Sporulation

The Provesteen process, developed by Phillips Petroleum Company, employs a proprietary 25,000-L continuous fermentor for producing Hansenu/a jejunii the sporulating form of C. utilis from glucose or sucrose at high cell concentrations up to 150 g/L. The fermentor is designed to provide optimum oxygen and heat transfer (69,70). 

Some mycehal fermentations exhibit early sporulation, breakup of mycehum, and low yields if the shear is excessive. A tip speed or 250 to 500 cm/s (8 to 16 ft/s) is considered permissible. Mixing time has been proposed as a scale-up consideration, but httle can be done to improve it in a large fermenter because gigantic motors would be required to get rapid mixing. Culturing cells from plants or animals is beset by mixing problems because these cell are easily damaged by shear. 

During the stationary phase, the growth rate is zero as a result of the depletion of nutrients and essential metabolites. Several important fermentation produets (ineluding most antibioties) are produeed in the stationary phase. The stationary phase is followed by a phase where eells die or sporulate. During the death phase, there is a deerease in live eell eoneentration, whieh results from the toxie byproduets eoupled with the depletion of the nutrient. The number of viable eells usually follows an exponential deeay eurve during this period. 

The slants were incubated for 7 days at 30°C, after which time sporulation was complete. The spores from the agar slants were used, in an aqueous suspension, to inoculate 100 ml of preseed medium in a 500 ml Erlenmeyer flask. The sterile preseed medium consisted of ... 

Sterile agar slants are prepared using the Streptomyces sporulation medium of Hickey and Tresner, J. Bact., vol. 64, pages 891-892 (1952). Four of these slants are inoculated with lyophilized spores of Streptomyces antibioticus NRRL 3238, incubated at 28°C for 7 days or until aerial spore growth is well-advanced, and then stored at 5°C. The spores from the four slants are suspended in 40 ml of 0.1% sterile sodium heptadecyl sulfate solution. A nutrient medium having the following composition is then prepared 2.0% glucose monohydrate 1.0% soybean meal, solvent extracted, 44% protein 0.5% animal peptone (Wilson s protopeptone 159) 0.2% ammonium chloride 0.5% sodium chloride 0.25% calcium carbonate and water to make 100%. 

The secretion of the polymer into the growth medium is not only a means to control nuclear /3-poly(L-malate) concentration but also is a prerequisite for spor-ulation. L-Malate, which is a co-inductor of sporulation (Renzel and Hildebrandt, personal communication), is obtained by spontaneous and polymalatase-catalyzed hydrolysis of the secreted polymer. 

Bacillus thuringiensis produces a variety of organic compounds which are toxic to the larvae of certain susceptible insect hosts. Among the toxic entities are proteinaceous crystals, probably three soluble toxins, and certain enzymes. The protein material is the major toxin active in killing lepidopterous larvae. The protein is formed by the cells apparently in close synchrony with sporulation, and its nature is a constant function of bacterial strain. The mode of action of the protein is under study. The sequence of events in the pathology observed is probably solubilization of the crystal (enzymatic or physical)—>liberation of toxic unit—>alteration of permeability of larval gut wall— change in hemolymph pH—>invasion of hemolymph by spores or vegetative cells of the bacterium. 

Microbial insecticides are very complex materials in their final formulation, because they are produced by fermentation of a variety of natural products. For growth, the bacteria must be provided with a source of carbon, nitrogen, and mineral salts. Sufficient nutrient is provided to take the strain of choice through its life cycle to complete sporulation with concomitant parasporal body formation. Certain crystalliferous bacilli require sources of preformed vitamins and/or amino acids for growth. Media for growing these bacilli may vary from completely soluble, defined formulations, usable for bench scale work, to rich media containing insoluble constituents for production situations (10,27). Complex natural materials such as cottonseed, soybean, and fish meal are commonly used. In fact, one such commercial production method (25) is based on use of a semisolid medium, a bran, which becomes part of the final product. 

The bacterial culture converts a portion of the supplied nutrient into vegetative cells, spores, crystalline protein toxin, soluble toxins, exoenzymes, and metabolic excretion products by the time of complete sporulation of the population. Although synchronous growth is not necessary, nearly simultaneous sporulation of the entire population is desired in order to obtain a uniform product. Depending on the manner of recovery of active material for the product, it will contain the insolubles including bacterial spores, crystals, cellular debris, and residual medium ingredients plus any soluble materials which may be carried with the fluid constituents. Diluents, vehicles, stickers, and chemical protectants, as the individual formulation procedure may dictate, are then added to the harvested fermentation products. The materials are used experimentally and commercially as dusts, wettable powders, and sprayable liquid formulations. Thus, a... 

The toxin kills larvae and/or pupae of some Diptera and Lepidoptera and acts primarily by prevention of completion of pupation. It is produced in cultures prior to sporulation (9,26) and remains in the supernatant liquid of sporulated cultures. Its chemical structure is not known, but initial isolation and purification studies are under way (9). One cannot, however, leave a discussion of this toxin with a feeling of certainty. Burgerjon and deBarjac (7) and Krieg and Herfs (20) reported the above-mentioned effectiveness of the soluble... 

The exotoxin reported by Smirnoff (31) is definitively different from other soluble toxins, as indicated by its reported heat lability. This soluble toxin was obtained from the supernatant of a sporulated B. thuringiensis culture. In testing, it was found to be very toxic by ingestion to 18 species of larch sawfly larvae. No further studies on this toxin have been reported at this time. 

BAL—see Propan-l-ol, 2,3-dimercapto-Bacillus cereus calcium transport, 6,572 Bacillus megaterium calcium efflux, 6, 570 calcium transport, 6,572 sporulation zinc transport, 6, 572 Bacillus spp. sodium ions, 6, 559 sporulation... 

Zur Transformation organischer Verbindungen konnen auch Pilzsporen verwendet werden. Sie werden durch Sporulation aus den Klassen der Ascomyceten, Deuteromyce-ten (Fungii imperfecti) und Phycomyceten (Mmorales) gewonnen2-5. 

Cell Death. Spontaneous death or sporulation of cells is commonly modeled as a first-order process. Equation (12.8) (or 12.12)) is modified to include a disappearance term ... 

The reason for heat resistance is thought to lie in the fact that the core or spore cytoplasm becomes dehydrated during sporulation. The mechanism for this dehydration... 

Approximate time (hours) following commencement of sporulation... 

Fig. 2.6 The moqjhological events of sporulation in Saccharomyces cerevisiae. (a) starved cell V, vacuole LG, lipid granule ER, endoplasmic reticulum CW, cell wall M, mitochondrion S, spindle pole SM, spindle microtubules N, nucleus NO, nucleolus, (b) Synaptonemal complex (SX) and development of polycomplex body (PB) along with division of spindle pole body in (c). (d) First meiotic division which is completed in (e). (f) Prepararation for meiosis II. (g) Enlargement of prospore wall, culminating in enclosure of separate haploid nuclei (h). (i) Spore coat (SC) materials produced and deposited, giving rise to the distinct outer spore coat (OSC) seen in the completed spores of the mature ascus (j). Reproduced from the review by Dickinson (1988) with permission from Blackwell Science Ltd.

Mycobacteria are more resistant than other non-sporulating bacteria to a wide range of biocides. Examples of such organisms axe Mycobacterium tuberculosis, theM avium-intracellulare (MAI) group andM. chelonae (M. chelonei). Of the bacteria, however, the most resistant of all to biocides are bacterial spores, e.g. Bacillus subtilis, B. cereus. 

In additional tests, the suppressor activities of pectins in the intact host/pathogen-interaction were demonstrated. Injection of suppressors rendered genetically highly resistant plants more susceptible, i. e., we observed increased growth of the fungus, and in some cases sporulation occured. Obviously, oligogalacturonides do not only have an... 

The basal medium of Mandels (Mandels et al., 1976) was used with the following modifications it was buffered with 3 g/1 of sodium nitrate to pH 5.5 and supplemented with 1% w/v citrus pectin " Sigma" or other carbon sources. For enzyme production, 50 ml medium in 250 ml erlemneyer flasks were inoculatedwith spores (10 spores /ml ) exept for the non sporulating Pol 6 strain, where mycelium was used. The culture were incubated at 30° C on a rotary shaker (150 rev mn -1) for 5 days. The culture broth was filtered (Millipore 0.45 pm ) and the supernatant was analysed for pectinolytic activities, reducing sugars and proteins. 

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