A copper-transport (copYAZ) operon was cloned from the oral bacterium Streptococcus mutans JH1005. DNA sequencing showed that the operon contained three genes (copY, copA and copZ), which were flanked by a single promoter and a factor-independent terminator. copY encoded a small protein of 147 aa with a heavy-metal-binding motif (CXCX 4 CXC) at the C-terminus. CopY shared extensive homology with other bacterial negative transcriptional regulators. copA encoded a 742 aa protein that shared extensive homology with P-type ATPases. copZ encoded a 67 aa protein that also contained a heavymetal-binding motif (CXXC) at the N-terminus. Northern blotting showed that a 32 kb transcript was produced by Cu 2M -induced Strep. mutans cells, suggesting that the genes were synthesized as a polycistronic message. The transcriptional start site of the cop operon was mapped and shown to lie within the inverted repeats of the promoter-operator region. Strep. mutans wild-type cells were resistant to 800 µM Cu 2M , whereas cells of a cop knock-out mutant were killed by 200 µM Cu 2M . Complementation of the cop knock-out mutant with the cop operon restored Cu 2M resistance to wild-type level. The wild-type and the mutant did not show any differences in susceptibility to other heavy metals, suggesting that the operon was specific for copper. By using a chloramphenicol acetyltransferase reporter gene fusion, the cop operon was shown to be negatively regulated by CopY and could be derepressed by Cu 2M .

One of the most abundant proteins synthesized by Legionella pneumophila, particularly during growth in a variety of eukaryotic host cells, is Hsp60, a member of the GroEL family of molecular chaperones. The present study was initiated in response to a growing number of reports suggesting that for some bacteria, includingL. pneumophila, Hsp60 may exist in extracytoplasmic locations. Immunolocalization techniques with Hsp60-specific monoclonal and polyclonal antibodies were used to define the subcellular location and distribution of Hsp60 in L. pneumophila grown in vitro, or in vivo inside of HeLa cells. For comparative purposesEscherichia coli, expressing recombinant L. pneumophila Hsp60, was employed. In contrast to E. coli, where Hsp60 was localized exclusively in the cytoplasm, inL. pneumophila Hsp60 was predominantly associated with the cell envelope, conforming to a distribution pattern typical of surface molecules that included the major outer membrane protein OmpS and lipopolysaccharide. Interestingly, heat-shocked L. pneumophila organisms exhibited decreased overall levels of cell-associated Hsp60 epitopes and increased relative levels of surface epitopes, suggesting that Hsp60 was released by stressed bacteria. Putative secretion of Hsp60 by L. pneumophila was further indicated by the accumulation of Hsp60 in the endosomal space, between replicating intracellular bacteria. These results are consistent with an extracytoplasmic location for Hsp60 in L. pneumophilaand further suggest both the existence of a novel secretion mechanism (not present in E. coli) and a potential role in pathogenesis.

The identification of genes associated with colonization and persistence of Helicobacter pylori in the gastric mucosa has been limited by the lack of robust animal models that support infection by strains whose genomes have been completely sequenced. Here we report that an interleukin-12 (IL-12)-deficient mouse (IL-12 ؊/؊ p40 subunit knockout in C57BL/6 mouse) is permissive for infection by a motile variant (KE88-3887) of The Institute For Genomic Research-sequenced strain (KE26695) of H. pylori. The IL-12-deficient mouse was also more permissive for colonization by the mouse-colonizing Sydney 1 strain of H. pylori than were wild-type C57BL/6 mice. Differences in colonization efficiency were demonstrated by mouse challenge with SS1 strains containing loss-of-function mutations in two genes (hspR and hrcA), whose products negatively regulate several heat shock genes. Helicobacter pylori colonizes the gastric mucosa of humans, producing a chronic gastritis that may remain asymptomatic for many years. In about 10% of individuals, more severe disease manifestations will occur such as duodenal and gastric ulcers, atrophic gastritis, and intestinal metaplasia, all risk factors for gastric cancer (20,40,42). The remarkable ability of H. pylori to establish lifelong infections is not well understood but likely involves evasion or modulation of host immune responses as well as adaptation (through mutation and selection) to the unique physiology of each individual host (3). In addition, different disease pathologies seem to correlate with particular H. pylori genotypes, with strains containing the cagassociated pathogenicity island and cagA gene (cytotoxinassociated gene) and vacA (vacuolating cytotoxin) correlating with more severe disease (11,14,39). There is much genetic diversity in these genes and in others such as the restriction modification genes (9) among strains from different geographic regions and people of different ethnicities (1, 10, 13). The evaluation of genetic diversity among strains and identification of genes associated with severity of infection have generally been hampered by the lack of good animal models (9,25,36).Robust animal models of infection are also a necessary component in the discovery process for new therapeutics or the evaluation of vaccine candidates (12,23,47). While several animal models have been developed, these models are limited to a few animal-adapted strains (9,22,36) or support only transient infection (25). Mouse-adapted strains such as the Sydney 1 strain (SS1), Hp1, and a few others are difficult to manipulate genetically (37) and usually require high infectious doses in order to establish infection (36). In such mouse models requiring high colonization thresholds, many genes scored as necessary for colonization may be dispensable in a more permissive animal. Another limitation of existing mouse-colonizing strains is that systematic whole-genome approaches to the study of virulence determinants cannot be performed, as the genomes of mouse-colonizing strains have not been se...

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The well-known finite-lattice method for the calculation of the properties of quantum spin systems on a two-dimensional lattice at zero temperature was introduced in 1978. The method has now been greatly improved for the square lattice by including finite lattices based on parallelogram tiles as well as the familiar finite lattices based on square tiles. Dozens of these new finite lattices have been tested and graded using the [Formula: see text] ferromagnet. In the process new and improved estimates have been obtained for the XY model's ground-state energy per spin, ε0 = −0.549 36(30) and spontaneous magnetization per spin, m = 0.4349(10). Other properties such as near-neighbour, zero-temperature spin–spin correlations, which appear not to have been calculated previously, have been estimated to high precision. Applications of the improved finite-lattice method to other models can readily be carried out.