Fungal bioconversion of 2,4-dichlorophenoxyacetic acid (2,4-D) and 2,4-dichlorophenol (2,4-DCP).

Ninety strains of fungi from the collection of our mycology laboratory were tested in Galzy and Slonimski (GS) synthetic liquid medium for their ability to degrade the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) and its by-product, 2,4-dichlorophenol (2,4-DCP) at 100 mg l(-1), each. Evolution of the amounts of each chemical in the culture media was monitored by HPLC. After 5 days of cultivation, the best results were obtained with Aspergillus penicilloides and Mortierella isabellina for 2,4-D and with Chrysosporium pannorum and Mucor genevensis for 2,4-DCP. The data collected seemed to prove, on one hand, that the strains responses varied with the taxonomic groups and the chemicals tested, and, on the other hand, that 2,4-D was less accessible to fungal degradation than 2,4-DCP. In each case, kinetics studies with the two most efficient strains revealed that there was a lag phase of 1 day before the onset of 2,4-D degradation, whereas there was none during 2,4-DCP degradation. Moreover, 2,4-DCP was detected transiently during 2,4-D degradation. Finally, M. isabellina improved its degradation potential in Tartaric Acid (TA) medium relative to GS and Malt Extract (ME) media.

Liquid chromatography study of pyrene degradation by two micromycetes in a freshwater sediment.

Pyrene biodegradation in a freshwater sediment without fungi supply, or inoculated with two sediment micromycetes, Mucor racemosus var. sphaerosporus and Phialophora alba was studied after 0, 5, 13, 28, 60 and 90 days. The influence of glucose addition was estimated, and a liquid chromatographic method for simultaneous quantitative determination of residual anthracene, fluoranthene and pyrene in the sediment was developed. Samples with PAHs were extracted in Soxhlet with ethyl acetate, and LC analysis was performed on a 5 microm Supelcosil column (150 x 4.6 mm I.D.) with gradient elution (2 ml min(-1)) of acetonitrile-water and UV detection at 254 nm. Recoveries of anthracene, fluoranthene and pyrene were 90.3%+/-1.1%, 93.2%+/-0.9% and 90.42%+/-1.9%, respectively, without interference. The native sediment microorganisms (with or without glucose added) have shown 35% pyrene degradation and sediment with glucose inoculated by the strains revealed 40%.

Effects of culture parameters on the degradation of 2,4-dichlorophenoxyacetic acid (2,4-D) and 2,4-dichlorophenol (2,4-DCP) by selected fungi. Groupe pour l'Etude du Devenir des Xénobiotiques dans l'Environnement (GEDEXE).

In order to enhance 2,4-D and 2,4-DCP degradation by four selected fungi (Cunninghamella elegans, C. echinulata, Rhizoctonia solani and Verticillium lecanii), three culture parameters (initial chemical concentration, amounts of glucose and nitrogen) were varied. The levels of both xenobiotics in the culture media were monitored by HPLC analysis after five days of cultivation. The best results were obtained at low initial concentration (20 mg.L-1 vs 100) and with low amounts of glucose (5 g.L-1 vs 10) and nitrogen (2.4 mM vs 24). When these two elements were lacking from the culture media, biodegradation was not suppressed, but took place to a lesser extent. Thus, initial chemical concentration and amounts of carbon and nitrogen, in the culture medium, were shown to strongly influence the extent of 2,4-D and 2,4-DCP removal by fungi.

Biodegradation of fluoranthene by soil fungi.

A selection of 39 strains of micromycetes known as good degraders of polychlorinated aromatic compounds, mostly isolated from soil and belonging to various taxonomic groups, have been investigated for fluoranthene degradation. Toxicity assays, first evaluated on solid medium MEA, have not shown any toxicity of fluoranthene (1-100 mg.L-1) towards fungi. Whereas, consumption assays on a solid synthetic medium showed a toxicity at 100 mg.L-1. The degradation of fluoranthene (10 mg.L-1) was then investigated in a liquid synthetic medium for 4 days and evaluated by HPLC. Among the 39 strains tested, 18 degraded fluoranthene at 60% or more. Zygomycetes appeared to be the most efficient group (mean degradation: 90%). Among 18 performant strains, 10 had not yet been reported in the literature: Sporormiella australis, Cryptococcus albidus, Cicinobolus cesatii, Pestalotia palmarum, beauveria alba, Aspergillus terreus. Cunninghamella blakesleeana, C. echinulata, Mortierella ramanniana and Rhizopus arrhizus. Fluoranthene adsorption on fungi was very low for the strains which degraded well fluoranthene (mean adsorption: 4%). Whereas, some strains adsorbed it much more such as Colletotrichum dematium (47%) and Penicillium italicum (43%).

Health risk assessment of a modern municipal waste incinerator.

During the modernization of the municipal waste incinerator (MWI, maximum capacity of 180,000 tons per year) of Metropolitan Grenoble (405,000 inhabitants), in France, a risk assessment was conducted, based on four tracer pollutants: two volatile organic compounds (benzene and 1, 1, 1 trichloroethane) and two heavy metals (nickel and cadmium, measured in particles). A Gaussian plume dispersion model, applied to maximum emissions measured at the MWI stacks, was used to estimate the distribution of these pollutants in the atmosphere throughout the metropolitan area. A random sample telephone survey (570 subjects) gathered data on time-activity patterns, according to demographic characteristics of the population. Life-long exposure was assessed as a time-weighted average of ambient air concentrations. Inhalation alone was considered because, in the Grenoble urban setting, other routes of exposure are not likely. A Monte Carlo simulation was used to describe probability distributions of exposures and risks. The median of the life-long personal exposures distribution to MWI benzene was 3.2 x 10(-5) micrograms/m3 (20th and 80th percentiles = 1.5 x 10(-5) and 6.5 x 10(-5) micrograms/m3), yielding a 2.6 x 10(-10) carcinogenic risk (1.2 x 10(-10)-5.4 x 10(-10)). For nickel, the corresponding life-time exposure and cancer risk were 1.8 x 10(-4) micrograms/m3 (0.9 x 10(-4)-3.6 x 10(-4) micrograms/m3) and 8.6 x 10(-8) (4.3 x 10(-8)-17.3 x 10(-8)); for cadmium they were respectively 8.3 x 10(-6) micrograms/m3 (4.0 x 10(-6)-17.6 x 10(-6)) and 1.5 x 10(-8) (7.2 x 10(-9)-3.1 x 10(-8)). Inhalation exposure to cadmium emitted by the MWI represented less than 1% of the WHO Air Quality Guideline (5 ng/m3), while there was a margin of exposure of more than 10(9) between the NOAEL (150 ppm) and exposure estimates to trichloroethane. Neither dioxins nor mercury, a volatile metal, were measured. This could lessen the attributable life-long risks estimated. The minute (VOCs and cadmium) to moderate (nickel) exposure and risk estimates are in accord with other studies on modern MWIs meeting recent emission regulations, however.

Microbial disinfection capacity of municipal solid waste (MSW) composting.

The disinfection capacity of a municipal solid waste (MSW) composting plant (Siloda) has been evaluated. In spring and summer, MSW was followed during the composting process from raw material to mature compost and long-term storage (1 year). Ascaris eggs, Salmonella, Shigella, total streptococci, faecal streptococci, total coliforms, faecal coliforms and Escherichia coli were studied. Disinfection was successful in terms of a decrease in faecal contamination indicators and disappearance of faecal pathogens. Faecal coliform concentration in raw waste reached 2.1 x 10(8) cfu g-1 dry weight in spring (CI 95%:5.2 x 10(7)-3.4 x 10(8)) and 7.2 x 10(8) cfu g-1 dry weight (1 x 10(8)-1.7 x 10(9)) in summer, and fell to less than 100 cfu g-1 dry weight within 20 d. Faecal streptococci concentrations reached 8.7 x 10(8) cfu g-1 dry weight (3.7 x 10(8)-1.3 x 10(9)) in spring and 2.0 x 10(9)cfu g-1 dry weight (5.6 x 10(8)-3.4 x 10(9)) in summer, and fell to 8.7 x 10(4) cfu g-1 dry weight (6.9 x 10(4)-1.0 x 10(5)). No seasonal pattern of contamination, mainly of animal origin, was observed. Microbiological quality of finished compost depends on the storage conditions. Therefore, the storage stage should be viewed as part of the composting process. Monitoring disinfection capacity of MSW composting needs to combine several microbial populations.

Biodegradation of anthracene by soil fungi. Groupe pour l'Etude du Devenir des Xénobiotiques dans l'Environnement (GEDEXE).

A selection of 39 strains of micromycetes known as good degraders of polychlorinated aromatic compounds, mostly isolated from soil and belonging to various taxonomic groups, have been investigated for anthracene degradation. Toxicity and consumption assays, first evaluated on solid media, have not shown any toxicity of anthracene (1-100 mg.L-1) towards fungi. Degradation of anthracene (10 mg.L-1) was then investigated in a liquid synthetic medium for 4 days and evaluated by HPLC. Among the 39 strains tested, 19 degraded anthracene at 50% or more. Zygomycetes appeared to be the most efficient group (mean degradation : 81%) while Melanconiales were the least efficient (mean: 14%). Among 19 efficient strains, 8 had not yet been reported in the literature: Cryphonectria parasitica, Ceriporiopsis subvermispora, Oxysporus sp., Cladosporium herbarum, Drechslera spicifera, Verticillium lecanii, Fusarium moniliforme var. subglutinans and Rhizopus arrhizus.

[Meta-analysis and dose-response functions of air pollution respiratory effects]. / Méta-analyse et fonctions dose-réponse des effets respiratoires de la pollution atmosphérique.

A Meta-analysis was conducted on 107 original epidemiological papers published between 1980 and 1993 on the respiratory health impact of suspended particles, sulfur dioxide, ozone and nitrogen dioxide. These are the usual air quality indicators that are monitored in urban ambient air. The range of exposures that were studied (averages and 90th percentiles are respectively up to 88, 83, 124 and 53 mu/m3 (averages) and 180, 147, 234 and 131 micrograms/m3 (90th percentile) for each of the four pollution indicators) allowed assessment of dose-response functions for irritative pulmonary conditions (cough and/or asthma episodes) and respiratory function (FEVI and peak expiratory flow). The dose-response functions seem linear in the range of observed concentrations, with nitrogen dioxide showing the least consistent association across different health indicators. When applied to exposures measured as a 100 micrograms/m3 increase of pollutant's concentrations compared to low background values, the average relative risks of airway irritation (cough and/or asthma épisodes) range, according to the pollutant, from 1.08 to 1.47; average relative decreases of respiratory function amount to 1.1 to 2.2%. The effects are most often stronger among young subjects. The results of this study, when applied to air pollution concentrations in the range of values that were observed in this study, should foster health risk assessment studies in places where population air pollution exposures are available.

Biodegradation of three substituted phenylurea herbicides (chlortoluron, diuron, and isoproturon) by soil fungi. A comparative study.

As a part of a study conducted on the fate of xenobiotics in the environment, a selection of 90 strains of micromycetes, mostly isolated from soil and belonging to various taxonomic groups, have been cultivated in liquid synthetic medium with chlortoluron (100 mg.L-1), diuron (20 mg.L-1), and isoproturon (100 mg.L-1) for 5 days. Evaluation of the chemicals in the culture media was made by HPLC. Our results show a wide variation not only with taxonomic groups but also with the species, and with the tested chemicals. On the whole, 4, 7, and 11% of the strains depleted respectively chlortoluron, diuron, and isoproturon, at 50% or over. Rhizoctonia solani was the only strain that depleted each of the 3 substituted phenylureas over 70%, nevertheless, the growth of this fungus was slightly inhibited by diuron. The very fast disappearance of the chemicals from the culture media was only due to biotic phenomena, as no adsorption occurred on the fungal biomass. So, depletion translated a real biodegradation of the tested substrates.

Influence of environmental adaptation on the sensitivity of micromycetes to pentachlorophenol toxicity in microcosms.

The search for fungal strains capable of pentachlorophenol degradation led to the isolation and identification of 98 different micromycetes. When these strains were submitted to toxicity tests in aquatic microcosms at concentrations of 10 mgL-1 and 100 mgL-1 PCP, inhibition of growth was 57% and 36% respectively of the total isolated and identified strains. Among the strains inhibited at 10 mgL-1, 6 can serve as bioindicators of PCP pollution while the strains resistant can be regarded as potential PCP biodegraders. It was confirmed that fungal strains isolated from sites contaminated by chemically different products manifest different levels of sensitivity to PCP toxicity and probably different biodegradation potentials.

Hazard to man and the environment posed by the use of urban waste compost: a review.

This review presents the current state of knowledge on the relationship between the environment and the use of municipal waste compost in terms of health risk assessment. The hazards stem from chemical and microbiological agents whose nature and magnitude depend heavily on the degree of sorting and on the composting methods. Three main routes of exposure can be determined and are quantified in the literature: (i) The ingestion of soil/compost mixtures by children, mostly in cases of pica, can be a threat because of the amount of lead, chromium, cadmium, PCDD/F and fecal streptococci that can be absorbed. (ii) Though concern about contamination through the food chain is weak when compost is used in agriculture, some authors anticipate accumulation of pollutants after several years of disposal, which might lead to future hazards. (iii) Exposure is also associated with atmospheric dispersion of compost organic dust that convey microorganisms and toxicants. Data on hazard posed by organic dust from municipal composts to the farmer or the private user is scarce. To date, microorganisms are only measured at composting plants, thus raising the issue of extrapolation to environmental situations. Lung damage and allergies may occur because of organic dust, Gram negative bacteria, actinomycetes and fungi. Further research is needed on the risk related to inhalation of chemical compounds.

Evaluation of acrodontiolamide, a chlorinated compound produced by Acrodontium salmoneum de Hoog for cytotoxicity and antimicrobial activity.

A new antifungal compound has been isolated from the culture medium of Acrodontium salmoneum de Hoog. Its structure was previously elucidated and was named acrodontiolamide. However, this compound is not characteristically produced by the genus Acrodontium, it is rather a feature of one isolate of A. Salmoneum coming from the soil of the grotto of La Pierre Saint Martin (France). Production, purification, cytotoxicity and antimicrobial activities of acrodontiolamide are described. Concerning microorganisms, inhibitory activity seems to be specifically restricted to phytopathogenic and entomapathogenic fungi. Acrodontiolamide is not cytotoxic to either normal human cultured cells or tumor cells.

Comparison of the toxicity of various lignin-related phenolic compounds toward selected fungi perfecti and fungi imperfecti.

A selection of 88 micromycetes belonging to various taxonomic groups was grown on malt-agar medium containing different lignin-related phenolic compounds (ferulic, protocatechuic, syringic and vanillic acids, catechol, guaiacol, and phenol) at concentrations ranging from 100 to 1000 mg/liter. Growth and morphological changes were recorded after 15 days of incubation. Behavior of fungi in contact with the different phenolics were compared. Some strains were very sensitive and might constitute specific or general bioindicators of pollution. The relationships between origin of isolates or constitutive phenoloxidase activity and resistance or sensitivity of fungi to phenolics were researched. No evidence of implication of these two parameters was found.

Sensitivity and specificity of phenoloxidase reactions of 1059 strains and species of Micromycetes cultivated on malt/agar medium.

The phenoloxidase (POx) activities of 1059 strains and species of micromycetes were determined on malt/agar medium. Overall, 600 (57%) of the isolates produced one or more POx. The sensitivity and specificity of the POx activities towards various substrates were used to group the isolates. Some 187 strains (31% of those producing POx) produced well-defined enzymes, 236 (39%) produced incompletely identified enzymes and 177 (30%) produced other, unidentified POx.

Antitumoral activity of patulin and patulin-cysteine adducts.

A comparison between the cytotoxicity and the anti-tumoral activity of patulin and patulin-cysteine adducts has been conducted. In vitro assays using L 1210 and P 388 cells showed that patulin-cysteine adducts had less cytotoxic activity than patulin (IC50 was 4 fold of IC50 patulin). In vivo, cysteine not only reduced the toxicity of patulin but also reduced its antitumoral activity against L 1210 and P 388 mouse leukemias (respectively of 25.7% and 46.6% with 3 mg.kg-1.d-1). By addition on patulin at the 4 or 7 position, cysteine might interfere with the lactone function and the -CH2- in in position 6 which are responsible for the antitumoral activity.

Diversity in phenol-metabolizing capability of 809 strains of micromycetes.

The property of 809 strains of micromycetes to grow in the presence of phenol (0.5 g/l) was investigated on solid media. Toxicity was determined on malt extract agar medium. Growth of the fungal strains on synthetic solid medium with phenol as the sole carbon source allowed evaluation of phenol consumption. Only 61 strains (8% of the whole) grew well under both conditions, which reflects the toxicity of 0.5 g/l of phenol upon micromycetes. Finally, Phanerochaete chrysosporium was chosen and cultivated in liquid synthetic medium at 24 degrees C and 37 degrees C with phenol (0.5 g/l). It consumed phenol fast and efficiently. Growth temperature was a key parameter. Catechol, cis-cis-muconic acid and beta-keto-adipate were characterized and quantified.

Extracellular phenoloxidase activity of micromycetes from various taxonomic groups.

The ability of Micromycetes strains to produce extracellular phenoloxidases was examined on solid malt agar medium using ten different reagents. We established a POx index summarizing the global activity given by the ten reagents used. The results indicated a wide variability depending on the taxonomic groups, the genera and the species. Some groups were relatively homogeneous, either no and low producers of phenoloxidases (Yeasts, Zygomycetes, genera Aspergillus and Penicillium) or medium and high producers of phenoloxidases (Basidiomycetes, Coelomycetes, Tuberculariales and Dematiaceae), while other groups were very heterogeneous (Ascomycetes, Mucedinaceae). The POx index was significantly higher for strains recently isolated than for strains kept in the fungi collection for a long time.

Relationship between the biodegradative capability of soil micromycetes for pentachlorophenol and for pentachloronitrobenzene.

A collection of 1056 strains in our laboratory were incubated with various xenobiotics among which were two potent fungicides: pentachlorophenol (PCP) and pentachloronitrobenzene (PCNB). The production of extracellular phenoloxidases were examined, using a series of ten different reagents. On the whole, PCNB is less accessible to fungal degradation than PCP. Although no correlation was found between the biodegradative capability of individual fungal strains for PCP or for PCNB, when taxonomic groups were considered as a whole, the same activity profiles were found. Zygomycetes were the most efficient; yeasts the least efficient towards both substrates. A more detailed study of the metabolism of both substrates on selected strains is in progress.

Antitumor activity of patulin and structural analogs.

A comparison between the cytotoxicity and the antitumor activity of patulin and five structural analogs (isopatulin, dehydroisopatulin, dimethylisopatulin, trimethylisopatulin and isopropylisopatulin) has been established. In vitro assays using L 1210 and P 388 cells showed that the structure of the pyranic ring as well as the nature of the substituents influenced the observed activities. Among the five structural analogs of patulin assayed in vivo against Ehrlich carcinoma, L 1210 and P 388 leukemias, dehydroisopatulin was the only one to be active on all 3 types of tumors at a dose of 100 mg.kg-1.d-1. The ratio between the LD50 in mice and the active dose was 5 while with patulin it was 10. It can be assumed that the lactone function is not solely responsible for the activity of patulin and its structural analogs.

Phenoloxidase production and vanillic acid metabolism by Zygomycetes.

The ability of 23 strains of Zygomycetes to produce extracellular phenoloxidases was examined on solid media by using 10 different reagents. The results varied depending on the reagent and indicated that most of the strains were devoid of phenoloxidase activity. The production of inducible phenoloxidases was demonstrated by the Bavendamm reaction. The study of the biotransformation of vanillic acid in synthetic medium indicated that the reaction most often obtained was the reduction of vanillic acid to vanillyl alcohol. Helicostylum piriforme and Rhizopus microsporus var. chinensis completely metabolized vanillic acid while good transformation was obtained with Absidia spinosa, Cunninghamella bainieri, Mucor bacilliformis, Mucor plumbeus, Rhizopus arrhizus, Rhizopus stolonifer, Syncephalastrum racemosum and Zygorhynchus moelleri. Other strains did not degrade or poorly degraded vanillic acid. Decarboxylation and demethoxylation of this compound was independent of the production of phenoloxidases as in the case of white-rot fungi. Other enzymatic systems might be implicated in this phenomenon.