Disentangling the impact of environmental and phylogenetic constraints on prokaryotic within-species diversity

Microbial organisms inhabit virtually all environments and encompass a vast biological diversity. The pangenome concept aims to facilitate an understanding of diversity within defined phylogenetic groups. Hence, pangenomes are increasingly used to characterize the strain diversity of prokaryotic species. To understand the interdependence of pangenome features (such as the number of core and accessory genes) and to study the impact of environmental and phylogenetic constraints on the evolution of conspecific strains, we computed pangenomes for 155 phylogenetically diverse species (from ten phyla) using 7,000 high-quality genomes to each of which the respective habitats were assigned. Species habitat ubiquity was associated with several pangenome features. In particular, core-genome size was more important for ubiquity than accessory genome size. In general, environmental preferences had a stronger impact on pangenome evolution than phylogenetic inertia. Environmental preferences explained up to 49% of the variance for pangenome features, compared with 18% by phylogenetic inertia. This observation was robust when the dataset was extended to 10,100 species (59 phyla). The importance of environmental preferences was further accentuated by convergent evolution of pangenome features in a given habitat type across different phylogenetic clades. For example, the soil environment promotes expansion of pangenome size, while host-associated habitats lead to its reduction. Taken together, we explored the global principles of pangenome evolution, quantified the influence of habitat, and phylogenetic inertia on the evolution of pangenomes and identified criteria governing species ubiquity and habitat specificity

proGenomes3: approaching one million accurately and consistently annotated high-quality prokaryotic genomes

The interpretation of genomic, transcriptomic and other microbial 'omics data is highly dependent on the availability of well-annotated genomes. As the number of publicly available microbial genomes continues to increase exponentially, the need for quality control and consistent annotation is becoming critical. We present proGenomes3, a database of 907 388 high-quality genomes containing 4 billion genes that passed stringent criteria and have been consistently annotated using multiple functional and taxonomic databases including mobile genetic elements and biosynthetic gene clusters. proGenomes3 encompasses 41 171 species-level clusters, defined based on universal single copy marker genes, for which pan-genomes and contextual habitat annotations are provided. The database is available at http://progenomes.embl.de/

Optimization Of Extracellular Lipase Production By Geotrichum Sp. Using Factorial Design

Response surface methodology was employed to study the effects of carbon source (soy oil, olive oil and glucose) and nitrogen source concentrations (corn steep liquor and NH4NO3) on the lipase production by Geotrichum sp. The experiment included a 24 central composite rotatable design (CCRD) and four others 23 CCRD. According to the responses from the experimental designs, the effects of each variable were calculated and the interactions between them were determined. The response surface methodology was applied for the optimization of the nutrient concentrations in the culture medium for the enzyme production, at 30 °C. The optimum medium composition for lipase production by Geotrichum sp. was ammonium nitrate 2.1-2.5%, corn steep liquor 13-15% and soy oil 0.6% as carbon source, which lead to a lipase activity of about 20 U/ml. Using olive oil as carbon source, the optimum composition was ammonium nitrate 0.8-1%, corn steep liquor 13-15% and olive oil 0.6%, leading to an activity of 17 U/ml. © 2003 Elsevier Ltd. All rights reserved.9117784Bertolini, M.C., Laramee, L., Thomas, D.Y., Cygler, M., Schrag, J.D., Vernet, T., Polymorphism in the lipase genes of Geotrichum candidum strains (1994) Europ. J. Biochem., 219, pp. 119-125Carvalho, C.M.L., Serralheiro, M.L.M., Cabral, J.M.S., Aires-Barros, M.R., Application of factorial design to the study of transesterification reactions using cutinase in AOT-reversed micelles (1997) Enzy. Microb. Technol., 21, pp. 117-123Cruz, P.M., Christen, P., Farres, A., Medium optimization by fractional factorial design for lipase production by Rizopus delemar (1993) J. Ferm. Bioeng., 76, pp. 94-97Elibol, M., Ozer, D., Influence of oxygen transfer on lipase production by Rizopus arrhizus (2000) Proc. Biochem., 36, pp. 325-329Freire, D.M., Teles, E.M.F., Bon, E.P.S., Lippel Sant'anna, G., Lipase production by Penicillium restrictum in a bench-scale fermenter (1997) Appl. Biochem. Biothec., 63-65, pp. 409-421Haaland, P.D., (1989) Experimental design in biotechnology, , New York: Marcel Dekker incHatzinikolaou, D.G., Macris, J.B., Christakopoulos, P., Kekos, D., Kolisis, F.N., Fountoukidis, G., Production and partial characterization of lipase from Aspergillus niger (1996) Biothecn. Lett., 18, pp. 547-552Jacobsen, T., Poulsen, O.M., Comparison of lipases from different strains of the fungus Geotrichum candidum (1995) Biochim. Biphys. Acta Lipids and Lipid Metabolism, 1257, pp. 96-102Kalil, S.J., Maugeri, F., Rodrigues, M.I., Response surface analysis and simulation as a tool for bioprocess design and optimization (2000) Proc. Biochem., 35, pp. 539-550Kamimura, E.S., Medieta, O., Rodrigues, M.I., Maugeri, F., Studies on lipase-affinity adsorption using response-surface analysis (2001) Biotechnol. Appl. Biochem., 33, pp. 153-159Kamini, N.R., Fujii, T., Kurosu, T., Iefuji, H., Production, purification and characterization of an extracellular lipase from yeast, Cryptococcus sp. S-2 (2000) Proc. Biochem., 36, pp. 317-324Khuri, A.I., Cornell, J.A., (1987) Response surface design and analyses, , New York: Marcel Dekker IncKui, T., Xu, X., He, C., Li, L., Lipase-catalysed modification of lard to produce human milk fat substitutes (2003) Food Cap Chem., 80, pp. 473-481Macêdo, G.A., Park, Y.K., Pastore, G.M., Partial purification and characterization of an extracellular lipase from a newly isolated strain of Geotrichum sp (1997) J. Brazil. Soc. Microbiol., 28, pp. 90-95Nagao, T., Shimada, Y., Sugihara, A., Tominaga, Y., Expression of lipase cDNA from Fusarium heterosporum by Saccharomyces cerevisae: High-level production and purification (1996) J. Ferment. Bioeng., 81, pp. 488-492Nagayama, K., Yamasaki, N., Imai, M., Fatty acid esterification catalyzed by Candida rugosa lipase in lecithin microemulsion-based organogels (2002) Biochem. Eng. J., 12, pp. 231-236Nini, L., Sarda, L., Coumeau, L.C., Boitard, E., Dubes, J.P., Chahinian, H., Lipase catalysed hydrolysis of short-chain substrates in solution and in emulsion: A kinetic study (2001) Biochim. Biphys. Acta Molecular and Cell Biology, 1534, pp. 34-44Phillips, A., Pretorius, G.H.J., Hester, G., Rensburg, V., Molecular characterization of a Galactomyces geotrichum lipase, another member of the cholinesterase/lipase family (1995) Biochim. Biphys. Acta Protein Struct. Molecular, 1252, pp. 305-311Piao, J., Kobayashi, T., Adachi, S., Nakanishi, K., Matsuno, R., Synthesis of mono- and dioleoyl erythritols through immobilized-lipase- catalyzed condensation of erythritol and oleic acid in acetone (2003) Biochem. Eng. J., 14, pp. 79-84Pokorny, D., Cinerman, A., Steiner, W., Aspergillus niger lipases: Induction, isolation and characterization of two lipases from MZKI A116 strain (1997) J. Molec. Cataly. B: Enzymatic, 2, pp. 215-222Rao, P.V., Jayaraman, K., Lakshmanan, C.M., Production of lipase by Candida rugosa in solid state fermentation 2: Medium optimization and effect of aeration (1993) Proces. Biochem., 28, pp. 391-395Raku, T., Kitagawa, M., Shimakawa, H., Tokiwa, Y., Enzymatic synthesis of threalose esters having lipophilicity (2003) J. Biotechnol., 100, pp. 203-208Saxena, R.K., Ghosh, P.K., Gupta, R., Davdison, W.S., Bradoo, S., Gulati, R., Microbial lipases: Potential biocatalysts for the future industry (1999) Curr. Sci., 77, pp. 101-115Shintre, M.S., Ghadge, R.S., Sawant, S.B., Lipolase catalyzed synthesis of benzyl esters of fatty acids (2002) Biochem. Eng. J., 12, pp. 131-141Vanot, G., Deyris, V., Guilhem, M.C., Phan Tan Luu, R., Comeau, L.C., Optimal design for the maximization of Penicillium cyclopium lipase production (2001) Appl. Microbiol. Biotechnol., 57, pp. 342-345Warwel, S., Borgdorf, R., Substrate selectivity of lipases in the esterification of cis-trans-isomers and positional isomers conjugated linoleic acid (CLA) (2000) Biotechnol. Lett., 22, pp. 1151-115

Optimization Of Medium Formulation And Seed Conditions For Expression Of Mature Psaa (pneumococcal Surface Adhesin A) In Escherichia Coli Using A Sequential Experimental Design Strategy And Response Surface Methodology

PsaA, a candidate antigen for a vaccine against pneumonia, is well-conserved in all Streptococcus pneumoniae serotypes. A sequence of two-level experimental designs was used to evaluate medium composition and seed conditions to optimize the expression of soluble mature PsaA in E. coli. A face-centered central composite design was first used to evaluate the effects of yeast extract (5 and 23.6 g/L), tryptone (0 and 10 g/L), and glucose (1 and 10 g/L), with replicate experiments at the central point (14.3 g/L yeast extract, 5 g/L tryptone, 5.5 g/L glucose). Next, a central composite design was used to analyze the influence of NaCl concentration (0, 5, and 10 g/L) compared with potassium salts (9.4 g/L K2HPO4/2.2 g/L KH2PO4), and seed growth (7 and 16 h). Tryptone had no significant effect and was removed from the medium. Yeast extract and glucose were optimized at their intermediate concentrations, resulting in an animal-derived material-free culture medium containing 15 g/L yeast extract, 8 g/L glucose, 50 μg/mL kanamycin, and 0.4% glycerol, yielding 1 g/L rPsaA after 16 h induction at 25°C in shake flasks at 200 rpm. All the seed age and salt conditions produced similar yields, indicating that no variation had a statistically significant effect on expression. Instead of growing the seed culture for 16 h (until saturation), the process can be conducted with 7 h seed growth until the exponential phase. These results enhanced the process productivity and reduced costs, with 5 g/L NaCl being used rather than potassium salts. © Society for Industrial Microbiology and Biotechnology 2012.396897908Bogaert, D., Hermans, P.W.M., Adrian, P.V., Rumke, H.C., De Groot, R., Pneumococcal vaccines: An update on current strategies (2004) Vaccine, 22 (17-18), pp. 2209-2220. , DOI 10.1016/j.vaccine.2003.11.038, PII S0264410X03008387Briles, D.E., Protection of the elderly from pneumococcal pneumonia with a protein-based vaccine? (2004) Mechanisms of Ageing and Development, 125 (2), pp. 129-131. , DOI 10.1016/j.mad.2003.11.008Briles, D.E., Ades, E., Paton, J.C., Sampson, J.S., Carlone, G.M., Huebner, R.C., Virolainen, A., Hollingshead, S.K., Intranasal immunization of mice with a mixture of the pneumococcal proteins PsaA and PspA is highly protective against nasopharyngeal carriage of Streptococcus pneumoniae (2000) Infection and Immunity, 68 (2), pp. 796-800. , DOI 10.1128/IAI.68.2.796-800.2000Chen, Y., Xing, X.-H., Ye, F., Kuang, Y., Luo, M., Production of MBP-HepA fusion protein in recombinant Escherichia coli by optimization of culture medium (2007) Biochemical Engineering Journal, 34 (2), pp. 114-121. , DOI 10.1016/j.bej.2006.11.020, PII S1369703X06003585Choi, J.H., Keum, K.C., Lee, S.Y., Production of recombinant proteins by high cell density culture of Escherichia coli (2006) Chemical Engineering Science, 61 (3), pp. 876-885. , DOI 10.1016/j.ces.2005.03.031, PII S0009250905002691, Biomolecular EngineeringChoi, W.C., Oh, B.C., Kim, H.K., Lee, E.S., Oh, T.K., Medium optimization for phytase production by recombinant escherichia coli using statistical experimental design (2002) J Microbiol Biotechnol, 12, pp. 490-496De, B.K., Sampson, J.S., Ades, E.W., Huebner, R.C., Jue, D.L., Johnson, S.E., Espina, M., Carlone, G.M., Purification and characterization of Streptococcus pneumoniae palmitoylated pneumococcal surface adhesin A expressed in Escherichia coli (2000) Vaccine, 18 (17), pp. 1811-1821. , DOI 10.1016/S0264-410X(99)00481-8, PII S0264410X99004818Douce, G., Ross, K., Cowan, G., Ma, J., Mitchell, T.J., Novel mucosal vaccines generated by genetic conjugation ofheterologous proteins to pneumolysin (ply) from streptococcus pneumoniae (2010) Vaccine, 28, pp. 3231-3237Fontani, S., Niccolai, A., Kapat, A., Olivieri, R., Studies on the maximization of recombinant Helicobacter pylori neutrophil-activating protein production in Escherichia coli: Application of Taguchi robust design and response surface methodology for process optimization (2003) World Journal of Microbiology and Biotechnology, 19 (7), pp. 711-717. , DOI 10.1023/A:1025104119260Gor, D.O., Ding, X., Li, Q., Schreiber, J.R., Dubinsky, M., Greenspan, N.S., Enhanced immunogenicity of pneumococcal surface adhesin a by genetic fusion to cytokines and evaluation of protective immunity in mice (2002) Infect Immun, 70, pp. 5589-5595Hao, D.C., Zhu, P.H., Yang, S.L., Yang, L., Optimization of recombinant cytochrome P450 2C9 protein production in Escherichia coli DH5α by statistically-based experimental design (2006) World Journal of Microbiology and Biotechnology, 22 (11), pp. 1169-1176. , DOI 10.1007/s11274-006-9158-9Ihssen, J., Kowarik, M., Dilettoso, S., Tanner, C., Wacker, M., Thöny-Meyer, L., Production of glycoprotein vaccines in escherichia coli (2010) Microb Cell Fact, 9, pp. 61-73Islam, R.S., Tisi, D., Levy, M.S., Lye, G.J., Framework for the rapid optimization of soluble protein expression in Escherichia coli combining microscale experiments and statistical experimental design (2007) Biotechnology Progress, 23 (4), pp. 785-793. , DOI 10.1021/bp070059aLarentis, A.L., Argondizzo, A.P.C., Esteves, G.S., Jessouron, E., Galler, R., Medeiros, M.A., Cloning and optimization of induction conditions for mature psaa (pneumococcal surface adhesin a) expression in escherichia coli and recombinant protein stability during long-term storage (2011) Protein Expr Purif, 78, pp. 38-47Laval, C.B., De Andrade, A.L.S.S., Pimenta, F.C., De Andrade, J.G., De Oliveira, R.M., Silva, S.A., De Lima, E.C., Brandileone, M.C.C., Serotypes of carriage and invasive isolates of Streptococcus pneumoniae in Brazilian children in the era of pneumococcal vaccines (2006) Clinical Microbiology and Infection, 12 (1), pp. 50-55. , DOI 10.1111/j.1469-0691.2005.01304.xLee, K.M., Rhee, C.H., Kang, C.K., Kim, J.H., Sequential and simultaneous statistical optimization by dynamic design of experiment for peptide overexpression in recombinant escherichia coli (2006) Appl Biochem Biotechnol, 135, pp. 59-80Lee, S.Y., High cell-density culture of Escherichia coli (1996) Trends in Biotechnology, 14 (3), pp. 98-105. , DOI 10.1016/0167-7799(96)80930-9Manderson, D., Dempster, R., Chisti, Y., A recombinant vaccine against hydatidosis: Production of the antigen in Escherichia coli (2006) Journal of Industrial Microbiology and Biotechnology, 33 (3), pp. 173-182. , DOI 10.1007/s10295-005-0046-3Niccolai, A., Fontani, S., Kapat, A., Olivieri, R., Maximization of recombinant Helicobacter pylori neutrophil activating protein production in Escherichia coli: Improvement of a chemically defined medium using response surface methodology (2003) FEMS Microbiology Letters, 221 (2), pp. 257-262. , DOI 10.1016/S0378-1097(03)00184-8Nikerel, I.E., Oner, E., Kirdar, B., Yildirim, R., Optimization of medium composition for biomass production of recombinant Escherichia coli cells using response surface methodology (2006) Biochemical Engineering Journal, 32 (1), pp. 1-6. , DOI 10.1016/j.bej.2006.08.009, PII S1369703X06002142Pan, H., Xie, Z., Bao, W., Zhang, J., Optimization of culture conditions to enhance cis-epoxysuccinate hydrolase production in escherichia coli by response surface methodology (2008) Biochem Eng J, 42, pp. 133-138Pimenta, F.C., Miyaji, E.N., Areas, A.P.M., Oliveira, M.L.S., De Andrade, A.L.S.S., Ho, P.L., Hollingshead, S.K., Leite, L.C.C., Intranasal immunization with the cholera toxin B subunit-pneumococcal surface antigen A fusion protein induces protection against colonization with Streptococcus pneumoniae and has negligible impact on the nasopharyngeal and oral microbiota of mice (2006) Infection and Immunity, 74 (8), pp. 4939-4944. , DOI 10.1128/IAI.00134-06Pistorino, M., Pfeifer, B.A., Efficient experimental design and micro-scale medium enhancement of 6-deoxyerythronolide b production through escherichia coli (2009) Biotechnol Prog, 25, pp. 1364-1371Rajam, G., Anderton, J.M., Carlone, G.M., Sampson, J.S., Ades, E.W., Pneumococcal surface adhesin a (psaa): A review (2008) Crit Rev Microbiol, 34, pp. 131-142Rodrigues, M.I., Iemma, A.F., Planejamento de experimentos e otimização de processos, 2nd edn (2009) Casa do Pão Editora, , CampinasSeo, J.-Y., Seong, S.Y., Ahn, B.-Y., Kwon, I.C., Chung, H., Jeong, S.Y., Cross-protective immunity of mice induced by oral immunization with pneumococcal surface adhesin A encapsulated in microspheres (2002) Infection and Immunity, 70 (3), pp. 1143-1149. , DOI 10.1128/IAI.70.3.1143-1149.2002Sunitha, K., Kim, Y.-O., Lee, J.-K., Oh, T.-K., Statistical optimization of seed and induction conditions to enhance phytase production by recombinant Escherichia coli (2000) Biochemical Engineering Journal, 5 (1), pp. 51-56. , DOI 10.1016/S1369-703X(99)00062-5, PII S1369703X99000625Tai, S.S., Streptococcus pneumoniae protein vaccine candidates: Properties, activities and animal studies (2006) Critical Reviews in Microbiology, 32 (3), pp. 139-153. , DOI 10.1080/10408410600822942, PII Q37714551L5247J0Volonté, F., Marinelli, F., Gastaldo, L., Sacchi, S., Pilone, M.S., Pollegioni, L., Molla, G., Optimization of glutaryl-7-aminocephalosporanic acid acylase expression in e. Coli (2008) Protein Expr Purif, 61, pp. 131-137Zhao, J., Wang, Y., Chu, J., Zhang, S., Zhuang, Y., Yuan, Z., Statistical optimization of medium for the production of pyruvate oxidase by the recombinant escherichia coli (2008) J Ind Microbiol Biotechnol, 35, pp. 257-26

Caracterização Morfofisiológica e Anatômica de Paspalum urvillei (Steudel) Morphophysiological and Anatomic Characterization of Paspalum urvillei (Steudel)

Este trabalho foi conduzido com o objetivo de caracterizar a dinâmica de afilhamento e acúmulo de biomassa, a fenologia e os aspectos anatômicos de P. urvillei. O ensaio foi desenvolvido em Passo Fundo, Rio Grande do Sul, onde foram colhidas quinzenalmente seis plantas, em um delineamento completamente casualizado, entre 21/10/99 a 21/04/00. As épocas foram avaliadas por unidades de graus-dia (GD) de crescimento, por intermédio da soma das temperaturas médias diárias (temperatura basal = 0&ordm;C). As características anatômicas de folha e colmo foram observadas por cortes histológicos à mão livre. O material foi corado com fucsina-azul de Astra e montado em glicerina, entre lâmina e lamínula. Observou-se um máximo de 70 afilhos/planta, sendo 60% basilares; a área foliar atingiu até 7066 cm²/planta, com 250 folhas verdes/planta. A altura do dossel vegetativo manteve-se em 50 cm. O florescimento foi precoce e longo, sugerindo a necessidade de manejo para encurtar tal estádio, a fim de prolongar o período de produção de folhas. Os estudos anatômicos comprovaram presença de células da bainha ao redor dos feixes vasculares, típico de plantas C4.<br>This work had the purpose to characterize the tillering and biomass accumulation dynamic, phenology and anatomic aspects. The experiment was conducted in Passo Fundo, Rio Grande do Sul, in which six plants were harvested every 15 days, between 21/10/99 to 21/04/00, in a completely randomized design. The times were accounted using units of growing degree-days (DD), calculated as the sum of the daily mean temperatures (threshold temperature = 0&ordm;C). Transverse hand sections were used to observe the anatomic characteristics of leaf and stem. The material was stained with basic fuchsin Astra blue. It was observed a peak of 70 tillers/plant, being 60% basal; the leaf area reached 7066 cm²/plant, with 250 green leaves/plant. The height of the vegetative canopy remained with 50 cm. The flowering time was early and long pointing out to a management practice to shorten this time to maintain the plants on continual leaf production. The anatomic evaluations confirmed the presence of sheath cells (Kranz) around of the vascular bundles, as C4 plants