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The aim was to isolate, characterize, and explore potentials of gut bacteria from the earthworm (Metaphire posthuma) and imply these bacteria for remediation of Cu(II) and Zn(II). An extracellular polymeric substance (EPS) producing gut bacteria (Bacillus licheniformis strain KX657843) was isolated and identified based on 16S rRNA sequencing and phylogenetic analysis. The strain showed maximum tolerance of 8 and 6 mM for Cu(II) and Zn(II) respectively. It removed 34.5% of Cu(II) and 54.4% of Zn(II) at 25 mg L after 72 and 96 h incubation respectively. The bacteria possessed a great potential to produce indole acetic acid (38.49 μg mL) at 5 mg mL L-tryptophan following 12 days incubation. The sterilized seeds of mung beans (Vigna radiata) displayed greater germination and growth under bacterium enriched condition. We observed that the bacterial strain phosphate solubilization ability with a maximum of 204.2 mg L in absence of Cu(II) and Zn(II). Endowed with biosurfactant property the bacterium exhibited 24% emulsification index. The bacterium offered significant potential of plant growth promotion, Cu(II) and Zn(II) removal, and as such this study is the first report on EPS producing B. licheniformis KX657843 from earthworm which can be applied as powerful tool in remediation programs of Cu(II) and Zn(II) contaminated sites.

The present study shows the potential of an extracellular polymeric substance (EPS) produced by Bacillus licheniformis strain KX657843 isolated from earthworm (Metaphire posthuma) gut in the sorption of Cu(II) and Zn(II) and in flocculation. After harvesting bacterial cells from sucrose supplemented denitrifying culture medium, the EPS was extracted following ethanolic extraction method. The Fourier Transform Infrared Spectroscopy (FTIR) and 1H and 13C Nuclear Magnetic Resonance (NMR) of EPS revealed its functional groups, electronegative constituents, unsaturated carbon, and carbonyl groups. The negatively charged functional groups of carbohydrates and protein moiety of the EPS endowed it with heavy metal binding capacity through electrostatic interactions. The highest flocculation activity (83%) of EPS was observed at 4 mg L−1 and pH 11. The metal sorption by EPS increased with increasing pH. At pH 8, the EPS was able to remove 86 and 81% Cu(II) and Zn(II), respectively, from a 25 mg L−1 metal solution. 94.8% of both the metals at 25 mg L−1 metal solutions were removed by EPS at EPS concentration of 100 mg L−1. From Langmuir isotherm model, the maximum sorption capacities of EPS were calculated to be 58.82 mg g−1 for Cu(II) and 52.45 mg g−1 for Zn(II). The bacterial EPS showed encouraging flocculating and metal sorption properties. The potential to remove Cu(II) and Zn(II) implies that the EPS obtained from the earthworm gut bacteria can be used as an effective agent for environmental remediation of heavy metals and in bioflocculation.

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