Metagenomic Study and Biodegrading Capability of Bacterial Community in Monocrotophos Treated Tea Soil

Abstract

Aims: The study was undertaken to reveal the diversity of bacteria in organophosphate (monocrotophos) pesticide-treated tea soil to provide new insights on monocrotophos degrading bacterial community. Study Design: A metagenomic study of monocrotophos treated and untreated soil to isolate and identify pesticide-degrading microflora. Place and Duration of Study: Tea soil was collected from Borbhetta Tea Estate of Tocklai Tea Research Institute, Jorhat, Assam and the experiments were carried out from December 2015 to December 2017. Methodology: Tea soil was enriched up to 300 ppm of monocrotophos for four weeks (TREATED sample) and 16S rRNA V3 region gene amplicon metagenomic sequencing was carried out on untreated (CONTROL) and spiked (TREATED) soil. The treated soil was cultured in mineral salt medium up to 600 ppm of monocrotophos and bacterial growth, and degrading capacity was studied for three isolated bacterial species at three different pH and identified by sequencing the 16s rRNA region. The bacterial species in the metagenome were also compared and grouped with the bacterial species in NCBI database based on the presence and absence of organophosphate hydrolase (OPH) gene. Results: Metagenomic sequencing revealed the presence of 20 bacterial phyla distributed across 119 families and 433 genera. 147559 sequences remained taxonomically unclassified suggesting the presence of unique undescribed bacteria. Pesticide-contaminated tea soil was mostly dominant with Acidobacteria, Actinobacteria, Firmicutes, Proteobacteria, Bacteroidetes, Nitrospirae and Verrucomicrobia phyla. The number of species observed in the control and treated soil was 1036 and 910 respectively. Three species were isolated and characterised from the TREATED soil in mineral salt medium with pH 5 to 7 and their monocrotophos degradation was determined by UV-Vis microplate reader and HPLC at 2-time points. Conclusion: Organophosphate-degrading bacteria namely, Pseudomonas pseudoalcaligenes, Pseudomonas fluorescence, Pseudomonas putida, Serratia species, Cupriavidis metallidurans, Burkholderia species, Achromobacter xylosoxidans, Achromobacter species, Sphingomonas species, Ochrobactrum gallinifaecis, and Brucella species were present in increased numbers in the treated sample. 52.2% monocrotophos degradation was observed in Serratia fonticola in 48 hrs in acidic pH of 5.