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Three distinct heterotrophic eubacterial strains were isolated from mixotrophic cultures of the filamentous cyanobacterium, Spirulina maxima (Gom) Geitl. Spirulina spp. are considered to be prime candidates for the phototrophic production of biomass protein, particularly in developing countries. These cyanobacteria are extreme alkaliphiles and halophiles, making their production in arid regions promising. Most previous studies on the eubacteria which live in Spirulina culture systems have focused on determining the possible presence of pathogenic species in biomass protein. Little has been done to understand the symbiotic relationships between the cyanobacterium and its eubacterial cosymbionts. From the perspective of a heterotrophic eubacterium, autotrophic cultural systems of Spirulina have limited carbon and energy resources, being limited to cyanobacterial exudates. In this study, three eubacterial strains were isolated and studied. One strain, a Gram-negative, non-sporing, motile rod, grew exceptionally well in a mineral salts medium where only a small amount of a single low molecular weight organic compound (e.g., acetate) was supplied as sole energy source. This strain was also extremely euryresponsive with respect to salinity and alkalinity as well. Two less well-adapted eubacterial strains are also described.

The literature of the last three years has seen a renaissance in the use of immobilized enzyme systems made practical by the immobilization of the entire cell. Such systems have proven to have a diversity of application unattainable with isolated pure enzymes and the operation of such systems in continuous reactors has proven to be less costly than more conventional processes based on free intact cells. Thus, whole cell immobilization as a technique should be of expanding practical and academic significance in the future.

Four species of Corynebacterium-C. fasciens, C. hydrocarboclastus, C. lepus and C. xerosis, were compared for growth and surfactant production from kerosene. Nitrate as sole nitrogen source resulted in a sharp peak in surfactant concentration during mid-exponential growth phase. Ammonium plus nitrate resulted in growth-associated surfactant production. Surfactant was produced at a concentration of 50-80 times the critical micelle concentration by these species; greatest concentrations were produced by C. lepus. All species were pleomorphic, with greatest cell length occurring in early exponential phase. C. hydrocarboclastus had distinctly longer cells (5 mu) than the other species (3 mu) when grown in shake-flask culture. Ammonium sulfate at concentrations greater than 0.1% (w/v) increased cell length in stationary phase.

Zahlreiche mikrobielle Polysaccharide sind auf ihre Faahigkeiten untersucht worden, Wasser- und OI-Emulsionen zu brechen oder aber zu stabilisieren. Negativ geladene schwach verzweigte o-Phosphonomannane mit einem relativ hohem Phosphorgehalt zeigen gute de-emulgierende Eigenschaften. Mannane stellen schlechte De-Emulgatoren dar, aber einige andere neutrale Polysaccharide erreichen fast so gute Werte wie die besten o-Phosphonomannane. Die untersuchten Polysaccharide konnen nicht als effektive Emulgatoren bezeichnet werden.

Microorganisms capable of growth under highly alkaline conditions have been isolated from natural habitats. The isolation process utilized pH levels of 9.7 to 11.0, salts of 4.0 to 5.0%, temperatures of 45–50°C, and anaerobic conditions. Four isolates are described as gram-positive, spore-forming, motile, catalase-positive rods. This indicates that they are in the genusBacillus. The culture characteristics of these and two other isolates were evaluated. All six strains tolerated up to 11.0% salts in the growth medium. The purpose of this study was to evaluate these microorganisms for their potential use in combination with an alkaline flood for enhanced oil recovery. Useful products in in situ microbial-enhanced oil recovery include gases, surfactants, polymers and acids. The best gas producer wasBacillus strain ACP-1 which evolved an average of 2.11 ml gas per ml medium in 96 h.Bacillus strain ACP-1 reduced the surface tension of the growth medium from 50.3 mN/m to an average of 44.1 mN/m. Another aspect of this research project included investigation of the movement of these microbes through a porous medium. Various synthetic surfactants were found to improve movement through a porous system, while other surfactants improved gas production.

Comparaison de la croissance de 3 souches en cultures pures ou en melange. Application a l'optimisation de la fermentation ethanolique des melasses

Series of synthetic neutral surfactants have been used to break emulsions of heavy oil, water and clay. The degree of de-emulsification was found to correlate with the hydrophilic-lipophilic balance (HLB) of the surfactant. The most effective agents for de-watering had HLB values between either 4 and 6 or 13 and 15. For clay removal, the most useful surfactants had HLB values above 20. Surfactants between 5 and 8 were also effective for removing clay, but the material released had appreciably more oil adhering than that removed by the higher HLB compounds. Other surfactant parameters such as surface tension, interfacial tension against hexadecane and spreading tension were unrelated to de-emulsifying ability. On a employe differents agents tensio-actifs neutres et synthetiques pour briser des emulsions formees d'huile lourde, d'eau et d'argile. On a trouve q'il y avait une correlation entre le degre de rupture de l'emulsion et le rapport hydro-phile-lipophile (HLB) de l'agent tensio-actif. Les agents les plus efficaces pour enlever l'eau avaient des valenrs de HLB comprises soit entre 4 et 6, soit entre 13 et 15. Dans le cas de l'enlevement de l'argile, on a obtenu le plus de succes avec les agents dont les valeurs du HLB excedaient 20; les agents tensio-actifs dont les valeurs du HLB variaient entre 5 et 8 se sont aussi averes efficaces pour enlever l'argile, mais le produit separe contenait bien plus d'huile adherente que dans le cas des agents dont les valeurs du HLB etaient plus elevees. On a trouve que d'autres parametres des agents tensio-actifs, tels que la tension superficielle, la tension interfaciale contre l'hexadecane et la tension d'expansion (spreading), n'avaient aucune relation avec l'aptitude de l'emulsion a se rompre.

This review discusses the currect state of the art on the subject of emulsifiers, particularly those of biological origins. The basic principles involved in the mode of action of emulsifier molecules are reviewed. Current ideas on the classification, physicochemical properties, stability, and rheological properties of emulsions are discussed. The literature review on bio-emulsifiers emphasizes those of microbial origins and their application in industry. Some of the more common methods for the study of emulsion properties are also outlined.

Ethane oxidation was studied in ethane-grown resting cells (mycelia) of an Acremonium sp. and in cell-free preparations of such mycelia. From resting cell experiments evidence was found for a pathway of ethane oxidation via ethanol, acetaldehyde, and acetic acid. In vitro studies indicated that ethane-oxidizing activity in such mycelia occurred predominantly in the microsomal fraction of crude homogenates. Microsomal preparations were inactive in the absence of added coenzyme. Marked stimulation of activity was obtained in such preparations with reduced nicotinamide adenine dinucleotide phosphate and to a much lesser degree with nicotinamide adenine dinucleotide phosphate. Ethane oxidation was inhibited by sodium azide and carbon monoxide.

Thirteen isolates ofZymomonas were analyzed for their ability to tolerate increasing concentrations of glucose and ethanol. In medium containing 5.0% (v/v) ethanol, four isolates grew well in 15.0% (w/v) glucose. Six cultures tolerated at least 6,0% ethanol. Of all the isolates, 7 preferred glucose and 4 preferred sucrose as a sugar substrate. In a nutrient medium containing mineral salts and high concentrations of pantothenate and biotin ethanol production for 2 isolates was approximately 7.0%. Continuous stirring and growth factors were responsible for this increased ethanol production.

A mixture of corynomycolic acids (R1-CH(OH)-CH(R2)-COOH) isolated fromCorynebacterium lepus was shown to have excellent surfactant properties. It caused significant lowering of surface tension in aqueous solution and the interfacial tension between water and hexadecane at all values of pH between 2 and 10. A series of carboxylic acids and some hydroxy-carboxylic acids and alcohols were also studied as a comparison. None of these caused as large a lowering of the surface and interfacial tensions as the corynomycolic acids. The series of carboxylic acids studied showed that surfactant properties depend on the length of the alkyl chain and the pH of the solution in a manner consistent with the hydrophiliclipophilic balance of these compounds. Hydroxyl substituents caused considerable enhancement of the surfactant properties of long chain carboxylic acids if they were located close to the carboxyl function.

Biodegradation of lignosulfonate-NH4 was tested under thermophilic conditions. A fungus, an Aspergillus sp., was cultivated in 2.0 or 3.0% lignosulfonate solutions in tap water at 45°C. Good growth of fungus was observed; however, the degradation of the lignosulfonate was quite limited within the 7 day test period used. In general the amounts of lignosulfonate utilized varied from 10–15% of the extent of biodegradation and the comparative results are reported.

Twelve cultures of Hyphomycetes capable of growth upon the ethane component of natural gas were isolated from raw sewage by a continuous enrichment technique. In addition, all isolates also grew on propane and n-butane but not methane. These cultures are described and their taxonomic position discussed. Four were tentatively placed in Graphium; three were identified as Phialophora jeanselmei (Langeron) Emmons; the remaining five are similar to Acremonium sensu Gams.

A variety of aeration and agitation conditions were used to investigate the oxygen demand of Erwinia aroideae for growth and L-asparaginase production in a 14-liter fermenter. Maximum L-asparaginase activity of 4.25 IU/ml was obtained when the fermentation medium was aerated at 0.5 volumes air per volume medium per minute and agitated at 500 rpm. However, the specific L-asparaginase activity was not affected significantly by the rate of oxygen transfer. In most instances the level of dissolved oxygen decreased to zero before harvesting. KLa rather than dissolved oxygen as measured during the fermentation cycle is a more satisfactory index to determine the oxygen requirement for cell growth and asparaginase production. The optimum level of KLa for L-asparaginase production was 0.98 min−1. However, a higher value of KLa of 1.2–1.9 min−1 was needed to achieve the highest growth.

The intracellular enzyme, L-asparaginase, from aerobically grown Erwinia aroideae NRRL B-138 has been purified and some of its properties studied. Sonic treatment permitted recovery of 95% of L-asparaginase from cells. The crude cell lysate was purified 167-fold by means of ammonium sulfate fractionation and column chromatography on hydroxylapatite–cellulose, and DEAE–Sephadex. The specific activity of the most active fraction of L-asparaginase is 256 IU/mg protein. The enzyme has a broad pH activity profile with maximum at pH 9.0–9.5. The optimum temperature for enzyme reaction was determined to be 41 °C. The apparent activation energy is 11 000 cal/mole. The molecular weight of L-asparaginase was estimated by gel filtration to be 108 000.

Isonicotinic acid hydrazide (INH) inhibits the growth of Corynebacterium lepus on hexadecane but has no effect on its growth on fructose. INH also inhibits the production of the mycolic acid containing lipopeptide bioemulsifier normally produced by C. lepus in response to an insoluble substrate. The primary effect of INH appears to be inhibition of mycolic acid synthesis, which limits the growth of C. lepus on hexadecane by reducing the concentration of bioemulsifier.

The hydrocarbon metabolizing Acinetobacter calcoaceticus sp. 2CA2 reduces the surface tension of the culture broth during growth on liquid hydrocarbons. This activity, which is not evident during growth on soluble substrates, is associated with the whole cells. Removing the cells from the culture broth increases the surface tension of the liquid phase. The cells when resuspended in water result in a dramatic lowering of the surface tension. Acinetobacter sp. 2CA2 tends to partition between the two liquid phases during growth on hydrocarbons. Both the hydrocarbon bound and nonadhering cells are equally surface active. The whole cells are also able to form and stabilize kerosene-water emulsions. This ability is not related to the lowering of the liquid surface or interfacial tension, since both surface active and nonsurface active cells demonstrated the same emulsifying properties. An extracellular lipopeptide produced during growth on hydrocarbons is not surface active but effectively forms and stabilizes kerosene-water emulsions. The cells and extracellular lipoptide are also effective in de-emulsifying surfactant stabilized test emulsions. The lipopeptide product reduced the half-life of a Tween-Span (TS) stabilized kerosene-water emulsion from 650 to 0.4 h at product concentrations of less than 1% (w/v).

This literature survey contains information pertaining to bacterial aerosol viability and electrostatic surface charges of bacteria. The variables involved in the survival of bacterial aerosols include the method of droplet formation and the intrinsic characteristics of the gas in which the bacteria-containing droplets are suspended. The survey contains information on overall microbial aerosol studies, assay techniques, cell ions, the effect of air composition, the cell surface, radiation effects on aerosols, and the action and the possible components of relative humidity involved in the viability of bacterial aerosols. A small portion of the investigation deals with the survival of mycoplasmas, L-forms, and algae in air.

The ability of Candida albicans and Candida spp. to adhere to inert polymeric surfaces may allow these organisms direct ingress into the human host. Biophysical characterization of this adherence shows that the forces responsible for such adherence are attractive London-van der Waals forces (or hydrophobic forces) and electrostatic forces. The hydrophobic affinity of yeasts was determined by (i) a water-hydrocarbon two-phase assay and by (ii) measurement of the contact angle (theta) of a liquid droplet on a monolayer of yeast cells. The hydrophobicity of the yeasts correlated with the tendency of yeasts to adhere to polystyrene and was reduced in the presence of Tween 20. The adherence of yeasts to polymers of increasing hydrophobicity (determined by the contact angle method) was directly proportional to theta. Yeast surface charges were altered by selectively blocking amino and carboxyl groups. The more positively charged yeasts adhered in greater numbers. Increasing the molarity of NaCl increased yeast adherence. These forces probably contribute to the negative cooperativity (determined by Scatchard and Hill plot) that characterizes the adherence of yeasts to polymers.

Analysis of gases evolved when Citrobacter intermedius and a Pseudomonas species are co-cultured on an acetate or glucose – nitrate – mineral salts medium indicates that acetate is anaerobically oxidized to CO2.

A fungus which grows well on a mineral salts solution with natural gas as the carbon source is described and provisionally identified as a Graphium species. Its taxonomic relation to several genera is presented. This organism was isolated from sewage after selection by enrichment techniques and continuous culture. The fermentor was operated at ambient temperatures, 28 °C ± 2, at a volume of 10 liters with a dilution rate of 10 liters/4 days to 10 liters/1.7 days. Coty's mineral salts medium gave the highest tissue yield. When the pH of the incoming mineral salts medium was decreased stepwise from 7.0 to 5.0 the pH of the reactor became self-adjusting, varying from around 2.7 to 3.5, and the dry weight of microbial tissue obtained varied from 65 to 275 mg/h. Also present in the continuous culture was an acid tolerant bacterium, which, when isolated, grew well on natural gas, methanol, and ethanol, and a strain of Trichoderma, which, when isolated, did not use natural gas as a carbon source. In mixed culture the Trichoderma is thought to grow on metabolites produced by either or both the Graphium and the acid-tolerant bacterium during oxidation of natural gas. The nature of the relationship is being investigated. The mixed culture has been under continuous cultivation for 18 months.

Classically all algae form their cellular carbon solely from carbon dioxide by photosynthesis. However, some are facultative heterotrophs and are able to utilize organic substrates as a source of carbon. Also there are obligate heterotrophic algae which must obtain at least some organic compounds from their surroundings. For example, the colorless alga Prototheca zopfii, which does not have the ability to photosynthesize, is unable to grow in the absence of organic materials (Barker, 1935). This alga utilizes ammonia, nitrogen from yeast autolyzate as well as glucose. It is unable to grow in the complete absence of yeast autolyzate.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . 89 1. Microorganisms . . . . . . . . . . . . . . . . . . . . . . . . 9 0 a) Historical Developments . . . . . . . . . . . . . . . . . . . 90 b) Classification of Methane Oxidizing Bacteria . . . . . . . . . . 96 2. Liquid Media . . . . . . . . . . . . . . . . . . . . . . . . . 101 pH and Temperature . . . . . . . . . . . . . . . . . . . . . . 102 3. Composit ion of the Gas Phase . . . . . . . . . . . . . . . . . . 103 4. Oxygen and Methane Requirements . . . . . . . . . . . . . . . . 103 5. Biochemistry of Methane Oxidation . . . . . . . . . . . . . . . . 108 a) Autotrophy-Heterotrophy . . . . . . . . . . . . . . . . . . . 108 b) Methane Oxidation Pathways . . . . . . . . . . . . . . . . . 109 c) Nitrogen Metabolism and Denitrification in Methaneand Methanol Oxidizers . . . . . . . . . . . . . . . . . . . . . . . . . . 110 d) Sulfate Reduction in Relation to Methane Oxidation . . . . . . . 112 6. Biosynthesis of Cell Components . . . . . . . . . . . . . . . . . 1 t3 7. Present and Possible Future Applications . . . . . . . . . . . . . 115 a) Single-Cell Protein Production . . . . . . . . . . . , . . . . . . 115 Quality of the SCP Derivcd from ttydrocarbons . . . . . . . . . 120 b) Removal of Methane from ('otll Nliiacs . . . . . . . . . . . . . . 12t c) Petroleum Prospecting . . . . . . . . . . . . . . . . . . . . 1 2 1 d) Microbial Fuel-Cell . . . . . . . . . . . . . . . . . . . . . . 122 References . . . . . . . . . . . . . . . . . . . . . . . . . . . . 122