Bacteria have been examined that have developed very efficient and different mechanisms for tolerating heavy metals. Often normal toxic levels of metals have no effect on cell growth of resistant strains. In many organisms, the genes controlling metal resistance are carried on plasmids, which provide the bacteria with a competitive advantage over other organisms when metals are present. Not all metal-resistant bacteria contain plasmids. For example, a Pseudomonas sp. (research at the Canada Centre for Inland Waters and the University of Guelph) resistant to 1000 #g/ml lead, also displays multiple antibiotic resistance. Vertical and horizontal agarose gel electrophoresis of cleared lysates revealed that plasmids were not present, even though the organism was tolerant to extremely high concentrations of lead.Plasmid-encoded resistance may provide organisms with efflux and bypass mechanisms, enzymes which catalyze the transformation of metals to volatile forms, or make the bacterial cell wall impermeable to the metal(s).A problem still remains in actually defining the concentrations that distinguish metal-resistant from metal-sensitive bacteria. Some researchers have proposed mathematical equations, and used statistical an.alysis to solve this problem. Nevertheless, standard concentrations have not been uni-versally proposed and/or accepted by the scientific community. This task is complicated by the various forms of metals used, the effect of media components, pH, and culture conditions, which have the capability of influencing the toxicity of the metal.Research on metal-bacteria interactions is in some ways still in its early stages of development. Nevertheless, significant advances in scientific knowledge have been achieved over the last 20 years. To fill the existing gaps in knowledge, research will need to be conducted on the extent of metal resistance in bacteria, the relationship between metal and antibiotic resistance, mechanisms specified by the plasmids, and the ecology, physiology, and genetics of gene transfer in the natural environment.Since some microorganisms are responsible for environmental metal transformations, they may also serve as bioassay indicator organisms in polluted and non-polluted environments. Moreover, there may be potential biotechnological applications for metal-resistant bacteria in the area of toxic metal control in waste-water treatment. INTRODUCTIONThe genetic basis for metal resistance in bacteria is an area of intensive research both in the en-0168-6445/85/$03.30

Thermograms of the exosporium-lacking dormant spores of Bacillus megaterium ATCC 33729, obtained by differential scanning calorimetry, showed three major irreversible endothermic transitions with peaks at 56, 100, and 114°C and a major irreversible exothermic transition with a peak at 119°C. The 114°C transition was identified with coat proteins, and the 56°C transition was identified with heat inactivation. Thermograms of the germinated spores and vegetative cells were much alike, including an endothermic transition attributable to DNA. The ascending part of the main endothermic 100°C transition in the dormant-spore thermograms corresponded to a first-order reaction and was correlated with spore death; i.e., >99.9%o of the spores were killed when the transition peak was reached. The maximum death rate of the dormant spores during calorimetry, calculated from separately measured D and z values, occurred at temperatures above the 73°C onset of thermal denaturation and was equivalent to the maximum inactivation rate calculated for the critical target. Most of the spore killing occurred before the release of most of the dipicolinic acid and other intraprotoplast materials. The exothermic 119°C transition was a consequence of the endothermic 100°C transition and probably represented the aggregation of intraprotoplast spore components. Taken together with prior evidence, the results suggest that a crucial protein is the rate-limiting primary target in the heat killing of dormant bacterial spores.How do bacterial spores resist heat, and, conversely, how are they killed when their thermal-defense mechanisms are overcome? Answering these two questions is fundamentally important to practical solutions of food spoilage, food poisoning, and infectious disease in which spores are causative. Furthermore, much of microbiological practice depends on heat sterilization, which is predicated on killing all spores in the material.The question of heat resistance mechanisms now seems mainly answered as follows: bacterial spores are able to resist heat because their protoplasts have become dehydrated to a water content that is low enough to immobilize and therefore to protect the vital macromolecules (e.g., proteins, RNA, and DNA) from denaturation and the supramolecular assemblies (e.g., membranes and ribosomes) from disruption. Although dehydration is the only property necessary and sufficient in itself to impart heat resistance, it is enhanced by mineralization (especially by calcification) and thermal adaptation. The physiological processes by which the spore attains the resistant state are less well understood, but the retention of resistance clearly is a function of an intact peptidoglycan cortex encasing the protoplast. The experimental evidence supporting these assertions has been reviewed by Gerhardt and Marquis (19).The question of heat killing mechanisms, however, remains mainly unanswered. In an effort to obtain experimental evidence about these mechanisms, we undertook the study of a selected spore morphotype, Bacillus m...

Eleven hundred Bacillus strains isolated from marine sediment from the Minas Basin, Nova Scotia, Canada, were purified on LB agar supplemented with ampicillin, chloramphenicol, erythromycin, streptomycin, tetracycline, or mercuric chloride. Seventy-seven isolates were examined for plasmid DNA, and for resistance to 11 antibiotics, HgCl2, and phenylmercuric acetate. Minimum inhibitory concentrations of Ag, Cd, Co, Cu, and Zn were also determined. Forty-three percent of antibiotic- and mercury-resistant strains contained one or more plasmids ranging from 1.9 to 210 MDa. Fifty-four percent carried plasmids greater than 20 MDa, and 97% were resistant to two or more metals. There was no correlation between plasmid content and resistance either to antibiotics or to mercurial compounds in these strains. Mercury-resistant isolates were unable to transform Hg2+ to volatile Hg0 by virtue of a mercuric reductase enzyme system (mer). Strains resistant to Hg2+ were investigated for their ability to produce H2S and intracellular acid-labile sulfide when grown in the absence and presence of HgCl2. Lower levels of H2S and intracellular sulfide were detected only in metal-resistant strains grown in the presence of HgCl2, suggesting that cellular sulfides complexed with Hg2+ in these strains.

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