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International Journal of Advanced Life Sciences (IJALS) 

Int. j. Adv. Lif. Sci., Available online on at www. 

Int. J. Adv. Lif. Sci., Available online on at www. ijals.com 

ISSN 

2277 – 758X 

Umamaheswari et al., IJALS, Vol.3. May - 2012. RESEARCH ARTICLE 

Introduction 

Studies on biodegradation of pesticides contaminants soil by using 

Tremetes versicolor and Phanerochaete chrysosporium 

G. Umamaheswari * , V. Ramamurthy ** , S. Raveendran *** , K. Radhika *** and A. Kavitha Amirthanayagi *** 

* Post Graduate and Research Department of Biotechnology, Marudupandiyar College, Vallam, 

Thanjavur – 403. Tamil Nadu. ** Post Graduate and Research Department of Biochemistry, 

Marudupandiyar College, Vallam, Thanjavur - 403, Thanjavur District, Tamil Nadu. 

*** P.G & Research Department of Zoology, Khadir Mohideen College, Adirampattinam - 614 701, 

Thanjavur District, Tamil Nadu. 

Email : v.ramamoorthy07@gmail.com 

Corresponding Author 

V. Ramamurthy 

Post Graduate and Research 

Department of Biochemistry, 

Marudupandiyar College, 

Vallam, Thanjavur - 613 403, 

E-mail: v.ramamoorthy07@gmail.com 

Article History 

Received on 12 March, 2012; 

Revised in revised form 4 April, 

2012; Accepted 22 April, 2012 

Abstract 

Currently more number of possible mechanisms 

is available to clean up of pesticides in soil, viz 

chemical treatment, volatilization and incineration. 

Chemical treatment and volatilization are feasible and 

are problematic as large volumes of acids and alkalis 

are produced and subsequently must be disposed of by 

incineration, which is a very reliable method for 

destruction of these compounds, has met serious 

public opposition, because of its potentially toxic 

emissions, and its elevated economic costs (Kearney 

and Wauchope, 1998; Zhang and Chiao, 2002). Overall 

most of these physico-chemical cleaning technologies 

Among the various soil contaminants, pesticides are of primary 

importance due to their continuous entry into the soil environment. However, in 

un-sterilized and untreated soils, pesticide degradation rate was at the medium 

when compared to the sterilized soil indicating that the soil microorganisms play 

an important role in pesticide degradation. The pesticides were reduced in their 

level after the treatment of bacterial isolates. Totally three different pesticide such 

as simazine, trifluralin and dieldrin was recorded in soil extract through HPLC. 

Among the pesticide simazine was recorded maximum than trifluralin and 

dieldrin. The results obtained in this study provide valuable knowledge on the 

abilities of T.versicolor and P.chrysosporium might serve as a sound basis for the 

further exploitation of these species as fungal inoculants in biological remediation 

processes. 

Keywords: Pesticides, simazine, trifluralin, dieldrin, Tremetes versicolor and 

Phanerochaete chrysosporium 

are expensive and rather inefficient (Nerud et al., 2003 

and Schoefs et al., 2004) because the contaminated 

soil has to be excavated at a site and moved to a storage 

area where it can be processed. Due to environmental 

concerns associated with the accumulation of pesticides 

in food products and water supplies there is a great 

need to develop safe, convenient and economically 

feasible methods for pesticide remediation (Zhang and 

Chiao, 2002). For this reason several biological 

techniques involving biodegradation of organic 

compounds by microorganisms have been developed 

(Schoefs et al., 2004). 

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The use of microorganisms (fungi or bacteria), 

either naturally occurring or introduced, to degrade 

pollutants is called bioremediation (Pointing, 2001). 

Microbial metabolism is probably the most important 

pesticide degradation process in soils (Kearney and 

Wauchope, 1998) and is the basis for bioremediation, 

as the degrading microorganisms obtain carbon, 

nitrogen or energy from the pesticide molecules (Gan 

and Koskinen, 1998). The goal of bioremediation is to 

at least reduce pollutant levels to undetectable, nontoxic 

or acceptable levels, i.e. within limits set by regulatory 

agencies (Pointing, 2001) or ideally completely mineralize 

organ is pollutants to carbon dioxide. From an 

environmental point of view this total mineralization is 

desirable as it represents complete detoxification (Gan 

and Koskinen, 1998). The use of bioremediation to 

remove pollutants is typically less expensive than the 

equivalent physico-chemical methods. This technology 

offers the potential to treat contaminated soil and 

groundwater on site without the need for excavation 

(Balba et al., 1998), it requires little energy input and 

preserves the soil structure (Hohener et al., 1998). 

Perhaps the most attractive feature of bioremediation is 

the reduced impact on the natural ecosystems, which 

should be more acceptable to the public. 

Application of fungal technology for the 

cleanup of contaminants has shown promise since 1985 

when the white rot species Phanerochaete chrysosporium 

was found to be able to metabolize a number of 

important environmental pollutants. This ability is 

generally attributed to the lignin degrading enzymatic 

system of the fungus and a similar degrading capacity 

was later described for other white rot fungal species 

(Sasek, 2003). An environmental factor that may well 

have a crucial effect on bioremediation is soil water 

availability, as it varies naturally throughout the 

year. Nevertheless, very few studies look at the effects 

of water availability on bioremediation. This study was 

aimed to isolate the fungi from pesticide-contaminated 

soil and to determine pesticide degradation rates in soil 

extract liquid broth after the incubation with fungal 

inoculation. 

Materials and Methods 

Collection of soil samples 

For this present study the soil samples were 

collected from agricultural field of Kurichi village, 

Pattukkottai taluk, Thanjavur District, Tamilnadu, 

India. The soil samples were collected from the study 

site at a depth within 15 cm from the surface of the soil 

at random during the study period. The collected soil 

samples were brought to the laboratory in sterilized 

polythene bags, handpicked, air dried and stored in 

containers for future use. 

Isolation of fungi 

One gram of the soil sample was suspended in 

100 ml of sterilized Minimal media, without sample 

considered as control. Both were incubated at 37 0 C for 

24 to 48 hrs. 0.1 ml of inoculum was transferred to 

petriplate having PDA medium by spread plate 

technique. Population of fungi was isolated from the 

soil samples by serial dilution technique. Fungi were 

identified by using standard manuals, viz Mannual of 

Soil Fungi (Gillman, 1947), Dematiaceous Hyphomycetes 

(Ellis, 1971). 

Physico-chemical properties of the soil 

Shallow, duplicate soil cores were collected 

from the study site with a piston corer (54.3 cm 3 , 12 cm 

long x 1.2 cm radius) and stored in watertight plastic 

bags until analysis of pH, nutrients and organic content. 

The soil sample was air dried and powdered to pass 

through a sieve (2 mm). They were collected in a plastic 

container, sealed and stored at 4°C for further use. 

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Determination of pH was done by a digital 

pH meter, electrical conductivity by a conductivity 

meter (Elico). The moisture content was determined 

after drying at 105°C for 24 hrs. Organic matter and 

organic carbon was determined titrimetrically (Walkley 

and Black, 1934) and expressed as percentage (%). The 

phosphorus content was estimated by Olsen et al., 

(1954). The nitrogen and potassium was estimated 

according to the method of Sankaram (1966). 

Micronutrients viz Zinc, Copper, Manganese and Iron 

were estimated by Lindsay and Norwell (1978). 

Inoculation with fungi, sampling and dry weight 

determination 

A soil extract liquid broth was used in this 

study. 100 ml of soil extract was taken in Erlenmeyer 

flasks (250 ml) to which four plugs of actively growing 

mycelium of Tremetes versicolor and Phanerochaete 

chrysosporium were inoculated in each flask. It was 

maintained at 27±1 o C, for 25 days with constant 

agitation at 150 rpm. After the incubation period the 

mycelium was filtered through whatman No. 1 filter 

paper and biomass determined by drying the mycelium 

for 48 hours at 80 o C. The fresh filtrate was frozen at 

20 o C and used later for pesticide quantification 

determination. 

Pesticide analysis 

The HPLC was used to determine the amount 

of simazine, trifluralin and dieldrin (Eliassy, 1997). 

Results 

The present study was undertaken to assess the 

distribution and occurrence of fungi from pesticides 

polluted soil, which were collected from agricultural 

field of Kurichi village, Pattukkottai, Thanjavur 

District, Tamilnadu, India. Totally 8 species of fungi 

belongs to 5 genus were isolated from the soil samples. 

Among them the genus Aspergillus was recorded with 

3 species viz., A. niger, A. terreus, and A. flavus followed 

by Tremetes with 2 species viz., T. socotrana and 

T.versicolor. The genus Phenerocheate, Cladosporium and 

Fusarium were recorded with single species each 

(Table - 1). 

The physico-chemical characteristics of soil 

samples of the study sites were given in the Table 2. 

The texture and colour of the soil was sandy and brown 

respectively in the soil samples. The pH and electrical 

conductivity values were reported viz., 7.8 and 0.32 

respectively in the soil samples. The percentage of 

organic matter was 0.56%. The macronutrients content 

in the study site were given in the Table - 2. 

The impact of the two fungal inoculants such 

as T. versicolor and P. chrysosporium on degradation 

of pesticides were observed and recorded for soil 

sample. Initially 3 different types of pesticides such 

as Simazine (5.1ppm), Trifluralin (3.34 ppm) and 

Dieldrin (1.76 ppm) were observed through HPLC. 

These pesticides were reduced in their level after the 

treatment of fungal isolates. Among the pesticides 

content the Simazine was reduced the maximum levels 

when compared to Trifluralin and Dieldrin. The fungi 

P. chrysosporium removed maximum amount of all 

pesticides than the T. versicolor in the present soil 

sample analysis (Table - 3). 

Discussion 

Successful bioremediation is dependent on an 

interdisciplinary approach involving such disciplines as 

microbiology, engineering, ecology, geology and 

chemistry (Boopathy, 2000). To evaluate the outcome 

of bioremediation it is critical to assess some 

microbiological and biochemical parameters all giving 

information on soil quality. It is difficult to choose 

which parameters are more reliable, as the relationship 

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between an individual biochemical property and the 

totalmicrobial activity is not always clear, in complex 

systems like soils where the microorganisms and 

processes involved in the degradation of organic 

compounds are highly diverse (Nannipieri et al., 1990). 

White rot fungi possess a number of advantages 

that can be exploited in bioremediation systems. Because 

key components of their lignin-degrading system are 

extracellular, these fungi can degrade insoluble 

chemicals such as lignin or an extremely diverse range 

of very persistent or toxic environmental pollutants 

(Barr and Aust, 1994). The mycelial growth habit is 

also advantageous as it allows rapid colonization of 

substrates, and hyphal extension enables penetration of 

soil reaching pollutants in ways that other organisms 

cannot do (Reddy and Mathew, 2001). This can 

maximise physical, mechanical and enzymatic contact 

with the surrounding environment (Maloney, 2001). In 

addition, these fungi use inexpensive and abundant 

lignocellulosic materials as a nutrient source. They can 

tolerate a wide range of environmental conditions, such 

as temperature, pH and moisture and do not require pre- 

conditioning to a particular pollutant, because their 

degradative system is induced by nutrient deprivation 

(Barr and Aust, 1994). 

In a more recent study by biological 

degradation of benzene and toluene by T.versicolor was 

analyzed and the biomass were determined by Demir 

(2004). Within an incubation period of 48 hrs, it was 

observed that, removal was completed after 4 hours 

when initial toluene concentration was 50 mg/l and was 

completed in 36 hrs when this was 300 mg/l. 

Biodegradation was completed by the end of the 4th hr 

at benzene concentrations of 50mgl -1 while it continued 

for 42 hrs at 300 mgl -1 . With the addition of veratryl 

alcohol, laccase inducers, to the basic feed medium, the 

operation of the enzyme system was enhanced and 

biodegradation completed in a shorter time period. 

Table - 1. Fungal flora in the soil sample 

Sl. No 

Name of the organism 

1 Aspergillus niger 

2 A. terreus 

3 A. flavus 

4 Phenerochete chrysosporium 

5 Tremetes socotrana 

6 T.versicolor 

7 Fusarium sp 

8 Cladosporium sp 

Table - 2. Physico-chemical characteristics of soil 

sample 

Sl.No Parameters Values 

1. Soils texture Sandy 

2. Colour Brown 

3. pH 7.8 

4. Ecdcm -1 0.32 

5. Organic carbon % 0. 56% 

6. Nitrogen (mg/g) 91.2 

7. Potassium (mg/g) 78.1 

8. Phosphorus (mg/g) 9.5 

9. Zinc (ppm) 0.93 

10. Copper (ppm) 0.36 

11. Iron (ppm) 3.23 

12. Manganese (ppm) 8.83 

The microbial population of a site 

contaminated with pesticides may be eliminated, 

significantly reduced or altered; but alternatively, 

microbes may adapt to the presence of toxic 

compounds and can survive by degrading them as some 

microorganisms can utilize pesticides as a nutrient 

source. As soil microorganisms are not equal resistant 

to xenobiotics, some of them are very sensitive and do 

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Table - 3. Analysis Pesticide level in soil extract by HPLC 

not grow when toxic compounds are present in high 

concentrations or constitute a low carbon and energy 

source, while other are able to adapt and grow well 

(Guirard et al., 2003). Han et al. (2004) studied the 

degradation of phenanthrene by T.versicolor and its 

laccase was purified. After 36 hrs incubation, about 

46 and 65% of 100mgl -1 of phenanthrene added in 

shaken and static fungal cultures were removed, 

respectively. Although the removal percentage was 

highest (76.7%) at 10mgl -1 of phenanthrene, the trans- 

formation rate was maximal (0.82mgh -1 ) at 100mgl -1 of 

phenanthrene in the fungal culture. When the purified 

laccase of T. versicolor reacted with phenanthrene, the 

compound was not transformed. Another interesting 

example of contaminant degradation and enzyme activity 

was in the study described by Barr and Aust (1994). 

They described cyanide to be quite toxic to spores of 

P.chrysosporium (50% inhibition of glucose metabolism 

at 2.6mgl -1 ). 

Min et al. (2001) reported that increasing 

concentrations of butachlor in soil enhanced the activity 

of dehydrogenase with the highest activity on the 

16 th day after application of 22mg/kg soil of but achlor. 

Baran et al. (2004) reported high dehydrogenase 

activity in soil contaminated with PAH. Felsot and 

Dzantor (1995) described the effect of alachlor and 

organic amendment on soil dehydrogenase activity 

and on pesticide degradation rates. Amendment of soil 

Control 

(ppm) 

with corn meal caused fasterdegradation of alachlor. 

At very high concentrations of alachlor (750mg/kg soil) 

dehydrogenase activities in amended soils surpassed 

levels in corresponding nopesticide controls after 

21days with coincident alachlor degradation >50% 

during the same period. They suggested that stimulation 

of microbial activity by addition of organic amendments 

might enhance co-metabolism of high concentrations of 

pesticides in soil. Totally three different pesticide such 

as simazine, trifluralin and dieldrin was recorded in soil 

extract through HPLC. Among the pesticide simazine 

was recorded maximum than trifluralin and dieldrin. 

The results obtained in this study provide valuable 

knowledge on the abilities of T.versicolor and 

P. chrysosporium might serve as a sound basis for the 

further exploitation of these species as fungal 

inoculants in biological remediation processes. 

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Corresponding Author : V. Ramamurthy, Post Graduate and Research Department of Biochemistry, 

Marudupandiyar College, Vallam, Thanjavur - 613 403, Thanjavur District, Tamil Nadu, India, E-mail : 

v.ramamoorthy07@gmail.com. 

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