Design of a Seven-GenomeEscherichia coliMicroarray for Comparative Genomic Profiling

Abstract

We describe the design and evaluate the use of a high-density oligonucleotide microarray covering seven sequenced Escherichia coli genomes in addition to several sequenced E. coli plasmids, bacteriophages, pathogenicity islands, and virulence genes. Its utility is demonstrated for comparative genomic profiling of two unsequenced strains, O175:H16 D1 and O157:H7 3538 (⌬stx 2 ::cat) as well as two well-known control strains, K-12 W3110 and O157:H7 EDL933. By using fluorescently labeled genomic DNA to query the microarrays and subsequently analyze common virulence genes and phage elements and perform whole-genome comparisons, we observed that O175:H16 D1 is a K-12-like strain and confirmed that its 3538 (⌬stx 2 ::cat) phage element originated from the E. coli 3538 (⌬stx 2 ::cat) strain, with which it shares a substantial proportion of phage elements. Moreover, a number of genes involved in DNA transfer and recombination was identified in both new strains, providing a likely explanation for their capability to transfer 3538 (⌬stx 2 ::cat) between them. Analyses of control samples demonstrated that results using our custom-designed microarray were representative of the true biology, e.g., by confirming the presence of all known chromosomal phage elements as well as 98.8 and 97.7% of queried chromosomal genes for the two control strains. Finally, we demonstrate that use of spatial information, in terms of the physical chromosomal locations of probes, improves the analysis.The Escherichia coli species is a complex group of bacteria comprising several intestinal and extraintestinal pathogroups as well as commensal bacteria that are normal inhabitants of the intestinal tract of all warm-blooded animals and humans. Shiga toxin-producing E. coli (STEC) has emerged as an important food-borne pathogen causing diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Healthy ruminants such as cattle and sheep are regarded as the primary reservoir of STEC, which may be pathogenic to humans depending on the genomic content and combination of pathogenicity factors.The Shiga toxins (Stx) are the main pathogenicity factors of STEC. Stx-encoding genes (stx) are located on lamboid bacteriophages known as stx phages (17, 30). The stx phages are not only passive vectors for the dissemination of stx but also genetic entities where the characteristics of the phage itself may influence toxin production and, thus, virulence of the host bacterium (35,36). Dissemination of stx genes by transduction is the most likely mechanism for intra-and intergenic spread of stx and subsequent development of new STEC. The host range of stx phages is highly variable, and phage transduction into E. coli and Shigella strains has been shown in different laboratory and animal experiments (1,12,16,28,34). Evidence for transduction of the bacteriophage 3538 (stx 2 ::cat) from E. coli O157:H7 3538 (⌬stx 2 ::cat) (28) has been shown in porcine loops (34) and recently by feeding sheep with E.

Explaining microbial phenotypes on a genomic scale: GWAS for microbes

“…These microarrays were spotted with probes that map to the genes present in a single genome (CGH arrays) or in a number of strains of the same species [16,17], coined pan-genome arrays [18]. The arrays could then be hybridized with fluorescently labeled genomic cDNA to determine the gene content of a query strain [19,20].…”

Comparative genomics of two ecotypes of the marine planktonic copiotroph Alteromonas macleodii suggests alternative lifestyles associated with different kinds of particulate organic matter

“…The most extensively studied cases are those of pathogenic bacteria in which the presence of specific gene clusters or pathogenicity islands are often linked to the clinical outcome of their infection (Tettelin et al, 2005;Dempsey et al, 2006;Hochhut et al, 2006;Petrosino et al, 2006;Willenbrock et al, 2006). However, free-living bacteria have similar genomic strategies when exploiting different habitats or niches and comparative genomics is a powerful tool to understand the ecological specialization of the different lineages (Ting et al, 2002;Kettler et al, 2007).…”

Genomic plasticity in prokaryotes: the case of the square haloarchaeon

“…Studies carried out mostly with pathogenic isolates (and sometimes non-pathogenic relatives) have revealed highly dynamic genomes indeed (Tettelin et al, 2005;Dempsey et al, 2006;Hochhut et al, 2006;Petrosino et al, 2006;Willenbrock et al, 2006). Bacterial genomes can change dramatically in size, gene repertoire and synteny among the different strains or environmental lineages characterized as belonging to specific, well-defined taxa (Lerat et al, 2005;Thompson et al, 2005;Dempsey et al, 2006;Green and Bohannan, 2006).…”