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S N Agathos

Publications and source records attributed to S N Agathos.

At least 19 recordsLinked to original sources
PubMed2006

Metabolic design of macroscopic bioreaction models: application to Chinese hamster ovary cells.

The aim of this paper is to present a systematic methodology to design macroscopic bioreaction models for cell cultures based upon metabolic networks. The cell culture is seen as a succession of phases. During each phase, a metabolic network represents the set of reactions occurring in the cell. Then, through the use of the elementary flux modes, these metabolic networks are used to derive macroscopic bioreactions linking the extracellular substrates and products. On this basis, as many separate models are obtained as there are phases. Then, a complete model is obtained by smoothly switching from model to model. This is illustrated with batch cultures of Chinese hamster ovary cells.

Animals↗
PubMed2004

Environmental genomics: exploring the unmined richness of microbes to degrade xenobiotics.

Increasing pollution of water and soils by xenobiotic compounds has led in the last few decades to an acute need for understanding the impact of toxic compounds on microbial populations, the catabolic degradation pathways of xenobiotics and the set-up and improvement of bioremediation processes. Recent advances in molecular techniques, including high-throughput approaches such as microarrays and metagenomics, have opened up new perspectives and pointed towards new opportunities in pollution abatement and environmental management. Compared with traditional molecular techniques dependent on the isolation of pure cultures in the laboratory, microarrays and metagenomics allow specific environmental questions to be answered by exploring and using the phenomenal resources of uncultivable and uncharacterized micro-organisms. This paper reviews the current potential of microarrays and metagenomics to investigate the genetic diversity of environmentally relevant micro-organisms and identify new functional genes involved in the catabolism of xenobiotics.

Bacteria↗
PubMed2004

Origin of rice protein hydrolysates added to protein-free media alters secretion and extracellular proteolysis of recombinant interferon-gamma as well as CHO-320 cell growth.

CHO-320 cells, cultivated in suspension in a protein-free medium supplemented with rice protein hydrolysates (peptones), secrete recombinant interferon-gamma (IFN-gamma) that undergo will or will not proteolysis, depending on the origin of the peptones. This proteolytic event, as well as the appearance of an unidentified 70 kDa gelatinase-like protease, are attributed to a cysteine protease. Casein zymographies revealed that one rice protein hydrolysate, but not another, contains a papain-like cysteine protease whose activity is undetectable in solution. This work underlines the significance of the origin of peptones when considered as supplements in serum- and protein-free media for overproduction of recombinant proteins.

Animals↗
PubMed2003

Insect cell culture for industrial production of recombinant proteins.

Insect cells used in conjunction with the baculovirus expression vector system (BEVS) are gaining ground rapidly as a platform for recombinant protein production. Insect cells present several comparative advantages to mammalian cells, such as ease of culture, higher tolerance to osmolality and by-product concentration and higher expression levels when infected with a recombinant baculovirus. Here we review some of the recent developments in protein expression by insect cells and their potential application in large-scale culture. Our current knowledge of insect cell metabolism is summarised and emphasis is placed on elements useful in the rational design of serum-free media. The culture of insect cells in the absence of serum is reaching maturity, and promising serum substitutes (hydrolysates, new growth and production-enhancing factors) are being evaluated. Proteolysis is a problem of the BEVS system due to its lytic nature, and can, therefore, be a critical issue in insect cell bioprocessing. Several cell- or baculovirus proteases are involved in degradation events during protein production by insect cells. Methods for proteolysis control, the optimal inhibitors and culture and storage conditions which affect proteolysis are discussed. Finally, engineering issues related to high-density culture (new bioreactor types, gas exchange, feeding strategies) are addressed in view of their relevance to large-scale culture.

Animals↗
PubMed2002

Use of the pAL5000 replicon in PAH-degrading mycobacteria: application for strain labelling and promoter probing.

Three environmental Mycobacterium strains (LB501T, LB307T and VM552) able to degrade anthracene, phenanthrene or pyrene, respectively, were successfully electroporated with pAL5000-based plasmids containing the green fluorescent protein (gfp) gene of Aequoria victoria under the control of the hsp60 promoter of Mycobacterium bovis following a slightly modified standard procedure. Transformants showed irregular gfp expression profiles. Four plasmid derivatives were constructed that contained gene promoters isolated from, and adapted to, gene expression in polycyclic aromatic hydrocarbon (PAH)-degrading mycobacteria. One derivative directed strong and homogeneous expression of GFP, allowing dual analysis of both GFP- and PAH-derived fluorescence as assessed by confocal laser scanning microscopy. The results reported here demonstrate the suitability of the pAL5000 replicon for the development of recombinant DNA-based studies in PAH-degrading Mycobacterium spp.

DNA, Bacterial↗
PubMed2002

Influence of electron donors and acceptors on the bioremediation of soil contaminated with trichloroethene and nickel: laboratory- and pilot-scale study.

Laboratory- and pilot-scale studies were conducted in order to adjust and optimize the in-situ conditions for bioremediation of a soil contaminated with trichlororethene (TCE) and nickel. Results from laboratory studies showed that the indigenous microorganisms of the soil were limited by the type of electron donor. A better TCE dechlorination was obtained when the electron donor was composed of a mixture of methanol and lactate, as compared to that with methanol alone. Addition of up to 10 mM of sulphate as external electron acceptor (in combination with TCE) and with a mixture of methanol and lactate as electron donor had no significant effect on the TCE reducing activity of indigenous microorganisms of the soil, while higher concentrations (15 and 20 mM) yielded a lower dechlorination. Long term operation of a large pilot-scale soil reactor demonstrated the feasibility of a single-process in situ soil remediation. Results showed that, on one hand, TCE was progressively and stepwise reduced to cis-dichloroethene (DCE), vinyl chloride (VC) and finally to ethene, using only the indigenous microorganisms of the soil. On the other hand, stimulating the activity of sulphate-reducing bacteria of the soil with the addition of sulphate as electron acceptor was efficient in precipitating nickel as nickel sulphide.

Bacteria, Anaerobic↗
PubMed2001

Implication of manganese (III), oxalate, and oxygen in the degradation of nitroaromatic compounds by manganese peroxidase (MnP).

The fungal ligninolytic enzyme manganese peroxidase (MnP) is known to function by oxidizing Mn(II) to Mn(III), a powerful oxidant. In this work, an abiotic system consisting of Mn(III) in oxalate buffer under aerobic conditions (Mn(III)/oxalate/O2 system) was shown to be capable of extensively transforming 2-amino-4,6-dinitrotoluene (2A46DNT)--one of the main reduction products of 2,4,6-trinitrotoluene (TNT). No significant transformation occurred in the presence of other organic acids or under anaerobic conditions. The Mn(III)/oxalate/O2 system was also able to transform other nitroaromatic compounds such as 2-nitrotoluene, 4-nitrotoluene, 2,4-dinitrotoluene, TNT - the latter to a lesser extent -, and their reduction derivatives. The Mn(III)/oxalate/O2 system mineralized 14C-U-ring labeled 2A46DNT slightly, while no significant mineralization of 14C-U-ring labeled TNT was observed. Unidentified 14C-transformation products were highly polar. Electron spin resonance experiments performed on the Mn(III)/oxalate/O2 system revealed the generation of formyl free radicals (*COO-). The oxygen requirement for the transformation of nitroaromatic compounds suggests the involvement of superoxide free radicals (O2-*). produced through autoxidation of *COO- by molecular oxygen. The implication of such a Mn(III)/oxalate/O2 system in the MnP-catalyzed degradation of nitroaromatic pollutants by white-rot fungi is further discussed.

Agaricales↗
PubMed2001

A PCR test to identify bacillus subtilis and closely related species and its application to the monitoring of wastewater biotreatment.

A PCR test based on the 16S rRNA gene was set up that could identify any of the five species of the 'Bacillus subtilis group' (B. subtilis, B. pumilus, B. atrophaeus, B. lichenijormis and B. amyloliquefaciens). The test was directly applicable to single colonies and showed excellent specificity. In the mixed population context of wastewater analysis, direct detection of the target Bacillus species by PCR on either crude or purified DNA extracts had poor sensitivity. When assayed on cell suspensions derived from enriched wastewater samples, sensitivity was increased. Using a simple calibration method, it was possible to estimate the proportion of the target organisms. This method was found suitable for easy monitoring of a wastewater bioaugmentation experiment carried out with a mixture of sporulated Bacillus strains.

Bacillus subtilis↗
PubMed2001

Abiotic transformation of catechol and 1-naphthol in aqueous solution-influence of environmental factors.

The abiotic transformation of catechol and 1-naphthol singly and in mixtures was tested in sterile Tris-HCl buffer with regard to several environmental factors including temperature (7 degrees C, 20 degrees C and 30 degrees C), lighting conditions, pH (between 7.0 and 8.5) and dissolved oxygen (at partial pressures of 0.0, 220, 2200, 11000 and 22000 Pa). Irrespective of lighting conditions. catechol autoxidation was confirmed in aerated medium with a rate independent of the presence of 1-naphthol but proportional to the dissolved oxygen concentration, to the pH (its half-disappearance occurred in 24h at pH 8.5) and, to a lesser extent, to the incubating temperature (at 20 degrees C, 20% disappeared in 10 days at pH 7.0). Under alkaline conditions, the reaction of the anionic form (catecholate) with an equimolar concentration of molecular oxygen (O2) led presumably to hydrogen peroxide anion (HO2-) and coloured polymerization products. When tested alone, 1-naphthol was not significantly influenced either by lighting conditions, incubating temperature or dissolved oxygen concentration. It was also found to be quite stable with respect to pH, with a 15-fold weaker transformation rate than for catechol at the highest pH used. When tested in a mixture with catechol, 1-naphthol was found to be involved in a new chemical oxidation reaction catalyzed by catecholate. The transformation of one mole of 1-naphthol consumes four moles of oxygen. In the presence of catechol, the stoichiometry of the 1-naphthol transformation, under the influence of oxygen, suggests the possible formation of 2,5,6,8-tetrahydroxy 1,4-naphthoquinone via Lawsone (2-hydroxy 1,4-naphthoquinone) and naphthopurpurine (2,5,8-trihydroxy 1,4-naphthoquinone) as hypothetic intermediates. This is the first report of the autoxidation of 1-naphthol, catalyzed by catechol, in aqueous solution, in the absence of UV irradiation.

Catechols↗
PubMed2001

Design of an efficient medium for insect cell growth and recombinant protein production.

We report the development of a new serum-free medium based on the use of factorial experiments. At first, a variety of hydrolysates were screened using a fractional factorial approach with High-Five cells. From this experiment yeastolate ultrafiltrate was found to have, by far, the most important effect on cell growth. Furthermore, Primatone RL was found to remarkably prolong the stationary phase of Sf-9 and High-Five cell cultures. The optimal concentrations for yeastolate and Primatone were determined to be 0.6 and 0.5%, respectively, on the basis of a complete factorial experiment. This new medium, called YPR, supported good growth of both Sf-9 and High-Five cells in batch cultures, with maximal densities of 5.4 and 6.1 x 10(6) cells/ml, respectively. In addition, both cell lines achieved good growth in bioreactor batch culture and had a prolonged stationary phase of 3-4 d in YPR medium compared to Insect-XPRESS medium. The ability of the new medium to support recombinant protein expression was also tested by infecting Sf-9 or High-Five cells at high density (2 x 10(6) cells/ml) with a baculovirus expressing secreted placental alkaline phosphatase (SEAP). The maximum total SEAP concentration after 7 d was about 43 lU/ml (58 mg/L) and 28 lU/ml (39 mg/L) for High-Five and Sf-9 cells, respectively.

Amino Acids↗
PubMed2000

ATR-FTIR sensor development for continuous on-line monitoring of chlorinated aliphatic hydrocarbons in a fixed-bed bioreactor.

This article describes the continuous on-line monitoring of a dechlorination process by a novel attenuated total reflection-Fourier transform infrared (ATR-FTIR) sensor. This optical sensor was developed to measure noninvasively part-per-million (ppm) concentrations of trichloroethylene (TCE), tetrachloroethylene (PCE), and carbon tetrachloride (CT) in the aqueous effluent of a fixed-bed dechlorinating bioreactor, without any prior sample preparation. The sensor was based on an ATR internal reflection element (IRE) coated with an extracting hydrophobic polymer, which prevented water molecules from interacting with the infrared (IR) radiation. The selective diffusion of chlorinated compound molecules from aqueous solution into the polymer made possible their detection by the IR beam. With the exclusion of water the detection limits were lowered, and measurements in the low ppm level became possible. The best extracting polymer was polyisobutylene (PIB) in the form of a 5.8-microm thick film, which afforded a detection limit of 2, 3, and 2. 5 mg/L (ppm) for TCE, PCE, and CT, respectively. Values of the enrichment factors between the polymer coating and the water matrix of these chloro-organics were determined experimentally and were compared individually with predictions obtained from the slopes of absorbance/concentration curves for the three analytes. Before coupling the ATR-FTIR sensor to the dechlorinating bioreactor, preliminary spectra of the chlorinated compounds were acquired on a laboratory scale configuration in stop-flow and flow-through closed-loop modes. In this way, it was possible to study the behavior and direct response of the optical sensor to any arbitrary concentration change of the analytes. Subsequently, the bioreactor was monitored with the infrared sensor coupled permanently to it. The sensor tracked the progression of the analytes' spectra over time without perturbing the dechlorinating process. To calibrate the ATR-FTIR sensor, a total of 13 standard mixtures of TCE, PCE and CT at concentrations ranging from 0 to 60 ppm were selected according to a closed symmetrical experimental design derived from a 3(2) full-factorial design. The above range of concentrations chosen for calibration reflected typical values during normal bioreactor operation. Several partial least squares (PLS) calibration models were generated to resolve overlapping absorption bands. The standard error of prediction (SEP) ranged between 0.6 and 1 ppm, with a relative standard error of prediction (RSEP) between 3 and 6% for the three analytes. The accuracy of this ATR-FTIR sensor was checked against gas chromatography (GC) measurements of the chlorocompounds in the bioreactor effluents. The results demonstrate the efficiency of this new sensor for routine continuous on-line monitoring of the dechlorinating bioreactor. This strategy is promising for bioprocess control and optimization.

Biodegradation, Environmental↗
PubMed2000

Biodegradation of atrazine in sand sediments and in a sand-filter.

The potential of a microbial consortium for treating waters contaminated with atrazine was considered. In conventional liquid culture, atrazine and its two dealkylated by-products were equally metabolised by the microbial consortium. Transient production of hydroxyatrazine was observed during atrazine catabolism, indicating that the catabolic pathway was similar to the one reported for isolates capable of atrazine mineralisation. This consortium was then inoculated to sediments sampled from an artificial recharge site. These sediments were contaminated by atrazine and diuron and exhibited only a slow endogenous herbicide dissipation. Inoculated microorganisms led to extensive atrazine degradation and survived for more than 10 weeks in the sediments. A rudimentary bioreactor was then setup using a soil core originating from the same recharge site. Degrading microorganisms rapidly colonised the core and expressed their degrading activity. The efficiency of the bioreactor was improved in the presence of spiked environmental surface waters. Atrazine degraders thus possibly benefited from the other organic sources in developing and expressing their activity. The microbial consortium did not initially exhibit the capacity to degrade diuron, which was used as reference compound. No change in this characteristic was detected throughout the study.

Atrazine↗
PubMed2000

Glutathione-mediated mineralization of 14C-labeled 2-amino-4,6-dinitrotoluene by manganese-dependent peroxidase H5 from the white-rot fungus Phanerochaete chrysosporium.

Manganese-dependent peroxidase (MnP) H5 from the white-rot fungus Phanerochaete chrysosporium, in the presence of either Mn(II) (10 mM) or GSH (10 mM). was able to mineralize 14C-U-ring-labeled 2-amino-4,6-dinitrotoluene (2-A-4,6-DNT) up to 29% in 12 days. When both Mn(II) and GSH were present, the mineralization extent reached 82%. On the other hand, no significant mineralization was observed in the absence of both Mn(II) and GSH, suggesting the requirement of a mediator [either Mn(II) or GSH] for the degradation of 2-A-4,6-DNT by MnP. Using electron spin resonance (ESR) techniques, it was found that the glutathionyl free radical (GS*) was produced through the oxidation of GSH by MnP in the presence as well as in the absence of Mn(II). GS* was also generated through the direct oxidation of GSH by Mn(III). Our results strongly suggest the involvement of GS* in the GSH-mediated mineralization of 2-A-4,6-DNT by MnP.

Aniline Compounds↗
PubMed2000

Biodegradation of nitroaromatic pollutants: from pathways to remediation.

Nitroaromatic compounds are important contaminants of the environment, mainly of anthropogenic origin. They are produced as intermediates and products in the industrial manufacturing of dyes, explosives, pesticides, etc. Their toxicity has been extensively demonstrated in a whole range of living organisms, and nitroaromatic contamination dating from World War II is the proof of the recalcitrance of such compounds to microbial recycling. In spite of this, bacteria have evolved diverse pathways that allow them to mineralize specific nitroaromatic compounds. Degradation sequences initiated by an oxidation, an attack by a hydride ion, or a partial reduction have been documented. Some of these reactions have been exploited in bioreactors. Although pathways and enzymes involved are rather well understood, the molecular basis of these pathways is still currently under investigation. However, productive metabolism is an exception. As a rule, most bacteria are only able to reduce the nitro group into an amino function. This reduction is cometabolic: the metabolism of exogenous carbon sources is required to provide reducing equivalents. Composting and processes in bioreactors have exploited the easy reduction of the nitroaromatic compounds. In the case an amino-aromatic compound is produced, it is important to incorporate it in the remediation scheme. Some processes dealing with both nitro- and amino-aromatic compounds have been described, the amino derivative being either mineralized by the same or, more often, another microorganism, or immobilized on soil particles. Depending on the nitroaromatic compound and the environment it is contaminating, a whole range of reactions and reactor studies are now available to help devise a successful remediation strategy.

Biodegradation, Environmental↗
PubMed1999

Continuous degradation of mixtures of 4-nitrobenzoate and 4-aminobenzoate by immobilized cells of Burkholderia cepacia strain PB4.

Although isolated on 4-aminobenzoate, Burkholderia cepacia strain PB4 is also able to grow on 4-nitrobenzoate. Degradation of an equimolar mixture of the nitroaromatic compound 4-nitrobenzoate and its corresponding aminoaromatic derivative 4-aminobenzoate by this strain was investigated. Batch experiments showed that, irrespective of preculturing conditions, both compounds were degraded simultaneously. The mixture-degrading ability of B. cepacia strain PB4 was subsequently tested in continuous packed bed reactors (PBR) with the strain immobilized on Celite grade R-633 or R-635. Higher degradation rates were achieved with the larger particles of Celite R-635. Maximum simultaneous degradation rates per liter of packed bed of 0.925 mmol 1(-1) h(-1) 4-nitrobenzoate and 4-aminobenzoate were obtained for an applied loading rate of the same value (0.925 mmol 1(-1) h(-1) of each compound). Even when the applied load was not removed in its entirety, neither of the two compounds was degraded preferentially but a percentage of both of them was mineralized. The present study shows the possibility for a pure strain to biodegrade not only a nitroaromatic compound (4-nitrobenzoate) but also its corresponding amino derivative (4-aminobenzoate) continuously and simultaneously.

4-Aminobenzoic Acid↗
PubMed1999

Transformation and mineralization of 2,4,6-trinitrotoluene (TNT) by manganese peroxidase from the white-rot basidiomycete Phlebia radiata.

The degradation of the nitroaromatic pollutant 2,4,6-trinitrotoluene (TNT) by the manganese-dependent peroxidase (MnP) of the white-rot fungus Phlebia radiata and the main reduction products formed were investigated. In the presence of small amounts of reduced glutathione (10 mM), a concentrated cell-free preparation of MnP from P. radiata exhibiting an activity of 36 nkat/ml (36 nmol Mn(II) oxidized per sec and per ml) transformed 10 mg/l of TNT within three days. The same preparation was capable of completely transforming the reduced derivatives of TNT. When present at 10 mg/l, the aminodinitrotoluenes were transformed in less than two days and the diaminonitrotoluenes in less than three hours. Experiments with 14C-U-ring labeled TNT and 2-amino-4,6-dinitrotoluene showed that these compounds were mineralized by 22% and 76%, respectively, within 5 days. Higher concentrations of reduced glutathione (50 mM) led to a severe inhibition of the degradation process. It is concluded that Phlebia radiata is a good candidate for the biodegradation of TNT as well as its reduction metabolites.

Basidiomycota↗