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In this study the plasmid pTC, a 90 kb self-conjugative virulence plasmid of the porcine enterotoxigenic Escherichia coli (ETEC) strain EC2173 encoding the STa and STb heat-stable enterotoxins and tetracycline resistance, has been sequenced in two steps. As a result we identified five main distinct regions of pTC: (i) the maintenance region responsible for the extreme stability of the plasmid, (ii) the TSL (toxin-specific locus comprising the estA and estB genes) which is unique and characteristic for pTC, (iii) a Tn10 transposon, encoding tetracycline resistance, (iv) the tra (plasmid transfer) region, and (v) the colE1-like origin of replication. It is concluded that pTC is a self-transmissible composite plasmid harbouring antibiotic resistance and virulence genes. pTC belongs to a group of large conjugative E. coli plasmids represented by NR1 with a widespread tra backbone which might have evolved from a common ancestor. This is the first report of a completely sequenced animal ETEC virulence plasmid containing an antimicrobial resistance locus, thereby representing a selection advantage for spread of pathogenicity in the presence of antimicrobials leading to increased disease potential., (Copyright © 2011. Published by Elsevier GmbH.)

The enterotoxigenic Escherichia coli (ETEC) strain Ec2173, causing post weaning diarrhoea in swine, harbours six plasmids ranging from 13 to 200 kb in size. The heat stable toxin genes sta, stb and a tetracycline resistance gene were located on a self conjugative 120-kb plasmid, called pTC. In the cloned ColE1 type origin of replication of pTC a deletion was detected compared to other ColE1 replicons affecting the replication modulator gene rom. Epidemiological studies on ETEC isolates showed that pTC-like plasmids are widely distributed among porcine ETEC strains; thus representing an example of co-evolution of antibacterial resistance and virulence in pathogenic E. coli.

Most virulence genes of enterotoxigenic Escherichia coli (ETEC) are located on plasmids. The gene for heat-stable enterotoxin I (sta) is part of the transposon Tn1681, and the heat-stable enterotoxin II (stb) gene was described to be part of the transposon Tn4521. In the studies presented here, we describe the linkage of the sta and stb genes on an approximately 10-kb fragment designated as toxin-specific locus (TSL). The TSL has been isolated from the 120-kb virulence plasmid pTC of the porcine ETEC strain 2173 that produces F18 fimbriae. The nucleotide sequence of the TSL fragment was determined. Sequences in the flanking regions of the sta gene indicated the presence of Tn1681, but--unexpectedly--flanking sequences of the neighbouring stb gene were completely different from those of Tn4521. The 10-kb TSL is part of a 40-kb fragment that contains the replication origin of pTC. This 40-kb fragment was mobilised into plasmid pACYC177 and the nucleotide sequence of the bordering fragments was determined. The 40-kb fragment was flanked by IS10 elements at both ends, indicating the existence of a new 40-kb pathogenicity island (PAI) in strain 2173. Several F4(K88)+ ETEC and F18+ ETEC as well as F18+ E. coli strains producing enterotoxins and verotoxin-2 (ETEC/VTEC) from weaned pigs of different geographical origin were tested for the flanking regions by PCR to see if they belong to the "Tn4521-like" or the "pTC-like" stb type. It turned out that the Tn4521-like stb-type was characteristic of F4(K88)+ ETEC, while the pTC-like stb type was present in most F18+ ETEC and F18+ ETEC/VTEC, although polymorphism was observed both in the K88 and F18 groups. These results suggest the existence and worldwide spread of a new plasmid-encoded pathogenicity island in porcine post weaning ETEC and ETEC/VTEC strains producing F18 fimbriae.

Purpose: Transposition of intestinal segments into the urinary tract predisposes to urinary tract infections. We characterized bacterial infections in these patients and examined the virulence genotype and persistence of Escherichia coli isolates., Materials and Methods: We followed 26 patients who underwent bladder reconstructive surgery using transposed intestinal segments. E. coli strains isolated from the urine of these patients were genotyped for established virulence determinants and the frequency of carriage was compared with E. coli strains isolated from community acquired urinary infections and the fecal flora of anonymous volunteers. A longitudinal study of E. coli strains in 9 patients was also done using pulsed field gel electrophoresis., Results: E. coli was the most frequently isolated organism, responsible for 59% (62 of 105) of monobacterial infections. Other bacteria isolated included Klebsiella species, Proteus species and Enterococcus faecalis. Community acquired E. coli strains were more likely to carry multiple determinants for particular adhesins (P and S fimbriae) and toxins (alpha-hemolysin and cytotoxic necrotizing factor) than fecal strains. Carriage frequency for bladder reconstruction strains was intermediary and not significantly different. The key finding was that E. coli strains persisted for prolonged periods, including 2 years in certain patients, often despite various antimicrobial treatments., Conclusions: This study highlights that further steps must be taken to prevent and treat urinary tract infections in this susceptible group. Particular attention should be given to the treatment of persistent infections.

Suppression subtractive hybridisation (SSH) was performed to identify genomic differences between the uropathogenic Escherichia coli strain 536 and the non-pathogenic E. coli K-12 strain MG1655. In total, 22 DNA fragments were isolated which were specific for strain 536. Five of these fragments showed homology to known virulence determinants and four fragments matched genes for lipopolysaccharide (LPS) or capsule biosynthesis and a siderophore receptor. Seven fragments did not show any homology to known genes. These fragments may represent parts of putative pathogenicity islands (PAIs). Whereas two fragments were highly specific for uropathogenic E. coli (UPEC), the other fragments could also be detected among the other tested wild-type strains.

Various genetic mechanisms including point mutations, genetic rearrangements and lateral gene transfer processes contribute to the evolution of microbes. Long-term processes leading to the development of new species or subspecies are termed macroevolution, and short-term developments, which occur during days or weeks, are considered as microevolution. Both processes, macro- and microevolution need horizontal gene transfer, which is particularly important for the development of pathogenic microorganisms. Plasmids, bacteriophages and so-called pathogenicity islands (PAIs) play a crucial role in the evolution of pathogens. During microevolution, genome variability of pathogenic microbes leads to new phenotypes, which play an important role in the acute development of an infectious disease. Infections due to Staphylococcus epidermidis, Candida albicans and Escherichia coli will be described with special emphasis on processes of microevolution. In contrast, the development of PAIs is a process involved in macroevolution. PAIs are especially important in processes leading to new pathotypes or even species. In this review, particular attention will be given to the fact that the evolution of pathogenic microbes can be considered as a specific example for microbial evolution in general.

Escherichia coli isolates from an immunocompetent woman with a history of repeated amnion infections and spontaneous abortion were characterized. Escherichia coli were isolated from stool, blood and cervix swab samples taken over a 21-month period after the last abortion which followed septicemia during pregnancy. All samples except the last cervix swab contained isolates of serotype O12:K1:H7, which produced adhesins, P fimbriae, type I fimbriae and the iron-chelator aerobactin. Pulsed-field gel electrophoresis revealed identical Xbal restriction patterns of the O12:K1:H7 isolates, suggesting that one particular Escherichia coli strain was responsible for the severe extraintestinal infections during pregnancy. The strain was able to persist in the intestine of the woman despite antibiotic therapy.