Dileep K. Singh

Biodegradation and bioremediation of pesticide in soil: concept, method and recent developments

Biodegradation is a natural process, where the degradation of a xenobiotic chemical or pesticide by an organism is primarily a strategy for their own survival. Most of these microbes work in natural environment but some modifications can be brought about to encourage the organisms to degrade the pesticide at a faster rate in a limited time frame. This capability of microbe is some times utilized as technology for removal of contaminant from actual site. Knowledge of physiology, biochemistry and genetics of the desired microbe may further enhance the microbial process to achieve bioremediation with precision and with limited or no scope for uncertainty and variability in microbe functioning. Gene encoding for enzyme has been identified for several pesticides, which will provide a new inputs in understanding the microbial capability to degrade a pesticide and develop a super strain to achieve the desired result of bioremediation in a short time.

Biodegradation of α and β endosulfan by Aspergillus sydoni

The biodegradation of endosulfan and the metabolites formed were studied using fungi both in broth culture as well as in soil microcosm. Fungal strains were isolated from soil and grown in broth Czapek-dox medium. The strain which utilized endosulfan and showed maximum growth was selected for detailed studies. Maximum degrading capability in shake flask culture was shown by Aspergillus sydoni which degraded 95% of endosulfan alpha and 97% of endosulfan beta in 18 d of incubation. Soil microcosm study was also carried out using this strain in six different treatments. Endosulfan sulfate was the main metabolite formed along with small quantity of endosulfan ether and endosulfan lactone both in broth culture and soil microcosm. This isolated fungal strain will be a potential source for endosulfan degrading enzymes and can be used for bioremediation at the contaminated sites.

Biodegradation of endosulfan and endosulfan sulfate by Achromobacter xylosoxidans strain C8B in broth medium

Endosulfan is one of the most widely used wide spectrum cyclodiene organochlorine insecticide. In environment, endosulfan can undergo either oxidation or hydrolysis reaction to form endosulfan sulfate and endosulfan diol respectively. Endosulfan sulfate is as toxic and as persistent as its parent isomers. In the present study, endosulfan degrading bacteria were isolated from soil through selective enrichment technique using sulfur free medium with endosulfan as sole sulfur source. Out of the 8 isolated bacterial strains, strain C8B was found to be the most efficient endosulfan degrader, degrading 94.12% α-endosulfan and 84.52% β-endosulfan. The bacterial strain was identified as Achromobacter xylosoxidans strain C8B on the basis of 16S rDNA sequence similarity. Achromobacter xylosoxidans strain C8B was also found to degrade 80.10% endosulfan sulfate using it as sulfur source. No known metabolites were found to be formed in the culture media during the entire course of degradation. Besides, the bacterial strain was found to degrade all the known endosulfan metabolites. There was marked increase in the quantity of released CO2 from the culture media with endosulfan as sulfur source as compared to MgSO4 suggesting that the bacterial strain, Achromobacter xylosoxidans strain C8B probably degraded endosulfan completely through the formation of endosulfan ether.

INSECTICIDE RESIDUES IN COTTON CROP SOIL

Dimethoate, monocrotophos, triazophos, deltamethrin, cypermethrin and endosulfan were applied to a cotton crop soil located at Nurpur village, Punjab, India. The insecticides were applied sequentially at recommended dosages in cotton fields by foliar application in 1995, 1996 and 1998. Soil samples were collected from the cotton crop farms and extracted with acetone. The extracted material was analysed by a gas liquid chromatograph (GLC) equipped with an 63Ni electron-capture detector (ECD-63Ni). Recovery data was obtained by fortifying soil with insecticide. The average recoveries from the fortified soil samples were 76–92% for organophosphorous compounds and 90 –98% for synthetic pyrethroids and organochlorines. The results showed that the insecticide residues under study were present in the range of 1.16 to 41.97 ng g 1 d.wt.soil. The pattern of dissipation of the insecticides used was similar for the duration of the crop. Half lives of the insecticides ranged from 7 to 22 days. Except endosulfan none of the other insecticides used were leached below 15 cm. Endosulfan was found to be rapidly degraded in the soil and formed a sulfate metabolite. Persistence and dissipation pattern in soils with history of exposure to the insecticide compared to non-history soils were similar.

Bacterial, Azotobacter, Actinomycetes, and Fungal Population in Soil after Diazinon, Imidacloprid, and Lindane Treatments in Groundnut (Arachis hypogaea L.) Fields

Bacterial, azotobacter, actinomycetes, and fungal populations were determined in groundnut (Arachis hypogaea L.) fields between July and November for three consecutive years (1997–1999) after insecticide treatments. Diazinon was applied for both seed and soil treatments. However, imidacloprid and lindane were used for seed treatments. An average half-life (t 1/2) of diazinon in seed- and soil-treated fields was found to be 29.32 and 34.87 days, respectively. Its residues were found for 60 days in both cases. In diazinon seed treatment, an increase in azotobacter, fungi, and actinomycetes populations was observed in samples from the 15th and 30th days, and this trend continued until crop harvest. However, the bacterial population had not been affected by this treatment. The diazinon soil treatment had indicated some significant adverse effects on fungi and actinomycetes population, which recovered after 30 days. The population of bacteria and azotobacter increased significantly in this treatment. The residues of imidacloprid and lindane were found for 90 and 120 days with an average half-life of 40.9 and 53.3 days, respectively. Imidacloprid had no significant effect on fungi and actinomycetes populations up to 15 days, and between 15 to 60 days some adverse effects were indicated. However, some significant increases in bacterial and azotobacter population were observed. Lindane had no effect on bacterial and fungal population. However, its adverse effects were observed in actinomycetes and azotobacter populations between 30 to 60 days.

Endosulfan and Quinalphos Residues and Toxicity to Soil Microarthropods after Repeated Applications in a Field Investigation

Endosulfan (1,4,5,6,7,7-hexachloro-8,9,10-trinorborn-5-en-2,3-ylenedimethylsulphite) and quinalphos (O,O-diethyl O-quinoxalin-2-yl phosphorothioate) persistence and their effect on soil microarthropods were studied after repeated applications in cotton fields. Dissipation behavior of insecticides after repeated applications was observed from 78 to 292 days after the first insecticide treatment. At any given time the concentrations of endosulfan β residues were always higher as compared to endosulfan α. From 78 to 85 days, 5.0% and 20.4% decrease in α and β endosulfan residues was observed, respectively. Endosulfan β isomer decreased up to 93.0% in 292 days. Endosulfan sulfate was detected as a major metabolite in the soil samples. Total endosulfan residues decreased by 86.6% from 78 to 292 days. The amounts of quinalphos residues were less as compared to endosulfan at any given time. The residues observed after 78 days of application were 0.88 ng g−1 d wt. soil. At the end of 145 days, a 35.0% decrease in quinalphos residue was observed, which decreased further by 50.9% in 292 days. Among the soil microarthropods studied, Acarina was more sensitive to the applied insecticides as compared to Collembola. Three days after the last treatment, up to 94.5% (p < 0.01) and 71.2% (p < 0.05) decrease in Acarina population was observed in endosulfan and quinalphos treated fields, respectively, compared to control field. In general, no noticeable change in Collembola population was observed after the insecticide treatments.

Synthesis and photophysical properties of β-triazole bridged porphyrin–coumarin dyads

Novel copper(II) β-triazole bridged porphyrin–coumarin conjugates have been synthesized in good to excellent yields via a copper(I)-catalyzed Huisgen 1,3-dipolar cycloaddition reaction of copper(II) 2-azido-5,10,15,20-tetraphenylporphyrin with various alkyne-substituted coumarins. These newly prepared copper complexes of β-triazole-bridged porphyrin–coumarin dyads were successfully demetallated in the presence of conc. H2SO4 at 0 °C to afford the corresponding free-base porphyrins which on zinc insertion by using Zn(OAc)2·2H2O in a chloroform–methanol mixture at 25 °C produced zinc(II) β-triazole bridged porphyrin–coumarin dyads in good yields. The synthesized products were characterized spectroscopically and evaluated for their photophysical properties by using UV-Vis and fluorescence spectroscopy. The preliminary results revealed a significant intramolecular energy transfer from the coumarin subunit to the porphyrin moiety in the case of some β-triazole bridged porphyrin–coumarin dyads.

Effect of UV radiation and temperature on mineralization and volatilization of coumaphos in water.

This study was undertaken to determine the dissipation and degradation of coumaphos [O-(3-chloro-4-methyl-2-oxo-2H-1-benzopyran-7-yl) O,O-diethyl phosphorothioate] under different sunlight conditions and at different temperatures. The effect of the ultra violet (UV) component of solar radiation was also studied using quartz tubes in addition to other radiation in the visible range using glass tubes and the results were compared with those obtained under the dark light conditions. Water suspensions of coumaphos were incubated at three temperatures viz. 22°C, 37°C and 53°C in closed systems to study the effect of temperature. Volatilization, mineralization and degradation of coumaphos increased with an increase in temperature and exposure to solar radiation, particularly under the UV component of the solar radiation. Major loss of the pesticide occurred through volatilization. The optimum temperature for the degradation of coumaphos was found to be at 37°C. The data obtained from the mineralization and degradation studies indicated that 53°C crosses the biological range for suitable growth of microorganism. UV radiation exposure along with maintaining temperature at 37°C may prove useful in the dissipation and/or degradation of coumaphos prior to its disposal as waste from cattle dipping vats.

Comparative performance evaluation of multi-metal resistant fungal strains for simultaneous removal of multiple hazardous metals

In the present study, five fungal strains viz., Aspergillus terreus AML02, Paecilomyces fumosoroseus 4099, Beauveria bassiana 4580, Aspergillus terreus PD-17, Aspergillus fumigatus PD-18, were screened for simultaneous multimetal removal. Highest metal tolerance index for each individual metal viz., Cd, Cr, Cu, Ni, Pb and Zn (500mg/L) was recorded for A. fumigatus for the metals (Cd, 0.72; Cu, 0.72; Pb, 1.02; Zn, 0.94) followed by B. bassiana for the metals (Cd, 0.56; Cu, 0.14; Ni, 0.29; Zn, 0.85). Next, the strains were exposed to multiple metal mixture (Cd, Cr, Cu, Ni, Pb and Zn) of various concentrations (6, 12, 18, 30mg/L). Compared to other strains, B. bassiana and A. fumigatus had higher cube root growth (k) constants indicating their better adaptability to multi metal stress. After 72h, multimetal accumulation potential of B. bassiana (26.94±0.07mg/L) and A. fumigatus (27.59±0.09mg/L) were higher than the other strains at initial multimetal concentration of 30mg/L. However, considering the post treatment concentrations of individual metals in multimetal mixture (at all the tested concentrations), A. fumigatus demonstrated exceptional performance and could bring down the concentrations of Cd, Cu, Ni, Pb and Zn below the threshold level for irrigation prescribed by Food and Agriculture Organization (FAO).

Ammonium, Nitrate and Nitrite Nitrogen and Nitrate Reductase Enzyme Activity in Groundnut (Arachis hypogaea L.) Fields After Diazinon, Imidacloprid and Lindane Treatments

Impacts of diazinon (O,O-diethyl O-2-isopropyl-6-methylpyrimidin-4-yl phosphorothioate), imidacloprid [1-(6-chloro-3-pyridylmethyl)-N-nitroimidazolidin-2-ylideneamine] and lindane (1,2,3,4,5.6-hexachlorocyclohexane) treatments on ammonium, nitrate, and nitrite nitrogen and nitrate reductase enzyme activities were determined in groundnut (Arachis hypogaea L.) field for three consecutive years (1997 to 1999). Diazinon was applied for both seed- and soil-treatments but imidacloprid and lindane were used for seed treatments only at recommended rates. Diazinon residues persisted for 60 days in both the cases. Average half-lives (t1/2) of diazinon were found 29.3 and 34.8 days respectively in seed and soil treatments. In diazinon seed treatment, NH4 +, NO3 −, and NO2 − nitrogen and nitrate reductase activity were not affected. Whereas, diazinon soil treatment indicated significant increase in NH4 +-N in a 1-day sample, which continued until 90 days. Some declines in NO3 −N were found from 15 to 60 days. Along with this decline, significant increases in NO2 −N and nitrate reductase activity were found between 1 and 30 days. Imidacloprid and lindane persisted for 90 and 120 days with average half-lives (t1/2) of 40.9 and 53.3 days, respectively. Within 90 days, imidacloprid residues lost by 73.17% to 82.49% while such losses for lindane residues were found 78.19% to 79.86 % within 120 days. In imidacloprid seed-treated field, stimulation of NO3 −N and the decline in NH4 +NO2 −-N and nitrate reductase enzyme activity were observed between 15 to 90 days. However, lindane seed treatment indicated significant increases in NH4 +-N, NO2 −-N and nitrate reductase activity and some adverse effects on NO3 −N between 15 and 90 days.