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Journal of Environmental Quality 30:418-426 (2001)
© 2001 American Society of Agronomy, Crop Science Society of America, and Soil Science Society of America

TECHNICAL REPORT
BIOREMEDIATION AND BIODEGRADATION

Removal of Herbicides from Liquid Media by Fungi Isolated from a Contaminated Soil

Ouahiba Bordjibaa, Régine Steimanb, Malika Kadrib, Ammar Semadia and Pascale Guiraudb

a Institut des Sciences de la Nature, Université d'Annaba, 23000 Annaba, Algérie
b EA 2945: Environnement-Santé, Groupe pour l'Etude du Devenir des Xénobiotiques dans l'Environnement (GEDEXE), UFR de Pharmacie, Université Joseph Fourier, Grenoble, France

Corresponding author (Pascale.Guiraud{at}ujf-grenoble.fr)

Received for publication June 21, 2000.

   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS AND DISCUSSION
 CONCLUSION
 REFERENCES
 
Fungi were isolated from soil samples corresponding to pesticide-contaminated soil (CS) and noncontaminated soil (NCS) in the Annaba vicinity (Algeria) and identified. The number of isolates obtained from CS and NCS were 263 and 288, respectively. The most frequent species (Aspergillus fumigatus, A. niger, A. terreus, Absidia corymbifera, and Rhizopus microsporus var microsporus) were not sensitive to the pesticides. The growth of the genus Trichoderma was inhibited by the pesticides, while genera Absidia and Fusarium were stimulated. The 53 species isolated were assayed for their ability to remove metribuzin from liquid medium. Only Botrytis cinerea from NCS and Sordaria superba and Absidia fusca from CS removed more than 50% of the compound after 5 d. Metamitron was very resistant. Among the 21 species tested, only Alternaria solani (from NCS), Drechslera australiensis (from CS and NCS), and Absidia fusca (from CS) reduced the concentration in the medium more than 10% (10–16%). Twelve species were grown with linuron, seven of them were inefficient in removing this compound. The two strains of Sordaria macrospora yielded 22 to 25% depletion, while Botrytis cinerea depleted linuron almost completely. Among the 31 species assayed for their ability to eliminate metobromuron, Botrytis cinerea (from CS and NCS) depleted almost completely the chemical from the medium. Rhizopus oryzae and Absidia fusca from CS removed 40 and 47% of the compound, respectively. No systematic relationships were observed between the soil contamination and herbicide elimination capacities of soil fungi. Absidia fusca and Botrytis cinerea were particularly interesting for bioremediation purposes because they were able to transform efficiently three of the four compounds assayed.

Abbreviations: CS, contaminated soil • GS, Galzy and Slonimski • HPLC, high performance liquid chromatography • MEA, malt extract agar • NCS, noncontaminated soil • RT, retention time


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS AND DISCUSSION
 CONCLUSION
 REFERENCES
 
ENVIRONMENTAL contamination by toxic xenobiotic chemicals, arising mainly from agricultural and industrial sources, and the consequences on food quality and human health are very serious worldwide problems. The extensive use of pesticides (fungicides, herbicides, insecticides) in various parts of the world for years without any control was responsible for a dramatic increase of pollution level in soil and water.

The empiric agricultural methods commonly employed in some countries, particularly in North Africa, led people to adopt short-cycle cultures such as Cucurbitaceous (pumpkin, melon) or Solanaceous (potato, tomato) without any considerations for soil, climate, or cultural requirements. As a result, numerous problems, such as fungal diseases and weed proliferation, occurred and limit considerably the crop yields. Moreover, fungicides and herbicides were often randomly used, resulting both in inefficiency and soil and ecosystem contamination.

In Algeria, the problem recently began to be taken into consideration. The National Institute for Plant Protection, located in El Krous (about 30 km east from Annaba), is administratively and technically responsible for authorized pesticide use and is in charge of both advising and monitoring during crop treatment. Experiments are now conducted to assess the efficiency but also the residues in soil and the environmental effect of the new proposed pesticides.

In collaboration with this institute, we have chosen experimental sites treated or not by pesticide mixtures in order to compare the soil fungal microflora. This is part of a program of investigation on the effect of industrial or agricultural chemical releases in the environment on natural soil fungal flora. Most studies investigating the effect of pesticides on soil microflora dealt with bacterial communities or total biomass evolution (Junnila et al., 1993; Assaf and Turco, 1994; Grigg et al., 1997; Shapir and Mandelbaum, 1997; Wenk et al., 1997; Engelen et al., 1998). Fungal communities were considerably less analyzed. Previous works were done by our team, either in situ, by comparisons of the mycoflora present in control and polluted soils (e.g., in Oued Sebou, Morocco; Sage et al., 1997), or in the laboratory, by studying the behavior of soil fungi toward various pesticides (Seigle-Murandi et al., 1992; Benoit-Guyod et al., 1994; Steiman et al., 1994; Vroumsia et al., 1996, 1999; Guiraud et al., 1999; Khadrani et al., 1999).

The purpose of the present study was to investigate the effect on the mycoflora of the accumulation of various herbicides (metribuzin, metamitron, metobromuron, linuron) and fungicides (propineb, mancozeb, and sandofan) in soil. The ability of the isolated fungal species and strains to remove some of the herbicides present at the highest concentrations from liquid media was researched. Our aim was to determine if soil fungi can adapt to critical ecological situations such that they may be useful in the area of soil chemical pollution either as bioindicators or as actors in detoxification and bioremediation.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS AND DISCUSSION
 CONCLUSION
 REFERENCES
 
Fungal Strain Isolation
Soil samples were taken from 1 to 10 cm depth and collected in sterile tubes. Contaminated soil (CS) came from a parcel treated by herbicides and fungicides where tomato (Lycopersicon esculentum var. esculentum) and potato (Solanum tuberosum L.) were cultivated (El Krous, 30 km east from Annaba). Noncontaminated control soil (NCS) came from an untreated parcel in the same area. Twenty samples were taken in both cases, and were mixed to obtain a representative mycoflora of CS and NCS. Samples were stored at +4°C until analysis. The isolation of the strains was accomplished by using the soil plates method of Warcup (Parkinson and Waid, 1960). Aliquots of the soil samples were dispatched into Petri dishes (90 mm diameter) and covered with a sterile medium: MEA [malt extract (1.5%)–agar (1.5%)] added by chloramphenicol (0.05%). Dishes were incubated at 22, 37, 45 and 55°C and the fungal strains were isolated as soon as colonies appeared. For each condition and each sample, five dishes were used and the whole experiment was done in triplicate. After identification, the strains were maintained in slant tubes on MEA medium. Those isolated at 22°C were stored at 4°C, while the others were kept at room temperature.

Chemicals
Metribuzin [4-amino-6-(1,1-dimethylethyl)-3-(methyl-thio)-1,2,4-triazin-5(4H)-one] and metamitron [4-amino-3methyl-6-phenyl-1,2,4-triazin-5(4H)-one] were from Bayer SA (Puteaux, France), metobromuron [N'-(4-bromophenyl)-N-methoxy-N-methylurea] was purchased from BASF (Ludwigshaffen, Germany), and linuron [N'-(3,4-dichlorophenyl)-N-methoxy-N-methylurea] was from Agrevo (Gif sur Yvette, France). The structural formulas of these compounds appear in Fig. 1 . Other chemicals were from Prolabo (Paris, France).



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  		Fig. 1. Structural formulas of linuron, metobromuron, metribuzin, and metamitron

 
Bioremediation Assays
Two triazines (metribuzin and metamitron) and two phenylureas (linuron and metobromuron) were added to cultures in liquid medium of the species isolated from CS and NCS. All fungi were tested with metribuzin while only selected strains were used in experiments with the other compounds. The disappearance of the compounds from culture media was monitored. When control strains were not isolated from NCS, they were taken out of the collection of our laboratory, Collection Mycology Pharmacy Grenoble (CMPG).

To obtain sufficient inoculum for liquid medium cultures, precultures of each strain were grown on MEA Petri dishes at their temperature of isolation for 8 to 15 d, depending on their growth rate. They were inoculated (mycelium and spores from one dish) into 100-mL Erlenmeyer flasks containing 25 mL of synthetic GS medium of Galzy and Slonimski (1957) with glucose (5 g/L) at pH 4.5. Flasks and liquid medium were previously sterilized by autoclaving for 20 min at 121°C. Cultures were incubated under shaking (180 rpm, orbital shaker) for 2 d to obtain enough biomass. At this time, herbicides (metribuzin, metamitron, linuron, or metobromuron) were added aseptically (stock solutions were prepared in ethanol and were sterilized by filtration through 0.22-µm Millipore [Bedford, MA] membranes, 100 µL were added in the cultures, corresponding to a final concentration in ethanol of 0.4%) at a final concentration of 100 mg/L. The depletion of pesticides was evaluated after five more days of cultivation at 24°C (except for Malbranchea cinnamomea, Thermoascus aurantiacus, and Thermomyces lanuginosus, which were grown at 37°C), under daylight lamps (photoperiod: 12 h/d, 1200 lux). Controls were medium and pesticide without fungi, analyzed after 5 d at 24 or 37°C (abiotic degration). Each assay was performed in triplicate.

Evaluation of the Depletion of Herbicides
The disappearance of the products was monitored by high performance liquid chromatography (HPLC) as previously described (Balinova, 1993; Khadrani et al., 1999; Vroumsia et al., 1999). Aliquots from the culture media were taken with a syringe, filtered through a Millipore membrane (0.45 µm), and injected directly. High performance liquid chromatography was performed with a liquid chromatograph (Shimadzu, Kyoto, Japan) equipped with a LC 6A pump, a SIL-9A automatic injector, and a SPD 6A UV detector. The separation column (4.0 mm i.d. x 300 mm long) was filled with µ-Bondapak C18 (Supelcosil LC 18) (Millipore, Milford, MA). The mobile phase was methanol and water (65:35). The flow rate was 1 mL/min and detection was recorded at 294 nm for metribuzin and metamitron, and at 247 nm for metobromuron and linuron. Three injections per sample were done, and quantification was obtained from calibration with a standard solution.


   RESULTS AND DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS AND DISCUSSION
CONCLUSION
 REFERENCES
 
Effect of Pesticides on the Soil Mycoflora
The study of the mycoflora present in both CS and NCS led to 551 isolates corresponding to 53 species of micromycetes. Table 1 gives the detailed list of the species isolated, the soil in which they were found, and the number of isolates (total cumulated number of colonies obtained during the isolation process). Five species were isolated with a high frequency (more than 20 isolates from each soil). On the other hand, some usually common species were not abundantly present (Acremonium chrysogenum, Alternaria alternata, Aspergillus carneus, A. flavus, Botrytis cinerea, Gliocladium roseum, Corynascus sepedonium, Penicillium citrinum, P. pinophilum, Phanerochaete chrysosporium, and Syncephalastrum racemosum).


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  		Table 1. List of the 53 species isolated from noncontaminated soil (NCS) and pesticide-contaminated soil (CS).{dagger}

 
The second column of Table 1 gives the 288 isolates obtained from NCS. The optimal isolation temperature was 45°C, it allowed us to obtain 72 isolates (at 22°C, 66 isolates; at 30°C, 64 isolates; at 37°C, 57 isolates; at 55°C, 21 thermophilous and thermotolerant isolates, namely Aspergillus fumigatus, Malbranchea cinnamomea, Paecilomyces lilacinus, Thermoascus aurantiacus, and Thermomyces lanuginosus). The purpose of this procedure was to obtain gradually a high number of species as a function of their growth rate at the different temperatures, making their isolation easier. However, it must be noted that the study of soil mycoflora is never exhaustive. Moreover, it is impossible to know the real state of fungi in soil (mycelium, spore, resistant forms, etc.). Concerning CS, some species were not sensitive to the pesticides since they were found with the same frequency than in NCS, which was the case for the five most abundant species. For some others, a dramatic decrease in frequency was noticed, for example 50 isolates of species belonging to the genus Trichoderma were obtained in NCS versus 13 in CS. Conversely, the genera Fusarium and Absidia seemed to be stimulated by the presence of pesticides since 19 isolates of Fusarium and 37 isolates of Absidia were obtained in CS versus respectively 13 and 26 in NCS. Moreover, one species of Absidia was found only in pesticide-treated soil. In CS, the best temperatures for isolation were 22 and 37°C and a total of 263 isolates was obtained.

The genus Penicillium was very poorly represented; only two common and ubiquitous species were isolated. Penicillium citrinum was found in northern (Nespiak, 1970) as well as in southern or tropical (Kobayasi et al., 1977) parts of the world and both in mineral and organic soils (Balasooriya and Parkinson, 1967). Penicillium pinophilum is an active biodeterioration agent in soil. These strains were not isolated from CS.

On the whole, 25% of the isolated species belonged to the Fungi Perfecti, and apart from Phanerochaete chrysosporium (Basidiomycete) and Absidia blakesleeana (Zygomycete), these species were all Ascomycetes. Moreover, these species were more abundant in CS (42 isolates versus 27 in NCS). Some species were only present in CS (Byssochlamys nivea, Corynascus sepedonium, Dichotomomyces cejpii, Sordaria macrospora, and S. superba) and the number of isolates of Emericella nidulans and Sordaria fimicola was higher.

The global distribution observed, particularly the presence of several perfect species, several Aspergillii and Zygomycetes, and only a few Penicillii, corresponded with the mycoflora described in hot parts of the world (Domsch et al., 1980; Abdel-Hafez, 1982; Mouchacca, 1982; Guiraud et al., 1995; Steiman et al., 1995, 1997).

The CS analyzed contained several compounds among which the herbicides metribuzin, metobromuron, linuron, and metamitron associated to the fungicides propineb [[(1-methyl-1,2-ethanediyl)bis(carbamodithioatol)](2-) zinc homopolymer], mancozeb [((1,2-ethanediylbis (carbamodithioato))(2-))manganese mixture with ((1,2-ethandiylbis(carbamodithioate))(2-))zinc], and sandofan [2-methoxy-N-(2-oxo-1,3-oxazolidin-3-yl) acet-2',6'-xylide] (data from the Algerian National Institute for Plant Protection, not shown). Results obtained showed that most species were resistant or able to adapt to this situation. The total amount of isolates was not very different between CS and NCS.

Bioremediation Ability of Species Isolated from Contaminated Soil and Noncontaminated Soil
Abiotic degradation of the compounds was negligible, since it did not exceed 2% after 5 d in all cases except for linuron (57%) (Fig. 2 and 3) . Table 2 gives the disappearance obtained in culture media. Some species (Sordaria superba, Ramichloridium subulatum, Acremonium chrysogenum, Aspergillus niger, Fusarium solani, F. ventricosum, Absidia corymbifera, A. fusca, and Rhizopus microsporus var microsporus) were stimulated by the presence of pesticides in soil and depleted at least two of the compounds at a higher level than the control when isolated from contaminated soil. The most significant results were obtained with A. fusca, for which an increased efficiency was noticed with all compounds. In other cases the strains from NCS gave better results than those from CS, this was only noticed with metribuzin and metobromuron for Byssochlamys nivea (metribuzin, metobromuron), Botrytis cinerea (metribuzin), Thermomyces lanuginosus (metobromuron), Aspergillus terreus (both), Malbranchea cinnamomea (both), and Trichoderma harzianum (metribuzin). No clear influence of the origin was noted for Corynascus sepedonium, Dichotomomyces cejpii var spinosus, Sordaria macrospora, Aspergillus fumigatus, A. niveus, Fusarium oxysporum, and Mucor hiemalis, whatever the compound considered. Some species exhibited very low bioremediation potentialities (results always below 20%), which was the case for C. sepedonium, D. cejpii, S. macrospora, Trichoderma hamatum, and M. hiemalis. Detailed effects of these herbicides on soil microflora and their biotransformation by soil microrganisms (bacteria and fungi) have been poorly documented.



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  		Fig. 2. High performance liquid chromatography (HPLC) chromatograms for metribuzin and metamitron showing abiotic controls and degradation by Absidia fusca and Botrytis cinerea. (A) metribuzin 1, retention time (RT): 2.99 min; (a) standard (100 mg/L), 1 = 100%; (b) abiotic control, 1 = 98%; (c) Absidia fusca NCS, 1 = 77%, main other peaks RT (min): 0.74–1.39–1.75–2.07–2.36; (d) Absidia fusca CS, 1 = 8%, main other peaks RT (min): 1.40–1.91–2.37; (e) Botrytis cinerea NCS, 1 = 35%, main other peaks RT (min): 1.44–1.62–2.09–2.39; (f) Botrytis cinerea CS, 1 = 75%, main other peaks RT (min): 1.45–1.75–2.10–2.39. (B) metamitron 2, RT: 2.46 min; (a) standard (100 mg/L), 2 = 100%; (b) abiotic control, 2 = 98%; (c) Absidia fusca NCS, 2 = 92%, main other peaks RT (min): 1.38–1.75–1.96; (d) Absidia fusca CS, 2 = 84%, main other peaks RT (min): 1.37–1.71–2.1. NCS, noncontaminated soil; CS, contaminated soil. Major peaks are underlined, secondary peaks are in italic type

 


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  		Fig. 3. High performance liquid chromatography (HPLC) chromatograms for metobromuron and linuron showing abiotic controls and degradation by Absidia fusca and Botrytis cinerea. (A) metobromuron 3, retention time (RT): 3.67 min; (a) standard (100 mg/L), 3 = 100%; (b) abiotic control, 3 = 99%; (c) Absidia fusca NCS, 3 = 78%, main other peaks RT (min): 1.44–2.00–2.68–2.86; (d) Absidia fusca CS, 3 = 54%, main other peaks RT (min): 1.42–1.99–2.67–2.85; (e) Botrytis cinerea NCS, 3 = 10%, main other peaks RT (min): 1.50–1.99–2.69–2.86; (f) Botrytis cinerea CS, 3 = 10.5%, main other peaks RT (min): 1.49–2.03–2.40–2.70–2.88. (B) linuron 4, RT: 2.98 min; (a) standard (100 mg/L), 4 = 100%; (b) abiotic control, 4 = 43%; (c) Absidia fusca NCS, 4 = 34.5%, main other peaks RT (min): 1.38–2.31; (d) Absidia fusca CS, 4 = 20%, main other peaks RT (min): 1.39–2.36; (e) Botrytis cinerea NCS, 4 = 0.5%, main other peaks RT (min): 1.36–1.53–1.79–2.11–2.33–3.33; (f) Botrytis cinerea CS, 4 = 7%, main other peaks RT (min): 1.50–2.12–2.34–3.27. NCS, noncontaminated soil; CS, contaminated soil. Major peaks are underlined, secondary peaks are in italic type

 

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  		Table 2. Depletion of metribuzin, metamitron, metobromuron, and linuron in the liquid culture medium of strains isolated from noncontaminated soil (NCS) and pesticide-contaminated soil (CS).{dagger}

 
Metribuzin is a pre- and postemergence triazine herbicide (Fig. 1). It is widely used to treat a variety of crops. This compound is absorbed by roots and acts by inhibiting photosynthesis with a wide spectrum (monocotyledones, dicotyledones). Kim and Feagley (1998) showed that metribuzin was weakly adsorbed to soil and mostly leached into the surface or ground water. However, other studies have shown that metribuzin can remain for a long time in soil (half-life: 17–377 d under aerobic conditions, depending on temperature, pH, and soil type), accumulates in plants, and interferes with other crops (Sharom and Stephenson, 1976; Bouchard et al., 1982; Peter and Weber, 1985; Jury et al., 1987; Weed et al., 1995). The influence of soil microbiological activity on the degradation rate of metribuzin was demonstrated (Sharom and Stephenson, 1976; Bouchard et al., 1982; Allen and Walker, 1988). Some toxic effects of metribuzin toward soil microflora have been described (Evans et al., 1965; Hill and Stratton, 1991; Junnila et al., 1993). The partial transformation of metribuzin by a strain of Streptomyces was reported by Shelton et al. (1996). Fungal degradation was studied in five species by Schilling et al. (1985).

Some preliminary assays were conducted on solid media to assess a potential toxic effect of metribuzin towards fungi. The strains were grown on solid media: MEA and GS–agar (1.5%). Results indicated the lack of toxicity of this compound up to the maximal final concentration tested of 500 mg/L (data not shown). The results obtained in liquid medium showed that only a few species (B. cinerea from NCS and S. superba and A. fusca from CS) decreased the level of the compound by more than 50% after 5 d (Table 2). However, the majority of the species were able to deplete partly the compound (between 15–40%). They could be potentially interesting after optimization of cultural conditions and a longer time of culture since Schilling et al. (1985) obtained a total transformation by Rhizopus japonicus and Cunninghamella echinulata and a depletion range of 27 to 45% with Aspergillus niger, Penicillium lilacinum, and Fusarium oxysporum after a 4-wk incubation period. Phanerochaete chrysosporium was considered highly efficient in numerous studies dealing with biodegradation or biotransformation of environmentally persistent pollutants (review by Aust, 1990). It was demonstrated to biotransform atrazine, one of the most widely used triazines (Mougin et al., 1994). Our results showed that this species is not among the best concerning the depletion of metribuzin (21% only); this observation was already made in the course of investigations about the degradation of other phenylureas or pentachlorophenols by fungi (Steiman et al., 1994; Vroumsia et al., 1996; Khadrani et al., 1999). One of the explanations could be that in our studies, P. chrysosporium was grown at 22°C like most other species, while most published assays were conducted at 37°C (optimal growth temperature for this fungus) although not reflecting the real conditions of natural ecosystems.

Only a few reports were published on metamitron, another triazine herbicide (Fig. 1). It was supposed to have the same behavior in soil as metribuzin, and it may also be found in ground water (Spliid and Koppen, 1998). A slight inhibition of soil biomass-related activities was observed with metamitron (Engelen et al., 1998). However, an inhibitory effect on nitrifying bacteria and on nitrification was reported by Gadkari (1985). Parekh et al. (1994) reported the high efficiency of a Rhodococcus sp. isolated from a treated soil. Nothing was available in the literature concerning degradation by fungi. The present results indicated that this compound was particularly resistant to fungal degradation or transformation. Among the 21 species tested, only Alternaria solani (from NCS) and the two strains of Drechslera australiensis and Absidia fusca (from CS) were able to reduce the concentration in the medium more than 10% (10–16%) (Table 2).

Linuron or methoxydiuron is a phenylurea, used as a selective pre- and postemergence herbicide (Fig. 1). This compound was also shown to modify the soil microbial community pattern (El Fantroussi et al., 1999). Shelton et al. (1996) reported a transformation exceeding 50% of linuron by a strain of Streptomyces. The adsorption of linuron by microorganisms (Aspergillus flavus and Pseudomonas aurantiaca) and its transformation to 3,4-dichloroaniline have been reported (Funtikova, 1979; Funtikova and Surovtseva, 1979). In the present work, 12 species were grown in a liquid medium with linuron as carbon source. Seven of them did not modify the initial concentration of the compound. The two strains of S. macrospora yielded a 22 to 25% reduction, while B. cinerea almost completely depleted the medium. An induction of the removal capacity seemingly due to soil contamination was observed for A. parasiticus, A. fusca, and to a lesser extent for F. solani.

The second phenylurea herbicide investigated was metobromuron (Fig. 1). This preemergence herbicide is absorbed through the plant roots and translocated in the transpiration stream to the leaves, where it interferes with photosynthesis (Park and Hamill, 1993). Talaromyces wortmanii and Fusarium oxysporum were reported to degrade respectively 37 and 11% of the metobromuron added to the culture medium (10 mg/L) after 18 d of incubation (Tweedy et al., 1970). In the present work, among the 31 strains assayed for their ability to degrade or transform metobromuron, Botrytis cinerea gave very interesting results since the two strains (from CS and NCS) almost completely depleted the compound. Rhizopus oryzae and Absidia fusca from CS removed respectively 40 and 47% of the compound.

Previous work with diuron, a bichlorinated phenylurea, showed that this compound was very resistant to fungal attack, while the monochlorinated herbicide chlortoluron was more easily metabolized, with the best results being obtained with isoproturon, devoided of chlorine (Vroumsia et al., 1996; Khadrani et al., 1999). In the present work, the same observation was made, since among the 11 species assayed both with linuron and metobromuron, 9 exhibited lower potentialities toward the chlorinated compound linuron than toward metobromuron, which does not contain chlorine.

On the whole, the HPLC profiles showed the presence of metabolites in culture medium extracts from all efficient fungi, as illustrated by Fig. 2 and 3. This indicates that the herbicide depletion observed is at least partly due to a biotransformation of the compounds. Figure 2A,B gives HPLC profiles obtained when growing Absidia fusca and Botrytis cinerea, from NCS and CS, in the presence of metribuzin and metamitron. Concerning metribuzin, the same major peaks were found for both species, regardless of their origin. The most abundant compound had a retention time (RT) of 1.39 to 1.45 min, the same was probably also present after transformation of metamitron by A. fusca. It must be noted that B. cinerea was unable to transform metamitron. For metobromuron (Fig. 3A), the main compounds produced had RT of 1.42 to 1.50, 2.67 to 2.70, and 2.85 to 2.88 min. The first was the most abundant in extracts from A. fusca culture medium, while B. cinerea produced mostly the others. These compounds were also seemingly obtained during linuron transformation (Fig. 3B).


   CONCLUSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS AND DISCUSSION
 CONCLUSION
REFERENCES
 
To our knowledge, this is the first report dealing with the bioremediation of metribuzin, metamitron, metobromuron, and linuron contaminated media by a selection of soil fungi from different taxonomic groups. The majority of HPLC profiles revealed the appearance of new peaks when the herbicides disappeared, indicating their biotransformation by fungi; however, from our results it cannot be excluded that another process (such as bioaccumulation) also may be involved. These points will be studied on the most efficient strains isolated. Sordaria superba, Aspergillus parasiticus, the three species of Absidia, and the two species of Rhizopus were interesting because of the probable inducibility of their degrading systems. Absidia fusca from CS was particularly efficient and exhibited a wide spectrum of activity. Further studies to compare strains from CS and strains from NCS will be conducted. Moreover, the degradation or biotransformation of xenobiotics by this species was never reported in the literature. Botrytis cinerea was very efficient whatever its origin, even on a resistant compound such as linuron. This species will also be submitted to detailed studies.


   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS AND DISCUSSION
 CONCLUSION
 REFERENCES
 





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