The mercury resistance (mer) operon of the gram-negative transposon Tn21 encodes not only a mercuric reductase and regulatory genes but also two inner membrane proteins (MerT and MerC) and a periplasmic protein (MerP). Although the merT, merP, and merC genes have been implicated in Hg(II) transport, the individual roles of these genes have not been established. We created in vitro precise deletion and frameshift mutations that eliminated each of the genes singly and in combination. Our results show that both merT and merP are required for Hg(II) binding but that merC is not. Both merT and merP are required for full expression of Hg(II) resistance, but loss of merP is less deleterious than loss of merT. Furthermore, mutations eliminating both merT and merP decrease resistance more than the single mutations do. In contrast, mutating merC had no effect on Hg(II) resistance. Both the merT and merP mutations increase the threshold Hg(II) concentration for induction of merA-lacZ transcriptional fusions and cause an increase in the maximal expression level. In contrast, the merC mutation had little effect on the threshold inducing concentration of Hg(II) but decreased the level of expression. Our results show that merT and merP alone are sufficient to specify a mercury transport system. The role of merC remains obscure.

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The detergent-resistance properties of 208 independent isolates of the Enterobacteriaceae have been examined. Of these bacterial strains, 200 were able to grow in the presence of greater than or equal to 5% sodium dodecyl sulfate, including all members of the Klebsielleae tribe. This resistance does not appear to be plasmid encoded. It is proposed that detergent-resistant organisms be termed saponotolerant or saponophilic, by analogy with other microorganisms occupying harsh ecological niches. In contrast to their prevalent resistance to anionic detergents, not one of the 208 strains tested was found to grow in the presence of three different cationic detergents. This sensitivity to cationic detergents may be of significance in combating nosocomial infections.

A physical map of the bacteriophage T5 genome was constructed by ordering the fragments produced by cleavage of T5 DNA with the restriction endonucleases SalI (4 fragments), SmaI (4 fragments), BamI (5 fragments), and HpaI (28 fragments). The following techniques were used to order the fragments. (i) Digestion of DNA from T5 heat-stable deletion mutants was used to identify fragments located in the deletable region. (ii) Fragments near the ends of the T5 DNA molecule were located by treating T5 DNA with X exonuclease before restriction endonuclease cleavage. (iii) Fragments spanning other restriction endonuclease cleavage sites were identified by combined digestion of T5 DNA with two restriction endonucleases. (iv) The general location of some fragments was determined by isolating individual restriction fragments from agarose gels and redigesting the isolated fragments with a second restriction enzyme. (v) Treatment of restriction digests with X exonuclease before digestion with a second restriction enzyme was used to identify fragments near, but not spanning, restriction cleavage sites. (vi) Exonuclease III treatment of T5 DNA before restriction endonuclease cleavage was used to locate fragments spanning or near the natural T5 single-chain interruptions. (vii) Analysis of the products of incomplete restriction endonuclease cleavage was used to identify adjacent fragments. MATERIALS AND METHODS Bacteriophage. Wild-type T5 and the heat-stable deletion mutants T5st(0) and T5st(102) (13, 15) were grown on Escherichia coli B23. EcoRI endonuclease. Purification and assay conditions for EcoRI have been described (11). Sally endonuclease. Streptomyces albus G (from G. Hayward) was grown at 30°C with shaking in a medium containing, in 1 liter: 0.33 g of yeast extract (Difco), 0.33 g of beef extract (Difco), 0.67 g of tryptone (Difco), trace FeSO4, and 3.33 g of glucose. After 2 days of growth, the mycelia were harvested by centrifugation and suspended in 0.01 M Tris, pH 7.9, containing 0.01 M 2-mercaptoethanol (approximately 10 ml per 15 g [wet weight] of mycelia). The cells were disrupted by sonication. Cell debris was removed from the extract by low-speed (8,000 rpm for 15 min in a Sorvall SS34 rotor) and high-speed centrifugation (33,000 rpm for 90 min in a Beckman SW50.1 rotor). The high-speed supernatant was made 1.0 M in NaCl and applied to a 200-ml column 249

An examination was made of the properties of T5HA4, a mutant of bacteriophage T5 that lacks the single-chain interruption that occurs at 7.9% from the left end of the genome. The DNAs of T5HA4 and the wild type were compared by electrophoresis in agarose gels of both single-stranded fragments produced by denaturation and duplex fragments generated by sequential treatment with