Abstract
The purposes of the study was to isolate, to characterize, and to identify rhizobacteria from plant rhizosphere growing in gold and copper mine. The isolation of rhizobacteria used N-free semisolid agar media, TSA, and SLP plus heavy metals (HMs), namely Pb, Cd, and or Cu. Isolated rhizobacteria were subsequently characterized for resistance to higher level of Pb, Cd, Cu in SLP media. Cultural and morphological characterization of rhizobacteria were conducted for cell morphology, motility, Gram staining, and biofilm formation. The rhizobacteria identification used sequence analysis of the 16S RNA gene fragments. The results showed that the majority of rhizobacterial from Cu mine site (66.7% of 21 isolates) were resistant to Cu (72150 ppm) while the majority of rhizobacteria from gold mine site (77.8% of 18 isolates) were sensitive to 72 ppm Cu. Majority of Cu in the soil was insoluble as granules attaching to gravel so that rhizobacteria of Cu mine site have been exposed and adapted to available Cu. This fact, explaining that the rhizobacteria’s MIC value was lower than the total Cu level in the soil. Three HMs-resistant rhizobacter (PbSM 2.1, MGR 334, and CuNFbM 4.1) formed biofilms, which was as one of the resistance mechanism to HMs. This research informed that HM contaminated-soil is better source for obtaining HM resistant rhizobacteria than HM uncontaminated-soil. The use four isolation media produce rhizobacteria which was more diverse than rhizobacteria from each isolation medium. Further characterization needs to be done to obtain HM resistant-rhizobacteria which can be used as biofertilizers and phytoremediation agent.
Highlights
Mining activity in many countries has brought about environmental issues such as heavy metal (HM) contamination and marginal land formation including low pH, loss of organic matter and nutrient, as well as the unstable situation of the land (Wong, 2003; Raymundo, 2006)
It has been known that media such as Trypticase Soy Agar (TSA) is for isolating gram positive and negative bacteria, N-free bromthymol medium (NFB), sucroseminimal salts low phosphate Agar (SLP), and soil extract agar (SEA) which exclude N, reduce P concentration, and contain soil extract respectively
Rhizobacteria from Marinduque were more tolerant than isolates from Antam Bogor to Cu, Cd, and Pb
Summary
Mining activity in many countries has brought about environmental issues such as heavy metal (HM) contamination and marginal land formation including low pH, loss of organic matter and nutrient, as well as the unstable situation of the land (Wong, 2003; Raymundo, 2006). Mine tailings are extreme environments, containing high concentrations of HMs and deficient in nitrogen and organic matter with extremely low pH of the soil In this such area, there are some plant species tolerant to HMs and capable of growing on it (He et al, 2010; Navarro-Noya et al, 2012). The use of dilute nutrient media suited for the growth of bacteria adapted to oligotrophic conditions, and the provision of simulated natural environmental conditions for bacterial culture. This has led to the recovery of ‘unculturables’ from environments, likely to be due to the inclusion of essential nutrients and/or signalling molecules from the native environment (Vartoukian, Palmer & Wade, 2010). The addition heavy metal to the each medium maymimic environment condition suitable for rare rhizobacteria growth from rhizosphere of plant growing on mine tailing
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