Mutants of Corynebacterium diphtheriae PW8 that produce toxin in medium with excess iron.

Two mutants that produce toxin in medium with excess iron were isolated from the PW8 strain of Corynebacterium diphtheriae. These mutants produced as much toxin in medium containing excess iron (3 mug of Fe2+ per ml) as did the parent PW8 strain in iron-depleted medium, and they will be very useful for easy production of toxin.     

The effect of the iron content in the culture medium on the nature of the protein spectrum in Neisseria meningitidis

The protein spectra of Neisseria meningitidis, strain A 208, in the process of its cultivation on solid culture medium based on peptone agar under the conditions of the surplus or deficiency of ions of trivalent iron in the medium. The creation of iron deficiency by the introduction of two iron-binding admixtures, desferol and diethylenetriaminepentaacetic acid (DTPA), was found to lead to the production of two additional proteins with molecular weights of 72 kD and 36-37 kD by meningococcal cells. Of these two admixtures, DTPA more readily stimulated the production of low-molecular protein with a molecular weight of 36-37 kD. This protein was found to be noticeably labile, while protein with a molecular weight of 72-73 kD showed no such lability. As the result of the cultivation of meningococci in iron-deficient medium, the content of protein in microbial residue was 2- to 3-fold greater than that obtained by the cultivation of meningococci in culture medium with the surplus of iron in the form of ferric nitrate.     

Regulation of toxinogenesis in Corynebacterium diphtheriae: mutations in the bacterial genome that alter the effects of iron on toxin production

Mutants of Corynebacterium diphtheriae C7(beta) that are resistant to the inhibitory effects of iron on toxinogenesis were identified by their ability to form colonies surrounded by toxin-antitoxin halos on agar medium containing both antitoxin and a high concentration of iron. Chromosomal mutations were essential for the altered phenotypes of four independently isolated mutant strains. During growth in deferrated liquid medium containing various amounts of added iron, these mutants differed from wild-type C. diphtheriae C7(beta) in several ways. Their growth rates were slower under low-iron conditions and were stimulated to various degrees under high-iron conditions. The concentrations of iron at which optimal toxin production occurred were higher for the mutants than for wild-type C. diphtheriae C7(beta). Toxin production by the mutants during growth in low-iron medium occurred throughout the period of exponential growth at nearly constant rates that were proportional to the bacterial growth rates. In contrast, toxin production by wild-type C. diphtheriae C7(beta) in similar low-iron cultures occurred predominantly during the late exponential phase, when iron was a growth-limiting nutrient. Additional studies demonstrated that these mutants had severe defects in their transport systems for ferric iron. We propose that the altered regulation of toxinogenesis by iron in our mutants was caused by the severe defects in their iron transport systems. As a consequence, the mutants exhibited a low-iron phenotype during growth under conditions that permitted wild-type C. diphtheriae C7(beta) to exhibit a high-iron phenotype.     

Effect of culture medium iron content on the biochemical composition and metabolism of Trichomonas vaginalis.

Trichomonas vaginalis grown in iron-enriched medium contained increased concentrations of iron-sulfur proteins, including ferredoxin and pyruvate-ferredoxin oxidoreductase. The increases in hydrogenosomal constituents correlated with increased in vivo hydrogenosomal metabolism.     

Effect of medium composition on the production of lysine byCorynebacterium sp. 9366-H-454

The strainCorynebacterium sp. 9366-H-454 grown on medium containing 10% sucrose and 4% corn-steep was found to produce as much as three times or four times (under laboratory conditions and on a pilot plant scale, respectively) more lysine than on the original. H-medium. The values are economical for a fermentation preparation of lysine.     

The effect of medium composition on the production of valine byCorynebacterium 9366-EMS/184

The auxotrophic mutantCorynebacterium 9366-EMS/184, requiring isoleuoine accumulates about 22 g of L-valine per litre after a 96 h cultivation in a medium containing 15% sucrose and 1% ammonium acetate.     

Cytotoxic activity of a recombinant chimaeric protein between Pseudomonas aeruginosa exotoxin A and Corynebacterium diphtheriae diphtheria toxin

A segment of the exotoxin A gene of Pseudomonas aeruginosa, coding for the N-terminal end of domain I and domain II of the toxin (ETA), was genetically fused to the diphtheria toxin gene of Corynebacterium diphtheriae, coding for the N-terminal end of A fragment of diphtheria toxin (DT). The resulting hybrid protein (termed CED1) was produced in large amounts and exported to the periplasm in Escherichia coli. This chimaeric protein reacted with both anti-ETA and anti-DT antisera. Furthermore, the chimaeric protein displayed ADP-ribosylation activity and exhibited cytotoxicity to mouse 3T6 fibroblasts. These results demonstrated that the chimaeric protein is cytotoxic, and that the toxic potential of DTA can be selectively internalized and translocated via domains I and II of exotoxin A, which are thus sufficient to direct and translocate an enzymatically active heterologous polypeptide segment into the cytosol of sensitive cells.     

Regulation of toxinogenesis in Corynebacterium diphtheriae. I. Mutations in bacteriophage beta that alter the effects of iron on toxin production

Diphtherial toxin is produced in maximal yields by Corynebacterium diphtheriae (C7(beta tox+) only when iron is present in growth-limiting amounts. Toxin production is markedly decreased under high-iron conditions. We studied the role of the bacteriophage beta genome in this apparent regulation of toxin production by iron. Using a passive immune hemolysis assay to detect toxin antigen production in individual plaques, we identified rare phage mutants that were toxinogenic in high-iron medium. Lysogenic derivatives of C. diphtheriae C7 harboring such phage mutants were constructed. The lysogens were compared with wild-type strain C7(beta) for their ability to produce toxin in deferrated liquid medium containing varying amounts of added iron. Quantitative tests for extracellular toxin were performed by competitive-binding radioimmunoassays. We identified phenotypically distinct mutant strains that produced slightly, moderately, or greatly increased yields of toxin antigen under high-iron conditions. The toxin produced by the mutant lysogens was biologically active and immunochemically indistinguishable from wild-type toxin. Complementation experiments demonstrated that the phage mutation designated tox-201 had a cis-dominant effect on the expression of the toxin structural gene of phage beta. The characteristics of the tox-201 mutation suggest that it defines a regulatory locus of phage beta that is involved in control of toxinogenesis by iron in C. diphtheriae.     

Biomass production and biochemical variability of the marine microalga Isochrysis galbana in relation to culture medium

We have studied the autotrophic growth of the marine microalga, Isochrysis galbana Parke, in a batch photobioreactor, comparing five different culture media and analysing the influence of each on growth kinetics as well as on the fatty-acid composition and protein content of the biomass. All the experiments were performed at 15 degrees C, with the culture medium at pH 8.0, a specific rate of air supply of 1vv(-1)min(-1) and a continuous illumination of 40-43Wm(-2). The results show no parallel between good nutritional characteristics and high values of the kinetic parameters. Nevertheless, a compromise between the nutritional factors and growth kinetics could be provided by Ukeles medium, which provided a biomass with a good composition in polyunsaturated fatty acids (quotient n3/n6=3.2), an adequate protein content (25.3%) and relatively high values, although not the highest registered, for maximum specific growth rate (μ(m)=0.018h(-1)) and biomass productivity (1.9x10(-3)kgm(-3)h(-1)).