Basfia succiniciproducens
| Basfia succiniciproducens | ||||||||||||
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| Systematics | ||||||||||||
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| Scientific name of the genus | ||||||||||||
| Basfia | ||||||||||||
| Kuhnert , Scholten , Haefner , Mayor & Frey 2009 | ||||||||||||
| Scientific name of the species | ||||||||||||
| Basfia succiniciproducens | ||||||||||||
| Kuhnert , Scholten , Haefner , Mayor & Frey 2009 |
Basfia succiniciproducens is a Gram stain negative, facultatively anaerobic and immobile bacterium from the Pasteurellaceae family. It was firstisolatedfrom the rumen of Holstein cattle in 2008and scientifically described in 2009. Because of its abilityto producerelatively large quantities of succinic acid through fermentation , it isof potential importancefor industrial biotechnology .
features
The bacterium is cocci to rod-shaped and, like all Pasteurellaceae, has no flagellum , which means that it cannot actively move. The bacterium is gram-negative, so it only has a thin, single-layer murein shell that does not contain any teichonic acids . Information about individual cell sizes is not scientifically documented, colonies reach a diameter of 0.1 to 0.5 millimeters after 24 hours of growth and are colored translucent gray.
Basfia succiniciproducens , like the majority of proteobacteria , was characterized solely by physiological or biochemical features, while it has no eidonomic peculiarities compared to other species and genera of the Pasteurellaceae. The first description thus fulfills the minimum standards applicable for characterizing a new species and genus within the Pasteurellaceae ; there are no further anatomical descriptions.
All known strains anaerobically produce significant amounts of succinic acid, acetic acid and formic acid as metabolic products during fermentation . In doing so, they can (unlike many other genera of the Pasteurellaceae) convert D - mannitol , D - mannose , D - galactose , D - trehalose , sucrose and D - xylose in addition to glucose and glycerine . In the indole test , the bacterium reacts negatively, which means that there is no detection of the enzyme tryptophanase for converting tryptophan to indole . The detection of urease to break down urea and catalase to break down hydrogen peroxide is also negative. The oxidase test for the detection of the enzyme cytochrome c oxidase in the respiratory chain is positive, as is the detection of the production of organic acids by methyl red .
insulation
The first strain (DD1, named after Dirk Dägele) of Basfia succiniciproducens was isolated from the rumen of the Holstein cattle by Edzard Scholten from BASF and Dirk Dägele from Vetter Pharma and examined for the ability to produce succinic acid. The bacterium was isolated during a screening for bacteria from the rumen of domestic cattle that are potentially of interest for the biotechnological production of succinic acid. A targeted search was made for bacteria with a maximum match with the bacterial strain MBEL55E ( Mannheimia succiniciproducens ), of which this property was already known. MBEL55E was isolated, presented and patented for the first time in 2002, but according to information from Kuhnert et al. 2009 not validly described.
For the first description of Basfia succiniciproducens , Peter Kuhnert isolated further strains from different individuals of different domestic cattle breeds in Switzerland. These were the type strain JF4016 T (= DSM 22022 T = CCUG 57335 T ) and JF4141 from the Simmentaler Fleckvieh , the strain JF4136 from the Piedmontese cattle , the strains JF4134 and JF4142 from the Red Holstein cattle and the strains JF42204213 from to the Limousin cattle .
ecology
Basfia succiniciproducens is part of the bacterial composition in the rumen of domestic cattle (rumen flora), although it has so far been detected in the rumen of Holstein cattle , Simmental Simmental cattle , Piemontese cattle , Red Holstein cattle and Limousin cattle .
The bacterium is facultatively anaerobic and draws its nutritional basis from the food pulp in the cow's stomach: the rumen flora, especially the fungi it contains, break the β-glycosidic bonds of the cellulose in the food. The resulting grape sugar (glucose) serves the microorganisms and thus B. succiniciproducens as a substrate, the products of the bacterial metabolism are short-chain carboxylic acids, especially succinic acid. In addition to glucose, the food pulp also contains nitrogen, which is available from the proteins and other nitrogen sources (non-protein nitrogen, NPN) contained in the feed. Proteins are largely split into peptides , amino acids or ammonia by the microorganisms in the rumen . These then serve the microorganisms as a source of nitrogen. The succinic acid produced by bacteria in turn serves as an energy source for other bacteria, which convert it into propionic acid.
B. succiniciproducens does not produce metabolic products that are toxic to cattle or other organisms and is also not (opportunistically) pathogenic , so it is regarded as a normal component of the rumen flora. The negative toxicity test leads to a classification as a non-hazardous organism, which means that the bacterium can be used without any technical restrictions.
Taxonomy
designation
The bacterial genus was their Erstbeschreibern after BASF SE as Basfia named. This name was justified by the fact that the first isolated bacterial strain came from the BASF laboratories in Ludwigshafen am Rhein .
The species name succiniciproducens is derived from the ability to produce succinic acid (Latin: acidum succinicum ), translated it means "producing succinic acid".
Phylogenetic classification
| Phylogenetic position of Basfia succiniciproducens | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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| Cladogram according to Kuhnert et al . 2009 based on the 16S rRNA sequences and the genes rpoB, infB and recN |
The first classification into the Pasteurellaceae carried out during the isolation of the bacterium was confirmed by the first description in 2009, and JF4016 T was defined as the type strain . This family currently (2009) consists of 13 recognized genera with around 60 validly described species, which were primarily isolated from various vertebrates and belong to the typical mucosal flora there; some species are facultatively pathogenic (opportunistic pathogenic ). In addition to B. succiniciproducens , the species Mannheimia ruminalis , Actinobacillus lignieresii , [Actinobacillus] succinogenes and Mannheimia succiniproducens also come from the rumen of domestic cattle; the last two are also succinic acid producers .
Basfia succiniciproducens was identified via the sequence comparison in the form of a so-called multi-locus sequence analysis (MLSA) of the 16S rRNA and the genes rpoB (encodes the β-segment of RNA polymerase ), infB (encoded for the initial factor 2 of translation ) and recN (encoded for a DNA repair protein) within the Pasteurellaceae. The selection of the genes was based on studies on the phylogeny of the Pasteurellaceae that had already been carried out in advance. It could be clearly demonstrated that all known strains of Basfia form a clade and accordingly a monophyletic group , which also includes the MBEL55E strain named Mannheimia succiniciproducens . The greatest agreement of the gene sequence was found with Aggregatibacter actinomycetemcomitans , which is accordingly regarded as a sister species; Due to the comparatively large genetic distance between the two species with a maximum agreement of 95% and the phenotypic differences, a separate genus Basfia was established for B. succiniciprocens . Other species that are closely related are Haemophilus influenzae , Lonepinella koalarum and the [Actinobacillus] porcinus .
Technical importance
Due to its ability to produce succinic acid through fermentation in an anaerobic environment, B. succiniciproducens, like other bacteria, is of great technical interest and is intensively investigated for its usability, especially in the BASF laboratory. A technical use of the bacterium is currently being carried out by BASF in cooperation with the Institute for Bioprocess Engineering at the Technical University of Braunschweig , whereby the metabolism of the bacterium is to be examined and used with the help of metabolic engineering to develop production organisms.
Succinic acid as a biotechnology product
Succinic acid is a platform chemical with an annual demand of around 15,000 tons and a market value of six to nine euros per kilogram. It is conventionally produced on the basis of n-butane and butadiene with maleic anhydride as an intermediate. Especially as a basis for various products in the chemical and pharmaceutical industry ( 1,4-butanediol , tetrahydrofuran, etc.) as well as for bio-based plastics such as polyamides (PA), polyesters and co-polyesters as well as polyester amides , succinic acid is of interest as a biotechnologically manufactured product a market potential of several hundred thousand tons is forecast. Together with other representatives of the C4-dicarboxylic acids such as fumaric and malic acid , succinic acid was identified by the US Department of Energy in 2004 as one of twelve platform chemicals with particular biotechnological manufacturing potential.
In addition to B. succiniciproducens , a number of other potential succinic acid producers are also being investigated, including above all Mannheimia succiniciproducens , Actinobacillus succinogenes , Corynebacterium glutamicum and Anaerobiospirillum succiniciproducens , but also the model organism Escherichia coli , which is to be optimized for the production of high amounts of amber acid via metabolic engineering .
Special features of B. succiniciproducens
| bacterium | Maximum yield (titer) (g / l) |
Productivity (g / l * h) |
Yield (g / g substrate) |
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Basfia succiniciproducens DD1 |
5.8 | 1.5 | 0.6 |
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Mannheimia succiniciproducens MBEL55E |
13.5 | 1.8 | 0.68 |
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Actinobacillus succinogenes 130Z; Wild type |
69-80 | 1.2-1.7 | 0.68-0.87 |
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Corynebacterium glutamicum R; Wild type |
23 | 3.8 | 0.19 |
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Anaerobiospirillum succiniciproducens wild type |
50.3 | 2.1 | 0.9 |
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Escherichia coli SBS550MG / pHL413 |
40 | 0.4-1.2 | 1.05 |
B. succiniciproducens can be cultivated biotechnologically both on the classic fermentation medium glucose (sugar syrup, starch) and on glycerine . Glycerine is obtained as raw glycerine in the production of rapeseed methyl ester ( biodiesel ) in large quantities (about 1:10 the quantity of biodiesel) and is therefore available as an inexpensive raw material for combined use .
In 2008, Scholten & Dägele were able to achieve yields of up to 5.8 g / l succinic acid based on D-glucose and sucrose with the newly isolated and at the time not yet described bacterium, with productivity of up to 1.5 g / l 1 · h and the yield was a maximum of 0.6 grams of succinic acid per gram of substrate. With raw glycerine they achieved yields of 8.4 g / l, 0.9 g / l · h and 1.2 g / g substrate. Compared to other bacteria, especially Actinobacillus succinogenes , Anaerobiospirillum succiniciproducens and modified Escherichia coli , the maximum yield per liter is comparatively low, but the yield based on the substrate input and productivity are comparable and can also be optimized by optimizing the strains. Bacteria based on raw glycerol as a substrate, for which the first approaches to continuous cultivation are in development, show particularly good properties.
Individual evidence
- ↑ H. Christensen, P. Kuhnert, HJ Busse, WC Frederiksen, M. Bisgaard: Proposed minimal standards for the description of genera, species and subspecies of the Pasteurellaceae. In: International Journal of Systematic and Evolutionary Microbiology. 57, 2007, p. 166, doi : 10.1099 / ijs.0.64838-0 . PMID 17220461 .
- ↑ a b c d Edzard Scholten, D. Dägele: Succinic acid production by a newly isolated bacterium. Biotechnology Letters 30 (12), 2008; Pp. 1243-1246. doi : 10.1007 / s10529-008-9806-2 .
- ↑ a b c P.C. Lee, SY Lee, SH Hong, HN Chang: Isolation and characterization of a new succinic acid-producing bacterium, Mannheimia succiniciproducens MBEL55E, from bovine rumen. Applied Microbiology and Biotechnology 58, 2002; Pp. 663-668. doi : 10.1007 / s00253-002-0935-6 .
- ↑ a b c d P. Kuhnert, E. Scholten, S. Haefner, D. Mayor, J. Frey: Basfia succiniciproducens gen. Nov., Sp. nov., a new member of the family Pasteurellaceae isolated from bovine rumen . In: International Journal of Systematic and Evolutionary Microbiology . tape 60 , no. 1 , January 1, 2010, p. 44-50 , doi : 10.1099 / ijs.0.011809-0 .
- ↑ "..derived from the chemical company BASF SE in Ludwigshafen, Germany, in reference to the origin of the first strain Characterized." Kuhnert et al. 2009; P. 7.
- ↑ Succinic acid - with biotech to the green platform chemical. Presentation from February 1, 2010 on the website of BIOPRO Baden-Württemberg GmbH.
- ↑ Production of polyesters based on succinic acid produced by fermentation or 1,4-butanediol Project description on the website of the Institute for Bioprocess Engineering . Retrieved February 21, 2012.
- ↑ a b c d e f James B. McKinley, C. Vieille, J. Gregory Zeikus: Prospects for a bio-based succinate industry. Applied Microbiology and Biotechnology 76, 2007; Pp. 727-740.
- ^ A b c I. Bechthold, K. Bretz, S. Kabasci, R. Kopitzky, A. Springer: Succinic Acid: A New Platform Chemical for Biobased Polymers from Renewable Resources. Chemical Engineering & Technology 31 (5), (2008); Pp. 647-654. doi : 10.1002 / ceat.200800063 .
- ↑ T. Werpy, G. Petersen: Top Value Added Chemicals from Biomass. Volume I — Results of Screening for Potential Candidates from Sugars and Synthesis Gas. Produced by the Staff at Pacific Northwest National Laboratory (PNNL); National Renewable Energy Laboratory (NREL), Office of Biomass Program (EERE), 2004 ( Download ; PDF; 1.5 MB)
- ↑ Sang Yup Lee, Ji Mahn Kim, Hyohak Song, Jeong Wook Lee, Tae Yong Kim, Yu-Sin Jang: From genome sequence to integrated bioprocess for succinic acid production by Mannheimia succiniciproducens. Applied Microbiology and Biotechnology 79 (1) May 2008; Pp. 11-22. doi : 10.1007 / s00253-008-1424-3 .
- ↑ a b A.M. Sanchez, GN Bennett, KY San: Novel pathway engineering design of the anaerobic central metabolic pathway in Escherichia coli to increase succinate yield and productivity. Metabolic Engineering 3, 2005; Pp. 229-239, doi: 10.1016 / j.ymben.2005.03.001 .
- ↑ a b Edzard Scholten, Torsten Renz, Jochen Thomas: Continuous cultivation approach for fermentative succinic acid production from crude glycerol by Basfia succiniciproducens DD1 Biotechnology Letters, 31, 2009, p. 1947, doi : 10.1007 / s10529-009-0104-4 .
- ↑ J. Hangebrauk, T. Fürch, G. von Abendroth, C. Wittmann: Optimization of the biotechnological production of succinic acid Abstract to the lecture at the ProcessNet annual conference / 27. Annual meeting of biotechnologists, published in CIT - Chemie Ingenieur Technik 81 (8), 2009; Pp. 1209-1210.
