Isolation of plant growth promoting bacteria from torch lake sediments and their role in phytoremediation of metal contaminated soil

In this study, we isolated eight copper-resistant bacteria from Torch Lake sediment contaminated by copper mine tailings (stamp sand). Sequence analysis of gyrB and rpoD genes revealed that these organisms are closer to various Pseudomonas species. These eight bacterial isolates were also resistant to zinc, cesium, lead, arsenate and mercury. Further characterization showed that all the strains produced plant growth promoting indole-3-acetic acid (IAA), iron chelating siderophore and solubilized mineral phosphate and metals. The effect of bacterial inoculation on plant growth and copper uptake by maize (Zea mays) and sunflower (Helianthus annuus) was investigated using one of the isolates (Pseudomonas sp. TLC 6-6.5-4) with higher IAA production and phosphate and metal soubilization, which resulted in a significant increase in copper accumulation in maize and sunflower, and an increase in the total biomass of maize. Genes involved in copper resistance of Pseudomonas sp. TLC 6-6.5-4 was analyzed by transposon mutational analysis. Two copper sensitive mutants with significant reduction in copper resistance were identified: CSM1, a mutant disrupted in trp A gene (tryptophan synthase alpha subunit); CSM2, a mutant disrupted in clpA gene (ATP-dependent Clp protease). Proteomic and metabolomic analysis were performed to identify biochemical and molecular mechanisms involved in copper resistance using CSM2 due to its lower minimum inhibitory concentration compared with CSM1 and the wild type. The effect of different bacterial inoculation methods on plant growth, copper uptake and soil enzyme activities was investigated. Four different delivery methods were used including soil inoculation (before or after plant emergence), seed coating and root dipping. Soil inoculation before sowing seeds and coating seeds with PGPB led to better growth of maize, higher copper uptake and an increase in soil invertase and dehydrogenase activities. Proteomic and metabolomic analyses were performed to investigate the effect of bacterial inoculation on maize grown in normal soil and stamp sand. Our results revealed that bacterial inoculation led to environment-dependent effects on maize proteome and metabolome.

REMEDIATION OF TETRACYCLINE FROM WATER SOURCES USING VETIVER GRASS (Chrysopogon zizanioides L. Nash) AND TETRACYCLINE-TOLERANT ROOT-ASSOCIATED BACTERIA

Antibiotics are emerging contaminants worldwide. Due to insufficient policy regulations, public awareness, and the constant exposure of the environment to antibiotic sources has created a major environmental concern. Wastewater treatment plants (WWTP) are not equipped to filter-out these compounds before the discharge of the disinfected effluent into water sources (e.g., lakes and streams) and current available technologies are not equipped to remediate these compounds from environmental sources. Hence, the challenge remains to establish a biological system to remove these antibiotics from wastewater. An invitro hydroponic remediation system was developed using vetiver grass (Chrysopogon zizanioides L. Nash) to remediate tetracycline (TC) from water. Comparative metabolomics studies were conducted to investigate the metabolites/pathways associated with tetracycline metabolism in plants and TC-degrading bacteria. The results show that vetiver plants effectively uptake tetracycline from water sources. Vetiver root-associated bacteria recovered during the hydroponic remediation trial were highly tolerant to TC (as high as 600 ppm) and could use TC as a sole carbon and energy source. Growth conditions (pH, temperature, and oxygen requirement) for TC-tolerant bacteria were optimized for higher TC remediation capability from water sources. The plant (roots and shoots) and bacterial species were further characterized for the metabolites produced during the TC degradation process using GC-MS to identify the possible biochemical mechanism involved. Also, the plant root zone was screened for metabolites/enzymes that were secreted during antibiotic degradation and could potentially enhance the degradation process. The root zone was selected for this analysis because this region of the plant has shown a greater capacity for antibiotic degradation compared to the shoot zone. The role of antioxidant enzymes in TC degradation process revealed glutathione-S-transferase (GSTs) as an important group of enzymes in both plant and bacteria potentially involved in TC degradation process. Metabolomics results also suggest potential GST activity in the TC remediation/ transformation process used by plants. This information could be useful in gaining insights for the application of biological remediation systems for the mitigation of antibiotics from waste-water.