Screening of isolates and strains of Rhizobium leguminosarum biovar trifolii for heavy metal resistance using buffered media

The computer program GEOCHEM-PC was used to calculate the metal ion activities of Cu, Zn, Cd, and Ni in defined media amended with various metal ion buffers at pH 6.6 or 6.0 so that the lowest-observed-effect concentrations (LOECs) of these metals to isolates and strains of Rhizobium leguminosarum biovar trifolii could be determined. Strains from the U.S. Department of Agriculture (USDA) Beltsville (MD) Rhizobium culture collection were generally more tolerant of the metals than any of the isolates from the sludge-treated soil (S-isolates) and those from farmyard manure-treated soil (F-isolates), although the S- were more metal resistant than the F-isolates. All isolates and strains, however, tolerated much larger concentrations in the buffered systems than those found in the solutions of soils from which they originated. Copper toxicity, using iminodiacetate (IDA), occurred for the F- and S-isolates and USDA strains at concentrations of 16, 47, and 430 mug ml-1, respectively, corresponding to predicted ion activities of 0.002, 0.006, and 0.06 mug ml-1, respectively. The Zn LOECs for the F- and S-isolates, without a buffer, occurred at concentrations of 47 and 207 mug ml-1, respectively, corresponding to predicted ion activities of 37 and 157 mug ml-1, respectively. No toxicity occurred when nitrilotriacetate was used for Cd up to concentrations of 356 mug ml-1 (Cd ion activity of 2.6 mug ml-1). Nickel concentrations up to 0.6 mug ml-1 had no effect with ethylene-bis(oxyethylenenitrilo)tetraacetate (EGTA; Ni ion activity of 0.006 mug ml-1) and up to 186 mug ml-1 with IDA (Ni ion activity of 11 mug ml-1).