Soil bacteria showing a potential of chlorpyrifos degradation and plant growth enhancement

ABSTRACT Background: Since 1960s, the organophosphate pesticide chlorpyrifos has been widely used for the purpose of pest control. However, given its persistence and toxicity towards life forms, the elimination of chlorpyrifos from contaminated sites has become an urgent issue. For this process bioremediation is the method of choice. Results: Two bacterial strains, JCp4 and FCp1, exhibiting chlorpyrifos-degradation potential were isolated from pesticide contaminated agricultural fields. These isolates were able to degrade 84.4% and 78.6% of the initial concentration of chlorpyrifos (100 mg L-1) within a period of only 10 days. Based on 16S rRNA sequence analysis, these strains were identified as Achromobacter xylosoxidans (JCp4) and Ochrobactrum sp. (FCp1). These strains exhibited the ability to degrade chlorpyrifos in sterilized as well as non-sterilized soils, and were able to degrade 93-100% of the input concentration (200 mg kg-1) within 42 days. The rate of degradation in inoculated soils ranged from 4.40 to 4.76 mg-1 kg-1 d-1 with rate constants varying between 0.047 and 0.069 d-1. These strains also displayed substantial plant growth promoting traits such as phosphate solubilization, indole acetic acid production and ammonia production both in absence as well as in the presence of chlorpyrifos. However, presence of chlorpyrifos (100 and 200 mg L-1) was found to have a negative effect on indole acetic acid production and phosphate solubilization with percentage reduction values ranging between 2.65-10.6% and 4.5-17.6%, respectively. Plant growth experiment demonstrated that chlorpyrifos has a negative effect on plant growth and causes a decrease in parameters such as percentage germination, plant height and biomass. Inoculation of soil with chlorpyrifos-degrading strains was found to enhance plant growth significantly in terms of plant length and weight. Moreover, it was noted that these strains degraded chlorpyrifos at an increased rate (5.69 mg-1 kg-1 d-1) in planted soil. Conclusion The results of this study clearly demonstrate that the chlorpyrifos-degrading strains have the potential to develop into promising candidates for raising the productivity of crops in pesticide contaminated soils.

Soil bacteria showing a potential of chlorpyrifos degradation and plant growth enhancement.

BACKGROUND: Since 1960s, the organophosphate pesticide chlorpyrifos has been widely used for the purpose of pest control. However, given its persistence and toxicity towards life forms, the elimination of chlorpyrifos from contaminated sites has become an urgent issue. For this process bioremediation is the method of choice. RESULTS: Two bacterial strains, JCp4 and FCp1, exhibiting chlorpyrifos-degradation potential were isolated from pesticide contaminated agricultural fields. These isolates were able to degrade 84.4% and 78.6% of the initial concentration of chlorpyrifos (100mgL(-1)) within a period of only 10 days. Based on 16S rRNA sequence analysis, these strains were identified as Achromobacter xylosoxidans (JCp4) and Ochrobactrum sp. (FCp1). These strains exhibited the ability to degrade chlorpyrifos in sterilized as well as non-sterilized soils, and were able to degrade 93-100% of the input concentration (200mgkg(-1)) within 42 days. The rate of degradation in inoculated soils ranged from 4.40 to 4.76mg(-1)kg(-1)d(-1) with rate constants varying between 0.047 and 0.069d(-1). These strains also displayed substantial plant growth promoting traits such as phosphate solubilization, indole acetic acid production and ammonia production both in absence as well as in the presence of chlorpyrifos. However, presence of chlorpyrifos (100 and 200mgL(-1)) was found to have a negative effect on indole acetic acid production and phosphate solubilization with percentage reduction values ranging between 2.65-10.6% and 4.5-17.6%, respectively. Plant growth experiment demonstrated that chlorpyrifos has a negative effect on plant growth and causes a decrease in parameters such as percentage germination, plant height and biomass. Inoculation of soil with chlorpyrifos-degrading strains was found to enhance plant growth significantly in terms of plant length and weight. Moreover, it was noted that these strains degraded chlorpyrifos at an increased rate (5.69mg(-1)kg(-1)d(-1)) in planted soil. CONCLUSION: The results of this study clearly demonstrate that the chlorpyrifos-degrading strains have the potential to develop into promising candidates for raising the productivity of crops in pesticide contaminated soils.

Bacterial community analysis of cypermethrin enrichment cultures and bioremediation of cypermethrin contaminated soils.

Cypermethrin is widely used for insect control; however, its toxicity toward aquatic life requires its complete removal from contaminated areas where the natural degradation ability of microbes can be utilized. Agricultural soil with extensive history of CM application was used to prepare enrichment cultures using cypermethrin as sole carbon source for isolation of cypermethrin degrading bacteria and bacterial community analysis using PCR-DGGE of 16 S rRNA gene. DGGE analysis revealed that dominant members of CM enrichment culture were associated with α-proteobacteria followed by γ-proteobacteria, Firmicutes, and Actinobacteria. Three potential CM-degrading isolates identified as Ochrobactrum anthropi JCm1, Bacillus megaterium JCm2, and Rhodococcus sp. JCm5 degraded 86-100% of CM (100 mg L(-1) ) within 10 days. These isolates were also able to degrade other pyrethroids, carbofuran, and cypermethrin degradation products. Enzyme activity assays revealed that enzymes involved in CM-degradation were inducible and showed activity when strains were grown on cypermethrin. Degradation kinetics of cypermethrin (200 mg kg(-1)) in soils inoculated with isolates JCm1, JCm2, and JCm5 suggested time-dependent disappearance of cypermethrin with rate constants of 0.0516, 0.0425, and 0.0807 d(-1), respectively, following first order rate kinetics. The isolated bacterial strains were among dominant genera selected under CM enriched conditions and represent valuable candidates for in situ bioremediation of contaminated soils and waters.

Determination of cypermethrin degradation potential of soil bacteria along with plant growth-promoting characteristics.

The pyrethroid insecticide cypermethrin is in extensive use since 1980s for insect control. However, its toxicity toward aquatic animals and humans requires its complete removal from contaminated areas that can be done using indigenous microbes through bioremediation. In this study, three bacterial strains isolated from agricultural soil and identified as Acinetobacter calcoaceticus MCm5, Brevibacillus parabrevis FCm9, and Sphingomonas sp. RCm6 were found highly efficient in degrading cypermethrin and other pyrethroids. These bacterial strains were able to degrade more than 85 % of cypermethrin (100 mg L(-1)) within 10 days. Degradation kinetics of cypermethrin (200 mg kg(-1)) in soils inoculated with isolates MCm5, FCm9, and RCm6 suggested time-dependent disappearance of cypermethrin with rate constants of 0.0406, 0.0722, and 0.0483 d(-1) following first-order rate kinetics. Enzyme assays for Carboxylesterase, 3-PBA dioxygenase, Phenol hydroxylase, and Catechol-1,2 dioxygenase showed higher activities with cypermethrin treated cell-free extracts compared to non-treated cell-free extracts. Meanwhile, SDS-PAGE analysis showed upregulation of some bands in cypermethrin-treated cells. This might suggest that cypermethrin degradation in these strains involves inducible enzymes. Besides, the isolates displayed substantial plant growth-promoting traits such as phosphate solubilization, Indole acetic acid production, and ammonia production. Implying the efficient biodegradation potential along with multiple biological properties, these isolates can be valuable candidates for the development of bioremediation strategies.

Bacterial community analysis in chlorpyrifos enrichment cultures via DGGE and use of bacterial consortium for CP biodegradation.

The organophosphate pesticide chlorpyrifos (CP) has been used extensively since the 1960s for insect control. However, its toxic effects on mammals and persistence in environment necessitate its removal from contaminated sites, biodegradation studies of CP-degrading microbes are therefore of immense importance. Samples from a Pakistani agricultural soil with an extensive history of CP application were used to prepare enrichment cultures using CP as sole carbon source for bacterial community analysis and isolation of CP metabolizing bacteria. Bacterial community analysis (denaturing gradient gel electrophoresis) revealed that the dominant genera enriched under these conditions were Pseudomonas, Acinetobacter and Stenotrophomonas, along with lower numbers of Sphingomonas, Agrobacterium and Burkholderia. Furthermore, it revealed that members of Bacteroidetes, Firmicutes, α- and γ-Proteobacteria and Actinobacteria were present at initial steps of enrichment whereas ß-Proteobacteria appeared in later steps and only Proteobacteria were selected by enrichment culturing. However, when CP-degrading strains were isolated from this enrichment culture, the most active organisms were strains of Acinetobacter calcoaceticus, Pseudomonas mendocina and Pseudomonas aeruginosa. These strains degraded 6-7.4 mg L(-1) day(-1) of CP when cultivated in mineral medium, while the consortium of all four strains degraded 9.2 mg L(-1) day(-1) of CP (100 mg L(-1)). Addition of glucose as an additional C source increased the degradation capacity by 8-14 %. After inoculation of contaminated soil with CP (200 mg kg(-1)) disappearance rates were 3.83-4.30 mg kg(-1) day(-1) for individual strains and 4.76 mg kg(-1) day(-1) for the consortium. These results indicate that these organisms are involved in the degradation of CP in soil and represent valuable candidates for in situ bioremediation of contaminated soils and waters.