Low Aeration Conditions Research Articles

Expression of multiple cytochromeb 556 species in aerobically grownEscherichia coli

Cytochrome with anα-band absorption maximum at 556 nm (77K) in the reduced minus oxidized spectrum is observed in membrnes fromEscherichia coli grown aerobically on most carbon sources. Previous work has suggested that this adsorption peak is due to cytochromeb556 of succinate dehydrogenase and to cytochromeo. We show here, by partial purification of the membrane cytochromes, that at least two other cytochromes with absorption maxima at 556 nm contribute to this peak. One of these cytochromes is associated with growth ondl-lactate. The other is formed under conditions of low aeration and has hydroperoxidase activity.

Growth of roots of differentPinus radiatagenotypes in soil at different strength and aeration

Summary The penetration and growth of roots of different genotypes of Pinus radiata into Stockdale loamy sand (it has a clay layer at 55–70 cm depth) at different soil strength and aeration were investigated. Soil aeration below 10% air-filled pore space significantly retarded root growth. There were significant differences between Pinus radiata families in root growth in the clay layer at high soil strength and low aeration. Families also differed in their sensitivity towards high soil strength and low aeration conditions in the clay layer. The results of this investigation suggest that there is scope for selecting genotypes that can penetrate and grow in clay layers of high strength or low aeration.

Physiological Properties of a Mutant of Pachysolen tannophilus Deficient in NADPH-Dependent d -Xylose Reductase

A d-xylose reductase mutant of Pachysolen tannophilus was isolated on the basis of its poor growth on d-xylose but normal growth on xylitol and d-glucose. Fractionation of cell extracts indicated that the mutant was deficient in d-xylose reductase activity that used NADPH exclusively as a cofactor, but not in activity that used both NADH and NADPH. Mutant cultures grown on d-xylose as the sole carbon source exhibited some properties that would be desired in improved strains. Growth rate, growth yield, and d-xylose consumption rate of the mutant were less sensitive than those of the wild type to changes in aeration rate. d-Xylose was utilized more efficiently in that less of a by-product, xylitol, was produced. In addition, under low aeration conditions, more ethanol was produced. A disadvantage was a relatively slow rate of d-xylose utilization.

Hyperproduction of Poly-β-Hydroxybutyrate during Exponential Growth of Azotobacter vinelandii UWD

The transformation of Azotobacter vinelandii UW with A. vinelandii 113 DNA resulted in the formation of rifampin-resistant colonies, 13% of which also inherited a previously unrecognized mutation in the respiratory NADH oxidase. These transformants produced colonies with a white-sectored phenotype after prolonged incubation. Cells from these sectors were separated and purified by streaking and were named UWD. The dense white phenotype was due to the production of a large amount of poly-beta-hydroxybutyrate during the exponential growth of strain UWD. The polymer accounted for 65 or 75% of the cell dry weight after 24 h of incubation of cultures containing glucose and either ammonium acetate or N(2), respectively, as the nitrogen source. Under the same conditions, strain UW cells contained 22 to 25% poly-beta-hydroxybutyrate, but O(2)-limited growth was required for these optimal production values. Polymer production was not dependent on O(2) limitation in strain UWD, but the efficiency of conversion of glucose to poly-beta-hydroxybutyrate was enhanced in O(2)-limited cultures. Conversion efficiencies were >0.25 and 0.33 mg of poly-beta-hydroxybutyrate per mg of glucose consumed under vigorous- and low-aeration conditions, respectively, compared with an efficiency of 0.05 achieved by strain UW. Strain UWD, therefore, appeared to from poly-beta-hydroxybutyrate under novel conditions, which may be useful in designing new methods for the industrial production of biodegradable plastics.

Growth of a Tryptophanase-producing Thermophile, Symbiobacterium thermophilum gen. nov., sp. nov., Is Dependent on Co-culture with a Bacillus sp.

SUMMARY: An obligately symbiotic bacterium, strain T, associating with a specific strain S of a thermophilic Bacillus sp. was isolated during screening for micro-organisms that produce heat-stable tryptophanase and β-tyrosinase. A colony containing both strains was finally isolated by successive selection with bacitracin. In mixed culture with Bacillus strain S under low aeration conditions at 60°C, growth of strain T was initiated following lysis of the Bacillus cells. No independent growth of strain T was observed in any of the media tested, even in the presence of other thermophilic Bacillus strains or cell-free extracts of strain S or its killed cells. No physical adhesion between cells of strains S and T was observed microscopically. Tryptophanase and β-tyrosinase production by strain T were induced by L-tryptophan and L-tyrosine, respectively. The name Symbiobacterium thermophilum gen. nov., sp. nov. is proposed for strain T which is a Gram-negative rod (0.25-0.35 pm diameter) with a multilayer surface structure containing meso-diaminopimelic acid. The G+C content of the DNA is 65.1 mol%; iso-C15:0, iso-c17:0 and anteiso-C17:0 acids are the major cellular fatty acids. The type strain is strain IAM 13621.

Myeloperoxidase-mediated damage to the succinate oxidase system of Escherichia coli. Evidence for selective inactivation of the dehydrogenase component.

Myeloperoxidase, a granule-associated enzyme of neutrophils and monocytes, combines with H2O2 and chloride to form a potent microbicidal system that contributes to phagocyte antimicrobial activity. The nature of the lesion or lesions induced by the myeloperoxidase system which are responsible for the loss of microbial replicative activity (viability) remains unknown. Using Escherichia coli grown to late log or stationary phase under conditions of low aeration with succinate as the sole carbon source, we found that myeloperoxidase-induced loss of microbial viability could be correlated with a decrease in succinate-dependent respiration (succinate oxidase activity). Succinate dehydrogenase activity fell rapidly to undetectable levels during incubation with the myeloperoxidase system, suggesting that damage to the dehydrogenase was a major factor in the loss of oxidase activity. Other components of the succinate oxidase system were resistant to the actions of myeloperoxidase. The ubiquinone-8 and cytochrome components of the respiratory chain remained nearly constant in amount despite reduction of respiration to undetectable levels. However, as expected from the loss of succinate dehydrogenase activity, succinate-ubiquinone reductase and succinate-cytochrome reductase activities were markedly impaired. We propose that the loss of E. coli viability induced by the myeloperoxidase-H2O2-chloride system is due in part to the loss of electron transport function consequent to the oxidation of critical catalytic centers in susceptible dehydrogenases.

Studies on the ventilation in submerged fermentations. Part VII. Influence of carbon dioxide partial pressure on pH in submerged cultures.

Bacillus subtilis AJ 3250 is capable of accumulating a remarkable amount of α-amylase in the culture liquid on pH-stat submerged fermentation. This microorganism requires sufficient oxygen as in other aerobic fermentations, and amylase production can only reach the maximum when the agitation and aeration conditions maintain the dissolved oxygen concentration above the critical level in pH controlled cultures. However, in pH uncontrolled cultures, amylase production as well as cell mass activity increase under low aeration conditions such as 1/20 vvm. Studies were carried out to analyse this phenomenon, and some new ways to express the relationship between the behaviour of pH and that of carbon dioxide partial pressure in a culture system were introduced. In this study, it was found that the differentiation in the carbon dioxide evolution rate reflects the dynamic characteristics of the pH change in this type of fermentation. An appropriate method of neutralization for managing pH-stat cultures on submerge...

Carbon monoxide-binding cytochromes in the respiratory chain of the thermophilic bacterium PS3 grown with sufficient or limited aeration.

The effects of aeration on the growth and cytochrome patterns of thermophilic bacterium PS3 were studied; bacteria grown with strong aeration synthesized cytochromes c, b, and aa3, while those grown with low aeration, showing non-exponential growth, synthesized higher amounts of cytochromes c and b including o, and a lower amount of cytochrome a (a3). The CO-difference spectra indicated that the terminal oxidase was cytochrome aa3 for high aeration conditions and the cytochrome o for low aeration conditions. Cytochrome o can be solubilized by Triton X-100 from the membrane fraction of bacteria grown under oxygen-limited conditions. The carbon monoxide complex of cytochrome o, obtained by exposing this extract to CO, was photolyzed and the subsequent rebinding of CO was analyzed; it followed first order kinetics with a rate constant of around 8 s-1 at 25 degrees C. At liquid nitrogen temperature, CO-rebinding did not occur. The CO-difference spectrum of purified cytochrome oxidase sample from the bacteria grown with strong aeration (Sone, N., et al. (1979) FEBS Lett. 106, 39-42) revealed the presence of a small amount of a cytochrome o-like pigment besides cytochrome aa3. Analysis of the CO complexes of these chromophores showed rate constants of 29-30 s-1 for cytochrome aa3 and 35-42 s-1 for the o-like pigment, indicating that the cytochrome o-like pigment contaminating the purified cytochrome oxidase preparation was not typical cytochrome o.

Effect of aeration and pH on gramicidin S production by Bacillus brevis

AbstractAlthough the biosynthesis of the antibiotic gramicidin S (GS) by Bacillus brevis ATCC 9999 has been studied extensively, almost no attention has been given to environmental control of its fermentation process. In this respect, GS fermentations conducted in a 7.5 dm3 fermentor in complex (YP) medium revealed that a high aeration rate resulted in a high biomass yield (12 g DCW dm−3) with very low GS levels (170 mg GS dm−3). Lowering the aeration rate (5 dm3 air min−1 at 300 rev min−1) caused a dramatic increase in GS formation (2100 mg GS dm−3) and comparable but slower biomass formation. In chemically‐defined (F3/6) medium fermentations, an aeration rate of 5 dm3 air min−1 at 300 rev min−1 was apparently too high as only 0.104 mg GS mg−1 DCW was produced. A much lower aeration rate (2 dm3 air min−1 at 250 rev min−1) was needed to arrive at a higher specific antibiotic level: 0.130 mg GS mg−1 DCW. These data seem compatible with the finding that oxygen is known to inactivate the GS‐synthetases. Furthermore, keeping the pH constant at 7.3 under low aeration conditions increased specific GS production up to 0.220 mg GS mg−1 DCW in YP, as well as in F3/6 fermentations. Both environmental pH and dissolved oxygen tension clearly affect growth pattern, growth extent and GS production in these high yielding media. These data stress the importance of controlling pH and aeration rate during GS fermentations.

Influence of cultural conditions on the lipopolysaccharide composition of Neisseria sicca.

SUMMARY: Growth rates of Neisseria sicca in batch cultures were controlled by changing aeration rates. Organisms cultured at high aeration rates grew more rapidly and had a greater proportion of lipopolysaccharide (LPS) than had organisms grown at low aeration rates. The LPS isolated from bacteria grown at a rapid rate had higher hexosamine and 3-deoxyoctulosonic acid (KDO) contents and higher galactosamine to glucosamine ratios than LPS from bacteria grown at a slower rate. These results indicated that environmental conditions affected the content and composition of the bacterial LPS. The LPS of organisms from highly aerated cultures had a larger percentage of unsaturated fatty acids than had that from organisms grown under conditions of low aeration, suggesting that oxygen-dependent desaturation of fatty acids had occurred.

The regulation of synthesis of iron and magnesium tetrapyrroles. Observations with mutant strains of Rhodopseudomonas spheroides

1. Cell suspensions of mutant strains of Rhodopseudomonas spheroides, which cannot form bacteriochlorophyll, have been examined for their ability to form other tetrapyrroles under conditions of low aeration. With the exception of strain L-57, the mutants could form carotenoids. 2. All strains, like the parent organism, formed iron protoporphyrin when incubated with delta-aminolaevulate, showing that the iron branch of the biosynthetic pathway operated. 3. Magnesium protoporphyrin or its monomethyl ester was also formed from delta-aminolaevulate by all strains with the exception of L-57. 4. Coproporphyrin and coproporphyrinogen were accumulated by the parent and by strains 2/73 and 2/21 when incubated with glycine and succinate in the presence of ethionine. Strain 2/33, which required methionine for growth, accumulated these compounds in the presence and absence of methionine. 5. Strain L-57 did not accumulate porphyrins from glycine and succinate under any conditions. However, the delta-aminolaevulate synthase of this mutant showed the same rise in activity in response to reduced aeration as did that of the parent organism. 6. Ethionine inhibited production of protoporphyrin and its derivatives from delta-aminolaevulate by the parent strain. 7. The accumulation of coproporphyrin(ogen) under conditions of methionine deficiency may reflect the presence of enzymes of the magnesium branch of the biosynthetic pathway. Strain L-57 may lack a genetic element which determines the development of the entire photosynthetic apparatus. Since this strain did not accumulate coproporphyrin(ogen), the possibility of a specific delta-aminolaevulate synthase, directed towards bacteriochlorophyll synthesis, should be considered.

The production of bacteriophage μ2

AbstractIn three series of experiments, 3‐l., 20‐l., and 150‐l. bacterial cultures were grown in stirred, deep culture vessels to average bacterial cell densities of 71 × 108, 63 × 108, and 43 × 108 viable organisms per milliliter, respectively, and then infected with phage. The average yield of progeny phage in each case was ca. 3000 mpfu (minimum plaque‐forming units) per cell. Thus, the average mass of phage obtained in the 3‐l. experiments was not less than 124 mg./l., calculated from the plaque counts, assuming a particle size of 3.6 × 106 Daltons for the μ2 phage. This is about twentyfold higher than is obtainable by conventional methods in aerated, shaken culture flasks. The actual phage yields are probably much higher than the minimum values calculated from plaque counts. For example, in the case of one of our culture lysates which was purified at King's College, the efficiency of plating was shown to be only 19%. The carbon dioxide evolution rate of cultures was measured and used as a guide to the time at which phage should be added. In this way, greater control of cultural conditions was obtained than is possible in shaken flasks. For the best yield of phage per milliliter of culture, the optimum time for phage infection was such that bacterial lysis just prevented the carbon dioxide evolution rate from reaching its potential maximum. The major factor influencing the phage yield per milliliter of culture was the aeration capacity of the culture vessel used. All had maximum aeration capacities much higher than those obtainable in shaken culture flasks. Cultures grown and infected in 3‐l. Vessel operated under conditions of low aeration gave poor yields of phage. The reason for this are discussed.

Ubiquinone concentrations in athiorhodaceae grown under various environmental conditions.

1. The nature and concentration of ubiquinone in six species of Athiorhodaceae have been examined after growth under aerobic and photosynthetic conditions. 2. Increase in ubiquinone concentration during adaptive synthesis of photosynthetic pigments by Rhodopseudomonas spheroides incubated under low-aeration conditions was observed. 3. The nature of the carbon source was found to have a marked effect on ubiquinone, as well as bacteriochlorophyll, concentrations.

The formation of ribulose 1:5-diphosphate carboxylase by growing cultures of Athiorhodaceae.

Summary: Ribulose l:5-diphosphate carboxylase, which catalyses the conversion of ribulose l:5-diphosphate and CO2 to 3-phosphoglyceric acid, is a key enzyme in the reductive pentose cycle. It was formed only in traces by Rhodopseudomonas spheroides growing in the dark under conditions of high or low aeration but organisms grown photosynthetically were rich in the enzyme. The activity of some other enzymes of the cycle, phosphoriboisomerase, phosphoglyceric acid kinase and triosephosphate dehydrogenase (diphosphopyridine nucleotide-linked) did not vary significantly with the growth conditions. The differential rate of synthesis (increase in enzyme/ increase in culture density) of the carboxylase was studied in cultures of R. spheroides growing exponentially under various conditions. It was influenced by the light intensity, being decreased when this was increased. Oxygen completely repressed formation of the enzyme even under continuous illumination; this effect was annulled by restoration of anaerobic conditions. Dark-grown organisms formed the enzyme at a high differential rate immediately on transfer to anaerobic + light conditions, provided that they contained a minimal amount of bacteriochlorophyll. Biotin deficiency did not affect the synthesis of the carboxylase. The conditions which promoted carboxylase formation were in many respects similar to those favourable to synthesis of enzymes required for the early stages of bacteriochlorophyll formation. The connexion of these observations with the regulation of pigment synthesis and with the possible physiological role of the enzyme in the Athiorhodaceae is discussed.