Reactions of 12 strains of soil bacteria to 15 metals

Reactions of 12 strains of common soil bacteria to 15 metals were tested in a laboratory experiment. Paper discs treated with metal ions were dried and placed on nutrient agar freshly inoculated with different varieties of bacteria. Growth patterns were observed at the end of 24 hours and at the end of 6 days. After 6 days, the.overall toxicity of the metals to the bacteria followed the order: Hg>Ag>Cd>V>Co>Sb>Bi. Only one or two strains of bacteria were sensitive to Al, Se, Te, and Zn, and none were sensitive to As, Cu, Ni, or Sn at the test concentration. Each species of bacteria exhibited a unique resistance pattern, indicating that bacterial reaction to a suite of heavy metals is complex and strain-specific. Such strain-specific heavy-metal resistances may provide a taxonomic basis for the study of three-dimensional metal distribution patterns in nature. INTRODUCTION A number of recent reports, some of which are concerned with environmental pollution, have established the feasibility of extending mineral-exploration geobotany to include surveys of metal adaptation in soil and sediment microflora (Marques et al., 1979; Timoney et al., 1978; Austin et al., 1977; Nelson and Colwell, 1975; Koval'skii et al., 1968; Kendrick, 1962 Tuovinen et al., 1971). For a recent review of the phenomena of microbial heavy-metal resistance, see Summers and Silver (1978). The present study was undertaken as part of an investigation of procedures applicable to the detection of microbial resistance to one or more of the metallic elements commonly associated with economic mineral deposits. A paper disc method (Bauer et al., 1966) is now commonly used to test microbial sensitivity to antibiotics. At least two previous studies report the use of this technique to test bacterial resistances to inorganic ions (Novick and Roth, 1968; Groves and Young, 1975). The metal-binding capacity of cellulose (Belford et al., 1958; Ogiwara and Kubota, 1969) was considered negligible in these tests at seven micrograms (yg) of metal per disc, but might become a factor in applications of disc-held metals at lower concentrations. Strains of bacteria tested for their natural resistance to metals in this experiment included Rhodospi rill urn rubrum, Bacillus megaten'um, Sarcina flava, Bacillus subtil is, Bacillus subtilis var. niger, Flavobacterium dem'trificans, Serratia marcescens, Serratia marcescens 933 (a colorless variant), Micrococcus coralinus, Sarcina aurantiaca, Micrococcus roseus. and Micrococcus luteus. Materials and sources are listed in table 1. PROCEDURE A loose-leaf binder hole punch was used to punch out filter paper discs about 6 mm in diameter from Whatman 41 filter paper. Two or three dozen discs were placed in each of an appropriate number of screw-cap glass vials and moistened to saturation with one of the metal standard solutions. Three additional vials of discs were saturated with 10 percent solutions of concentrated HC1, HNO^ and aqua regia (7:3 HCliHNO^) as carrier acid "blanks. " The moistened discs were dried in the vials, with caps loosened, in a drying oven at approximately 90°C for 3 days prior to use. Discs containing Ag, Al, Sb, Sn, and Zn darkened to some extent during drying. Twenty discs absorbed 0.14 ml of distilled water; each disc therefore held about 7 yl of liquid prior to drying, or 7 yg of metal. All strains were subcultured from supply house cultures by evenly streaking the bacteria over 100 x 15 mm petri plates containing 45 ml of nutrient agar. After a week, one or two additional subcultures of each strain were prepared by repeating the process; ten days Table 1 Materials and Sources