Chemically Defined Medium for the Growth of Clostridium perfringens

A defined medium which supports growth of Clostridium perfringens at low inoculum levels was developed. Generation time for strain 8797 was 1.5 times greater than previously reported for growth in purged fluid thioglycolate medium.     

Oxytetracycline Formation by Streptomyces rimosus in Chemically Defined Media

Details in the fermentation of oxytetracycline in a synthetic medium with Streptomyces rimosus have been presented. In these studies, an organic nitrogen source was shown to be essential for the production of significant amounts of antibiotic activity. Of the amino acids tested, aspartic acid, proline, threonine, valine, and beta-alanine were utilized well for both growth and antibiotic production. Markedly different fermentation patterns were observed with aspartic acid and beta-alanine. Glycerol and glucose supported antibiotic yields superior to those found with other carbohydrates tested. Short chain organic acids were not effectively utilized for growth in the absence of a readily fermentable carbohydrate.     

A Chemically Defined Medium for Rabbit Embryo Cryopreservation

This study evaluates a new synthetic substitute (CRYO3, Ref. 5617, Stem Alpha, France) for animal-based products in rabbit embryo cryopreservation solutions. This evaluation was performed using two approaches: a thermodynamic approach using differential scanning calorimetry and a biological approach using rabbit embryo slow-freezing. During the experiment, foetal calf serum (FCS) was used as a reference. Because FCS varies widely by supplier, three different FCS were selected for the thermodynamic approach. The rabbit embryo slow-freezing solutions were made from Dulbecco''s phosphate buffer saline containing 1.5 M Dimethyl Sulfoxide and 18% (v.v⁻¹) of CRYO3 or 18% (v.v⁻¹) of FCS. These solutions were evaluated using four characteristics: the end of melting temperature, the enthalpy of crystallisation (thermodynamic approach) and the embryo survival rates after culture and embryo transfer (biological approach). In the thermodynamic approach, the solutions containing one of the three different FCS had similar mean thermodynamic characteristics but had different variabilities in the overall data with aberrant values. The solution containing CRYO3 had similar thermodynamic properties when compared to those containing FCS. Moreover, no aberrant value was measured in the solution containing CRYO3. This solution appears to be more stable than the solutions containing a FCS. In the biological approach, the in vitro embryo survival rates obtained with the solution containing CRYO3 (73.7% and 81.3%) and with the solution containing a FCS (77.6% and 71.9%) were similar (p = 0.7). Nevertheless, during the in vivo evaluation, the implantation rate (21.8%) and the live-foetuses rate (18.8%) of the CRYO3 group were significantly higher than the implantation rate (7.1%, p = 0.0002) and the live-foetuses rate (5.3%, p = 0.0002) of the FCS group. The pregnancy rate was also higher in the CRYO3 group compared to the FCS group (81.3% and 43.8%, respectively, p = 0.066). We conclude that CRYO3 can be used as a chemically defined substitute for animal-based products in rabbit embryo cryopreservation solutions.     

Lovastatin Biosynthesis by Aspergillus terreus in a Chemically Defined Medium

Lovastatin is a secondary metabolite produced by Aspergillus terreus. A chemically defined medium was developed in order to investigate the influence of carbon and nitrogen sources on lovastatin biosynthesis. Among several organic and inorganic defined nitrogen sources metabolized by A. terreus, glutamate and histidine gave the highest lovastatin biosynthesis level. For cultures on glucose and glutamate, lovastatin synthesis initiated when glucose consumption levelled off. When A. terreus was grown on lactose, lovastatin production initiated in the presence of residual lactose. Experimental results showed that carbon source starvation is required in addition to relief of glucose repression, while glutamate did not repress biosynthesis. A threefold-higher specific productivity was found with the defined medium on glucose and glutamate, compared to growth on complex medium with glucose, peptonized milk, and yeast extract.     

Chemically Defined Medium for Growth and Sporulation of Clostridium perfringens

A chemically defined medium was developed that could support sporulation and growth of Clostridium perfringens strains ATCC 12916 and H9. This medium consisted of a modification of the basal medium of Boyd et al. plus 0.1% sodium thioglycolate and 0.5% monosodium glutamate. Five other strains grew, but did not sporulate, in this medium. With the addition of more vatamins into the medium, two more strains grew but did not sporulate. The effects of glucose, monosodium glutamate, ammonium glutamate, and sodium thioglycolate on growth and sporulation of C. perfringens ATCC 12916 in the defined medium was investigated.     

Production of Hemolysin by Vibrio parahaemolyticus in a Chemically Defined Medium

A chemically defined medium capable of supporting the production of heat-stable and heat-labile hemolysins by Vibrio parahaemolyticus is described. The indispensability of serine and glutamic acid for hemolysin production is also demonstrated.     

Development of a Chemically Defined Medium for the Growth of Leuconostoc mesenteroides

A chemically defined medium for the growth of Leuconostoc mesenteroides was developed. This medium contained lactose, Mn(sup2+), Mg(sup2+), 12 amino acids, eight vitamins, adenine, uracil, and Tween 80. We showed the beneficial effect of aerobic conditions on growth and that potassium phosphate (135 mM) is a suitable buffer. The growth rate in this medium was 0.85 (plusmn) 0.10 h(sup-1) for the six strains examined, and cell densities up to 3.5 x 10(sup9) CFU/ml were attained.     

High Aflatoxin Production on a Chemically Defined Medium

Aspergillus parasiticus ATCC 15517 produced 28 to 30 mg of aflatoxin per 100 ml of a medium containing sucrose, asparagine, and salts in stationary and shaken cultures. In the absence of asparagine in the medium, the toxin yields fell drastically, and the thin-layer chromatograms of the chloroform extracts of the cultures indicated the total absence of aflatoxin G₁ and the presence of new intense blue and green fluorescent bands having R_F values lower than aflatoxins. Initial pH was critical and had to be around 4.5 for good growth and high toxin production on this medium. Optimum concentrations of KH₂PO₄ and MgSO₄·7H₂O in the medium were much lower than those normally used in fungal growth media.     

Heat-stable Chemically Defined Medium for Growth of Animal Cells in Suspension

Procedures are described for preparation of a chemically defined medium, capable of being autoclaved, for the cultivation of animal cells in suspension. Yields of 34.1 x 10(5), 33.8 x 10(5), and 47.1 x 10(5) cells per ml were recorded for cat kidney, HeLa, and mouse fibroblast (L) cells after 6, 10, and 10 days of incubation, respectively.     

Chemically Defined Medium for Cultivation of Several Epiphytic and Phytopathogenic Spiroplasmas

A chemically defined medium, LD82, was formulated for in vitro cultivation of spiroplasmas. Medium LD82 supported good growth for four epiphytic and insect-pathogenic spiroplasmas, Spiroplasma floricola 23-6^T, Spiroplasma sp. strain SR3, Spiroplasma sp. strain brevi, and Spiroplasma sp. strain AS576, and of the phytopathogenic spiroplasmas Spiroplasma citri Maroc R8A2^T and PC1. Titers of all six strains grown in defined medium LD82 reached 2.0 × 10⁹ to 6.0 × 10⁹ CFU/ml of culture. All spiroplasma strains tested formed colonies readily on agar medium LD82. None of the spiroplasmas formed typical fried-egg colonies. All formed diffuse colonies, but the forms of colonies differed somewhat among the spiroplasma strains. In preliminary studies of nutritional requirements, phospholipids slightly enhanced the growth of the epiphytic and insect-pathogenic strains in medium LD82 and were found essential for good growth of S. citri.     

Chemically Defined Minimal Medium for Growth of the Anaerobic Cellulolytic Thermophile Clostridium thermocellum

A minimal chemically defined medium has been developed for Clostridium thermocellum. The growth factors required are biotin, pyridoxamine, vitamin B₁₂, and p-aminobenzoic acid.     

Protein Production in Chemically Defined Medium by Bacillus brevis No. 47

A simple chemically defined medium was devised for exoprotein production by Bacillus brevis No. 47. About 2 mg/ml of proteins was produced in the synthetic medium containing 4% glucose and 1% ammonium sulfate. An essential component of fermentation medium was Ca salt which is required by this organism for assimilating glucose.

Studies on the effects of various medium components on protein production revealed that the conditions appropriate for growth are also suitable for protein accumulation. Some compounds, especially inhibitors of cell wall synthesis and certain detergents, were found to enhance protein production.     

On the Iron Requirement of Lactobacilli Grown in Chemically Defined Medium

The iron requirement of four strains of lactobacilli (L. acidophilus, L. delbrueckii subsp. bulgaricus, L. plantarum, and L. pentosus) was studied in a synthetic medium under aerobic or anaerobic conditions. Effects of iron salt and iron-chelated compounds were tested on bacterial growth in manganese-free or -supplemented media. No significant growth stimulation was observed in any condition. These results support the absolute manganese requirement for optimum growth of lactobacilli and the needless incorporation of iron in growth media. Received: 5 November 1997 / Accepted: 20 January 1998     

Microcycle Sporogenesis of Bacillus cereus in a Chemically Defined Medium

A chemically defined medium which allowed germination, outgrowth, and subsequent resporulation of Bacillus cereus T spores, without intervening cell division (microcycle sporogenesis), is described. No medium replacement was required. The second-stage spores were heat-stable and had similar germination characteristics and dipicolinic acid content to primary spores. Deoxyribonucleic acid (DNA) replication began soon after germination and there was a doubling in the DNA content of the cells within 2 hr.     

Acetoin Catabolism and Acetylbutanediol Formation by Bacillus pumilus in a Chemically Defined Medium

Background

Most low molecular diols are highly water-soluble, hygroscopic, and reactive with many organic compounds. In the past decades, microbial research to produce diols, e.g. 1,3-propanediol and 2,3-butanediol, were considerably expanded due to their versatile usages especially in polymer synthesis and as possible alternatives to fossil based feedstocks from the bioconversion of renewable natural resources. This study aimed to provide a new way for bacterial production of an acetylated diol, i.e. acetylbutanediol (ABD, 3,4-dihydroxy-3-methylpentan-2-one), by acetoin metabolism.

Methodology/Principal Findings

When Bacillus pumilus ATCC 14884 was aerobically cultured in a chemically defined medium with acetoin as the sole carbon and energy source, ABD was produced and identified by gas chromatography – chemical ionization mass spectrometry and NMR spectroscopy.

Conclusions/Significance

Although the key enzyme leading to ABD from acetoin has not been identified yet at this stage, this study proposed a new metabolic pathawy to produce ABD in vivo from using renewable resources – in this case acetoin, which could be reproduced from glucose in this study – making it the first facility in the world to prepare this new bio-based diol product.     

Biotransformation of benzyl thiocyanate by Streptomyces aureofaciens

Benzyl thiocyanate, a specific stimulator of chlortetracycline biosynthesis, was transformed into dibenzyl disulphide by Streptomyces aureofaciens. The disulphide stimulated chlortetracycline production to a lesser extent than did benzyl thiocyanate.     

Vero细胞微载体培养的化学成分明确无血清培养基的设计

目的：设计适用于Vero细胞微载体培养的化学成分明确无血清培养基。方法：以商品化的DMEM／F12合成培养基为基础培养基,应用Plackett—Burman实验设计和响应面分析法设计支持Vero细胞微载体培养的化学成分明确无血清培养基。结果：以细胞密度为评价指标,在单因素实验的基础上采用Plackett-Burman实验设计考察10种培养基添加成分对Vero细胞生长的影响,确定了3种对Vero细胞生长起明显促进作用的培养基添加成分,为胰岛素、血清素和腐胺。继而利用响应面法分析了这3种添加成分的最佳水平范围,设计了一种支持Vero细胞贴附培养的无血清培养基（VERO—SFM—A）。在Bellco搅拌式培养瓶中采用VERO-SFM．A和Cytodex1微载体培养Vero细胞,细胞密度由接种时的4×10^5cells／ml增加到培养6d后的22．3×10^cells／ml,细胞活力保持在96％以上。结论：VERO—SFM—A能够有效地支持Vero细胞在微载体表面固定化生长并达到较高的细胞密度,具有实际应用于Vero细胞微载体规模化培养的应用潜力。     

Nutritional Studies on Acanthamoeba culbertsoni and Development of Chemically Defined Medium

A chemically defined medium containing 11 amino acids, 3 vitamins, 6 inorganic salts and glucose, yielding maximum cell densities of 1.5-2.5 × 10⁷ cells/ml, has been developed for Acanthamoeba culbertsoni with a mean generation time (MGT) of 10 h. A medium containing six amino acids viz. arginine, methionine, leucine, isoleucine, valine and glycine along with other components could also support good albeit slower growth (MGT 27 h) of the amoeba. Acetate did not serve as a suitable carbon/energy source for A. culbertsoni. This organism bears close resemblance in its nutritional requirements to other Acanthamoeba especially A. polyphaga.     

Inhibitor of Clostridium perfringens Formed by Heating Sodium Nitrite in a Chemically Defined Medium

An inhibitor of Clostridium perfringens formed when low levels of nitrite were autoclaved with a defined chemical medium. A systematic study of the medium revealed that only amino acids and mineral salts were involved in the production of this inhibitor, which was proven to be a toxic compound formed from cysteine, ferrous sulfate, and sodium nitrite. The inhibitor was compared to several known compounds. S-nitrosocysteine inhibited the test organism, but would not form in the test system in amounts large enough to explain the observed inhibition. Roussin red salt was unstable in the test system and therefore was not the inhibitor. Roussin black salt, which was also inhibitory, could form in sufficient amounts to explain the inhibition. A complex of cysteine, iron, and nitric oxide was detected in the autoclaved solution of cysteine, ferrous sulfate, and sodium nitrite; this cysteine complex did not appear to be inhibitory, however, at levels which could form in the autoclaved medium. The observed inhibition may have been due to the combined effects of sublethal concentrations of each compound.