Anaerobic digestion of cellulose by pure and mixed bacterial cultures

Summary By enrichment technique, nine anaerobic mixed bacterial cultures were isolated, five of which showed stable cellulolysis. All cultures fermented cellulose and produced different fermentative products. Mixed culture BOC 25 yielded major levels of acetate and ethanol (39.6 and 12.0 mmol/l, respectively) and minor levels of propionate (2.5 mmol/l) and digested filter paper cellulose to the extent of 32.5% w/v. BOC 25 digested cellulosic and lignocellulosic substrates and produced filter paper cellulase, carboxymethyl cellulase, Avicelase and -glucosidase. Strain DC 25, a cellulolyticClostridium was purified from one of the mixed cultures. The fermentation products of DC 25 from cellulose, cellobiose or glucose were ethanol, acetate, formate, H₂ and CO₂.     

Utilization of dimeric lignin model compounds by mixed bacterial cultures

Summary The degradation of dimeric phenylpropanoid lignin model compounds using mixed bacterial cultures was studied. The six model compounds contained the most common linkages of lignin: -O-4, -, -5, and -1. The results indicate that it is possible to enrich bacteria which are able to degrade all these compounds. Bacteria were also able to use these dimers as the sole source of carbon for growth. In view of these results it seems probable that bacterial inability to degrade polymeric lignin is due to the physical properties such as the molecular size of lignin.     

Alkaline treatment of wheat straw for increasing anaerobic biodegradability

Wheat straw was treated with NaOH and anaerobically digested for methane production. Alkaline treatment resulted in a greater than 100% increase in biodegradability of wheat straw. The potential of a process flow scheme employing high alkali concentration at ambient temperature with solids separation and recycle of filtrate containing residual alkali was explored. The effect of NaOH on the solubilization of cell wall constituents and potential problems of toxicity are discussed. A solubilization model was developed which is used to predict biodegradability of whole samples based on solids and filtrate biodegradabilities. Energy requirements and chemical costs are also addressed.     

Biodegradation potential of oily sludge by pure and mixed bacterial cultures

The biodegradation capacity of aliphatic and aromatic hydrocarbons of petrochemical oily sludge in liquid medium by a bacterial consortium and five pure bacterial cultures was analyzed. Three bacteria isolated from petrochemical oily sludge, identified as Stenotrophomonas acidaminiphila, Bacillus megaterium and Bacillus cibi, and two bacteria isolated from a soil contaminated by petrochemical waste, identified as Pseudomonas aeruginosa and Bacillus cereus demonstrated efficiency in oily sludge degradation when cultivated during 40 days. The bacterial consortium demonstrated an excellent oily sludge degradation capacity, reducing 90.7% of the aliphatic fraction and 51.8% of the aromatic fraction, as well as biosurfactant production capacity, achieving 39.4% reduction of surface tension of the culture medium and an emulsifying activity of 55.1%. The results indicated that the bacterial consortium has potential to be applied in bioremediation of petrochemical oily sludge contaminated environments, favoring the reduction of environmental passives and increasing industrial productivity.     

Mineralization of the s-triazine ring of atrazine by stable bacterial mixed cultures.

Enrichment cultures containing atrazine (2-chloro-4-ethylamino-6-isopropylamino-1,3,5-triazine) at a concentration of 100 ppm (0.46 mM) as a sole nitrogen source were obtained from soils exposed to repeated spills of atrazine, alachlor, and metolachlor. Bacterial growth occurred concomitantly with formation of metabolites from atrazine and subsequent biosynthesis of protein. When ring-labeled [14C]atrazine was used, 80% or more of the s-triazine ring carbon atoms were liberated as 14CO2. Hydroxyatrazine may be an intermediate in the atrazine mineralization pathway. More than 200 pure cultures isolated from the enrichment cultures failed to utilize atrazine as a nitrogen source. Mixing pure cultures restored atrazine-mineralizing activity. Repeated transfer of the mixed cultures led to increased rates of atrazine metabolism. The rate of atrazine degradation, even at the elevated concentrations used, far exceeded the rates previously reported in soils, waters, and mixed and pure cultures of bacteria.     

Properties and roles of bacterial symbionts of polyvinyl alcohol-utilizing mixed cultures

From several polyvinyl alcohol (PVA)-utilizing mixed cultures, two component bacterial strains essential for PVA utilization were isolated, and their properties and roles in PVA utilization were studied. Each pair of essential component strains consisted of a type I strain, which produced a PVA-degrading enzyme and constituted the predominant population of the mixed culture in PVA, and a type II strain, which produced a certain growth stimulant for the former strain. All of the type I strains were taxonomically identical and assigned as Pseudomonas sp. In contrast, type II strains were taxonomically different from each other, belonging to Pseudomonas spp. and Alcaligenes sp. PVA utilization occurred in each mixed culture of a type I strain with Pseudomonas putida VM15A as a substitute for the type II strain of the original pair and also in each mixed culture of a type II strain with Pseudomonas sp. VM15C. The growth rates of these substituted, mixed cultures differed from each other.     

A study of mixed continuous cultures of sulfate-reducing and methane-producing bacteria

Ecological relationships between sulfate-reducing and methane-producing bacteria in mud of Lake Vechten have been studied by continuous culture studies using the chemostat technique. The maximum specific growth rate (μ _max) and saturation constant (K _s) were, respectively, 0.36 hr⁻¹ and 0.047 mM for lactate-limited growth ofDesulfovibrio desulfuricans and 0,011 hr⁻¹ and 0.17 mM for acetate-limited growth ofMethanobacterium sp. Calculated values for the true molar growth yieldsY _G) and maintenance coefficients (m) were 30.6 g bacterial mass/mole of lactate and 0.53 g substrate/g dry wt hr forD. desulfuricans and 37.8 g bacterial mass/mole of acetate and 0.54 g substrate/g dry wt hr forMethanobacterium. No growth ofMethanobacterium was observed at apS²⁻ value (the hydrogen sulfide potential) of more than 11 and there was no effect on the growth atpS²⁻ values above 13. In mixed continuous culture experiments the concentration of acetate decreased in the secondstage growth vessel, whereas that of methane increased stoichiometrically. If the substrate concentration in the reservoirs (S _r) was increased from 0.1 to 0.5 mg/ml, the population ofDesulfovibrio increased and that ofMethanobacterium was washed out of the culture vessel, since the concentration of hydrogen sulfide reached apS²⁻ value of 10.5. From the mixed continuous culture experiments a commensalism between the two species can be described, i.e., the acetate-fermentingMethanobacterium benefits from the acetate released byDesulfovibrio which is, in turn, not affected in the presence of the former.     

Comparison of methods for measurement of bacterial growth rates in mixed batch cultures.

Eight methods of assessing growth rate constants of bacteria were compared in batch cultures of 3-micrometers-filtered estuarine water from the Skidaway River in Ga. Mixed assemblages of bacteria were grown under four nutrient regimes of added yeast extract ranging from 0 to 100 mg/liter. Linear and exponential growth rate constants were computed from changes in cell densities, biovolumes, and ATP concentrations. Exponential growth rate constants were obtained from the frequency of dividing cells and RNA synthesis as measured by [3H]adenine uptake. Rate constants obtained during lag, exponential, and stationary growth phases depended largely on the method used. Constants calculated from changes in cell densities, frequency of dividing cells, and adenine uptake correlated most closely with each other, whereas constants calculated from changes in ATP concentrations and biovolumes correlated best with each other. Estimates of in situ bacterial productivity and growth vary depending on the method used and the assumptions made regarding the growth state of bacteria.     

Neuronal cell cultures: A tool for investigations in developmental neurobiology

The aim of this review is to describe environmental requirements for survival of neuronal cells in culture, and secondly to survey the complex interplay between hormones, neurotrophic factors, transport- and extracellular matrix- proteins, which characterize the developmental program of differentiating neurons. An overall reconsideration of the literature in this vast field is above the limits of the present paper; since progress and refinement in the techniques of neuronal cell cultures have paralleled the advancement in Developmental Neurobiology, we will run instead through the main steps which form the conceptual framework of neuronal cell cultures.     

Improved method for determination of ammonia and nitrite oxidation activities in mixed bacterial cultures

A simple and reliable method to measure the activity of ammonia and nitrite oxidisers in mixed bacterial cultures was developed. The developed method differentiates between the ammonia and nitrite oxidisers by consecutive injection of NO₂^– and NH₄⁺. The main advantage of this method is that it avoids the use of metabolic inhibitors for ammonia or nitrite oxidisers, as used by other methods. Moreover, it allows measuring of the short-term effect of an inhibitor on both the ammonia and nitrite oxidisers in one test under controlled environmental conditions (pH, temperature). The developed method was applied to determine the inhibitory effects of salt (NaCl up to 15 g Cl/l) on an enriched culture of nitrifying bacteria. The results of the method demonstrate its potential to accurately determine the individual activities of nitrite and ammonia oxidisers.     

Transformation of steroids by mixed cultures

Some results of our studios on transformation of steroids by mixed culture fermentation are presented in this paper. Arthrobacter simplex was paired in turn with each of the following: Streptomyces roseochromogenes, Curvularia lunata, Absidia coerulea, and Aspergillus ochraceus. The steroid substrates examined for multiple transformation were 16α-hydroxy-cortexolone, 16α-hydroxy-cortexolone 16,17-acetonide, 9α-fluorohydrocortisone, 9α-fluorohydrocortisone 21-acetate, and 9α-fluorohydrocortisone 21-hemisuccinate. The effects of media, steroid substrate, and microbial interaction in a mixed culture on the induction and repression of steroid transforming enzymes were unique to each case studied. The reaction mechanism of the multiple steroid transformation was also found to vary from one mixed culture system to another. Two different reaction mechanisms were observed, namely, consecutive and parallel. In the former, one of the two enzymatic reactions always preceded the other, while in the latter, two different enzyme reactions occurred simultaneously, thereby giving rise to two different intermediates. Multiple transformation of steroids by a single step mixed culture fermentation has potential economic advantages.     

Mass spectrometry: A tool for on-line monitoring of animal cell cultures

The magnetic sector mass spectrometer is able to detect oxygen uptake and carbon dioxide production rates from animal cell cultivations performed in 101 biorectors. Such data have not been available with the use of classic exhaust gas analysis applying paramagnetic analyzers and infra-red sensors due to the insensitivity of the apparatus available. In the course of the present work we were able to demonstrate, that the oxygen uptake rate correlates to the number of viable cells. Additionally oxygen uptake rates supplied on-line information about the actual physiology of the cells: When the rates changed during the cultivation process, this immediately indicated the occurrence of limitations of components in the medium. The information could be useful in timing key events, such as performing splits or harvesting the bioreactor.Abbreviations OUR oxygen uptake rate - CDPR carbon dioxide production rate - RQ respiratory quotient This publication is dedicated to the 65 ^th birthday of Prof. Dr. F. Wagner, University of Braunschweig.     

Sulfolane degradation by mixed cultures and a bacterial isolate identified as a Variovorax sp.

Sulfolane (tetrahydrothiophene-1,1-dioxide) is used in the Sulfinol process for natural gas sweetening. At many sour-gas processing plants spills, landfills and leakage from unlined surface storage ponds have contaminated groundwaters with sulfolane. Due to its high water solubility and mobility in aquifers, sulfolane poses a risk for off-site contamination. This study investigated the aerobic biodegradation of sulfolane by two mixed microbial enrichment cultures and by three bacterial isolates. Sulfolane served as the sole C, S and energy source for these cultures. In the two mixed cultures, 60% and 80% of the sulfolane C was recovered as CO2, whereas in cultures of the three isolates only 40-42% of the substrate C was recovered as CO,. In the mixed cultures, 81% and 97% of the sulfolane S was converted to sulfate, and in the pure isolates, 55-90% of the substrate S was converted to sulfate. Thus, the mixed cultures were capable of greater mineralization than the pure isolates. One isolate, strain WP1, was identified using a combination of 16S rRNA gene sequencing, physiological traits and cell morphology. WP1 was determined to be most similar to Varioivorax paradoxus.     

Production of polyhydroxyalkanoates by mixed microbial cultures

Polyhydroxyalkanoates (PHAs) are biodegradable bioplastics formed from renewable resources, like sugars, with similar characteristics of polypropylene. These bioplastics are industrially produced by pure cultures using expensive pure substrates. These factors lead to a much higher selling price of PHAs compared to petroleum-based plastics, like polypropylene. The use of mixed cultures and cheap substrates (waste materials) can reduce costs of PHA production by more than 50%. Storage of PHAs by mixed populations occurs under transient conditions mainly caused by discontinuous feeding and variation in the electron donor/acceptor presence. In the last years the mechanisms of storage, metabolism and kinetics of mixed cultures have been studied. The maximum capacity of PHA storage and production rate is dependent on the substrate and on the operating conditions used. In this paper an overview and discussion of various mechanisms and processes for PHA production by mixed cultures is presented.