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Sample records for actinobacillus succinogenes atcc

  1. Actinobacillus succinogenes ATCC 55618 Fermentation Medium Optimization for the Production of Succinic Acid by Response Surface Methodology

    PubMed Central

    Zhu, Li-Wen; Wang, Cheng-Cheng; Liu, Rui-Sang; Li, Hong-Mei; Wan, Duan-Ji; Tang, Ya-Jie

    2012-01-01

    As a potential intermediary feedstock, succinic acid takes an important place in bulk chemical productions. For the first time, a method combining Plackett-Burman design (PBD), steepest ascent method (SA), and Box-Behnken design (BBD) was developed to optimize Actinobacillus succinogenes ATCC 55618 fermentation medium. First, glucose, yeast extract, and MgCO3 were identified to be key medium components by PBD. Second, preliminary optimization was run by SA method to access the optimal region of the key medium components. Finally, the responses, that is, the production of succinic acid, were optimized simultaneously by using BBD, and the optimal concentration was located to be 84.6 g L−1 of glucose, 14.5 g L−1 of yeast extract, and 64.7 g L−1 of MgCO3. Verification experiment indicated that the maximal succinic acid production of 52.7 ± 0.8 g L−1 was obtained under the identified optimal conditions. The result agreed with the predicted value well. Compared with that of the basic medium, the production of succinic acid and yield of succinic acid against glucose were enhanced by 67.3% and 111.1%, respectively. The results obtained in this study may be useful for the industrial commercial production of succinic acid. PMID:23093852

  2. Performance analyses of a neutralizing agent combination strategy for the production of succinic acid by Actinobacillus succinogenes ATCC 55618.

    PubMed

    Wang, Cheng-Cheng; Zhu, Li-Wen; Li, Hong-Mei; Tang, Ya-Jie

    2012-05-01

    A neutralizing agent combination strategy was developed to enhance the succinic acid production by Actinobacillus succinogenes ATCC 55618. First, a maximal succinic acid production of 48.2 g/L was obtained at a culture pH of 7.5. Second, NaOH and KOH were screened to identify the optimal neutralizing agent for pH control. However, the production of succinic acid did not increase, and severe cell flocculation was observed due to a high concentration of metal ions when only one neutralizing agent was used to control pH. Finally, a neutralizing agent combination strategy was developed with a supply of neutralizing agents with OH(-) and carbonate. The cell flocculation was eliminated, and a maximum succinic acid production of 59.2 g/L was obtained with 5 M NaOH and 40 g/L of MgCO(3); this production was 27.9% higher than that obtained with NaOH alone. The results obtained in this study may be useful for the large-scale industrial production of succinic acid. PMID:22002101

  3. Actinobacillus succinogenes ATCC 55618 fermentation medium optimization for the production of succinic acid by response surface methodology.

    PubMed

    Zhu, Li-Wen; Wang, Cheng-Cheng; Liu, Rui-Sang; Li, Hong-Mei; Wan, Duan-Ji; Tang, Ya-Jie

    2012-01-01

    As a potential intermediary feedstock, succinic acid takes an important place in bulk chemical productions. For the first time, a method combining Plackett-Burman design (PBD), steepest ascent method (SA), and Box-Behnken design (BBD) was developed to optimize Actinobacillus succinogenes ATCC 55618 fermentation medium. First, glucose, yeast extract, and MgCO(3) were identified to be key medium components by PBD. Second, preliminary optimization was run by SA method to access the optimal region of the key medium components. Finally, the responses, that is, the production of succinic acid, were optimized simultaneously by using BBD, and the optimal concentration was located to be 84.6 g L(-1) of glucose, 14.5 g L(-1) of yeast extract, and 64.7 g L(-1) of MgCO(3). Verification experiment indicated that the maximal succinic acid production of 52.7 ± 0.8 g L(-1) was obtained under the identified optimal conditions. The result agreed with the predicted value well. Compared with that of the basic medium, the production of succinic acid and yield of succinic acid against glucose were enhanced by 67.3% and 111.1%, respectively. The results obtained in this study may be useful for the industrial commercial production of succinic acid. PMID:23093852

  4. Economical succinic acid production from cane molasses by Actinobacillus succinogenes.

    PubMed

    Liu, Yu-Peng; Zheng, Pu; Sun, Zhi-Hao; Ni, Ye; Dong, Jin-Jun; Zhu, Lei-Lei

    2008-04-01

    In this work, production of succinic acid by Actinobacillus succinogenes CGMCC1593 using cane molasses as a low cost carbon source was developed. In anaerobic bottles fermentation, succinic acid concentration of 50.6+/-0.9 g l(-1) was attained at 60 h using an optimum medium containing molasses pretreated with sulfuric acid, resulting in a succinic acid yield of 79.5+/-1.1% and sugar utilization of 97.1+/-0.6%. When batch fermentation was carried out in a 5-l stirred bioreactor with pretreated molasses, 46.4 g l(-1) of succinic acid was attained at 48 h and faster cells growth was also observed. Fed batch fermentation was performed to minimize the substrate (sugar) inhibition effect, giving 55.2 g l(-1) of succinic acid and 1.15 g l(-1)h(-1) of productivity at 48 h. The present study suggests that the inexpensive cane molasses could be utilized for the economical and efficient production of succinic acid by A. succinogenes. PMID:17532626

  5. Respiratory glycerol metabolism of Actinobacillus succinogenes 130Z for succinate production.

    PubMed

    Schindler, Bryan D; Joshi, Rajasi V; Vieille, Claire

    2014-09-01

    Actinobacillus succinogenes 130Z naturally produces among the highest levels of succinate from a variety of inexpensive carbon substrates. A few studies have demonstrated that A. succinogenes can anaerobically metabolize glycerol, a waste product of biodiesel manufacture and an inexpensive feedstock, to produce high yields of succinate. However, all these studies were performed in the presence of yeast extract, which largely removes the redox constraints associated with fermenting glycerol, a highly reduced molecule. We demonstrated that A. succinogenes cannot ferment glycerol in minimal medium, but that it can metabolize glycerol by aerobic or anaerobic respiration. These results were expected based on the A. succinogenes genome, which encodes respiratory enzymes, but no pathway for 1,3-propanediol production. We investigated A. succinogenes's glycerol metabolism in minimal medium in a variety of respiratory conditions by comparing growth, metabolite production, and in vitro activity of terminal oxidoreductases. Nitrate inhibited succinate production by inhibiting fumarate reductase expression. In contrast, growth in the presence of dimethylsulfoxide and in microaerobic conditions allowed high succinate yields. The highest succinate yield was 0.75 mol/mol glycerol (75 % of the maximum theoretical yield) in continuous microaerobic cultures. A. succinogenes could also grow and produce succinate on partially refined glycerols obtained directly from biodiesel manufacture. Finally, by expressing a heterologous 1,3-propanediol synthesis pathway in A. succinogenes, we provide the first proof of concept that A. succinogenes can be engineered to grow fermentatively on glycerol. PMID:25047181

  6. A genomic perspective on the potential of Actinobacillus succinogenes for industrial succinate production

    PubMed Central

    2010-01-01

    Background Succinate is produced petrochemically from maleic anhydride to satisfy a small specialty chemical market. If succinate could be produced fermentatively at a price competitive with that of maleic anhydride, though, it could replace maleic anhydride as the precursor of many bulk chemicals, transforming a multi-billion dollar petrochemical market into one based on renewable resources. Actinobacillus succinogenes naturally converts sugars and CO2 into high concentrations of succinic acid as part of a mixed-acid fermentation. Efforts are ongoing to maximize carbon flux to succinate to achieve an industrial process. Results Described here is the 2.3 Mb A. succinogenes genome sequence with emphasis on A. succinogenes's potential for genetic engineering, its metabolic attributes and capabilities, and its lack of pathogenicity. The genome sequence contains 1,690 DNA uptake signal sequence repeats and a nearly complete set of natural competence proteins, suggesting that A. succinogenes is capable of natural transformation. A. succinogenes lacks a complete tricarboxylic acid cycle as well as a glyoxylate pathway, and it appears to be able to transport and degrade about twenty different carbohydrates. The genomes of A. succinogenes and its closest known relative, Mannheimia succiniciproducens, were compared for the presence of known Pasteurellaceae virulence factors. Both species appear to lack the virulence traits of toxin production, sialic acid and choline incorporation into lipopolysaccharide, and utilization of hemoglobin and transferrin as iron sources. Perspectives are also given on the conservation of A. succinogenes genomic features in other sequenced Pasteurellaceae. Conclusions Both A. succinogenes and M. succiniciproducens genome sequences lack many of the virulence genes used by their pathogenic Pasteurellaceae relatives. The lack of pathogenicity of these two succinogens is an exciting prospect, because comparisons with pathogenic Pasteurellaceae could

  7. Production of succinic acid from oil palm empty fruit bunch cellulose using Actinobacillus succinogenes

    NASA Astrophysics Data System (ADS)

    Pasma, Satriani Aga; Daik, Rusli; Maskat, Mohamad Yusof

    2013-11-01

    Succinic acid is a common metabolite in plants, animals and microorganisms. It has been used widely in agricultural, food and pharmaceutical industries. Enzymatic hydrolysate glucose from oil palm empty fruit bunch (OPEFB) cellulose was used as a substrate for succinic acid production using Actinobacillus succinogenes. Using cellulose extraction from OPEFB can enhance the production of glucose as a main substrate for succinic acid production. The highest concentration of glucose produced from enzymatic hydrolysis is 167 mg/mL and the sugar recovery is 0.73 g/g of OPEFB. By optimizing the culture medium for succinic acid fermentation with enzymatic hydrolysate of OPEFB cellulose, the nitrogen sources could be reduced to just only 2.5 g yeast extract and 2.5 g corn step liquor. Batch fermentation was carried out using enzymatic hydrolysate of OPEFB cellulose with yeast extract, corn steep liquor and the salts mixture, 23.5 g/L succinic acid was obtained with consumption of 72 g/L glucose in enzymatic hydrolysate of OPEFB cellulose at 38 hours and 37°C. This study suggests that enzymatic hydrolysate of OPEFB cellulose maybe an alternative substrate for the efficient production of succinic acid by Actinobacillus succinogenes.

  8. Succinic acid production from duckweed (Landoltia punctata) hydrolysate by batch fermentation of Actinobacillus succinogenes GXAS137.

    PubMed

    Shen, Naikun; Wang, Qingyan; Zhu, Jing; Qin, Yan; Liao, Siming; Li, Yi; Zhu, Qixia; Jin, Yanling; Du, Liqin; Huang, Ribo

    2016-07-01

    Duckweed is potentially an ideal succinic acid (SA) feedstock due to its high proportion of starch and low lignin content. Pretreatment methods, substrate content and nitrogen source were investigated to enhance the bioconversion of duckweed to SA and to reduce the costs of production. Results showed that acid hydrolysis was an effective pretreatment method because of its high SA yield. The optimum substrate concentration was 140g/L. The optimum substrate concentration was 140g/L. Corn steep liquor powder could be considered a feasible and inexpensive alternative to yeast extract as a nitrogen source. Approximately 57.85g/L of SA was produced when batch fermentation was conducted in a 1.3L stirred bioreactor. Therefore, inexpensive duckweed can be a promising feedstock for the economical and efficient production of SA through fermentation by Actinobacillus succinogenes GXAS137. PMID:27023386

  9. Immobilization of Actinobacillus succinogenes by adhesion or entrapment for the production of succinic acid.

    PubMed

    Corona-González, Rosa Isela; Miramontes-Murillo, Ricardo; Arriola-Guevara, Enrique; Guatemala-Morales, Guadalupe; Toriz, Guillermo; Pelayo-Ortiz, Carlos

    2014-07-01

    The production of succinic acid was studied with entrapped and adsorbed Actinobacillus succinogenes. The adsorption of fermentation products (organic acids in the concentration range of 1-20 g/L) on different supports was evaluated. It was found that succinic acid was adsorbed in small quantities on diatomite and zeolite (12.6 mg/g support). The highest production of succinic acid was achieved with A. succinogenes entrapped in agar beads. Batch fermentations with immobilized cells were carried out with glucose concentrations ranging from 20 to 80 g/L. Succinic acid (43.4 g/L) was obtained from 78.3g/L glucose, and a high productivity (2.83 g/Lh) was obtained with a glucose concentration of 37.6g/L. For repeated batch fermentations (5 cycles in 72 h) with immobilized cells in agar, the total glucose consumed was 147.55 g/L, while the production of succinic acid was 107 g/L. Immobilized cells reduced significantly the fermentation time, yield, productivity and final concentration of succinic acid. PMID:24844165

  10. Succinic acid production from acid hydrolysate of corn fiber by Actinobacillus succinogenes.

    PubMed

    Chen, Kequan; Jiang, Min; Wei, Ping; Yao, Jiaming; Wu, Hao

    2010-01-01

    Dilute acid hydrolysate of corn fiber was used as carbon source for the production of succinic acid by Actinobacillus succinogenes NJ113. The optimized hydrolysis conditions were obtained by orthogonal experiments. When corn fiber particles were of 20 mesh in size and treated with 1.0% sulfuric acid at 121 degrees C for 2 h, the total sugar yield could reach 63.3%. It was found that CaCO(3) neutralization combined with activated carbon adsorption was an effective method to remove fermentation inhibitors especially furfural that presented in the acid hydrolysate of corn fiber. Only 5.2% of the total sugar was lost, while 91.9% of furfural was removed. The yield of succinic acid was higher than 72.0% with the detoxified corn fiber hydrolysate as the carbon source in anaerobic bottles or 7.5 L fermentor cultures. It was proved that the corn fiber hydrolysate could be an alternative to glucose for the production of succinic acid by A. succinogenes NJ113. PMID:18830824

  11. Improving succinic acid production by Actinobacillus succinogenes from raw industrial carob pods.

    PubMed

    Carvalho, Margarida; Roca, Christophe; Reis, Maria A M

    2016-10-01

    Carob pods are an inexpensive by-product of locust bean gum industry that can be used as renewable feedstock for bio-based succinic acid. Here, for the first time, unprocessed raw carob pods were used to extract a highly enriched sugar solution, afterwards used as substrate to produce succinic acid using Actinobacillus succinogenes. Batch fermentations containing 30g/L sugars resulted in a production rate of 1.67gSA/L.h and a yield of 0.39gSA/g sugars. Taking advantage of A. succinogenes' metabolism, uncoupling cell growth from succinic acid production, a fed-batch mode was implemented to increase succinic acid yield and reduce by-products formation. This strategy resulted in a succinic acid yield of 0.94gSA/g sugars, the highest yield reported in the literature for fed-batch and continuous experiments, while maintaining by-products at residual values. Results demonstrate that raw carob pods are a highly efficient feedstock for bio-based succinic acid production. PMID:27394995

  12. Characterization of bifunctional L-glutathione synthetases from Actinobacillus pleuropneumoniae and Actinobacillus succinogenes for efficient glutathione biosynthesis.

    PubMed

    Yang, Jianhua; Li, Wei; Wang, Dezheng; Wu, Hui; Li, Zhimin; Ye, Qin

    2016-07-01

    Glutathione (GSH), an important bioactive substance, is widely applied in pharmaceutical and food industries. In this work, two bifunctional L-glutathione synthetases (GshF) from Actinobacillus pleuropneumoniae (GshFAp) and Actinobacillus succinogenes (GshFAs) were successfully expressed in Escherichia coli BL-21(DE3). Similar to the GshF from Streptococcus thermophilus (GshFSt), GshFAp and GshFAs can be applied for high titer GSH production because they are less sensitive to end-product inhibition (Ki values 33 and 43 mM, respectively). The active catalytic forms of GshFAs and GshFAp are dimers, consistent with those of GshFPm (GshF from Pasteurella multocida) and GshFSa (GshF from Streptococcus agalactiae), but are different from GshFSt (GshF from S. thermophilus) which is an active monomer. The analysis of the protein sequences and three dimensional structures of GshFs suggested that the binding sites of GshFs for substrates, L-cysteine, L-glutamate, γ-glutamylcysteine, adenosine-triphosphate, and glycine are highly conserved with only very few differences. With sufficient supply of the precursors, the recombinant strains BL-21(DE3)/pET28a-gshFas and BL-21(DE3)/pET28a-gshFap were able to produce 36.6 and 34.1 mM GSH, with the molar yield of 0.92 and 0.85 mol/mol, respectively, based on the added L-cysteine. The results showed that GshFAp and GshFAs are potentially good candidates for industrial GSH production. PMID:26996628

  13. A complete industrial system for economical succinic acid production by Actinobacillus succinogenes.

    PubMed

    Li, Jian; Zheng, Xiao-Yu; Fang, Xiao-Jiang; Liu, Shu-Wen; Chen, Ke-Quan; Jiang, Min; Wei, Ping; Ouyang, Ping-Kai

    2011-05-01

    An industrial fermentation system using lignocellulosic hydrolysate, waste yeast hydrolysate, and mixed alkali to achieve high-yield, economical succinic acid production by Actinobacillus succinogenes was developed. Lignocellulosic hydrolysate and waste yeast hydrolysate were used efficiently as carbon sources and nitrogen source instead of the expensive glucose and yeast extract. Moreover, as a novel method for regulating pH mixed alkalis (Mg(OH)(2) and NaOH) were first used to replace the expensive MgCO(3) for succinic acid production. Using the three aforementioned substitutions, the total fermentation cost decreased by 55.9%, and 56.4 g/L succinic acid with yield of 0.73 g/g was obtained, which are almost the same production level as fermentation with glucose, yeast extract and MgCO(3). Therefore, the cheap carbon and nitrogen sources, as well as the mixed alkaline neutralize could be efficiently used instead of expensive composition for industrial succinic acid production. PMID:21470857

  14. Carob pod water extracts as feedstock for succinic acid production by Actinobacillus succinogenes 130Z.

    PubMed

    Carvalho, Margarida; Roca, Christophe; Reis, Maria A M

    2014-10-01

    Carob pods are a by-product of locust bean gum industry containing more than 50% (w/w) sucrose, glucose and fructose. In this work, carob pod water extracts were used, for the first time, for succinic acid production by Actinobacillus succinogenes 130Z. Kinetic studies of glucose, fructose and sucrose consumption as individual carbon sources till 30g/L showed no inhibition on cell growth, sugar consumption and SA production rates. Sugar extraction from carob pods was optimized varying solid/liquid ratio and extraction time, maximizing sugar recovery while minimizing the extraction of polyphenols. Batch fermentations containing 10-15g/L total sugars resulted in a maximum specific SA production rate of 0.61Cmol/Cmol X.h, with a yield of 0.55Cmol SA/Cmol sugar and a volumetric productivity of 1.61g SA/L.h. Results demonstrate that carob pods can be a promising low cost feedstock for bio-based SA production. PMID:25164341

  15. Succinic acid production by Actinobacillus succinogenes from batch fermentation of mixed sugars.

    PubMed

    Almqvist, Henrik; Pateraki, Chrysanthi; Alexandri, Maria; Koutinas, Apostolis; Lidén, Gunnar

    2016-08-01

    Succinic acid production from the monosaccharides xylose, arabinose, glucose, mannose and galactose was studied using the bacterium Actinobacillus succinogenes. In Duran bottle cultures, containing 10 g/L of each of sugar, succinic acid was produced from all sugars except for galactose. The highest succinate yield, 0.56 g/g, was obtained with glucose, whereas the succinate yield was 0.42, 0.38 and 0.44 g/g for xylose, mannose and arabinose, respectively. The specific succinate productivity was 0.7 g/g h for glucose, but below 0.2 g/g h for the other sugars. Batch bioreactor fermentations were carried out using a sugar mixture of the five sugars giving a total concentration of 50 g/L, mimicking the distribution of sugars in spent sulfite liquor (SSL) from Eucalyptus which is rich in xylose. In this mixture, an almost complete conversion of all sugars (except galactose) was achieved resulting in a final succinate concentration of 21.8-26.8 g/L and a total yield of 0.59-0.68 g/g. There was evidence of co-consumption of glucose and xylose, whereas mannose was consumed after glucose. The main by-products were acetate 0.14-0.20 g/g and formate 0.08-0.13 g/g. NADH balance calculations suggested that NADH required for succinate production was not met solely from formate and acetate production, but other means of NADH production was necessary. Results from mixed sugar fermentations were verified using SSL as substrate resulting in a succinate yield of 0.60 g/g. In addition, it was found that CO2 sparging could replace carbonate supply in the form of MgCO3 without affecting the succinate yield. PMID:27255975

  16. Bagasse hydrolyzates from Agave tequilana as substrates for succinic acid production by Actinobacillus succinogenes in batch and repeated batch reactor.

    PubMed

    Corona-González, Rosa Isela; Varela-Almanza, Karla María; Arriola-Guevara, Enrique; Martínez-Gómez, Álvaro de Jesús; Pelayo-Ortiz, Carlos; Toriz, Guillermo

    2016-04-01

    The aim of this work was to obtain fermentable sugars by enzymatic or acid hydrolyses of Agave tequilana Weber bagasse in order to produce succinic acid with Actinobacillus succinogenes. Hydrolyses were carried out with mineral acids (sulfuric and hydrochloric acids) or a commercial cellulolytic enzyme, and were optimized statistically by a response surface methodology, having as factors the concentration of acid/enzyme and time of hydrolysis. The concentration of sugars obtained at optimal conditions for each hydrolysis were 21.7, 22.4y 19.8g/L for H2SO4, HCl and the enzymatic preparation respectively. Concerning succinic acid production, the enzymatic hydrolyzates resulted in the highest yield (0.446g/g) and productivity (0.57g/Lh) using A. succinogenes in a batch reactor system. Repeated batch fermentation with immobilized A. succinogenes in agar and with the enzymatic hydrolyzates resulted in a maximum concentration of succinic acid of 33.6g/L from 87.2g/L monosaccharides after 5 cycles in 40h, obtaining a productivity of 1.32g/Lh. PMID:26802183

  17. Utilization of CO2 fixating bacterium Actinobacillus succinogenes 130Z for simultaneous biogas upgrading and biosuccinic acid production.

    PubMed

    Gunnarsson, Ingólfur B; Alvarado-Morales, Merlin; Angelidaki, Irini

    2014-10-21

    Biogas is an attractive renewable energy carrier. However, it contains CO2 which limits its use for certain applications. Here we report a novel approach for removing CO2 from biogas and capturing it as a biochemical through a biological process. This approach entails converting CO2 into biosuccinic acid using the bacterial strain Actinobacillus succinogenes 130 Z, and simultaneously producing high-purity CH4 (> 95%). Results showed that when pressure during fermentation was increased from 101.325 to 140 kPa, higher CO2 solubility was achieved, thereby positively affecting final succinic acid yield and titer, CO2 consumption rate, and CH4 purity. When using biogas as the only CO2 source at 140 kPa, the CO2 consumption rate corresponded to 2.59 L CO2 L(-1) d(-1) with a final succinic acid titer of 14.4 g L(-1). Under this pressure condition, the highest succinic acid yield and biogas quality reached corresponded to 0.635 g g(-1) and 95.4% (v v(-1)) CH4 content, respectively, after 24 h fermentation. This work represents the first successful attempt to develop a system capable of upgrading biogas to vehicle fuel/gas grid quality and simultaneously produce biosuccinic acid, a valuable building block with large market potential in the near term. PMID:25275929

  18. Use of corn steep liquor as an economical nitrogen source for biosuccinic acid production by Actinobacillus succinogenes

    NASA Astrophysics Data System (ADS)

    Tan, J. P.; Jahim, J. M.; Wu, T. Y.; Harun, S.; Mumtaz, T.

    2016-06-01

    Expensive raw materials are the driving force that leads to the shifting of the petroleum-based succinic acid production into bio-based succinic acid production by microorganisms. Cost of fermentation medium is among the main factors contributing to the total production cost of bio-succinic acid. After carbon source, nitrogen source is the second largest component of the fermentation medium, the cost of which has been overlooked for the past years. The current study aimed at replacing yeast extract- a costly nitrogen source with corn steep liquor for economical production of bio-succinic acid by Actinobacillus succinogenes 130Z. In this study, a final succinic acid concentration of 20.6 g/L was obtained from the use of corn steep liquor as the nitrogen source, which was comparable with the use of yeast extract as the nitrogen source that had a final succinate concentration of 21.4 g/l. In terms of economical wise, corn steep liquor was priced at 200 /ton, which was one fifth of the cost of yeast extract at 1000 /ton. Therefore, corn steep liquor can be considered as a potential nitrogen source in biochemical industries instead of the costly yeast extract.

  19. Succinic acid production on xylose-enriched biorefinery streams by Actinobacillus succinogenes in batch fermentation

    DOE PAGESBeta

    Salvachua, Davinia; Mohagheghi, Ali; Smith, Holly; Bradfield, Michael F. A.; Nicol, Willie; Black, Brenna A.; Biddy, Mary J.; Dowe, Nancy; Beckham, Gregg T.

    2016-02-02

    Co-production of chemicals from lignocellulosic biomass alongside fuels holds promise for improving the economic outlook of integrated biorefineries. In current biochemical conversion processes that use thermochemical pretreatment and enzymatic hydrolysis, fractionation of hemicellulose-derived and cellulose-derived sugar streams is possible using hydrothermal or dilute acid pretreatment (DAP), which then offers a route to parallel trains for fuel and chemical production from xylose- and glucose-enriched streams. Succinic acid (SA) is a co-product of particular interest in biorefineries because it could potentially displace petroleum-derived chemicals and polymer precursors for myriad applications. Furthermore, SA production from biomass-derived hydrolysates has not yet been fully exploredmore » or developed.« less

  20. Continuous Succinic Acid Production by Actinobacillus succinogenes on Xylose-Enriched Hydrolysate

    DOE PAGESBeta

    Bradfield, Michael F. A.; Mohagheghi, Ali; Salvachua, Davinia; Smith, Holly; Black, Brenna A.; Dowe, Nancy; Beckham, Gregg T.; Nicol, Willie

    2015-11-14

    Bio-manufacturing of high-value chemicals in parallel to renewable biofuels has the potential to dramatically improve the overall economic landscape of integrated lignocellulosic biorefineries. However, this will require the generation of carbohydrate streams from lignocellulose in a form suitable for efficient microbial conversion and downstream processing appropriate to the desired end use, making overall process development, along with selection of appropriate target molecules, crucial to the integrated biorefinery. Succinic acid (SA), a high-value target molecule, can be biologically produced from sugars and has the potential to serve as a platform chemical for various chemical and polymer applications. However, the feasibility ofmore » microbial SA production at industrially relevant productivities and yields from lignocellulosic biorefinery streams has not yet been reported.« less

  1. Continuous Succinic Acid Production by Actinobacillus succinogenes on Xylose-Enriched Hydrolysate

    SciTech Connect

    Bradfield, Michael F. A.; Mohagheghi, Ali; Salvachua, Davinia; Smith, Holly; Black, Brenna A.; Dowe, Nancy; Beckham, Gregg T.; Nicol, Willie

    2015-11-14

    Bio-manufacturing of high-value chemicals in parallel to renewable biofuels has the potential to dramatically improve the overall economic landscape of integrated lignocellulosic biorefineries. However, this will require the generation of carbohydrate streams from lignocellulose in a form suitable for efficient microbial conversion and downstream processing appropriate to the desired end use, making overall process development, along with selection of appropriate target molecules, crucial to the integrated biorefinery. Succinic acid (SA), a high-value target molecule, can be biologically produced from sugars and has the potential to serve as a platform chemical for various chemical and polymer applications. However, the feasibility of microbial SA production at industrially relevant productivities and yields from lignocellulosic biorefinery streams has not yet been reported.

  2. Functional annotation of Fibrobacter succinogenes carbohydrate active enzymes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fibrobacter succinogenes (Fisuc) is a cellulolytic bacterium that degrades plant cell wall biomass in ruminant animals, and is among the most rapidly fibrolytic of all mesophilic bacteria. The complete genome sequence of Fisuc was completed by the DOE Joint Genome Institute in late 2009. Using new e...

  3. Functional annotation of Fibrobacter succinogenes S85 carbohydrate active enzymes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fibrobacter succinogenes is a cellulolytic bacterium that degrades plant cell wall biomass in ruminant animals and is among the most rapidly fibrolytic of all mesophilic bacteria. The complete genome sequence of Fisuc was completed by the DOE Joint Genome Institute in late 2009. Using new expression...

  4. Kinetics of Cellulose Digestion by Fibrobacter succinogenes S85

    PubMed Central

    Maglione, G.; Russell, J. B.; Wilson, D. B.

    1997-01-01

    Growing cultures of Fibrobacter succinogenes S85 digested cellulose at a rapid rate, but nongrowing cells and cell extracts did not have detectable crystalline cellulase activity. Cells that had been growing exponentially on cellobiose initiated cellulose digestion and succinate production immediately, and cellulose-dependent succinate production could be used as an index of enzyme activity against crystalline cellulose. Cells incubated with cellulose never produced detectable cellobiose, and cells that were preincubated for a short time with thiocellobiose lost their ability to digest cellulose (competitive inhibition [K(infi)] of only 0.2 mg/ml or 0.56 mM). Based on these results, the crystalline cellulases of F. succinogenes were very sensitive to feedback inhibition. Different cellulose sources bound different amounts of Congo red, and the binding capacity was HCl-regenerated cellulose > ball-milled cellulose > Sigmacel > Avicel > filter paper. Congo red binding capacity was highly correlated with the maximum rates of metabolism of cellulose digestion and inversely related to K(infm). Congo red (250 (mu)g/ml) did not inhibit the growth of F. succinogenes S85 on cellobiose, but this concentration of Congo red inhibited the rate of ball-milled cellulose digestion. A Lineweaver-Burk plot of ball-milled cellulose digestion rate versus the amount of cellulose indicated that Congo red was a competitive inhibitor of cellulose digestion (K(infi) was 250 (mu)g/ml). PMID:16535519

  5. An unconventional anaerobic membrane protein production system based on Wolinella succinogenes.

    PubMed

    Lafontaine, Michael; Lancaster, C Roy D

    2015-01-01

    In cases where membrane protein production attempts in more conventional Escherichia coli-based systems have failed, a solution is to resort to a system based on the nonpathogenic epsilon-proteobacterium Wolinella succinogenes. This approach has been demonstrated to be successful for structural and mechanistic analyses not only for homologous production of W. succinogenes membrane proteins but also for the heterologous production of membrane protein complexes from the human pathogens Helicobacter pylori and Campylobacter jejuni. The procedure to establish a system for the production of native and variant enzymes in W. succinogenes is presented in detail for the examples of the quinol:fumarate reductase and the SdhABE complexes of W. succinogenes. Subsequently, further projects using W. succinogenes as expression host are covered. PMID:25857779

  6. Evaluating Models of Cellulose Degradation by Fibrobacter succinogenes S85

    PubMed Central

    Burnet, Meagan C.; Dohnalkova, Alice C.; Neumann, Anthony P.; Lipton, Mary S.; Smith, Richard D.; Suen, Garret; Callister, Stephen J.

    2015-01-01

    Fibrobacter succinogenes S85 is an anaerobic non-cellulosome utilizing cellulolytic bacterium originally isolated from the cow rumen microbial community. Efforts to elucidate its cellulolytic machinery have resulted in the proposal of numerous models which involve cell-surface attachment via a combination of cellulose-binding fibro-slime proteins and pili, the production of cellulolytic vesicles, and the entry of cellulose fibers into the periplasmic space. Here, we used a combination of RNA-sequencing, proteomics, and transmission electron microscopy (TEM) to further clarify the cellulolytic mechanism of F. succinogenes. Our RNA-sequence analysis shows that genes encoding type II and III secretion systems, fibro-slime proteins, and pili are differentially expressed on cellulose, relative to glucose. A subcellular fractionation of cells grown on cellulose revealed that carbohydrate active enzymes associated with cellulose deconstruction and fibro-slime proteins were greater in the extracellular medium, as compared to the periplasm and outer membrane fractions. TEMs of samples harvested at mid-exponential and stationary phases of growth on cellulose and glucose showed the presence of grooves in the cellulose between the bacterial cells and substrate, suggesting enzymes work extracellularly for cellulose degradation. Membrane vesicles were only observed in stationary phase cultures grown on cellulose. These results provide evidence that F. succinogenes attaches to cellulose fibers using fibro-slime and pili, produces cellulases, such as endoglucanases, that are secreted extracellularly using type II and III secretion systems, and degrades the cellulose into cellodextrins that are then imported back into the periplasm for further digestion by β-glucanases and other cellulases. PMID:26629814

  7. Evaluating models of cellulose degradation by Fibrobacter succinogenes S85

    DOE PAGESBeta

    Burnet, Meagan C.; Dohnalkova, Alice C.; Neumann, Anthony P.; Lipton, Mary S.; Smith, Richard D.; Suen, Garret; Callister, Stephen J.

    2015-12-02

    Fibrobacter succinogenes S85 is an anaerobic non-cellulosome utilizing cellulolytic bacterium originally isolated from the cow rumen microbial community. Efforts to elucidate its cellulolytic machinery have resulted in the proposal of numerous models which involve a combination of cell-surface attachment via a combination of cellulose-binding fibro-slime proteins and pili, the production of cellulolytic vesicles, and the entry of cellulose fibers into the periplasmic space. Here, we used a combination of RNA-sequencing, proteomics, and transmission electron microscopy (TEM) to further elucidate the cellulolytic mechanism of F. succinogenes. Our RNA-sequence analysis shows that genes encoding Type II and III secretion systems, fibro-slime proteins,more » and pili are differentially expressed on cellulose, relative to glucose. A subcellular fractionation of cells grown on cellulose revealed that carbohydrate active enzymes associated with cellulose deconstruction and fibro-slime proteins were greater in the extracellular media, as compared to the periplasm and outer membrane fractions. TEMs of samples harvested at mid-exponential and stationary phases of growth on cellulose and glucose showed the presence of grooves in the cellulose between the bacterial cells and substrate, suggesting enzymes work extracellularly for cellulose degradation. Membrane vesicles were only observed in stationary phase cultures grown on cellulose. Furthermore, these results provide evidence that F. succinogenes attaches to cellulose fibers using fibro-slime and pili, produces cellulases, such as endoglucanases, that are secreted extracellularly using type II and III secretion systems, and degrades the cellulose into cellodextrins that are then imported back into the periplasm for further digestion by β-glucanases and other cellulases.« less

  8. Actinobacillus suis septicaemia in two foals.

    PubMed

    Nelson, K M; Darien, B J; Konkle, D M; Hartmann, F A

    1996-01-13

    A 24-hour-old Hackney ony filly developed signs of weakness, depression and a poor suck reflex, with harsh lung sounds over both fields, and a 48-hour-old Arabian colt from a normal birth which had sucked vigorously developed loose stools and became depressed, weak and anorectic. Both foals had serum IgG concentrations greater than 800 mg/dl, but each had a severe neutropenia with a left shift, and blood cultures from both of them yielded Actinobacillus suis. The A suis isolates had different antimicrobial susceptibility patterns and, in the case of the Arabian, the isolate was resistant to commonly used broad spectrum antimicrobial agents. PMID:8629322

  9. Identification of an anaerobic bacterium which reduces perchlorate and chlorate as Wolinella succinogenes

    SciTech Connect

    Wallace, W.; Attaway, H. |

    1995-12-31

    Perchlorate and chlorate salts are widely used by the chemical, aerospace and defense industries as oxidizers in propellant, explosives and pyrotechnics. The authors have isolated a anaerobic bacterium which is capable of the dissimilatory reduction of both perchlorate and chlorate for energy and growth. Strain HAP-1 is a gram negative, thin rod, non-sporeforming, highly motile strict anaerobe. Antibiotic resistance profiles, utilization of carbon substrates and electron acceptors demonstrated similar physiological characteristics to Wolinella succinogenes. Pairwise comparisons of 16S RNA sequences showed only a 0.75% divergence between strain HAP-1 and W. succinogenes. Physiological, morphological and 16S RRNA sequence data indicate strain HAP-1 is a subspecies of W. succinogenes that can utilize perchlorate and chlorate as terminal electron acceptors.

  10. The Complete Genome Sequence of Fibrobacter succinogenes S85 Reveals a Cellulolytic and Metabolic Specialist

    PubMed Central

    Suen, Garret; Weimer, Paul J.; Stevenson, David M.; Aylward, Frank O.; Boyum, Julie; Deneke, Jan; Drinkwater, Colleen; Ivanova, Natalia N.; Mikhailova, Natalia; Chertkov, Olga; Goodwin, Lynne A.; Currie, Cameron R.; Mead, David; Brumm, Phillip J.

    2011-01-01

    Fibrobacter succinogenes is an important member of the rumen microbial community that converts plant biomass into nutrients usable by its host. This bacterium, which is also one of only two cultivated species in its phylum, is an efficient and prolific degrader of cellulose. Specifically, it has a particularly high activity against crystalline cellulose that requires close physical contact with this substrate. However, unlike other known cellulolytic microbes, it does not degrade cellulose using a cellulosome or by producing high extracellular titers of cellulase enzymes. To better understand the biology of F. succinogenes, we sequenced the genome of the type strain S85 to completion. A total of 3,085 open reading frames were predicted from its 3.84 Mbp genome. Analysis of sequences predicted to encode for carbohydrate-degrading enzymes revealed an unusually high number of genes that were classified into 49 different families of glycoside hydrolases, carbohydrate binding modules (CBMs), carbohydrate esterases, and polysaccharide lyases. Of the 31 identified cellulases, none contain CBMs in families 1, 2, and 3, typically associated with crystalline cellulose degradation. Polysaccharide hydrolysis and utilization assays showed that F. succinogenes was able to hydrolyze a number of polysaccharides, but could only utilize the hydrolytic products of cellulose. This suggests that F. succinogenes uses its array of hemicellulose-degrading enzymes to remove hemicelluloses to gain access to cellulose. This is reflected in its genome, as F. succinogenes lacks many of the genes necessary to transport and metabolize the hydrolytic products of non-cellulose polysaccharides. The F. succinogenes genome reveals a bacterium that specializes in cellulose as its sole energy source, and provides insight into a novel strategy for cellulose degradation. PMID:21526192

  11. Cellulase producing microorganism ATCC 55702

    DOEpatents

    Dees, H. Craig

    1997-01-01

    Bacteria which produce large amounts of cellulase--containing cell-free fermentate have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase producing bacterium (ATCC 55702), which was identified through replicate plating. ATCC 55702 has improved characteristics and qualifies for the degradation of cellulosic waste materials for fuel production, food processing, textile processing, and other industrial applications. ATCC 55702 is an improved bacterial host for genetic manipulations using recombinant DNA techniques, and is less likely to destroy genetic manipulations using standard mutagenesis techniques.

  12. Cellulase producing microorganism ATCC 55702

    DOEpatents

    Dees, H.C.

    1997-12-30

    Bacteria which produce large amounts of cellulase--containing cell-free fermentate have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase producing bacterium (ATCC 55702), which was identified through replicate plating. ATCC 55702 has improved characteristics and qualifies for the degradation of cellulosic waste materials for fuel production, food processing, textile processing, and other industrial applications. ATCC 55702 is an improved bacterial host for genetic manipulations using recombinant DNA techniques, and is less likely to destroy genetic manipulations using standard mutagenesis techniques. 5 figs.

  13. Survival of Actinobacillus pleuropneumoniae outside the pig.

    PubMed

    Assavacheep, P; Rycroft, A N

    2013-02-01

    Transmission of Actinobacillus pleuropneumoniae is primarily thought to be via direct transfer of mucus from pig to pig. For transfer between farms, the organism may need to persist in the wet or dried state to be carried on an inanimate surface. The survival of A. pleuropneumoniae was investigated under controlled laboratory conditions. In aqueous suspension, survival was improved by the presence of NaCl and mucin; it was prolonged at lower temperature. In dry state, it survived best on a hydrophobic surface either under desiccated conditions or saturated humidity. Detectable viability was maintained for 3-4 days. When frozen, A. pleuropneumoniae survived for more than 17 weeks at -20 °C, but the viability declined to 0.01% during that time. Survival at -70 °C was effective for long term storage. Results obtained from this investigation would be applicable for sampling method, transport techniques, epidemiological study, and biosecurity implementation. PMID:22892250

  14. Release of toxic microvesicles by Actinobacillus actinomycetemcomitans.

    PubMed Central

    Nowotny, A; Behling, U H; Hammond, B; Lai, C H; Listgarten, M; Pham, P H; Sanavi, F

    1982-01-01

    Oral isolates of Actinobacillus actinomycetemcomitans (strain Y4) release spherical microvesicles in large numbers during normal growth. The biological activities of these products were studied, and it was estimated that approximately 1/10 of their dry weight was made up of heat- and proteolysis-resistant endotoxin. The chicken embryo lethality and bone-resorbing activity of the microvesicles were heat stable but proteolysis sensitive. Other laboratories have reported the presence of a heat- and proteolysis-sensitive leukotoxin in similar preparations. Accordingly, the microvesicles released by strain Y4 may contain, in addition to endotoxin, several potent substances which are highly toxic and active in bone resorption, and these may be significant factors in the pathogenesis of periodontal diseases. PMID:7049947

  15. Release of toxic microvesicles by Actinobacillus actinomycetemcomitans.

    PubMed

    Nowotny, A; Behling, U H; Hammond, B; Lai, C H; Listgarten, M; Pham, P H; Sanavi, F

    1982-07-01

    Oral isolates of Actinobacillus actinomycetemcomitans (strain Y4) release spherical microvesicles in large numbers during normal growth. The biological activities of these products were studied, and it was estimated that approximately 1/10 of their dry weight was made up of heat- and proteolysis-resistant endotoxin. The chicken embryo lethality and bone-resorbing activity of the microvesicles were heat stable but proteolysis sensitive. Other laboratories have reported the presence of a heat- and proteolysis-sensitive leukotoxin in similar preparations. Accordingly, the microvesicles released by strain Y4 may contain, in addition to endotoxin, several potent substances which are highly toxic and active in bone resorption, and these may be significant factors in the pathogenesis of periodontal diseases. PMID:7049947

  16. The complete genome sequence of Fibrobacter succinogenes S85 reveals a cellulolytic and metabolic specialist

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fibrobacter succinogenes S85 is an important member of the rumen microbial community that converts plant biomass into nutrients usable by its host. This bacterium, which is also one of two known species in its phylum, is an efficient and prolific degrader of cellulose. Specifically, it has a particu...

  17. Molecular cloning and expression in Escherichia coli of a cellodextrinase gene from Bacteroides succinogenes S85.

    PubMed Central

    Gong, J H; Lo, R Y; Forsberg, C W

    1989-01-01

    A DNA fragment coding for a cellodextrinase of Bacteroides succinogenes S85 was isolated by screening of a pBR322 gene library in Escherichia coli HB101. Of 100,000 colonies screened on a complex medium with methylumbelliferyl-beta-D-cellobioside as the indicator substrate, two cellodextrinase-positive clones (CB1 and CB2) were isolated. The DNA inserts from the two recombinant plasmids were 7.7 kilobase pairs in size and had similar restriction maps. After subcloning from pCB2, a 2.5-kilobase-pair insert which coded for cellodextrinase activity was isolated. The enzyme was located in the cytoplasm of the E. coli host. It exhibited no activity on carboxymethyl cellulose, Avicel microcrystalline cellulose, acid-swollen cellulose, or cellobiose but hydrolyzed p-nitrophenyl-beta-D-cellobioside and p-nitrophenyl-beta-D-lactoside. The Km (0.1 mM) for the hydrolysis of p-nitrophenyl-cellobioside by the enzyme expressed in E. coli was similar to that reported for the purified enzyme from B. succinogenes. Expression of the cellodextrinase gene was subjected to catabolite repression by glucose and was not induced by cellobiose. The origin of the DNA insert from B. succinogenes was confirmed by Southern blot analysis. Western blotting (immunoblotting) using antibodies raised against the purified B. succinogenes cellodextrinase revealed a protein with a molecular weight of approximately 50,000 in E. coli clones which comigrated with the native enzyme isolated from B. succinogenes. These data indicate that the cellodextrinase gene expressed in E. coli is fully functional and codes for an enzyme with properties similar to those of the native enzyme. Images PMID:2650617

  18. Inhibition of fibroblast proliferation by Actinobacillus actinomycetemcomitans.

    PubMed Central

    Shenker, B J; Kushner, M E; Tsai, C C

    1982-01-01

    We have examined soluble sonic extracts of Actinobacillus actinomycetemcomitans for their ability to alter human and murine fibroblast proliferation. We found that extracts of all A. actinomycetemcomitans strains examined (both leukotoxic and nonleukotoxic) caused a dose-dependent inhibition of both murine and human fibroblast proliferation as assessed by DNA synthesis ([3H]thymidine incorporation). Addition of sonic extract simultaneously with [3H]thymidine had no effect on incorporation, indicating that suppression was not due to the presence of excessive amounts of cold thymidine. Inhibition of DNA synthesis was also paralleled by decreased RNA synthesis ([3H]uridine incorporation) and by a decrease in cell growth as assessed by direct cell counts; there was no effect on cell viability. The suppressive factor(s) is heat labile; preliminary purification and characterization studies indicate that it is a distinct and separate moiety from other A. actinomycetemcomitans mediators previously reported, including leukotoxin, immune suppressive factor, and endotoxin. Although it is not clear how A. actinomycetemcomitans acts to cause disease, we propose that one aspect of the pathogenicity of this organism rests in its ability to inhibit fibroblast growth, which in turn could contribute to the collagen loss associated with certain forms of periodontal disease, in particular juvenile periodontitis. PMID:7152684

  19. Killing of Actinobacillus actinomycetemcomitans by human lactoferrin.

    PubMed Central

    Kalmar, J R; Arnold, R R

    1988-01-01

    Actinobacillus actinomycetemcomitans is a fastidious, facultative gram-negative rod associated with endocarditis, certain forms of periodontal disease, and other focal infections. Human neutrophils have demonstrated bactericidal activity against A. actinomycetemcomitans, and much of the oxygen-dependent killing has been attributed to the myeloperoxidase-H2O2-halide system. However, the contribution of other neutrophil components to killing activity is obscure. Lactoferrin, an iron-binding glycoprotein, is a major constituent of neutrophil-specific granules and is also found in mucosal secretions. In this report, we show that human lactoferrin is bactericidal for A. actinomycetemcomitans. Killing activity required an unsaturated (iron- and anion-free) molecule that produced a 2-log decrease in viability within 120 min at 37 degrees C at a concentration of 1.9 microM. Besides exhibiting concentration dependence, killing kinetics were affected by minor variations in temperature and pH. Magnesium, a divalent cation thought to stabilize lipopolysaccharide interactions on the surface of gram-negative organisms, enhanced lactoferrin killing of A. actinomycetemcomitans, while other cations, such as potassium and calcium, had no effect. Our data suggest that lactoferrin contributes to killing of A. actinomycetemcomitans by human neutrophils and that it may also play a significant role in innate secretory defense against this potential periodontopathogen. PMID:3417349

  20. Immunosuppressive properties of Actinobacillus actinomycetemcomitans leukotoxin.

    PubMed Central

    Rabie, G; Lally, E T; Shenker, B J

    1988-01-01

    Actinobacillus actinomycetemcomitans produces a leukotoxin that kills human polymorphonuclear cells (PMNs) and monocytes but not lymphocytes. In this study, we examined A. actinomycetemcomitans leukotoxin for its ability to alter human peripheral blood lymphocyte (HPBL) responsiveness. After a 90-min exposure to the leukotoxin, all monocytes were killed and HPBL responsiveness to mitogens and antigens was significantly inhibited. The ability of the leukotoxin to inhibit HPBL responses was not surprising, since monocytes and macrophages are required for many lymphocyte functions. However, we were unable to totally restore HPBL responsiveness when adherent autologous monocytes were added back to cultures of leukotoxin-treated lymphocytes. These studies demonstrate that A. actinomycetemcomitans leukotoxin may also exert nonlethal effects directly on lymphocytes. Furthermore, impaired lymphocyte function did not appear to be the result of indirect effects of products released by dying monocytes. Although it is not clear how A. actinomycetemcomitans acts to cause disease, several investigators have proposed that impaired host defenses may play a pivotal role. Several studies have demonstrated defects in PMN, monocyte, and lymphocyte function in patients with periodontal disease. These findings, along with the data presented in this paper, support the hypothesis that patients who harbor A. actinomycetemcomitans could suffer from local or systemic immune suppression. The effects of this suppression may be to enhance the pathogenicity of A. actinomycetemcomitans itself or that of some other opportunistic organism. PMID:3335399

  1. Rapid detection of Actinobacillus actinomycetemcomitans, Prevotella intermedia and Porphyromona gingivalis by multiplex PCR.

    PubMed

    García, L; Tercero, J C; Legido, B; Ramos, J A; Alemany, J; Sanz, M

    1998-01-01

    The identification of specific periodontal pathogens by conventional methods, mainly anaerobic cultivation, is difficult, time consuming and even sometimes unreliable. Therefore, a multiplex PCR method for simultaneous detection of Actinobacillus actinomycetemcomitans (A.a.), Porphyromona gingivalis (P.g.) and Prevotella intermedia (P.i.) was developed for rapid and easy identification of these specific bacterial pathogens in subgingival plaque samples. In this paper, there is a detailed description of the oligonucleotide primer selection, DNA extraction and PCR conditions and the sequencing of the amplified products. The locus chosen to be amplified is a highly variable region in the 16S ribosomal DNA. For the development of this technique ATCC cultures and pure cultures from subgingival plaque samples taken from periodontitis patients were used. As an internal positive control a recombinant plasmid was developed. This simple DNA extraction procedure and the DNA amplification and visualization of the amplified product permits the detection of the bacteria in a working day. Thus, this multiplex PCR method is a rapid and effective detection method for specific periodontal pathogens. PMID:9524322

  2. Isolation of Actinobacillus suis from a cat's lung.

    PubMed Central

    Daignault, D; Chouinard, L; Møller, K; Ahrens, P; Messier, S; Higgins, R

    1999-01-01

    Actinobacillus suis has been isolated from the lungs of 9-month-old cat. The bacterium was characterized biochemically as well as genetically, and its sensitivity profile to different antimicrobial agents was established. The role of this isolate in the cat's condition is discussed. PMID:9919368

  3. Biofilm Growth and Detachment of Actinobacillus actinomycetemcomitans

    PubMed Central

    Kaplan, Jeffrey B.; Meyenhofer, Markus F.; Fine, Daniel H.

    2003-01-01

    The gram-negative, oral bacterium Actinobacillus actinomycetemcomitans has been implicated as the causative agent of several forms of periodontal disease in humans. When cultured in broth, fresh clinical isolates of A. actinomycetemcomitans form tenacious biofilms on surfaces such as glass, plastic, and saliva-coated hydroxyapatite, a property that probably plays an important role in the ability of this bacterium to colonize the oral cavity and cause disease. We examined the morphology of A. actinomycetemcomitans biofilm colonies grown on glass slides and in polystyrene petri dishes by using light microscopy and scanning and transmission electron microscopy. We found that A. actinomycetemcomitans developed asymmetric, lobed biofilm colonies that displayed complex architectural features, including a layer of densely packed cells on the outside of the colony and nonaggregated cells and large, transparent cavities on the inside of the colony. Mature biofilm colonies released single cells or small clusters of cells into the medium. These released cells adhered to the surface of the culture vessel and formed new colonies, enabling the biofilm to spread. We isolated three transposon insertion mutants which produced biofilm colonies that lacked internal, nonaggregated cells and were unable to release cells into the medium. All three transposon insertions mapped to genes required for the synthesis of the O polysaccharide (O-PS) component of lipopolysaccharide. Plasmids carrying the complementary wild-type genes restored the ability of mutant strains to synthesize O-PS and release cells into the medium. Our findings suggest that A. actinomycetemcomitans biofilm growth and detachment are discrete processes and that biofilm cell detachment evidently involves the formation of nonaggregated cells inside the biofilm colony that are destined for release from the colony. PMID:12562811

  4. Lipopolysaccharides of Actinobacillus pleuropneumoniae bind pig hemoglobin.

    PubMed Central

    Bélanger, M; Bégin, C; Jacques, M

    1995-01-01

    A previous study indicated that lipopolysaccharides (LPS) extracted from Actinobacillus pleuropneumoniae bind two low-molecular-mass proteins, of approximately 10 and 11 kDa, present in porcine respiratory tract secretions (M. Bélanger, D. Dubreuil, and M. Jacques, Infect. Immun. 62:868-873, 1994). In the present study, we determined the N-terminal amino acid sequences of these two proteins, which revealed high homology with the alpha and beta chains of pig hemoglobin. Some isolates of A. pleuropneumoniae were able to use hemoglobin from various animal species as well as other heme compounds as sole sources of iron for growth, while other isolates were unable to use them. Immunoelectron microscopy showed binding of pig hemoglobin at the surface of all A. pleuropneumoniae isolates as well as labeling of outer membrane blebs. We observed, using Western blotting (immunoblotting), that the lipid A-core region of LPS of all isolates was binding pig hemoglobin. Furthermore, lipid A obtained after acid hydrolysis of LPS extracted from A. pleuropneumoniae was able to bind pig hemoglobin and this binding was completely abolished by preincubation of lipid A with polymyxin B but was not inhibited by preincubation with glucosamines. Fatty acids constituting the lipid A of A. pleuropneumoniae, namely, dodecanoic acid, tetradecanoic acid, 3-hydroxytetradecanoic acid, hexadecanoic acid, and octadecanoic acid, were also binding pig hemoglobin. Our results indicate that LPS of all A. pleuropneumoniae isolates tested bind pig hemoglobin and that lipid A is involved in this binding. Our results also indicate that some A. pleuropneumoniae isolates are, in addition, able to use hemoglobin for growth. Binding of hemoglobin to LPS might represent an important means by which A. pleuropneumoniae acquires iron in vivo from hemoglobin released from erythrocytes lysed by the action of its hemolysins. PMID:7822035

  5. Evaluating models of cellulose degradation by Fibrobacter succinogenes S85

    SciTech Connect

    Burnet, Meagan C.; Dohnalkova, Alice C.; Neumann, Anthony P.; Lipton, Mary S.; Smith, Richard D.; Suen, Garret; Callister, Stephen J.

    2015-12-02

    Fibrobacter succinogenes S85 is an anaerobic non-cellulosome utilizing cellulolytic bacterium originally isolated from the cow rumen microbial community. Efforts to elucidate its cellulolytic machinery have resulted in the proposal of numerous models which involve a combination of cell-surface attachment via a combination of cellulose-binding fibro-slime proteins and pili, the production of cellulolytic vesicles, and the entry of cellulose fibers into the periplasmic space. Here, we used a combination of RNA-sequencing, proteomics, and transmission electron microscopy (TEM) to further elucidate the cellulolytic mechanism of F. succinogenes. Our RNA-sequence analysis shows that genes encoding Type II and III secretion systems, fibro-slime proteins, and pili are differentially expressed on cellulose, relative to glucose. A subcellular fractionation of cells grown on cellulose revealed that carbohydrate active enzymes associated with cellulose deconstruction and fibro-slime proteins were greater in the extracellular media, as compared to the periplasm and outer membrane fractions. TEMs of samples harvested at mid-exponential and stationary phases of growth on cellulose and glucose showed the presence of grooves in the cellulose between the bacterial cells and substrate, suggesting enzymes work extracellularly for cellulose degradation. Membrane vesicles were only observed in stationary phase cultures grown on cellulose. Furthermore, these results provide evidence that F. succinogenes attaches to cellulose fibers using fibro-slime and pili, produces cellulases, such as endoglucanases, that are secreted extracellularly using type II and III secretion systems, and degrades the cellulose into cellodextrins that are then imported back into the periplasm for further digestion by β-glucanases and other cellulases.

  6. Production and consumption of nitrous oxide in nitrate-ammonifying Wolinella succinogenes cells.

    PubMed

    Luckmann, Monique; Mania, Daniel; Kern, Melanie; Bakken, Lars R; Frostegård, Asa; Simon, Jörg

    2014-08-01

    Global warming is moving more and more into the public consciousness. Besides the commonly mentioned carbon dioxide and methane, nitrous oxide (N2O) is a powerful greenhouse gas in addition to its contribution to depletion of stratospheric ozone. The increasing concern about N2O emission has focused interest on underlying microbial energy-converting processes and organisms harbouring N2O reductase (NosZ), such as denitrifiers and ammonifiers of nitrate and nitrite. Here, the epsilonproteobacterial model organism Wolinella succinogenes is investigated with regard to its capacity to produce and consume N2O during growth by anaerobic nitrate ammonification. This organism synthesizes an unconventional cytochrome c nitrous oxide reductase (cNosZ), which is encoded by the first gene of an atypical nos gene cluster. However, W. succinogenes lacks a nitric oxide (NO)-producing nitrite reductase of the NirS- or NirK-type as well as an NO reductase of the Nor-type. Using a robotized incubation system, the wild-type strain and suitable mutants of W. succinogenes that either produced or lacked cNosZ were analysed as to their production of NO, N2O and N2 in both nitrate-sufficient and nitrate-limited growth medium using formate as electron donor. It was found that cells growing in nitrate-sufficient medium produced small amounts of N2O, which derived from nitrite and, most likely, from the presence of NO. Furthermore, cells employing cNosZ were able to reduce N2O to N2. This reaction, which was fully inhibited by acetylene, was also observed after adding N2O to the culture headspace. The results indicate that W. succinogenes cells are competent in N2O and N2 production despite being correctly grouped as respiratory nitrate ammonifiers. N2O production is assumed to result from NO detoxification and nitrosative stress defence, while N2O serves as a terminal electron acceptor in anaerobic respiration. The ecological implications of these findings are discussed. PMID:24781903

  7. First isolation of Actinobacillus genomospecies 2 in Japan

    PubMed Central

    MURAKAMI, Miyuki; SHIMONISHI, Yoshimasa; HOBO, Seiji; NIWA, Hidekazu; ITO, Hiroya

    2015-01-01

    We describe here the first isolation of Actinobacillus genomospecies 2 in Japan. The isolate was found in a septicemic foal and characterized by phenotypic and genetic analyses, with the latter consisting of 16S rDNA nucleotide sequence analysis plus multilocus sequence analysis using three housekeeping genes, recN, rpoA and thdF, that have been proposed for use as a genomic tool in place of DNA-DNA hybridization. PMID:26668165

  8. First Human Case of Meningitis and Sepsis in a Child Caused by Actinobacillus suis or Actinobacillus equuli

    PubMed Central

    Montagnani, Carlotta; Pecile, Patrizia; Moriondo, Maria; Petricci, Patrizia; Becciani, Sabrina; Chiappini, Elena; Indolfi, Giuseppe; Rossolini, Gian Maria; de Martino, Maurizio

    2015-01-01

    We report the first human case of meningitis and sepsis caused in a child by Actinobacillus suis or A. equuli, a common opportunistic pathogen of swine or horses, respectively. Identification was performed by matrix-assisted laser desorption ionization–time of flight mass spectrometry and real-time PCR assay. A previous visit to a farm was suspected as the source of infection. PMID:25878346

  9. Molecular cloning, expression, and characterization of endoglucanase genes from Fibrobacter succinogenes AR1.

    PubMed Central

    Cavicchioli, R; Watson, K

    1991-01-01

    A cosmid gene library was constructed in Escherichia coli from genomic DNA isolated from the ruminal anaerobe Fibrobacter succinogenes AR1. Clones were screened on carboxymethyl cellulose, and 8 colonies that produced large clearing zones and 25 colonies that produced small clearing zones were identified. Southern blot hybridization revealed the existence of at least three separate genes encoding cellulase activity. pRC093, which is representative of cosmid clones that produce large clearing zones, was subcloned in pGem-1, and the resulting hybrid pRCEH directed synthesis of endoglucanase activity localized on a 2.1-kb EcoRI-HindIII insert. Activity was expressed from this fragment when it was cloned in both orientations in pGem-1 and pGem-2, indicating that F. succinogenes promoters functioned successfully in E. coli. A high level of endoglucanase activity was detected on acid-swollen cellulose, ball-milled cellulose, and carboxymethyl cellulose; and a moderate level was detected on filter paper, Avicel, lichenan, and xylan. Most activity (80%) was localized in the periplasm of E. coli, with low but significant levels (16%) being detected in the extracellular medium. The periplasmic endoglucanase had an estimated molecular weight of 46,500, had an optimum temperature of 39 degrees C, and exhibited activity over a broad pH range, with a maximum at pH 5.0. Images PMID:2014986

  10. Analysis of a continuous culture of Fibrobacter succinogenes S85 on a standardized glucose medium.

    PubMed

    Guiavarch, E; Pons, A; Christophe, G; Creuly, C; Dussap, C-G

    2010-05-01

    Continuous cultures of Fibrobacter succinogenes S85 were performed on a standardized fully synthetic culture medium with glucose as carbon source at a dilution rate (D = 0.02 h(-1)) in a 5-L bioreactor. The culture was stabilized during 20 days and demonstrated the ability of Fibrobacter succinogenes to grow in this synthetic medium. CO(2) partial pressure and redox potential probes were used to check the anaerobic state of the culture. The biomass yield was calculated 0.206 g (g glucose)(-1) and the production yield of succinate, the major end-product, was 0.63 mol (mol glucose)(-1). The consistency of the experimental data was checked by proton and mass (C, N) balances. The results were satisfactory (90-110% recovery) leading to derive a stoichiometric equation representative of the growth on glucose. The stoichiometric coefficients were calculated using data reconciliation and linear algebra methods enabling to obtain a complete modeling of all conversion yields possible. PMID:19548008

  11. Reclassification of Actinobacillus muris as Muribacter muris gen. nov., comb. nov.

    PubMed

    Nicklas, Werner; Bisgaard, Magne; Aalbæk, Bent; Kuhnert, Peter; Christensen, Henrik

    2015-10-01

    To reinvestigate the taxonomy of [Actinobacillus] muris, 474 strains, mainly from mice and rats, were characterized by phenotype and 130 strains selected for genotypic characterization by 16S rRNA and partial rpoB gene sequencing. The type strain was further investigated by whole-genome sequencing. Phylogenetic analysis of the DNA sequences showed one monophyletic group with intragroup similarities of 96.7 and 97.2 % for the 16S rRNA and rpoB genes, respectively. The highest 16S rRNA gene sequence similarity to a taxon with a validly published name outside the group was 95.9 %, to the type strain of [Pasteurella] pneumotropica. The closest related taxon based on rpoB sequence comparison was 'Haemophilus influenzae-murium', with 88.4 % similarity. A new genus and a new combination, Muribacter muris gen. nov., comb. nov., are proposed based on a distinct phylogenetic position based on 16S rRNA and rpoB gene sequence comparisons, with major divergence from the existing genera of the family Pasteurellaceae. The new genus has the characteristics of [A.] muris with the emendation that acid formation from ( - )-d-mannitol and hydrolysis of aesculin are variable, while the α-glucosidase test is positive. There is no requirement for exogenously supplied NAD (V factor) for the majority of strains investigated; however, one strain was found to require NAD. The major fatty acids of the type strain of Muribacter muris were C14 : 0, C14 : 0 3-OH/iso-C16 : 1 I, C16 : 1ω7c and C16 : 0, which is in line with most genera of the Pasteurellaceae. The type strain of Muribacter muris is CCUG 16938T ( = NCTC 12432T = ATCC 49577T). PMID:26296776

  12. Influence of Substrates on the Surface Characteristics and Membrane Proteome of Fibrobacter succinogenes S85

    PubMed Central

    Raut, Mahendra P.; Karunakaran, Esther; Mukherjee, Joy; Biggs, Catherine A.; Wright, Phillip C.

    2015-01-01

    Although Fibrobacter succinogenes S85 is one of the most proficient cellulose degrading bacteria among all mesophilic organisms in the rumen of herbivores, the molecular mechanism behind cellulose degradation by this bacterium is not fully elucidated. Previous studies have indicated that cell surface proteins might play a role in adhesion to and subsequent degradation of cellulose in this bacterium. It has also been suggested that cellulose degradation machinery on the surface may be selectively expressed in response to the presence of cellulose. Based on the genome sequence, several models of cellulose degradation have been suggested. The aim of this study is to evaluate the role of the cell envelope proteins in adhesion to cellulose and to gain a better understanding of the subsequent cellulose degradation mechanism in this bacterium. Comparative analysis of the surface (exposed outer membrane) chemistry of the cells grown in glucose, acid-swollen cellulose and microcrystalline cellulose using physico-chemical characterisation techniques such as electrophoretic mobility analysis, microbial adhesion to hydrocarbons assay and Fourier transform infra-red spectroscopy, suggest that adhesion to cellulose is a consequence of an increase in protein display and a concomitant reduction in the cell surface polysaccharides in the presence of cellulose. In order to gain further understanding of the molecular mechanism of cellulose degradation in this bacterium, the cell envelope-associated proteins were enriched using affinity purification and identified by tandem mass spectrometry. In total, 185 cell envelope-associated proteins were confidently identified. Of these, 25 proteins are predicted to be involved in cellulose adhesion and degradation, and 43 proteins are involved in solute transport and energy generation. Our results supports the model that cellulose degradation in F. succinogenes occurs at the outer membrane with active transport of cellodextrins across for

  13. Electron transport to periplasmic nitrate reductase (NapA) of Wolinella succinogenes is independent of a NapC protein.

    PubMed

    Simon, Jörg; Sänger, Monica; Schuster, Stephan C; Gross, Roland

    2003-07-01

    The rumen bacterium Wolinella succinogenes grows by respiratory nitrate ammonification with formate as electron donor. Whereas the enzymology and coupling mechanism of nitrite respiration is well known, nitrate reduction to nitrite has not yet been examined. We report here that intact cells and cell fractions catalyse nitrate and chlorate reduction by reduced viologen dyes with high specific activities. A gene cluster encoding components of a putative periplasmic nitrate reductase system (napA, G, H, B, F, L, D) was sequenced. The napA gene was inactivated by inserting a kanamycin resistance gene cassette. The resulting mutant did not grow by nitrate respiration and did not reduce nitrate during growth by fumarate respiration, in contrast to the wild type. An antigen was detected in wild-type cells using an antiserum raised against the periplasmic nitrate reductase (NapA) from Paracoccus pantotrophus. This antigen was absent in the W. succinogenes napA mutant. It is concluded that the periplasmic nitrate reductase NapA is the only respiratory nitrate reductase in W. succinogenes, although a second nitrate-reducing enzyme is apparently induced in the napA mutant. The nap cluster of W. succinogenes lacks a napC gene whose product is thought to function in quinol oxidation and electron transfer to NapA in other bacteria. The W. succinogenes genome encodes two members of the NapC/NirT family, NrfH and FccC. Characterization of corresponding deletion mutants indicates that neither of these two proteins is required for nitrate respiration. A mutant lacking the genes encoding respiratory nitrite reductase (nrfHA) had wild-type properties with respect to nitrate respiration. A model of the electron transport chain of nitrate respiration is proposed in which one or more of the napF, G, H and L gene products mediate electron transport from menaquinol to the periplasmic NapAB complex. Inspection of the W. succinogenes genome sequence suggests that ammonia formation from

  14. Draft Genome Sequences of Rhodosporidium toruloides Strains ATCC 10788 and ATCC 10657 with Compatible Mating Types

    PubMed Central

    Hu, Jie

    2016-01-01

    Rhodosporidium toruloides ATCC 10788 (haploid, A1 mating type) and ATCC 10657 (haploid, A2 mating type) were derived from the same diploid parent strain Rhodotorula glutinis ATCC 90781 and are important strains for metabolic engineering. Draft genome sequences of both strains are reported here. The current assembly of strain ATCC 10788 comprises 61 scaffolds with a total size of 20.75 Mbp and a GC content of 62.01%, while that of strain ATCC 10657 comprises 137 scaffolds with a total size of 21.49 Mbp and a GC content of 61.81%. Genome annotation predicts 7,730 and 7,800 protein encoding genes for strain ATCC 10788 and strain ATCC 10657, respectively. PMID:26966203

  15. Draft Genome Sequences of Rhodosporidium toruloides Strains ATCC 10788 and ATCC 10657 with Compatible Mating Types.

    PubMed

    Hu, Jie; Ji, Lianghui

    2016-01-01

    Rhodosporidium toruloides ATCC 10788 (haploid, A1 mating type) and ATCC 10657 (haploid, A2 mating type) were derived from the same diploid parent strain Rhodotorula glutinis ATCC 90781 and are important strains for metabolic engineering. Draft genome sequences of both strains are reported here. The current assembly of strain ATCC 10788 comprises 61 scaffolds with a total size of 20.75 Mbp and a GC content of 62.01%, while that of strain ATCC 10657 comprises 137 scaffolds with a total size of 21.49 Mbp and a GC content of 61.81%. Genome annotation predicts 7,730 and 7,800 protein encoding genes for strain ATCC 10788 and strain ATCC 10657, respectively. PMID:26966203

  16. Evidence that extracellular components function in adherence of Actinobacillus actinomycetemcomitans to epithelial cells.

    PubMed Central

    Meyer, D H; Fives-Taylor, P M

    1993-01-01

    Extracellular microvesicles and a highly proteinaceous polymer associated with a leukotoxin-producing strain, Actinobacillus actinomycetemcomitans SUNY 75, were shown to increase adherence of other weakly adherent A. actinomycetemcomitans strains to KB epithelial cells. Images PMID:8406899

  17. Actinobacillus spp. and related bacteria in infected wounds of humans bitten by horses and sheep.

    PubMed Central

    Peel, M M; Hornidge, K A; Luppino, M; Stacpoole, A M; Weaver, R E

    1991-01-01

    We describe the isolation of Actinobacillus lignieresii and an A. equuli-like bacterium from an infected horse-bite wound in a 22-year-old stable foreman and A. suis from a bite injury in a 35-year-old man who had been attacked by a horse. A. lignieresii was also isolated in pure culture from an infected sheep-bite wound in a rural worker. These species of the genus Actinobacillus are primarily associated with animals and animal diseases and are rarely isolated from humans. The purpose of this report is to raise awareness of the possible occurrence of Actinobacillus spp. in bite wounds inflicted by farm animals and to discuss the difficulties encountered in the identification of species of Actinobacillus and related bacteria. PMID:1774260

  18. Actinobacillus actinomycetemcomitans contamination of toothbrushes from patients harbouring the organism.

    PubMed

    Müller, H P; Lange, D E; Müller, R F

    1989-07-01

    The main ecological niche of Actinobacillus actinomycetemcomitans (A.a.) seems to be the periodontal pocket, but it can also be isolated from supragingival plaque, buccal and tongue mucosa, or saliva. We examined toothbrushes from 21 patients, all identified as harbouring moderate to large numbers of A.a. in subgingival plaque, for contamination with this organism. 29% of the toothbrushes presented by our patients yielded detectable numbers of A.a. Immediately after toothbrushing this figure rose to 62%, but dropped to 50% after 1 h. Numbers of isolated A.a. on toothbrushes were weakly correlated with the degree of periodontal destruction, and significantly more numbers of A.a. on toothbrushes could be detected if the organism was found on mucous membranes or in saliva. There was no association with gingival inflammation, supragingival plaque nor mean numbers of isolated subgingival A.a. PMID:2760252

  19. Experimental aerosol transmission of Actinobacillus pleuropneumoniae to pigs.

    PubMed Central

    Jobert, J L; Savoye, C; Cariolet, R; Kobisch, M; Madec, F

    2000-01-01

    In order to demonstrate the possible role of aerosol in the transmission of Actinobacillus pleuropneumoniae, an experiment including 18 specific pathogen-free (SPF), 10-week-old piglets, randomly distributed into 2 adjacent units, was carried out. In these facilities, air was forced through absolute filters to prevent any contact with infectious agents. During the first 6 d post inoculation, the 2 units were connected by a rectangular opening and the air circulation was forced by the ventilation system from unit A (inoculated pigs) to unit B (non-inoculated pigs). The A. pleuropneumoniae strain (biovar 1 serovar 9) was isolated in France from an outbreak of porcine pleuropneumonia. Two different infecting doses, 10(7) cfu/animal and 10(8) cfu/animal, were inoculated by intranasal route in 6 pigs of unit A. The infection spread quickly from the inoculated pigs to the non-inoculated pigs. Clinical signs were acute during the 4 d post inoculation: hyperthermia, respiratory distress and, sometimes, death (6 pigs of the unit A and 2 pigs of the unit B). All pigs seroconverted against A. pleuropneumoniae serovar 9 within 2 weeks. Lung lesions were severe: fibrinous pleurisy and lung hemorrhages in the acute stage, pleural adherences and focal pulmonary necrosis in the chronic stage. Actinobacillus pleuropneumoniae was isolated from the tonsils and/or lungs in 16 animals. It could be also isolated from the air of the experimental unit. This study showed that A. pleuropneumoniae was readily transmitted through aerosol over a distance of at least 2.5 m. Images Figure 1. Figure 2. PMID:10680652

  20. Biosynthetic Pathways of Vibrio succinogenes growing with fumarate as terminal electron acceptor and sole carbon source.

    PubMed

    Bronder, M; Mell, H; Stupperich, E; Kröger, A

    1982-05-01

    1. With fumarate as the terminal electron acceptor and either H2 or formate as donor, Vibrio succinogenes could grow anaerobically in a mineral medium using fumarate as the sole carbon source. Both the growth rate and the cell yield were increased when glutamate was also present in the medium. 2. Glutamate was incorporated only into the amino acids of the glutamate family (glutamate, glutamine, proline and arginine) of the protein. The residual cell constituents were synthesized from fumarate. 3. Pyruvate and phosphoenolpyruvate, as the central intermediates of most of the cell constituents, were formed through the action of malic enzyme and phosphoenolpyruvate synthetase. Fructose-1,6-bisphosphate aldolase was present in the bacterium suggesting that this enzyme is involved in carbohydrate synthesis. 4. In the absence of added glutamate the amino acids of the glutamate family were synthesized from fumarate via citrate. The enzymes involved in glutamate synthesis were present. 5. During growth in the presence of glutamate, net reducing equivalents were needed for cell synthesis. Glutamate and not H2 or formate was used as the source of these reducing equivalents. For this purpose part of the glutamate was oxidized to yield succinate and CO2. 6. The alpha-ketoglutarate dehydrogenase involved in this reaction was found to use ferredoxin as the electron acceptor. The ferredoxin of the bacterium was reoxidized by means of a NADP-ferredoxin oxidoreductase. Enzymes catalyzing the reduction of NAD, NADP or ferredoxin by H2 or formate were not detected in the bacterium. PMID:7103660

  1. Application of a data reconciliation method to the stoichiometric analysis of Fibrobacter succinogenes growth.

    PubMed

    Guiavarch, Erell; Pons, Agnes; Creuly, Catherine; Dussap, Claude-Gilles

    2008-12-01

    Fibrobacter succinogenes S85, a strictly anaerobic Gram-negative bacterium, was grown in continuous culture in a bioreactor at different dilution rates (0.02 to 0.092 h(-1)) on a fully synthetic culture medium with glucose as carbon source. Glucose and ammonium sulfate consumption, as well as biomass, succinate, acetate, formate, and carbohydrate production were regularly measured. The relevant biomass elemental compositions were established for each dilution rate. Robustness of the experimental information was checked by C and N mass balances estimation, which were satisfactory. A detailed overall stoichiometry analysis of the process, including all substrates and products of the culture, was proposed. Online and off-line parameters measured during the culture brought a large number of data which were weighted by their respective variance associated to the measured value. The material balance resulted in an overdetermined linear system of equations made of weighted relationships including experimental data, elemental balances (C, H, O, N, S, Na), and an additional constraint. The mass balances involved in stoichiometric equations were solved using data reconciliation and linear algebra methods to take into account error measurements. This methodology allowed to establish the overall stoichiometric equation for each dilution rate studied. PMID:18581268

  2. Genome sequence of Lactobacillus rhamnosus ATCC 8530.

    PubMed

    Pittet, Vanessa; Ewen, Emily; Bushell, Barry R; Ziola, Barry

    2012-02-01

    Lactobacillus rhamnosus is found in the human gastrointestinal tract and is important for probiotics. We became interested in L. rhamnosus isolate ATCC 8530 in relation to beer spoilage and hops resistance. We report here the genome sequence of this isolate, along with a brief comparison to other available L. rhamnosus genome sequences. PMID:22247527

  3. Genome Sequence of Propionibacterium acidipropionici ATCC 55737.

    PubMed

    Luna-Flores, Carlos H; Nielsen, Lars K; Marcellin, Esteban

    2016-01-01

    Propionibacterium acidipropionici produces propionic acid as its main fermentation product. Traditionally derived from fossil fuels, environmental and sustainable issues have revived the interest in producing propionic acid using biological resources. Here, we present the closed sequence of Propionibacterium acidipropionici ATCC 55737, an efficient propionic acid producer. PMID:27198010

  4. Identification and expression of the Actinobacillus actinomycetemcomitans leukotoxin gene.

    PubMed

    Lally, E T; Kieba, I R; Demuth, D R; Rosenbloom, J; Golub, E E; Taichman, N S; Gibson, C W

    1989-02-28

    The leukotoxin produced by the oral bacterium Actinobacillus actinomycetemcomitans has been implicated in the pathogenesis of juvenile periodontitis. In order to elucidate the structure of the leukotoxin, molecular cloning of the leukotoxin gene was carried out. A DNA library of A. actinomycetemcomitans, strain JP2, was constructed by partial digestion of genomic DNA with Sau3AI and ligation of 0.5 to 5.0 kilobase pair fragments into the Bam HI site of the plasmid vector pENN-vrf. After transformation into E. coli RR1 (lambda cI857), the clones were screened for the production of A. actinomycetemcomitans leukotoxin with polyclonal antibody. Six immunoreactive clones were identified. The clones expressed proteins which ranged from 21-80 kilodaltons, and the clone designated pII-2, producing the largest protein was selected for further study. Antibodies eluted from immobilized pII-2 protein also recognized the native A. actinomycetemcomitans leukotoxin molecule indicating that both molecules shared at least one epitope. DNA sequence analysis demonstrated that there are regions of significant amino acid sequence homology between the cloned A. actinomycetemcomitans leukotoxin and two other cytolysins, Escherichia coli alpha-hemolysin and Pasteurella haemolytica leukotoxin, suggesting that a family of cytolysins may exist which share a common mechanism of killing but vary in their target cell specificity. PMID:2647082

  5. Antimicrobial resistance of Actinobacillus pleuropneumoniae isolated from swine.

    PubMed

    Vanni, Michele; Merenda, Marianna; Barigazzi, Giuseppe; Garbarino, Chiara; Luppi, Andrea; Tognetti, Rosalba; Intorre, Luigi

    2012-04-23

    The aim of this retrospective study was to evaluate the antimicrobial resistance rates and the trend in resistance of Actinobacillus pleuropneumoniae isolated from pigs in Italy from 1994 to 2009. A total of 992 A. pleuropneumoniae isolates were tested for their susceptibility to a panel of antimicrobial agents in a disk diffusion method. Resistance to 7 drugs (amoxicillin, amoxicillin/clavulanic acid, ampicillin, cefquinome, cotrimoxazole, penicillin G and tilmicosin) showed a significant increasing trend over the time, while for 2 drugs (gentamycin and marbofloxacin) a significant decrease was observed. Resistance to the remaining 14 antimicrobial agents tested did not change significantly over the study period. Most of the isolates retained high susceptibility to antimicrobials usually effective against A. pleuropneumoniae such as amphenicols, fluoroquinolones and ceftiofur. However, high rates of resistance were observed for potentiated sulfa drugs, tetracyclines and penicillins which are currently recommended antimicrobials for pig pleuropneumonia therapy. Our results suggest the importance of continued monitoring of A. pleuropneumoniae clinical isolates in order to choose the most appropriate treatment of infections and to control the increase of resistance to currently used antimicrobials. PMID:22104584

  6. Cell Yields of Vibrio succinogenes growing with formate and fumarate as sole carbon and energy sources in chemostat culture.

    PubMed

    Mell, H; Bronder, M; Kröger, A

    1982-05-01

    Vibrio succinogenes which gains all the ATP by anaerobic electron transport phosphorylation, was grown in continuous culture on a defined medium with formate and fumarate as sole energy sources. The growth yield at infinite dilution rate (Ymax) was obtained by extrapolation from the growth yields measured at various dilution rates. With formate as the growth limiting substrate, Ymax was found as 14 g dry cells/mol formate. Under these conditions growth was limited by the rate of energy supply, because formate is used only as a catabolic substrate (Bronder et al. 1982). The YmaxATP calculated from the ATP requirement for cell synthesis was 18 g dry cells/mol ATP. This gives an ATP/2e ratio of 0.8. The ATP/2e ratio in vitro had been measured as 1 (Kröger and Winkler 1981). It is concluded that growing V. succinogenes gain at least 80% the stoichiometrically possible amount of ATP, when growth is limited by energy supply. PMID:7103661

  7. Draft Genome Assemblies of Proteus mirabilis ATCC 7002 and Proteus vulgaris ATCC 49132.

    PubMed

    Minogue, T D; Daligault, H E; Davenport, K W; Bishop-Lilly, K A; Bruce, D C; Chain, P S; Coyne, S R; Chertkov, O; Freitas, T; Frey, K G; Jaissle, J; Koroleva, G I; Ladner, J T; Palacios, G F; Redden, C L; Xu, Y; Johnson, S L

    2014-01-01

    The pleomorphic swarming bacilli of the genus Proteus are common human gut commensal organisms but also the causative agents of recurrent urinary tract infections and bacteremia. We sequenced and assembled the 3.99-Mbp genome of Proteus mirabilis ATCC 7002 (accession no. JOVJ00000000) and the 3.97-Mbp genome of Proteus vulgaris ATCC 49132 (accession no. JPIX00000000), both of which are commonly used reference strains. PMID:25342681

  8. Adhesion of Bacteroides succinogenes in pure culture and in the presence of Ruminococcus flavefaciens to cell walls in leaves of perennial ryegrass (Lolium perenne).

    PubMed Central

    Latham, M J; Brooker, B E; Pettipher, G L; Harris, P J

    1978-01-01

    Bacteroides succinogenes and Ruminococcus flavefaciens are two of the most important cellulolytic bacteria in the rumen. Adhesion of B. succinogenes in pure culture, and in mixed culture with R. flavefaciens, to the various types of cell walls in sections of perennial ryegrass (Lolium perenne L. cultivar S24) leaves was examined by transmission and scanning electron microscopy. B. succinogenes adhered to the cut edges of most plant cell walls except those of the meta- and protoxylem. It also adhered, though in much smaller numbers, to the uncut surfaces of mesophyll, epidermal, and phloem cell walls. In mixed culture, both species adhered in significant numbers to the cut edges of most types of plant cell wall, but R. flavefaciens predominated on the epidermis, phloem, and sclerenchyma cell walls. B. succinogenes predominated on the cut edges and on the uncut surfaces of the mesophyll cell walls, and its ability to adhere to uncut surfaces of other cell walls was not affected by the presence of the ruminococcus. Both organisms rapidly digested the epidermal, mesophyll, and phloem cell walls. Zones of digestion were observed around bacteria of both species when attached to the lignified cell walls of the sclerenchyma, but not when attached to the lignified xylem vessels. Images PMID:567035

  9. Pathogenesis of porcine Actinobacillus pleuropneumonia: Part I. Effects of surface components of Actinobacillus pleuropneumoniae in vitro and in vivo.

    PubMed Central

    Huang, H; Potter, A A; Campos, M; Leighton, F A; Willson, P J; Yates, W D

    1998-01-01

    To understand the role of non-secreted components of Actinobacillus pleuropneumoniae in virulence, we investigated in vitro cytotoxicity and in vivo pulmonary changes in pigs due to various A. pleuropneumoniae (serotype 1) fractions. Following 1.5 h incubation, lipopolysaccharide (LPS), 2 crude extracts and bacterial culture supernatant (BCS) at high concentrations were cytotoxic to porcine pulmonary alveolar macrophages (PAM), peripheral blood mononuclear leucocytes, neutrophils and a cultured porcine bone marrow cell line. Heat-killed bacteria were cytotoxic to PAM after 24 h incubation. The 2 crude extracts were prepared by shaking either intact bacteria after removing culture supernatants (crude surface extract, CSE), or whole bacterial culture (crude surface plus culture supernatant extract, CSSE) with glass beads in saline at 60 degrees C. Further experiments showed that proteins from the bacterial membrane were partially involved in cytotoxicities of these 2 extracts. Both BCS and CSSE caused multivocal hemorrhage and neutrophil infiltration when inoculated into porcine lungs, but CSE did not. The lung:whole body weight ratios of the pigs treated with CSSE were significantly higher (P < 0.05) than those of pigs treated with BCS, CSE, or control solution. It is concluded that beside the secreted proteins, bacterial surface components including LPS and non-secreted proteins were cytotoxic in vitro; and secreted and non-secreted components act synergistically to cause lung lesions. PMID:9553707

  10. Actinobacillus actinomycetemcomitans induces apoptosis in human monocytic THP-1 cells.

    PubMed

    Kato, Satsuki; Sugimura, Norihiko; Nakashima, Keisuke; Nishihara, Tatsuji; Kowashi, Yusuke

    2005-03-01

    It has previously been reported that the murine macrophage cell line J774.1 and the human oral epithelial cell line KB undergo apoptosis as a result of Actinobacillus actinomycetemcomitans infection. Recent studies have demonstrated that apoptosis regulation is modulated by multiple phosphorylation of several different protein kinases, including the major subtypes of the mitogen-activated protein kinase (MAPK) family. The MAPK family promotes cell survival and/or proliferation in response to growth factor stimulation, or apoptosis in response to various stress stimuli. The primary objective of the present investigation was to clarify whether human immune cells undergo apoptosis following A. actinomycetemcomitans infection and, if so, to establish the involvement of the MAPK family. Human monocytic THP-1 cells were infected with A. actinomycetemcomitans in microtubes. Lactate dehydrogenase release into the culture supernatant and DNA fragmentation in the cells were monitored. DNA fragmentation was also identified by agarose gel electrophoresis. Cell death following A. actinomycetemcomitans infection occurred by apoptosis, shown by an increase in the proportion of fragmented DNA and the typical ladder pattern of DNA fragmentation indicative of apoptosis. Furthermore, p38 MAPK activity and tumour necrosis factor alpha (TNF-alpha) levels increased following A. actinomycetemcomitans infection. In contrast, cell death and TNF-alpha levels in infected cells decreased upon addition of a p38 inhibitor or an anti-TNF-alpha antibody. However, exogenous TNF-alpha could not induce apoptosis in uninfected THP-1 cells. Interestingly, p38 MAPK activity diminished in the presence of anti-TNF-alpha antibody. These findings indicated that A. actinomycetemcomitans infection induces apoptosis in THP-1 cells and that p38 MAPK activity is directly involved in apoptosis. TNF-alpha may play an indirect role in apoptosis via enhanced p38 MAPK activity. A. actinomycetemcomitans

  11. A cohort study on Actinobacillus pleuropneumoniae colonisation in suckling piglets.

    PubMed

    Tobias, T J; Klinkenberg, D; Bouma, A; van den Broek, J; Daemen, A J J M; Wagenaar, J A; Stegeman, J A

    2014-06-01

    Actinobacillus pleuropneumoniae causes respiratory disease in pigs and despite the use of preventive measures such as vaccination and antimicrobials clinical outbreaks still occur. At weaning often many piglets are not colonised. If differences in prevalence between litters are large and if factors were known that could explain these differences, this may provide an opportunity to raise groups of A. pleuropneumoniae free piglets. To this end, a cohort study was performed on two endemically infected farrow-to-finish farms. Seventy-six of 133 sows were selected using stratified random selection by parity. Farmers complied with a strict hygiene and animal management protocol to prevent transmission between litters. Tonsil brush and serum samples taken three weeks before parturition were tested for antigen with an apxIVA qPCR and antibodies with Apx and Omp ELISAs, respectively. Three days before weaning tonsil brush samples from all piglets (n=871) were collected and tested for antigen. Whereas all sows tested positive both in serology tests as well as qPCR, 0.41 of the litters tested fully negative and 0.73 of all piglets tested negative. The proportion of positively tested piglets in positive litters ranged from 0.08-1.0 (median=0.36). A grouped logistic regression model with a beta binomial distribution of the probability for piglets to become infected was fitted to the data and associations with explanatory variables were explored. To test the possibility that alternatively the clustering was caused by onwards transmission among the piglets, a transmission model was fitted to the data incorporating sow-piglet and piglet-piglet transmission, but this model did not fit better. The results of this study showed that the number of colonised suckling piglets was highly clustered and mainly attributable to the variability of infectiousness of the dam, but no dam related risk factor for colonisation status of litter or piglets within litters could be identified. PMID

  12. The gingival immune response to Actinobacillus actinomycetemcomitans in juvenile periodontitis.

    PubMed

    Hall, E R; Falkler, W A; Martin, S A; Suzuki, J B

    1991-12-01

    The established and advanced lesions of juvenile periodontitis-localized form (JP) are predominated by B-lymphocytes and plasma cells. Local immune processes may participate in protective or immunopathologic roles in the pathogenesis of this disease. Actinobacillus actinomycetemcomitans (A.a.) is implicated as a primary etiologic agent in JP. An in vitro gingival explant culture system was utilized to study the specificity of immunoglobulins produced by diseased JP tissues. A dot-immunobinding assay demonstrated that 46% of the supernatant fluids (SF) from explant cultures of diseased tissues (n = 39) were positive for the presence of antibody to A.a. Y4, while 61% of autologous JP sera (n = 39) tested positive. For rapidly progressive (RP) and adult periodontitis (AP) SF, 50% and 40% were positive for A.a. Y4, respectively. Seventeen percent of SF from healthy tissue were positive for A.a. Y4. There was no significant difference between JP SF reactivities to A.a. Y4 when compared to reactivities of SF from AP and RP patients. Only 10% of JP SF were positive for Porphyromonas asaccharolytica, a non-oral control microorganism. The de novo biosynthesis of antibody in JP tissue, reactive with A.a. Y4, was demonstrated with Staph Protein A isolated 14C-labeled IgG (SPAG) and the use of a dot-immunobinding assay and autoradiography. The in vitro gingival tissue explant culture system described provides a useful model for the study of the synthesis and specificity of localized immunoglobulins produced by diseased tissues of JP patients. PMID:1765941

  13. Biogenesis of the Actinobacillus actinomycetemcomitans Cytolethal Distending Toxin Holotoxin

    PubMed Central

    Ueno, Yoko; Ohara, Masaru; Kawamoto, Toru; Fujiwara, Tamaki; Komatsuzawa, Hitoshi; Oswald, Eric; Sugai, Motoyuki

    2006-01-01

    The cell cycle G2/M specific inhibitor cytolethal distending toxin (CDT) from Actinobacillus actinomycetemcomitans is composed of CdtA, CdtB, and CdtC coded on the cdtA, cdtB, and cdtC genes that are tandem on the chromosomal cdt locus. A. actinomycetemcomitans CdtA has the lipid binding consensus domain, the so-called “lipobox”, at the N-terminal signal sequence. Using Escherichia coli carrying plasmid pTK3022, we show that the 16th residue, cysteine, of CdtA bound [3H]palmitate or [3H]glycerol. Further, posttranslational processing of the signal peptide, CdtA, was inhibited using globomycin, an inhibitor of lipoprotein-specific signal peptidase II. Fractionation and immunoblotting show the lipid-modified CdtA is present in the outer membrane. Immunoprecipitation and the pull-down assay of the CDT complex from E. coli carrying a plasmid containing cdtABC demonstrated that the CDT complex in the periplasm is composed of CdtA, CdtB, and CdtC and that the CDT complex in culture supernatant is an N-terminally truncated (36 to 43 amino acids) form of CdtA (CdtA′), CdtB, and CdtC. This suggests that CDT is present as a complex both in the periplasm and the supernatant where CdtA undergoes posttranslation processing to CdtA′ in the process of biogenesis and secretion of CDT holotoxin into the culture supernatant. Site-directed mutagenesis of the 16th cysteine residue to glycine in CdtA altered localization of CdtA in the cell and reduced the amount of CDT activity in the culture supernatant. This suggests that CDT forms a complex inside the periplasm for lipid modification where posttranslational processing of CdtA plays an important role for the efficient production of CDT holotoxin into the culture supernatant. PMID:16714579

  14. Identification of Actinobacillus actinomycetemcomitans in subgingival plaque by PCR.

    PubMed Central

    Flemmig, T F; Rüdiger, S; Hofmann, U; Schmidt, H; Plaschke, B; Strätz, A; Klaiber, B; Karch, H

    1995-01-01

    The purpose of this study was to assess the sensitivity and specificity of the PCR in detecting Actinobacillus actinomycetemcomitans. The PCR's detection capability was compared with those of three other methods: culture-enhanced PCR (CE-PCR), colony hybridization (CH), and conventional culture with presumptive biochemical identification. A 285-bp stretch of the leukotoxin gene lktA of A. actinomycetemcomitans was amplified by PCR with primers TT-15 and TT-16. For CH, the PCR product was labeled with digoxigenin and used as a hybridization probe. Nucleotide sequence analysis of the PCR product of A. actinomycetemcomitans 1D4 and 1664 and three clinical isolates revealed complete homology among the tested strains, with only one base substitution (at position 1344) in comparison with the published sequence. With artificially infected subgingival plaque, the detection limit of PCR for A. actinomycetemcomitans was 10(3) CFU/ml of plaque suspension. Culturing subgingival plaque on tryptic soy-serum-bacitracin-vancomycin agar prior to PCR (CE-PCR) improved the limit of detection to 10(2) CFU/ml. Analysis of subgingival plaque samples from 35 patients with periodontal disease and 10 periodontally healthy subjects revealed that CE-PCR and CH had the highest overall rate of A. actinomycetemcomitans detection (both 58%), followed by PCR and culture (both 42%). With CH as the "gold standard", the sensitivities of CE-PCR, PCR, and culture were 88, 65, and 58%, respectively; the specificities were 84, 89, and 79%, respectively. The CE-PCR provided acceptable positive and negative predictive values (> or = 70%) when the prevalence of A. actinomycetemcomitans varied between 30 and 70%. PCR alone provided comparable predictive values over a narrower range of prevalence rates (30 to 50%), while culture did not afford acceptable predictive values at any prevalence rate. PCR and CE-PCR were found to be superior to culture with presumptive biochemical identification and should be the

  15. Proteomic Analysis of the Secretome of Cellulomonas fimi ATCC 484 and Cellulomonas flavigena ATCC 482

    PubMed Central

    Wakarchuk, Warren W.; Brochu, Denis; Foote, Simon; Robotham, Anna; Saxena, Hirak; Erak, Tamara; Kelly, John

    2016-01-01

    The bacteria in the genus Cellulomonas are known for their ability to degrade plant cell wall biomass. Cellulomonas fimi ATCC 484 and C. flavigena ATCC 482 have been the subject of much research into secreted cellulases and hemicellulases. Recently the genome sequences of both C. fimi ATCC 484 and C. flavigena ATCC 482 were published, and a genome comparison has revealed their full spectrum of possible carbohydrate-active enzymes (CAZymes). Using mass spectrometry, we have compared the proteins secreted by C. fimi and C. flavigena during growth on the soluble cellulose substrate, carboxymethylcellulose (CMC), as well as a soluble xylan fraction. Many known C. fimi CAZymes were detected, which validated our analysis, as were a number of new CAZymes and other proteins that, though identified in the genome, have not previously been observed in the secretome of either organism. Our data also shows that many of these are co-expressed on growth of either CMC or xylan. This analysis provides a new perspective on Cellulomonas enzymes and provides many new CAZyme targets for characterization. PMID:26950732

  16. Biochemical and Domain Analyses of FSUAxe6B, a Modular Acetyl Xylan Esterase, Identify a Unique Carbohydrate Binding Module in Fibrobacter succinogenes S85▿ †

    PubMed Central

    Yoshida, Shosuke; Mackie, Roderick I.; Cann, Isaac K. O.

    2010-01-01

    Acetyl xylan esterase (EC 3.1.1.72) is a member of a set of enzymes required to depolymerize hemicellulose, especially xylan that is composed of a main chain of β-1,4-linked xylopyranoside residues decorated with acetyl side groups. Fibrobacter succinogenes S85 Axe6B (FSUAxe6B) is an acetyl xylan esterase encoded in the genome of this rumen bacterium. The enzyme is a modular protein comprised of an esterase domain, a carbohydrate-binding module, and a region of unknown function. Sequences that are homologous to the region of unknown function are paralogously distributed, thus far, only in F. succinogenes. Therefore, the sequences were designated Fibrobacter succinogenes-specific paralogous module 1 (FPm-1). The FPm-1s are associated with at least 24 polypeptides in the genome of F. succinogenes S85. A bioinformatics search showed that most of the FPm-1-appended polypeptides are putative carbohydrate-active enzymes, suggesting a potential role in carbohydrate metabolism. Truncational analysis of FSUAxe6B, together with catalytic and substrate binding studies, has allowed us to delineate the functional modules in the polypeptide. The N-terminal half of FSUAxe6B harbors the activity that cleaves side chain acetyl groups from xylan-like substrates, and the binding of insoluble xylan was determined to originate from FPm-1. Site-directed mutagenesis studies of highly conserved active-site residues in the esterase domain suggested that the esterase activity is derived from a tetrad composed of Ser44, His273, Glu194, and Asp270, with both Glu194 and Asp270 functioning as helper acids, instead of a single carboxylate residue proposed to initiate catalysis. PMID:19897648

  17. Construction of Actinobacillus pleuropneumoniae-Escherichia coli shuttle vectors: expression of antibiotic-resistance genes.

    PubMed

    West, S E; Romero, M J; Regassa, L B; Zielinski, N A; Welch, R A

    1995-07-01

    We constructed several cloning vectors, designated pGZRS-18/19 and pGZRS-38/39, which were based on an endogenous Actinobacillus pleuropneumoniae (Apl) 4.3-kb plasmid. They carry the lacZ alpha-complementation fragment and MCS from pUC18/19, and either the bla gene under the control of a putative Apl promoter or the KmR gene from Tn903. These vectors replicate in representative strains of Apl serotypes 1 and 7, Escherichia coli, Pasteurella haemolytica (Ph) and Haemophilus (Actinobacillus) actinomycetemcomitans. We also found that Apl and Ph did not express genes under the control of the lacZ or bla promoters, suggesting that their RNA polymerases may not utilize these promoters. PMID:7628722

  18. Substrate specificity of three cytochrome c haem lyase isoenzymes from Wolinella succinogenes: unconventional haem c binding motifs are not sufficient for haem c attachment by NrfI and CcsA1

    PubMed Central

    Kern, Melanie; Eisel, Florian; Scheithauer, Juliane; Kranz, Robert G.; Simon, Jörg

    2012-01-01

    Summary Bacterial c-type cytochrome maturation is dependent on a complex enzymic machinery. The key reaction is catalysed by cytochrome c haem lyase (CCHL) that usually forms two thioether bonds to attach haem b to the cysteine residues of a haem c binding motif (HBM) which is, in most cases, a CX2CH sequence. Here, the HBM specificity of three distinct CCHL isoenzymes (NrfI, CcsA1 and CcsA2) from the Epsilonproteobacterium Wolinella succinogenes was investigated using either W. succinogenes or Escherichia coli as host organism. Several reporter c-type cytochromes were employed including cytochrome c nitrite reductases (NrfA) from E. coli and Campylobacter jejuni that differ in their active site HBMs (CX2CK or CX2CH). W. succinogenes CcsA2 was found to attach haem to standard CX2CH motifs in various cytochromes whereas other HBMs were not recognized. NrfI was able to attach haem c to the active site CX2CK motif of both W. succinogenes and E. coli NrfA, but not to NrfA from C. jejuni. Different apo-cytochrome variants carrying the CX15CH motif, assumed to be recognized by CcsA1 during maturation of the octahaem cytochrome MccA, were not processed by CcsA1 in either W. succinogenes or E. coli. It is concluded that the dedicated CCHLs NrfI and CcsA1 attach haem to non-standard HBMs only in the presence of further, as yet uncharacterised structural features. Interestingly, it proved impossible to delete the ccsA2 gene from the W. succinogenes genome; a finding that is discussed in the light of the available genomic, proteomic and functional data on W. succinogenes c-type cytochromes. PMID:19919672

  19. Recent allopolyploid origin of Zygosaccharomyces rouxii strain ATCC 42981.

    PubMed

    Gordon, Jonathan L; Wolfe, Kenneth H

    2008-06-01

    Zygosaccharomyces rouxii strain ATCC 42981 has been reported to have two copies of several genes including HOG1 and SOD2, whereas the type strain of Z. rouxii (CBS 732) has only one. To investigate the structure of the ATCC 42981 genome we sequenced random fragments from this genome and compared the data to the type strain. We found that ATCC 42981 contains two versions of the ribosomal RNA array, one of which is identical in the ITS1-ITS2 and 26S D1/D2 regions to Z. rouxii CBS 732, while the other is almost identical to a species provisionally named Z. pseudorouxii. We found that most genomic regions from Z. rouxii CBS 732 map in a one-to-two fashion to pairs of regions in ATCC 42981, with one of the ATCC 42981 regions having 97-100% DNA sequence identity to CBS 732 and the other having about 80-90% identity. Complete sequencing of regions containing 30 pairs of genes from ATCC 42981 and their orthologues in CBS 732 showed no evidence of the gene deletions or pseudogene formation that might be expected if ATCC 42981 had undergone whole-genome duplication several million years ago and was in the early stages of gene loss. Instead, we conclude that ATCC 42981 is a Z. rouxii-Z. pseudorouxii interspecies hybrid that was formed so recently that its genome has not had time to decay. PMID:18509846

  20. Draft Genome Sequence of Type Strain Streptococcus gordonii ATCC 10558

    PubMed Central

    Rasmussen, Louise H.; Dargis, Rimtas; Skovgaard, Ole

    2016-01-01

    Streptococcus gordonii ATCC 10558T was isolated from a patient with infective endocarditis in 1946 and announced as a type strain in 1989. Here, we report the 2,154,510-bp draft genome sequence of S. gordonii ATCC 10558T. This sequence will contribute to knowledge about the pathogenesis of infective endocarditis. PMID:26893427

  1. Manganese transport in Brevibacterium ammoniagenes ATCC 6872.

    PubMed Central

    Schmid, J; Auling, G

    1987-01-01

    Uptake of manganese by Brevibacterium ammoniagenes ATCC 6872 was energy dependent and obeyed saturation kinetics (Km = 0.65 microM; Vmax = 0.12 mumol/min per g [dry weight]). Uptake showed optima at 27 degrees C and pH 9.5. 54Mn2+ accumulated by the cells was released by treatment with toluene or by exchange for unlabeled manganese ions, via an energy-dependent process. Co2+, Fe2+, Cd2+, and Zn2+ inhibited manganese uptake. Inhibition by Cd2+ and Zn2+ was competitive (Ki = 0.15 microM Cd2+ and 1.2 microM Zn2+). Experiments with 65Zn2+ provided no evidence for Zn2+ uptake via the Mn2+ transport system. PMID:3597325

  2. The xynC gene from Fibrobacter succinogenes S85 codes for a xylanase with two similar catalytic domains.

    PubMed Central

    Paradis, F W; Zhu, H; Krell, P J; Phillips, J P; Forsberg, C W

    1993-01-01

    The xynC gene of Fibrobacter succinogenes S85 codes for a 66.4-kDa xylanase which consists of three distinct domains separated by two flexible regions rich in serine residues. Domains A and B of XynC code for catalytic domains with 56.5% identity and 9.6% similarity with each other, and both domains share homology with xylanases of Ruminococcus flavefaciens, Neocallimastix patriciarum, Clostridium acetobutylicum, Bacillus pumilus, Bacillus subtilis, and Bacillus circulans. More than 88% of the xylanase activity of Escherichia coli cells carrying the original 13-kb recombinant plasmid was released from intact cells by cold water washes. The major products of hydrolysis of xylan by both domains were xylose and xylobiose, indicating that the xynC gene product exhibits catalytic properties similar to those of the XynA xylanases from R. flavefaciens and N. patriciarum. So far, these features are not shared broadly with bacteria from other environments and may indicate specific selection for this domain structure in the highly competitive environment of the rumen. Images PMID:8244936

  3. Draft Genome Sequence of Mycobacterium brumae ATCC 51384

    PubMed Central

    D'Auria, Giuseppe

    2016-01-01

    Here, we report the draft genome sequence of Mycobacterium brumae type strain ATCC 51384. This is the first draft genome sequence of M. brumae, a nonpathogenic, rapidly growing, nonchromogenic mycobacterium, with immunotherapeutic capacities. PMID:27125480

  4. Genome Sequence of Streptomyces aureofaciens ATCC Strain 10762

    PubMed Central

    Gradnigo, Julien S.; Somerville, Greg A.; Huether, Michael J.; Kemmy, Richard J.; Johnson, Craig M.; Oliver, Michael G.

    2016-01-01

    Streptomyces aureofaciens is a Gram-positive actinomycete that produces the antibiotics tetracycline and chlortetracycline. Here, we report the assembly and initial annotation of the draft genome sequence of S. aureofaciens ATCC strain 10762. PMID:27340076

  5. Genome sequence of the fish pathogen Flavobacterium columnare ATCC 49512

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Flavobacterium columnare is a Gram-negative, rod shaped, motile, and highly prevalent fish pathogen causing columnaris disease in freshwater fish worldwide. Here, we present the complete genome sequence of F. columnare strain ATCC 49512. ...

  6. Effects of alkaline hydrogen peroxide treatment on in vitro degradation of cellulosic substrates by mixed ruminal microorganisms and Bacteroides succinogenes S85.

    PubMed Central

    Lewis, S M; Montgomery, L; Garleb, K A; Berger, L L; Fahey, G C

    1988-01-01

    The effects of sodium hydroxide (NaOH) and alkaline hydrogen peroxide (AHP) treatments on wheat straw (WS) and various cellulosic substrates were determined by measuring susceptibility to degradation by mixed ruminal organisms or Bacteroides succinogenes S85. In vitro incubations were used to measure differences in fermentation resulting from each successive step in the AHP treatment process. In vitro incubations through 48 or 108 h were conducted to measure these differences. The AHP treatment of WS increased (P less than 0.05) dry matter, neutral detergent fiber, and acid detergent fiber degradation over control WS when these substrates were incubated with mixed ruminal microorganisms or B. succinogenes S85. Fermentations containing AHP-treated WS had greater (P less than 0.05) microbial purine (RNA) and volatile fatty acid concentrations by 12 h compared with those containing untreated or NaOH-treated WS. Xylose in AHP-treated WS was utilized more extensively (P less than 0.05) by 12 h compared with the xylose of untreated or NaOH-treated WS. Treatment with AHP removed 23% of the alkali-labile phenolic compounds from WS. When substrates with high levels of crystalline cellulose (raw cotton fiber, Solka floc, and Sigmacell-50) were treated with NaOH or AHP and incubated for 108 h with B. succinogenes S85, extent of acid detergent fiber degradation of cotton fiber and Sigmacell-50 was similar to that of their respective controls. Sodium hydroxide and AHP treatments were effective in increasing acid detergent fiber degradation of the Solka floc which contained, on average, 3.3 and 4.8 percentage units more acid detergent lignin and hemicellulose, respectively, than cotton fiber and Sigmacell-50.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:3291761

  7. Actinobacillus actinomycetemcomitans strains Y4 and N27 adhere to hydroxyapatite by distinctive mechanisms.

    PubMed Central

    Kagermeier, A S; London, J

    1985-01-01

    Actinobacillus actinomycetemcomitans strains Y4 and N27 absorb to spheroidal hydroxyapatite in roughly the same numbers per milligram of substrate and with the same tenacity as two previously tested Cytophaga species. Although the two strains of A. actinomycetemcomitans exhibited similar affinities and number of binding sites for SHA, their response to enzyme treatment and heating were very different. The capacity of strain Y4 to attach to spheroidal hydroxyapatite was diminished by treatment with proteases and phospholipases and was unaffected by neuraminidase, while strain N27 was unaffected by proteases and phospholipases and lost its binding capabilities when treated with neuraminidase. Images PMID:3972445

  8. Nucleic Acid Similarities Among Pseudomonas pseudomallei, Pseudomonas multivorans, and Actinobacillus mallei1

    PubMed Central

    Rogul, M.; Brendle, J. J.; Haapala, D. K.; Alexander, A. D.

    1970-01-01

    Annealing experiments on membrane filters were carried out with deoxyribonucleic acids (DNA) from selected strains of the nomen-species of Pseudomonas, Actinobacillus, Chromobacterium, and Micrococcus, with the use of DNA of Pseudomonas pseudomallei and Actinobacillus mallei as reference materials. Under the usual conditions employed in these experiments, the results were not quantitatively reproducible. Incorporation of dimethylsulfoxide (DMSO) into the incubation medium greatly increased differences in comparative binding. DNA binding in agar matrices was examined in the presence and absence of DMSO at various incubation temperatures. It was found that the greatest specificity, stability, and total binding for DNA containing high amounts of guanine and cytosine occurred in the presence of DMSO. Under the most stringent annealing conditions permitted in agar, DNA species from P. pseudomallei and A. mallei in the presence of DMSO demonstrated interspecific relative bindings of 76 to 86% when compared to the homologous reactions. The thermal elution midpoints (Em) of these duplexed interspecific DNA species were quite close to the homologous Em values. The relative bindings of P. multivorans DNA types to either reference DNA ranged between 6 to 27%, and the Em values were 4 to 7 C less than those for the homologous reactions. PMID:5438051

  9. Crystallization and preliminary X-ray crystallographic analysis of MacA from Actinobacillus actinomycetemcomitans

    SciTech Connect

    Piao, Shunfu; Xu, Yongbin; Ha, Nam-Chul

    2008-05-01

    A periplasmic membrane-fusion protein MacA from Actinobacillus actinomycetemcomitans, an essential component of the multidrug efflux pump in Gram-negative bacteria, was crystallized. Periplasmic membrane-fusion proteins (MFPs) are an essential component of the multidrug efflux pump in Gram-negative bacteria. They play a crucial role in bridging the outer membrane porin TolC and two distinct types of inner membrane transporters. The MFP MacA bridges the inner membrane ABC-type multidrug transporter MacB and the outer membrane porin TolC. MacA from the pathogenic bacterium Actinobacillus actinomycetemcomitans was expressed in Escherichia coli B834 (DE3) and the recombinant protein was purified using Ni–NTA affinity, Q anion-exchange and gel-filtration chromatography. The purified MacA protein was crystallized using the vapour-diffusion method. A MAD diffraction data set was collected to a resolution of 3.0 Å at 100 K. The crystal belongs to space group P622, with unit-cell parameters a = b = 109.2, c = 255.4 Å, α = β = 90, γ = 120°, and contains one molecule in the asymmetric unit.

  10. Use of bean husk as an easily digestible fiber source for activating the fibrolytic rumen bacterium Fibrobacter succinogenes and rice straw digestion.

    PubMed

    Fuma, Ryosuke; Oyaizu, Shinya; Nukui, Yoko; Ngwe, Tin; Shinkai, Takumi; Koike, Satoshi; Kobayashi, Yasuo

    2012-10-01

    A series of in sacco and in vitro studies were carried out to evaluate bean husks for activation of fibrolytic rumen bacteria and rice straw digestion. First, lablab bean husk, chickpea husk and rice straw were suspended in the rumen of sheep to analyze the bacterial consortium developed on each fiber source. Known members of fiber-associating bacteria were found on both lablab bean husk and rice straw, but some of these bacteria were lacking on chickpea husk. Second, a pure culture study was carried out using six strains of Fibrobacter succinogenes. Both husks stimulated the growth of all tested strains, including a strain that did not grow on rice straw. The strain OS128 that showed the highest growth on rice straw displayed even higher growth on lablab bean husk without a time lag. Finally, two-step incubations were carried out to determine whether prior incubation of rumen fluid with husks stimulates subsequent rice straw digestion. Higher digestibility of rice straw was recorded in the second-round incubation following the first incubation with bean husks. These results suggest that the tested bean husks improve the digestion of rice straw by activating fibrolytic F. succinogenes and other associated bacteria. PMID:23035709

  11. Reclassification of Lactobacillus casei subsp. casei ATCC 393 and Lactobacillus rhamnosus ATCC 15820 as Lactobacillus zeae nom. rev., designation of ATCC 334 as the neotype of L. casei subsp. casei, and rejection of the name Lactobacillus paracasei.

    PubMed

    Dicks, L M; Du Plessis, E M; Dellaglio, F; Lauer, E

    1996-01-01

    The type strain of Lactobacillus casei subsp. casei (ATCC 393) exhibits low levels of DNA homology with other strains of L. casei subsp. casei (8 to 46%) and strains of Lactobacillus paracasei (30 to 50%), but exhibits a level of DNA similarity of 80% with Lactobacillus rhamnosus ATCC 15820, the original type strain of "Lactobacterium zeae" Kuznetsov 1959. Strains ATCC 393T (T = type strain) and ATCC 15820T are members of one protein profile cluster that is separate from the other Lactobacillus spp. The randomly amplified polymorphic DNA PCR profile of strain ATCC 393T is also different from the profiles obtained for the other species. L. casei ATCC 334T is genetically closely related to L. casei subsp. casei strains (71 to 97%) and L. paracasei strains (71 to 91%), is a member of the same protein profile cluster as these organisms, and shares several DNA amplicons with L. paracasei strains. On the basis of these results, we propose that L. casei subsp. casei ATCC 393T and L. rhamnosus ATCC 15820 should be reclassified as members of Lactobacillus zeae nom. rev. (type strain, ATCC 15820), that strain ATCC 334 should be designated the neotype strain of L. casei subsp. casei, and that the name L. paracasei should be rejected. PMID:8573516

  12. Antibodies to Aqx toxin of Actinobacillus equuli in horses and foals.

    PubMed

    Berthoud, H; Frey, J; Sternberg, S; Straub, R; Kuhnert, P

    2004-08-21

    Actinobacillus equuli is found in the normal oral flora of horses, but has been associated with several diseases, and particularly with the usually fatal septicaemia in neonatal foals which is thought to be associated with a failure of the passive transfer of immunoglobulins via the colostrum. The Aqx protein of A equuli, belonging to the RTX family of pore-forming toxins, is also cytotoxic to horse lymphocytes. The presence of antibodies to Aqx was investigated in sera from individual horses from different regions; the sera from adult horses and foals 24 hours after birth reacted with Aqx, and sera from foals sampled shortly after an intake of colostrum also reacted with Aqx, but sera from foals taken before an intake of colostrum did not react with Aqx. PMID:15384504

  13. Serotyping reanalysis of unserotypable Actinobacillus pleuropneumoniae isolates by agar gel diffusion test.

    PubMed

    Morioka, Ayako; Shimazaki, Yoko; Uchiyama, Mariko; Suzuki, Shoko

    2016-05-01

    We observed increasing unserotypable (UT) Actinobacillus pleuropneumoniae isolates using agar gel diffusion (AGD) test. To reanalyze their serovar, we performed rapid slide agglutination (RSA) test and multiplex PCR for 47 UT isolates. Of these, 25 were serovar 1 (UT-serovar 1), 20 were serovar 2 (UT-serovar 2) and 2 were serovar 15 (UT-serovar 15). We examined serotyping antigen extraction temperature to determine heat influence. UT-serovar 1 and 15 were influenced by heat, because their precipitation lines were observed in the case of low antigen extraction temperature. To investigate the relationship between antigenicity and genotype, we performed pulsed-field gel electrophoresis (PFGE) analysis using UT-serovar 2 and 15. The predominant PFGE pattern of UT-serovar 2 was identical to that of serovar 2. PMID:26726101

  14. Capsular polysaccharide antigens for detection of serotype-specific antibodies to Actinobacillus pleuropneumoniae.

    PubMed Central

    Bossé, J T; Johnson, R P; Rosendal, S

    1990-01-01

    Capsular polysaccharides (CPS) of serotypes 1, 2, 5 and 7 of Actinobacillus (Haemophilus) pleuropneumoniae were obtained from 18 h culture supernatants by precipitation with hexadecyltrimethyl-ammonium bromide (Cetavlon) followed by extraction with sodium chloride and reprecipitation in ethanol. These crude extracts, and portions purified further by phenol extraction to remove contaminating proteins, were evaluated as antigens for the detection of serotype-specific antibodies to A. pleuropneumoniae in sera from immunized rabbits and swine by an enzyme-linked immunosorbent assay. The crude extracts reacted strongly with homologous antisera, but except for serotype 1 showed considerable cross-reactivity with antisera to other serotypes. Phenol extraction greatly improved the serospecificity of the antigens from serotypes 1, 7 and, to a lesser extent, 5. The serotype 2 CPS antigen showed poor reactivity following phenol extraction, and did not appear as useful for detection of serotype-specific antibodies. PMID:2379111

  15. Isolation of Actinobacillus seminis from a goat with clinical epididymo-orchitis in Brazil

    PubMed Central

    dos Santos, Fabrine Alexandre; de Azevedo, Edísio Oliveira; de Azevedo, Sérgio Santos; Júnior, Felício Garino; Mota, Rinaldo Aparecido; de Cássia Peixoto Kim, Pomy; Gomes, Ana Lisa Vale; Alves, Clebert José

    2014-01-01

    The present study reports the first isolation of Actinobacillus seminis from a goat in Brazil. A four-year-old Moxotó breeding goat in a flock of 70 goats and 65 sheep reared together in the county of Patos, semiarid region of Northeastern Brazil, showed clinical signs of unilateral orchitis and epididymitis. Diagnosis of A. seminis infection was confirmed by association of clinical findings, bacterial isolation and 16S rRNA gene sequencing. This result suggests that A. seminis may be an additional cause of infertility in goats, and that sheep may be the source of infection because the mixed farming system allows the contact between sheep and goats in the semiarid region of Northeastern Brazil. PMID:24948932

  16. Treatment of pigs experimentally infected with Mycoplasma hyopneumoniae, Pasteurella multocida, and Actinobacillus pleuropneumoniae with various antibiotics.

    PubMed Central

    Stipkovits, L; Miller, D; Glavits, R; Fodor, L; Burch, D

    2001-01-01

    The authors have performed a comparative study of the efficacy of various in-feed medications for the treatment of 5- to 6-week-old specific pathogen-free (SPF) piglets experimentally infected on day 1 with Mycoplasma hyopneumoniae, on day 8 with Pasteurella multocida (serotype A), and on day 15 with Actinobacillus pleuropneumoniae (serotype 2). The treatment started on day 9 and continued for 12 consecutive days, then the piglets were euthanized for examination of macroscopic, histologic, and pathologic lesions and for the presence of mycoplasmas and bacteria in the lungs. Based on the results of clinical observations (respiratory signs, rectal temperature, body weight gain, and feed conversion efficiency), macroscopic and histologic lesions of the lungs, and microbiologic findings, the best results were obtained by treatment of pigs with Econor + chlortetracycline, followed by Tetramutin, Pulmotil, Cyfac, and lincomycin + chlortetracycline. PMID:11768127

  17. Serotyping reanalysis of unserotypable Actinobacillus pleuropneumoniae isolates by agar gel diffusion test

    PubMed Central

    MORIOKA, Ayako; SHIMAZAKI, Yoko; UCHIYAMA, Mariko; SUZUKI, Shoko

    2016-01-01

    We observed increasing unserotypable (UT) Actinobacillus pleuropneumoniae isolates using agar gel diffusion (AGD) test. To reanalyze their serovar, we performed rapid slide agglutination (RSA) test and multiplex PCR for 47 UT isolates. Of these, 25 were serovar 1 (UT-serovar 1), 20 were serovar 2 (UT-serovar 2) and 2 were serovar 15 (UT-serovar 15). We examined serotyping antigen extraction temperature to determine heat influence. UT-serovar 1 and 15 were influenced by heat, because their precipitation lines were observed in the case of low antigen extraction temperature. To investigate the relationship between antigenicity and genotype, we performed pulsed-field gel electrophoresis (PFGE) analysis using UT-serovar 2 and 15. The predominant PFGE pattern of UT-serovar 2 was identical to that of serovar 2. PMID:26726101

  18. Microgravity Alters the Physiological Characteristics of Escherichia coli O157:H7 ATCC 35150, ATCC 43889, and ATCC 43895 under Different Nutrient Conditions

    PubMed Central

    Kim, H. W.; Matin, A.

    2014-01-01

    The aim of this study is to provide understanding of microgravity effects on important food-borne bacteria, Escherichia coli O157:H7 ATCC 35150, ATCC 43889, and ATCC 43895, cultured in nutrient-rich or minimal medium. Physiological characteristics, such as growth (measured by optical density and plating), cell morphology, and pH, were monitored under low-shear modeled microgravity (LSMMG; space conditions) and normal gravity (NG; Earth conditions). In nutrient-rich medium, all strains except ATCC 35150 showed significantly higher optical density after 6 h of culture under LSMMG conditions than under NG conditions (P < 0.05). LSMMG-cultured cells were approximately 1.8 times larger than NG-cultured cells at 24 h; therefore, it was assumed that the increase in optical density was due to the size of individual cells rather than an increase in the cell population. The higher pH of the NG cultures relative to that of the LSMMG cultures suggests that nitrogen metabolism was slower in the latter. After 24 h of culturing in minimal media, LSMMG-cultured cells had an optical density 1.3 times higher than that of NG-cultured cells; thus, the higher optical density in the LSMMG cultures may be due to an increase in both cell size and number. Since bacteria actively grew under LSMMG conditions in minimal medium despite the lower pH, it is of some concern that LSMMG-cultured E. coli O157:H7 may be able to adapt well to acidic environments. These changes may be caused by changes in nutrient metabolism under LSMMG conditions, although this needs to be demonstrated in future studies. PMID:24487539

  19. Proteins found within porcine respiratory tract secretions bind lipopolysaccharides of Actinobacillus pleuropneumoniae.

    PubMed Central

    Bélanger, M; Dubreuil, D; Jacques, M

    1994-01-01

    Affinity for porcine respiratory tract secretions was found in some isolates of Actinobacillus pleuropneumoniae and involved lipopolysaccharides (LPS) (M. Bélanger, S. Rioux, B. Foiry, and M. Jacques, FEMS Microbiol. Lett. 97:119-126, 1992). In the present study, the affinity for a crude preparation of porcine respiratory tract mucus of isolates of the Pasteurellaceae family, i.e., Actinobacillus, Haemophilus, and Pasteurella spp., and of some unrelated gram-negative bacteria was examined. Affinity for crude porcine respiratory tract mucus was not a property shared by all Pasteurellaceae isolates tested. Furthermore, affinity for the porcine crude mucus preparation was not unique to the Pasteurellaceae group and did not seem to be restricted to bacteria originating from pigs. Different surface properties of A. pleuropneumoniae isolates in relation to their adherence to crude mucus were examined. The capsular layer seemed to mask the adhesin and interfered with adherence to crude mucus. Two poorly capsulated isolates, which had a more hydrophobic surface and bound Congo red, were also heavily labeled by gold particles coated with polymyxin, which is known to interact with the lipid A-core region of LPS, and adhered strongly to respiratory tract secretions. Tetramethylurea, charged polymers, divalent cations, chelators, monosaccharides and amino sugars, or lectins were unable to inhibit adherence of A. pleuropneumoniae to the crude mucus preparation. To identify the receptor(s) recognized by the lipopolysaccharidic adhesin of A. pleuropneumoniae, affinity chromatography was used. Two bands, which were proteinaceous in nature, of 10 and 11 kDa were recovered. Our results suggest that two low-molecular-mass proteins present in porcine respiratory tract secretions bind A. pleuropneumoniae LPS. Images PMID:8112857

  20. Genome Sequence of Ureaplasma diversum Strain ATCC 49782.

    PubMed

    Marques, Lucas M; Guimarães, Ana M S; Martins, Hellen B; Rezende, Izadora S; Barbosa, Maysa S; Campos, Guilherme B; do Nascimento, Naíla C; Dos Santos, Andrea P; Amorim, Aline T; Santos, Verena M; Messick, Joanne B; Timenetsky, Jorge

    2015-01-01

    Here, we report the complete genome sequence of Ureaplasma diversum strain ATCC 49782. This species is of bovine origin, having an association with reproductive disorders in cattle, including placentitis, fetal alveolitis, abortion, and birth of weak calves. It has a small circular chromosome of 975,425 bp. PMID:25883297

  1. Draft Genome Sequence of Zymomonas mobilis ZM481 (ATCC 31823)

    PubMed Central

    Zhao, Ning; Pan, Yongxu

    2016-01-01

    Zymomonas mobilis ZM481 (ATCC 31823) is an ethanol-tolerant strain that can produce the highest level of ethanol in Z. mobilis from glucose in the shortest time. Here, we report a draft genome sequence of ZM481, which can help us understand the genes related to the ethanol tolerance of this strain. PMID:27056218

  2. Genome Assembly of Serratia marcescens Type Strain ATCC 13880.

    PubMed

    Daligault, H E; Davenport, K W; Minogue, T D; Broomall, S M; Bruce, D C; Chain, P S; Coyne, S R; Gibbons, H S; Jaissle, J; Rosenzweig, C N; Scholz, M; Teshima, H; Johnson, S L

    2014-01-01

    Serratia marcescens ATCC 13880 is the type strain of the species and a commonly used quality control strain. Here, we present the annotated genome assembly of 5.13 Mbp (59.8% G+C content) as submitted to NCBI under accession no. JOVM00000000. PMID:25291774

  3. Draft Genome Sequence of Alicyclobacillus acidoterrestris Strain ATCC 49025

    PubMed Central

    Pasvolsky, Ronit; Sela, Noa; Green, Stefan J.; Zakin, Varda

    2013-01-01

    Alicyclobacillus acidoterrestris is a spore-forming Gram-positive, thermo-acidophilic, nonpathogenic bacterium which contaminates commercial pasteurized fruit juices. The draft genome sequence for A. acidoterrestris strain ATCC 49025 is reported here, providing genetic data relevant to the successful adaptation and survival of this strain in its ecological niche. PMID:24009113

  4. 3-Methylindole production is regulated in Clostridium scatologenes ATCC 25775

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aims: 3-Methylindole (3-MI) is a degradation product of L-tryptophan and is both an animal waste malodorant and threat to ruminant health. Culture conditions which influence 3-MI production in Clostridium scatologenes ATCC 25775 were investigated. Methods and Results: Cells cultured in anaerobic ...

  5. Genome Sequence of the Oleaginous Yeast Rhodotorula glutinis ATCC 204091.

    PubMed

    Paul, Debarati; Magbanua, Zenaida; Arick, Mark; French, Todd; Bridges, Susan M; Burgess, Shane C; Lawrence, Mark L

    2014-01-01

    Rhodotorula glutinis ATCC 204091 is an oleaginous oxidative red yeast that can accumulate lipids to >50% of its biomass when grown with appropriate carbon and nitrogen ratios. It produces a red pigment consisting of useful antioxidants, such as carotenoids, torulene, and torularhodin, when cultivated under carbon-deficient conditions. PMID:24526636

  6. Draft Genome Sequence of Mycobacterium interjectum Strain ATCC 51457T

    PubMed Central

    Levasseur, Anthony; Asmar, Shady; Robert, Catherine

    2016-01-01

    Mycobacterium interjectum is a nontuberculosis species rarely responsible for human infection. The draft genome of M. interjectum ATCC 51457T comprises 5,927,979 bp, exhibiting 67.91% G+C content, 5,314 protein-coding genes, and 51 predicted RNA genes. PMID:27231376

  7. Complete genome sequence of Campylobacter gracilis ATCC 33236T

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The human oral pathogen Campylobacter gracilis has been isolated from periodontal and endodontal infections, and also from non-oral head, neck or lung infections. This study describes the whole-genome sequence of the human periodontal isolate ATCC 33236T (=FDC 1084), which is the first closed genome...

  8. Draft Genome Sequence of Alternaria alternata ATCC 34957.

    PubMed

    Nguyen, Hai D T; Lewis, Christopher T; Lévesque, C André; Gräfenhan, Tom

    2016-01-01

    We report the draft genome sequence of Alternaria alternata ATCC 34957. This strain was previously reported to produce alternariol and alternariol monomethyl ether on weathered grain sorghum. The genome was sequenced with PacBio technology and assembled into 27 scaffolds with a total genome size of 33.5 Mb. PMID:26769939

  9. Draft Genome Sequence of Rhodococcus rhodochrous Strain ATCC 21198

    SciTech Connect

    Shields-Menard, Sara A.; Brown, Steven D; Klingeman, Dawn Marie; Indest, Karl; Hancock, Dawn; Wewalwela, Jayani; French, Todd; Donaldson, Janet

    2014-01-01

    Rhodococcus rhodochrous is a Gram-positive red-pigmented bacterium commonly found in the soil. The draft genome sequence for R. rhodochrous strain ATCC 21198 is presented here to provide genetic data for a better understanding of its lipid-accumulating capabilities.

  10. Draft Genome Sequence of Mycobacterium houstonense Strain ATCC 49403T

    PubMed Central

    Levasseur, Anthony; Asmar, Shady; Robert, Catherine

    2016-01-01

    Mycobacterium houstonense is a nontuberculous species rarely responsible for human infection. The draft genome of M. houstonense ATCC 49403T comprises 6,451,020 bp, exhibiting a 66.96% G+C content, 5,881 protein-coding genes, and 65 predicted RNA genes. PMID:27231371

  11. Genome Assembly of Serratia marcescens Type Strain ATCC 13880

    PubMed Central

    Daligault, H. E.; Davenport, K. W.; Minogue, T. D.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Coyne, S. R.; Gibbons, H. S.; Jaissle, J.; Rosenzweig, C. N.; Scholz, M.; Teshima, H.

    2014-01-01

    Serratia marcescens ATCC 13880 is the type strain of the species and a commonly used quality control strain. Here, we present the annotated genome assembly of 5.13 Mbp (59.8% G+C content) as submitted to NCBI under accession no. JOVM00000000. PMID:25291774

  12. Iron-Chelating Activity of Tetracyclines and Its Impact on the Susceptibility of Actinobacillus actinomycetemcomitans to These Antibiotics

    PubMed Central

    Grenier, Daniel; Huot, Marie-Pierre; Mayrand, Denis

    2000-01-01

    Three tetracyclines (tetracycline, doxycycline, and minocycline) were found to possess iron-chelating activity in a colorimetric siderophore assay. Determination of MICs indicated that the activity of doxycycline against the periodontopathogen Actinobacillus actinomycetemcomitans was only slightly influenced by the presence of an excess of iron that likely saturates the antibiotic. On the other hand, the MICs of doxycycline and minocycline were significantly lower for A. actinomycetemcomitans cultivated under iron-poor conditions than under iron-rich conditions. PMID:10681353

  13. Draft Genome Sequence of Strain ATCC 33958, Reported To Be Elizabethkingia miricola

    PubMed Central

    Matyi, Stephanie A.; Hoyt, Peter R.; Ayoubi-Canaan, Patricia; Hasan, Nabeeh A.

    2015-01-01

    We report the draft genome of Elizabethkingia strain ATCC 33958, which has been classified as Elizabethkingia miricola. Similar to other Elizabethkingia species, the ATCC 33958 draft genome contains numerous β-lactamase genes. ATCC 33958 also harbors a urease gene cluster which supports classification as E. miricola. PMID:26205869

  14. 40 CFR 180.1205 - Beauveria bassiana ATCC #74040; exemption from the requirements of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Beauveria bassiana ATCC #74040... RESIDUES IN FOOD Exemptions From Tolerances § 180.1205 Beauveria bassiana ATCC #74040; exemption from the... the insecticide Beauveria bassiana (ATCC #74040) in or on all food commodities when applied or used...

  15. 40 CFR 180.1205 - Beauveria bassiana ATCC #74040; exemption from the requirements of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Beauveria bassiana ATCC #74040... RESIDUES IN FOOD Exemptions From Tolerances § 180.1205 Beauveria bassiana ATCC #74040; exemption from the... the insecticide Beauveria bassiana (ATCC #74040) in or on all food commodities when applied or used...

  16. Spectrophotometric evaluation of selenium binding by Saccharomyces cerevisiae ATCC MYA-2200 and Candida utilis ATCC 9950 yeast.

    PubMed

    Kieliszek, Marek; Błażejak, Stanisław; Płaczek, Maciej

    2016-05-01

    In this study, the ability of selenium binding the biomas of Saccharomyces cerevisiae ATCC MYA-2200 and Candida utilis ATCC 9950 was investigated. Sodium selenite(IV) salts were added to the experimental media at concentrations of 10, 20, 40, and 60mg Se(4+) L(-1). In the tested concentration range, one concentration reported a significant reduction in the biomass yield of both yeast strains. Intense growth was observed for C. utilis yeast, which reached the highest biomass yield of 15gd.w.L(-1) after 24h cultivation in the presence of 10mg Se(4+) L(-1). Based on the use of spectrophotometric method for the determination of selenium content by using Variamine Blue as a chromogenic agent, efficient accumulation of this element in the biomass of the investigated yeast was observed. The highest amount of selenium, that is, 5.64mg Se(4+)gd.w.(-1), was bound from the environment by S. cerevisiae ATCC MYA-2200 cultured in the presence of 60mg Se(4+) L(-1) medium 72h Slightly less amount, 5.47mg Se(4+) gd.w.(-1), was absorbed by C. utilis ATCC 9950 during similar cultural conditions. Based on the results of the biomass yield and the use of selenium from the medium, it can be observed that yeasts of the genus Candida are more efficient in binding this element, and this property finds practical application in the production of selenium-enriched yeast. PMID:27049131

  17. Closed Genome Sequence of Clostridium pasteurianum ATCC 6013

    PubMed Central

    Rotta, Carlo; Poehlein, Anja; Schwarz, Katrin; McClure, Peter; Daniel, Rolf

    2015-01-01

    We report here the closed genome of Clostridium pasteurianum ATCC 6013, a saccharolytic, nitrogen-fixing, and spore-forming Gram-positive obligate anaerobe. The organism is of biotechnological interest due to the production of solvents (butanol and 1,3-propanediol) but can be associated with food spoilage. The genome comprises a total of 4,351,223 bp. PMID:25700419

  18. Thermostable purified endoglucanase II from Acidothermus cellulolyticus ATCC

    DOEpatents

    Adney, William S.; Thomas, Steven R.; Nieves, Rafael A.; Himmel, Michael E.

    1994-01-01

    A purified low molecular weight endoglucanase II from Acidothermus cellulolyticus (ATCC 43068) is disclosed. The endoglucanase is water soluble, possesses both C.sub.1, and C.sub.x types of enzyme activity, a high degree of stability toward heat, and exhibits optimum temperature activity at about 81.degree. C. at pH's from about 2 to about 9, and at a inactivation temperature of about 100.degree. C. at pH's from about 2 to about 9.

  19. Magnetic response in cultures of Streptococcus mutans ATCC-27607.

    PubMed

    Adamkiewicz, V W; Bassous, C; Morency, D; Lorrain, P; Lepage, J L

    1987-01-01

    Streptococcus mutans ATCC-27607 produces exopolysaccharides that adhere to glass. In the normal geomagnetic field about 50% more polysaccharide adhere preferentially to glass surfaces facing North as compared to South facing surfaces. Reversal of the direction of the magnetic field by 180 degrees produces a similar reversal in the direction of the preferential accumulation. Reduction of the field by 90% abolishes the preferential accumulation. PMID:3582582

  20. Thermostable purified endoglucanase II from Acidothermus cellulolyticus ATCC

    DOEpatents

    Adney, W.S.; Thomas, S.R.; Nieves, R.A.; Himmel, M.E.

    1994-11-22

    A purified low molecular weight endoglucanase II from Acidothermus cellulolyticus (ATCC 43068) is disclosed. The endoglucanase is water soluble, possesses both C[sub 1], and C[sub x] types of enzyme activity, a high degree of stability toward heat, and exhibits optimum temperature activity at about 81 C at pH's from about 2 to about 9, and at a inactivation temperature of about 100 C at pH's from about 2 to about 9. 9 figs.

  1. Pharmacokinetics of tildipirosin in porcine plasma, lung tissue, and bronchial fluid and effects of test conditions on in vitro activity against reference strains and field isolates of Actinobacillus pleuropneumoniae.

    PubMed

    Rose, M; Menge, M; Bohland, C; Zschiesche, E; Wilhelm, C; Kilp, S; Metz, W; Allan, M; Röpke, R; Nürnberger, M

    2013-04-01

    The pharmacokinetics of tildipirosin (Zuprevo(®) 40 mg/mL solution for injection for pigs), a novel 16-membered-ring macrolide for the treatment for swine respiratory disease (SRD), was investigated in studies collecting blood plasma and postmortem samples of lung tissue and bronchial fluid (BF) from swine. In view of factors influencing the in vitro activity of macrolides, and for the interpretation of tildipirosin pharmacokinetics in relation to minimum inhibitory concentrations (MIC), additional experiments were conducted to study the effects of pH, carbon dioxide-enriched atmosphere, buffers, and serum on tildipirosin MICs for various reference strains and Actinobacillus (A.) pleuropneumoniae field isolates. After single intramuscular (i.m.) injection at 4 mg/kg body weight, maximum plasma concentration (Cmax) was 0.9 μg/mL observed within 23 min (Tmax ). Mean residence time from the time of dosing to the time of last measurable concentration (MRTlast) and terminal half-life (T1/2) both were about 4 days. A dose-response relationship with no significant sex effect is observed for area under the plasma concentration-time curve from time 0 to the last sampling time with a quantifiable drug concentration (AUClast) over the range of doses up to 6 mg/kg. However, linear dose proportionality could not be proven with statistical methods. The time-concentration profile of tildipirosin in BF and lung far exceeded that in blood plasma. In lung, tildipirosin concentrations reached 3.1 μg/g at 2 h, peaked at 4.3 μg/g at day 1, and slowly declined to 0.8 μg/g at day 17. In BF, tildipirosin levels were 14.3, 7.0, and 6.5 μg/g at days 5, 10, and 14. T1/2 in lung was ∼7 days. Tildipirosin is rapidly and extensively distributed to the respiratory tract followed by slow elimination. Culture media pH and carbon dioxide-enriched atmosphere (CO2 -EA) had a marked impact on in vitro activity of tildipirosin in reference strains of various rapidly growing aerobic and

  2. Cellulose produced by Gluconacetobacter xylinus strains ATCC 53524 and ATCC 23768: Pellicle formation, post-synthesis aggregation and fiber density.

    PubMed

    Lee, Christopher M; Gu, Jin; Kafle, Kabindra; Catchmark, Jeffrey; Kim, Seong H

    2015-11-20

    The pellicle formation, crystallinity, and bundling of cellulose microfibrils produced by bacterium Gluconacetobacter xylinus were studied. Cellulose pellicles were produced by two strains (ATCC 53524 and ATCC 23769) for 1 and 7 days; pellicles were analyzed with scanning electron microscopy (SEM), X-ray diffraction (XRD), vibrational sum-frequency-generation (SFG) spectroscopy, and attenuated total reflectance infrared (ATR-IR) spectroscopy. The bacterial cell population was higher at the surface exposed to air, indicating that the newly synthesized cellulose is deposited at the top of the pellicle. XRD, ATR-IR, and SFG analyses found no significant changes in the cellulose crystallinity, crystal size or polymorphic distribution with the culture time. However, SEM and SFG analyses revealed cellulose macrofibrils produced for 7 days had a higher packing density at the top of the pellicle, compared to the bottom. These findings suggest that the physical properties of cellulose microfibrils are different locally within the bacterial pellicles. PMID:26344281

  3. PCR-based identification of serotype 2 isolates of Actinobacillus pleuropneumoniae biovars I and II.

    PubMed

    Hüssy, Daniela; Schlatter, Yvonne; Miserez, Raymond; Inzana, Thomas; Frey, Joachim

    2004-04-19

    A genetic typing method utilizing PCR for the identification of Actinobacillus pleuropneumoniae serotype 2 isolates has been developed based on the in vitro amplification of a 1.4 kb DNA segment of the serotype 2 capsular polysaccharide genes cps2AB. The assay was tested with all serotype reference strains and a collection of 92 different A. pleuropneumoniae strains of all 15 serotypes of both biovars I and II, originating from 18 different countries worldwide. The cps2 based PCR identified the serotype 2 reference strain and all 12 serotype 2 collection strains contained in this set. DNA was not amplified from the remaining A. pleuropneumoniae reference and collection strains, indicating the PCR assay was highly specific. Furthermore, the PCR method detected all 31 A. pleuropneumoniae serotype 2 field isolates from diseased pigs that were identified in parallel as serotype 2 by agar gel diffusion. The serotype 2 PCR assay proved to be highly specific and reliable for the identification of serotype 2 isolates of A. pleuropneumoniae. PMID:15066734

  4. Opsonic capacity of foal serum for the two neonatal pathogens Escherichia coli and Actinobacillus equuli.

    PubMed

    Gröndahl, G; Sternberg, S; Jensen-Waern, M; Johannisson, A

    2001-11-01

    Two of the most commonly isolated foal pathogens are Escherichia coli and Actinobacillus equuli. The hypothesis tested in this study was that young foals carry a lower opsonic capacity for these bacteria compared to adult horses. A flow-cytometric method for the phagocytosis of these by equine neutrophils was established. The opsonic capacity of serum from healthy foals from birth to age 6 weeks was evaluated and related to the concentrations of IgGa and IgGb. Phagocytosis of yeast was used as a control. Serum was required for phagocytosis, with higher concentrations for E. coli than for A. equuli. Ingestion of colostrum led to a significantly higher serum opsonic capacity. After that, there was no consistent age-related trend for opsonic capacity for the different microbes. Foal serum showed similar or higher opsonisation of E. coli and A. equuli compared to serum from mature individuals. During the studied period, the predominance among IgG subisotypes switched from IgGb to IgGa. Although the overall correlation between concentrations of IgG subisotypes and serum opsonic capacity was poor, sera with IgGb levels below 1.9 mg/ml induced lower opsonisation of E. coli and yeast, but not of A. equuli. Complement activation was important for opsonisation of all tested microbes. The results of this study are significant to the understanding of a key immunological facet in the pathophysiology of equine neonatal septicaemia in clinical practice. PMID:11770988

  5. Characterization of an Actinobacillus pleuropneumoniae seeder pig challenge-exposure model.

    PubMed

    Lechtenberg, K F; Shryock, T R; Moore, G

    1994-12-01

    Five strains of Actinobacillus pleuropneumoniae serotype 1 were used to intranasally infect 5 groups of pigs. Using each bacterial strain, infected pigs (termed seeder pigs) were commingled for 48 hours with 5 groups of noninfected test pigs, then were removed. Seeder and test pigs were maintained in isolation and were observed for 14 days. Seeder pigs had mortality that was threefold greater than that of test pigs (24% vs 8%). Rectal temperature in excess of 40.3 C was achieved for 84% of test pigs and 88% of seeder pigs. Neither of these 2 variables was statistically different between the 2 groups of pigs. Clinical impression scores > or = 2 (on a 0 to 3 scale) were three-fold (64% vs 20%) greater for seeder than for test pigs (P < 0.05). The total number of bacterial isolations or nonrecoverable isolates was tabulated for test and seeder pigs' lungs at necropsy, irrespective of the amount of lesions. The number of A pleuropneumoniae isolations was not statistically different between test and seeder pig populations. Recovery of Pasteurella multocida or other bacteria was greater from the seeder pigs (P < 0.05), whereas the number of non-recoverable isolates was greater from test pigs than from seeder pigs (P < 0.05). Assessment of lung lesions at necropsy by either visual estimation or on a weight basis were in agreement.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7887514

  6. Racial tropism of a highly toxic clone of Actinobacillus actinomycetemcomitans associated with juvenile periodontitis.

    PubMed Central

    Haubek, D; Dirienzo, J M; Tinoco, E M; Westergaard, J; López, N J; Chung, C P; Poulsen, K; Kilian, M

    1997-01-01

    Actinobacillus actinomycetemcomitans strains with enhanced levels of production of leukotoxin are characterized by a 530-bp deletion from the promoter region of the leukotoxin gene operon. Previous isolates with this deletion constituted a single clone belonging to serotype b, although they displayed minor differences among each other. We have analyzed the geographic dissemination of this clone by examining 326 A. actinomycetemcomitans isolates from healthy and periodontally diseased individuals as well as from patients with different types of extraoral infections originating from countries worldwide. A total of 38 isolates, all belonging to the same clone, showed the 530-bp deletion. Comparison of a 440-bp sequence from the promoter region of the leukotoxin gene operon from 10 of these strains revealed complete identity, which indicates that the deletion originates from a single mutational event. This particular clone was exclusively associated with localized juvenile periodontitis (LJP). In at least 12 of 28 families from which the clone was isolated, more than one family member had LJP. Notably, all the subjects carrying this clone had a genetic affiliation with the African population. These observations suggest that juvenile periodontitis in some adolescents with an African origin is associated with a disseminating clone of A. actinomycetemcomitans. PMID:9399490

  7. Secreted proteases from Actinobacillus pleuropneumoniae serotype 1 degrade porcine gelatin, hemoglobin and immunoglobulin A.

    PubMed Central

    Negrete-Abascal, E; Tenorio, V R; Serrano, J J; Garcia, C; de la Garza, M

    1994-01-01

    It was found that 48 hour cultures of Actinobacillus pleuropneumoniae secreted proteases into the medium. Electrophoresis in polyacrylamide gels (10%) copolymerized with porcine gelatin (0.1%), of the 70% (NH4)2SO4 precipitate from the culture supernatants, displayed protease activities of different molecular weights: > 200, 200, 90, 80, 70 and 50 kDa. They had activity over a broad range of pHs (4-8), with an optimal pH of 6-7. All were inhibited by 10 mM EDTA, and reactivated by 10 mM calcium. They were stable at -20 degrees C for more than a month. The proteases also degraded porcine IgA and porcine, human, and bovine hemoglobin, although they appeared to be less active against the hemoglobins. The IgA was totally cleaved in 48 h, using supernatants concentrated with polyvinyl pyrrolidone or the 70% (NH4)2SO4. Extracellular proteases could play a role in virulence. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:8004545

  8. [Generation of nalidixic acid-resistant strains and signature-tagged mutants of Actinobacillus pleuropneumoniae].

    PubMed

    Shang, Lin; Li, Wei; Li, Liangjun; Li, Lu; Zhang, Sihua; Li, Tingting; Li, Yaokun; Liu, Lei; Guo, Zhiwei; Zhou, Rui; Chen, Huanchun

    2008-01-01

    Actinobacillus pleuropneumoniae is a very important respiratory pathogen for swine and causes great economic losses in pig industry worldwide. Signature-tagged mutagenesis (STM) is an effective method to identify virulence genes in bacteria. In this study, we selected nalidixic acid-resistant strains of APP serotypes 1 and 3 by in vitro cultivation, and used as receipt strains for constructing transposon mutants by mating with E. coli CC 118 lambdapir or S17-1 lambdapir containing mini-Tn10 tag plasmids pLOF/TAG1-48, with or without the help of E. coli DH5alpha (pRK2073). We screened mutant strains by antibiotics selection, PCR and Southern blot identification. Our data revealed that nalidixic acid-resistance of APP strains could easily be induced in vitro and the resistance was due to the mutation in the DNA gyrase A subunit gene gyrA. In the mating experiments, the bi-parental mating was more effective and easier than tri-parental mating. Different APP strains showed a different mating and transposon efficiency in the bi-parental mating, with the strains of serotype 1 much higher than serotype 3 and the reference strain of serotype 3 higher than the field strains. These data were helpful for the construction of STM mutants and pickup of virulence genes of APP. PMID:18338580

  9. Identification of an immunogenic protein of Actinobacillus seminis that is present in microvesicles

    PubMed Central

    2006-01-01

    Abstract Actinobacillus seminis is a gram-negative bacterium of the Pasteurellaceae family that is involved in ovine epididymitis. Looking for a protein specific to this species, we determined the protein profile of subcellular fractions of A. seminis (American Type Culture Collection number 15768): proteins from the outer membrane (OMPs), inner membrane (IMPs), and cytoplasm (CPs). These profiles provide the first data, to our knowledge, regarding subcellular fractions of A. seminis. In the OMP fraction, we identified a protein with a molecular mass of 75 kDa that proved to be immunogenic and apparently specific for A. seminis. This conclusion was based on the reaction of hyperimmune serum of rabbits inoculated with whole cells of A. seminis that was tested against sonicated complete cells of reference strains and field isolates of Brucella ovis, Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni. No protein of these bacteria cross-reacted with the 75-kDa protein of A. seminis. Furthermore, when each type of hyperimmune serum was tested against the sonicated cells and each of the subcellular fractions of A. seminis, it did not recognize the A. seminis 75-kDa protein. We also isolated and identified this protein in microvesicles released to the culture supernatant. The results suggest that the 75-kDa protein could be used to establish a diagnostic test specific for ovine epididymitis caused by A. seminis. PMID:16548331

  10. Preparation, characterization, and immunogenicity of conjugate vaccines directed against Actinobacillus pleuropneumoniae virulence determinants.

    PubMed

    Byrd, W; Kadis, S

    1992-08-01

    Conjugate vaccines were prepared in an attempt to protect pigs against swine pleuropneumonia induced by Actinobacillus pleuropneumoniae (SPAP). Two subunit conjugates were prepared by coupling the A. pleuropneumoniae 4074 serotype 1 capsular polysaccharide (CP) to the hemolysin protein (HP) and the lipopolysaccharide (LPS) to the HP. Adipic acid dihydrazide was used as a spacer to facilitate the conjugation in a carbodiimide-mediated reaction. The CP and the LPS were found to be covalently coupled to the HP in the conjugates as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and detergent gel chromatography analyses. Following a booster vaccination, pigs exhibited significantly high (P less than 0.05) immunoglobulin G antibodies against CP, LPS, and HP. The anti-CP and anti-LPS immunoglobulin G antibodies were found to function as opsonins in the phagocytosis of A. pleuropneumoniae by polymorphonuclear leukocytes, whereas antibodies to the HP neutralized the cytotoxic effect of the HP on polymorphonuclear leukocytes. No killing of A. pleuropneumoniae was observed when the effects of the antibodies were tested in the presence of complement. Thus, polysaccharide-protein A. pleuropneumoniae conjugates elicit significant antibody responses against each component of each conjugate, which could be instrumental in protecting swine against SPAP. PMID:1639471

  11. Overexpression of Porcine Beta-Defensin 2 Enhances Resistance to Actinobacillus pleuropneumoniae Infection in Pigs

    PubMed Central

    Yang, Xi; Cheng, Yu-Ting; Tan, Mei-Fang; Zhang, Hua-Wei; Liu, Wan-Quan; Zou, Geng; Zhang, Liang-Sheng; Zhang, Chun-Yan; Deng, Si-Min; Yu, Lei; Hu, Xue-Ying; Li, Lu

    2015-01-01

    To reduce the need for antibiotics in animal production, alternative approaches are needed to control infection. We hypothesized that overexpression of native defensin genes will provide food animals with enhanced resistance to bacterial infections. In this study, recombinant porcine beta-defensin 2 (PBD-2) was overexpressed in stably transfected PK-15 porcine kidney cells. PBD-2 antibacterial activities against Actinobacillus pleuropneumoniae, an important respiratory pathogen causing porcine contagious pleuropneumonia, were evaluated on agar plates. Transgenic pigs constitutively overexpressing PBD-2 were produced by a somatic cell cloning method, and their resistance to bacterial infection was evaluated by direct or cohabitation infection with A. pleuropneumoniae. Recombinant PBD-2 peptide that was overexpressed in the PK-15 cells showed antibacterial activity against A. pleuropneumoniae. PBD-2 was overexpressed in the heart, liver, spleen, lungs, kidneys, and jejunum of the transgenic pigs, which showed significantly lower bacterial loads in the lungs and reduced lung lesions after direct or cohabitation infection with A. pleuropneumoniae. The results demonstrate that transgenic overexpression of PBD-2 in pigs confers enhanced resistance against A. pleuropneumoniae infection. PMID:25916992

  12. Actinobacillus actinomycetemcomitans Keratitis After Glaucoma Infiltration Surgery: A Clinical Report and Literature Review.

    PubMed

    Hong, Jiaxu; Xu, Jianjiang; Cao, Wenjun; Ji, Jian; Sun, Xinghuai

    2016-01-01

    Actinobacillus actinomycetemcomitans infection is a rare and easily misdiagnosed ocular disease. In this article, the authors report a chronic, purulent, and difficult-to-treat case of A actinomycetemcomitans keratitis following a glaucoma infiltration surgery.A 56-year-old man with a long-standing history of open-angle glaucoma in both eyes presented with a 12-week history of ocular pain, redness, and blurred vision in his right eye. He underwent a glaucoma infiltration surgery in his right eye 6 months ago. Three months postoperatively, he developed peripheral corneal stromal opacities associated with a white, thin, cystic bleb, and conjunctival injection. These opacities grew despite topical treatment with topical tobramycin, levofloxacin, natamycin, amikacin, and metronidazole eye drops.Multiple corneal scrapings revealed no organisms, and no organisms grew on aerobic, anaerobic, fungal, or mycobacterial cultures. The patient's right eye developed a severe purulent corneal ulcer with a dense hypopyon and required a corneal transplantation. Histopathologic analysis and 16S ribosomalribonucleic acid polymerase chain reaction sequencing revealed A actinomycetemcomitans as the causative organism. Postoperatively, treatment was initiated with topical levofloxacin and cyclosporine, as well as oral levofloxacin and cyclosporine. Graft and host corneal transparency were maintained at the checkup 1 month after surgery.Although it is a rare cause of corneal disease, A actinomycetemcomitans should be suspected in patients with keratitis refractory to topical antibiotic therapy. Delay in diagnosis and appropriate treatment can result in vision loss. PMID:26817919

  13. Cloning and expression of a transferrin-binding protein from Actinobacillus pleuropneumoniae.

    PubMed Central

    Gerlach, G F; Anderson, C; Potter, A A; Klashinsky, S; Willson, P J

    1992-01-01

    An expression library was constructed from Actinobacillus pleuropneumoniae serotype 7. Escherichia coli transformants expressing recombinant proteins were identified by immunoscreening with porcine convalescent serum. One transformant expressing a 60-kDa protein (60K protein) in aggregated form was identified. Serum raised against the recombinant protein recognized a polypeptide with an indistinguishable electrophoretic mobility in the A. pleuropneumoniae wild type after iron-restricted growth only. The recombinant protein bound transferrin after blotting onto nitrocellulose. Using a competitive enzyme-linked immunosorbent assay (ELISA), the specificity of this binding for the amino-terminal half of iron-saturated porcine transferrin was established. Also, the 60K wild-type protein bound hemin as assessed by hemin-agarose chromatography. Hemin could inhibit transferrin binding of the recombinant protein in the competitive ELISA, whereas hemoglobin and synthetic iron chelators failed to do so. Southern blot analysis of several other A. pleuropneumoniae strains indicated that highly homologous sequence is present in eight of eight isolates of serotype 7 and in some isolates of serotypes 2, 3, and 4. Images PMID:1541562

  14. High-molecular-mass lipopolysaccharides are involved in Actinobacillus pleuropneumoniae adherence to porcine respiratory tract cells.

    PubMed Central

    Paradis, S E; Dubreuil, D; Rioux, S; Gottschalk, M; Jacques, M

    1994-01-01

    Actinobacillus pleuropneumoniae is the causative agent of porcine pleuropneumonia. The major adhesin of A. pleuropneumoniae has been identified as the lipopolysaccharides (LPSs) (M. Bélanger, D. Dubreuil, J. Harel, C. Girard, and M. Jacques, Infect. Immun. 58:3523-3530, 1990). Using immunoelectron microscopy and flow cytometry, we showed in the present study that LPSs were well exposed at the surface of this encapsulated microorganism. Immunolocalization with porcine lung and tracheal frozen sections showed that extracted LPS bound to the lung mesenchyme and vascular endothelium and to the tracheal epithelium, respectively. Inhibition of adherence of A. pleuropneumoniae with extracted LPS was also performed with lung and tracheal frozen sections. Acid hydrolysis of LPS revealed that the active component of LPS was not lipid A but the polysaccharides. LPSs from A. pleuropneumoniae serotypes 1 and 2 were separated by chromatography on Sephacryl S-300 SF, in the presence of sodium deoxycholate, according to their molecular masses. The adherence-inhibitory activity was found in the high-molecular-mass fractions. These high-molecular-mass fractions contained 2-keto-3-deoxyoctulosonic acid and neutral sugars, and they were recognized by a monoclonal antibody directed against A. pleuropneumoniae O antigen but not recognized by a monoclonal antibody against capsular antigen. Images PMID:8039902

  15. Draft genome sequence of Rhodococcus rhodochrous strain ATCC 17895

    PubMed Central

    Chen, Bi-Shuang; Otten, Linda G.; Resch, Verena; Muyzer, Gerard; Hanefeld, Ulf

    2013-01-01

    Rhodococcus rhodochrous ATCC 17895 possesses an array of mono- and dioxygenases, as well as hydratases, which makes it an interesting organism for biocatalysis. R. rhodochrous is a Gram-positive aerobic bacterium with a rod-like morphology. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 6,869,887 bp long genome contains 6,609 protein-coding genes and 53 RNA genes. Based on small subunit rRNA analysis, the strain is more likely to be a strain of Rhodococcus erythropolis rather than Rhodococcus rhodochrous. PMID:24501654

  16. Thermostable purified endoglucanas from acidothermus cellulolyticus ATCC 43068

    DOEpatents

    Himmel, Michael E.; Adney, William S.; Tucker, Melvin P.; Grohmann, Karel

    1994-01-01

    A purified low molecular weight cellulase endoglucanase I having a molecular weight of between about 57,420 to about 74,580 daltons from Acidothermus cellulolyticus (ATCC 43068). The cellulase is water soluble, possesses both C.sub.1 and C.sub.x types of enzyme activity, a high degree of stability toward heat, and exhibits optimum temperature activity at about 83.degree. C. at pH's from about 2 to about 9, and in inactivation temperature of about 110.degree. C. at pH's from about 2 to about 9.

  17. Thermostable purified endoglucanase from Acidothermus cellulolyticus ATCC 43068

    DOEpatents

    Himmel, M.E.; Adney, W.S.; Tucker, M.P.; Grohmann, K.

    1994-01-04

    A purified low molecular weight cellulase endoglucanase I having a molecular weight of between about 57,420 to about 74,580 daltons from Acidothermus cellulolyticus (ATCC 43068) is presented. The cellulase is water soluble, possesses both C[sub 1] and C[sub x] types of enzyme activity, a high degree of stability toward heat, and exhibits optimum temperature activity at about 83 C at pH's from about 2 to about 9, and in inactivation temperature of about 110 C at pH's from about 2 to about 9. 7 figures.

  18. Method of producing a cellulase-containing cell-free fermentate produced from microorganism ATCC 55702

    DOEpatents

    Dees, H. Craig

    1998-01-01

    Bacteria which produce large amounts of cellulose-containing cell-free fermentate have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase producing bacterium (ATCC 55702), which was identified through replicate plating. ATCC 55702 has improved characteristics and qualities for the degradation of cellulosic waste materials for fuel production, food processing, textile processing, and other industrial applications. ATCC 55702 is an improved bacterial host for genetic manipulations using recombinant DNA techniques, and is less likely to destroy genetic manipulations using standard mutagenesis techniques.

  19. Method of producing a cellulase-containing cell-free fermentate produced from microorganism ATCC 55702

    DOEpatents

    Dees, H.C.

    1998-05-26

    Bacteria which produce large amounts of cellulose-containing cell-free fermentate have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase producing bacterium (ATCC 55702), which was identified through replicate plating. ATCC 55702 has improved characteristics and qualities for the degradation of cellulosic waste materials for fuel production, food processing, textile processing, and other industrial applications. ATCC 55702 is an improved bacterial host for genetic manipulations using recombinant DNA techniques, and is less likely to destroy genetic manipulations using standard mutagenesis techniques. 5 figs.

  20. Cellulase-containing cell-free fermentate produced from microorganism ATCC 55702

    DOEpatents

    Dees, H. Craig

    1997-12-16

    Bacteria which produce large amounts of cellulase-containing cell-free fermentate have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase producing bacterium (ATCC 55702), which was identified through replicate plating. ATCC 55702 has improved characteristics and qualities for the degradation of cellulosic waste materials for fuel production, food processing, textile processing, and other industrial applications. ATCC 55702 is an improved bacterial host for genetic manipulations using recombinant DNA techniques, and is less likely to destroy genetic manipulations using standard mutagenesis techniques.

  1. Cellulase-containing cell-free fermentate produced from microorganism ATCC 55702

    DOEpatents

    Dees, H.C.

    1997-12-16

    Bacteria which produce large amounts of cellulase-containing cell-free fermentate have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase producing bacterium (ATCC 55702), which was identified through replicate plating. ATCC 55702 has improved characteristics and qualities for the degradation of cellulosic waste materials for fuel production, food processing, textile processing, and other industrial applications. ATCC 55702 is an improved bacterial host for genetic manipulations using recombinant DNA techniques, and is less likely to destroy genetic manipulations using standard mutagenesis techniques. 5 figs.

  2. Pharmacokinetics of tulathromycin in edible tissues of healthy and experimentally infected pigs with Actinobacillus pleuropneumoniae.

    PubMed

    Bladek, Tomasz; Posyniak, Andrzej; Jablonski, Artur; Gajda, Anna

    2015-01-01

    The aim of this study was the comparison of the tissue pharmacokinetics of tulathromycin in healthy pigs and pigs experimentally infected with Actinobacillus pleuropneumoniae (App). Tulathromycin was given to 24 healthy and 24 infected pigs by intramuscular injection at a single dosage of 2.5 mg kg(-1) body weight (b.w.). Pigs were euthanised at each group and then samples of liver, kidney, muscle, injection site and skin with fat were taken at scheduled time points. Drug concentrations were determined by LC-MS/MS. In this study, higher values of the area under the concentration-time curves (AUC) were calculated in all tissue samples taken from infected than healthy pigs. In pigs with App the AUCs of liver, kidney, muscle, skin with fat and injection site were 1111, 1973, 235, 181 and 2931 mg kg(-1) h, while in pigs without inflammation they were 509, 1295, 151, 111 and 1587 mg kg(-1) h, respectively. Maximum drug tissue concentrations (Cmax) in infected animals were 2370, 6650, 2016, 666 and 83,870 µg kg(-1), while in healthy pigs they were 1483, 6677, 1733, 509 and 55,006 µg kg(-1), respectively. The eliminations half-times (T1/2) were respectively longer in all tissue samples taken from infected animals (from 157.3 to 187.3 h) than in healthy ones (from 138.6 to 161.2 h). The tulathromycin tissue concentrations were significantly higher (p < 0.05) in all tissue samples of the infected pigs compared with the healthy animals at 360 h (from 0.0014 to 0.0280) and at 792 h (from 0.0007 to 0.0242) after drug administration. The results suggest that the tissue pharmacokinetic properties and residue depletion of tulathromycin can be influenced by the disease state of animals. PMID:26247868

  3. Oral and systemic immunoglobulin G-subclass antibodies to Actinobacillus actinomycetemcomitans leukotoxin.

    PubMed

    Engström, P E; George, M; Larsson, P; Lally, E T; Taichman, N S; Norhagen, G

    1999-04-01

    Salivary, gingival crevicular fluid and serum-specific immunoglobulin G (IgG)-subclass antibodies to Actinobacillus actinomycetemcomitans leuktoxin were quantified by enzyme-linked immunosorbent assay. Samples were taken from six patients with periodontal pockets > or = 5 mm, harboring A. actinomycetemcomitans in subgingival plaque and from six healthy, sex- and age-matched controls, who did not harbor A. actinomycetemcomitans. In individuals suffering from periodontitis, the median values of specific IgG1- and IgG2-subclass antibodies in saliva, gingival crevicular fluid and serum were, respectively IgG1 147 ng/ml, 5226 ng/ml and 7318 ng/ml and IgG2 4.8 ng/ml, 934 ng/ml and 860 ng/ml. In the patients, specific IgG3 antibodies were detected in one out of six individuals in saliva, in two individuals in gingival crevicular fluid and in five out of six patients in serum with a median value of 561 ng/ml. The median values of specific IgG4 antibodies in saliva, gingival crevicular fluid and serum were below detectable levels. The median values of the total IgG subclasses in saliva and serum were 14622 ng/ml and 10.3 g/l respectively. Individuals with periodontitis had, compared with controls, a higher ratio of specific IgG1 antibodies to total IgG1 in saliva (P < 0.05) and in serum (P < 0.05) and a higher ratio of specific IgG antibodies to total IgG in saliva (P < 0.05) and in serum (P < 0.01). The results show an elevation of both oral and systemic specific antibodies to A. actinomycetemcomitans leukotoxin. PMID:10219169

  4. Structural analysis of the Actinobacillus pleuropneumoniae-RTX-toxin I (ApxI) operon.

    PubMed Central

    Jansen, R; Briaire, J; Kamp, E M; Gielkens, A L; Smits, M A

    1993-01-01

    Actinobacillus pleuropneumoniae-RTX-toxin I (ApxI), an important virulence factor, is secreted by serotypes 1, 5, 9, 10, and 11 of A. pleuropneumoniae. However, sequences homologous to the secretion genes apxIBD of the ApxI operon are present in all 12 serotypes except serotype 3. The purpose of this study was to determine and compare the structures of the ApxI operons of the 12 A. pleuropneumoniae serotypes. We focused on the nucleotide sequence comparison of the ApxI-coding genes, the structures of the ApxI operons, and the transcription of the ApxI operons. We determined the nucleotide sequences of the toxin-encoding apxICA genes of serotype 9 and found that the gene for the structural toxin, apxIA, was almost identical to the apxIA gene of serotype 1. The toxin-encoding genes of the other serotypes are also similar for the main part; nevertheless, two variants were identified, one in serotypes 1, 9, and 11 and one in serotypes 5 and 10. The two apxIA variants differ mainly within the distal 110 nucleotides. Structural analysis demonstrated that intact ApxI operons, consisting of the four contiguous genes apxICABD, are present in serotypes 1, 5, 9, 10, and 11. ApxI operons with a major deletion in the apxICA genes are present in serotypes 2, 4, 6, 7, 8, and 12. Serotype 3 does not contain ApxI operon sequences. We found that all ApxI operons are transcriptionally active despite the partial deletion of the operon in some serotypes. The implications of these data for the expression and secretion of ApxI and the other Apx-toxins, ApxII and ApxIII, as well as for the development of a subunit vaccine against A. pleuropneumoniae will be discussed. Images PMID:8359891

  5. Sub-inhibitory concentrations of penicillin G induce biofilm formation by field isolates of Actinobacillus pleuropneumoniae.

    PubMed

    Hathroubi, S; Fontaine-Gosselin, S-È; Tremblay, Y D N; Labrie, J; Jacques, M

    2015-09-30

    Actinobacillus pleuropneumoniae is a Gram-negative bacterium and causative agent of porcine pleuropneumonia. This is a highly contagious disease that causes important economic losses to the swine industry worldwide. Penicillins are extensively used in swine production and these antibiotics are associated with high systemic clearance and low oral bioavailability. This may expose A. pleuropneumoniae to sub-inhibitory concentrations of penicillin G when the antibiotic is administered orally. Our goal was to evaluate the effect of sub-minimum inhibitory concentration (MIC) of penicillin G on the biofilm formation of A. pleuropneumoniae. Biofilm production of 13 field isolates from serotypes 1, 5a, 7 and 15 was tested in the presence of sub-MIC of penicillin G using a polystyrene microtiter plate assay. Using microscopy techniques and enzymatic digestion, biofilm architecture and composition were also characterized after exposure to sub-MIC of penicillin G. Sub-MIC of penicillin G significantly induced biofilm formation of nine isolates. The penicillin G-induced biofilms contained more poly-N-acetyl-D-glucosamine (PGA), extracellular DNA and proteins when compared to control biofilms grown without penicillin G. Additionally, penicillin G-induced biofilms were sensitive to DNase which was not observed with the untreated controls. Furthermore, sub-MIC of penicillin G up-regulated the expression of pgaA, which encodes a protein involved in PGA synthesis, and the genes encoding the envelope-stress sensing two-component regulatory system CpxRA. In conclusion, sub-MICs of penicillin G significantly induce biofilm formation and this is likely the result of a cell envelope stress sensed by the CpxRA system resulting in an increased production of PGA and other matrix components. PMID:26130517

  6. A computational strategy for the search of regulatory small RNAs in Actinobacillus pleuropneumoniae.

    PubMed

    Rossi, Ciro C; Bossé, Janine T; Li, Yanwen; Witney, Adam A; Gould, Kate A; Langford, Paul R; Bazzolli, Denise M S

    2016-09-01

    Bacterial regulatory small RNAs (sRNAs) play important roles in gene regulation and are frequently connected to the expression of virulence factors in diverse bacteria. Only a few sRNAs have been described for Pasteurellaceae pathogens and no in-depth analysis of sRNAs has been described for Actinobacillus pleuropneumoniae, the causative agent of porcine pleuropneumonia, responsible for considerable losses in the swine industry. To search for sRNAs in A. pleuropneumoniae, we developed a strategy for the computational analysis of the bacterial genome by using four algorithms with different approaches, followed by experimental validation. The coding strand and expression of 17 out of 23 RNA candidates were confirmed by Northern blotting, RT-PCR, and RNA sequencing. Among them, two are likely riboswitches, three are housekeeping regulatory RNAs, two are the widely studied GcvB and 6S sRNAs, and 10 are putative novel trans-acting sRNAs, never before described for any bacteria. The latter group has several potential mRNA targets, many of which are involved with virulence, stress resistance, or metabolism, and connect the sRNAs in a complex gene regulatory network. The sRNAs identified are well conserved among the Pasteurellaceae that are evolutionarily closer to A. pleuropneumoniae and/or share the same host. Our results show that the combination of newly developed computational programs can be successfully utilized for the discovery of novel sRNAs and indicate an intricate system of gene regulation through sRNAs in A. pleuropneumoniae and in other Pasteurellaceae, thus providing clues for novel aspects of virulence that will be explored in further studies. PMID:27402897

  7. Role of (p)ppGpp in Viability and Biofilm Formation of Actinobacillus pleuropneumoniae S8.

    PubMed

    Li, Gang; Xie, Fang; Zhang, Yanhe; Bossé, Janine T; Langford, Paul R; Wang, Chunlai

    2015-01-01

    Actinobacillus pleuropneumoniae is a Gram-negative bacterium and the cause of porcine pleuropneumonia. When the bacterium encounters nutritional starvation, the relA-dependent (p)ppGpp-mediated stringent response is activated. The modified nucleotides guanosine 5'-diphosphate 3'-diphosphate (ppGpp) and guanosine 5'-triphosphate 3'-diphosphate (pppGpp) are known to be signaling molecules in other prokaryotes. Here, to investigate the role of (p)ppGpp in A. pleuropneumoniae, we created a mutant A. pleuropneumoniae strain, S8ΔrelA, which lacks the (p)ppGpp-synthesizing enzyme RelA, and investigated its phenotype in vitro. S8ΔrelA did not survive after stationary phase (starvation condition) and grew exclusively as non-extended cells. Compared to the wild-type (WT) strain, the S8ΔrelA mutant had an increased ability to form a biofilm. Transcriptional profiles of early stationary phase cultures revealed that a total of 405 bacterial genes were differentially expressed (including 380 up-regulated and 25 down-regulated genes) in S8ΔrelA as compared with the WT strain. Most of the up-regulated genes are involved in ribosomal structure and biogenesis, amino acid transport and metabolism, translation cell wall/membrane/envelope biogenesis. The data indicate that (p)ppGpp coordinates the growth, viability, morphology, biofilm formation and metabolic ability of A. pleuropneumoniae in starvation conditions. Furthermore, S8ΔrelA could not use certain sugars nor produce urease which has been associated with the virulence of A. pleuropneumoniae, suggesting that (p)ppGpp may directly or indirectly affect the pathogenesis of A. pleuropneumoniae during the infection process. In summary, (p)ppGpp signaling represents an essential component of the regulatory network governing stress adaptation and virulence in A. pleuropneumoniae. PMID:26509499

  8. Surface Polysaccharide Mutants Reveal that Absence of O Antigen Reduces Biofilm Formation of Actinobacillus pleuropneumoniae

    PubMed Central

    Hathroubi, S.; Hancock, M. A.; Langford, P. R.; Tremblay, Y. D. N.; Labrie, J.

    2015-01-01

    Actinobacillus pleuropneumoniae is a Gram-negative bacterium belonging to the Pasteurellaceae family and the causative agent of porcine pleuropneumonia, a highly contagious lung disease causing important economic losses. Surface polysaccharides, including lipopolysaccharides (LPS) and capsular polysaccharides (CPS), are implicated in the adhesion and virulence of A. pleuropneumoniae, but their role in biofilm formation is still unclear. In this study, we investigated the requirement for these surface polysaccharides in biofilm formation by A. pleuropneumoniae serotype 1. Well-characterized mutants were used: an O-antigen LPS mutant, a truncated core LPS mutant with an intact O antigen, a capsule mutant, and a poly-N-acetylglucosamine (PGA) mutant. We compared the amount of biofilm produced by the parental strain and the isogenic mutants using static and dynamic systems. Compared to the findings for the biofilm of the parental or other strains, the biofilm of the O antigen and the PGA mutants was dramatically reduced, and it had less cell-associated PGA. Real-time PCR analyses revealed a significant reduction in the level of pgaA, cpxR, and cpxA mRNA in the biofilm cells of the O-antigen mutant compared to that in the biofilm cells of the parental strain. Specific binding between PGA and LPS was consistently detected by surface plasmon resonance, but the lack of O antigen did not abolish these interactions. In conclusion, the absence of the O antigen reduces the ability of A. pleuropneumoniae to form a biofilm, and this is associated with the reduced expression and production of PGA. PMID:26483403

  9. Role of (p)ppGpp in Viability and Biofilm Formation of Actinobacillus pleuropneumoniae S8

    PubMed Central

    Li, Gang; Xie, Fang; Zhang, Yanhe; Bossé, Janine T.; Langford, Paul R.; Wang, Chunlai

    2015-01-01

    Actinobacillus pleuropneumoniae is a Gram-negative bacterium and the cause of porcine pleuropneumonia. When the bacterium encounters nutritional starvation, the relA-dependent (p)ppGpp-mediated stringent response is activated. The modified nucleotides guanosine 5’-diphosphate 3’-diphosphate (ppGpp) and guanosine 5’-triphosphate 3’-diphosphate (pppGpp) are known to be signaling molecules in other prokaryotes. Here, to investigate the role of (p)ppGpp in A. pleuropneumoniae, we created a mutant A. pleuropneumoniae strain, S8ΔrelA, which lacks the (p)ppGpp-synthesizing enzyme RelA, and investigated its phenotype in vitro. S8ΔrelA did not survive after stationary phase (starvation condition) and grew exclusively as non-extended cells. Compared to the wild-type (WT) strain, the S8ΔrelA mutant had an increased ability to form a biofilm. Transcriptional profiles of early stationary phase cultures revealed that a total of 405 bacterial genes were differentially expressed (including 380 up-regulated and 25 down-regulated genes) in S8ΔrelA as compared with the WT strain. Most of the up-regulated genes are involved in ribosomal structure and biogenesis, amino acid transport and metabolism, translation cell wall/membrane/envelope biogenesis. The data indicate that (p)ppGpp coordinates the growth, viability, morphology, biofilm formation and metabolic ability of A. pleuropneumoniae in starvation conditions. Furthermore, S8ΔrelA could not use certain sugars nor produce urease which has been associated with the virulence of A. pleuropneumoniae, suggesting that (p)ppGpp may directly or indirectly affect the pathogenesis of A. pleuropneumoniae during the infection process. In summary, (p)ppGpp signaling represents an essential component of the regulatory network governing stress adaptation and virulence in A. pleuropneumoniae. PMID:26509499

  10. New plasmid tools for genetic analysis of Actinobacillus pleuropneumoniae and other pasteurellaceae.

    PubMed

    Bossé, Janine T; Durham, Andrew L; Rycroft, Andrew N; Kroll, J Simon; Langford, Paul R

    2009-10-01

    We have generated a set of plasmids, based on the mobilizable shuttle vector pMIDG100, which can be used as tools for genetic manipulation of Actinobacillus pleuropneumoniae and other members of the Pasteurellaceae. A tandem reporter plasmid, pMC-Tandem, carrying promoterless xylE and gfpmut3 genes downstream of a multiple-cloning site (MCS), can be used for identification of transcriptional regulators and conditions which favor gene expression from different cloned promoters. The ability to detect transcriptional regulators using the tandem reporter system was validated in A. pleuropneumoniae using the cloned rpoE (sigma(E)) promoter (P). The resulting plasmid, pMCrpoEP, was used to identify a mutant defective in production of RseA, the negative regulator of sigma(E), among a bank of random transposon mutants, as well as to detect induction of sigma(E) following exposure of A. pleuropneumoniae to ethanol or heat shock. pMCsodCP, carrying the cloned sodC promoter of A. pleuropneumoniae, was functional in A. pleuropneumoniae, Haemophilus influenzae, Haemophilus parasuis, Mannheimia haemolytica, and Pasteurella multocida. Two general expression vectors, pMK-Express and pMC-Express, which differ in their antibiotic resistance markers (kanamycin and chloramphenicol, respectively), were constructed for the Pasteurellaceae. Both plasmids have the A. pleuropneumoniae sodC promoter upstream of the gfpmut3 gene and an extended MCS. Replacement of gfpmut3 with a gene of interest allows complementation and heterologous gene expression, as evidenced by expression of the Haemophilus ducreyi nadV gene in A. pleuropneumoniae, rendering the latter NAD independent. PMID:19666733

  11. Actinobacillus pleuropneumoniae Possesses an Antiviral Activity against Porcine Reproductive and Respiratory Syndrome Virus

    PubMed Central

    Labrie, Josée; Hernandez Reyes, Yenney; Burciaga Nava, Jorge A.; Gagnon, Carl A.; Jacques, Mario

    2014-01-01

    Pigs are often colonized by more than one bacterial and/or viral species during respiratory tract infections. This phenomenon is known as the porcine respiratory disease complex (PRDC). Actinobacillus pleuropneumoniae (App) and porcine reproductive and respiratory syndrome virus (PRRSV) are pathogens that are frequently involved in PRDC. The main objective of this project was to study the in vitro interactions between these two pathogens and the host cells in the context of mixed infections. To fulfill this objective, PRRSV permissive cell lines such as MARC-145, SJPL, and porcine alveolar macrophages (PAM) were used. A pre-infection with PRRSV was performed at 0.5 multiplicity of infection (MOI) followed by an infection with App at 10 MOI. Bacterial adherence and cell death were compared. Results showed that PRRSV pre-infection did not affect bacterial adherence to the cells. PRRSV and App co-infection produced an additive cytotoxicity effect. Interestingly, a pre-infection of SJPL and PAM cells with App blocked completely PRRSV infection. Incubation of SJPL and PAM cells with an App cell-free culture supernatant is also sufficient to significantly block PRRSV infection. This antiviral activity is not due to LPS but rather by small molecular weight, heat-resistant App metabolites (<1 kDa). The antiviral activity was also observed in SJPL cells infected with swine influenza virus but to a much lower extent compared to PRRSV. More importantly, the PRRSV antiviral activity of App was also seen with PAM, the cells targeted by the virus in vivo during infection in pigs. The antiviral activity might be due, at least in part, to the production of interferon γ. The use of in vitro experimental models to study viral and bacterial co-infections will lead to a better understanding of the interactions between pathogens and their host cells, and could allow the development of novel prophylactic and therapeutic tools. PMID:24878741

  12. A computational strategy for the search of regulatory small RNAs in Actinobacillus pleuropneumoniae

    PubMed Central

    Rossi, Ciro C.; Bossé, Janine T.; Li, Yanwen; Witney, Adam A.; Gould, Kate A.; Langford, Paul R.; Bazzolli, Denise M.S.

    2016-01-01

    Bacterial regulatory small RNAs (sRNAs) play important roles in gene regulation and are frequently connected to the expression of virulence factors in diverse bacteria. Only a few sRNAs have been described for Pasteurellaceae pathogens and no in-depth analysis of sRNAs has been described for Actinobacillus pleuropneumoniae, the causative agent of porcine pleuropneumonia, responsible for considerable losses in the swine industry. To search for sRNAs in A. pleuropneumoniae, we developed a strategy for the computational analysis of the bacterial genome by using four algorithms with different approaches, followed by experimental validation. The coding strand and expression of 17 out of 23 RNA candidates were confirmed by Northern blotting, RT-PCR, and RNA sequencing. Among them, two are likely riboswitches, three are housekeeping regulatory RNAs, two are the widely studied GcvB and 6S sRNAs, and 10 are putative novel trans-acting sRNAs, never before described for any bacteria. The latter group has several potential mRNA targets, many of which are involved with virulence, stress resistance, or metabolism, and connect the sRNAs in a complex gene regulatory network. The sRNAs identified are well conserved among the Pasteurellaceae that are evolutionarily closer to A. pleuropneumoniae and/or share the same host. Our results show that the combination of newly developed computational programs can be successfully utilized for the discovery of novel sRNAs and indicate an intricate system of gene regulation through sRNAs in A. pleuropneumoniae and in other Pasteurellaceae, thus providing clues for novel aspects of virulence that will be explored in further studies. PMID:27402897

  13. Effect of bovine apo-lactoferrin on the growth and virulence of Actinobacillus pleuropneumoniae.

    PubMed

    Luna-Castro, Sarahí; Aguilar-Romero, Francisco; Samaniego-Barrón, Luisa; Godínez-Vargas, Delfino; de la Garza, Mireya

    2014-10-01

    Actinobacillus pleuropneumoniae (App) is a Gram-negative bacterium that causes porcine pleuropneumonia, leading to economic losses in the swine industry. Due to bacterial resistance to antibiotics, new treatments for this disease are currently being sought. Lactoferrin (Lf) is an innate immune system glycoprotein of mammals that is microbiostatic and microbicidal and affects several bacterial virulence factors. The aim of this study was to investigate whether bovine iron-free Lf (BapoLf) has an effect on the growth and virulence of App. Two serotype 1 strains (reference strain S4074 and the isolate BC52) and a serotype 7 reference strain (WF83) were analyzed. First, the ability of App to grow in iron-charged BLf was discarded because in vivo, BapoLf sequesters iron and could be a potential source of this element favoring the infection. The minimum inhibitory concentration of BapoLf was 14.62, 11.78 and 10.56 µM for the strain BC52, S4074 and WF83, respectively. A subinhibitory concentration (0.8 µM) was tested by assessing App adhesion to porcine buccal epithelial cells, biofilm production, and the secretion and function of toxins and proteases. Decrease in adhesion (24-42 %) was found in the serotype 1 strains. Biofilm production decreased (27 %) for only the strain 4074 of serotype 1. Interestingly, biofilm was decreased (60-70 %) in the three strains by BholoLf. Hemolysis of erythrocytes and toxicity towards HeLa cells were not affected by BapoLf. In contrast, proteolytic activity in all strains was suppressed in the presence of BapoLf. Finally, oxytetracycline produced synergistic effect with BapoLf against App. Our results suggest that BapoLf affects the growth and several of the virulence factors in App. PMID:24878848

  14. Identification of dfrA14 in two distinct plasmids conferring trimethoprim resistance in Actinobacillus pleuropneumoniae

    PubMed Central

    Bossé, Janine T.; Li, Yanwen; Walker, Stephanie; Atherton, Tom; Fernandez Crespo, Roberto; Williamson, Susanna M.; Rogers, Jon; Chaudhuri, Roy R.; Weinert, Lucy A.; Oshota, Olusegun; Holden, Matt T. G.; Maskell, Duncan J.; Tucker, Alexander W.; Wren, Brendan W.; Rycroft, Andrew N.; Langford, Paul R.

    2015-01-01

    Objectives The objective of this study was to determine the distribution and genetic basis of trimethoprim resistance in Actinobacillus pleuropneumoniae isolates from pigs in England. Methods Clinical isolates collected between 1998 and 2011 were tested for resistance to trimethoprim and sulphonamide. The genetic basis of trimethoprim resistance was determined by shotgun WGS analysis and the subsequent isolation and sequencing of plasmids. Results A total of 16 (out of 106) A. pleuropneumoniae isolates were resistant to both trimethoprim (MIC >32 mg/L) and sulfisoxazole (MIC ≥256 mg/L), and a further 32 were resistant only to sulfisoxazole (MIC ≥256 mg/L). Genome sequence data for the trimethoprim-resistant isolates revealed the presence of the dfrA14 dihydrofolate reductase gene. The distribution of plasmid sequences in multiple contigs suggested the presence of two distinct dfrA14-containing plasmids in different isolates, which was confirmed by plasmid isolation and sequencing. Both plasmids encoded mobilization genes, the sulphonamide resistance gene sul2, as well as dfrA14 inserted into strA, a streptomycin-resistance-associated gene, although the gene order differed between the two plasmids. One of the plasmids further encoded the strB streptomycin-resistance-associated gene. Conclusions This is the first description of mobilizable plasmids conferring trimethoprim resistance in A. pleuropneumoniae and, to our knowledge, the first report of dfrA14 in any member of the Pasteurellaceae. The identification of dfrA14 conferring trimethoprim resistance in A. pleuropneumoniae isolates will facilitate PCR screens for resistance to this important antimicrobial. PMID:25957382

  15. Nitric oxide production by murine spleen cells stimulated with lipopolysaccharide from Actinobacillus actinomycetemcomitans.

    PubMed

    Sosroseno, Wihaskoro; Herminajeng, Endang; Susilowati, Heni; Budiarti, Sri

    2002-12-01

    The aim of this study was to determine whether Actinobacillus actinomycetemcomitans lipopolysaccharide (LPS-A. actinomycetemcomitans) could induce murine spleen cells to produce nitric oxide (NO). Spleen cells derived from Balb/c mice were stimulated with LPS-A. actinomycetemcomitans or LPS from Escherichia coli for 4 days. The effects of N(G)-monomethyl-L-arginine (NMMA), polymyxin B, and cytokines (IFN-gamma and IL-4) on the production of NO were also assessed. The NO production from the carrageenan-treated spleen cells stimulated with LPS-A. actinomycetemcomitans or both LPS-A. actinomycetemcomitans and IFN-gamma was determined. The carrageenan-treated mice were transferred with splenic macrophages and the NO production was assessed from the spleen cells stimulated with LPS-A. actinomycetemcomitans or LPS-A. actinomycetemcomitans and IFN-gamma. The results showed that NO production was detectable in the cultures of spleen cells stimulated with LPS-A. actinomycetemcomitans in a dose-dependent fashion, but was lower than in the cells stimulated with LPS from E. coli. The NO production was blocked by NMMA and polymyxin B. IFN-gamma up-regulated but IL-4 suppressed the production of NO by the spleen cells stimulated with LPS-A. actinomycetemcomitans. The carrageenan-treated spleen cells failed to produce NO after stimulation with LPS-A. actinomycetemcomitans or both LPS-A. actinomycetemcomitans and IFN-gamma. Adoptive transfer of splenic macrophages to the carrageenan-treated mice could restore the ability of the spleen cells to produce NO. The results of the present study suggest that LPS-A. actinomycetemcomitans under the regulatory control of cytokines induces murine spleen cells to produce NO and that splenic macrophages are the cellular source of the NO production. Therefore, these results may support the view that NO production by LPS-A. actinomycetemcomitans-stimulated macrophages may play a role in the course of periodontal diseases. PMID:16887678

  16. Growth of Actinobacillus pleuropneumoniae is promoted by exogenous hydroxamate and catechol siderophores.

    PubMed Central

    Diarra, M S; Dolence, J A; Dolence, E K; Darwish, I; Miller, M J; Malouin, F; Jacques, M

    1996-01-01

    Siderophores bind ferric ions and are involved in receptor-specific iron transport into bacteria. Six types of siderophores were tested against strains representing the 12 different serotypes of Actinobacillus pleuropneumoniae. Ferrichrome and bis-catechol-based siderophores showed strong growth-promoting activities for A. pleuropneumoniae in a disk diffusion assay. Most strains of A. pleuropneumoniae tested were able to use ferrichrome (21 of 22 or 95%), ferrichrome A (20 of 22 or 90%), and lysine-based bis-catechol (20 of 22 or 90%), while growth of 36% (8 of 22) was promoted by a synthetic hydroxamate, N5-acetyl-N5-hydroxy-L-ornithine tripeptide. A. pleuropneumoniae serotype 1 (strain FMV 87-682) and serotype 5 (strain 2245) exhibited a distinct yellow halo around colonies on Chrome Azurol S agar plates, suggesting that both strains can produce an iron chelator (siderophore) in response to iron stress. The siderophore was found to be neither a phenolate nor a hydroxamate by the chemical tests of Arnow and Csaky, respectively. This is the first report demonstrating the production of an iron chelator and the use of exogenous siderophores by A. pleuropneumoniae. A spermidine-based bis-catechol siderophore conjugated to a carbacephalosporin was shown to inhibit growth of A. pleuropneumoniae. A siderophore-antibiotic-resistant strain was isolated and shown to have lost the ability to use ferrichrome, synthetic hydroxamate, or catechol-based siderophores when grown under conditions of iron restriction. This observation indicated that a common iron uptake pathway, or a common intermediate, for hydroxamate- and catechol-based siderophores may exist in A. pleuropneumoniae. PMID:8975614

  17. Experimental Actinobacillus pleuropneumoniae challenge in swine: Comparison of computed tomographic and radiographic findings during disease

    PubMed Central

    2012-01-01

    Background In pigs, diseases of the respiratory tract like pleuropneumonia due to Actinobacillus pleuropneumoniae (App) infection have led to high economic losses for decades. Further research on disease pathogenesis, pathogen-host-interactions and new prophylactic and therapeutic approaches are needed. In most studies, a large number of experimental animals are required to assess lung alterations at different stages of the disease. In order to reduce the required number of animals but nevertheless gather information on the nature and extent of lung alterations in living pigs, a computed tomographic scoring system for quantifying gross pathological findings was developed. In this study, five healthy pigs served as control animals while 24 pigs were infected with App, the causative agent of pleuropneumonia in pigs, in an established model for respiratory tract disease. Results Computed tomographic (CT) findings during the course of App challenge were verified by radiological imaging, clinical, serological, gross pathology and histological examinations. Findings from clinical examinations and both CT and radiological imaging, were recorded on day 7 and day 21 after challenge. Clinical signs after experimental App challenge were indicative of acute to chronic disease. Lung CT findings of infected pigs comprised ground-glass opacities and consolidation. On day 7 and 21 the clinical scores significantly correlated with the scores of both imaging techniques. At day 21, significant correlations were found between clinical scores, CT scores and lung lesion scores. In 19 out of 22 challenged pigs the determined disease grades (not affected, slightly affected, moderately affected, severely affected) from CT and gross pathological examination were in accordance. Disease classification by radiography and gross pathology agreed in 11 out of 24 pigs. Conclusions High-resolution, high-contrast CT examination with no overlapping of organs is superior to radiography in the

  18. Modulation of Gene Expression in Actinobacillus pleuropneumoniae Exposed to Bronchoalveolar Fluid

    PubMed Central

    Lone, Abdul G.; Deslandes, Vincent; Nash, John H. E.; Jacques, Mario; MacInnes, Janet I.

    2009-01-01

    Background Actinobacillus pleuropneumoniae, the causative agent of porcine contagious pleuropneumonia, is an important pathogen of swine throughout the world. It must rapidly overcome the innate pulmonary immune defenses of the pig to cause disease. To better understand this process, the objective of this study was to identify genes that are differentially expressed in a medium that mimics the lung environment early in the infection process. Methods and Principal Findings Since bronchoalveolar lavage fluid (BALF) contains innate immune and other components found in the lungs, we examined gene expression of a virulent serovar 1 strain of A. pleuropneumoniae after a 30 min exposure to BALF, using DNA microarrays and real-time PCR. The functional classes of genes found to be up-regulated most often in BALF were those encoding proteins involved in energy metabolism, especially anaerobic metabolism, and in cell envelope, DNA, and protein biosynthesis. Transcription of a number of known virulence genes including apxIVA and the gene for SapF, a protein which is involved in resistance to antimicrobial peptides, was also up-regulated in BALF. Seventy-nine percent of the genes that were up-regulated in BALF encoded a known protein product, and of these, 44% had been reported to be either expressed in vivo and/or involved in virulence. Conclusions The results of this study suggest that in early stages of infection, A. pleuropneumoniae may modulate expression of genes involved in anaerobic energy generation and in the synthesis of proteins involved in cell wall biogenesis, as well as established virulence factors. Given that many of these genes are thought to be expressed in vivo or involved in virulence, incubation in BALF appears, at least partially, to simulate in vivo conditions and may provide a useful medium for the discovery of novel vaccine or therapeutic targets. PMID:19578537

  19. Production of Biohydrogen from Wastewater by Klebsiella oxytoca ATCC 13182.

    PubMed

    Thakur, Veena; Tiwari, K L; Jadhav, S K

    2015-08-01

    Production of biohydrogen from distillery effluent was carried out by using Klebsiella oxytoca ATCC 13182. The work focuses on optimization of pH, temperature, and state of bacteria, which are the various affecting factors for fermentative biohydrogen production. Results indicates that at 35 °C for suspended cultures, the production was at its maximum (i.e., 91.33 ± 0.88 mL) when compared with other temperatures. At 35 °C and at pH 5 and 6, maximum productions of 117.67 ± 1.45 and 111.67 ± 2.72 mL were observed with no significant difference. When immobilized, Klebsiella oxytoca ATCC 13182 was used for biohydrogen production at optimized conditions, production was 186.33 ± 3.17 mL. Hence, immobilized cells were found to be more advantageous for biological hydrogen production over suspended form. Physicochemical analysis of the effluent was conducted before and after fermentation and the values suggested that the fermentative process is an efficient method for biological treatment of wastewater. PMID:26237683

  20. Induction of natural competence in Bacillus cereus ATCC14579

    PubMed Central

    Mirończuk, Aleksandra M.; Kovács, Ákos T.; Kuipers, Oscar P.

    2008-01-01

    Summary Natural competence is the ability of certain microbes to take up exogenous DNA from the environment and integrate it in their genome. Competence development has been described for a variety of bacteria, but has so far not been shown to occur in Bacillus cereus. However, orthologues of most proteins involved in natural DNA uptake in Bacillus subtiliscould be identified in B. cereus. Here, we report that B. cereus ATCC14579 can become naturally competent. When expressing the B. subtilis ComK protein using an IPTG‐inducible system in B. cereus ATCC14579, cells grown in minimal medium displayed natural competence, as either genomic DNA or plasmid DNA was shown to be taken up by the cells and integrated into the genome or stably maintained respectively. This work proves that a sufficient structural system for DNA uptake exists in B. cereus. Bacillus cereus can be employed as a model system to investigate the mechanism of DNA uptake in related bacteria such as Bacillus anthracis and Bacillus thuringiensis. Moreover, natural competence provides an important tool for biotechnology, as it will allow more efficient transformation of B. cereus and related organisms, e.g. to knockout genes in a high‐throughput way. PMID:21261842

  1. Comparison of conventional and long-acting oxytetracyclines in prevention of induced Actinobacillus (Haemophilus) pleuropneumoniae infection of growing swine.

    PubMed Central

    Kiorpes, A L; Bäckström, L R; Collins, M T; Kruse, G O

    1989-01-01

    These experiments tested the hypothesis that long-acting oxytetracycline (oxytetracycline-LA) was more effective than regular oxytetracycline in preventing porcine pleuropneumonia when administered either 24 or 48 h prior to experimental challenge with virulent strains of Actinobacillus pleuropneumoniae. Two experiments (1 and 2) were conducted using growing pigs (average weight 12-15 kg). Antibiotic treatments were administered once intramuscularly at 20 mg/kg body weight; controls received an equivalent volume of saline. Clinical signs were recorded over seven days, and mortality rates and pathological lesions were analyzed using analysis of variance. Serum oxytetracycline levels were compared 48 and 72 h postinjection. All pigs developed clinical disease following experimental infection. Actinobacillus pleuropneumoniae was recovered from 42% of experiment 1 pigs and all of experiment 2 pigs. The data showed that both oxytetracycline and oxytetracycline-LA given at the same dose protected pigs against experimental infection when given 24 h prior to challenge, and there was no difference between the efficacy of the two drugs in this experiment. When administered 48 h prior to challenge, only oxytetracycline-LA reduced the clinical signs and pathological changes following A. pleuropneumoniae challenge. Between 48 and 72 h postinjection, oxytetracycline-LA blood levels were significantly greater compared to oxytetracycline-treated pigs. PMID:2531629

  2. Species-specific multiplex PCR for the diagnosis of Brucella ovis, Actinobacillus seminis, and Histophilus somni infection in rams

    PubMed Central

    2013-01-01

    Background Infectious ovine epididymitis results in substantial economic losses worldwide due to reproductive failure and culling of breeders. The most common causative agents of these infections are Brucella ovis, Actinobacillus seminis, and Histophilus somni. The aim of this study was to develop a multiplex PCR assay for simultaneous detection of Brucella ovis, Actinobacillus seminis, and Histophilus somni with species-specific primers applied to biological samples for molecular diagnosis of these infections. Results The multiplex assay was capable of detecting B. ovis, A. seminis, and H. somni DNA simultaneously from genomic bacterial DNA samples and pool of semen samples from experimentally infected rams. The method was highly specific since it did not amplify DNA from other bacterial species that can potentially cause epididymitis in rams as well as species phylogenetically related to B. ovis. All negative control samples were negative in PCR multiplex assay. Urine can be used as an alternative to semen samples. Conclusions The species-specific multiplex PCR assay developed in this study can be successfully used for the detection of three of the most common bacterial causes of ovine epididymitis. PMID:23514236

  3. Antibacterial Activity of Synthetic Peptides Derived from Lactoferricin against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212

    PubMed Central

    León-Calvijo, María A.; Leal-Castro, Aura L.; Almanzar-Reina, Giovanni A.; Rosas-Pérez, Jaiver E.; García-Castañeda, Javier E.; Rivera-Monroy, Zuly J.

    2015-01-01

    Peptides derived from human and bovine lactoferricin were designed, synthesized, purified, and characterized using RP-HPLC and MALDI-TOF-MS. Specific changes in the sequences were designed as (i) the incorporation of unnatural amino acids in the sequence, the (ii) reduction or (iii) elongation of the peptide chain length, and (iv) synthesis of molecules with different number of branches containing the same sequence. For each peptide, the antibacterial activity against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 was evaluated. Our results showed that Peptides I.2 (RWQWRWQWR) and I.4 ((RRWQWR)4K2Ahx2C2) exhibit bigger or similar activity against E. coli (MIC 4–33 μM) and E. faecalis (MIC 10–33 μM) when they were compared with lactoferricin protein (LF) and some of its derivate peptides as II.1 (FKCRRWQWRMKKLGA) and IV.1 (FKCRRWQWRMKKLGAPSITCVRRAE). It should be pointed out that Peptides I.2 and I.4, containing the RWQWR motif, are short and easy to synthesize; our results demonstrate that it is possible to design and obtain synthetic peptides that exhibit enhanced antibacterial activity using a methodology that is fast and low-cost and that allows obtaining products with a high degree of purity and high yield. PMID:25815317

  4. Antibacterial activity of synthetic peptides derived from lactoferricin against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212.

    PubMed

    León-Calvijo, María A; Leal-Castro, Aura L; Almanzar-Reina, Giovanni A; Rosas-Pérez, Jaiver E; García-Castañeda, Javier E; Rivera-Monroy, Zuly J

    2015-01-01

    Peptides derived from human and bovine lactoferricin were designed, synthesized, purified, and characterized using RP-HPLC and MALDI-TOF-MS. Specific changes in the sequences were designed as (i) the incorporation of unnatural amino acids in the sequence, the (ii) reduction or (iii) elongation of the peptide chain length, and (iv) synthesis of molecules with different number of branches containing the same sequence. For each peptide, the antibacterial activity against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 was evaluated. Our results showed that Peptides I.2 (RWQWRWQWR) and I.4 ((RRWQWR)4K2Ahx2C2) exhibit bigger or similar activity against E. coli (MIC 4-33 μM) and E. faecalis (MIC 10-33 μM) when they were compared with lactoferricin protein (LF) and some of its derivate peptides as II.1 (FKCRRWQWRMKKLGA) and IV.1 (FKCRRWQWRMKKLGAPSITCVRRAE). It should be pointed out that Peptides I.2 and I.4, containing the RWQWR motif, are short and easy to synthesize; our results demonstrate that it is possible to design and obtain synthetic peptides that exhibit enhanced antibacterial activity using a methodology that is fast and low-cost and that allows obtaining products with a high degree of purity and high yield. PMID:25815317

  5. Complete Genome and Methylome Sequences of Salmonella enterica subsp. enterica Serovar Panama (ATCC 7378) and Salmonella enterica subsp. enterica Serovar Sloterdijk (ATCC 15791).

    PubMed

    Yao, Kuan; Muruvanda, Tim; Roberts, Richard J; Payne, Justin; Allard, Marc W; Hoffmann, Maria

    2016-01-01

    Salmonella enterica spp. are pathogenic bacteria commonly associated with food-borne outbreaks in human and animals. Salmonella enterica spp. are characterized into more than 2,500 different serotypes, which makes epidemiological surveillance and outbreak control more difficult. In this report, we announce the first complete genome and methylome sequences from two Salmonella type strains associated with food-borne outbreaks, Salmonella enterica subsp. enterica serovar Panama (ATCC 7378) and Salmonella enterica subsp. enterica serovar Sloterdijk (ATCC 15791). PMID:26988049

  6. Pseudomonas oleovorans subsp. lubricantis subsp. nov., and reclassification of Pseudomonas pseudoalcaligenes ATCC 17440T as later synonym of Pseudomonas oleovorans ATCC 8062 T.

    PubMed

    Saha, Ratul; Spröer, Cathrin; Beck, Brian; Bagley, Susan

    2010-04-01

    Isolate RS1(T) isolated from used metalworking fluid was found to be a Gram-negative, motile, and non-spore forming rod. Based on phylogenetic analyses with 16S rRNA, isolate RS1(T) was placed into the mendocina sublineage of Pseudomonas. The major whole cell fatty acids were C(18:1)omega7c (32.6%), C(16:0) (25.5%), and C(15:0) ISO 2OH/C(16:1)omega7c (14.4%). The sequence similarities of isolate RS1(T) based on gyrB and rpoD genes were 98.9 and 98.0% with Pseudomonas pseudoalcaligenes, and 98.5 and 98.1% with Pseudomonas oleovorans, respectively. The ribotyping pattern showed a 0.60 similarity with P. oleovorans ATCC 8062(T) and 0.63 with P. pseudoalcaligenes ATCC17440(T). The DNA G + C content of isolate RS1(T) was 62.2 mol.%. The DNA-DNA relatedness was 73.0% with P. oleovorans ATCC 8062(T) and 79.1% with P. pseudoalcaligenes ATCC 17440(T). On the basis of morphological, biochemical, and molecular studies, isolate RS1(T) is considered to represent a new subspecies of P. oleovorans. Furthermore, based on the DNA-DNA relatedness (>70%), chemotaxonomic, and molecular profile, P. pseudoalcaligenes ATCC 17440(T) and P. oleovorans ATCC 8062(T) should be united under the same name; according to the rules of priority, P. oleovorans, the first described species, is the earlier synonym and P. pseudoalcaligenes is the later synonym. As a consequence, the division of the species P. oleovorans into two novel subspecies is proposed: P. oleovorans subsp. oleovorans subsp. nov. (type strain ATCC 8062(T) = DSM 1045(T) = NCIB 6576(T)), P. oleovorans subsp. lubricantis subsp. nov. (type strain RS1(T) = ATCC BAA-1494(T) = DSM 21016(T)). PMID:19936829

  7. Complete Genome and Methylome Sequences of Salmonella enterica subsp. enterica Serovar Panama (ATCC 7378) and Salmonella enterica subsp. enterica Serovar Sloterdijk (ATCC 15791)

    PubMed Central

    Yao, Kuan; Muruvanda, Tim; Roberts, Richard J.; Payne, Justin; Allard, Marc W.

    2016-01-01

    Salmonella enterica spp. are pathogenic bacteria commonly associated with food-borne outbreaks in human and animals. Salmonella enterica spp. are characterized into more than 2,500 different serotypes, which makes epidemiological surveillance and outbreak control more difficult. In this report, we announce the first complete genome and methylome sequences from two Salmonella type strains associated with food-borne outbreaks, Salmonella enterica subsp. enterica serovar Panama (ATCC 7378) and Salmonella enterica subsp. enterica serovar Sloterdijk (ATCC 15791). PMID:26988049

  8. Complete Genome Sequence of Arthrobacter sp. ATCC 21022, a Host for Bacteriophage Discovery

    PubMed Central

    Russell, Daniel A.

    2016-01-01

    We report the complete genome sequence of Arthrobacter sp. ATCC 21022, a strain maintained by ATCC and a commonly used host for bacteriophage isolation and genomic analysis. The strain is prophage-free and CRISPR-free but codes for two predicted restriction-modification systems. PMID:27013048

  9. Growth of Lactobacillus paracasei ATCC334 in a cheese model system: A biochemical approach

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Growth of Lactobacillus paracasei ATCC 334, in a cheese-ripening model system based upon a medium prepared from ripening Cheddar cheese extract (CCE) was evaluated. Lactobacillus paracasei ATCC 334 grows in CCE made from cheese ripened for 2 (2mCCE), 6 (6mCCE), and 8 (8mCCE) mo, to final cell densit...

  10. Identification of the Herboxidiene Biosynthetic Gene Cluster in Streptomyces chromofuscus ATCC 49982

    PubMed Central

    Shao, Lei; Zi, Jiachen; Zeng, Jia

    2012-01-01

    The 53-kb biosynthetic gene cluster for the novel anticholesterol natural product herboxidiene was identified in Streptomyces chromofuscus ATCC 49982 by genome sequencing and gene inactivation. In addition to herboxidiene, a biosynthetic intermediate, 18-deoxy-herboxidiene, was also isolated from the fermentation broth of S. chromofuscus ATCC 49982 as a minor metabolite. PMID:22247174

  11. Draft Genome Sequence of Chemolithoautotrophic Acetogenic Butanol-Producing Eubacterium limosum ATCC 8486

    PubMed Central

    Song, Yoseb

    2015-01-01

    Eubacterium limosum ATCC 8486 is an anaerobic chemolithoautotrophic acetogenic bacterium that converts and transforms syngas and isoflavonoids to butanol and phytoestrogens, respectively. Here, we report the draft genome sequence of the E. limosum ATCC 8486 (4.37 Mb) strain and its annotation information, including syngas fermentation and denitrification metabolic pathways. PMID:25676768

  12. Complete Genome Sequence of Klebsiella quasipneumoniae subsp. similipneumoniae Strain ATCC 700603

    PubMed Central

    Elliott, Alysha G.; Ganesamoorthy, Devika; Coin, Lachlan; Cooper, Matthew A.

    2016-01-01

    Klebsiella quasipneumoniae subsp. similipneumoniae strain ATCC 700603, formerly known as K. pneumoniae K6, is known for producing extended-spectrum β-lactamase (ESBL) enzymes that can hydrolyze oxyimino-β-lactams, resulting in resistance to these drugs. We herein report the complete genome of strain ATCC 700603 and show that the ESBL genes are plasmid-encoded. PMID:27231369

  13. Osmoresistant yeast Zygosaccharomyces rouxii: the two most studied wild-type strains (ATCC 2623 and ATCC 42981) differ in osmotolerance and glycerol metabolism.

    PubMed

    Pribylova, Lenka; de Montigny, Jacky; Sychrova, Hana

    2007-03-01

    The yeast Zygosaccharomyces rouxii is known for its high tolerance to osmotic stress, which is thought to be caused by sets of specific genes. Relatively few Z. rouxii genes have been identified so far, all of them having homologues in Saccharomyces cerevisiae; none of them was Z. rouxii-specific. Most of the known Z. rouxii genes were isolated from two wild-type strains, ATCC 2623 and ATCC 42981. In this study, we compared these two strains with regard to some of their morphological, physiological and genomic properties. Important differences were found in their salt tolerance and assimilation of glycerol and karyotype; slight differences were also present in their cell morphology. The ATCC 42981 strain showed a higher resistance to salts, higher glycerol production and, unlike ATCC 2623, was able to assimilate glycerol. Under conditions of osmotic stress, the glycerol production in both Z. rouxii strains was much lower than in a S. cerevisiae S288c culture, which suggested the presence of a system that efficiently retains glycerol inside Z. rouxii cells. The karyotype analysis revealed that ATCC 42981 cells contain more chromosomes and have a bigger genome size than those of ATCC 2623. PMID:17351908

  14. Complete genome sequence of Anabaena variabilis ATCC 29413

    SciTech Connect

    Thiel, Teresa; Pratte, Brenda S.; Zhong, Jinshun; Goodwin, Lynne A.; Copeland, A; Lucas, Susan; Han, Cliff; Pitluck, Sam; Land, Miriam L; Kyrpides, Nikos C; Woyke, Tanja

    2013-01-01

    Anabaena variabilis ATCC 29413 is a filamentous, heterocyst-forming cyanobacterium that has served as a model organism, with an extensive literature extending over 40 years. The strain has three distinct nitrogenases that function under different environmental conditions and is capable of photoautotrophic growth in the light and true heterotrophic growth in the dark using fructose as both carbon and energy source. While this strain was first isolated in 1964 in Mississippi and named Ana-baena flos-aquae MSU A-37, it clusters phylogenetically with cyanobacteria of the genus Nostoc. The strain is a moderate thermophile, growing well at approximately 40 C. Here we provide some additional characteristics of the strain, and an analysis of the complete genome sequence.

  15. Xanthomonas campestris atcc 31601 and process for use

    SciTech Connect

    Weisrock, W.P.; McCarthy, E.F.

    1983-11-29

    A degenerative-resistant strain of Xanthomonas campestris has been developed and a process for using this strain to effectively overcome the problems of continuous xanthan production. This strain of X. campestris, designated X. campestris XCP-19 ATCC 31601, is capable of continuously producing xanthan at high specific productivities, i.e., 0.24 to 0.32 gm xanthan/gm cells/hr, for several hundred hours without culture degeneration from inexpensive aqueous nutrient media such as, for example, a minimal medium consisting primarily of inorganic salts, glucose, and NH4Cl. The medium may or may not also contain a yeast extract or yeast autolysate as a supplemental nitrogen source. Any medium having assimilable sources of carbon, nitrogen, and inorganic substances will serve satisfactorily for use with this new organism. 14 claims.

  16. Complete genome sequence of Anabaena variabilis ATCC 29413.

    PubMed

    Thiel, Teresa; Pratte, Brenda S; Zhong, Jinshun; Goodwin, Lynne; Copeland, Alex; Lucas, Susan; Han, Cliff; Pitluck, Sam; Land, Miriam L; Kyrpides, Nikos C; Woyke, Tanja

    2014-06-15

    Anabaena variabilis ATCC 29413 is a filamentous, heterocyst-forming cyanobacterium that has served as a model organism, with an extensive literature extending over 40 years. The strain has three distinct nitrogenases that function under different environmental conditions and is capable of photoautotrophic growth in the light and true heterotrophic growth in the dark using fructose as both carbon and energy source. While this strain was first isolated in 1964 in Mississippi and named Anabaena flos-aquae MSU A-37, it clusters phylogenetically with cyanobacteria of the genus Nostoc. The strain is a moderate thermophile, growing well at approximately 40(°) C. Here we provide some additional characteristics of the strain, and an analysis of the complete genome sequence. PMID:25197444

  17. Thermostable Cyanuric Acid Hydrolase from Moorella thermoacetica ATCC 39073▿

    PubMed Central

    Li, Qingyan; Seffernick, Jennifer L.; Sadowsky, Michael J.; Wackett, Lawrence P.

    2009-01-01

    Cyanuric acid, a metabolic intermediate in the degradation of many s-triazine compounds, is further metabolized by cyanuric acid hydrolase. Cyanuric acid also accumulates in swimming pools due to the breakdown of the sanitizing agents di- and trichloroisocyanuric acid. Structurally stable cyanuric acid hydrolases are being considered for usage in pool water remediation. In this study, cyanuric acid hydrolase from the thermophile Moorella thermoacetica ATCC 39073 was cloned, expressed in Escherichia coli, and purified to homogeneity. The recombinant enzyme was found to have a broader temperature range and greater stability, at both elevated and low temperatures, than previously described cyanuric acid hydrolases. The enzyme had a narrow substrate specificity, acting only on cyanuric acid and N-methylisocyanuric acid. The M. thermoacetica enzyme did not require metals or other discernible cofactors for activity. Cyanuric acid hydrolase from M. thermoacetica is the most promising enzyme to use for cyanuric acid remediation applications. PMID:19767460

  18. Enzymatic specificities and modes of action of the two catalytic domains of the XynC xylanase from Fibrobacter succinogenes S85.

    PubMed Central

    Zhu, H; Paradis, F W; Krell, P J; Phillips, J P; Forsberg, C W

    1994-01-01

    The xylanase XynC of Fibrobacter succinogenes S85 was recently shown to contain three distinct domains, A, B, and C (F. W. Paradis, H. Zhu, P. J. Krell, J. P. Phillips, and C. W. Forsberg, J. Bacteriol. 175:7666-7672, 1993). Domains A and B each bear an active site capable of hydrolyzing xylan, while domain C has no enzymatic activity. Two truncated proteins, each containing a single catalytic domain, named XynC-A and XynC-B were purified to homogeneity. The catalytic domains A and B had similar pH and temperature parameters of 6.0 and 50 degrees C for maximum hydrolytic activity and extensively degraded birch wood xylan to xylose and xylobiose. The Km and Vmax values, respectively, were 2.0 mg ml-1 and 6.1 U mg-1 for the intact enzyme, 1.83 mg ml-1 and 689 U mg-1 for domain A, and 2.38 mg ml-1 and 91.8 U mg-1 for domain B. Although domain A had a higher specific activity than domain B, domain B exhibited a broader substrate specificity and hydrolyzed rye arabinoxylan to a greater extent than domain A. Furthermore, domain B, but not domain A, was able to release xylose at the initial stage of the hydrolysis. Both catalytic domains cleaved xylotriose, xylotetraose, and xylopentaose but had no activity on xylobiose. Bond cleavage frequencies obtained from hydrolysis of xylo-alditol substrates suggest that while both domains have a strong preference for internal linkages of the xylan backbone, domain B has fewer subsites for substrate binding than domain A and cleaves arabinoxylan more efficiently. Chemical modification with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide methiodide and N-bromosuccinimide inactivated both XynC-A and XynC-B in the absence of xylan, indicating that carboxyl groups and tryptophan residues in the catalytic site of each domain have essential roles. Images PMID:8021170

  19. Comparative in vitro activity of 16 antimicrobial agents against Actinobacillus pleuropneumoniae.

    PubMed

    Yoshimura, H; Takagi, M; Ishimura, M; Endoh, Y S

    2002-01-01

    Sixteen antimicrobial agents were tested for their activity against 68 isolates of Actinobacillus pleuropneumoniae by determining the minimum inhibitory concentrations (MICs). Ceftiofur and the fluoroquinolones danofloxacin and enrofloxacin were the most active compounds, with a MIC for 90% of the isolates (MIC90) of (0.05 microg/ml. The MIC90 values of benzylpenicillin, amoxicillin and aspoxicillin were 0.78 units/ml, 0.39 microg/ml and < or = 0.05 microg/ml, respectively. Three isolates (4.4%) were resistant to penicillins, but aspoxicillin was as active as ceftiofur against the susceptible isolates, with MICs of < or = 0.05 microg/ml for all isolates. Resistance to oxytetracycline, chloramphenicol and thiamphenicol occurred in 22 (32.4%), 14 (20.6%) and 15 (22.1%) of the isolates, respectively. Doxycycline was more active than oxytetracycline, with a MIC90 of 1.56 microg/ml as against 25 microg/ml. Florfenicol was not only as active as thiamphenicol, with a MIC for 50% of the isolates (MIC50) of 0.39 microg/ml, but also active against thiamphenicol-resistant isolates. All the isolates were susceptible to florfenicol. All the isolates were also susceptible to gentamicin, spectinomycin, tilmicosin, colistin and tiamulin. Of these, spectinomycin was the least active, with a MIC50 of 25 microg/ml, followed by tiamulin, with a MIC50 of 6.25 microg/ml. Of the 68 isolates tested, 49 (72.0%) were of serotype 2; 14 (20.5%) were of serotype 1; 2 each (3.0%) were of serotypes 5 and 6; and one was of serotype 7. Of the isolates, 23 (33.8%) were resistant to one or more of the major antibiotics. Antibiotic resistance was found only infrequently among serotype 2, with 5 (10.2%) of 49 isolates being resistant to chloramphenicol and/or oxytetracycline, while it occurred in 18 (94.7%) of the 19 isolates of other serotypes. PMID:11860083

  20. The survival rate of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, and Bacteroides forsythus following 4 randomized treatment modalities.

    PubMed

    Shiloah, J; Patters, M R; Dean, J W; Bland, P; Toledo, G

    1997-08-01

    The overall goal of this clinical study was to determine the short-term anti-infective effects of four randomized treatment modalities on Actinobacillus actinomycetemcomitans (Aa), Porphyromonas gingivalis (Pg), and Bacteroides forsythus (Bf) and determine the effects of bacterial survival on treatment outcomes in patients with adult periodontitis. Twelve adult patients requiring therapy for moderate periodontitis were selected for this study. All patients had at least one tooth in each quadrant that had an inflamed pocket of probing depth > or =5 mm with probing attachment loss that harbored at least one of the following three periodontal pathogens: Aa, Pg, or Bf. The number of target organisms per site was determined pre-operatively, at 1 week, and 1 month and 3 months postoperatively utilizing DNA probes. One quadrant in each patient was randomly assigned to each one of the following four treatment groups: 1) scaling and root planing (SRP group); 2) pocket reduction through osseous surgery and apically-positioned flap (OS group); 3) modified Widman flap (MWF group); and 4) modified Widman flap and topical application of saturated citric acid at pH 1 for 3 minutes (CA group). The 4 treatment modalities were performed in one appointment. No postoperative antibiotics were used. Patients were instructed to supplement their daily oral hygiene with chlorohexidine oral rinse during the study. The results of this investigation indicated that: 1) none of the treatment modalities was effective in eliminating the target species; 2) the incidence of infected sites for all groups was 100% preoperatively; 62.5%, 33.3%, and 31.3% at 1 week, and 1 and 3 months postoperatively, respectively; 3) these infected sites lost 1.1 +/- 0.4 mm of probing attachment compared to gain of 0.0 +/- 0.3 mm for uninfected sites; 4) the infected sites had higher plaque and bleeding on probing 0.9 +/- 0.3, 73 +/- 12%, respectively, compared to 0.3 +/- 0.1 and 30 +/- 8% for the uninfected sites

  1. Evaluation and Field Validation of PCR Tests for Detection of Actinobacillus pleuropneumoniae in Subclinically Infected Pigs

    PubMed Central

    Fittipaldi, Nahuel; Broes, André; Harel, Josée; Kobisch, Marylène; Gottschalk, Marcelo

    2003-01-01

    Eight PCR tests were evaluated for their abilities to detect Actinobacillus pleuropneumoniae in swine tonsils. At first they were compared regarding their specificities by using A. pleuropneumoniae and related bacterial species and their analytical sensitivities by using tonsils experimentally infected in vitro. PCRs were carried out both directly with tonsil homogenates (direct PCR) and after culture of the sample (after-culture PCR). Most tests demonstrated good specificities; however, some tests gave false-positive results with some non-A. pleuropneumoniae species. High degrees of variation in the analytical sensitivities among the tests were observed for the direct PCRs (109 to 102 CFU/g of tonsil), whereas those of most of the after-culture PCRs were similar (102 CFU/g of tonsil). In a second phase, the effects of sample storage time and storage conditions were evaluated by using tonsils from experimentally infected animals. Storage at −20°C allowed the detection of the organism for at least 4 months. Finally, the omlA PCR test described by Savoye et al. (C. Savoye et al., Vet. Microbiol. 73:337-347, 2000) and the commercially available Adiavet App PCR test were further validated with field samples. Their effectiveness was compared to those of standard and immunomagnetic separation-based methods of bacterial isolation. In addition, a comparison of tonsil biopsy specimens (from living animals) and whole tonsils (collected at the slaughterhouse) was also conducted. A. pleuropneumoniae was neither isolated nor detected by PCR from a herd serologically negative for A. pleuropneumoniae. PCR was more sensitive than the standard isolation method with whole tonsils from three infected herds. After-culture PCR offered the highest degree of sensitivity (93 and 83% for the omlA and Adiavet App PCRs, respectively). The PCR detection rate was higher with whole tonsils than with tonsil biopsy specimens. Good agreement (κ = 0.65) was found between the presence of A

  2. A Transcriptome Map of Actinobacillus pleuropneumoniae at Single-Nucleotide Resolution Using Deep RNA-Seq

    PubMed Central

    Su, Zhipeng; Zhu, Jiawen; Xu, Zhuofei; Xiao, Ran; Zhou, Rui; Li, Lu; Chen, Huanchun

    2016-01-01

    Actinobacillus pleuropneumoniae is the pathogen of porcine contagious pleuropneumoniae, a highly contagious respiratory disease of swine. Although the genome of A. pleuropneumoniae was sequenced several years ago, limited information is available on the genome-wide transcriptional analysis to accurately annotate the gene structures and regulatory elements. High-throughput RNA sequencing (RNA-seq) has been applied to study the transcriptional landscape of bacteria, which can efficiently and accurately identify gene expression regions and unknown transcriptional units, especially small non-coding RNAs (sRNAs), UTRs and regulatory regions. The aim of this study is to comprehensively analyze the transcriptome of A. pleuropneumoniae by RNA-seq in order to improve the existing genome annotation and promote our understanding of A. pleuropneumoniae gene structures and RNA-based regulation. In this study, we utilized RNA-seq to construct a single nucleotide resolution transcriptome map of A. pleuropneumoniae. More than 3.8 million high-quality reads (average length ~90 bp) from a cDNA library were generated and aligned to the reference genome. We identified 32 open reading frames encoding novel proteins that were mis-annotated in the previous genome annotations. The start sites for 35 genes based on the current genome annotation were corrected. Furthermore, 51 sRNAs in the A. pleuropneumoniae genome were discovered, of which 40 sRNAs were never reported in previous studies. The transcriptome map also enabled visualization of 5'- and 3'-UTR regions, in which contained 11 sRNAs. In addition, 351 operons covering 1230 genes throughout the whole genome were identified. The RNA-Seq based transcriptome map validated annotated genes and corrected annotations of open reading frames in the genome, and led to the identification of many functional elements (e.g. regions encoding novel proteins, non-coding sRNAs and operon structures). The transcriptional units described in this study

  3. Effect of tulathromycin on the carrier status of Actinobacillus pleuropneumoniae serotype 2 in the tonsils of pigs.

    PubMed

    Angen, Ø; Andreasen, M; Nielsen, E O; Stockmarr, A; Baekbo, P

    2008-10-11

    The effect of a single or double dose of tulathromycin was evaluated in pigs carrying Actinobacillus pleuropneumoniae serotype 2 in their tonsils. Twenty-nine pigs from a reinfected specific pathogen-free-herd were selected from animals testing positive in an A pleuropneumoniae serotype 2-specific pcr test on tonsil scrapings and they were divided into three groups. The pigs in group 1 were treated subcutaneously with 2.5 mg/kg tulathromycin on day 0, the pigs in group 2 were treated with 2.5 mg/kg tulathromycin on days 0 and 4, and the pigs in group 3 were left untreated as controls. The pigs were tested by pcr on tonsil scrapings on days 0, 4, 11 and 33, and on day 33 all the animals were euthanased. There were no significant differences between the numbers of PCR-positive animals in the three groups on any of the sampling dates. PMID:18849576

  4. Genetic and Functional Analyses of the Actinobacillus actinomycetemcomitans AfeABCD Siderophore-Independent Iron Acquisition System

    PubMed Central

    Rhodes, Eric R.; Tomaras, Andrew P.; McGillivary, Glen; Connerly, Pamela L.; Actis, Luis A.

    2005-01-01

    The Actinobacillus actinomycetemcomitans afeABCD iron transport system, the expression of which is controlled by iron and Fur, was identified in three different isolates. The protein products of this locus are related to bacterial ABC transporters involved in metal transport. Transformation of the Escherichia coli 1017 iron acquisition mutant with a plasmid harboring afeABCD promoted cell growth under iron-chelated conditions. However, insertion disruption of each of the afeABCD coding regions abolished this growth-relieving effect. The replacement of the parental afeA allele with the derivative afeA::EZ::TN drastically reduced the ability of A. actinomycetemcomitans cells to grow under iron-chelated conditions. PMID:15908408

  5. A genomic island provides Acidithiobacillus ferrooxidans ATCC 53993 additional copper resistance: a possible competitive advantage.

    PubMed

    Orellana, Luis H; Jerez, Carlos A

    2011-11-01

    There is great interest in understanding how extremophilic biomining bacteria adapt to exceptionally high copper concentrations in their environment. Acidithiobacillus ferrooxidans ATCC 53993 genome possesses the same copper resistance determinants as strain ATCC 23270. However, the former strain contains in its genome a 160-kb genomic island (GI), which is absent in ATCC 23270. This GI contains, amongst other genes, several genes coding for an additional putative copper ATPase and a Cus system. A. ferrooxidans ATCC 53993 showed a much higher resistance to CuSO(4) (>100 mM) than that of strain ATCC 23270 (<25 mM). When a similar number of bacteria from each strain were mixed and allowed to grow in the absence of copper, their respective final numbers remained approximately equal. However, in the presence of copper, there was a clear overgrowth of strain ATCC 53993 compared to ATCC 23270. This behavior is most likely explained by the presence of the additional copper-resistance genes in the GI of strain ATCC 53993. As determined by qRT-PCR, it was demonstrated that these genes are upregulated when A. ferrooxidans ATCC 53993 is grown in the presence of copper and were shown to be functional when expressed in copper-sensitive Escherichia coli mutants. Thus, the reason for resistance to copper of two strains of the same acidophilic microorganism could be determined by slight differences in their genomes, which may not only lead to changes in their capacities to adapt to their environment, but may also help to select the more fit microorganisms for industrial biomining operations. PMID:21789491

  6. Molecular cloning and expression of ptxA, the gene encoding the 120-kilodalton cytotoxin of Actinobacillus pleuropneumoniae serotype 2.

    PubMed

    MacDonald, J; Rycroft, A N

    1992-07-01

    The genetic determinants of the 120-kDa cytotoxin of Actinobacillus pleuropneumoniae serotype 2 were isolated from a lambda DNA library by a plaque immunoblot technique. Expression of the 120-kDa polypeptide was confirmed by Western immunoblot analysis of infected Escherichia coli cell lysates, which were shown to be toxic for porcine alveolar macrophages in vitro. The genetic determinants of the toxin were subcloned into the plasmid vector pUC18. This plasmid (pPTX1) directed the synthesis and secretion of the active 120-kDa cytotoxin in E. coli. The recombinant toxin was indistinguishable from native cytotoxin from A. pleuropneumoniae serotype 2 with respect to molecular size, reaction in Western blot analysis, heat lability, cytotoxic activity, and neutralization by serum antibody. A restriction endonuclease cleavage map of pPTX1 was prepared, and deletion mutants were used to locate the minimal region of DNA required for production of intracellular toxin; this gene was termed ptxA. Southern hybridization analysis with a 1.7-kb PvuII fragment located within the ptxA gene revealed sequences with a high degree of homology in serotype reference strains 2, 3, 4, 6, and 8. Other reference strains did not contain sequences that were recognized by this probe. However, related sequences (greater than 71% homology) were detected in Actinobacillus actinomycetemcomitans and A. equuli. Weak hybridization was observed between the ptxA probe and pLKT5, which carries the lktAC genes of Pasteurella haemolytica, and between the ptxA probe and pAPH1, which carries the structural gene for type II hemolysin from A. pleuropneumoniae. The isolation of the genetic determinants of this cytotoxin will enable investigations of the structure and organization of the ptx DNA region and further analysis of its role in the pathogenesis of pleuropneumonia. PMID:1612740

  7. Genome Sequence of the Ethanol-Producing Zymomonas mobilis subsp. mobilis Lectotype Strain ATCC 10988 ▿

    PubMed Central

    Pappas, Katherine M.; Kouvelis, Vassili N.; Saunders, Elizabeth; Brettin, Thomas S.; Bruce, David; Detter, Chris; Balakireva, Mariya; Han, Cliff S.; Savvakis, Giannis; Kyrpides, Nikos C.; Typas, Milton A.

    2011-01-01

    Zymomonas mobilis ATCC 10988 is the type strain of the Z. mobilis subsp. mobilis taxon, members of which are some of the most rigorous ethanol-producing bacteria. Isolated from Agave cactus fermentations in Mexico, ATCC 10988 is one of the first Z. mobilis strains to be described and studied. Its robustness in sucrose-substrate fermentations, physiological characteristics, large number of plasmids, and overall genomic plasticity render this strain important to the study of the species. Here we report the finishing and annotation of the ATCC 10988 chromosomal and plasmid genome. PMID:21725006

  8. Dielectric characterization of forespores isolated from Bacillus megaterium ATCC 19213.

    PubMed Central

    Marquis, R E; Bender, G R; Carstensen, E L; Child, S Z

    1983-01-01

    Isolated stage III forespores of Bacillus megaterium ATCC 19213 in aqueous suspensions were nearly as dehydrated as mature spores, as indicated by low dextran-impermeable volumes of ca. 3.0 ml per g (dry weight) of cells compared with values of ca. 2.6 for mature spores and 7.3 for vegetative cells. The forespores lacked dipicolinate, had only minimal levels of calcium, magnesium, manganese, potassium, and sodium, and were more heat sensitive than vegetative cells. The effective homogeneous conductivities and dielectric constants measured over a frequency range of 1 to 200 MHz indicated that the inherent conductivities of the forespores were unusually low, in keeping with their low mineral contents, but that the forespores could be invaded by environmental ions which could penetrate dielectrically effective membranes. Overall, our findings support the view that the dehydration of a forespore during stage III of sporogenesis may be the result of ion movements out of the forespore into the sporangium. PMID:6401285

  9. L-Lactic Acid Production by Lactobacillus rhamnosus ATCC 10863

    PubMed Central

    Senedese, Ana Lívia Chemeli; Maciel Filho, Rubens; Maciel, Maria Regina Wolf

    2015-01-01

    Lactic acid has been shown to have the most promising application in biomaterials as poly(lactic acid). L. rhamnosus ATCC 10863 that produces L-lactic acid was used to perform the fermentation and molasses was used as substrate. A solution containing 27.6 g/L of sucrose (main composition of molasses) and 3.0 g/L of yeast extract was prepared, considering the final volume of 3,571 mL (14.0% (v/v) inoculum). Batch and fed batch fermentations were performed with temperature of 43.4°C and pH of 5.0. At the fed batch, three molasses feed were applied at 12, 24, and 36 hours. Samples were taken every two hours and the amounts of lactic acid, sucrose, glucose, and fructose were determined by HPLC. The sucrose was barely consumed at both processes; otherwise the glucose and fructose were almost entirely consumed. 16.5 g/L of lactic acid was produced at batch and 22.0 g/L at fed batch. Considering that lactic acid was produced due to the low concentration of the well consumed sugars, the final amount was considerable. The cell growth was checked and no substrate inhibition was observed. A sucrose molasses hydrolysis is suggested to better avail the molasses fermentation with this strain, surely increasing the L-lactic acid. PMID:25922852

  10. New Insights into Chloramphenicol Biosynthesis in Streptomyces venezuelae ATCC 10712

    PubMed Central

    Fernández-Martínez, Lorena T.; Borsetto, Chiara; Gomez-Escribano, Juan Pablo; Bibb, Maureen J.; Al-Bassam, Mahmoud M.; Chandra, Govind

    2014-01-01

    Comparative genome analysis revealed seven uncharacterized genes, sven0909 to sven0915, adjacent to the previously identified chloramphenicol biosynthetic gene cluster (sven0916–sven0928) of Streptomyces venezuelae strain ATCC 10712 that was absent in a closely related Streptomyces strain that does not produce chloramphenicol. Transcriptional analysis suggested that three of these genes might be involved in chloramphenicol production, a prediction confirmed by the construction of deletion mutants. These three genes encode a cluster-associated transcriptional activator (Sven0913), a phosphopantetheinyl transferase (Sven0914), and a Na+/H+ antiporter (Sven0915). Bioinformatic analysis also revealed the presence of a previously undetected gene, sven0925, embedded within the chloramphenicol biosynthetic gene cluster that appears to encode an acyl carrier protein, bringing the number of new genes likely to be involved in chloramphenicol production to four. Microarray experiments and synteny comparisons also suggest that sven0929 is part of the biosynthetic gene cluster. This has allowed us to propose an updated and revised version of the chloramphenicol biosynthetic pathway. PMID:25267678

  11. Highly Hydrolytic Reuteransucrase from Probiotic Lactobacillus reuteri Strain ATCC 55730

    PubMed Central

    Kralj, S.; Stripling, E.; Sanders, P.; van Geel-Schutten, G. H.; Dijkhuizen, L.

    2005-01-01

    Lactobacillus reuteri strain ATCC 55730 (LB BIO) was isolated as a pure culture from a Reuteri tablet purchased from the BioGaia company. This probiotic strain produces a soluble glucan (reuteran), in which the majority of the linkages are of the α-(1→4) glucosidic type (∼70%). This reuteran also contains α-(1→6)- linked glucosyl units and 4,6-disubstituted α-glucosyl units at the branching points. The LB BIO glucansucrase gene (gtfO) was cloned and expressed in Escherichia coli, and the GTFO enzyme was purified. The recombinant GTFO enzyme and the LB BIO culture supernatants synthesized identical glucan polymers with respect to linkage type and size distribution. GTFO thus is a reuteransucrase, responsible for synthesis of this reuteran polymer in LB BIO. The preference of GTFO for synthesizing α-(1→4) linkages is also evident from the oligosaccharides produced from sucrose with different acceptor substrates, e.g., isopanose from isomaltose. GTFO has a relatively high hydrolysis/transferase activity ratio. Complete conversion of 100 mM sucrose by GTFO nevertheless yielded large amounts of reuteran, although more than 50% of sucrose was converted into glucose. This is only the second example of the isolation and characterization of a reuteransucrase and its reuteran product, both found in different L. reuteri strains. GTFO synthesizes a reuteran with the highest amount of α-(1→4) linkages reported to date. PMID:16000808

  12. Microbial ecology of Actinobacillus actinomycetemcomitans, Eikenella corrodens and Capnocytophaga spp. in adult periodontitis.

    PubMed

    Müller, H P; Heinecke, A; Borneff, M; Knopf, A; Kiencke, C; Pohl, S

    1997-08-01

    Information on intraoral distribution of putative periodontal pathogens might be essential for controlling different forms of periodontal disease. Colonization may be either promoted or impeded by other bacteria competing in the subgingival ecosystem. In recent investigations microbial associations between dental organisms have been determined in a multitude of subgingival plaque samples within multiple patients and described by odds ratios, in most circumstances without taking into account the correlated structure of the observations within a single individual. The present investigation had 3 major objectives: (i) to describe the intraoral distribution of some facultatively anaerobic, Gram-negative rods, i.e. Actinobacillus actinomycetemcomitans, Eikenella corrodens-like organisms and Capnocytophaga spp., in a multitude of subgingival and extracrevicular samples of 10 adult subjects with A. actinomycetemcomitans-associated periodontitis; (ii) to analyse possible inconsistencies of microbial associations between these periodontal organisms; and (iii) to determine factors increasing the likelihood of isolating these bacteria in a given subgingival site by employing Generalized Estimation Equation (GEE) methods. Clinical examinations were carried out at 6 sites of every tooth present. In each subject, 13 extracrevicular (2 cheek mucosa, 3 tongue, 4 gingival, 2 tonsillar samples, 1 palatinal, 1 saliva sample) and between 22 and 44 subgingival samples from deepest sites of every tooth present (n = 296) were selectively cultivated for A. actinomycetemcomitans, E. corrodens and Capnocytophaga spp. In extracrevicular material, A. actinomycetemcomitans, Capnocytophaga spp. and E. corrodens were isolated in 9, 10 and 6 patients, and from 65, 82 and 15% samples, respectively. The organisms were recovered from 51, 62 and 27% subgingival plaque samples, respectively. Heterogeneity tests did not reveal significant inconsistencies of microbial associations between bacteria in

  13. Antifungal effects of citronella oil against Aspergillus niger ATCC 16404.

    PubMed

    Li, Wen-Ru; Shi, Qing-Shan; Ouyang, You-Sheng; Chen, Yi-Ben; Duan, Shun-Shan

    2013-08-01

    Essential oils are aromatic oily liquids obtained from some aromatic plant materials. Certain essential oils such as citronella oil contain antifungal activity, but the antifungal effect is still unknown. In this study, we explored the antifungal effect of citronella oil with Aspergillus niger ATCC 16404. The antifungal activity of citronella oil on conidia of A. niger was determined by poisoned food technique, broth dilution method, and disc volatility method. Experimental results indicated that the citronella oil has strong antifungal activity: 0.125 (v/v) and 0.25 % (v/v) citronella oil inhibited the growth of 5 × 10⁵ spore/ml conidia separately for 7 and 28 days while 0.5 % (v/v) citronella oil could completely kill the conidia of 5 × 10⁵ spore/ml. Moreover, the fungicidal kinetic curves revealed that more than 90 % conidia (initial concentration is 5 × 10⁵ spore/ml) were killed in all the treatments with 0.125 to 2 % citronella oil after 24 h. Furthermore, with increase of citronella oil concentration and treatment time, the antifungal activity was increased correspondingly. The 0.5 % (v/v) concentration of citronella oil was a threshold to kill the conidia thoroughly. The surviving conidia treated with 0.5 to 2 % citronella oil decreased by an order of magnitude every day, and no fungus survived after 10 days. With light microscope, scanning electron microscope, and transmission electron microscope, we found that citronella oil could lead to irreversible alteration of the hyphae and conidia. Based on our observation, we hypothesized that the citronella oil destroyed the cell wall of the A. niger hyphae, passed through the cell membrane, penetrated into the cytoplasm, and acted on the main organelles. Subsequently, the hyphae was collapsed and squashed due to large cytoplasm loss, and the organelles were severely destroyed. Similarly, citronella oil could lead to the rupture of hard cell wall and then act on the sporoplasm to kill the

  14. The Zur regulon of Corynebacterium glutamicum ATCC 13032

    PubMed Central

    2010-01-01

    Background Zinc is considered as an essential element for all living organisms, but it can be toxic at large concentrations. Bacteria therefore tightly regulate zinc metabolism. The Cg2502 protein of Corynebacterium glutamicum was a candidate to control zinc metabolism in this species, since it was classified as metalloregulator of the zinc uptake regulator (Zur) subgroup of the ferric uptake regulator (Fur) family of DNA-binding transcription regulators. Results The cg2502 (zur) gene was deleted in the chromosome of C. glutamicum ATCC 13032 by an allelic exchange procedure to generate the zur-deficient mutant C. glutamicum JS2502. Whole-genome DNA microarray hybridizations and real-time RT-PCR assays comparing the gene expression in C. glutamicum JS2502 with that of the wild-type strain detected 18 genes with enhanced expression in the zur mutant. The expression data were combined with results from cross-genome comparisons of shared regulatory sites, revealing the presence of candidate Zur-binding sites in the mapped promoter regions of five transcription units encoding components of potential zinc ABC-type transporters (cg0041-cg0042/cg0043; cg2911-cg2912-cg2913), a putative secreted protein (cg0040), a putative oxidoreductase (cg0795), and a putative P-loop GTPase of the COG0523 protein family (cg0794). Enhanced transcript levels of the respective genes in C. glutamicum JS2502 were verified by real-time RT-PCR, and complementation of the mutant with a wild-type zur gene reversed the effect of differential gene expression. The zinc-dependent expression of the putative cg0042 and cg2911 operons was detected in vivo with a gfp reporter system. Moreover, the zinc-dependent binding of purified Zur protein to double-stranded 40-mer oligonucleotides containing candidate Zur-binding sites was demonstrated in vitro by DNA band shift assays. Conclusion Whole-genome expression profiling and DNA band shift assays demonstrated that Zur directly represses in a zinc

  15. Transcriptomic analysis of Clostridium thermocellum ATCC 27405 cellulose fermentation

    SciTech Connect

    McKeown, Catherine K; Brown, Steven D

    2011-01-01

    The ability of Clostridium thermocellum ATCC 27405 wild-type strain to hydrolyze cellulose and ferment the degradation products directly to ethanol and other metabolic byproducts makes it an attractive candidate for consolidated bioprocessing of cellulosic biomass to biofuels. In this study, whole-genome microarrays were used to investigate the expression of C. thermocellum mRNA during growth on crystalline cellulose in controlled replicate batch fermentations. A time-series analysis of gene expression revealed changes in transcript levels of {approx}40% of genes ({approx}1300 out of 3198 ORFs encoded in the genome) during transition from early-exponential to late-stationary phase. K-means clustering of genes with statistically significant changes in transcript levels identified six distinct clusters of temporal expression. Broadly, genes involved in energy production, translation, glycolysis and amino acid, nucleotide and coenzyme metabolism displayed a decreasing trend in gene expression as cells entered stationary phase. In comparison, genes involved in cell structure and motility, chemotaxis, signal transduction and transcription showed an increasing trend in gene expression. Hierarchical clustering of cellulosome-related genes highlighted temporal changes in composition of this multi-enzyme complex during batch growth on crystalline cellulose, with increased expression of several genes encoding hydrolytic enzymes involved in degradation of non-cellulosic substrates in stationary phase. Overall, the results suggest that under low substrate availability, growth slows due to decreased metabolic potential and C. thermocellum alters its gene expression to (i) modulate the composition of cellulosomes that are released into the environment with an increased proportion of enzymes than can efficiently degrade plant polysaccharides other than cellulose, (ii) enhance signal transduction and chemotaxis mechanisms perhaps to sense the oligosaccharide hydrolysis products

  16. Detergent composition comprising a cellulase containing cell-free fermentate produced from microorganism ATCC 55702 or mutant thereof

    DOEpatents

    Dees, H. Craig

    1998-01-01

    Bacteria which produce large amounts of a cellulase-containing cell-free fermentate have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase producing bacterium (ATCC 55702), which was identified through replicate plating. ATCC 55702 has improved characteristics and qualities for the degradation of cellulosic waste materials for fuel production, food processing, textile processing, and other industrial applications. ATCC 55702 is an improved bacterial host for genetic manipulations using recombinant DNA techniques, and is less likely to destroy genetic manipulations using standard mutagenesis techniques.

  17. Detergent composition comprising a cellulase containing cell-free fermentate produced from microorganism ATCC 55702 or mutant thereof

    DOEpatents

    Dees, H.C.

    1998-07-14

    Bacteria which produce large amounts of a cellulase-containing cell-free fermentate have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase producing bacterium (ATCC 55702), which was identified through replicate plating. ATCC 55702 has improved characteristics and qualities for the degradation of cellulosic waste materials for fuel production, food processing, textile processing, and other industrial applications. ATCC 55702 is an improved bacterial host for genetic manipulations using recombinant DNA techniques, and is less likely to destroy genetic manipulations using standard mutagenesis techniques. 5 figs.

  18. Impact of Actinobacillus pleuropneumoniae biofilm mode of growth on the lipid A structures and stimulation of immune cells.

    PubMed

    Hathroubi, Skander; Beaudry, Francis; Provost, Chantale; Martelet, Léa; Segura, Mariela; Gagnon, Carl A; Jacques, Mario

    2016-07-01

    Actinobacillus pleuropneumoniae (APP), the etiologic agent of porcine pleuropneumonia, forms biofilms on biotic and abiotic surfaces. APP biofilms confers resistance to antibiotics. To our knowledge, no studies have examined the role of APP biofilm in immune evasion and infection persistence. This study was undertaken to (i) investigate biofilm-associated LPS modifications occurring during the switch to biofilm mode of growth; and (ii) characterize pro-inflammatory cytokines expression in porcine pulmonary alveolar macrophages (PAMs) and proliferation in porcine PBMCs challenged with planktonic or biofilm APP cells. Extracted lipid A samples from biofilm and planktonic cultures were analyzed by HPLC high-resolution, accurate mass spectrometry. Biofilm cells displayed significant changes in lipid A profiles when compared with their planktonic counterparts. Furthermore, in vitro experiments were conducted to examine the inflammatory response of PAMs exposed to UV-inactivated APP grown in biofilm or in suspension. Relative mRNA expression of pro-inflammatory genes IL1, IL6, IL8 and MCP1 decreased in PAMs when exposed to biofilm cells compared to planktonic cells. Additionally, the biofilm state reduced PBMCs proliferation. Taken together, APP biofilm cells show a weaker ability to stimulate innate immune cells, which could be due, in part, to lipid A structure modifications. PMID:27226465

  19. Construction and immunogenicity of a ∆apxIC/ompP2 mutant of Actinobacillus pleuropneumoniae and Haemophilus parasuis.

    PubMed

    Liu, Qiong; Gong, Yuheng; Cao, Yuqin; Wen, Xintian; Huang, Xiaobo; Yan, Qigui; Huang, Yong; Cao, Sanjie

    2013-01-01

    The apxIC genes of the Actinobacillus pleuropneumoniae serovar 5 (SC-1), encoding the ApxIactivating proteins, was deleted by a method involving sucrose counter-selection. In this study, a mutant strain of A. pleuropneumoniae (SC-1) was constructed and named DapxIC/ ompP2. The mutant strain contained foreign DNA in the deletion site of ompP2 gene of Haemophilus parasuis. It showed no haemolytic activity and lower virulence of cytotoxicity in mice compared with the parent strain, and its safety and immunogenicity were also evaluated in mice. The LD50 data shown that the mutant strain was attenuated 30-fold, compared with the parent strain (LD50 of the mutant strain and parent strain in mice were determined to be 1.0 × 10(7) CFU and 3.5 × 10(5) CFU respectively). The mutant strain that was attenuated could secrete inactivated ApxIA RTX toxins with complete antigenicity and could be used as a candidate live vaccine strain against infections of A. pleuropneumoniae and H. parasuis. PMID:23718128

  20. Molecular Analysis of an Alternative N-Glycosylation Machinery by Functional Transfer from Actinobacillus pleuropneumoniae to Escherichia coli*

    PubMed Central

    Naegeli, Andreas; Neupert, Christine; Fan, Yao-Yun; Lin, Chia-Wei; Poljak, Kristina; Papini, Anna Maria; Schwarz, Flavio; Aebi, Markus

    2014-01-01

    N-Linked protein glycosylation is a frequent post-translational modification that can be found in all three domains of life. In a canonical, highly conserved pathway, an oligosaccharide is transferred by a membrane-bound oligosaccharyltransferase from a lipid donor to asparagines in the sequon NX(S/T) of secreted polypeptides. The δ-proteobacterium Actinobacillus pleuropneumoniae encodes an unusual pathway for N-linked protein glycosylation. This pathway takes place in the cytoplasm and is mediated by a soluble N-glycosyltransferase (NGT) that uses nucleotide-activated monosaccharides to glycosylate asparagine residues. To characterize the process of cytoplasmic N-glycosylation in more detail, we studied the glycosylation in A. pleuropneumoniae and functionally transferred the glycosylation system to Escherichia coli. N-Linked glucose specific human sera were used for the analysis of the glycosylation process. We identified autotransporter adhesins as the preferred protein substrate of NGT in vivo, and in depth analysis of the modified sites in E. coli revealed a surprisingly relaxed peptide substrate specificity. Although NX(S/T) is the preferred acceptor sequon, we detected glycosylation of alternative sequons, including modification of glutamine and serine residues. We also demonstrate the use of NGT to glycosylate heterologous proteins. Therefore, our study could provide the basis for a novel route for the engineering of N-glycoproteins in bacteria. PMID:24275653

  1. Genomic relatedness among Actinobacillus pleuropneumoniae field strains of sterotypes 1 and 5 isolated from healthy and diseased pigs.

    PubMed Central

    Chatellier, S; Harel, J; Dugourd, D; Chevallier, B; Kobisch, M; Gottschalk, M

    1999-01-01

    Forty-four Actinobacillus pleuropneumoniae isolates recovered from both healthy and diseased pigs were characterized by random amplified polymorphic DNA analysis (RAPD), pulsed field gel electrophoresis (PFGE) and apx toxin gene typing. Nine RAPD types and 14 PFGE patterns were identified. No common RAPD or PFGE patterns were found between strains of serotype 1 and those of serotype 5. The RAPD analysis indicated that the 15 serotype 1 strains isolated from diseased pigs were assigned to 4 RAPD types, with 66% of strains characterized by the same RAPD type. By contrast, the 5 strains of serotype 1 isolated from healthy carriers were dispersed in 4 RAPD types. These data suggest that the diversity of strains isolated from healthy pigs could be higher than that of strains recovered from diseased pigs. In addition, all serotype 5 strains exhibited a unique RAPD type. Unlike RAPD, PFGE analysis allowed discrimination among isolates of serotype 1 and among those of serotype 5. All but 3 isolates showed the same apx genotype as their respective serotype reference strain. These data indicate that RAPD analysis is a valuable rapid tool for routine subtyping of strains of serotype 1. For strains of serotype 5, a combination of several typing methods, such as PFGE and apx gene typing, is needed to provide useful information on the molecular epidemiology of swine pleuropneumonia. Images Figure 1. Figure 3. PMID:10480458

  2. Construction of a broad host range shuttle vector for gene cloning and expression in Actinobacillus pleuropneumoniae and other Pasteurellaceae.

    PubMed

    Frey, J

    1992-01-01

    We have constructed a pair of broad host range expression vectors, pJFF224-NX and pJFF224-XN, based on plasmid RSF1010, which enable cloning and efficient expression of genes in Actinobacillus pleuropneumoniae and Pasteurella haemolytica and in Escherichia coli. The vectors consist of the minimal autonomous replicon of the broad host range plasmid RSF1010 and a type II chloramphenicol acetyl transferase gene for chloramphenicol resistance selection. In addition, they contain a gene expression cassette based on the E. coli bacteriophage T4 gene 32 promoter region and a transcription stop signal, which are separated by a segment of multiple cloning sites in both orientations. Electroporation and subsequent selection for chloramphenicol resistance was used for the introduction of the vectors in A. pleuropneumoniae and P. haemolytica. A promoterless xy/E gene from the Pseudomonas putida TOL plasmid was cloned onto pJFF224-NX. This plasmid enabled efficient expression of active catechol2,3oxygenase in A. pleuropneumoniae and P. haemolytica. It was stably maintained in A. pleuropneumoniae without antibiotic selection, showing less than 0.1% loss after 100 generations, while native RSF1010 and other RSF1010-based vectors were unstable in this host. PMID:1448612

  3. ICEApl1, an Integrative Conjugative Element Related to ICEHin1056, Identified in the Pig Pathogen Actinobacillus pleuropneumoniae

    PubMed Central

    Bossé, Janine T.; Li, Yanwen; Fernandez Crespo, Roberto; Chaudhuri, Roy R.; Rogers, Jon; Holden, Matthew T. G.; Maskell, Duncan J.; Tucker, Alexander W.; Wren, Brendan W.; Rycroft, Andrew N.; Langford, Paul R.; Consortium, the BRaDP1T

    2016-01-01

    ICEApl1 was identified in the whole genome sequence of MIDG2331, a tetracycline-resistant (MIC = 8 mg/L) serovar 8 clinical isolate of Actinobacillus pleuropneumoniae, the causative agent of porcine pleuropneumonia. PCR amplification of virB4, one of the core genes involved in conjugation, was used to identify other A. pleuropneumoniae isolates potentially carrying ICEApl1. MICs for tetracycline were determined for virB4 positive isolates, and shotgun whole genome sequence analysis was used to confirm presence of the complete ICEApl1. The sequence of ICEApl1 is 56083 bp long and contains 67 genes including a Tn10 element encoding tetracycline resistance. Comparative sequence analysis was performed with similar integrative conjugative elements (ICEs) found in other members of the Pasteurellaceae. ICEApl1 is most similar to the 59393 bp ICEHin1056, from Haemophilus influenzae strain 1056. Although initially identified only in serovar 8 isolates of A. pleuropneumoniae (31 from the UK and 1 from Cyprus), conjugal transfer of ICEApl1 to representative isolates of other serovars was confirmed. All isolates carrying ICEApl1 had a MIC for tetracycline of 8 mg/L. This is, to our knowledge, the first description of an ICE in A. pleuropneumoniae, and the first report of a member of the ICEHin1056 subfamily in a non-human pathogen. ICEApl1 confers resistance to tetracycline, currently one of the more commonly used antibiotics for treatment and control of porcine pleuropneumonia. PMID:27379024

  4. Actinobacillus actinomycetemcomitans serotype b-specific polysaccharide antigen stimulates production of chemotactic factors and inflammatory cytokines by human monocytes.

    PubMed Central

    Yamaguchi, N; Yamashita, Y; Ikeda, D; Koga, T

    1996-01-01

    Serotype b-specific polysaccharide antigen (SPA) was extracted from whole cells of Actinobacillus actinomycetemcomitans Y4 by autoclaving and purified by chromatography on DEAE-Sephadex A-25 and Sephacryl S-300. SPA induced the release of monocyte and leukocyte chemotactic factors by human monocytes. Polymyxin B had almost no effect on the release of monocyte chemotactic factor, but a monoclonal antibody against SPA markedly inhibited it. Human monocytes stimulated with SPA exhibited the increased mRNA expression of monocyte chemoattractant protein 1 (MCP-1) and a neutrophil chemotactic factor, interleukin-8 (IL-8). On the other hand, SPA induced the release of IL-1, IL-6, and tumor necrosis factor (TNF) and enhanced the expression of IL-1alpha, IL-1beta, IL-6, and TNF alpha (TNF-alpha) mRNAs. Human monocytes expressed MCP-1 and IL-8 mRNAs when stimulated by human recombinant IL-1alpha, I1-1beta, IL-6, and TNF-alpha, suggesting that these inflammatory cytokines induced by SPA might participate in the production of chemotactic factors in human monocytes. PMID:8698480

  5. Identification of Actinobacillus pleuropneumoniae Genes Preferentially Expressed During Infection Using In Vivo-Induced Antigen Technology (IVIAT).

    PubMed

    Zhang, Fei; Zhang, Yangyi; Wen, Xintian; Huang, Xiaobo; Wen, Yiping; Wu, Rui; Yan, Qigui; Huang, Yong; Ma, Xiaoping; Zhao, Qin; Cao, Sanjie

    2015-10-01

    Porcine pleuropneumonia is an infectious disease caused by Actinobacillus pleuropneumoniae. The identification of A. pleuropneumoniae genes, specially expressed in vivo, is a useful tool to reveal the mechanism of infection. IVIAT was used in this work to identify antigens expressed in vivo during A. pleuropneumoniae infection, using sera from individuals with chronic porcine pleuropneumonia. Sequencing of DNA inserts from positive clones showed 11 open reading frames with high homology to A. pleuropneumoniae genes. Based on sequence analysis, proteins encoded by these genes were involved in metabolism, replication, transcription regulation, and signal transduction. Moreover, three function-unknown proteins were also indentified in this work. Expression analysis using quantitative real-time PCR showed that most of the genes tested were up-regulated in vivo relative to their expression levels in vitro. IVI (in vivoinduced) genes that were amplified by PCR in different A. pleuropneumoniae strains showed that these genes could be detected in almost all of the strains. It is demonstrated that the identified IVI antigen may have important roles in the infection of A. pleuropneumoniae. PMID:26059519

  6. Viability of Actinobacillus pleuropneumoniae in frozen pig lung samples and comparison of different methods of direct diagnosis in fresh samples.

    PubMed

    Gutierrez, C B; Rodriguez Barbosa, J I; Gonzalez, O R; Tascon, R I; Rodriguez Ferri, E F

    1992-04-01

    A comparative study on different methods of diagnosis of Actinobacillus pleuropneumoniae from both fresh and frozen pig lungs is described. A total of 196 lung tissues with pneumonic lesions were examined for culture isolation on chocolate blood agar, as well as for antigen detection by means of the coagglutination test, the immunodiffusion test and the indirect ELISA. These samples were subsequently frozen for 1 yr and then they were recultured. A. pleuropneumoniae was recovered from fresh lung specimens in 30 cases (15.3%) and from frozen samples in only two cases (0.9%). Such a different degree of isolation demonstrates that long freezing had an adverse effect on the viability of this organism in lung samples. A pleuropneumoniae detection was positive in 134 samples (68.4%) by at least one of the immunological techniques examined. The indirect ELISA was the most sensitive and specific test, with antigen detected in 125 lungs (63.8%). In comparison with the coagglutination and immunodiffusion tests, the sensitivities of the indirect ELISA were 95.8 and 93.7%, and the specificities were 67.0 and 63.4%, respectively. PMID:1563263

  7. Susceptibility testing of Actinobacillus pleuropneumoniae in Denmark. Evaluation of three different media of MIC-determinations and tablet diffusion tests.

    PubMed

    Aarestrup, F M; Jensen, N E

    1999-02-12

    This study was conducted to compare the applicability of three different media in sensitivity testing of Actinobacillus pleuropneumoniae by means of MIC and tablet diffusion tests. The media used were: modified PPLO agar, chocolatized Mueller-Hinton-II and Columbia agar supplemented with NAD. Seven antimicrobial agents were tested: ceftiofur, enrofloxacin, penicillin, spectinomycin, tiamulin, trimethoprim + sulfadiazine and tylosin, against 40 randomly selected A. pleuropneumoniae isolates. In general, good agreement was found between results obtained with all combinations of media, most antimicrobials tested and the two-test systems. Some variations between media were observed for spectinomycin, tiamulin and tylosin. For ceftiofur and trimethoprim + sulfadiazine some isolates with low MIC-values were classified as resistant using tablet diffusion, indicating that the break points of resistance for these antimicrobials using the tablet diffusion tests need adjustment. Using current break points for resistance with MIC-determinations, all isolates tested susceptible to ceftiofur, enrofloxacin, penicillin, tiamulin and trimethoprim + sulfadiazine. A larger number of isolates tested resistant to spectinomycin and tylosin on all three media using both MIC determinations and tablet diffusion. PMID:10063535

  8. Identification of a hemolysin from Actinobacillus pleuropneumoniae and characterization of its channel properties in planar phospholipid bilayers.

    PubMed

    Lalonde, G; McDonald, T V; Gardner, P; O'Hanley, P D

    1989-08-15

    A proteinaceous hemolysin secreted by strain 4074 of serotype 1 of Actinobacillus pleuropneumoniae was purified by diafiltration and ion exchange chromatographic techniques. The hemolytic activity is associated with a 107-kDa band as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and confirmed by Western blotting and immunoprecipitation. This hemolysin produces pores in membranes as demonstrated by osmotic protection studies using red blood cells and carbohydrate compounds of various molecular weights. These assays suggest a pore diameter in the order of 2 nm. Phospholipid bilayers composed of 1:1 w/w phosphotidylserine:phosphotidylethanolamine exposed to this toxin display discrete current flow events typical of transmembrane channels and consistent with the interpretation that this toxin acts by forming pores in phospholipid membranes. The linear relationship of current amplitude to holding potential when examined over the -60 to +60 mV range indicates that this pore has a constant mean single channel conductance level of 350-400 pS. PMID:2474533

  9. Detection of cytolethal distending toxin activity and cdt genes in Actinobacillus actinomycetemcomitans isolates from geographically diverse populations

    PubMed Central

    Fabris, A. S.; DiRienzo, J. M.; Wïkstrom, M.; Mayer, M. P. A.

    2008-01-01

    A cytolethal distending toxin (CDT) found in Actinobacillus actinomycetemcomitans inhibits the eukaryotic cell cycle, which may contribute to the pathogenic potential of the bacterium. The presence of the cdtABC genes and CDT activity were examined in 40 clinical isolates of A. actinomycetemcomitans from Brazil, Kenya, Japan and Sweden. Thirty-nine of 40 cell lysates caused distension of Chinese hamster ovary cells. At least one of the cdt genes was detected in all strains examined. The three cdt genes were detected, by PCR, in 34 DNA samples. DNA from one strain from Kenya did not yield amplicons of the cdtA and cdtB genes and did not express toxic activity. Restriction analysis was performed on every amplicon obtained. PCR-RFLP patterns revealed that the three cdt genes were conserved. These data provided evidence that the cdt genes are found and expressed in the majority of the A. actinomycetemcomitans isolates. Although a quantitative difference in cytotoxicity was observed, indicating variation in expression of CDT among strains, no clear relationship between CDT activity and periodontal status was found. PMID:12121473

  10. Specific point mutations in Lactobacillus casei ATCC 27139 cause a phenotype switch from Lac- to Lac+.

    PubMed

    Tsai, Yu-Kuo; Chen, Hung-Wen; Lo, Ta-Chun; Lin, Thy-Hou

    2009-03-01

    Lactose metabolism is a changeable phenotype in strains of Lactobacillus casei. In this study, we found that L. casei ATCC 27139 was unable to utilize lactose. However, when exposed to lactose as the sole carbon source, spontaneous Lac(+) clones could be obtained. A gene cluster (lacTEGF-galKETRM) involved in the metabolism of lactose and galactose in L. casei ATCC 27139 (Lac(-)) and its Lac(+) revertant (designated strain R1) was sequenced and characterized. We found that only one nucleotide, located in the lacTEGF promoter (lacTp), of the two lac-gal gene clusters was different. The protein sequence identity between the lac-gal gene cluster and those reported previously for some L. casei (Lac(+)) strains was high; namely, 96-100 % identity was found and no premature stop codon was identified. A single point mutation located within the lacTp promoter region was also detected for each of the 41 other independently isolated Lac(+) revertants of L. casei ATCC 27139. The revertants could be divided into six classes based on the positions of the point mutations detected. Primer extension experiments conducted on transcription from lacTp revealed that the lacTp promoter of these six classes of Lac(+) revertants was functional, while that of L. casei ATCC 27139 was not. Northern blotting experiments further confirmed that the lacTEGF operon of strain R1 was induced by lactose but suppressed by glucose, whereas no blotting signal was ever detected for L. casei ATCC 27139. These results suggest that a single point mutation in the lacTp promoter was able to restore the transcription of a fully functional lacTEGF operon and cause a phenotype switch from Lac(-) to Lac(+) for L. casei ATCC 27139. PMID:19246746

  11. 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

  12. Relationship between Glycopeptide Production and Resistance in the Actinomycete Nonomuraea sp. ATCC 39727

    PubMed Central

    Binda, Elisa; Carrano, Lucia; Bibb, Mervyn; Marinelli, Flavia

    2014-01-01

    Glycopeptides and β-lactams inhibit bacterial peptidoglycan synthesis in Gram-positive bacteria; resistance to these antibiotics is studied intensively in enterococci and staphylococci because of their relevance to infectious disease. Much less is known about antibiotic resistance in glycopeptide-producing actinomycetes that are likely to represent the evolutionary source of resistance determinants found in bacterial pathogens. Nonomuraea sp. ATCC 39727, the producer of A40926 (the precursor for the semisynthetic dalbavancin), does not harbor the canonical vanHAX genes. Consequently, we investigated the role of the β-lactam-sensitive d,d-peptidase/d,d-carboxypeptidase encoded by vanYn, the only van-like gene found in the A40926 biosynthetic gene cluster, in conferring immunity to the antibiotic in Nonomuraea sp. ATCC 39727. Taking advantage of the tools developed recently to genetically manipulate this uncommon actinomycete, we varied vanYn gene dosage and expressed vanHatAatXat from the teicoplanin producer Actinoplanes teichomyceticus in Nonomuraea sp. ATCC 39727. Knocking out vanYn, complementing a vanYn mutant, or duplicating vanYn had no effect on growth but influenced antibiotic resistance and, in the cases of complementation and duplication, antibiotic production. Nonomuraea sp. ATCC 39727 was found to be resistant to penicillins, but its glycopeptide resistance was diminished in the presence of penicillin G, which inhibits VanYn activity. The heterologous expression of vanHatAatXat increased A40926 resistance in Nonomuraea sp. ATCC 39727 but did not increase antibiotic production, indicating that the level of antibiotic production is not directly determined by the level of resistance. The vanYn-based self-resistance in Nonomuraea sp. ATCC 39727 resembles the glycopeptide resistance mechanism described recently in mutants of Enterococcus faecium selected in vitro for high-level resistance to glycopeptides and penicillins. PMID:24957828

  13. Genome sequence and comparative genome analysis of Pseudomonas syringae pv. syringae type strain ATCC 19310.

    PubMed

    Park, Yong-Soon; Jeong, Haeyoung; Sim, Young Mi; Yi, Hwe-Su; Ryu, Choong-Min

    2014-04-01

    Pseudomonas syringae pv. syringae (Psy) is a major bacterial pathogen of many economically important plant species. Despite the severity of its impact, the genome sequence of the type strain has not been reported. Here, we present the draft genome sequence of Psy ATCC 19310. Comparative genomic analysis revealed that Psy ATCC 19310 is closely related to Psy B728a. However, only a few type III effectors, which are key virulence factors, are shared by the two strains, indicating the possibility of host-pathogen specificity and genome dynamics, even under the pathovar level. PMID:24444998

  14. Genome sequence and plasmid transformation of the model high-yield bacterial cellulose producer Gluconacetobacter hansenii ATCC 53582

    NASA Astrophysics Data System (ADS)

    Florea, Michael; Reeve, Benjamin; Abbott, James; Freemont, Paul S.; Ellis, Tom

    2016-03-01

    Bacterial cellulose is a strong, highly pure form of cellulose that is used in a range of applications in industry, consumer goods and medicine. Gluconacetobacter hansenii ATCC 53582 is one of the highest reported bacterial cellulose producing strains and has been used as a model organism in numerous studies of bacterial cellulose production and studies aiming to increased cellulose productivity. Here we present a high-quality draft genome sequence for G. hansenii ATCC 53582 and find that in addition to the previously described cellulose synthase operon, ATCC 53582 contains two additional cellulose synthase operons and several previously undescribed genes associated with cellulose production. In parallel, we also develop optimized protocols and identify plasmid backbones suitable for transformation of ATCC 53582, albeit with low efficiencies. Together, these results provide important information for further studies into cellulose synthesis and for future studies aiming to genetically engineer G. hansenii ATCC 53582 for increased cellulose productivity.

  15. Genome sequence and plasmid transformation of the model high-yield bacterial cellulose producer Gluconacetobacter hansenii ATCC 53582.

    PubMed

    Florea, Michael; Reeve, Benjamin; Abbott, James; Freemont, Paul S; Ellis, Tom

    2016-01-01

    Bacterial cellulose is a strong, highly pure form of cellulose that is used in a range of applications in industry, consumer goods and medicine. Gluconacetobacter hansenii ATCC 53582 is one of the highest reported bacterial cellulose producing strains and has been used as a model organism in numerous studies of bacterial cellulose production and studies aiming to increased cellulose productivity. Here we present a high-quality draft genome sequence for G. hansenii ATCC 53582 and find that in addition to the previously described cellulose synthase operon, ATCC 53582 contains two additional cellulose synthase operons and several previously undescribed genes associated with cellulose production. In parallel, we also develop optimized protocols and identify plasmid backbones suitable for transformation of ATCC 53582, albeit with low efficiencies. Together, these results provide important information for further studies into cellulose synthesis and for future studies aiming to genetically engineer G. hansenii ATCC 53582 for increased cellulose productivity. PMID:27010592

  16. Genome sequence and plasmid transformation of the model high-yield bacterial cellulose producer Gluconacetobacter hansenii ATCC 53582

    PubMed Central

    Florea, Michael; Reeve, Benjamin; Abbott, James; Freemont, Paul S.; Ellis, Tom

    2016-01-01

    Bacterial cellulose is a strong, highly pure form of cellulose that is used in a range of applications in industry, consumer goods and medicine. Gluconacetobacter hansenii ATCC 53582 is one of the highest reported bacterial cellulose producing strains and has been used as a model organism in numerous studies of bacterial cellulose production and studies aiming to increased cellulose productivity. Here we present a high-quality draft genome sequence for G. hansenii ATCC 53582 and find that in addition to the previously described cellulose synthase operon, ATCC 53582 contains two additional cellulose synthase operons and several previously undescribed genes associated with cellulose production. In parallel, we also develop optimized protocols and identify plasmid backbones suitable for transformation of ATCC 53582, albeit with low efficiencies. Together, these results provide important information for further studies into cellulose synthesis and for future studies aiming to genetically engineer G. hansenii ATCC 53582 for increased cellulose productivity. PMID:27010592

  17. Multigene disruption in undomesticated Bacillus subtilis ATCC 6051a using the CRISPR/Cas9 system.

    PubMed

    Zhang, Kang; Duan, Xuguo; Wu, Jing

    2016-01-01

    Bacillus subtilis ATCC 6051a is an undomesticated strain used in the industrial production of enzymes. Because it is poorly transformable, genetic manipulation in this strain requires a highly efficient genome editing method. In this study, a Streptococcus pyogenes CRISPR/Cas9 system consisting of an all-in-one knockout plasmid containing a target-specific guide RNA, cas9, and a homologous repair template was established for highly efficient gene disruption in B. subtilis ATCC 6051a. With an efficiency of 33% to 53%, this system was used to disrupt the srfC, spoIIAC, nprE, aprE and amyE genes of B. subtilis ATCC 6051a, which hamper its use in industrial fermentation. Compared with B. subtilis ATCC 6051a, the final mutant, BS5 (ΔsrfC, ΔspoIIAC, ΔnprE, ΔaprE, ΔamyE), produces much less foam during fermentation, displays greater resistant to spore formation, and secretes 2.5-fold more β-cyclodextrin glycosyltransferase into the fermentation medium. Thus, the CRISPR/Cas9 system proved to be a powerful tool for targeted genome editing in an industrially relevant, poorly transformable strain. PMID:27305971

  18. Genome sequence of the vertebrate gut symbiont Lactobacillus reuteri ATCC 53608.

    PubMed

    Heavens, Darren; Tailford, Louise E; Crossman, Lisa; Jeffers, Faye; Mackenzie, Donald A; Caccamo, Mario; Juge, Nathalie

    2011-08-01

    Lactobacillus reuteri, inhabiting the gastrointestinal tracts of a range of vertebrates, is a true symbiont with effects established as beneficial to the host. Here we describe the draft genome of L. reuteri ATCC 53608, isolated from a pig. The genome sequence provides important insights into the evolutionary changes underlying host specialization. PMID:21622738

  19. Multigene disruption in undomesticated Bacillus subtilis ATCC 6051a using the CRISPR/Cas9 system

    PubMed Central

    Zhang, Kang; Duan, Xuguo; Wu, Jing

    2016-01-01

    Bacillus subtilis ATCC 6051a is an undomesticated strain used in the industrial production of enzymes. Because it is poorly transformable, genetic manipulation in this strain requires a highly efficient genome editing method. In this study, a Streptococcus pyogenes CRISPR/Cas9 system consisting of an all-in-one knockout plasmid containing a target-specific guide RNA, cas9, and a homologous repair template was established for highly efficient gene disruption in B. subtilis ATCC 6051a. With an efficiency of 33% to 53%, this system was used to disrupt the srfC, spoIIAC, nprE, aprE and amyE genes of B. subtilis ATCC 6051a, which hamper its use in industrial fermentation. Compared with B. subtilis ATCC 6051a, the final mutant, BS5 (ΔsrfC, ΔspoIIAC, ΔnprE, ΔaprE, ΔamyE), produces much less foam during fermentation, displays greater resistant to spore formation, and secretes 2.5-fold more β-cyclodextrin glycosyltransferase into the fermentation medium. Thus, the CRISPR/Cas9 system proved to be a powerful tool for targeted genome editing in an industrially relevant, poorly transformable strain. PMID:27305971

  20. Interaction of Pseudomonas putida ATCC 12633 and Bacteriophage gh-1 in Berea Sandstone Rock.

    PubMed

    Chang, P L; Yen, T F

    1985-12-01

    Measurements of the passage of Pseudomonas putida ATCC 12633 and a phage-resistant mutant through Berea sandstone rock were made. When bacteriophage gh-1 was adsorbed within the rock matrix, a reduction in the passage of the susceptible but not the resistant cells through the rock was observed. PMID:16346956

  1. Interaction of Pseudomonas putida ATCC 12633 and Bacteriophage gh-1 in Berea Sandstone Rock

    PubMed Central

    Chang, Philip Lee; Yen, Teh Fu

    1985-01-01

    Measurements of the passage of Pseudomonas putida ATCC 12633 and a phage-resistant mutant through Berea sandstone rock were made. When bacteriophage gh-1 was adsorbed within the rock matrix, a reduction in the passage of the susceptible but not the resistant cells through the rock was observed. PMID:16346956

  2. Complete genome sequence of Streptococcus equi subsp. zooepidemicus strain ATCC 35246.

    PubMed

    Ma, Zhe; Geng, Jianing; Zhang, Hui; Yu, Haiying; Yi, Li; Lei, Meng; Lu, Cheng-ping; Fan, Hong-jie; Hu, Songnian

    2011-10-01

    Streptococcus equi subsp. zooepidemicus is an opportunistic pathogen. It has caused a very large economic loss in the swine industry of China and has become a threat to human health. We announce the complete genome sequence of S. equi subsp. zooepidemicus strain ATCC 35246, which provides opportunities to understand its pathogenesis mechanism and genetic basis. PMID:21914890

  3. Draft Genome Assembly of Ralstonia pickettii Type Strain K-288 (ATCC 27853)

    PubMed Central

    Daligault, H. E.; Davenport, K. W.; Minogue, T. D.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Coyne, S. R.; Gibbons, H. S.; Jaissle, J.; Lo, C.-C.; Meincke, L.; Munk, A. C.; Rosenzweig, C. N.

    2014-01-01

    We present the genome assembly of Ralstonia pickettii K-288 (ATCC 27511), consisting of 27 contigs placed into a single scaffold. This 4.76-Mbp genome has 64.0% G+C content and 4,425 coding sequences. Because this is the type strain, inclusion of its data set among other Ralstonia genomes should provide a historical genomic perspective. PMID:25258272

  4. Draft Genome Assembly of Ralstonia pickettii Type Strain K-288 (ATCC 27853).

    PubMed

    Daligault, H E; Davenport, K W; Minogue, T D; Broomall, S M; Bruce, D C; Chain, P S; Coyne, S R; Gibbons, H S; Jaissle, J; Lo, C-C; Meincke, L; Munk, A C; Rosenzweig, C N; Johnson, S L

    2014-01-01

    We present the genome assembly of Ralstonia pickettii K-288 (ATCC 27511), consisting of 27 contigs placed into a single scaffold. This 4.76-Mbp genome has 64.0% G+C content and 4,425 coding sequences. Because this is the type strain, inclusion of its data set among other Ralstonia genomes should provide a historical genomic perspective. PMID:25258272

  5. Genome Assembly of Shigella flexneri ATCC 12022, a Quality Control Reference Strain

    PubMed Central

    Daligault, H. E.; Davenport, K. W.; Minogue, T. D.; Bishop-Lilly, K. A.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Coyne, S. R.; Freitas, T.; Frey, K. G.; Gibbons, H. S.; Jaissle, J.; Lo, C.-C.; Meincke, L.; Munk, A. C.; Redden, C. L.; Rosenzweig, C. N.

    2014-01-01

    Shigella flexneri causes shigellosis, severe and potentially life-threatening diarrhea, and accounts for 18% of shigellosis cases in the United States. Here, we present the 4.51-Mbp genome assembly of S. flexneri ATCC 12022, a quality control and reference strain, in 10 scaffolds. PMID:25359907

  6. Complete Genome Assembly of Escherichia coli ATCC 25922, a Serotype O6 Reference Strain.

    PubMed

    Minogue, T D; Daligault, H A; Davenport, K W; Bishop-Lilly, K A; Broomall, S M; Bruce, D C; Chain, P S; Chertkov, O; Coyne, S R; Freitas, T; Frey, K G; Gibbons, H S; Jaissle, J; Redden, C L; Rosenzweig, C N; Xu, Y; Johnson, S L

    2014-01-01

    We present the complete genome assembly of Escherichia coli ATCC 25922 as submitted to NCBI under accession no. CP009072. This strain was originally isolated from a clinical sample in Seattle, Washington (1946), and is often used in quality control testing. The assembled genome is 5.20 Mb (50.4% G+C content) and includes two plasmids. PMID:25291776

  7. Complete Genome Assembly of Escherichia coli ATCC 25922, a Serotype O6 Reference Strain

    PubMed Central

    Minogue, T. D.; Daligault, H. A.; Davenport, K. W.; Bishop-Lilly, K. A.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Chertkov, O.; Coyne, S. R.; Freitas, T.; Frey, K. G.; Gibbons, H. S.; Jaissle, J.; Redden, C. L.; Rosenzweig, C. N.; Xu, Y.

    2014-01-01

    We present the complete genome assembly of Escherichia coli ATCC 25922 as submitted to NCBI under accession no. CP009072. This strain was originally isolated from a clinical sample in Seattle, Washington (1946), and is often used in quality control testing. The assembled genome is 5.20 Mb (50.4% G+C content) and includes two plasmids. PMID:25291776

  8. Genome Assembly of Shigella flexneri ATCC 12022, a Quality Control Reference Strain.

    PubMed

    Daligault, H E; Davenport, K W; Minogue, T D; Bishop-Lilly, K A; Broomall, S M; Bruce, D C; Chain, P S; Coyne, S R; Freitas, T; Frey, K G; Gibbons, H S; Jaissle, J; Lo, C-C; Meincke, L; Munk, A C; Redden, C L; Rosenzweig, C N; Johnson, S L

    2014-01-01

    Shigella flexneri causes shigellosis, severe and potentially life-threatening diarrhea, and accounts for 18% of shigellosis cases in the United States. Here, we present the 4.51-Mbp genome assembly of S. flexneri ATCC 12022, a quality control and reference strain, in 10 scaffolds. PMID:25359907

  9. Complete Genome Sequence of Stenotrophomonas maltophilia Type Strain 810-2 (ATCC 13637).

    PubMed

    Davenport, K W; Daligault, H E; Minogue, T D; Broomall, S M; Bruce, D C; Chain, P S; Coyne, S R; Gibbons, H S; Jaissle, J; Li, P-E; Rosenzweig, C N; Scholz, M B; Johnson, S L

    2014-01-01

    An emerging nosocomial pathogen, Stenotrophomonas maltophila has a high mortality rate in those it infects. Here, we present the complete genome sequence of Stenotrophomonas maltophilia 810-2 (ATCC 13637), the type strain of the species. The 5-Mb (66.1% G+C content) genome has been deposited in NCBI under accession number CP008838. PMID:25258273

  10. Complete Genome Sequence of Stenotrophomonas maltophilia Type Strain 810-2 (ATCC 13637)

    PubMed Central

    Davenport, K. W.; Daligault, H. E.; Minogue, T. D.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Coyne, S. R.; Gibbons, H. S.; Jaissle, J.; Li, P.-E.; Rosenzweig, C. N.; Scholz, M. B.

    2014-01-01

    An emerging nosocomial pathogen, Stenotrophomonas maltophila has a high mortality rate in those it infects. Here, we present the complete genome sequence of Stenotrophomonas maltophilia 810-2 (ATCC 13637), the type strain of the species. The 5-Mb (66.1% G+C content) genome has been deposited in NCBI under accession number CP008838. PMID:25258273

  11. Complete Genome Sequence of Thiostrepton-Producing Streptomyces laurentii ATCC 31255.

    PubMed

    Doi, Katsumi; Fujino, Yasuhiro; Nagayoshi, Yuko; Ohshima, Toshihisa; Ogata, Seiya

    2016-01-01

    Streptomyces laurentii ATCC 31255 produces thiostrepton, a thiopeptide class antibiotic. Here, we report the complete genome sequence for this strain, which contains a total of 8,032,664 bp, 7,452 predicted coding sequences, and a G+C content of 72.3%. PMID:27257211

  12. Draft Genome Sequence of the Probiotic Yeast Saccharomyces cerevisiae var. boulardii Strain ATCC MYA-796

    PubMed Central

    Marques, E. T. A.; Franco, G. R.

    2014-01-01

    Saccharomyces boulardii is the only yeast approved as a probiotic for human consumption. Here, we report the draft genome sequence of the strain ATCC MYA-796, derived from the French Ultra Levure probiotic drug. The genome has a size of 11.6 Mb with 5,305 putative open reading frames predicted. PMID:25523784

  13. Genome Sequence of meso-2,3-Butanediol-Producing Strain Serratia marcescens ATCC 14041

    PubMed Central

    Li, Lixiang; Wang, Yu; Li, Kun; Su, Fei; Ma, Cuiqing

    2014-01-01

    Serratia marcescens strain ATCC 14041 was found to be an efficient meso-2,3-butanediol (meso-2,3-BD) producer from glucose and sucrose. Here we present a 5.0-Mb assembly of its genome. We have annotated 4 coding sequences (CDSs) for meso-2,3-BD fermentation and 2 complete operons including 6 CDSs for sucrose utilization. PMID:24948764

  14. Complete Genome Sequence of Brachyspira hyodysenteriae Type Strain B-78 (ATCC 27164)

    PubMed Central

    Mirajkar, Nandita S.; Johnson, Timothy J.

    2016-01-01

    Reported herein is the complete genome sequence of the type strain B-78 (ATCC 27164) of Brachyspira hyodysenteriae, the etiological agent of swine dysentery. The 3.1-Mb genome consists of a 3.056-Mb chromosome and a 45-kb plasmid, with 2,617 protein-coding genes, 39 RNA genes, and 40 pseudogenes. PMID:27540064

  15. Complete Genome Sequence of the Bioluminescent Marine Bacterium Vibrio harveyi ATCC 33843 (392 [MAV]).

    PubMed

    Wang, Zheng; Hervey, W Judson; Kim, Seongwon; Lin, Baochuan; Vora, Gary J

    2015-01-01

    Vibrio harveyi is a Gram-negative marine γ-proteobacterium that is known to be a formidable pathogen of aquatic animals and is a model organism for the study of bacterial bioluminescence and quorum sensing. In this report, we describe the complete genome sequence of the most studied strain of this species: V. harveyi ATCC 33843 (392 [MAV]). PMID:25635019

  16. Complete Genome Sequence of the Bioluminescent Marine Bacterium Vibrio harveyi ATCC 33843 (392 [MAV])

    PubMed Central

    Wang, Zheng; Hervey, W. Judson; Kim, Seongwon; Lin, Baochuan

    2015-01-01

    Vibrio harveyi is a Gram-negative marine γ-proteobacterium that is known to be a formidable pathogen of aquatic animals and is a model organism for the study of bacterial bioluminescence and quorum sensing. In this report, we describe the complete genome sequence of the most studied strain of this species: V. harveyi ATCC 33843 (392 [MAV]). PMID:25635019

  17. Draft Genome Sequence of a Metronidazole-Resistant Derivative of Gardnerella vaginalis Strain ATCC 14019

    PubMed Central

    Schuyler, Jessica A.; Mordechai, Eli; Adelson, Martin E.; Gygax, Scott E.

    2015-01-01

    We report the genome sequence of a metronidazole-resistant derivative of Gardnerella vaginalis ATCC 14019. This strain was obtained after serial selection to increase the MIC from 4 to ≥500 µg/ml. Two coding changes, in genes encoding a response regulator and an NAD+ synthetase, arose during selection. PMID:26564054

  18. Genome Sequence of Streptomyces viridosporus Strain T7A ATCC 39115, a Lignin-Degrading Actinomycete

    SciTech Connect

    Davis, Jennifer R.; Goodwin, Lynne A.; Teshima, Hazuki; Detter, J. Chris; Tapia, Roxanne; Han, Cliff; Huntemann, Marcel; Wei, Chia-Lin; Han, James; Chen, Amy; Kyrpides, Nikos C; Mavromatis, K; Szeto, Ernest; Markowitz, Victor; Ivanova, N; Mikhailova, Natalia; Ovchinnikova, Galina; Pagani, Ioanna; Pati, Amrita; Woyke, Tanja; Pitluck, Sam; Peters, Lin; Nolan, Matt; Land, Miriam L; Sello, Jason K.

    2013-01-01

    We announce the availability of the genome sequence of Streptomyces viridosporus strain T7A ATCC 39115, a plant biomass- degrading actinomycete. This bacterium is of special interest because of its capacity to degrade lignin, an underutilized compo- nent of plants in the context of bioenergy. It has a full complement of genes for plant biomass catabolism.

  19. Effect of Calcium in Assay Medium on D Value of Bacillus stearothermophilus ATCC 7953 Spores

    PubMed Central

    Sasaki, Koichi; Shintani, Hideharu; Itoh, Junpei; Kamogawa, Takuji; Kajihara, Yousei

    2000-01-01

    The D value of commercial biological indicator spore strips using Bacillus stearothermophilus ATCC 7953 was increased by higher calcium concentrations in assay media. The calcium concentration in assay media varied among the manufacturers. The calcium concentration in assay media is an important factor to consider to minimize the variation of D value. PMID:11097939

  20. Draft Genome Sequence of the Probiotic Strain Lactobacillus acidophilus ATCC 4356

    PubMed Central

    Palomino, Maria Mercedes; Allievi, Mariana C.; Fina Martin, Joaquina; Waehner, Pablo M.; Prado Acosta, Mariano; Sanchez Rivas, Carmen

    2015-01-01

    We present the 1,956,699-bp draft genome sequence of Lactobacillus acidophilus strain ATCC 4356. Comparative genomic analysis revealed 99.96% similarity with L. acidophilus NCFM NC_006814.3 and 99.97% with La-14 NC_021181.2 genomes. PMID:25593259

  1. Effect of calcium in assay medium on D value of Bacillus stearothermophilus ATCC 7953 spores.

    PubMed

    Sasaki, K; Shintani, H; Itoh, J; Kamogawa, T; Kajihara, Y

    2000-12-01

    The D value of commercial biological indicator spore strips using Bacillus stearothermophilus ATCC 7953 was increased by higher calcium concentrations in assay media. The calcium concentration in assay media varied among the manufacturers. The calcium concentration in assay media is an important factor to consider to minimize the variation of D value. PMID:11097939

  2. Antimicrobial mechanism of flavonoids against Escherichia coli ATCC 25922 by model membrane study

    NASA Astrophysics Data System (ADS)

    He, Mengying; Wu, Ting; Pan, Siyi; Xu, Xiaoyun

    2014-06-01

    Antimicrobial mechanism of four flavonoids (kaempferol, hesperitin, (+)-catechin hydrate, biochanin A) against Escherichia coli ATCC 25922 was investigated through cell membranes and a liposome model. The release of bacterial protein and images from transmission electron microscopy demonstrated damage to the E. coli ATCC 25922 membrane. A liposome model with dipalmitoylphosphatidylethanolamine (DPPE) (0.6 molar ratio) and dipalmitoylphosphatidylglycerol (DPPG) (0.4 molar ratio), representative of the phospholipid membrane of E. coli ATCC 25922, was used to specify the mode of action of four selected flavonoids through Raman spectroscopy and differential scanning calorimetry. It is suggested that for flavonoids, to be effective antimicrobials, interaction with the polar head-group of the model membrane followed by penetration into the hydrophobic regions must occur. The antimicrobial efficacies of the flavonoids were consistent with liposome interaction activities, kaempferol > hesperitin > (+)-catechin hydrate > biochanin A. This study provides a liposome model capable of mimicking the cell membrane of E. coli ATCC 25922. The findings are important in understanding the antibacterial mechanism on cell membranes.

  3. Draft Genome Sequence of the Probiotic Strain Lactobacillus acidophilus ATCC 4356.

    PubMed

    Palomino, Maria Mercedes; Allievi, Mariana C; Fina Martin, Joaquina; Waehner, Pablo M; Prado Acosta, Mariano; Sanchez Rivas, Carmen; Ruzal, Sandra M

    2015-01-01

    We present the 1,956,699-bp draft genome sequence of Lactobacillus acidophilus strain ATCC 4356. Comparative genomic analysis revealed 99.96% similarity with L. acidophilus NCFM NC_006814.3 and 99.97% with La-14 NC_021181.2 genomes. PMID:25593259

  4. Draft Genome Sequence of Streptomyces fradiae ATCC 19609, a Strain Highly Sensitive to Antibiotics.

    PubMed

    Bekker, Olga B; Klimina, Ksenia M; Vatlin, Aleksey A; Zakharevich, Natalia V; Kasianov, Artem S; Danilenko, Valery N

    2014-01-01

    We report here a sequence of the genome of the Streptomyces fradiae ATCC 19609 strain, initially isolated from the soil, which produces tylosin. S. fradiae is highly sensitive to different classes of antibiotics, compared to the sensitivities of other bacteria. We have identified 9 groups of genes directly or indirectly involved in the resistome formation. PMID:25477406

  5. Complete Genome Sequence and Methylome Analysis of Bacillus globigii ATCC 49760.

    PubMed

    Morgan, Richard D

    2016-01-01

    Bacillus subtilis (Ehrenburg) Cohn ATCC 49760, deposited as Bacillus globigii, is the source strain for the restriction enzymes BglI and BglII. Its complete sequence and full methylome were determined using single-molecule real-time (SMRT) sequencing. PMID:27231364

  6. Complete Genome Sequence and Methylome Analysis of Bacillus globigii ATCC 49760

    PubMed Central

    2016-01-01

    Bacillus subtilis (Ehrenburg) Cohn ATCC 49760, deposited as Bacillus globigii, is the source strain for the restriction enzymes BglI and BglII. Its complete sequence and full methylome were determined using single-molecule real-time (SMRT) sequencing. PMID:27231364

  7. Complete Genome Sequence of Thiostrepton-Producing Streptomyces laurentii ATCC 31255

    PubMed Central

    Fujino, Yasuhiro; Nagayoshi, Yuko; Ohshima, Toshihisa; Ogata, Seiya

    2016-01-01

    Streptomyces laurentii ATCC 31255 produces thiostrepton, a thiopeptide class antibiotic. Here, we report the complete genome sequence for this strain, which contains a total of 8,032,664 bp, 7,452 predicted coding sequences, and a G+C content of 72.3%. PMID:27257211

  8. Complete Genome Sequence of Brachyspira hyodysenteriae Type Strain B-78 (ATCC 27164).

    PubMed

    Mirajkar, Nandita S; Johnson, Timothy J; Gebhart, Connie J

    2016-01-01

    Reported herein is the complete genome sequence of the type strain B-78 (ATCC 27164) of Brachyspira hyodysenteriae, the etiological agent of swine dysentery. The 3.1-Mb genome consists of a 3.056-Mb chromosome and a 45-kb plasmid, with 2,617 protein-coding genes, 39 RNA genes, and 40 pseudogenes. PMID:27540064

  9. Optimization and standardization of an enzyme-linked immunosorbent assay protocol for serodiagnosis of Actinobacillus pleuropneumoniae serotype 5.

    PubMed Central

    Trottier, Y L; Wright, P F; Larivière, S

    1992-01-01

    An indirect enzyme-linked immunosorbent assay protocol has been optimized with special emphasis given to assay standardization and quality control. Technical aspects such as choice of a microplate, antigen immobilization, buffer composition, optimal screening dilution of sera, and kinetics of the enzymatic reaction were studied and evaluated in order to design a standard protocol offering maximal analytical sensitivity and specificity, as well as to obtain minimal within- and between-plate variability. Among the 27 plates tested, the Nunc 475-094 and 269-620 immunoplates were found to be the best in terms of high positive-to-negative ratio and low variability. No significant differences in antigen immobilization were found by using buffers of various compositions or pHs; however, the presence of magnesium ions (Mg2+; 0.02 M) resulted in a twofold increase in nonspecific background. An optimal screening dilution of sera was established at 1:200. A 1-h incubation period for test serum was found to be optimal. Maximum enzymatic activity for peroxidase was obtained by adjusting both substrate (H2O2) and hydrogen donor [2,2' -azinobis(3-ethylbenz-thiazoline sulfonic acid)] concentrations to 4 and 1 mM, respectively. To control between-plate variability, a timing protocol was adopted. Within-plate variability was also controlled by using a sample placement configuration pattern. Sliding scales were determined by repeated testing of a cross section of samples to set acceptance limits for both within- and between-plate variability. These limits were used in a quality control program to monitor assay performance. The results obtained suggest that this standardized protocol might be useful in the serodiagnosis of Actinobacillus pleuropneumoniae serotype 5. PMID:1734068

  10. Purification and characterization of a protease from Actinobacillus pleuropneumoniae serotype 1, an antigen common to all the serotypes.

    PubMed Central

    Negrete-Abascal, E; Tenorio, V R; Guerrero, A L; García, R M; Reyes, M E; de la Garza, M

    1998-01-01

    A high molecular-mass proteolytic enzyme of Actinobacillus pleuropneumoniae serotype 1, was purified from culture supernatants (CSN) by using DEAE-cellulose and sepharose-4B-gelatin chromatography. In 10% SDS-polyacrylamide gels copolymerized with porcine gelatin, the protease showed a single band of activity of > 200 kDa. However, minor molecular-mass proteolytic bands were observed when the protease was electrophoresed in the presence of either 5% beta-mercaptoethanol, 50 mM dithiothreitol, or 0.25 M urea. Furthermore, when the > 200-kDa purified protein was passed through a sucrose gradient, several bands with proteolytic activity were found: 62, 90, 190, and 540 kDa. The proteolytic activity was increased in the presence of calcium or zinc and was not affected after being heated at 90 degrees C for 5 min. Proteolytic activities were also observed in CSN from all A. pleuropneumoniae serotypes and biotypes. The purified protease hydrolyzed porcine IgA and IgG in vitro. In addition, by immunoblot the protease was recognized by serum of naturally infected pigs with serotypes 1 and 5, and by serum of pigs experimentally infected with serotypes 1, 2, 8, or 9. Serum of a pig vaccinated with CSN of a serotype 3 strain also recognized the protease, but not sera of pigs vaccinated with a bacterin (serotype 1). Proteins from CSN of all the serotypes, which were precipitated with 70% (NH4)2SO4, were recognized by a polyclonal antibody raised against the purified protease. Taken together these results indicate that an antigenic protease is produced in vivo by all the serotypes of A. pleuropneumoniae. The results indicate that proteases could have a role in the disease and in the immune response of pigs infected with A. pleuropneumoniae. Images Figure 2A. Figure 2B. Figure 3. Figure 4. Figure 5A. Figure 5B. Figure 6A. Figure 6B. PMID:9684047

  11. The ClpP Protease Is Required for the Stress Tolerance and Biofilm Formation in Actinobacillus pleuropneumoniae

    PubMed Central

    Xie, Fang; Zhang, Yanhe; Li, Gang; Zhou, Long; Liu, Siguo; Wang, Chunlai

    2013-01-01

    In the respiratory tract and lung tissue, a balanced physiological response is essential for Actinobacillus pleuropneumoniae to survive various types of challenges. ClpP, the catalytic core of the Clp proteolytic complex, is involved in various stresses response and regulation of biofilm formation in many pathogenic bacteria. To investigate the role of ClpP in the virulence of A. pleuropneumoniae, the clpP gene was deleted by homologous recombination, resulting in the mutant strain S8ΔclpP. The reduced growth of S8ΔclpP mutant at high temperatures and under several other stress conditions suggests that the ClpP protein is required for the stress tolerance of A. pleuropneumoniae. Interestingly, we observed that the S8ΔclpP mutant exhibited an increased ability to take up iron in vitro compared to the wild-type strain. We also found that the cells without ClpP displayed rough and irregular surfaces and increased cell volume relative to the wild-type strain using scanning electron microscopy (SEM). Confocal laser scanning microscopy (CLSM) revealed that the S8ΔclpP mutant showed decreased biofilm formation compared to the wild-type strain. We examined the transcriptional profiles of the wild type S8 and the S8ΔclpP mutant strains of A. pleuropneumoniae using RNA sequencing. Our analysis revealed that the expression of 16 genes was changed by the deletion of the clpP gene. The data presented in this study illustrate the important role of ClpP protease in the stress response, iron acquisition, cell morphology and biofilm formation related to A. pleuropneumoniae and further suggest a putative role of ClpP protease in virulence regulation. PMID:23326465

  12. [Construction and immunogenicity of an attenuated mutant of Actinobacillus pleuropneumoniae by insertional inactivation of apxIC].

    PubMed

    Xu, Fu-Zhou; Shi, Ai-Hua; Chen, Xiao-Ling; Yang, Bing; Wang, Jin-Luo

    2007-10-01

    Actinobacillus pleuropneumoniae is the aetiological agent of porcine pleuropneumonia. Apx toxin, an exotoxin secreted by A. pleuropneumoniae, is one of the most important virulence factors. To construct an avirulent mutant strain by inactivation of ApxI toxin, the apxIC gene of A. pleuropneumoniae serovar 10 was inactivated by inserting a chloramphenicol resistance gene cassette into the downstream XhoI site of the apxIC gene for constructing the transfer plasmid. The transfer plasmid was introduced into the electrocompetent A. pleuropneumoniae serovar 10 for homologous recombination by electroporation. The mutant strain was obtained and identified by PCR and Southern blotting. The mutant strain was phenotypically identical to the parent strain except that it showed no haemolytic activity. The mutant strain was also able to secret the same ApxI toxin as the parent strain. In the intra-peritoneal mouse model, the virulence of the mutant strain decreased at least 100 fold compared with the parent strain. The mutant was evaluated as a potential vaccine using a vaccination-challenge trial in which pigs were given two intra-nasal doses of the mutant with 14 days' interval and then challenged 14 days after the last vaccination with A. pleuropneumoniae serovar 1 and serovar 10 reference strains respectively. The death number and lung lesion score in the vaccinated pigs given the serovar 1 challenge were obviously lower than those in the unvaccinated pigs. And the lower lung lesion score was also observed in the vaccinated pigs challenged with serovar 10. And the positive numbers of A. pleuropneumoniae re-isolation and PCR detection showed the same consistency. The vaccination-challenge trial suggested that the mutant strain could offer partial cross-protection as a live attenuated vaccine against A . pleuropneumoniae infection. PMID:18062275

  13. Transcriptional Portrait of Actinobacillus pleuropneumoniae during Acute Disease - Potential Strategies for Survival and Persistence in the Host

    PubMed Central

    Klitgaard, Kirstine; Friis, Carsten; Jensen, Tim K.; Angen, Øystein; Boye, Mette

    2012-01-01

    Background Gene expression profiles of bacteria in their natural hosts can provide novel insight into the host-pathogen interactions and molecular determinants of bacterial infections. In the present study, the transcriptional profile of the porcine lung pathogen Actinobacillus pleuropneumoniae was monitored during the acute phase of infection in its natural host. Methodology/Principal Findings Bacterial expression profiles of A. pleuropneumoniae isolated from lung lesions of 25 infected pigs were compared in samples taken 6, 12, 24 and 48 hours post experimental challenge. Within 6 hours, focal, fibrino hemorrhagic lesions could be observed in the pig lungs, indicating that A. pleuropneumoniae had managed to establish itself successfully in the host. We identified 237 differentially regulated genes likely to encode functions required by the bacteria for colonization and survival in the host. This group was dominated by genes involved in various aspects of energy metabolism, especially anaerobic respiration and carbohydrate metabolism. Remodeling of the bacterial envelope and modifications of posttranslational processing of proteins also appeared to be of importance during early infection. The results suggested that A. pleuropneumoniae is using various strategies to increase its fitness, such as applying Na+ pumps as an alternative way of gaining energy. Furthermore, the transcriptional data provided potential clues as to how A. pleuropneumoniae is able to circumvent host immune factors and survive within the hostile environment of host macrophages. This persistence within macrophages may be related to urease activity, mobilization of various stress responses and active evasion of the host defenses by cell surface sialylation. Conclusions/Significance The data presented here highlight the importance of metabolic adjustments to host conditions as virulence factors of infecting microorganisms and help to provide insight into the mechanisms behind the efficient

  14. Isolation and characterization of flagellar filaments from Bacillus cereus ATCC 14579.

    PubMed

    Tagawa, Yuichi

    2014-12-01

    Isolated flagellar filaments from the type strain of Bacillus cereus, ATCC 14579, were shown to consist of 34, 32 and 31 kDa proteins in similar proportions as judged by band intensities on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The N-terminal amino acid sequences of these three proteins of strain ATCC 14579 were identical with the deduced sequences of three flagellin genes BC1657, BC1658 and BC1659 in the whole genome sequence. Strain ATCC 14579 was classified into serotype T2 by a flagellar serotyping scheme for B. cereus strains that are untypeable into known flagellar serotypes H1 to H23. Flagellar filaments from a reference strain of serotype T2 contained two protein bands at 34 and 32 kDa, but a single protein band at 39 kDa was detected in flagellar filaments of a reference strain of serotype H1. Two murine monoclonal antibodies, 1A5 and 2A5, which recognize both the 34 and 32 kDa flagellins and a single flagellin of 32 kDa, respectively, were specifically reactive with B. cereus strains ATCC 14579 and serotype T2 in whole-cell ELISA and bacterial motility inhibition tests. In immunoelectron microscopy with monoclonal antibodies 1A5 and 2A5, colloidal gold spheres were shown to localize almost evenly over the entire part of flagellar filaments. Since strain ATCC 14579, and presumably strain serotype T2, are unusual among B. cereus strains in possessing multiple genes that encode flagellin subunits, a possible unique mechanism may contribute to assembly of multiple flagellin subunits into the filament over its entire length. PMID:25227778

  15. Growth of Lactobacillus paracasei ATCC 334 in a cheese model system: a biochemical approach.

    PubMed

    Budinich, M F; Perez-Díaz, I; Cai, H; Rankin, S A; Broadbent, J R; Steele, J L

    2011-11-01

    Growth of Lactobacillus paracasei ATCC 334, in a cheese-ripening model system based upon a medium prepared from ripening Cheddar cheese extract (CCE) was evaluated. Lactobacillus paracasei ATCC 334 grows in CCE made from cheese ripened for 2 (2mCCE), 6 (6mCCE), and 8 (8mCCE) mo, to final cell densities of 5.9×10(8), 1.2×10(8), and 2.1×10(7)cfu/mL, respectively. Biochemical analysis and mass balance equations were used to determine substrate consumption patterns and products formed in 2mCCE. The products formed included formate, acetate, and D-lactate. These data allowed us to identify the pathways likely used and to initiate metabolic flux analysis. The production of volatiles during growth of Lb. paracasei ATCC 334 in 8mCCE was monitored to evaluate the metabolic pathways utilized by Lb. paracasei during the later stages of ripening Cheddar cheese. The 2 volatiles detected at high levels were ethanol and acetate. The remaining detected volatiles are present in significantly lower amounts and likely result from amino acid, pyruvate, and acetyl-coenzyme A metabolism. Carbon balance of galactose, lactose, citrate, and phosphoserine/phosphoserine-containing peptides in terms of D-lactate, acetate, and formate are in agreement with the amounts of substrates observed in 2mCCE; however, this was not the case for 6mCCE and 8mCCE, suggesting that additional energy sources are utilized during growth of Lb. paracasei ATCC 334 in these CCE. This study provides valuable information on the biochemistry and physiology of Lb. paracasei ATCC 334 in ripening cheese. PMID:22032349

  16. Processing of cellulosic material by a cellulase-containing cell-free fermentate produced from cellulase-producing bacteria, ATCC 55702

    DOEpatents

    Dees, H. Craig

    1998-01-01

    Bacteria which produce large amounts of a cellulase-containing cell-free fermentate, have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase degrading bacterium ATCC 55702, which was identified through replicate plating. ATCC 55702 has improved characteristics and qualities for the degradation of cellulosic materials.

  17. Processing of cellulosic material by a cellulase-containing cell-free fermentate produced from cellulase-producing bacteria, ATCC 55702

    DOEpatents

    Dees, H.C.

    1998-08-04

    Bacteria which produce large amounts of a cellulase-containing cell-free fermentate, have been identified. The original bacterium (ATCC 55703) was genetically altered using nitrosoguanidine (MNNG) treatment to produce the enhanced cellulase degrading bacterium ATCC 55702, which was identified through replicate plating. ATCC 55702 has improved characteristics and qualities for the degradation of cellulosic materials. 5 figs.

  18. Lactobacillus acidophilus ATCC 4356 prevents atherosclerosis via inhibition of intestinal cholesterol absorption in apolipoprotein E-knockout mice.

    PubMed

    Huang, Ying; Wang, Jinfeng; Quan, Guihua; Wang, Xiaojun; Yang, Longfei; Zhong, Lili

    2014-12-01

    The objective of this study was to investigate the effect of Lactobacillus acidophilus ATCC 4356 on the development of atherosclerosis in apolipoprotein E-knockout (ApoE(-/-)) mice. Eight-week-old ApoE(-/-) mice were fed a Western diet with or without L. acidophilus ATCC 4356 daily for 16 weeks. L. acidophilus ATCC 4356 protected ApoE(-/-) mice from atherosclerosis by reducing their plasma cholesterol levels from 923 ± 44 to 581 ± 18 mg/dl, likely via a marked decrease in cholesterol absorption caused by modulation of Niemann-Pick C1-like 1 (NPC1L1). In addition, suppression of cholesterol absorption induced reverse cholesterol transport (RCT) in macrophages through the peroxisome proliferator-activated receptor/liver X receptor (PPAR/LXR) pathway. Fecal lactobacillus and bifidobacterium counts were significantly (P < 0.05) higher in the L. acidophilus ATCC 4356 treatment groups than in the control groups. Furthermore, L. acidophilus ATCC 4356 was detected in the rat small intestine, colon, and feces during the feeding trial. The bacterial levels remained high even after the administration of lactic acid bacteria had been stopped for 2 weeks. These results suggest that administration of L. acidophilus ATCC 4356 can protect against atherosclerosis through the inhibition of intestinal cholesterol absorption. Therefore, L. acidophilus ATCC 4356 may be a potential therapeutic material for preventing the progression of atherosclerosis. PMID:25261526

  19. Lactobacillus acidophilus ATCC 4356 Prevents Atherosclerosis via Inhibition of Intestinal Cholesterol Absorption in Apolipoprotein E-Knockout Mice

    PubMed Central

    Wang, Jinfeng; Quan, Guihua; Wang, Xiaojun; Yang, Longfei; Zhong, Lili

    2014-01-01

    The objective of this study was to investigate the effect of Lactobacillus acidophilus ATCC 4356 on the development of atherosclerosis in apolipoprotein E-knockout (ApoE−/−) mice. Eight-week-old ApoE−/− mice were fed a Western diet with or without L. acidophilus ATCC 4356 daily for 16 weeks. L. acidophilus ATCC 4356 protected ApoE−/− mice from atherosclerosis by reducing their plasma cholesterol levels from 923 ± 44 to 581 ± 18 mg/dl, likely via a marked decrease in cholesterol absorption caused by modulation of Niemann-Pick C1-like 1 (NPC1L1). In addition, suppression of cholesterol absorption induced reverse cholesterol transport (RCT) in macrophages through the peroxisome proliferator-activated receptor/liver X receptor (PPAR/LXR) pathway. Fecal lactobacillus and bifidobacterium counts were significantly (P < 0.05) higher in the L. acidophilus ATCC 4356 treatment groups than in the control groups. Furthermore, L. acidophilus ATCC 4356 was detected in the rat small intestine, colon, and feces during the feeding trial. The bacterial levels remained high even after the administration of lactic acid bacteria had been stopped for 2 weeks. These results suggest that administration of L. acidophilus ATCC 4356 can protect against atherosclerosis through the inhibition of intestinal cholesterol absorption. Therefore, L. acidophilus ATCC 4356 may be a potential therapeutic material for preventing the progression of atherosclerosis. PMID:25261526

  20. Stability of free and encapsulated Lactobacillus acidophilus ATCC 4356 in yogurt and in an artificial human gastric digestion system.

    PubMed

    Ortakci, F; Sert, S

    2012-12-01

    The objective of this study was to determine the effect of encapsulation on survival of probiotic Lactobacillus acidophilus ATCC 4356 (ATCC 4356) in yogurt and during artificial gastric digestion. Strain ATCC 4356 was added to yogurt either encapsulated in calcium alginate or in free form (unencapsulated) at levels of 8.26 and 9.47 log cfu/g, respectively, and the influence of alginate capsules (1.5 to 2.5mm) on the sensorial characteristics of yogurts was investigated. The ATCC 4356 strain was introduced into an artificial gastric solution consisting of 0.08 N HCl (pH 1.5) containing 0.2% NaCl or into artificial bile juice consisting of 1.2% bile salts in de Man, Rogosa, and Sharpe broth to determine the stability of the probiotic bacteria. When incubated for 2h in artificial gastric juice, the free ATCC 4356 did not survive (reduction of >7 log cfu/g). We observed, however, greater survival of encapsulated ATCC 4356, with a reduction of only 3 log cfu/g. Incubation in artificial bile juice (6 h) did not significantly affect the viability of free or encapsulated ATCC 4356. Moreover, statistically significant reductions (~1 log cfu/g) of both free and encapsulated ATCC 4356 were observed during 4-wk refrigerated storage of yogurts. The addition of probiotic cultures in free or alginate-encapsulated form did not significantly affect appearance/color or flavor/odor of the yogurts. However, significant deficiencies were found in body/texture of yogurts containing encapsulated ATCC 4356. We concluded that incorporation of free and encapsulated probiotic bacteria did not substantially change the overall sensory properties of yogurts, and encapsulation in alginate using the extrusion method greatly enhanced the survival of probiotic bacteria against an artificial human gastric digestive system. PMID:23021757

  1. Significant differences between Lactobacillus casei subsp. casei ATCC 393T and a commonly used plasmid-cured derivative revealed by a polyphasic study.

    PubMed

    Acedo-Félix, Evelia; Pérez-Martínez, Gaspar

    2003-01-01

    Many studies on Lactobacillus casei subsp. casei (L. casei) have been carried out using strain ATCC 393 (pLZ15-). Four strains of L. casei ATCC 393T and three of ATCC 393 (pLZ15-) were compared using phenotypic methods and many of the available genotyping techniques. These tests showed that strains of ATCC 393T obtained from independent public type-culture collections were significantly different from the plasmid-free (pLZ15-) strains of ATCC 393T. These findings were confirmed by sequencing the first 580 nt (domain I) of the 16S and 23S rDNAs of the strains. Complete sequencing of the 16S rDNA of one representative strain from each group revealed that strain ATCC 393T from culture collections was 99% similar to Lactobacillus zeae ATCC 15820T and that the strain so far considered as L. casei ATCC 393 (pLZ15-) was, in turn, 100% similar to L. casei ATCC 334 and Lactobacillus paracasei subsp. paracasei ATCC 4022. All data obtained in this work indicate that the ancestral strain of ATCC 393 (pLZ15-) might never have been the strain that is now available from culture collections. PMID:12656154

  2. Conservation and antigenic cross-reactivity of the transferrin-binding proteins of Haemophilus influenzae, Actinobacillus pleuropneumoniae and Neisseria meningitidis.

    PubMed

    Holland, J; Parsons, T R; Hasan, A A; Cook, S M; Stevenson, P; Griffiths, E; Williams, P

    1996-12-01

    Haemophilus influenzae acquires iron from the iron-transporting glycoprotein transferrin via a receptor-mediated process. This involves two outer-membrane transferrin-binding proteins (Tbps) termed Tbp1 and Tbp2 which show considerable preference for the human form of transferrin. Since the Tbps are attracting considerable attention as potential vaccine components, we used transferrin affinity chromatography to examine their conservation amongst 28 H. influenzae type b strains belonging to different outer-membrane-protein subtypes as well as six non-typable strains. Whole cells of all type b and non-typable strains examined bound human transferrin; whilst most strains possessed a Tbp1 of approximately 105 kDa, the molecular mass of Tbp2 varied from 79 to 94 kDa. Antisera raised against affinity-purified native H. influenzae Tbp1/Tbp2 receptor complex cross-reacted on Western blots with the respective Tbps of all the Haemophilus strains examined. When used to probe Neisseria meningitidis Tbps, sera from each of four mice immunized with the Haemophilus Tbp1/2 complex recognized the 68 kDa Tbp2 of N. meningitidis strain B16B6 but not the 78 kDa Tbp2 of N. meningitidis strain 70942. Serum from one mouse also reacted weakly with Tbp1 of strain B16B6. Apart from a weak reaction with the Tbp2 of a serotype 5 strain, this mouse antiserum failed to recognize the Tbps of the porcine pathogen A. pleuropneumoniae. However, a monospecific polyclonal antiserum raised against the denatured Tbp2 of Neisseria meningitidis B16B6 recognized the Tbps of all Haemophilus and Actinobacillus strains examined. Since H. influenzae forms part of the natural flora of the upper respiratory tract, human sera were screened for the presence of antibodies to the Tbps. Sera from healthy adults contained antibodies which recognized both Tbp1 and Tbp2 from H. influenzae but not N. meningitidis. Convalescent sera from meningococcal meningitis patients contained antibodies which, on Western blots

  3. malT knockout mutation invokes a stringent type gene-expression profile in Actinobacillus pleuropneumoniae in bronchoalveolar fluid

    PubMed Central

    2009-01-01

    Background Actinobacillus pleuropneumoniae causes contagious pleuropneumonia, an economically important disease of commercially reared pigs throughout the world. To cause this disease, A. pleuropneumoniae must rapidly overcome porcine pulmonary innate immune defenses. Since bronchoalveolar fluid (BALF) contains many of the innate immune and other components found in the lungs, we examined the gene expression of a virulent serovar 1 strain of A. pleuropneumoniae after exposure to concentrated BALF for 30 min. Results In reverse transcription PCR differential display (RT-PCR DD) experiments, A. pleuropneumoniae CM5 exposed to BALF up-regulated, among other genes, a gene predicted to encode LamB, an outer-membrane transport protein of the maltose regulon. To determine the role of the lamB and other genes of the maltose regulon in the pathogenesis of A. pleuropneumoniae, knockout mutations were created in the lamB and malT genes, the latter being the positive transcriptional regulator of the maltose regulon. Relative to the lamB mutant and the wild type, the malT mutant had a significant (P < 0.05) decrease in growth rate and an increased sensitivity to fresh porcine serum and high concentrations (more than 0.5 M) of sodium chloride. In DNA microarray experiments, the BALF-exposed malT mutant exhibited a gene-expression profile resembling that of a stringent type gene-expression profile seen in bacteria facing amino acid or carbon starvation. Genes encoding proteins for protein synthesis, energy metabolism, and DNA replication were down-regulated, while genes involved in stringent response (e.g., relA), amino acid and nucleotide biosynthesis, biofilm formation, DNA transformation, and stress response were up-regulated. Conclusion These results suggest that MalT may be involved in protection against some stressors and in the transport of one or more essential nutrients in BALF. Moreover, if MalT is directly or indirectly linked to the stringent response, an important

  4. Actinobacillus pleuropneumoniae genes expression in biofilms cultured under static conditions and in a drip-flow apparatus

    PubMed Central

    2013-01-01

    Background Actinobacillus pleuropneumoniae is the Gram-negative bacterium responsible for porcine pleuropneumonia. This respiratory infection is highly contagious and characterized by high morbidity and mortality. The objectives of our study were to study the transcriptome of A. pleuropneumoniae biofilms at different stages and to develop a protocol to grow an A. pleuropneumoniae biofilm in a drip-flow apparatus. This biofilm reactor is a system with an air-liquid interface modeling lung-like environment. Bacteria attached to a surface (biofilm) and free floating bacteria (plankton) were harvested for RNA isolation. Labelled cDNA was hybridized to a microarray to compare the expression profiles of planktonic cells and biofilm cells. Results It was observed that 47 genes were differentially expressed (22 up, 25 down) in a 4 h-static growing/maturing biofilm and 117 genes were differentially expressed (49 up, 68 down) in a 6h-static dispersing biofilm. The transcriptomes of a 4 h biofilm and a 6 h biofilm were also compared and 456 genes (235 up, 221 down) were identified as differently expressed. Among the genes identified in the 4 h vs 6h biofilm experiment, several regulators of stress response were down-regulated and energy metabolism associated genes were up-regulated. Biofilm bacteria cultured using the drip-flow apparatus differentially expressed 161 genes (68 up, 93 down) compared to the effluent bacteria. Cross-referencing of differentially transcribed genes in the different assays revealed that drip-flow biofilms shared few differentially expressed genes with static biofilms (4 h or 6 h) but shared several differentially expressed genes with natural or experimental infections in pigs. Conclusion The formation of a static biofilm by A. pleuropneumoniae strain S4074 is a rapid process and transcriptional analysis indicated that dispersal observed at 6 h is driven by nutritional stresses. Furthermore, A. pleuropneumoniae can form a biofilm under low

  5. Effect of nitrogen source on curdlan production by Alcaligenes faecalis ATCC 31749.

    PubMed

    Jiang, Longfa

    2013-01-01

    This study aims to investigate the effect of nitrogen source on curdlan production by Alcaligenes faecalis ATCC 31749. Curdlan production fell when excess nitrogen source was present, while biomass accumulation increased as the level of nitrogen source raised. Curdlan production and biomass accumulation were greater with urea compared with those with other nitrogen sources. The highest production of curdlan and biomass accumulation by A. faecalis ATCC 31749 was 28.16 g L(-1) and 9.58 g L(-1), respectively, with urea, whereas those with NH(4)Cl were 15.17 g L(-1) and 6.25 g L(-1), respectively. The optimum fermentation time for curdlan production was also affected by the nitrogen source in the medium. PMID:23085490

  6. Effects of phosphoenolpyruvate carboxylase desensitization on glutamic acid production in Corynebacterium glutamicum ATCC 13032.

    PubMed

    Wada, Masaru; Sawada, Kazunori; Ogura, Kotaro; Shimono, Yuta; Hagiwara, Takuya; Sugimoto, Masakazu; Onuki, Akiko; Yokota, Atsushi

    2016-02-01

    Phosphoenolpyruvate carboxylase (PEPC) in Corynebacterium glutamicum ATCC13032, a glutamic-acid producing actinobacterium, is subject to feedback inhibition by metabolic intermediates such as aspartic acid and 2-oxoglutaric acid, which implies the importance of PEPC in replenishing oxaloacetic acid into the TCA cycle. Here, we investigated the effects of feedback-insensitive PEPC on glutamic acid production. A single amino-acid substitution in PEPC, D299N, was found to relieve the feedback control by aspartic acid, but not by 2-oxoglutaric acid. A simple mutant, strain R1, having the D299N substitution in PEPC was constructed from ATCC 13032 using the double-crossover chromosome replacement technique. Strain R1 produced glutamic acid at a concentration of 31.0 g/L from 100 g/L glucose in a jar fermentor culture under biotin-limited conditions, which was significantly higher than that of the parent, 26.0 g/L (1.19-fold), indicative of the positive effect of desensitized PEPC on glutamic acid production. Another mutant, strain DR1, having both desensitized PEPC and PYK-gene deleted mutations, was constructed in a similar manner using strain D1 with a PYK-gene deleted mutation as the parent. This mutation had been shown to enhance glutamic acid production in our previous study. Although marginal, strain D1 produced higher glutamic acid, 28.8 g/L, than ATCC13032 (1.11-fold). In contrast, glutamic acid production by strain DR-1 was elevated up to 36.9 g/L, which was 1.42-fold higher than ATCC13032 and significantly higher than the other three strains. The results showed a synergistic effect of these two mutations on glutamic acid production in C. glutamicum. PMID:26168906

  7. Genome sequence of the methanotrophic Alphaproteobacterium, Methylocystis sp. Rockwell (ATCC 49242)

    SciTech Connect

    Stein, Lisa Y.; Bringel, Francoise O.; DiSpiritto, Alan A.; Han, Sukkyun; Jetten, MSM; Kalyuzhnaya, Marina G.; Kits, K. Dimitri; Klotz, Martin G; Op den Camp, HJM; Semrau, Jeremy D.; Vuilleumier, Stephane; Bruce, David; Cheng, Jan-Fang; Copeland, A; Davenport, Karen W.; Goodwin, Lynne A.; Han, Cliff; Hauser, Loren John; Lajus, Aurelie; Lapidus, Alla L.; Lucas, Susan; Medigue, Claudine

    2011-01-01

    Methylocystis sp. strain Rockwell (ATCC 49242) is an aerobic methane-oxidizing Alphaproteobacterium isolated from an aquifer in southern California. Unlike most methanotrophs in the Methylocystaceae family, this strain has a single pmo operon encoding particulate methane monooxygenase and no evidence of the genes encoding soluble methane monooxygenase. This is the first reported genome sequence of a member of the Methylocystis species of the Methylocystaceae family in the order Rhizobiales.

  8. Complete Genome Assembly of Streptococcus pyogenes ATCC 19615, a Group A β-Hemolytic Reference Strain.

    PubMed

    Minogue, T D; Daligault, H A; Davenport, K W; Bishop-Lilly, K A; Broomall, S M; Bruce, D C; Chain, P S; Chertkov, O; Coyne, S R; Freitas, T; Frey, K G; Gibbons, H S; Jaissle, J; Redden, C L; Rosenzweig, C N; Xu, Y; Johnson, S L

    2014-01-01

    We present the complete genome assembly of Streptococcus pyogenes ATCC 19615 (Rosenbach) as submitted to GenBank under accession number CP008926. This group A nonmotile β-hemolytic clinical isolate is used for quality control in a variety of commercially available tests. The assembled genome is 1.84 Mb (38.5% G+C content) and contains 1,788 coding regions. PMID:25258274

  9. Complete Genome Assembly of Streptococcus pyogenes ATCC 19615, a Group A β-Hemolytic Reference Strain

    PubMed Central

    Minogue, T. D.; Daligault, H. A.; Davenport, K. W.; Bishop-Lilly, K. A.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Chertkov, O.; Coyne, S. R.; Freitas, T.; Frey, K. G.; Gibbons, H. S.; Jaissle, J.; Redden, C. L.; Rosenzweig, C. N.; Xu, Y.

    2014-01-01

    We present the complete genome assembly of Streptococcus pyogenes ATCC 19615 (Rosenbach) as submitted to GenBank under accession number CP008926. This group A nonmotile β-hemolytic clinical isolate is used for quality control in a variety of commercially available tests. The assembled genome is 1.84 Mb (38.5% G+C content) and contains 1,788 coding regions. PMID:25258274

  10. Efficacy of oral Bifidobacterium bifidum ATCC 29521 on microflora and antioxidant in mice.

    PubMed

    Wang, Bao-Gui; Xu, Hai-Bo; Xu, Feng; Zeng, Zhe-Ling; Wei, Hua

    2016-03-01

    This study aimed to examine whether Bifidobacterium bifidum ATCC 29521, a species of colonic microflora in humans, is involved in the intestinal tract of mice. This study was also conducted to determine the antioxidant activity of this species by evaluating different microbial populations and reactive oxygen species isolated from feces and intestinal contents for 28 days of oral administration. Microbial diversities were assessed through bacterial culture techniques, PCR-DGGE, and real-time PCR. This study showed that the intake of B. bifidum ATCC 29521 significantly (p < 0.05) improved the ecosystem of the intestinal tract of BALB/c mice by increasing the amount of probiotics (Lactobacillus intestinalis and Lactobacillus crispatus) and by reducing unwanted bacterial populations (Enterobacter, Escherichia coli). Antioxidative activities of incubated cell-free extracts were evaluated through various assays, including the scavenging ability of DPPH radical (64.5% and 67.54% (p < 0.05), respectively, at 21 days in nutrients and 28 days in MRS broth), superoxide anion, and hydroxyl radical (85% and 61.5% (p < 0.05), respectively, at intestinal contents in nutrients and 21 days in MRS broth). Total reducing power (231.5 μmol/L (p < 0.05), 14 days in MRS broth) and mRNA level of genes related to oxidative stress were also determined. Results indicated that B. bifidum ATCC 29521 elicits a beneficial effect on murine gut microbiota and antioxidant activities compared with the control samples. This species can be considered as a potential bioresource antioxidant to promote health. Bifidobacterium bifidum ATCC 29521 may also be used as a promising material in microbiological and food applications. PMID:26863255

  11. Complete Genome Sequence of Pseudomonas syringae pv. lapsa Strain ATCC 10859, Isolated from Infected Wheat

    PubMed Central

    Kong, Jun; Jiang, Hongshan; Li, Baiyun; Zhao, Wenjun

    2016-01-01

    Pseudomonas syringae pv. lapsa is a pathovar of Pseudomonas syringae that can infect wheat. The complete genome of P. syringae pv. lapsa strain ATCC 10859 contains a 5,918,899-bp circular chromosome with 4,973 coding sequences, 16 rRNAs, 69 tRNAs, and an average GC content of 59.13%. The analysis of this genome revealed several gene clusters that are related to pathogenesis and virulence. PMID:26941133

  12. Cloning, characterization, and production of three α-l-fucosidases from Clostridium perfringens ATCC 13124.

    PubMed

    Fan, Shuquan; Zhang, Huaqin; Chen, Xiaodi; Lu, Lili; Xu, Li; Xiao, Min

    2016-04-01

    α-l-Fucosidases are key enzymes for the degradation of intestinal glycans by gut microbes. In this work, three putative α-l-fucosidases (Afc1, Afc2, and Afc3) genes from Clostridium perfringens ATCC 13124 were cloned and expressed in Escherichia coli. Afc1 had the α-l-fucosidase domain of glycoside hydrolase (GH) 29 family but showed no enzyme activity toward all the substrates examined. The putative acid/base residue of Afc1, Ser205, was replaced by a glutamic acid which is conserved in GH29-B α-l-fucosidases. However, the mutant Afc1-S205E still failed to show enzyme activity. Afc2 and Afc3 were determined to be 1,3-1,4-α-l-fucosidase of GH29-B subfamily and 1,2-α-l-fucosidase of GH95 family, respectively, and both of them could release fucose from porcine gastric mucin (PGM). When C. perfringens ATCC 13124 grew with the presence of PGM, the transcription of afc1 decreased slightly, while those of afc2 and afc3 increased to 2.2-fold and 1.4-fold, respectively, and the enzyme activities of Afc2 and Afc3 in the culture increased to 2.2-fold and 2.6-fold, respectively. These results suggest that Afc2 and Afc3 are involved in the degradation of intestinal fucosyl glycans by C. perfringens ATCC 13124. PMID:26663202

  13. Genome-scale reconstruction of metabolic networks of Lactobacillus casei ATCC 334 and 12A.

    PubMed

    Vinay-Lara, Elena; Hamilton, Joshua J; Stahl, Buffy; Broadbent, Jeff R; Reed, Jennifer L; Steele, James L

    2014-01-01

    Lactobacillus casei strains are widely used in industry and the utility of this organism in these industrial applications is strain dependent. Hence, tools capable of predicting strain specific phenotypes would have utility in the selection of strains for specific industrial processes. Genome-scale metabolic models can be utilized to better understand genotype-phenotype relationships and to compare different organisms. To assist in the selection and development of strains with enhanced industrial utility, genome-scale models for L. casei ATCC 334, a well characterized strain, and strain 12A, a corn silage isolate, were constructed. Draft models were generated from RAST genome annotations using the Model SEED database and refined by evaluating ATP generating cycles, mass-and-charge-balances of reactions, and growth phenotypes. After the validation process was finished, we compared the metabolic networks of these two strains to identify metabolic, genetic and ortholog differences that may lead to different phenotypic behaviors. We conclude that the metabolic capabilities of the two networks are highly similar. The L. casei ATCC 334 model accounts for 1,040 reactions, 959 metabolites and 548 genes, while the L. casei 12A model accounts for 1,076 reactions, 979 metabolites and 640 genes. The developed L. casei ATCC 334 and 12A metabolic models will enable better understanding of the physiology of these organisms and be valuable tools in the development and selection of strains with enhanced utility in a variety of industrial applications. PMID:25365062

  14. Cloning, Purification and Characterization of the Collagenase ColA Expressed by Bacillus cereus ATCC 14579.

    PubMed

    Abfalter, Carmen M; Schönauer, Esther; Ponnuraj, Karthe; Huemer, Markus; Gadermaier, Gabriele; Regl, Christof; Briza, Peter; Ferreira, Fatima; Huber, Christian G; Brandstetter, Hans; Posselt, Gernot; Wessler, Silja

    2016-01-01

    Bacterial collagenases differ considerably in their structure and functions. The collagenases ColH and ColG from Clostridium histolyticum and ColA expressed by Clostridium perfringens are well-characterized collagenases that cleave triple-helical collagen, which were therefore termed as ´true´ collagenases. ColA from Bacillus cereus (B. cereus) has been added to the collection of true collagenases. However, the molecular characteristics of B. cereus ColA are less understood. In this study, we identified ColA as a secreted true collagenase from B. cereus ATCC 14579, which is transcriptionally controlled by the regulon phospholipase C regulator (PlcR). B. cereus ATCC 14579 ColA was cloned to express recombinant wildtype ColA (ColAwt) and mutated to a proteolytically inactive (ColAE501A) version. Recombinant ColAwt was tested for gelatinolytic and collagenolytic activities and ColAE501A was used for the production of a polyclonal anti-ColA antibody. Comparison of ColAwt activity with homologous proteases in additional strains of B. cereus sensu lato (B. cereus s.l.) and related clostridial collagenases revealed that B. cereus ATCC 14579 ColA is a highly active peptidolytic and collagenolytic protease. These findings could lead to a deeper insight into the function and mechanism of bacterial collagenases which are used in medical and biotechnological applications. PMID:27588686

  15. The sim operon facilitates the transport and metabolism of sucrose isomers in Lactobacillus casei ATCC 334.

    PubMed

    Thompson, John; Jakubovics, Nicholas; Abraham, Bindu; Hess, Sonja; Pikis, Andreas

    2008-05-01

    Inspection of the genome sequence of Lactobacillus casei ATCC 334 revealed two operons that might dissimilate the five isomers of sucrose. To test this hypothesis, cells of L. casei ATCC 334 were grown in a defined medium supplemented with various sugars, including each of the five isomeric disaccharides. Extracts prepared from cells grown on the sucrose isomers contained high levels of two polypeptides with M(r)s of approximately 50,000 and approximately 17,500. Neither protein was present in cells grown on glucose, maltose or sucrose. Proteomic, enzymatic, and Western blot analyses identified the approximately 50-kDa protein as an NAD(+)- and metal ion-dependent phospho-alpha-glucosidase. The oligomeric enzyme was purified, and a catalytic mechanism is proposed. The smaller polypeptide represented an EIIA component of the phosphoenolpyruvate-dependent sugar phosphotransferase system. Phospho-alpha-glucosidase and EIIA are encoded by genes at the LSEI_0369 (simA) and LSEI_0374 (simF) loci, respectively, in a block of seven genes comprising the sucrose isomer metabolism (sim) operon. Northern blot analyses provided evidence that three mRNA transcripts were up-regulated during logarithmic growth of L. casei ATCC 334 on sucrose isomers. Internal simA and simF gene probes hybridized to approximately 1.5- and approximately 1.3-kb transcripts, respectively. A 6.8-kb mRNA transcript was detected by both probes, which was indicative of cotranscription of the entire sim operon. PMID:18310337

  16. Dynamic proteome analysis of Cyanothece sp. ATCC 51142 under constant light

    SciTech Connect

    Aryal, Uma K.; Stockel, Jana; Welsh, Eric A.; Gritsenko, Marina A.; Nicora, Carrie D.; Koppenaal, David W.; Smith, Richard D.; Pakrasi, Himadri B.; Jacobs, Jon M.

    2012-02-03

    Understanding the dynamic nature of protein abundances provides insights into protein turnover not readily apparent from conventional, static mass spectrometry measurements. This level of data is particularly informative when surveying protein abundances in biological systems subjected to large perturbations or alterations in environment such as cyanobacteria. Our current analysis expands upon conventional proteomic approaches in cyanobacteria by measuring dynamic changes of the proteome using a 13C15N-L-leucine metabolic labeling in Cyanothece ATCC51142. Metabolically labeled Cyanothece ATCC51142 cells grown under nitrogen sufficient conditions in continuous light were monitored longitudinally for isotope incorporation over a 48 h period, revealing 422 proteins with dynamic changes in abundances. In particular, proteins involved in carbon fixation, pentose phosphate pathway, cellular protection, redox regulation, protein folding, assembly and degradation showed higher levels of isotope incorporation suggesting that these biochemical pathways are important for growth under non-diazotrophic conditions. Calculation of relative isotope abundances (RIA) values allowed to measure actual active protein synthesis over time for different biochemical pathways under non-diazotrophic conditions. Overall results demonstrated the utility of 'non-steady state' pulsed metabolic labeling for systems-wide dynamic quantification of the proteome in Cyanothece ATCC51142 that can also be applied to other cyanobacteria.

  17. Dynamic proteomic profiling of a unicellular cyanobacterium Cyanothece ATCC51142 across light-dark diurnal cycles

    SciTech Connect

    Aryal, Uma K.; Stockel, Jana; Krovvidi, Ravi K.; Gritsenko, Marina A.; Monroe, Matthew E.; Moore, Ronald J.; Koppenaal, David W.; Smith, Richard D.; Pakrasi, Himadri B.; Jacobs, Jon M.

    2011-12-01

    Unicellular cyanobacteria of the genus Cyanothece are recognized for their ability to execute nitrogen (N2)-fixation in the dark and photosynthesis in the light. Systems-wide dynamic proteomic profiling with mass spectrometry (MS) analysis reveals fundamental insights into the control and regulation of these functions. To expand upon the current knowledge of protein expression patterns in Cyanothece ATCC51142, we performed quantitative proteomic analysis using partial ("unsaturated") metabolic labeling and high mass accuracy LC-MS analysis. This dynamic proteomic profiling identified 721 actively synthesized proteins with significant temporal changes in expression throughout the light-dark cycles, of which 425 proteins matched with previously characterized cycling transcripts. The remaining 296 proteins contained a cluster of proteins uniquely involved in DNA replication and repair, protein degradation, tRNA synthesis and modification, transport and binding, and regulatory functions. Analysis of protein functions revealed that the expression of nitrogenase in the dark is mediated by higher respiration and glycogen metabolism. We have also shown that Cyanothece ATCC51142 utilizes alternative pathways for carbon (C) and nitrogen (N) acquisition, particularly, aspartic acid and glutamate as substrates of C and N, respectively. Utilization of phosphoketolase (PHK) pathway for the conversion of xylulose-5P to pyruvate and acetyl-P likely constitutes an alternative strategy to compensate higher ATP and NADPH demand. In conclusion, this study provides a deeper insight into how Cyanothece ATCC51142 modulates cellular functions to accommodate photosynthesis and N2-fixation within the single cell.

  18. Genome –Scale Reconstruction of Metabolic Networks of Lactobacillus casei ATCC 334 and 12A

    PubMed Central

    Vinay-Lara, Elena; Hamilton, Joshua J.; Stahl, Buffy; Broadbent, Jeff R.; Reed, Jennifer L.; Steele, James L.

    2014-01-01

    Lactobacillus casei strains are widely used in industry and the utility of this organism in these industrial applications is strain dependent. Hence, tools capable of predicting strain specific phenotypes would have utility in the selection of strains for specific industrial processes. Genome-scale metabolic models can be utilized to better understand genotype-phenotype relationships and to compare different organisms. To assist in the selection and development of strains with enhanced industrial utility, genome-scale models for L. casei ATCC 334, a well characterized strain, and strain 12A, a corn silage isolate, were constructed. Draft models were generated from RAST genome annotations using the Model SEED database and refined by evaluating ATP generating cycles, mass-and-charge-balances of reactions, and growth phenotypes. After the validation process was finished, we compared the metabolic networks of these two strains to identify metabolic, genetic and ortholog differences that may lead to different phenotypic behaviors. We conclude that the metabolic capabilities of the two networks are highly similar. The L. casei ATCC 334 model accounts for 1,040 reactions, 959 metabolites and 548 genes, while the L. casei 12A model accounts for 1,076 reactions, 979 metabolites and 640 genes. The developed L. casei ATCC 334 and 12A metabolic models will enable better understanding of the physiology of these organisms and be valuable tools in the development and selection of strains with enhanced utility in a variety of industrial applications. PMID:25365062

  19. Characterization of KfrA proteins encoded by a plasmid of Paenibacillus popilliae ATCC 14706T

    PubMed Central

    Iiyama, Kazuhiro; Mon, Hiroaki; Mori, Kazuki; Mitsudome, Takumi; Lee, Jae Man; Kusakabe, Takahiro; Tashiro, Kousuke; Asano, Shin-ichiro; Yasunaga-Aoki, Chisa

    2015-01-01

    A scaffold obtained from whole-genome shotgun sequencing of Paenibacillus popilliae ATCC 14706T shares partial homology with plasmids found in other strains of P. popilliae. PCR and sequencing for gap enclosure indicated that the scaffold originated from a 15,929-bp circular DNA. The restriction patterns of a plasmid isolated from P. popilliae ATCC 14706T were identical to those expected from the sequence; thus, this circular DNA was identified as a plasmid of ATCC 14706T and designated pPOP15.9. The plasmid encodes 17 putative open reading frames. Orfs 1, 5, 7, 8, and 9 are homologous to Orfs 11, 12, 15, 16, and 17, respectively. Orf1 and Orf11 are annotated as replication initiation proteins. Orf8 and Orf16 are homologs of KfrA, a plasmid-stabilizing protein in Gram-negative bacteria. Recombinant Orf8 and Orf16 proteins were assessed for the properties of KfrA. Indeed, they formed multimers and bound to inverted repeat sequences in upstream regions of both orf8 and orf16. A phylogenetic tree based on amino acid sequences of Orf8, Orf16 and Kfr proteins did not correlate with species lineage. PMID:25853059

  20. Economic impacts of reduced pork production associated with the diagnosis of Actinobacillus pleuropneumoniae on grower/finisher swine operations in the United States.

    PubMed

    Losinger, Willard C

    2005-05-10

    An examination of the economic impacts of the diagnosis of Actinobacillus pleuropneumoniae on grower/finisher swine operations indicated that reduced pork production, associated with the diagnosis of A. pleuropneumoniae on the operation, diminished consumer surplus by $53+/-52 million, and resulted in a total loss of $32+/-30 million to the US economy in 1995. Most of the economic surplus lost by consumers was transferred to producers, whose economic surplus increased by $21+/-25 million (which was not significantly different from zero). Uncertainty analysis showed that an estimate of the decline in production associated with the diagnosis of A. pleuropneumoniae accounted for most of the uncertainty of the change in consumer surplus and of the total loss to the economy. The estimate of the price elasticity of demand for pork also contributed towards a lot of the uncertainty in the estimated change in producer surplus. PMID:15820115

  1. Minimum inhibitory concentration breakpoints and disk diffusion inhibitory zone interpretive criteria for tilmicosin susceptibility testing against Pasteurella multocida and Actinobacillus pleuropneumoniae associated with porcine respiratory disease.

    PubMed

    Shryock, Thomas R; Staples, J Mitchell; DeRosa, David C

    2002-09-01

    Tilmicosin is a novel macrolide antibiotic developed for exclusive use in veterinary medicine. Tilmicosin has been approved as a feed premix to control porcine respiratory disease associated with Pasteurella multocida and Actinobacillus pleuropneumoniae. The development of antimicrobial susceptibility testing guidelines for tilmicosin was predicated on the relationship of clinical efficacy studies that demonstrated a favorable therapeutic outcome, on pharmacokinetic data, and on in vitro test data, as recommended by the National Committee for Clinical Laboratory Standards (NCCLS). The approved breakpoints for the minimum inhibitory concentration dilution testing for both species are resistant, > or = 32 microg/ml, and susceptible, < or = 16 microg/ml. The zone of inhibition interpretive criteria for disk diffusion testing with a 15-microg tilmicosin disk are resistant, < or = 10 mm, and susceptible, > or = 11 mm. PMID:12296390

  2. Use of recombinant ApxIV in serodiagnosis of Actinobacillus pleuropneumoniae infections, development and prevalidation of the ApxIV ELISA.

    PubMed

    Dreyfus, A; Schaller, A; Nivollet, S; Segers, R P A M; Kobisch, M; Mieli, L; Soerensen, V; Hüssy, D; Miserez, R; Zimmermann, W; Inderbitzin, F; Frey, J

    2004-04-19

    Actinobacillus pleuropneumoniae is the etiological agent of porcine pleuropneumonia, which causes worldwide severe losses in pig farming. The virulence of the 15 serotypes of A. pleuropneumoniae is mainly determined by the three major RTX toxins ApxI, ApxII and ApxIII, which are secreted by the different serotypes in various combinations. A fourth RTX toxin, ApxIV, is produced by all 15 serotypes only during infection of pigs, but not under in vitro conditions. Pigs infected with A. pleuropneumoniae show specific antibodies directed against ApxIV. In contrast, antibodies against the other three toxins ApxI, ApxII and ApxIII are also found in pigs free of A. pleuropneumoniae. The antibodies to the three latter might result from other, less pathogenic Actinobacillus species such as A. rossii and A. suis. We used a recombinant protein based on the N'-terminal part of ApxIV to serologically detect A. pleuropneumoniae infections in pigs by immunoblot analysis. The analysis of sera of experimentally infected pigs revealed that ApxIV-immunoblots detected A. pleuropneumoniae infections in the second to third week post infection. We developed an indirect ELISA based on the purified recombinant N'-terminal moiety of ApxIV. The analysis of sera from pigs that were experimentally or naturally infected by A. pleuropneumoniae, and of sera of pigs that were free of A. pleuropneumoniae, revealed that the ELISA had a specificity of 100% and a sensitivity of 93.8%. The pre-validation study of the ApxIV-ELISA revealed that the latter was able to detect A. pleuropneumoniae-positive herds, even when clinical and pathological signs of porcine pleuropneumonia were not evident. Pigs vaccinated with a subunit vaccine Porcilis App were serologically negative in the ApxIV-ELISA. PMID:15066725

  3. Serodiagnosis of pleuropneumonia using enzyme-linked immunosorbent assay with capsular polysaccharide antigens of Actinobacillus pleuropneumoniae serotypes 1, 2, 5 and 7.

    PubMed Central

    Bossé, J T; Johnson, R P; Rosendal, S

    1990-01-01

    Capsular polysaccharide antigens of serotypes 1, 2, 5 and 7 of Actinobacillus pleuropneumoniae were used in enzyme-linked immunosorbent assays (ELISAs) to test sera from experimentally infected and field pigs. Specific reactions were found in sera of experimental pigs with antigens of serotypes 1, 5 and 7 whereas the serotype 2 antigen was cross-reactive. A 1:200 serum dilution was used for testing of 300 sera from 21 swine herds in southern Ontario. Cases of pleuropneumonia had occurred in 11 of these herds, but not in the others. The negative cut-off value was the mean optical density at 405 nm (OD405) + three standard deviations (SD) for 16 negative reference sera. Sera from four pigs naturally infected with Actinobacillus suis were tested and found to react to varying degrees with each of the antigens. Therefore a second cut-off value was determined as the mean OD405 + 2 SD for the A. suis sera. Sera which, in the ELISA produced OD readings above the latter cut-off were considered positive for antibodies to A. pleuropneumoniae; those which were lower than the former cut-off were considered negative. Readings between the two cut-off values may have been due to low positive titers or cross-reactivity, possibly with A. suis, and could not be used to predict pleuropneumonia. Of the pleuropneumonia-free herds, none had positive reactors to serotypes 5 or 7, whereas one and two herds had positive reactors to serotypes 1 and 2, respectively. Of the pleuropneumonia positive herds, six had positive reactors to serotype 1, one to serotype 2, four to serotype 5, and eight to serotype 7. PMID:2249177

  4. Effects of oakmoss and its components on Acanthamoeba castellanii ATCC 30234 and the uptake of Legionella pneumophila JCM 7571 (ATCC 33152) into A. castellanii.

    PubMed

    Nomura, Harue; Isshiki, Yasunori; Sakuda, Keisuke; Sakuma, Katsuya; Kondo, Seiichi

    2015-01-01

    Acanthamoeba castellanii, a ubiquitous organism in water environments, is pathogenic toward humans and also is a host for bacteria of the genus Legionella, a causative agent of legionellosis. Oakmoss, a natural fragrance ingredient, and its components are antibacterial agents specifically against the genus Legionella. In the present study, oakmoss and its components were investigated for their amoebicidal activity against A. castellanii ATCC 30234 and the inhibitory effect on the uptake of L. pneumophila JCM 7571 (ATCC 33152) into A. castellanii. The oakmoss and its components 3-hydroxy-5-methylphenyl 2,4-dihydroxy-6-methylbenzoate(5), and 6,8-dihydroxy-3-pentyl-1H-isochromen-1-one (12) exhibited high amoebicidal activity (IC50 values; 10.5 ± 2.3, 16.3 ± 4.0 and 17.5 ± 2.8 μg/mL, respectively) after 48 h of treatment, which were equivalent to that of the reference compound, chlorhexidine gluconate. Pretreatment of L. pneumophila with sub-minimal inhibitory concentration of oakmoss, compound 5, 3-hydroxy-5-methylphenyl 2-hydroxy-4-methoxy-6-methylbenzoate (10) and 8-(2,4-dihydroxy-6-pentylphenoxy)-6-hydroxy-3-pentyl-1H-isochromen-1-one (14) obviously reduced the uptake of L. pneumophila into A.castellanii (p < 0.05).The inhibitory effect of compound 5 on the uptake of L. pneumophila was almost equivalent to that of ampicillin used as a reference. Thus, the oakmoss and its components were considered to be good candidates for disinfectants against not only genus Legionella but also A. castellanii. PMID:25817814

  5. Extraction, purification, and characterization of major outer membrane proteins from Wolinella recta ATCC 33238.

    PubMed Central

    Kennell, W L; Holt, S C

    1991-01-01

    The outer membrane of Wolinella recta ATCC 33238 was isolated by French pressure cell disruption and differential centrifugation. Outer membrane proteins (OMPs) were solubilized by Zwittergent 3.14 extraction and separated by DEAE-Sephacel ion-exchange chromatography. The major OMPs that were found in W. recta ATCC 33238 and in several other Wolinella spp. consisted of proteins with apparent molecular masses of 51, 45, and 43 kDa. These three conserved proteins were purified to essential homogeneity by one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and characterized chemically. Heating at between 75 and 100 degrees C revealed both the 43- and 51-kDa proteins to be heat modified from apparent molecular masses of 32 and 38 kDa, respectively. The 45-kDa protein was unmodified at all temperatures tested. Two-dimensional isoelectric focusing-SDS-PAGE revealed the 51-kDa protein to be composed of multiple pIs between a pH of 5.0 and greater than 8.0 while the 43- and 45-kDa proteins had a pI of approximately 6.0. N'-terminal amino acid sequence analysis of the first 30 to 40 amino acids and search of the Protein Identification Resource data base for similar proteins only revealed the 43-kDa protein to be similar to the P.69 OMP of Bordetella pertussis; however, the homology was weak (33%). Amino acid analysis revealed the 43-kDa protein to be noncharged and the 45- and 51-kDa proteins to be hydrophilic, containing between 38 to 42% polar residues but no cysteine. This study reports the purification and partial characterization of three conserved proteins in W. recta ATCC 33238. Images PMID:1894372

  6. Dynamic proteomic profiling of a unicellular cyanobacterium Cyanothece ATCC51142 across light-dark diurnal cycles

    PubMed Central

    2011-01-01

    Background Unicellular cyanobacteria of the genus Cyanothece are recognized for their ability to execute nitrogen (N2)-fixation in the dark and photosynthesis in the light. An understanding of these mechanistic processes in an integrated systems context should provide insights into how Cyanothece might be optimized for specialized environments and/or industrial purposes. Systems-wide dynamic proteomic profiling with mass spectrometry (MS) analysis should reveal fundamental insights into the control and regulation of these functions. Results To expand upon the current knowledge of protein expression patterns in Cyanothece ATCC51142, we performed quantitative proteomic analysis using partial ("unsaturated") metabolic labeling and high mass accuracy LC-MS analysis. This dynamic proteomic profiling identified 721 actively synthesized proteins with significant temporal changes in expression throughout the light-dark cycles, of which 425 proteins matched with previously characterized cycling transcripts. The remaining 296 proteins contained a cluster of proteins uniquely involved in DNA replication and repair, protein degradation, tRNA synthesis and modification, transport and binding, and regulatory functions. Functional classification of labeled proteins suggested that proteins involved in respiration and glycogen metabolism showed increased expression in the dark cycle together with nitrogenase, suggesting that N2-fixation is mediated by higher respiration and glycogen metabolism. Results indicated that Cyanothece ATCC51142 might utilize alternative pathways for carbon (C) and nitrogen (N) acquisition, particularly, aspartic acid and glutamate as substrates of C and N, respectively. Utilization of phosphoketolase (PHK) pathway for the conversion of xylulose-5P to pyruvate and acetyl-P likely constitutes an alternative strategy to compensate higher ATP and NADPH demand. Conclusion This study provides a deeper systems level insight into how Cyanothece ATCC51142

  7. Improved penicillin amidase production using a genetically engineered mutant of escherichia coli ATCC 11105

    SciTech Connect

    Robas, N.; Zouheiry, H.; Branlant, G.; Branlant, C. )

    1993-01-05

    Penicillin G amidase (PGA) is a key enzyme for the industrial production of penicillin G derivatives used in therapeutics. Escherichia coli ATCC 11105 is the more commonly used strain for PGA production. To improve enzyme yield, the authors constructed various recombinant E. coli HB 101 and ATCC 11105 strains. For each strain, PGA production was determined for various concentrations of glucose and phenylacetic acid (PAA) in the medium. The E. coli strain, G271, was identified as the best performer (800 U NIPAB/L). This strain was obtained as follows: an E. coli ATCC 11105 mutant (E. coli G133) was first selected based on a low negative effect of glucose on PGA production. This mutant was then transformed with a pBR322 derivative containing the PGA gene. Various experiments were made to try to understand the reason for the high productivity of E. coli G271. The host strain, E. coli G133, was found to be mutated in one (or more) gene(s) whose product(s) act(s) in trans on the PGA gene expression. Its growth is not inhibited by high glucose concentration in the medium. Interestingly, whereas glucose still exerts some negative effect on the PGA production by E. coli G133, PGA production by its transformant (E. coli G271) is stimulated by glucose. The reason for this stimulation is discussed. Transformation of E. coli G133 with a pBR322 derivative containing the HindIII fragment of the PGA gene, showed that the performance of E. coli G271 depends both upon the host strain properties and the plasmid structure. Study of the production by the less efficient E. coli HB101 derivatives brought some light on the mechanism of regulation of the PGA gene.

  8. Evaluating Chemical Mitigation of Salmonella Typhimurium ATCC 14028 in Animal Feed Ingredients.

    PubMed

    Cochrane, Roger A; Huss, Anne R; Aldrich, Gregory C; Stark, Charles R; Jones, Cassandra K

    2016-04-01

    Salmonella Typhimurium is a potential feed safety hazard in animal feed ingredients. Thermal mitigation of Salmonella spp. during rendering is effective but does not eliminate the potential for cross-contamination. Therefore, the objective of this experiment was to evaluate the effectiveness of chemicals to mitigate postrendering Salmonella Typhimurium ATCC 14028 contamination in rendered proteins over time. Treatments were arranged in a 6 × 4 factorial with six chemical treatments and four rendered protein meals. The chemical treatments included (i) control without chemical treatment, (ii) 0.3% commercial formaldehyde product, (iii) 2% essential oil blend, (iv) 2% medium chain fatty acid blend, (v) 3% organic acid blend, and (vi) 1% sodium bisulfate. The four rendered protein meals included (i) feather meal, (ii) blood meal, (iii) meat and bone meal, and (iv) poultry by-product meal. After matrices were chemically treated, they were inoculated with Salmonella Typhimurium ATCC 14028, stored at room temperature, and enumerated via plate counts on days 0, 1, 3, 7, 14, 21, and 42 postinoculation. The Salmonella concentration in ingredients treated with medium chain fatty acid and commercial formaldehyde were similar to one another (P = 0.23) but were 2 log lower than the control (P < 0.05). Ingredients treated with organic acids and essential oils also had lower Salmonella concentrations than the control (P < 0.05). Time also played a significant role in Salmonella mitigation, because all days except days 14 and 21 (P = 0.92) differed from one another. Rendered protein matrix also affected Salmonella stability, because concentrations in meat and bone meal and blood meal were similar to one another (P = 0.36) but were greater than levels in feather meal and poultry by-product meal (P < 0.05). In summary, chemical treatment and time both mitigated Salmonella Typhimurium ATCC 14028, but their effectiveness was matrix dependent. Time and chemical treatment with medium

  9. Complete genome sequence of Vibrio alginolyticus ATCC 33787(T) isolated from seawater with three native megaplasmids.

    PubMed

    Wang, Pengxia; Wen, Zhongling; Li, Baiyuan; Zeng, Zhenshun; Wang, Xiaoxue

    2016-08-01

    Vibrio alginolyticus, an opportunistic pathogen, is commonly associated with vibriosis in fish and shellfish and can also cause superficial and ear infections in humans. V. alginolyticus ATCC 33787(T) was originally isolated from seawater and has been used as one of the type strains for exploring the virulence factors of marine bacteria and for developing vaccine against vibriosis. Here we sequenced and assembled the whole genome of this strain, and identified three megaplasmids and three Type VI secretion systems, thus providing useful information for the study of virulence factors and for the development of vaccine for Vibrio. PMID:27211073

  10. Multicenter Investigation of Gepotidacin (GSK2140944) Agar Dilution Quality Control Determinations for Neisseria gonorrhoeae ATCC 49226.

    PubMed

    Jones, Ronald N; Fedler, Kelley A; Scangarella-Oman, Nicole E; Ross, James E; Flamm, Robert K

    2016-07-01

    Gepotidacin, a novel triazaacenaphthylene antibacterial agent, is the first in a new class of type IIA topoisomerase inhibitors with activity against many biothreat and conventional pathogens, including Neisseria gonorrhoeae To assist ongoing clinical studies of gepotidacin to treat gonorrhea, a multilaboratory quality assurance investigation determined the reference organism (N. gonorrhoeae ATCC 49226) quality control MIC range to be 0.25 to 1 μg/ml (88.8% of gepotidacin MIC results at the 0.5 μg/ml mode). PMID:27161642

  11. Genomic and genetic characterization of the bile stress response of probiotic Lactobacillus reuteri ATCC 55730.

    PubMed

    Whitehead, Kristi; Versalovic, James; Roos, Stefan; Britton, Robert A

    2008-03-01

    Probiotic bacteria encounter various stresses after ingestion by the host, including exposure to the low pH in the stomach and bile in the small intestine. The probiotic microorganism Lactobacillus reuteri ATCC 55730 has previously been shown to survive in the human small intestine. To address how L. reuteri can resist bile stress, we performed microarray experiments to determine gene expression changes that occur when the organism is exposed to physiological concentrations of bile. A wide variety of genes that displayed differential expression in the presence of bile indicated that the cells were dealing with several types of stress, including cell envelope stress, protein denaturation, and DNA damage. Mutations in three genes were found to decrease the strain's ability to survive bile exposure: lr1864, a Clp chaperone; lr0085, a gene of unknown function; and lr1516, a putative esterase. Mutations in two genes that form an operon, lr1584 (a multidrug resistance transporter in the major facilitator superfamily) and lr1582 (unknown function), were found to impair the strain's ability to restart growth in the presence of bile. This study provides insight into the possible mechanisms that L. reuteri ATCC 55730 may use to survive and grow in the presence of bile in the small intestine. PMID:18245259

  12. Phosphoketolase pathway dominates in Lactobacillus reuteri ATCC 55730 containing dual pathways for glycolysis.

    PubMed

    Arsköld, Emma; Lohmeier-Vogel, Elke; Cao, Rong; Roos, Stefan; Rådström, Peter; van Niel, Ed W J

    2008-01-01

    Metabolic flux analysis indicated that the heterofermentative Lactobacillus reuteri strain ATCC 55730 uses both the Embden-Meyerhof pathway (EMP) and phosphoketolase pathway (PKP) when glucose or sucrose is converted into the three-carbon intermediate stage of glycolysis. In all cases studied, the main flux is through the PKP, while the EMP is used as a shunt. In the exponential growth phase, 70%, 73%, and 84% of the flux goes through the PKP in cells metabolizing (i) glucose plus fructose, (ii) glucose alone, and (iii) sucrose alone, respectively. Analysis of the genome of L. reuteri ATCC 55730 confirmed the presence of the genes for both pathways. Further evidence for the simultaneous operation of two central carbon metabolic pathways was found through the detection of fructose-1,6-bisphosphate aldolase, phosphofructokinase, and phosphoglucoisomerase activities and the presence of phosphorylated EMP and PKP intermediates using in vitro 31P NMR. The maximum specific growth rate and biomass yield obtained on glucose were twice as low as on sucrose. This was the result of low ATP levels being present in glucose-metabolizing cells, although the ATP production flux was as high as in sucrose-metabolizing cells due to a twofold increase of enzyme activities in both glycolytic pathways. Growth performance on glucose could be improved by adding fructose as an external electron acceptor, suggesting that the observed behavior is due to a redox imbalance causing energy starvation. PMID:17965151

  13. Crystallization and crystallographic analysis of branching enzymes from Cyanothece sp. ATCC 51142.

    PubMed

    Hayashi, Mari; Suzuki, Ryuichiro; Colleoni, Christophe; Ball, Steven G; Fujita, Naoko; Suzuki, Eiji

    2015-08-01

    Several cyanobacterial species, including Cyanothece sp. ATCC 51142, remarkably have four isoforms of α-glucan branching enzymes (BEs). Based on their primary structures, they are classified into glycoside hydrolase (GH) family 13 (BE1, BE2 and BE3) or family 57 (GH57 BE). In the present study, GH13-type BEs from Cyanothece sp. ATCC 51142 (BE1, BE2 and BE3) have been overexpressed in Escherichia coli and biochemically characterized. The recombinant BE1 was crystallized by the hanging-drop vapour-diffusion method. Crystals of BE1 were obtained at 293 K in the presence of 0.2 M Mg(2+), 7-10%(w/v) ethanol, 0.1 M HEPES-NaOH pH 7.2-7.9. The crystals belonged to the tetragonal space group P41212, with unit-cell parameters a = b = 133.75, c = 185.90 Å, and diffracted to beyond 1.85 Å resolution. Matthews coefficient calculations suggested that the crystals of BE1 contained two molecules in the asymmetric unit. PMID:26249708

  14. In silico analysis of Clostridium acetobutylicum ATCC 824 metabolic response to an external electron supply.

    PubMed

    Gallardo, Roberto; Acevedo, Alejandro; Quintero, Julián; Paredes, Ivan; Conejeros, Raúl; Aroca, Germán

    2016-02-01

    The biological production of butanol has become an important research field and thanks to genome sequencing and annotation; genome-scale metabolic reconstructions have been developed for several Clostridium species. This work makes use of the iCAC490 model of Clostridium acetobutylicum ATCC 824 to analyze its metabolic capabilities and response to an external electron supply through a constraint-based approach using the Constraint-Based Reconstruction Analysis Toolbox. Several analyses were conducted, which included sensitivity, production envelope, and phenotypic phase planes. The model showed that the use of an external electron supply, which acts as co-reducing agent along with glucose-derived reducing power (electrofermentation), results in an increase in the butanol-specific productivity. However, a proportional increase in the butyrate uptake flux is required. Besides, the uptake of external butyrate leads to the coupling of butanol production and growth, which coincides with results reported in literature. Phenotypic phase planes showed that the reducing capacity becomes more limiting for growth at high butyrate uptake fluxes. An electron uptake flux allows the metabolism to reach the growth optimality line. Although the maximum butanol flux does not coincide with the growth optimality line, a butyrate uptake combined with an electron uptake flux would result in an increased butanol volumetric productivity, being a potential strategy to optimize the production of butanol by C. acetobutylicum ATCC 824. PMID:26650720

  15. Distinct adhesion of probiotic strain Lactobacillus casei ATCC 393 to rat intestinal mucosa.

    PubMed

    Saxami, Georgia; Ypsilantis, Petros; Sidira, Marianthi; Simopoulos, Constantinos; Kourkoutas, Yiannis; Galanis, Alex

    2012-08-01

    Adhesion to the intestine represents a critical parameter for probiotic action. In this study, the adhesion ability of Lactobacillus casei ATCC 393 to the gastrointestinal tract of Wistar rats was examined after single and daily administration of fermented milk containing either free or immobilized cells on apple pieces. The adhesion of the probiotic cells at the large intestine (cecum and colon) was recorded at levels ≥6 logCFU/g (suggested minimum levels for conferring a probiotic effect) following daily administration for 7 days by combining microbiological and strain-specific multiplex PCR analysis. Single dose administration resulted in slightly reduced counts (5 logCFU/g), while they were lower at the small intestine (duodenum, jejunum, ileum) (≤3 logCFU/g), indicating that adhesion was a targeted process. Of note, the levels of L. casei ATCC 393 were enhanced in the cecal and colon fluids both at single and daily administration of immobilized cells (6 and 7 logCFU/g, respectively). The adhesion of the GI tract was transient and thus daily consumption of probiotic products containing the specific strain is suggested as an important prerequisite for retaining its levels at an effective concentration. PMID:22554894

  16. Global transcriptome analysis of Bacillus cereus ATCC 14579 in response to silver nitrate stress

    PubMed Central

    2011-01-01

    Silver nanoparticles (AgNPs) were synthesized using Bacillus cereus strains. Earlier, we had synthesized monodispersive crystalline silver nanoparticles using B. cereus PGN1 and ATCC14579 strains. These strains have showed high level of resistance to silver nitrate (1 mM) but their global transcriptomic response has not been studied earlier. In this study, we investigated the cellular and metabolic response of B. cereus ATCC14579 treated with 1 mM silver nitrate for 30 & 60 min. Global expression profiling using genomic DNA microarray indicated that 10% (n = 524) of the total genes (n = 5234) represented on the microarray were up-regulated in the cells treated with silver nitrate. The majority of genes encoding for chaperones (GroEL), nutrient transporters, DNA replication, membrane proteins, etc. were up-regulated. A substantial number of the genes encoding chemotaxis and flagellar proteins were observed to be down-regulated. Motility assay of the silver nitrate treated cells revealed reduction in their chemotactic activity compared to the control cells. In addition, 14 distinct transcripts overexpressed from the 'empty' intergenic regions were also identified and proposed as stress-responsive non-coding small RNAs. PMID:22071005

  17. Using an established pre adipose cell line (ATCC CL 173) for studying chromium metabolism and biopotency

    SciTech Connect

    Hendricks, D.G.; Azuka, C. ); Johnson, B. )

    1991-03-15

    Studies concerning the mechanism by which chromium enhances insulin action and identification of chromium compounds which have high glucose tolerance factor activity have been limited by biological testing methods. The isolated rat adipose tissue assay has been commonly used. The pre-adipose cell line (ATCC CL 173) has many advantages over the isolated rat adipose tissue method including: uniformity of cells; increased viability of the cell cultures; can readily be grown in a low chromium media with a cell doubling time of about 18 hours; studies can be made on rapidly multiplying cells or on mature adipose cells and using rate of cell number increase, percent attachment of cells or cultures in an Omni Spec instrument avoid the use of radio labeled compounds. The ATCC CL 173 pre adipose cell line has been used to evaluate various chromium compounds for insulin like and insulin enhancing activity using physiological and pharmacological chromium levels. Rapidly dividing cells and mature adipose cells do not always react similarly with the same chromium compound. Cell response to a chromium compound and/or insulin also differs based on the growth media in which the cells have been maintained. Duration of metabolic studies as well as metabolite chosen also influence cellular response to chromium and insulin. Using this method for biological testing several chromium compounds have shown insulin like activity and a few show actual insulin potentiation.

  18. Ferulic acid transformation into the main vanilla aroma compounds by Amycolatopsis sp. ATCC 39116.

    PubMed

    Pérez-Rodríguez, Noelia; Oliveira, Ricardo Pinheiro de Souza; Agrasar, Ana María Torrado; Domínguez, José Manuel

    2016-02-01

    The wild strain Amycolatopsis sp. ATCC 39116 was explored in ferulic acid-based media to produce naturally the aroma components of the cured vanilla pod, namely vanillin,vanillic acid, and vanillyl alcohol. Other phenolic compounds(4-vinyl guaiacol, guaiacol, and protocatechuic acid) were also evaluated. The influence of medium composition,fermentation technology (batch or fed-batch), supplementation with vanillic acid, and inoculum concentration on ferulic acid biotransformation were evaluated. The results postulate the initial concentration of cell mass as the variable with the strongest impact on ferulic acid metabolization under the studied conditions. The highest amounts of vanillin and vanillic acid were achieved at intermediate values of cell mass.Vanillyl alcohol and protocatechuic acid were more closely linked to high cell mass concentrations. Conversely, 4-vinyl guaiacol reached its highest amount at the lowest amount of cell mass. Guaiacol was not detected in any case. Therefore,the initial cell concentration must be considered a critical parameter when using Amycolaptosis sp. ATCC 39116 for the production of vanillin and related compounds. PMID:26476645

  19. Production of fructosyltransferase by Aureobasidium sp. ATCC 20524 in batch and two-step batch cultures.

    PubMed

    Salinas, Martín A; Perotti, Nora I

    2009-01-01

    A comparison of fructosyltransferase (EC 2.4.1.9) production by Aureobasidium sp. ATCC 20524 in batch and two step batch cultures was investigated in a 1-l stirred tank reactor using a sucrose supply of 200 g/l. Results showed that the innovative cultivation in two step of Aureobasidium sp. produced more fructosyltransferase (FFase) than the single batch culture at the same sucrose concentration with a maximal enzyme production of 523 U/ml, which was 80.5% higher than the one obtained in the batch culture. The production of fructooligosaccharides (FOSs) was also analyzed; their concentration reached a maximum value of 160 g/l the first day in the two-step culture and 127 g/l in the single-batch mode. The use of the two-step batch culture with Aureobasidium sp. ATCC 20524 in allowing the microorganism to grow up prior to the induction of sucrose (second step), proved to be a powerful method for producing fructosyltransferase and FOSs. PMID:18810518

  20. Assessment of in vitro removal of cholesterol oxidation products by Lactobacillus casei ATCC334.

    PubMed

    Machorro-Méndez, I A; Hernández-Mendoza, A; Cardenia, V; Rodriguez-Estrada, M T; Lercker, G; Spinelli, F; Cellini, A; García, H S

    2013-11-01

    Cholesterol oxidation products (COPs) are a group of compounds formed during processing and storage of foods from animal origin. After ingestion, COPs are absorbed in the intestine and can be distributed to serum and various tissues, potentially promoting a variety of toxic effects. Therefore, inhibition of their intestinal absorption may contribute to reduce the health risks associated with dietary intake of COPs. Some studies have shown that drugs and dietary compounds may inhibit the intestinal absorption of dietary COPs. However, proven cholesterol- and/or food toxins-binding lactic acid bacteria have not been previously evaluated as potential COPs removal agents. The aim of this study was to assess the ability of Lactobacillus casei ATCC334 to remove COPs in aqueous solution. Results showed the ability of both growing and resting cells to remove COPs (ca. 30-60%). All COPs-bacterium interactions were specific and partly reversible, being resting cells the most efficient for COPs removal in a ranking order of 7-KC > 7α-OH/7β-OH > triol > 5,6β-EP > 5,6α-EP > 25-OH. Binding to the cell wall and/or cell membrane incorporation appears to be the most likely mechanisms involved on COPs removal by L. casei ATCC 334. PMID:23848962

  1. Composition of the carbohydrate granules of the cyanobacterium, Cyanothece sp. strain ATCC 51142

    NASA Technical Reports Server (NTRS)

    Schneegurt, M. A.; Sherman, D. M.; Sherman, L. A.; Mitchell, C. A. (Principal Investigator)

    1997-01-01

    Cyanothece sp. strain ATCC 51142 is an aerobic, unicellular, diazotrophic cyanobacterium that temporally separates O2-sensitive N2 fixation from oxygenic photosynthesis. The energy and reducing power needed for N2 fixation appears to be generated by an active respiratory apparatus that utilizes the contents of large interthylakoidal carbohydrate granules. We report here on the carbohydrate and protein composition of the granules of Cyanothece sp. strain ATCC 51142. The carbohydrate component is a glucose homopolymer with branches every nine residues and is chemically identical to glycogen. Granule-associated protein fractions showed temporal changes in the number of proteins and their abundance during the metabolic oscillations observed under diazotrophic conditions. There also were temporal changes in the protein pattern of the granule-depleted supernatant fractions from diazotrophic cultures. None of the granule-associated proteins crossreacted with antisera directed against several glycogen-metabolizing enzymes or nitrogenase, although these proteins were tentatively identified in supernatant fractions. It is suggested that the granule-associated proteins are structural proteins required to maintain a complex granule architecture.

  2. Isolation and Characterization of a Novel Virulent Phage of Lactobacillus casei ATCC 393.

    PubMed

    Zhang, Xi; Lan, Yu; Jiao, Wenchao; Li, Yijing; Tang, Lijie; Jiang, Yanping; Cui, Wen; Qiao, Xinyuan

    2015-12-01

    A new virulent phage (Lcb) of Lactobacillus casei ATCC 393 was isolated from Chinese sauerkraut. It was specific to L. casei ATCC 393. Electron micrograph revealed that it had an icosahedral head (60.2 ± 0.8 nm in diameter) and a long tail (251 ± 2.6 nm). It belonged to the Siphoviridae family. The genome of phage Lcb was estimated to be approximately 40 kb and did not contain cohesive ends. One-step growth kinetics of its lytic development revealed latent and burst periods of 75 and 45 min, respectively, with a burst size of 16 PFU per infected cell. The phage was able to survive in a pH range between 4 and 11. However, a treatment of 70 °C for 30 min and 75% ethanol or isopropanol for 20 min was observed to inactivate phage Lcb thoroughly. The presence of both Ca(2+) and Mg(2+) showed a little influence on phage adsorption, but they were indispensable to gain complete lysis and improve plaque formation. The adsorption kinetics were similar on viable or nonviable cells, and high adsorption rates maintained between 10 and 37 °C. The highest adsorption rate was at 30 °C. This study increased the knowledge on phages of L. casei. The characterization of phage Lcb is helpful to establish a basis for adopting effective strategies to control phage attack in industry. PMID:26123178

  3. Heterologous expression of Streptomyces clavuligerus ATCC 27064 cephamycin C gene cluster.

    PubMed

    Martínez-Burgo, Y; Álvarez-Álvarez, R; Pérez-Redondo, R; Liras, P

    2014-09-30

    The Streptomyces clavuligerus cephamycin C gene cluster has been subcloned in a SuperCos-derived cosmid and introduced in Streptomyces flavogriseus ATCC 33331, Streptomyces coelicolor M1146 and Streptomyces albus J1074. The exconjugant strains were supplemented with an additional copy of the S. clavuligerus cephamycin regulatory activator gene, ccaRC, expressed from the constitutive Pfur promoter. Only S. flavogriseus-derived exconjugants produced a compound active against Escherichia coli ESS22-31 that was characterized by HPLC-MS as cephamycin C. The presence of an additional ccaR copy resulted in about 40-fold increase in cephamycin C production. Optimal heterologous cephamycin C production was in the order of 9% in relation to that of S. clavuligerus ATCC 27064. RT-qPCR studies indicated that ccaRC expression in S. flavogriseus::[SCos-CF] was 7% of that in S. clavuligerus and increased to 47% when supplemented with a copy of Pfur-ccaR. The effect on cephamycin biosynthesis gene expression was thus improved but not in an uniform manner for every gene. In heterologous strains, integration of the cephamycin cluster results in a ccaR-independent increased resistance to penicillin, cephalosporin and cefoxitin, what corresponds well to the strong expression of the pcbR and pbpA genes in S. flavogriseus-derived strains. PMID:24975573

  4. Next-generation sequencing-based transcriptome analysis of L-lysine-producing Corynebacterium glutamicum ATCC 21300 strain.

    PubMed

    Kim, Hong-Il; Nam, Jae-Young; Cho, Jae-Yong; Lee, Chang-Soo; Park, Young-Jin

    2013-12-01

    In the present study, 151 genes showed a significant change in their expression levels in Corynebacterium glutamicum ATCC 21300 compared with those of C. glutamicum ATCC 13032. Of these 151 genes, 56 genes (2%) were up-regulated and 95 genes (3%) were down-regulated. RNA sequencing analysis also revealed that 11 genes, involved in the L-lysine biosynthetic pathway of C. glutamicum, were up- or down-regulated compared with those of C. glutamicum ATCC 13032. Of the 151 genes, 10 genes were identified to have mutations including SNP (9 genes) and InDel (1 gene). This information will be useful for genome breeding of C. glutamicum to develop an industrial amino acid-producing strain with minimal mutation. PMID:24385368

  5. High-Quality Draft Genome Sequence of Aneurinibacillus migulanus ATCC 9999T (DSM 2895), a Gramicidin S-Producing Bacterium Isolated from Garden Soil

    PubMed Central

    Wang, Jie-ping; Liu, Guo-hong; Ge, Ci-bin; Xiao, Rong-feng; Zheng, Xue-fang; Shi, Huai

    2015-01-01

    Aneurinibacillus migulanus ATCC 9999T (DSM 2895) is a Gram-positive, round-spore-forming, and gramicidin S-producing bacterium. Here, we report the 6.35-Mb high-quality draft genome sequence of A. migulanus ATCC 9999T, which will provide useful information for the genomic taxonomy and phylogenomics of Bacillus-like bacteria. PMID:26494674

  6. High-Quality Draft Genome Sequence of Aneurinibacillus migulanus ATCC 9999T (DSM 2895), a Gramicidin S-Producing Bacterium Isolated from Garden Soil.

    PubMed

    Wang, Jie-Ping; Liu, Bo; Liu, Guo-Hong; Ge, Ci-Bin; Xiao, Rong-Feng; Zheng, Xue-Fang; Shi, Huai

    2015-01-01

    Aneurinibacillus migulanus ATCC 9999(T) (DSM 2895) is a Gram-positive, round-spore-forming, and gramicidin S-producing bacterium. Here, we report the 6.35-Mb high-quality draft genome sequence of A. migulanus ATCC 9999(T), which will provide useful information for the genomic taxonomy and phylogenomics of Bacillus-like bacteria. PMID:26494674

  7. Effect of continuous light on diurnal rhythms in Cyanothece sp. ATCC 51142

    PubMed Central

    Elvitigala, Thanura; Stöckel, Jana; Ghosh, Bijoy K; Pakrasi, Himadri B

    2009-01-01

    Background Life on earth is strongly affected by alternating day and night cycles. Accordingly, many organisms have evolved an internal timekeeping system with a period of approximately 24 hours. Cyanobacteria are the only known prokaryotes with robust rhythms under control of a central clock. Numerous studies have been conducted to elucidate components of the circadian clock and to identify circadian-controlled genes. However, the complex interactions between endogenous circadian rhythms and external cues are currently not well understood, and a direct and mathematical based comparison between light-mediated and circadian-controlled gene expression is still outstanding. Therefore, we combined and analyzed data from two independent microarray experiments, previously performed under alternating light-dark and continuous light conditions in Cyanothece sp. ATCC 51142, and sought to classify light responsive and circadian controlled genes. Results Fourier Score-based methods together with random permutations and False Discovery Rates were used to identify genes with oscillatory expression patterns, and an angular distance based criterion was applied to recognize transient behaviors in gene expression under constant light conditions. Compared to previously reported mathematical approaches, the combination of these methods also facilitated the detection of modified amplitudes and phase-shifts of gene expression. Our analysis showed that the majority of diurnally regulated genes, essentially those genes that are maximally expressed during the middle of the light and dark period, are in fact light responsive. In contrast, most of the circadian controlled genes are up-regulated during the beginning of the dark or subjective dark, and are greatly enriched for genes associated with energy metabolism. Many of the circadian controlled and light responsive genes are found in gene clusters within the Cyanothece sp. ATCC 51142 genome. Interestingly, in addition to cyclic

  8. Adh enhances Actinobacillus pleuropneumoniae pathogenicity by binding to OR5M11 and activating p38 which induces apoptosis of PAMs and IL-8 release

    PubMed Central

    Wang, Lei; Qin, Wanhai; Zhang, Jing; Bao, Chuntong; Zhang, Hu; Che, Yanyi; Sun, Changjiang; Gu, Jingmin; Feng, Xin; Du, Chongtao; Han, Wenyu; Richard, Paul Langford; Lei, Liancheng

    2016-01-01

    Members of the Trimeric Autotransporter Adhesin (TAA) family play a crucial role in the adhesion of Gram-negative pathogens to host cells, but the immunopathogenesis of TAAs remains unknown. Our previous studies demonstrated that Adh from Actinobacillus pleuropneumoniae (A. pleuropneumoniae) is required for full bacterial pathogenicity. Alveolar macrophages are the first line of defense against respiratory infections. This study compared the interactions between porcine alveolar macrophages (PAMs) and wild-type A. pleuropneumoniae (5b WT) or an Adh-deletion strain (5b ΔAdh) via gene microarray, immunoprecipitation and other technologies. We found that Adh was shown to interact with the PAMs membrane protein OR5M11, an olfactory receptor, resulting in the high-level secretion of IL-8 by activation of p38 MAPK signaling pathway. Subsequently, PAMs apoptosis via the activation of the Fax and Bax signaling pathways was observed, followed by activation of caspases 8, 9, and 3. The immunological pathogenic roles of Adh were also confirmed in both murine and piglets infectious models in vivo. These results identify a novel immunological strategy for TAAs to boost the pathogenicity of A. pleuropneumoniae. Together, these datas reveal the high versatility of the Adh protein as a virulence factor and provide novel insight into the immunological pathogenic role of TAAs. PMID:27046446

  9. Pharmacokinetic/pharmacodynamic evaluation of marbofloxacin in the treatment of Haemophilus parasuis and Actinobacillus pleuropneumoniae infections in nursery and fattener pigs using Monte Carlo simulations.

    PubMed

    Vilalta, C; Giboin, H; Schneider, M; El Garch, F; Fraile, L

    2014-12-01

    This study evaluated the theoretical clinical outcome of three marbofloxacin posology regimens in two groups of pigs (weaners and fatteners) for the treatment of Actinobacillus pleuropneumoniae (App) and Haemophilus parasuis (Hp) infection and the appearance of resistant bacteria due to the antibiotic treatment. The probability of target attainment (PTA) for pharmacokinetic/pharmacodynamics (PK/PD) ratios associated with clinical efficacy and with the appearance of antimicrobial resistance for fluoroquinolones at each minimum inhibitory concentration (MIC) or mutant prevention concentration (MPC) were calculated, respectively. The cumulative fraction of response (CFR) was calculated for the three posology regimens against App and they ranged from 91.12% to 96.37% in weaners and from 93% to 97.43% in fatteners, respectively. In the case of Hp, they ranged from 80.52% to 85.14% in weaners and from 82.01% to 88.49% in fatteners, respectively. Regarding the PTA of the PK/PD threshold associated with the appearance of antimicrobial resistance, results showed that marbofloxacin would prevent resistances in most of the animals up to the MPC value of 1 μg/mL. PMID:24903473

  10. Identification and preliminary characterization of a 75-kDa hemin- and hemoglobin-binding outer membrane protein of Actinobacillus pleuropneumoniae serotype 1.

    PubMed

    Archambault, Marie; Labrie, Josée; Rioux, Clément R; Dumas, France; Thibault, Pierre; Elkins, Christopher; Jacques, Mario

    2003-10-01

    The reference strains representing serotypes 1 to 12 of Actinobacillus pleuropneumoniae biotype 1 were examined for their ability to utilize porcine hemoglobin (Hb) or porcine hemin (Hm) as iron sources for growth. In a growth promotion assay, all of the reference strains were able to use porcine Hb, and all strains except 2 were able to use porcine Hm. Using a preliminary characterization procedure with Hm- or Hb-agarose, Hm- and Hb-binding outer membrane proteins (OMPs) of approximately 75 kDa were isolated from A. pleuropneumoniae serotype 1 strain 4074 grown under iron-restricted conditions. Matrix-assisted laser desorption ionization/time-of-flight (MALDI-TOF) analysis revealed a number of common tryptic peptides between the Hb-agarose- and Hm-agarose-purified 75 kDa OMPs, strongly suggesting that these peptides originate from the same protein. A database search of these peptide sequences revealed identities with proteins from various Gram-negative bacteria, including iron-regulated OMPs, transporter proteins, as well as TonB-dependent receptors. Taken together, our data suggest that A. pleuropneumoniae synthesizes potential Hm- and Hb-binding proteins that could be implicated in the iron uptake from porcine Hb and Hm. PMID:14620863

  11. Nasal immunization with M cell-targeting ligand-conjugated ApxIIA toxin fragment induces protective immunity against Actinobacillus pleuropneumoniae infection in a murine model.

    PubMed

    Park, Jisang; Seo, Ki-Weon; Kim, Sae-Hae; Lee, Ha-Yan; Kim, Bumseok; Lim, Chae Woong; Kim, Jin-Hee; Yoo, Han Sang; Jang, Yong-Suk

    2015-05-15

    Actinobacillus pleuropneumoniae is the causative agent of porcine pleuropneumonia and severe economic loss in the swine industry has been caused by the infection. Therefore, the development of an effective vaccine against the bacteria is necessary. ApxII toxin, among several virulence factors expressed by the bacteria, is considered to be a promising vaccine candidate because ApxII toxin not only accompanies cytotoxic and hemolytic activities, but is also expressed in all 15 serotypes of bacteria except serotypes 10 and 14. In this study, we identified the peptide ligand capable of targeting the ligand-conjugated ApxIIA #5 fragment antigen to nasopharynx-associated lymphoid tissue. It was found that nasal immunization with ligand-conjugated ApxIIA #5 induced efficient mucosal and systemic immune responses measured at the levels of antigen-specific antibodies, cytokine-secreting cells after antigen exposure, and antigen-specific lymphocyte proliferation. More importantly, the nasal immunization induced protective immunity against nasal challenge infection of the bacteria, which was confirmed by histopathological studies and bacterial clearance after challenge infection. Collectively, we confirmed that the ligand capable of targeting the ligand-conjugated antigen to nasopharynx-associated lymphoid tissue can be used as an effective nasal vaccine adjuvant to induce protective immunity against A. pleuropneumoniae infection. PMID:25818577

  12. Lactobacillus reuteri ATCC 55730 and L22 display probiotic potential in vitro and protect against Salmonella-induced pullorum disease in a chick model of infection.

    PubMed

    Zhang, Dexian; Li, Rui; Li, Jichang

    2012-08-01

    Lactobacillus reuteri ATCC 55730 (L. reuteri ATCC 55730) and L. reuteri L22 were studied for their probiotic potential. These two strains were able to produce an antimicrobial substance, termed reuterin, the maximum production of reuterin by these two strains was detected in the late logarithmic growth phase (16 h in MRS and 20 h in LB broths). These two strains could significantly reduce the growth of Salmonella pullorum ATCC 9120 in MRS broth, L. reuteri ATCC 55730 with a reduction of 48.2±4.15% (in 5 log) and 89.7±2.59% (in 4 log) respectively, at the same time, L. reuteri L22 was 69.4±3.48% (in 5 log) and 80.4±3.22% respectively. L. reuteri ATCC 55730 was active against the majority of the pathogenic species, including S. pullorum ATCC 9120 and Escherichia coli O(78), while L. reuteri L22 was not as effective as L. reuteri ATCC 55730. The two potential strains were found to survive variably at pH 2.5 and were unaffected by bile salts, while neither of the strains was haemolytic. Moreover, L. reuteri ATCC 55730 exhibited variable susceptibility towards commonly used antibiotics; but L. reuteri L22 showed resistant to most antibiotics in this study. L. reuteri ATCC 55730 consequently was found to significantly increase survival rate in a Salmonella-induced pullorum disease model in chick. To conclude, strain L. reuteri ATCC 55730 possesses desirable probiotic properties, such as antimicrobial activity and immunomodulation in vitro, which were confirmed in vivo by the use of animal models. PMID:21764090

  13. Pore-forming ability of major outer membrane proteins from Wolinella recta ATCC 33238.

    PubMed Central

    Kennell, W L; Egli, C; Hancock, R E; Holt, S C

    1992-01-01

    Three major outer membrane proteins with apparent molecular masses of 43, 45, and 51 kDa were purified from Wolinella recta ATCC 33238, and their pore-forming abilities were determined by the black lipid bilayer method. The non-heat-modifiable 45-kDa protein (Omp 45) showed no pore-forming activity even at high KCl concentrations. The single-channel conductances in 1 M KCl of the heat-modifiable proteins with apparent molecular masses of 43 kDa (Omp 43) and 51 kDa (Omp 51) were 0.49 and 0.60 nS, respectively. The proteins formed nonselective channels and, as determined by experiments of ion selectivity and zero-current potential, were weakly anion selective. Images PMID:1370429

  14. Effects of Salt Stress on Carbohydrate Metabolism of Lactobacillus plantarum ATCC 14917.

    PubMed

    Wang, Pingping; Wu, Zhen; Wu, Jing; Pan, Daodong; Zeng, Xiaoqun; Cheng, Kemeng

    2016-10-01

    Lactic acid bacteria are widely used in fermented foods, especially cheese products. In this study, we observed the salt tolerance of Lactobacillus plantarum ATCC 14917 after exposure to different concentrations of NaCl in MRS medium. Quantitative proteomic profiles using two-dimensional electrophoresis identified 384 proteins, of which 26 were upregulated and 31 downregulated. Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry was then used to identify 11 proteins, of which three were linked to carbohydrate metabolism. The downregulation of carbamoyl phosphate synthase in carbohydrate metabolism revealed a bacterial regulation mechanism to save energy in order to survive during the salt tolerance. Other proteins were found involved in transcription-translation processes, fatty acid biosynthesis, and the primary metabolic process. PMID:27342422

  15. Ala(0)-actagardine, a new lantibiotic from cultures of Actinoplanes liguriae ATCC 31048.

    PubMed

    Vértesy, L; Aretz, W; Bonnefoy, A; Ehlers, E; Kurz, M; Markus, A; Schiell, M; Vogel, M; Wink, J; Kogler, H

    1999-08-01

    The actagardine-producing strain Actinoplanes liguriae ATCC 31048, forms an additional lantibiotic when it is cultured on mannitol and soya meal. The new compound, Ala(0)-actagardine (1), has been isolated by solid-phase extraction followed by a two-step chromatographic separation. The molecular formula of 1 is C84H129N21O25S4. Its chemical structure was determined by 2D-NMR analysis and was further confirmed by an amino acid analysis, Edman degradation, and partial synthesis from actagardine. 1 exhibits a slightly higher biological activity than the parent compound actagardine. The synthetic analogs Lys(0)-actagardine (2) and Ile(0)-actagardine (3) demonstrate also antibacterial activities and emphasize the importance of the N-terminus for further derivatization. PMID:10580386

  16. Characterization of the pyoverdines of Azotobacter vinelandii ATCC 12837 with regard to heterogeneity.

    PubMed

    Menhart, N; Thariath, A; Viswanatha, T

    1991-01-01

    Azotobacter vinelandii strain ATCC 12837 produces peptide siderophores of the general class known as pyoverdines. In the past, it was assumed that a single well-defined pyoverdine was produced by each parent microorganism. However, there are a number of reports of incompletely characterized pyoverdines that demonstrate heterogeneity in pyoverdine preparations obtained from a single organism, but the nature of this phenomena has not been explained. This study shows that A. vinelandii does indeed produce more than one pyoverdine and that these compounds differ in their peptide components. The metabolism of these siderophores suggests that only one of them is a true siderophore while the others are metabolic byproducts. It was demonstrated that this phenomenon is likely due to intrinsic limitations of the synthetase complex involved in the biosynthesis of these compounds. Characterization of two of the major pyoverdines produced demonstrated that they are novel compounds, although they belonged to the Azotobacter-type family of pyoverdines. PMID:1838001

  17. Pullulan Production by Aureobasidium pullulans ATCC 201253 Cells Adsorbed onto Cellulose Anion and Cation Exchangers

    PubMed Central

    West, Thomas P.

    2012-01-01

    The anion exchanger phosphocellulose and the cation exchanger triethylaminoethyl cellulose were used to immobilize cells of the fungus Aureobasidium pullulans ATCC 201253 and the adsorbed cells were subsequently investigated for their ability to produce the polysaccharide pullulan using batch fermentation. The cells adsorbed on the triethylaminoethyl cellulose at pH 7.5 produced higher pullulan levels than those cells immobilized on phosphocellulose at pH 4.0 for 2 cycles of 168 h at 30 °C. Relative to the initial cycle of 168 h, pullulan production by the cells immobilized on the triethylaminoethyl cellulose decreased slightly after 168 h of the second production cycle while pullulan production by the phosphocellulose-immobilized cells remained about the same after 168 h of the second production cycle. PMID:23762749

  18. Isolation of a mutant strain of Pseudomonas sp ATCC 31461 exhibiting elevated polysaccharide production.

    PubMed

    West, T P

    2002-10-01

    A mutant strain of the bacterium Pseudomonas sp. ATCC 31461 that exhibited elevated production of the polysaccharide gellan on glucose or corn syrup as a carbon source was isolated. Gellan production by the mutant strain was about twofold higher than its parent strain on glucose or corn syrup after 48 h of growth, and about 1.4-fold higher after 72 h. An increase in biomass production was not correlated with enhanced gellan synthesis by the mutant strain. The increased gellan production by the mutant strain on either carbon source resulted in an increase in its culture medium viscosity and the viscosity of the isolated polysaccharide produced by glucose-grown cells. No differences in the glucuronic acid content of the polysaccharides produced by the mutant and parent strains were observed. PMID:12355317

  19. Response of electrically stimulated cells of Pseudomonas oleovorans strain ATCC 29347 suspended in silicone oil.

    PubMed

    Anglade, J; Hirschler, A; Le Petit, J; Matheron, R; Scarpitta, A; Iacazio, G

    2001-05-15

    A high intensity direct current was applied for more than 10 min onto a bacterial suspension of Pseudomonas oleovorans ATCC 29347 suspended in silicone oil. The application of a gradually increased electric field from 0 to 2500 V x cm(-1) resulted in a decrease of the optical density of the bacterial suspension and the occurrence of a peak current of several hundred microA for living cells instead of a linear increase (few microA) for killed or lyophilised cells. This procedure is not only a rapid way of investigating the living state of cell cultures but also an efficient experimental tool to study the cellular effects of a controlled electrical stress. PMID:11356578

  20. Desulfurization and denitrogenation of heavy gas oil by Rhodococcus erythropolis ATCC 4277.

    PubMed

    Maass, D; Todescato, D; Moritz, D E; Oliveira, J Vladimir; Oliveira, D; Ulson de Souza, A A; Guelli Souza, S M A

    2015-08-01

    Some of the noxious atmospheric pollutants such as nitrogen and sulfur dioxides come from the fossil fuel combustion. Biodesulfurization and biodenitrogenation are processes which remove those pollutants through the action of microorganisms. The ability of sulfur and nitrogen removal by the strain Rhodococcus erythropolis ATCC 4277 was tested in a biphasic system containing different heavy gas oil concentrations in a batch reactor. Heavy gas oil is an important fraction of petroleum, because after passing through, the vacuum distillation is incorporated into diesel oil. This strain was able to remove about 40% of the nitrogen and sulfur present in the gas heavy oil. Additionally, no growth inhibition occurred even when in the presence of pure heavy gas oil. Results present in this work are considered relevant for the development of biocatalytic processes for nitrogen and sulfur removal toward building feasible industrial applications. PMID:25759162

  1. 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.

  2. Biodegradation of bisphenol A and its halogenated analogues by Cunninghamella elegans ATCC36112.

    PubMed

    Keum, Young Soo; Lee, Hye Ri; Park, Hee Won; Kim, Jeong-Han

    2010-11-01

    Bisphenol A and its halogenated analogues are commonly used industrial chemicals with strong toxicological effects over many organisms. In this study, metabolic fate of bisphenol A and its halogenated analogues were evaluated with Cunninghamella elegans ATCC36112. Bisphenol A and related analogues were rapidly transformed into several metabolites by C. elegans within 2-4 days. Detailed analysis of metabolites reveals that both phase I and II metabolism occurred in C. elegans. Cytochrome P450-dependent hydroxylation was observed in BPA. However, major reaction with bisphenol A and analogues with 1-2 halogen atoms were the formation of glucose-conjugate, not being inhibited by cytochrome P450 inhibitor. Overall metabolic rates decreased with increasing number of substitution at 2- and 6-position of BPA structures, which may be consequences of limited bioavailability or steric hindrance to conjugate-forming reaction. Information from the current study will provide detailed insights over the fungal metabolism of BPA and analogues. PMID:20455075

  3. Nisin production from Lactococcus lactis A.T.C.C. 7962 using supplemented whey permeate.

    PubMed

    Flôres, S H; Alegre, R M

    2001-10-01

    The influence of pH control and aeration (20% dissolved oxygen) on nisin production in a supplemented cheese whey permeate was examined during batch fermentation with Lactococcus lactis subsp. lactis A.T.C.C. 7962. A maximum nisin activity of 5280 i.u./ml of medium was observed in the raw extract of nisin after 9 h of fermentation with a constant pH at 4.9. However, the fermentation was continued until 24 h, when a decrease in the nisin activity was observed. The pH control did not influence the nisin production and aeration of the culture medium increased cell growth (biomass) but not nisin activity. The yeast Kluyveromyces marxianus, used as an alternative method to control pH, has not been efficient. PMID:11592916

  4. Genome Assembly of Methicillin-Resistant Quality Control Strain Staphylococcus aureus CDC73-57501 (ATCC 29247).

    PubMed

    Daligault, H E; Davenport, K W; Minogue, T D; Bishop-Lilly, K A; Broomall, S M; Bruce, D C; Chain, P S; Coyne, S R; Freitas, T; Frey, K G; Gibbons, H S; Jaissle, J; Lo, C-C; Meincke, L; Munk, A C; Redden, C L; Rosenzweig, C N; Johnson, S L

    2014-01-01

    Staphylococcus aureus is a major cause of bacterial infections in the United States, with high percentages of serious infections resistant to a variety of β-lactam antibiotics. Here, we present the scaffolded genome assembly into 16 contigs of S. aureus CDC73-57501 (ATCC 29247), a methicillin-resistant quality control strain. PMID:25278527

  5. Genome Assembly of Methicillin-Resistant Quality Control Strain Staphylococcus aureus CDC73-57501 (ATCC 29247)

    PubMed Central

    Daligault, H. E.; Davenport, K. W.; Minogue, T. D.; Bishop-Lilly, K. A.; Broomall, S. M.; Bruce, D. C.; Chain, P. S.; Coyne, S. R.; Freitas, T.; Frey, K. G.; Gibbons, H. S.; Jaissle, J.; Lo, C.-C.; Meincke, L.; Munk, A. C.; Redden, C. L.; Rosenzweig, C. N.

    2014-01-01

    Staphylococcus aureus is a major cause of bacterial infections in the United States, with high percentages of serious infections resistant to a variety of β-lactam antibiotics. Here, we present the scaffolded genome assembly into 16 contigs of S. aureus CDC73-57501 (ATCC 29247), a methicillin-resistant quality control strain. PMID:25278527

  6. Complete Genome Sequence of Gluconacetobacter hansenii Strain NQ5 (ATCC 53582), an Efficient Producer of Bacterial Cellulose

    PubMed Central

    Pfeffer, Sarah; Mehta, Kalpa

    2016-01-01

    This study reports the release of the complete nucleotide sequence of Gluconacetobacter hansenii strain NQ5 (ATCC 53582). This strain was isolated by R. Malcolm Brown, Jr. in a sugar mill in North Queensland, Australia, and is an efficient producer of bacterial cellulose. The elucidation of the genome will contribute to the study of the molecular mechanisms necessary for cellulose biosynthesis. PMID:27516505

  7. A murine oral model for Mycobacterium avium subsp. paratuberculosis infection and immunomodulation with Lactobacillus casei ATCC 334.

    PubMed

    Cooney, Meagan A; Steele, James L; Steinberg, Howard; Talaat, Adel M

    2014-01-01

    Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) the causative agent of Johne's disease, is one of the most serious infectious diseases in dairy cattle worldwide. Due to the chronic nature of this disease and no feasible control strategy, it is essential to have an efficient animal model which is representative of the natural route of infection as well as a viable treatment option. In this report, we evaluated the effect of different doses of M. paratuberculosis in their ability to colonize murine tissues following oral delivery and the ability of Lactobacillus casei ATCC 334, a nascent probiotic, to combat paratuberculosis. Oral inoculation of mice was able to establish paratuberculosis in a dose-dependent manner. Two consecutive doses of approximately 10(9) CFU per mouse resulted in a disseminated infection, whereas lower doses were not efficient to establish infection. All inoculated mice were colonized with M. paratuberculosis, maintained infection for up to 24 weeks post infection and generated immune responses that reflect M. paratuberculosis infection in cattle. Notably, oral administration of L. casei ATCC 334 did not reduce the level of M. paratuberculosis colonization in treated animals. Interestingly, cytokine responses and histology indicated a trend for the immunomodulation and reduction of pathology in animals receiving L. casei ATCC 334 treatment. Overall, a reproducible oral model of paratuberculosis in mice was established that could be used for future vaccine experiments. Although the L. casei ATCC 334 was not a promising candidate for controlling paratuberculosis, we established a protocol to screen other probiotic candidates. PMID:24551602

  8. A murine oral model for Mycobacterium avium subsp. paratuberculosis infection and immunomodulation with Lactobacillus casei ATCC 334

    PubMed Central

    Cooney, Meagan A.; Steele, James L.; Steinberg, Howard; Talaat, Adel M.

    2014-01-01

    Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) the causative agent of Johne's disease, is one of the most serious infectious diseases in dairy cattle worldwide. Due to the chronic nature of this disease and no feasible control strategy, it is essential to have an efficient animal model which is representative of the natural route of infection as well as a viable treatment option. In this report, we evaluated the effect of different doses of M. paratuberculosis in their ability to colonize murine tissues following oral delivery and the ability of Lactobacillus casei ATCC 334, a nascent probiotic, to combat paratuberculosis. Oral inoculation of mice was able to establish paratuberculosis in a dose-dependent manner. Two consecutive doses of approximately 109 CFU per mouse resulted in a disseminated infection, whereas lower doses were not efficient to establish infection. All inoculated mice were colonized with M. paratuberculosis, maintained infection for up to 24 weeks post infection and generated immune responses that reflect M. paratuberculosis infection in cattle. Notably, oral administration of L. casei ATCC 334 did not reduce the level of M. paratuberculosis colonization in treated animals. Interestingly, cytokine responses and histology indicated a trend for the immunomodulation and reduction of pathology in animals receiving L. casei ATCC 334 treatment. Overall, a reproducible oral model of paratuberculosis in mice was established that could be used for future vaccine experiments. Although the L. casei ATCC 334 was not a promising candidate for controlling paratuberculosis, we established a protocol to screen other probiotic candidates. PMID:24551602

  9. Fe(III) stimulates 3-methylindole and 4-methylphenol production in swine lagoon enrichments and Clostridium scatologenes ATCC 25775

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aims: To determine the effects of anaerobic electron acceptors on 3-methylindole (3-MI) and 4-methylphenol (4-MP) production in swine waste lagoon enrichments and Clostridium scatologenes ATCC 25775. Methods and Results: Swine waste lagoon sediment was incubated anaerobically in tryptone-yeast ext...

  10. Genome Sequence of Streptococcus phocae subsp. phocae Strain ATCC 51973T Isolated from a Harbor Seal (Phoca vitulina)

    PubMed Central

    Poblete-Morales, Matías

    2015-01-01

    Streptococcus phocae subsp. phocae is a pathogen that affects different pinniped and mammalian species. This announcement reports the genome sequence of the type strain ATCC 51973 isolated in Norway from clinical specimens of harbor seal (Phoca vitulina), revealing interesting genes related to possible virulence factors. PMID:26586875

  11. Altered Composition of Ralstonia eutropha Poly(hydroxyalkanoate) through Expression of PHA Synthase from Allochromatium vinosum ATCC 35206

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The class III poly(hydroxyalkanoate) synthase (PHAS) genes (phaC and phaE) of a photosynthetic bacterium, Allochromatium vinosum ATCC 35206, were cloned, sequenced and expressed in a heterologous host. We employed a PCR technique coupled with a chromosomal gene-walking method to clone and subsequen...

  12. Construction of expression vectors for metabolic engineering of the vanillin-producing actinomycete Amycolatopsis sp. ATCC 39116.

    PubMed

    Fleige, Christian; Steinbüchel, Alexander

    2014-01-01

    Amycolatopsis sp. ATCC 39116 is able to synthesize the important flavoring agent vanillin from cheap natural substrates. The bacterium is therefore of great interest for the industry and used for the fermentative production of vanillin. In order to improve the production of natural vanillin with Amycolatopsis sp. ATCC 39116, the strain has been genetically engineered to optimize the metabolic flux towards the desired product. Extensive metabolic engineering was hitherto hampered, due to the lack of genetic tools like functional promoters and expression vectors. In this study, we report the establishment of a plasmid-based gene expression system for Amycolatopsis sp. ATCC 39116 that allows a further manipulation of the genotype. Four new Escherichia coli-Amycolatopsis shuttle vectors harboring different promoter elements were constructed, and the functionality of these regulatory elements was proven by the expression of the reporter gene gusA, encoding a β-glucuronidase. Glucuronidase activity was detected in all plasmid-harboring strains, and remarkable differences in the expression strength of the reporter gene depending on the used promoter were observed. The new expression vectors will promote the further genetic engineering of Amycolatopsis sp. ATCC 39116 to get insight into the metabolic network and to improve the strain for a more efficient industrial use. PMID:24743982

  13. Complete Genome Sequence of Gluconacetobacter hansenii Strain NQ5 (ATCC 53582), an Efficient Producer of Bacterial Cellulose.

    PubMed

    Pfeffer, Sarah; Mehta, Kalpa; Brown, R Malcolm

    2016-01-01

    This study reports the release of the complete nucleotide sequence of Gluconacetobacter hansenii strain NQ5 (ATCC 53582). This strain was isolated by R. Malcolm Brown, Jr. in a sugar mill in North Queensland, Australia, and is an efficient producer of bacterial cellulose. The elucidation of the genome will contribute to the study of the molecular mechanisms necessary for cellulose biosynthesis. PMID:27516505

  14. Effect of Lactobacillus brevis ATCC 8287 as a feeding supplement on the performance and immune function of piglets

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Lactobacillus brevis ATCC 8287, a surface (S-layer) strain, possesses a variety of functional properties that make it both a potential probiotic and a good vaccine vector candidate. With this in mind, our aim was to study the survival of L. brevis in the porcine gut and investigate the effect of th...

  15. Complete genome sequence of Streptomyces venezuelae ATCC 15439, a promising cell factory for production of secondary metabolites.

    PubMed

    Song, Ju Yeon; Yoo, Young Ji; Lim, Si-Kyu; Cha, Sun Ho; Kim, Ji-Eun; Roe, Jung-Hye; Kim, Jihyun F; Yoon, Yeo Joon

    2016-02-10

    Streptomyces venezuelae ATCC 15439, which produces 12- and 14-membered ring macrolide antibiotics, is a platform strain for heterologous expression of secondary metabolites. Its 9.05-Mb genome sequence revealed an abundance of genes involved in the biosynthesis of secondary metabolites and their precursors, which should be useful for the production of bioactive compounds. PMID:26718561

  16. Ca2+-Citrate Uptake and Metabolism in Lactobacillus casei ATCC 334

    PubMed Central

    Mortera, Pablo; Pudlik, Agata; Magni, Christian; Alarcón, Sergio

    2013-01-01

    The putative citrate metabolic pathway in Lactobacillus casei ATCC 334 consists of the transporter CitH, a proton symporter of the citrate-divalent metal ion family of transporters CitMHS, citrate lyase, and the membrane-bound oxaloacetate decarboxylase complex OAD-ABDH. Resting cells of Lactobacillus casei ATCC 334 metabolized citrate in complex with Ca2+ and not as free citrate or the Mg2+-citrate complex, thereby identifying Ca2+-citrate as the substrate of the transporter CitH. The pathway was induced in the presence of Ca2+ and citrate during growth and repressed by the presence of glucose and of galactose, most likely by a carbon catabolite repression mechanism. The end products of Ca2+-citrate metabolism by resting cells of Lb. casei were pyruvate, acetate, and acetoin, demonstrating the activity of the membrane-bound oxaloacetate decarboxylase complex OAD-ABDH. Following pyruvate, the pathway splits into two branches. One branch is the classical citrate fermentation pathway producing acetoin by α-acetolactate synthase and α-acetolactate decarboxylase. The other branch yields acetate, for which the route is still obscure. Ca2+-citrate metabolism in a modified MRS medium lacking a carbohydrate did not significantly affect the growth characteristics, and generation of metabolic energy in the form of proton motive force (PMF) was not observed in resting cells. In contrast, carbohydrate/Ca2+-citrate cometabolism resulted in a higher biomass yield in batch culture. However, also with these cells, no generation of PMF was associated with Ca2+-citrate metabolism. It is concluded that citrate metabolism in Lb. casei is beneficial when it counteracts acidification by carbohydrate metabolism in later growth stages. PMID:23709502

  17. Ca2+-citrate uptake and metabolism in Lactobacillus casei ATCC 334.

    PubMed

    Mortera, Pablo; Pudlik, Agata; Magni, Christian; Alarcón, Sergio; Lolkema, Juke S

    2013-08-01

    The putative citrate metabolic pathway in Lactobacillus casei ATCC 334 consists of the transporter CitH, a proton symporter of the citrate-divalent metal ion family of transporters CitMHS, citrate lyase, and the membrane-bound oxaloacetate decarboxylase complex OAD-ABDH. Resting cells of Lactobacillus casei ATCC 334 metabolized citrate in complex with Ca(2+) and not as free citrate or the Mg(2+)-citrate complex, thereby identifying Ca(2+)-citrate as the substrate of the transporter CitH. The pathway was induced in the presence of Ca(2+) and citrate during growth and repressed by the presence of glucose and of galactose, most likely by a carbon catabolite repression mechanism. The end products of Ca(2+)-citrate metabolism by resting cells of Lb. casei were pyruvate, acetate, and acetoin, demonstrating the activity of the membrane-bound oxaloacetate decarboxylase complex OAD-ABDH. Following pyruvate, the pathway splits into two branches. One branch is the classical citrate fermentation pathway producing acetoin by α-acetolactate synthase and α-acetolactate decarboxylase. The other branch yields acetate, for which the route is still obscure. Ca(2+)-citrate metabolism in a modified MRS medium lacking a carbohydrate did not significantly affect the growth characteristics, and generation of metabolic energy in the form of proton motive force (PMF) was not observed in resting cells. In contrast, carbohydrate/Ca(2+)-citrate cometabolism resulted in a higher biomass yield in batch culture. However, also with these cells, no generation of PMF was associated with Ca(2+)-citrate metabolism. It is concluded that citrate metabolism in Lb. casei is beneficial when it counteracts acidification by carbohydrate metabolism in later growth stages. PMID:23709502

  18. Cloning and characterization of the genes encoding nitrilotriacetate monooxygenase of Chelatobacter heintzii ATCC 29600.

    PubMed Central

    Knobel, H R; Egli, T; van der Meer, J R

    1996-01-01

    A 6.2-kb DNA fragment containing the genes for the nitrilotriacetate (NTA) monooxygenase of Chelatobacter heintzii ATCC 29600 was cloned and characterized by DNA sequencing and expression studies. The nucleotide sequence contained three major open reading frames (ORFs). Two of the ORFs, which were oriented divergently with an intergenic region of 307 bp, could be assigned to the NTA monooxygenase components A and B. The predicted N-terminal amino acid sequences of these ORFs were identical with those determined for the purified components. We therefore named these genes ntaA (for component A of NTA monooxygenase) and ntaB (for component B). The ntaA and ntaB genes could be expressed in Escherichia coli DH5alpha, and the gene products were visualized after Western blotting (immunoblotting) and incubation with polyclonal antibodies against component A or B. By mixing overproduced NtaB from E. coli and purified component A from C. heintzii ATCC 29600, reconstitution of a functional NTA monooxygenase complex was possible. The deduced gene product of ntaA showed only significant homology to SoxA (involved in dibenzothiophene degradation) and to SnaA (involved in pristamycin synthesis); that of ntaB shared weak homologies in one domain with other NADH:flavine mononucleotide oxidoreductases. These homologies provide no conclusive answer as to the possible evolutionary origin of the NTA monooxygenase. The deduced gene product of the third ORF (ORF1) had homology in the N-terminal region with the GntR class of bacterial regulator proteins and therefore may encode a regulator protein, possibly involved in regulation of ntaA and ntaB expression. PMID:8892809

  19. Enhancement of linear gramicidin expression from Bacillus brevis ATCC 8185 by casein peptide.

    PubMed

    Nakai, Tomonori; Yamauchi, Daisuke; Kubota, Kou

    2005-04-01

    Bacillus brevis (Brevibacillus parabrevis) ATCC 8185 synthesizes two kinds of antibiotic peptides, cyclopeptide tyrocidine and linear gramicidin. The production of linear gramicidin can be induced by the standard method (using a skim milk medium for pre-culture and beef broth for the main culture) employed for the induction of tyrocidine. In this study, we tried to determine the optimal growth medium for B. brevis ATCC 8185 for synthesizing linear gramicidin. The yield of linear gramicidin produced by the standard method was 3.11 microg/ml. When beef broth was used both as the pre-medium and the main medium, the yield of the antibiotic was only 0.59 microg/ml. To confirm the influence of skim milk, the strain was grown in a 1% skim milk medium. As a result, the amount of linear gramicidin produced reached 20.3 microg/ml. These findings show the importance of skim milk in the production of linear gramicidin. In the skim milk medium, the cells produced an extracellular protease 2 h before the linear gramicidin was expressed. The 1% skim milk medium pretreated by this protease did not allow the induction of linear gramicidin into the cells, and protease activity was not detected in the supernatant of the culture. When the cells were cultivated in a 1% egg albumin medium, protease activity from the supernatant of the culture was detected, but production of linear gramicidin was not observed. Therefore, a 1% casein medium was used for production of linear gramicidin. As a result, the yield of linear gramicidin produced in the medium reached 6.69 microg/ml. We concluded that a digested product of the extracellular protease from casein enhances linear gramicidin production. PMID:15849407

  20. Meta-analysis: Lactobacillus reuteri strain DSM 17938 (and the original strain ATCC 55730) for treating acute gastroenteritis in children.

    PubMed

    Szajewska, H; Urbańska, M; Chmielewska, A; Weizman, Z; Shamir, R

    2014-09-01

    Lactobacillus reuteri ATCC 55730 has been shown to provide a moderate clinical effect in the treatment of acute gastroenteritis (AGE) in children. However, as the L. reuteri ATCC 55730 strain was found to carry potentially transferable resistance traits for tetracycline and lincomycin, it was replaced by a new strain, L. reuteri DSM 17938, without unwanted plasmid-borne antibiotic resistance. Bioequivalence of the two strains has been suggested. We aimed to systematically evaluate data on the effectiveness of L. reuteri DSM 17938 and the original strain, L. reuteri ATCC 55730, in the treatment of AGE in children. The Cochrane Library, MEDLINE, and EMBASE databases, reference lists, and abstract books of major scientific meetings were searched in August 2013, with no language restrictions, for relevant randomised controlled trials (RCTs). Two RCTs (n=196) that evaluated L. reuteri DSM 17938 and three RCTs (n=156) that evaluated L. reuteri ATCC 55730, which involved hospitalised children aged 3 to 60 months, met the inclusion criteria. Compared with placebo or no treatment, DSM 17938 significantly reduced the duration of diarrhoea (mean difference -32 h, 95% confidence interval (CI): -41 to -24) and increased the chance of cure on day 3 (relative risk: 3.5, 95% CI: 1.2 to 10.8, random effects model). Similar results were obtained with the original strain, L. reuteri ATCC 55730. In conclusion, in hospitalised children, use of both strains of L. reuteri reduced the duration of diarrhoea, and more children were cured within 3 days. Data from outpatients and countryspecific cost-effectiveness analyses are needed. Given the limited data and the methodological limitations of the included trials, the evidence should be viewed with caution. PMID:24463209

  1. 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. PMID

  2. Investigation of the Amycolatopsis sp. Strain ATCC 39116 Vanillin Dehydrogenase and Its Impact on the Biotechnical Production of Vanillin

    PubMed Central

    Fleige, Christian; Hansen, Gunda; Kroll, Jens

    2013-01-01

    The actinomycete Amycolatopsis sp. strain ATCC 39116 is capable of synthesizing large amounts of vanillin from ferulic acid, which is a natural cell wall component of higher plants. The desired intermediate vanillin is subject to undesired catabolism caused by the metabolic activity of a hitherto unknown vanillin dehydrogenase (VDHATCC 39116). In order to prevent the oxidation of vanillin to vanillic acid and thereby to obtain higher yields and concentrations of vanillin, the responsible vanillin dehydrogenase in Amycolatopsis sp. ATCC 39116 was investigated for the first time by using data from our genome sequence analysis and further bioinformatic approaches. The vdh gene was heterologously expressed in Escherichia coli, and the encoded vanillin dehydrogenase was characterized in detail. VDHATCC 39116 was purified to apparent electrophoretic homogeneity and exhibited NAD+-dependent activity toward vanillin, coniferylaldehyde, cinnamaldehyde, and benzaldehyde. The enzyme showed its highest level of activity toward vanillin at pH 8.0 and at a temperature of 44°C. In a next step, a precise vdh deletion mutant of Amycolatopsis sp. ATCC 39116 was generated. The mutant lost its ability to grow on vanillin and did not show vanillin dehydrogenase activity. A 2.3-times-higher vanillin concentration and a substantially reduced amount of vanillic acid occurred with the Amycolatopsis sp. ATCC 39116 Δvdh::Kmr mutant when ferulic acid was provided for biotransformation in a cultivation experiment on a 2-liter-bioreactor scale. Based on these results and taking further metabolic engineering into account, the Amycolatopsis sp. ATCC 39116 Δvdh::Kmr mutant represents an optimized and industrially applicable platform for the biotechnological production of natural vanillin. PMID:23064333

  3. Heterologous expression of endo-beta-1,4-D-glucanase from Clostridium cellulovorans in Clostridium acetobutylicum ATCC 824 following transformation of the engB gene.

    PubMed Central

    Kim, A Y; Attwood, G T; Holt, S M; White, B A; Blaschek, H P

    1994-01-01

    Heterologous expression of the Clostridium cellulovorans engB gene by Clostridium acetobutylicum BKW-1 was detected as zones of hydrolysis on carboxymethyl cellulose (CMC) Trypticase glucose yeast plates stained with Congo red. The extracellular cellulase preparation from C. acetobutylicum BKW-1 has a specific activity towards CMC which is more than fourfold that present in C. acetobutylicum ATCC 824. Western blot (immunoblot) analysis using the C. cellulovorans anti-EngB primary antibody demonstrated that an additional 44-kDa protein band was present in the supernatant derived from C. acetobutylicum BKW-1 but was not present in ATCC 824 or ATCC 824(pMTL500E). Images PMID:8117087

  4. Two Master Switch Regulators Trigger A40926 Biosynthesis in Nonomuraea sp. Strain ATCC 39727

    PubMed Central

    Lo Grasso, Letizia; Maffioli, Sonia; Sosio, Margherita; Bibb, Mervyn; Puglia, Anna Maria

    2015-01-01

    ABSTRACT The actinomycete Nonomuraea sp. strain ATCC 39727 produces the glycopeptide A40926, the precursor of dalbavancin. Biosynthesis of A40926 is encoded by the dbv gene cluster, which contains 37 protein-coding sequences that participate in antibiotic biosynthesis, regulation, immunity, and export. In addition to the positive regulatory protein Dbv4, the A40926-biosynthetic gene cluster encodes two additional putative regulators, Dbv3 and Dbv6. Independent mutations in these genes, combined with bioassays and liquid chromatography-mass spectrometry (LC-MS) analyses, demonstrated that Dbv3 and Dbv4 are both required for antibiotic production, while inactivation of dbv6 had no effect. In addition, overexpression of dbv3 led to higher levels of A40926 production. Transcriptional and quantitative reverse transcription (RT)-PCR analyses showed that Dbv4 is essential for the transcription of two operons, dbv14-dbv8 and dbv30-dbv35, while Dbv3 positively controls the expression of four monocistronic transcription units (dbv4, dbv29, dbv36, and dbv37) and of six operons (dbv2-dbv1, dbv14-dbv8, dbv17-dbv15, dbv21-dbv20, dbv24-dbv28, and dbv30-dbv35). We propose a complex and coordinated model of regulation in which Dbv3 directly or indirectly activates transcription of dbv4 and controls biosynthesis of 4-hydroxyphenylglycine and the heptapeptide backbone, A40926 export, and some tailoring reactions (mannosylation and hexose oxidation), while Dbv4 directly regulates biosynthesis of 3,5-dihydroxyphenylglycine and other tailoring reactions, including the four cross-links, halogenation, glycosylation, and acylation. IMPORTANCE This report expands knowledge of the regulatory mechanisms used to control the biosynthesis of the glycopeptide antibiotic A40926 in the actinomycete Nonomuraea sp. strain ATCC 39727. A40926 is the precursor of dalbavancin, approved for treatment of skin infections by Gram-positive bacteria. Therefore, understanding the regulation of its biosynthesis

  5. Azotobacter Genomes: The Genome of Azotobacter chroococcum NCIMB 8003 (ATCC 4412)

    PubMed Central

    Robson, Robert L.; Jones, Robert; Robson, R. Moyra; Schwartz, Ariel; Richardson, Toby H.

    2015-01-01

    The genome of the soil-dwelling heterotrophic N2-fixing Gram-negative bacterium Azotobacter chroococcum NCIMB 8003 (ATCC 4412) (Ac-8003) has been determined. It consists of 7 circular replicons totalling 5,192,291 bp comprising a circular chromosome of 4,591,803 bp and six plasmids pAcX50a, b, c, d, e, f of 10,435 bp, 13,852, 62,783, 69,713, 132,724, and 311,724 bp respectively. The chromosome has a G+C content of 66.27% and the six plasmids have G+C contents of 58.1, 55.3, 56.7, 59.2, 61.9, and 62.6% respectively. The methylome has also been determined and 5 methylation motifs have been identified. The genome also contains a very high number of transposase/inactivated transposase genes from at least 12 of the 17 recognised insertion sequence families. The Ac-8003 genome has been compared with that of Azotobacter vinelandii ATCC BAA-1303 (Av-DJ), a derivative of strain O, the only other member of the Azotobacteraceae determined so far which has a single chromosome of 5,365,318 bp and no plasmids. The chromosomes show significant stretches of synteny throughout but also reveal a history of many deletion/insertion events. The Ac-8003 genome encodes 4628 predicted protein-encoding genes of which 568 (12.2%) are plasmid borne. 3048 (65%) of these show > 85% identity to the 5050 protein-encoding genes identified in Av-DJ, and of these 99 are plasmid-borne. The core biosynthetic and metabolic pathways and macromolecular architectures and machineries of these organisms appear largely conserved including genes for CO-dehydrogenase, formate dehydrogenase and a soluble NiFe-hydrogenase. The genetic bases for many of the detailed phenotypic differences reported for these organisms have also been identified. Also many other potential phenotypic differences have been uncovered. Properties endowed by the plasmids are described including the presence of an entire aerobic corrin synthesis pathway in pAcX50f and the presence of genes for retro-conjugation in pAcX50c. All these

  6. Actinoplanes teichomyceticus ATCC 31121 as a cell factory for producing teicoplanin

    PubMed Central

    2011-01-01

    Background Teicoplanin is a glycopeptide antibiotic used clinically in Europe and in Japan for the treatment of multi-resistant Gram-positive infections. It is produced by fermenting Actinoplanes teichomyceticus. The pharmaceutically active principle is teicoplanin A2, a complex of compounds designated T-A2-1-A2-5 differing in the length and branching of the fatty acid moiety linked to the glucosamine residue on the heptapeptide scaffold. According to European and Japanese Pharmacopoeia, components of the drug must be reproduced in fixed amounts to be authorized for clinical use. Results We report our studies on optimizing the fermentation process to produce teicoplanin A2 in A. teichomyceticus ATCC 31121. Robustness of the process was assessed on scales from a miniaturized deep-well microtiter system to flasks and 3-L bioreactor fermenters. The production of individual factors T-A2-1-A2-5 was modulated by adding suitable precursors to the cultivation medium. Specific production of T-A2-1, characterized by a linear C10:1 acyl moiety, is enhanced by adding methyl linoleate, trilinoleate, and crude oils such as corn and cottonseed oils. Accumulation of T-A2-3, characterized by a linear C10:0 acyl chain, is stimulated by adding methyl oleate, trioleate, and oils such as olive and lard oils. Percentages of T-A2-2, T-A2-4, and, T-A2-5 bearing the iso-C10:0, anteiso-C11:0, and iso-C11:0 acyl moieties, respectively, are significantly increased by adding precursor amino acids L-valine, L-isoleucine, and L-leucine. Along with the stimulatory effect on specific complex components, fatty acid esters, oils, and amino acids (with the exception of L-valine) inhibit total antibiotic productivity overall. By adding industrial oils to medium containing L-valine the total production is comparable, giving unusual complex compositions. Conclusions Since the cost and the quality of teicoplanin production depend mainly on the fermentation process, we developed a robust and scalable

  7. 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

  8. Comparative activities of selected fluoroquinolones against dynamic populations of Actinobacillus pleuropneumoniae in an in vitro model of time-kill continuous culture experiment.

    PubMed

    Damte, Dereje; Lee, Seung-Jin; Yohannes, Sileshi B; Hossain, Md Akil; Suh, Joo-Won; Park, Seung-Chun

    2013-12-01

    The aim of the current study was to demonstrate and compare the impact of different pharmacokinetics of marbofloxacin, enrofloxacin and difloxacin on their antimicrobial effects, their killing and re-growth kinetics, and the population dynamics of Actinobacillus pleuropneumoniae clinical isolates in an in vitro dynamic model. Selected clinical isolates of A. pleuropneumoniae and three fluoroquinolones at a range of simulated AUC(24)/MIC ratios of multiple doses were investigated. At the same simulated AUC(24)/MIC ratios of the three fluoroquinolones, the killing re-growth profile and I(E) values (intensity of the antimicrobial effect) revealed strain- and fluoroquinolone-specific effects. For example, a 31% lower I(E) of difloxacin was observed in AppK5 (biofilm-former) than in AppK2 (biofilm-non-former) at the same AUC(24)/MIC ratio of 120 h. In addition, losses in A. pleuropneumoniae susceptibility of both strains by the three fluoroquinolones were observed. AUC(24)/MPC ratios of 20.89 and 39.81 for marbofloxacin, 17.32 and 19.49 for enrofloxacin and 31.62 and 60.25 for difloxacin were estimated to be protective against the selection of AppK2 and AppK5 strain mutants, respectively. Integration of these in vitro data with published pharmacokinetics revealed the inadequacy of the conventional clinical doses of the three drugs to attain the above protective values for minimum biofilm eradication concentration (MBEC) and concentration to prevent growth of 90% of the mutant subpopulation (MPC(90)). In conclusion, the results suggest optimising doses could suffice for resistant mutants control, while for biofilm-forming strains combination with biofilm-disrupting agents to reduce the MBEC to achieve AUC/MBEC ratios within the possible dosing regimens is desired. PMID:24139884

  9. Nasal immunization with mannan-decorated mucoadhesive HPMCP microspheres containing ApxIIA toxin induces protective immunity against challenge infection with Actinobacillus pleuropneumoiae in mice.

    PubMed

    Li, Hui-Shan; Shin, Min-Kyoung; Singh, Bijay; Maharjan, Sushila; Park, Tae-Eun; Kang, Sang-Kee; Yoo, Han-Sang; Hong, Zhong-Shan; Cho, Chong-Su; Choi, Yun-Jaie

    2016-07-10

    The development of subunit mucosal vaccines requires an appropriate delivery system or an immune modulator such as an adjuvant to improve antigen immunogenicity. The nasal route for vaccine delivery by microparticles has attracted considerable interest, although challenges such as the rapid mucociliary clearance in the respiratory mucosa and the low immunogenicity of subunit vaccine still remain. Here, we aimed to develop mannan-decorated mucoadhesive thiolated hydroxypropylmethyl cellulose phthalate (HPMCP) microspheres (Man-THM) that contain ApxIIA subunit vaccine - an exotoxin fragment as a candidate for a subunit nasal vaccine against Actinobacillus pleuropneumoniae. For adjuvant activity, mucoadhesive thiolated HPMCP microspheres decorated with mannan could be targeted to the PRRs (pathogen recognition receptors) and mannose receptors (MR) of antigen presenting cells (APCs) in the respiratory immune system. The potential adjuvant ability of Man-THM for intranasal immunization was confirmed by in vitro and in vivo experiments. In a mechanistic study using APCs in vitro, it was found that Man-THM enhanced receptor-mediated endocytosis by stimulating the MR of APCs. In vivo, the nasal vaccination of ApxIIA-loaded Man-THM in mice resulted in higher levels of mucosal sIgA and serum IgG than mice in the ApxIIA and ApxIIA-loaded THM groups due to the specific recognition of the mannan in the Man-THM by the MRs of the APCs. Moreover, ApxIIA-containing Man-THM protected immunized mice when challenged with strains of A. pleuropneumoniae serotype 5. These results suggest that mucoadhesive Man-THM may be a promising candidate for a nasal vaccine delivery system to elicit systemic and mucosal immunity that can protect from pathogenic bacteria infection. PMID:27189136

  10. Improved Multiplex PCR Using Conserved and Species-Specific 16S rRNA Gene Primers for Simultaneous Detection of Actinobacillus actinomycetemcomitans, Bacteroides forsythus, and Porphyromonas gingivalis

    PubMed Central

    Tran, Simon Dangtuan; Rudney, Joel D.

    1999-01-01

    Among putative periodontal pathogens, Actinobacillus actinomycetemcomitans, Bacteroides forsythus, and Porphyromonas gingivalis are most convincingly implicated as etiological agents in periodontitis. Therefore, techniques for detection of those three species would be of value. We previously published a description of a multiplex PCR that detects A. actinomycetemcomitans and P. gingivalis. The present paper presents an improvement on that technique, which now allows more sensitive detection of all three periodontal pathogens. Sensitivity was determined by testing serial dilutions of A. actinomycetemcomitans, B. forsythus, and P. gingivalis cells. Primer specificity was tested against (i) all gene sequences from the GenBank-EMBL database, (ii) six A. actinomycetemcomitans, one B. forsythus, and four P. gingivalis strains, (iii) eight different species of oral bacteria, and (iv) supra- and subgingival plaque samples from 20 healthy subjects and subgingival plaque samples from 10 patients with periodontitis. The multiplex PCR had a detection limit of 10 A. actinomycetemcomitans, 10 P. gingivalis, and 100 B. forsythus cells. Specificity was confirmed by the fact that (i) none of our forward primers were homologous to the 16S rRNA genes of other oral species, (ii) amplicons of predicted size were detected for all A. actinomycetemcomitans, B. forsythus, and P. gingivalis strains tested, and (iii) no amplicons were detected for the eight other bacterial species. A. actinomycetemcomitans, B. forsythus, and P. gingivalis were detected in 6 of 20, 1 of 20, and 11 of 20 of supragingival plaque samples, respectively, and 4 of 20, 7 of 20, and 13 of 20 of subgingival plaque samples, respectively, from periodontally healthy subjects. Among patients with periodontitis, the organisms were detected in 7 of 10, 10 of 10, and 7 of 10 samples, respectively. The simultaneous detection of three periodontal pathogens is an advantage of this technique over conventional PCR assays. PMID

  11. The type strain of Lactobacillus casei is ATCC 393, ATCC 334 cannot serve as the type because it represents a different taxon, the name Lactobacillus paracasei and its subspecies names are not rejected and the revival of the name 'Lactobacillus zeae' contravenes Rules 51b (1) and (2) of the International Code of Nomenclature of Bacteria. Opinion 82.

    PubMed

    2008-07-01

    The Judicial Commission affirms that typification of Lactobacillus casei is based on ATCC 393, that ATCC 334 is a member of a different taxon and that the publication rejecting the name Lactobacillus paracasei (and its included subspecies) together with the revival of the name 'Lactobacillus zeae' contravenes Rules 51b (1) and (2) of the International Code of Nomenclature of Bacteria. PMID:18599731

  12. An ATP-grasp ligase involved in the last biosynthetic step of the iminomycosporine shinorine in Nostoc punctiforme ATCC 29133.

    PubMed

    Gao, Qunjie; Garcia-Pichel, Ferran

    2011-11-01

    We investigated the genetic basis for mycosporine sunscreen biosynthesis by the cyanobacterium Nostoc punctiforme ATCC 29133. Heterologous expression in Escherichia coli of three contiguous N. punctiforme genes (NpR5600, NpR5599, and NpR5598, here named mysA, mysB, and mysC, respectively) led to the production of mycosporine-glycine, an oxomycosporine. Additional expression of gene NpF5597 (mysD) led to the conversion of mycosporine-glycine into iminomycosporines (preferentially shinorine but also others like mycosporine-2-glycine and porphyra-334). This represents a new mode of enzymatic synthesis for iminomycosporines, one that differs in genetic origin, mechanism, and apparent substrate specificity from that known in Anabaena variabilis ATCC 29413. These results add to the emerging profile of the protein family of ATP-dependent ligases, to which the mysC product belongs, as important condensation enzymes in microbial secondary metabolism. PMID:21890703

  13. Isolation and characterisation of dipeptidyl peptidase IV from Prevotella loescheii ATCC 15930.

    PubMed

    Koreeda, Y; Hayakawa, M; Ikemi, T; Abiko, Y

    2001-08-01

    A proline-specific dipeptidyl aminopeptidase, dipeptidyl peptidase IV (EC 3.4.14.5), was purified from a cell sonicate soluble fraction of Prevotella loescheii ATCC 15930 by sequential column chromatography. The molecular mass of the native enzyme was estimated as 160 kDa by high-pressure liquid gel filtration column chromatography and unheated sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The subunit molecular mass was 80 kDa when the enzyme was heated to 100 degrees C in the presence of 2-mercaptoethanol before SDS-PAGE, suggesting that the native enzyme consists of two identical subunits and is folded in 2% SDS. The optimum pH, with glycyl-prolyl-4-methyl-coumaryl-7-amide as the substrate, was 8.0; the isoelectric point was 5.2. Purified enzyme showed a strong preference for dipeptide substrates containing proline and, less efficiently, alanine in the P1 position. The enzyme was markedly inhibited by Cd(2+), Zn(2+), Hg(2+), Co(2+), and serine proteinase inhibitor di-isopropylfluorophosphate. PMID:11389867

  14. Cloning, expression and bioinformatics analysis of ATP sulfurylase from Acidithiobacillus ferrooxidans ATCC 23270 in Escherichia coli

    PubMed Central

    Jaramillo, Michael L; Abanto, Michel; Quispe, Ruth L; Calderón, Julio; del Valle, Luís J; Talledo, Miguel; Ramírez, Pablo

    2012-01-01

    Molecular studies of enzymes involved in sulfite oxidation in Acidithiobacillus ferrooxidans have not yet been developed, especially in the ATP sulfurylase (ATPS) of these acidophilus tiobacilli that have importance in biomining. This enzyme synthesizes ATP and sulfate from adenosine phosphosulfate (APS) and pyrophosphate (PPi), final stage of the sulfite oxidation by these organisms in order to obtain energy. The atpS gene (1674 bp) encoding the ATPS from Acidithiobacillus ferrooxidans ATCC 23270 was amplified using PCR, cloned in the pET101-TOPO plasmid, sequenced and expressed in Escherichia coli obtaining a 63.5 kDa ATPS recombinant protein according to SDS-PAGE analysis. The bioinformatics and phylogenetic analyses determined that the ATPS from A. ferrooxidans presents ATP sulfurylase (ATS) and APS kinase (ASK) domains similar to ATPS of Aquifex aeolicus, probably of a more ancestral origin. Enzyme activity towards ATP formation was determined by quantification of ATP formed from E. coli cell extracts, using a bioluminescence assay based on light emission by the luciferase enzyme. Our results demonstrate that the recombinant ATP sulfurylase from A. ferrooxidans presents an enzymatic activity for the formation of ATP and sulfate, and possibly is a bifunctional enzyme due to its high homology to the ASK domain from A. aeolicus and true kinases. PMID:23055613

  15. Identification of galacto-N-biose phosphorylase from Clostridium perfringens ATCC13124.

    PubMed

    Nakajima, Masahiro; Nihira, Takanori; Nishimoto, Mamoru; Kitaoka, Motomitsu

    2008-03-01

    Lacto-N-biose phosphorylase (LNBP) from bifidobacteria is involved in the metabolism of lacto-N-biose I (Galbeta1-->3GlcNAc, LNB) and galacto-N-biose (Galbeta1-->3GalNAc, GNB). A homologous gene of LNBP (CPF0553 protein) was identified in the genome of Clostridium perfringens ATCC13124, which is a gram-positive anaerobic intestinal bacterium. In the present study, we cloned the gene and compared the substrate specificity of the CPF0553 protein with LNBP from Bifidobacterium longum JCM1217 (LNBPBl). In the presence of alpha-galactose 1-phosphate (Gal 1-P) as a donor, the CPF0553 protein acted only on GlcNAc and GalNAc, and GalNAc was a more effective acceptor than GlcNAc. The reaction product from GlcNAc/GalNAc and Gal 1-P was identified as LNB or GNB. The CPF0553 protein also phosphorolyzed GNB much faster than LNB, which suggests that the protein should be named galacto-N-biose phosphorylase (GNBP). GNBP showed a kcat/Km value for GNB that was approximately 50 times higher than that for LNB, whereas LNBPBl showed similar kcat/Km values for both GNB and LNB. Because C. perfringens possesses a gene coding endo-alpha-N-acetylgalactosaminidase, GNBP may play a role in the intestinal residence by metabolizing GNB that is available as a mucin core sugar. PMID:18183385

  16. Analysis of the mechanism and regulation of lactose transport and metabolism in Clostridium acetobutylicum ATCC 824.

    PubMed

    Yu, Yang; Tangney, Martin; Aass, Hans C; Mitchell, Wilfrid J

    2007-03-01

    Although the acetone-butanol-ethanol fermentation of Clostridium acetobutylicum is currently uneconomic, the ability of the bacterium to metabolize a wide range of carbohydrates offers the potential for revival based on the use of cheap, low-grade substrates. We have investigated the uptake and metabolism of lactose, the major sugar in industrial whey waste, by C. acetobutylicum ATCC 824. Lactose is taken up via a phosphoenolpyruvate-dependent phosphotransferase system (PTS) comprising both soluble and membrane-associated components, and the resulting phosphorylated derivative is hydrolyzed by a phospho-beta-galactosidase. These activities are induced during growth on lactose but are absent in glucose-grown cells. Analysis of the C. acetobutylicum genome sequence identified a gene system, lacRFEG, encoding a transcriptional regulator of the DeoR family, IIA and IICB components of a lactose PTS, and phospho-beta-galactosidase. During growth in medium containing both glucose and lactose, C. acetobutylicum exhibited a classical diauxic growth, and the lac operon was not expressed until glucose was exhausted from the medium. The presence upstream of lacR of a potential catabolite responsive element (cre) encompassing the transcriptional start site is indicative of the mechanism of carbon catabolite repression characteristic of low-GC gram-positive bacteria. A pathway for the uptake and metabolism of lactose by this industrially important organism is proposed. PMID:17209069

  17. Heterologous expression and localization of gentisate transporter Ncg12922 from Corynebacterium glutamicum ATCC 13032

    SciTech Connect

    Xu Ying; Yan Dazhong; Zhou Ningyi . E-mail: n.zhou@pentium.whiov.ac.cn

    2006-07-28

    Ralstonia sp. strain U2 metabolizes naphthalene via gentisate (2,5-dihydroxybenzoate) to central metabolites, but it was found unable to utilize gentisate as growth substrate. A putative gentisate transporter encoded by ncg12922 from Corynebacterium glutamicum ATCC 13032 was functionally expressed in Ralstonia sp. strain U2, converting strain U2 to a gentisate utilizer. After ncg12922 was inserted into plasmid pGFPe with green fluorescence protein gene gfp, the expressed fusion protein Ncg12922-GFP could be visualized in the periphery of Escherichia coli cells under confocal microscope, consistent with a cytoplasmic membrane location. In contrast, GFP was ubiquitous in the cytoplasm of E. coli cells carrying pGFPe only. Gentisate 1,2-dioxygenase activity was present in the cell extract from strain U2 induced with gentisate but at a much lower level (one-fifth) than that obtained with salicylate. However, it exhibited a similar level in strain U2 containing Ncg12922 induced either by salicylate or gentisate.

  18. Metabolic engineering of Corynebacterium glutamicum ATCC13032 to produce S-adenosyl-L-methionine.

    PubMed

    Han, Guoqiang; Hu, Xiaoqing; Qin, Tianyu; Li, Ye; Wang, Xiaoyuan

    2016-02-01

    As an important biological methyl group donor, S-adenosyl-L-methionine is used as nutritional supplement or drug for various diseases, but bacterial strains that can efficiently produce S-adenosyl-L-methionine are not available. In this study, Corynebacterium glutamicum strain HW104 which can accumulate S-adenosyl-L-methionine was constructed from C. glutamicum ATCC13032 by deleting four genes thrB, metB, mcbR and Ncgl2640, and six genes metK, vgb, lysC(m), hom(m), metX and metY were overexpressed in HW104 in different combinations, forming strains HW104/pJYW-4-metK-vgb, HW104/pJYW-4-SAM2C-vgb, HW104/pJYW-4-metK-vgb-metYX, and HW104/pJYW-4-metK-vgb-metYX-hom(m)-lysC(m). Fermentation experiments showed that HW104/pJYW-4-metK-vgb produced more S-adenosyl-L-methionine than other strains, and the yield achieved 196.7 mg/L (12.15 mg/g DCW) after 48h. The results demonstrate the potential application of C. glutamicum for production of S-adenosyl-L-methionine without addition of L-methionine. PMID:26777246

  19. Biosurfactant Production by Cultivation of Bacillus atrophaeus ATCC 9372 in Semidefined Glucose/Casein-Based Media

    NASA Astrophysics Data System (ADS)

    Das Neves, Luiz Carlos Martins; de Oliveira, Kátia Silva; Kobayashi, Márcio Junji; Vessoni Penna, Thereza Christina; Converti, Attilio

    Biosurfactants are proteins with detergent, emulsifier, and antimicrobial actions that have potential application in environmental applications such as the treatment of organic pollutants and oil recovery. Bacillus atrophaeus strains are nonpathogenic and are suitable source of biosurfactants, among which is surfactin. The aim of this work is to establish a culture medium composition able to stimulate biosurfactants production by B. atrophaeus ATCC 9372. Batch cultivations were carried out in a rotary shaker at 150 rpm and 35°C for 24 h on glucose- and/or casein-based semidefined culture media also containing sodium chloride, dibasic sodium phosphate, and soy flour. The addition of 14.0 g/L glucose in a culture medium containing 10.0 g/L of casein resulted in 17 times higher biosurfactant production (B max=635.0 mg/L). Besides, the simultaneous presence of digested casein (10.0 g/L), digested soy flour (3.0 g/L), and glucose (18.0 g/L) in the medium was responsible for a diauxic effect during cell growth. Once the diauxie started, the average biosurfactants concentration was 16.8% less than that observed before this phenomenon. The capability of B. atrophaeus strain to adapt its own metabolism to use several nutrients as energy sources and to preserve high levels of biosurfactants in the medium during the stationary phase is a promising feature for its possible application in biological treatments.

  20. Production of Surfactant from Bacillus subtilis ATCC 21332 using Potato substrates

    SciTech Connect

    Fox, Sandra Lynn; Bala, Greg Alan

    2000-12-01

    Surfactin, a lipopeptide biosurfactant, produced by Bacillus subtilis is known to reduce the surface tension of water from 72 to 27 mN/m. Potato substrates were evaluated as a carbon source for surfactant production by B. subtilis ATCC 21332. An established potato medium, simulated liquid and solid potato waste media, and a commercially prepared potato starch in a mineral salts medium were evaluated in shake flask experiments to verify growth, surface tension reduction, and carbohydrate reduction capabilities. Total carbohydrate assays and glucose monitoring indicated that B. subtilis was able to degrade potato substrates to produce surfactant. Surface tensions dropped from 71.3±0.1 to 28.3±0.3 mN/m (simulated solid potato medium) and to 27.5±0.3 mN/m (mineral salts medium). A critical micelle concentration (CMC) of 0.10 g/l was obtained from a methylene chloride extract of the simulated solid potato medium.

  1. A novel meta-cleavage product hydrolase from Flavobacterium sp. ATCC27551

    SciTech Connect

    Khajamohiddin, Syed; Babu, Pakala Suresh; Chakka, Deviprasanna; Merrick, Mike; Bhaduri, Anirban; Sowdhamini, Ramanathan; Siddavattam, Dayananda . E-mail: sdsl@uohyd.ernet.in

    2006-12-22

    The organophosphate degrading (opd) gene cluster of plasmid pPDL2 of Flavobacterium sp. ATCC27551 contains a novel open-reading frame, orf243. This was predicted to encode an {alpha}/{beta} hydrolase distantly related to the meta-fission product (MFP) hydrolases such as XylF, PhnD, and CumD. By homology modeling Orf243 has most of the structural features of MFP hydrolases including the characteristic active site catalytic triad. The purified protein (designated MfhA) is a homotetramer and shows similar affinity for 2-hydroxy-6-oxohepta-2,4-dienoate (HOHD), 2-hydroxymuconic semialdehyde (HMSA), and 2-hydroxy-5-methylmuconic semialdehyde (HMMSA), the meta-fission products of 3-methyl catechol, catechol, and 4-methyl catechol. The unique catalytic properties of MfhA and the presence near its structural gene of cis-elements required for transposition suggest that mfhA has evolved towards encoding a common hydrolase that can act on meta-fission products containing either aldehyde or ketone groups.

  2. Bismuth(III) deferiprone effectively inhibits growth of Desulfovibrio desulfuricans ATCC 27774.

    PubMed

    Barton, Larry L; Lyle, Daniel A; Ritz, Nathaniel L; Granat, Alex S; Khurshid, Ali N; Kherbik, Nada; Hider, Robert; Lin, Henry C

    2016-04-01

    Sulfate-reducing bacteria have been implicated in inflammatory bowel diseases and ulcerative colitis in humans and there is an interest in inhibiting the growth of these sulfide-producing bacteria. This research explores the use of several chelators of bismuth to determine the most effective chelator to inhibit the growth of sulfate-reducing bacteria. For our studies, Desulfovibrio desulfuricans ATCC 27774 was grown with nitrate as the electron acceptor and chelated bismuth compounds were added to test for inhibition of growth. Varying levels of inhibition were attributed to bismuth chelated with subsalicylate or citrate but the most effective inhibition of growth by D. desulfuricans was with bismuth chelated by deferiprone, 3-hydroxy-1,2-dimethyl-4(1H)-pyridone. Growth of D. desulfuricans was inhibited by 10 μM bismuth as deferiprone:bismuth with either nitrate or sulfate respiration. Our studies indicate deferiprone:bismuth has bacteriostatic activity on D. desulfuricans because the inhibition can be reversed following exposure to 1 mM bismuth for 1 h at 32 °C. We suggest that deferiprone is an appropriate chelator for bismuth to control growth of sulfate-reducing bacteria because deferiprone is relatively nontoxic to animals, including humans, and has been used for many years to bind Fe(III) in the treatment of β-thalassemia. PMID:26896170

  3. Biosurfactant-mediated biodegradation of straight and methyl-branched alkanes by Pseudomonas aeruginosa ATCC 55925

    PubMed Central

    2011-01-01

    Accidental oil spills and waste disposal are important sources for environmental pollution. We investigated the biodegradation of alkanes by Pseudomonas aeruginosa ATCC 55925 in relation to a rhamnolipid surfactant produced by the same bacterial strain. Results showed that the linear C11-C21 compounds in a heating oil sample degraded from 6% to 100%, whereas the iso-alkanes tended to be recalcitrant unless they were exposed to the biosurfactant; under such condition total biodegradation was achieved. Only the biodegradation of the commercial C12-C19 alkanes could be demonstrated, ranging from 23% to 100%, depending on the experimental conditions. Pristane (a C19 branched alkane) only biodegraded when present alone with the biosurfactant and when included in an artificial mixture even without the biosurfactant. In all cases the biosurfactant significantly enhanced biodegradation. The electron scanning microscopy showed that cells depicted several adaptations to growth on hydrocarbons, such as biopolymeric spheres with embedded cells distributed over different layers on the spherical surfaces and cells linked to each other by extracellular appendages. Electron transmission microscopy revealed transparent inclusions, which were associated with hydrocarbon based-culture cells. These patterns of hydrocarbon biodegradation and cell adaptations depended on the substrate bioavailability, type and length of hydrocarbon. PMID:21906343

  4. Proteome data to explore the impact of pBClin15 on Bacillus cereus ATCC 14579.

    PubMed

    Madeira, Jean-Paul; Alpha-Bazin, Béatrice; Armengaud, Jean; Omer, Hélène; Duport, Catherine

    2016-09-01

    This data article reports changes in the cellular and exoproteome of B. cereus cured from pBClin15.Time-course changes of proteins were assessed by high-throughput nanoLC-MS/MS. We report all the peptides and proteins identified and quantified in B. cereus with and without pBClin15. Proteins were classified into functional groups using the information available in the KEGG classification and we reported their abundance in term of normalized spectral abundance factor. The repertoire of experimentally confirmed proteins of B. cereus presented here is the largest ever reported, and provides new insights into the interplay between pBClin15 and its host B. cereus ATCC 14579. The data reported here is related to a published shotgun proteomics analysis regarding the role of pBClin15, "Deciphering the interactions between the Bacillus cereus linear plasmid, pBClin15, and its host by high-throughput comparative proteomics" Madeira et al. [1]. All the associated mass spectrometry data have been deposited in the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org) via the PRIDE partner repository (http://www.ebi.ac.uk/pride/), with the dataset identifier PRIDE: PXD001568, PRIDE: PXD002788 and PRIDE: PXD002789. PMID:27547804

  5. SpoIIE Regulates Sporulation but Does Not Directly Affect Solventogenesis in Clostridium acetobutylicum ATCC 824

    PubMed Central

    Scotcher, Miles C.; Bennett, George N.

    2005-01-01

    Using gene expression reporter vectors, we examined the activity of the spoIIE promoter in wild-type and spo0A-deleted strains of Clostridium acetobutylicum ATCC 824. In wild-type cells, the spoIIE promoter is active in a transient manner during late solventogenesis, but in strain SKO1, where the sporulation initiator spo0A is disrupted, no spoIIE promoter activity is detectable at any stage of growth. Strains 824(pMSpo) and 824(pASspo) were created to overexpress spoIIE and to decrease spoIIE expression via antisense RNA targeted against spoIIE, respectively. Some cultures of strains 824(pMSpo) degenerated during fermentations by losing the pSOL1 megaplasmid and hence did not produce the solvents ethanol, acetone, and butanol. The frequent degeneration event was shown to require an intact copy of spoIIE. Nondegenerate cultures of 824(pMSpo) exhibited normal growth and solvent production. Strain 824(pASspo) exhibited prolonged solventogenesis characterized by increased production of ethanol (225%), acetone (43%), and butanol (110%). Sporulation in strains harboring pASspo was significantly delayed, with sporulating cells exhibiting altered morphology. These results suggest that SpoIIE has no direct effect on the control of solventogenesis and that the changes in solvent production in spoIIE-downregulated cells are mediated by effects on the cell during sporulation. PMID:15743939

  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. Effect of Low Shear Modeled Microgravity (LSMMG) on the Probiotic Lactobacillus Acidophilus ATCC 4356

    NASA Technical Reports Server (NTRS)

    Stahl, S.; Voorhies, A.; Lorenzi, H.; Castro-Wallace, S.; Douglas, G.

    2016-01-01

    The introduction of generally recognized as safe (GRAS) probiotic microbes into the spaceflight food system has the potential for use as a safe, non-invasive, daily countermeasure to crew microbiome and immune dysregulation. However, the microgravity effects on the stress tolerances and genetic expression of probiotic bacteria must be determined to confirm translation of strain benefits and to identify potential for optimization of growth, survival, and strain selection for spaceflight. The work presented here demonstrates the translation of characteristics of a GRAS probiotic bacteria to a microgravity analog environment. Lactobacillus acidophilus ATCC 4356 was grown in the low shear modeled microgravity (LSMMG) orientation and the control orientation in the rotating wall vessel (RWV) to determine the effect of LSMMG on the growth, survival through stress challenge, and gene expression of the strain. No differences were observed between the LSMMG and control grown L. acidophilus, suggesting that the strain will behave similarly in spaceflight and may be expected to confer Earth-based benefits.

  8. Mutation of aspartic acid residues in the fructosyltransferase of Streptococcus salivarius ATCC 25975.

    PubMed Central

    Song, D D; Jacques, N A

    1999-01-01

    The site-directed mutated fructosyltransferases (Ftfs) of Streptococcus salivarius ATCC 25975, D312E, D312S, D312N and D312K were all active at 37 degrees C, indicating that Asp-312 present in the 'sucrose box' was not the nucleophilic Asp residue responsible for the formation of a covalent fructosyl-enzyme intermediate required for enzyme activity. Analysis of the kinetic constants of the purified mutated forms of the enzyme showed that Asp-312 was most likely an essential amino acid involved in determining acceptor recognition and/or stabilizing a beta-turn in the protein. In contrast, when the Asp-397 of the Ftf present in the conserved triplet RDP motif of all 60 bacterial and plant family-32 glycosylhydrolases was mutated to a Ser residue, both sucrose hydrolysis and polymerization ceased. Tryptophan emission spectra confirmed that this mutation did not alter protein structure. Comparison of published data from other site-directed mutated enzymes implicated the Asp residue in the RDP motif as the one that may form a transient covalent fructosyl intermediate during the catalysis of sucrose by the Ftf of S. salivarius. PMID:10548559

  9. Construction and Evaluation of a Clostridium thermocellum ATCC 27405 Whole-Genome Oligonucleotide Microarray

    NASA Astrophysics Data System (ADS)

    Brown, Steven D.; Raman, Babu; McKeown, Catherine K.; Kale, Shubha P.; He, Zhili; Mielenz, Jonathan R.

    Clostridium thermocellum is an anaerobic, thermophilic bacterium that can directly convert cellulosic substrates into ethanol. Microarray technology is a powerful tool to gain insights into cellular processes by examining gene expression under various physiological states. Oligonucleotide microarray probes were designed for 96.7% of the 3163 C. thermocellum ATCC 27405 candidate protein-encoding genes and then a partial-genome microarray containing 70 C. thermocellum specific probes was constructed and evaluated. We detected a signal-to-noise ratio of three with as little as 1.0 ng of genomic DNA and only low signals from negative control probes (nonclostridial DNA), indicating the probes were sensitive and specific. In order to further test the specificity of the array we amplified and hybridized 10 C. thermocellum polymerase chain reaction products that represented different genes and found gene specific hybridization in each case. We also constructed a whole-genome microarray and prepared total cellular RNA from the same point in early-logarithmic growth phase from two technical replicates during cellobiose fermentation. The reliability of the microarray data was assessed by cohybridization of labeled complementary DNA from the cellobiose fermentation samples and the pattern of hybridization revealed a linear correlation. These results taken together suggest that our oligonucleotide probe set can be used for sensitive and specific C. thermocellum transcriptomic studies in the future.

  10. Cloning, expression and characterization of a eukaryotic cycloalkanone monooxygenase from Cylindrocarpon radicicola ATCC 11011.

    PubMed

    Leipold, Friedemann; Wardenga, Rainer; Bornscheuer, Uwe T

    2012-05-01

    In this study, we have cloned and characterized a cycloalkanone monooxygenase (CAMO) from the ascomycete Cylindrocarpon radicicola ATCC 11011 (identical to Cylindrocarpon destructans DSM 837). The primary structure of this Baeyer-Villiger monooxygenase (BMVO) revealed 531 residues with around 45% sequence identity to known cyclohexanone monooxygenases. The enzyme was functionally overexpressed in Escherichia coli and investigated with respect to substrate spectrum and kinetic parameters. Substrate specificity studies revealed that a large variety of cycloaliphatic and bicycloaliphatic ketones are converted by this CAMO. A high catalytic efficiency against cyclobutanone was observed and seems to be a particular property of this BVMO. The thus produced butyrolactone derivatives are valuable building blocks for the synthesis of a variety of natural products and bioactive compounds. Furthermore, the enzyme revealed activity against open-chain ketones such as cyclobutyl, cyclopentyl and cyclohexyl methyl ketone which have not been reported to be accepted by typical cyclohexanone monooxygenases. These results suggest that the BVMO from C. radicicola indeed might be rather unique and since no BVMOs originating from eukaryotic organisms have been produced recombinantly so far, this study provides the first example for such an enzyme. PMID:22075635

  11. Modeling for Gellan Gum Production by Sphingomonas paucimobilis ATCC 31461 in a Simplified Medium

    PubMed Central

    Wang, Xia; Xu, Ping; Yuan, Yong; Liu, Changlong; Zhang, Dezhong; Yang, Zhengting; Yang, Chunyu; Ma, Cuiqing

    2006-01-01

    Gellan gum production was carried out by Sphingomonas paucimobilis ATCC 31461 in a simplified medium with a short incubation time, and a kinetic model for understanding, controlling, and optimizing the fermentation process was proposed. The results revealed that glucose was the best carbon source and that the optimal concentration was 30 g liter−1. As for the fermenting parameters, considerably large amounts of gellan gum were yielded by an 8-h-old culture and a 4% inoculum at 200 rpm on a rotary shaker. Under the optimized conditions, the maximum level of gellan gum (14.75 g liter−1) and the highest conversion efficiency (49.17%) were obtained in a 30-liter fermentor in batch fermentation. Logistic and Luedeking-Piret models were confirmed to provide a good description of gellan gum fermentation, which gave some support for the study of gellan gum fermentation kinetics. Additionally, this study is the first demonstration that gellan gum production is largely growth associated by analysis of kinetics in its batch fermentation process. Based on model prediction, higher gellan gum production (17.71 g liter−1) and higher conversion efficiency (57.12%) were obtained in fed-batch fermentation at the same total glucose concentration (30 g liter−1). PMID:16672479

  12. Modeling for gellan gum production by Sphingomonas paucimobilis ATCC 31461 in a simplified medium.

    PubMed

    Wang, Xia; Xu, Ping; Yuan, Yong; Liu, Changlong; Zhang, Dezhong; Yang, Zhengting; Yang, Chunyu; Ma, Cuiqing

    2006-05-01

    Gellan gum production was carried out by Sphingomonas paucimobilis ATCC 31461 in a simplified medium with a short incubation time, and a kinetic model for understanding, controlling, and optimizing the fermentation process was proposed. The results revealed that glucose was the best carbon source and that the optimal concentration was 30 g liter(-1). As for the fermenting parameters, considerably large amounts of gellan gum were yielded by an 8-h-old culture and a 4% inoculum at 200 rpm on a rotary shaker. Under the optimized conditions, the maximum level of gellan gum (14.75 g liter(-1)) and the highest conversion efficiency (49.17%) were obtained in a 30-liter fermentor in batch fermentation. Logistic and Luedeking-Piret models were confirmed to provide a good description of gellan gum fermentation, which gave some support for the study of gellan gum fermentation kinetics. Additionally, this study is the first demonstration that gellan gum production is largely growth associated by analysis of kinetics in its batch fermentation process. Based on model prediction, higher gellan gum production (17.71 g liter(-1)) and higher conversion efficiency (57.12%) were obtained in fed-batch fermentation at the same total glucose concentration (30 g liter(-1)). PMID:16672479

  13. Sulphate production by Paracoccus pantotrophus ATCC 35512 from different sulphur substrates: sodium thiosulphate, sulphite and sulphide.

    PubMed

    Meyer, Daniel Derrossi; Andrino, Felipe Gabriel; Possedente de Lira, Simone; Fornaro, Adalgiza; Corção, Gertrudes; Brandelli, Adriano

    2016-03-01

    One of the problems in waste water treatment plants (WWTPs) is the increase in emissions of hydrogen sulphide (H2S), which can cause damage to the health of human populations and ecosystems. To control emissions of this gas, sulphur-oxidizing bacteria can be used to convert H2S to sulphate. In this work, sulphate detection was performed by spectrophotometry, ion chromatography and atomic absorption spectrometry, using Paracoccus pantotrophus ATCC 35512 as a reference strain growing in an inorganic broth supplemented with sodium thiosulphate (Na2S2O3·5H2O), sodium sulphide (Na2S) or sodium sulphite (Na2SO3), separately. The strain was metabolically competent in sulphate production. However, it was only possible to observe significant differences in sulphate production compared to abiotic control when the inorganic medium was supplemented with sodium thiosulphate. The three methods for sulphate detection showed similar patterns, although the chromatographic method was the most sensitive for this study. This strain can be used as a reference for sulphate production in studies with sulphur-oxidizing bacteria originating from environmental samples of WWTPs. PMID:26269005

  14. Ca2+/calmodulin dependent protein kinase from Mycobacterium smegmatis ATCC 607.

    PubMed

    Sharma, S; Giri, S; Khuller, G K

    1998-06-01

    A soluble Ca2+/calmodulin dependent protein kinase has been partially purified (approximately 400 fold) from Mycobacterium smegmatis ATCC 607 using several purification steps like ammonium sulphate precipitation (30-60%), Sepharose CL-6B gel filtration, DEAE-cellulose and finally calmodulin-agarose affinity chromatography. On SDS-PAGE, this enzyme preparation showed a major protein band of molecular mass 35 kD and its activity was dependent on calcium, calmodulin and ATP when measured under saturating histone IIs (exogenous substrate) concentration. Phosphorylation of histone IIs was inhibited by W-7 (calmodulin inhibitor) and KN-62 (CaM-kinase inhibitor) with IC50 of 1.5 and 0.25 microm respectively, but was not affected by inhibitors of PKA (Sigma P5015) and PKC (H-7). All these results confirm that purified enzyme is Ca2+/calmodulin dependent protein kinase of M. smegmatis. The protein kinase of M. smegmatis demonstrated a narrow substrate specificity for both exogenous as well as endogenous substrates. These results suggest that purified CaM-kinase must be involved in regulating specific function(s) in this organism. PMID:9655195

  15. Production and structural analysis of the polysaccharide secreted by Trametes (Coriolus) versicolor ATCC 200801.

    PubMed

    Rau, Udo; Kuenz, Anja; Wray, Victor; Nimtz, Manfred; Wrenger, Julika; Cicek, Hasan

    2009-01-01

    Trametes versicolor ATCC 200801 secretes 4.1 g L(-1) of exopolysaccharide (EPS) when synthetic minimal medium and low-shear bioreactor cultivation technique are used. Structural and compositional analyses by thin layer chromatography, gas chromatography-mass spectrometry, electrospray ionization tandem mass spectrometry, and nuclear magnetic resonance spectroscopy yielded predominantly glucose and small amounts of galactose, mannose, arabinose, and xylose. The main EPS is composed of beta-1,3/beta-1,6-linked D-glucose molecules which is identical with Schizophyllan but does not possess a triple helical arrangement as secondary structure. Two molar mass fractions were detected by size exclusion chromatography yielding weight-average molecular weights of 4,100 and 2.6 kDa. Protein content varies between 2-3.6% (w/w). The exopolysaccharide is different in the nature of the glycosidic linkage, composition of monosaccharides, protein content, and weight-average molecular weight compared to the well-known polysaccharopeptide (PSP) and polysaccharopeptide Krestin (PSK). PMID:18800181

  16. Optimization of culture medium and conditions for penicillin acylase production by Streptomyces lavendulae ATCC 13664.

    PubMed

    Torres-Bacete, Jesús; Arroyo, Miguel; Torres-Guzmán, Raquel; De La Mata, Isabel; Acebal, Carmen; Castillón, M Pilar

    2005-08-01

    The culture medium for Streptomyces lavendulae ATCC 13664 was optimized on a shake-flask scale by using a statistical factorial design for enhanced production of penicillin acylase. This extracellular enzyme recently has been reported to be a penicillin K acylase, presenting also high hydrolytic activity against penicillin V and other natural aliphatic penicillins such as penicillin K, penicillin F, and penicillin dihydroF. The factorial design indicated that the main factors that positively affect penicillin acylase production by S. lavendulae were the concentration of yeast extract and the presence of oligoelements in the fermentation medium, whereas the presence of olive oil in the medium had no effect on enzyme production. An initial concentration of 2.5% (w/v) yeast extract and 3 microg/mL of CuSO4 x 5H2O was found to be best for acylase production. In such optimized culture medium, fermentation of the microorganism yielded 289 IU/L of enzyme in 72 h when employing a volume medium/volume flask ratio of 0.4 and a 300-rpm shaking speed. The presence of copper, alone and in combination with other metals, stimulated biomass as well as penicillin acylase production. The time course of penicillin acylase production was also studied in the optimized medium and conditions. Enzyme production showed catabolite repression by different carbon sources such as glucose, lactose, citrate, glycerol, and glycine. PMID:16118466

  17. Multi-functional glycoside hydrolase: Blon_0625 from Bifidobacterium longum subsp. infantis ATCC 15697.

    PubMed

    Matsumoto, Takuya; Shimada, Shota; Hata, Yuto; Tanaka, Tsutomu; Kondo, Akihiko

    2015-01-01

    We here describe a unique β-D-glucosidase (BGL; Blon_0625) derived from Bifidobacterium longum subsp. infantis ATCC 15697. The Blon_0625 gene was expressed by recombinant Escherichia coli. Purified recombinant Blon_0625 retains hydrolyzing activity against both p-nitrophenyl-β-D-glucopyranoside (pNPG; 17.3±0.24Umg(-1)) and p-nitrophenyl-β-D-xylopyranoside (pNPX; 16.7±0.32Umg(-1)) at pH 6.0, 30°C. To best of our knowledge, no previously described BGL retains the same level of both pNPGase and pNPXase activity. Furthermore, Blon_0625 also retains the activity against 4-nitrophenyl-α-l-arabinofranoside (pNPAf; 5.6±0.09Umg(-1)). In addition, the results of the degradation of phosphoric acid swollen cellulose (PASC) or xylan using endoglucanase from Thermobifida fusca YX (Tfu_0901) or xylanase from Kitasatospora setae KM-6054 (KSE_59480) show that Blon_0625 acts as a BGL and as a β-D-xylosidase (XYL) for hydrolyzing oligosaccharides. These results clearly indicate that Blon_0625 is a multi-functional glycoside hydrolase which retains the activity of BGL, XYL, and also α-l-arabinofuranosidase. Therefore, the utilization of multi-functional Blon_0625 may contribute to facilitating the efficient degradation of lignocellulosic materials and help enhance bioconversion processes. PMID:25435500

  18. Identification and characterization of the afsR homologue regulatory gene from Streptomyces peucetius ATCC 27952.

    PubMed

    Parajuli, Niranjan; Viet, Hung Trinh; Ishida, Kenji; Tong, Hang Thi; Lee, Hei Chan; Liou, Kwangkyoung; Sohng, Jae Kyung

    2005-01-01

    We have isolated an afsR homologue, called afsR-p, through genome analysis of Streptomyces peucetius ATCC 27952. AfsR-p shares 60% sequence identity with AfsR from Streptomyces coelicolor A3 (2). afsR-p was expressed under the control of the ermE* promoter in its hosts S. peucetius, Streptomyces lividans TK 24, Streptomyces clavuligerus and Streptomyces griseus. We observed overproduction of doxorubicin (4-fold) in S. peucetius, gamma-actinorhodin (2.6-fold) in S. lividans, clavulanic acid (1.5-fold) in S. clavuligerus and streptomycin (slight) in S. griseus. Overproduction was due to expression of the gene in these strains as compared to the wild-type strains harboring the vector only. Comparative study of the expression of afsR-p revealed that regulatory networking in Streptomyces is not uniform. We speculate that phosphorylated AfsR-p becomes bound to the promoter region of afsS. The latter activates other regulatory genes, including pathway regulatory genes, and induces the production of secondary metabolites including antibiotics. We identified specific conserved amino acids and exploited them for the isolation of the partial sequence of the afsR homologue from S. clavuligerus and Streptomyces achromogens (rubradirin producer). Such findings provide additional evidence for the presence of a serine/threonine and tyrosine kinase-dependent global regulatory network in Streptomyces. PMID:15921897

  19. Response to copper of Acidithiobacillus ferrooxidans ATCC 23270 grown in elemental sulfur.

    PubMed

    Almárcegui, Rodrigo J; Navarro, Claudio A; Paradela, Alberto; Albar, Juan Pablo; von Bernath, Diego; Jerez, Carlos A

    2014-11-01

    The response of Acidithiobacillus ferrooxidans ATCC 23270 to copper was analyzed in sulfur-grown cells by using quantitative proteomics. Forty-seven proteins showed altered levels in cells grown in the presence of 50 mM copper sulfate. Of these proteins, 24 were up-regulated and 23 down-regulated. As seen before in ferrous iron-grown cells, there was a notorious up-regulation of RND-type Cus systems and different RND-type efflux pumps, indicating that these proteins are very important in copper resistance. Copper also triggered the down-regulation of the major outer membrane porin of A. ferrooxidans in sulfur-grown bacteria, suggesting they respond to the metal by decreasing the influx of cations into the cell. On the contrary, copper in sulfur-grown cells caused an overexpression of putative TadA and TadB proteins known to be essential for biofilm formation in bacteria. Surprisingly, sulfur-grown microorganisms showed increased levels of proteins related with energy generation (rus and petII operons) in the presence of copper. Although rus operon is overexpressed mainly in cells grown in ferrous iron, the up-regulation of rusticyanin in sulfur indicates a possible role for this protein in copper resistance as well. Finally, copper response in A. ferrooxidans appears to be influenced by the substrate being oxidized by the microorganism. PMID:25041950

  20. Cloning, expression and bioinformatics analysis of ATP sulfurylase from Acidithiobacillus ferrooxidans ATCC 23270 in Escherichia coli.

    PubMed

    Jaramillo, Michael L; Abanto, Michel; Quispe, Ruth L; Calderón, Julio; Del Valle, Luís J; Talledo, Miguel; Ramírez, Pablo

    2012-01-01

    Molecular studies of enzymes involved in sulfite oxidation in Acidithiobacillus ferrooxidans have not yet been developed, especially in the ATP sulfurylase (ATPS) of these acidophilus tiobacilli that have importance in biomining. This enzyme synthesizes ATP and sulfate from adenosine phosphosulfate (APS) and pyrophosphate (PPi), final stage of the sulfite oxidation by these organisms in order to obtain energy. The atpS gene (1674 bp) encoding the ATPS from Acidithiobacillus ferrooxidans ATCC 23270 was amplified using PCR, cloned in the pET101-TOPO plasmid, sequenced and expressed in Escherichia coli obtaining a 63.5 kDa ATPS recombinant protein according to SDS-PAGE analysis. The bioinformatics and phylogenetic analyses determined that the ATPS from A. ferrooxidans presents ATP sulfurylase (ATS) and APS kinase (ASK) domains similar to ATPS of Aquifex aeolicus, probably of a more ancestral origin. Enzyme activity towards ATP formation was determined by quantification of ATP formed from E. coli cell extracts, using a bioluminescence assay based on light emission by the luciferase enzyme. Our results demonstrate that the recombinant ATP sulfurylase from A. ferrooxidans presents an enzymatic activity for the formation of ATP and sulfate, and possibly is a bifunctional enzyme due to its high homology to the ASK domain from A. aeolicus and true kinases. PMID:23055613

  1. Antimicrobial potential of immobilized Lactococcus lactis subsp. lactis ATCC 11454 against selected bacteria.

    PubMed

    Millette, M; Smoragiewicz, W; Lacroix, M

    2004-06-01

    Immobilization of living cells of lactic acid bacteria could be an alternative or complementary method of immobilizing organic acids and bacteriocins and inhibit undesirable bacteria in foods. This study evaluated the inhibition potential of immobilized Lactococcus lactis subsp. lactis ATCC 11454 on selected bacteria by a modified method of the agar spot test. L. lactis was immobilized in calcium alginate (1 to 2%)-whey protein concentrate (0 and 1%) beads. The antimicrobial potential of immobilized L. lactis was evaluated in microbiological media against pathogenic bacteria (Escherichia coli, Salmonella, and Staphylococcus aureus) or Pseudomonas putida, a natural meat contaminant, and against seven gram-positive bacteria used as indicator strains. Results obtained in this study indicated that immobilized L. lactis inhibited the growth of S. aureus, Enterococcus faecalis, Enterococcus faecium, Lactobacillus curvatus, Lactobacillus sakei, Kocuria varians, and Pediococcus acidilactici. Only 4 h of incubation at 35 degrees C resulted in a clear inhibition zone around the beads that increased with time. With the addition of 10 mM of a chelating agent (EDTA) to the media, results showed growth inhibition of E. coli; however, P. putida and Salmonella Typhi were unaffected by this treatment. These results indicate that immobilized lactic acid bacteria strains can be successfully used to produce nisin and inhibit bacterial growth in semisolid synthetic media. PMID:15222547

  2. Compositional and toxicological evaluation of the diazotrophic cyanobacterium, Cyanothece sp. strain ATCC 51142

    NASA Technical Reports Server (NTRS)

    Schneegurt, M. A.; Arieli, B.; McKeehen, J. D.; Stephens, S. D.; Nielsen, S. S.; Saha, P. R.; Trumbo, P. R.; Sherman, L. A.; Mitchell, C. A. (Principal Investigator)

    1995-01-01

    Compositional analyses of Cyanothece sp. strain ATCC 51142 showed high protein (50-60%) and low fat (0.4-1%) content, and the ability to synthesize vitamin B12. The amino acid profile indicated that Cyanothece sp. was a balanced protein source. Fatty acids of the 18:3n-3 type were also present. Mineral analyses indicated that the cellular biomass may be a good source of Fe, Zn and Na. Caloric content was 4.5 to 5.1 kcal g dry weight-1 and the carbon content was approximately 40% on a dry weight basis. Nitrogen content was 8 to 9% on a dry weight basis and total nucleic acids were 1.3% on a dry weight basis. Short-term feeding studies in rats followed by histopathology found no toxicity or dietary incompatibility problems. The level of uric acid and allantoin in urine and tissues was low, suggesting no excess of nucleic acids, as sometimes reported in the past for a cyanobacteria-containing diet. The current work discusses the potential implications of these results for human nutrition applications.

  3. Transcriptional and Mutational Analysis of the Uptake Hydrogenase of the Filamentous Cyanobacterium Anabaena variabilis ATCC 29413

    PubMed Central

    Happe, Thomas; Schütz, Kathrin; Böhme, Herbert

    2000-01-01

    A 10-kb DNA region of the cyanobacterium Anabaena variabilis ATCC 29413 containing the structural genes of the uptake hydrogenase (hupSL) was cloned and sequenced. In contrast to the hupL gene of Anabaena sp. strain PCC 7120, which is interrupted by a 10.5-kb DNA fragment in vegetative cells, there is no programmed rearrangement within the hupL gene during the heterocyst differentiation of A. variabilis. The hupSL genes were transcribed as a 2.7-kb operon and were induced only under nitrogen-fixing conditions, as shown by Northern blot experiments and reverse transcriptase PCR. Primer extension experiments with a fluorescence-labeled oligonucleotide primer confirmed these results and identified the 5′ start of the mRNA transcript 103 bp upstream of the ATG initiation codon. A consensus sequence in the promoter that is recognized by the fumarate nitrate reductase regulator (Fnr) could be detected. The hupSL operon in A. variabilis was interrupted by an interposon deletion (mutant strain AVM13). Under N2-fixing conditions, the mutant strain exhibited significantly increased rates in H2 accumulation and produced three times more hydrogen than the wild type. These results indicate that the uptake hydrogenase is catalytically active in the wild type and that the enzyme reoxidizes the H2 developed by the nitrogenase. The Nif phenotype of the mutant strain showed a slight decrease of acetylene reduction compared to that of the wild type. PMID:10692368

  4. Improved welan gum production by Alcaligenes sp. ATCC31555 from pretreated cane molasses.

    PubMed

    Ai, Hongxia; Liu, Min; Yu, Pingru; Zhang, Shaozhi; Suo, Yukai; Luo, Ping; Li, Shuang; Wang, Jufang

    2015-09-20

    Welan gum production by Alcaligenes sp. ATCC31555 from cane molasses was studied in batch fermentation to reduce production costs and enhance gum production. The pretreatment of cane molasses, agitation speed and the addition of supplements were investigated to optimize the process. Sulfuric acid hydrolysis was found to be the optimal pretreatment, resulting in a maximum gum concentration of 33.5 g/L, which is 50.0% higher than those obtained from the molasses' mother liquor. Agitation at 600 rpm at 30°C and addition of 10% n-dodecane following fermentation for 36 h increased the maximum gum production up to 41.0 ± 1.41 g/L, which is 49.1% higher than the greatest welan gum concentration in the literature so far. The welan gum product showed an acceptable molecular weight, similar rheological properties and better thermal stability to that obtained from glucose. These results indicate that cane molasses may be a suitable and inexpensive substrate for cost-effective industrial-scale welan gum production. PMID:26050885

  5. Pivotal role of anthranilate dioxygenase genes in the adaptation of Burkholderia multivorans ATCC 17616 in soil.

    PubMed

    Nishiyama, Eri; Ohtsubo, Yoshiyuki; Yamamoto, Yasuhiro; Nagata, Yuji; Tsuda, Masataka

    2012-05-01

    In our recent screen for soil-induced genes, the expression of andA operon (andAcAdAbAa) for anthranilate catabolism in Burkholderia multivorans ATCC 17616 was found to increase dramatically in a soil sample (Nishiyama et al., Environ Microbiol 12: 2539, 2010). The operon was preceded by andR encoding a putative transcriptional regulator for the andA operon. In this study, the andA promoter was induced by tryptophan and anthranilate in an andR-dependent manner. The andA promoter in a deletion mutant lacking tryptophan dioxygenase (one of enzymes for the catabolism of tryptophan to anthranilate) did not respond to tryptophan, indicating that not tryptophan but anthranilate is the effector of AndR. Although both anthranilate and tryptophan were under the detection levels in the soil sample, andA promoter showed higher activity in the soil sample than in a laboratory medium. Such induction required andR and was moderately dependent on the ferric uptake regulator (Fur). The proliferation ability of andAc mutant in the sterile soil was low compared with the co-incubated wild-type cells. These findings suggested that in the soil environment, anthranilate dioxygenase genes are induced by AndR and Fur, and play a pivotal role in the proliferation in the soil environment. PMID:22360670

  6. Purification and characterization of acidolysin, an acidic metalloprotease produced by Clostridium acetobutylicum ATCC 824.

    PubMed Central

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

    1990-01-01

    Acidolysin an extracellular protease produced by Clostridium acetobutylicum ATCC 824 was purified to homogeneity by anion-exchange chromatography with a recovery of 91%. The enzyme was a monomeric protein with a molecular weight of 44,000 as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an acidic isoelectric point of 3.3. Acidolysin was very sensitive to metal-chelating agents and phosphoramidon and was unaffected by sulfhydryl reagents. It was shown to be a calcium- and zinc-containing protease. It exhibited optimal activity against Azocoll at pH 5 and 45 degrees C. It was stable at low pH and heat labile above 50 degrees C. It exhibited specificity toward peptide bonds formed by the amino group of hydrophobic amino acids (isoleucine, leucine, and phenylalanine) and its NH2-terminal amino acid sequence showed a high degree of similarity with that of Bacillus subtilis neutral metalloprotease A. Acidolysin is the first phosphoramidon-sensitive, acidic zinc metalloprotease reported. Images PMID:2082818

  7. Construction and evaluation of a Clostridium thermocellum ATCC 27405 whole-genome oligonucleotide microarray

    SciTech Connect

    Brown, Steven David; Raman, Babu; McKeown, Catherine K; Kale, Shubhangi P; He, Zhili; Mielenz, Jonathan R

    2007-04-01

    Clostridium thermocellum is an anaerobic, thermophilic bacterium that can directly convert cellulosic substrates into ethanol. Microarray technology is a powerful tool to gain insights into cellular processes by examining gene expression under various physiological states. Oligonucleotide microarray probes were designed for 96.7% of the 3163 C. thermocellum ATCC 27405 candidate protein-encoding genes and then a partial-genome microarray containing 70 C. thermocellum specific probes was constructed and evaluated. We detected a signal-to-noise ratio of three with as little as 1.0 ng of genomic DNA and only low signals from negative control probes (nonclostridial DNA), indicating the probes were sensitive and specific. In order to further test the specificity of the array we amplified and hybridized 10 C. thermocellum polymerase chain reaction products that represented different genes and found gene specific hybridization in each case. We also constructed a whole-genome microarray and prepared total cellular RNA from the same point in early-logarithmic growth phase from two technical replicates during cellobiose fermentation. The reliability of the microarray data was assessed by cohybridization of labeled complementary DNA from the cellobiose fermentation samples and the pattern of hybridization revealed a linear correlation. These results taken together suggest that our oligonucleotide probe set can be used for sensitive and specific C. thermocellum transcriptomic studies in the future.

  8. Sucrose fermentation by Fusobacterium mortiferum ATCC 25557: transport, catabolism, and products.

    PubMed Central

    Thompson, J; Nguyen, N Y; Robrish, S A

    1992-01-01

    Studies of sucrose utilization by Fusobacterium mortiferum ATCC 25557 have provided the first definitive evidence for phosphoenolpyruvate-dependent sugar:phosphotransferase activity in the family Bacteroidaceae. The phosphoenolpyruvate-dependent sucrose:phosphotransferase system and the two enzymes required for the dissimilation of sucrose 6-phosphate are induced specifically by growth of F. mortiferum on the disaccharide. Monomeric sucrose 6-phosphate hydrolase (M(r), 52,000) and a dimeric ATP-dependent fructokinase (subunit M(r), 32,000) have been purified to electrophoretic homogeneity. The physicochemical and catalytic properties of these enzymes have been examined, and the N-terminal amino acid sequences for both proteins are reported. The characteristics of sucrose 6-phosphate hydrolase and fructokinase from F. mortiferum are compared with the same enzymes from both gram-positive and gram-negative species. Butyric, acetic, and D-lactic acids are the end products of sucrose fermentation by F. mortiferum. A pathway is proposed for the translocation, phosphorylation, and metabolism of sucrose by this anaerobic pathogen. Images PMID:1533618

  9. Production of single cell oil from Lipomyces starkeyi ATCC 56304 using biorefinery by-products.

    PubMed

    Probst, Kyle V; Vadlani, Praveen V

    2015-12-01

    Single cell oil (SCO) is a valuable noncrop-based renewable oil source. Hemicellulose derived sugars can be utilized to produce SCO using the oleaginous yeast Lipomyces starkeyi ATCC 56304. Bran by-products were tested as hemicellulose-rich feedstocks for the production of SCO. Whole and destarched corn and wheat bran hydrolysates were produced using hydrothermal and dilute sulfuric acid (0%, 0.5%, 1.0%, v/v) pretreatment along with enzymatic hydrolysis. Whole bran hydrolysates produced from hydrothermal pretreatment generated the highest average oil yields of 126.7 and 124.3 mg oil/g sugar for both wheat and corn bran, respectively. 1.0% acid pretreatment was effective for the destarched bran generating a hemicellulose hydrolysis efficiency of 94% and 84% for wheat and corn bran, respectively, resulting in the highest oil yield of 70.7 mg oil/g sugar. The results indicate pretreated corn and wheat bran hydrolysates can serve as viable feedstocks for oleaginous yeast SCO bioconversion. PMID:26402869

  10. Catabolite repression in Lactobacillus casei ATCC 393 is mediated by CcpA.

    PubMed Central

    Monedero, V; Gosalbes, M J; Pérez-Martínez, G

    1997-01-01

    The chromosomal ccpA gene from Lactobacillus casei ATCC 393 has been cloned and sequenced. It encodes the CcpA protein, a central catabolite regulator belonging to the LacI-GalR family of bacterial repressors, and shows 54% identity with CcpA proteins from Bacillus subtilis and Bacillus megaterium. The L. casei ccpA gene was able to complement a B. subtilis ccpA mutant. An L. casei ccpA mutant showed increased doubling times and a relief of the catabolite repression of some enzymatic activities, such as N-acetylglucosaminidase and phospho-beta-galactosidase. Detailed analysis of CcpA activity was performed by using the promoter region of the L. casei chromosomal lacTEGF operon which is subject to catabolite repression and contains a catabolite responsive element (cre) consensus sequence. Deletion of this cre site or the presence of the ccpA mutation abolished the catabolite repression of a lacp::gusA fusion. These data support the role of CcpA as a common regulatory element mediating catabolite repression in low-GC-content gram-positive bacteria. PMID:9352913

  11. Immunobiological activities of a porin fraction isolated from Fusobacterium nucleatum ATCC 10953.

    PubMed Central

    Takada, H; Ogawa, T; Yoshimura, F; Otsuka, K; Kokeguchi, S; Kato, K; Umemoto, T; Kotani, S

    1988-01-01

    From Fusobacterium nucleatum ATCC 10953 cell envelope fraction whose inner membranes had been removed by treatment with sodium N-lauroyl sarcosinate, an outer membrane protein (37,000 Mr in a native state) was prepared by extraction with lithium dodecyl sulfate. The protein thus obtained showed distinct porin activity, namely, the ability to form hydrophilic diffusion pores by incorporation into the artificial liposome membrane. The porin fraction exhibited strong immunobiological activities in the in vitro assays: B-cell mitogenicity and polyclonal B-cell activation on murine splenocytes, stimulatory effects on guinea pig peritoneal macrophages, and enhancement of the migration of human blood monocytes. The porin fraction also exhibited immunoadjuvant activity to increase the antibody production against sheep erythrocytes in the spleen of mice that were immunized by sheep erythrocytes with porin. Although chemical analyses revealed that the test porin fraction contained a considerable amount of lipopolysaccharide (LPS) (around 12% of the fraction), the studies with LPS-nonresponding C3H/HeJ mice and on the inhibitory effects of polymyxin B strongly suggest that most of the above bioactivities are due to porin protein itself, not to coexistent LPS in the porin fraction. Images PMID:2831155

  12. Evaluation of Cyanothece sp. ATCC 51142 as a candidate for inclusion in a CELSS

    NASA Technical Reports Server (NTRS)

    Schneegurt, M. A.; Arieli, B.; Nielsen, S. S.; Trumbo, P. R.; Sherman, L. A.; Mitchell, C. A. (Principal Investigator)

    1996-01-01

    Controlled ecological life support systems (CELSS) have been proposed to make long-duration manned space flights more cost-effective. Higher plants will presumably provide food and a breathable atmosphere for the crew. It has been suggested that imbalances between the CO2/O2 gas exchange ratios of the heterotrophic and autotrophic components of the system will inevitably lead to an unstable system, and the loss of O2 from the atmosphere. Ratio imbalances may be corrected by including a second autotroph with an appropriate CO2/O2 gas exchange ratio. Cyanothece sp. ATCC 51142 is a large unicellular N2-fixing cyanobacterium, exhibiting high growth rates under diverse physiological conditions. A rat-feeding study showed the biomass to be edible. Furthermore, it may have a CO2/O2 gas exchange ratio that theoretically can compensate for ratio imbalances. It is suggested that Cyanothece spp. could fulfill several roles in a CELSS: supplementing atmosphere recycling, generating fixed N from the air, providing a balanced protein supplement, and protecting a CELSS in case of catastrophic crop failure.

  13. Evaluation of Cyanothece sp. ATCC 51142 as a candidate for inclusion in a CELSS.

    PubMed

    Schneegurt, M A; Arieli, B; Nielsen, S S; Trumbo, P R; Sherman, L A

    1996-01-01

    Controlled ecological life support systems (CELSS) have been proposed to make long-duration manned space flights more cost-effective. Higher plants will presumably provide food and a breathable atmosphere for the crew. It has been suggested that imbalances between the CO2/O2 gas exchange ratios of the heterotrophic and autotrophic components of the system will inevitably lead to an unstable system, and the loss of O2 from the atmosphere. Ratio imbalances may be corrected by including a second autotroph with an appropriate CO2/O2 gas exchange ratio. Cyanothece sp. ATCC 51142 is a large unicellular N2-fixing cyanobacterium, exhibiting high growth rates under diverse physiological conditions. A rat-feeding study showed the biomass to be edible. Furthermore, it may have a CO2/O2 gas exchange ratio that theoretically can compensate for ratio imbalances. It is suggested that Cyanothece spp. could fulfill several roles in a CELSS: supplementing atmosphere recycling, generating fixed N from the air, providing a balanced protein supplement, and protecting a CELSS in case of catastrophic crop failure. PMID:11538794

  14. Sophorolipids Production by Candida bombicola ATCC 22214 and its Potential Application in Microbial Enhanced Oil Recovery

    PubMed Central

    Elshafie, Abdulkadir E.; Joshi, Sanket J.; Al-Wahaibi, Yahya M.; Al-Bemani, Ali S.; Al-Bahry, Saif N.; Al-Maqbali, Dua’a; Banat, Ibrahim M.

    2015-01-01

    Biosurfactant production using Candida bombicola ATCC 22214, its characterization and potential applications in enhancing oil recovery were studied at laboratory scale. The seed media and the production media were standardized for optimal growth and biosurfactant production. The production media were tested with different carbon sources: glucose (2%w/v) and corn oil (10%v/v) added separately or concurrently. The samples were collected at 24 h interval up to 120 h and checked for growth (OD660), and biosurfactant production [surface tension (ST) and interfacial tension (IFT)]. The medium with both glucose and corn oil gave better biosurfactant production and reduced both ST and IFT to 28.56 + 0.42mN/m and 2.13 + 0.09mN/m, respectively within 72 h. The produced biosurfactant was quite stable at 13–15% salinity, pH range of 2–12, and at temperature up to 100°C. It also produced stable emulsions (%E24) with different hydrocarbons (pentane, hexane, heptane, tridecane, tetradecane, hexadecane, 1-methylnaphthalene, 2,2,4,4,6,8-heptamethylnonane, light and heavy crude oil). The produced biosurfactant was extracted using ethyl acetate and characterized as a mixture of sophorolipids (SPLs). The potential of SPLs in enhancing oil recovery was tested using core-flooding experiments under reservoir conditions, where additional 27.27% of residual oil (Sor) was recovered. This confirmed the potential of SPLs for applications in microbial enhanced oil recovery. PMID:26635782

  15. A model of cyclic transcriptomic behavior in the cyanobacterium Cyanothece sp. ATCC 51142.

    PubMed

    McDermott, Jason E; Oehmen, Christopher S; McCue, Lee Ann; Hill, Eric; Choi, Daniel M; Stöckel, Jana; Liberton, Michelle; Pakrasi, Himadri B; Sherman, Louis A

    2011-08-01

    Systems biology attempts to reconcile large amounts of disparate data with existing knowledge to provide models of functioning biological systems. The cyanobacterium Cyanothece sp. ATCC 51142 is an excellent candidate for such systems biology studies because: (i) it displays tight functional regulation between photosynthesis and nitrogen fixation; (ii) it has robust cyclic patterns at the genetic, protein and metabolomic levels; and (iii) it has potential applications for bioenergy production and carbon sequestration. We have represented the transcriptomic data from Cyanothece 51142 under diurnal light/dark cycles as a high-level functional abstraction and describe development of a predictive in silico model of diurnal and circadian behavior in terms of regulatory and metabolic processes in this organism. We show that incorporating network topology into the model improves performance in terms of our ability to explain the behavior of the system under new conditions. The model presented robustly describes transcriptomic behavior of Cyanothece 51142 under different cyclic and non-cyclic growth conditions, and represents a significant advance in the understanding of gene regulation in this important organism. PMID:21698331

  16. Clavulanic acid production by the MMS 150 mutant obtained from wild type Streptomyces clavuligerus ATCC 27064

    PubMed Central

    da Silva Vasconcelos, Eliton; de Lima, Vanderlei Aparecido; Goto, Leandro Seiji; Cruz-Hernández, Isara Lourdes; Hokka, Carlos Osamu

    2013-01-01

    Clavulanic acid (CA) is a powerful inhibitor of the beta-lactamases, enzymes produced by bacteria resistants to penicillin and cefalosporin. This molecule is produced industrially by strains of Streptomyces clavuligerus in complex media which carbon and nitrogen resources are supplied by inexpensive compounds still providing high productivity. The genetic production improvement using physical and chemical mutagenic agents is an important strategy in programs of industrial production development of bioactive metabolites. However, parental strains are susceptible to loss of their original productivity due genetic instability phenomenona. In this work, some S. clavuligerus mutant strains obtained by treatment with UV light and with MMS are compared with the wild type (Streptomyces clavuligerus ATCC 27064). The results indicated that the random mutations originated some strains with different phenotypes, most divergent demonstrated by the mutants strains named AC116, MMS 150 and MMS 54, that exhibited lack of pigmentation in their mature spores. Also, the strain MMS 150 presented a larger production of CA when cultivated in semi-synthetics media. Using other media, the wild type strain obtained a larger CA production. Besides, using the modifed complex media the MMS 150 strain showed changes in its lipolitic activity and a larger production of CA. The studies also allowed finding the best conditions for a lipase activity exhibited by wild type S. clavuligerus and the MMS150 mutant. PMID:24688492

  17. Physical and genetic map of the Clostridium acetobutylicum ATCC 824 chromosome.

    PubMed Central

    Cornillot, E; Croux, C; Soucaille, P

    1997-01-01

    A physical and genetic map of the Clostridium acetobutylicum ATCC 824 chromosome was constructed. The macrorestriction map for CeuI, EagI, and SstII was created by ordering the 38 restriction sites by one- and two-dimensional pulsed-field gel electrophoresis (PFGE) and by using an original strategy based on the CeuI enzyme and indirect end labelling by hybridization on both sides of the CeuI sites with rrs (16S RNA) and 3' rrl (23S RNA) probes. The circular chromosome was estimated to be 4.15 Mb in size, and the average resolution of the physical map is 110 kb. The chromosome contains 11 rrn loci, which are localized on 44% of the chromosome in a divergent transcriptional orientation regarding the presumed location of the replication origin. In addition to these 11 rrn operons, a total of 40 identified genes were mapped by hybridization experiments with genes from C. acetobutylicum and from various other clostridia as probes. The genetic map of C. acetobutylicum was compared to that of the three other endospore-forming bacteria characterized so far: Bacillus subtilis, Clostridium beijerinckii, and Clostridium perfringens. Parodoxically, the chromosomal backbone of C. acetobutylicum showed more similarity to that of B. subtilis than to those of the clostridia. PMID:9393708

  18. In vitro and in vivo activities of ticarcillin-loaded nanoliposomes with different surface charges against Pseudomonas aeruginosa (ATCC 29248)

    PubMed Central

    2012-01-01

    Background Pseudomonas aeruginosa exhibits multiple antibiotic resistance mechanisms. Different studies have shown that entrapment of antibiotics into liposomes could increase their anti-Pseudomonas activity. The objectives of this study were to prepare ticarcillin loaded-nanoliposomes with variable surface charges and evaluate their in vitro and in vivo efficacies against Pseudomonas aeruginosa (ATCC 29248). Methods Ticarcillin-loaded nanoliposomes with positive, negative and neutral surface charges were prepared by extrusion method. Ticarcillin encapsulation efficacies for different formulations were measured by HPLC method. Minimum inhibitory concentration (MIC) of ticarcillin nanoliposomal forms against strain ATCC 29248 were determined by broth dilution method. The killing rate of Pseudomonas aeruginosa was exposed to various concentrations of ticarcillin in free and nanoliposomal forms were analyzed. Ultimately, in vivo therapeutic efficacy of nanoliposomes in burned mice skin infected with strain ATCC 29248 was investigated. Results The encapsulation efficacies for ticarcillin-loaded cationic nanoliposomes were significantly higher (76% ± 0.17) than those of neutral (55% ± 0.14) and anionic (43% ± 0.14) nanoliposomes. The MIC of free, cationic, neutral and anionic nanoliposomal forms of ticarcillin against ATCC 29248 were to 24, 3, 6 and 48 mg/L, respectively. The killing rates of ticarcillin-loaded cationic nanoliposomes were higher than those of free and other drug formulations. Treatment by ticarcillin-loaded nanoliposomes with positive, neutral and negative surface charges resulted in almost 100, 60 and 20% survival rates, respectively. Conclusion Our data suggested that cationic ticarcillin-loaded nanoliposomes because of high effectiveness would be a good choice to treatment of Pseudomonas aeruginosa infections. PMID:23351156

  19. Classification of Nonomuraea sp. ATCC 39727, an actinomycete that produces the glycopeptide antibiotic A40926, as Nonomuraea gerenzanensis sp. nov.

    PubMed

    Dalmastri, Claudia; Gastaldo, Luciano; Marcone, Giorgia Letizia; Binda, Elisa; Congiu, Terenzio; Marinelli, Flavia

    2016-02-01

    Strain ATCC 39727, which produces the antibiotic A40926 (the natural precursor of the antibiotic dalbavancin), was isolated from a soil sample collected in India, and it was originally classified as a member of the genus Actinomadura on the base of morphology and cell-wall composition. A phylogenetic analysis based on 16S rRNA gene sequences indicates that the strain forms a distinct clade within the genus Nonomuraea, and it is most closely related to Nonomuraea angiospora DSM 43173T (98.72 % similarity) and Nonomuraea jabiensis A4036T (98.69 %). The strain forms an extensively branched substrate mycelium and aerial hyphae that form spiral chains of spores with ridged surfaces. The cell wall contains meso-diaminopimelic acid and the whole-cell sugars are glucose, ribose, galactose, mannose and madurose (madurose as the diagnostic sugar). The N-acyl type of muramic acid is acetyl. The predominant menaquinone is MK-9(H4), with minor amounts of MK-9(H2), MK-9(H6) and MK-9(H0). The polar-lipid profile includes diphosphatidylglycerol, phosphatidylethanolamine, hydroxyphosphatidylethanolamine, phosphatidylmethylethanolamine, hydroxyphosphatidylmethylethanolamine, phosphatidylinositol and a series of uncharacterized phospholipids, glycolipids and phosphoglycolipids. The major cellular fatty acids are iso-C16 : 0 and 10-methyl C17 : 0. The genomic DNA G+C content is 71.2 mol%. Significant differences in the morphological, chemotaxonomic and biochemical data, together with DNA-DNA relatedness between strain ATCC 39727 and closely related type strains, clearly demonstrated that strain ATCC 39727 represents a novel species of the genus Nonomuraea, for which the name Nonomuraea gerenzanensis sp. nov. is proposed. The type strain is ATCC 39727T ( = DSM 100948T). PMID:26944798

  20. Complete Genome Sequence of Mycoplasma hominis Strain Sprott (ATCC 33131), Isolated from a Patient with Nongonococcal Urethritis

    PubMed Central

    Foecking, Mark F.

    2015-01-01

    Presented here is the complete and annotated genome sequence of Mycoplasma hominis Sprott (ATCC 33131). The chromosome comprises 695,214 bp, which is approximately 30 kb larger than the syntenic genome of M. hominis PG21T. Tetracycline resistance of strain Sprott is most probably conferred by the tetM determinant, harbored on a mosaic transposon-like structure. PMID:26159538

  1. Co-culturing a novel Bacillus strain with Clostridium tyrobutyricum ATCC 25755 to produce butyric acid from sucrose

    PubMed Central

    2013-01-01

    Background Currently, the most promising microorganism used for the bio-production of butyric acid is Clostridium tyrobutyricum ATCC 25755T; however, it is unable to use sucrose as a sole carbon source. Consequently, a newly isolated strain, Bacillus sp. SGP1, that was found to produce a levansucrase enzyme, which hydrolyzes sucrose into fructose and glucose, was used in a co-culture with this strain, permitting C. tyrobutyricum ATCC 25755T to ferment sucrose to butyric acid. Results B. sp. SGP1 alone did not show any butyric acid production and the main metabolite produced was lactic acid. This allowed C. tyrobutyricum ATCC 25755T to utilize the monosaccharides resulting from the activity of levansucrase together with the lactic acid produced by B. sp. SGP1 to generate butyric acid, which was the main fermentative product within the co-culture. Furthermore, the final acetic acid concentration in the co-culture was significantly lower when compared with pure C. tyrobutyricum ATCC 25755T cultures grown on glucose. In fed-batch fermentations, the optimum conditions for the production of butyric acid were around pH 5.50 and a temperature of 37°C. Under these conditions, the final butyrate concentration was 34.2±1.8 g/L with yields of 0.35±0.03 g butyrate/g sucrose and maximum productivity of 0.3±0.04 g/L/h. Conclusions Using this co-culture, sucrose can be utilized as a carbon source for butyric acid production at a relatively high yield. In addition, this co-culture offers also the benefit of a greater selectivity, with butyric acid constituting 92.8% of the acids when the fermentation was terminated. PMID:23452443

  2. Pelczaria aurantia ATCC 49321T (=DSM 12801T) is a strain of Kocuria rosea (Flügge 1886) Stackebrandt et al. 1995.

    PubMed

    Schumann, P; Tindall, B J; Mendrock, U; Kramer, I; Stackebrandt, E

    2000-07-01

    Phylogenetic and chemotaxonomic analyses of Pelczaria aurantia ATCC 49321T (= DSM 12801T) indicate that this species is very closely related to Kocuria rosea. The DNA-DNA reassociation value of 87.1% determined for the type strains of the two species supports this finding. The results of phylogenetic analysis of the 16S rDNA of a subculture of the original strain of Pelczaria aurantia, deposited at the National Institutes of Health, Bethesda, MD, USA, as 'Neisseria aurantia', are identical to those for strain ATCC 49321T and indicate that Pelczaria aurantia ATCC 49321T is an authentic subculture of the original culture described by Poston (1993). On the basis of these findings it is concluded that P. aurantia ATCC 49321T and K. rosea DSM 20447T are members of the same taxon. The taxonomic consequences of this union are discussed. PMID:10939645

  3. Expression of arsenic resistance genes in the obligate anaerobe Bacteroides vulgatus ATCC 8482, a gut microbiome bacterium.

    PubMed

    Li, Jiaojiao; Mandal, Goutam; Rosen, Barry P

    2016-06-01

    The response of the obligate anaerobe Bacteroides vulgatus ATCC 8482, a common human gut microbiota, to arsenic was determined. B. vulgatus ATCC 8482 is highly resistant to pentavalent As(V) and methylarsenate (MAs(V)). It is somewhat more sensitive to trivalent inorganic As(III) but 100-fold more sensitive to methylarsenite (MAs(III)) than to As(III). B. vulgatus ATCC 8482 has eight continuous genes in its genome that we demonstrate form an arsenical-inducible transcriptional unit. The first gene of this ars operon, arsR, encodes a putative ArsR As(III)-responsive transcriptional repressor. The next three genes encode proteins of unknown function. The remaining genes, arsDABC, have well-characterized roles in detoxification of inorganic arsenic, but there are no known genes for MAs(III) resistance. Expression of each gene after exposure to trivalent and pentavalent inorganic and methylarsenicals was analyzed. MAs(III) was the most effective inducer. The arsD gene was the most highly expressed of the ars operon genes. These results demonstrate that this anaerobic microbiome bacterium has arsenic-responsive genes that confer resistance to inorganic arsenic and may be responsible for the organism's ability to maintain its prevalence in the gut following dietary exposure to inorganic arsenic. PMID:27040269

  4. Transcriptome analysis of Cronobacter sakazakii ATCC BAA-894 after interaction with human intestinal epithelial cell line HCT-8.

    PubMed

    Jing, Chun-e; Du, Xin-jun; Li, Ping; Wang, Shuo

    2016-01-01

    Cronobacter spp. are opportunistic pathogens that are responsible for infections including severe meningitis, septicemia, and necrotizing enterocolitis in neonates and infants. To date, questions still remain regarding the mechanisms of pathogenicity and virulence determinants for each bacterial strain. In this study, we established an in vitro model for Cronobacter sakazakii ATCC BAA-894 infection of HCT-8 human colorectal epithelial cells. The transcriptome profile of C. sakazakii ATCC BAA-894 after interaction with HCT-8 cells was determined using high-throughput whole-transcriptome sequencing (RNA sequencing (RNA-seq)). Gene expression profiles indicated that 139 genes were upregulated and 72 genes were downregulated in the adherent C. sakazakii ATCC BAA-894 strain on HCT-8 cells compared to the cultured bacteria in the cell-free medium. Expressions of some flagella genes and virulence factors involved in adherence were upregulated. High osmolarity and osmotic stress-associated genes were highly upregulated, as well as genes responsible for the synthesis of lipopolysaccharides and outer membrane proteins, iron acquisition systems, and glycerol and glycerophospholipid metabolism. In sum, our study provides further insight into the mechanisms underlying C. sakazakii pathogenesis in the human gastrointestinal tract. PMID:26481623

  5. Selection of the Strain Lactobacillus acidophilus ATCC 43121 and Its Application to Brewers' Spent Grain Conversion into Lactic Acid

    PubMed Central

    Liguori, Rossana; Soccol, Carlos Ricardo; Vandenberghe, Luciana Porto de Souza; Woiciechowski, Adenise Lorenci; Ionata, Elena; Marcolongo, Loredana; Faraco, Vincenza

    2015-01-01

    Six Lactobacillus strains were analyzed to select a bacterium for conversion of brewers' spent grain (BSG) into lactic acid. Among the investigated strains, L. acidophilus ATCC 43121 showed the highest yield of lactic acid production (16.1 g/L after 48 hours) when grown in a synthetic medium. It was then analyzed for its ability to grow on the hydrolysates obtained from BSG after acid-alkaline (AAT) or aqueous ammonia soaking (AAS) pretreatment. The lactic acid production by L. acidophilus ATCC 43121 through fermentation of the hydrolysate from AAS treated BSG was 96% higher than that from the AAT treated one, although similar yields of lactic acid per consumed glucose were achieved due to a higher (46%) glucose consumption by L. acidophilus ATCC 43121 in the AAS BSG hydrolysate. It is worth noting that adding yeast extract to the BSG hydrolysates increased both the yield of lactic acid per substrate consumed and the volumetric productivity. The best results were obtained by fermentation of AAS BSG hydrolysate supplemented by yeast extract, in which the strain produced 22.16 g/L of lactic acid (yield of 0.61 g/g), 27% higher than the value (17.49 g/L) obtained in the absence of a nitrogen source. PMID:26640784

  6. Genome mining of astaxanthin biosynthetic genes from Sphingomonas sp. ATCC 55669 for heterologous overproduction in Escherichia coli.

    PubMed

    Ma, Tian; Zhou, Yuanjie; Li, Xiaowei; Zhu, Fayin; Cheng, Yongbo; Liu, Yi; Deng, Zixin; Liu, Tiangang

    2016-02-01

    As a highly valued keto-carotenoid, astaxanthin is widely used in nutritional supplements and pharmaceuticals. Therefore, the demand for biosynthetic astaxanthin and improved efficiency of astaxanthin biosynthesis has driven the investigation of metabolic engineering of native astaxanthin producers and heterologous hosts. However, microbial resources for astaxanthin are limited. In this study, we found that the α-Proteobacterium Sphingomonas sp. ATCC 55669 could produce astaxanthin naturally. We used whole-genome sequencing to identify the astaxanthin biosynthetic pathway using a combined PacBio-Illumina approach. The putative astaxanthin biosynthetic pathway in Sphingomonas sp. ATCC 55669 was predicted. For further confirmation, a high-efficiency targeted engineering carotenoid synthesis platform was constructed in E. coli for identifying the functional roles of candidate genes. All genes involved in astaxanthin biosynthesis showed discrete distributions on the chromosome. Moreover, the overexpression of exogenous E. coli idi in Sphingomonas sp. ATCC 55669 increased astaxanthin production by 5.4-fold. This study described a new astaxanthin producer and provided more biosynthesis components for bioengineering of astaxanthin in the future. PMID:26580858

  7. Lactobacillus acidophilus ATCC 4356 inhibits biofilm formation by C. albicans and attenuates the experimental candidiasis in Galleria mellonella

    PubMed Central

    Vilela, Simone FG; Barbosa, Júnia O; Rossoni, Rodnei D; Santos, Jéssica D; Prata, Marcia CA; Anbinder, Ana Lia; Jorge, Antonio OC; Junqueira, Juliana C

    2015-01-01

    Probiotic strains of Lactobacillus have been studied for their inhibitory effects on Candida albicans. However, few studies have investigated the effect of these strains on biofilm formation, filamentation and C. albicans infection. The objective of this study was to evaluate the influence of Lactobacillus acidophilus ATCC 4356 on C. albicans ATCC 18804 using in vitro and in vivo models. In vitro analysis evaluated the effects of L. acidophilus on the biofilm formation and on the capacity of C. albicans filamentation. For in vivo study, Galleria mellonella was used as an infection model to evaluate the effects of L. acidophilus on candidiasis by survival analysis, quantification of C. albicans CFU/mL, and histological analysis. The direct effects of L. acidophilus cells on C. albicans, as well as the indirect effects using only a Lactobacillus culture filtrate, were evaluated in both tests. The in vitro results showed that both L. acidophilus cells and filtrate were able to inhibit C. albicans biofilm formation and filamentation. In the in vivo study, injection of L. acidophilus into G. mellonella larvae infected with C. albicans increased the survival of these animals. Furthermore, the number of C. albicans CFU/mL recovered from the larval hemolymph was lower in the group inoculated with L. acidophilus compared to the control group. In conclusion, L. acidophilus ATCC 4356 inhibited in vitro biofilm formation by C. albicans and protected G. mellonella against experimental candidiasis in vivo. PMID:25654408

  8. Genome Sequences of Ralstonia insidiosa Type Strain ATCC 49129 and Strain FC1138, a Strong Biofilm Producer Isolated from a Fresh-Cut Produce-Processing Plant

    PubMed Central

    Xu, Yunfeng; Nagy, Attila; Yan, Xianghe; Haley, Bradd J.; Kim, Seon Woo; Liu, Nancy T.

    2016-01-01

    Ralstonia insidiosa is an opportunistic pathogen and a strong biofilm producer. Here, we present the complete genome sequences of R. insidiosa FC1138 and ATCC 49129. Both strains have two circular chromosomes of approximately 3.9 and 1.9 Mb and a 50-kb plasmid. ATCC 49129 also possesses a megaplasmid of approximately 318 kb. PMID:27540070

  9. Antibacterial activity of antagonistic bacterium Bacillus subtilis DJM-51 against phytopathogenic Clavibacter michiganense subsp. michiganense ATCC 7429 in vitro.

    PubMed

    Jung, W J; Mabood, F; Souleimanov, A; Whyte, L G; Niederberger, T D; Smith, D L

    2014-12-01

    To investigate antibacterial activity against the tomato pathogen Clavibacter michiganense subsp. michiganense ATCC 7429 (Cmm ATCC 7429), Bacillus subtilis DJM-51 was isolated from rhizosphere soil. For isolation of bacteria, samples were taken from rhizosphere soil. The isolate, DJA-51, had strong antagonistic ability against Tomato pathogen Cmm ATCC 7429 on nutrient-broth yeast extract agar (NBYA) as indicated by inhibition zones around colonies. On the basis of the nucleotide sequence of a conserved segment of the 16S rRNA gene, the bacterium has been identified as B. subtilis DJM-51. The growth of Cmm ATCC 7429 on NBYA plates was inhibited by culture broth of B. subtilis DJM-51 including cells, by the supernatant of culture broth of B. subtilis DJM-51, and by the liquid material resulting from butanol extract of bacterial cultures. The OD value in co-culture mixture was lower than the control throughout the entire incubation period. Antibiotics obtained from B. subtilis DJM-51 inhibited the growth of Tomato pathogen Cmm ATCC 7429. These results provide potentially information about the protection of tomato from pathogen Cmm ATCC 7429 under greenhouse conditions in Quebec. PMID:25457795

  10. Differential proteomic analysis of Clostridium perfringens ATCC13124; identification of dominant, surface and structure associated proteins

    PubMed Central

    2009-01-01

    Background Clostridium perfringens is a medically important clostridial pathogen causing diseases in man and animals. To invade, multiply and colonize tissues of the host, a pathogen must be able to evade host immune system, and obtain nutrients essential for growth. The factors involved in these complex processes are largely unknown and of crucial importance to understanding microbial pathogenesis. Many of the virulence determinants and putative vaccine candidates for bacterial pathogens are known to be surface localized. Results Using 2-DE mass spectrometry strategy, we identified major surface (22) and cell envelope (10) proteins from Clostridium perfringens ATCC13124 and those differentially expressed (11) in cells grown on cooked meat medium (CMM) in comparison with cells grown in reference state (tryptose-yeast extract-glucose medium). Riboflavin biosynthesis protein, ornithine carbamoyltransferase, cystathionine beta-lyase, and threonine dehydratase were the predominant proteins that exhibited 2.19 to 8.5 fold increase in the expression level in cells growing on CMM. Conclusion Ornithine carbamoyltransferase and cystathionine beta-lyase were over-expressed in cells grown on cooked meat medium and also identified in the surface protein fraction and the former was immunogenic; making them potential vaccine candidates. Based upon bioinformatic analysis; choloylglycine hydrolase family protein, cell wall-associated serine proteinase, and rhomboid family protein were predicted as surface protein markers for specific detection of C. perfringens from the environment and food. Most of the proteins over-expressed in CMM were shown to have putative function in metabolism, of which seven were involved in amino acid transport and metabolism or lipid metabolism. PMID:19664283

  11. Complete Genome Sequence of the Marine, Chemolithoautotrophic, Ammonia-Oxidizing Bacterium Nitrosococcus oceani ATCC 19707

    SciTech Connect

    Klots, Martin G.; Arp, D J; Chain, Patrick S; El-Sheikh, Amal F.; Hauser, Loren John; Hommes, Norman G.; Larimer, Frank W; Malfatti, Stephanie; Norton, Jeanette M.; Poret-Peterson, Amisha T.; Vergez, Lisa; Ward, Bess B.

    2006-01-01

    The gammaproteobacterium Nitrosococcus oceani (ATCC 19707) is a gram-negative obligate chemolithoautotroph capable of extracting energy and reducing power from the oxidation of ammonia to nitrite. Sequencing and annotation of the genome revealed a single circular chromosome (3,481,691 bp; G+C content of 50.4%) and a plasmid (40,420 bp) that contain 3,052 and 41 candidate protein-encoding genes, respectively. The genes encoding proteins necessary for the function of known modes of lithotrophy and autotrophy were identified. Contrary to betaproteobacterial nitrifier genomes, the N. oceani genome contained two complete rrn operons. In contrast, only one copy of the genes needed to synthesize functional ammonia monooxygenase and hydroxylamine oxidoreductase, as well as the proteins that relay the extracted electrons to a terminal electron acceptor, were identified. The N. oceani genome contained genes for 13 complete two-component systems. The genome also contained all the genes needed to reconstruct complete central pathways, the tricarboxylic acid cycle, and the Embden-Meyerhof-Parnass and pentose phosphate pathways. The N. oceani genome contains the genes required to store and utilize energy from glycogen inclusion bodies and sucrose. Polyphosphate and pyrophosphate appear to be integrated in this bacterium's energy metabolism, stress tolerance, and ability to assimilate carbon via gluconeogenesis. One set of genes for type I ribulose-1,5-bisphosphate carboxylase/oxygenase was identified, while genes necessary for methanotrophy and for carboxysome formation were not identified. The N. oceani genome contains two copies each of the genes or operons necessary to assemble functional complexes I and IV as well as ATP synthase (one H+-dependent F0F1 type, one Na+-dependent V type).

  12. Multicopy Integration and Expression of Heterologous Genes in Methylobacterium extorquens ATCC 55366†

    PubMed Central

    Choi, Young J.; Bourque, Denis; Morel, Lyne; Groleau, Denis; Míguez, Carlos B.

    2006-01-01

    High-level expression of chromosomally integrated genes in Methylobacterium extorquens ATCC 55366 was achieved under the control of the strong M. extorquens AM1 methanol dehydrogenase promoter (PmxaF) using the mini-Tn7 transposon system. Stable maintenance and expression of the integrated genes were obtained in the absence of antibiotic selective pressure. Furthermore, using this technology, a multicopy integration protocol for M. extorquens was also developed. Chromosomal integration of one to five copies of the gene encoding the green fluorescent protein (gfp) was achieved. The multicopy-based expression system permitted expression of a preset number of gene copies. A unique specific Tn7 integration locus in the chromosome of M. extorquens, known as the Tn7 attachment site (attTn7 site), was identified. This single attTn7 site was identified in an intergenic region between glmS, which encodes the essential enzyme glucosamine-6-phosphate synthetase, and dhaT, which encodes 1,3-propanediol dehydrogenase. The fact that the integration event is site specific and the fact that the attTn7 site is a noncoding region of the chromosome make the mini-Tn7 transposon system very useful for insertion of target genes and subsequent expression. In all transformants tested, expression and segregation of the transforming gene were stable without generation of secondary mutations in the host. In this paper, we describe single and multicopy chromosome integration and stable expression of heterologous genes (bgl [β-galactosidase], est [esterase], and gfp [green fluorescent protein]) in M. extorquens. PMID:16391115

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

    PubMed

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

    2015-08-01

    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

  14. Mutational Studies of Putative Biosynthetic Genes for the Cyanobacterial Sunscreen Scytonemin in Nostoc punctiforme ATCC 29133

    PubMed Central

    Ferreira, Daniela; Garcia-Pichel, Ferran

    2016-01-01

    The heterocyclic indole-alkaloid scytonemin is a sunscreen found exclusively among cyanobacteria. An 18-gene cluster is responsible for scytonemin production in Nostoc punctiforme ATCC 29133. The upstream genes scyABCDEF in the cluster are proposed to be responsible for scytonemin biosynthesis from aromatic amino acid substrates. In vitro studies of ScyA, ScyB, and ScyC proved that these enzymes indeed catalyze initial pathway reactions. Here we characterize the role of ScyD, ScyE, and ScyF, which were logically predicted to be responsible for late biosynthetic steps, in the biological context of N. punctiforme. In-frame deletion mutants of each were constructed (ΔscyD, ΔscyE, and ΔscyF) and their phenotypes studied. Expectedly, ΔscyE presents a scytoneminless phenotype, but no accumulation of the predicted intermediaries. Surprisingly, ΔscyD retains scytonemin production, implying that it is not required for biosynthesis. Indeed, scyD presents an interesting evolutionary paradox: it likely originated in a duplication event from scyE, and unlike other genes in the operon, it has not been subjected to purifying selection. This would suggest that it is a pseudogene, and yet scyD is highly conserved in the scytonemin operon of cyanobacteria. ΔscyF also retains scytonemin production, albeit exhibiting a reduction of the production yield compared with the wild-type. This indicates that ScyF is not essential but may play an adjuvant role for scytonemin synthesis. Altogether, our findings suggest that these downstream genes are not responsible, as expected, for the late steps of scytonemin synthesis and we must look for those functions elsewhere. These findings are particularly important for biotechnological production of this sunscreen through heterologous expression of its genes in more tractable organisms. PMID:27242750

  15. Streptococcus salivarius ATCC 25975 possesses at least two genes coding for primer-independent glucosyltransferases.

    PubMed Central

    Simpson, C L; Giffard, P M; Jacques, N A

    1995-01-01

    Fractionation of the culture medium showed that Streptococcus salivarius ATCC 25975 secreted a glucosyltransferase (Gtf) that was primer independent. On the basis of this observation, a gene library of S. salivarius chromosomal DNA cloned into lambda L47.1 was screened for a gene(s) coding for such an activity. As a result of this screening process, two new gtf genes, gtfL and gtfM, both of which coded for primer-independent Gtf activities, were isolated. GtfL produced an insoluble glucan that was refractory to digestion by the endo-(1-->6)-alpha-D-glucanase. of Chaetonium gracile, while GtfM produced a soluble glucan that was readily degraded by the glucanase. Comparison of the deduced amino acid sequences of gtfL and gtfM with 10 other available Gtf sequences allowed the relatedness of the conserved catalytic regions to be assessed. This analysis showed that the 12 enzymes did not form clusters based on their primer dependencies or on their product solubilities. Further analysis of the YG repeats in the C-terminal glucan-binding domains of GtfJ, GtfK, GtfL, and GtfM from S. salivarius showed that there was strong homology between a block of contiguous triplet YG repeats present in the four alleles. These blocks of YG repeats were coded for by a region of each gene that appeared to have arisen as a result of a recent duplication event(s). PMID:7822030

  16. Induction of secondary metabolism of Aspergillus terreus ATCC 20542 in the batch bioreactor cultures.

    PubMed

    Boruta, Tomasz; Bizukojc, Marcin

    2016-04-01

    Cultivation of Aspergillus terreus ATCC 20542 in a stirred tank bioreactor was performed to induce the biosynthesis of secondary metabolites and provide the bioprocess-related insights into the metabolic capabilities of the investigated strain. The activation of biosynthetic routes was attempted by the diversification of process conditions and growth media. Several strategies were tested, including the addition of rapeseed oil or inulin, changing the concentration of nitrogen source, reduction of chlorine supply, cultivation under saline conditions, and using various aeration schemes. Fifteen secondary metabolites were identified in the course of the study by using ultra-high performance liquid chromatography coupled with mass spectrometry, namely mevinolinic acid, 4a,5-dihydromevinolinic acid, 3α-hydroxy-3,5-dihydromonacolin L acid, terrein, aspulvinone E, dihydroisoflavipucine, (+)-geodin, (+)-bisdechlorogeodin, (+)-erdin, asterric acid, butyrolactone I, desmethylsulochrin, questin, sulochrin, and demethylasterric acid. The study also presents the collection of mass spectra that can serve as a resource for future experiments. The growth in a salt-rich environment turned out to be strongly inhibitory for secondary metabolism and the formation of dense and compact pellets was observed. Generally, the addition of inulin, reducing the oxygen supply, and increasing the content of nitrogen source did not enhance the production of examined molecules. The most successful strategy involved the addition of rapeseed oil to the chlorine-deficient medium. Under these conditions, the highest levels of butyrolactone I, asterric acid, and mevinolinic acid were achieved and the presence of desmethylsulochrin and (+)-bisdechlorogeodin was detected in the broth. The constant and relatively high aeration rate in the idiophase was shown to be beneficial for terrein and (+)-geodin biosynthesis. PMID:26603760

  17. The Complete Genome Sequence of the Marine, Chemolithoautotrophic, Ammonia-Oxidizing Bacterium Nitrosococcus oceani ATCC19707

    SciTech Connect

    Klotz, M G; Arp, D J; Chain, P S; El-Sheikh, A F; Hauser, L J; Hommes, N G; Larimer, F W; Malfatti, S A; Norton, J M; Poret-Peterson, A T; Vergez, L M; Ward, B B

    2006-08-03

    The Gammaproteobacterium, Nitrosococcus oceani (ATCC 19707), is a Gram-negative obligate chemolithoautotroph capable of extracting energy and reducing power from the oxidation of ammonia to nitrite. Sequencing and annotation of the genome revealed a single circular chromosome (3,481,691 bp; 50.4% G+C) and a plasmid (40,420 bp) that contain 3052 and 41 candidate protein-encoding genes, respectively. The genes encoding proteins necessary for the function of known modes of lithotrophy and autotrophy were identified. In contrast to betaproteobacterial nitrifier genomes, the N. oceani genome contained two complete rrn operons. In contrast, only one copy of the genes needed to synthesize functional ammonia monooxygenase and hydroxylamine oxidoreductase, as well as the proteins that relay the extracted electrons to a terminal electron acceptor were identified. The N. oceani genome contained genes for 13 complete two-component systems. The genome also contained all the genes needed to reconstruct complete central pathways, the tricarboxylic acid cycle and the Embden-Meyerhof-Parnass and pentose phosphate pathways. The N. oceani genome contains the genes required to store and utilize energy from glycogen inclusion bodies and sucrose. Polyphosphate and pyrophosphate appear to be integrated in this bacterium's energy metabolism, stress tolerance and the ability to assimilate carbon via gluconeogenesis. One set of genes for type I RuBisCO was identified, while genes necessary for methanotrophy and for carboxysome formation were not identified. The N. oceani genome contains two copies each of the genes or operons necessary to assemble functional complexes I and IV as well as ATP synthase (one H{sup +}-dependent F{sub 0}F{sub 1}-type, one Na{sup +}-dependent V-type).

  18. Mutational Studies of Putative Biosynthetic Genes for the Cyanobacterial Sunscreen Scytonemin in Nostoc punctiforme ATCC 29133.

    PubMed

    Ferreira, Daniela; Garcia-Pichel, Ferran

    2016-01-01

    The heterocyclic indole-alkaloid scytonemin is a sunscreen found exclusively among cyanobacteria. An 18-gene cluster is responsible for scytonemin production in Nostoc punctiforme ATCC 29133. The upstream genes scyABCDEF in the cluster are proposed to be responsible for scytonemin biosynthesis from aromatic amino acid substrates. In vitro studies of ScyA, ScyB, and ScyC proved that these enzymes indeed catalyze initial pathway reactions. Here we characterize the role of ScyD, ScyE, and ScyF, which were logically predicted to be responsible for late biosynthetic steps, in the biological context of N. punctiforme. In-frame deletion mutants of each were constructed (ΔscyD, ΔscyE, and ΔscyF) and their phenotypes studied. Expectedly, ΔscyE presents a scytoneminless phenotype, but no accumulation of the predicted intermediaries. Surprisingly, ΔscyD retains scytonemin production, implying that it is not required for biosynthesis. Indeed, scyD presents an interesting evolutionary paradox: it likely originated in a duplication event from scyE, and unlike other genes in the operon, it has not been subjected to purifying selection. This would suggest that it is a pseudogene, and yet scyD is highly conserved in the scytonemin operon of cyanobacteria. ΔscyF also retains scytonemin production, albeit exhibiting a reduction of the production yield compared with the wild-type. This indicates that ScyF is not essential but may play an adjuvant role for scytonemin synthesis. Altogether, our findings suggest that these downstream genes are not responsible, as expected, for the late steps of scytonemin synthesis and we must look for those functions elsewhere. These findings are particularly important for biotechnological production of this sunscreen through heterologous expression of its genes in more tractable organisms. PMID:27242750

  19. Regulation of Three Nitrogenase Gene Clusters in the Cyanobacterium Anabaena variabilis ATCC 29413

    PubMed Central

    Thiel, Teresa; Pratte, Brenda S.

    2014-01-01

    The filamentous cyanobacterium Anabaena variabilis ATCC 29413 fixes nitrogen under aerobic conditions in specialized cells called heterocysts that form in response to an environmental deficiency in combined nitrogen. Nitrogen fixation is mediated by the enzyme nitrogenase, which is very sensitive to oxygen. Heterocysts are microxic cells that allow nitrogenase to function in a filament comprised primarily of vegetative cells that produce oxygen by photosynthesis. A. variabilis is unique among well-characterized cyanobacteria in that it has three nitrogenase gene clusters that encode different nitrogenases, which function under different environmental conditions. The nif1 genes encode a Mo-nitrogenase that functions only in heterocysts, even in filaments grown anaerobically. The nif2 genes encode a different Mo-nitrogenase that functions in vegetative cells, but only in filaments grown under anoxic conditions. An alternative V-nitrogenase is encoded by vnf genes that are expressed only in heterocysts in an environment that is deficient in Mo. Thus, these three nitrogenases are expressed differentially in response to environmental conditions. The entire nif1 gene cluster, comprising at least 15 genes, is primarily under the control of the promoter for the first gene, nifB1. Transcriptional control of many of the downstream nif1 genes occurs by a combination of weak promoters within the coding regions of some downstream genes and by RNA processing, which is associated with increased transcript stability. The vnf genes show a similar pattern of transcriptional and post-transcriptional control of expression suggesting that the complex pattern of regulation of the nif1 cluster is conserved in other cyanobacterial nitrogenase gene clusters. PMID:25513762

  20. Characterization of Five β-Glycoside Hydrolases from Cellulomonas fimi ATCC 484

    PubMed Central

    Gao, Juan

    2014-01-01

    The Gram-positive bacterium Cellulomonas fimi produces a large array of carbohydrate-active enzymes. Analysis of the collection of carbohydrate-active enzymes from the recent genome sequence of C. fimi ATCC 484 shows a large number of uncharacterized genes for glycoside hydrolase (GH) enzymes potentially involved in biomass utilization. To investigate the enzymatic activity of potential β-glucosidases in C. fimi, genes encoding several GH3 enzymes and one GH1 enzyme were cloned and recombinant proteins were expressed in Escherichia coli. Biochemical analysis of these proteins revealed that the enzymes exhibited different substrate specificities for para-nitrophenol-linked substrates (pNP), disaccharides, and oligosaccharides. Celf_2726 encoded a bifunctional enzyme with β-d-xylopyranosidase and α-l-arabinofuranosidase activities, based on pNP-linked substrates (CfXyl3A). Celf_0140 encoded a β-d-glucosidase with activity on β-1,3- and β-1,6-linked glucosyl disaccharides as well as pNP-β-Glc (CfBgl3A). Celf_0468 encoded a β-d-glucosidase with hydrolysis of pNP-β-Glc and hydrolysis/transglycosylation activities only on β-1,6-linked glucosyl disaccharide (CfBgl3B). Celf_3372 encoded a GH3 family member with broad aryl-β-d-glycosidase substrate specificity. Celf_2783 encoded the GH1 family member (CfBgl1), which was found to hydrolyze pNP-β-Glc/Fuc/Gal, as well as cellotetraose and cellopentaose. CfBgl1 also had good activity on β-1,2- and β-1,3-linked disaccharides but had only very weak activity on β-1,4/6-linked glucose. PMID:25225266

  1. Genome Sequence and Analysis of the Oral Bacterium Fusobacterium nucleatum Strain ATCC 25586

    PubMed Central

    Kapatral, Vinayak; Anderson, Iain; Ivanova, Natalia; Reznik, Gary; Los, Tamara; Lykidis, Athanasios; Bhattacharyya, Anamitra; Bartman, Allen; Gardner, Warren; Grechkin, Galina; Zhu, Lihua; Vasieva, Olga; Chu, Lien; Kogan, Yakov; Chaga, Oleg; Goltsman, Eugene; Bernal, Axel; Larsen, Niels; D'Souza, Mark; Walunas, Theresa; Pusch, Gordon; Haselkorn, Robert; Fonstein, Michael; Kyrpides, Nikos; Overbeek, Ross

    2002-01-01

    We present a complete DNA sequence and metabolic analysis of the dominant oral bacterium Fusobacterium nucleatum. Although not considered a major dental pathogen on its own, this anaerobe facilitates the aggregation and establishment of several other species including the dental pathogens Porphyromonas gingivalis and Bacteroides forsythus. The F. nucleatum strain ATCC 25586 genome was assembled from shotgun sequences and analyzed using the ERGO bioinformatics suite (http://www.integratedgenomics.com). The genome contains 2.17 Mb encoding 2,067 open reading frames, organized on a single circular chromosome with 27% GC content. Despite its taxonomic position among the gram-negative bacteria, several features of its core metabolism are similar to that of gram-positive Clostridium spp., Enterococcus spp., and Lactococcus spp. The genome analysis has revealed several key aspects of the pathways of organic acid, amino acid, carbohydrate, and lipid metabolism. Nine very-high-molecular-weight outer membrane proteins are predicted from the sequence, none of which has been reported in the literature. More than 137 transporters for the uptake of a variety of substrates such as peptides, sugars, metal ions, and cofactors have been identified. Biosynthetic pathways exist for only three amino acids: glutamate, aspartate, and asparagine. The remaining amino acids are imported as such or as di- or oligopeptides that are subsequently degraded in the cytoplasm. A principal source of energy appears to be the fermentation of glutamate to butyrate. Additionally, desulfuration of cysteine and methionine yields ammonia, H2S, methyl mercaptan, and butyrate, which are capable of arresting fibroblast growth, thus preventing wound healing and aiding penetration of the gingival epithelium. The metabolic capabilities of F. nucleatum revealed by its genome are therefore consistent with its specialized niche in the mouth. PMID:11889109

  2. Genome sequence and analysis of the oral bacterium Fusobacterium nucleatum strain ATCC 25586.

    PubMed

    Kapatral, Vinayak; Anderson, Iain; Ivanova, Natalia; Reznik, Gary; Los, Tamara; Lykidis, Athanasios; Bhattacharyya, Anamitra; Bartman, Allen; Gardner, Warren; Grechkin, Galina; Zhu, Lihua; Vasieva, Olga; Chu, Lien; Kogan, Yakov; Chaga, Oleg; Goltsman, Eugene; Bernal, Axel; Larsen, Niels; D'Souza, Mark; Walunas, Theresa; Pusch, Gordon; Haselkorn, Robert; Fonstein, Michael; Kyrpides, Nikos; Overbeek, Ross

    2002-04-01

    We present a complete DNA sequence and metabolic analysis of the dominant oral bacterium Fusobacterium nucleatum. Although not considered a major dental pathogen on its own, this anaerobe facilitates the aggregation and establishment of several other species including the dental pathogens Porphyromonas gingivalis and Bacteroides forsythus. The F. nucleatum strain ATCC 25586 genome was assembled from shotgun sequences and analyzed using the ERGO bioinformatics suite (http://www.integratedgenomics.com). The genome contains 2.17 Mb encoding 2,067 open reading frames, organized on a single circular chromosome with 27% GC content. Despite its taxonomic position among the gram-negative bacteria, several features of its core metabolism are similar to that of gram-positive Clostridium spp., Enterococcus spp., and Lactococcus spp. The genome analysis has revealed several key aspects of the pathways of organic acid, amino acid, carbohydrate, and lipid metabolism. Nine very-high-molecular-weight outer membrane proteins are predicted from the sequence, none of which has been reported in the literature. More than 137 transporters for the uptake of a variety of substrates such as peptides, sugars, metal ions, and cofactors have been identified. Biosynthetic pathways exist for only three amino acids: glutamate, aspartate, and asparagine. The remaining amino acids are imported as such or as di- or oligopeptides that are subsequently degraded in the cytoplasm. A principal source of energy appears to be the fermentation of glutamate to butyrate. Additionally, desulfuration of cysteine and methionine yields ammonia, H(2)S, methyl mercaptan, and butyrate, which are capable of arresting fibroblast growth, thus preventing wound healing and aiding penetration of the gingival epithelium. The metabolic capabilities of F. nucleatum revealed by its genome are therefore consistent with its specialized niche in the mouth. PMID:11889109

  3. Regulation of fructose metabolism and polymer synthesis by Fusobacterium nucleatum ATCC 10953.

    PubMed Central

    Robrish, S A; Thompson, J

    1990-01-01

    Energy for the anaerobic growth of Fusobacterium nucleatum ATCC 10953 can be derived from the fermentation of sugar (fructose) or amino acid (glutamate). During growth on fructose, the cells formed large intracellular granules which after extraction yielded glucose by either acid or enzymatic hydrolysis. The endogenous polymer was subsequently metabolized, and after overnight incubation of the cells in buffer, the glucan granules were no longer detectable by electron microscopy. Anaerobically, washed cells grown previously on fructose fermented this sugar to a mixture of lactic, acetic, and butyric acids, and little intracellular glucan was formed. Aerobically, the cells slowly metabolized fructose to acetate. Provision of glutamic acid as an additional energy (ATP) source elicited rapid synthesis of polymer by glycolyzing cells. Intracellular granules were not present in glutamate-grown cells, and under anaerobic conditions, the resting cells failed to metabolize [14C] fructose. However, the addition of glutamic acid to the suspension resulted in the rapid accumulation of sugar by the cells. Approximately 15% of the 14C-labeled material was extractable with boiling water, and by 31P nuclear magnetic resonance spectroscopy, this phosphorylated derivative was identified as [14C]fructose-1-phosphate. The nonextractable material represented [14C]glucan polymer. Fructose-1-phosphate kinase activity in fructose-grown cells was fivefold greater than that in glutamate-grown cells. We suggest that the activity of fructose-1-phosphate kinase and the availability of ATP regulate the flow of fructose into either the glycolytic or polymer-synthesizing pathway in F. nucleatum. Images PMID:2211506

  4. Cloning and Characterization of Three Fatty Alcohol Oxidase Genes from Candida tropicalis Strain ATCC 20336

    PubMed Central

    Eirich, L. Dudley; Craft, David L.; Steinberg, Lisa; Asif, Afreen; Eschenfeldt, William H.; Stols, Lucy; Donnelly, Mark I.; Wilson, C. Ron

    2004-01-01

    Candida tropicalis (ATCC 20336) converts fatty acids to long-chain dicarboxylic acids via a pathway that includes among other reactions the oxidation of ω-hydroxy fatty acids to ω-aldehydes by a fatty alcohol oxidase (FAO). Three FAO genes (one gene designated FAO1 and two putative allelic genes designated FAO2a and FAO2b), have been cloned and sequenced from this strain. A comparison of the DNA sequence homology and derived amino acid sequence homology between these three genes and previously published Candida FAO genes indicates that FAO1 and FAO2 are distinct genes. Both genes were individually cloned and expressed in Escherichia coli. The substrate specificity and Km values for the recombinant FAO1 and FAO2 were significantly different. Particularly striking is the fact that FAO1 oxidizes ω-hydroxy fatty acids but not 2-alkanols, whereas FAO2 oxidizes 2-alkanols but not ω-hydroxy fatty acids. Analysis of extracts of strain H5343 during growth on fatty acids indicated that only FAO1 was highly induced under these conditions. FAO2 contains one CTG codon, which codes for serine (amino acid 177) in C. tropicalis but codes for leucine in E. coli. An FAO2a construct, with a TCG codon (codes for serine in E. coli) substituted for the CTG codon, was prepared and expressed in E. coli. Neither the substrate specificity nor the Km values for the FAO2a variant with a serine at position 177 were radically different from those of the variant with a leucine at that position. PMID:15294826

  5. Physiological and transcriptional response of Lactobacillus casei ATCC 334 to acid stress.

    PubMed

    Broadbent, Jeff R; Larsen, Rebecca L; Deibel, Virginia; Steele, James L

    2010-05-01

    This study investigated features of the acid tolerance response (ATR) in Lactobacillus casei ATCC 334. To optimize ATR induction, cells were acid adapted for 10 or 20 min at different pH values (range, 3.0 to 5.0) and then acid challenged at pH 2.0. Adaptation over a broad range of pHs improved acid tolerance, but the highest survival was noted in cells acid adapted for 10 or 20 min at pH 4.5. Analysis of cytoplasmic membrane fatty acids (CMFAs) in acid-adapted cells showed that they had significantly (P < 0.05) higher total percentages of saturated and cyclopropane fatty acids than did control cells. Specifically, large increases in the percentages of C(14:0), C(16:1n(9)), C(16:0), and C(19:0(11c)) were noted in the CMFAs of acid-adapted and acid-adapted, acid-challenged cells, while C(18:1n(9)) and C(18:1n(11)) showed the greatest decrease. Comparison of the transcriptome from control cells (grown at pH 6.0) against that from cells acid adapted for 20 min at pH 4.5 indicated that acid adaption invoked a stringent-type response that was accompanied by other functions which likely helped these cells resist acid damage, including malolactic fermentation and intracellular accumulation of His. Validation of microarray data was provided by experiments that showed that L. casei survival at pH 2.5 was improved at least 100-fold by chemical induction of the stringent response or by the addition of 30 mM malate or 30 mM histidine to the acid challenge medium. To our knowledge, this is the first report that intracellular histidine accumulation may be involved in bacterial acid resistance. PMID:20207759

  6. DNA sequence and comparison of virulence plasmids from Rhodococcus equi ATCC 33701 and 103.

    PubMed

    Takai, S; Hines, S A; Sekizaki, T; Nicholson, V M; Alperin, D A; Osaki, M; Takamatsu, D; Nakamura, M; Suzuki, K; Ogino, N; Kakuda, T; Dan, H; Prescott, J F

    2000-12-01

    The virulence plasmids of the equine virulent strains Rhodococcus equi ATCC 33701 and 103 were sequenced, and their genetic structure was analyzed. p33701 was 80,610 bp in length, and p103 was 1 bp shorter; their sequences were virtually identical. The plasmids contained 64 open reading frames (ORFs), 22 of which were homologous with genes of known function and 3 of which were homologous with putative genes of unknown function in other species. Putative functions were assigned to five ORFs based on protein family characteristics. The most striking feature of the virulence plasmids was the presence of a 27,536-bp pathogenicity island containing seven virulence-associated protein (vap) genes, including vapA. These vap genes have extensive homology to vapA, which encodes a thermoregulated and surface-expressed protein. The pathogenicity island contained a LysR family transcriptional regulator and a two-component response regulator upstream of six of the vap genes. The vap genes were present as a cluster of three (vapA, vapC, and vapD), as a pair (vapE and vapF), or individually (vapG; vapH). A region of extensive direct repeats of unknown function, possibly associated with thermoregulation, was present immediately upstream of the clustered and the paired genes but not the individual vap genes. There was extensive homology among the C-terminal halves of all vap genes but not generally among the N-terminal halves. The remainder of the plasmid consisted of a large region which appears to be associated with conjugation functions and a large region which appears to be associated with replication and partitioning functions. PMID:11083803

  7. A Model of Cyclic Transcriptomic Behavior in Cyanobacterium Cyanothece sp. ATCC 51142

    SciTech Connect

    McDermott, Jason E.; Oehmen, Christopher S.; McCue, Lee Ann; Hill, Eric A.; Choi, Daniel M.; Stockel, Jana; Liberton, Michelle L.; Pakrasi, Himadri B.; Sherman, Louis A.

    2011-07-01

    Systems biology attempts to reconcile large amounts of disparate data with existing knowledge to provide models of functioning biological systems. Useful and predictive models aim to summarize complex and dynamic processes and represent the relationships between these processes. The cyanobacterial Cyanothece species Strain sp. ATCC 51142 is an excellent candidate for such systems studies because: (i) it displays tight functional regulation as it must separate the opposing processes of oxygen-generating photosynthesis and oxygen-sensitive nitrogen fixation temporally in the same cell, ; (ii) it has robust cyclic patterns at the genetic, protein and metabolomic levels, ; and (iii) and it has potential applications for bioenergy and carbon sequestration, and thus a predictive model of its function is of practical use. We have represented the transcriptomic data from Cyanothece 51142 under diurnal light/dark cycles as a high-level functional abstraction and describe development of a predictive in silico model of diurnal and circadian behavior in terms of regulatory and metabolic processes in Cyanothece 51142. Our model provides a way to integrate disparate data types into a framework that can be used to explain behavior, generate high-quality predictions for validation, and to suggest future experiments. We show that incorporating network topology into the model improves performance in terms of our ability to explain the behavior of the system under new conditions. The model presented robustly describes transcriptomic behavior of Cyanothece 51142 under different cyclic and non-cyclic growth conditions robustly, and represents a significant advance in the understanding of gene regulation in this important organism.

  8. Phosphotransbutyrylase from Clostridium acetobutylicum ATCC 824 and its role in acidogenesis.

    PubMed Central

    Wiesenborn, D P; Rudolph, F B; Papoutsakis, E T

    1989-01-01

    Phosphotransbutyrylase (phosphate butyryltransferase [EC 2.3.1.19]) from Clostridium acetobutylicum ATCC 824 was purified approximately 200-fold to homogeneity with a yield of 13%. Steps used in the purification procedure were fractional precipitation with (NH4)2SO4, Phenyl Sepharose CL-4B chromatography, DEAE-Sephacel chromatography, high-pressure liquid chromatography with an anion-exchange column, and high-pressure liquid chromatography with a hydrophobic-interaction column. Gel filtration and denaturing gel electrophoresis data were consistent with a native enzyme having eight 31,000-molecular-weight subunits. Within the physiological range of pH 5.5 to 7, the enzyme was very sensitive to pH change in the butyryl phosphate-forming direction and showed virtually no activity below pH 6. This finding indicates that a change in internal pH may be one important factor in the regulation of the enzyme. The enzyme was less sensitive to pH change in the reverse direction. The enzyme could use a number of substrates in addition to butyryl coenzyme A (butyryl-CoA) but had the highest relative activity with butyryl-CoA, isovaleryl-CoA, and valeryl-CoA. The Km values at 30 degrees C and pH 8.0 for butyryl-CoA, phosphate, butyryl phosphate, and CoASH (reduced form of CoA) were 0.11, 14, 0.26, and 0.077 mM, respectively. Results of product inhibition studies were consistent with a random Bi Bi binding mechanism in which phosphate binds at more than one site. Images PMID:2719475

  9. gamma-Glutamyltransferase from the outer cell envelope of Treponema denticola ATCC 35405.

    PubMed Central

    Mäkinen, P L; Mäkinen, K K

    1997-01-01

    The human oral spirochete Treponema denticola ATCC 35405 was shown to exhibit relatively high enzyme activity toward the gamma-glutamyl amide bond present in N-gamma-L-glutamyl-4-nitroaniline. The enzyme responsible for this catalysis (gamma-glutamyltransferase [GGT]; EC 2.3.2.2) was purified by means of fast protein liquid chromatography to two sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE)-pure forms from a mild (0.1%) Triton X-100 extract of washed cells. The GGT was studied primarily with regard to its hydrolytic activity by using N-gamma-L-glutamyl-4-nitroaniline as a substrate, although the GGT was shown to catalyze transpeptidation reactions. The high-molecular-mass form of the GGT gave a value of about 213 kDa by SDS-PAGE when heat treatment was omitted and one of 26 kDa after heat treatment; mass spectrometry gave a value of 26.877. The larger form may represent an aggregate with nonprotein structures (possibly of a carbohydrate nature). The preliminary N-terminal sequence of the GGT is MKKPLIGITGSXLYETSQXXF. The enzyme was highly active on glutathione, transferring its Glu residue either to a water molecule or to the Gly-L-Leu dipeptide. The GGT stability was absolutely dependent on the presence of free thiol(s), while no evidence of metalloenzyme nature was obtained. The proposed location of the GGT in the outer cell envelope and its high activity on glutathione, a major nonprotein thiol present in virtually all cells, suggest that the GGT may play a role in the propagation of T. denticola within inflamed periodontal tissues. PMID:9009331

  10. Dechlorination of chlorophenols using extracellular peroxidases produced by streptomyces albus ATCC 3005.

    PubMed

    Antonopoulos, V T.; Rob, A; Ball, A S.; Wilson, M T.

    2001-07-01

    Streptomyces albus ATCC 3005 was found to produce higher levels of extracellular peroxidase activity (3.420 U mg(-1)) than previously reported for any other actinomycete. Maximum peroxidase activity was obtained after 72 h of incubation at a temperature of 30 degrees C in a liquid medium (pH 7.6) containing (in w/v) 0.8% to 0.9% oat spelts xylan and 0.6% yeast extract, corresponding to a C:N ratio of around 8.4:1. Characterization of the peroxidases revealed that the optimal temperature for peroxidase activity, using the standard 2,4-dichlorophenol (2,4-DCP) assay was 53 degrees C, when the enzyme reaction was performed at pH 7.2. A study of the effect of temperature on the stability of peroxidase over time, showed that the enzyme was stable at 40 degrees C, with a half-life of 224 min, while at higher temperatures the stability and activity was reduced such that at 50 degrees C and 70 degrees C the half-life of the enzyme was 50 min and 9 min respectively. The optimum pH for the activity of the enzyme occurred between pH 8.1 and 10.4. In terms of substrate specificity, the peroxidase was able to catalyze a broad range of substrates including 2,4-DCP, L-3,4-dihydroxyphenylalanine (L-DOPA), 2,4,5-trichlorophenol and other chlorophenols in the presence of hydrogen peroxide. Ion exchange chromatography was used to confirm that the enzyme was able to release chloride ions from a range of chlorophenols. PMID:11427236

  11. Regulation of Three Nitrogenase Gene Clusters in the Cyanobacterium Anabaena variabilis ATCC 29413.

    PubMed

    Thiel, Teresa; Pratte, Brenda S

    2014-01-01

    The filamentous cyanobacterium Anabaena variabilis ATCC 29413 fixes nitrogen under aerobic conditions in specialized cells called heterocysts that form in response to an environmental deficiency in combined nitrogen. Nitrogen fixation is mediated by the enzyme nitrogenase, which is very sensitive to oxygen. Heterocysts are microxic cells that allow nitrogenase to function in a filament comprised primarily of vegetative cells that produce oxygen by photosynthesis. A. variabilis is unique among well-characterized cyanobacteria in that it has three nitrogenase gene clusters that encode different nitrogenases, which function under different environmental conditions. The nif1 genes encode a Mo-nitrogenase that functions only in heterocysts, even in filaments grown anaerobically. The nif2 genes encode a different Mo-nitrogenase that functions in vegetative cells, but only in filaments grown under anoxic conditions. An alternative V-nitrogenase is encoded by vnf genes that are expressed only in heterocysts in an environment that is deficient in Mo. Thus, these three nitrogenases are expressed differentially in response to environmental conditions. The entire nif1 gene cluster, comprising at least 15 genes, is primarily under the control of the promoter for the first gene, nifB1. Transcriptional control of many of the downstream nif1 genes occurs by a combination of weak promoters within the coding regions of some downstream genes and by RNA processing, which is associated with increased transcript stability. The vnf genes show a similar pattern of transcriptional and post-transcriptional control of expression suggesting that the complex pattern of regulation of the nif1 cluster is conserved in other cyanobacterial nitrogenase gene clusters. PMID:25513762

  12. Metabolic flux analysis of Cyanothece sp. ATCC 51142 under mixotrophic conditions.

    PubMed

    Alagesan, Swathi; Gaudana, Sandeep B; Sinha, Avinash; Wangikar, Pramod P

    2013-11-01

    Cyanobacteria are a group of photosynthetic prokaryotes capable of utilizing solar energy to fix atmospheric carbon dioxide to biomass. Despite several "proof of principle" studies, low product yield is an impediment in commercialization of cyanobacteria-derived biofuels. Estimation of intracellular reaction rates by (13)C metabolic flux analysis ((13)C-MFA) would be a step toward enhancing biofuel yield via metabolic engineering. We report (13)C-MFA for Cyanothece sp. ATCC 51142, a unicellular nitrogen-fixing cyanobacterium, known for enhanced hydrogen yield under mixotrophic conditions. Rates of reactions in the central carbon metabolism under nitrogen-fixing and -non-fixing conditions were estimated by monitoring the competitive incorporation of (12)C and (13)C from unlabeled CO2 and uniformly labeled glycerol, respectively, into terminal metabolites such as amino acids. The observed labeling patterns suggest mixotrophic growth under both the conditions, with a larger fraction of unlabeled carbon in nitrate-sufficient cultures asserting a greater contribution of carbon fixation by photosynthesis and an anaplerotic pathway. Indeed, flux analysis complements the higher growth observed under nitrate-sufficient conditions. On the other hand, the flux through the oxidative pentose phosphate pathway and tricarboxylic acid cycle was greater in nitrate-deficient conditions, possibly to supply the precursors and reducing equivalents needed for nitrogen fixation. In addition, an enhanced flux through fructose-6-phosphate phosphoketolase possibly suggests the organism's preferred mode under nitrogen-fixing conditions. The (13)C-MFA results complement the reported predictions by flux balance analysis and provide quantitative insight into the organism's distinct metabolic features under nitrogen-fixing and -non-fixing conditions. PMID:23954952

  13. 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.

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

    PubMed Central

    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 degrees C. The alpha-amylase was stable at an acidic pH but very sensitive to thermal inactivation. It hydrolyzed soluble starch, with a Km of 3.6 g . liter-1 and a Kcat of 122 mol of reducing sugars . s-1 . mol-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. Images PMID:8967771

  15. 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

  16. Fluorene and phenanthrene uptake by Pseudomonas putida ATCC 17514: kinetics and physiological aspects.

    PubMed

    Rodrigues, Ana C; Wuertz, Stefan; Brito, António G; Melo, Luís F

    2005-05-01

    Pseudomonas putida ATCC 17514 was used as a model strain to investigate the characteristics of bacterial growth in the presence of solid fluorene and phenanthrene. Despite the lower water-solubility of phenanthrene, P. putida degraded this polycyclic aromatic hydrocarbon (PAH) at a maximum observed rate of 1.4 +/- 0.1 mg L(-1) h(-1), higher than the apparent degradation rate of fluorene, 0.8 +/- 0.07 mg L(-1) h(-1). The role of physiological processes on the biodegradation of these PAHs was analyzed and two different uptake strategies were identified. Zeta potential measurements revealed that phenanthrene-grown cells were slightly more negatively charged (-57.5 +/- 4.7 mV) than fluorene-grown cells (-51.6 +/- 4.9 mV), but much more negatively charged than glucose-grown cells (-26.8 +/- 3.3 mV), suggesting that the PAH substrate induced modifications on the physical properties of bacterial surfaces. Furthermore, protein-to-exopolysaccharide ratios detected during bacterial growth on phenanthrene were typical of biofilms developed under physicochemical stress conditions, caused by the presence of sparingly water-soluble chemicals as the sole carbon and energy source for growth, the maximum value for TP/EPS during growth on phenanthrene (1.9) being lower than the one obtained with fluorene (5.5). Finally, confocal laser microscopy observations using a gfp-labeled derivative strain revealed that, in the presence of phenanthrene, P. putida::gfp cells formed a biofilm on accessible crystal surfaces, whereas in the presence of fluorene the strain grew randomly between the crystal clusters. The results showed that P. putida was able to overcome the lower aqueous solubility of phenanthrene by adhering to the solid PAH throughout the production of extracellular polymeric substances, thus promoting the availability and uptake of such a hydrophobic compound. PMID:15800860

  17. Surface-associated material from the bacterium Actinobacillus actinomycetemcomitans contains a peptide which, in contrast to lipopolysaccharide, directly stimulates fibroblast interleukin-6 gene transcription.

    PubMed

    Reddi, K; Nair, S P; White, P A; Hodges, S; Tabona, P; Meghji, S; Poole, S; Wilson, M; Henderson, B

    1996-03-15

    The oral commensal Gram-negative bacterium Actinobacillus actinomycetemcomitans is believed to be the causative organism of localized juvenile periodontitis, a disease in which there is rapid loss of alveolar bone supporting the teeth. Previously, we have reported that gentle saline extraction of this bacterium removed a loosely adherent proteinaceous fraction from the cell surface of the bacterium, which we have termed surface-associated material. This material contained potent bone-resorbing activity. We now report that surface-associated material is also a potent stimulator of cytokines, and in particular, interleukin-6 (IL-6) synthesis, while the lipopolysaccharide from this bacterium is only a weak stimulator of IL-6 synthesis by fibroblasts and monocytes. In contrast to enteric lipopolysaccharide (LPS), which induces fibroblast IL-1, IL-6 and tumour necrosis factor (TNF) alpha synthesis, surface-associated material stimulated gingival fibroblasts to synthesize only IL-6, with no induction of IL-1 or TNF (the normal inducers of IL-6 synthesis). Reverse transcriptase PCR also failed to detect mRNA for IL-1 or TNF in surface-associated-material-stimulated fibroblasts, although both mRNAs were present in Escherichia coli LPS-stimulated cells. Neutralizing antibodies to IL-1 and/or TNF or the natural IL-1 receptor antagonist (IL-1ra) inhibited enteric LPS-induced IL-6 synthesis, but did not inhibit surface-associated-material-induced synthesis. In addition, dexamethasone, which completely suppressed LPS-induced IL-6 synthesis, only inhibited surface-associated-material-induced IL-6 synthesis by 50%. This suggests that the active constituent in the surface-associated material stimulates IL-6 gene transcription by a transcriptional control mechanism distinct to that of E. coli LPS. The IL-6 stimulating activity of the surface-associated material is inhibited by both heat and trypsin, suggesting that it is proteinaceous. The activity has been isolated using anion

  18. Comparative genomics of citric-acid producing Aspergillus niger ATCC 1015 versus enzyme-producing CBS 513.88

    SciTech Connect

    Andersen, Mikael R.; Salazar, Margarita; Schaap, Peter; van de Vondervoort, Peter; Culley, David E.; Thykaer, Jette; Frisvad, Jens C.; Nielsen, Kristian F.; Albang, Richard; Albermann, Kaj; Berka, Randy; Braus, Gerhard; Braus-Stromeyer, Susanna A.; Corrochano, Luis; Dai, Ziyu; van Dijck, Piet; Hofmann, Gerald; Lasure, Linda L.; Magnuson, Jon K.; Menke, Hildegard; Meijer, Martin; Meijer, Susan; Nielsen, Jakob B.; Nielsen, Michael L.; van Ooyen, Albert; Pel, Herman J.; Poulsen, Lars; Samson, Rob; Stam, Hein; Tsang, Adrian; van den Brink, Johannes M.; ATkins, Alex; Aerts, Andrea; Shapiro, Harris; Pangilinan, Jasmyn; Salamov, Asaf; Lou, Yigong; Lindquist, Erika; Lucas, Susan; Grimwood, Jane; Grigoriev, Igor V.; Kubicek, Christian P.; Martinez, Diego; van Peij, Noel; Roubos, Johannes A.; Nielsen, Jens B.; Baker, Scott E.

    2011-06-01

    The filamentous fungus Aspergillus niger exhibits great diversity in its phenotype. It is found globally, both as marine and terrestrial strains, produces both organic acids and hydrolytic enzymes in high amounts, and some isolates exhibit pathogenicity. Although the genome of an industrial enzyme-producing A. niger strain (CBS 513.88) has already been sequenced, the versatility and diversity of this species compels additional exploration. We therefore undertook whole genome sequencing of the acidogenic A. niger wild type strain (ATCC 1015), and produced a genome sequence of very high quality. Only 15 gaps are present in the sequence and half the telomeric regions have been elucidated. Moreover, sequence information from ATCC 1015 was utilized to improve the genome sequence of CBS 513.88. Chromosome-level comparisons uncovered several genome rearrangements, deletions, a clear case of strain-specific horizontal gene transfer, and identification of 0.8 megabase of novel sequence. Single nucleotide polymorphisms per kilobase (SNPs/kb) between the two strains were found to be exceptionally high (average: 7.8, maximum: 160 SNPs/kb). High variation within the species was confirmed with exo-metabolite profiling and phylogenetics. Detailed lists of alleles were generated, and genotypic differences were observed to accumulate in metabolic pathways essential to acid production and protein synthesis. A transcriptome analysis revealed up-regulation of the electron transport chain, specifically the alternative oxidative pathway in ATCC 1015, while CBS 513.88 showed significant up regulation of genes associated with biosynthesis of amino acids that are abundant in glucoamylase A, tRNA-synthases and protein transporters.

  19. Analysis of the genome sequence of Lactobacillus gasseri ATCC 33323 reveals the molecular basis of an autochthonous intestinal organism.

    PubMed

    Azcarate-Peril, M Andrea; Altermann, Eric; Goh, Yong Jun; Tallon, Richard; Sanozky-Dawes, Rosemary B; Pfeiler, Erika A; O'Flaherty, Sarah; Buck, B Logan; Dobson, Alleson; Duong, Tri; Miller, Michael J; Barrangou, Rodolphe; Klaenhammer, Todd R

    2008-08-01

    This study presents the complete genome sequence of Lactobacillus gasseri ATCC 33323, a neotype strain of human origin and a native species found commonly in the gastrointestinal tracts of neonates and adults. The plasmid-free genome was 1,894,360 bp in size and predicted to encode 1,810 genes. The GC content was 35.3%, similar to the GC content of its closest relatives, L. johnsonii NCC 533 (34%) and L. acidophilus NCFM (34%). Two identical copies of the prophage LgaI (40,086 bp), of the Sfi11-like Siphoviridae phage family, were integrated tandomly in the chromosome. A number of unique features were identified in the genome of L. gasseri that were likely acquired by horizontal gene transfer and may contribute to the survival of this bacterium in its ecological niche. L. gasseri encodes two restriction and modification systems, which may limit bacteriophage infection. L. gasseri also encodes an operon for production of heteropolysaccharides of high complexity. A unique alternative sigma factor was present similar to that of B. caccae ATCC 43185, a bacterial species isolated from human feces. In addition, L. gasseri encoded the highest number of putative mucus-binding proteins (14) among lactobacilli sequenced to date. Selected phenotypic characteristics that were compared between ATCC 33323 and other human L. gasseri strains included carbohydrate fermentation patterns, growth and survival in bile, oxalate degradation, and adhesion to intestinal epithelial cells, in vitro. The results from this study indicated high intraspecies variability from a genome encoding traits important for survival and retention in the gastrointestinal tract. PMID:18539810

  20. Design and production of functionalized biopolyesters by Methylobacterium extorquens ATCC 55366: Toward new tissue engineering materials

    NASA Astrophysics Data System (ADS)

    Hoefer, Heinrich Friedrich Philipp Till Nikolaus

    Vascular networks are required to support the formation and function of three-dimensional tissues. Biodegradable scaffolds are being considered in order to promote vascularization where natural regeneration of lost or destroyed vascular networks fails. Particularly; composite materials are expected to fulfill the complex demands of a patient's body to support wound healing. Microbial biopolyesters are being regarded as such second and third generation biomaterials. Methylobacterium extorquens is one of several microorganisms that should be considered for the production of advanced polyhydroxyalkanoates (PHAs). M. extorquens displays a distinct advantage in that it is able to utilize methanol as an inexpensive substrate for growth and biopolyester production. The design of functionalized PHAs, which would be made of both saturated short-chain-length (scl, C ≤ 5) and unsaturated medium-chain-length (mcl, 6 ≤ C ≤ 14) monomeric units, aimed at combining desirable material properties of inert scl/mcl-PHAs with those of functionalized mcl-PHAs. By independently inserting the phaC1 or the phaC2 gene from Pseudomonas fluorescens GK13, recombinant M. extorquens strains were obtained which were capable of producing PHAs containing C-C double bonds. A fermentation process was developed to obtain gram quantities of biopolyesters employing the recombinant M. extorquens ATCC 55366 strain which harbored the phaC2 gene of P. fluorescens GK13, the better one of the two strains at incorporating unsaturated monomeric units. The PHAs produced were found in a blend of scl-PHAs and functionalized scl/mcl-PHAs (4 ≤ C ≤ 6), which were the products of the native and of the recombinant PHA synthase, respectively. Thermo-mechanical analysis confirmed that the functionalized scl/mcl-PHAs exhibited the desirable material properties expected. This project contributed to current research on polyhydroxyalkanoates at different levels. The terminal double bonds of the functionalized scl

  1. Transcriptomic and genomic analysis of cellulose fermentation by Clostridium thermocellum ATCC 27405

    SciTech Connect

    Raman, Babu; McKeown, Catherine K; Rodriguez, Jr., Miguel; Brown, Steven D; Mielenz, Jonathan R

    2011-01-01

    The ability of Clostridium thermocellum ATCC 27405 wild-type strain to hydrolyze cellulose and ferment the degradation products directly to ethanol and other metabolic byproducts makes it an attractive candidate for consolidated bioprocessing of cellulosic biomass to biofuels. In this study, whole-genome microarrays were used to investigate the expression of C. thermocellum mRNA during growth on crystalline cellulose in controlled replicate batch fermentations. A time-series analysis of gene expression revealed changes in transcript levels of {approx}40% of genes ({approx}1300 out of 3198 ORFs encoded in the genome) during transition from early-exponential to late-stationary phase. K-means clustering of genes with statistically significant changes in transcript levels identified six distinct clusters of temporal expression. Broadly, genes involved in energy production, translation, glycolysis and amino acid, nucleotide and coenzyme metabolism displayed a decreasing trend in gene expression as cells entered stationary phase. In comparison, genes involved in cell structure and motility, chemotaxis, signal transduction and transcription showed an increasing trend in gene expression. Hierarchical clustering of cellulosome-related genes highlighted temporal changes in composition of this multi-enzyme complex during batch growth on crystalline cellulose, with increased expression of several genes encoding hydrolytic enzymes involved in degradation of non-cellulosic substrates in stationary phase. Overall, the results suggest that under low substrate availability, growth slows due to decreased metabolic potential and C. thermocellum alters its gene expression to (i) modulate the composition of cellulosomes that are released into the environment with an increased proportion of enzymes than can efficiently degrade plant polysaccharides other than cellulose, (ii) enhance signal transduction and chemotaxis mechanisms perhaps to sense the oligosaccharide hydrolysis products

  2. Characterizations of Metal Binding in the Active Sites of Acireductone Dioxygenase Isoforms from Klebsiella ATCC 8724

    SciTech Connect

    Chai,S.; Ju, T.; Dang, M.; Goldsmith, R.; Maroney, M.; Pochapsky, T.

    2008-01-01

    The two acireductone dioxygenase (ARD) isozymes from the methionine salvage pathway of Klebsiella ATCC 8724 present an unusual case in which two enzymes with different structures and distinct activities toward their common substrates (1, 2-dihydroxy-3-oxo-5-(methylthio)pent-1-ene and dioxygen) are derived from the same polypeptide chain. Structural and functional differences between the two isozymes are determined by the type of M2+ metal ion bound in the active site. The Ni2+-bound NiARD catalyzes an off-pathway shunt from the methionine salvage pathway leading to the production of formate, methylthiopropionate, and carbon monoxide, while the Fe2+-bound FeARD' catalyzes the on-pathway formation of methionine precursor 2-keto-4-methylthiobutyrate and formate. Four potential protein-based metal ligands were identified by sequence homology and structural considerations. Based on the results of site-directed mutagenesis experiments, X-ray absorption spectroscopy (XAS), and isothermal calorimetry measurements, it is concluded that the same four residues, His96, His98, Glu102 and His140, provide the protein-based ligands for the metal in both the Ni- and Fe-containing forms of the enzyme, and subtle differences in the local backbone conformations trigger the observed structural and functional differences between the FeARD' and NiARD isozymes. Furthermore, both forms of the enzyme bind their respective metals with pseudo-octahedral geometry, and both may lose a histidine ligand upon binding of substrate under anaerobic conditions. However, mutations at two conserved nonligand acidic residues, Glu95 and Glu100, result in low metal contents for the mutant proteins as isolated, suggesting that some of the conserved charged residues may aid in transfer of metal from in vivo sources or prevent the loss of metal to stronger chelators. The Glu100 mutant reconstitutes readily but has low activity. Mutation of Asp101 results in an active enzyme that incorporates metal in vivo but

  3. Characterization of Metal Binding in the Active Sites of acireductone dioxygenase Isoforms from Klebsiella ATCC 8724

    SciTech Connect

    S Chai; T Ju; M Dang; R Goldsmith; M Maroney; T Pochapsky

    2011-12-31

    The two acireductone dioxygenase (ARD) isozymes from the methionine salvage pathway of Klebsiella ATCC 8724 present an unusual case in which two enzymes with different structures and distinct activities toward their common substrates (1,2-dihydroxy-3-oxo-5-(methylthio)pent-1-ene and dioxygen) are derived from the same polypeptide chain. Structural and functional differences between the two isozymes are determined by the type of M{sup 2+} metal ion bound in the active site. The Ni{sup 2+}-bound NiARD catalyzes an off-pathway shunt from the methionine salvage pathway leading to the production of formate, methylthiopropionate, and carbon monoxide, while the Fe{sup 2+}-bound FeARD catalyzes the on-pathway formation of methionine precursor 2-keto-4-methylthiobutyrate and formate. Four potential protein-based metal ligands were identified by sequence homology and structural considerations. Based on the results of site-directed mutagenesis experiments, X-ray absorption spectroscopy (XAS), and isothermal calorimetry measurements, it is concluded that the same four residues, His96, His98, Glu102 and His140, provide the protein-based ligands for the metal in both the Ni- and Fe-containing forms of the enzyme, and subtle differences in the local backbone conformations trigger the observed structural and functional differences between the FeARD and NiARD isozymes. Furthermore, both forms of the enzyme bind their respective metals with pseudo-octahedral geometry, and both may lose a histidine ligand upon binding of substrate under anaerobic conditions. However, mutations at two conserved nonligand acidic residues, Glu95 and Glu100, result in low metal contents for the mutant proteins as isolated, suggesting that some of the conserved charged residues may aid in transfer of metal from in vivo sources or prevent the loss of metal to stronger chelators. The Glu100 mutant reconstitutes readily but has low activity. Mutation of Asp101 results in an active enzyme that incorporates

  4. Interaction of Wild Strains of Aspergilla with Aspergillus parasiticus ATCC15517 and Aflatoxin Production †

    PubMed Central

    Martins, H. Marina; Almeida, Inês; Marques, Marta; Bernardo, Fernando

    2008-01-01

    Aflatoxins are secondary metabolites produced by some competent mould strains of Aspergillus flavus, A. parasiticus and A. nomius. These compounds have been extensively studied with regards to their toxicity for animals and humans; they are able to induce liver cancer and may cause a wide range of adverse effects in living organisms. Aflatoxins are found as natural contaminants of food and feed; the main line of the strategy to control them is based on the prevention of the mould growth in raw vegetable or during its storage and monitoring of each crop batch. Mould growth is conditioned by many ecological factors, including biotic ones. Hazard characterization models for aflatoxins in crops must take into consideration biotic interactions between moulds and their potential effects on growth development. The aim of this work is to study the effect of the biotic interaction of 14 different wild strains of Aspergilla (different species), with a competent strain (Aspergillus parasiticus ATCC 15517) using an in vitro production model. The laboratory model used was a natural matrix (humidified cracked corn), on which each wild strain challenged the aflatoxin production of a producer strain. Cultures were incubated at 28°C for 12 days and sampled at the 8th and 12th. Aflatoxin detection and quantification was performed by HPLC using a procedure with a MRPL = 1 μg/kg. Results of those interactive cultures revealed both synergic and antagonistic effects on aflatoxin biosynthesis. Productivity increases were particularly evident on the 8th day of incubation with wild strains of A. flavipes (+ 70.4 %), A. versicolor (+ 54.9 %) and A. flavus 3 (+ 62.6 %). Antagonistic effects were found with A. niger (− 69.5%), A. fumigatus (− 47.6 %) and A. terreus (− 47.6 %) on the 12th day. The increased effects were more evident on the 8th of incubation and the decreases were more patent on the 12th day. Results show that the development of Aspergilla strains concomitantly with

  5. Inhibition of Listeria monocytogenes ATCC 19115 on ham steak by tea bioactive compounds incorporated into chitosan-coated plastic films

    PubMed Central

    2012-01-01

    Background The consumer demands for better quality and safety of food products have given rise to the development and implementation of edible films. The use of antimicrobial films can be a promising tool for controlling L. monocytogenes on ready to eat products. The aim of this study was to develop effective antimicrobial films incorporating bioactive compounds from green and black teas into chitosan, for controlling L. monocytogenes ATCC 19115 on vacuum-packaged ham steak. The effectiveness of these antimicrobial films was evaluated at room temperature (20°C) for 10 days and at refrigerated temperature (4°C) for 8 weeks. Results The HPLC results clearly show that relative concentrations of catechins and caffeine in green tea ranked EGCG>EGC>CAF>ECG>EC>C while in black tea extracts ranked CAF>EGCG>ECG>EGC>EC>C. The chitosan-coated plastic films incorporating green tea and black tea extracts shows specific markers identified by FTIR. Incorporating natural extracts into chitosan showed that the growth of L monocytogenes ATCC 19115 was inhibited. The efficacy of antimicrobial effect of tea extracts incorporated into chitosan-coated plastic film was dose dependent. However, chitosan-coated films without addition of tea extracts did not inhibit the growth of L. monocytogenes ATCC 19115. Chitosan-coated plastic films incorporating 4% Green tea extract was the most effective antimicrobial, reducing the initial counts from 3.2 to 2.65 log CFU/cm2 during room temperature storage and from 3.2 to 1–1.5 log CFU/cm2 during refrigerated storage. Conclusions Incorporation of tea extracts into the chitosan-coated films considerably enhanced their effectiveness against L. monocytogenes ATCC 19115. 4% Green tea incorporated into chitosan-coated plastic film had a better antilisterial effect than 2% green tea or 2% and 4% black tea. Data from this study would provide new formulation options for developing antimicrobial packaging films using tea extracts to improve the

  6. Isolation, sequence analysis, and comparison of two plasmids (28 and 29 kilobases) from the biomining bacterium Leptospirillum ferrooxidans ATCC 49879.

    PubMed

    Coram, Nicolette J; van Zyl, Leonardo J; Rawlings, Douglas E

    2005-11-01

    Two plasmids, of 28,878 bp and 28,012 bp, were isolated from Leptospirillum ferrooxidans ATCC 49879. Altogether, a total of 67 open reading frames (ORFs) were identified on both plasmids, of which 32 had predicted products with high homology to proteins of known function, while 11 ORFs had predicted products with homology to previously identified proteins of unknown function. Twenty-four ORFs had products with no homologues in the GenBank/NCBI database. An analysis of the ORFs and other features of the two plasmids, the first to be isolated from a bacterium of the genus Leptospirillum, is presented. PMID:16269793

  7. Comparative genomics of citric-acid producing Aspergillus niger ATCC 1015 versus enzyme-producing CBS 513.88

    SciTech Connect

    Grigoriev, Igor V.; Baker, Scott E.; Andersen, Mikael R.; Salazar, Margarita P.; Schaap, Peter J.; Vondervoot, Peter J.I. van de; Culley, David; Thykaer, Jette; Frisvad, Jens C.; Nielsen, Kristen F.; Albang, Richard; Albermann, Kaj; Berka, Randy M.; Braus, Gerhard H.; Braus-Stromeyer, Susanna A.; Corrochano, Luis M.; Dai, Ziyu; Dijck, Piet W.M. van; Hofmann, Gerald; Lasure, Linda L.; Magnusson, Jon K.; Meijer, Susan L.; Nielsen, Jakob B.; Nielsen, Michael L.; Ooyen, Albert J.J. van; Panther, Kathyrn S.; Pel, Herman J.; Poulsen, Lars; Samson, Rob A.; Stam, Hen; Tsang, Adrian; Brink, Johannes M. van den; Atkins, Alex; Aerts, Andrea; Shapiro, Harris; Pangilinan, Jasmyn; Salamov, Asaf; Lou, Yigong; Lindquist, Erika; Lucas, Susan; Grimwood, Jane; Kubicek, Christian P.; Martinez, Diego; Peij, Noel N.M.E. van; Roubos, Johannes A.; Nielsen, Jens

    2011-04-28

    The filamentous fungus Aspergillus niger exhibits great diversity in its phenotype. It is found globally, both as marine and terrestrial strains, produces both organic acids and hydrolytic enzymes in high amounts, and some isolates exhibit pathogenicity. Although the genome of an industrial enzyme-producing A. niger strain (CBS 513.88) has already been sequenced, the versatility and diversity of this species compels additional exploration. We therefore undertook whole genome sequencing of the acidogenic A. niger wild type strain (ATCC 1015), and produced a genome sequence of very high quality. Only 15 gaps are present in the sequence and half the telomeric regions have been elucidated. Moreover, sequence information from ATCC 1015 was utilized to improve the genome sequence of CBS 513.88. Chromosome-level comparisons uncovered several genome rearrangements, deletions, a clear case of strain-specific horizontal gene transfer, and identification of 0.8 megabase of novel sequence. Single nucleotide polymorphisms per kilobase (SNPs/kb) between the two strains were found to be exceptionally high (average: 7.8, maximum: 160 SNPs/kb). High variation within the species was confirmed with exo-metabolite profiling and phylogenetics. Detailed lists of alleles were generated, and genotypic differences were observed to accumulate in metabolic pathways essential to acid production and protein synthesis. A transcriptome analysis revealed up-regulation of the electron transport chain, specifically the alternative oxidative pathway in ATCC 1015, while CBS 513.88 showed significant up-regulation of genes relevant to glucoamylase A production, such as tRNA-synthases and protein transporters. Our results and datasets from this integrative systems biology analysis resulted in a snapshot of fungal evolution and will support further optimization of cell factories based on filamentous fungi.[Supplemental materials (10 figures, three text documents and 16 tables) have been made available

  8. Microbial biotransformation of 16α,17-epoxy-ent-kaurane-19-oic acid by Beauveria sulfurescens ATCC 7159-F.

    PubMed

    Furtado, Ricardo A; Gunaherath, G M Kamal B; Bastos, Jairo K; Gunatilak, A A Leslie

    2013-08-01

    Biotransformation of 16alpha,17-epoxy-ent-kaurane-19-oic acid (1) by Beauveria sulfurescens ATCC 7159-F led to the production of a new ent-kaurane diterpenoid, 7beta,17-dihydroxy-ent-kaur-15-en-19-oic acid (7), and four other ent-kauranes (8 - 11), all of which were identified as their methyl esters. Compounds 9 and 10 were found to be new stereoisomers. Structures of these were established by the extensive usage of their spectroscopic characteristics. PMID:24079162

  9. Next-generation sequencing-based genome-wide mutation analysis of L-lysine-producing Corynebacterium glutamicum ATCC 21300 strain.

    PubMed

    Lee, Chang-Soo; Nam, Jae-Young; Son, Eun-Suk; Kwon, O-Chul; Han, Woorijarang; Cho, Jae-Yong; Park, Young-Jin

    2012-10-01

    In order to identify single nucleotide polymorphism and insertion/deletion mutations, we performed whole-genome re-sequencing of the enhanced L-lysine-producing Corynebacterium glutamicum ATCC 21300 strain. In total, 142 single nucleotide polymorphisms and 477 insertion/deletion mutations were identified in the ATCC 21300 strain when compared to 3,434 predicted genes of the wild-type C. glutamicum ATCC 13032 strain. Among them, 110 transitions and 29 transversions of single nucleotide polymorphisms were found from genes of the ATCC 21300 strain. In addition, 11 genes, involved in the L-lysine biosynthetic pathway and central carbohydrate metabolism, contained mutations including single nucleotide polymorphisms and insertions/deletions. Interestingly, RT-PCR analysis of these 11 genes indicated that they were normally expressed in the ATCC 21300 strain. This information of genome-wide gene-associated variations will be useful for genome breeding of C. glutamicum in order to develop an industrial amino acid-producing strain with minimal mutation. PMID:23124757

  10. Development of a potential functional food prepared with pigeon pea (Cajanus cajan), oats and Lactobacillus reuteri ATCC 55730.

    PubMed

    Barboza, Yasmina; Márquez, Enrique; Parra, Katynna; Piñero, M Patricia; Medina, Luis M

    2012-11-01

    The purpose of this study was to investigate the survival of Lactobacillus reuteri ATCC 55730 in creams, prepared with pigeon peas and oat. Products were analysed to determine their content of protein, fibre, fat, carbohydrates and degree of likeness. Viable numbers of L. reuteri and pH were determined after 1, 7, 14, 21 and 28 days of storage at 4°C. Results showed significant differences (P < 0.05) in protein, fat, fibre and carbohydrate content between creams. No significant differences (P > 0.05) were found on sensory quality between control and creams with L. reuteri. After 28 days, the cell viability was above 7 log cfu/g in all creams. L. reuteri ATCC 55730 had the highest viability in cream with 40% pigeon pea and 20% oat (8.16 log cfu/g). In conclusion, due to its acceptability and highly nutritious value, the product could be used so as to support the growth of L. reuteri. PMID:22533458

  11. Complete structure of the cell surface polysaccharide of Streptococcus oralis ATCC 10557: A receptor for lectin-mediated interbacterial adherence

    SciTech Connect

    Abeygunawardana, C.; Bush, C.A. ); Cisar, J.O. )

    1991-07-02

    Lectin-carbohydrate binding is known to play an important role in a number of different cell-cell interactions including those between certain species of oral streptococci and actinomyces that colonize teeth. The cell wall polysaccharides of Streptococcus oralis ATCC 10557, S. oralis 34, and Streptococcus mitis J22, although not identical antigenically, each function as a receptor molecule for the galactose and N-acetylgalactosamine reactive fimbrial lectins of Actinomyces viscosus and Actinomyces naeslundii. Carbohydrate analysis of the receptor polysaccharide isolated from S. oralis ATCC 10557 shows galactose (3 mol), glucose (1 mol), GalNAc (1 mol), and rhamnose (1 mol). {sup 1}H NMR spectra of the polysaccharide show that is partially O-acetylated. Analysis of the {sup 1}H NMR spectrum of the de-O-acetylated polysaccharide shows that it is composed of repeating subunits containing six monosaccharides and that the subunits are joined by a phosphodiester linkage. The {sup 1}H and {sup 13}C NMR spectra were completely assigned by two-dimensional homonuclear correlation methods and by {sup 1}H-detected heteronuclear multiple-quantum correlation ({sup 1}H({sup 13}C)HMQC). The complete {sup 1}H and {sup 13}C assignment of the native polysaccharide was carried out by the same techniques augmented by a {sup 13}C-coupled hybrid HMQC-COSY method, which is shown to be especially useful for carbohydrates in which strong coupling and overlapping peaks in the {sup 1}H spectrum pose difficulties.

  12. A Novel Metallo-β-Lactamase Involved in the Ampicillin Resistance of Streptococcus pneumoniae ATCC 49136 Strain

    PubMed Central

    Chang, Chia-Yu; Lin, Hui-Jen; Li, Yaw-Kuen

    2016-01-01

    Streptococcus pneumoniae, a penicillin-sensitive bacterium, is recognized as a major cause of pneumonia and is treated clinically with penicillin-based antibiotics. The rapid increase in resistance to penicillin and other antibiotics affects 450 million people globally and results in 4 million deaths every year. To unveil the mechanism of resistance of S. pneumoniae is thus an important issue to treat streptococcal disease that might consequently save millions of lives around the world. In this work, we isolated a streptococci-conserved L-ascorbate 6-phosphate lactonase, from S. pneumoniae ATCC 49136. This protein reveals a metallo-β-lactamase activity in vitro, which is able to deactivate an ampicillin-based antibiotic by hydrolyzing the amide bond of the β-lactam ring. The Michaelis parameter (Km) = 25 μM and turnover number (kcat) = 2 s-1 were obtained when nitrocefin was utilized as an optically measurable substrate. Through confocal images and western blot analyses with a specific antibody, the indigenous protein was recognized in S. pneumoniae ATCC 49136. The protein-overexpressed S. pneumonia exhibits a high ampicillin-tolerance ability in vivo. In contrast, the protein-knockout S. pneumonia reveals the ampicillin-sensitive feature relative to the wild type strain. Based on these results, we propose that this protein is a membrane-associated metallo-β-lactamase (MBL) involved in the antibiotic-resistant property of S. pneumoniae. PMID:27214294

  13. Inactivation of Listeria monocytogenes ATCC 7644 on fresh-cut tomato using nisin in combinations with organic salts.

    PubMed

    Oladunjoye, Adebola O; Singh, Suren; Ijabadeniyi, Oluwatosin A

    2016-01-01

    The inhibition of Listeria monocytogenes ATCC 7644 on fresh-cut tomato was investigated using nisin alone, and in combinations with organic salts. Nisin at a concentration of 5000UI/mL was introduced alone or in combination with an organic salt (sodium citrate or sodium acetate each at 3 and 5g/100mL each) on fresh-cut tomato previously inoculated with 10(8)CFU/mL of L. monocytogenes ATCC 7644. Chlorine at 200ppm was used as a control. The inoculated samples were incubated at different temperatures (4, 10 and 25°C) and examined at 0, 24, 48 and 72h. The effects of the antimicrobial treatments on quality parameters of tomato (pH, soluble solids, titratable acidity and vitamin C) were also evaluated, and colour parameters were observed at the lowest storage temperature for 10 days. Both nisin and the organic salts inhibited growth of L. monocytogenes, but the combinations of two compounds were more effective. The nisin-sodium citrate (5%) combination was significantly (p≤0.05) effective, while chlorine was least effective against L. monocytogenes. The quality parameters were substantially retained, especially at 4°C, suggesting good shelf stability at a low temperature. These results substantiate the use of the cheap and eco-friendly approach to reducing this pathogen of health concern in common fresh produce. PMID:27261167

  14. Oscillating behavior of carbohydrate granule formation and dinitrogen fixation in the cyanobacterium Cyanothece sp. strain ATCC 51142.

    PubMed Central

    Schneegurt, M A; Sherman, D M; Nayar, S; Sherman, L A

    1994-01-01

    It has been shown that some aerobic, unicellular, diazotrophic cyanobacteria temporally separate photosynthetic O2 evolution and oxygen-sensitive N2 fixation. Cyanothece sp. ATCC strain 51142 is an aerobic, unicellular, diazotrophic cyanobacterium that fixes N2 during discrete periods of its cell cycle. When the bacteria are maintained under diurnal light-dark cycles, N2 fixation occurs in the dark. Similar cycling is observed in continuous light, implicating a circadian rhythm. Under N2-fixing conditions, large inclusion granules form between the thylakoid membranes. Maximum granulation, as observed by electron microscopy, occurs before the onset of N2 fixation, and the granules decrease in number during the period of N2 fixation. The granules can be purified from cell homogenates by differential centrifugation. Biochemical analyses of the granules indicate that these structures are primarily carbohydrate, with some protein. Further analyses of the carbohydrate have shown that it is a glucose polymer with some characteristics of glycogen. It is proposed that N2 fixation is driven by energy and reducing power stored in these inclusion granules. Cyanothece sp. strain ATCC 51142 represents an excellent experimental organism for the study of the protective mechanisms of nitrogenase, metabolic events in cyanobacteria under normal and stress conditions, the partitioning of resources between growth and storage, and biological rhythms. Images PMID:8132452

  15. Growth and nitrite and nitrous oxide accumulation of Paracoccus denitrificans ATCC 19367 in the presence of selected pesticides.

    PubMed

    Sáez, Florentina; Pozo, Clementina; Gómez, Miguel Angel; Rodelas, Belén; Gónzalez-López, Jesús

    2003-09-01

    The effects of the application of eight pesticides (aldrin, lindane, dimetoate, methylparathion, methidation, atrazine, simazine, and captan) on growth, respiratory activity (as CO2 production), denitrifying activity (as N2O released), and nitrite accumulation in the culture medium by Paracoccus denitrificans strain ATCC 19367 were studied. The fungicide captan totally inhibited growth and biological activity of P. denitrificans, while the rest of the tested pesticides delayed the growth and CO2 release of P. denitrificans but did not drastically affect the bacterial growth or respiratory capacity after 96 h of culture. The denitrifying activity of P. denitrificans ATCC 19367 (as N2O released) was negatively affected by all tested pesticides. The release of N2O was strongly inhibited by several organochlorinated and organophosphorated insecticides (aldrin, lindane, dimetoate, and methidation), which led to high accumulation of nitrite in the surrounding medium. Atrazine decreased N2O release after 48 h of culture because of negative effects on growth, and methylparathion and simazine delayed the onset of N2O release by P. denitrificans. These three pesticides reduced the accumulation of NO2- compared to unamended control cultures. PMID:12959522

  16. Cytochrome cb-type nitric oxide reductase with cytochrome c oxidase activity from Paracoccus denitrificans ATCC 35512.

    PubMed

    Fujiwara, T; Fukumori, Y

    1996-04-01

    A highly active nitric oxide reductase was purified from Paracoccus denitrificans ATCC 35512, formerly named Thiosphaera pantotropha, which was anaerobically cultivated in the presence of nitrate. The enzyme was composed of two subunits with molecular masses of 34 and 15 kDa and contained two hemes b and one heme c per molecule. Copper was not found in the enzyme. The spectral properties suggested that one of the two hemes b and heme c were in six-coordinated low-spin states and another heme b was in a five-coordinated high-spin state and reacted with carbon monoxide. The enzyme showed high cytochrome c-nitric oxide oxidoreductase activity and formed nitrous oxide from nitric oxide with the expected stoichiometry when P. denitrificans ATCC 35512 ferrocytochrome c-550 was used as the electron donor. The V max and Km values for nitric oxide were 84 micromol of nitric oxide per min/mg of protein and 0.25 microM, respectively. Furthermore, the enzyme showed ferrocytochrome c-550-O2 oxidoreductase activity with a V max of 8.4 micromol of O2 per min/mg of protein and a Km value of 0.9 mM. Both activities were 50% inhibited by about 0.3 mM KCN. PMID:8606159

  17. Efficient simultaneous saccharification and fermentation of inulin to 2,3-butanediol by thermophilic Bacillus licheniformis ATCC 14580.

    PubMed

    Li, Lixiang; Chen, Chao; Li, Kun; Wang, Yu; Gao, Chao; Ma, Cuiqing; Xu, Ping

    2014-10-01

    2,3-Butanediol (2,3-BD) is an important starting material for the manufacture of bulk chemicals. For efficient and large-scale production of 2,3-BD through fermentation, low-cost substrates are required. One such substrate, inulin, is a polydisperse fructan found in a wide variety of plants. In this study, a levanase with high inulinase activity and high pH and temperature stability was identified in Bacillus licheniformis strain ATCC 14580. B. licheniformis strain ATCC 14580 was found to efficiently produce 2,3-BD from fructose at 50°C. Then, the levanase was used for simultaneous saccharification and fermentation (SSF) of inulin to 2,3-BD. A fed-batch SSF yielded 103.0 g/liter 2,3-BD in 30 h, with a high productivity of 3.4 g/liter · h. The results suggest that the SSF process developed with the thermophilic B. licheniformis strain used might be a promising alternative for efficient 2,3-BD production from the favorable substrate inulin. PMID:25107977

  18. Oncocin Onc72 is efficacious against antibiotic-susceptible Klebsiella pneumoniae ATCC 43816 in a murine thigh infection model.

    PubMed

    Knappe, Daniel; Adermann, Knut; Hoffmann, Ralf

    2015-11-01

    Oncocins and apidaecins are short proline-rich antimicrobial peptides (PrAMPs) representing novel antibiotic drug lead compounds that kill bacteria after internalization and inhibition of intracellular targets (e.g. 70S ribosome and DnaK). Oncocin Onc72 is highly active against Gram-negative bacteria in vitro and in vivo protecting mice in systemic infection models with Escherichia coli and KPC-producing Klebsiella pneumoniae. Here we studied its efficacy in a murine thigh infection model using meropenem as antibiotic comparator that had a 44-fold higher molar in vitro activity than Onc72. Male CD1 mice were rendered neutropenic using cyclophosphamide for four days before intramuscular infection with K. pneumoniae ATCC 43816. After 75 min oncocin Onc72 or the antibiotic comparator meropenem were administered subcutaneously with 100 mg (43 µmol) and 25 mg (65 µmol) per kg of body weight, respectively, six times every 75 min. Onc72 and meropenem administered subcutaneously reduced the thigh tissue burden of K. pneumoniae ATCC 43816 in neutropenic mice significantly by 4.14 and 4.65 a log10 cfu/g, respectively. The bacterial counts were ∼0.5 and ∼1 log10 below the pre-treatment burden, respectively, indicating bactericidal effects for both compounds. Thus, Onc72 was as efficacious as meropenem in vivo despite its much lower in vitro activity determined according to CLSI standard antimicrobial activity tests. PMID:25968331

  19. Efficient Simultaneous Saccharification and Fermentation of Inulin to 2,3-Butanediol by Thermophilic Bacillus licheniformis ATCC 14580

    PubMed Central

    Li, Lixiang; Chen, Chao; Li, Kun; Wang, Yu; Gao, Chao; Ma, Cuiqing

    2014-01-01

    2,3-Butanediol (2,3-BD) is an important starting material for the manufacture of bulk chemicals. For efficient and large-scale production of 2,3-BD through fermentation, low-cost substrates are required. One such substrate, inulin, is a polydisperse fructan found in a wide variety of plants. In this study, a levanase with high inulinase activity and high pH and temperature stability was identified in Bacillus licheniformis strain ATCC 14580. B. licheniformis strain ATCC 14580 was found to efficiently produce 2,3-BD from fructose at 50°C. Then, the levanase was used for simultaneous saccharification and fermentation (SSF) of inulin to 2,3-BD. A fed-batch SSF yielded 103.0 g/liter 2,3-BD in 30 h, with a high productivity of 3.4 g/liter · h. The results suggest that the SSF process developed with the thermophilic B. licheniformis strain used might be a promising alternative for efficient 2,3-BD production from the favorable substrate inulin. PMID:25107977

  20. Production and characterization of thermostable alkaline protease of Bacillus subtilis (ATCC 6633) from optimized solid-state fermentation.

    PubMed

    Chatterjee, Joyee; Giri, Sudipta; Maity, Sujan; Sinha, Ankan; Ranjan, Ashish; Rajshekhar; Gupta, Suvroma

    2015-01-01

    Proteases are the most important group of enzymes utilized commercially in various arenas of industries, such as food, detergent, leather, dairy, pharmaceutical, diagnostics, and waste management, accounting for nearly 20% of the world enzyme market. Microorganisms of specially Bacillus genera serve as a vast repository of diverse set of industrially important enzymes and utilized for the large-scale enzyme production using a fermentation technology. Approximately 30%-40% of the cost of industrial enzymes originates from the cost of the growth medium. This study is attempted to produce protease from Bacillus subtilis (ATCC 6633) after optimization of various process parameters with the aid of solid-state fermentation using a cheap nutrient source such as wheat bran. B. subtilis (ATCC 6633) produces proteases of molecular weight 36 and 20 kDa, respectively, in the fermented medium as evident from SDS zymogram. Alkaline protease activity has been detected with optimum temperature at 50 °C and is insensitive to ethylenediaminetetraacetic acid. This thermostable alkaline protease exhibits dual pH optimum at 7 and 10 with moderate pH stability at alkaline pH range. It preserves its activity in the presence of detergent such as SDS, Tween 20, and Triton X-100 and may be considered as an effective additive to detergent formulation with some industrial importance. PMID:25323045

  1. Free and immobilized Lactobacillus casei ATCC 393 on whey protein as starter cultures for probiotic Feta-type cheese production.

    PubMed

    Dimitrellou, Dimitra; Kandylis, Panagiotis; Sidira, Marianthi; Koutinas, Athanasios A; Kourkoutas, Yiannis

    2014-01-01

    The use of free and immobilized Lactobacillus casei ATCC 393 on whey protein as starter culture in probiotic Feta-type cheese production was evaluated. The probiotic cultures resulted in significantly higher acidity; lower pH; reduced counts of coliforms, enterobacteria, and staphylococci; and improved quality characteristics compared with cheese with no culture. Microbiological and strain-specific multiplex PCR analysis showed that both free and immobilized L. casei ATCC 393 were detected in the novel products at levels required for conferring a probiotic effect at the end of the ripening. The effect of starter culture on production of volatile compounds was investigated by the solid-phase microextraction gas chromatography-mass spectrometry analysis technique. The immobilized cells resulted in an improved profile of aroma-related compounds and the overall high quality of the novel products was ascertained by the preliminary sensory test. Finally, the high added value produced by exploitation of whey, which is an extremely polluting industrial waste, was highlighted and assessed. PMID:24931523

  2. Oscillating behavior of carbohydrate granule formation and dinitrogen fixation in the cyanobacterium Cyanothece sp. strain ATCC 51142

    NASA Technical Reports Server (NTRS)

    Schneegurt, M. A.; Sherman, D. M.; Nayar, S.; Sherman, L. A.; Mitchell, C. A. (Principal Investigator)

    1994-01-01

    It has been shown that some aerobic, unicellular, diazotrophic cyanobacteria temporally separate photosynthetic O2 evolution and oxygen-sensitive N2 fixation. Cyanothece sp. ATCC strain 51142 is an aerobic, unicellular, diazotrophic cyanobacterium that fixes N2 during discrete periods of its cell cycle. When the bacteria are maintained under diurnal light-dark cycles, N2 fixation occurs in the dark. Similar cycling is observed in continuous light, implicating a circadian rhythm. Under N2-fixing conditions, large inclusion granules form between the thylakoid membranes. Maximum granulation, as observed by electron microscopy, occurs before the onset of N2 fixation, and the granules decrease in number during the period of N2 fixation. The granules can be purified from cell homogenates by differential centrifugation. Biochemical analyses of the granules indicate that these structures are primarily carbohydrate, with some protein. Further analyses of the carbohydrate have shown that it is a glucose polymer with some characteristics of glycogen. It is proposed that N2 fixation is driven by energy and reducing power stored in these inclusion granules. Cyanothece sp. strain ATCC 51142 represents an excellent experimental organism for the study of the protective mechanisms of nitrogenase, metabolic events in cyanobacteria under normal and stress conditions, the partitioning of resources between growth and storage, and biological rhythms.

  3. Purification of acetoacetate decarboxylase from Clostridium acetobutylicum ATCC 824 and cloning of the acetoacetate decarboxylase gene in Escherichia coli

    SciTech Connect

    Petersen, D.J.; Bennett, G.N. )

    1990-11-01

    In Clostridium acetobutylicum ATCC 824, acetoacetate decarboxylase (EC 4.1.1.4) is essential for solvent production, catalyzing the decarboxylation of acetoacetate to acetone. We report here the purification of the enzyme from C. acetobutylicum ATCC 824 and the cloning and expression of the gene encoding the acetoacetate decarboxylase enzyme in Escherichia coli. A bacteriophage lambda EMBL3 library of C. acetobutylicum DNA was screened by plaque hybridization, using oligodeoxynucleotide probes derived from the N-terminal amino acid sequence obtained from the purified protein. Phage DNA from positive plaques was analyzed by Southern hybridization. Restriction mapping and subsequent subcloning of DNA fragments hybridizing to the probes localized the gene within an {approximately}2.1-kb EcoRI/BglII fragment. A polypeptide with a molecular weight of {approximately}28,000 corresponding to that of the purified acetoacetate decarboxylase was observed in both Western blots (immunoblots) and maxicell analysis of whole-cell extracts of E. coli harboring the clostridial gene. Although the expression of the gene is tightly regulated in C. acetobutylicum, it was well expressed in E. coli, although from a promoter sequence of clostridial origin.

  4. Cytochrome cb-type nitric oxide reductase with cytochrome c oxidase activity from Paracoccus denitrificans ATCC 35512.

    PubMed Central

    Fujiwara, T; Fukumori, Y

    1996-01-01

    A highly active nitric oxide reductase was purified from Paracoccus denitrificans ATCC 35512, formerly named Thiosphaera pantotropha, which was anaerobically cultivated in the presence of nitrate. The enzyme was composed of two subunits with molecular masses of 34 and 15 kDa and contained two hemes b and one heme c per molecule. Copper was not found in the enzyme. The spectral properties suggested that one of the two hemes b and heme c were in six-coordinated low-spin states and another heme b was in a five-coordinated high-spin state and reacted with carbon monoxide. The enzyme showed high cytochrome c-nitric oxide oxidoreductase activity and formed nitrous oxide from nitric oxide with the expected stoichiometry when P. denitrificans ATCC 35512 ferrocytochrome c-550 was used as the electron donor. The V max and Km values for nitric oxide were 84 micromol of nitric oxide per min/mg of protein and 0.25 microM, respectively. Furthermore, the enzyme showed ferrocytochrome c-550-O2 oxidoreductase activity with a V max of 8.4 micromol of O2 per min/mg of protein and a Km value of 0.9 mM. Both activities were 50% inhibited by about 0.3 mM KCN. PMID:8606159

  5. Effect of probiotic-fermented milk administration on gastrointestinal survival of Lactobacillus casei ATCC 393 and modulation of intestinal microbial flora.

    PubMed

    Sidira, Marianthi; Galanis, Alex; Ypsilantis, Petros; Karapetsas, Athanasios; Progaki, Zoi; Simopoulos, Constantinos; Kourkoutas, Yiannis

    2010-01-01

    The aim of the present study was to assess the survival of free and immobilized Lactobacillus casei ATCC 393 on apple pieces, contained in probiotic-fermented milk, after gastrointestinal (GI) transit and to investigate the potential regulation of intestinal microbial flora in a rat model. In in vitro GI stress tolerance tests, immobilized L. casei ATCC 393 exhibited significantly higher survival rates compared to free cells. At a second stage, probiotic-fermented milk produced by either free or immobilized cells was administered orally at a single dose or daily for 9 days in Wistar rats. By 12 h after single-dose administration, both free and immobilized cells were detected by microbiological and molecular analysis at levels ≥6 logCFU/g of feces. Moreover, daily administration led to significant reduction of staphylococci, enterobacteria, coliforms and streptococci counts. In conclusion, L. casei ATCC 393 contained in fermented milk survived GI transit and modulated intestinal microbiota. PMID:21160205

  6. Supporting data for comparative proteomic analysis of Listeria monocytogenes ATCC 7644 exposed to a sublethal concentration of nisin

    PubMed Central

    Miyamoto, Kendi Nishino; Mariante Monteiro, Karina; da Silva Caumo, Karin; Rodrigues Lorenzatto, Karina; Bunselmeyer Ferreira, Henrique; Brandelli, Adriano

    2015-01-01

    Here we provide the LC–MS/MS data from a comparative analysis of Listeria monocytogenes ATCC 7644 treated and non-treated with a sublethal concentration of nisin (10−3 mg/mL). Protein samples were analyzed by multidimensional protein identification technology (MudPIT) approach, in an off-line configuration. The raw MS/MS data allowed the detection of 49,591 spectra which resulted in 576 protein identifications. After Scaffold validation, 179 proteins were identified with high confidence. A label-free quantitative analysis based of normalized spectral abundance factor (NSAF) was used and 13 proteins were found differentially expressed between nisin-treated and non-treated cells. Gene ontology analysis of differentially expressed proteins revealed that most of them are correlated to metabolic process, oxidative stress response mechanisms and molecular binding. A detailed analysis and discussion of these data may be found in Miyamoto et al. [1]. PMID:26217729

  7. A refined model for the structure of acireductone dioxygenase from Klebsiella ATCC 8724 incorporating residual dipolar couplings

    PubMed Central

    Pochapsky, Thomas C.; Pochapsky, Susan S.; Ju, Tingting; Hoefler, Chris; Liang, Jue

    2006-01-01

    Summary Acireductone dioxygenase (ARD) from Klebsiella ATCC 8724 is a metalloenzyme that is capable of catalyzing different reactions with the same substrates (acireductone and O2) depending upon the metal bound in the active site. A model for the solution structure of the paramagnetic Ni2+-containing ARD has been refined using residual dipolar couplings (RDCs) measured in two media. Additional dihedral restraints based on chemical shift (TALOS) were included in the refinement, and backbone structure in the vicinity of the active site was modeled from a crystallographic structure of the mouse homolog of ARD. The incorporation of residual dipolar couplings into the structural refinement alters the relative orientations of several structural features significantly, and improves local secondary structure determination. Comparisons between the solution structures obtained with and without RDCs are made, and structural similarities and differences between mouse and bacterial enzymes are described. Finally, the biological significance of these differences is considered. PMID:16518698

  8. Complete genome sequence of Piscirickettsia salmonis LF-89 (ATCC VR-1361) a major pathogen of farmed salmonid fish.

    PubMed

    Pulgar, Rodrigo; Travisany, Dante; Zuñiga, Alejandro; Maass, Alejandro; Cambiazo, Verónica

    2015-10-20

    Piscirickettsia salmonis, the causative agent of salmonid rickettsial septicemia (SRS), is a significant threat to the healthy and sustainable production of salmonid farming industry. This Gram-negative bacterium, originally isolated from a coho salmon in Southern Chile, produces a systemic infection characterized by colonization of several fish organs. P. salmonis is able to infect, survive, and replicate inside salmonid macrophages however little is known about its mechanisms of pathogenesis. Here, we present the whole genome sequence and annotation of the P. salmonis reference strain LF-89 (ATCC VR-1361). The genome contains one circular chromosome of 3,184,851 bp and three plasmids, pPSLF89-1 (180,124 bp), pPSLF89-2 (33,516 bp) and pPSLF89-3 (51,573 bp). A total of 2850 protein-coding genes, 56 tRNAs and six copies of 5S-16S-23S rRNA. PMID:26220311

  9. Supporting data for comparative proteomic analysis of Listeria monocytogenes ATCC 7644 exposed to a sublethal concentration of nisin.

    PubMed

    Miyamoto, Kendi Nishino; Mariante Monteiro, Karina; da Silva Caumo, Karin; Rodrigues Lorenzatto, Karina; Bunselmeyer Ferreira, Henrique; Brandelli, Adriano

    2015-06-01

    Here we provide the LC-MS/MS data from a comparative analysis of Listeria monocytogenes ATCC 7644 treated and non-treated with a sublethal concentration of nisin (10(-3) mg/mL). Protein samples were analyzed by multidimensional protein identification technology (MudPIT) approach, in an off-line configuration. The raw MS/MS data allowed the detection of 49,591 spectra which resulted in 576 protein identifications. After Scaffold validation, 179 proteins were identified with high confidence. A label-free quantitative analysis based of normalized spectral abundance factor (NSAF) was used and 13 proteins were found differentially expressed between nisin-treated and non-treated cells. Gene ontology analysis of differentially expressed proteins revealed that most of them are correlated to metabolic process, oxidative stress response mechanisms and molecular binding. A detailed analysis and discussion of these data may be found in Miyamoto et al. [1]. PMID:26217729

  10. Multi-Omic Dynamics Associate Oxygenic Photosynthesis with Nitrogenase-Mediated H2 Production in Cyanothece sp. ATCC 51142

    PubMed Central

    Bernstein, Hans C.; Charania, Moiz A.; McClure, Ryan S.; Sadler, Natalie C.; Melnicki, Matthew R.; Hill, Eric A.; Markillie, Lye Meng; Nicora, Carrie D.; Wright, Aaron T.; Romine, Margaret F.; Beliaev, Alexander S.

    2015-01-01

    To date, the proposed mechanisms of nitrogenase-driven photosynthetic H2 production by the diazotrophic unicellular cyanobacterium Cyanothece sp. ATCC 51142 have assumed that reductant and ATP requirements are derived solely from glycogen oxidation and cyclic-electron flow around photosystem I. Through genome-scale transcript and protein profiling, this study presents and tests a new hypothesis on the metabolic relationship between oxygenic photosynthesis and nitrogenase-mediated H2 production in Cyanothece 51142. Our results show that net-positive rates of oxygenic photosynthesis and increased expression of photosystem II reaction centers correspond and are synchronized with nitrogenase expression and H2 production. These findings provide a new and more complete view on the metabolic processes contributing to the energy budget of photosynthetic H2 production and highlight the role of concurrent photocatalytic H2O oxidation as a participating process. PMID:26525576

  11. Characterization and replication mode determination of the minimal replicon of Tetragenococcus halophila ATCC33315 plasmid pUCL287.

    PubMed

    Benachour, A; Frère, J; Boutibonnes, P; Auffray, Y

    1995-01-01

    pUCL287 is a cryptic plasmid of Tetragenococcus halophila (formerly Pediococcus halophilus) ATCC33315 of relatively small size (8.7 kb). Its minimal replicon was located on a 1235 bp MamI-EcoRI fragment. This minimal replicon contains a non-translated region, followed by a gene encoding a putative 311 amino acid protein. Deletion experiments showed that the non-translated region corresponds to the replication origin. Determination of the replication mode was carried out in Enterococcus faecalis JH2-2 harboring pUCL287 minimal replicon. The replicating intermediates detected revealed that pUCL287 minimal replicon follows a bidirectional theta replicating mode. PMID:8824766

  12. Biochemical Characterization of the FEZ-1 Metallo-β-Lactamase of Legionella gormanii ATCC 33297T Produced in Escherichia coli

    PubMed Central

    Mercuri, Paola Sandra; Bouillenne, Fabrice; Boschi, Letizia; Lamotte-Brasseur, Josette; Amicosante, Gianfranco; Devreese, Bart; van Beeumen, Jozef; Frère, Jean-Marie; Rossolini, Gian Maria; Galleni, Moreno

    2001-01-01

    The blaFEZ-1 gene coding for the metallo-β-lactamase of Legionella (Fluoribacter) gormanii ATCC 33297T was overexpressed via a T7 expression system in Escherichia coli BL21(DE3)(pLysS). The product was purified to homogeneity in two steps with a yield of 53%. The FEZ-1 metallo-β-lactamase exhibited a broad-spectrum activity profile, with a preference for cephalosporins such as cephalothin, cefuroxime, and cefotaxime. Monobactams were not hydrolyzed. The β-lactamase was inhibited by metal chelators. FEZ-1 is a monomeric enzyme with a molecular mass of 29,440 Da which possesses two zinc-binding sites. Its zinc content did not vary in the pH range of 5 to 9, but the presence of zinc ions modified the catalytic efficiency of the enzyme. A model of the FEZ-1 three-dimensional structure was built. PMID:11257043

  13. Multi-omic dynamics associate oxygenic photosynthesis with nitrogenase-mediated H2 production in Cyanothece sp. ATCC 51142

    DOE PAGESBeta

    Bernstein, Hans C.; Charania, Moiz A.; McClure, Ryan S.; Sadler, Natalie C.; Melnicki, Matthew R.; Hill, Eric A.; Markillie, Lye Meng; Nicora, Carrie D.; Wright, Aaron T.; Romine, Margaret F.; et al

    2015-11-03

    This study combines transcriptomic and proteomic profiling to provide new insights on the metabolic relationship between oxygenic photosynthesis and nitrogenase-mediated H2 production in the model cyanobacterium, Cyanothece sp. ATCC 51142. To date, the proposed mechanisms used to describe the energy metabolism processes that support H2 production in Cyanothece 51142 have assumed that ATP and reductant requirements are derived solely from glycogen oxidation and/or cyclic-electron flow around photosystem I. The results from this study present and test an alternative hypothesis by showing that net-positive rates of oxygenic photosynthesis and increased expression of photosystem II reaction centers correspond and are synchronized withmore » nitrogenase expression and H2 production. These findings provide a new and more complete view on the metabolic processes contributing to the energy budget of photosynthetic H2 production and highlight the likely role of photocatalytic H2O oxidation as a major participating process.« less

  14. High quality permanent draft genome sequence of Phaseolibacter flectens ATCC 12775T, a plant pathogen of French bean pods

    DOE PAGESBeta

    Aizenberg-Gershtein, Yana; Izhaki, Ido; Lapidus, Alla; Copeland, Alex; Reddy, TBK; Huntemann, Marcel; Pillay, Manoj; Markowitz, Victor; Göker, Markus; Woyke, Tanja; et al

    2016-01-13

    We report that the Phaseolibacter flectens strain ATCC 12775T (Halpern et al., Int J Syst Evol Microbiol 63:268–273, 2013) is a Gram-negative, rod shaped, motile, aerobic, chemoorganotroph bacterium. Ph. flectens is as a plant-pathogenic bacterium on pods of French bean and was first identified by Johnson (1956) as Pseudomonas flectens. After its phylogenetic position was reexamined, Pseudomonas flectens was transferred to the family Enterobacteriaceae as Phaseolibacter flectens gen. nov., comb. nov. Here we describe the features of this organism, together with the draft genome sequence and annotation. The DNA GC content is 44.34 mol%. The chromosome length is 2,748,442 bp.more » It encodes 2,437 proteins and 89 RNA genes. Ph. flectens genome is part of the Genomic Encyclopedia of Type Strains, Phase I: the one thousand microbial genomes study.« less

  15. Identification of a cryptic type III polyketide synthase (1,3,6,8-tetrahydroxynaphthalene synthase) from Streptomyces peucetius ATCC 27952.

    PubMed

    Ghimire, Gopal Prasad; Oh, Tae-Jin; Liou, Kwangkyoung; Sohng, Jae Kyung

    2008-10-31

    We identified a 1,134-bp putative type III polyketide synthase from the sequence analysis of Streptomyces peucetius ATCC 27952, named Sp-RppA, which is characterized as 1,3,6,8-tetrahydroxynaphthalene synthase and shares 33% identity with SCO1206 from S. coelicolor A3(2) and 32% identity with RppA from S. griseus. The 1,3,6,8-tetrahydroxynaphthalene synthase is known to catalyze the sequential decarboxylative condensation, intramolecular cyclization, and aromatization of an oligoketide derived from five units of malonyl-CoA to give 1,3,6,8-tetrahydroxynaphthalene, which spontaneously oxidizes to form 2,5,7-trihydroxy-1,4-naphthoquinone (flaviolin). In this study, we report the in vivo expression and in vitro synthesis of flaviolin from purified gene product (Sp-RppA). PMID:18612244

  16. Cloning and sequencing of the trpE gene from Arthrobacter globiformis ATCC 8010 and several related subsurface Arthrobacter isolates

    SciTech Connect

    Chernova, T.; Viswanathan, V.K.; Austria, N.; Nichols, B.P.

    1998-09-01

    Tryptophan dependent mutants of Arthrobacter globiformis ATCC 8010 were isolated and trp genes were cloned by complementation and marker rescue of the auxotrophic strains. Rescue studies and preliminary sequence analysis reveal that at least the genes trpE, trpC, and trpB are clustered together in this organism. In addition, sequence analysis of the entire trpE gene, which encodes component I of anthranilate synthase, is described. Segments of the trpE gene from 17 subsurface isolates of Arthrobacter sp. were amplified by PCR and sequenced. The partial trpE sequences from the various strains were aligned and subjected to phylogenetic analysis. The data suggest that in addition to single base changes, recombination and genetic exchange play a major role in the evolution of the Arthrobacter genome.

  17. Development of an Unnatural Amino Acid Incorporation System in the Actinobacterial Natural Product Producer Streptomyces venezuelae ATCC 15439.

    PubMed

    He, Jingxuan; Van Treeck, Briana; Nguyen, Han B; Melançon, Charles E

    2016-02-19

    Many Actinobacteria, most notably Streptomyces, produce structurally diverse bioactive natural products, including ribosomally synthesized peptides, by multistep enzymatic pathways. The use of site-specific genetic incorporation of unnatural amino acids to investigate and manipulate the functions of natural product biosynthetic enzymes, enzyme complexes, and ribosomally derived peptides in these organisms would have important implications for drug discovery and development efforts. Here, we have designed, constructed, and optimized unnatural amino acid systems capable of incorporating p-iodo-l-phenylalanine and p-azido-l-phenylalanine site-specifically into proteins in the model natural product producer Streptomyces venezuelae ATCC 15439. We observed notable differences in the fidelity and efficiency of these systems between S. venezuelae and previously used hosts. Our findings serve as a foundation for using an expanded genetic code in Streptomyces to address questions related to natural product biosynthesis and mechanism of action that are relevant to drug discovery and development. PMID:26562751

  18. Factors affecting the photoproduction of ammonia from dinitrogen and water by the cyanobacterium Anabaena sp. strain ATCC 33047

    SciTech Connect

    Ramos, J.L.; Guerrero, M.G.; Losada, M.

    1987-04-01

    Synthesis of ammonia from dinitrogen and water by suspensions of Anabaena sp. strain ATCC 33047 treated with the glutamine synthetase inhibitor L-methionine-D,L-sulfoximine is strictly dependent on light. Under otherwise optimal conditions, the yield of ammonia production is influenced by irradiance, as well as by the density, depth, and turbulence of the cell suspension. The interaction among these factors seems to determine the actual amount of light available to each single cell or filament in the suspension for the photoproduction process. Under convenient illumination, the limiting factor in the synthesis of ammonia seems to be the cellular nitrogenase activity level, but under limiting light conditions the limiting factor could, however, be the assimilatory power required for nitrogen fixation. Photosynthetic ammonia production from atmospheric nitrogen and water can operate with an efficiency of ca. 10% of its theoretical maximum, representing a remarkable process for the conversion of light energy into chemical energy.

  19. AcsA-AcsB: The core of the cellulose synthase complex from Gluconacetobacter hansenii ATCC23769.

    PubMed

    McManus, John B; Deng, Ying; Nagachar, Nivedita; Kao, Teh-hui; Tien, Ming

    2016-01-01

    The gram-negative bacterium, Gluconacetobacter hansenii, produces cellulose of exceptionally high crystallinity in comparison to the cellulose of higher plants. This bacterial cellulose is synthesized and extruded into the extracellular medium by the cellulose synthase complex (CSC). The catalytic component of this complex is encoded by the gene AcsAB. However, several other genes are known to encode proteins critical to cellulose synthesis and are likely components of the bacterial CSC. We have purified an active heterodimer AcsA-AcsB from G. hansenii ATCC23769 to homogeneity by two different methods. With the purified protein, we have determined how it is post-translationally processed, forming the active heterodimer AcsA-AcsB. Additionally, we have performed steady-state kinetic studies on the AcsA-AcsB complex. Finally through mutagenesis studies, we have explored the roles of the postulated CSC proteins AcsC, AcsD, and CcpAx. PMID:26672449

  20. The Biosynthetic Gene Cluster of Zorbamycin, a Member of the Bleomycin Family of Antitumor Antibiotics, from Streptomyces flavoviridis ATCC 21892

    PubMed Central

    Galm, Ute; Wendt-Pienkowski, Evelyn; Wang, Liyan; George, Nicholas P.; Oh, Tae-Jin; Yi, Fan; Tao, Meifeng; Coughlin, Jane M.; Shen, Ben

    2011-01-01

    The biosynthetic gene cluster for the glycopeptide-derived antitumor antibiotic zorbamycin (ZBM) was cloned by screening a cosmid library of Streptomyces flavoviridis ATCC 21892. Sequence analysis revealed 40 ORFs belonging to the ZBM biosynthetic gene cluster. However, only 23 and 22 ORFs showed striking similarities to the biosynthetic gene clusters for the bleomycins (BLMs) and tallysomycins (TLMs), respectively; the remaining ORFs do not show significant homology to ORFs from the related BLM and TLM clusters. The ZBM gene cluster consists of 16 nonribosomal peptide synthetase (NRPS) genes encoding eight complete NRPS modules, three incomplete didomain NRPS modules, and eight freestanding single NRPS domains or associated enzymes, a polyketide synthase (PKS) gene encoding one PKS module, six sugar biosynthesis genes, as well as genes encoding other biosynthesis and resistance proteins. A genetic system using Escherichia coli-Streptomyces flavoviridis intergeneric conjugation was developed to enable ZBM gene cluster boundary determinations and biosynthetic pathway manipulations. PMID:19081934

  1. High quality permanent draft genome sequence of Phaseolibacter flectens ATCC 12775(T), a plant pathogen of French bean pods.

    PubMed

    Aizenberg-Gershtein, Yana; Izhaki, Ido; Lapidus, Alla; Copeland, Alex; Reddy, Tbk; Huntemann, Marcel; Pillay, Manoj; Markowitz, Victor; Göker, Markus; Woyke, Tanja; Klenk, Hans-Peter; Kyrpides, Nikos C; Halpern, Malka

    2016-01-01

    Phaseolibacter flectens strain ATCC 12775(T) (Halpern et al., Int J Syst Evol Microbiol 63:268-273, 2013) is a Gram-negative, rod shaped, motile, aerobic, chemoorganotroph bacterium. Ph. flectens is as a plant-pathogenic bacterium on pods of French bean and was first identified by Johnson (1956) as Pseudomonas flectens. After its phylogenetic position was reexamined, Pseudomonas flectens was transferred to the family Enterobacteriaceae as Phaseolibacter flectens gen. nov., comb. nov. Here we describe the features of this organism, together with the draft genome sequence and annotation. The DNA GC content is 44.34 mol%. The chromosome length is 2,748,442 bp. It encodes 2,437 proteins and 89 RNA genes. Ph. flectens genome is part of the Genomic Encyclopedia of Type Strains, Phase I: the one thousand microbial genomes study. PMID:26767091

  2. Control of nitrogenase recovery from oxygen inactivation by ammonia in the cyanobacterium Anabaena sp. strain CA (ATCC 33047).

    PubMed Central

    Smith, R L; Van Baalen, C; Tabita, F R

    1990-01-01

    The control of nitrogenase recovery from inactivation by oxygen was studied in Anabaena sp. strain CA (ATCC 33047). Nitrogenase activity (acetylene reduction) in cultures grown in 1% CO2 in air was inhibited by exposure to 1% CO2-99% O2 and allowed to recover in the presence of high oxygen tensions. Cultures exposed to hyperbaric levels of oxygen in the presence of 10 mM NH4NO3 were incapable of regaining nitrogenase activity, whereas control cultures returned to 65 to 80% of their original activity within about 3 h after exposure to high oxygen tension. In contrast to the regulation of heterocyst differentiation and nitrogenase synthesis, recovery from oxygen inactivation in this organism was shown to be under the control of NH4+ rather than NO3-. PMID:2110151

  3. Genome sequence of Victivallis vadensis ATCC BAA-548, an anaerobic bacterium from the phylum Lentisphaerae, isolated from the human gastro-intestinal tract

    SciTech Connect

    Van Passel, Mark W.J.; Kant, Ravi; Palva, Airi; Lucas, Susan; Copeland, A; Lapidus, Alla L.; Glavina Del Rio, Tijana; Dalin, Eileen; Tice, Hope; Bruce, David; Goodwin, Lynne A.; Pitluck, Sam; Davenport, Karen W.; Sims, David; Detter, J. Chris; Han, Cliff; Larimer, Frank W; Land, Miriam L; Hauser, Loren John; Kyrpides, Nikos C; Ovchinnikova, Galina; Richardson, Paul; De Vos, Willem M.; Smidt, Hauke; Zoetendal, Erwin G.

    2011-01-01

    Victivallis vadensis ATCC BAA-548 represents the first cultured representative from the novel phylum Lentisphaerae, a deep-branching bacterial lineage. Few cultured bacteria from this phylum are known, and V. vadensis therefore represents an important organism for evolutionary studies. V. vadensis is a strictly anaerobic sugar-fermenting isolate from the human gastro-intestinal tract.

  4. The effects of different light-dark cycles on the metabolism of the diazotrophic, unicellular cyanobacteria Cyanothece sp. ATCC 51142, and Cyanothecesp. PCC 7822.

    PubMed

    Arshad, Sarah; Mishra, Sujata; Sherman, Louis A

    2014-10-01

    The diazotrophic unicellular cyanobacterium Cyanothece sp. ATCC 51142 demonstrates circadian patterns in nitrogenase activity, H2 production and glycogen storage when grown under nitrogen-fixing, 12:12 light:dark (L:D) conditions. In this study, we grew Cyanothece sp. ATCC 51142, and another strain in this genus, Cyanothece sp. PCC 7822, under long-day (16:8 L:D) and short-day (8:16 L:D) nitrogen-fixing conditions to determine if they continued to display circadian rhythms. Both strains demonstrated similar circadian patterns for all three metabolic parameters when grown under long-day conditions. However, the strains responded differently to short-day growth conditions. Cyanothece sp. ATCC 51142 retained reasonable circadian patterns under 8:16 L:D conditions, whereas Cyanothece sp. PCC 7822 had quite damped patterns without a clear circadian pattern. In particular, glycogen storage changed very little throughout the day and we ascribe this to the difference in the type of glycogen granules in Cyanothece sp. PCC 7822 which has small β-granules, compared to the large, starch-like granules in Cyanothece sp. ATCC 51142. The results suggested that both mechanistic and regulatory processes play a role in establishing the basis for these metabolic oscillations. PMID:26988646

  5. Complete Genome Sequence of a Gluconacetobacter hansenii ATCC 23769 Isolate, AY201, Producer of Bacterial Cellulose and Important Model Organism for the Study of Cellulose Biosynthesis

    PubMed Central

    Mehta, Kalpa

    2016-01-01

    The cellulose producer and model organism used for the study of cellulose biosynthesis, Gluconacetobacter hansenii AY201, is a variant of G. hansenii ATCC 23769. We report here the complete nucleotide sequence of G. hansenii AY201, information which may be utilized to further the research into understanding the genes necessary for cellulose biosynthesis. PMID:27516506

  6. Draft genome sequence of the extremely acidophilic biomining bacterium Acidithiobacillus thiooxidans ATCC 19377 provides insights into the evolution of the Acidithiobacillus genus.

    PubMed

    Valdes, Jorge; Ossandon, Francisco; Quatrini, Raquel; Dopson, Mark; Holmes, David S

    2011-12-01

    Acidithiobacillus thiooxidans is a mesophilic, extremely acidophilic, chemolithoautotrophic gammaproteobacterium that derives energy from the oxidation of sulfur and inorganic sulfur compounds. Here we present the draft genome sequence of A. thiooxidans ATCC 19377, which has allowed the identification of genes for survival and colonization of extremely acidic environments. PMID:22123759

  7. Draft Genome Sequences of Salmonella enterica subsp. enterica Serovar Berta ATCC 8392 and a Nalidixic Acid-Resistant Isolate of This Strain

    PubMed Central

    Cooper, Ashley; Koziol, Adam G.; Carrillo, Catherine D.

    2016-01-01

    Salmonella enterica subspecies enterica serovar Berta has been isolated in multiple animal species and has been implicated in human disease. Here, we report a 4.7-Mbp draft genome sequence of S. enterica serovar Berta (ATCC strain 8392) and a nalidixic acid-resistant isolate derived from this strain. PMID:27103707

  8. Complete Genome Sequence of a Gluconacetobacter hansenii ATCC 23769 Isolate, AY201, Producer of Bacterial Cellulose and Important Model Organism for the Study of Cellulose Biosynthesis.

    PubMed

    Pfeffer, Sarah; Mehta, Kalpa; Brown, R Malcolm

    2016-01-01

    The cellulose producer and model organism used for the study of cellulose biosynthesis, Gluconacetobacter hansenii AY201, is a variant of G. hansenii ATCC 23769. We report here the complete nucleotide sequence of G. hansenii AY201, information which may be utilized to further the research into understanding the genes necessary for cellulose biosynthesis. PMID:27516506

  9. No evidence of harms of probiotic Lactobacillus rhamnosus GG ATCC 53103 in healthy elderly-a Phase I Open Label Study to assess safety, tolerability and cytokine responses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Although Lactobacillus rhamnosus GG ATCC 53103 (LGG) has been consumed since the mid 1990s by between 2 and 5 million people daily, the scientific literature lacks rigorous clinical trials that describe the potential harms of LGG, particularly in the elderly. The primary objective of this open label...

  10. Complete Genome Sequence of Streptomyces venezuelae ATCC 15439, Producer of the Methymycin/Pikromycin Family of Macrolide Antibiotics, Using PacBio Technology

    PubMed Central

    He, Jingxuan; Sundararajan, Anitha; Devitt, Nicholas P.; Schilkey, Faye D.; Ramaraj, Thiruvarangan

    2016-01-01

    Here, we report the complete genome sequence of Streptomyces venezuelae ATCC 15439, a producer of the methymycin/pikromycin family of macrolide antibiotics and a model host for natural product studies, obtained exclusively using PacBio sequencing technology. The 9.03-Mbp genome harbors 8,775 genes and 11 polyketide and nonribosomal peptide natural product gene clusters. PMID:27151802

  11. Difference in cellular damage and cell death in thermal death time disks and high hydrostatic pressure treated Salmonella Enteritidis (ATCC13076) in liquid whole egg

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Differences in membrane damage including leakage of intracellular UV-materials and loss of viability of Salmonella Enteritidis (ATCC13076) in liquid whole egg (LWE) following thermal-death-time (TDT) disk and high hydrostatic pressure treatments were examined. Salmonella enteritidis was inoculated ...

  12. Genome sequences of three tunicamycin-producing Streptomyces strains; S. chartreusis NRRL 12338, S. chartreusis NRRL 3882, and S. lysosuperificus ATCC 31396

    Technology Transfer Automated Retrieval System (TEKTRAN)

    S. chartreusis strains NRRL 12338 and NRRL 3882, S. clavuligerus NRRL 3585, and S. lysosuperificus ATCC 31396, are known producers of tunicamycins, and also of charteusins, clavulinate, cephalosporins, holomycins, and calcimycin. Here we announce the sequencing of the S. lysosuperificus and the two...

  13. Evaluation of molecular techniques for identification and enumeration of Raoultella terrigena ATCC 33257 in water purifier efficacy testing.

    PubMed

    Saha, Ratul; Bechanko, Robin; Bestervelt, Lorelle L; Donofrio, Robert S

    2011-09-01

    Raoultella terrigena ATCC 33257, a representative of the coliform group, is commonly used as a challenge organism in water purifier efficacy testing. In addition to being time consuming, traditional culturing techniques and metabolic identification systems (including automated systems) also fail to accurately differentiate this organism from its closely related neighbors belonging to the Enterobacteriaceae group. Molecular-based techniques, such as real-time quantitative polymerase chain reaction (qPCR) and enterobacterial repetitive intergenic consensus (ERIC)-PCR fingerprinting, are preferred methods of detection because of their accuracy, reproducibility, specificity, and sensitivity, along with shorter turnaround time. ERIC-PCR performed with the 1R primer set demonstrated stable unique banding patterns (~800, ~300 bp) for R. terrigena ATCC 33257 different from patterns observed for R. planticola and R. ornithinolytica. The primer pair developed from gyraseA (gyrA) sequence of R. terrigena for the SYBR Green qPCR assay using the AlleleID(®) 7.0 primer probe design software was highly specific and sensitive for the target organism. The sensitivity of the assay was 10(1) colony forming units (CFU)/ml for whole cells and 4.7 fg with genomic DNA. The primer pair was successful in determining the concentration (5.5 ± 0.3 × 10(6) CFU/ml) of R. terrigena from water samples spiked with equal concentration of Escherichia coli and R. terrigena. Based on these results from the ERIC-PCR and the SYBR Green qPCR assay, these molecular techniques can be efficiently used for rapid identification and quantification of R. terrigena during water purifier testing. PMID:21132347

  14. Toxic Accumulation of LPS Pathway Intermediates Underlies the Requirement of LpxH for Growth of Acinetobacter baumannii ATCC 19606.

    PubMed

    Richie, Daryl L; Takeoka, Kenneth T; Bojkovic, Jade; Metzger, Louis E; Rath, Christopher M; Sawyer, William S; Wei, Jun-Rong; Dean, Charles R

    2016-01-01

    The lipid A moiety of lipopolysaccharide (LPS) is the main constituent of the outer leaflet of the Gram-negative bacterial outer membrane (OM) and is essential in many Gram-negative pathogens. An exception is Acinetobacter baumannii ATCC 19606, where mutants lacking enzymes occurring early in lipid A biosynthesis (LpxA, LpxC or LpxD), and correspondingly lacking LPS, can grow. In contrast, we show here that LpxH, an enzyme that occurs downstream of LpxD in the lipid A biosynthetic pathway, is essential for growth in this strain. Multiple attempts to disrupt lpxH on the genome were unsuccessful, and when LpxH expression was controlled by an isopropyl β-d-1-thiogalactopyranoside (IPTG) inducible promoter, cell growth under typical laboratory conditions required IPTG induction. Mass spectrometry analysis of cells shifted from LpxH-induced to uninduced (and whose growth was correspondingly slowing as LpxH was depleted) showed a large cellular accumulation of UDP-2,3-diacyl-GlcN (substrate of LpxH), a C14:0(3-OH) acyl variant of the LpxD substrate (UDP-3-O-[(R)-3-OH-C14]-GlcN), and disaccharide 1-monophosphate (DSMP). Furthermore, the viable cell counts of the LpxH depleted cultures dropped modestly, and electron microscopy revealed clear defects at the cell (inner) membrane, suggesting lipid A intermediate accumulation was toxic. Consistent with this, blocking the synthesis of these intermediates by inhibition of the upstream LpxC enzyme using CHIR-090 abrogated the requirement for IPTG induction of LpxH. Taken together, these data indicate that LpxH is essential for growth in A. baumannii ATCC19606, because, unlike earlier pathway steps like LpxA or LpxC, blockage of LpxH causes accumulation of detergent-like pathway intermediates that prevents cell growth. PMID:27526195

  15. Characterization of the major dehydrogenase related to d-lactic acid synthesis in Leuconostoc mesenteroides subsp. mesenteroides ATCC 8293.

    PubMed

    Li, Ling; Eom, Hyun-Ju; Park, Jung-Mi; Seo, Eunyoung; Ahn, Ji Eun; Kim, Tae-Jip; Kim, Jeong Hwan; Han, Nam Soo

    2012-10-10

    Leuconostoc mesenteroides subsp. mesenteroides ATCC 8293 is a lactic acid bacterium that converts pyruvate mainly to d-(-)-lactic acid by using d-(-)-lactate dehydrogenase (ldhD). The aim of this study was to identify the gene responsible for d-lactic acid formation in this organism and to characterize the enzyme to facilitate the production of optically pure d-lactic acid. A genomic analysis of L. mesenteroides ATCC 8293 revealed that 7 genes encode lactate-related dehydrogenase. According to transcriptomic, proteomic, and phylogenetic analyses, LEUM_1756 was the major gene responsible for the production of d-lactic acid. The LEUM_1756 gene, of 996bp and encoding 332 amino acids (36.5kDa), was cloned and overexpressed in Escherichia coli BL21(DE3) Star from an inducible pET-21a(+) vector. The enzyme was purified by Ni-NTA column chromatography and showed a specific activity of 4450U/mg, significantly higher than those of other previously reported ldhDs. The gel permeation chromatography analysis showed that the purified enzyme exists as tetramers in solution and this was the first report among lactic acid bacteria. The pH and temperature optima were pH 8.0 and 30°C, respectively, for the pyruvate reduction reaction, and pH 11.0 and 20°C, respectively, for the lactate oxidation reaction. The K(m) kinetic parameters for pyruvate and lactate were 0.58mM and 260mM, respectively. In addition, the k(cat) values for pyruvate and lactate were 2900s(-1) and 2280s(-1), respectively. The enzyme was not inhibited by Ca(2+), Co(2+), Cu(2+), Mg(2+), Mn(2+), Na(+), or urea, but was inhibited by 1mM Zn(2+) and 1mM SDS. PMID:22975125

  16. Molecular cloning and heterologous expression of the isopullulanase gene from Aspergillus niger A.T.C.C. 9642.

    PubMed Central

    Aoki, H; Yopi; Sakano, Y

    1997-01-01

    Isopullulanase (IPU) from Aspergillus niger A.T.C.C. (American Type Culture Collection) 9642 hydrolyses pullulan to isopanose. IPU is important for the production of isopanose and is used in the structural analysis of oligosaccharides with alpha-1,4 and alpha-1,6 glucosidic linkages. We have isolated the ipuA gene encoding IPU from the filamentous fungi A. niger A.T.C.C. 9642. The ipuA gene encodes an open reading frame of 1695 bp (564 amino acids). IPU contained a signal sequence of 19 amino acids, and the molecular mass of the mature form was calculated to be 59 kDa. IPU has no amino-acid-sequence similarity with the other pullulan-hydrolysing enzymes, which are pullulanase, neopullulanase and glucoamylase. However, IPU showed a high amino-acid-sequence similarity with dextranases from Penicillium minioluteum (61%) and Arthrobacter sp. (56%). When the ipuA gene was expressed in Aspergillus oryzae, the expressed protein (recombinant IPU) had IPU activity and was immunologically reactive with antibodies raised against native IPU. The substrate specificity, thermostability and pH profile of recombinant IPU were identical with those of the native enzyme, but recombinant IPU (90 kDa) was larger than the native enzyme (69-71 kDa). After deglycosylation with peptide-N-glycosidase F, the deglycosylated recombinant IPU had the same molecular mass as deglycosylated native enzyme (59 kDa). This result suggests that the carbohydrate chain of recombinant IPU differed from that of the native enzyme. PMID:9169610

  17. Toxic Accumulation of LPS Pathway Intermediates Underlies the Requirement of LpxH for Growth of Acinetobacter baumannii ATCC 19606

    PubMed Central

    Richie, Daryl L.; Takeoka, Kenneth T.; Bojkovic, Jade; Metzger, Louis E.; Rath, Christopher M.; Sawyer, William S.; Wei, Jun-Rong; Dean, Charles R.

    2016-01-01

    The lipid A moiety of lipopolysaccharide (LPS) is the main constituent of the outer leaflet of the Gram-negative bacterial outer membrane (OM) and is essential in many Gram-negative pathogens. An exception is Acinetobacter baumannii ATCC 19606, where mutants lacking enzymes occurring early in lipid A biosynthesis (LpxA, LpxC or LpxD), and correspondingly lacking LPS, can grow. In contrast, we show here that LpxH, an enzyme that occurs downstream of LpxD in the lipid A biosynthetic pathway, is essential for growth in this strain. Multiple attempts to disrupt lpxH on the genome were unsuccessful, and when LpxH expression was controlled by an isopropyl β-d-1-thiogalactopyranoside (IPTG) inducible promoter, cell growth under typical laboratory conditions required IPTG induction. Mass spectrometry analysis of cells shifted from LpxH-induced to uninduced (and whose growth was correspondingly slowing as LpxH was depleted) showed a large cellular accumulation of UDP-2,3-diacyl-GlcN (substrate of LpxH), a C14:0(3-OH) acyl variant of the LpxD substrate (UDP-3-O-[(R)-3-OH-C14]-GlcN), and disaccharide 1-monophosphate (DSMP). Furthermore, the viable cell counts of the LpxH depleted cultures dropped modestly, and electron microscopy revealed clear defects at the cell (inner) membrane, suggesting lipid A intermediate accumulation was toxic. Consistent with this, blocking the synthesis of these intermediates by inhibition of the upstream LpxC enzyme using CHIR-090 abrogated the requirement for IPTG induction of LpxH. Taken together, these data indicate that LpxH is essential for growth in A. baumannii ATCC19606, because, unlike earlier pathway steps like LpxA or LpxC, blockage of LpxH causes accumulation of detergent-like pathway intermediates that prevents cell growth. PMID:27526195

  18. Simultaneous production of isopropanol, butanol, ethanol and 2,3-butanediol by Clostridium acetobutylicum ATCC 824 engineered strains.

    PubMed

    Collas, Florent; Kuit, Wouter; Clément, Benjamin; Marchal, Rémy; López-Contreras, Ana M; Monot, Frederic

    2012-01-01

    Isopropanol represents a widely-used commercial alcohol which is currently produced from petroleum. In nature, isopropanol is excreted by some strains of Clostridium beijerinckii, simultaneously with butanol and ethanol during the isopropanol butanol ethanol (IBE) fermentation. In order to increase isopropanol production, the gene encoding the secondary-alcohol dehydrogenase enzyme from C. beijerinckii NRRL B593 (adh) which catalyzes the reduction of acetone to isopropanol, was cloned into the acetone, butanol and ethanol (ABE)-producing strain C. acetobutylicum ATCC 824. The transformants showed high capacity for conversion of acetone into isopropanol (> 95%). To increase isopropanol production levels in ATCC 824, polycistronic transcription units containing, in addition to the adh gene, homologous genes of the acetoacetate decarboxylase (adc), and/or the acetoacetyl-CoA:acetate/butyrate:CoA transferase subunits A and B (ctfA and ctfB) were constructed and introduced into the wild-type strain. Combined overexpression of the ctfA and ctfB genes resulted in enhanced solvent production. In non-pH-controlled batch cultures, the total solvents excreted by the transformant overexpressing the adh, ctfA, ctfB and adc genes were 24.4 g/L IBE (including 8.8 g/L isopropanol), while the control strain harbouring an empty plasmid produced only 20.2 g/L ABE (including 7.6 g/L acetone). The overexpression of the adc gene had limited effect on IBE production. Interestingly, all transformants with the adh gene converted acetoin (a minor fermentation product) into 2,3-butanediol, highlighting the wide metabolic versatility of solvent-producing Clostridia. PMID:22909015

  19. Simultaneous production of isopropanol, butanol, ethanol and 2,3-butanediol by Clostridium acetobutylicum ATCC 824 engineered strains

    PubMed Central

    2012-01-01

    Isopropanol represents a widely-used commercial alcohol which is currently produced from petroleum. In nature, isopropanol is excreted by some strains of Clostridium beijerinckii, simultaneously with butanol and ethanol during the isopropanol butanol ethanol (IBE) fermentation. In order to increase isopropanol production, the gene encoding the secondary-alcohol dehydrogenase enzyme from C. beijerinckii NRRL B593 (adh) which catalyzes the reduction of acetone to isopropanol, was cloned into the acetone, butanol and ethanol (ABE)-producing strain C. acetobutylicum ATCC 824. The transformants showed high capacity for conversion of acetone into isopropanol (> 95%). To increase isopropanol production levels in ATCC 824, polycistronic transcription units containing, in addition to the adh gene, homologous genes of the acetoacetate decarboxylase (adc), and/or the acetoacetyl-CoA:acetate/butyrate:CoA transferase subunits A and B (ctfA and ctfB) were constructed and introduced into the wild-type strain. Combined overexpression of the ctfA and ctfB genes resulted in enhanced solvent production. In non-pH-controlled batch cultures, the total solvents excreted by the transformant overexpressing the adh, ctfA, ctfB and adc genes were 24.4 g/L IBE (including 8.8 g/L isopropanol), while the control strain harbouring an empty plasmid produced only 20.2 g/L ABE (including 7.6 g/L acetone). The overexpression of the adc gene had limited effect on IBE production. Interestingly, all transformants with the adh gene converted acetoin (a minor fermentation product) into 2,3-butanediol, highlighting the wide metabolic versatility of solvent-producing Clostridia. PMID:22909015

  20. Multiple Mechanisms Contribute to Lateral Transfer of an Organophosphate Degradation (opd) Island in Sphingobium fuliginis ATCC 27551

    PubMed Central

    Pandeeti, Emmanuel Vijay Paul; Longkumer, Toshisangba; Chakka, Deviprasanna; Muthyala, Venkateswar Reddy; Parthasarathy, Sunil; Madugundu, Anil Kumar; Ghanta, Sujana; Medipally, Srikanth Reddy; Pantula, Surat Chameli; Yekkala, Harshita; Siddavattam, Dayananda

    2012-01-01

    The complete sequence of pPDL2 (37,317 bp), an indigenous plasmid of Sphingobium fuliginis ATCC 27551 that encodes genes for organophosphate degradation (opd), revealed the existence of a site-specific integrase (int) gene with an attachment site attP, typically seen in integrative mobilizable elements (IME). In agreement with this sequence information, site-specific recombination was observed between pPDL2 and an artificial plasmid having a temperature-sensitive replicon and a cloned attB site at the 3′ end of the seryl tRNA gene of Sphingobium japonicum. The opd gene cluster on pPDL2 was found to be part of an active catabolic transposon with mobile elements y4qE and Tn3 at its flanking ends. Besides the previously reported opd cluster, this transposon contains genes coding for protocatechuate dioxygenase and for two transport proteins from the major facilitator family that are predicted to be involved in transport and metabolism of aromatic compounds. A pPDL2 derivative, pPDL2-K, was horizontally transferred into Escherichia coli and Acinetobacter strains, suggesting that the oriT identified in pPDL2 is functional. A well-defined replicative origin (oriV), repA was identified along with a plasmid addiction module relB/relE that would support stable maintenance of pPDL2 in Sphingobium fuliginis ATCC 27551. However, if pPDL2 is laterally transferred into hosts that do not support its replication, the opd cluster appears to integrate into the host chromosome, either through transposition or through site-specific integration. The data presented in this study help to explain the existence of identical opd genes among soil bacteria. PMID:23275877