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The O157:H7 lineage of enterohemorrhagic Escherichia coli is a geographically disseminated complex of highly related genotypes that share common ancestry. The common clone that is found worldwide carries several markers of events in its evolution, including markers for acquisition of virulence genes and loss of physiological characteristics, such as sorbitol fermentation ability and ␤-glucuronidase production. Populations of variants that are distinct with respect to motility and the sorbitol and ␤-glucuronidase markers appear to have diverged at several points along the inferred evolutionary pathway. In addition to these variants, distinct subpopulations of the contemporary non-sorbitol-fermenting, ␤-glucuronidase-negative O157:H7 clone were recently detected among bovine and human clinical isolates in the United Stares by using highresolution genome comparison. In order to determine if these recently described subpopulations were derived from a regional or ancestral divergence event, we used octamer-based genome scanning, marker sorting, and DNA sequence analysis to examine their phylogenetic relationship to populations of non-sorbitol-fermenting, ␤-glucuronidase negative O157:H7 and O157:H؊ strains from Australia. The inferred phylogeny is consistent with the hypothesis that subpopulations on each continent resulted from geographic spread of an ancestral divergence event and subsequent expansion of distinct subpopulations. Marker sorting and DNA sequence analyses identified sets of monophyletic markers consistent with the pattern of divergence and demonstrated that phylogeographic variation occurred through emergence of regional subclones and concentration of regional polymorphisms among distinct subpopulations. DNA sequence analysis of representative polyphyletic markers showed that genome diversity accrued through random drift and bacteriophage-mediated events.Hemorrhagic colitis is caused by a number of serotypes of Shiga toxin-producing Escherichia coli (STEC) (14). Among the clinical STEC strains that have been isolated, a subset of enterohemorrhagic E. coli (EHEC) strains has been found which carry common sets of virulence genes that encode factors for attachment to host cells, elaboration of effector molecules, and production of two different types of Shiga toxins (22). The sets of virulence genes are found in the locus of enterocyte effacement (LEE) pathogenicity island, lambdoid bacteriophages, and a large virulence-associated plasmid (8,9,23,25,26,31,32). Population genetic analysis of EHEC and STEC strains has shown that EHEC strains comprise two divergent lineages, termed EHEC 1 and EHEC 2, that are only distantly related but apparently experienced similar pathways of virulence gene acquisition (24,28,38). The EHEC 1 lineage is comprised solely of a geographically disseminated cluster of strains with related genotypes bearing O157:H7 and O157:HϪ serotypes, while the EHEC 2 lineage is serotypically and genotypically more diverse.The O157 serotype can be found in genetically diverse populations of E...

Thirteen different serotypes of Listeria monocytogenes can be distinguished on the basis of variation in somatic and flagellar antigens. Although the known virulence genes are present in all serotypes, greater than 90% of human cases of listeriosis are caused by serotypes 1/2a, 1/2b, and 4b and nearly all outbreaks of food-borne listeriosis have been caused by serotype 4b strains. Phylogenetic analysis of these three common clinical serotypes places them into two different lineages, with serotypes 1/2b and 4b belonging to lineage I and 1/2a belonging to lineage II. To begin examining evolution of the genome in these serotypes, DNA microarray analysis was used to identify lineage-specific and serotype-specific differences in genome content. A set of 44 strains representing serotypes 1/2a, 1/2b, and 4b was probed with a shotgun DNA microarray constructed from the serotype 1/2a strain 10403s. Clones spanning 47 different genes in 16 different contiguous segments relative to the lineage II 1/2a genome were found to be absent in all lineage I strains tested (serotype 4b and 1/2b) and an additional nine were altered exclusively in 4b strains. Southern hybridization confirmed that conserved alterations were, in all but two loci, due to absence of the segments from the genome. Genes within these contiguous segments comprise five functional categories, including genes involved in synthesis of cell surface molecules and regulation of virulence gene expression. Phylogenetic reconstruction and examination of compositional bias in the regions of difference are consistent with a model in which the ancestor of the two lineages had the 1/2 somatic serotype and the regions absent in the lineage I genome arose by loss of ancestral sequences.Listeria monocytogenes is a ubiquitous gram-positive bacterium that can cause life-threatening infections including meningitis, septicemia, abortion, and fetal death. Its primary route of transmission to humans is through contaminated food and despite the relatively low incidence of listeriosis in humans, outbreaks of listeriosis often have high associated morbidity, particularly among pregnant women, unborn fetuses, and immunocompromised individuals (24). These characteristics, coupled with its physiological durability and its ubiquitous distribution in nature have propelled L. monocytogenes to the forefront of food safety research and the regulatory arena.Pathogenesis of listeriosis is a consequence of the organism's ability to invade and replicate within several different cell types in mammalian tissues, including intestinal, liver, and neural tissues. During the course of food-borne infections, the bacteria penetrate the intestinal lining through cell invasion and translocation and use the lymphatic system as a conduit to reach the main target tissues within the liver and spleen (41). Prolonged replication in the liver, facilitated by depressed cell mediated immunity, is thought to be an antecedent to spread of the bacteria to brain tissue and breach of the placental barrier in pregnant wo...

Endospore formation in Bacillus subtilis begins with an asymmetric cell division that partitions the bacterium into mother cell and forespore compartments. Mother cell-specific gene expression is initiated by E , a transcription factor that is active only in the mother cell but which existed as an inactive precursor (pro-E ) in the predivisional cell. Activation of pro-E involves the removal of 27 amino acids from its amino terminus. A chimera of pro-E and the green fluorescent protein (GFP) was expressed from either the normal sigE promoter (P spoIIG ), which places pro-E ::GFP in both mother cell and forespore compartments, or the forespore-specific promoter (P dacF ), which produces pro-E ::GFP only in the forespore compartment. The pro-E ::GFP expressed from P spoIIG , but not P dacF , was converted to a lower-molecular-weight form by a mechanism dependent on gene products (SpoIIGA and F

As Bacillus subtilis proceeds through sporulation, the principal vegetative cell ς subunit (ςA) persists in the cell but is replaced in the extractable RNA polymerase (RNAP) by sporulation-specific ς factors. To explore how this holoenzyme changeover might occur, velocity centrifugation techniques were used in conjunction with Western blot analyses to monitor the associations of RNAP with ςA and two mother cell ς factors, ςE and ςK, which successively replace ςA on RNAP. Although the relative abundance of ςA with respect to RNAP remained virtually unchanged during sporulation, the percentage of the detectable ςAwhich cosedimented with RNAP fell from approximately 50% at the onset of sporulation (T 0) to 2 to 8% by 3 h into the process (T 3). In a strain that failed to synthesize ςE, the first of the mother cell-specific ς factors, approximately 40% of the ςA remained associated with RNAP at T 3. The level of ςA-RNAP cosedimentation dropped to less than 10% in a strain which synthesized a ςE variant (ςECR119) that could bind to RNAP but was unable to direct ςE-dependent transcription. The E-ςE-to-E-ςK changeover was characterized by both the displacement of ςE from RNAP and the disappearance of ςE from the cell. Analyses of extracts from wild-type and mutant B. subtilis showed that the ςK protein is required for the displacement of ςE from RNAP and also confirmed that ςK is needed for the loss of the ςE protein. The results indicate that the successive appearance of mother cell ς factors, but not necessarily their activities, is an important element in the displacement of preexisting ς factors from RNAP. It suggests that competition for RNAP by consecutive sporulation ς factors may be an important feature of the holoenzyme changeovers that occur during sporulation.