg; Bahl, Hubert

    2007-02-01

    We report on the development of a Standard Operating Procedure (SOP) for extraction and handling of intra- and extracellular protein fractions of Clostridium acetobutylicum ATCC 824 for reproducible high quality two-dimensional gel electrophoresis (2-DE) analyses. Standardized cells from a phosphate-limited chemostat were used to evaluate different protein preparation methods. For the preparation of the secretome, a dialysis/ultrafiltration procedure resulted in higher protein yields and proved to be more reliable compared to different precipitation methods using TCA, DOC-TCA, acetone, and PEG 6000. Sonication was found to be the most efficient method among different tested techniques of cell disruption for the analysis of the intracellular proteome. Furthermore, the effect of protease inhibitors and sample storage conditions were tested for both intra- and extracellular protein samples. Significant changes in the protein pattern were observed depending on the addition of protease inhibitors. 2-DE gels with a pH gradient from 4 to 7 prepared according to the developed SOP contained at least 736 intracellular and 324 extracellular protein spots.

  9. Integrated bioprocess for long-term continuous cultivation of Clostridium acetobutylicum coupled to pervaporation with PDMS composite membranes.

    PubMed

    Van Hecke, Wouter; Vandezande, Pieter; Claes, Stan; Vangeel, Silvia; Beckers, Herman; Diels, Ludo; De Wever, Heleen

    2012-05-01

    A continuous cultivation of Clostridium acetobutylicum ATCC 824 is described using a two-stage design to mimic the two phases of batch culture growth of the organism. A hydrophobic pervaporation unit was coupled to the second fermentor containing the highest solvent titers. This in situ product recovery technology efficiently decreased butanol toxicity in the fermentor while the permeate was enriched to 57-195 g L(-1) total solvents depending on the solvent concentrations in the fermentor. By the alleviation of product inhibition, the glucose concentration could be increased from 60 to 126 g L(-1) while the productivity increased concomitantly from 0.13 to 0.30 g L(-1)h(-1). The continuous fermentation was conducted for 1172 h during which the pervaporation was coupled to the second fermentor for 475 h with an average flux of 367 g m(-2)h(-1). The energy consumption was calculated for a 2 wt.% n-butanol fermentation broth and compared with the conventional process.

  10. Continuous two stage acetone-butanol-ethanol fermentation with integrated solvent removal using Clostridium acetobutylicum B 5313.

    PubMed

    Bankar, Sandip B; Survase, Shrikant A; Singhal, Rekha S; Granström, Tom

    2012-02-01

    The objective of this study was to optimize continuous acetone-butanol-ethanol (ABE) fermentation using a two stage chemostat system integrated with liquid-liquid extraction of solvents produced in the first stage. This minimized end product inhibition by butanol and subsequently enhanced glucose utilization and solvent production in continuous cultures of Clostridium acetobutylicum B 5313. During continuous two-stage ABE fermentation, sugarcane bagasse was used as the cell holding material for the both stages and liquid-liquid extraction was performed using an oleyl alcohol and decanol mixture. An overall solvent production of 25.32g/L (acetone 5.93g/L, butanol 16.90g/L and ethanol 2.48g/L) was observed as compared to 15.98g/L in the single stage chemostat with highest solvent productivity and solvent yield of 2.5g/Lh and of 0.35g/g, respectively. Maximum glucose utilization (83.21%) at a dilution rate of 0.051/h was observed as compared to 54.38% in the single stage chemostat.

  11. Crystal structure of Clostridium acetobutylicum Aspartate kinase (CaAK): An important allosteric enzyme for amino acids production.

    PubMed

    Manjasetty, Babu A; Chance, Mark R; Burley, Stephen K; Panjikar, Santosh; Almo, Steven C

    2014-09-01

    Aspartate kinase (AK) is an enzyme which is tightly regulated through feedback control and responsible for the synthesis of 4-phospho-L-aspartate from L-aspartate. This intermediate step is at an important branch point where one path leads to the synthesis of lysine and the other to threonine, methionine and isoleucine. Concerted feedback inhibition of AK is mediated by threonine and lysine and varies between the species. The crystal structure of biotechnologically important Clostridium acetobutylicum aspartate kinase (CaAK; E.C. 2.7.2.4; Mw=48,030Da; 437aa; SwissProt: Q97MC0) has been determined to 3Å resolution. CaAK acquires a protein fold similar to the other known structures of AKs despite the low sequence identity (<30%). It is composed of two domains: an N-terminal catalytic domain (kinase) domain and a C-terminal regulatory domain further comprised of two small domains belonging to the ACT domain family. Pairwise comparison of 12 molecules in the asymmetric unit helped to identify the bending regions which are in the vicinity of ATP binding site involved in domain movements between the catalytic and regulatory domains. All 12 CaAK molecules adopt fully open T-state conformation leading to the formation of three tetramers unique among other similar AK structures. On the basis of comparative structural analysis, we discuss tetramer formation based on the large conformational changes in the catalytic domain associated with the lysine binding at the regulatory domains. The structure described herein is homologous to a target in wide-spread pathogenic (toxin producing) bacteria such as Clostridium tetani (64% sequence identity) suggesting the potential of the structure solved here to be applied for modeling drug interactions. CaAK structure may serve as a guide to better understand and engineer lysine biosynthesis for the biotechnology industry.

  12. Synergistic dark and photo-fermentation continuous system for hydrogen production from molasses by Clostridium acetobutylicum ATCC 824 and Rhodobacter capsulatus DSM 1710.

    PubMed

    Morsy, Fatthy Mohamed

    2017-04-01

    This study investigated synergistic dark and photo-fermentation using continuous fermentation system (CFS). The system relies on connecting several fermenters from bottom of one to top culture level of the next in a manner that allows for delaying movement of the substrate and thus for its full consumption. While H2 was collected, CFS allowed for moving liquid byproducts toward the outlet and hence continuous productivity. CFS could be efficiently used for: (1) Continuous dark and photo-fermentation H2 production by Clostridium acetobutylicum and Rhodobacter capsulatus producing 5.65moleH2mole(-1) hexose; (2) Continuous dark-fermentation synergistic H2, acetone, butanol and ethanol (ABE) production by C. acetobutylicum which produced per mole hexose, 2.43mol H2 along with 73.08g ABE (3) Continuous H2 and methane production by C. acetobutylicum and bacterial sludge producing, per mole hexose, 1.64mol pure H2 and 2.56mol CH4 mixed with 0.37mol H2·The hydraulic retention time (HRT) for whole system was short where organic acids produced in dark-fermentation in first fermenter were synergistically utilized for H2 production by R. capsulatus in subsequent fermenters. CFS is suitable for fast-digestible sugars but not lignocelluloses or other hard-digestible organics, requiring prolonged HRT, unless such polymeric organics were hydrolyzed prior to fermentation.

  13. The mechanism of switching from an acidogenic to butanol-acetone fermentation by Clostridium acetobutylicum. Technical progress report, July 1990--June 1993

    SciTech Connect

    Rogers, P.

    1994-11-01

    The overall objective of this project was to elucidate the detailed mechanism by which solvent-forming bacteria such as Clostridium acetobutylicum regulate the well-known shift in fermentation pathway between alcohol-acetone and organic acid production. We eventually want to isolate and describe: (1) the regulatory genes and protein elements that determine induction of synthesis of the solvent-pathway enzymes; and (2) how this regulation system interacts with the sporulation induction and development program and with related pathways such as granulose and exopolysaccharide formation in clostridia. A working model for how clostridial control systems work can be derived from recent research on stress systems in E. coli and sporulation in Bacillus subtilis. This research was centered upon the technique of employing transposable elements that create gene fusions and mutations due to insertion in the chromosome of gram positive bacteria. Our approach was based on recent demonstration in our laboratory and by others of transconjugation of Tn916 into C. acetobutylicum and its insertion into the chromosome. A panel of strains with Tn916 inserts that are also solvent-negative and/or asporogenic were used to identify specific regulatory genes. A second approach was based upon electroporative transformation of plasmid PTV1 DNA carrying transposon Tn917 into C. acetobutylicum. Insertion of Tn917 lac to report activity of genes and functions in vegetative and stationary or slow-growing cells will be investigated.

  14. Enhancing acetone biosynthesis and acetone-butanol-ethanol fermentation performance by co-culturing Clostridium acetobutylicum/Saccharomyces cerevisiae integrated with exogenous acetate addition.

    PubMed

    Luo, Hongzhen; Ge, Laibing; Zhang, Jingshu; Ding, Jian; Chen, Rui; Shi, Zhongping

    2016-01-01

    Acetone is the major by-product in ABE fermentations, most researches focused on increasing butanol/acetone ratio by decreasing acetone biosynthesis. However, economics of ABE fermentation industry strongly relies on evaluating acetone as a valuable platform chemical. Therefore, a novel ABE fermentation strategy focusing on bio-acetone production by co-culturing Clostridium acetobutylicum/Saccharomyces cerevisiae with exogenous acetate addition was proposed. Experimental and theoretical analysis revealed the strategy could, enhance C. acetobutylicum survival oriented amino acids assimilation in the cells; control NADH regeneration rate at moderately lower level to enhance acetone synthesis but without sacrificing butanol production; enhance the utilization ability of C. acetobutylicum on glucose and direct most of extra consumed glucose into acetone/butanol synthesis routes. By implementing the strategy using synthetic or acetate fermentative supernatant, acetone concentrations increased to 8.27-8.55g/L from 5.86g/L of the control, while butanol concentrations also elevated to the higher levels of 13.91-14.23g/L from 11.63g/L simultaneously.

  15. Isolation of Clostridium tetani from anaerobic empyema.

    PubMed

    Mayall, B C; Snashall, E A; Peel, M M

    1998-11-01

    We report the isolation of Clostridium tetani (along with Fusobacterium mortiferum) from empyema pus. The patient, a 68 year old retired farmer from rural NSW, had recently undergone cholecystectomy, had heart failure and developed an empyema. He improved after drainage of the empyema and penicillin therapy, but died suddenly during convalescence.

  16. Structural Characterization of Clostridium acetobutylicum 8-Oxoguanine DNA Glycosylase in Its Apo Form and in Complex with 8-Oxodeoxyguanosine

    SciTech Connect

    Faucher, Frédérick; Robey-Bond, Susan M.; Wallace, Susan S.; Doublié, Sylvie

    2009-06-30

    DNA is subject to a multitude of oxidative damages generated by oxidizing agents from metabolism and exogenous sources and by ionizing radiation. Guanine is particularly vulnerable to oxidation, and the most common oxidative product 8-oxoguanine (8-oxoG) is the most prevalent lesion observed in DNA molecules. 8-OxoG can form a normal Watson-Crick pair with cytosine (8-oxoG:C), but it can also form a stable Hoogsteen pair with adenine (8-oxoG:A), leading to a G:C {yields} T:A transversion after replication. Fortunately, 8-oxoG is recognized and excised by either of two DNA glycosylases of the base excision repair pathway: formamidopyrimidine-DNA glycosylase and 8-oxoguanine DNA glycosylase (Ogg). While Clostridium acetobutylicum Ogg (CacOgg) DNA glycosylase can specifically recognize and remove 8-oxoG, it displays little preference for the base opposite the lesion, which is unusual for a member of the Ogg1 family. This work describes the crystal structures of CacOgg in its apo form and in complex with 8-oxo-2'-deoxyguanosine. A structural comparison between the apo form and the liganded form of the enzyme reveals a structural reorganization of the C-terminal domain upon binding of 8-oxoG, similar to that reported for human OGG1. A structural comparison of CacOgg with human OGG1, in complex with 8-oxoG containing DNA, provides a structural rationale for the lack of opposite base specificity displayed by CacOgg.

  17. Clostridium acetobutylicum 8-Oxoguanine DNA Glycosylase (Ogg) Differs from Eukaryotic Oggs with Respect to Opposite Base Discrimination†

    PubMed Central

    Robey-Bond, Susan M.; Barrantes-Reynolds, Ramiro; Bond, Jeffrey P.; Wallace, Susan S.; Bandaru, Viswanath

    2008-01-01

    During repair of damaged DNA, the oxidized base 8-oxoguanine (8-oxoG) is removed by 8-oxoguanine—DNA glycosylase (Ogg) in eukaryotes and most archaea, whereas in most bacteria it is removed by formamidopyrimidine—DNA glycosylase (Fpg). We report the first characterization of a bacterial Ogg, Clostridium acetobutylicum Ogg (CacOgg). Like human OGG1 and Escherichia coli Fpg (EcoFpg), CacOgg excised 8-oxoguanine. However, unlike hOGG1 and EcoFpg, CacOgg showed little preference for the base opposite the damage during base excision and removed 8-oxoguanine from single-stranded DNA. Thus, our results showed unambiguous qualitative functional differences in vitro between CacOgg and both hOGG1 and EcoFpg. CacOgg differs in sequence from the eukaryotic enzymes at two sequence positions, M132 and F179, which align with amino acids (R154 and Y203) in human OGG1 (hOGG1) found to be involved in opposite base interaction. To address the sequence basis for functional differences with respect to opposite base interactions, we prepared three CacOgg variants, M132R, F179Y, and M132R/F179Y. All three variants showed a substantial increase in specificity for 8-oxoG·C relative to 8-oxoG·A. While we were unable to definitively associate these qualitative functional differences with differences in selective pressure between eukaryotes, Clostridia, and other bacteria, our results are consistent with the idea that evolution of Ogg function is based on kinetic control of repair. PMID:18578506

  18. Introducing a single secondary alcohol dehydrogenase into butanol-tolerant Clostridium acetobutylicum Rh8 switches ABE fermentation to high level IBE fermentation

    PubMed Central

    2012-01-01

    Background Previously we have developed a butanol tolerant mutant of Clostridium acetobutylicum Rh8, from the wild type strain DSM 1731. Strain Rh8 can tolerate up to 19 g/L butanol, with solvent titer improved accordingly, thus exhibiting industrial application potential. To test if strain Rh8 can be used for production of high level mixed alcohols, a single secondary alcohol dehydrogenase from Clostridium beijerinckii NRRL B593 was overexpressed in strain Rh8 under the control of thl promoter. Results The heterogenous gene sADH was functionally expressed in C. acetobutylicum Rh8. This simple, one-step engineering approach switched the traditional ABE (acetone-butanol-ethanol) fermentation to IBE (isopropanol-butanol-ethanol) fermentation. The total alcohol titer reached 23.88 g/l (7.6 g/l isopropanol, 15 g/l butanol, and 1.28 g/l ethanol) with a yield to glucose of 31.42%. The acid (butyrate and acetate) assimilation rate in isopropanol producing strain Rh8(psADH) was increased. Conclusions The improved butanol tolerance and the enhanced solvent biosynthesis machinery in strain Rh8 is beneficial for production of high concentration of mixed alcohols. Strain Rh8 can thus be considered as a good host for further engineering of solvent/alcohol production. PMID:22742819

  19. Mutant strain of C. acetobutylicum and process for making butanol

    DOEpatents

    Jain, Mahendra K.; Beacom, Daniel; Datta, Rathin

    1993-01-01

    A biologically pure asporogenic mutant of Clostridium acetobutylicum is produced by growing sporogenic C. acetobutylicum ATCC 4259 and treating the parent strain with ethane methane sulfonate. The mutant which as been designated C. acetobutylicum ATCC 55025 is useful in an improved ABE fermentation process, and produces high concentrations of butanol and total solvents.

  20. Northern, morphological, and fermentation analysis of spo0A inactivation and overexpression in Clostridium acetobutylicum ATCC 824.

    PubMed

    Harris, Latonia M; Welker, Neil E; Papoutsakis, Eleftherios T

    2002-07-01

    The Clostridium acetobutylicum ATCC 824 spo0A gene was cloned, and two recombinant strains were generated, an spo0A inactivation strain (SKO1) and an spo0A overexpression strain [824(pMPSOA)]. SKO1 was developed by targeted gene inactivation with a replicative plasmid capable of double-crossover chromosomal integration--a technique never used before with solventogenic clostridia. SKO1 was severely deficient in solvent formation: it produced only 2 mM acetone and 13 mM butanol, compared to the 92 mM acetone and 172 mM butanol produced by the parental strain. After 72 h of growth on solid media, SKO1 formed long filaments of rod-shaped cells that failed to septate. SKO1 cells never achieved the swollen clostridial form typical of the parental strain and did not form endospores. No spo0A transcripts were detected in SKO1, while transcription of two solvent formation operons (aad-ctfA-ctfB and adc; both containing 0A boxes in their promoter regions) was limited. Strain 824(pMSPOA) produced higher butanol concentrations than the control strain [824(pIMP1)] and dramatically elevated spo0A transcript levels and displayed a bimodal pattern of spo0A transcription similar to that of B. subtilis. Microscopic studies indicated that sporulation was both enhanced and accelerated due to spo0A overexpression compared to that of both the 824(pIMP1) and parental strains. Consistent with that, expression of the key solvent formation genes (aad-ctfA-ctfB and adc) and three sporulation-specific genes (spoIIGA, sigE, and sigG) was observed earlier in strain 824(pMSPOA) than in the plasmid control. These data support the hypothesis that Spo0A is a transcriptional regulator that positively controls sporulation and solvent production. Its effect on solvent formation is a balancing act in regulating sporulation versus solvent gene expression: its overexpression apparently tips the balance in favor of accelerated and enhanced sporulation at the expense of overall solvent production.

  1. Cloning and expression of clostridium acetobutylicum ATCC 824 acetoacetyl-coenzyme A:acetate/butyrate:coenzyme A-transferase in Escherichia coli

    SciTech Connect

    Cary, J.W.; Petersen, D.J.; Bennett, G.N. ); Papoutsakis, E.T. )

    1990-06-01

    Coenzyme A (CoA)-transferase (acetoacetyl-CoA:acetate/butyrate:CoA-transferase (butyrate-acetoacetate CoA-transferase) (EC 2.8.3.9)) of Clostridium acetobutylicum ATCC 824 is an important enzyme in the metabolic shift between the acid-producing and solvent-forming states of this organism. The genes encoding the two subunits of this enzyme have been cloned and subsequent subcloning experiments established the position of the structural genes for CoA-transferase. Complementation of Escherichia coli ato mutants with the recombinant plasmid pCoAT4 (pUC19 carrying a 1.8-kilobase insert of C. acetobutylicum DNA encoding CoA-transferase activity) enabled the transformants to grow on butyrate as a sole carbon source. Despite the ability of CoA-transferase to complement the ato defect in E. coli mutants, Southern blot and Western blot (immunoblot) analyses showed showed that neither the C. acetobutylicum genes encoding CoA-transferase nor the enzyme itself shared any apparent homology with its E. coli counterpart. Polypeptides of M{sub r} of the purified CoA-transferase subunits were observed by Western blot and maxicell analysis of whole-cell extracts of E.coli harboring pCoAT4. The proximity and orientation of the genes suggest that the genes encoding the two subunits of CoA-transferase may form an operon similar to that found in E. coli. In the plasmid, however, transcription appears to be primarily from the lac promoter of the vector.

  2. Cloning, expression, and purification of glutamine synthetase from Clostridum acetobutylicum

    SciTech Connect

    Usdin, K.P.; Zappe, H.; Jones, D.T.; Woods, D.R.

    1986-09-01

    A glutamine synthetase (GS) gene, glnA, from the gram-positive obligate anaerobe Clostridium acetobutylicum was cloned on recombinant plasmid pHZ200 and enabled Escherichia coli glnA deletion mutants to utilize (NH/sub 4/)/sub 2/ as a sole source of nitrogen. The cloned C. acetobutylicum gene was expressed from a regulatory region contained within the cloned DNA fragment. glnA expression was subject to nitrogen regulation in E. coli. This cloned glnA DNA did not enable an E. coli glnA ntrB ntrC deletion mutant to utilize arginine or low levels of glutamine as sole nitrogen sources, and failed to activate histidase activity in this strain which contained the Klebsiella aerogenes hut operon. The GS produced by pHZ200 was purified and had an apparent subunit molecular weight of approximately 59,000. There was no DNA or protein homology between the cloned C. acetobutylicum glnA gene and GS and the corresponding gene and GS from E. coli. The C. acetobutylicum GS was inhibited by Mg/sup 2 +/ in the ..gamma..-glutamyl transferase assay, but there was no evidence that the GS was adenylylated.

  3. Regulation of the sol locus genes for butanol and acetone formation in Clostridium acetobutylicum ATCC 824 by a putative transcriptional repressor.

    PubMed

    Nair, R V; Green, E M; Watson, D E; Bennett, G N; Papoutsakis, E T

    1999-01-01

    A gene (orf1, now designated solR) previously identified upstream of the aldehyde/alcohol dehydrogenase gene aad (R. V. Nair, G. N. Bennett, and E. T. Papoutsakis, J. Bacteriol. 176:871-885, 1994) was found to encode a repressor of the sol locus (aad, ctfA, ctfB and adc) genes for butanol and acetone formation in Clostridium acetobutylicum ATCC 824. Primer extension analysis identified a transcriptional start site 35 bp upstream of the solR start codon. Amino acid comparisons of SolR identified a potential helix-turn-helix DNA-binding motif in the C-terminal half towards the center of the protein, suggesting a regulatory role. Overexpression of SolR in strain ATCC 824(pCO1) resulted in a solvent-negative phenotype owing to its deleterious effect on the transcription of the sol locus genes. Inactivation of solR in C. acetobutylicum via homologous recombination yielded mutants B and H (ATCC 824 solR::pO1X) which exhibited deregulated solvent production characterized by increased flux towards butanol and acetone formation, earlier induction of aad, lower overall acid production, markedly improved yields of solvents on glucose, a prolonged solvent production phase, and increased biomass accumulation compared to those of the wild-type strain.

  4. Integrative modelling of pH-dependent enzyme activity and transcriptomic regulation of the acetone-butanol-ethanol fermentation of Clostridium acetobutylicum in continuous culture.

    PubMed

    Millat, Thomas; Janssen, Holger; Bahl, Hubert; Fischer, Ralf-Jörg; Wolkenhauer, Olaf

    2013-09-01

    In a continuous culture under phosphate limitation the metabolism of Clostridium acetobutylicum depends on the external pH level. By comparing seven steady-state conditions between pH 5.7 and pH 4.5 we show that the switch from acidogenesis to solventogenesis occurs between pH 5.3 and pH 5.0 with an intermediate state at pH 5.1. Here, an integrative study is presented investigating how a changing external pH level affects the clostridial acetone-butanol-ethanol (ABE) fermentation pathway. This is of particular interest as the biotechnological production of n-butanol as biofuel has recently returned into the focus of industrial applications. One prerequisite is the furthering of the knowledge of the factors determining the solvent production and their integrative regulations. We have mathematically analysed the influence of pH-dependent specific enzyme activities of branch points of the metabolism on the product formation. This kinetic regulation was compared with transcriptomic regulation regarding gene transcription and the proteomic profile. Furthermore, both regulatory mechanisms were combined yielding a detailed projection of their individual and joint effects on the product formation. The resulting model represents an important platform for future developments of industrial butanol production based on C. acetobutylicum.

  5. The mechanism of switching from an acidogenic to butanol-acetone fermentation by Clostridium acetobutylicum. Technical progress report, July 1990--December 1992

    SciTech Connect

    Rogers, P.

    1992-12-31

    The overall objective of this project is to elucidate the detailed mechanism by which solvent-forming bacteria such as Clostridium acetobutylicum regulate the well-known shift in fermentation pathway between alcohol-acetone and organic acid production. It is desired to eventually isolate and describe: (1) the regulatory genes and protein elements that determine induction of synthesis of the solvent-pathway enzymes; and (2) how this regulation system interacts with the sporulatin induction and development program and with related pathways such as granulse and exopolysaccharide formation in clostridia. A working model forhow clostridial control systems work can be derived from recent research on stress systems in E. coli and sporulation in Bacillus subtilis.

  6. Comparative genomic and proteomic analyses of Clostridium acetobutylicum Rh8 and its parent strain DSM 1731 revealed new understandings on butanol tolerance

    SciTech Connect

    Bao, Guanhui; Dong, Hongjun; Zhu, Yan; Mao, Shaoming; Zhang, Tianrui; Zhang, Yanping; Chen, Zugen; Li, Yin

    2014-08-08

    Highlights: • Genomes of a butanol tolerant strain and its parent strain were deciphered. • Comparative genomic and proteomic was applied to understand butanol tolerance. • None differentially expressed proteins have mutations in its corresponding genes. • Mutations in ribosome might be responsible for the global difference of proteomics. - Abstract: Clostridium acetobutylicum strain Rh8 is a butanol-tolerant mutant which can tolerate up to 19 g/L butanol, 46% higher than that of its parent strain DSM 1731. We previously performed comparative cytoplasm- and membrane-proteomic analyses to understand the mechanism underlying the improved butanol tolerance of strain Rh8. In this work, we further extended this comparison to the genomic level. Compared with the genome of the parent strain DSM 1731, two insertion sites, four deletion sites, and 67 single nucleotide variations (SNVs) are distributed throughout the genome of strain Rh8. Among the 67 SNVs, 16 SNVs are located in the predicted promoters and intergenic regions; while 29 SNVs are located in the coding sequence, affecting a total of 21 proteins involved in transport, cell structure, DNA replication, and protein translation. The remaining 22 SNVs are located in the ribosomal genes, affecting a total of 12 rRNA genes in different operons. Analysis of previous comparative proteomic data indicated that none of the differentially expressed proteins have mutations in its corresponding genes. Rchange Algorithms analysis indicated that the mutations occurred in the ribosomal genes might change the ribosome RNA thermodynamic characteristics, thus affect the translation strength of these proteins. Take together, the improved butanol tolerance of C. acetobutylicum strain Rh8 might be acquired through regulating the translational process to achieve different expression strength of genes involved in butanol tolerance.

  7. Molecular Characterization and Transcriptional Analysis of adhE2, the Gene Encoding the NADH-Dependent Aldehyde/Alcohol Dehydrogenase Responsible for Butanol Production in Alcohologenic Cultures of Clostridium acetobutylicum ATCC 824

    PubMed Central

    Fontaine, Lisa; Meynial-Salles, Isabelle; Girbal, Laurence; Yang, Xinghong; Croux, Christian; Soucaille, Philippe

    2002-01-01

    The adhE2 gene of Clostridium acetobutylicum ATCC 824, coding for an aldehyde/alcohol dehydrogenase (AADH), was characterized from molecular and biochemical points of view. The 2,577-bp adhE2 codes for a 94.4-kDa protein. adhE2 is expressed, as a monocistronic operon, in alcohologenic cultures and not in solventogenic cultures. Primer extension analysis identified two transcriptional start sites 160 and 215 bp upstream of the adhE2 start codon. The expression of adhE2 from a plasmid in the DG1 mutant of C. acetobutylicum, a mutant cured of the pSOL1 megaplasmid, restored butanol production and provided elevated activities of NADH-dependent butyraldehyde and butanol dehydrogenases. The recombinant AdhE2 protein expressed in E. coli as a Strep-tag fusion protein and purified to homogeneity also demonstrated NADH-dependent butyraldehyde and butanol dehydrogenase activities. This is the second AADH identified in C. acetobutylicum ATCC 824, and to our knowledge this is the first example of a bacterium with two AADHs. It is noteworthy that the two corresponding genes, adhE and adhE2, are carried by the pSOL1 megaplasmid of C. acetobutylicum ATCC 824. PMID:11790753

  8. Isolation and characterisation of non-anaerobic butanol-producing symbiotic system TSH06.

    PubMed

    Wang, Genyu; Wu, Pengfei; Liu, Ya; Mi, Shuo; Mai, Shuai; Gu, Chunkai; Wang, Gehua; Liu, Hongjuan; Zhang, Jianan; Børresen, Børre Tore; Mellemsæther, Evy; Kotlar, Hans Kristian

    2015-10-01

    Butanol-producing microorganisms are all obligate anaerobes. In this study, a unique symbiotic system TSH06 was isolated to be capable of producing butanol under non-anaerobic condition. Denaturing gradient gel electrophoresis (DGGE) analysis of 16S ribosomal RNA (rRNA) revealed that two strains coexist in TSH06. The two strains were identical to Clostridium acetobutylicum and Bacillus cereus, respectively. They were isolated individually and named as C. acetobutylicum TSH1 and B. cereus TSH2. C. acetobutylicum TSH1 is a butanol-producing, obligate anaerobic strain. Facultative anaerobic B. cereus TSH2 did not possess the ability of butanol production; however, it offered C. acetobutylicum TSH1 the viability under non-anaerobic condition. Moreover, B. cereus TSH2 enhanced butanol yield and speed of fermentation. TSH06 produced 12.97 g/L butanol and 15.39 g/L total solvent under non-anaerobic condition, which is 25 and 24 %, respectively, higher than those of C. acetobutylicum TSH1. In addition, TSH06 produced butanol faster under non-anaerobic condition than under anaerobic condition. Butanol accounted for more than 80 % of total solvent, which is higher than the known report. TSH06 was stable during passage. In all, TSH06 is a promising candidate for industrialisation of biobutanol with high yield, high butanol proportion, easy-handling and time-saving system. These results demonstrated the potential advantage of symbiosis. This study also provides a promising strategy for butanol fermentation.

  9. Acetone-butanol-ethanol (ABE) fermentation using Clostridium acetobutylicum XY16 and in situ recovery by PDMS/ceramic composite membrane.

    PubMed

    Wu, Hao; Chen, Xiao-Peng; Liu, Gong-Ping; Jiang, Min; Guo, Ting; Jin, Wan-Qin; Wei, Ping; Zhu, Da-Wei

    2012-09-01

    PDMS/ceramic composite membrane was directly integrated with acetone-butanol-ethanol (ABE) fermentation using Clostridium acetobutylicum XY16 at 37 °C and in situ removing ABE from fermentation broth. The membrane was integrated with batch fermentation, and approximately 46 % solvent was extracted. The solvent in permeates was 118 g/L, and solvent productivity was 0.303 g/(L/h), which was approximately 33 % higher compared with the batch fermentation without in situ recovery. The fed-batch fermentation with in situ recovery by pervaporation continued for more than 200 h, 61 % solvent was extracted, and the solvent in penetration was 96.2 g/L. The total flux ranged from 0.338 to 0.847 kg/(m(2)/h) and the separation factor of butanol ranged from 5.1 to 27.1 in this process. The membrane was fouled by the active fermentation broth, nevertheless the separation performances were partially recovered by offline membrane cleaning, and the solvent productivity was increased to 0.252 g/(L/h), which was 19 % higher compared with that in situ recovery process without membrane cleaning.

  10. Direct fermentation of gelatinized cassava starch to acetone, butanol, and ethanol using Clostridium acetobutylicum mutant obtained by atmospheric and room temperature plasma.

    PubMed

    Li, Han-guang; Luo, Wei; Wang, Qiang; Yu, Xiao-bin

    2014-04-01

    The mutant strain designated as ART18, obtained from the wild-type strain Clostridium acetobutylicum PW12 treated by atmospheric and room temperature plasma, showed higher solvent tolerance and butanol production than that of the wild-type strain. The production of butanol was 11.3 ± 0.5 g/L, 31 % higher than that of the wild-type strain when it was used for acetone, butanol, and ethanol fermentation in P2 medium. Furthermore, the effects of cassava flour concentration, pH regulators, and vitamins on the ABE production were also investigated. The highest butanol production of 15.8 ± 0.8 g/L and butanol yield (0.31 g/g) were achieved after the above factors were optimized. When acetone, butanol, and ethanol fermentation by ART18 was carried out in a 15-L bioreactor, the butanol production, the productivity of butanol, and the total solvent were 16.3 ± 0.9, 0.19, and 0.28 g/L(/)h, respectively. These results indicate that ART18 is a promising industrial producer in ABE fermentation.

  11. Clostridium thermoalcaliphilum sp. nov., an anaerobic and thermotolerant facultative alkaliphile.

    PubMed

    Li, Y; Engle, M; Weiss, N; Mandelco, L; Wiegel, J

    1994-01-01

    An anaerobic and thermophilic alkaliphile, strain JW/YL23-2T (T = type strain), was isolated from sewage sludge obtained from a sewage plant in Atlanta, Ga. At pH 10.1 and 50 degrees C, the doubling time of this strain was 19 min. Strain JW/YL23-2T, a motile rod-shaped bacterium with 2 to 12 peritrichous flagella, exhibited a negative Gram stain reaction but was gram-type positive as judged by the polymyxin B test. No heat-stable (85 degrees C, 15 min) endospores were detected. At 50 degrees C, growth occurred at pH values ranging from 7.0 to 11.0; the optimum pH was 9.6 to 10.1. The temperature range for growth ranged from 27 to 57.5 degrees C; the optimum temperature was 48 to 51 degrees C (pH 10.1). Dissimilatory sulfate reduction was not detected. The organism utilized glucose, fructose, sucrose, maltose, cellobiose, and Casamino Acids. The DNA G+C content was 32 mol% (as determined by chemical analysis). A 16S rRNA sequence analysis revealed a 2% inferred evolutionary distance to Clostridium paradoxum. However, the cell wall type of strain JW/YL23-2T was A4 beta (L-Orn-D-Asp), while that of C. paradoxum was A1 tau (m-diaminopimelic acid direct). On the basis of the alkaline pH values and high temperatures for optimal growth, the inability to form spores, and other characteristics different from C. paradoxum characteristics, strain JW/YL-23-2 was placed in a new species, Clostridium thermoalcaliphilum; JW/YL23-2 (= DSM 7309) is the type strain of this new species.

  12. Clostridium thermoalcaliphilum sp. nov., an anaerobic and thermotolerant facultative alkaliphile

    SciTech Connect

    Youhong Li; Engle, M.; Wiegel, J.

    1994-01-01

    An anaerobic and thermophilic alkaliphile, strain JW/YL23-2{sub T} (T = type strain), was isolated from sewage sludge obtained from a sewage plant in Atlanta, Ga. at pH 10.1 and 50{degrees}C, the doubling time of this strain was 19 min. Strain JW/YL23-2{sub T}, a motile rod-shaped bacterium with 2 to 12 peritrichous flagella, exhibited a negative Gram stain reaction but was gram-type positive as judged by the polymyxin B test. No heat-stable (85{degrees}C, 15 min) endospores were detected. At 50{degrees}C, growth occurred at pH values ranging from 7.0 to 11.0; the optimum pH was 9.6 to 10.1. The temperature range for growth ranged from 27 to 57.5{degrees}C; the optimum temperature was 48 to 51{degrees}C (pH 10.1). Dissimilatory sulfate reduction was not detected. The organism utilized glucose, fructose, sucrose, maltose, cellobiose, and Casamino Acids. The DNA G+C content was 32 mol% (as determined by chemical analysis). A 16S rRNA sequence analysis revealed a 2% inferred evolutionary distance to Clostridium paradoxum. However, the cell wall type of strain JW/YL23-2{sup T} was A4{beta} (L-Orn-D-Asp), while that of C. paradoxum was Al{sub {tau}} (m-diaminopimelic acid direct). On the basis of the alkaline pH values and high temperatures for optimal growth, the inability to form spores, and other characteristics different from C. paradoxum characteristics, strain JW/YL-23-2 was placed in a new species, Clostridium thermoalcaliphilum; JW/YL23-2 (= DSM 7309) is the type strain of this new species.

  13. Common Mesophilic Anaerobes, Including Clostridium botulinum and Clostridium tetani, in 21 Soil Specimens

    PubMed Central

    Smith, Louis Ds.

    1975-01-01

    A relatively rich medium was markedly superior to a dilute medium for the isolation of anaerobic bacteria from soil. The obligate anaerobes isolated from 21 soil samples were all clostridia and the counts ranged from 2.7 × 102 to 3.3 × 106 per g. The organisms most frequently isolated were Clostridium subterminale, C. sordellii, C. sporogenes, C. indolis, C. bifermentans, C. mangenoti, and C. perfringens. Seventeen other species were also recognized but almost one-third of the isolates could not be identified with any known species of Clostridum. C. botulinum type A was demonstrated in six soil samples, and type B in one. These soils were neutral to alkaline in reaction (average pH 7.9) and low in organic matter content (1.4%). The association of C. botulinum types A and B with neutral to alkaline soils was statistically significant (P = 0.001) as was their association with soils low in organic matter (P = 0.005). C. botulinum types E and F were found in one soil sample, pH 4.5, with organic matter 13.7%. C. tetani was isolated from two soil samples, both of intermediate pH value and higher than average organic matter content. PMID:238468

  14. σK of Clostridium acetobutylicum is the first known sporulation-specific sigma factor with two developmentally separated roles, one early and one late in sporulation.

    PubMed

    Al-Hinai, Mohab A; Jones, Shawn W; Papoutsakis, Eleftherios T

    2014-01-01

    Sporulation in the model endospore-forming organism Bacillus subtilis proceeds via the sequential and stage-specific activation of the sporulation-specific sigma factors, σ(H) (early), σ(F), σ(E), σ(G), and σ(K) (late). Here we show that the Clostridium acetobutylicum σ(K) acts both early, prior to Spo0A expression, and late, past σ(G) activation, thus departing from the B. subtilis model. The C. acetobutylicum sigK deletion (ΔsigK) mutant was unable to sporulate, and solventogenesis, the characteristic stationary-phase phenomenon for this organism, was severely diminished. Transmission electron microscopy demonstrated that the ΔsigK mutant does not develop an asymmetric septum and produces no granulose. Complementation of sigK restored sporulation and solventogenesis to wild-type levels. Spo0A and σ(G) proteins were not detectable by Western analysis, while σ(F) protein levels were significantly reduced in the ΔsigK mutant. spo0A, sigF, sigE, sigG, spoIIE, and adhE1 transcript levels were all downregulated in the ΔsigK mutant, while those of the sigH transcript were unaffected during the exponential and transitional phases of culture. These data show that σ(K) is necessary for sporulation prior to spo0A expression. Plasmid-based expression of spo0A in the ΔsigK mutant from a nonnative promoter restored solventogenesis and the production of Spo0A, σ(F), σ(E), and σ(G), but not sporulation, which was blocked past the σ(G) stage of development, thus demonstrating that σ(K) is also necessary in late sporulation. sigK is expressed very early at low levels in exponential phase but is strongly upregulated during the middle to late stationary phase. This is the first sporulation-specific sigma factor shown to have two developmentally separated roles.

  15. ClosTron-mediated engineering of Clostridium.

    PubMed

    Kuehne, Sarah A; Heap, John T; Cooksley, Clare M; Cartman, Stephen T; Minton, Nigel P

    2011-01-01

    The genus Clostridium is a diverse assemblage of Gram positive, anaerobic, endospore-forming bacteria. Whilst certain species have achieved notoriety as important animal and human pathogens (e.g. Clostridium difficile, Clostridium botulinum, Clostridium tetani, and Clostridium perfringens), the vast majority of the genus are entirely benign, and are able to undertake all manner of useful biotransformations. Prominent amongst them are those species able to produce the biofuels, butanol and ethanol from biomass-derived residues, such as Clostridium acetobutylicum, Clostridium beijerinkii, Clostridium thermocellum, and Clostridium phytofermentans. The prominence of the genus in disease and biotechnology has led to the need for more effective means of genetic modification. The historical absence of methods based on conventional strategies for "knock-in" and "knock-out" in Clostridium has led to the adoption of recombination-independent procedures, typified by ClosTron technology. The ClosTron uses a retargeted group II intron and a retro-transposition-activated marker to selectively insert DNA into defined sites within the genome, to bring about gene inactivation and/or cargo DNA delivery. The procedure is extremely efficient, rapid, and requires minimal effort by the operator.

  16. Butyric acid from anaerobic fermentation of lignocellulosic biomass hydrolysates by Clostridium sp. strain RPT-4213

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A novel Clostridium sp. strain RPT-4213 was found producing butyrate under strict anaerobic conditions. This strain produced 9.47 g L-1 butyric acid from MRS media (0.48 g/g glucose). RPT-4213 was also used to ferment dilute acid pretreated hydrolysates including wheat straw (WSH), corn fiber (CFH...

  17. Butyric acid from anaerobic fermentation of lignocellulosic biomass hydrolysates by Clostridium tyrobutyricum strain RPT-4213

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A newly isolated Clostridium sp. strain RPT-4213 was found to produce butyrate under anaerobic conditions. Fermentations using Lactobacilli MRS Broth produced 9.47 g L-1 butyric acid from glucose (0.48 g/g glucose). However, the strain was not capable of utilizing five carbon sugars. To assess the a...

  18. A quantitative metabolomics study of high sodium response in Clostridium acetobutylicum ATCC 824 acetone-butanol-ethanol (ABE) fermentation

    PubMed Central

    Zhao, Xinhe; Condruz, Stefan; Chen, Jingkui; Jolicoeur, Mario

    2016-01-01

    Hemicellulose hydrolysates, sugar-rich feedstocks used in biobutanol refinery, are normally obtained by adding sodium hydroxide in the hydrolyze process. However, the resulting high sodium concentration in the hydrolysate inhibits ABE (acetone-butanol-ethanol) fermentation, and thus limits the use of these low-cost feedstocks. We have thus studied the effect of high sodium on the metabolic behavior of Clostridium acetobutyricum ATCC 824, with xylose as the carbon source. At a threshold sodium concentration of 200 mM, a decrease of the maximum cell dry weight (−19.50 ± 0.85%) and of ABE yield (−35.14 ± 3.50% acetone, −33.37 ± 0.74% butanol, −22.95 ± 1.81% ethanol) were observed compared to control culture. However, solvents specific productivities were not affected by supplementing sodium. The main effects of high sodium on cell metabolism were observed in acidogenesis, during which we observed the accumulation of ATP and NADH, and the inhibition of the pentose phosphate (PPP) and the glycolytic pathways with up to 80.73 ± 1.47% and 68.84 ± 3.42% decrease of the associated metabolic intermediates, respectively. However, the NADP+-to-NADPH ratio was constant for the whole culture duration, a phenomenon explaining the robustness of solvents specific productivities. Therefore, high sodium, which inhibited biomass growth through coordinated metabolic effects, interestingly triggered cell robustness on solvents specific productivity. PMID:27321153

  19. Anaerobic decomposition of humic substances by Clostridium from the deep subsurface.

    PubMed

    Ueno, Akio; Shimizu, Satoru; Tamamura, Shuji; Okuyama, Hidetoshi; Naganuma, Takeshi; Kaneko, Katsuhiko

    2016-01-08

    Decomposition of humic substances (HSs) is a slow and cryptic but non-negligible component of carbon cycling in sediments. Aerobic decomposition of HSs by microorganisms in the surface environment has been well documented; however, the mechanism of anaerobic microbial decomposition of HSs is not completely understood. Moreover, no microorganisms capable of anaerobic decomposition of HSs have been isolated. Here, we report the anaerobic decomposition of humic acids (HAs) by the anaerobic bacterium Clostridium sp. HSAI-1 isolated from the deep terrestrial subsurface. The use of (14)C-labelled polycatechol as an HA analogue demonstrated that the bacterium decomposed this substance up to 7.4% over 14 days. The decomposition of commercial and natural HAs by the bacterium yielded lower molecular mass fractions, as determined using high-performance size-exclusion chromatography. Fourier transform infrared spectroscopy revealed the removal of carboxyl groups and polysaccharide-related substances, as well as the generation of aliphatic components, amide and aromatic groups. Therefore, our results suggest that Clostridium sp. HSAI-1 anaerobically decomposes and transforms HSs. This study improves our understanding of the anaerobic decomposition of HSs in the hidden carbon cycling in the Earth's subsurface.

  20. Anaerobic decomposition of humic substances by Clostridium from the deep subsurface

    PubMed Central

    Ueno, Akio; Shimizu, Satoru; Tamamura, Shuji; Okuyama, Hidetoshi; Naganuma, Takeshi; Kaneko, Katsuhiko

    2016-01-01

    Decomposition of humic substances (HSs) is a slow and cryptic but non-negligible component of carbon cycling in sediments. Aerobic decomposition of HSs by microorganisms in the surface environment has been well documented; however, the mechanism of anaerobic microbial decomposition of HSs is not completely understood. Moreover, no microorganisms capable of anaerobic decomposition of HSs have been isolated. Here, we report the anaerobic decomposition of humic acids (HAs) by the anaerobic bacterium Clostridium sp. HSAI-1 isolated from the deep terrestrial subsurface. The use of 14C-labelled polycatechol as an HA analogue demonstrated that the bacterium decomposed this substance up to 7.4% over 14 days. The decomposition of commercial and natural HAs by the bacterium yielded lower molecular mass fractions, as determined using high-performance size-exclusion chromatography. Fourier transform infrared spectroscopy revealed the removal of carboxyl groups and polysaccharide-related substances, as well as the generation of aliphatic components, amide and aromatic groups. Therefore, our results suggest that Clostridium sp. HSAI-1 anaerobically decomposes and transforms HSs. This study improves our understanding of the anaerobic decomposition of HSs in the hidden carbon cycling in the Earth’s subsurface. PMID:26743007

  1. Improvement of the butanol production selectivity and butanol to acetone ratio (B:A) by addition of electron carriers in the batch culture of a new local isolate of Clostridium acetobutylicum YM1.

    PubMed

    Nasser Al-Shorgani, Najeeb Kaid; Kalil, Mohd Sahaid; Wan Yusoff, Wan Mohtar; Shukor, Hafiza; Hamid, Aidil Abdul

    2015-12-01

    Improvement in the butanol production selectivity or enhanced butanol:acetone ratio (B:A) is desirable in acetone-butanol-ethanol (ABE) fermentation by Clostridium strains. In this study, artificial electron carriers were added to the fermentation medium of a new isolate of Clostridium acetobutylicum YM1 in order to improve the butanol yield and B:A ratio. The results revealed that medium supplementation with electron carriers changed the metabolism flux of electron and carbon in ABE fermentation by YM1. A decrease in acetone production, which subsequently improved the B:A ratio, was observed. Further improvement in the butanol production and B:A ratios were obtained when the fermentation medium was supplemented with butyric acid. The maximum butanol production (18.20 ± 1.38 g/L) was gained when a combination of methyl red and butyric acid was added. Although the addition of benzyl viologen (0.1 mM) and butyric acid resulted in high a B:A ratio of 16:1 (800% increment compared with the conventional 2:1 ratio), the addition of benzyl viologen to the culture after 4 h resulted in the production of 18.05 g/L butanol. Manipulating the metabolic flux to butanol through the addition of electron carriers could become an alternative strategy to achieve higher butanol productivity and improve the B:A ratio.

  2. An Esterase from Anaerobic Clostridium hathewayi Can Hydrolyze Aliphatic-Aromatic Polyesters.

    PubMed

    Perz, Veronika; Hromic, Altijana; Baumschlager, Armin; Steinkellner, Georg; Pavkov-Keller, Tea; Gruber, Karl; Bleymaier, Klaus; Zitzenbacher, Sabine; Zankel, Armin; Mayrhofer, Claudia; Sinkel, Carsten; Kueper, Ulf; Schlegel, Katharina; Ribitsch, Doris; Guebitz, Georg M

    2016-03-15

    Recently, a variety of biodegradable polymers have been developed as alternatives to recalcitrant materials. Although many studies on polyester biodegradability have focused on aerobic environments, there is much less known on biodegradation of polyesters in natural and artificial anaerobic habitats. Consequently, the potential of anaerobic biogas sludge to hydrolyze the synthetic compostable polyester PBAT (poly(butylene adipate-co-butylene terephthalate) was evaluated in this study. On the basis of reverse-phase high-performance liquid chromatography (RP-HPLC) analysis, accumulation of terephthalic acid (Ta) was observed in all anaerobic batches within the first 14 days. Thereafter, a decline of Ta was observed, which occurred presumably due to consumption by the microbial population. The esterase Chath_Est1 from the anaerobic risk 1 strain Clostridium hathewayi DSM-13479 was found to hydrolyze PBAT. Detailed characterization of this esterase including elucidation of the crystal structure was performed. The crystal structure indicates that Chath_Est1 belongs to the α/β-hydrolases family. This study gives a clear hint that also micro-organisms in anaerobic habitats can degrade manmade PBAT.

  3. Clostridium algifaecis sp. nov., an anaerobic bacterial species from decomposing algal scum.

    PubMed

    Wu, Yu-Fan; Zheng, Hui; Wu, Qing-Long; Yang, Hong; Liu, Shuang-Jiang

    2014-11-01

    Two anaerobic bacterial strains, MB9-7(T) and MB9-9, were isolated from decomposing algal scum and were characterized using a polyphasic approach. Phylogenetic analysis of 16S rRNA gene sequences showed that strains MB9-7(T) and MB9-9 are closely related to each other (99.7% similarity) and they are also closely related to Clostridium tyrobutyricum (96.5%). The two strains were Gram-stain positive and rod-shaped. Growth occurred at 20-45 °C, at pH 4.0-8.0 and at NaCl concentrations of up to 2% (w/v). Acid was produced from glucose, xylose and mannose. Products of fermentation in PYG medium were mainly butyrate, acetate, carbon dioxide and hydrogen. The predominant cellular fatty acids were C(14:0) and C(16:0). The cellular polar lipids comprised phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, two glycolipids, one phospholipid, one aminophospholipid and two aminolipids. The DNA G+C contents of strain MB9-7(T) and MB9-9 were 27.9 and 28.7 mol%, respectively. These results support the assignment of the new isolates to the genus Clostridium and also distinguish them from other species of the genus Clostridium. Hence, it is proposed that strains MB9-7(T) and MB9-9 represent a novel species of the genus Clostridium, with the suggested name Clostridium algifaecis sp. nov. The type strain is MB9-7(T) ( =CGMCC 1.5188(T) =DSM 28783(T)).

  4. Clostridium aldrichii sp. nov., a cellulolytic mesophile inhabiting a wood-fermenting anaerobic digester.

    PubMed

    Yang, J C; Chynoweth, D P; Williams, D S; Li, A

    1990-07-01

    An anaerobic, mesophilic, spore-forming, cellulolytic bacterium was repeatedly isolated from a wood-fermenting anaerobic digester. Cells of this organism were gram-positive rods, motile with a bundle of polar flagella, and formed subterminal oblong spores. The colonies in agar had an irregular shape with many platelike structures and were greyish white. Cellulose, xylan, and cellobiose served as substrates for growth. Acetate, propionate, butyrate, isobutyrate, isovalerate, lactate, succinate, H2, and CO2 were products of cellobiose fermentation. The optimal temperature and pH for growth were 35 degrees C and 7, respectively. The DNA composition was 40 mol% G + C. The name Clostridium aldrichii sp. nov. is proposed. The type strain is P-1 (= OGI 112, = ATCC 49358).

  5. Butanol production employing fed-batch fermentation by Clostridium acetobutylicum GX01 using alkali-pretreated sugarcane bagasse hydrolysed by enzymes from Thermoascus aurantiacus QS 7-2-4.

    PubMed

    Pang, Zong-Wen; Lu, Wei; Zhang, Hui; Liang, Zheng-Wu; Liang, Jing-Juan; Du, Liang-Wei; Duan, Cheng-Jie; Feng, Jia-Xun

    2016-07-01

    Sugarcane bagasse (SB) is a potential feedstock for butanol production. However, biological production of butanol from SB is less economically viable. In this study, evaluation of eight pretreatments on SB showed that alkali pretreatment efficiently removed lignin from SB while retaining the intact native structure of the released microfibrils. In total, 99% of cellulose and 100% of hemicellulose in alkali-pretreated SB were hydrolysed by enzymes from Thermoascus aurantiacus. The hydrolysate was used to produce butanol in a fed-batch fermentation by Clostridium acetobutylicum. At 60h, 14.17 and 21.11gL(-1) of butanol and acetone-butanol-ethanol (ABE) were produced from 68.89gL(-1) of total sugars, respectively, yielding 0.22 and 0.33gg(-1) of sugars. The maximum yield of butanol and ABE reached 15.4g and 22.9g per 100g raw SB, respectively. This established process may have potential application for butanol production from SB.

  6. Isolation and characterization of an anaerobic, cellulolytic bacterium, Clostridium cellulovorans sp. nov

    SciTech Connect

    Sleat, R.; Mah, R.A.; Robinson, R.

    1984-07-01

    A new anaerobic, mesophilic, spore-forming cellulolytic bacterium is described. Cellulose is cleared within 24 to 48 h around colonies formed in cellulose agar roll tubes. Cells stain gram negative and are nonmotile rods which form oblong spores either centrally or subterminally in a clostridial swelling. Colonies are irregular with an opaque edge and a center devoid of both vegetative cells and spores. Cellulose, xylan, pectin, cellubiose, glucose, maltose, galactose, sucrose, lactose, and mannose serve as substrates for growth. H/sub 2/, CO/sub 2/, acetate, butyrate, formate, and lactate are produced during fermentation of cellulose or cellobiose. The temperature and pH for optimum growth are 37/sup 0/C and 7.0, respectively. The DNA composition is 26 to 27 mol% guanine plus cytosine. This bacterium resembles Clostridium lochhheadii in morphological and some biochemical characteristics but is not identical to it. The name Clostridium cellulovorans sp. nov. is proposed. The type strain is 743B (ATCC 35296).

  7. Investigation of the relation between anaerobic bacteria genus clostridium and late-onset autism etiology in children.

    PubMed

    Keşli, Recep; Gökçen, Cem; Buluğ, Ufuk; Terzi, Yüksel

    2014-01-01

    The aim of this study was to investigate the relation between the etiology of late-onset childhood autism and anaerobic bacteria. Thirty children diagnosed with autistic disorder and control group have been included in the study. 3-(3-hydroxy phenyl)-3-hydroxypropionic acid (HPHPA) excretion rates which is a metabolic product of the genus Clostridium, were measured via mass spectrometry-gas chromatography (MS-GC) method from urine samples. When the assayed average HPHPA values compared with each group, a statistically significant difference was found (p < 0.05). Data obtained from this study support the existence of a significant correlation between autism etiology and anaerobic bacteria.

  8. Reductive Disslocation of Pu(IV) by Clostridium sp. Under Anaerobic Conditions

    SciTech Connect

    Francis,A.; Dodge, C.; Gillow, J.

    2008-01-01

    An anaerobic, gram positive, spore-forming bacterium Clostridium sp., common in soils and wastes, capable of reduction of Fe(III) to Fe(II), Mn(IV) to Mn(II), Tc(VII) to Tc(IV), and U(VI) to U(IV), reduced Pu(IV) to Pu(III). Addition of 242Pu (IV)-nitrate to the bacterial growth medium at pH 6.4 resulted in the precipitation of Pu as amorphous Pu(OH)4 due to hydrolysis and polymerization reactions. The Pu (1 x 10-5 M) had no effect upon growth of the bacterium as evidenced by glucose consumption; carbon dioxide and hydrogen production; a decrease in pH of the medium from 6.4 to 3.0 due to production of acetic and butyric acids from glucose fermentation; and a change in the Eh of the culture medium from +50 to -180 mV. Commensurate with bacterial growth, Pu was rapidly solubilized as evidenced by an increase in Pu concentration in solution which passed through a 0.03 {mu}m filtration. Selective solvent extraction of the culture by thenoyltrifluoroacetone (TTA) indicated the presence of a reduced Pu species in the soluble fraction. X-ray absorption near edge spectroscopic (XANES) analysis of Pu in the culture sample at the Pu LIII absorption edge (18.054 keV) showed a shift of -3 eV compared to a Pu(IV) standard indicating reduction of Pu(IV) to Pu(III). These results suggest that, although Pu generally exists as insoluble Pu(IV) in the environment, under appropriate conditions, anaerobic microbial activity could affect the long-term stability and mobility of Pu by its reductive dissolution.

  9. Selective medium for isolation of Clostridium butyricum from human feces.

    PubMed Central

    Popoff, M R

    1984-01-01

    A selective medium, Clostridium butyricum isolation medium (BIM), is described for the isolation of C. butyricum from human feces. The BIM is a synthetic minimal medium and contains trimethoprim (16 micrograms/ml), D-cycloserine (10 micrograms/ml), and polymyxin B sulfate (20 micrograms/ml) as selective inhibitory agents. Qualitative tests indicated that C. butyricum and other butyric acid-producing clostridia grew on BIM, Clostridium sphenoides and Bacillus cereus produced small colonies, and other clostridia and other obligate anaerobic or facultatively anerobic bacteria were inhibited. Quantitative recovery of C. butyricum from cultures or seeded fecal samples was comparable with BIM and with complex medium, but the quantitative recovery of the other butyric acid-producing clostridia tested (C. beijerinckii, C. acetobutylicum) was lower with BIM than with complex medium. The BIM should aid the rapid isolation of C. butyricum from fecal samples and should be useful for bacteriological investigation of neonatal necrotizing enterocolitis. PMID:6490827

  10. Facultative Anaerobe Caldibacillus debilis GB1: Characterization and Use in a Designed Aerotolerant, Cellulose-Degrading Coculture with Clostridium thermocellum.

    PubMed

    Wushke, Scott; Levin, David B; Cicek, Nazim; Sparling, Richard

    2015-08-15

    Development of a designed coculture that can achieve aerotolerant ethanogenic biofuel production from cellulose can reduce the costs of maintaining anaerobic conditions during industrial consolidated bioprocessing (CBP). To this end, a strain of Caldibacillus debilis isolated from an air-tolerant cellulolytic consortium which included a Clostridium thermocellum strain was characterized and compared with the C. debilis type strain. Characterization of isolate C. debilis GB1 and comparisons with the type strain of C. debilis revealed significant physiological differences, including (i) the absence of anaerobic metabolism in the type strain and (ii) different end product synthesis profiles under the experimental conditions used. The designed cocultures displayed unique responses to oxidative conditions, including an increase in lactate production. We show here that when the two species were cultured together, the noncellulolytic facultative anaerobe C. debilis GB1 provided respiratory protection for C. thermocellum, allowing the synergistic utilization of cellulose even under an aerobic atmosphere.

  11. Inactivation of Clostridium difficile in sewage sludge by anaerobic thermophilic digestion.

    PubMed

    Xu, Changyun; Salsali, Hamidreza; Weese, Scott; Warriner, Keith

    2016-01-01

    There has been an increase in community-associated Clostridium difficile infections with biosolids derived from wastewater treatment being identified as one potential source. The current study evaluated the efficacy of thermophilic digestion in decreasing levels of C. difficile ribotype 078 associated with sewage sludge. Five isolates of C. difficile 078 were introduced (final density of 5 log CFU/g) into digested sludge and subjected to anaerobic digestion at mesophilic (36 or 42 °C) or thermophilic (55 °C) temperatures for up to 60 days. It was found that mesophilic digestion at 36 °C did not result in a significant reduction in C. difficile spore levels. In contrast, thermophilic sludge digestion reduced endospore levels at a rate of 0.19-2.68 log CFU/day, depending on the strain tested. The mechanism of lethality was indirect - by stimulating germination then inactivating the resultant vegetative cells. Acidification of sludge by adding acetic acid (6 g/L) inhibited the germination of spores regardless of the sludge digestion temperature. In conclusion, thermophilic digestion can be applied to reduce C. difficile in biosolids, thereby reducing the environmental burden of the enteric pathogen.

  12. Isolation and characterization of a moderately thermophilic anaerobic alkaliphile, Clostridium paradoxum sp. nov.

    SciTech Connect

    Youhong Li; Wiegel, J.; Mandelco, L.

    1993-07-01

    Alkaliphilic, moderately thermophilic anaerobic bacteria able to grow above pH 10.5 and 55{degrees}C were isolated from various sewage plants in the United States. The strains were motile with two to six peritrichous flagella and formed round to slightly oval terminal spores in terminally distended and slightly enlarged cells. Sporulated cells remained motile. The pH range for growth was between 7.0 and 11.1, with an optimum of around 10.1. At pH 10.1 the temperature range for growth was between 30 and 63{degrees}C, with an optimum of 56{degrees}C. The shortest observed doubling time (glucose) was around 16 min at 56{degrees}C and pH 10.1. No dissimilatory sulfate reduction was detected. The organism utilized glucose, fructose, sucrose, maltose, and pyruvate but required yeast extract or tryptone for growth. Optimal NaCl concentrations for growth were between 50 and 200 mM. The guanine-plus-cytosine content was 30.0 {+-} 0.10 mol%. On the basis of unique properties and 16S rRNA analysis, the strains are placed in a new species, Clostridium paradoxum, referring to the unusual retainment of motility by sporulated cells. Strain JW-YL-7 (DSM 7308) is designated as the type strain.

  13. The Selenoproteome of Clostridium sp. OhILAs: Characterization of Anaerobic Bacterial Selenoprotein Methionine Sulfoxide Reductase A

    PubMed Central

    Kim, Hwa-Young; Zhang, Yan; Lee, Byung Cheon; Kim, Jae-Ryong; Gladyshev, Vadim N.

    2008-01-01

    SUMMARY Selenocysteine (Sec) is incorporated into proteins in response to UGA codons. This residue is frequently found at the catalytic sites of oxidoreductases. In the present study, we characterized the selenoproteome of an anaerobic bacterium, Clostridium sp. (also known as Alkaliphilus oremlandii) OhILA, and identified 13 selenoprotein genes, 5 of which have not been previously described. One of the detected selenoproteins was methionine sulfoxide reductase A (MsrA), an antioxidant enzyme that repairs oxidatively damaged methionines in a stereospecific manner. To date, little is known about MsrA from anaerobes. We characterized this selenoprotein MsrA which had a single Sec residue at the catalytic site but no cysteine (Cys) residues in the protein sequence. Its SECIS (Sec insertion sequence) element did not resemble those in Escherichia coli. Although with low translational efficiency, the expression of the Clostridium selenoprotein msrA gene in E. coli could be demonstrated by 75Se metabolic labeling, immunoblot analyses, and enzyme assays, indicating that its SECIS element was recognized by the E. coli Sec insertion machinery. We found that the Sec-containing MsrA exhibited at least a 20-fold higher activity than its Cys mutant form, indicating a critical role of Sec in the catalytic activity of the enzyme. Furthermore, our data revealed that the Clostridium MsrA was inefficiently reducible by thioredoxin, which is a typical reducing agent for MsrA, suggesting the use of alternative electron donors in this anaerobic bacterium that directly act on the selenenic acid intermediate and do not require resolving Cys residues. PMID:18767149

  14. Isolation and characterization of an active mannanase-producing anaerobic bacterium, Clostridium tertium KT-5A, from lotus soil.

    PubMed

    Kataoka, N; Tokiwa, Y

    1998-03-01

    Of 10 strains of mannanase-producing anaerobic bacteria isolated from soils and methanogenic sludges, Clostridium tertium KT-5A, which was isolated from lotus soil, produced high amounts of extracellular beta-1,4-mannanase. The isolate was an aerotolerant anaerobe without quinon systems; the cell growth cultivated with no addition of reducing agents was also stable. High yields of mannanase were obtained by inducing enzyme production with galactomannan guar gum and beef extract/peptone as carbon and nitrogen sources, respectively. Fermentation end products on galactomannan fermentation were formate, acetate, lactate, butyrate, carbon dioxide and hydrogen. The extracellular mannanase displayed high activity on galactomannans of locust bean gum galactose/mannose (G/M) ratio 1:4 and spino gum (G/M 1:3), but weak activity on guar gum galactomannan (G/M 1:2) and konjac glucomannan. As far as is known, this is the first report on the isolation of an active mannanase-producing anaerobic bacterium from natural environments.

  15. Deciphering Clostridium tyrobutyricum Metabolism Based on the Whole-Genome Sequence and Proteome Analyses

    PubMed Central

    Lee, Joungmin; Jang, Yu-Sin; Han, Mee-Jung; Kim, Jin Young

    2016-01-01

    ABSTRACT Clostridium tyrobutyricum is a Gram-positive anaerobic bacterium that efficiently produces butyric acid and is considered a promising host for anaerobic production of bulk chemicals. Due to limited knowledge on the genetic and metabolic characteristics of this strain, however, little progress has been made in metabolic engineering of this strain. Here we report the complete genome sequence of C. tyrobutyricum KCTC 5387 (ATCC 25755), which consists of a 3.07-Mbp chromosome and a 63-kbp plasmid. The results of genomic analyses suggested that C. tyrobutyricum produces butyrate from butyryl-coenzyme A (butyryl-CoA) through acetate reassimilation by CoA transferase, differently from Clostridium acetobutylicum, which uses the phosphotransbutyrylase-butyrate kinase pathway; this was validated by reverse transcription-PCR (RT-PCR) of related genes, protein expression levels, in vitro CoA transferase assay, and fed-batch fermentation. In addition, the changes in protein expression levels during the course of batch fermentations on glucose were examined by shotgun proteomics. Unlike C. acetobutylicum, the expression levels of proteins involved in glycolytic and fermentative pathways in C. tyrobutyricum did not decrease even at the stationary phase. Proteins related to energy conservation mechanisms, including Rnf complex, NfnAB, and pyruvate-phosphate dikinase that are absent in C. acetobutylicum, were identified. Such features explain why this organism can produce butyric acid to a much higher titer and better tolerate toxic metabolites. This study presenting the complete genome sequence, global protein expression profiles, and genome-based metabolic characteristics during the batch fermentation of C. tyrobutyricum will be valuable in designing strategies for metabolic engineering of this strain. PMID:27302759

  16. Ferredoxin and Formyltetrahydrofolate Synthetase: Comparative Studies with Clostridium acidiurici, Clostridium cylindrosporum, and Newly Isolated Anaerobic Uric Acid-Fermenting Strains

    PubMed Central

    Champion, Alexander B.; Rabinowitz, Jesse C.

    1977-01-01

    Six strains of Clostridium acidiurici and three strains of C. cylindrosporum were isolated from soil samples by enrichment culture with uric acid as the source of carbon, nitrogen, and energy. The newly isolated strains were characterized by their spore morphology and the amounts of glycine and formate formed by the fermentation of uric acid. The strains were easily identified as belonging to one species or the other on the basis of spore morphology and formate production. The crystal properties and spectra of the native ferredoxins of all the strains isolated and the amino acid composition and partial carboxy-terminal sequence of all their apoferredoxins were determined. All the ferredoxins were tested for cross-reactivity with antiserum to C. acidiurici ferredoxin by microcomplement fixation. Five of the six C. acidiurici strains, which had ferredoxins with amino acid compositions identical to that from C. acidiurici, also showed immunological identity (immunological distance = 0.0). These results suggest sequence identity. The one strain with a different amino acid composition failed to show complete cross-reactivity. Two of the three C. cylindrosporum strains have ferredoxin amino acid compositions identical to that from C. cylindrosporum. The third strain had a minimum of five differences in sequence. All C. cylindrosporum strains had ferredoxins that differed considerably from C. acidiurici strains (minimum of eight to nine differences), and none of these ferredoxins cross-reacted with antisera to C. acidiurici ferredoxin. Antisera were prepared to formyltetrahydrofolate synthetase from C. acidiurici and C. cylindrosporum, and all possible comparisons were made by using immunodiffusion and microcomplement fixation. There is more intraspecies variation in the synthetases than in the ferredoxins; however, the results suggest considerable interspecies differences in both proteins. These results suggest a low degree of genomic relatedness between the two species

  17. Clostridium guangxiense sp. nov. and Clostridium neuense sp. nov., two phylogenetically closely related hydrogen-producing species isolated from lake sediment.

    PubMed

    Zhao, Xin; Li, Danyang; Xu, Shuhong; Guo, Zhanghao; Zhang, Yan; Man, Lin; Jiang, Binhui; Hu, Xiaomin

    2017-03-01

    Two novel anaerobic, mesophilic, biohydrogen-producing bacteria, designated strains ZGM211T and G1T, were isolated from lake sediment. 16S rRNA and ATP synthase beta subunit (atpD) gene sequences and phylogenetic analysis of strains ZGM211T and G1T revealed an affiliation to the genus Clostridium sensu stricto (cluster I of the clostridia), with Clostridium acetobutylicum as the closest characterized species, showing the same sequence similarity of 96.4 % to the type strain (98.9 % between the two isolates). Cells of the two strains were rod shaped. Growth occurred at 20-45 °C, pH 4.0-8.0 and NaCl concentrations up to 2 % (w/v). Grown on glucose, the main fermentation products were H2, CO2, acetate and butyrate. The major fatty acids were C14 : 0 and C16 : 0. The DNA G+C contents of strains ZGM211T and G1T were 40.7 and 41.5 mol%, respectively. Based on phenotypic, chemotaxonomic and phylogenetic differences, strains ZGM211T (=CICC 24070T=BCRC 80950T) and G1T (=CICC 24069T=BCRC 80949T) are proposed as the type strains of novel species of the genus Clostridium with the names Clostridium guangxiense sp. nov. and Clostridium neuense sp. nov., respectively.

  18. Anaerobic

    MedlinePlus

    ... shock. Anaerobic is the opposite of aerobic . In exercise, our bodies need to perform both anaerobic and aerobic reactions ... during shorter, more intense activities like sprinting. Anaerobic ... removing the lactic acid by providing oxygen to their bodies.

  19. Physiology and Sporulation in Clostridium.

    PubMed

    Dürre, Peter

    2014-08-01

    Clostridia are Gram-positive, anaerobic, endospore-forming bacteria, incapable of dissimilatory sulfate reduction. Comprising approximately 180 species, the genus Clostridium is one of the largest bacterial genera. Physiology is mostly devoted to acid production. Numerous pathways are known, such as the homoacetate fermentation by acetogens, the propionate fermentation by Clostridium propionicum, and the butyrate/butanol fermentation by C. acetobutylicum, a well-known solvent producer. Clostridia degrade sugars, alcohols, amino acids, purines, pyrimidines, and polymers such as starch and cellulose. Energy conservation can be performed by substrate-level phosphorylation as well as by the generation of ion gradients. Endospore formation resembles the mechanism elucidated in Bacillus. Morphology, contents, and properties of spores are very similar to bacilli endospores. Sporulating clostridia usually form swollen mother cells and accumulate the storage substance granulose. However, clostridial sporulation differs by not employing the so-called phosphorelay. Initiation starts by direct phosphorylation of the master regulator Spo0A. The cascade of sporulation-specific sigma factors is again identical to what is known from Bacillus. The onset of sporulation is coupled in some species to either solvent (acetone, butanol) or toxin (e.g., C. perfringens enterotoxin) formation. The germination of spores is often induced by various amino acids, often in combination with phosphate and sodium ions. In medical applications, C. butyricum spores are used as a C. difficile prophylaxis and as treatment against diarrhea. Recombinant spores are currently under investigation and testing as antitumor agents, because they germinate only in hypoxic tissues (i.e., tumor tissue), allowing precise targeting and direct killing of tumor cells.

  20. Co-utilization of glycerol and lignocellulosic hydrolysates enhances anaerobic 1,3-propanediol production by Clostridium diolis

    PubMed Central

    Xin, Bo; Wang, Yu; Tao, Fei; Li, Lixiang; Ma, Cuiqing; Xu, Ping

    2016-01-01

    Anaerobic fermentation using lignocellulosic hydrolysates as co-substrates is an economically attractive method to enhance 1,3-propanediol (1,3-PD) production by increasing the conversion yield from glycerol. Lignocellulosic hydrolysates contain the mixed sugars that are primarily glucose, xylose, and arabinose. Therefore, these three individual sugars were used, separately, as co-substrates with glycerol, in 1,3-PD production by a Clostridium diolis strain DSM 15410, resulting in an 18%–28% increase in the 1,3-PD yield. Co-fermentation of the mixed sugars and glycerol obtained a higher intracellular NADH/NAD+ ratio and increased the 1,3-PD yield by 22% relative to fermentation of glycerol alone. Thereafter, two kinds of lignocellulosic hydrolysates, corn stover hydrolysate and corncob molasses, were individually co-fermented with glycerol. The maximum 1,3-PD yield from glycerol reached 0.85 mol/mol. Fed-batch co-fermentation was also performed, improving the 1,3-PD yield (from 0.62 mol/mol to 0.82 mol/mol). These results demonstrate that the co-fermentation strategy is an efficient and economical way to produce 1,3-PD from glycerol. PMID:26750307

  1. Co-utilization of glycerol and lignocellulosic hydrolysates enhances anaerobic 1,3-propanediol production by Clostridium diolis.

    PubMed

    Xin, Bo; Wang, Yu; Tao, Fei; Li, Lixiang; Ma, Cuiqing; Xu, Ping

    2016-01-11

    Anaerobic fermentation using lignocellulosic hydrolysates as co-substrates is an economically attractive method to enhance 1,3-propanediol (1,3-PD) production by increasing the conversion yield from glycerol. Lignocellulosic hydrolysates contain the mixed sugars that are primarily glucose, xylose, and arabinose. Therefore, these three individual sugars were used, separately, as co-substrates with glycerol, in 1,3-PD production by a Clostridium diolis strain DSM 15410, resulting in an 18%-28% increase in the 1,3-PD yield. Co-fermentation of the mixed sugars and glycerol obtained a higher intracellular NADH/NAD(+) ratio and increased the 1,3-PD yield by 22% relative to fermentation of glycerol alone. Thereafter, two kinds of lignocellulosic hydrolysates, corn stover hydrolysate and corncob molasses, were individually co-fermented with glycerol. The maximum 1,3-PD yield from glycerol reached 0.85 mol/mol. Fed-batch co-fermentation was also performed, improving the 1,3-PD yield (from 0.62 mol/mol to 0.82 mol/mol). These results demonstrate that the co-fermentation strategy is an efficient and economical way to produce 1,3-PD from glycerol.

  2. Laboratory identification of anaerobic bacteria isolated on Clostridium difficile selective medium.

    PubMed

    Rodriguez, Cristina; Warszawski, Nathalie; Korsak, Nicolas; Taminiau, Bernard; Van Broeck, Johan; Delmée, Michel; Daube, Georges

    2016-06-01

    Despite increasing interest in the bacterium, the methodology for Clostridium difficile recovery has not yet been standardized. Cycloserine-cefoxitin fructose taurocholate (CCFT) has historically been the most used medium for C. difficile isolation from human, animal, environmental, and food samples, and presumptive identification is usually based on colony morphologies. However, CCFT is not totally selective. This study describes the recovery of 24 bacteria species belonging to 10 different genera other than C. difficile, present in the environment and foods of a retirement establishment that were not inhibited in the C. difficile selective medium. These findings provide insight for further environmental and food studies as well as for the isolation of C. difficile on supplemented CCFT.

  3. Wukongibacter baidiensis gen. nov., sp. nov., an anaerobic bacterium isolated from hydrothermal sulfides, and proposal for the reclassification of the closely related Clostridium halophilum and Clostridium caminithermale within Maledivibacter gen. nov. and Paramaledivibacter gen. nov., respectively.

    PubMed

    Li, Guangyu; Zeng, Xiang; Liu, Xiupian; Zhang, Xiaobo; Shao, Zongze

    2016-11-01

    An anaerobic, Gram-stain-positive, spore-forming bacterium, designated DY30321T, was isolated from a sample of mixed hydrothermal sulfides collected during cruise DY30 of R/V Da Yang Yi Hao. Cells of strain DY30321T were rod-shaped with rounded ends, and were not motile. Strain DY30321T grew optimally at pH 8.0, at 30 °C and at a salinity (sea salts) of 30-40 g l-1. The principal fatty acids of strain DY30321T were C14 : 0 and summed feature 1 (comprising iso H-C15 : 1/C13 : 0 3-OH). The predominant polar lipids of strain DY30321T were diphosphatidylglycerol, phosphatidylcholine and phosphatidylethanolamine. No respiratory quinone was detected. The G+C content of the genomic DNA of strain DY30321T was 33.4 mol%. Phylogenetically, strain DY30321T branched within the family Peptostreptococcaceae, with (misclassified) Clostridium halophilum M1T being its closest phylogenetic relative (94.6 % 16S rRNA gene sequence similarity), followed by (misclassified) Clostridium caminithermale DVird3T (92.1 %). These strains showed very low 16S rRNA gene sequence similarity (<84 %) to Clostrdium butyricum ATCC 19398T, the type species of the genus Clostridium sensu stricto. On the basis of its phenotypic, phylogenetic and chemotaxonomic characteristics, strain DY30321T (=KCTC 15549T=MCCC 1A01532T) is considered as the type strain of a novel species of a new genus in the family Peptostreptococcaceae, for which the name Wukongibacterbaidiensis gen. nov., sp. nov. is proposed. Maledivibacter gen. nov. is proposed to accommodate Clostridium halophilum as Maledivibacter halophilus comb. nov. (type species of the genus), and Paramaledivibacter gen. nov. to accommodate Clostridium caminithermale as Paramaledivibacter caminithermalis comb. nov. (type species of the genus).

  4. Uncoupler-Resistant Glucose Uptake by the Thermophilic Glycolytic Anaerobe Thermoanaerobacter thermosulfuricus (Clostridium thermohydrosulfuricum)

    PubMed Central

    Cook, Gregory M.; Janssen, Peter H.; Morgan, Hugh W.

    1993-01-01

    The transport of glucose across the bacterial cell membrane of Thermoanaerobacter thermosulfuricus (Clostridium thermohydrosulfuricum) Rt8.B1 was governed by a permease which did not catalyze concomitant substrate transport and phosphorylation and thus was not a phosphoenolpyruvate-dependent phosphotransferase. Glucose uptake was carrier mediated, could not be driven by an artificial membrane potential (Δψ) in the presence or absence of sodium, and was not sensitive to inhibitors which dissipate the proton motive force (Δp; tetrachlorosalicylanilide, N,N-dicyclohexylcarboiimide, and 2,4-dinitrophenol), and no uptake of the nonmetabolizable analog 2-deoxyglucose could be demonstrated. The glucokinase apparent Km for glucose (0.21 mM) was similar to the Kt (affinity constant) for glucose uptake (0.15 mM), suggesting that glucokinase controls the rate of glucose uptake. Inhibitors of ATP synthesis (iodoacetate and sodium fluoride) also inhibited glucose uptake, and this effect was due to a reduction in the level of ATP available to glucokinase for glucose phosphorylation. These results indicated that T. thermosulfuricus Rt8.B1 lacks a concentrative uptake system for glucose and that uptake is via facilitated diffusion, followed by ATP-dependent phosphorylation by glucokinase. In T. thermosulfuricus Rt8.B1, glucose is metabolized by the Embden-Meyerhof-Parnas pathway, which yields 2 mol of ATP (G. M. Cook, unpublished data). Since only 1 mol of ATP is used to transport 1 mol of glucose, the energetics of this system are therefore similar to those found in bacteria which possess a phosphotransferase. PMID:16349043

  5. The genome sequence of Clostridium tetani, the causative agent of tetanus disease

    PubMed Central

    Brüggemann, Holger; Bäumer, Sebastian; Fricke, Wolfgang Florian; Wiezer, Arnim; Liesegang, Heiko; Decker, Iwona; Herzberg, Christina; Martínez-Arias, Rosa; Merkl, Rainer; Henne, Anke; Gottschalk, Gerhard

    2003-01-01

    Tetanus disease is one of the most dramatic and globally prevalent diseases of humans and vertebrate animals, and has been reported for over 24 centuries. The manifestation of the disease, spastic paralysis, is caused by the second most poisonous substance known, the tetanus toxin, with a human lethal dose of ≈1 ng/kg. Fortunately, this disease is successfully controlled through immunization with tetanus toxoid; nevertheless, according to the World Health Organization, an estimated 400,000 cases still occur each year, mainly of neonatal tetanus. The causative agent of tetanus disease is Clostridium tetani, an anaerobic spore-forming bacterium, whose natural habitat is soil, dust, and intestinal tracts of various animals. Here we report the complete genome sequence of toxigenic C. tetani E88, a variant of strain Massachusetts. The genome consists of a 2,799,250-bp chromosome encoding 2,372 ORFs. The tetanus toxin and a collagenase are encoded on a 74,082-bp plasmid, containing 61 ORFs. Additional virulence-related factors could be identified, such as an array of surface-layer and adhesion proteins (35 ORFs), some of them unique to C. tetani. Comparative genomics with the genomes of Clostridium perfringens, the causative agent of gas gangrene, and Clostridium acetobutylicum, a nonpathogenic solvent producer, revealed a remarkable capacity of C. tetani: The organism can rely on an extensive sodium ion bioenergetics. Additional candidate genes involved in the establishment and maintenance of a pathogenic lifestyle of C. tetani are presented. PMID:12552129

  6. The genome sequence of Clostridium tetani, the causative agent of tetanus disease.

    PubMed

    Bruggemann, Holger; Baumer, Sebastian; Fricke, Wolfgang Florian; Wiezer, Arnim; Liesegang, Heiko; Decker, Iwona; Herzberg, Christina; Martinez-Arias, Rosa; Merkl, Rainer; Henne, Anke; Gottschalk, Gerhard

    2003-02-04

    Tetanus disease is one of the most dramatic and globally prevalent diseases of humans and vertebrate animals, and has been reported for over 24 centuries. The manifestation of the disease, spastic paralysis, is caused by the second most poisonous substance known, the tetanus toxin, with a human lethal dose of approximately 1 ng/kg. Fortunately, this disease is successfully controlled through immunization with tetanus toxoid; nevertheless, according to the World Health Organization, an estimated 400,000 cases still occur each year, mainly of neonatal tetanus. The causative agent of tetanus disease is Clostridium tetani, an anaerobic spore-forming bacterium, whose natural habitat is soil, dust, and intestinal tracts of various animals. Here we report the complete genome sequence of toxigenic C. tetani E88, a variant of strain Massachusetts. The genome consists of a 2,799,250-bp chromosome encoding 2,372 ORFs. The tetanus toxin and a collagenase are encoded on a 74,082-bp plasmid, containing 61 ORFs. Additional virulence-related factors could be identified, such as an array of surface-layer and adhesion proteins (35 ORFs), some of them unique to C. tetani. Comparative genomics with the genomes of Clostridium perfringens, the causative agent of gas gangrene, and Clostridium acetobutylicum, a nonpathogenic solvent producer, revealed a remarkable capacity of C. tetani: The organism can rely on an extensive sodium ion bioenergetics. Additional candidate genes involved in the establishment and maintenance of a pathogenic lifestyle of C. tetani are presented.

  7. FT-IR spectroscopic analysis for studying Clostridium cell response to conversion of enzymatically hydrolyzed hay

    NASA Astrophysics Data System (ADS)

    Grube, Mara; Gavare, Marita; Nescerecka, Alina; Tihomirova, Kristina; Mezule, Linda; Juhna, Talis

    2013-07-01

    Grass hay is one of assailable cellulose containing non-food agricultural wastes that can be used as a carbohydrate source by microorganisms producing biofuels. In this study three Clostridium strains Clostridium acetobutylicum, Clostridium beijerinckii and Clostridium tetanomorphum, capable of producing acetone, butanol and ethanol (ABE) were adapted to convert enzymatically hydrolyzed hay used as a growth media additive. The results of growth curves, substrate degradation kinetics and FT-IR analyses of bacterial biomass macromolecular composition showed diverse strain-specific cell response to the growth medium composition.

  8. Engineering Clostridium Strain to Accept Unmethylated DNA

    PubMed Central

    Dong, Hongjun; Zhang, Yanping; Dai, Zongjie; Li, Yin

    2010-01-01

    It is difficult to genetically manipulate the medically and biotechnologically important genus Clostridium due to the existence of the restriction and modification (RM) systems. We identified and engineered the RM system of a model clostridial species, C. acetobutylicum, with the aim to allow the host to accept the unmethylated DNA efficiently. A gene CAC1502 putatively encoding the type II restriction endonuclease Cac824I was identified from the genome of C. acetobutylicum DSM1731, and disrupted using the ClosTron system based on group II intron insertion. The resulting strain SMB009 lost the type II restriction endonuclease activity, and can be transformed with unmethylated DNA as efficiently as with methylated DNA. The strategy reported here makes it easy to genetically modify the clostridial species using unmethylated DNA, which will help to advance the understanding of the clostridial physiology from the molecular level. PMID:20161730

  9. The VirS/VirR two-component system regulates the anaerobic cytotoxicity, intestinal pathogenicity, and enterotoxemic lethality of Clostridium perfringens type C isolate CN3685.

    PubMed

    Ma, Menglin; Vidal, Jorge; Saputo, Juliann; McClane, Bruce A; Uzal, Francisco

    2011-01-25

    Clostridium perfringens vegetative cells cause both histotoxic infections (e.g., gas gangrene) and diseases originating in the intestines (e.g., hemorrhagic necrotizing enteritis or lethal enterotoxemia). Despite their medical and veterinary importance, the molecular pathogenicity of C. perfringens vegetative cells causing diseases of intestinal origin remains poorly understood. However, C. perfringens beta toxin (CPB) was recently shown to be important when vegetative cells of C. perfringens type C strain CN3685 induce hemorrhagic necrotizing enteritis and lethal enterotoxemia. Additionally, the VirS/VirR two-component regulatory system was found to control CPB production by CN3685 vegetative cells during aerobic infection of cultured enterocyte-like Caco-2 cells. Using an isogenic virR null mutant, the current study now reports that the VirS/VirR system also regulates CN3685 cytotoxicity during infection of Caco-2 cells under anaerobic conditions, as found in the intestines. More importantly, the virR mutant lost the ability to cause hemorrhagic necrotic enteritis in rabbit small intestinal loops. Western blot analyses demonstrated that the VirS/VirR system mediates necrotizing enteritis, at least in part, by controlling in vivo CPB production. In addition, vegetative cells of the isogenic virR null mutant were, relative to wild-type vegetative cells, strongly attenuated in their lethality in a mouse enterotoxemia model. Collectively, these results identify the first regulator of in vivo pathogenicity for C. perfringens vegetative cells causing disease originating in the complex intestinal environment. Since VirS/VirR also mediates histotoxic infections, this two-component regulatory system now assumes a global role in regulating a spectrum of infections caused by C. perfringens vegetative cells.

  10. Metabolic properties of Eubacterium pyruvativorans, a ruminal 'hyper-ammonia-producing' anaerobe with metabolic properties analogous to those of Clostridium kluyveri.

    PubMed

    Wallace, R John; Chaudhary, Lal C; Miyagawa, Eiichi; McKain, N; Walker, Nicola D

    2004-09-01

    Eubacterium pyruvativorans I-6(T) is a non-saccharolytic, amino-acid-fermenting anaerobe from the rumen, isolated by its ability to grow on pancreatic casein hydrolysate (PCH) as sole C source. This study investigated its metabolic properties and its likely ecological niche. Additional growth was supported by pyruvate, vinyl acetate, and, to a lesser extent, lactate and crotonate, and also by a mixture of amino acids (alanine, glycine, serine and threonine) predicted to be catabolized to pyruvate. No single amino acid supported growth, and peptides were required for growth on amino acids. Alanine, followed by leucine, serine and proline, were used most extensively during growth, but only alanine and asparate were extensively modified before incorporation. Growth on PCH, but not on pyruvate, was increased by the addition of acetate, propionate and butyrate. l-Lactate was fermented incompletely, mainly to acetate, but no lactate-C was incorporated. Propionate and butyrate were utilized during growth, forming valerate and caproate, respectively. Labelling experiments suggested a metabolic pattern where two C atoms of butyrate, valerate and caproate were derived from amino acids, with the others being formed from acetate, propionate and butyrate. The metabolic strategy of E. pyruvativorans therefore resembles that of Clostridium kluyveri, which ferments ethanol only when the reaction is coupled to acetate, propionate or butyrate utilization. The fermentative niche of E. pyruvativorans appears to be to scavenge amino acids, lactate and possibly other metabolites in order to generate ATP via acetate formation, using volatile fatty acid elongation with C(2) units derived from other substrates to dispose of reducing equivalents.

  11. The VirS/VirR Two-Component System Regulates the Anaerobic Cytotoxicity, Intestinal Pathogenicity, and Enterotoxemic Lethality of Clostridium perfringens Type C Isolate CN3685

    PubMed Central

    Ma, Menglin; Vidal, Jorge; Saputo, Juliann; McClane, Bruce A.; Uzal, Francisco

    2011-01-01

    Clostridium perfringens vegetative cells cause both histotoxic infections (e.g., gas gangrene) and diseases originating in the intestines (e.g., hemorrhagic necrotizing enteritis or lethal enterotoxemia). Despite their medical and veterinary importance, the molecular pathogenicity of C. perfringens vegetative cells causing diseases of intestinal origin remains poorly understood. However, C. perfringens beta toxin (CPB) was recently shown to be important when vegetative cells of C. perfringens type C strain CN3685 induce hemorrhagic necrotizing enteritis and lethal enterotoxemia. Additionally, the VirS/VirR two-component regulatory system was found to control CPB production by CN3685 vegetative cells during aerobic infection of cultured enterocyte-like Caco-2 cells. Using an isogenic virR null mutant, the current study now reports that the VirS/VirR system also regulates CN3685 cytotoxicity during infection of Caco-2 cells under anaerobic conditions, as found in the intestines. More importantly, the virR mutant lost the ability to cause hemorrhagic necrotic enteritis in rabbit small intestinal loops. Western blot analyses demonstrated that the VirS/VirR system mediates necrotizing enteritis, at least in part, by controlling in vivo CPB production. In addition, vegetative cells of the isogenic virR null mutant were, relative to wild-type vegetative cells, strongly attenuated in their lethality in a mouse enterotoxemia model. Collectively, these results identify the first regulator of in vivo pathogenicity for C. perfringens vegetative cells causing disease originating in the complex intestinal environment. Since VirS/VirR also mediates histotoxic infections, this two-component regulatory system now assumes a global role in regulating a spectrum of infections caused by C. perfringens vegetative cells. PMID:21264065

  12. Scale-up of anaerobic 1,3-propanediol production by Clostridium butyricum DSP1 from crude glycerol

    PubMed Central

    2014-01-01

    Background As the production of biofuels from raw materials continuously increases, optimization of production processes is necessary. A very important issue is the development of wasteless methods of biodiesel production. One way of utilization of glycerol generated in biodiesel production is its microbial conversion to 1,3-PD (1,3-propanediol). Results The study investigated the scale-up of 1,3-PD synthesis from crude glycerol by Clostridium butyricum. Batch fermentations were carried out in 6.6 L, 42 L and 150 L bioreactors. It was observed that cultivation of C. butyricum on a pilot scale did not decrease the efficiency of 1,3-PD production. The highest concentrations of 1,3-PD, 37 g/L for batch fermentation and 71 g/L for fed-batch fermentation, were obtained in the 6.6 L bioreactor. The kinetic parameters of 1,3-PD synthesis from crude glycerol established for batch fermentation were similar regarding all three bioreactor capacities. During fed-batch fermentation, the concentration of 1,3-PD in the 150 L bioreactor was lower and the substrate was not completely utilized. That suggested the presence of multifunctional environmental stresses in the 150 L bioreactor, which was confirmed by protein analysis. Conclusion The values of effectivity parameters for 1,3-PD synthesis in batch fermentations carried out in 6.6 L, 42 L and 150 L bioreactors were similar. The parameters obtained during fed-batch fermentations in the 150 L bioreactor differed in the rate and percentage of substrate utilization. The analysis of cell proteins demonstrated that a number of multifunctional stresses occurred during fed-batch fermentations in the 150 L bioreactor, which suggests the possibility of identifying the key stages in the biochemical process where inhibition of 1,3-PD synthesis pathways can be observed. PMID:24555775

  13. Draft Genome Sequence of the Cellulolytic Strain Clostridium sp. Bc-iso-3 Isolated from an Industrial-Scale Anaerobic Digester

    PubMed Central

    2016-01-01

    Clostridium sp. Bc-iso-3 is a cellulolytic strain isolated from a Swedish industrial-scale biogas digester. Here, we present the draft genome sequence of this strain, which consists of four contigs with a total length of 4,327,139 bp and an average coverage of 312.97×. PMID:27789641

  14. Anaerobic thermophilic culture system

    DOEpatents

    Ljungdahl, Lars G.; Wiegel, Jurgen K. W.

    1981-01-01

    A mixed culture system of the newly discovered microorganism Thermoanaerobacter ethanolicus ATCC31550 and the microorganism Clostridium thermocellum ATCC31549 is described. In a mixed nutrient culture medium that contains cellulose, these microorganisms have been coupled and cultivated to efficiently ferment cellulose to produce recoverable quantities of ethanol under anaerobic, thermophilic conditions.

  15. An unexpected negative influence of light intensity on hydrogen production by dark fermentative bacteria Clostridium beijerinckii.

    PubMed

    Zagrodnik, R; Laniecki, M

    2016-01-01

    The role of light intensity on biohydrogen production from glucose by Clostridium beijerinckii, Clostridium acetobutylicum, and Rhodobacter sphaeroides was studied to evaluate the performance and possible application in co-culture fermentation system. The applied source of light had spectrum similar to the solar radiation. The influence of light intensity on hydrogen production in dark process by C. acetobutylicum was negligible. In contrast, dark fermentation by C. beijerinckii bacteria showed a significant decrease (83%) in produced hydrogen at light intensity of 540W/m(2). Here, the redirection of metabolism from acetic and butyric acid formation towards lactic acid was observed. This not yet reported effect was probably caused by irradiation of these bacteria by light within UVA range, which is an important component of the solar radiation. The excessive illumination with light of intensity higher than 200W/m(2) resulted in decrease in hydrogen production with photofermentative bacteria as well.

  16. Small RNAs in the genus Clostridium.

    PubMed

    Chen, Yili; Indurthi, Dinesh C; Jones, Shawn W; Papoutsakis, Eleftherios T

    2011-01-25

    The genus Clostridium includes major human pathogens and species important to cellulose degradation, the carbon cycle, and biotechnology. Small RNAs (sRNAs) are emerging as crucial regulatory molecules in all organisms, but they have not been investigated in clostridia. Research on sRNAs in clostridia is hindered by the absence of a systematic method to identify sRNA candidates, thus delegating clostridial sRNA research to a hit-and-miss process. Thus, we wanted to develop a method to identify potential sRNAs in the Clostridium genus to open up the field of sRNA research in clostridia. Using comparative genomics analyses combined with predictions of rho-independent terminators and promoters, we predicted sRNAs in 21 clostridial genomes: Clostridium acetobutylicum, C. beijerinckii, C. botulinum (eight strains), C. cellulolyticum, C. difficile, C. kluyveri (two strains), C. novyi, C. perfringens (three strains), C. phytofermentans, C. tetani, and C. thermocellum. Although more than one-third of predicted sRNAs have Shine-Dalgarno (SD) sequences, only one-sixth have a start codon downstream of SD sequences; thus, most of the predicted sRNAs are noncoding RNAs. Quantitative reverse transcription-PCR (Q-RT-PCR) and Northern analysis were employed to test the presence of a randomly chosen set of sRNAs in C. acetobutylicum and several C. botulinum strains, leading to the confirmation of a large fraction of the tested sRNAs. We identified a conserved, novel sRNA which, together with the downstream gene coding for an ATP-binding cassette (ABC) transporter gene, responds to the antibiotic clindamycin. The number of predicted sRNAs correlated with the physiological function of the species (high for pathogens, low for cellulolytic, and intermediate for solventogenic), but not with 16S rRNA-based phylogeny.

  17. Extending CRISPR-Cas9 Technology from Genome Editing to Transcriptional Engineering in the Genus Clostridium.

    PubMed

    Bruder, Mark R; Pyne, Michael E; Moo-Young, Murray; Chung, Duane A; Chou, C Perry

    2016-10-15

    The discovery and exploitation of the prokaryotic adaptive immunity system based on clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins have revolutionized genetic engineering. CRISPR-Cas tools have enabled extensive genome editing as well as efficient modulation of the transcriptional program in a multitude of organisms. Progress in the development of genetic engineering tools for the genus Clostridium has lagged behind that of many other prokaryotes, presenting the CRISPR-Cas technology an opportunity to resolve a long-existing issue. Here, we applied the Streptococcus pyogenes type II CRISPR-Cas9 (SpCRISPR-Cas9) system for genome editing in Clostridium acetobutylicum DSM792. We further explored the utility of the SpCRISPR-Cas9 machinery for gene-specific transcriptional repression. For proof-of-concept demonstration, a plasmid-encoded fluorescent protein gene was used for transcriptional repression in C. acetobutylicum Subsequently, we targeted the carbon catabolite repression (CCR) system of C. acetobutylicum through transcriptional repression of the hprK gene encoding HPr kinase/phosphorylase, leading to the coutilization of glucose and xylose, which are two abundant carbon sources from lignocellulosic feedstocks. Similar approaches based on SpCRISPR-Cas9 for genome editing and transcriptional repression were also demonstrated in Clostridium pasteurianum ATCC 6013. As such, this work lays a foundation for the derivation of clostridial strains for industrial purposes.

  18. Complementation of a Clostridium perfringens spo0A mutant with wild-type spo0A from other Clostridium species.

    PubMed

    Huang, I-Hsiu; Sarker, Mahfuzur R

    2006-09-01

    To evaluate whether C. perfringens can be used as a model organism for studying the sporulation process in other clostridia, C. perfringens spo0A mutant IH101 was complemented with wild-type spo0A from four different Clostridium species. Wild-type spo0A from C. acetobutylicum or C. tetani, but not from C. botulinum or C. difficile, restored sporulation and enterotoxin production in IH101. The ability of spo0A from C. botulinum or C. difficile to complement the lack of spore formation in IH101 might be due, at least in part, to the low levels of spo0A transcription and Spo0A production.

  19. Complete Genome Sequence of Clostridium clariflavum DSM 19732

    SciTech Connect

    Goodwin, Lynne A.; Davenport, Karen W.; Teshima, Hazuki; Bruce, David; Detter, J. Chris; Tapia, Roxanne; Han, Cliff; Land, Miriam L; Hauser, Loren John; Jeffries, Cynthia; Han, James; Pitluck, Sam; Nolan, Matt; Chen, Amy; Huntemann, Marcel; Mavromatis, K; Mikhailova, Natalia; Liolios, Konstantinos; Woyke, Tanja; Lynd, Lee R

    2012-01-01

    Clostridium clariflavum is a Cluster III Clostridium within the family Clostridiaceae isolated from thermophilic anaerobic sludge (Shiratori et al, 2009). This species is of interest because of its similarity to the model cellulolytic organism Clostridium thermocellum and for the ability of environmental isolates to break down cellulose and hemicellulose. Here we describe features of the 4,897,678 bp long genome and its annotation, consisting of 4,131 proteincoding and 98 RNA genes, for the type strain DSM 19732.

  20. Genome Sequence of Clostridium tunisiense TJ, Isolated from Drain Sediment from a Pesticide Factory

    PubMed Central

    Sun, Lili; Wang, Yu; Yu, Chunyan; Zhao, Yongqin

    2012-01-01

    Clostridium tunisiense is a Gram-positive, obligate anaerobe that was first isolated in an anaerobic evironment under eutrophication. Here we report the first genome sequence of the Clostridium tunisiense TJ isolated from drain sediment of a pesticide factory in Tianjin, China. The genome is of great importance for both basic and application research. PMID:23209212

  1. Genome sequence of Clostridium tunisiense TJ, isolated from drain sediment from a pesticide factory.

    PubMed

    Sun, Lili; Wang, Yu; Yu, Chunyan; Zhao, Yongqin; Gan, Yinbo

    2012-12-01

    Clostridium tunisiense is a Gram-positive, obligate anaerobe that was first isolated in an anaerobic environment under eutrophication. Here we report the first genome sequence of the Clostridium tunisiense TJ isolated from drain sediment of a pesticide factory in Tianjin, China. The genome is of great importance for both basic and application research.

  2. Influence of high gas production during thermophilic anaerobic digestion in pilot-scale and lab-scale reactors on survival of the thermotolerant pathogens Clostridium perfringens and Campylobacter jejuni in piggery wastewater.

    PubMed

    Skillman, L C; Bajsa, O; Ho, L; Santhanam, B; Kumar, M; Ho, G

    2009-07-01

    Safe reuse of animal wastes to capture energy and nutrients, through anaerobic digestion processes, is becoming an increasingly desirable solution to environmental pollution. Pathogen decay is the most important safety consideration and is in general, improved at elevated temperatures and longer hydraulic residence times. During routine sampling to assess pathogen decay in thermophilic digestion, an inversely proportional relationship between levels of Clostridium perfringens and gas production was observed. Further samples were collected from pilot-scale, bench-scale thermophilic reactors and batch scale vials to assess whether gas production (predominantly methane) could be a useful indicator of decay of the thermotolerant pathogens C. perfringens and Campylobacter jejuni. Pathogen levels did appear to be lower where gas production and levels of methanogens were higher. This was evident at each operating temperature (50, 57, 65 degrees C) in the pilot-scale thermophilic digesters, although higher temperatures also reduced the numbers of pathogens detected. When methane production was higher, either when feed rate was increased, or pH was lowered from 8.2 (piggery wastewater) to 6.5, lower numbers of pathogens were detected. Although a number of related factors are known to influence the amount and rate of methane production, it may be a useful indicator of the removal of the pathogens C. perfringens and C. jejuni.

  3. Antipathogenic activity of probiotics against Salmonella Typhimurium and Clostridium difficile in anaerobic batch culture systems: is it due to synergies in probiotic mixtures or the specificity of single strains?

    PubMed

    Tejero-Sariñena, Sandra; Barlow, Janine; Costabile, Adele; Gibson, Glenn R; Rowland, Ian

    2013-12-01

    Probiotics are currently being investigated for prevention of infections caused by enteric pathogens. The aim of this in vitro study was to evaluate the influence of three single probiotics: Lactobacillus casei NCIMB 30185 (PXN 37), Lactobacillus acidophilus NCIMB 30184 (PXN 35), Bifidobacterium breve NCIMB 30180 (PXN 25) and a probiotic mixture containing the above strains plus twelve other strains belonging to the Lactobacillus, Bifidobacterium, Lactococcus, Streptococcus and Bacillus genera on the survival of Salmonella Typhimurium and Clostridium difficile using pH-controlled anaerobic batch cultures containing mixed faecal bacteria. Changes in relevant bacterial groups and effects of probiotic addition on survival of the two pathogens were assessed over 24 h. Quantitative analysis of bacterial populations revealed that there was a significant increase in lactobacilli and/or bifidobacteria numbers, depending on probiotic addition, compared with the control (no added probiotic). There was also a significant reduction in S. Typhimurium and C. difficile numbers in the presence of certain probiotics compared with controls. Of the probiotic treatments, two single strains namely L. casei NCIMB 30185 (PXN 37), and B. breve NCIMB 30180 (PXN 25) were the most potent in reducing the numbers of S. Typhimurium and C. difficile. In addition, the supplementation with probiotics into the systems influenced some fermentations parameters. Acetate was found in the largest concentrations in all vessels and lactate and formate were generally detected in higher amounts in vessels with probiotic addition compared to controls.

  4. Biotechnological potential of Clostridium butyricum bacteria

    PubMed Central

    Szymanowska-Powałowska, Daria; Orczyk, Dorota; Leja, Katarzyna

    2014-01-01

    In response to demand from industry for microorganisms with auspicious biotechnological potential, a worldwide interest has developed in bacteria and fungi isolation. Microorganisms of interesting metabolic properties include non-pathogenic bacteria of the genus Clostridium, particularly C. acetobutylicum, C. butyricum and C. pasteurianum. A well-known property of C. butyricum is their ability to produce butyric acid, as well as effectively convert glycerol to 1,3-propanediol (38.2 g/L). A conversion rate of 0.66 mol 1,3-propanediol/mol of glycerol has been obtained. Results of the studies described in the present paper broaden our knowledge of characteristic features of C. butyricum specific isolates in terms of their phylogenetic affiliation, fermentation capacity and antibacterial properties. PMID:25477923

  5. First Report of Clostridium lavalense Isolated in Human Blood Cultures

    PubMed Central

    Bourque, Christine; Thibault, Louise; Côté, Jean-Charles; Domingo, Marc-Christian

    2016-01-01

    An 88-year-old man was admitted to the hospital with worsening malaise, fever, and weakness. Anaerobic blood culture bottles revealed the presence of an anaerobic, Gram-positive sporulated bacillus. Empirical antibiotherapy with intravenous piperacillin-tazobactam was initiated. The patient defervesced after four days and was switched to oral amoxicillin on his 6th day of antibiotic therapy and later discharged from the hospital. Four months later, he had recovered. The bacterium was initially identified as Clostridium butyricum using anaerobic manual identification panel. 16S rRNA gene sequence and phylogenetic analysis showed the bacterium to be Clostridium lavalense, a recently described species with no previously published case of isolation in human diagnostic samples so far. This is the first report of Clostridium lavalense isolation from human blood cultures. Further studies are needed in order to elucidate the role of Clostridium lavalense in human disease and its virulence factors. PMID:27478446

  6. Comparative Analysis of Clostridium perfringens Bacteriophage

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: Clostridium perfringens are Gram-positive bacteria that are a major bacterial cause of food-borne disease and gas gangrene among humans. These anaerobic bacteria are also the presumptive etiologic agent of necrotic enteritis among chickens. Pathogenesis and symptoms of a necrotic enterit...

  7. Coculture Production of Butanol by Clostridium Bacteria

    NASA Technical Reports Server (NTRS)

    Bergstrom, S. L.; Foutch, G. L.

    1985-01-01

    Production of butanol by anaerobic fermentation of sugars enhanced by use of two Clostridium species, one of which feeds on metabolic product of other. Renewed interest in fermentation process for making butanol stimulated by potential use of butanol as surfactant in enhanced oil recovery. Butanol also used as fuel or as chemical feedstock and currently produced synthetically from petroleum.

  8. Physiology, biochemistry, and genetics of a pure culture of an obligatory anaerobic bacterium that utilizes 2,4,-6-trinitrotoluene (TNT) and biodegradation of RDX by pure cultures of obligatory anaerobic bacteria of the genus clostridium. Final report, 1 September 1993-31 August 1996

    SciTech Connect

    Crawford, R.L.; Crawford, D.L.

    1996-09-01

    In work supported by the US AFOSR (grant F49620-94-1-0306) we are conducting detailed biochemical and genetic studies of three strains of Clostridium bifernientans, obligatory anaerobic bacteria that appear to completely degrade a variety of nitroaromatic compounds, including 2,4,6-trinitrotoluene (TNT). We are determining the optimal physiological conditions for the degradative activities of C. bifermentans strains; and identifying and characterizing enzymes and genes involved in the biotransformation of nitroaromatic compounds by C. bifermentans. In our AASERT supplemental grant(AFOSR-93-1-O464) we expanded these goals to the explosive RDX (1,3,5-triaza-1, 3,5-trinitrocyclohexane). The AASERT grant funded two graduate students, who characterized the ability of C. bifermentans to degrade RDX (Regan, K. N., and R.L. Crawford, 1994. Biotechnol. Kett. 16: 1081- 1086), and prepared both genomic and plasmid DNA libraries from C. bifermentans. This genetic work will accelerate our progress toward our goal of characterizing the genetics of TNT/RDx degradation by our clostridia (K. Diedrich, M.S. thesis, University of Idaho; in preparation).

  9. The acetate kinase of Clostridum acetobutylicum strain P262.

    PubMed

    Diez-Gonzalez, F; Russell, J B; Hunter, J B

    1996-12-01

    Clostridum acetobutylicum strain P262 fermented glucose, pyruvate, or lactate, and the butyrate production was substrate-dependent. Differences in butyrate yield could not be explained by changes in butyrate kinase activities, but the butyrate production was inversely related to acetate kinase activity. The acetate kinase had a pH optimum of 8.0, a Km for acetate of 160 mM, and a kcat of 16, 800 min-1. The enyzme had a native molecular mass of 78 kDa; the size of 42 kDa on SDS-PAGE indicated that the acetate kinase of strain P262 was a homodimer.

  10. Draft Genome Sequence of the Cellulolytic and Xylanolytic Thermophile Clostridium clariflavum Strain 4-2a.

    PubMed

    Rooney, Elise A; Rowe, Kenneth T; Guseva, Anna; Huntemann, Marcel; Han, James K; Chen, Amy; Kyrpides, Nikos C; Mavromatis, Konstantinos; Markowitz, Victor M; Palaniappan, Krishna; Ivanova, Natalia; Pati, Amrita; Liolios, Konstantinos; Nordberg, Henrik P; Cantor, Michael N; Hua, Susan X; Shapiro, Nicole; Woyke, Tanja; Lynd, Lee R; Izquierdo, Javier A

    2015-07-23

    Clostridium clariflavum strain 4-2a, a novel strain isolated from a thermophilic biocompost pile, has demonstrated an extensive capability to utilize both cellulose and hemicellulose under thermophilic anaerobic conditions. Here, we report the draft genome of this strain.

  11. Metabolic control of Clostridium thermocellum via selective inhibition and compensatory product formation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Clostridium thermocellum is a thermophilic, anaerobic bacterium that catabolizes recalcitrant plant fibers such as cellulose. Cellulose is depolymerized by an extracellular, membrane-associated enzyme system, and the sugars are then transported across the cell membrane for fermentation. C. thermoc...

  12. Acetone-butanol-ethanol production from substandard and surplus dates by Egyptian native Clostridium strains.

    PubMed

    Abd-Alla, Mohamed Hemida; Zohri, Abdel-Naser Ahmed; El-Enany, Abdel-Wahab Elsadek; Ali, Shimaa Mohamed

    2015-04-01

    One hundred and seven mesophilic isolates of Clostridium were isolated from agricultural soils cultivated with different plants in Assuit Governorate, Egypt. Eighty isolates (out of 107) showed the ability to produce ABE (Acetone, butanol and ethanol) on T6 medium ranging from 0.036 to 31.89 g/L. The highest numbers of ABE producing isolates were obtained from soil samples of potato contributing 27 isolates, followed by 18 isolates from wheat and 10 isolates from onion. On the other hand, there were three native isolates that produced ABE more than those produced by the reference isolate Clostridium acetobutylicum ATCC 824 (11.543 g/L). The three isolates were identified based on phenotypic and gene encoding 16S rRNA as Clostridium beijerinckii ASU10 (KF372577), Clostridium chauvoei ASU55 (KF372580) and Clostridium roseum ASU58 (KF372581). The highest ABE level from substandard and surplus dates was produced by C. beijerinckii ASU10 (24.07 g/L) comprising butanol 67.15% (16.16 g/L), acetone 30.73% (7.4 g/L) and ethanol 2.12% (0.51 g/L), while C. roseum ASU58 and C. chauvoei ASU55 produced ABE contributing 20.20 and 13.79 g/L, respectively. ABE production by C. acetobutylicum ATCC 824 was 15.01 g/L. This study proved that the native strains C. beijerinckii ASU10 and C. roseum ASU58 have high competitive efficacy on ABE production from economical substrate as substandard and surplus date fruits. Additionally, using this substrate without any nutritional components is considered to be a commercial substrate for desired ABE production.

  13. Two Serious Cases of Infection with Clostridium celatum after 40 Years in Hiding?

    PubMed Central

    Hoegh, Silje Vermedal; Holt, Hanne Marie; Justesen, Ulrik Stenz

    2015-01-01

    Clostridium celatum [ce.la'tum. L. adj. celatum hidden] has been known since 1974, when it was isolated from human feces. In 40 years, no association with human infection has been reported. In this work, we present two serious cases of infection with the anaerobic Gram-positive rod Clostridium celatum. PMID:26560535

  14. Clostridium novyi, sordellii, and tetani: mechanisms of disease.

    PubMed

    Aronoff, David M

    2013-12-01

    Clostridia represent a diverse group of spore-forming gram positive anaerobes that include several pathogenic species. In general, diseases caused by clostridia are a result of intoxication of the infected host. Thus, clostridial toxins have been targeted for diagnostic, therapeutic, and preventive strategies against infection. Studying the mechanisms of action of clostridial toxins has not only shed light on the pathogenesis of infection but has provided important new insights into cell biology and immunology. A primary purpose of this manuscript is to provide a succinct review on the mechanisms of disease caused by intoxication by the pathogens Clostridium tetani, Clostridium novyi, and Clostridium sordellii.

  15. Alternative non-chromatographic method for alcohols determination in Clostridium acetobutylicum fermentations.

    PubMed

    Noriega-Medrano, Laura J; Vega-Estrada, Jesús; Ortega-López, Jaime; Ruiz-Medrano, Roberto; Cristiani-Urbina, Eliseo; Montes-Horcasitas, Maria Del Carmen

    2016-07-01

    An economic, simple, quantitative, and non-chromatographic method for the determination of alcohols using microdiffusion principle has been adapted and validated for acetone-butanol-ethanol (ABE) fermentation samples. This method, based on alcohols oxidation using potassium dichromate in acid medium, and detection by spectrophotometry, was evaluated varying, both, temperature (35°C, 45°C, and 55°C) and reaction time (0 to 125min). With a sample analysis time of 90min at 45°C, a limit of detection (LOD), and a limit of quantification (LOQ) of 0.10, and 0.40g/L, respectively. The proposed method has been successfully applied to determine butanol and ethanol concentrations in ABE fermentation samples with the advantage that multiple samples can be analyzed simultaneously. The measurements obtained with the proposed method were in good agreement with those obtained with the Gas Chromatography Method (GCM). This proposed method is useful for routine analysis of alcohols and screening samples in laboratories and industries.

  16. TRANSFORMATION OF TNT AND RELATED NITROAROMATIC COMPOUNDS BY CLOSTRIDIUM ACETOBUTYLICUM. (R825513C006)

    EPA Science Inventory

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  17. Biobutanol production by Clostridium acetobutylicum using xylose recovered from birch Kraft black liquor.

    PubMed

    Kudahettige-Nilsson, Rasika L; Helmerius, Jonas; Nilsson, Robert T; Sjöblom, Magnus; Hodge, David B; Rova, Ulrika

    2015-01-01

    Acetone-butanol-ethanol (ABE) fermentation was studied using acid-hydrolyzed xylan recovered from hardwood Kraft black liquor by CO2 acidification as the only carbon source. Detoxification of hydrolyzate using activated carbon was conducted to evaluate the impact of inhibitor removal and fermentation. Xylose hydrolysis yields as high as 18.4% were demonstrated at the highest severity hydrolysis condition. Detoxification using active carbon was effective for removal of both phenolics (76-81%) and HMF (38-52%). Batch fermentation of the hydrolyzate and semi-defined P2 media resulted in a total solvent yield of 0.12-0.13g/g and 0.34g/g, corresponding to a butanol concentration of 1.8-2.1g/L and 7.3g/L respectively. This work is the first study of a process for the production of a biologically-derived biofuel from hemicelluloses solubilized during Kraft pulping and demonstrates the feasibility of utilizing xylan recovered directly from industrial Kraft pulping liquors as a feedstock for biological production of biofuels such as butanol.

  18. Molecular characterization of two Clostridium acetobutylicum ATCC 824 butanol dehydrogenase isozyme genes.

    PubMed Central

    Walter, K A; Bennett, G N; Papoutsakis, E T

    1992-01-01

    A 4-kb segment of DNA containing two previously cloned butanol dehydrogenase (BDH) isozyme genes (D. Petersen, R. Welch, F. Rudolph, and G. Bennett, J. Bacteriol. 173:1831-1834, 1991) was sequenced. Two complete open reading frames (ORFs) were identified (bdhA and bdhB), along with a third truncated ORF (ORF1). The translation products of bdhA and bdhB corresponded to the N-terminal sequences of the purified BDH I and BDH II proteins, respectively. The two isozymes had a high amino acid identity (73%) and showed homology to a newly described class of alcohol dehydrogenases. Northern blots revealed that bdhA and bdhB did not form an operon. Primer extension experiments located single transcriptional start sites 37 and 58 bp upstream of the start codons of bdhA and bdhB, respectively. The -10 and -35 promoter regions for these genes were almost identical. bdhA and bdhB were found to be induced or derepressed immediately prior to significant butanol production in controlled pH 5.0 batch fermentations. Images PMID:1385386

  19. Recent progress on industrial fermentative production of acetone-butanol-ethanol by Clostridium acetobutylicum in China.

    PubMed

    Ni, Ye; Sun, Zhihao

    2009-06-01

    China is one of the few countries, which maintained the fermentative acetone-butanol-ethanol (ABE) production for several decades. Until the end of the last century, the ABE fermentation from grain was operated in a few industrial scale plants. Due to the strong competition from the petrochemical industries, the fermentative ABE production lost its position in the 1990s, when all the solvent fermentation plants in China were closed. Under the current circumstances of concern about energy limitations and environmental pollution, new opportunities have emerged for the traditional ABE fermentation industry since it could again be potentially competitive with chemical synthesis. From 2006, several ABE fermentation plants in China have resumed production. The total solvent (acetone, butanol, and ethanol) production capacity from ten plants reached 210,000 tons, and the total solvent production is expected to be extended to 1,000,000 tons (based on the available data as of Sept. 2008). This article reviews current work in strain development, the continuous fermentation process, solvent recovery, and economic evaluation of ABE process in China. Challenges for an economically competitive ABE process in the future are also discussed.

  20. Clostridium difficile: from obscurity to superbug.

    PubMed

    Brazier, J S

    2008-01-01

    According to the UK media and popular press, Clostridium difficile is now a fully fledged member of that notorious but ill-defined group of microorganisms portrayed to the general public as superbugs. Following the trail blazed by methicillin-resistant Staphylococcus aureus (MRSA), C. difficile has made the transition from being an obscure anaerobic bacterium, mainly of interest to specialist anaerobic microbiologists, to that of an infamous superbug responsible for outbreaks of hospital-acquired infection that commonly result in serious disease and death. This review tracks the rise in scientific knowledge and public awareness of this organism.

  1. Identification, distribution, and toxigenicity of obligate anaerobes in polluted waters.

    PubMed Central

    Daily, O P; Joseph, S W; Gillmore, J D; Colwell, R R; Seidler, R J

    1981-01-01

    A seasonal occurrence of obligately anaerobic bacteria, predominantly of the genera Bacteroides and Clostridium, in a polluted water site has been observed. The number of anaerobes varied from 1.8 X 10(3) cells/ml in the warmer months to 10 cells/ml in winter. Several isolates were toxigenic, indicating a potential human health hazard. PMID:7235706

  2. Variability in DPA and Calcium Content in the Spores of Clostridium Species

    PubMed Central

    Jamroskovic, Jan; Chromikova, Zuzana; List, Cornelia; Bartova, Barbora; Barak, Imrich; Bernier-Latmani, Rizlan

    2016-01-01

    Spores of a number of clostridial species, and their resistance to thermal treatment is a major concern for the food industry. Spore resistance to wet heat is related to the level of spore hydration, which is inversely correlated with the content of calcium and dipicolinic acid (DPA) in the spore core. It is widely believed that the accumulation of DPA and calcium in the spore core is a fundamental component of the sporulation process for all endospore forming species. We have noticed heterogeneity in the heat resistance capacity and overall DPA/calcium content among the spores of several species belonging to Clostridium sensu stricto group: two C. acetobutylicum strains (DSM 792 and 1731), two C. beijerinckii strains (DSM 791 and NCIMB 8052), and a C. collagenovorans strain (DSM 3089). A C. beijerinckii strain (DSM 791) and a C. acetobutylicum strain (DSM 792) display low Ca and DPA levels. In addition, these two species, with the lowest average Ca/DPA content amongst the strains considered, also exhibit minimal heat resistance. There appears to be no correlation between the Ca/DPA content and the phylogenetic distribution of the C. acetobutylicum and C. beijerinckii species based either on the 16S rRNA or the spoVA gene. This finding suggests that a subset of Clostridium sensu stricto species produce spores with low resistance to wet heat. Additionally, analysis of individual spores using STEM-EDS and STXM revealed that DPA and calcium levels can also vary amongst individual spores in a single spore population. PMID:27891119

  3. Characterization of an acetoin reductase/2,3-butanediol dehydrogenase from Clostridium ljungdahlii DSM 13528.

    PubMed

    Tan, Yang; Liu, Zi-Yong; Liu, Zhen; Li, Fu-Li

    2015-11-01

    Acetoin reductase catalyzes the formation of 2,3-butanediol from acetoin. In Clostridium ljungdahlii DSM 13528, the gene CLJU_c23220 encoding the putative Zn(2+)-dependent alcohol dehydrogenase was cloned and expressed in Escherichia coli. The recombinant enzyme, CLAR, can catalyze the conversion of acetoin to 2,3-butanediol with NADPH as the cofactor. Furthermore, the gene CLJU_c23220 was introduced into Clostridium acetobutylicum ATCC 824 and the transformant was conferred the capacity of 2,3-butanediol production. In batch fermentation the transformant produced up to 3.1g/L of 2,3-butanediol, as well as acetone, butanol and ethanol (ABE, 17.8 g/L) in amounts similar to those produced by the wild type strain. This study provides conclusive evidence at the protein level that CLJU_c23220 is the key gene responsible for the conversion of acetoin to 2,3-butanediol in C. ljungdahlii DSM 13528. Moreover, the C. acetobutylicum ATCC 824 was modified via one-step metabolic engineering to produce 2,3-butanediol without influencing the ABE production.

  4. Characterization of Clostridium sp. RKD producing botulinum-like neurotoxin.

    PubMed

    Dixit, Aparna; Dhaked, Ram Kumar; Alam, Syed Imteyaz; Singh, Lokendra

    2005-07-01

    A Gram positive, motile, rod-shaped, strictly anaerobic bacterium isolated from intestine of decaying fish was identified as Clostridium sp. RKD and produced a botulinum type B-like neurotoxin as suggested by mouse bioassay and protection with anti botulinum antibodies. The neurotoxicity was functionally characterized by the phrenic nerve hemi-diaphragm assay. Phylogenetic analysis based on 16S rDNA sequence, placed it at a different position from the reported strains of Clostridium botulinum. The strain exhibited differences from both Clostridium botulinum and Clostridium tetani with respect to morphological, biochemical and chemotaxonomic characteristics. Botulinum group specific and serotype specific primers amplified the DNA fragments of 260 and 727 bp, respectively, indicating presence of botulinum type 'B' toxin gene. Sequence of nearly 700 bp amplified using primers specific for botulinum neurotoxin type B gene, did not show any significant match in the database when subjected to BLAST search.

  5. Anaerobic bacteria

    MedlinePlus

    Anaerobic bacteria are bacteria that do not live or grow when oxygen is present. In humans, these bacteria ... Goldstein EJ. Diseases caused by non-spore forming anaerobic bacteria. In: Goldman L, Schafer AI, eds. Goldman's Cecil ...

  6. Biofilms of Clostridium species.

    PubMed

    Pantaléon, Véronique; Bouttier, Sylvie; Soavelomandroso, Anna Philibertine; Janoir, Claire; Candela, Thomas

    2014-12-01

    The biofilm is a microbial community embedded in a synthesized matrix and is the main bacterial way of life. A biofilm adheres on surfaces or is found on interfaces. It protects bacteria from the environment, toxic molecules and may have a role in virulence. Clostridium species are spread throughout both environments and hosts, but their biofilms have not been extensively described in comparison with other bacterial species. In this review we describe all biofilms formed by Clostridium species during both industrial processes and in mammals where biofilms may be formed either during infections or associated to microbiota in the gut. We have specifically focussed on Clostridium difficile and Clostridium perfringens biofilms, which have been studied in vitro. Regulatory processes including sporulation and germination highlight how these Clostridium species live in biofilms. Furthermore, biofilms may have a role in the survival and spreading of Clostridium species.

  7. Autism and Clostridium tetani.

    PubMed

    Bolte, E R

    1998-08-01

    Autism is a severe developmental disability believed to have multiple etiologies. This paper outlines the possibility of a subacute, chronic tetanus infection of the intestinal tract as the underlying cause for symptoms of autism observed in some individuals. A significant percentage of individuals with autism have a history of extensive antibiotic use. Oral antibiotics significantly disrupt protective intestinal microbiota, creating a favorable environment for colonization by opportunistic pathogens. Clostridium tetani is an ubiquitous anaerobic bacillus that produces a potent neurotoxin. Intestinal colonization by C. tetani, and subsequent neurotoxin release, have been demonstrated in laboratory animals which were fed vegetative cells. The vagus nerve is capable of transporting tetanus neurotoxin (TeNT) and provides a route of ascent from the intestinal tract to the CNS. This route bypasses TeNT's normal preferential binding sites in the spinal cord, and therefore the symptoms of a typical tetanus infection are not evident. Once in the brain, TeNT disrupts the release of neurotransmitters by the proteolytic cleavage of synaptobrevin, a synaptic vesicle membrane protein. This inhibition of neurotransmitter release would explain a wide variety of behavioral deficits apparent in autism. Lab animals injected in the brain with TeNT have exhibited many of these behaviors. Some children with autism have also shown a significant reduction in stereotyped behaviors when treated with antimicrobials effective against intestinal clostridia. When viewed as sequelae to a subacute, chronic tetanus infection, many of the puzzling abnormalities of autism have a logical basis. A review of atypical tetanus cases, and strategies to test the validity of this paper's hypothesis, are included.

  8. Clostridium phytofermentans sp. nov., a cellulolytic mesophile from forest soil.

    PubMed

    Warnick, Thomas A; Methé, Barbara A; Leschine, Susan B

    2002-07-01

    An obligately anaerobic, mesophilic, cellulolytic bacterium, strain ISDgT, was isolated from forest soil. Cells of this isolate stained Gram-negative, despite possessing a Gram-positive cell-wall ultrastructure, and were motile, straight rods that formed spherical terminal spores that swelled the sporangium. Cellulose, pectin, polygalacturonic acid, starch, xylan, arabinose, cellobiose, fructose, galactose, gentiobiose, glucose, lactose, maltose, mannose, ribose and xylose supported growth. The major end products of fermentation were ethanol, acetate, CO2 and H2; formate and lactate were minor products. The optimum temperature for growth was 35-37 degrees C. Phylogenetic analyses based on 16S rRNA sequence comparisons showed that strain ISDgT was related to a group of anaerobes that included Clostridium herbivorans, Clostridium polysaccharolyticum and Clostridium populeti. The G+C content of this strain was 35.9 mol%. On the basis of numerous genotypic and phenotypic differences between strain ISDgT and its close relatives, strain ISDgT is proposed as a novel species in the genus Clostridium, for which the name Clostridium phytofermentans sp. nov. is proposed. The type strain is ISDgT (= ATCC 700394T).

  9. Purification of Clostridium toxoids.

    PubMed

    Buchowicz, I; Hay, M; Schiller, B; Korbecki, M; Sochańska, R

    1977-01-01

    A two-step fractionation procedure was applied for purification and concentration of the individual Clostridium toxoids. The toxoids were precipitated with hydrochloric acid in the presence of sodium sextametaphosphate, then antigenic fractions were separated from inactive contaminants by Sephadex G-75 filtration. Specific activity of the preparations thus obtained, as determined by Mancini radial immunodiffusion, was 150--565 binding units per mg of protein nitrogen for Clostridium perfringens toxoid, 204--352 binding units for Clostridium oedematiens toxoid and 26.6 -- 51.2 binding units for Clostridium septicum toxoid.

  10. Description of Anaerobacterium chartisolvens gen. nov., sp. nov., an obligately anaerobic bacterium from Clostridium rRNA cluster III isolated from soil of a Japanese rice field, and reclassification of Bacteroides cellulosolvens Murray et al. 1984 as Pseudobacteroides cellulosolvens gen. nov., comb. nov.

    PubMed

    Horino, Haruka; Fujita, Takashi; Tonouchi, Akio

    2014-04-01

    An obligately anaerobic bacterial strain designated T-1-35(T) was isolated as a dominant cultivable cellulose-degrading bacterium from soil of a Japanese rice field as an anaerobic filter-paper degrader. Cells of strain T-1-35(T) stained Gram-positive and were non-spore-forming rods with rounded ends, 0.8-1.0×3.5-15.0 µm, and motile by means of two to four polar flagella. Cells of strain T-1-35(T) exhibited pleomorphism: in aged cultures (over 90 days of incubation), almost all cells were irregularly shaped. Although no spore formation was observed, cells tolerated high temperatures, up to 90 °C for 10 min. The temperature range for growth was 15-40 °C, with an optimum at 35 °C. The pH range for growth was 5.5-9.0, with an optimum at pH 8.0-8.5 (slightly alkaliphilic). Strain T-1-35(T) fermented some carbohydrates to produce ethanol and lactate as the major products. Major cellular fatty acids were iso-C16 : 0 and iso-C13 : 0 3-OH. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain T-1-35(T) belonged to Clostridium rRNA cluster III. The closest relative of strain T-1-35(T) was Bacteroides cellulosolvens WM2(T), with 16S rRNA gene sequence similarity of 93.4 %. Phenotypic, physiological and molecular genetic methods demonstrated that strain T-1-35(T) was distinct from its phylogenetic relatives (members of Clostridium rRNA cluster III) because it predominantly produced ethanol, iso-C13 : 0 3-OH was a major cellular fatty acid and it always exhibited pleomorphism. On the basis of the results of a polyphasic taxonomic study, strain T-1-35(T) is considered to represent a novel genus and species, Anaerobacterium chartisolvens gen. nov., sp. nov. The type strain of Anaerobacterium chartisolvens is T-1-35(T) ( = DSM 27016(T) = NBRC 109520(T)). In addition, from the results of our phylogenetic analysis and its phenotypic features, the species Bacteroides cellulosolvens Murray et al. 1984 is proposed to be reclassified

  11. Metabolic engineering of Clostridium tyrobutyricum for n-butanol production from maltose and soluble starch by overexpressing α-glucosidase.

    PubMed

    Yu, Le; Xu, Mengmeng; Tang, I-Ching; Yang, Shang-Tian

    2015-07-01

    Clostridium tyrobutyricum does not have the enzymes needed for using maltose or starch. Two extracellular α-glucosidases encoded by agluI and agluII from Clostridium acetobutylicum ATCC 824 catalyzing the hydrolysis of α-1,4-glycosidic bonds in maltose and starch from the non-reducing end were cloned and expressed in C. tyrobutyricum (Δack, adhE2), and their effects on n-butanol production from maltose and soluble starch in batch fermentations were studied. Compared to the parental strain grown on glucose, mutants expressing agluI showed robust activity in breaking down maltose and produced more butanol (17.2 vs. 9.5 g/L) with a higher butanol yield (0.20 vs. 0.10 g/g) and productivity (0.29 vs. 0.16 g/L h). The mutant was also able to use soluble starch as substrate, although at a slower rate compared to maltose. Compared to C. acetobutylicum ATCC 824, the mutant produced more butanol from maltose (17.2 vs. 11.2 g/L) and soluble starch (16.2 vs. 8.8 g/L) in batch fermentations. The mutant was stable in batch fermentation without adding antibiotics, achieving a high butanol productivity of 0.40 g/L h. This mutant strain thus can be used in industrial production of n-butanol from maltose and soluble starch.

  12. Regulation of Toxin Production in Clostridium perfringens

    PubMed Central

    Ohtani, Kaori; Shimizu, Tohru

    2016-01-01

    The Gram-positive anaerobic bacterium Clostridium perfringens is widely distributed in nature, especially in soil and the gastrointestinal tracts of humans and animals. C. perfringens causes gas gangrene and food poisoning, and it produces extracellular enzymes and toxins that are thought to act synergistically and contribute to its pathogenesis. A complicated regulatory network of toxin genes has been reported that includes a two-component system for regulatory RNA and cell-cell communication. It is necessary to clarify the global regulatory system of these genes in order to understand and treat the virulence of C. perfringens. We summarize the existing knowledge about the regulatory mechanisms here. PMID:27399773

  13. Isolation and Identification of Psychrophilic Species of Clostridium from Milk

    PubMed Central

    Bhadsavle, C. H.; Shehata, T. E.; Collins, E. B.

    1972-01-01

    Four of 48 raw milk samples contained catalase-negative, gram-positive, motile, sporeforming, rod-shaped bacteria that grew optimally at 22 to 30 C and slowly at low temperatures. Isolates from two samples had a minimal growth temperature of 4 C, were anaerobic, and had characteristics similar to Clostridium hastiforme; those from the other two samples had a minimal growth temperature of 0 ± 1 C, were anaerobic, aerotolerant, and had characteristics similar to C. carnis. Specific growth rates, doubling times, ability to grow in pasteurized milk stored in commercial cartons, and resistance of spores to heating were determined for one strain of C. hastiforme. PMID:4565634

  14. Felled oil palm trunk as a renewable source for biobutanol production by Clostridium spp.

    PubMed

    Komonkiat, Itsara; Cheirsilp, Benjamas

    2013-10-01

    This study aimed to convert felled oil palm trunk to biobutanol by Clostridium spp. For efficient utilization of oil palm trunk, it was separated into sap and trunk fiber. The sap was used directly while the trunk fiber was hydrolyzed to fermentable sugars before use. Among five clostridia strains screened, Clostridium acetobutylicum DSM 1731 was the most suitable strain for butanol production from the sap without any supplementation of nutrients. It produced the highest amount of butanol (14.4 g/L) from the sap (sugar concentration of 50 g/L) with butanol yield of 0.35 g/g. When hydrolysate from the trunk fiber was used as an alternative carbon source (sugar concentration of 30 g/L), of the strains tested Clostridium beijerinckii TISTR 1461 produced the highest amount of butanol (10.0 g/L) with butanol yield of 0.41 g/g. The results presented herein suggest that oil palm trunk is a promising renewable substrate for biobutanol production.

  15. Characterization of a butanol-acetone-producing Clostridium strain and identification of its solventogenic genes.

    PubMed

    Chua, Teck Khiang; Liang, Da-Wei; Qi, Chao; Yang, Kun-Lin; He, Jianzhong

    2013-05-01

    A unique Clostridium species strain G117 was obtained in this study to be capable of producing dominant butanol from glucose. Butanol of 13.50 g/L was produced when culture G117 was fed with 60 g/L glucose, which is ~20% higher than previously reported butanol production by wild-type Clostridium acetobutylicum ATCC 824 under similar conditions. Strain G117 also distinguishes itself by generating negligible amount of ethanol, but producing butanol and acetone as biosolvent end-products. A butanol dehydrogenase gene (bdh gene) was identified in strain G117, which demonstrated a ~200-fold increase in transcription level measured by quantitative real-time PCR after 10h of culture growth. The high transcription suggests that this bdh gene could be a putative gene involved in butanol production. In all, Clostridium sp. strain G117 serves as a potential candidate for industrial biobutanol production while the absence of ethanol ensures an economic-efficient separation and purification of butanol.

  16. Regulation of toxin and bacteriocin gene expression in Clostridium by interchangeable RNA polymerase sigma factors.

    PubMed

    Dupuy, Bruno; Raffestin, Stéphanie; Matamouros, Susana; Mani, Nagraj; Popoff, Michel R; Sonenshein, Abraham L

    2006-05-01

    The production of major extracellular toxins by pathogenic strains of Clostridium botulinum, Clostridium tetani and Clostridium difficile, and a bacteriocin by Clostridium perfringens is dependent on a related group of RNA polymerase sigma-factors. These sigma-factors (BotR, TetR, TcdR and UviA) were shown to be sufficiently similar that they could substitute for one another in in vitro DNA binding and run-off transcription experiments. In cells, however, the sigma-factors fell into two subclasses. BotR and TetR were able to direct transcription of their target genes in a fully reciprocal manner. Similarly, UviA and TcdR were fully interchangeable. Neither BotR nor TetR could substitute for UviA or TcdR, however, and neither UviA nor TcdR could direct transcription of the natural targets of BotR or TetR. The extent of functional interchangeability of the sigma-factors was attributed to the strong conservation of their subregion 4.2 sequences and the conserved -35 sequences of their target promoters, while restrictions on interchangeability were attributed to variations in their subregion 2.4 sequences and the target site -10 sequences. The four sigma-factors have been assigned to group 5 of the sigma(70) family and seem to have arisen from a common ancestral protein that may have co-evolved with the genes whose transcription they direct. A fifth Clostridiumsigma-factor, sigma(Y) of Clostridium acetobutylicum, resembles the TcdR family, but was not functionally interchangeable with members of this family.

  17. Collagenase Clostridium Histolyticum Injection

    MedlinePlus

    ... disease (a thickening of tissue [plaque] inside the penis that causes the penis to curve). Collagenase Clostridium histolyticum injection is in ... the plaque of thickened tissue and allows the penis to be straightened.

  18. Clostridium Difficile Infections

    MedlinePlus

    Clostridium difficile (C. difficile) is a bacterium that causes diarrhea and more serious intestinal conditions such as colitis. Symptoms include Watery ... Nausea Abdominal pain or tenderness You might get C. difficile disease if you have an illness that ...

  19. Formation of involatile methylantimony species by Clostridium spp.

    PubMed

    Smith, L M; Craig, P J; Jenkins, R O

    2002-04-01

    Trimethylantimony was detected by gas chromatography-mass spectrometry (GC-MS) in the headspace of a soil enrichment culture designed to promote growth of clostridia. Clostridial isolates from the soil enrichment culture were shown to biomethylate inorganic antimony in monseptic culture, using hydride generation-gas chromatographyatomic absorption spectrometry (HG-GC-AAS). GC-MS profiles of headspace gases from soil enrichment cultures shown to generate trimethylantimony, were used to select characterised Clostridium spp for assessment of antimony biomethylation capability. Involatile methylantimony species (up to 21 microg Sb dm(-3)) were detected by HG-GC-AAS in the medium of monoseptic cultures of C. acetobutylicum, C. butyricum and C. cochlearium. The relative quantities of involatile mono-, di- and trimethylantimony species produced over the course of a 28-day cultivation period is consistent with trimethylantimony oxide being a final product of antimony biomethylation by these bacteria, with mono- and dimethylantimony species appearing transiently in the cultures as intermediates of an antimony biomethylation pathway. Clostridia may be the principal agents of antimony biomethylation in methanogenic environments and could give rise to methylated forms of antimony in both the aqueous and gaseous phases.

  20. Draft Genome Sequences of Clostridium Strains Native to Colombia with the Potential To Produce Solvents

    PubMed Central

    Rosas-Morales, Juan Pablo; Perez-Mancilla, Ximena; López-Kleine, Liliana

    2015-01-01

    Genomes from four Clostridium sp. strains considered to be mesophilic anaerobic bacteria, isolated from crop soil in Colombia, with a strong potential to produce alcohols like 1,3-propanediol, were analyzed. We present the draft genome of these strains, which will be useful for developing genetic engineering strategies. PMID:25999575

  1. First Insights into the Draft Genome of Clostridium colicanis DSM 13634, Isolated from Canine Feces

    PubMed Central

    Poehlein, Anja; Schilling, Tobias; Bhaskar Sathya Narayanan, Udhaya

    2016-01-01

    Clostridium colicanis DSM 13634 is a strictly anaerobic, rod-shaped, and spore-forming bacterium. It produces acids from common sugars such as glucose and fructose. The draft genome consists of one chromosome (2.6 Mbp) and contains 2,159 predicted protein-encoding genes. PMID:27198021

  2. Draft Genome Sequence of the Cellulolytic and Xylanolytic Thermophile Clostridium clariflavum Strain 4-2a

    PubMed Central

    Rooney, Elise A.; Rowe, Kenneth T.; Guseva, Anna; Huntemann, Marcel; Han, James K.; Chen, Amy; Kyrpides, Nikos C.; Mavromatis, Konstantinos; Markowitz, Victor M.; Palaniappan, Krishna; Ivanova, Natalia; Pati, Amrita; Liolios, Konstantinos; Nordberg, Henrik P.; Cantor, Michael N.; Hua, Susan X.; Shapiro, Nicole; Woyke, Tanja; Lynd, Lee R.

    2015-01-01

    Clostridium clariflavum strain 4-2a, a novel strain isolated from a thermophilic biocompost pile, has demonstrated an extensive capability to utilize both cellulose and hemicellulose under thermophilic anaerobic conditions. Here, we report the draft genome of this strain. PMID:26205857

  3. Draft genome sequences of clostridium strains native to Colombia with the potential to produce solvents.

    PubMed

    Rosas-Morales, Juan Pablo; Perez-Mancilla, Ximena; López-Kleine, Liliana; Montoya Castaño, Dolly; Riaño-Pachón, Diego Mauricio

    2015-05-21

    Genomes from four Clostridium sp. strains considered to be mesophilic anaerobic bacteria, isolated from crop soil in Colombia, with a strong potential to produce alcohols like 1,3-propanediol, were analyzed. We present the draft genome of these strains, which will be useful for developing genetic engineering strategies.

  4. Near complete genome sequence of Clostridium paradoxum strain JW-YL-7

    SciTech Connect

    Lancaster, Andrew; Utturkar, Sagar M.; Poole, Farris; Klingeman, Dawn Marie; Elias, Dwayne A.; Adams, Michael W. W.; Brown, Steven D.

    2016-05-05

    Clostridium paradoxum strain JW-YL-7 is a moderately thermophilic anaerobic alkaliphile isolated from the municipal sewage treatment plant in Athens, GA. We report the near-complete genome sequence of C. paradoxum strain JW-YL-7 obtained by using PacBio DNA sequencing and Pilon for sequence assembly refinement with Illumina data.

  5. Complete Genome Sequence of the Cellulolytic Thermophile Clostridium thermocellum DSM1313

    SciTech Connect

    Feinberg, Lawrence F; Foden, Justine; Barrett, Trisha; Davenport, Karen W.; Bruce, David; Detter, J. Chris; Tapia, Roxanne; Han, Cliff; Lapidus, Alla L.; Lucas, Susan; Cheng, Jan-Fang; Pitluck, Sam; Woyke, Tanja; Ivanova, N; Mikhailova, Natalia; Land, Miriam L; Hauser, Loren John; Argyros, Aaron; Goodwin, Lynne A.; Hogsett, David; Caiazza, Nicky

    2011-01-01

    Clostridium thermocellum DSM1313 is a thermophilic, anaerobic bacterium with some of the highest rates of cellulose hydrolysis reported. The complete genome sequence reveals a suite of carbohydrate-active enzymes and demonstrates a level of diversity at the species level distinguishing it from the type strain ATCC27405.

  6. Molecular Characterization of Podoviridae Bacteriophages Virulent for Clostridium perfringens and Comparison of Their Predicted Lytic Proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium that plays a significant role in human food-borne disease as well as non-food-borne human, animal and poultry diseases. There has been a resurgent interest in the use of bacteriophages or their gene products to control ba...

  7. Clostridium chauvoei in hens.

    PubMed

    Prukner-Radovcic, E; Milakovic-Novak, L; Ivesa-Petricevic, S; Grgic, N

    1995-03-01

    The bacterium Clostridium chauvoei causes disease in certain animals, most frequently in cattle and sheep. It occurs rarely in pigs, while equines and poultry appear to be resistant to infection. Two cases are presented in which C. chauvoei was isolated from disease of complex aetiology in hens. In Case I, 15-week-old light hybrid chickens were affected with chronic respiratory disease, coccidiosis, ascariasis and inflammation of the skin on the head, with necrosis of the comb. Growth was uneven and mortality reached 24%. Clostridium chauvoei was isolated from two of three combs examined. In Case II a flock of broiler breeders aged 11 weeks developed coccidiosis and, owing to disease or death, 60% were excluded from production. Clostridium chauvoei was isolated from all of 10 livers examined. These results demonstrate that C. chauvoei can infect chickens and that its possible role as a pathogen under certain circumstances should be further investigated.

  8. Genetic and biochemical analysis of solvent formation in Clostridium acetobutylicum. Progress report, September 1, 1992--July 31, 1996

    SciTech Connect

    Bennett, G.N.; Rudolph, F.B.

    1997-01-01

    Several degenerate strains were isolated and characterized by sporulation, motility and growth properties. Cell appearance and colony morphology were also recorded. Enzymatic assays revealed reduced butyraldehyde dehydrogenase and Co-A transferase enzyme activities in the degenerates. DNA analysis revealed that in complete degenerate strains the genes of the solvent locus were absent. Gyrase inhibitors slightly reduced the growth rate and decreased acetone formation preferentially. In an effort to analyze the role of sporulation sigma factors in solvent gene expression, recombination experiments were conducted and led to strains with increased solvent production. Analysis of redox systems has resulted in the sequence analysis of a cluster encoding formyl transferase proteins and an oxidoreductase-like gene. The genes for the two subunits of an apparent electron transfer flavoprotein were sequenced and suggest this factor acts to carry electrons to the butyryl-CoA dehydrogenase. The genes encoding the Fo subunits of the membrane ATPase have been sequenced.

  9. Impact of sweet sorghum cuticular waxes (SSCW) on acetone-butanol-ethanol fermentation using Clostridium acetobutylicum ABE1201.

    PubMed

    Cai, Di; Chang, Zhen; Wang, Chengyu; Ren, Wenqiang; Wang, Zheng; Qin, Peiyong; Tan, Tianwei

    2013-12-01

    The effect of cuticular waxes of sweet sorghum stem on acetone-butanol-ethanol (ABE) fermentation process was investigated. About 22.9% of butanol and 25.4% of ABE were decreased with fermentation period extended when SSCW was added. The inhibition of SSCW militate against both acidogenesis and solventogenesis phase, which were inconsistent with the inhibition of lignocellulose hydrolysate. Further studies on the composition of SSCW were performed. Regulations of inhibition with different carbon chain length of main compositions of SSCW on ABE fermentation were also investigated.

  10. REDUCTION OF 2,4,6-TRINITROTOLUENE BY CLOSTRIDIUM ACETOBUTYLICUM THROUGH HYDROXYLAMINO-NITROTOLUENE INTERMEDIATES. (R825513C006)

    EPA Science Inventory

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  11. Analysis of Redox Responses During TNT Transformation by Clostridium acetobutylicum ATCC 824 and Mutants Exhibiting Altered Metabolism

    DTIC Science & Technology

    2012-01-01

    2006) Functional studies of [ FeFe ] hydrogenase maturation in an Escherichia coli biosynthetic system. J Bacteriol 188(6):2163–2172 Kutty R, Bennett...activation of [ FeFe ] hydrogenase: new insights into hydrogenase maturation. J Biol Inorg Chem 12(4):443–447 Mermelstein LD, Papoutsakis ET (1993) In

  12. An active principle of Nigella sativa L., thymoquinone, showing significant antimicrobial activity against anaerobic bacteria

    PubMed Central

    Randhawa, Mohammad Akram; Alenazy, Awwad Khalaf; Alrowaili, Majed Gorayan; Basha, Jamith

    2017-01-01

    Aim/Background: Thymoquinone (TQ) is the major active principle of Nigella sativa seed (black seed) and is known to control many fungi, bacteria, and some viruses. However, the activity of TQ against anaerobic bacteria is not well demonstrated. Anaerobic bacteria can cause severe infections, including diarrhea, aspiration pneumonia, and brain abscess, particularly in immunodeficient individuals. The present study aimed to investigate the in vitro antimicrobial activity of TQ against some anaerobic pathogens in comparison to metronidazole. Methods: Standard, ATCC, strains of four anaerobic bacteria (Clostridium difficile, Clostridium perfringens, Bacteroides fragilis, and Bacteroides thetaiotaomicron), were initially isolated on special Brucella agar base (with hemin and vitamin K). Then, minimum inhibitory concentrations (MICs) of TQ and metronidazole were determined against these anaerobes when grown in Brucella agar, using serial agar dilution method according to the recommended guidelines for anaerobic organisms instructed by the Clinical and Laboratory Standards Institute. Results: TQ showed a significant antimicrobial activity against anaerobic bacteria although much weaker than metronidazole. MICs of TQ and metronidazole against various anaerobic human pathogens tested were found to be between 10-160 mg/L and 0.19-6.25 mg/L, respectively. Conclusions: TQ controlled the anaerobic human pathogenic bacteria, which supports the use of N. sativa in the treatment of diarrhea in folk medicine. Further investigations are in need for determination of the synergistic effect of TQ in combination with metronidazole and the activity of derivatives of TQ against anaerobic infections. PMID:28163966

  13. Bacteriophages of Clostridium perfringens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The specific aims of the book chapter are to: (1) Briefly review the nomenclature of bacteriophages and how these agents are classified. (2) Discuss the problems associated with addition/removal of antibiotics in commercial animal feeds. (3) Provide a brief overview of Clostridium perfringens biolog...

  14. Clostridium tetani bacteraemia.

    PubMed

    Hallit, Rabih Riad; Afridi, Muhammad; Sison, Raymund; Salem, Elie; Boghossian, Jack; Slim, Jihad

    2013-01-01

    Tetanus is a neuromuscular disease in which Clostridium tetani exotoxin (tetanospasmin) produces muscle spasms, incapacitating its host. To our knowledge, C. tetani bacteraemia has never been reported in the literature. The ideal management of this entity remains unresolved given that there is no literature to guide the therapy.

  15. Microbiology and physiology of anaerobic fermentations of cellulose. Progress report, September 1982-March 1983

    SciTech Connect

    Peck, H.D. Jr.; Ljungdahl, L.G.

    1983-01-01

    Research progress for the period September 1982-March 1983 is reported. The cellulose enzyme system of the anaerobic thermophile Clostridium thermocellum is being studied. Mutants have been obtained from thermoanaerobacter ethanolicus which produce higher amounts of ethanol than does the wild type. Clostridium thermoautotrophicum has been studied with respect to the pathway of acetate synthesis from CO/sub 2/. Progress has been achieved on properties of the NADP-dependent formate dehydrogenase and the CO dehydrogenase of Clostridium thermoaceticum. The CO dehydrogenase of Acetobacterium woodii has been isolated. Research has continued on the bioenergetics of dissimilatory sulfate reduction in sulfate reducing and methanogenic bacteria.

  16. Toxin Plasmids of Clostridium perfringens

    PubMed Central

    Li, Jihong; Adams, Vicki; Bannam, Trudi L.; Miyamoto, Kazuaki; Garcia, Jorge P.; Uzal, Francisco A.; Rood, Julian I.

    2013-01-01

    SUMMARY In both humans and animals, Clostridium perfringens is an important cause of histotoxic infections and diseases originating in the intestines, such as enteritis and enterotoxemia. The virulence of this Gram-positive, anaerobic bacterium is heavily dependent upon its prolific toxin-producing ability. Many of the ∼16 toxins produced by C. perfringens are encoded by large plasmids that range in size from ∼45 kb to ∼140 kb. These plasmid-encoded toxins are often closely associated with mobile elements. A C. perfringens strain can carry up to three different toxin plasmids, with a single plasmid carrying up to three distinct toxin genes. Molecular Koch's postulate analyses have established the importance of several plasmid-encoded toxins when C. perfringens disease strains cause enteritis or enterotoxemias. Many toxin plasmids are closely related, suggesting a common evolutionary origin. In particular, most toxin plasmids and some antibiotic resistance plasmids of C. perfringens share an ∼35-kb region containing a Tn916-related conjugation locus named tcp (transfer of clostridial plasmids). This tcp locus can mediate highly efficient conjugative transfer of these toxin or resistance plasmids. For example, conjugative transfer of a toxin plasmid from an infecting strain to C. perfringens normal intestinal flora strains may help to amplify and prolong an infection. Therefore, the presence of toxin genes on conjugative plasmids, particularly in association with insertion sequences that may mobilize these toxin genes, likely provides C. perfringens with considerable virulence plasticity and adaptability when it causes diseases originating in the gastrointestinal tract. PMID:23699255

  17. Bacterial ecology of abattoir wastewater treated by an anaerobic digestor

    PubMed Central

    Jabari, Linda; Gannoun, Hana; Khelifi, Eltaief; Cayol, Jean-Luc; Godon, Jean-Jacques; Hamdi, Moktar; Fardeau, Marie-Laure

    2016-01-01

    Wastewater from an anaerobic treatment plant at a slaughterhouse was analysed to determine the bacterial biodiversity present. Molecular analysis of the anaerobic sludge obtained from the treatment plant showed significant diversity, as 27 different phyla were identified. Firmicutes, Proteobacteria, Bacteroidetes, Thermotogae, Euryarchaeota (methanogens), and msbl6 (candidate division) were the dominant phyla of the anaerobic treatment plant and represented 21.7%, 18.5%, 11.5%, 9.4%, 8.9%, and 8.8% of the total bacteria identified, respectively. The dominant bacteria isolated were Clostridium, Bacteroides, Desulfobulbus, Desulfomicrobium, Desulfovibrio and Desulfotomaculum. Our results revealed the presence of new species, genera and families of microorganisms. The most interesting strains were characterised. Three new bacteria involved in anaerobic digestion of abattoir wastewater were published. PMID:26887229

  18. Genome sequence of Clostridium sporogenes DSM 795(T), an amino acid-degrading, nontoxic surrogate of neurotoxin-producing Clostridium botulinum.

    PubMed

    Poehlein, Anja; Riegel, Karin; König, Sandra M; Leimbach, Andreas; Daniel, Rolf; Dürre, Peter

    2015-01-01

    Clostridium sporogenes DSM 795 is the type strain of the species Clostridium sporogenes, first described by Metchnikoff in 1908. It is a Gram-positive, rod-shaped, anaerobic bacterium isolated from human faeces and belongs to the proteolytic branch of clostridia. C. sporogenes attracts special interest because of its potential use in a bacterial therapy for certain cancer types. Genome sequencing and annotation revealed several gene clusters coding for proteins involved in anaerobic degradation of amino acids, such as glycine and betaine via Stickland reaction. Genome comparison showed that C. sporogenes is closely related to C. botulinum. The genome of C. sporogenes DSM 795 consists of a circular chromosome of 4.1 Mb with an overall GC content of 27.81 mol% harboring 3,744 protein-coding genes, and 80 RNAs.

  19. Anaerobic Digestion.

    PubMed

    Liebetrau, Jan; Sträuber, Heike; Kretzschmar, Jörg; Denysenko, Velina; Nelles, Michael

    2017-04-09

    The term anaerobic digestion usually refers to the microbial conversion of organic material to biogas, which mainly consists of methane and carbon dioxide. The technical application of the naturally-occurring process is used to provide a renewable energy carrier and - as the substrate is often waste material - to reduce the organic matter content of the substrate prior to disposal.Applications can be found in sewage sludge treatment, the treatment of industrial and municipal solid wastes and wastewaters (including landfill gas utilization), and the conversion of agricultural residues and energy crops.For biorefinery concepts, the anaerobic digestion (AD) process is, on the one hand, an option to treat organic residues from other production processes. Concomitant effects are the reduction of organic carbon within the treated substance, the conversion of nitrogen and sulfur components, and the production of an energy-rich gas - the biogas. On the other hand, the multistep conversion of complex organic material offers the possibility of interrupting the conversion chain and locking out intermediates for utilization as basic material within the chemical industry.

  20. The prospect for vaccines to prevent Clostridium difficile infection.

    PubMed

    Ghose, Chandrabali; Kelly, Ciarán P

    2015-03-01

    Clostridium difficile is a spore-forming anaerobic gram-positive organism that is the leading cause of antibiotic-associated nosocomial infectious diarrhea in the Western world. This article describes the evolving epidemiology of C difficile infection (CDI) in the twenty-first century, evaluates the importance of vaccines against the disease, and defines the roles of both innate and adaptive host immune responses in CDI. The effects of passive immunotherapy and active vaccination against CDI in both humans and animals are also discussed.

  1. [Characteristics of Clostridium tetani and laboratory diagnosis of tetanus].

    PubMed

    Smietańska, Karolina; Rokosz-Chudziak, Natalia; Rastawicki, Waldemar

    2013-01-01

    The causative agent of tetanus is the obligate anaerobic bacterium--Clostridium tetani. These bacteria form endospores that are able to survive long periods of exposure to air and other adverse environmental conditions. Infection generally occurs through wound contamination. We can distinguish several forms of tetanus: generalized, local and neonatal. Diagnosis of tetanus is based primarily on the patient's clinical symptoms (muscle cramps, painful back muscle spasms, generalized contractions of the arcuate curvature of the body) as well as on microbiological diagnosis. This article is a brief review of C. tetani and diagnosis of infections caused by these organisms in humans.

  2. Myonecrosis by Clostridium septicum in a dog, diagnosed by a new multiplex-PCR.

    PubMed

    Ribeiro, Márcio Garcia; Silva, Rodrigo Otávio Silveira; Pires, Prhiscylla Sadanã; Martinho, Anna Paula Vitirito; Lucas, Thays Mizuki; Teixeira, Ana Izabel Passarela; Paes, Antonio Carlos; Barros, Claudenice Batista; Lobato, Francisco Carlos Faria

    2012-10-01

    Clostridial myositis is an acute, generally fatal toxemia that is considered to be rare in pet animals. The present report describes an unusual canine clostridial myositis that was diagnosed by a new multiplex-PCR (mPCR) designed for simultaneous identification of Clostridium sordellii, Clostridium septicum, Clostridium perfringens type A, Clostridium chauvoei, and Clostridium novyi type A. A ten-month-old male Rottweiler dog, that had displayed lameness and swelling of the left limb for 12 h, was admitted to a veterinary hospital. The animal was weak, dyspneic and hyperthermic, and a clinical examination indicated the presence of gas and edema in the limb. Despite emergency treatment, the animal died in only a few minutes. Samples of muscular tissue from the necrotic area were aseptically collected and plated onto defibrinated sheep blood agar (5%) in anaerobic conditions. Colonies suggestive of Clostridium spp. were submitted to testing by multiplex-PCR. Impression smears of the tissues, visualized with Gram and also with panoptic stains, revealed long rod-shaped organisms, and specimens also tested positive using the fluorescent antibody technique (FAT). The FAT and mPCR tests enabled a diagnosis of C. septicum myonecrosis in the dog.

  3. Molecular Characterization of Podoviral Bacteriophages Virulent for Clostridium perfringens and Their Comparison with Members of the Picovirinae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Clostridium perfringens is a Gram positive, spore-forming anaerobic bacterium that plays a significant role in human food-borne disease as well as non-food-borne human, animal, and poultry diseases. There has been a resurgent interest in the use of bacteriophages or their gene products to control b...

  4. Anaerobic Thermophiles

    PubMed Central

    Canganella, Francesco; Wiegel, Juergen

    2014-01-01

    The term “extremophile” was introduced to describe any organism capable of living and growing under extreme conditions. With the further development of studies on microbial ecology and taxonomy, a variety of “extreme” environments have been found and an increasing number of extremophiles are being described. Extremophiles have also been investigated as far as regarding the search for life on other planets and even evaluating the hypothesis that life on Earth originally came from space. The first extreme environments to be largely investigated were those characterized by elevated temperatures. The naturally “hot environments” on Earth range from solar heated surface soils and water with temperatures up to 65 °C, subterranean sites such as oil reserves and terrestrial geothermal with temperatures ranging from slightly above ambient to above 100 °C, to submarine hydrothermal systems with temperatures exceeding 300 °C. There are also human-made environments with elevated temperatures such as compost piles, slag heaps, industrial processes and water heaters. Thermophilic anaerobic microorganisms have been known for a long time, but scientists have often resisted the belief that some organisms do not only survive at high temperatures, but actually thrive under those hot conditions. They are perhaps one of the most interesting varieties of extremophilic organisms. These microorganisms can thrive at temperatures over 50 °C and, based on their optimal temperature, anaerobic thermophiles can be subdivided into three main groups: thermophiles with an optimal temperature between 50 °C and 64 °C and a maximum at 70 °C, extreme thermophiles with an optimal temperature between 65 °C and 80 °C, and finally hyperthermophiles with an optimal temperature above 80 °C and a maximum above 90 °C. The finding of novel extremely thermophilic and hyperthermophilic anaerobic bacteria in recent years, and the fact that a large fraction of them belong to the Archaea has

  5. Prevalence of Clostridium species and behaviour of Clostridium botulinum in gnocchi, a REPFED of italian origin.

    PubMed

    Del Torre, M; Stecchini, M L; Braconnier, A; Peck, M W

    2004-11-01

    Sales and consumption of refrigerated processed foods of extended durability (REPFEDs) have increased many-fold in Europe over the last 10 years. The safety and quality of these convenient ready-to-eat foods relies on a combination of mild heat treatment and refrigerated storage, sometimes in combination with other hurdles such as mild preservative factors. The major hazard to the microbiological safety of these foods is Clostridium botulinum. This paper reports on the prevalence and behaviour of proteolytic C. botulinum and non-proteolytic C. botulinum in gnocchi, a potato-based REPFED of Italian origin. Attempts to isolate proteolytic C. botulinum and non-proteolytic C. botulinum from gnocchi and its ingredients were unsuccessful. Based on assessment of the adequacy of the methods used, it was estimated that for proteolytic C. botulinum there was < 25 spores/kg of gnocchi and < 70 spores/kg of ingredients. The total anaerobic microbial load of gnocchi and its ingredients was low, with an estimated 1 MPN/g in processed gnocchi. Most of the anaerobic flora was facultatively anaerobic. A few obligately anaerobic bacteria were isolated from gnocchi and its ingredients and belonged to different Clostridium species. The protection factor, number of decimal reductions in the probability of toxigenesis from a single spore, was determined for eight different gnocchi formulations by challenge test studies. For all gnocchi stored at 8 degrees C (as recommended by the manufacturer) or 12 degrees C (mild temperature abuse), growth and toxin production were not detected in 75 days. The protection factor was >4.2 for proteolytic C. botulinum, and >6.2 for non-proteolytic C. botulinum. When inoculated packs were stored at 20 degrees C (severe temperature abuse), toxin production in 75 days was prevented by the inclusion of 0.09% (w/w) sorbic acid (protection factors as above), however in the absence of sorbic acid the packs became toxic before the end of the intended shelf

  6. Clostridium difficile infection: molecular pathogenesis and novel therapeutics

    PubMed Central

    Rineh, Ardeshir; Kelso, Michael J; Vatansever, Fatma; Tegos, George P; Hamblin, Michael R

    2015-01-01

    The Gram-positive anaerobic bacterium Clostridium difficile produces toxins A and B, which can cause a spectrum of diseases from pseudomembranous colitis to C. difficile-associated diarrhea. A limited number of C. difficile strains also produce a binary toxin that exhibits ADP ribosyltransferase activity. Here, the structure and the mechanism of action of these toxins as well as their role in disease are reviewed. Nosocomial C. difficile infection is often contracted in hospital when patients treated with antibiotics suffer a disturbance in normal gut microflora. C. difficile spores can persist on dry, inanimate surface for months. Metronidazole and oral vancomycin are clinically used for treatment of C. difficile infection but clinical failure and concern about promotion of resistance are motivating the search for novel non-antibiotic therapeutics. Methods for controlling both toxins and spores, replacing gut microflora by probiotics or fecal transplant, and killing bacteria in the anaerobic gut by photodynamic therapy are discussed. PMID:24410618

  7. Cultivation of Anaerobic and Facultatively Anaerobic Bacteria from Spacecraft-Associated Clean Rooms▿

    PubMed Central

    Stieglmeier, Michaela; Wirth, Reinhard; Kminek, Gerhard; Moissl-Eichinger, Christine

    2009-01-01

    In the course of this biodiversity study, the cultivable microbial community of European spacecraft-associated clean rooms and the Herschel Space Observatory located therein were analyzed during routine assembly operations. Here, we focused on microorganisms capable of growing without oxygen. Anaerobes play a significant role in planetary protection considerations since extraterrestrial environments like Mars probably do not provide enough oxygen for fully aerobic microbial growth. A broad assortment of anaerobic media was used in our cultivation strategies, which focused on microorganisms with special metabolic skills. The majority of the isolated strains grew on anaerobic, complex, nutrient-rich media. Autotrophic microorganisms or microbes capable of fixing nitrogen were also cultivated. A broad range of facultatively anaerobic bacteria was detected during this study and also, for the first time, some strictly anaerobic bacteria (Clostridium and Propionibacterium) were isolated from spacecraft-associated clean rooms. The multiassay cultivation approach was the basis for the detection of several bacteria that had not been cultivated from these special environments before and also led to the discovery of two novel microbial species of Pseudomonas and Paenibacillus. PMID:19363082

  8. Peptoniphilus methioninivorax sp. nov., A Novel Gram-Positive Anaerobic Coccus Isolated from Retail Ground Beef

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Strain NRRL B-23883 was isolated from retail ground beef as part of a study on the genetic diversity of Clostridium perfringens. The strain was found to be a strictly anaerobic, gram-type positive coccus that was able to utilize peptone as a sole carbon source. Subsequent to sequencing the 16S rib...

  9. Clostridium difficile Infection

    PubMed Central

    Heinlen, Latisha; Ballard, Jimmy D.

    2010-01-01

    Clostridium difficile is the leading cause of hospital-acquired diarrhea in Europe and North America and is a serious re-emerging pathogen. Recent outbreaks have led to increasing morbidity and mortality and have been associated with a new strain (BI/NAP1/027) of C. difficile that produces more toxin than historical strains. With the increasing incidence of C. difficile infection, clinicians have also seen a change in the epidemiology with increased infections in previously low-risk populations. This chapter highlights the current knowledge on C. difficile virulence, human disease, epidemic outbreaks, and optimal treatment strategies. PMID:20697257

  10. Degradation of natural and synthetic polyesters under anaerobic conditions.

    PubMed

    Abou-Zeid, D M; Müller, R J; Deckwer, W D

    2001-03-30

    Often, degradability under anaerobic conditions is desirable for plastics claimed to be biodegradable, e.g. in anaerobic biowaste treatment plants, landfills and in natural anaerobic sediments. The biodegradation of the natural polyesters poly(beta-hydroxybutyrate) (PHB), poly(beta-hydroxybutyrate-co-11.6%-beta-hydroxyvalerate) (PHBV) and the synthetic polyester poly(epsilon-caprolactone) (PCL) was studied in two anaerobic sludges and individual polyester degrading anaerobic strains were isolated, characterized and used for degradation experiments under controlled laboratory conditions. Incubation of PHB and PHBV films in two anaerobic sludges exhibited significant degradation in a time scale of 6-10 weeks monitored by weight loss and biogas formation. In contrast to aerobic conditions, PHB was degraded anaerobically more rapidly than the copolyester PHBV, when tested with either mixed cultures or a single strained isolate. PCL tends to degrade slower than the natural polyesters PHB and PHBV. Four PHB and PCL degrading isolates were taxonomically identified and are obviously new species belonging to the genus Clostridium group I. The depolymerizing enzyme systems of PHB and PCL degrading isolates are supposed to be different. Using one isolated strain in an optimized laboratory degradation test with PHB powder, the degradation time was drastically reduced compared to the degradation in sludges (2 days vs. 6-10 weeks).

  11. Ultrastructure and extreme heat resistance of spores from thermophilic Clostridium species.

    PubMed Central

    Hyun, H H; Zeikus, J G; Longin, R; Millet, J; Ryter, A

    1983-01-01

    The heat resistance and ultrastructural features of spore suspensions prepared from Clostridium thermocellum LQRI, Clostridium thermosulfurogenes 4B, and Clostridium thermohydrosulfuricum 39E were compared as a function of decimal reduction time. The decimal reduction times at 121 degrees C for strains LQRI, 4B, and 39E were 0.5, 2.5, and 11 min. The higher degree of spore heat resistance was associated with a spore architecture displaying a thicker cortex layer. Heat resistance of these spores was proportional to the ratio of spore cortex volume to cytoplasmic volume. These ratios for spores of strains LQRI, 4B, and 39E were 1.4, 1.6, and 6.6, respectively. The extreme heat resistance and autoclavable nature of C. thermohydrosulfuricum spores under routine sterilization procedures is suggested as a common cause of laboratory contamination with pure cultures of thermophilic, saccharide-fermenting anaerobes. Images PMID:6643392

  12. [Toxins of Clostridium perfringens as a natural and bioterroristic threats].

    PubMed

    Omernik, Andrzej; Płusa, Tadeusz

    2015-09-01

    Clostridium perfringens is absolutely anaerobic rod-shaped, sporeforming bacterium. The morbidity is connected with producing toxins. Depending on the type of toxin produced Clostridium perfringens can be divided into five serotypes:A-E. Under natural conditions, this bacterium is responsible for local outbreaks of food poisoning associated with eating contaminated food which which was improperly heat treated. Some countries with lower economic level are endemic foci of necrotizing enteritis caused by Clostridium perfringens. The bacterium is also a major cause of gas gangrene. It is a disease, associated with wound infection, with potentially fatal prognosis in the case of treatment's delays. In the absence of early radical surgery, antibiotic therapy and (if available) hyperbaric treatment leads to the spread of toxins in the body causing shock, coma and death. Due to the force of produced toxins is a pathogen that poses a substrate for the production of biological weapons. It could potentially be used to induce outbreaks of food poisoning and by missiles contamination by spore lead to increased morbidity of gas gangrene in injured soldiers. C. perfringens types B and D produce epsilon toxin considered to be the third most powerful bacterial toxin. Because of the ability to disperse the toxin as an aerosol and a lack of methods of treatment and prevention of poisoning possible factors it is a potential tool for bioterrorism It is advisable to continue research into vaccines and treatments for poisoning toxins of C. perfringens.

  13. Rapid Confirmation of Clostridium perfringens by Using Chromogenic and Fluorogenic Substrates

    PubMed Central

    Adcock, Philip W.; Saint, Christopher P.

    2001-01-01

    The use of 4-methylumbelliferyl phosphate (MUP) and ortho-nitrophenyl-β-d-galactopyranoside (ONPG) for the identification of Clostridium perfringens was investigated. A liquid assay containing both MUP and ONPG was a highly specific alternative method for C. perfringens confirmation, reducing incubation time from 48 to only 4 h. The assay solution is easy to prepare, does not require anaerobic conditions for use, and has an extended shelf life. PMID:11526053

  14. [Efficacy of tinidazole against anaerobes in comparison with metronidazole, ornidazole, cefoxitin and lamoxactam].

    PubMed

    Werner, H; Kuchenbecker, K; Hammann, R

    1983-06-01

    The in vitro inhibitory activity of tinidazole, metronidazole, ornidazole, cefoxitin and moxalactam was determined against 150 isolates of clinically important anaerobes including Bacteroides fragilis, Bacteroides bivius and Clostridium perfringens by means of agar dilution tests. The members of 18 gramnegative and 14 grampositive species were inhibited by tinidazole at less than or equal to 0,01-8 micrograms/ml thus being without exception susceptible to the drug. A similar in vitro activity was recorded for metronidazole and ornidazole. Though cefoxitin and moxalactam generally had a good in vitro activity against most anaerobic species, single strains of Bacteroides thetaiotaomicron, B. distasonis, Clostridium difficile and Eubacterium rectale were resistant to these drugs. With regard to its in vitro activity, tinidazole seems to be a promising substance for the therapy of anaerobic infections.

  15. Microbiology and physiology of anaerobic fermentations of cellulose

    SciTech Connect

    Wiegel, J.

    1991-05-01

    The biochemistry and physiology of four major groups of anaerobic bacteria involved in the conversion of cellulose to methane or chemical feedstocks are examined. Aspects of metabolism which are relevant to the interactions and bioenergetics of consortia are being studied. Properties of the cellulolytic enzyme cluster of Clostridium thermocellum are investigated. Five different hydrogenases have been characterized in detail from anaerobic bacteria. Genes for different hydrogenases are being cloned and sequenced to determine their structural relationships. The role of metal clusters in activation of H{sub 2} is being investigated, as is the structure and role of metal clusters in formate metabolism. The function of formate in the total synthesis of acetate from CO{sub 2} and the role of this primary in anaerobes will be examined as well. Finally, these enzyme studies will be performed on thermophilic bacteria and new, pertinent species will be isolated. 50 refs., 3 figs., 1 tab.

  16. My Lifelong Passion for Biochemistry and Anaerobic Microorganisms.

    PubMed

    Thauer, Rudolf Kurt

    2015-01-01

    Early parental influence led me first to medical school, but after developing a passion for biochemistry and sensing the need for a deeper foundation, I changed to chemistry. During breaks between semesters, I worked in various biochemistry labs to acquire a feeling for the different areas of investigation. The scientific puzzle that fascinated me most was the metabolism of the anaerobic bacterium Clostridium kluyveri, which I took on in 1965 in Karl Decker's lab in Freiburg, Germany. I quickly realized that little was known about the biochemistry of strict anaerobes such as clostridia, methanogens, acetogens, and sulfate-reducing bacteria and that these were ideal model organisms to study fundamental questions of energy conservation, CO2 fixation, and the evolution of metabolic pathways. My passion for anaerobes was born then and is unabated even after 50 years of study.

  17. Vaccines against Clostridium difficile.

    PubMed

    Leuzzi, Rosanna; Adamo, Roberto; Scarselli, Maria

    2014-01-01

    Clostridium difficile infection (CDI) is recognized as a major cause of nosocomial diseases ranging from antibiotic related diarrhea to fulminant colitis. Emergence during the last 2 decades of C. difficile strains associated with high incidence, severity and lethal outcomes has increased the challenges for CDI treatment. A limited number of drugs have proven to be effective against CDI and concerns about antibiotic resistance as well as recurring disease solicited the search for novel therapeutic strategies. Active vaccination provides the attractive opportunity to prevent CDI, and intense research in recent years led to development of experimental vaccines, 3 of which are currently under clinical evaluation. This review summarizes recent achievements and remaining challenges in the field of C. difficile vaccines, and discusses future perspectives in view of newly-identified candidate antigens.

  18. Livestock Anaerobic Digester Database

    EPA Pesticide Factsheets

    The Anaerobic Digester Database provides basic information about anaerobic digesters on livestock farms in the United States, organized in Excel spreadsheets. It includes projects that are under construction, operating, or shut down.

  19. Quantitative real-time PCR assay for Clostridium septicum in poultry gangrenous dermatitis associated samples.

    PubMed

    Neumann, A P; Dunham, S M; Rehberger, T G; Siragusa, G R

    2010-08-01

    Clostridium septicum is a spore-forming anaerobe frequently implicated in cases of gangrenous dermatitis (GD) and other spontaneously occurring myonecrotic infections of poultry. Although C. septicum is readily cultured from diseased tissues it can be difficult to enumerate due to its tendency to swarm over the surface of agar plates. In this study a quantitative real-time PCR assay was developed in order to more accurately measure the levels of C. septicum in healthy as well as GD associated poultry samples. The assay was specifically designed to target the C. septicum alpha toxin gene, csa, which is, to our knowledge, carried by all strains of C. septicum and has been shown to be essential for virulence. Genomic DNAs from a diverse collection of bacterial species, including closely related Clostridium chauvoei, Clostridium carnis, Clostridium tertium as well as several strains of Clostridium perfringens, all failed to produce a positive reaction. An approximate reproducible limit of detection in spiked extracts of at least 10(3) cfu/g of C. septicum was observed for a variety of different sample types. C. septicum levels in broiler chicken field samples estimated from the results of qPCR were statistically correlated to culture based enumerations obtained from those same tissues.

  20. Clostridium polynesiense sp. nov., a new member of the human gut microbiota in French Polynesia.

    PubMed

    Sankar, Senthil Alias; Rathored, Jaishriram; Metidji, Sarah; Lagier, Jean-Christophe; Khelaifia, Saber; Labas, Noemie; Musso, Didier; Raoult, Didier; Fournier, Pierre-Edouard

    2015-12-01

    Strain MS1, a Gram-positive, obligately anaerobic, motile and spore-forming rod belonging to the Clostridium genus, was isolated from the feces of a healthy Polynesian male living in French Polynesia. The temperature range for growth was 30-45 °C. We sequenced its complete genome and studied its phenotypic characteristics. The 3,560,738-bp long genome (one chromosome, no plasmid, G + C content 34%) contained 3535 protein-coding and 70 RNA genes. Strain MS1 exhibited a 98.24% 16S rRNA similarity with Clostridium amylolyticum, the phylogenetically closest species. When compared with other Clostridium species with standing in nomenclature, it had an average genomic similarity of 68.8-70%, a unique MALDI-TOF spectrum, and differed in nitrate reduction, motility and L-arabinose and D-lactose metabolism with most of the closest species. Therefore, strain MS1 is sufficiently distinct from type strains of the genus Clostridium to represent a novel species within this genus, for which the name Clostridium polynesiense sp. nov. is proposed. The type strain of C. polynesiense is MS1(T) (= CSUR P630 = DSM 27072).

  1. Physical Characterization of Clostridium Botulinum Neurotoxin Genes

    DTIC Science & Technology

    1992-02-17

    type A toxin of Clostridium difficile ; Von Eichel-Streiber, 1989) or total failure (eg., the bacteriocir, of the Clostridium perf•ingens plasmid... CLOSTRIDIUM BOTULINUM NEUROTOXIN GENES PRINCIPAL INVESTIGATIR: NIGEL P. MINTON PI ADDRESS: Public Health Laboratory Service Center for Applied...NUMBERS Physical Characterization of Clostridium botulinum DAMDl7-90-Z-0033 Neurotoxin Genes 61102A 6. AUTHOR(S) 3Ml61102BSI2 AA Nigel P. Minton DA335530

  2. Action of nitroheterocyclic drugs against Clostridium difficile

    PubMed Central

    Kumar, Manish; Adhikari, Sudip; Hurdle, Julian G.

    2014-01-01

    The nitroheterocyclic classes of drugs have a long history of use in treating anaerobic infections, as exemplified by metronidazole as a first-line treatment for mild-to-moderate Clostridium difficile infection (CDI). Since direct comparisons of the three major classes of nitroheterocyclic drugs (i.e. nitroimidazole, nitazoxanide and nitrofurans) and nitrosating agents against C. difficile are under-examined, in this study their actions against C. difficile were compared. Results show that whilst transient resistance occurs to metronidazole and nitazoxanide, stable resistance arises to nitrofurans upon serial passage. All compounds killed C. difficile at high concentrations in addition to the host defence nitrosating agent S-nitrosoglutathione (GSNO). This suggests that GSNO killing of C. difficile contributes to its efficacy in murine CDI. Although nitric oxide production could not be detected for the nitroheterocyclic drugs, the cellular response to metronidazole and nitrofurans has some overlap with the response to GSNO, causing significant upregulation of the hybrid-cluster protein Hcp that responds to nitrosative stress. These findings provide new insights into the action of nitroheterocyclic drugs against C. difficile. PMID:25129314

  3. Perfringolysin O: The Underrated Clostridium perfringens Toxin?

    PubMed Central

    Verherstraeten, Stefanie; Goossens, Evy; Valgaeren, Bonnie; Pardon, Bart; Timbermont, Leen; Haesebrouck, Freddy; Ducatelle, Richard; Deprez, Piet; Wade, Kristin R.; Tweten, Rodney; Van Immerseel, Filip

    2015-01-01

    The anaerobic bacterium Clostridium perfringens expresses multiple toxins that promote disease development in both humans and animals. One such toxin is perfringolysin O (PFO, classically referred to as θ toxin), a pore-forming cholesterol-dependent cytolysin (CDC). PFO is secreted as a water-soluble monomer that recognizes and binds membranes via cholesterol. Membrane-bound monomers undergo structural changes that culminate in the formation of an oligomerized prepore complex on the membrane surface. The prepore then undergoes conversion into the bilayer-spanning pore measuring approximately 250–300 Å in diameter. PFO is expressed in nearly all identified C. perfringens strains and harbors interesting traits that suggest a potential undefined role for PFO in disease development. Research has demonstrated a role for PFO in gas gangrene progression and bovine necrohemorrhagic enteritis, but there is limited data available to determine if PFO also functions in additional disease presentations caused by C. perfringens. This review summarizes the known structural and functional characteristics of PFO, while highlighting recent insights into the potential contributions of PFO to disease pathogenesis. PMID:26008232

  4. 2,4,6-Trinitrotoluene reduction by carbon monoxide dehydrogenase from Clostridium thermoaceticum

    SciTech Connect

    Huang, S.; Lindahl, P.A.; Wang, C.; Bennett, G.N.; Rudolph, F.B.; Hughes, J.B.

    2000-03-01

    Purified CO dehydrogenase (CODH) from Clostridium thermoaceticum catalyzed the transformation of 2,4,6-trinitrotoluene (TNT). The intermediates and reduced products of TNT transformation were separated and appear to be identical to the compounds formed by C. acetobutylicum, namely, 2-hydroxylamino-4,6-dinitrotoluene (2 HA46DNT), 4-hydroxylamino-2,6-dinitrotoluene (4HA26DNT), 2,4-dihydroxylamino-6-nitrotoluene (24DHANT), and the Bamberger rearrangement product of 2,4-dihydroxylamino-6-nitrotoluene. In the presence of saturating CO, CODH catalyzed the conversion of TNT to two monohydroxylamino derivatives (2HA46DNT and 4HA26DNT), with 4HA26DNT as the dominant isomer. These derivatives were then converted to 24DHANT, which slowly converted to the Bamberger rearrangement product. Apparent K{sub m} and k{sub cat} values of TNT reduction were 165 {+-} 43 {micro}M for TNT and 400 {+-} 94 s{sup {minus}1}, respectively. Cyanide, an inhibitor for the CO/CO{sub 2} oxidation/reduction activity of CODH, inhibited the TNT degradation activity of CODH.

  5. 2,4,6-trinitrotoluene reduction by carbon monoxide dehydrogenase from Clostridium thermoaceticum.

    PubMed

    Huang, S; Lindahl, P A; Wang, C; Bennett, G N; Rudolph, F B; Hughes, J B

    2000-04-01

    Purified CO dehydrogenase (CODH) from Clostridium thermoaceticum catalyzed the transformation of 2,4,6-trinitrotoluene (TNT). The intermediates and reduced products of TNT transformation were separated and appear to be identical to the compounds formed by C. acetobutylicum, namely, 2-hydroxylamino-4,6-dinitrotoluene (2HA46DNT), 4-hydroxylamino-2,6-dinitrotoluene (4HA26DNT), 2, 4-dihydroxylamino-6-nitrotoluene (24DHANT), and the Bamberger rearrangement product of 2,4-dihydroxylamino-6-nitrotoluene. In the presence of saturating CO, CODH catalyzed the conversion of TNT to two monohydroxylamino derivatives (2HA46DNT and 4HA26DNT), with 4HA26DNT as the dominant isomer. These derivatives were then converted to 24DHANT, which slowly converted to the Bamberger rearrangement product. Apparent K(m) and k(cat) values of TNT reduction were 165 +/- 43 microM for TNT and 400 +/- 94 s(-1), respectively. Cyanide, an inhibitor for the CO/CO(2) oxidation/reduction activity of CODH, inhibited the TNT degradation activity of CODH.

  6. Construction of heterologous gene expression cassettes for the development of recombinant Clostridium beijerinckii.

    PubMed

    Oh, Young Hoon; Eom, Gyeong Tae; Kang, Kyoung Hee; Joo, Jeong Chan; Jang, Young-Ah; Choi, Jae Woo; Song, Bong Keun; Lee, Seung Hwan; Park, Si Jae

    2016-04-01

    Gene-expression cassettes for the construction of recombinant Clostridium beijerinckii were developed as potential tools for metabolic engineering of C. beijerinckii. Gene expression cassettes containing ColE1 origin and pAMB origin along with the erythromycin resistance gene were constructed, in which promoters from Escherichia coli, Lactococcus lactis, Ralstonia eutropha, C. acetobutylicum, and C. beijerinckii are examined as potential promoters in C. beijerinckii. Zymogram analysis of the cell extracts and comparison of lipase activities of the recombinant C. beijerinckii strains expressing Pseudomonas fluorescens tliA gene suggested that the tliA gene was functionally expressed by all the examined promoters with different expression level. Also, recombinant C. beijerinckii expressing C. beijerinckii secondary alcohol dehydrogenase by the constructed expression cassettes successfully produced 2-propanol from glucose. The best promoter for TliA expression was the R. eutropha phaP promoter while that for 2-propanol production was the putative C. beijerinckii pta promoter. Gene expression cassettes developed in this study may be useful tools for the construction of recombinant C. beijerinckii strains as host strains for the valuable chemicals and fuels from renewable resources.

  7. Comparative genomic analysis of dha regulon and related genes for anaerobic glycerol metabolism in bacteria.

    PubMed

    Sun, Jibin; van den Heuvel, Joop; Soucaille, Philippe; Qu, Yinbo; Zeng, An-Ping

    2003-01-01

    The dihydroxyacetone (dha) regulon of bacteria encodes genes for the anaerobic metabolism of glycerol. In this work, genomic data are used to analyze and compare the dha regulon and related genes in different organisms in silico with respect to gene organization, sequence similarity, and possible functions. Database searches showed that among the organisms, the genomes of which have been sequenced so far, only two, i.e., Klebsiella pneumoniae MGH 78578 and Clostridium perfringens contain a complete dha regulon bearing all known enzymes. The components and their organization in the dha regulon of these two organisms differ considerably from each other and also from the previously partially sequenced dha regulons in Citrobacter freundii, Clostridium pasteurianum, and Clostridium butyricum. Unlike all of the other organisms, genes for the oxidative and reductive pathways of anaerobic glycerol metabolism in C. perfringens are located in two separate organization units on the chromosome. Comparisons of deduced protein sequences of genes with similar functions showed that the dha regulon components in K. pneumoniae and C. freundii have high similarities (80-95%) but lower similarities to those of the Clostridium species (30-80%). Interestingly, the protein sequence similarities among the dha genes of the Clostridium species are in many cases even lower than those between the Clostridium species and K. pneumoniae or C. freundii, suggesting two different types of dha regulon in the Clostridium species studied. The in silico reconstruction and comparison of dha regulons revealed several new genes in the microorganisms studied. In particular, a novel dha kinase that is phosphoenolpyruvate-dependent is identified and experimentally confirmed for K. pneumoniae in addition to the known ATP-dependent dha kinase. This finding gives new insights into the regulation of glycerol metabolism in K. pneumoniae and explains some hitherto not well understood experimental observations.

  8. Draft Genome Sequences for Clostridium thermocellum Wild-Type Strain YS and Derived Cellulose Adhesion-Defective Mutant Strain AD2

    SciTech Connect

    Brown, Steven D; Lamed, Raphael; Morag, Ely; Borovok, Ilya; Shoham, Yuval; Klingeman, Dawn Marie; Johnson, Courtney M; Yang, Zamin; Land, Miriam L; Utturkar, Sagar M; Keller, Martin; Bayer, Edward A

    2012-01-01

    Clostridium thermocellum wild-type strain YS is an anaerobic, thermophilic, cellulolytic bacterium capable of directly converting cellulosic substrates into ethanol. Strain YS and a derived cellulose adhesion-defective mutant strain AD2 played pivotal roles in describing the original cellulosome concept. We present their draft genome sequences.

  9. Characterization of Bacteriophages Virulent for Clostridium perfringens and Identification of Phage Lytic Enzymes as Alternatives to Antibiotics for Potential Control of the Bacterium.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium that plays a significant role in human food-borne disease as well as non-food-borne human, animal, and poultry diseases. There has been a resurgent interest in the use of bacteriophages or their gene products to control b...

  10. Characterization of Bacteriophages Virulent for Clostridium perfringens and Identification of Phage Lytic Enzymes as Alternatives to Antibiotics for Potential Control of the Bacterium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium that plays a significant role in human food-borne disease as well as non-food-borne human, animal, and poultry diseases. There has been a resurgent interest in the use of bacteriophages or their gene products to control b...

  11. Expression of a Clostridium perfringens genome-encoded putative N-acetylmuramoyl-L-alanine amidase as a potential antimicrobial to control the bacterium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium that plays a substantial role in non-foodborne human, animal and avian diseases as well as human foodborne disease. Previously discovered C. perfringens bacteriophage lytic enzyme amino acid sequences were utilized to iden...

  12. Draft Genome Sequence of Clostridium sp. Strain W14A Isolated from a Cellulose-Degrading Biofilm in a Landfill Leachate Microcosm

    PubMed Central

    2016-01-01

    Here, we report the draft genome of Clostridium sp. strain W14A, isolated from the anaerobic, cellulolytic biofilm of a cotton string sample incubated in a landfill leachate microcosm. The draft genome comprises 131 contigs, 3,823,510 bp, 51.5% G+C content, and 4,119 predicted coding domain sequences. PMID:27660778

  13. Characterization of bacteriophages virulent for Clostridium perfringens and identification of phage lytic enzymes as alternatives to antibiotics for potential control of the bacterium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There has been a resurgent interest in the use of bacteriophages or their gene products to control bacterial pathogens as alternatives to currently utilized antibiotics. Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium that plays a significant role in human food-borne di...

  14. Characterization of bacteriophages virulent for Clostridium perfringens and identification of phage lytic enzymes as alternatives to antibiotics for potential control of the bacterium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There has been a resurgent interest in the use of bacteriophages or their gene products to control bacterial pathogens as alternatives to currently utilized antibiotics. Clostridium perfringens is a Gram-positive, spore-forming anaerobic bacterium that plays a significant role in human food-borne d...

  15. Paraclostridium benzoelyticum gen. nov. sp. nov., isolated from marine sediment and reclassification of Clostridium bifermentans as Paraclostridium bifermentans comb. nov. Proposal of a new genus Paeniclostridium gen. nov. to accommodate Clostridium sordellii and Clostridium ghonii.

    PubMed

    T S, Sasi Jyothsna; L, Tushar; Ch, Sasikala; Ch V, Ramana

    2016-01-05

    Twenty three rod shaped, endospore forming, Gram-stain-positive, obligately anaerobic bacteria were isolated from different marine sediment samples of Gujarat. All the twenty three strains have 16S rRNA gene sequence similarity of ~100%. Strain JC272T was designated as the type strain and has sequence similarity with Clostridium bifermentans ATCC638T (99.8%), Clostridium ghonii JCM1400T (98.0%), Clostridium sordellii ATCC9714T (97.9%) and other members of the genus Clostridium (<96.4%). C16:0, C18:0, C17:0, C16:1ω9C and iso-C16:0 are the major (>5%) fatty acids. Strain JC272T contains diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine and unidentified amino lipids (AL1&AL2). However, genome based analysis of ANI and in silico DDH of strain JC272T with C. bifermentans ATCC 638T yielded values of 94.35% and 58.5+2.8%, respectively. G+C mol% of strain JC272T was 28.3%. Strain JC272T together with C. bifermentans fall outside Clostridium rRNA cluster I considered as Clostridium senso stricto. Based on ANI value, in-silico DDH, distinct morphological and physiological differences from the previously described taxa, we propose strain JC272T as a representative of a new genus and species in the family Clostridiaceae, for which the name Paraclostridium benzoelyticum gen. nov., sp. nov. is proposed. Type strain is JC272T (=KCTC15476T =LMG28745T). It is also proposed to transfer C. bifermentans to this new genus, as Paraclostridium bifermentans comb. nov. (type strain is ATCC638T =DSM14991T =JCM1386T). We also propose the genus Paeniclostridium gen. nov. to accommodate Clostridium sordellii and Clostridium ghonii as Paeniclostridium sordellii comb. nov. (type strain is ATCC9714T =LMG15708T =JCM3814T) and Paeniclostridium ghonii comb. nov. (type strain is ATCC25757T = DSM15049T =JCM1400T).

  16. Genomics of Clostridium tetani.

    PubMed

    Brüggemann, Holger; Brzuszkiewicz, Elzbieta; Chapeton-Montes, Diana; Plourde, Lucile; Speck, Denis; Popoff, Michel R

    2015-05-01

    Genomic information about Clostridium tetani, the causative agent of the tetanus disease, is scarce. The genome of strain E88, a strain used in vaccine production, was sequenced about 10 years ago. One additional genome (strain 12124569) has recently been released. Here we report three new genomes of C. tetani and describe major differences among all five C. tetani genomes. They all harbor tetanus-toxin-encoding plasmids that contain highly conserved genes for TeNT (tetanus toxin), TetR (transcriptional regulator of TeNT) and ColT (collagenase), but substantially differ in other plasmid regions. The chromosomes share a large core genome that contains about 85% of all genes of a given chromosome. The non-core chromosome comprises mainly prophage-like genomic regions and genes encoding environmental interaction and defense functions (e.g. surface proteins, restriction-modification systems, toxin-antitoxin systems, CRISPR/Cas systems) and other fitness functions (e.g. transport systems, metabolic activities). This new genome information will help to assess the level of genome plasticity of the species C. tetani and provide the basis for detailed comparative studies.

  17. [Antimicrobial susceptibility testing of anaerobic bacteria].

    PubMed

    García-Sánchez, José E; García-Sánchez, Enrique; García-García, María Inmaculada

    2014-02-01

    The anaerobic bacteria resistance to antibiotics is increasing, and even has appeared against the most active of those, like metronidazol and carbapenems. This fact forces to make and periodical sensibility tests -at least in the most aggressive and virulent species, in cases that they are isolated from life locations and in the absence of therapeutic response- to check the local sensibility and to establish suitable empiric therapies, all based on multicentric studies carried out in order to this or well to check the activity of new antibiotics. For the laboratory routine, the easiest sensibility method is the E-test/MIC evaluator. Another alternative is microdilution, that's only normalized for Bacteroides. There are preliminary facts that allow the use of disc diffusion method in some species of Bacteroides and Clostridium. For the temporal and multicentric studies, the procedure is dilution in agar plate, the reference method.

  18. Development of a High-Efficiency Transformation Method and Implementation of Rational Metabolic Engineering for the Industrial Butanol Hyperproducer Clostridium saccharoperbutylacetonicum Strain N1-4.

    PubMed

    Herman, Nicolaus A; Li, Jeffrey; Bedi, Ripika; Turchi, Barbara; Liu, Xiaoji; Miller, Michael J; Zhang, Wenjun

    2017-01-15

    While a majority of academic studies concerning acetone, butanol, and ethanol (ABE) production by Clostridium have focused on Clostridium acetobutylicum, other members of this genus have proven to be effective industrial workhorses despite the inability to perform genetic manipulations on many of these strains. To further improve the industrial performance of these strains in areas such as substrate usage, solvent production, and end product versatility, transformation methods and genetic tools are needed to overcome the genetic intractability displayed by these species. In this study, we present the development of a high-efficiency transformation method for the industrial butanol hyperproducer Clostridium saccharoperbutylacetonicum strain N1-4 (HMT) ATCC 27021. Following initial failures, we found that the key to creating a successful transformation method was the identification of three distinct colony morphologies (types S, R, and I), which displayed significant differences in transformability. Working with the readily transformable type I cells (transformation efficiency, 1.1 × 10(6) CFU/μg DNA), we performed targeted gene deletions in C. saccharoperbutylacetonicum N1-4 using a homologous recombination-mediated allelic exchange method. Using plasmid-based gene overexpression and targeted knockouts of key genes in the native acetone-butanol-ethanol (ABE) metabolic pathway, we successfully implemented rational metabolic engineering strategies, yielding in the best case an engineered strain (Clostridium saccharoperbutylacetonicum strain N1-4/pWIS13) displaying an 18% increase in butanol titers and 30% increase in total ABE titer (0.35 g ABE/g sucrose) in batch fermentations. Additionally, two engineered strains overexpressing aldehyde/alcohol dehydrogenases (encoded by adh11 and adh5) displayed 8.5- and 11.8-fold increases (respectively) in batch ethanol production.

  19. Genomics of Clostridium

    NASA Astrophysics Data System (ADS)

    Jacobson, Mark Joseph; Johnson, Eric A.

    The clostridia have a rich history and contemporary importance in industrial, environmental, and medical microbiology. Due to their ability to form endospores, clostridia are ubiquitous in nature and are found in many environments, especially in soils and the intestinal tract of animals including humans. Many clostridia cause devastating diseases of humans and animals, such as botulism, tetanus, and gas gangrene, through the production of protein toxins. The clostridia produce more protein toxins that are lethal for humans and animals than any other bacterial genus (Johnson, 2005; Van Heyningen, 1950). Other species are important in the formation of solvents and organic acids by anaerobic fermentations or as a source of unique enzymes for biocatalysis (Bradshaw and Johnson, 2010; Hatheway and Johnson, 1998).

  20. Anaerobes as Sources of Bioactive Compounds and Health Promoting Tools.

    PubMed

    Mamo, Gashaw

    Aerobic microorganisms have been sources of medicinal agents for several decades and an impressive variety of drugs have been isolated from their cultures, studied and formulated to treat or prevent diseases. On the other hand, anaerobes, which are believed to be the oldest life forms on earth and evolved remarkably diverse physiological functions, have largely been neglected as sources of bioactive compounds. However, results obtained from the limited research done so far show that anaerobes are capable of producing a range of interesting bioactive compounds that can promote human health. In fact, some of these bioactive compounds are found to be novel in their structure and/or mode of action.Anaerobes play health-promoting roles through their bioactive products as well as application of whole cells. The bioactive compounds produced by these microorganisms include antimicrobial agents and substances such as immunomodulators and vitamins. Bacteriocins produced by anaerobes have been in use as preservatives for about 40 years. Because these substances are effective at low concentrations, encounter relatively less resistance from bacteria and are safe to use, there is a growing interest in these antimicrobial agents. Moreover, several antibiotics have been reported from the cultures of anaerobes. Closthioamide and andrimid produced by Clostridium cellulolyticum and Pantoea agglomerans, respectively, are examples of novel antibiotics of anaerobe origin. The discovery of such novel bioactive compounds is expected to encourage further studies which can potentially lead to tapping of the antibiotic production potential of this fascinating group of microorganisms.Anaerobes are widely used in preparation of fermented foods and beverages. During the fermentation processes, these organisms produce a number of bioactive compounds including anticancer, antihypertensive and antioxidant substances. The well-known health promoting effect of fermented food is mostly due to these

  1. Sequential degradation of chlorophenols in anaerobic freshwater sediments

    SciTech Connect

    Zhang, X.

    1993-01-01

    Anaerobic degradation of 2,4-dichlorophenol and 3-chloro-4-hydroxybenzoate in the freshwater sediment samples was investigated. Studies of the enrichment cultures and a pure culture, adaptation times, correlation of substrate degradation and product accumulation, maximal observed transformation rates, temperature and pH ranges for the transformation provided the bases for the proposed sequential pathway for degradation of 2,4-dichlorophenol. At least six different bacterial species were required to catalyze following reactions: (1) the dechlorination of 2,4-dichlorophenol; (2) the dechlorination of 4-chlorophenol; (3) the para-carboxylation of phenol; (4) the reductive dehydroxylation of para-hydroxybenzoate; (5) the degradation of benzoate to acetate, H[sub 2] and CO[sub 2]; and (6) the conversion of H[sub 2]/CO[sub 2] and acetate to methane. The rate limiting reaction in the pathway was the dechlorination of 4-chlorophenol. A new species, Clostridium [open quote]hydroxybenzoicum[close quote], isolated from the enrichment, catalyzed the carboxylation of phenol at the para-position to 4-hydroxybenzoate by a reversible decarboxylation/carboxylation enzyme. 3,4-Dihydroxybenzoate was decarboxylated by a second enzyme in this organism. The activities were biotin and ATP independent. The bacterium, in a pure culture, did not benefit from the decarboxylation reaction but apparently it benefited in the phenol-degrading enrichment culture. Of 46 strains (42 species) tested, only three exhibited hydroxybenzoate decarboxylation activities:Clostridium thermoaceticum, Clostridium thermoautotrophicum,Clostridium scatologenes. The history of the sediment determined the first step in the anaerobic degradation.

  2. Descriptions of Anaerotaenia torta gen. nov., sp. nov. and Anaerocolumna cellulosilytica gen. nov., sp. nov. isolated from a methanogenic reactor of cattle waste and reclassification of Clostridium aminovalericum, Clostridium jejuense and Clostridium xylanovorans as Anaerocolumna species.

    PubMed

    Ueki, Atsuko; Ohtaki, Yoshimi; Kaku, Nobuo; Ueki, Katsuji

    2016-09-01

    Strictly anaerobic bacterial strains (FH052T and SN021T) belonging to clostridial cluster XIVa were isolated from a methanogenic reactor. Cells of the two strains were Gram-stain-positive, slender or curved rods producing terminal spores. The strains were slightly alkaliphilic. They fermented various carbohydrates including xylan and produced acetate, ethanol and H2. Strain SN021T decomposed cellulose. The genomic DNA G+C contents were 47.2 mol% for strain FH052T and 38.1 mol% for strain SN021T. The two strains had common cellular fatty acids such as C16 : 0, C16 : 0 dimethylacetal and C18 : 1ω7c dimethylacetal as major components. 16S rRNA gene sequence similarity between the two strains was 94.3 % and they shared closely related species such as Clostridium jejuense, Clostridium xylanovorans and Clostridium aminovalericum (92.6-95.7 % sequence similarity). Phenotypic and phylogenetic analyses suggested that these two isolates should be assigned to novel genera other than the genus Clostridium, and thus the genera Anaerotaenia gen. nov. and Anaerocolumna gen. nov. in the family Lachnospiraceae were proposed to accommodate them as Anaerotaenia torta gen. nov., sp. nov. for strain FH052T (=JCM 30820T=DSM 100431T) and Anaerocolumna cellulosilytica gen. nov., sp. nov. for strain SN021T (=JCM 30819T=DSM 100423T). For the three related Clostridium species, Anaerocolumna aminovalerica DSM 1283T (=JCM 11016T=ATCC 13725T) comb. nov., Anaerocolumna jejuensis HY-35-12T (=DSM 15929T=KCTC 5026T) comb. nov. and Anaerocolumna xylanovoransstrain HESP1T (=DSM 12503T=JCM 31057T) comb. nov. are proposed with emended descriptions of these species.

  3. Policy development for Clostridium difficile.

    PubMed

    Wilcox, Mark H

    2012-07-01

    The Advisory Committee on Antimicrobial Resistance and Healthcare Associated Infection (ARHAI) was created at the height of the incidence of Clostridium difficile infection (CDI). This article describes the role of ARHAI in the evaluation of laboratory testing for CDI, a related consultation on the legal requirements for manufacturers of in vitro diagnostic medical devices, a CDI healthcare bundle and surveillance of CDI in children.

  4. Clostridiumm ljungdahlii, an anaerobic ethanol and acetate producing microorganism

    DOEpatents

    Gaddy, James L.; Clausen, Edgar C.

    1992-01-01

    A newly discovered microorganism was isolated in a biologically pure culture and designated Clostridium ljungdahlii, having the identifying characteristics of ATCC No. 49587. Cultured in an aqueous nutrient medium under anaerobic conditions, this microorganism is capable of producing ethanol and acetate from CO and H.sub.2 O and/or CO.sub.2 and H.sub.2 in synthesis gas. Under optimal growth conditions, the microorganism produces acetate in preference to ethanol. Conversely, under non-growth conditions, ethanol production is favored over acetate.

  5. Clostridiumm ljungdahlii, an anaerobic ethanol and acetate producing microorganism

    DOEpatents

    Gaddy, J.L.; Clausen, E.C.

    1992-12-22

    A newly discovered microorganism was isolated in a biologically pure culture and designated Clostridium ljungdahlii, having the identifying characteristics of ATCC No. 49587. Cultured in an aqueous nutrient medium under anaerobic conditions, this microorganism is capable of producing ethanol and acetate from CO and H[sub 2]O and/or CO[sub 2] and H[sub 2] in synthesis gas. Under optimal growth conditions, the microorganism produces acetate in preference to ethanol. Conversely, under non-growth conditions, ethanol production is favored over acetate. 3 figs.

  6. Proline-Dependent Regulation of Clostridium difficile Stickland Metabolism

    PubMed Central

    Bouillaut, Laurent; Self, William T.

    2013-01-01

    Clostridium difficile, a proteolytic Gram-positive anaerobe, has emerged as a significant nosocomial pathogen. Stickland fermentation reactions are thought to be important for growth of C. difficile and appear to influence toxin production. In Stickland reactions, pairs of amino acids donate and accept electrons, generating ATP and reducing power in the process. Reduction of the electron acceptors proline and glycine requires the d-proline reductase (PR) and the glycine reductase (GR) enzyme complexes, respectively. Addition of proline in the medium increases the level of PR protein but decreases the level of GR. We report the identification of PrdR, a protein that activates transcription of the PR-encoding genes in the presence of proline and negatively regulates the GR-encoding genes. The results suggest that PrdR is a central metabolism regulator that controls preferential utilization of proline and glycine to produce energy via the Stickland reactions. PMID:23222730

  7. Methods for detection of Clostridium botulinum toxin in foods.

    PubMed

    Sharma, Shashi K; Whiting, Richard C

    2005-06-01

    Botulism is a deadly disease caused by ingestion of the preformed neurotoxin produced from the anaerobic spore-forming bacteria Clostridium botulinum. Botulinum neurotoxins are the most poisonous toxins known and have been a concern in the food industry for a long time. Therefore, rapid identification of botulinum neurotoxin using molecular and biochemical techniques is an essential component in the establishment of coordinated laboratory response systems and is the focus of current research and development. Because of the extreme toxicity of botulinum neurotoxin, some confirmatory testing with the mouse bioassay is still necessary, but rapid methods capable of screening large numbers of samples are also needed. This review is focused on the development of several detection methods for botulinum neurotoxins in foods.

  8. Clostridium botulinum: an increasing complication of heroin misuse.

    PubMed

    Cooper, Jamie G; Spilke, Cord E; Denton, Miles; Jamieson, Stuart

    2005-10-01

    Wound botulism is a rare infectious disease due to neurotoxin release from the anaerobic, spore-forming bacterium Clostridium botulinum that is becoming an ever more frequent complication of parenteral drug abuse in the Western world. Before the year 2000, no such cases had been reported in the UK and Ireland, but since then the number of proven and suspected cases of wound botulism occurring in parenteral drug users has increased markedly. The diagnosis is often difficult, based on a high degree of clinical suspicion and if not considered in the initial differential diagnosis, then considerable delays in treatment may result. This is the case report of a male heroin user who presented three times to an Emergency Department in the UK before a diagnosis of wound botulism was made and treatment commenced. It is important that emergency clinicians are aware of the possibility of wound botulism in parenteral drug users that present with unusual neurological or respiratory symptomatology.

  9. Metronidazole-triazole conjugates: Activity against Clostridium difficile and parasites

    PubMed Central

    Jarrad, Angie M.; Karoli, Tomislav; Debnath, Anjan; Tay, Chin Yen; Huang, Johnny X.; Kaeslin, Geraldine; Elliott, Alysha G.; Miyamoto, Yukiko; Ramu, Soumya; Kavanagh, Angela M.; Zuegg, Johannes; Eckmann, Lars; Blaskovich, Mark A.T.; Cooper, Matthew A.

    2015-01-01

    Metronidazole has been used clinically for over 50 years as an antiparasitic and broad-spectrum antibacterial agent effective against anaerobic bacteria. However resistance to metronidazole in parasites and bacteria has been reported, and improved second-generation metronidazole analogues are needed. The copper catalysed Huigsen azide-alkyne 1,3-dipolar cycloaddition offers a way to efficiently assemble new libraries of metronidazole analogues. Several new metronidazole-triazole conjugates (Mtz-triazoles) have been identified with excellent broad spectrum antimicrobial and antiparasitic activity targeting Clostridium difficile, Entamoeba histolytica and Giardia lamblia. Cross resistance to metronidazole was observed against stable metronidazole resistant C. difficile and G. lamblia strains. However for the most potent Mtz-triazoles, the activity remained in a therapeutically relevant window. PMID:26117821

  10. Metronidazole-triazole conjugates: activity against Clostridium difficile and parasites.

    PubMed

    Jarrad, Angie M; Karoli, Tomislav; Debnath, Anjan; Tay, Chin Yen; Huang, Johnny X; Kaeslin, Geraldine; Elliott, Alysha G; Miyamoto, Yukiko; Ramu, Soumya; Kavanagh, Angela M; Zuegg, Johannes; Eckmann, Lars; Blaskovich, Mark A T; Cooper, Matthew A

    2015-08-28

    Metronidazole has been used clinically for over 50 years as an antiparasitic and broad-spectrum antibacterial agent effective against anaerobic bacteria. However resistance to metronidazole in parasites and bacteria has been reported, and improved second-generation metronidazole analogues are needed. The copper catalysed Huigsen azide-alkyne 1,3-dipolar cycloaddition offers a way to efficiently assemble new libraries of metronidazole analogues. Several new metronidazole-triazole conjugates (Mtz-triazoles) have been identified with excellent broad spectrum antimicrobial and antiparasitic activity targeting Clostridium difficile, Entamoeba histolytica and Giardia lamblia. Cross resistance to metronidazole was observed against stable metronidazole resistant C. difficile and G. lamblia strains. However for the most potent Mtz-triazoles, the activity remained in a therapeutically relevant window.

  11. [Tetanus and Clostridium tetani--a brief review].

    PubMed

    Stock, Ingo

    2015-02-01

    Tetanus is an acute, often fatal, disease caused by an exotoxin (tetanospasmin) produced by the anaerobic, gram-positive spore-forming bacterium Clostridium tetani. It is characterized by generalized rigidity and convulsive spasms of skeletal muscles. In most industrialized countries, tetanus is a rare disease. However, in many tropical and subtropical countries with low vaccination coverage and poor medical care, it is still widely distributed. This applies in particular for neonatal tetanus. About 50 000 newborns and infants die each year from consequences from this severe illness. Management of tetanus involves neutralization of free circulating toxin, adequate antibacterial and symptomatic therapy as well as intensive care of the patient. For prophylaxis of the disease, active tetanus toxoid vaccination is the method of choice.

  12. Traits of selected Clostridium strains for syngas fermentation to ethanol.

    PubMed

    Martin, Michael E; Richter, Hanno; Saha, Surya; Angenent, Largus T

    2016-03-01

    Syngas fermentation is an anaerobic bioprocess that could become industrially relevant as a biorefinery platform for sustainable production of fuels and chemicals. An important prerequisite for commercialization is adequate performance of the biocatalyst (i.e., sufficiently high production rate, titer, selectivity, yield, and stability of the fermentation). Here, we compared the performance of three potential candidate Clostridium strains in syngas-to-ethanol conversion: Clostridium ljungdahlii PETC, C. ljungdahlii ERI-2, and Clostridium autoethanogenum JA1-1. Experiments were conducted in a two-stage, continuously fed syngas-fermentation system that had been optimized for stable ethanol production. The two C. ljungdahlii strains performed similar to each other but different from C. autoethanogenum. When the pH value was lowered from 5.5 to 4.5 to induce solventogenesis, the cell-specific carbon monoxide and hydrogen consumption (similar rate for all strains at pH 5.5), severely decreased in JA1-1, but hardly in PETC and ERI-2. Ethanol production in strains PETC and ERI-2 remained relatively stable while the rate of acetate production decreased, resulting in a high ethanol/acetate ratio, but lower overall productivities. With JA1-1, lowering the pH severely lowered rates of both ethanol and acetate production; and as a consequence, no pronounced shift to solventogenesis was observed. The highest overall ethanol production rate of 0.301 g · L(-1)  · h(-1) was achieved with PETC at pH 4.5 with a corresponding 19 g/L (1.9% w/v) ethanol concentration and a 5.5:1 ethanol/acetate molar ratio. A comparison of the genes relevant for ethanol metabolism revealed differences between C. ljungdahlii and C. autoethanogenum that, however, did not conclusively explain the different phenotypes.

  13. Microbiology of wetwood: importance of pectin degradation and Clostridium species in living trees. [Eastern Cottonwood; Block Poplar; American Elm

    SciTech Connect

    Schink, B.; Ward, J.C.; Zeikus, J.G.

    1981-09-01

    Wetwood samples from standing trees of eastern cottonwood (Populus deltoides), black poplar (Populus nigra), and American elm (Ulmus americana) contained high numbers of aerobic and anaerobic pectin-degrading bacteria (10 to the power of 4 to 10 to the power of 6 cells per g of wood). High activity of polygalacturonate lyase (is less than or equal to 0.5 U/ml) was also detected in the fetid liquid that spurted from wetwood zones in the lower trunk when the trees were bored. A prevalent pectin-degrading obligately anaerobic bacterium isolated from these wetwoods was identified as Clostridium butyricum. Pectin decomposition by Clostridium butyricum strain 4P1 was associated with an inducible polygalacturonate lyase and pectin methylesterase, the same types of pectinolytic activity expressed in the wetwood of these trees. The pH optimum of the extracellular polygalacturonate lyase was alkaline (near pH 8.5). In vitro tests with sapwood samples from a conifer (Douglas fir, Pseudotsuga menziesii) showed that tori in membranes of bordered pits are degraded by pure cultures of strain 4P1, polygalacturonate lyase enzyme preparations of strain 4P1, and mixed methanogenic cultures from the tree samples of wetwood. These results provide evidence that pectin in xylem tissue is actively degraded by Clostridium butyricum strain 4P1 via polygalacturonate lyase activity. The importance of pectin degradation by bacteria, including Clostridium species, appears paramount in the formation and maintenance of the wetwood syndrome in certain living trees. (Refs. 38).

  14. Comparative investigation on microbial community and electricity generation in aerobic and anaerobic enriched MFCs.

    PubMed

    Quan, Xiang-chun; Quan, Yan-ping; Tao, Kun; Jiang, Xiao-man

    2013-01-01

    This study compared the difference in microbial community and power generation capacity of air-cathode MFCs enriched under anode aerobic and anaerobic conditions. Results showed that MFCs successfully started with continuous air inputting to anode chamber. The aerobic enriched MFC produced comparable and even more electricity with the fuels of acetate, glucose and ethanol compared to the anaerobic MFC when returning to anaerobic condition. The two MFCs showed a slightly different microbial community for anode biofilms (a similarity of 77%), but a highly similar microbial community (a similarity of 97%) for anolyte microbes. The anode biofilm of aerobic enriched MFC showed the presence of some specific bacteria closely related to Clostridium sticklandii, Leucobacter komagatae and Microbacterium laevaniformans. The anaerobic enriched MFC found the presence of a large number of yeast Trichosporon sp. This research demonstrates that it is possible to enrich oxygen-tolerant anode respiring bacteria through purposely aeration in anode chamber.

  15. Insights into the global regulation of anaerobic metabolism for improved biohydrogen production.

    PubMed

    Lu, Yuan; Zhao, Hongxin; Zhang, Chong; Xing, Xin-Hui

    2016-01-01

    To improve the biohydrogen yield in bacterial dark fermentation, a new approach of global anaerobic regulation was introduced. Two cellular global regulators FNR and NarP were overexpressed in two model organisms: facultatively anaerobic Enterobacter aerogenes (Ea) and strictly anaerobic Clostridium paraputrificum (Cp). The overexpression of FNR and NarP greatly altered anaerobic metabolism and increased the hydrogen yield by 40%. Metabolic analysis showed that the global regulation caused more reducing environment inside the cell. To get a thorough understanding of the global metabolic regulation, more genes (fdhF, fhlA, ppk, Cb-fdh1, and Sc-fdh1) were overexpressed in different Ea and Cp mutants. For the first time, it demonstrated that there were approximately linear relationships between the relative change of hydrogen yield and the relative change of NADH yield or ATP yield. It implied that cellular reducing power and energy level played vital roles in the biohydrogen production.

  16. Survey of neuraminidase production by Clostridium butyricum, Clostridium beijerinckii, and Clostridium difficile strains from clinical and nonclinical sources.

    PubMed Central

    Popoff, M R; Dodin, A

    1985-01-01

    Neuraminidase production was investigated in 57 Clostridium butyricum strains, 16 Clostridium beijerinckii strains, and 25 Clostridium difficile strains. Neuraminidase activity was found only in C. butyricum strains originating from one human newborn with neonatal necrotizing enterocolitis, two newborns with hemorrhagic colitis, one infected placenta, and one adult with peritonitis, It was concluded that neuraminidase was not a major virulence factor in C. butyricum strains. PMID:4056013

  17. Antibiotic Susceptibility Pattern of Aerobic and Anaerobic Bacteria Isolated From Surgical Site Infection of Hospitalized Patients

    PubMed Central

    Akhi, Mohammad Taghi; Ghotaslou, Reza; Beheshtirouy, Samad; Asgharzadeh, Mohammad; Pirzadeh, Tahereh; Asghari, Babak; Alizadeh, Naser; Toloue Ostadgavahi, Ali; Sorayaei Somesaraei, Vida; Memar, Mohammad Yousef

    2015-01-01

    Background: Surgical Site Infections (SSIs) are infections of incision or deep tissue at operation sites. These infections prolong hospitalization, delay wound healing, and increase the overall cost and morbidity. Objectives: This study aimed to investigate anaerobic and aerobic bacteria prevalence in surgical site infections and determinate antibiotic susceptibility pattern in these isolates. Materials and Methods: One hundred SSIs specimens were obtained by needle aspiration from purulent material in depth of infected site. These specimens were cultured and incubated in both aerobic and anaerobic condition. For detection of antibiotic susceptibility pattern in aerobic and anaerobic bacteria, we used disk diffusion, agar dilution, and E-test methods. Results: A total of 194 bacterial strains were isolated from 100 samples of surgical sites. Predominant aerobic and facultative anaerobic bacteria isolated from these specimens were the members of Enterobacteriaceae family (66, 34.03%) followed by Pseudomonas aeruginosa (26, 13.4%), Staphylococcus aureus (24, 12.37%), Acinetobacter spp. (18, 9.28%), Enterococcus spp. (16, 8.24%), coagulase negative Staphylococcus spp. (14, 7.22%) and nonhemolytic streptococci (2, 1.03%). Bacteroides fragilis (26, 13.4%), and Clostridium perfringens (2, 1.03%) were isolated as anaerobic bacteria. The most resistant bacteria among anaerobic isolates were B. fragilis. All Gram-positive isolates were susceptible to vancomycin and linezolid while most of Enterobacteriaceae showed sensitivity to imipenem. Conclusions: Most SSIs specimens were polymicrobial and predominant anaerobic isolate was B. fragilis. Isolated aerobic and anaerobic strains showed high level of resistance to antibiotics. PMID:26421133

  18. Anaerobic thermophilic culture

    DOEpatents

    Ljungdahl, Lars G.; Wiegel, Jurgen K. W.

    1981-01-01

    A newly discovered thermophilic anaerobe is described that was isolated in a biologically pure culture and designated Thermoanaerobacter ethanolicus ATCC 3/550. T. Ethanolicus is cultured in aqueous nutrient medium under anaerobic, thermophilic conditions and is used in a novel process for producing ethanol by subjecting carbohydrates, particularly the saccharides, to fermentation action of the new microorganism in a biologically pure culture.

  19. Antimicrobial susceptibility of clinical isolates of anaerobic bacteria in Ontario, 2010-2011.

    PubMed

    Marchand-Austin, Alex; Rawte, Prasad; Toye, Baldwin; Jamieson, Frances B; Farrell, David J; Patel, Samir N

    2014-08-01

    The local epidemiology of antimicrobial susceptibility patterns in anaerobic bacteria is important in guiding the empiric treatment of infections. However, susceptibility data are very limited on anaerobic organisms, particularly among non-Bacteroides organisms. To determine susceptibility profiles of clinically-significant anaerobic bacteria in Ontario Canada, anaerobic isolates from sterile sites submitted to Public Health Ontario Laboratory (PHOL) for identification and susceptibility testing were included in this study. Using the E-test method, isolates were tested for various antimicrobials including, penicillin, cefoxitin, clindamycin, meropenem, piperacillin-tazobactam and metronidazole. The MIC results were interpreted based on guidelines published by Clinical and Laboratory Standards Institute. Of 2527 anaerobic isolates submitted to PHOL, 1412 were either from sterile sites or bronchial lavage, and underwent susceptibility testing. Among Bacteroides fragilis, 98.2%, 24.7%, 1.6%, and 1.2% were resistant to penicillin, clindamycin, piperacillin-tazobactam, and metronidazole, respectively. Clostridium perfringens was universally susceptible to penicillin, piperacillin-tazobactam, and meropenem, whereas 14.2% of other Clostridium spp. were resistant to penicillin. Among Gram-positive anaerobes, Actinomyces spp., Parvimonas micra and Propionibacterium spp. were universally susceptible to β-lactams. Eggerthella spp., Collinsella spp., and Eubacterium spp. showed variable resistance to penicillin. Among Gram-negative anaerobes, Fusobacterium spp., Prevotella spp., and Veillonella spp. showed high resistance to penicillin but were universally susceptible to meropenem and piperacillin-tazobactam. The detection of metronidazole resistant B. fragilis is concerning as occurrence of these isolates is extremely rare. These data highlight the importance of ongoing surveillance to provide clinically relevant information to clinicians for empiric management of

  20. Physical Characterization of Clostridium Botulinum Neurotoxin Genes

    DTIC Science & Technology

    1993-10-01

    jacewoburylicwn was obtained. 14. SUBJECT TERMS 115. NUMBER OF PAGES Foreign, BD, Clostridium , Cloning, Vaccines , BL3, ________ DNAotrahnsfery, Gn...met with either very limited success (eg., type A toxin of Clostridium difficile ; von Eichel-Streiber, 1989) or total failure (eg., the bacteriocin of...AD-A27 2 939 GRANT NO: DAMDl7-90-Z-0033 TITLE: PHYSICAL CHARACTERIZATION OF CLOSTRIDIUM BOTULINUM NEUROTOXIN GENES PRINCIPAL INVESTIGATOR: Nigel P

  1. [Antimicrobial activity of ornidazole and 6 other antibiotics against anaerobic bacteria].

    PubMed

    Alados, J C; Martínez-Brocal, A; Miranda, C; Rojo, M D; García, V; Domínguez, M C; de la Rosa, M

    1991-04-01

    The antimicrobial susceptibility of 235 anaerobic bacterial strains to ornidazole, metronidazole, chloramphenicol, clindamycin, penicillin, cefoxitin and imipenem has been studied using agar-dilution technique. Ornidazole and metronidazole were active against 88.6% and 86% of gram-positive cocci. Overall, 99.1% of Bacteroides group fragilis, and 91.3% of non-fragilis Bacteroides were also sensitive to both drugs. We did not find any Clostridium perfringens resistant strain. Cefoxitin and penicillin showed good activity against all Clostridium perfringens strains, and also against 97.7% and 92.5% of gram-positive cocci. We found one single imipenem resistant strain among gram-positive bacteria. Bacteroides fragilis also showed sensitivity to penicillin (41.5%), cefoxitin (85.7%) and imipenem (97.1%). Clindamycin was active against Clostridium perfringens (90.9%), gram-positive cocci (86.7%) and imipenem (68.6%). Chloramphenicol showed good activity against Clostridium perfringens (100%), gram-positive cocci (95.5%) and Bacteroides spp. (99.4%). Our results showed an overall good activity of all the seven drugs tested against anaerobic gram-positive microorganisms. Of notice, we found a good activity of chloramphenicol, imipenem, metronidazole and ornidazole against Bacteroides spp.

  2. Clostridium perfringens Epsilon Toxin: A Malevolent Molecule for Animals and Man?

    PubMed Central

    Stiles, Bradley G.; Barth, Gillian; Barth, Holger; Popoff, Michel R.

    2013-01-01

    Clostridium perfringens is a prolific, toxin-producing anaerobe causing multiple diseases in humans and animals. One of these toxins is epsilon, a 33 kDa protein produced by Clostridium perfringens (types B and D) that induces fatal enteric disease of goats, sheep and cattle. Epsilon toxin (Etx) belongs to the aerolysin-like toxin family. It contains three distinct domains, is proteolytically-activated and forms oligomeric pores on cell surfaces via a lipid raft-associated protein(s). Vaccination controls Etx-induced disease in the field. However, therapeutic measures are currently lacking. This review initially introduces C. perfringens toxins, subsequently focusing upon the Etx and its biochemistry, disease characteristics in various animals that include laboratory models (in vitro and in vivo), and finally control mechanisms (vaccines and therapeutics). PMID:24284826

  3. Electrophoretic study of Clostridium species.

    PubMed Central

    Cato, E P; Hash, D E; Holdeman, L V; Moore, W E

    1982-01-01

    Polyacrylamide gel electrophoretic analysis of soluble cellular proteins (without sodium dodecyl sulfate) of 70 Clostridium species indicated that the procedure was readily applicable to the differentiation of species in the genus. The protein patterns correlated well with the available DNA homology data and with most accepted differential tests. Results indicated that several earlier names for species were synonyms of those of accepted species and that two accepted species may be synonymous. Images PMID:6175658

  4. Anaerobic performance at altitude.

    PubMed

    Coudert, J

    1992-10-01

    Anaerobic metabolism is usually evaluated by the determination of the anaerobic capacity and the maximal anaerobic mechanical external power (Wmax). Conflicting results are reported on anaerobic capacity evaluated by maximal oxygen deficit and debt, and maximal blood lactate concentration during acute or chronic hypoxia (acclimatized subjects). Data on muscle biopsies (lactate concentration, changes in ATP, phosphocreatine and glycogen stores, glycolytic enzyme activities) and the few studies on lactate flux give in most cases evidence of a non-alteration of the anaerobic capacity for altitudes up to 5,500 m. No differences are observed in Wmax measured at high altitudes up to 5,200 m during intense short-term exercises: (1) jumps on a force platform which is a good indicator of alactic Wmax, and (2) 7-10 s sprints (i.e. force-velocity test) which solicit alactic metabolism but also lactic pathway. For exercises of duration equal or more than 30 s (i.e. Wingate test), there are conflicting results because a lower participation of aerobic metabolism during this test at high altitude can interfere with anaerobic performance. In conclusion, we can admit that anaerobic performances are not altered by high altitudes up to 5,200 m if the length of exposure does not exceed 5 weeks. After this period, muscle mass begins to decrease.

  5. Anaerobic Digestion and its Applications

    EPA Science Inventory

    Anaerobic digestion is a natural biological process. The initials "AD" may refer to the process of anaerobic digestion, or the built systems of anaerobic digesters. While there are many kinds of digesters, the biology is basically the same for all. Anaerobic digesters are built...

  6. Clostridium ljungdahlii represents a microbial production platform based on syngas

    PubMed Central

    Köpke, Michael; Held, Claudia; Hujer, Sandra; Liesegang, Heiko; Wiezer, Arnim; Wollherr, Antje; Ehrenreich, Armin; Liebl, Wolfgang; Gottschalk, Gerhard; Dürre, Peter

    2010-01-01

    Clostridium ljungdahlii is an anaerobic homoacetogen, able to ferment sugars, other organic compounds, or CO2/H2 and synthesis gas (CO/H2). The latter feature makes it an interesting microbe for the biotech industry, as important bulk chemicals and proteins can be produced at the expense of CO2, thus combining industrial needs with sustained reduction of CO and CO2 in the atmosphere. Sequencing the complete genome of C. ljungdahlii revealed that it comprises 4,630,065 bp and is one of the largest clostridial genomes known to date. Experimental data and in silico comparisons revealed a third mode of anaerobic homoacetogenic metabolism. Unlike other organisms such as Moorella thermoacetica or Acetobacterium woodii, neither cytochromes nor sodium ions are involved in energy generation. Instead, an Rnf system is present, by which proton translocation can be performed. An electroporation procedure has been developed to transform the organism with plasmids bearing heterologous genes for butanol production. Successful expression of these genes could be demonstrated, leading to formation of the biofuel. Thus, C. ljungdahlii can be used as a unique microbial production platform based on synthesis gas and carbon dioxide/hydrogen mixtures. PMID:20616070

  7. Anaerobic microbial dissolution of lead and production of organic acids

    DOEpatents

    Francis, A.J.; Dodge, C.; Chendrayan, K.; Quinby, H.L.

    1987-04-16

    The present invention related to an anaerobic bacterial culture of Clostridium sp. ATCC No. 53464 which solubilizes lead oxide under anaerobic conditions in coal and industrial wastes and therefore presents a method of removing lead from such wastes before they are dumped into the environment. The rat of lead dissolution during logarithmic growth of the bacteria in 40 ml medium containing 3.32 ..mu..moles of lead as lead oxide was 0.042 ..mu..moles m1/sup /-/1/ hr/sup /-/1/. Dissolution of lead oxide by the bacterial isolate is due to the production of metabolites and acidity in the culture medium. The major metabolites are acetic, butyric and lactic acid. The major metabolites are acetic, butyric and lactic acid. Clostridium sp. ATCC No. 53464 can be used in the recovery of the strategic metals from ores and wastes and also for the production of lactic acid for commercial purposes. The process yields large quantities of lactic acid as well as lead complexed in a stable form with said acids. 4 figs., 3 tabs.

  8. Anaerobic microbial dissolution of lead and production of organic acids

    DOEpatents

    Francis, Arokiasamy J.; Dodge, Cleveland; Chendrayan, Krishnachetty; Quinby, Helen L.

    1988-01-01

    The present invention relates to an anaerobic bacterial culture of Clostridium sp. ATCC No. 53464 which solubilizes lead oxide under anaerobic conditions in coal and industrial wastes and therefore presents a method of removing lead from such wastes before they are dumped into the environment. The rate of lead dissolution during logarithmic growth of the bacteria in 40 ml medium containing 3.32 .mu.moles of lead as lead oxide was 0.042 .mu.moles ml.sup.-1 hr.sup.-1. Dissolution of lead oxide by the bacterial isolate is due to the production of metabolites and acidity in the culture medium. The major metabolites are acetic, butyric and lactic acid. Clostridium sp. ATCC No. 53464 can be used in the recovery of strategic metals from ores and wastes and also for the production of lactic acid for commercial purposes. The process yields large quantities of lactic acid as well as lead complexed in a stable form with said acids.

  9. Hydrogen production from starch by co-culture of Clostridium acetobutylicum and Rhodobacter sphaeroides in one step hybrid dark- and photofermentation in repeated fed-batch reactor.

    PubMed

    Zagrodnik, R; Łaniecki, M

    2017-01-01

    Hydrogen production from starch by a co-culture hybrid dark and photofermentation under repeated fed-batch conditions at different organic loading rates (OLR) was studied. Effective cooperation between bacteria in co-culture during initial days was observed at controlled pH 7.0. However, at pH above 6.5 dark fermentation phase was redirected from H2 formation towards production of formic acid, lactic acid and ethanol (which are not coupled with hydrogen production) with simultaneous lower starch removal efficiency. This resulted in decrease in the hydrogen production rate. The highest H2 production in co-culture process (3.23LH2/Lmedium - after 11days) was achieved at OLR of 1.5gstarch/L/day, and it was twofold higher than for dark fermentation process (1.59LH2/Lmedium). The highest H2 yield in the co-culture (2.62molH2/molhexose) was obtained at the OLR of 0.375gstarch/L/day. Different pH requirements of bacteria were proven to be a key limitation in co-culture system.

  10. Use of the composite membrane of poly(ether-block-amide) and carbon nanotubes (CNTs) in a pervaporation system incorporated with fermentation for butanol production by Clostridium acetobutylicum.

    PubMed

    Yen, Hong-Wei; Chen, Zhi-Heng; Yang, I-Kuan

    2012-04-01

    Fermentation incorporated with pervaporation was regarded as an efficient way to relieve the feedback inhibition of butanol in acetone-butanol-ethanol (ABE) fermentation. The addition of CNTs (carbon nanotubes) to PEBA (poly(ether-block-amide)) could greatly enhance the removal flux of solvents (acetone, butanol and ethanol) in a model solution test. The butanol removing rate results in a 61% increase in the batch with PEBA+CNTs (5%) membrane compared with that of the batch with PEBA alone. Besides the increase of removal flux, the addition of CNTs enforces the mechanical strength of the pervaporation membrane, which leads to more resistance for a longer operational time. The combination of a 5-L fermenter with the pervaporation membrane of PEBA+CNTs (10%) indicates a 20% increase both in productivity and yield compared to using PEBA. In conclusion, the addition of CNTs to a PEBA pervaporation membrane has great potential when applied in the ABE fermentation industry.

  11. Use of poly(ether-block-amide) in pervaporation coupling with a fermentor to enhance butanol production in the cultivation of Clostridium acetobutylicum.

    PubMed

    Yen, Hong-Wei; Lin, Shang-Fu; Yang, I-Kuan

    2012-03-01

    The toxicity of the end-products of acetone-butanol-ethanol (ABE) process, mainly butanol, is recognized as the major problem contributing to the low productivity of butanol. The pervaporation technique was regarded as one of the ways to efficiently remove organic components. The results of pervaporation performance of poly(ether-block-amide) (PEBA) and polydimethylsiloxane (PDMS) membrane in a model solution indicated that PEBA membrane owned a higher butanol permeation flux of 9.975 gm(-2)h(-1) as opposed to 3.911 gm(-2)h(-1) using a PDMS membrane. Moreover, a higher temperature would result in a higher permeation flux, but has a lower separation factor (α) obtained, while using PEBA membrane. The batch fermentor operation connected to the pervaporation with PEBA membrane created 43% and 34% of increase in the butanol productivity and in the yield as compared to that of the simple batch. The fed-batch fermentation mode by glucose feeding combined with PEBA pervaporation lasting for 24h could achieve 39% increase of butanol productivity as compared to a simple batch. Conclusively, the pervaporation with PEBA membrane coupling with fermentor was presumed to be capable of enhancing butanol production in ABE fermentation, which might have the potential applied in the commercialized ABE fermentation process.

  12. Effect of chemical pretreatments on corn stalk bagasse as immobilizing carrier of Clostridium acetobutylicum in the performance of a fermentation-pervaporation coupled system.

    PubMed

    Cai, Di; Li, Ping; Chen, Changjing; Wang, Yong; Hu, Song; Cui, Caixia; Qin, Peiyong; Tan, Tianwei

    2016-11-01

    In this study, different pretreatment methods were evaluated for modified the corn stalk bagasse and further used the pretreated bagasse as immobilized carrier in acetone-butanol-ethanol fermentation process. Structural changes of the bagasses pretreated by different methods were analyzed by Fourier transform infrared, crystallinity index and scanning pictures by electron microscope. And the performances of batch fermentation using the corn stalk based carriers were evaluated. Results indicated that the highest ABE concentration of 23.86g/L was achieved using NaOH pretreated carrier in batch fermentation. Immobilized fermentation-pervaporation integration process was further carried out. The integration process showed long-term stability with 225-394g/L of ABE solvents on the permeate side of pervaporation membrane. This novel integration process was found to be an efficient method for biobutanol production.

  13. Clostridium perfringens: A review of enteric diseases in dogs, cats and wild animals.

    PubMed

    Silva, Rodrigo Otávio Silveira; Lobato, Francisco Carlos Faria

    2015-06-01

    Clostridium perfringens is a gram-positive anaerobic bacillus that is commonly part of the microbiota of humans and animals. It is considered a common enteric pathogen, but the pathogenesis and the predisposing factors of the disease commonly differ between host species. Thus, specific research is necessary to understand the role of this pathogen, how to diagnose it, and which control measures are applicable. The aim of this paper is to review the current knowledge of C. perfringens infections in dogs, cats and wild animals.

  14. Complete Genome Sequence of Clostridium septicum Strain CSUR P1044, Isolated from the Human Gut Microbiota

    PubMed Central

    Benamar, Samia; Cassir, Nadim; Caputo, Aurélia; Cadoret, Frédéric

    2016-01-01

    Clostridium septicum is one of the first pathogenic anaerobes to be identified. Here, we announce the genome draft sequence of C. septicum strain CSUR P1044 isolated from the gut of a healthy adult. Its chromosome genome consists of 3.2 Mbp with a plasmid of 32 Kbp. C. septicum strain CSUR P1044 has a G+C content of 27.5%, and is composed of 3,125 protein-coding genes together with 103 RNA genes, including 22 rRNA genes. PMID:27609912

  15. The inhibition of Clostridium botulinum type C by other bacteria in wetland sediments

    USGS Publications Warehouse

    Sandler, Renee J.; Rocke, Tonie E.; Yuill, Thomas M.

    1998-01-01

    Bacteria with inhibitory activity against Clostridium botulinum type C were isolated from 32% of sediment samples (n = 1600) collected from 10 marshes in a northern California wetland over a 12 mo period. Aerobic and anaerobic bacteria with inhibitory activity were isolated from 12% and 23% of the samples, respectively. Bacteria with inhibitory activity were isolated from all 10 study sites and throughout the year. This study demonstrates that bacteria with inhibitory activity against C. botulinum type C occur naturally in wetland sediments.

  16. Phylogenetic analysis and PCR detection of Clostridium chauvoei, Clostridium haemolyticum, Clostridium novyi types A and B, and Clostridium septicum based on the flagellin gene.

    PubMed

    Sasaki, Yoshimasa; Kojima, Akemi; Aoki, Hiroshi; Ogikubo, Yasuaki; Takikawa, Noriyasu; Tamura, Yutaka

    2002-05-01

    The flagellin genes (fliC) of Clostridium chauvoei, Clostridium haemolyticum, Clostridium novyi types A and B, and Clostridium septicum were analysed by PCR amplification and DNA sequencing. The five Clostridium species have at least two copies of the flagellin gene (fliC) arranged in tandem on the chromosome. The deduced N- and C-terminal aminoacid sequences of the flagellin proteins (FliCs) of these clostridia are well conserved but their central region aminoacid sequences are not. Phylogenic analysis based on the N-terminal aminoacid sequence of the FliC protein revealed that these clostridia, which belong to Clostridium 16S rDNA phylogenic cluster I (), are more closely related to Bacillus subtilis than to Clostridium difficile, which belongs to the cluster XI. Moreover, a multiplex polymerase reaction (PCR) system based on the fliC sequence was developed to rapidly identify C. chauvoei, C. haemolyticum, C. novyi types A and B, and C. septicum. PCR of each Clostridium amplified a species-specific band. The multiplex PCR system may be useful for rapid identification of pathogenic clostridia.

  17. Safety assessment of the Clostridium butyricum MIYAIRI 588® probiotic strain including evaluation of antimicrobial sensitivity and presence of Clostridium toxin genes in vitro and teratogenicity in vivo.

    PubMed

    Isa, K; Oka, K; Beauchamp, N; Sato, M; Wada, K; Ohtani, K; Nakanishi, S; McCartney, E; Tanaka, M; Shimizu, T; Kamiya, S; Kruger, C; Takahashi, M

    2016-08-01

    Probiotics are live microorganisms ingested for the purpose of conferring a health benefit on the host. Development of new probiotics includes the need for safety evaluations that should consider factors such as pathogenicity, infectivity, virulence factors, toxicity, and metabolic activity. Clostridium butyricum MIYAIRI 588(®) (CBM 588(®)), an anaerobic spore-forming bacterium, has been developed as a probiotic for use by humans and food animals. Safety studies of this probiotic strain have been conducted and include assessment of antimicrobial sensitivity, documentation of the lack of Clostridium toxin genes, and evaluation of CBM 588(®) on reproductive and developmental toxicity in a rodent model. With the exception of aminoglycosides, to which anaerobes are intrinsically resistant, CBM 588(®) showed sensitivity to all antibiotic classes important in human and animal therapeutics. In addition, analysis of the CBM 588(®) genome established the absence of genes for encoding for α, β, or ε toxins and botulin neurotoxins types A, B, E, or F. There were no deleterious reproductive and developmental effects observed in mice associated with the administration of CBM 588(®) These data provide further support for the safety of CBM 588(®) for use as a probiotic in animals and humans.

  18. Anaerobic brain abscess

    PubMed Central

    Sudhaharan, Sukanya; Chavali, Padmasri

    2016-01-01

    Background and Objectives: Brain abscess remains a potentially fatal central nervous system (CNS) disease, especially in developing countries. Anaerobic abscess is difficult to diagnose because of cumbersome procedures associated with the isolation of anaerobes. Materials and Methods: This is a hospital-based retrospective microbiological analysis of 430 brain abscess materials (purulent aspirates and/or tissue), for anaerobic organisms, that were received between 1987–2014, by the Microbiology Laboratory in our Institute. Results: Culture showed growth of bacteria 116/430 (27%) of the cases of which anaerobes were isolated in 48/116 (41.1%) of the cases. Peptostreptococcus (51.4 %), was the predominant organism isolated in four cases followed by Bacteroides and Peptococcus species. Conclusion: Early diagnosis and detection of these organisms would help in the appropriate management of these patients. PMID:27307977

  19. Biotransformation of 1,1,1-trichloroethane, trichloromethane, and tetrachloromethane by a Clostridium sp.

    PubMed Central

    Gälli, R; McCarty, P L

    1989-01-01

    A gram-positive, strictly anaerobic, motile, endospore-forming rod, tentatively identified as a proteolytic Clostridium sp., was isolated from the effluent of an anaerobic suspended-growth bioreactor. The organism was able to biotransform 1,1,1-trichloroethane, trichloromethane, and tetrachloromethane. 1,1,1-Trichloroethane was completely transformed (greater than or equal to 99.5%) by reductive dehalogenation to 1,1-dichloroethane (30 to 40%) and, presumably by other mechanisms, to acetic acid (7%) and unidentified products. The reductive dehalogenation of tetrachloromethane led to the intermediate trichloromethane, which was further transformed to dichloromethane (8%) and unidentified products. The biotransformation occurred during the exponential growth phase, as well as during the stationary phase. Tetrachlorethene, trichloroethene, 1,1-dichloroethene, chloroethane, 1,1-dichloroethane, and dichloromethane were not biotransformed significantly by the organism. PMID:2729985

  20. Impact of bioaugmentation on biochemical methane potential for wheat straw with addition of Clostridium cellulolyticum.

    PubMed

    Peng, Xiaowei; Börner, Rosa Aragão; Nges, Ivo Achu; Liu, Jing

    2014-01-01

    Hydrolysis is usually the rate-limited step for methane production from lignocellulosic substrate. Two bioaugmentation strategies, using the cellulolytic anaerobic bacteria Clostridium cellulolyticum, were adopted to enhance the hydrolysis of wheat straw with the purpose of improving the biochemical methane potential (BMP). Namely, the 24-h-incubated seed (C24S) with cellobiose as carbon source and the 60-h-incubated seed (WS60S) with wheat straw as carbon source were respectively used as the bioaugmentation agents. As a result, the BMPs were respectively 342.5 and 326.3 ml g(-1) VS of wheat straw, with an increase of 13.0% and 7.6% comparing to the no-bioaugmentation BMP of 303.3 ml g(-1) VS. The result indicates that the anaerobic digestion efficiency can be improved by bioaugmentation, which therefore may be a promising method for improving methane production from lignocellulosic substrate.

  1. Clostridium difficile and the microbiota

    PubMed Central

    Seekatz, Anna M.; Young, Vincent B.

    2014-01-01

    Clostridium difficile infection (CDI) is the leading health care–associated illness. Both human and animal models have demonstrated the importance of the gut microbiota’s capability of providing colonization resistance against C. difficile. Risk factors for disease development include antibiotic use, which disrupts the gut microbiota, leading to the loss of colonization resistance and subsequent CDI. Identification of the specific microbes capable of restoring this function remains elusive. Future studies directed at how microbial communities influence the metabolic environment may help elucidate the role of the microbiota in disease development. These findings will improve current biotherapeutics for patients with CDI, particularly those with recurrent disease. PMID:25036699

  2. Regulation of toxin synthesis in Clostridium botulinum and Clostridium tetani.

    PubMed

    Connan, Chloé; Denève, Cécile; Mazuet, Christelle; Popoff, Michel R

    2013-12-01

    Botulinum and tetanus neurotoxins are structurally and functionally related proteins that are potent inhibitors of neuroexocytosis. Botulinum neurotoxin (BoNT) associates with non-toxic proteins (ANTPs) to form complexes of various sizes, whereas tetanus toxin (TeNT) does not form any complex. The BoNT and ANTP genes are clustered in a DNA segment called the botulinum locus, which has different genomic localization (chromosome, plasmid, phage) in the various Clostridium botulinum types and subtypes. The botulinum locus genes are organized in two polycistronic operons (ntnh-bont and ha/orfX operons) transcribed in opposite orientations. A gene called botR lying between the two operons in C. botulinum type A encodes an alternative sigma factor which regulates positively the synthesis of BoNT and ANTPs at the late exponential growth phase and beginning of the stationary phase. In Clostridium tetani, the gene located immediately upstream of tent encodes a positive regulatory protein, TetR, which is related to BotR. C. botulinum and C. tetani genomes contain several two-component systems and predicted regulatory orphan genes. In C. botulinum type A, four two-component systems have been found that positively or negatively regulate the synthesis of BoNT and ANTPs independently of BotR/A. The synthesis of neurotoxin in Clostridia seems to be under the control of complex network of regulation.

  3. Gender comparisons in anaerobic power and anaerobic capacity tests.

    PubMed Central

    Maud, P J; Shultz, B B

    1986-01-01

    The purpose of the study was to compare anaerobic power and anaerobic capacity test scores between young active men and women. Three performance measures of anaerobic power and two of anaerobic capacity were administered to a sample comprising 52 male and 50 female college students (means age = 21.4 yrs). Results indicated significant differences between men and women in body height, weight and per cent fat, in fat free mass (FFM), anaerobic power, and anaerobic capacity when recorded as gross work completed and relative to body weight. However, these differences are reduced when data is adjusted for body weight and further reduced when corrected for FFM. The study found no significant differences between men and women in either anaerobic power or anaerobic capacity when values were given relative to FFM. PMID:3730753

  4. Hygienization of sludge through anaerobic digestion at 35, 55 and 60 °C.

    PubMed

    Kjerstadius, H; la Cour Jansen, J; De Vrieze, J; Haghighatafshar, S; Davidsson, A

    2013-01-01

    Legislation in Sweden and the European Union concerning the use of sewage sludge in agriculture is under revision and future concentration limits for pathogens in treated sludge are likely to be expected. The aim of this study was to evaluate the hygienization of Salmonella, Escherichia coli, Enterococcus and Clostridium perfringens through continuous anaerobic digestion at 35, 55 or 60 °C, as well as to investigate process stability and methane production at 60 °C. The results indicated that digestion at 55 or 60 °C with a minimum exposure time of 2 h resulted in good reduction of Salmonella, E. coli and Enterococcus and that anaerobic digestion could thus be used to reach the concentration limits suggested for the EU, as well as Sweden. Furthermore, stable continuous anaerobic digestion of sludge was achieved at 60 °C, albeit with 10% less methane production compared to digestion at 35 and 55 °C.

  5. [Anaerobic bacteria 150 years after their discovery by Pasteur].

    PubMed

    García-Sánchez, José Elías; García-Sánchez, Enrique; Martín-Del-Rey, Ángel; García-Merino, Enrique

    2015-02-01

    In 2011 we celebrated the 150th anniversary of the discovery of anaerobic bacteria by Louis Pasteur. The interest of the biomedical community on such bacteria is still maintained, and is particularly focused on Clostridium difficile. In the past few years important advances in taxonomy have been made due to the genetic, technological and computing developments. Thus, a significant number of new species related to human infections have been characterised, and some already known have been reclassified. At pathogenic level some specimens of anaerobic microflora, that had not been isolated from human infections, have been now isolated in some clinical conditions. There was emergence (or re-emergence) of some species and clinical conditions. Certain anaerobic bacteria have been associated with established infectious syndromes. The virulence of certain strains has increased, and some hypotheses on their participation in certain diseases have been given. In terms of diagnosis, the routine use of MALDI-TOF has led to a shortening of time and a cost reduction in the identification, with an improvement directly related to the improvement of data bases. The application of real-time PCR has been another major progress, and the sequencing of 16srRNA gene and others is currently a reality for several laboratories. Anaerobes have increased their resistance to antimicrobial agents, and the emergence of resistance to carbapenems and metronidazole, and multi-resistance is a current reality. In this situation, linezolid could be an effective alternative for Bacteroides. Fidaxomicin is the only anti-anaerobic agent introduced in the recent years, specifically for the diarrhoea caused by C.difficile. Moreover, some mathematical models have also been proposed in relation with this species.

  6. Microbiology and physiology of anaerobic fermentation of cellulose. Annual report for 1990, 1992, 1993 and final report

    SciTech Connect

    Ljungdahl, L.G.; Wiegel, J.; Peck, H.D. Jr.; Mortenson, L.E.

    1993-08-31

    This report focuses on the bioconversion of cellulose to methane by various anaerobes. The structure and enzymatic activity of cellulosome and polycellulosome was studied in Clostridium thermocellum. The extracellular enzymes involved in the degradation of plant material and the physiology of fermentation was investigated in anaerobic fungi. Enzymes dealing with CO, CO{sub 2}, H{sub 2}, CH{sub 3}OH, as well as electron transport and energy generation coupled to the acetyl-CoA autotrophic pathway was studied in acetogenic clostridia.

  7. Prognostic factors and impact of antibiotherapy in 117 cases of anaerobic bacteraemia.

    PubMed

    Robert, R; Deraignac, A; Le Moal, G; Ragot, S; Grollier, G

    2008-08-01

    Bacteraemia due to anaerobic bacteria occurs infrequently, making the systematic use of an anaerobic blood sample bottle in patients with sepsis controversial. We retrospectively reviewed the clinical and microbiological data from all cases of anaerobic bacteraemia in a teaching hospital over 2 years and determined the prognostic factors and antibiotic management. With the goal of evaluating the morbidity and mortality of bacteraemia due to anaerobic bacteria, a case-control study was also performed. One hundred eighty-four blood cultures from 125 patients grew at least one anaerobic bacterium, representing 0.5% of all and 7.0% of the positive blood cultures. One hundred seventeen patients were studied. In 24 cases, anaerobic blood cultures were associated with concomitant aerobic bacteria isolation. The most frequently isolated anaerobic species were Bacteroides sp. (n = 62), Clostridium sp. (n = 25), and Fusobacterium sp. (n = 12). The most frequent site of origin was the digestive tract (n = 61). In 51 cases, patients did not receive adequate empirical antianaerobic therapy. The mortality rate was 27%. Age [odds ratio (OR) 1.059; 95% confidence interval (CI) 1.021-1.100], cancer history (OR 3.21, 95% CI 1.126-9.156), and ineffective definitive antibiotherapy (OR 19.292, 95% CI 5.330-69.832) were independently associated with increased hospital mortality. The 72 patients that could be matched with patients without anaerobic bacteria according to their primary diagnosis had a longer hospitalisation and a trend toward increased mortality (P = 0.08). Anaerobic bacteraemia contributed significantly to the morbidity of the patients, and adequate empirical antibiotherapy may play an important role in the clinical outcomes.

  8. Stress hormone epinephrine (adrenaline) and norepinephrine (noradrenaline) effects on the anaerobic bacteria.

    PubMed

    Boyanova, Lyudmila

    2017-01-08

    Microbial endocrinology is a relatively new research area that already encompasses the anaerobes. Stress hormones, epinephrine and norepinephrine, can affect the growth of anaerobic bacteria such as Fusobacterium nucleatum, Prevotella spp., Porhyromonas spp., Tanerella forsythia and Propionibacterium acnes and can increase virulence gene expression, iron acquisition and many virulence factors of some anaerobic species such as Clostridium perfringens, Porphyromonas gingivalis and Brachyspira pilosicoli. Epinephrine and norepinephrine effects can lead to a growth increase or decrease, or no effect on the growth of the anaerobes. The effects are species-specific and perhaps strain-specific. Discrepancies in the results of some studies can be due to the different methods and media used, catecholamine concentrations, measurement techniques and the low number of strains tested. Biological effects of the stress hormones on the anaerobes may range from halitosis and a worsening of periodontal diseases to tissue damages and atherosclerotic plaque ruptures. Optimizations of the research methods and a detailed assessment of the catecholamine effects in conditions mimicking those in affected organs and tissues, as well as the effects on the quorum sensing and virulence of the anaerobes and the full spectrum of biological consequences of the effects are interesting topics for further evaluation.

  9. Lighting Up Clostridium Difficile: Reporting Gene Expression Using Fluorescent Lov Domains

    PubMed Central

    Buckley, Anthony M.; Jukes, Caitlin; Candlish, Denise; Irvine, June J.; Spencer, Janice; Fagan, Robert P.; Roe, Andrew J.; Christie, John M.; Fairweather, Neil F.; Douce, Gillian R.

    2016-01-01

    The uses of fluorescent reporters derived from green fluorescent protein have proved invaluable for the visualisation of biological processes in bacteria grown under aerobic conditions. However, their requirement for oxygen has limited their application in obligate anaerobes such as Clostridium difficile. Fluorescent proteins derived from Light, Oxygen or Voltage sensing (LOV) domains have been shown to bridge this limitation, but their utility as translational fusions to monitor protein expression and localisation in a strict anaerobic bacterium has not been reported. Here we demonstrate the utility of phiLOV in three species of Clostridium and its application as a marker of real-time protein translation and dynamics through genetic fusion with the cell division protein, FtsZ. Time lapse microscopy of dividing cells suggests that Z ring assembly arises through the extension of the FtsZ arc starting from one point on the circumference. Furthermore, through incorporation of phiLOV into the flagella subunit, FliC, we show the potential of bacterial LOV-based fusion proteins to be successfully exported to the extracellular environment. PMID:26996606

  10. Lytic Clostridium perfringens Bacteriophage 39-O Genomic

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Screening for bacteriophages lytic for Clostridium perfringens was completed utilizing filtered samples obtained from poultry (intestinal material), soil, sewage and poultry processing drainage water. Following limit dilution cloning and three rounds of plaque purification lytic phage preparations ...

  11. Evidence for antibiotic induced Clostridium perfringens diarrhoea

    PubMed Central

    Modi, N; Wilcox, M

    2001-01-01

    Clostridium difficile is a well documented cause of antibiotic associated diarrhoea in hospitalised patients, but may account for only approximately 20% of all cases. This leader reviews the current knowledge and understanding of the pathogenesis, epidemiology, and diagnosis of non-food borne Clostridium perfringens diarrhoea. Although enterotoxigenic C perfringens has been implicated in some C difficile negative cases of antibiotic associated diarrhoea, C perfringens enterotoxin detection methods are not part of the routine laboratory investigation of such cases. Testing for C perfringens enterotoxin in faecal samples from patients with antibiotic associated diarrhoea and sporadic diarrhoea on a routine basis would have considerable resource implications. Therefore, criteria for initiating investigations and optimum laboratory tests need to be established. In addition, establishing the true burden of C perfringens antibiotic associated diarrhoea is important before optimum control and treatment measures can be defined. Key Words: Clostridium perfringens • Clostridium difficile • hospital acquired infective diarrhoea PMID:11577119

  12. Clostridium difficile and C. difficile Toxin Testing

    MedlinePlus

    ... C diff antigen; GDH Formal name: Clostridium difficile Culture; C. difficile Toxin, A and B; C. difficile Cytotoxin Assay; Glutamate Dehydrogenase Test Related tests: Stool Culture ; O&P At a Glance Test Sample The ...

  13. Structure of a cellulose degrading bacterial community during anaerobic digestion.

    PubMed

    O'Sullivan, Cathryn A; Burrell, Paul C; Clarke, William P; Blackall, Linda L

    2005-12-30

    It is widely accepted that cellulose is the rate-limiting substrate in the anaerobic digestion of organic solid wastes and that cellulose solubilisation is largely mediated by surface attached bacteria. However, little is known about the identity or the ecophysiology of cellulolytic microorganisms from landfills and anaerobic digesters. The aim of this study was to investigate an enriched cellulolytic microbial community from an anaerobic batch reactor. Chemical oxygen demand balancing was used to calculate the cellulose solubilisation rate and the degree of cellulose solubilisation. Fluorescence in situ hybridisation (FISH) was used to assess the relative abundance and physical location of three groups of bacteria belonging to the Clostridium lineage of the Firmicutes that have been implicated as the dominant cellulose degraders in this system. Quantitation of the relative abundance using FISH showed that there were changes in the microbial community structure throughout the digestion. However, comparison of these results to the process data reveals that these changes had no impact on the cellulose solubilisation in the reactor. The rate of cellulose solubilisation was approximately stable for much of the digestion despite changes in the cellulolytic population. The solubilisation rate appears to be most strongly affected by the rate of surface area colonisation and the biofilm architecture with the accepted model of first order kinetics due to surface area limitation applying only when the cellulose particles are fully covered with a thin layer of cells.

  14. Effect of ozonolysis parameters on the inhibitory compound generation and on the production of ethanol by Pichia stipitis and acetone-butanol-ethanol by Clostridium from ozonated and water washed sugarcane bagasse.

    PubMed

    Travaini, Rodolfo; Barrado, Enrique; Bolado-Rodríguez, Silvia

    2016-10-01

    Sugarcane bagasse (SCB) was ozone pretreated and detoxified by water washing, applying a L9(3)(4) orthogonal array (OA) design of experiments to study the effect of pretreatment parameters (moisture content, ozone concentration, ozone/oxygen flow and particle size) on the generation of inhibitory compounds and on the composition of hydrolysates of ozonated-washed samples. Ozone concentration resulted the highest influence process parameter on delignification and sugar release after washing; while, for inhibitory compound formation, moisture content also had an important role. Ozone expended in pretreatment related directly with sugar release and inhibitory compound formation. Washing detoxification was effective, providing non-inhibitory hydrolysates. Maximum glucose and xylose release yields obtained were 84% and 67%, respectively, for ozonated-washed SCB. Sugar concentration resulted in the decisive factor for biofuels yields. Ethanol production achieved an 88% yield by Pichia stipitis, whereas Clostridium acetobutylicum produced 0.072gBUTANOL/gSUGAR and 0.188gABE/gSUGAR, and, Clostridium beijerinckii 0.165gBUTANOL/gSUGAR and 0.257gABE/gSUGAR.

  15. Interspecies Interactions between Clostridium difficile and Candida albicans

    PubMed Central

    van Leeuwen, Pim T.; van der Peet, Jasper M.; Bikker, Floris J.; Hoogenkamp, Michel A.; Oliveira Paiva, Ana M.; Kostidis, Sarantos; Mayboroda, Oleg A.

    2016-01-01

    ABSTRACT The facultative anaerobic polymorphic fungus Candida albicans and the strictly anaerobic Gram-positive bacterium Clostridium difficile are two opportunistic pathogens residing in the human gut. While a few studies have focused on the prevalence of C. albicans in C. difficile-infected patients, the nature of the interactions between these two microbes has not been studied thus far. In the current study, both chemical and physical interactions between C. albicans and C. difficile were investigated. In the presence of C. albicans, C. difficile was able to grow under aerobic, normally toxic, conditions. This phenomenon was neither linked to adherence of bacteria to hyphae nor to biofilm formation by C. albicans. Conditioned medium of C. difficile inhibited hyphal growth of C. albicans, which is an important virulence factor of the fungus. In addition, it induced hypha-to-yeast conversion. p-Cresol, a fermentation product of tyrosine produced by C. difficile, also induced morphological effects and was identified as an active component of the conditioned medium. This study shows that in the presence of C. albicans, C. difficile can persist and grow under aerobic conditions. Furthermore, p-cresol, produced by C. difficile, is involved in inhibiting hypha formation of C. albicans, directly affecting the biofilm formation and virulence of C. albicans. This study is the first detailed characterization of the interactions between these two gut pathogens. IMPORTANCE Candida albicans and Clostridium difficile are two opportunistic pathogens that reside in the human gut. A few studies have focused on the prevalence of C. albicans in C. difficile-infected patients, but none have shown the interaction(s) that these two organisms may or may not have with each other. In this study, we used a wide range of different techniques to better understand this interaction at a macroscopic and microscopic level. We found that in the presence of C. albicans, C

  16. The anaerobic digestion process

    SciTech Connect

    Rivard, C.J.; Boone, D.R.

    1996-01-01

    The microbial process of converting organic matter into methane and carbon dioxide is so complex that anaerobic digesters have long been treated as {open_quotes}black boxes.{close_quotes} Research into this process during the past few decades has gradually unraveled this complexity, but many questions remain. The major biochemical reactions for forming methane by methanogens are largely understood, and evolutionary studies indicate that these microbes are as different from bacteria as they are from plants and animals. In anaerobic digesters, methanogens are at the terminus of a metabolic web, in which the reactions of myriads of other microbes produce a very limited range of compounds - mainly acetate, hydrogen, and formate - on which the methanogens grow and from which they form methane. {open_quotes}Interspecies hydrogen-transfer{close_quotes} and {open_quotes}interspecies formate-transfer{close_quotes} are major mechanisms by which methanogens obtain their substrates and by which volatile fatty acids are degraded. Present understanding of these reactions and other complex interactions among the bacteria involved in anaerobic digestion is only now to the point where anaerobic digesters need no longer be treated as black boxes.

  17. Conditions associated with Clostridium sporogenes growth as a surrogate for Clostridium botulinum in nonthermally processed canned butter.

    PubMed

    Taylor, R H; Dunn, M L; Ogden, L V; Jefferies, L K; Eggett, D L; Steele, F M

    2013-05-01

    The objective of this study was to better understand the effect of butter composition and emulsion structure on growth and survival of Clostridium sporogenes, used as a surrogate for C. botulinum in canned butter. The lack of a thermal process step in commercially available canned butter raises questions of potential safety, because it is hermetically sealed and generally exhibits anaerobic growth conditions, which are optimal for Clostridium botulinum growth. Without thermal processing, low-acid canned foods must have inhibitory factors present to prevent C. botulinum growth. Some potential intrinsic inhibitory factors, or hurdles, within butter include: reduced water activity, acidity in cultured products, elevated salt content, and the micro-droplet nature of the aqueous phase in the butter emulsion. It was hypothesized that a normal, intact butter emulsion would have sufficient hurdles to prevent C. botulinum growth, whereas a broken butter emulsion would result in a coalesced aqueous phase that would allow for C. botulinum growth. Batch-churned butter was inoculated with C. sporogenes; butter samples with varying salt contents (0, 0.8, 1.6, and 2.4% wt/wt NaCl) were prepared and stored in coated steel cans for varying times (1 or 2 wk) and temperatures (22 or 41°C) to determine temperature and emulsion structure effects on C. sporogenes growth. Samples stored at 41°C showed a significant increase in C. sporogenes growth compared with those stored at 22°C. Furthermore, NaCl addition was found to have a significant effect on C. sporogenes growth, with 0.8% NaCl promoting more growth than 0%, but with decreases in growth observed at 1.6 and 2.4%. Uninoculated control plates were also found to have bacterial growth; this growth was attributed to other anaerobic bacteria present within the cream. It was concluded that removal of the hurdle created by the micro-droplet size of the emulsion aqueous phase could result in C. botulinum growth even at elevated salt

  18. [Clostridium-difficile-associated diarrhea].

    PubMed

    Bujanda, Luis; Cosme, Angel

    2009-01-01

    Clostridium difficile is the most frequent cause of nosocomial diarrhea and is a significant cause of morbidity among hospitalized patients. The inflammation is produced as a result of a non-specific response to toxins. In the last few years, a hypervirulent strain, NAP1/BI/027, has been reported. Symptoms usually consist of abdominal pain and diarrhea. The diagnosis should be suspected in any patient who develops diarrhea during antibiotic therapy or 6-8 weeks after treatment. Diagnosis should be confirmed by the detection of CD toxin in stool and by colonoscopy in special situations. The treatment of choice is metronidazole or vancomycin. In some patients who do not respond to this therapy or who have complications, subtotal colectomy may be required. Relapse is frequent and must be distinguished from reinfection. Prevention and control in healthcare settings requires careful attention.

  19. Identification of Clostridium Species and DNA Fingerprinting of Clostridium perfringens by Amplified Fragment Length Polymorphism Analysis▿

    PubMed Central

    Keto-Timonen, Riikka; Heikinheimo, Annamari; Eerola, Erkki; Korkeala, Hannu

    2006-01-01

    An amplified fragment length polymorphism (AFLP) method was applied to 129 strains representing 24 different Clostridium species, with special emphasis on pathogenic clostridia of medical or veterinary interest, to assess the potential of AFLP for identification of clostridia. In addition, the ability of the same AFLP protocol to type clostridia at the strain level was assessed by focusing on Clostridium perfringens strains. All strains were typeable by AFLP, so the method seemed to overcome the problem of extracellular DNase production. AFLP differentiated all Clostridium species tested, except for Clostridium ramosum and Clostridium limosum, which clustered together with a 45% similarity level. Other Clostridium species were divided into species-specific clusters or occupied separate positions. Wide genetic diversity was observed among Clostridium botulinum strains, which were divided into seven species-specific clusters. The same AFLP protocol was also suitable for typing C. perfringens at the strain level. A total of 29 different AFLP types were identified for 37 strains of C. perfringens; strains initially originating from the same isolate showed identical fingerprinting patterns and were distinguished from unrelated strains. AFLP proved to be a highly reproducible, easy-to-perform, and relatively fast method which enables high throughput of samples and can serve in the generation of identification libraries. These results indicate that the AFLP method provides a promising tool for the identification and characterization of Clostridium species. PMID:16971642

  20. Bifidobacterium, Bacteroides, and Clostridium spp. in fecal samples from breast-fed and bottle-fed infants with and without iron supplement.

    PubMed Central

    Mevissen-Verhage, E A; Marcelis, J H; de Vos, M N; Harmsen-van Amerongen, W C; Verhoef, J

    1987-01-01

    Bifidobacterium, Bacteroides, and Clostridium spp. isolated from the feces of 23 neonates during the first 3 months of life were identified. Of the 23 neonates, 10 were breast fed, 6 received an infant formula with iron supplement (5 mg/liter), and 7 received the formula without iron supplement (iron concentration, less than 0.5 mg/liter). The Bifidobacterium spp. most frequently isolated from the three groups of infants were B. longum, B. breve, B. adolescentis, and B. bifidum. The bacteroides spp. most frequently isolated were B. fragilis and B. vulgatus. The most common Clostridium sp. in the three groups of infants was C. perfringens. The type of milk did not select for species of Bifidobacterium, Bacteroides, or Clostridium, except for Clostridium butyricum, which was isolated significantly more often from bottle-fed infants with iron supplement than from the other groups, and Clostridium tertium, which was more often isolated from breast-fed infants. The species of the three anaerobic genera did not persist for a long period of time in the three groups of infants. PMID:3818925

  1. Anaerobic and aerobic degradation of cyanophycin by the denitrifying bacterium Pseudomonas alcaligenes strain DIP1 and role of three other coisolates in a mixed bacterial consortium.

    PubMed

    Sallam, Ahmed; Steinbüchel, Alexander

    2008-06-01

    Four bacterial strains were isolated from a cyanophycin granule polypeptide (CGP)-degrading anaerobic consortium, identified by 16S rRNA gene sequencing, and assigned to species of the genera Pseudomonas, Enterococcus, Clostridium, and Paenibacillus. The consortium member responsible for CGP degradation was assigned as Pseudomonas alcaligenes strain DIP1. The growth of and CGP degradation by strain DIP1 under anaerobic conditions were enhanced but not dependent on the presence of nitrate as an electron acceptor. CGP was hydrolyzed to its constituting beta-Asp-Arg dipeptides, which were then completely utilized within 25 and 4 days under anaerobic and aerobic conditions, respectively. The end products of CGP degradation by strain DIP1 were alanine, succinate, and ornithine as determined by high-performance liquid chromatography analysis. The facultative anaerobic Enterococcus casseliflavus strain ELS3 and the strictly anaerobic Clostridium sulfidogenes strain SGB2 were coisolates and utilized the beta-linked isodipeptides from the common pool available to the mixed consortium, while the fourth isolate, Paenibacillus odorifer strain PNF4, did not play a direct role in the biodegradation of CGP. Several syntrophic interactions affecting CGP degradation, such as substrate utilization, the reduction of electron acceptors, and aeration, were elucidated. This study demonstrates the first investigation of CGP degradation under both anaerobic and aerobic conditions by one bacterial strain, with regard to the physiological role of other bacteria in a mixed consortium.

  2. Functional analysis of an feoB mutant in Clostridium perfringens strain 13.

    PubMed

    Awad, Milena M; Cheung, Jackie K; Tan, Joanne E; McEwan, Alastair G; Lyras, Dena; Rood, Julian I

    2016-10-01

    Bacterial pathogens have adopted numerous mechanisms for acquiring iron from host proteins during an infection, including the direct acquisition of ferric iron from heme-associated proteins or from iron-scavenging siderophores. Ferric iron then is transported into the cytosol, where it can be utilized by the bacterial pathogen. Under anaerobic conditions bacteria can also transport ferrous iron using the transmembrane complex FeoAB, but little is known about iron transport systems in anaerobic bacteria such as the pathogenic clostridia. In this study we sought to characterize the iron acquisition process in Clostridium perfringens. Bioinformatic analysis of the Clostridium perfringens strain 13 genome sequence revealed that it has seven potential iron acquisition systems: three siderophore-mediated systems, one ferric citrate uptake system, two heme-associated acquisition systems and one ferrous iron uptake system (FeoAB). The relative level of expression of these systems was determined using quantitative real-time RT-PCR assays that were specific for one gene from each system. Each of these genes was expressed, with the feoAB genes generating the most abundant iron-uptake related transcripts. To further examine the role of this system in the growth of C. perfringens, insertional inactivation was used to isolate a chromosomal feoB mutant. Growth of this mutant in the presence and absence of iron revealed that it had altered growth properties and a markedly reduced total iron and manganese content compared to the wild type; effects that were reversed upon complementation with the wild-type feoB gene. These studies suggest that under anaerobic conditions FeoB is the major protein required for the uptake of iron into the cell and that it may play an important role in the pathogenesis of C. perfringens infections.

  3. Occurrence and molecular characterization of cultivable mesophilic and thermophilic obligate anaerobic bacteria isolated from paper mills.

    PubMed

    Suihko, Maija-Liisa; Partanen, Laila; Mattila-Sandholm, Tiina; Raaska, Laura

    2005-08-01

    The aim of this work was to characterize the cultivable obligate anaerobic bacterial population in paper mill environments. A total of 177 anaerobically grown bacterial isolates were screened for aerotolerance, from which 67 obligate anaerobes were characterized by automated ribotyping and 41 were further identified by partial 16S rDNA sequencing. The mesophilic isolates indicated 11 different taxa (species) within the genus Clostridium and the thermophilic isolates four taxa within the genus Thermoanaerobacterium and one within Thermoanaerobacter (both formerly Clostridium). The most widespread mesophilic bacterium was closely related to C. magnum and occurred in three of four mills. One mill was contaminated with a novel mesophilic bacterium most closely related to C. thiosulfatireducens. The most common thermophile was T. thermosaccharolyticum, occurring in all four mills. The genetic relationships of the mill isolates to described species indicated that most of them are potential members of new species. On the basis of identical ribotypes clay could be identified to be the contamination source of thermophilic bacteria. Automated ribotyping can be a useful tool for the identification of clostridia as soon as comprehensive identification libraries are available.

  4. Spore Coat Architecture of Clostridium novyi-NT spores

    SciTech Connect

    Plomp, M; McCafferey, J; Cheong, I; Huang, X; Bettegowda, C; Kinzler, K; Zhou, S; Vogelstein, B; Malkin, A

    2007-05-07

    Spores of the anaerobic bacterium Clostridium novyi-NT are able to germinate in and destroy hypoxic regions of tumors in experimental animals. Future progress in this area will benefit from a better understanding of the germination and outgrowth processes that are essential for the tumorilytic properties of these spores. Towards this end, we have used both transmission electron microscopy and atomic force microscopy to determine the structure of dormant as well as germinating spores. We found that the spores are surrounded by an amorphous layer intertwined with honeycomb parasporal layers. Moreover, the spore coat layers had apparently self-assembled and this assembly was likely to be governed by crystal growth principles. During germination and outgrowth, the honeycomb layers as well as the underlying spore coat and undercoat layers sequentially dissolved until the vegetative cell was released. In addition to their implications for understanding the biology of C. novyi-NT, these studies document the presence of proteinaceous growth spirals in a biological organism.

  5. Development of Photodynamic Antimicrobial Chemotherapy (PACT) for Clostridium difficile

    PubMed Central

    Pye, Hayley; Kohoutova, Darina; Mosse, Charles A.; Yahioglu, Gokhan; Stamati, Ioanna; Deonarain, Mahendra; Battah, Sinan; Ready, Derren; Allan, Elaine; Mullany, Peter; Lovat, Laurence B.

    2015-01-01

    Background Clostridium difficile is the leading cause of antibiotic-associated diarrhoea and pseudo membranous colitis in the developed world. The aim of this study was to explore whether Photodynamic Antimicrobial Chemotherapy (PACT) could be used as a novel approach to treating C. difficile infections. Methods PACT utilises the ability of light-activated photosensitisers (PS) to produce reactive oxygen species (ROS) such as free radical species and singlet oxygen, which are lethal to cells. We screened thirteen PS against C. difficile planktonic cells, biofilm and germinating spores in vitro, and cytotoxicity of effective compounds was tested on the colorectal adenocarcinoma cell-line HT-29. Results Three PS were able to kill 99.9% of bacteria in both aerobic and anaerobic conditions, both in the planktonic state and in a biofilm, after exposure to red laser light (0.2 J/cm2) without harming model colon cells. The applicability of PACT to eradicate C. difficile germinative spores indirectly was also shown, by first inducing germination with the bile salt taurocholate, followed by PACT. Conclusion This innovative and simple approach offers the prospect of a new antimicrobial therapy using light to treat C. difficile infection of the colon. PMID:26313448

  6. Using a Novel Lysin To Help Control Clostridium difficile Infections

    PubMed Central

    Wang, Qiong; Euler, Chad W.; Delaune, Aurelia

    2015-01-01

    As a consequence of excessive antibiotic therapies in hospitalized patients, Clostridium difficile, a Gram-positive anaerobic spore-forming intestinal pathogen, is the leading cause of hospital-acquired diarrhea and colitis. Drug treatments for these diseases are often complicated by antibiotic-resistant strains and a high frequency of treatment failures and relapse; therefore, novel nonantibiotic approaches may prove to be more effective. In this study, we recombinantly expressed a prophage lysin identified from a C. difficile strain, CD630, which we named PlyCD. PlyCD was found to have lytic activity against specific C. difficile strains. However, the recombinantly expressed catalytic domain of this protein, PlyCD1–174, displayed significantly greater lytic activity (>4-log kill) and a broader lytic spectrum against C. difficile strains while still retaining a high degree of specificity toward C. difficile versus commensal clostridia and other bacterial species. Our data also indicated that noneffective doses of vancomycin and PlyCD1–174 when combined in vitro could be significantly more bactericidal against C. difficile. In an ex vivo treatment model of mouse colon infection, we found that PlyCD1–174 functioned in the presence of intestinal contents, significantly decreasing colonizing C. difficile compared to controls. Together, these data suggest that PlyCD1–174 has potential as a novel therapeutic for clinical application against C. difficile infection, either alone or in combination with other preexisting treatments to improve their efficacy. PMID:26392484

  7. Current knowledge on the laboratory diagnosis of Clostridium difficile infection.

    PubMed

    Martínez-Meléndez, Adrián; Camacho-Ortiz, Adrián; Morfin-Otero, Rayo; Maldonado-Garza, Héctor Jesús; Villarreal-Treviño, Licet; Garza-González, Elvira

    2017-03-07

    Clostridium difficile (C. difficile) is a spore-forming, toxin-producing, gram-positive anaerobic bacterium that is the principal etiologic agent of antibiotic-associated diarrhea. Infection with C. difficile (CDI) is characterized by diarrhea in clinical syndromes that vary from self-limited to mild or severe. Since its initial recognition as the causative agent of pseudomembranous colitis, C. difficile has spread around the world. CDI is one of the most common healthcare-associated infections and a significant cause of morbidity and mortality among older adult hospitalized patients. Due to extensive antibiotic usage, the number of CDIs has increased. Diagnosis of CDI is often difficult and has a substantial impact on the management of patients with the disease, mainly with regards to antibiotic management. The diagnosis of CDI is primarily based on the clinical signs and symptoms and is only confirmed by laboratory testing. Despite the high burden of CDI and the increasing interest in the disease, episodes of CDI are often misdiagnosed. The reasons for misdiagnosis are the lack of clinical suspicion or the use of inappropriate tests. The proper diagnosis of CDI reduces transmission, prevents inadequate or unnecessary treatments, and assures best antibiotic treatment. We review the options for the laboratory diagnosis of CDI within the settings of the most accepted guidelines for CDI diagnosis, treatment, and prevention of CDI.

  8. Metabolism of lactose by Clostridium thermolacticum growing in continuous culture.

    PubMed

    Collet, Christophe; Girbal, Laurence; Péringer, Paul; Schwitzguébel, Jean-Paul; Soucaille, Philippe

    2006-06-01

    The objective of the present study was to characterize the metabolism of Clostridium thermolacticum, a thermophilic anaerobic bacterium, growing continuously on lactose (10 g l(-1)) and to determine the enzymes involved in the pathways leading to the formation of the fermentation products. Biomass and metabolites concentration were measured at steady-state for different dilution rates, from 0.013 to 0.19 h(-1). Acetate, ethanol, hydrogen and carbon dioxide were produced at all dilution rates, whereas lactate was detected only for dilution rates below 0.06 h(-1). The presence of several key enzymes involved in lactose metabolism, including beta-galactosidase, glyceraldehyde-3-phosphate dehydrogenase, pyruvate:ferredoxin oxidoreductase, acetate kinase, ethanol dehydrogenase and lactate dehydrogenase, was demonstrated. Finally, the intracellular level of NADH, NAD+, ATP and ADP was also measured for different dilution rates. The production of ethanol and lactate appeared to be linked with the re-oxidation of NADH produced during glycolysis, whereas hydrogen produced should come from reduced ferredoxin generated during pyruvate decarboxylation. To produce more hydrogen or more acetate from lactose, it thus appears that an efficient H2 removal system should be used, based on a physical (membrane) or a biological approach, respectively, by cultivating C. thermolacticum with efficient H2 scavenging and acetate producing microorganisms.

  9. Hydrolysis of synthetic polyesters by Clostridium botulinum esterases.

    PubMed

    Perz, Veronika; Baumschlager, Armin; Bleymaier, Klaus; Zitzenbacher, Sabine; Hromic, Altijana; Steinkellner, Georg; Pairitsch, Andris; Łyskowski, Andrzej; Gruber, Karl; Sinkel, Carsten; Küper, Ulf; Ribitsch, Doris; Guebitz, Georg M

    2016-05-01

    Two novel esterases from the anaerobe Clostridium botulinum ATCC 3502 (Cbotu_EstA and Cbotu_EstB) were expressed in Escherichia coli BL21-Gold(DE3) and were found to hydrolyze the polyester poly(butylene adipate-co-butylene terephthalate) (PBAT). The active site residues (triad Ser, Asp, His) are present in both enzymes at the same location only with some amino acid variations near the active site at the surrounding of aspartate. Yet, Cbotu_EstA showed higher kcat values on para-nitrophenyl butyrate and para-nitrophenyl acetate and was considerably more active (sixfold) on PBAT. The entrance to the active site of the modeled Cbotu_EstB appears more narrowed compared to the crystal structure of Cbotu_EstA and the N-terminus is shorter which could explain its lower activity on PBAT. The Cbotu_EstA crystal structure consists of two regions that may act as movable cap domains and a zinc metal binding site.

  10. Immunogenicity and protective efficacy of Clostridium difficile spore proteins.

    PubMed

    Ghose, Chandrabali; Eugenis, Ioannis; Edwards, Adrianne N; Sun, Xingmin; McBride, Shonna M; Ho, David D

    2016-02-01

    Clostridium difficile is a spore-forming, anaerobic, Gram-positive organism that is the leading cause of antibiotic-associated infectious diarrhea, commonly known as C. difficile infection (CDI). C. difficile spores play an important role in the pathogenesis of CDI. Spore proteins, especially those that are surface-bound may play an essential role in the germination, colonization and persistence of C. difficile in the human gut. In our current study, we report the identification of two surface-bound spore proteins, CdeC and CdeM that may be utilized as immunization candidates against C. difficile. These spore proteins are immunogenic in mice and are able to protect mice against challenge with C. difficile UK1, a clinically-relevant 027/B1/NAP1 strain. These spore proteins are also able to afford high levels of protection against challenge with C. difficile 630Δerm in golden Syrian hamsters. This unprecedented study shows the vaccination potential of C. difficile spore exosporium proteins.

  11. Current knowledge on the laboratory diagnosis of Clostridium difficile infection

    PubMed Central

    Martínez-Meléndez, Adrián; Camacho-Ortiz, Adrián; Morfin-Otero, Rayo; Maldonado-Garza, Héctor Jesús; Villarreal-Treviño, Licet; Garza-González, Elvira

    2017-01-01

    Clostridium difficile (C. difficile) is a spore-forming, toxin-producing, gram-positive anaerobic bacterium that is the principal etiologic agent of antibiotic-associated diarrhea. Infection with C. difficile (CDI) is characterized by diarrhea in clinical syndromes that vary from self-limited to mild or severe. Since its initial recognition as the causative agent of pseudomembranous colitis, C. difficile has spread around the world. CDI is one of the most common healthcare-associated infections and a significant cause of morbidity and mortality among older adult hospitalized patients. Due to extensive antibiotic usage, the number of CDIs has increased. Diagnosis of CDI is often difficult and has a substantial impact on the management of patients with the disease, mainly with regards to antibiotic management. The diagnosis of CDI is primarily based on the clinical signs and symptoms and is only confirmed by laboratory testing. Despite the high burden of CDI and the increasing interest in the disease, episodes of CDI are often misdiagnosed. The reasons for misdiagnosis are the lack of clinical suspicion or the use of inappropriate tests. The proper diagnosis of CDI reduces transmission, prevents inadequate or unnecessary treatments, and assures best antibiotic treatment. We review the options for the laboratory diagnosis of CDI within the settings of the most accepted guidelines for CDI diagnosis, treatment, and prevention of CDI. PMID:28321156

  12. Anaerobic digestion process

    SciTech Connect

    Ishida, M.; Haga, R.; Odawara, Y.

    1982-10-19

    An algae culture grown on the water from the digested slurry of a biogasification plant serves as a means of removing CO/sub 2/ from the methane stream while purifying the wastewater and providing more biomass for the anaerobic digestion plant. Tested on a sewage-sludge digestion system, the proposed process improved the methane yield by 32% and methane concentration by 53-98 vol % while lowering the concentration of nitrogen and phosphorus in the final water.

  13. Adjuvants for Clostridium tetani and Clostridium diphtheriae vaccines updating.

    PubMed

    Alshanqiti, Fatimah M; Al-Masaudi, Saad B; Al-Hejin, Ahmed M; Redwan, Elrashdy M

    2017-01-01

    It's known that diphtheria and tetanus are a contagious lethal diseases over the years, they caused by pathogenic microbes corynebacterium diphtheria and Clostridium tetani, respectively. The diseases result from the production of bacterial toxin. Vaccination with bacterial toxoid vaccines adsorbed on particulates adjuvants still are the best way to prevent this epidemic diseases from spread. The particulate vaccines have been shown to be more efficient than soluble one for the induction of the immune responses. Nanoparticles can be engineered to enhance the immune responses. As well known the immune response to inactivate killed and subunit vaccine enhances by alum adjuvants. The adjuvants examined and tested after reducing its size to particle size, thus mimic size of viruses which is considered smallest units can derive the immune system. The major issue is minimizing the adjuvant particles, to gain insight of resulting immunity types and impact on immune response. The adjuvant effect of micro/nanoparticles appears to largely be a consequence of their uptake into antigen presenting cells.

  14. Early anaerobic metabolisms

    PubMed Central

    Canfield, Don E; Rosing, Minik T; Bjerrum, Christian

    2006-01-01

    Before the advent of oxygenic photosynthesis, the biosphere was driven by anaerobic metabolisms. We catalogue and quantify the source strengths of the most probable electron donors and electron acceptors that would have been available to fuel early-Earth ecosystems. The most active ecosystems were probably driven by the cycling of H2 and Fe2+ through primary production conducted by anoxygenic phototrophs. Interesting and dynamic ecosystems would have also been driven by the microbial cycling of sulphur and nitrogen species, but their activity levels were probably not so great. Despite the diversity of potential early ecosystems, rates of primary production in the early-Earth anaerobic biosphere were probably well below those rates observed in the marine environment. We shift our attention to the Earth environment at 3.8 Gyr ago, where the earliest marine sediments are preserved. We calculate, consistent with the carbon isotope record and other considerations of the carbon cycle, that marine rates of primary production at this time were probably an order of magnitude (or more) less than today. We conclude that the flux of reduced species to the Earth surface at this time may have been sufficient to drive anaerobic ecosystems of sufficient activity to be consistent with the carbon isotope record. Conversely, an ecosystem based on oxygenic photosynthesis was also possible with complete removal of the oxygen by reaction with reduced species from the mantle. PMID:17008221

  15. Display of Clostridium cellulovorans xylose isomerase on the cell surface of Saccharomyces cerevisiae and its direct application to xylose fermentation.

    PubMed

    Ota, Miki; Sakuragi, Hiroshi; Morisaka, Hironobu; Kuroda, Kouichi; Miyake, Hideo; Tamaru, Yutaka; Ueda, Mitsuyoshi

    2013-01-01

    Xylose isomerase (XI) is a key enzyme in the conversion of D-xylose, which is a major component of lignocellulosic biomass, to D-xylulose. Genomic analysis of the bacterium Clostridium cellulovorans revealed the presence of XI-related genes. In this study, XI derived from C. cellulovorans was produced and displayed using the yeast cell-surface display system, and the xylose assimilation and fermentation properties of this XI-displaying yeast were examined. XI-displaying yeast grew well in medium containing xylose as the sole carbon source and directly produced ethanol from xylose under anaerobic conditions.

  16. Consumption of freons CFC-11 and CFC-12 by anaerobic sediments and soils

    USGS Publications Warehouse

    Lovley, D.R.; Woodward, J.C.

    1992-01-01

    A variety of anaerobic sediments and soils consumed CFC-11 (CFCl3) and CFC-12 (CF2Cl2). An aerobic soil did not. Active microbial metabolism was required for CFC-12 uptake in all of the sediments examined. CFC-11 uptake was faster in the presence of microbial activity, but reduced components in the sediments also resulted in nonenzymatic CFC-11 consumption in most instances. CFC-12 uptake in a culture of Clostridium pasteurianum provided a model for the sediment uptake of CFC-11 and CFC-12 that required active microbial metabolism. Consumption of CFC-11 in the presence of reduced hematin demonstrated a potential mechanism for nonenzymatic CFC-11 consumption. These findings demonstrate that CFC-11 and CFC-12 are not biochemically inert under anaerobic conditions. This suggests that anaerobic degradation of CFC-11 and CFC-12 in anaerobic landfills might prevent some disposed CFC-11 and CFC-12 from entering the atmosphere. The results also suggest that CFC-11 and CFC-12 cannot be used as stable tracers in anaerobic environments. Furthermore, although the microbial sink for atmospheric CFC-11 and CFC-12 is much less than current anthropogenic release, this sink could have a significant long-term effect on the amount of CFC-11 and CFC-12 reaching the stratosphere.

  17. Simple Method for Culturing Anaerobes

    PubMed Central

    Davis, C. E.; Hunter, W. J.; Ryan, J. L.; Braude, A. I.

    1973-01-01

    A simple, effective method is needed for growing obligate anaerobes in the clinical laboratory. This report describes a pre-reduced anaerobic bottle that can be taken to the bedside for direct inoculation, provides a flat agar surface for evaluation of number and morphology of colonies, and can be incubated in conventional bacteriological incubators. Each anaerobic culture set consisted of two bottles containing brain heart infusion agar and CO2. Gentamicin sulfate (50 μg/ml) was added to one of these to inhibit facultative enteric bacilli. Comparison of the anaerobic bottles with an identical aerobic bottle which was also routinely inoculated permitted early identification of anaerobic colonies. Representative species of most anaerobic genera of proven pathogenicity for man have been isolated from this system during 10 months of routine use. Images PMID:4571657

  18. Clostridium difficile infection in Thailand.

    PubMed

    Putsathit, Papanin; Kiratisin, Pattarachai; Ngamwongsatit, Puriya; Riley, Thomas V

    2015-01-01

    Clostridium difficile is the aetiological agent in ca. 20% of cases of antimicrobial-associated diarrhoea in hospitalised adults. Diseases caused by this organism range from mild diarrhoea to occasional fatal pseudomembranous colitis. The epidemiology of C. difficile infection (CDI) has changed notably in the past decade, following epidemics in the early 2000s of PCR ribotype (RT) 027 infection in North America and Europe, where there was an increase in disease severity and mortality. Another major event has been the emergence of RT 078, initially as the predominant ribotype in production animals in the USA and Europe, and then in humans in Europe. Although there have been numerous investigations of the epidemiology of CDI in North America and Europe, limited studies have been undertaken elsewhere, particularly in Asia. Antimicrobial exposure remains the major risk factor for CDI. Given the high prevalence of indiscriminate and inappropriate use of antimicrobials in Asia, it is conceivable that CDI is relatively common among humans and animals. This review describes the level of knowledge in Thailand regarding C. difficile detection methods, prevalence and antimicrobial susceptibility profile, as well as the clinical features of, treatment options for and outcomes of the disease. In addition, antimicrobial usage in livestock in Thailand will be reviewed. A literature search yielded 18 studies mentioning C. difficile in Thailand, a greater number than from any other Asian country. It is possible that the situation in Thailand in relation to CDI may mirror the situation in other developing Asians countries.

  19. Thermostable chaperonin from Clostridium thermocellum.

    PubMed

    Cross, S J; Ciruela, A; Poomputsa, K; Romaniec, M P; Freedman, R B

    1996-06-01

    Homologues of the chaperonins Cpn60 and Cpn10 have been purified from the Gram-positive cellulolytic thermophile Clostridium thermocellum. The Cpn60 protein was purified by ATP-affinity chromatography and the Cpn10 protein was purified by gel-filtration, ion-exchange and hydrophobic interaction chromatographies. The identities of the proteins were confirmed by N-terminal sequence analysis and antigenic cross-reactivity. The Cpn60 homologue is a weak, thermostable ATPase (t1/2 at 70 decrees C more than 90 min) with optimum activity (Kcat 0.07 S-1) between 60 degrees C and 70 degrees C. The ATPase activity of the authentic Cpn60 was inhibited by Escherichia coli GroES. The catalytic properties of a recombinant C. thermocellum Cpn60 purified from a GST-Cpn60 fusion protein expressed in E. coli [Ciruela (1995) Ph.D. Thesis, University of Kent] were identical with those of the authentic C. thermocellum Cpn60. Gel-filtration studies show that at room temperature the Cpn60 migrates mainly as a heptamer. Electron microscopy confirms the presence of complexes showing 7-fold rotational symmetry and also reveals a small number of particles that seem to be tetradecamers with a similar structure to E. coli GroEL complexes.

  20. Probiotics in Clostridium difficile Infection

    PubMed Central

    Na, Xi; Kelly, Ciaran

    2017-01-01

    Clostridium difficile infection (CDI) is one of the most prevalent nosocomial infections. A dramatic increase in the incidence and severity of CDI has been noted in the past decade. Current recommendations suggest metronidazole as first-line therapy in mild to moderately severe CDI and oral vancomycin in individuals with severe CDI, or when metronidazole fails or is contradicted. Alterations of the colonic microbiota, usually caused by antimicrobial therapy, seem to play a critical role in CDI pathogenesis. Probiotics are live microorganisms that confer a health benefit to the host, and have been used in CDI. Although a wide variety of probiotics have been studied, the exact role of probiotics in preventing and treating CDI is not clear. In this study, we reviewed the current literature and recommendations on the most commonly studied protiotic agents (Saccharomyces boulardii, Lactobacillus species, and probiotic mixtures) used to prevent or treat CDI. Lactobacillus-containing probiotic mixtures and S. boulardii may be effective in the prevention of CDI in high-risk antibiotic recipients but this finding is based on small, individual studies, and further, larger, well-controlled studies are needed to confirm preliminary positive findings and to better delineate the efficacy of probiotics in CDI prevention or treatment. PMID:21992956

  1. Management of Clostridium difficile Infection

    PubMed Central

    Al-Jashaami, Layth S.

    2016-01-01

    Since the discovery of Clostridium difficile infection (CDI) in the 1970s, there has been an increase in the incidence, severity, and recurrence rate of the disease. We reviewed the recent CDI literature in PubMed published before February 28, 2016 that focused on advances in therapy. Despite a large number of studies describing methods for diagnosing the disease, there is currently no definitive test that identifies this infection with certainty, which complicates therapy. Recommended therapy for CDI includes oral metronidazole for mild cases and oral vancomycin or fidaxomicin for moderate to severe cases, each given for 10 to 14 days. For infection with spore-forming C difficile, this length of treatment may be insufficient to lead to cure; however, continuing antibiotics for longer periods of time may unfavorably alter the microbiome, preventing recovery. Treatment with metronidazole has been associated with an increasing failure rate, and the only clear recommended form of metronidazole for treatment of CDI is the intravenous formulation for patients unable to take oral medications. For vancomycin or fidaxomicin treatment of first CDI recurrences, the drug used in the initial bout can be repeated. For second or future recurrences, vancomycin can be given in pulsed or tapered doses. New modalities of treatment, such as bacteriotherapy and immunotherapy, show promise for the treatment of recurrent CDI. PMID:27917075

  2. Fidaxomicin: in Clostridium difficile infection.

    PubMed

    Duggan, Sean T

    2011-12-24

    Fidaxomicin is a first-in-class macrocyclic antibacterial that primarily demonstrates activity against species of clostridia, predominantly Clostridium difficile, while having limited or no activity against normal faecal microflora. Fidaxomicin is minimally absorbed following oral administration and is excreted almost solely in the faeces. Fidaxomicin displayed a high level of antibacterial activity against C. difficile in vitro, with a minimum inhibitory concentration required to inhibit 90% of C. difficile strains of 0.125-0.5 μg/mL, and was ≈2- to 8-fold more active than vancomycin or metronidazole. Fidaxomicin demonstrated a prolonged postantibiotic effect against C. difficile relative to vancomycin and metronidazole. In two randomized, double-blind, phase III trials, oral fidaxomicin 200 mg every 12 hours for 10 days was no less effective than oral vancomycin 125 mg every 6 hours for 10 days in the treatment of C. difficile infection, based on noninferiority analyses of clinical cure rates (primary endpoint). Fidaxomicin therapy was associated with a significantly lower rate of recurrence, as well as a significantly higher rate of global cure (i.e. sustained clinical response; resolution of diarrhoea without recurrence) compared with vancomycin therapy in the two clinical trials. Fidaxomicin was generally well tolerated in patients with C. difficile infection, with a tolerability profile generally similar to that of vancomycin.

  3. The microbial ecology of anaerobic cellulose degradation in municipal waste landfill sites: evidence of a role for fibrobacters.

    PubMed

    McDonald, James E; Houghton, James N I; Rooks, David J; Allison, Heather E; McCarthy, Alan J

    2012-04-01

    Cellulose is reputedly the most abundant organic polymer in the biosphere, yet despite the fundamental role of cellulolytic microorganisms in global carbon cycling and as potential sources of novel enzymes for biotechnology, their identity and ecology is not well established. Cellulose is a major component of landfill waste and its degradation is therefore a key feature of the anaerobic microbial decomposition process. Here, we targeted a number of taxa containing known cellulolytic anaerobes (members of the bacterial genus Fibrobacter, lineages of Clostridium clusters I, III, IV and XIV, and anaerobic fungi of the Neocallimastigales) in landfill leachate and colonized cellulose 'baits' via PCR and quantitative PCR (qPCR). Fibrobacter spp. and Clostridium clusters III, IV and XIV were detected in almost all leachate samples and cluster III and XIV clostridia were the most abundant (1-6% and 1-17% of total bacterial 16S rRNA gene copies respectively). Two landfill leachate microcosms were constructed to specifically assess those microbial communities that colonize and degrade cellulose substrates in situ. Scanning electron microscopy (SEM) of colonized cotton revealed extensive cellulose degradation in one microcosm, and Fibrobacter spp. and Clostridium cluster III represented 29% and 17%, respectively, of total bacterial 16S rRNA gene copies in the biofilm. Visible cellulose degradation was not observed in the second microcosm, and this correlated with negligible relative abundances of Clostridium cluster III and Fibrobacter spp. (≤ 0.1%), providing the first evidence that the novel fibrobacters recently detected in landfill sites and other non-gut environments colonize and degrade cellulose substrates in situ.

  4. Clostridium thermocellum releases coumaric acid during degradation of untreated grasses by the action of an unknown enzyme.

    PubMed

    Herring, Christopher D; Thorne, Philip G; Lynd, Lee R

    2016-03-01

    Clostridium thermocellum is an anaerobic thermophile with the ability to digest lignocellulosic biomass that has not been pretreated with high temperatures. Thermophilic anaerobes have previously been shown to more readily degrade grasses than wood. Part of the explanation for this may be the presence of relatively large amounts of coumaric acid in grasses, with linkages to both hemicellulose and lignin. We found that C. thermocellum and cell-free cellulase preparations both release coumaric acid from bagasse and switchgrass. Cellulase preparations from a mutant strain lacking the scaffoldin cipA still showed activity, though diminished. Deletion of all three proteins in C. thermocellum with ferulic acid esterase domains, either singly or in combination, did not eliminate the activity. Further work will be needed to identify the novel enzyme(s) responsible for the release of coumaric acid from grasses and to determine whether these enzymes are important factors of microbial biomass degradation.

  5. Prevention of Infection Due to Clostridium difficile.

    PubMed

    Cooper, Christopher C; Jump, Robin L P; Chopra, Teena

    2016-12-01

    Clostridium difficile is one of the foremost nosocomial pathogens. Preventing infection is particularly challenging. Effective prevention efforts typically require a multifaceted bundled approach. A variety of infection control procedures may be advantageous, including strict hand decontamination with soap and water, contact precautions, and using chlorine-containing decontamination agents. Additionally, risk factor reduction can help reduce the burden of disease. The risk factor modification is principally accomplished though antibiotic stewardship programs. Unfortunately, most of the current evidence for prevention is in acute care settings. This review focuses on preventative approaches to reduce the incidence of Clostridium difficile infection in healthcare settings.

  6. ISOLATION OF CLOSTRIDIUM TETANI FROM SOIL.

    PubMed

    SANADA, I; NISHIDA, S

    1965-03-01

    Sanada, Ichiro (Kanazawa University, Kanazawa, Japan), and Shoki Nishida. Isolation of Clostridium tetani from soil. J. Bacteriol. 89:626-629. 1965.-The higher the temperatures applied to soil specimens, the weaker the toxigenicity of Clostridium tetani strains isolated from them. The glucose- and maltose-fermenting ability of these isolates was inversely proportional to their toxigenicity. The biological properties of atoxic strains were indistinguishable from those of C. tetanomorphum. Since a considerable number of toxic strains fermented glucose and maltose, these criteria are of doubtful value for differentiating C. tetani from C. tetanomorphum.

  7. Enhanced coproduction of hydrogen and methane from cornstalks by a three-stage anaerobic fermentation process integrated with alkaline hydrolysis.

    PubMed

    Cheng, Xi-Yu; Liu, Chun-Zhao

    2012-01-01

    A three-stage anaerobic fermentation process including H(2) fermentation I, H(2) fermentation II, methane fermentation was developed for the coproduction of hydrogen and methane from cornstalks. Hydrogen production from cornstalks using direct microbial conversion by Clostridium thermocellum 7072 was markedly enhanced in the two-stage thermophilic hydrogen fermentation process integrated with alkaline treatment. The highest total hydrogen yield from cornstalks in the two-stage fermentation process reached 74.4 mL/g-cornstalk. The hydrogen fermentation effluents and alkaline hydrolyzate were further used for methane fermentation by anaerobic granular sludge, and the total methane yield reached 205.8 mL/g-cornstalk. The total energy recovery in the three-stage anaerobic fermentation process integrated with alkaline hydrolysis reached 70.0%.

  8. Anaerobic wastewater treatment using anaerobic baffled bioreactor: a review

    NASA Astrophysics Data System (ADS)

    Hassan, Siti Roshayu; Dahlan, Irvan

    2013-09-01

    Anaerobic wastewater treatment is receiving renewed interest because it offers a means to treat wastewater with lower energy investment. Because the microorganisms involved grow more slowly, such systems require clever design so that the microbes have sufficient time with the substrate to complete treatment without requiring enormous reactor volumes. The anaerobic baffled reactor has inherent advantages over single compartment reactors due to its circulation pattern that approaches a plug flow reactor. The physical configuration of the anaerobic baffled reactor enables significant modifications to be made; resulting in a reactor which is proficient of treating complex wastewaters which presently require only one unit, ultimately significant reducing capital costs. This paper also concerns about mechanism, kinetic and hydrodynamic studies of anaerobic digestion for future application of the anaerobic baffled reactor for wastewater treatment.

  9. Isolation, Culture Characteristics, and Identification of Anaerobic Bacteria from the Chicken Cecum

    PubMed Central

    Salanitro, J. P.; Fairchilds, I. G.; Zgornicki, Y. D.

    1974-01-01

    Studies on the anaerobic cecal microflora of the 5-week-old chicken were made to determine a suitable roll-tube medium for enumeration and isolation of the bacterial population, to determine effects of medium components on recovery of total anaerobes, and to identify the predominant bacterial groups. The total number of microorganisms in cecal contents determined by direct microscope cell counts varied (among six samples) from 3.83 × 1010 to 7.64 × 1010 per g. Comparison of different nonselective media indicated that 60% of the direct microscope count could be recovered with a rumen fluid medium (M98-5) and 45% with medium 10. Deletion of rumen fluid from M98-5 reduced the total anaerobic count by half. Colony counts were lower if chicken cecal extract was substituted for rumen fluid in M98-5. Supplementing medium 10 with liver, chicken fecal, or cecal extracts improved recovery of anaerobes slightly. Prereduced blood agar media were inferior to M98-5. At least 11 groups of bacteria were isolated from high dilutions (10-9) of cecal material. Data on morphology and physiological and fermentation characteristics of 90% of the 298 isolated strains indicated that these bacteria represented species of anaerobic gram-negative cocci, facultatively anaerobic cocci and streptococci, Peptostreptococcus, Propionibacterium, Eubacterium, Bacteroides, and Clostridium. The growth of many of these strains was enhanced by rumen fluid, yeast extract, and cecal extract additions to basal media. These studies indicate that some of the more numerous anaerobic bacteria present in chicken cecal digesta can be isolated and cultured when media and methods that have been developed for ruminal bacteria are employed. PMID:4596749

  10. Identification and antimicrobial susceptibility of obligate anaerobic bacteria from clinical samples of animal origin.

    PubMed

    Mayorga, Melissa; Rodríguez-Cavallini, Evelyn; López-Ureña, Diana; Barquero-Calvo, Elías; Quesada-Gómez, Carlos

    2015-12-01

    The etiology of veterinary infectious diseases has been the focus of considerable research, yet relatively little is known about the causative agents of anaerobic infections. Susceptibility studies have documented the emergence of antimicrobial resistance and indicate distinct differences in resistance patterns related to veterinary hospitals, geographic regions, and antibiotic-prescribing regimens. The aim of the present study was to identify the obligate anaerobic bacteria from veterinary clinical samples and to determinate the in vitro susceptibility to eight antimicrobials and their resistance-associated genes. 81 clinical specimens obtained from food-producing animals, pets and wild animals were examined to determine the relative prevalence of obligate anaerobic bacteria, and the species represented. Bacteroides spp, Prevotella spp and Clostridium spp represented approximately 80% of all anaerobic isolates. Resistance to metronidazole, clindamycin, tetracycline and fluoroquinolones was found in strains isolated from food-producing animals. Ciprofloxacin, enrofloxacin and cephalotin showed the highest resistance in all isolates. In 17%, 4% and 14% of tetracycline-resistant isolates, the resistance genes tetL, tetM and tetW were respectively amplified by PCR whereas in 4% of clindamycin-resistant strains the ermG gene was detected. 26% of the isolates were positive for cepA, while only 6% harbored the cfxA (resistance-conferring genes to beta-lactams). In this study, the obligate anaerobic bacteria from Costa Rica showed a high degree of resistance to most antimicrobials tested. Nevertheless, in the majority of cases this resistance was not related to the resistance acquired genes usually described in anaerobes. It is important to address and regulate the use of antimicrobials in the agricultural industry and the empirical therapy in anaerobic bacterial infections in veterinary medicine, especially since antibiotics and resistant bacteria can persist in the

  11. EGA Protects Mammalian Cells from Clostridium difficile CDT, Clostridium perfringens Iota Toxin and Clostridium botulinum C2 Toxin.

    PubMed

    Schnell, Leonie; Mittler, Ann-Katrin; Sadi, Mirko; Popoff, Michel R; Schwan, Carsten; Aktories, Klaus; Mattarei, Andrea; Azarnia Tehran, Domenico; Montecucco, Cesare; Barth, Holger

    2016-04-01

    The pathogenic bacteria Clostridium difficile, Clostridium perfringens and Clostridium botulinum produce the binary actin ADP-ribosylating toxins CDT, iota and C2, respectively. These toxins are composed of a transport component (B) and a separate enzyme component (A). When both components assemble on the surface of mammalian target cells, the B components mediate the entry of the A components via endosomes into the cytosol. Here, the A components ADP-ribosylate G-actin, resulting in depolymerization of F-actin, cell-rounding and eventually death. In the present study, we demonstrate that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone (EGA), a compound that protects cells from multiple toxins and viruses, also protects different mammalian epithelial cells from all three binary actin ADP-ribosylating toxins. In contrast, EGA did not inhibit the intoxication of cells with Clostridium difficile toxins A and B, indicating a possible different entry route for this toxin. EGA does not affect either the binding of the C2 toxin to the cells surface or the enzyme activity of the A components of CDT, iota and C2, suggesting that this compound interferes with cellular uptake of the toxins. Moreover, for C2 toxin, we demonstrated that EGA inhibits the pH-dependent transport of the A component across cell membranes. EGA is not cytotoxic, and therefore, we propose it as a lead compound for the development of novel pharmacological inhibitors against clostridial binary actin ADP-ribosylating toxins.

  12. Identification of Clostridium botulinum, Clostridium argentinense, and related organisms by cellular fatty acid analysis.

    PubMed Central

    Ghanem, F M; Ridpath, A C; Moore, W E; Moore, L V

    1991-01-01

    On the basis of 686 analyses of 285 strains of Clostridium botulinum, Clostridium argentinense (formerly C. botulinum type G), and phenotypically related organisms, 14 cellular fatty acid (CFA) groups of toxic organisms and 6 CFA groups of nontoxic organisms were delineated. The CFA groups of toxic strains included two of type A, three of proteolytic strains of type B, two of proteolytic strains of type F, one each of nonproteolytic strains of types B, E, and F, and one each of types C alpha, C beta, and D and C. argentinense. The groups of phenotypically similar nontoxic strains included Clostridium sporogenes, Clostridium putrificum, nontoxic strains with phenotypic characteristics similar to those of nonproteolytic strains of C. botulinum types B, E, and F (BEF-like), two groups of nontoxigenic organisms with phenotypic characteristics similar to those of C. botulinum types C and D and Clostridium novyi (CDN-like), and Clostridium subterminale, which has phenotypic characteristics similar to those of C. argentinense. Within the toxin types, 89 to 100% of the strains were correctly identified by CFA analysis, and 74 to 100% of the analyses were correct. Of 36 strains of C. sporogenes, 30 (83%) were correctly identified; 17% of the strains of C. sporogenes were incorrectly identified as C. botulinum type A or B. All analyses of C. putrificum and C. subterminale were correctly identified. There was no significant level of similarity between strains of C. botulinum and phenotypically similar organisms and 85 other species of clostridia or 407 other taxa of gram-positive and gram-negative bacteria. Additionally, the one strain each of Clostridium baratii and Clostridium butyricum previously reported to produce C. botulinum toxin could be differentiated from C.botulinum types as well as from strains of C. baratii and C. butyricum that did not produce neurotoxin. PMID:1864927

  13. Phylogenetic positions of Clostridium chauvoei and Clostridium septicum based on 16S rRNA gene sequences.

    PubMed

    Kuhnert, P; Capaul, S E; Nicolet, J; Frey, J

    1996-10-01

    The sequences of the 16S rRNA genes (rrs genes) of Clostridium chauvoei, the causative agent of blackleg in cattle, and the phenotypically related organism Clostridium septicum were determined. After amplification of 1,507-bp PCR fragments from the corresponding rrs genes, the sequences were determined in a single round of sequencing by using conserved region primers. A sequence similarity analysis of the sequences revealed the close phylogenetic relationship of C. chauvoei and C. septicum in Clostridium cluster I (M. D. Collins, P. A. Lawson, A. Willems, J. J. Cordoba, J. Fernandez-Garayzabal, P. Garcia, J. Cai, H. Hippe, and J. A. E. Farrow, Int. J. Syst. Bacteriol. 44:812-826, 1994), which includes Clostridium carnis, Clostridium perfringens, Clostridium botulinum, and Clostridium tetani. We found that 99.3% of the nucleotides in the genes of C. chauvoei and C. septicum are identical.

  14. Clostridium difficile phages: still difficult?

    PubMed Central

    Hargreaves, Katherine R.; Clokie, Martha R. J.

    2014-01-01

    Phages that infect Clostridium difficile were first isolated for typing purposes in the 1980s, but their use was short lived. However, the rise of C. difficile epidemics over the last decade has triggered a resurgence of interest in using phages to combat this pathogen. Phage therapy is an attractive treatment option for C. difficile infection, however, developing suitable phages is challenging. In this review we summarize the difficulties faced by researchers in this field, and we discuss the solutions and strategies used for the development of C. difficile phages for use as novel therapeutics. Epidemiological data has highlighted the diversity and distribution of C. difficile, and shown that novel strains continue to emerge in clinical settings. In parallel with epidemiological studies, advances in molecular biology have bolstered our understanding of C. difficile biology, and our knowledge of phage–host interactions in other bacterial species. These three fields of biology have therefore paved the way for future work on C. difficile phages to progress and develop. Benefits of using C. difficile phages as therapeutic agents include the fact that they have highly specific interactions with their bacterial hosts. Studies also show that they can reduce bacterial numbers in both in vitro and in vivo systems. Genetic analysis has revealed the genomic diversity among these phages and provided an insight into their taxonomy and evolution. No strictly virulent C. difficile phages have been reported and this contributes to the difficulties with their therapeutic exploitation. Although treatment approaches using the phage-encoded endolysin protein have been explored, the benefits of using “whole-phages” are such that they remain a major research focus. Whilst we don’t envisage working with C. difficile phages will be problem-free, sufficient study should inform future strategies to facilitate their development to combat this problematic pathogen. PMID:24808893

  15. Economic viability of anaerobic digestion

    SciTech Connect

    Wellinger, A.

    1996-01-01

    The industrial application of anaerobic digestion is a relatively new, yet proven waste treatment technology. Anaerobic digestion reduces and upgrades organic waste, and is a good way to control air pollution as it reduces methane and nitrous gas emissions. For environmental and energy considerations, anaerobic digestion is a nearly perfect waste treatment process. However, its economic viability is still in question. A number of parameters - type of waste (solid or liquid), digester system, facility size, product quality and end use, environmental requirements, cost of alternative treatments (including labor), and interest rates - define the investment and operating costs of an anaerobic digestion facility. Therefore, identical facilities that treat the same amount and type of waste may, depending on location, legislation, and end product characteristics, reveal radically different costs. A good approach for evaluating the economics of anaerobic digestion is to compare it to treatment techniques such as aeration or conventional sewage treatment (for industrial wastewater), or composting and incineration (for solid organic waste). For example, the cost (per ton of waste) of in-vessel composting with biofilters is somewhat higher than that of anaerobic digestion, but the investment costs 1 1/2 to 2 times more than either composting or anaerobic digestion. Two distinct advantages of anaerobic digestion are: (1) it requires less land than either composting or incinerating, which translates into lower costs and milder environmental and community impacts (especially in densely populated areas); and (2) it produces net energy, which can be used to operate the facility or sold to nearby industries.

  16. Key microbial communities steering the functioning of anaerobic digesters during hydraulic and organic overloading shocks.

    PubMed

    Regueiro, Leticia; Lema, Juan M; Carballa, Marta

    2015-12-01

    Overloading is one of the most typical process disturbance in anaerobic digesters, resulting in volatile fatty acids (VFAs) accumulation. This work aimed to study the microbial community dynamics during hydraulic (decreasing the hydraulic retention time (HRT)) and organic (increasing the organic loading rate maintaining the HRT constant) overload shocks in anaerobic reactors treating agro-industrial wastes, as well as during the recovery period. In both cases, the organic loading rate increased from 2 to 10gCODL(-1)d(-1), resulting in VFAs accumulation up to 9gL(-1). Both overloads were correlated to an increase in Bacteroidetes and Actinobacteria phyla and with a drop in Syntrophomonadaceae and Pseudomonadaceae families. In contrast, Tissierellaceae family only increased during the organic shock. Active Archaea decreased in both overloads, going from Methanosaeta dominance to Methanosarcina prevalence. During the recovery period, Porphyromonadaceae family increased its presence and Clostridium genus recovered values prior to perturbation.

  17. Discovery of clostrubin, an exceptional polyphenolic polyketide antibiotic from a strictly anaerobic bacterium.

    PubMed

    Pidot, Sacha; Ishida, Keishi; Cyrulies, Michael; Hertweck, Christian

    2014-07-21

    Genome mining of the strictly anaerobic bacterium Clostridium beijerinckii, an industrial producer of solvents, revealed the presence of several cryptic gene clusters for secondary metabolite biosynthesis. To unearth its metabolic potential, a C. beijerinckii strain was cultured under various conditions, which led to the discovery of a deep purple pigment. This novel metabolite, named clostrubin (1), was isolated and its structure was fully elucidated. The pentacyclic polyphenol features a benzo[a]tetraphene ring topology that is unprecedented for natural products. Stable-isotope labeling experiments showed that 1 is an aromatic polyketide that folds in a noncanonical manner to form the unusual perifused ring system. In addition to being the first reported polyketide from an anaerobic bacterium, 1 is a potent antibiotic with pronounced activity against various pathogenic bacteria, such as MRSA, VRE, and mycobacteria, with minimum inhibitory concentrations (MIC) of 0.12-0.97 μM.

  18. MALDI-TOF MS identification of anaerobic bacteria: assessment of pre-analytical variables and specimen preparation techniques.

    PubMed

    Hsu, Yen-Michael S; Burnham, Carey-Ann D

    2014-06-01

    Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has emerged as a tool for identifying clinically relevant anaerobes. We evaluated the analytical performance characteristics of the Bruker Microflex with Biotyper 3.0 software system for identification of anaerobes and examined the impact of direct formic acid (FA) treatment and other pre-analytical factors on MALDI-TOF MS performance. A collection of 101 anaerobic bacteria were evaluated, including Clostridium spp., Propionibacterium spp., Fusobacterium spp., Bacteroides spp., and other anaerobic bacterial of clinical relevance. The results of our study indicate that an on-target extraction with 100% FA improves the rate of accurate identification without introducing misidentification (P<0.05). In addition, we modify the reporting cutoffs for the Biotyper "score" yielding acceptable identification. We found that a score of ≥1.700 can maximize the rate of identification. Of interest, MALDI-TOF MS can correctly identify anaerobes grown in suboptimal conditions, such as on selective culture media and following oxygen exposure. In conclusion, we report on a number of simple and cost-effective pre- and post-analytical modifications could enhance MALDI-TOF MS identification for anaerobic bacteria.

  19. Clostridium difficile in poultry and poultry meat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The incidence and severity of disease associated with toxigenic Clostridium difficile have increased in hospitals in North America from the emergence of newer, more virulent strains. Toxigenic C. difficile has been isolated from food animals and retail meat with potential implications of transfer t...

  20. Insights in metabolism and toxin production from the complete genome sequence of Clostridium tetani.

    PubMed

    Brüggemann, Holger; Gottschalk, Gerhard

    2004-04-01

    The decryption of prokaryotic genome sequences progresses rapidly and provides the scientific community with an enormous amount of information. Clostridial genome sequencing projects have been finished only recently, starting with the genome of the solvent-producing Clostridium acetobutylicum in 2001. A lot of attention has been devoted to the genomes of pathogenic clostridia. In 2002, the genome sequence of C. perfringens, the causative agent of gas gangrene, has been released. Currently in the finishing stage and prior to publication are the genomes of the foodborne botulism-causing C. botulinum and of C. difficile, the causative agent of a wide spectrum of clinical manifestations such as antibiotic-associated diarrhea. Our team sequenced the genome of neuropathogenic C. tetani, a Gram-positive spore-forming bacterium predominantly found in the soil. In deep wound infections it occasionally causes spastic paralysis in humans and vertebrate animals, known as tetanus disease, by the secretion of potent neurotoxin, designated tetanus toxin. The toxin blocks the release of neurotransmitters from presynaptic membranes of interneurons of the spinal cord and the brainstem, thus preventing muscle relaxation. Fortunately, this disease is successfully controlled through immunization with tetanus toxoid, a formaldehyde-treated tetanus toxin, but nevertheless, an estimated 400,000 cases still occur each year, mainly of neonatal tetanus. The World Health Organization has stated that neonatal tetanus is the second leading cause of death from vaccine preventable diseases among children worldwide. This minireview focuses on an analysis of the genome sequence of C. tetani E88, a vaccine production strain, which is a toxigenic non-sporulating variant of strain Massachusetts. The genome consists of a 2,799,250 bp chromosome encoding 2618 open reading frames. The tetanus toxin is encoded on a 74,082 kb plasmid, containing 61 genes. Additional virulence-related factors as well as an

  1. Whole genome sequence of Clostridium bornimense strain M2/40 isolated from a lab-scale mesophilic two-phase biogas reactor digesting maize silage and wheat straw.

    PubMed

    Hahnke, Sarah; Wibberg, Daniel; Tomazetto, Geizecler; Pühler, Alfred; Klocke, Michael; Schlüter, Andreas

    2014-08-20

    The bacterium Clostridium bornimense M2/40 is a mesophilic, anaerobic bacterium isolated from a two-phase biogas reactor continuously fed with maize silage and 5% wheat straw. Grown on glucose, it produced H2, CO2, formiate, lactate and propionate as the main fermentation products, of which some compounds serve as substrates for methanogenic Archaea to form methane. Here, the whole genome sequence of the bacterium consisting of two circular replicons is reported. This genome information provides the basis for further studies addressing metabolic features of the isolate and its role in anaerobic biomass degradation.

  2. Proteomic analysis and identification of cell surface-associated proteins of Clostridium chauvoei.

    PubMed

    Jayaramaiah, Usharani; Singh, Neetu; Thankappan, Sabarinath; Mohanty, Ashok Kumar; Chaudhuri, Pallab; Singh, Vijendra Pal; Nagaleekar, Viswas Konasagara

    2016-06-01

    Blackleg is a highly fatal disease of cattle and sheep, caused by Clostridium chauvoei, a Gram positive, anaerobic, spore forming bacteria. Cell surface-associated proteins play a major role in inducing the protective immunity. However, the identity of a majority of cell surface-associated proteins of C. chauvoei is not known. In the present investigation, we have used SDS-PAGE, 2D-gel electrophoresis and Western blotting followed by mass spectrometry to identify cell surface-associated proteins of C. chauvoei. Among the identified proteins, which have shown to offer protective antigencity in other bacteria, Enolase, Chaperonin, Ribosomal protein L10, Glycosyl Hydrolase and Flavoprotein were characterized by sequencing and their overexpression in Escherichia coli. In conclusion, cell surface-associated proteins were identified using proteomic approach and the genes for the immunoreactive proteins were expressed, which may prove to be potential diagnostic or vaccine candidates.

  3. Mutant alcohol dehydrogenase leads to improved ethanol tolerance in Clostridium thermocellum

    SciTech Connect

    Brown, Steven D; Guss, Adam M; Karpinets, Tatiana V; Parks, Jerry M; Smolin, Nikolai; Yang, Shihui; Land, Miriam L; Klingeman, Dawn Marie; Bhandiwad, Ashwini; Rodriguez, Jr., Miguel; Raman, Babu; Shao, Xiongjun; Mielenz, Jonathan R; Smith, Jeremy C; Keller, Martin; Lynd, Lee R

    2011-01-01

    Clostridium thermocellum is a thermophilic, obligately anaerobic, Gram-positive bacterium that is a candidate microorganism for converting cellulosic biomass into ethanol through consolidated bioprocessing. Ethanol intolerance is an important metric in terms of process economics, and tolerance has often been described as a complex and likely multigenic trait for which complex gene interactions come into play. Here, we resequence the genome of an ethanol-tolerant mutant, show that the tolerant phenotype is primarily due to a mutated bifunctional acetaldehyde-CoA/alcohol dehydrogenase gene (adhE), hypothesize based on structural analysis that cofactor specificity may be affected, and confirm this hypothesis using enzyme assays. Biochemical assays confirm a complete loss of NADH-dependent activity with concomitant acquisition of NADPH-dependent activity, which likely affects electron flow in the mutant. The simplicity of the genetic basis for the ethanol-tolerant phenotype observed here informs rational engineering of mutant microbial strains for cellulosic ethanol production.

  4. Closing the Carbon Balance for Fermentation by Clostridium thermocellum (ATCC 27405)

    SciTech Connect

    Ellis, Lucas D; Holwerda, Evert K; Hogsett, David; Rogers, Steve; Shao, Xiongjun; Tschaplinski, Timothy J; Thorne, Phil; Lynd, L.

    2012-01-01

    Our lab and most others have not been able to close a carbon balance for fermentation by the thermophilic, cellulolytic anaerobe, Clostridium thermocellum. We undertook a detailed accounting of product formation in C. thermocellum ATCC 27405. Elemental analysis revealed that for both cellulose (Avicel) and cellobiose, {>=}92% of the substrate carbon utilized could be accounted for in the pellet, supernatant and off-gas when including sampling. However, 11.1% of the original substrate carbon was found in the liquid phase and not in the form of commonly-measured fermentation products - ethanol, acetate, lactate, and formate. Further detailed analysis revealed all the products to be <720 da and have not usually been associated with C. thermocellum fermentation, including malate, pyruvate, uracil, soluble glucans, and extracellular free amino acids. By accounting for these products, 92.9% and 93.2% of the final product carbon was identified during growth on cellobiose and Avicel, respectively.

  5. Clostridium hastiforme bacteraemia secondary to pyometra in a 64-year-old woman.

    PubMed

    Ørum, Matilde; Fuglsang-Damgaard, David; Nielsen, Hans Linde

    2017-01-18

    The Gram-negative, rod-shaped, anaerobe bacteria Tissierella praeacuta was first described in 1908 by Tissier. However, during the past decade, Clostridium hastiforme has been recognised as a later synonym of T. praeacuta. C. hastiforme/T. praeacuta has only rarely been described in previous literature as a cause of human infection. We present here a case report of C. hastiforme/T. praeacuta bacteraemia secondary to pyometra in a 64-year-old woman with a history of multiple sclerosis and an intrauterine device inserted three decades earlier. C. hastiforme/T. praeacuta was isolated from blood as well as pus from the site of infection. The patient was cured with a combination of drainage and antibiotic therapy.

  6. Fatal clostridium septicum myonecrosis in a captive canada lynx (Lynx canadensis).

    PubMed

    Izer, Jenelle M; Wilson, Ronald P; Cooper, Timothy K

    2014-09-01

    A 1-yr-old female Canada lynx (Lynx canadensis) presented for sudden onset of rapidly progressive bilateral pelvic limb paralysis. The lynx was chemically immobilized to perform a physical examination but expired shortly thereafter. On postmortem radiographs, there were myriad small irregular, round-to-spherical gas densities within the skeletal muscle of the right thigh and epaxial musculature. At gross necropsy, the muscles of the right thigh, right lateral abdominal wall, and epaxial region were emphysematous and necrohemorrhagic, with subcutaneous and muscular crepitant swelling. Multiple skin puncture wounds, consistent with bites, were present over the affected tissues. Clostridium septicum was isolated in pure anaerobic culture from the musculature of the right hind limb. Histopathologic examination confirmed the diagnosis of acute, severe necrohemorrhagic and gangrenous myositis and cellulitis. Gram stains demonstrated large gram-positive bacilli with subterminal spores. This is the first known documented case of C. septicum myonecrosis in a nondomestic felid.

  7. An update on antibody-based immunotherapies for Clostridium difficile infection

    PubMed Central

    Hussack, Greg; Tanha, Jamshid

    2016-01-01

    Clostridium difficile continues to be one of the most prevalent hospital-acquired bacterial infections in the developed world, despite the recent introduction of a novel and effective antibiotic agent (fidaxomicin). Alternative approaches under investigation to combat the anaerobic Gram-positive bacteria include fecal transplantation therapy, vaccines, and antibody-based immunotherapies. In this review, we catalog the recent advances in antibody-based approaches under development and in the clinic for the treatment of C. difficile infection. By and large, inhibitory antibodies that recognize the primary C. difficile virulence factors, toxin A and toxin B, are the most popular passive immunotherapies under investigation. We provide a detailed summary of the toxin epitopes recognized by various antitoxin antibodies and discuss general trends on toxin inhibition efficacy. In addition, antibodies to other C. difficile targets, such as surface-layer proteins, binary toxin, motility factors, and adherence and colonization factors, are introduced in this review. PMID:27536153

  8. Characterization of a heme oxygenase of Clostridium tetani and its possible role in oxygen tolerance.

    PubMed

    Brüggemann, Holger; Bauer, Rosalie; Raffestin, Stéphanie; Gottschalk, Gerhard

    2004-10-01

    In order to colonize mammalian wounds, the anaerobic bacterium Clostridium tetani must presumably cope with temporary oxic conditions. Therefore, the recently decoded genome sequence was searched for genes which could confer oxygen tolerance. A few identified systems such as superoxide dismutases and peroxidases are probably responsible for this protection against toxic oxygen species. Another system was detected, a heme oxygenase which could have a role in establishing or maintaining an anoxic microenvironment in the process of wound colonization. The hemT gene encoding the heme oxygenase is expressed in C. tetani, as shown by reverse transcription-PCR. When overexpressed in Escherichia coli, the enzyme converts heme to biliverdin under strict oxic conditions.

  9. Purification and properties of 4-hydroxybutyrate coenzyme A transferase from Clostridium aminobutyricum.

    PubMed Central

    Scherf, U; Buckel, W

    1991-01-01

    A new coenzyme A (CoA)-transferase from the anaerobe Clostridium aminobutyricum catalyzing the formation of 4-hydroxybutyryl-CoA from 4-hydroxybutyrate and acetyl-CoA is described. The enzyme was purified to homogeneity by standard techniques, including fast protein liquid chromatography under aerobic conditions. Its molecular mass was determined to be 110 kDa, and that of the only subunit was determined to be 54 kDa, indicating a homodimeric structure. Besides acetate and acetyl-CoA, the following substrates were detected (in order of decreasing kcat/Km): 4-hydroxybutyryl-CoA, butyryl-CoA and propionyl-CoA, vinyl-acetyl-CoA (3-butenoyl-CoA), and 5-hydroxyvaleryl-CoA. In an indirect assay the corresponding acids were also found to be substrates; however, DL-lactate, DL-2-hydroxybutyrate, DL-3-hydroxybutyrate, crotonate, and various dicarboxylates were not. PMID:1768145

  10. MLST analysis reveals a highly conserved core genome among poultry isolates of Clostridium septicum.

    PubMed

    Neumann, Anthony P; Rehberger, Thomas G

    2009-06-01

    Clostridium septicum is a highly virulent, anaerobic bacterium capable of establishing necrotizing tissue infections and forming heat resistant endospores. Disease is primarily facilitated by secretion of numerous toxic products including a lethal pore-forming cytolysin. Spontaneously occurring clostridial myonecrosis involving C. septicum has recently reemerged as a concern for many poultry producers. However, despite its increasing prevalence, the epidemiology of infection and population structure of C. septicum remains largely unknown. In this study a multilocus sequence typing (MLST) approach was utilized to examine evolutionary relationships within a diverse collection of C. septicum isolates recovered from poultry flocks experiencing episodes of gangrenous dermatitis. The 109 isolates examined represented 42 turkey flocks and 24 different flocks of broiler chickens as well as C. septicum type strain, ATCC 12464. Isolates were recovered predominantly from gangrenous lesions although isolates from livers, gastrointestinal tracts, spleens and blood were included. The loci analyzed were csa, the major lethal toxin produced by C. septicum, and the housekeeping genes gyrA, groEL, dnaK, recA, tpi, ddl, colA and glpK. These loci were included in part because of their previous use in MLST analysis of Clostridium perfringens and Clostridium difficile. Results indicated a high level of conservation present within these housekeeping gene fragments when compared to what has been previously reported for the aforementioned clostridia. Of the 5352 bp of sequence data examined for each isolate, 99.7% (5335/5352) was absolutely conserved among the 109 isolates. Only one of the ten unique sequence types, or allelic profiles, identified among the isolates was recovered from both turkeys and broiler chickens suggesting some host species preference. Phylogenetic analyses identified two unique clusters, or clonal complexes, among these poultry isolates which may have important

  11. Anaerobic Metabolism of Indoleacetate

    PubMed Central

    Ebenau-Jehle, Christa; Thomas, Markus; Scharf, Gernot; Kockelkorn, Daniel; Knapp, Bettina; Schühle, Karola; Heider, Johann

    2012-01-01

    The anaerobic metabolism of indoleacetate (indole-3-acetic acid [IAA]) in the denitrifying betaproteobacterium Azoarcus evansii was studied. The strain oxidized IAA completely and grew with a generation time of 10 h. Enzyme activities that transformed IAA were present in the soluble cell fraction of IAA-grown cells but were 10-fold downregulated in cells grown on 2-aminobenzoate or benzoate. The transformation of IAA did not require molecular oxygen but required electron acceptors like NAD+ or artificial dyes. The first products identified were the enol and keto forms of 2-oxo-IAA. Later, polar products were observed, which could not yet be identified. The first steps likely consist of the anaerobic hydroxylation of the N-heterocyclic pyrrole ring to the enol form of 2-oxo-IAA, which is catalyzed by a molybdenum cofactor-containing dehydrogenase. This step is probably followed by the hydrolytic ring opening of the keto form, which is catalyzed by a hydantoinase-like enzyme. A comparison of the proteome of IAA- and benzoate-grown cells identified IAA-induced proteins. Owing to the high similarity of A. evansii with strain EbN1, whose genome is known, we identified a cluster of 14 genes that code for IAA-induced proteins involved in the early steps of IAA metabolism. These genes include a molybdenum cofactor-dependent dehydrogenase of the xanthine oxidase/aldehyde dehydrogenase family, a hydantoinase, a coenzyme A (CoA) ligase, a CoA transferase, a coenzyme B12-dependent mutase, an acyl-CoA dehydrogenase, a fusion protein of an enoyl-CoA hydratase and a 3-hydroxyacyl-CoA dehydrogenase, a beta-ketothiolase, and a periplasmic substrate binding protein for ABC transport as well as a transcriptional regulator of the GntR family. Five predicted enzymes form or act on CoA thioesters, indicating that soon after the initial oxidation of IAA and possibly ring opening, CoA thioesters are formed, and the carbon skeleton is rearranged, followed by a CoA-dependent thiolytic

  12. Development of a recombinant flagellin based ELISA for the detection of Clostridium chauvoei.

    PubMed

    Usharani, J; Nagaleekar, Viswas Konasagara; Thomas, Prasad; Gupta, Santosh K; Bhure, Sanjeev K; Dandapat, Premanshu; Agarwal, Rajesh K; Singh, Vijendra P

    2015-06-01

    Blackleg, an economically important and highly fatal disease of ruminants, is caused by anaerobic bacillus, Clostridium chauvoei. Identification and differentiation of the causative agent is crucial for implementation of therapeutic and control measures in real time. Most of the diagnostic tests available for blackleg are PCR based, and only a couple of serological tests have been reported. In this study, we targeted flagellin, an important immunogenic protein of C. chauvoei, to develop a sandwich ELISA for detection of C. chauvoei. Sequence analysis of flagellin gene of related Clostridium species showed that central region of flagellin gene is unique to C. chauvoei. Hence, we cloned and expressed central region of flagellin in a prokaryotic expression system. Antiserum against recombinant flagellin was generated in rabbits and chickens. A sandwich ELISA was developed, in which rabbit anti-flagellin antibodies were used as capture antibodies and chicken anti-flagellin antibodies as detecting antibodies. The test was specific and sensitive in detection of up to 10(4) CFU/ml of C. chauvoei. This study shows that assay developed can be used for detection of C. chauvoei in suspected samples.

  13. Use of Clostridium botulinum toxin in gastrointestinal motility disorders in children

    PubMed Central

    Arbizu, Ricardo A; Rodriguez, Leonel

    2015-01-01

    More than a century has elapsed since the identification of Clostridia neurotoxins as the cause of paralytic diseases. Clostridium botulinum is a heterogeneous group of Gram-positive, rod-shaped, spore-forming, obligate anaerobic bacteria that produce a potent neurotoxin. Eight different Clostridium botulinum neurotoxins have been described (A-H) and 5 of those cause disease in humans. These toxins cause paralysis by blocking the presynaptic release of acetylcholine at the neuromuscular junction. Advantage can be taken of this blockade to alleviate muscle spams due to excessive neural activity of central origin or to weaken a muscle for treatment purposes. In therapeutic applications, minute quantities of botulinum neurotoxin type A are injected directly into selected muscles. The Food and Drug Administration first approved botulinum toxin (BT) type A in 1989 for the treatment of strabismus and blepharospasm associated with dystonia in patients 12 years of age or older. Ever since, therapeutic applications of BT have expanded to other systems, including the gastrointestinal tract. Although only a single fatality has been reported to our knowledge with use of BT for gastroenterological conditions, there are significant complications ranging from minor pain, rash and allergic reactions to pneumothorax, bowel perforation and significant paralysis of tissues surrounding the injection (including vocal cord paralysis and dysphagia). This editorial describes the clinical experience and evidence for the use BT in gastrointestinal motility disorders in children. PMID:25992183

  14. Complete Genome Sequence of Clostridium estertheticum DSM 8809, a Microbe Identified in Spoiled Vacuum Packed Beef

    PubMed Central

    Yu, Zhongyi; Gunn, Lynda; Brennan, Evan; Reid, Rachael; Wall, Patrick G.; Gaora, Peadar Ó.; Hurley, Daniel; Bolton, Declan; Fanning, Séamus

    2016-01-01

    Blown pack spoilage (BPS) is a major issue for the beef industry. Etiological agents of BPS involve members of a group of Clostridium species, including Clostridium estertheticum which has the ability to produce gas, mostly carbon dioxide, under anaerobic psychotrophic growth conditions. This spore-forming bacterium grows slowly under laboratory conditions, and it can take up to 3 months to produce a workable culture. These characteristics have limited the study of this commercially challenging bacterium. Consequently information on this bacterium is limited and no effective controls are currently available to confidently detect and manage this production risk. In this study the complete genome of C. estertheticum DSM 8809 was determined by SMRT® sequencing. The genome consists of a circular chromosome of 4.7 Mbp along with a single plasmid carrying a potential tellurite resistance gene tehB and a Tn3-like resolvase-encoding gene tnpR. The genome sequence was searched for central metabolic pathways that would support its biochemical profile and several enzymes contributing to this phenotype were identified. Several putative antibiotic/biocide/metal resistance-encoding genes and virulence factors were also identified in the genome, a feature that requires further research. The availability of the genome sequence will provide a basic blueprint from which to develop valuable biomarkers that could support and improve the detection and control of this bacterium along the beef production chain. PMID:27891116

  15. Microbial bioproducts from cheese whey through fermentation with wastewater sludge Clostridium isolates.

    PubMed

    Ellis, Joshua T; Sims, Ronald C; Miller, Charles D

    2014-07-01

    We demonstrated the production of hydrogen, ethanol, and a variety of acids by several Clostridium species using cheese whey as substrate. These species were isolated from the anaerobic sediments of a municipal wastewater stabilization pond. Eight isolates were obtained and all were classified taxonomically as Clostridium spp. based on 16S rRNA sequencing. Sludge isolates showed maximum bioproduct production yields and productivities after approximately 24 h of batch cultivation with 6% (m/v) cheese whey. Fermentation byproducts measured included hydrogen, ethanol, acetic acid, butyric acid, and lactic acid. The maximum yields of bioproducts were 0.59 mol H(2)/mol lactose, 0.071 g ethanol/g, 0.204 g acetic acid/g, 0.218 g butyric acid/g, and 0.144 g lactic acid/g. The production of these high value biofuels and biofuel intermediates from cheese whey could have significant implications for conversion of waste to high value bioproducts to enhance domestic energy economies.

  16. Spoilage of an acid food product by Clostridium perfringens, C. barati and C. butyricum.

    PubMed

    de Jong, J

    1989-05-01

    Spoilage of canned pasteurized brined mung bean sprouts, acidified with citric acid to pH 4.0-4.5, was found to be caused by acid tolerant Clostridium spp. including the species barati, perfringens and butyricum. The pH limit for growth in the brine used were estimated 3.7, 3.7 and 4.0 respectively. Some of the isolated C. perfringens strains produced enterotoxins in sporulation media. The spores of the isolated anaerobes appeared to originate from mung beans, but C. barati and C. perfringens strains freshly isolated from dry beans, were unable to grow in acidified brine. During germination and sprouting of mung beans, the oxygen concentration decreased, while carbon dioxide concentration increased considerably, due to respiration of the sprouts and actively growing Enterobacteriaceae and lactobacilli. It was assumed that this allowed C. barati and C. perfringens strains to grow and acquire the observed unusual acid tolerance. After increasing aerobicity during sprouting, no growth of Clostridium spp. was observed, substantiating the assumption.

  17. Anaerobic Bacteremia: Impact of Inappropriate Therapy on Mortality

    PubMed Central

    Lee, Yangsoon; Park, Yongjung; Kim, Myungsook; Choi, Jun Yong; Yong, Dongeun; Jeong, Seok Hoon

    2016-01-01

    Background Investigation on incidence and mortality of anaerobic bacteremia (AB) is clinically relevant in spite of its infrequent occurrence and not often explored, which report varies according to period and institutions. Therefore, it is necessary to analyze the incidence and risk factors related to mortality and assess clinical outcomes of AB in current aspect. Materials and Methods Characteristics of AB patients and anaerobic bacteria from blood culture at a university hospital in 2012 were reviewed retrospectively. The correlation between risk factors and 28-day patient mortality was analyzed. Results A total of 70 non-duplicated anaerobic bacteria were isolated from blood of 70 bacteremia patients in 2012. The history of cardiovascular disease as host's risk factor was statistically significant (P = 0.0344) in univariate and multivariate analysis. Although the inappropriate therapy was not statistically significant in univariate and multivariate analysis, the survival rate of bacteremia was significantly worse in patients who had inappropriate therapy compared with those underwent appropriate therapy (hazard ratio, 5.4; 95% confidence interval, 1.7–6.9; P = 0.004). The most frequently isolated organism was Bacteroides fragilis (32 isolates, 46%), followed by Bacteroides thetaiotaomicron (10, 14%), and non-perfringens Clostridium (7, 10%). Conclusion The incidence of AB in 2012 was 2.3% (number of AB patients per 100 positive blood culture patients) and the mortality rate in patients with clinically significant AB was 21.4%. In addition, AB was frequently noted in patients having malignancy and the survival rate of AB was significantly worse in patients who received inappropriate therapy compared with those underwent appropriate therapy. PMID:27433379

  18. PCB breakdown by anaerobic microorganisms

    SciTech Connect

    Not Available

    1989-03-01

    Recently, altered PCB cogener distribution patterns observed in anaerobic sediment samples from the upper Hudson River are being attributed to biologically mediated reductive dechlorination. The authors report their successful demonstration of biologically mediated reductive dechlorination of an Aroclor mixture. In their investigation, they assessed the ability of microorganisms from PCB-contaminated Hudson River sediments (60-562 ppm PCBs) to dechlorinate Aroclor 1242 under anaerobic conditions by eluting microorganisms from the PCB- contaminated sediments and transferring them to a slurry of reduced anaerobic mineral medium and PCB-free sediments in tightly stoppered bottles. They observed dechlorination to be the most rapid at the highest PCB concentration tried by them.

  19. Antarctic strict anaerobic microbiota from Deschampsia antarctica vascular plants rhizosphere reveals high ecology and biotechnology relevance.

    PubMed

    Peixoto, Rafael José Marques; Miranda, Karla Rodrigues; Lobo, Leandro Araujo; Granato, Alessandra; de Carvalho Maalouf, Pedro; de Jesus, Hugo Emiliano; Rachid, Caio T C C; Moraes, Saulo Roni; Dos Santos, Henrique Fragoso; Peixoto, Raquel Silva; Rosado, Alexandre Soares; Domingues, Regina Maria Cavalcanti Pilotto

    2016-11-01

    The Antarctic soil microbial community has a crucial role in the growth and stabilization of higher organisms, such as vascular plants. Analysis of the soil microbiota composition in that extreme environmental condition is crucial to understand the ecological importance and biotechnological potential. We evaluated the efficiency of isolation and abundance of strict anaerobes in the vascular plant Deschampsia antarctica rhizosphere collected in the Antarctic's Admiralty Bay and associated biodiversity to metabolic perspective and enzymatic activity. Using anaerobic cultivation methods, we identified and isolated a range of microbial taxa whose abundance was associated with Plant Growth-Promoting Bacteria (PGPB) and presences were exclusively endemic to the Antarctic continent. Firmicutes was the most abundant phylum (73 %), with the genus Clostridium found as the most isolated taxa. Here, we describe two soil treatments (oxygen gradient and heat shock) and 27 physicochemical culture conditions were able to increase the diversity of anaerobic bacteria isolates. Heat shock treatment allowed to isolate a high percentage of new species (63.63 %), as well as isolation of species with high enzymatic activity (80.77 %), which would have potential industry application. Our findings contribute to the understanding of the role of anaerobic microbes regarding ecology, evolutionary, and biotechnological features essential to the Antarctic ecosystem.

  20. Effects of anaerobic/aerobic incubation and storage temperature on preservation and deodorization of kitchen garbage.

    PubMed

    Wang, Qunhui; Narita, Jun-ya; Xie, Weimin; Ohsumi, Yukihide; Kusano, Kohji; Shirai, Yoshihito; Ogawa, Hiroaki I

    2002-09-01

    To develop a garbage recycling system for the purpose of the production of lactic acid (LA) to use as raw material for producing biodegradable plastics, the preservation and deodorization of garbage during storage are very important. Anaerobic incubation (i.e., storage) was prove to be more suitable than aerobic incubation during the garbage storage in terms of concentration of LA and soluble sugar, pH value, viable bacteria counts and offensive odour substances. This difference is due to a fact that the growth of putrefactive bacteria such as coliforms and Clostridium spp. appeared to be inhibited by anaerobic fermentation during the storage, because the fermentation caused a drop of garbage pH and generated inhibitory substances, i.e., bacteriocins. Under anaerobic condition, LA concentration in the stored garbage was found to be higher in the order: 37 > 25 > 50 > 5 degrees C, and the concentration of sugar accumulated during the 50 degrees C-storage was the highest. Among the conditions employed, the optimum condition for the storage of kitchen garbage was anaerobic at 5 degrees C.

  1. Epidemiology and Antimicrobial Susceptibilities of Wound Isolates of Obligate Anaerobes from Combat Casualties

    PubMed Central

    White, Brian K.; Mende, Katrin; Weintrob, Amy C.; Beckius, Miriam L.; Zera, Wendy C.; Lu, Dan; Bradley, William; Tribble, David R.; Schnaubelt, Elizabeth R.; Murray, Clinton K.

    2015-01-01

    Data from recent conflicts related to war wounds and obligate anaerobes are limited. We define the epidemiology and antimicrobial susceptibility of obligate anaerobes from Iraq and Afghanistan casualties (6/2009–12/2013), as well as their association with clinical outcomes. Susceptibility against eleven antibiotics (7 classes) was tested. Overall, 59 patients had 119 obligate anaerobes identified (83 were first isolates). Obligate anaerobes were isolated 7–13 days post-injury, primarily from lower extremity wounds (43%), and were largely Bacteroides spp. (42%) and Clostridium spp. (19%). Patients with pelvic wounds were more likely to have Bacteroides spp. and concomitant resistant gram-negative aerobes. Seventy-three percent of isolates were resistant to ≥1 antimicrobials. Bacteroides spp. demonstrated the most resistance (16% of first isolates). Patients with resistant isolates had similar outcomes to those with susceptible strains. Serial recovery of isolates occurred in 15% of patients and was significantly associated with isolation of Bacteroides spp., along with resistant gram-negative aerobes. PMID:26607420

  2. Microbial community dynamics in batch high-solid anaerobic digestion of food waste under mesophilic conditions.

    PubMed

    Yi, Jing; Dong, Bin; Xue, Yonggang; Li, Ning; Gao, Peng; Zhao, Yuxin; Dai, Lingling; Dai, Xiaohu

    2014-02-28

    Microbial community shifts, associated with performance data, were investigated in an anaerobic batch digester treating high-solid food waste under mesophilic conditions using, a combination of molecular techniques and chemical analysis methods. The batch process was successfully operated with an organic removal efficiency of 44.5% associated with a biogas yield of 0.82 L/g VSremoval. Microbial community structures were examined by denaturing gel gradient electrophoresis. Clostridium and Symbiobacterium organisms were suggested to be mainly responsible for the organic matter catabolism in hydrolysis and acidogenesis reactions. The dynamics of archaeal and methanogenic populations were monitored using real-time PCR targeting 16S rRNA genes. Methanosarcina was the predominant methanogen, suggesting that the methanogenesis took place mainly via an aceticlastic pathway. Hydrogenotrophic methanogens were also supported in high-solid anaerobic digestion of food waste through syntrophism with syntrophic bacterium. Microbial community shifts showed good agreement with the performance parameters in anaerobic digestion, implying the possibility of diagnosing a high-solid anaerobic digestion process by monitoring microbial community shifts. On the other hand, the batch results could be relevant to the start-up period of a continuous system and could also provide useful information to set up a continuous operation.

  3. Anaerobic sporeformers and their significance with respect to milk and dairy products.

    PubMed

    Doyle, Conor J; Gleeson, David; Jordan, Kieran; Beresford, Tom P; Ross, R Paul; Fitzgerald, Gerald F; Cotter, Paul D

    2015-03-16

    Sporeforming bacteria are a significant concern for the international dairy industry. Spores present in milk survive heat treatments and can persist during downstream processing. If they are present in sufficient numbers in dairy products they can cause spoilage or lead to illness as a result of toxin production. While many reviews have highlighted the threat posed by spores of aerobic bacteria to the dairy industry, few have focused on problems caused by the array of different species of anaerobic sporeformers (Clostridium and related genera) that can be found in milk. This is despite of the fact that members of these bacteria are found throughout the dairy farm environment, and can be toxigenic, neurotoxigenic or spoilage bacteria. This makes the possible presence of Clostridium and related spores in bulk tank milk (BTM) important from both a financial and a public health perspective. In this review dairy associated anaerobic sporeformers are assessed from a number of perspectives. This includes the taxonomy of this group of bacteria, the important subgroup of this genus the "sulphite reducing clostridia" (SRC), how these bacteria are detected in milk products, the epidemiological data regarding pathogenic species and strains within the SRC group as well as the influence of farming practices on the presence of SRC in BTM.

  4. Screening of anaerobic activities in sediments of an acidic environment: Tinto River.

    PubMed

    Sánchez-Andrea, Irene; Rojas-Ojeda, Patricia; Amils, Ricardo; Sanz, José Luis

    2012-11-01

    The Tinto River (Huelva, Spain) is a natural acidic rock drainage environment produced by the bio-oxidation of metallic sulfides from the Iberian Pyritic Belt. A geomicrobiological model of the different microbial cycles operating in the sediments was recently developed through molecular biological methods, suggesting the presence of iron reducers, methanogens, nitrate reducers and hydrogen producers. In this study, we used a combination of molecular biological methods and targeted enrichment incubations to validate this model and prove the existence of those potential anaerobic activities in the acidic sediments of Tinto River. Methanogenic, sulfate-reducing, denitrifying and hydrogen-producing enrichments were all positive at pH between 5 and 7. Methanogenic enrichments revealed the presence of methanogenic archaea belonging to the genera Methanosarcina and Methanobrevibacter. Enrichments for sulfate-reducing microorganisms were dominated by Desulfotomaculum spp. Denitrifying enrichments showed a broad diversity of bacteria belonging to the genera Paenibacillus, Bacillus, Sedimentibacter, Lysinibacillus, Delftia, Alcaligenes, Clostridium and Desulfitobacterium. Hydrogen-producing enrichments were dominated by Clostridium spp. These enrichments confirm the presence of anaerobic activities in the acidic sediments of the Tinto River that are normally assumed to take place exclusively at neutral pH.

  5. Diagnostic trends in Clostridium difficile detection in Finnish microbiology laboratories.

    PubMed

    Könönen, Eija; Rasinperä, Marja; Virolainen, Anni; Mentula, Silja; Lyytikäinen, Outi

    2009-12-01

    Due to increased interest directed to Clostridium difficile-associated infections, a questionnaire survey of laboratory diagnostics of toxin-producing C. difficile was conducted in Finland in June 2006. Different aspects pertaining to C. difficile diagnosis, such as requests and criteria used for testing, methods used for its detection, yearly changes in diagnostics since 1996, and the total number of investigations positive for C. difficile in 2005, were asked in the questionnaire, which was sent to 32 clinical microbiology laboratories, including all hospital-affiliated and the relevant private clinical microbiology laboratories in Finland. The situation was updated by phone and email correspondence in September 2008. In June 2006, 28 (88%) laboratories responded to the questionnaire survey; 24 of them reported routinely testing requested stool specimens for C. difficile. Main laboratory methods included toxin detection (21/24; 88%) and/or anaerobic culture (19/24; 79%). In June 2006, 18 (86%) of the 21 laboratories detecting toxins directly from feces, from the isolate, or both used methods for both toxin A (TcdA) and B (TcdB), whereas only one laboratory did so in 1996. By September 2008, all of the 23 laboratories performing diagnostics for C. difficile used methods for both TcdA and TcdB. In 2006, the number of specimens processed per 100,000 population varied remarkably between different hospital districts. In conclusion, culturing C. difficile is common and there has been a favorable shift in toxin detection practice in Finnish clinical microbiology laboratories. However, the variability in diagnostic activity reported in 2006 creates a challenge for national monitoring of the epidemiology of C. difficile and related diseases.

  6. Analysis of Proline Reduction in the Nosocomial Pathogen Clostridium difficile▿

    PubMed Central

    Jackson, Sarah; Calos, Mary; Myers, Andrew; Self, William T.

    2006-01-01

    Clostridium difficile, a proteolytic strict anaerobe, has emerged as a clinically significant nosocomial pathogen in recent years. Pathogenesis is due to the production of lethal toxins, A and B, members of the large clostridial cytotoxin family. Although it has been established that alterations in the amino acid content of the growth medium affect toxin production, the molecular mechanism for this observed effect is not yet known. Since there is a paucity of information on the amino acid fermentation pathways used by this pathogen, we investigated whether Stickland reactions might be at the heart of its bioenergetic pathways. Growth of C. difficile on Stickland pairs yielded large increases in cell density in a limiting basal medium, demonstrating that these reactions are tied to ATP production. Selenium supplementation was required for this increase in cell yield. Analysis of genome sequence data reveals genes encoding the protein components of two key selenoenzyme reductases, glycine reductase and d-proline reductase (PR). These selenoenzymes were expressed upon the addition of the corresponding Stickland acceptor (glycine, proline, or hydroxyproline). Purification of the selenoenzyme d-proline reductase revealed a mixed complex of PrdA and PrdB (SeCys-containing) proteins. PR utilized only d-proline but not l-hydroxyproline, even in the presence of an expressed and purified proline racemase. PR was found to be independent of divalent cations, and zinc was a potent inhibitor of PR. These results show that Stickland reactions are key to the growth of C. difficile and that the mechanism of PR may differ significantly from that of previously studied PR from nonpathogenic species. PMID:17041035

  7. Lactobacillus acidophilus modulates the virulence of Clostridium difficile.

    PubMed

    Yun, B; Oh, S; Griffiths, M W

    2014-01-01

    Clostridium difficile is a spore-forming, toxin-producing, anaerobic bacterium that colonizes the human gastrointestinal tract. This pathogen causes antibiotic-associated diarrhea and colitis in animals and humans. Antibiotic-associated diseases may be treated with probiotics, and interest is increasing in such uses of probiotics. This study investigated the effect of Lactobacillus strains on the quorum-sensing signals and toxin production of C. difficile. In addition, an in vivo experiment was designed to assess whether Lactobacillus acidophilus GP1B is able to control C. difficile-associated disease. Autoinducer-2 activity was measured for C. difficile using the Vibrio harveyi coupled bioluminescent assay. Cell extract (10μg/mL) of L. acidophilus GP1B exhibited the highest inhibitory activity among 5 to 40μg/mL cell-extract concentrations. Real-time PCR data indicated decreased transcriptional levels in luxS, tcdA, tcdB, and txeR genes in the presence of 10μg/mL of cell extract of L. acidophilus GP1B. Survival rates at 5d for mice given the pathogen alone with L. acidophilus GP1B cell extract or L. acidophilus GP1B were 10, 70, and 80%, respectively. In addition, the lactic acid-produced L. acidophilus GP1B exhibits an inhibitory effect against the growth of C. difficile. Both the L. acidophilus GP1B and GP1B cell extract have significant antipathogenic effects on C. difficile.

  8. Cellodextrin and Laminaribiose ABC Transporters in Clostridium thermocellum▿

    PubMed Central

    Nataf, Yakir; Yaron, Sima; Stahl, Frank; Lamed, Raphael; Bayer, Edward A.; Scheper, Thomas-Helmut; Sonenshein, Abraham L.; Shoham, Yuval

    2009-01-01

    Clostridium thermocellum is an anaerobic thermophilic bacterium that grows efficiently on cellulosic biomass. This bacterium produces and secretes a highly active multienzyme complex, the cellulosome, that mediates the cell attachment to and hydrolysis of the crystalline cellulosic substrate. C. thermocellum can efficiently utilize only β-1,3 and β-1,4 glucans and prefers long cellodextrins. Since the bacterium can also produce ethanol, it is considered an attractive candidate for a consolidated fermentation process in which cellulose hydrolysis and ethanol fermentation occur in a single process. In this study, we have identified and characterized five sugar ABC transporter systems in C. thermocellum. The putative transporters were identified by sequence homology of the putative solute-binding lipoprotein to known sugar-binding proteins. Each of these systems is transcribed from a gene cluster, which includes an extracellular solute-binding protein, one or two integral membrane proteins, and, in most cases, an ATP-binding protein. The genes of the five solute-binding proteins were cloned, fused to His tags, overexpressed, and purified, and their abilities to interact with different sugars was examined by isothermal titration calorimetry. Three of the sugar-binding lipoproteins (CbpB to -D) interacted with different lengths of cellodextrins (G2 to G5), with disassociation constants in the micromolar range. One protein, CbpA, binds only cellotriose (G3), while another protein, Lbp (laminaribiose-binding protein) interacts with laminaribiose. The sugar specificity of the different binding lipoproteins is consistent with the observed substrate preference of C. thermocellum, in which cellodextrins (G3 to G5) are assimilated faster than cellobiose. PMID:18952792

  9. Mechanisms of Toxin Production of Food Bacteria (Clostridium botulinum)

    DTIC Science & Technology

    1983-12-01

    l~ V- 9;-iC -’.1,- r, 4. •, . . . . . MECHANISMS OF TOXIN PRODUCTION OF FOOD BACTERIA ( CLOSTRIDIUM BOTULINUM) FINAL REPORT DR. H. U. EKLUND F. T...Mechanisms of Toxin Production of Food Bacteria Clostridium botulinum Final Y,’v/ ’ "D30 • ’q• 6, PERFORM G ORG. REPORT NUMBER 7. AUTHOR(.) S...WORDS (Continue on reverse aide If necessary and Identify by block number) Clostridium botulinum Bacteriophages Plasmids Food Poisoning Toxins

  10. Genetic Engineering of Clostridium difficile Toxin A Vaccine

    DTIC Science & Technology

    1988-07-14

    AD N o GENETIC ENGINEERING OF CLOSTRIDIUM DIFFICILE TOXIN A VACCINE 0 C%" ANNUAL REPORT ! Lycurgus L. Muldrow Joe Johnson July 14, 1988 Supported by...17. COSATI CODES 18. SUBJECT TERMS (Continue on reverse if necessary and identify by block number) FIELD GROUP SUB-GROUP Clostridium difficile Vaccine ...development of vaccines . Improvement of vaccine biotechnology in the area of recombinant DNA studies using Clostridium difficile toxin A as the model, is

  11. Binary Interactions of Antagonistic Bacteria with Candida albicans Under Aerobic and Anaerobic Conditions.

    PubMed

    Benadé, Eliska; Stone, Wendy; Mouton, Marnel; Postma, Ferdinand; Wilsenach, Jac; Botha, Alfred

    2016-04-01

    We used both aerobic and anaerobic liquid co-cultures, prepared with Luria Bertani broth, to study the effect of bacteria on the survival of Candida albicans in the external environment, away from an animal host. The bacteria were represented by Aeromonas hydrophila, Bacillus cereus, Bacillus subtilis, Clostridium, Enterobacter, Klebsiella pneumoniae, Kluyvera ascorbata and Serratia marcescens. Under aerobic conditions, the yeast's growth was inhibited in the presence of bacterial growth; however, under anaerobic conditions, yeast and bacterial growth in co-cultures was similar to that observed for pure cultures. Subsequent assays revealed that the majority of bacterial strains aerobically produced extracellular hydrolytic enzymes capable of yeast cell wall hydrolysis, including chitinases and mannan-degrading enzymes. In contrast, except for the A. hydrophila strain, these enzymes were not detected in anaerobic bacterial cultures, nor was the antimicrobial compound prodigiosin found in anaerobic cultures of S. marcescens. When we suspended C. albicans cells in crude extracellular enzyme preparations from K. pneumoniae and S. marcescens, we detected no negative effect on yeast viability. However, we found that these preparations enhance the toxicity of prodigiosin towards the yeast, especially in combination with mannan-degrading enzymes. Analyses of the chitin and mannan content of yeast cell walls revealed that less chitin was produced under anaerobic than aerobic conditions; however, the levels of mannan, known for its low permeability, remained the same. The latter phenomenon, as well as reduced production of the bacterial enzymes and prodigiosin, may contribute to anaerobic growth and survival of C. albicans in the presence of bacteria.

  12. Array-based transcriptional analysis of Clostridium sporogenes UC9000 during germination, cell outgrowth and vegetative life.

    PubMed

    Bassi, Daniela; Cappa, Fabrizio; Cocconcelli, Pier Sandro

    2013-02-01

    The members of the genus Clostridium, including the spore-forming anaerobic bacteria, have a complex and strictly regulated life cycle, but very little is known about the genetic pathways involved in the different stages of their life cycle. Clostridium sporogenes, a Gram-positive bacterium usually involved in food spoilage and frequently isolated from late blowing cheese, is genetically indistinguishable from the proteolytic Clostridium botulinum. As the non-neurotoxic counterpart, it is often used as an exemplar for the toxic subtypes. In this work, we performed a microscopic study combined with a custom array-based analysis of the C. sporogenes cycle, from dormant spores to the early stationary phase. We identified a total of 211 transcripts in spores, validating the hypothesis that mRNAs are abundant in spores and the pattern of mRNA expression is strikingly different from that present in growing cells. The spore transcripts included genes responsible for different life-sustaining functions, suggesting there was transcript entrapment or basic poly-functional gene activation for future steps. In addition, 3 h after the beginning of the germination process, 20% of the total up-regulated genes were temporally expressed in germinating spores. The vegetative condition appeared to be more active in terms of gene transcription and protein synthesis than the spore, and genes coding for germination and sporulation factors seemed to be expressed at this point. These results suggest that spores are not silent entities, and a broader knowledge of the genetic pathways involved in the Clostridium life cycle could provide a better understanding of pathogenic clostridia types.

  13. Entérite nécrotique chez le poulet de gril II. Caractères des souches de Clostridium perfringens isolées

    PubMed Central

    Bernier, G.; Filion, R.; Malo, R.; Phaneuf, J.-B.

    1974-01-01

    A Gram positive bacillus, strictly anaerobic, was isolated from the viscera of all diseased birds showing lesions of necrotic enteritis. Its morphology and biochemical reactions, the presence of alpha and thêta hemolysins and the production of a lecithinase-C in vitro, all these characteristics indicated a similarity to those belonging to the group of Clostridium perfringens. The two hemolysins were neutralized in vitro only by the antitoxin A. Broiler chickens injected I.V. with a Viande-Foie (VF) broth culture of Clostridium perfringens together with the antitoxin A survived, whereas those receiving antitoxin C died. These results seem to indicate that this organism belongs to the type A. This bacillus was sensitive to a great variety of antibiotics, except neomycin. PMID:4368193

  14. A Clostridium Group IV Species Dominates and Suppresses a Mixed Culture Fermentation by Tolerance to Medium Chain Fatty Acids Products

    PubMed Central

    Andersen, Stephen J.; De Groof, Vicky; Khor, Way Cern; Roume, Hugo; Props, Ruben; Coma, Marta; Rabaey, Korneel

    2017-01-01

    A microbial community is engaged in a complex economy of cooperation and competition for carbon and energy. In engineered systems such as anaerobic digestion and fermentation, these relationships are exploited for conversion of a broad range of substrates into products, such as biogas, ethanol, and carboxylic acids. Medium chain fatty acids (MCFAs), for example, hexanoic acid, are valuable, energy dense microbial fermentation products, however, MCFA tend to exhibit microbial toxicity to a broad range of microorganisms at low concentrations. Here, we operated continuous mixed population MCFA fermentations on biorefinery thin stillage to investigate the community response associated with the production and toxicity of MCFA. In this study, an uncultured species from the Clostridium group IV (related to Clostridium sp. BS-1) became enriched in two independent reactors that produced hexanoic acid (up to 8.1 g L−1), octanoic acid (up to 3.2 g L−1), and trace concentrations of decanoic acid. Decanoic acid is reported here for the first time as a possible product of a Clostridium group IV species. Other significant species in the community, Lactobacillus spp. and Acetobacterium sp., generate intermediates in MCFA production, and their collapse in relative abundance resulted in an overall production decrease. A strong correlation was present between the community composition and both the hexanoic acid concentration (p = 0.026) and total volatile fatty acid concentration (p = 0.003). MCFA suppressed species related to Clostridium sp. CPB-6 and Lactobacillus spp. to a greater extent than others. The proportion of the species related to Clostridium sp. BS-1 over Clostridium sp. CPB-6 had a strong correlation with the concentration of octanoic acid (p = 0.003). The dominance of this species and the increase in MCFA resulted in an overall toxic effect on the mixed community, most significantly on the Lactobacillus spp., which resulted in a decrease in total

  15. Characteristics of hydrogen and methane production from cornstalks by an augmented two- or three-stage anaerobic fermentation process.

    PubMed

    Lu, Yuan; Lai, Qiheng; Zhang, Chong; Zhao, Hongxin; Ma, Kun; Zhao, Xuebing; Chen, Hongzhang; Liu, Dehua; Xing, Xin-Hui

    2009-06-01

    This paper presents the co-production of hydrogen and methane from cornstalks by a two- or three-stage anaerobic fermentation process augmented with effective artificial microbial community. Two-stage fermentation by using the anaerobic sludge and DGGE analysis showed that effective and stable strains should be introduced into the system. We introduced Enterobacter aerogens or Clostridium paraputrificum into the hydrogen stage, and C. paraputrificum was proven to be more effective. In the three-stage process consisting of the improved hydrolysis, hydrogen and methane production stages, the highest soluble sugars (0.482 kg/kg cornstalks) were obtained after the introduction of Clostridium thermocellum in the hydrolysis stage, under the thermophilic (55 degrees C) and acidic (pH 5.0) conditions. Hydrolysates from 1 kg of cornstalks could produce 2.61 mol (63.7 l) hydrogen by augmentation with C. paraputrificum and 4.69 mol (114.6 l) methane by anaerobic granular sludge, corresponding to 54.1% energy recovery.

  16. Dance--Aerobic and Anaerobic.

    ERIC Educational Resources Information Center

    Cohen, Arlette

    1984-01-01

    This article defines and explains aerobic exercise and its effects on the cardiovascular system. Various studies on dancers are cited indicating that dance is an anaerobic activity with some small degree of aerobic benefit. (DF)

  17. Implementing Livestock Anaerobic Digestion Projects

    EPA Pesticide Factsheets

    Page provides information to help make an informed decision about installing an anaerobic digester. Is it a good match for a farm’s organic waste, project financing, development guidelines and permit requirements?

  18. Saccharofermentans acetigenes gen. nov., sp. nov., an anaerobic bacterium isolated from sludge treating brewery wastewater.

    PubMed

    Chen, Shuangya; Niu, Lili; Zhang, Yongxiang

    2010-12-01

    A spore-forming anaerobic bacterium, designated strain P6(T), was isolated from the sludge of an up-flow anaerobic sludge blanket reactor treating brewery wastewater. Cells were Gram-positive, oval and 0.6-0.9 μm by 1.2-1.8 μm in size. Growth was observed at 20-42 °C and at pH 5.0-7.5. It fermented several hexoses, polysaccharides and alcohols. Sucrose and aesculin could also be fermented. The main end products of fermentation from glucose were acetate, lactate and fumarate; trace CO(2) and H(2) were also produced. The DNA G+C content of strain P6(T) was 55.6 mol%. The major cellular fatty acids were iso-C(15 : 0), anteiso-C(15 : 0) and iso-C(14 : 0) 3-OH. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain P6(T) represented a novel phyletic sublineage in clostridial cluster III, and showed <91 % similarity to the type strains of recognized species in this cluster. Phenotypically, the new isolate was distinguished from its phylogenetic relatives (e.g. Clostridium straminisolvens, Clostridium thermocellum, Acetivibrio cellulolyticus and Clostridium aldrichii) by producing acid from glucose and its inability to degrade cellulose. On the basis of evidence from this polyphasic study, strain P6(T) is considered to represent a novel species of a new genus, for which the name Saccharofermentans acetigenes gen. nov., sp. nov. is proposed. The type strain of Saccharofermentans acetigenes is P6(T) (=JCM 14006(T) =AS 1.5064(T)).

  19. Immunization strategies for Clostridium difficile infections.

    PubMed

    Rebeaud, Fabien; Bachmann, Martin F

    2012-04-01

    Clostridium difficile infection is a major cause of nosocomial disease in Western countries. The recent emergence of hypervirulent strains resistant to most antibiotics correlates with increasing disease incidence, severity and lethal outcomes. Current treatments rely on metronidazol and vancomycin, but the limited ability of these antibiotics to cure infection and prevent relapse highlights the need for new strategies. A better knowledge of the molecular mechanisms of the disease, the host immune response and identification of key virulence factors of Clostridium difficile now permits the development of new products specifically targeting the pathogen. Immune-based strategies relying on active vaccination or passive administration of antibody products are the focus of intense research and, today, the efficacy of monoclonal antibodies and of two vaccines are evaluated clinically. This review presents recent data, discusses the different strategies and highlights the challenges linked to the development of immunization strategies against this emerging threat.

  20. Genetic Analysis of Nitroaromatic Degradation by Clostridium

    DTIC Science & Technology

    2013-07-30

    Phenazine , a molecule produced by some soil bacteria was found to have a significant effect on metabolite pattern in two clostridium test strains...potential Effect on butyrate levels Methylene blue +0.011 no phenazine -1-carboxylic acid -0.116 More butyrate TNT -0.253 More butyrate Neutral red -0.325...interesting in light of the analysis of natural mobile soluble electron carriers in natural soil ecosystems where molecules such as quinones, and phenazines

  1. The anticancer drug tirapazamine has antimicrobial activity against Escherichia coli, Staphylococcus aureus and Clostridium difficile.

    PubMed

    Shah, Zarna; Mahbuba, Raya; Turcotte, Bernard

    2013-10-01

    Rapidly increasing bacterial resistance to existing therapies creates an urgent need for the development of new antibacterials. Tirapazamine (TPZ, 3-amino-1,2,4-benzotriazine 1,4 dioxide) is a prodrug undergoing clinical trials for various types of cancers. In this study, we showed that TPZ has antibacterial activity, particularly at low oxygen levels. With Escherichia coli, TPZ was bactericidal under both aerobic and anaerobic conditions. Escherichia coli mutants deficient in homologous recombination were hypersusceptible to TPZ, suggesting that drug toxicity may be due to DNA damage. Moreover, E. coli strains deleted for genes encoding putative reductases were resistant to TPZ, implying that these enzymes are responsible for conversion of the prodrug to a toxic compound. Fluoroquinolone-resistant E. coli strains were as susceptible to TPZ as a wild-type strain. Methicillin-resistant Staphylococcus aureus strains were also susceptible to TPZ (MIC = 0.5 μg mL(-1) ), as were pathogenic strains of Clostridium difficile (MIC = 7.5 ng mL(-1) ). TPZ may merit additional study as a broad-spectrum antibacterial, particularly for anaerobes.

  2. Clostridium septicum Gas Gangrene in Colon Cancer: Importance of Early Diagnosis.

    PubMed

    Nanjappa, Sowmya; Shah, Sweta; Pabbathi, Smitha

    2015-01-01

    The Clostridia species are responsible for some of the deadliest diseases including gas gangrene, tetanus, and botulism. Clostridium septicum is a rare subgroup known to cause atraumatic myonecrosis and is associated with colonic malignancy or immunosuppression. It is a Gram-positive, anaerobic, spore-forming bacillus found in the gastrointestinal tract and can lead to direct, spontaneous infections of the bowel and peritoneal cavity. The anaerobic glycolysis of the tumor produces an acidic, hypoxic environment favoring germination of clostridial spores. Tumor-induced mucosal ulceration allows for translocation of sporulated bacteria from the bowel into the bloodstream, leading to fulminant sepsis. C. septicum bacteremia can have a variable presentation and is associated with greater than 60% mortality rate. The majority of deaths occur within the first 24 hours if diagnosis and appropriate treatment measures are not promptly started. We report a case of abdominal myonecrosis in a patient with newly diagnosed colon cancer. The aim of this study is to stress the importance of maintaining a high suspicion of C. septicum infection in patients with underlying colonic malignancy.

  3. Identification, Isolation and characterization of a novel azoreductase from Clostridium perfringens.

    PubMed

    Morrison, Jessica M; Wright, Cristee M; John, Gilbert H

    2012-04-01

    Azo dyes are used widely in the textile, pharmaceutical, cosmetic and food industries as colorants and are often sources of environmental pollution. There are many microorganisms that are able to reduce azo dyes by use of an azoreductase enzyme. It is through the reduction of the azo bonds of the dyes that carcinogenic metabolites are produced thereby a concern for human health. The field of research on azoreductases is growing, but there is very little information available on azoreductases from strict anaerobic bacteria. In this study, the azoreductase gene was identified in Clostridium perfringens, a pathogen that is commonly found in the human intestinal tract. C. perfringens shows high azoreductase activity, especially in the presence of the common dye Direct Blue 15. A gene that encodes for a flavoprotein was isolated and expressed in Escherichia coli, and further purified and tested for azoreductase activity. The azoreductase (known as AzoC) was characterized by enzymatic reaction assays using different dyes. AzoC activity was highest in the presence of two cofactors, NADH and FAD. A strong cofactor effect was shown with some dyes, as dye reduction occurred without the presence of the AzoC (cofactors alone). AzoC was shown to perform best at a pH of 9, at room temperature, and in an anaerobic environment. Enzyme kinetics studies suggested that the association between enzyme and substrate is strong. Our results show that AzoC from C. perfringens has azoreductase activity.

  4. Characterization of a Unique Class C Acid Phosphatase from Clostridium perfringens▿

    PubMed Central

    Reilly, Thomas J.; Chance, Deborah L.; Calcutt, Michael J.; Tanner, John J.; Felts, Richard L.; Waller, Stephen C.; Henzl, Michael T.; Mawhinney, Thomas P.; Ganjam, Irene K.; Fales, William H.

    2009-01-01

    Clostridium perfringens is a gram-positive anaerobe and a pathogen of medical importance. The detection of acid phosphatase activity is a powerful diagnostic indicator of the presence of C. perfringens among anaerobic isolates; however, characterization of the enzyme has not previously been reported. Provided here are details of the characterization of a soluble recombinant form of this cell-associated enzyme. The denatured enzyme was ∼31 kDa and a homodimer in solution. It catalyzed the hydrolysis of several substrates, including para-nitrophenyl phosphate, 4-methylumbelliferyl phosphate, and 3′ and 5′ nucleoside monophosphates at pH 6. Calculated Kms ranged from 0.2 to 0.6 mM with maximum velocity ranging from 0.8 to 1.6 μmol of Pi/s/mg. Activity was enhanced in the presence of some divalent cations but diminished in the presence of others. Wild-type enzyme was detected in all clinical C. perfringens isolates tested and found to be cell associated. The described enzyme belongs to nonspecific acid phosphatase class C but is devoid of lipid modification commonly attributed to this class. PMID:19363079

  5. Isolation and characterization of an Fe,-S8 ferredoxin (ferredoxin II) from Clostridium thermoaceticum.

    PubMed Central

    Elliott, J I; Ljungdahl, L G

    1982-01-01

    A second ferredoxin protein was isolated from the thermophilic anaerobic bacterium Clostridium thermoaceticum and termed ferredoxin II. This ferredoxin was found to contain 7.9 +/- 0.3 iron atoms and 7.4 +/- 0.4 acid-labile sulfur atoms per mol of protein. Extrusion studies of the iron-sulfur centers showed the presence of two [Fe4-S4] centers per mol of protein and accounted for all of the iron present. The absorption spectrum was characterized by maxima at 390 nm (epsilon 390 = 30,400 M-1cm-1) and 280 nm (epsilon 280 = 41.400 M-1 cm-1) and by a shoulder at 300 nm. The ration of the absorbance of the pure protein at 390 nm to the absorbance at 280 nm was 0.74. Electron paramagnetic resonance data showed a weak signal in the oxidized state, and the reduced ferredoxin exhibited a spectrum typical of [Fe4-S4] clusters. Double integration of the reduced spectra showed that two electrons were necessary for the complete reduction of ferredoxin II. Amino histidine, and 1 arginine, and a molecular weight of 6,748 for the native protein. The ferredoxin is stable under anaerobic conditions for 60 min at 70 degrees C. The average oxidation-reduction potential for the two [Fe4-S4] centers was measured as -365 mV. PMID:6282807

  6. Development of a cyclic voltammetry method for the detection of Clostridium novyi in black disease.

    PubMed

    Liu, L L; Jiang, D N; Xiang, G M; Liu, C; Yu, J C; Pu, X Y

    2014-03-17

    Black disease is an acute disease of sheep and cattle. The pathogen is the obligate anaerobe, Clostridium novyi. Due to difficulties of anaerobic culturing in the country or disaster sites, a simple, rapid, and sensitive method is required. In this study, an electrochemical method, the cyclic voltammetry method, basing on loop-mediated isothermal amplification (LAMP), electrochemical ion bonding (positive dye, methylene blue), was introduced. DNA extracted from C. novyi specimens was amplified through the LAMP reaction. Then the products combined were with methylene blue, which lead to a reduction in the oxidation peak current (ipA) and the reduction peak current (ipC) of the cyclic voltammetry. The changes of ipA/ipC were real-time measured by special designed electrode, so the DNA was quantitatively detected. The results displayed that this electrochemical detection of C. novyi could be completed in 1-2 h with the lowest bacterial concentration of 10(2) colony forming units/mL, and high accuracy (96.5%), sensitivity (96%), and specificity (97%) compared to polymerase chain reation. The cyclic voltammetry method was a simple and fast method, with high sensitivity and high specificity, and has great potential to be a usable molecular tool for fast diagnosis of Black disease.

  7. Genome shuffling in Clostridium diolis DSM 15410 for improved 1,3-propanediol production.

    PubMed

    Otte, Burkhard; Grunwaldt, Eike; Mahmoud, Osama; Jennewein, Stefan

    2009-12-01

    Several microorganisms are known for their efficient anaerobic conversion of glycerol to 1,3-propanediol, with Clostridium diolis DSM 15410 as one of the better performers in terms of molar yield and volumetric productivity. However, this performance is still insufficient to compete with established chemical processes. Previous studies have shown that high concentrations of 1,3-propanediol, glycerol, and fermentation side products can limit the productivity of C. diolis DSM 15410. Here, we describe the use of genome shuffling for improved 1,3-propanediol fermentation by the strict anaerobe C. diolis DSM 15410. By using chemical mutagenesis, strains with superior substrate and product tolerance levels were isolated and higher product yields were obtained. These superior strains were then used for genome shuffling and selection for 1,3-propanediol and organic acid tolerance. After four rounds of genome shuffling and selection, significant improvements were observed, with one strain attaining a 1,3-propanediol volumetric yield of 85 g/liter. This result represents an 80% improvement compared to the yield from the parental wild-type strain.

  8. A phosphoethanolamine-modified glycosyl diradylglycerol in the polar lipids of Clostridium tetani.

    PubMed

    Johnston, Norah C; Aygun-Sunar, Semra; Guan, Ziqiang; Ribeiro, Anthony A; Daldal, Fevzi; Raetz, Christian R H; Goldfine, Howard

    2010-07-01

    The polar lipids of the anaerobic bacterium Clostridium tetani, the causative agent of tetanus, have been examined by two-dimensional thin layer chromatography, ESI mass spectrometry, and NMR spectroscopy. Plasmalogen and di- and tetra-acylated species of phosphatidylethanolamine, phosphatidylglycerol, cardiolipin, and N-acetylglucosaminyl diradylglycerol were the major lipids present in most strains examined except for strain ATCC 10779, the parent of strain E88, the first C. tetani strain to have its genome sequenced. This strain contained the same di- and tetra-acylated species but did not contain plasmalogens. All strains contained a novel derivative of N-acetylglucosaminyl diradylglycerol in which a phosphoethanolamine unit is attached to the 6'-position of the sugar, as judged by selective 31P-decoupled, 1H-detected NMR difference spectroscopy. The N-acetylglucosamine (GlcNAc) residue is presumably linked to the 3-positon of the diradylglycerol moiety, and it has the beta-anomeric configuration. Very little plasmalogen component was detected by mass spectrometry in the precursors phosphatidic acid and phosphatidylserine, consistent with the idea that plasmalogens are formed from diacylated phospholipids at a late stage of phospholipid assembly in anaerobic clostridia.

  9. Rapid, sensitive, and specific detection of Clostridium tetani by loop-mediated isothermal amplification assay.

    PubMed

    Jiang, Dongneng; Pu, Xiaoyun; Wu, Jiehong; Li, Meng; Liu, Ping

    2013-01-01

    Tetanus is a specific infectious disease, which is often associated with catastrophic events such as earthquakes, traumas, and war wounds. The obligate anaerobe Clostridium tetani is the pathogen that causes tetanus. Once the infection of tetanus progresses to an advanced stage within the wounds of limbs, the rates of amputation and mortality increase manifold. Therefore, it is necessary to devise a rapid and sensitive point-of-care detection method for C. tetani so as to ensure an early diagnosis and clinical treatment of tetanus. In this study, we developed a detection method for C. tetani using loop-mediated isothermal amplification (LAMP) assay, wherein the C. tetani tetanus toxin gene was used as the target gene. The method was highly specific and sensitive, with a detection limit of 10 colony forming units (CFU)/ml, and allowed quantitative analysis. While detecting C. tetani in clinical samples, it was found that the LAMP results completely agreed with those of the traditional API 20A anaerobic bacteria identification test. As compared with the traditional API test and PCR assay, LAMP detection of C. tetani is simple and rapid, and the results can be identified through naked-eye observation. Therefore, it is an ideal and rapid point-of-care testing method for tetanus.

  10. Improving the performance of solventogenic clostridia by reinforcing the biotin synthetic pathway.

    PubMed

    Yang, Yunpeng; Lang, Nannan; Yang, Gaohua; Yang, Sheng; Jiang, Weihong; Gu, Yang

    2016-05-01

    An efficient production process is important for industrial microorganisms. The cellular efficiency of solventogenic clostridia, a group of anaerobes capable of producing a wealth of bulk chemicals and biofuels, must be improved for competitive commercialization. Here, using Clostridium acetobutylicum, a species of solventogenic clostridia, we revealed that the insufficient biosynthesis of biotin, a pivotal coenzyme for many important biological processes, is a major limiting bottleneck in this anaerobe's performance. To address this problem, we strengthened the biotin synthesis of C. acetobutylicum by overexpressing four relevant genes involved in biotin transport and biosynthesis. This strategy led to faster growth and improved the titer and productivity of acetone, butanol and ethanol (ABE solvents) of C. acetobutylicum in both biotin-containing and biotin-free media. Expressionally modulating these four genes by modifying the ribosome binding site further promoted cellular performance, achieving ABE solvent titer and productivity as high as 21.9g/L and 0.30g/L/h, respectively, in biotin-free medium; these values exceeded those of the wild-type strain by over 30%. More importantly, biotin synthesis reinforcement also conferred improved ability of C. acetobutylicum to use hexose and pentose sugars, further demonstrating the potential of this metabolic-engineering strategy in solventogenic clostridia.

  11. Mechanisms of Antibacterial Action of Quinoxaline 1,4-di-N-oxides against Clostridium perfringens and Brachyspira hyodysenteriae

    PubMed Central

    Xu, Fanfan; Cheng, Guyue; Hao, Haihong; Wang, Yulian; Wang, Xu; Chen, Dongmei; Peng, Dapeng; Liu, Zhenli; Yuan, Zonghui; Dai, Menghong

    2016-01-01

    Quinoxaline 1,4-di-N-oxides (QdNOs) are a class of bioreductive compounds, however, their antibacterial mechanisms are still unclarified. The aim of this study was to assess the ability of two representative QdNO drugs, cyadox (CYA) and olaquindox (OLA), to produce reactive oxide species (ROS) in Gram-positive anaerobe Clostridium perfringens CVCC1125 and Gram-negative anaerobe Brachyspira hyodysenteriae B204. In addition, the effects of QdNOs on the integrity of bacterial cell walls and membranes as well as the morphological alterations and DNA oxidative damage in C. perfringens and B. hyodysenteriae were analyzed. It was demonstrated that under anaerobic conditions, QdNOs were metabolized into the reduced products which did not show any antibacterial activity. A significant dose-related increase of intracellular ROS level and intracellular hydroxyl radicals were evident in bacteria exposed to QdNOs. The result of biochemical assay showed that the cell walls and membranes of the bacteria treated with QdNOs were damaged. After exposure to 1/2MIC to 4MIC of CYA and OLA, C. perfringens and B. hyodysenteriae became elongated and filamentous. Morphological observation with scanning and transmission electron microscopes revealed rupture, loss of cytoplasmic material and cell lysis in QdNO-treated bacteria, indicating serious damage of cells. There was an increase of 8-OHdG in the two strains treated by QdNOs, but it was lower in C. perfringens CVCC1125 than in B. hyodysenteriae B204. Agarose gel electrophoresis showed the degradation of chromosomal DNA in both of the two anaerobes treated by QdNOs. The results suggest that QdNOs may kill C. perfringens and B. hyodysenteriae via the generation of ROS and hydroxyl radicals from the bacterial metabolism of QdNOs, which cause oxidative damage in bacteria under anaerobic conditions. PMID:28018297

  12. Microbial communities mediating algal detritus turnover under anaerobic conditions

    PubMed Central

    Morrison, Jessica M.; Murphy, Chelsea L.; Baker, Kristina; Zamor, Richard M.; Nikolai, Steve J.; Wilder, Shawn; Elshahed, Mostafa S.

    2017-01-01

    Background Algae encompass a wide array of photosynthetic organisms that are ubiquitously distributed in aquatic and terrestrial habitats. Algal species often bloom in aquatic ecosystems, providing a significant autochthonous carbon input to the deeper anoxic layers in stratified water bodies. In addition, various algal species have been touted as promising candidates for anaerobic biogas production from biomass. Surprisingly, in spite of its ecological and economic relevance, the microbial community involved in algal detritus turnover under anaerobic conditions remains largely unexplored. Results Here, we characterized the microbial communities mediating the degradation of Chlorella vulgaris (Chlorophyta), Chara sp. strain IWP1 (Charophyceae), and kelp Ascophyllum nodosum (phylum Phaeophyceae), using sediments from an anaerobic spring (Zodlteone spring, OK; ZDT), sludge from a secondary digester in a local wastewater treatment plant (Stillwater, OK; WWT), and deeper anoxic layers from a seasonally stratified lake (Grand Lake O’ the Cherokees, OK; GL) as inoculum sources. Within all enrichments, the majority of algal biomass was metabolized within 13–16 weeks, and the process was accompanied by an increase in cell numbers and a decrease in community diversity. Community surveys based on the V4 region of the 16S rRNA gene identified different lineages belonging to the phyla Bacteroidetes, Proteobacteria (alpha, delta, gamma, and epsilon classes), Spirochaetes, and Firmicutes that were selectively abundant under various substrate and inoculum conditions. Within all kelp enrichments, the microbial communities structures at the conclusion of the experiment were highly similar regardless of the enrichment source, and were dominated by the genus Clostridium, or family Veillonellaceae within the Firmicutes. In all other enrichments the final microbial community was dependent on the inoculum source, rather than the type of algae utilized as substrate. Lineages enriched

  13. Nitrogen-gas bubbling during the cultivation of Clostridium tetani produces a higher yield of tetanus toxin for the preparation of its toxoid.

    PubMed

    De Luca, M M; Abeiro, H D; Bernagozzi, J A; Basualdo, J A

    1997-01-01

    We investigated the effect of exposing cultures of Clostridium tetani to nitrogen (N2) gas on the recovery of tetanus toxin to be processed for the preparation of its toxoid. N2 was bubbled through nine 10-liter cultures during the growth of the bacteria, while nine parallel control incubations were maintained without bubbling. We found that treatment of the C. tetani anaerobes with an inert gas in this manner during cultivation produced a highly significant increase in the yield of tetanus toxin from them in comparison with the standard procedure.

  14. Fecal Microbiota Transplantation for Clostridium difficile-Associated Diarrhea.

    PubMed

    Cohen, Nathaniel A; Ben Ami, Ronen; Guzner-Gur, Hanan; Santo, Moshe E; Halpern, Zamir; Maharshak, Nitsan

    2015-08-01

    Clostridium difficile-associated diarrhea is a problem most hospital-based physicians will face in their career. This review aims to refresh current knowledge with regard to Clostridium difficile infection and bring physicians up to date with the latest developments in the growing field of fecal microbiota transplantation, the benefits it offers, and the promise this and other developments hold for the future.

  15. Anaerobic benzene degradation by bacteria

    PubMed Central

    Vogt, Carsten; Kleinsteuber, Sabine; Richnow, Hans‐Hermann

    2011-01-01

    Summary Benzene is a widespread and toxic contaminant. The fate of benzene in contaminated aquifers seems to be primarily controlled by the abundance of oxygen: benzene is aerobically degraded at high rates by ubiquitous microorganisms, and the oxygen‐dependent pathways for its breakdown were elucidated more than 50 years ago. In contrast, benzene was thought to be persistent under anoxic conditions until 25 years ago. Nevertheless, within the last 15 years, several benzene‐degrading cultures have been enriched under varying electron acceptor conditions in laboratories around the world, and organisms involved in anaerobic benzene degradation have been identified, indicating that anaerobic benzene degradation is a relevant environmental process. However, only a few benzene degraders have been isolated in pure culture so far, and they all use nitrate as an electron acceptor. In some highly enriched strictly anaerobic cultures, benzene has been described to be mineralized cooperatively by two or more different organisms. Despite great efforts, the biochemical mechanism by which the aromatic ring of benzene is activated in the absence of oxygen is still not fully elucidated; methylation, hydroxylation and carboxylation are discussed as likely reactions. This review summarizes the current knowledge about the ‘key players’ of anaerobic benzene degradation under different electron acceptor conditions and the possible pathway(s) of anaerobic benzene degradation. PMID:21450012

  16. Development of a microarray for identification of pathogenic Clostridium species

    PubMed Central

    Janvilisri, Tavan; Scaria, Joy; Gleed, Robin; Fubini, Susan; Bonkosky, Michelle M.; Gröhn, Yrjö T.; Chang, Yung-Fu

    2009-01-01

    In recent years, Clostridium species have rapidly reemerged as human and animal pathogens. The detection and identification of pathogenic Clostridium species is therefore critical for clinical diagnosis and antimicrobial therapy. Traditional diagnostic techniques for clostridia are laborious, time-consuming and may adversely affect the therapeutic outcome. In this study, we developed an oligonucleotide diagnostic microarray for pathogenic Clostridium species. The microarray specificity was tested against 65 Clostridium isolates. The applicability of this microarray in a clinical setting was assessed with the use of mock stool samples. The microarray was successful in discriminating at least four species with the limit of detection as low as 104 CFU/ml. In addition, the pattern of virulence and antibiotic resistance genes of tested strains were determined through the microarrays. This approach demonstrates the high-throughput detection and identification of Clostridium species and provides advantages over traditional methods. Microarray-based techniques are promising applications for clinical diagnosis and epidemiological investigations. PMID:19879710

  17. [Spontaneous gas gangrene in a diabetic patient with Clostridium septicum].

    PubMed

    Mischke, A; Besier, S; Walcher, F; Waibel, H; Brade, V; Brandt, C

    2005-10-01

    Atraumatic infections due to Clostridium septicum are known to be associated with immunosuppression or even malignancy. In this case report, we present a patient with severe Clostridium septicum infection related to advanced colon cancer that had not previously been diagnosed. The case demonstrates the strong association between Clostridium septicum infections and malignancy, particularly in the presence of other predisposing diseases such as diabetes mellitus. It strongly suggests excluding malignant neoplasms, especially of the gastrointestinal tract, when severe Clostridium septicum infections occur. Moreover, if patients with known colorectal or other malignancy develop septicaemia or spontaneous gas gangrene, clinicians should be aware of Clostridium septicum as one of the main causative agents, as early diagnosis and aggressive treatment are important to improve prognosis.

  18. Plasmidome Interchange between Clostridium botulinum, Clostridium novyi and Clostridium haemolyticum Converts Strains of Independent Lineages into Distinctly Different Pathogens

    PubMed Central

    Skarin, Hanna; Segerman, Bo

    2014-01-01

    Clostridium botulinum (group III), Clostridium novyi and Clostridium haemolyticum are well-known pathogens causing animal botulism, gas gangrene/black disease, and bacillary hemoglobinuria, respectively. A close genetic relationship exists between the species, which has resulted in the collective term C. novyi sensu lato. The pathogenic traits in these species, e.g., the botulinum neurotoxin and the novyi alpha toxin, are mainly linked to a large plasmidome consisting of plasmids and circular prophages. The plasmidome of C. novyi sensu lato has so far been poorly characterized. In this study we explored the genomic relationship of a wide range of strains of C. novyi sensu lato with a special focus on the dynamics of the plasmidome. Twenty-four genomes were sequenced from strains selected to represent as much as possible the genetic diversity in C. novyi sensu lato. Sixty-one plasmids were identified in these genomes and 28 of them were completed. The genomic comparisons revealed four separate lineages, which did not strictly correlate with the species designations. The plasmids were categorized into 13 different plasmid groups on the basis of their similarity and conservation of plasmid replication or partitioning genes. The plasmid groups, lineages and species were to a large extent entwined because plasmids and toxin genes had moved across the lineage boundaries. This dynamic process appears to be primarily driven by phages. We here present a comprehensive characterization of the complex species group C. novyi sensu lato, explaining the intermixed genetic properties. This study also provides examples how the reorganization of the botulinum toxin and the novyi alpha toxin genes within the plasmidome has affected the pathogenesis of the strains. PMID:25254374

  19. Plasmidome interchange between Clostridium botulinum, Clostridium novyi and Clostridium haemolyticum converts strains of independent lineages into distinctly different pathogens.

    PubMed

    Skarin, Hanna; Segerman, Bo

    2014-01-01

    Clostridium botulinum (group III), Clostridium novyi and Clostridium haemolyticum are well-known pathogens causing animal botulism, gas gangrene/black disease, and bacillary hemoglobinuria, respectively. A close genetic relationship exists between the species, which has resulted in the collective term C. novyi sensu lato. The pathogenic traits in these species, e.g., the botulinum neurotoxin and the novyi alpha toxin, are mainly linked to a large plasmidome consisting of plasmids and circular prophages. The plasmidome of C. novyi sensu lato has so far been poorly characterized. In this study we explored the genomic relationship of a wide range of strains of C. novyi sensu lato with a special focus on the dynamics of the plasmidome. Twenty-four genomes were sequenced from strains selected to represent as much as possible the genetic diversity in C. novyi sensu lato. Sixty-one plasmids were identified in these genomes and 28 of them were completed. The genomic comparisons revealed four separate lineages, which did not strictly correlate with the species designations. The plasmids were categorized into 13 different plasmid groups on the basis of their similarity and conservation of plasmid replication or partitioning genes. The plasmid groups, lineages and species were to a large extent entwined because plasmids and toxin genes had moved across the lineage boundaries. This dynamic process appears to be primarily driven by phages. We here present a comprehensive characterization of the complex species group C. novyi sensu lato, explaining the intermixed genetic properties. This study also provides examples how the reorganization of the botulinum toxin and the novyi alpha toxin genes within the plasmidome has affected the pathogenesis of the strains.

  20. An Atypical Clostridium Strain Related to the Clostridium botulinum Group III Strain Isolated from a Human Blood Culture

    PubMed Central

    Ruimy, Raymond; Bouchier, Christiane; Faucher, Nathalie; Mazuet, Christelle; Popoff, Michel R.

    2014-01-01

    A nontoxigenic strain isolated from a fatal human case of bacterial sepsis was identified as a Clostridium strain from Clostridium botulinum group III, based on the phenotypic characters and 16S rRNA gene sequence, and was found to be related to the mosaic C. botulinum D/C strain according to a multilocus sequence analysis of 5 housekeeping genes. PMID:24088855

  1. An atypical Clostridium strain related to the Clostridium botulinum group III strain isolated from a human blood culture.

    PubMed

    Bouvet, Philippe; Ruimy, Raymond; Bouchier, Christiane; Faucher, Nathalie; Mazuet, Christelle; Popoff, Michel R

    2014-01-01

    A nontoxigenic strain isolated from a fatal human case of bacterial sepsis was identified as a Clostridium strain from Clostridium botulinum group III, based on the phenotypic characters and 16S rRNA gene sequence, and was found to be related to the mosaic C. botulinum D/C strain according to a multilocus sequence analysis of 5 housekeeping genes.

  2. Factors affecting growth and toxin production by Clostridium botulinum type E on irradiated (0. 3 Mrad) chicken skins

    SciTech Connect

    Firstenberg-Eden, R.; Rowley, D.B.; Shattuck, G.E.

    1982-05-01

    A model system (chicken skins with chicken exudate) was used to determine if Clostridium botulinum type E (Beluga) spores, stressed by low dose irradiation, would develop and produce toxin at abuse temperatures of 10 and 30/sup 0/C in the absence of characteristic spoilage. Unstressed spores germinated, multiplied, and produced toxin on vacuum-packed chicken skins, stored at either 30 or 10/sup 0/C. Cell numbers increased faster and toxin was evident sooner at 30/sup 0/C than at 10/sup 0/C. At 30/sup 0/C, growth occurred and toxin was produced more slowly when samples were incubated aerobically than anaerobically. When samples were incubated aerobically at 10/sup 0/C, no toxin was detected within a test period of 14 days. An irradiation dose of 0.3 Mrad at 5/sup 0/C reduced a spore population on vacuum-sealed chicken skins by about 90%. The surviving population produced toxin at 30/sup 0/C under either aerobic or anaerobic conditions, at 10/sup 0/C no toxin was detected even on skins incubated anaerobically. Under the worst conditions (30/sup 0/C, vacuum packed) toxin was not detected prior to characteristic spoilage caused by the natural flora surviving 0.3 Mrad.

  3. The Transition from Aerobic to Anaerobic Metabolism.

    ERIC Educational Resources Information Center

    Skinner, James S.; McLellan, Thomas H.

    1980-01-01

    The transition from aerobic to anaerobic metabolism is discussed. More research is needed on different kinds of athletes and athletic activities and how they may affect aerobic and anaerobic metabolisms. (CJ)

  4. Arsenic, Anaerobes, and Astrobiology

    NASA Astrophysics Data System (ADS)

    Stolz, J. F.; Oremland, R. S.; Switzer Blum, J.; Hoeft, S. E.; Baesman, S. M.; Bennett, S.; Miller, L. G.; Kulp, T. R.; Saltikov, C.

    2013-12-01

    Arsenic is an element best known for its highly poisonous nature, so it is not something one would associate with being a well-spring for life. Yet discoveries made over the past two decades have delineated that not only are some microbes resistant to arsenic, but that this element's primary redox states can be exploited to conserve energy and support prokaryotic growth ('arsenotrophy') in the absence of oxygen. Hence, arsenite [As(III)] can serve as an electron donor for chemo- or photo-autotrophy while arsenate [As(V)] will serve as an electron acceptor for chemo-heterotrophs and chemo-autotrophs. The phylogenetic diversity of these microbes is broad, encompassing many individual species from diverse taxonomic groups in the Domain Bacteria, with fewer representatives in the Domain Archaea. Speculation with regard to the evolutionary origins of the key functional genes in anaerobic arsenic transformations (arrA and arxA) and aerobic oxidation (aioB) has led to a disputation as to which gene and function is the most ancient and whether arsenic metabolism extended back into the Archaean. Regardless of its origin, robust arsenic metabolism has been documented in extreme environments that are rich in their arsenic content, such as hot springs and especially hypersaline soda lakes associated with volcanic regions. Searles Lake, CA is an extreme, salt-saturated end member where vigorous arsenic metabolism occurs, but there is no detectable sulfate-reduction or methanogenesis. The latter processes are too weak bio-energetically to survive as compared with arsenotrophy, and are also highly sensitive to the abundance of borate ions present in these locales. These observations have implications with respect to the search for microbial life elsewhere in the Solar System where volcanic-like processes have been operative. Hence, because of the likelihood of encountering dense brines in the regolith of Mars (formed by evapo-concentration) or beneath the ice layers of Europa

  5. Anaerobic bioprocessing of organic wastes.

    PubMed

    Verstraete, W; de Beer, D; Pena, M; Lettinga, G; Lens, P

    1996-05-01

    Anaerobic digestion of dissolved, suspended and solid organics has rapidly evolved in the last decades but nevertheless still faces several scientific unknowns. In this review, some fundamentals of bacterial conversions and adhesion are addressed initially. It is argued in the light of ΔG-values of reactions, and in view of the minimum energy quantum per mol, that anaerobic syntrophs must have special survival strategies in order to support their existence: redistributing the available energy between the partners, reduced end-product fermentation reactions and special cell-to-cell physiological interactions. In terms of kinetics, it appears that both reaction rates and residual substrate thresholds are strongly related to minimum ΔG-values. These new fundamental insights open perspectives for efficient design and operation of anaerobic bioprocesses. Subsequently, an overview is given of the current anaerobic biotechnology. For treating wastewaters, a novel and high performance new system has been introduced during the last decade; the upflow anaerobic sludge blanket system (UASB). This reactor concept requires anaerobic consortia to grow in a dense and eco-physiologically well-organized way. The microbial principles of such granular sludge growth are presented. Using a thermodynamic approach, the formation of different types of aggregates is explained. The application of this bioprocess in worldwide wastewater treatment is indicated. Due to the long retention times of the active biomass, the UASB is also suitable for the development of bacterial consortia capable of degrading xenobiotics. Operating granular sludge reactors at high upflow velocities (5-6 m/h) in expanded granular sludge bed (EGSB) systems enlarges the application field to very low strength wastewaters (chemical oxygen demand < 1 g/l) and psychrophilic temperatures (10°C). For the treatment of organic suspensions, there is currently a tendency to evolve from the conventional mesophilic

  6. How anaerobic is the Wingate Anaerobic Test for humans?

    PubMed

    Beneke, R; Pollmann, C; Bleif, I; Leithäuser, R M; Hütler, M

    2002-08-01

    The Wingate Anaerobic Test (WAnT) is generally used to evaluate anaerobic cycling performance, but knowledge of the metabolic profile of WAnT is limited. Therefore the energetics of WAnT was analysed with respect to working efficiency and performance. A group of 11 male subjects [mean (SD), age 21.6 (3.8) years, height 178.6 (6.6) cm, body mass 82.2 (12.1) kg] performed a maximal incremental exercise test and a WAnT. Lactic and alactic anaerobic energy outputs were calculated from net lactate production and the fast component of the kinetics of post-exercise oxygen uptake. Aerobic metabolism was determined from oxygen uptake during exercise. The WAnT mean power of 683 (96.0) W resulted from a total energy output above the value at rest of 128.1 (23.2) kJ x 30 s(-1) [mean metabolic power=4.3 (0.8) kW] corresponding to a working efficiency of 16.2 (1.6)%. The WAnT working efficiency was lower (P < 0.01) than the corresponding value of 24.1 (1.7)% at 362 (41) W at the end of an incremental exercise test. During WAnT the fractions of the energy from aerobic, anaerobic alactic and lactic acid metabolism were 18.6 (2.5)%, 31.1 (4.6)%, and 50.3 (5.1)%, respectively. Energy from metabolism of anaerobic lactic acid explained 83% and 81% of the variance of WAnT peak and mean power, respectively. The results indicate firstly that WAnT requires the use of more anaerobically derived energy than previously estimated, secondly that anaerobic metabolism is dominated by glycolysis, thirdly that WAnT mechanical efficiency is lower than that found in aerobic exercise tests, and fourthly that the latter finding partly explains discrepancies between previously published and the present data about the metabolic profile of WAnT.

  7. Detection and identification by PCR of Clostridium chauvoei in clinical isolates, bovine faeces and substrates from biogas plant

    PubMed Central

    Bagge, E; Lewerin, S Sternberg; Johansson, K-E

    2009-01-01

    Background Clostridium chauvoei causes blackleg, an acute disease associated with high mortality in ruminants. The apparent primary port of entry is oral, during grazing on pasture contaminated by spores. Cases of blackleg can occur year after year on contaminated pastures. A method to determine the prevalence of C. chauvoei spores on pasture would be useful. The standard method for C. chauvoei detection is culture and biochemical identification, which requires a pure culture. In most muscle samples from cattle dead from blackleg the amount of C. chauvoei in samples is high and the bacterium can easily be cultured, although some samples may be contaminated. Detection by PCR would be faster and independent of contaminating flora. Digested residues from biogas plants provide an excellent fertiliser, but it is known that spore-forming baeria such as Clostridium spp. are not reduced by pasteurisation. The use of digested residues as fertiliser may contribute to the spread of C. chauvoei. Soil, manure and substrate from biogas plants are contaminated with other anaerobic bacteria which outgrow C. chauvoei. Therefore, detection by PCR is would be useful. This study applied a PCR-based method to detect of C. chauvoei in 25 muscle and blood samples, 114 manure samples, 84 soil samples and 33 samples from the biogas process. Methods Muscle tissues from suspected cases of blackleg were analysed both by the standard culture method followed by biochemical identification and by PCR, with and without preculture. To investigate whether muscle tissue samples are necessary, samples taken by swabs were also investigated. Samples from a biogas plant and manure and soil from farms were analysed by culture followed by PCR. The farms had proven cases of blackleg. For detection of C. chauvoei in the samples, a specific PCR primer pair complementary to the spacer region of the 16S-23S rRNA gene was used. Results Clostridium chauvoei was detected in 32% of muscle samples analysed by

  8. Protective cellular antigen of Clostridium chauvoei.

    PubMed

    Stevenson, J R; Stonger, K A

    1980-04-01

    Cellular antigens of Clostridium chauvoei, strain IRP-128, were demonstrated to be important in induction of immunity against this bacterium in guinea pigs. At least one major component of the cellular antigen complex was heat-labile. Acid extraction of the bacterial cells, followed by selective purification for flagella, led to the preparation of an acid extract antigen that possessed a high degree of immunogenicity. The acid extract antigen contained flagellar components and was resolved into two major and approximately five minor protein components by polyacrylamide-gel electrophoresis.

  9. An Update on Clostridium difficile Toxinotyping

    PubMed Central

    Janezic, Sandra

    2015-01-01

    Toxinotyping is a PCR-restriction fragment length polymorphism (RFLP)-based method for differentiation of Clostridium difficile strains according to the changes in the pathogenicity locus (PaLoc), a region coding for toxins A and B. Toxinotypes are a heterogenous group of strains that are important in the development of molecular diagnostic tests and vaccines and are a good basis for C. difficile phylogenetic studies. Here we describe an overview of the 34 currently known toxinotypes (I to XXXIV) and some changes in nomenclature. PMID:26511734

  10. Chronic Clostridium botulinum infections in farmers.

    PubMed

    Rodloff, Arne C; Krüger, Monika

    2012-04-01

    Although botulism is usually an acute, often lethal disease that is caused by the ingestion of botulinum neurotoxin, there are also recognized forms like infant botulism, wound botulism, or "botulism of undefined origin" that are characterized by the fact that Clostridium botulinum colonizes the host and produces its toxin in the host. Evidence is presented here that a disease in cattle and in human care takers of diseased animals that has evolved over the past two decades, may be a chronic, visceral form of C. botulinum infection.

  11. Diagnosis of Clostridium difficile Infections in Children

    PubMed Central

    Leber, Amy L.

    2016-01-01

    The detection and diagnosis of Clostridium difficile infection in pediatric populations have some unique considerations in comparison to testing in adults. The testing methodologies, including toxigenic culture, cell cytotoxicity, antigen detection, and, more recently, molecular testing, are the same in all age groups. However, limited data exist on the specific performance characteristics in children. In this review, we focus on the challenges of testing in pediatric populations and assess the available data on test performance in these populations. Additionally, a review of the existing guidance for testing is provided. PMID:26912759

  12. An Update on Clostridium difficile Toxinotyping.

    PubMed

    Rupnik, Maja; Janezic, Sandra

    2016-01-01

    Toxinotyping is a PCR-restriction fragment length polymorphism (RFLP)-based method for differentiation of Clostridium difficile strains according to the changes in the pathogenicity locus (PaLoc), a region coding for toxins A and B. Toxinotypes are a heterogenous group of strains that are important in the development of molecular diagnostic tests and vaccines and are a good basis for C. difficile phylogenetic studies. Here we describe an overview of the 34 currently known toxinotypes (I to XXXIV) and some changes in nomenclature.

  13. Clostridium difficile infection: Updates in management.

    PubMed

    Tariq, Raseen; Khanna, Sahil

    2017-01-01

    Clostridium difficile was first identified in 1978 as a diarrhea-causing bacterium in humans. In the last three decades, C. difficile infection (CDI) has reached an epidemic state, both in health care and community settings worldwide. There has been substantial progress in the field of CDI, including identification of novel risk factors, presence of CDI in individuals not considered at risk previously, and treatment options including new drugs, monoclonal antibodies, and fecal microbiota transplantation. This review discusses epidemiology, novel and traditional risk factors, and updates in management for CDI.

  14. Clostridium difficile colitis: pathogenesis and host defence.

    PubMed

    Abt, Michael C; McKenney, Peter T; Pamer, Eric G

    2016-10-01

    Clostridium difficile is a major cause of intestinal infection and diarrhoea in individuals following antibiotic treatment. Recent studies have begun to elucidate the mechanisms that induce spore formation and germination and have determined the roles of C. difficile toxins in disease pathogenesis. Exciting progress has also been made in defining the role of the microbiome, specific commensal bacterial species and host immunity in defence against infection with C. difficile. This Review will summarize the recent discoveries and developments in our understanding of C. difficile infection and pathogenesis.

  15. [In vitro activity of roxithromycin, new semisynthetic macrolide against obligate anaerobes].

    PubMed

    Dubreuil, L; Devos, J; Romond, C; Bryskier, A

    1986-05-01

    The "in vitro" susceptibility to roxithromycin and three other macrolides of 236 anaerobes isolated from clinical samples in 1984/1985 was determined by an agar-dilution method on Wilkins Chalgren medium. 90% of Gram positive cocci were susceptible to both roxithromycin and josamycin (MIC less than 1 mg/l, whereas 1 mg/l erythromycin and 2 mg/l spiramycin were able to inhibit respectively 46 and 86% of the same tested strains. No resistance to the four macrolides was observed among Eubacterium, propionibacterium and Bifidobacterium. Two C. perfringens strains and one C. difficile strain were resistant to all four macrolides, while 97% of Clostridium sp. strains were inhibited by 4 mg/l erythromycin, josamycin or roxithromycin. Against Gram positive anaerobes, roxithromycin was equal or superior to erythromycin and spiramycin. At a concentration of 4 mg/l, roxithromycin inhibited 82% of B. fragilis strains. Roxithromycin and josamycin were more active against Gram negative bacilli that erythromycin and spiramycin. Macrolides had no effect on Fusobacterium strains. In this study, 4 mg/l roxithromycin inhibited 217 of the 236 anaerobic strains investigated (92%).

  16. [In vitro activities of sulopenem, a new parenteral penem, against anaerobes].

    PubMed

    Watanabe, K; Kato, N; Tanaka-Bandoh, K; Tanaka, Y; Kato, H; Ueno, K

    1996-04-01

    In vitro activities of sulopenem, a novel parenteral penem, was compared with those of imipenem, flomoxef, cefuzonam, cefoperazone and sulbactam/ampicillin against 66 reference strains (19 genera, 61 species) and 392 recent clinical isolates of anaerobic bacteria and fastidious aerobic bacteria. Sulopenem had a very broad spectrum against anaerobic bacteria. In general, this compound was active against anaerobic reference strains with MICs of < or = 0.78 micrograms/ml, while being the least active against Bifidobacterium spp. and less active than imipenem against Lactobacillus spp. Sulopenem was more active against Bacteroides fragilis isolates than imipenem and had the highest activities against Bacteroides thetaiotaomicron, Prevotella intermedia, Porphyromonas gingivalis, Fusobacterium spp. and Peptostreptococcus spp. among the antibiotics tested. Sulopenem was not hydrolyzed by oxyiminocephalosporinase type 1 produced by B. fragilis GAI-0558, GAI-7955 and GAI-10150 and its stability was comparable to imipenem. Its susceptibilities to hydrolysis by a metallo-beta-lactamase from B. fragilis GAI-30144 was less than imipenem. Sulopenem (120 mg/kg, 3 times a day for 4 days) was as effective as imipenem/cilastatin against a mixed intraabdominal mice infection due to E. coli and B. fragilis. Sulopenem (20 mg/kg twice a day for 5 days) did not induce an overgrowth of Clostridium difficile in the caecum of mice.

  17. Anaerobic High-Throughput Cultivation Method for Isolation of Thermophiles Using Biomass-Derived Substrates

    SciTech Connect

    Hamilton-Brehm, Scott; Vishnivetskaya, Tatiana A; Allman, Steve L; Mielenz, Jonathan R; Elkins, James G

    2012-01-01

    Flow cytometry (FCM) techniques have been developed for sorting mesophilic organisms, but the difficulty increases if the target microbes are thermophilic anaerobes. We demonstrate a reliable, high-throughput method of screening thermophilic anaerobic organisms using FCM and 96-well plates for growth on biomass-relevant substrates. The method was tested using the cellulolytic thermophiles Clostridium ther- mocellum (Topt = 55 C), Caldicellulosiruptor obsidiansis (Topt = 78 C) and the fermentative hyperthermo- philes, Pyrococcus furiosus (Topt = 100 C) and Thermotoga maritima (Topt = 80 C). Multi-well plates were incubated at various temperatures for approximately 72 120 h and then tested for growth. Positive growth resulting from single cells sorted into individual wells containing an anaerobic medium was verified by OD600. Depending on the growth substrate, up to 80 % of the wells contained viable cultures, which could be transferred to fresh media. This method was used to isolate thermophilic microbes from Rabbit Creek, Yellowstone National Park (YNP), Wyoming. Substrates for enrichment cultures including crystalline cellulose (Avicel), xylan (from Birchwood), pretreated switchgrass and Populus were used to cultivate organisms that may be of interest to lignocellulosic biofuel production.

  18. Clostridium difficile in the Military Population

    DTIC Science & Technology

    2016-08-05

    obligate anaerobic, spore forming rod bacteria , remains one of the major causes of health-care associated diarrhea1,2. From recognition in the 1970...from other bacteria ). This study had some limitations. Mild or asymptomatic cases of C. difficile may have not been tested or diagnosed. Data

  19. Genetic, phenotypic and matrix-assisted laser desorption ionization time-of-flight mass spectrometry-based identification of anaerobic bacteria and determination of their antimicrobial susceptibility at a University Hospital in Japan.

    PubMed

    Yunoki, Tomoyuki; Matsumura, Yasufumi; Nakano, Satoshi; Kato, Karin; Hotta, Go; Noguchi, Taro; Yamamoto, Masaki; Nagao, Miki; Takakura, Shunji; Ichiyama, Satoshi

    2016-05-01

    The accuracies of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and the phenotypic method using VITEK 2 were compared to the accuracy of 16S rRNA sequence analysis for the identification of 170 clinically isolated anaerobes. The antimicrobial susceptibility of the isolates was also evaluated. Genetic analysis identified 21 Gram-positive species in 14 genera and 29 Gram-negative species in 11 genera. The most frequently isolated genera were Prevotella spp. (n = 46), Bacteroides spp. (n = 25) and Clostridium spp. (n = 25). MALDI-TOF MS correctly identified more isolates compared with VITEK 2 at the species (80 vs. 58%, respectively; p < 0.01) and genus (85 vs. 71%, respectively; p < 0.01) levels. More than 90% of the isolates of the three major genera identified (Prevotella, Bacteroides, and Clostridium species other than Clostridium difficile) were susceptible to beta-lactam/beta-lactamase inhibitor combinations, carbapenems, metronidazole and chloramphenicol. MALDI-TOF MS provided better identification results than VITEK2. Commonly used anti-anaerobic agents indicated that the isolates of the three most frequently identified anaerobic genera exhibited good antimicrobial susceptibility.

  20. Pathogen Characterization of Fresh and Stored Mesophilic Anaerobically Digested Biosolids.

    PubMed

    A, Cecily

    2017-01-13

    Culturable bacterial pathogens (Campylobacter, Salmonella, Listeria, Yersinia) and indicators (E. coli, enterococci, Clostridium perfringens) were quantified at six wastewater treatment plants that land apply anaerobically digested biosolids in Ontario, Canada. Cryptosporidium parvum and Giardia lamblia were also quantified by PCR. Salmonella and Listeria were frequently detected in sludge and liquid biosolids (70-100% of samples) but less often in fresh dewatered cake biosolids (50-60%); with low levels in fresh cake (<100 cells/g dw). Yersinia were in 20-30% of samples, typically at very low levels (<10 cell/g dw). Giardia and Cryptosporidium were detected in 80% and 20% of cake biosolids at geometric means of 270 cysts/g dw and 70 oocysts/g dw, respectively. E. coli reduction was typically >2-log10 while pathogen reduction was variable. "Sudden increase" of pathogens was not observed, however, Salmonella and E. coli showed regrowth (at 1 to 3 orders of magnitude) after 2 to 3 day storage at 30°C.

  1. Thioredoxin system of the phototsynthetic anaerobe Chromatium vinosum

    SciTech Connect

    Johnson, T.C.; Crawford, N.A.; Buchanan, B.B.

    1984-06-01

    Chromatium vinosum, an anaerobic photosynthetic purple sulfur bacterium, resembles aerobic bacterial cells in that it has an NADP-thioredoxin system composed of a single thioredoxin which is reduced by NADPH via NADP-thioredoxin reductase. Both protein components were purified to homogeneity, and some of their properties were determined. Chromatium vinosum thioredoxin was slightly larger than other bacterial thioredoxins (13 versus 12 kilodaltons) but was similar in its specificity (ability to activate chloroplasts NADP-malate dehydrogenase more effectively than chloroplast fructose-1,6-bisphosphatase) and immunological properties. As in other bacteria, Chromatium vinosum NADP-thioredoxin reductase was an arsenite-sensitive flavoprotein composed of two 33.5-kilodalton subunits, that required thioredoxin for the NADPH-linked reduction of 5,5'-dithiobis(2-nitrobenzoic acid). Chromatium vinosum NADP-thioredoxin reductase very effectively reduced several different bacterial-type thioredoxins (Escherichia coli, Chlorobium thiosulfatophilum (this name has not been approved by the International Committee of Systematic Bacteriolgy), Rhizobium meliloti) but not others (Clostridium pasteurianum, spinach chloroplast thioredoxin m). The results show that Chromatium vinosum contains an NADP-thioredoxin system typical of evolutionary more advanced microorganisms.

  2. Prevalence and characterization of Clostridium perfringens from spices in Argentina.

    PubMed

    Aguilera, Milton Osmar; Stagnitta, Patricia Virginia; Micalizzi, Blas; de Guzmán, Ana María Stefanini

    2005-12-01

    Spices can present high microbial counts and Clostridium perfringens, Bacillus cereus, Salmonella and Shigella, among others have been isolated from spices. C. perfringens is an important pathogen agent causing, among other diseases, enteritis in humans caused by C. perfringens enterotoxin (CPE) which causes human food poisoning and enterotoxemia in domestic animals. The aims of the present work were (i) to establish the hygienic sanitary quality of some spices in San Luis, Argentina; (ii) to determine the presence of C. perfringens in these spices by means of the most probable number (MPN) and count on plate methods; (iii) to characterize the enterotoxigenic strains of C. perfringens by PCR and immunological methods such as reverse passive latex agglutination (RPLA) and (iv) to type by PCR C. perfringens strains isolated. A total of 115 samples of spices, 67 of which were purchased in local retail stores and 48 domestically collected were analysed. Total aerobe counts on tryptone glucose yeast extract agar medium of the 115 samples were between <10 and 10(6) CFU/g. The colifecal counts using Mac Conkey broth of the 115 samples were <4-10(3)CFU/g, with 28 samples (24.34%) exceeding the limit established by the Spanish Alimentary Code (10 CFU/g) while 2 samples (1.73%) had a sulfite reducing anaerobe load above standard limits. A total of 14 C. perfringens strains (12.17%) were isolated and characterized from 115 samples by the standard biochemical tests. Four of which (28.60%) turned out to be enterotoxigenic by PCR and RPLA. In order to type C. perfringens strains based on their main toxins, the 14 strains were analysed by PCR. All strains belonged to type A. All RPLA positive strains were cpe(+) by PCR. The percentage of enterotoxigenic strains was more elevated that those reported in other studies for this type of sample. These results indicate that sanitary conditions in different production stages of species must be improved to reduce health hazards. The high

  3. Genetic Engineering of Clostridium Difficile Toxin A Vaccine

    DTIC Science & Technology

    1991-09-04

    AD-A242 265 AD GENETIC ENGINEERING OF CLOSTRIDIUM DIFFICILE TOXIN A VACCINE ANNUAL/FINAL REPORT DTIC LYCJRGUS L. MULDROW F EIECTE JOE JOHNSON ’ N OVI...62770A 62770A871 AA DA314471 (U) Genetic Engineering of Clostridium difficile Toxin A Vaccine 12. PERSONAL AUTHOR(S) Lycurgus L. Muldrow and Joe... Clostridium difficile Vaccine 06 o2 Recombinant DNA 06 o3 RA 1 19. ABSTRACT (Continue on revere if n.ece•x••y and itd•entify by 0o/ r ou er).. .... Recombinant

  4. Secretion of clostridium cellulase by E. coli

    DOEpatents

    Yu, Ida Kuo

    1998-01-01

    A gene, encoding an endocellulase from a newly isolated mesophilic Clostridium strain IY-2 which can digest bamboo fibers, cellulose, rice straw, and sawdust, was isolated by shotgun cloning in an E. coli expression plasmid pLC2833. E. coli positive clones were selected based on their ability to hydrolyze milled bamboo fibers and cellulose present in agar plates. One clone contained a 2.8 kb DNA fragment that was responsible for cellulase activity. Western blot analyses indicated that the positive clone produced a secreted cellulase with a mass of about 58,000 daltons that was identical in size to the subunit of one of the three major Clostridium cellulases. The products of cellulose digestion by this cloned cellulase were cellotetraose and soluble higher polymers. The cloned DNA contained signal sequences capable of directing the secretion of heterologous proteins from an E. coli host. The invention describes a bioprocess for the treatment of cellulosic plant materials to produce cellular growth substrates and fermentation end products suitable for production of liquid fuels, solvents, and acids.

  5. The Pangenome of the genus Clostridium.

    PubMed

    Udaondo, Zulema; Duque, Estrella; Ramos, Juan Luis

    2017-03-21

    We present the pangenome for the genus Clostridium sensu stricto, which was obtained using highly curated and annotated genomes from 16 species, some of these cause disease, while others are used for the production of added-value chemicals. Multilocus sequencing analysis revealed that species of this genus group into at least two clades that include non-pathogenic and pathogenic strains, suggesting that pathogenicity is dispersed across the phylogenetic tree. The core genome of the genus includes 546 protein families, which mainly comprise those involved in protein translation and DNA repair. The GS-GOGAT may represent the central pathway for generating organic nitrogen from inorganic nitrogen sources. Glycerol and glucose metabolism genes are well represented in the core genome together with a set of energy conservation systems. A metabolic network comprising proteins/enzymes, RNAs and metabolites, whose topological structure is a non-random and scale-free network with hierarchically structured modules was built. These modules shed light on the interactions between RNAs, proteins and metabolites, revealing biological features of transcription and translation, cell wall biosynthesis, C1 metabolism and N metabolism. Network analysis identified four nodes that function as hubs and bottlenecks, namely, coenzyme A, HPr kinases, S-adenosylmethionine and the ribonuclease P-protein, suggesting pivotal roles for them in Clostridium. This article is protected by copyright. All rights reserved.

  6. Tea and Recurrent Clostridium difficile Infection

    PubMed Central

    Starley, Brad; Galagan, Jack Carl; Yabes, Joseph Michael; Evans, Sara

    2016-01-01

    Background and Aims. Studies have shown effects of diet on gut microbiota. We aimed to identify foods associated with recurrent Clostridium difficile infection (CDI). Methods. In this cross-sectional survey, consecutive patients diagnosed with CDI were identified by electronic medical records. Colitis symptoms and positive Clostridium difficile assay were confirmed. Health-care onset-health-care facility associated CDI was excluded. Food surveys were mailed to 411 patients. Survey responses served as the primary outcome measure. Spearman's rank correlation identified risk factors for CDI recurrence. Results. Surveys were returned by 68 patients. Nineteen patients experienced CDI recurrence. Compared to patients without CDI recurrence, patients with CDI recurrence had more antibiotics prescribed preceding their infection (p = 0.003). Greater numbers of the latter also listed tea (p = 0.002), coffee (p = 0.013), and eggs (p = 0.013), on their 24-hour food recall. Logistic regression identified tea as the only food risk factor for CDI recurrence (adjusted OR: 5.71; 95% CI: 1.26–25.89). Conclusion. The present results indicate a possible association between tea and CDI recurrence. Additional studies are needed to characterize and confirm this association. PMID:27651790

  7. [Selected aspects of Clostridium difficile infection].

    PubMed

    Mehlich, Agnieszka; Górska, Sabina; Gamian, Andrzej; Myc, Andrzej

    2015-05-05

    Clostridium difficile pathogen is a cause of the most frequent nosocomial infection, which is antibiotic-associated diarrhea. Antibiotic treatment causes disruption of the microbiome balance, which makes the gut a friendly environment for the pathogen. It leads to pseudomembranous colitis, toxic megacolon and even death. Clostridium difficile infection (CDI) is particularly dangerous to elderly patients, leading to the highest mortality rate. C. difficile is equipped with many virulence factors such as toxin A and B, binary toxin CDT, flagellum, S-layer proteins, Cwp66 and GroEL proteins, protease Cwp84, fibronectin-binding protein and the ability to form biofilm and spores. Problems with anti-CDI therapy prompt researchers and clinicians to seek alternative ways of therapy. Identification of immunological epitopes in outer layer proteins and the use of them as antigens for anti-CDI vaccines would be a rational approach to prevent the disease, but unfortunately such vaccines are not available yet. In this article we review the course of the disease, virulence and risk factors. We summarize briefly epidemiological data and the latest achievements in CDI treatment.

  8. Isolation and identification of Clostridium spp. from infections associated with the injection of drugs: experiences of a microbiological investigation team.

    PubMed

    Brazier, J S; Duerden, B I; Hall, V; Salmon, J E; Hood, J; Brett, M M; McLauchlin, J; George, R C

    2002-11-01

    Pathogenic species of the genus Clostridium may contaminate the materials used in the injection of drugs and under the right conditions may cause serious or life-threatening disease. C. novyi type A was implicated in an outbreak of severe infection with high mortality in injecting drug users who injected heroin extravascularly. The isolation of such highly oxygen-sensitive clostridia from clinical material may require adherence to enhanced methods and, once isolated, commercially available anaerobe identification kits alone may not give an accurate identification. Additional phenotypic tests that are useful in recognising the main pathogenic species are described. Differentiation of C. novyi type A from C. botulinum type C in reference laboratories was based on 16S rDNA sequence data and specific neutralisation of cytopathic effects in tissue culture.

  9. Clostridium baratii: a rare case of pneumonia associated with an Alzheimer patient in Rio de Janeiro, Brazil

    PubMed Central

    Lima, Carla Ormundo Gonçalves Ximenes; da Rocha, Vinicius Magno; Filho, Joaquim Santos; Serradas, Lucia Rodrigues; Silva, Rodrigo Otávio Silveira; Lobato, Francisco Carlos Faria; Domingues, Regina Maria Cavalcanti Pilotto

    2016-01-01

    Introduction: Clostridium baratii is rarely associated with human diseases. Infection is usuallcaused by ingestion of contaminated food, and infant botulism is the most common clinical presentation. Case Report: Here we report a case of pneumonia by a non-toxigenic strain of C. baratii in an Alzheimer 70-year-old male with sepsis in Rio de Janeiro, Brazil. The micro-organism was identified by phenotypical tests, mass spectrometry (MALDI-TOF), DNA amplification (PCR) and sequencing of the 16S rRNA gene. Testing for the presence of botulinum F toxin was made using multiplex PCR. Bioassay for a large number of colonies was performed in mice to evaluate the production of any lethal toxin, but the results were negative. Conclusion: To our knowledge, there are no cases of C. baratii infection reported in Brazil and we highlight the importance of anaerobic lab tests in the standard routine of diagnosis. PMID:28348769

  10. Bilateral abscessed orchiepididymitis associated with sepsis caused by Veillonella parvula and Clostridium perfringens: case report and review of the literature.

    PubMed Central

    Arrosagaray, P M; Salas, C; Morales, M; Correas, M; Barros, J M; Cordon, M L

    1987-01-01

    Veillonella species is a gram-negative coccus which is part of the anaerobic normal flora in the oral cavity, small intestine, upper respiratory tract, vagina, and urinary tract. The role that this organism plays in infection is not well known, and it is generally associated with other bacteria. We present a case of bilateral abscessed orchiepididymitis associated with septicemia due to Veillonella parvula and, later, to Clostridium perfringens, with the development of severe renal insufficiency and septic shock, which resolved favorably with antibiotic therapy, treatment of shock, and hyperbaric oxygen therapy. In reviewing the literature, we have not found any other case of sepsis due to Veillonella sp. associated with urological disorders. PMID:2887584

  11. Processing anaerobic sludge for extended storage as anaerobic digester inoculum.

    PubMed

    Li, Jiajia; Zicari, Steven M; Cui, Zongjun; Zhang, Ruihong

    2014-08-01

    Thermophilic anaerobic sludge was processed to reduce the volume and moisture content in order to reduce costs for storing and transporting the sludge as microbial inoculum for anaerobic digester startup. The moisture content of the sludge was reduced from 98.7% to 82.0% via centrifugation and further to 71.5% via vacuum evaporation. The processed sludge was stored for 2 and 4 months and compared with the fresh sludge for the biogas and methane production using food waste and non-fat dry milk as substrates. It was found that fresh unprocessed sludge had the highest methane yield and the yields of both unprocessed and processed sludges decreased during storage by 1-34%, however processed sludges seemed to regain some activity after 4 months of storage as compared to samples stored for only 2 months. Maximum methane production rates obtained from modified Gompertz model application also increased between the 2-month and 4-month processed samples.

  12. Detection of toxigenic Clostridium perfringens and Clostridium botulinum from food sold in Lagos, Nigeria.

    PubMed

    Chukwu, Emelda E; Nwaokorie, Francisca O; Coker, Akitoye O; Avila-Campos, Mario J; Solis, Rosa L; Llanco, Luis A; Ogunsola, Folasade T

    2016-12-01

    Food-borne diseases contribute to the huge burden of sickness and death globally and in the last decade, have become more frequently reported in Africa. In line with this, food safety is becoming a significant and growing public health problem in Nigeria. Diarrhoea is a common problem in Nigeria and has been reported but there has been little data on the possibility of clostridia as aetiological agents. Clostridium species are ubiquitous in the environment and in the gastrointestinal tract of man and animals and can serve as a marker for faecal contamination. We set out to determine the potential of these foods to transmit Clostridium species. A total of 220 food commodities from six local governments in Lagos State were sampled. Isolates obtained were identified based on cultural, morphological and biochemical characteristics. Toxinotyping was done using multiplex-PCR with primers specific for alpha, beta, epsilon and iota-toxin genes, enterotoxigenic cpe gene and neurotoxigenic BoNt gene. Fifty (22.7%) clostridial species were isolated of which 29 (58%) were identified as C. perfringens. Toxinotyping of the 29 strains showed that 28 (96.6%) were toxin producing C. perfringens type A while one (3.4%) was C. perfringens type D. Two (4%) C. botulinum species were isolated and identified by 16S rRNA sequencing, both harbouring BoNt/A gene. The contamination rates of food with Clostridium species show that food hygiene is a problem and Clostridium species may be a source of food borne disease in Lagos State, Nigeria.

  13. Detection of Clostridium sordellii strains expressing hemorrhagic toxin (TcsH) and implications for diagnostics and regulation of veterinary vaccines.

    PubMed

    Thiele, Teri L; Stuber, Tod P; Hauer, Paul J

    2013-10-17

    Clostridium sordellii is a Gram positive anaerobic bacterium that causes multiple disease syndromes in both humans and animals. As with many clostridial pathogens, toxins contribute to the virulence of C. sordellii. Two large toxins have been identified: a lethal toxin (TcsL) and a hemorrhagic toxin (TcsH) which are similar in structure and function to Clostridium difficile toxin B (TcdB) and toxin A (TcdA), respectively. While TcdA, TcdB, and TcsL have been extensively studied, relatively little is known about TcsH. This study elucidated the TcsH gene sequence using whole genome sequencing, compared the genotype with toxin expression of 52 C. sordellii strains, and examined the role of TcsH in batch release potency tests required for veterinary vaccines licensed in the United States and other testing utilizing WHO standard antitoxin. Data from this study will assist in future research to clarify the TcsH contribution to the pathogenesis of C. sordellii infections and may aid in the development of improved vaccines.

  14. Comparative genomics of the mesophilic cellulosome-producing Clostridium cellulovorans and its application to biofuel production via consolidated bioprocessing.

    PubMed

    Tamaru, Yutaka; Miyake, Hideo; Kuroda, Kouichi; Ueda, Mitsuyoshi; Doi, Roy H

    2010-01-01

    Clostridium cellulovorans is an anaerobic, mesophilic bacterium that efficiently degrades native substrates in soft biomass such as corn fibre and rice straw by producing an extracellular enzyme complex called the cellulosomes. By examining genome sequences from multiple Clostridium species, comparative genomics offers new insight into genome evolution and the way natural selection moulds functional DNA sequence evolution. Recently, we reported the whole genome sequence of C. cellulovorans. A total of 57 cellulosomal genes were found in the C. cellulovorans genome and coded for not only carbohydrate-active enzymes but also lipase, peptidase and proteinase inhibitors, in addition to two novel genes encoding scaffolding proteins CbpB and CbpC. Interestingly, the genome size of C. cellulovorans was about 1 Mbp larger than that of other cellulosome-producing clostridia: mesophilic C. cellulolyticum and thermophilic C. thermocellum. Since the C. cellulovorans genome included not only cellulosomal genes but also a large number of genes encoding non-cellulosomal enzymes, the genome expansion of C. cellulovorans included genes more related to degradation of polysaccharides, such as hemicelluloses and pectins, than to cellulose. In this review, we propose a strategy for industrial applications such as biofuel production using enhanced mesophilic cellulosome- and solvent-producing clostridia.

  15. [Clostridium alkalicellum sp. nov., an obligately alkaliphilic cellulolytic bacterium from a soda lake in the Baikal region].

    PubMed

    Zhilina, T N; Kevbrin, V V; Turova, T P; Lysenko, A M; Kostrikina, N A; Zavarzin, G A

    2005-01-01

    The first anaerobic alkaliphilic cellulolytic microorganism has been isolated from the Verkhnee Beloe soda lake (Buryatiya, Russia) with pH 10.2 and a salt content of up to 24 g/l. Five strains were characterized. Strain Z-7026 was chosen as the type strain. The cells of the isolate are gram-positive spore-forming rods. A mucous external capsule is produced. The microorganism is obligately alkaliphilic, growing in a pH range of 8.0-10.2, with an optimum at pH 9.0. Sodium ions and, in carbonate-buffered media, sodium chloride are obligately required. The microorganism is slightly halophilic; it grows at 0.017-0.4 M Na+ with an optimum at 0.15-0.3 M Na+. The metabolism is fermentative and strictly anaerobic. Cellulose, cellobiose, and xylan can be used as growth substrates. Plant and algal debris can be fermented. Lactate, ethanol, acetate, hydrogen, and traces of formate are produced during cellulose or cellobiose fermentation. Yeast extract or vitamins are required for anabolic purposes. The microorganism fixes dinitrogen and is nitrogenase-positive. It is tolerant to up to 48 mM Na2S. Growth is not inhibited by kanamycin or neomycin. Chloramphenicol, streptomycin, penicillin, ampicillin, ampiox, bacillin, novobiocin, and bacitracin suppress growth. The DNA G+C content is 29.9 mol %. According to the nucleotide sequence of its 16S rRNA gene, strain Z-7026 is phylogenetically close to the neutrophilic cellulolytic bacteria Clostridium thermocellum (95.5%), C. aldrichii (94.9%), and Acetivibrio cellulolyticus (94.8%). It is proposed as a new species: Clostridium alkalicellum sp. nov.

  16. Production and counting of spores of Clostridium chauvoei.

    PubMed

    Bagadi, H O

    1977-06-01

    The concentration and viability of spores produced by four different strains of Clostridium chauvoei (C. feseri) grown in a modified medium for 18 days are described. The medium yielded enough viable spores for experimental work.

  17. Flooding and Clostridium difficile infection: a case-crossover analysis

    EPA Science Inventory

    Clostridium difficile is a bacterium that can spread by water. It often causes acute gastrointestinal illness in older adults who are hospttalized and/or receiving antibiotics; however, community­ associated infections affecting otherwise healthy individuals have become more ...

  18. Characterization of Clostridium difficile Spores Lacking Either SpoVAC or Dipicolinic Acid Synthetase

    PubMed Central

    Donnelly, M. Lauren; Fimlaid, Kelly A.

    2016-01-01

    ABSTRACT The spore-forming obligate anaerobe Clostridium difficile is a leading cause of antibiotic-associated diarrhea around the world. In order for C. difficile to cause infection, its metabolically dormant spores must germinate in the gastrointestinal tract. During germination, spores degrade their protective cortex peptidoglycan layers, release dipicolinic acid (DPA), and hydrate their cores. In C. difficile, cortex hydrolysis is necessary for DPA release, whereas in Bacillus subtilis, DPA release is necessary for cortex hydrolysis. Given this difference, we tested whether DPA synthesis and/or release was required for C. difficile spore germination by constructing mutations in either spoVAC or dpaAB, which encode an ion channel predicted to transport DPA into the forespore and the enzyme complex predicted to synthesize DPA, respectively. C. difficile spoVAC and dpaAB mutant spores lacked DPA but could be stably purified and were more hydrated than wild-type spores; in contrast, B. subtilis spoVAC and dpaAB mutant spores were unstable. Although C. difficile spoVAC and dpaAB mutant spores exhibited wild-type germination responses, they were more readily killed by wet heat. Cortex hydrolysis was not affected by this treatment, indicating that wet heat inhibits a stage downstream of this event. Interestingly, C. difficile spoVAC mutant spores were significantly more sensitive to heat treatment than dpaAB mutant spores, indicating that SpoVAC plays additional roles in conferring heat resistance. Taken together, our results demonstrate that SpoVAC and DPA synthetase control C. difficile spore resistance and reveal differential requirements for these proteins among the Firmicutes. IMPORTANCE Clostridium difficile is a spore-forming obligate anaerobe that causes ∼500,000 infections per year in the United States. Although spore germination is essential for C. difficile to cause disease, the factors required for this process have been only partially characterized

  19. Culturable prokaryotic diversity of deep, gas hydrate sediments: first use of a continuous high-pressure, anaerobic, enrichment and isolation system for subseafloor sediments (DeepIsoBUG)

    PubMed Central

    Parkes, R John; Sellek, Gerard; Webster, Gordon; Martin, Derek; Anders, Erik; Weightman, Andrew J; Sass, Henrik

    2009-01-01

    Deep subseafloor sediments may contain depressurization-sensitive, anaerobic, piezophilic prokaryotes. To test this we developed the DeepIsoBUG system, which when coupled with the HYACINTH pressure-retaining drilling and core storage system and the PRESS core cutting and processing system, enables deep sediments to be handled without depressurization (up to 25 MPa) and anaerobic prokaryotic enrichments and isolation to be conducted up to 100 MPa. Here, we describe the system and its first use with subsurface gas hydrate sediments from the Indian Continental Shelf, Cascadia Margin and Gulf of Mexico. Generally, highest cell concentrations in enrichments occurred close to in situ pressures (14 MPa) in a variety of media, although growth continued up to at least 80 MPa. Predominant sequences in enrichments were Carnobacterium, Clostridium, Marinilactibacillus and Pseudomonas, plus Acetobacterium and Bacteroidetes in Indian samples, largely independent of media and pressures. Related 16S rRNA gene sequences for all of these Bacteria have been detected in deep, subsurface environments, although isolated strains were piezotolerant, being able to grow at atmospheric pressure. Only the Clostridium and Acetobacterium were obligate anaerobes. No Archaea were enriched. It may be that these sediment samples were not deep enough (total depth 1126–1527 m) to obtain obligate piezophiles. PMID:19694787

  20. The effect of probiotics on Clostridium difficile diarrhea.

    PubMed

    Pochapin, M

    2000-01-01

    Clostridium difficile is the leading cause of nosocomially acquired intestinal infection in the United States, affecting virtually all cases of pseudomembranous colitis and up to 20% of cases of antibiotic-associated diarrhea. Even after receiving antibiotic treatment with either metronidazole or vancomycin, 20% of patients will have recurrent Clostridium difficile diarrhea. An innovative approach to the problem involves the introduction of competing, nonpathogenic (probiotic) organisms into the intestinal tract to restore microbial balance. The theoretical premise behind this approach is that the protective intestinal microflora is damaged by antibiotic treatment; the initial antibiotic exposure thus leaves the host susceptible to colonization and subsequent infection by Clostridium difficile. A so-called "second-hit" to the intestinal microflora occurs when the infected host is treated with flagyl or vancomycin, further destroying susceptible bacterial flora. Probiotic agents, such as Lactobacillus GG and Saccharomyces boulardii, have been studied for the treatment of Clostridium difficile. We are currently running a prospective, randomized, placebo-controlled trial of Lactobacillus GG in combination with standard antibiotics for the treatment of Clostridium difficile infection. Although it is too early to draw statistically significant conclusions, two patterns seem to be emerging: Lactobacillus GG is effective in reducing the 3-wk recurrence rate of Clostridium difficile, and patients feel better when taking Lactobacillus GG, as compared with the placebo, with early disappearance of abdominal cramps and diarrhea. In conclusion, the use of probiotics for the treatment of primary and recurrent Clostridium difficile diarrhea looks promising. Patients seem to have less recurrent Clostridium difficile diarrhea and early symptomatic improvement when using the probiotic Lactobacillus GG.

  1. In vitro activity of moxifloxacin against 923 anaerobes isolated from human intra-abdominal infections.

    PubMed

    Goldstein, Ellie J C; Citron, Diane M; Warren, Yumi A; Tyrrell, Kerin L; Merriam, C Vreni; Fernandez, Helen

    2006-01-01

    The in vitro activity of moxifloxacin against 923 recent anaerobic isolates obtained from pretreatment cultures in patients with complicated intra-abdominal infections was studied using the CLSI M11-A-6 agar dilution method. Moxifloxacin was active against 87% (96 of 110) Bacteroides fragilis strains at < or = 1 microg/ml and 87% (79 of 90) B. thetaiotaomicron strains at < or = 2 microg/ml. Species variation was seen, with B. uniformis, B. vulgatus, Clostridium clostridioforme, and C. symbiosum being least susceptible and accounting for most of the resistant isolates; excluding the aforementioned four resistant species, 86% (303 of 363) of Bacteroides species isolates and 94% (417 of 450) of all other genera and species were susceptible to < or = 2 microg/ml of moxifloxacin. Overall, moxifloxacin was active against 763 of 923 (83%) of strains at < or = 2 microg/ml, supporting its use as a monotherapy for some community-acquired intra-abdominal infections.

  2. Susceptibilities of anaerobic bacteria isolated from animals with ovine foot rot to 28 antimicrobial agents.

    PubMed Central

    Piriz, S; Cuenca, R; Valle, J; Vadillo, S

    1992-01-01

    The agar dilution method was used to determine the inhibitory activities of 28 antimicrobial agents against 35 strains of the genus Peptostreptococcus, 4 strains of the species Peptococcus niger, 20 strains of the species Megasphaera elsdenii, 7 strains from the species Acidaminococcus fermentans, 8 strains of the genus Clostridium, 11 strains of the genus Eubacterium, and 1 strain of the species Propionibacterium acidipropionici, all of which were isolated from 125 clinical cases of ovine foot rot between January 1987 and December 1988. The three unreidopenicillins studied proved to be the most active antimicrobial agents, with a high percentage of strains being susceptible at a concentration of 64 micrograms/ml. Penicillin G, ampicillin, and the three cephalosporins studied also had good activity. Fosfomycin showed a high degree of activity among the 116 anaerobic bacteria tested. PMID:1590689

  3. Bioaugmentation with hydrolytic microbes to improve the anaerobic biodegradability of lignocellulosic agricultural residues.

    PubMed

    Tsapekos, P; Kougias, P G; Vasileiou, S A; Treu, L; Campanaro, S; Lyberatos, G; Angelidaki, I

    2017-03-12

    Bioaugmentation with hydrolytic microbes was applied to improve the methane yield of bioreactors fed with agricultural wastes. The efficiency of Clostridium thermocellum and Melioribacter roseus to degrade lignocellulosic matter was evaluated in batch and continuously stirred tank reactors (CSTRs). Results from batch assays showed that C. thermocellum enhanced the methane yield by 34%. A similar increase was recorded in CSTR during the bioaugmentation period; however, at steady-state the effect was noticeably lower (7.5%). In contrast, the bioaugmentation with M. roseus did not promote markedly the anaerobic biodegradability, as the methane yield was increased up to 10% in batch and no effect was shown in CSTR. High-throughput 16S rRNA amplicon sequencing was used to assess the effect of bioaugmentation strategies on bacterial and archaeal populations. The microbial analysis revealed that both strains were not markedly resided into biogas microbiome. Additionally, the applied strategies did not alter significantly the microbial communities.

  4. Fecal microbiota transplantation in the treatment of Clostridium difficile infections.

    PubMed

    Austin, Matthew; Mellow, Mark; Tierney, William M

    2014-06-01

    In recent years, Clostridium difficile infections have become more frequent, more severe, more refractory to standard treatment, and more likely to recur. Current antibiotic treatment regimens for Clostridium difficile infection alter the normal gut flora, which provide colonization resistance against Clostridium difficile. Over the past few years, there has been a marked increase in the knowledge of the gut microbiota and its role in health maintenance and disease causation. This has, fortuitously, coincided with the use of a unique microbial replacement therapy, fecal microbiota transplantation, in the treatment of patients with multiple recurrent Clostridium difficile infections. We briefly review current knowledge of the gut microbiota's functions. We then review the indications for use of fecal microbiota transplantation in Clostridium difficile infection, the techniques employed, and results of treatment. Fecal microbiota transplantation has been shown to be efficacious for patients with multiply recurrent Clostridium difficile infections (reported cure rates of 90%), with an excellent short-term safety profile, and has been included in the American College of Gastroenterology treatment guidelines for this troublesome disease.

  5. Chemical extraction versus direct smear for MALDI-TOF mass spectrometry identification of anaerobic bacteria.

    PubMed

    Fournier, Rémi; Wallet, Frédéric; Grandbastien, Bruno; Dubreuil, Luc; Courcol, René; Neut, Christel; Dessein, Rodrigue

    2012-06-01

    In the present study, two pre-analytic processes for mass spectrometric bacterial identification were compared: the time-consuming reference method, chemical extraction, and the direct smear technique directly using cultured colonies without any further preparation. These pre-analytic processes were compared in the identification of a total of 238 strains of anaerobic bacteria representing 34 species. The results showed that 218/238 strains were identified following chemical extraction, 185 identifications (77.7%) were secured to both genus and species [log(score) > 2.0] whereas 33 identifications (14%) were secured to genus only [log(score) between 1.7 and 2.0]. Following direct smear, 207/238 anaerobic bacteria were identified, 158 identifications (66.4%) were secured to both genus and species [log(score) > 2.0] whereas 49 identifications were secured to genus only [log(score) between 1.7 and 2.0]. Twenty strains were not identified [log(score) < 1.7] by MALDI-TOF MS following chemical extraction whereas 31 strains were not identified with the direct smear technique. Although direct smear led to a significant decrease of the log(score) values for the Clostridium genus and the Gram positive anaerobic bacteria (GPAC) group (p < 0.0001, Wilcoxon test), identification to both species and genus were not changed. However these differences were not statistically significant (p = 0.1, Chi square). Therefore, MALDI-TOF MS identification following the direct smear technique appears to both non-inferior to the reference method and relevant for anaerobic bacteria identification.

  6. Novel Real-Time PCR Assay for Simultaneous Detection and Differentiation of Clostridium chauvoei and Clostridium septicum in Clostridial Myonecrosis▿

    PubMed Central

    Halm, Anna; Wagner, Martin; Köfer, Josef; Hein, Ingeborg

    2010-01-01

    A real-time PCR assay based on the 16S rRNA gene sequence was designed for differentiation of blackleg-causing Clostridium chauvoei and Clostridium septicum, a phylogenetically closely related bacterium responsible for malignant edema. In order to exclude false-negative results, an internal amplification control was included in the assay. A set of three probes, one specific for C. chauvoei, one specific for C. septicum, and one specific for both species, permitted unequivocal detection of C. chauvoei in tests of 32 Clostridium sp. strains and 10 non-Clostridium strains. The assay proved to be sensitive, detecting one genome of C. chauvoei or C. septicum per PCR and 1.79 × 103 C. chauvoei cells/g artificially contaminated muscle tissue. In tests of 11 clinical specimens, the real-time PCR assay yielded the same results as an established conventional PCR method. PMID:20129968

  7. Novel real-time PCR assay for simultaneous detection and differentiation of Clostridium chauvoei and Clostridium septicum in clostridial myonecrosis.

    PubMed

    Halm, Anna; Wagner, Martin; Köfer, Josef; Hein, Ingeborg

    2010-04-01

    A real-time PCR assay based on the 16S rRNA gene sequence was designed for differentiation of blackleg-causing Clostridium chauvoei and Clostridium septicum, a phylogenetically closely related bacterium responsible for malignant edema. In order to exclude false-negative results, an internal amplification control was included in the assay. A set of three probes, one specific for C. chauvoei, one specific for C. septicum, and one specific for both species, permitted unequivocal detection of C. chauvoei in tests of 32 Clostridium sp. strains and 10 non-Clostridium strains. The assay proved to be sensitive, detecting one genome of C. chauvoei or C. septicum per PCR and 1.79 x 10(3) C. chauvoei cells/g artificially contaminated muscle tissue. In tests of 11 clinical specimens, the real-time PCR assay yielded the same results as an established conventional PCR method.

  8. Characterization of flagellin from Clostridium chauvoei.

    PubMed

    Kojima, A; Amimoto, K; Ohgitani, T; Tamura, Y

    1999-06-30

    Differential centrifugation and cesium chloride-equilibrium centrifugation were used to purify the flagella from the strain Okinawa of the formalin-fixed Clostridium chauvoei. SDS-PAGE profile of purified flagella showed that a major protein band with a molecular mass of 46 kDa, corresponding to the flagellin monomer, and at least two minor protein bands with molecular masses of approximately 73 and 100 kDa were found. The amino acid composition of C. chauvoei flagellin was similar to the flagellin of Salmonella typhimurium and Bacillus subtilis. In addition, C. chauvoei flagellin monomer shared limited sequence homology with the N-terminal amino acid sequence reported for other bacterial flagellins. N-terminal sequences of two minor bands corresponded to the flagellin monomer, indicating that higher molecular mass bands were polymeric forms of the flagellin monomer.

  9. Patho-genetics of Clostridium chauvoei.

    PubMed

    Frey, Joachim; Falquet, Laurent

    2015-05-01

    The genomic sequence of Clostridium chauvoei, the etiological agent of blackleg, a severe disease of ruminants with high mortality specified by a myonecrosis reveals a chromosome of 2.8 million base-pairs and a cryptic plasmid of 5.5 kilo base-pairs. The chromosome contains the main pathways like glycolysis/gluconeogenesis, sugar metabolism, purine and pyrimidine metabolisms, but the notable absence of genes of the citric acid cycle and deficient or partially deficient amino acid metabolism for Histidine, Tyrosine, Phenylalanine, and Tryptophan. These essential amino acids might be acquired from host tissue damage caused by various toxins and by protein metabolism that includes 57 genes for peptidases, and several ABC transporters for amino acids import.

  10. [Laboratory diagnosis of Clostridium difficile infection].

    PubMed

    Alcalá-Hernández, Luis; Mena-Ribas, Ana; Niubó-Bosh, Jordi; Marín-Arriaza, Mercedes

    2016-11-01

    Clostridium difficile is the leading cause of nosocomial diarrhoea in developed countries, and is one of the main aetiologic agents of community diarrhea. The eruption of the hypervirulent strain BI/NAP1/027 has given rise to an increase in the morbidity and mortality of C.difficile infection (CDI). This document aims to review the main clinical pictures of CDI and the laboratory diagnosis, including sampling, transport and storage of specimens, specimen processing, diagnostic procedures, antimicrobial susceptibility testing, and molecular characterisation of the isolates. The main purpose of the article is to develop a practical document that provides answers to the main questions that arise in the laboratory diagnosis of CDI.

  11. Clostridium difficile infection in older adults

    PubMed Central

    Jump, Robin LP

    2014-01-01

    Clostridium difficile infection, the most frequent cause of nosocomial diarrhea, disproportionately affects older adults. The two most important risk factors for developing C. difficile infection are antimicrobial exposure and age >65 years old. Risk factors specific to older adults are frequent interactions with healthcare systems and age-related changes in physiology, including immune senescence and changes to the gut microbiome. Metronidazole and oral vancomcyin are the mainstays of conventional treatment for C. difficile infection. Alternative therapies include fidaxomicin, a narrow-spectrum macrocyclic antibiotic, and fectal bacteriotherapy, which offers an excellent therapeutic outcome. Strategies to prevent C. difficile infections include enhanced infection control measures and reducing inappropriate antimicrobial use through stewardship. PMID:24955106

  12. Clostridium difficile Infection and Fecal Microbiota Transplant.

    PubMed

    Liubakka, Alyssa; Vaughn, Byron P

    2016-07-01

    Clostridium difficile infection (CDI) is a major source of morbidity and mortality for hospitalized patients. Although most patients have a clinical response to existing antimicrobial therapies, recurrent infection develops in up to 30% of patients. Fecal microbiota transplant is a novel approach to this complex problem, with an efficacy rate of nearly 90% in the setting of multiple recurrent CDI. This review covers the current epidemiology of CDI (including toxigenic and nontoxigenic strains, risk factors for infection, and recurrent infection), methods of diagnosis, existing first-line therapies in CDI, the role of fecal microbiota transplant for multiple recurrent CDIs, and the potential use of fecal microbial transplant for patients with severe or refractory infection.

  13. Carbohydrate-based Clostridium difficile vaccines.

    PubMed

    Monteiro, Mario A; Ma, Zuchao; Bertolo, Lisa; Jiao, Yuening; Arroyo, Luis; Hodgins, Douglas; Mallozzi, Michael; Vedantam, Gayatri; Sagermann, Martin; Sundsmo, John; Chow, Herbert

    2013-04-01

    Clostridium difficile is responsible for thousands of deaths each year and a vaccine would be welcomed, especially one that would disrupt bacterial maintenance, colonization and persistence in carriers and convalescent patients. Structural explorations at the University of Guelph (ON, Canada) discovered that C. difficile may express three phosphorylated polysaccharides, named PSI, PSII and PSIII; this review captures our recent efforts to create vaccines based on these glycans, especially PSII, the common antigen that has precipitated immediate attention. The authors describe the design and immunogenicity of vaccines composed of raw polysaccharides and conjugates thereof. So far, it has been observed that anti-PSII antibodies can be raised in farm animals, mice and hamster models; humans and horses carry anti-PSII IgA and IgG antibodies from natural exposure to C. difficile, respectively; phosphate is an indispensable immunogenic epitope and vaccine-induced PSII antibodies recognize PSII on C. difficile outer surface.

  14. Therapeutic approaches for Clostridium difficile infections.

    PubMed

    Marsh, Jane W; Curry, Scott R

    2013-10-02

    Metronidazole and vancomycin remain the front-line therapies for most Clostridium difficile infections (CDI). However, recurrent CDI occurs in ∼ 25% of patients, causing significant morbidity and mortality and healthcare costs. For this population, traditional antibiotic therapies fail and new treatment options are greatly needed. The US Food and Drug Administration recently approved fidaxomicin for CDI treatment. This narrow-spectrum antibiotic preserves the normal gut microbiota and shows promise as a treatment for severe and recurrent CDI. Monoclonal antibodies and vaccines directed against toxin are currently in clinical trials and represent alternative, non-antibiotic therapies. Less traditional therapeutic interventions include bacteriotherapy with non-toxigenic C. difficile and fecal transplant. This commentary will provide an overview of current and forthcoming CDI therapies.

  15. Investigational new treatments for Clostridium difficile infection.

    PubMed

    Ivarsson, Mattias E; Leroux, Jean-Christophe; Castagner, Bastien

    2015-05-01

    Significant progress has been made by industry and academia in the past two years to address the medical threats posed by Clostridium difficile infection. These developments provide an excellent example of how patient need has driven a surge of innovation in drug discovery. Indeed, only two drugs were approved for the infection in the past 30 years but there are 13 treatment candidates in clinical trials today. What makes the latter number even more remarkable is the diversity in the strategies represented (antibiotics, microbiota supplements, vaccines, antibiotic quenchers and passive immunization). In this review, we provide a snapshot of the current stage of these breakthroughs and argue that there is still room for further innovation in treating C. difficile infection.

  16. Clostridium difficile: clinical disease and diagnosis.

    PubMed Central

    Knoop, F C; Owens, M; Crocker, I C

    1993-01-01

    Clostridium difficile is an opportunistic pathogen that causes a spectrum of disease ranging from antibiotic-associated diarrhea to pseudomembranous colitis. Although the disease was first described in 1893, the etiologic agent was not isolated and identified until 1978. Since clinical and pathological features of C. difficile-associated disease are not easily distinguished from those of other gastrointestinal diseases, including ulcerative colitis, chronic inflammatory bowel disease, and Crohn's disease, diagnostic methods have relied on either isolation and identification of the microorganism or direct detection of bacterial antigens or toxins in stool specimens. The current review focuses on the sensitivity, specificity, and practical use of several diagnostic tests, including methods for culture of the etiologic agent, cellular cytotoxicity assays, latex agglutination tests, enzyme immunoassay systems, counterimmunoelectrophoresis, fluorescent-antibody assays, and polymerase chain reactions. PMID:8358706

  17. Clostridium difficile outbreaks: prevention and treatment strategies

    PubMed Central

    Martinez, Fernando J; Leffler, Daniel A; Kelly, Ciaran P

    2012-01-01

    The incidence and severity of Clostridium difficile infection (CDI) have increased dramatically over the past decade. Its treatment, however, has largely remained the same with the exception of oral vancomycin use as a first-line agent in severe disease. From 1999 to 2004, 20,642 deaths were attributed to CDI in the United States, almost 7 times the rate of all other intestinal infections combined. Worldwide, several major CDI outbreaks have occurred, and many of these were associated with the NAP1 strain. This ‘epidemic’ strain has contributed to the rising incidence and mortality of CDI. The purpose of this article is to review the current management, treatment, infection control, and prevention strategies that are needed to combat this increasingly morbid disease. PMID:22826646

  18. Anaerobic digestion of aliphatic polyesters.

    PubMed

    Šmejkalová, Pavla; Kužníková, Veronika; Merna, Jan; Hermanová, Soňa

    2016-01-01

    Anaerobic processes for the treatment of plastic materials waste represent versatile and effective approach in environmental protection and solid waste management. In this work, anaerobic biodegradability of model aliphatic polyesters, poly(L-lactic acid) (PLA), and poly(ɛ-caprolactone) (PCL), in the form of powder and melt-pressed films with varying molar mass, was studied. Biogas production was explored in batch laboratory trials at 55 ± 1°C under a nitrogen atmosphere. The inoculum used was thermophilic digested sludge (total solids concentration of 2.9%) from operating digesters at the Central Waste Water Treatment Plant in Prague, Czech Republic. Methanogenic biodegradation of PCLs typically yielded from 54 to 60% of the theoretical biogas yield. The biodegradability of PLAs achieved from 56 to 84% of the theoretical value. High biogas yield (up to 677 mL/g TS) with high methane content (more than 60%), comparable with conventionally processed materials, confirmed the potential of polyester samples for anaerobic treatment in the case of their exploitation in agriculture or as a packaging material in the food industry.

  19. Clostridium difficile infection in horses: a review.

    PubMed

    Diab, S S; Songer, G; Uzal, F A

    2013-11-29

    Clostridium difficile is considered one of the most important causes of diarrhea and enterocolitis in horses. Foals and adult horses are equally susceptible to the infection. The highly resistant spore of C. difficile is the infectious unit of transmission, which occurs primarily via the fecal-oral route, with sources of infection including equine feces, contaminated soil, animal hospitals, and feces of other animals. Two major risk factors for the development of C. difficile associated disease (CDAD) in adult horses are hospitalization and antimicrobial treatment, although sporadically, cases of CDAD can occur in horses that have not received antimicrobials or been hospitalized. The most common antibiotics associated with CDAD in horses are erythromycin, trimethoprim/sulfonamides, β-lactam antimicrobials, clindamycin, rifampicin, and gentamicin. Clinical signs and intestinal lesions of CDAD infection are not specific and they cannot be used to distinguish infections by C. difficile from infections by other agents, such as Clostridium perfringens or Salmonella sp. The distribution of lesions throughout the intestinal tract seems to be age-dependent. Small intestine is invariably affected, and colon and cecum may or may not have lesions in foals<1-month old. Naturally acquired disease in older foals and adult horses has a more aboral distribution, affecting colon and sometimes cecum, but rarely the small intestine. Detection of toxin A, toxin B or both in intestinal contents or feces is considered the most reliable diagnostic criterion for CDAD in horses. Isolation of toxigenic strains of C. difficile from horses with intestinal disease is highly suggestive of CDAD. A better understanding of pathogenesis, reservoirs of infection, and vaccines and other methods of control is needed. Also further studies are recommended to investigate other possible predisposing factors and/or etiological agents of enteric diseases of horses.

  20. 454-Pyrosequencing analysis of highly adapted azo dye-degrading microbial communities in a two-stage anaerobic-aerobic bioreactor treating textile effluent.

    PubMed

    Köchling, Thorsten; Ferraz, Antônio Djalma Nunes; Florencio, Lourdinha; Kato, Mario Takayuki; Gavazza, Sávia

    2017-03-01

    Azo dyes, which are widely used in the textile industry, exhibit significant toxic characteristics for the environment and the human population. Sequential anaerobic-aerobic reactor systems are efficient for the degradation of dyes and the mineralization of intermediate compounds; however, little is known about the composition of the microbial communities responsible for dye degradation in these systems. 454-Pyrosequencing of the 16S rRNA gene was employed to assess the bacterial biodiversity and composition of a two-stage (anaerobic-aerobic) pilot-scale reactor that treats effluent from a denim factory. The anaerobic reactor was inoculated with anaerobic sludge from a domestic sewage treatment plant. Due to the selective composition of the textile wastewater, after 210 days of operation, the anaerobic reactor was dominated by the single genus Clostridium, affiliated with the Firmicutes phylum. The aerobic biofilter harbored a diverse bacterial community. The most abundant phylum in the aerobic biofilter was Proteobacteria, which was primarily represented by the Gamma, Delta and Epsilon classes followed by Firmicutes and other phyla. Several bacterial genera were identified that most likely played an essential role in azo dye degradation in the investigated system.