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Ulrich Dobrindt

Publications and source records attributed to Ulrich Dobrindt.

36 records · Page 2Linked to original sources
PubMed2004

Pathogenomics of mobile genetic elements of toxigenic bacteria.

The growing knowledge of genetic diversity and whole genome organization in bacteria shows that pathogenicity islands (PAIs) represent a subtype of a more general genetic element, termed genomic island (GEI), which is widespread among pathogenic and non-pathogenic microbes. These findings mirror the importance of horizontal gene transfer, genome reduction and recombination events as fundamental mechanisms involved in evolution of bacterial variants. GEIs are part of the flexible gene pool and carry selfish genes, but also determinants which may be beneficial under certain conditions thus increasing bacterial fitness and consequently their survival or transmission. In this review, we focus on the role of mobile genetic elements that may also contain toxin-encoding genes for genome variability and evolution of bacteria.

Bacterial Toxins↗
PubMed2004

Excision of the high-pathogenicity island of Yersinia pseudotuberculosis requires the combined actions of its cognate integrase and Hef, a new recombination directionality factor.

The Yersinia high-pathogenicity island (HPI) encodes the siderophore yersiniabactin-mediated iron uptake system. The HPI of Yersinia pseudotuberculosis I has previously been shown to be able to excise precisely from the bacterial chromosome by recombination between the attB-R and attB-L sites flanking the island. However, the nature of the Y. pseudotuberculosis HPI excision machinery remained unknown. We show here that, upon excision, the HPI forms an episomal circular molecule. The island thus has the ability to excise from the chromosome, circularize and reintegrate itself, either in the same location or in another asn tRNA copy. We also demonstrate that the HPI-encoded bacteriophage P4-like integrase (Int) plays a critical role in HPI excision and that, like phage integrases, it acts as a site-specific recombinase that catalyses both excision and integration reactions. However, Int alone cannot efficiently promote recombination between the attB-R and attB-L sites, and we demonstrate that a newly identified HPI-borne factor, designated Hef (for HPI excision factor) is also required for this activity. Hef belongs to a family of recombination directionality factors. Like the other members of this family, Hef probably plays an architectural rather than a catalytic role and promotes HPI excision from the chromosome by driving the function of Int towards an excisionase activity. The fact that the HPI, and probably several other pathogenicity islands, carry a machinery of integration/excision highly similar to those of bacteriophages argues for a phage-mediated acquisition and transfer of these elements.

Amino Acid Sequence↗
PubMed2004

The pathogenicity island-associated K15 capsule determinant exhibits a novel genetic structure and correlates with virulence in uropathogenic Escherichia coli strain 536.

The K15 capsule determinant of uropathogenic Escherichia coli strain 536 (O6:K15:H31) is part of a novel 79.6-kb pathogenicity island (PAI) designated PAI V536 that is absent from the genome of nonpathogenic E. coli K-12 strain MG1655. PAI V536 shows typical characteristics of a composite PAI that is associated with the pheV tRNA gene and contains the pix fimbriae determinant as well as genes coding for a putative phosphoglycerate transport system, an autotransporter protein, and hypothetical open reading frames. A gene cluster coding for a putative general secretion pathway system, together with a kps(K15) determinant, is localized downstream of a truncated pheV gene ('pheV) also present in this chromosomal region. The distribution of genes present on PAI V536 was studied by PCR in different pathogenic and nonpathogenic E. coli isolates of various sources. Analysis of the 20-kb kps locus revealed a so far unknown genetic organization. Generally, the kps(K15) gene cluster resembles that of group 2 and 3 capsules, where two conserved regions (regions 1 and 3) are located up- or downstream of a highly variable serotype-specific region (region 2). Interestingly, recombination of a group 2 and 3 determinant may have been involved in the evolution of the K15 capsule-encoding gene cluster. Expression of the K15 capsule is important for virulence in a murine model of ascending urinary tract infection but not for serum resistance of E. coli strain 536.

Animals↗
PubMed2004

Oral immunization with an rfaH mutant elicits protection against salmonellosis in mice.

Loss of the transcriptional antiterminator RfaH results in virulence attenuation (>10(4)-fold increase in 50% lethal dose) of the archetypal Salmonella enterica serovar Typhimurium strain SL1344 by both orogastric and intraperitoneal routes of infection in BALB/c mice. Oral immunization with the mutant efficiently protects mice against a subsequent oral infection with the wild-type strain. Interestingly, in vitro immunoreactivity is not confined to strain SL1344; rather, it is directed also towards other serovars of S. enterica and even Salmonella bongori strains.

Administration, Oral↗
PubMed2004

Instability of pathogenicity islands in uropathogenic Escherichia coli 536.

The uropathogenic Escherichia coli strain 536 carries at least five genetic elements on its chromosome that meet all criteria characteristic of pathogenicity islands (PAIs). One main feature of these distinct DNA regions is their instability. We applied the so-called island-probing approach and individually labeled all five PAIs of E. coli 536 with the counterselectable marker sacB to evaluate the frequency of PAI-negative colonies under the influence of different environmental conditions. Furthermore, we investigated the boundaries of these PAIs. According to our experiments, PAI II536 and PAI III536 were the most unstable islands followed by PAI I536 and PAI V536, whereas PAI IV536 was stable. In addition, we found that deletion of PAI II536 and PAI III536 was induced by several environmental stimuli. Whereas excision of PAI I536, PAI II536, and PAI V536 was based on site-specific recombination between short direct repeat sequences at their boundaries, PAI III536 was deleted either by site-specific recombination or by homologous recombination between two IS100-specific sequences. In all cases, deletion is thought to lead to the formation of nonreplicative circular intermediates. Such extrachromosomal derivatives of PAI II536 and PAI III536 were detected by a specific PCR assay. Our data indicate that the genome content of uropathogenic E. coli can be modulated by deletion of PAIs.

Chromosomal Instability↗
PubMed2004

Analysis of the genome structure of the nonpathogenic probiotic Escherichia coli strain Nissle 1917.

Nonpathogenic Escherichia coli strain Nissle 1917 (O6:K5:H1) is used as a probiotic agent in medicine, mainly for the treatment of various gastroenterological diseases. To gain insight on the genetic level into its properties of colonization and commensalism, this strain's genome structure has been analyzed by three approaches: (i) sequence context screening of tRNA genes as a potential indication of chromosomal integration of horizontally acquired DNA, (ii) sequence analysis of 280 kb of genomic islands (GEIs) coding for important fitness factors, and (iii) comparison of Nissle 1917 genome content with that of other E. coli strains by DNA-DNA hybridization. PCR-based screening of 324 nonpathogenic and pathogenic E. coli isolates of different origins revealed that some chromosomal regions are frequently detectable in nonpathogenic E. coli and also among extraintestinal and intestinal pathogenic strains. Many known fitness factor determinants of strain Nissle 1917 are localized on four GEIs which have been partially sequenced and analyzed. Comparison of these data with the available knowledge of the genome structure of E. coli K-12 strain MG1655 and of uropathogenic E. coli O6 strains CFT073 and 536 revealed structural similarities on the genomic level, especially between the E. coli O6 strains. The lack of defined virulence factors (i.e., alpha-hemolysin, P-fimbrial adhesins, and the semirough lipopolysaccharide phenotype) combined with the expression of fitness factors such as microcins, different iron uptake systems, adhesins, and proteases, which may support its survival and successful colonization of the human gut, most likely contributes to the probiotic character of E. coli strain Nissle 1917.

Adhesins, Bacterial↗
PubMed2003

Prokaryotic chromosomes and disease.

Recent insights into bacterial genome organization and function have improved our understanding of the nature of pathogenic bacteria and their ability to cause disease. It is becoming increasingly clear that the bacterial chromosome constantly undergoes structural changes due to gene acquisition and loss, recombination, and mutational events that have an impact on the pathogenic potential of the bacterium. Even though the bacterial genome includes additional genetic elements, the chromosome represents the most important entity in this context. Here, we will show that various processes of genomic instability have an influence on the many manifestations of infectious disease.

Adaptation, Physiological↗
PubMed2003

Bacteroides vulgatus protects against Escherichia coli-induced colitis in gnotobiotic interleukin-2-deficient mice.

BACKGROUND & AIMS: The microflora plays a crucial role in inflammatory bowel diseases (IBDs). Specific pathogen-free (SPF), but not germ-free, interleukin (IL)-2-deficient (IL-2-/-) mice develop colitis. The colitogenicity of commensal bacteria was determined. METHODS: Gnotobiotic IL-2-/- and IL-2+/+ mice were colonized with Escherichia coli mpk, Bacteroides vulgatus mpk, or both bacterial strains, or with E. coli strain Nissle 1917. DNA arrays were used to characterize E. coli mpk. Colitis was analyzed by histology and real-time reverse-transcription polymerase chain reaction (RT-PCR) for interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, IL-10, and CD14 messenger RNA (mRNA) expression. Bacterial numbers in feces and bacterial localization in the colon was determined by culture and fluorescence in situ hybridization (FISH). RESULTS: IL-2-/- but not IL-2+/+ mice monocolonized with E. coli mpk developed colitis, whereas mono-association with B. vulgatus mpk, or E. coli Nissle, or co-colonization with E. coli mpk and B. vulgatus mpk, did not induce colitis. DNA array experiments and cellular studies revealed that E. coli mpk is a nonpathogenic strain. FISH and culture methods revealed that the anticolitogenic effect of B. vulgatus mpk on E. coli mpk cannot be explained by a significant reduction in numbers of E. coli in the colon. E. coli mpk-induced colitis was associated with increased IFN-gamma, TNF-alpha, CD14, and IL-10 mRNA expression in the colon. CONCLUSIONS: In IL-2-/- mice, B. vulgatus mpk protects against E. coli mpk-triggered colitis by an unknown mechanism. E. coli Nissle does not induce colitis. Various bacterial species common to the microflora differ in their ability to trigger IBD.

Animals↗
PubMed2003

Commensal bacteria make a difference.

Nowhere is the relationship between microorganisms and eukaryotes as diverse, intimate and clinically relevant as in the gastrointestinal tract. An estimated 500-1000 mostly anaerobic species reside in the intestine, approaching enormous densities of 10(11)-10(12) organisms per gram colon content. The commensal interactions between intestinal microorganisms and animal hosts have been difficult to study in the past because of the diversity of microorganisms involved and because of the lack of culturability that accompanies many microbial consortia. However, recent work has provided new insights into these interactions.

Anaerobiosis↗
PubMed2003

Cytolethal distending toxin gene cluster in enterohemorrhagic Escherichia coli O157:H- and O157:H7: characterization and evolutionary considerations.

We identified a cytolethal distending toxin (cdt) gene cluster in 87, 6, and 0% of sorbitol-fermenting (SF) enterohemorrhagic Escherichia coli (EHEC) O157:H(-), EHEC O157:H7, and E. coli O55:H7/H(-) strains, respectively. The toxin was expressed by the wild-type EHEC O157 strains and by a cdt-containing cosmid from a library of SF EHEC O157:H(-) strain 493/89. The cdt flanks in strain 493/89 were homologous to bacteriophages P2 and lambda. Our data demonstrate that cdt, encoding a potential virulence factor, is present in the EHEC O157 complex and suggest that cdt may have been acquired by phage transduction.

Bacterial Toxins↗
PubMed2003

Analysis of genome plasticity in pathogenic and commensal Escherichia coli isolates by use of DNA arrays.

Genomes of prokaryotes differ significantly in size and DNA composition. Escherichia coli is considered a model organism to analyze the processes involved in bacterial genome evolution, as the species comprises numerous pathogenic and commensal variants. Pathogenic and nonpathogenic E. coli strains differ in the presence and absence of additional DNA elements contributing to specific virulence traits and also in the presence and absence of additional genetic information. To analyze the genetic diversity of pathogenic and commensal E. coli isolates, a whole-genome approach was applied. Using DNA arrays, the presence of all translatable open reading frames (ORFs) of nonpathogenic E. coli K-12 strain MG1655 was investigated in 26 E. coli isolates, including various extraintestinal and intestinal pathogenic E. coli isolates, 3 pathogenicity island deletion mutants, and commensal and laboratory strains. Additionally, the presence of virulence-associated genes of E. coli was determined using a DNA "pathoarray" developed in our laboratory. The frequency and distributional pattern of genomic variations vary widely in different E. coli strains. Up to 10% of the E. coli K-12-specific ORFs were not detectable in the genomes of the different strains. DNA sequences described for extraintestinal or intestinal pathogenic E. coli are more frequently detectable in isolates of the same origin than in other pathotypes. Several genes coding for virulence or fitness factors are also present in commensal E. coli isolates. Based on these results, the conserved E. coli core genome is estimated to consist of at least 3,100 translatable ORFs. The absence of K-12-specific ORFs was detectable in all chromosomal regions. These data demonstrate the great genome heterogeneity and genetic diversity among E. coli strains and underline the fact that both the acquisition and deletion of DNA elements are important processes involved in the evolution of prokaryotes.

Electrophoresis, Gel, Pulsed-Field↗
PubMed2002

Identification and distribution of the enterohemorrhagic Escherichia coli factor for adherence (efa1) gene in sorbitol-fermenting Escherichia coli O157: H-.

Sorbitol-fermenting (SF) Shiga toxin (Stx)-producing Escherichia coli (STEC) O157:H- strains are emerging as causes of hemorrhagic colitis and the hemolytic-uremic syndrome in Europe. Using subtractive hybridization between SF STEC O157:H- strain 493/89 and STEC O157:H7 strain EDL933, three different fragments, of approximately 700 bp in length, were identified. Each demonstrated > 99% homology to genes encoding the enterohemorrhagic E. coli factor for adherence (efa1) and lymphostatin (lifA). Therefore, a cosmid library was constructed from SF STEC O157:H- strain 493/89, and one clone containing these fragments was sequenced. This sequencing demonstrated a 9669-bp open reading frame (ORF) that had 99.9% sequence homology to efa1 of STEC O111:H- strain E45035 and to lifA of an enteropathogenic E. coli O127:H6 strain E2348/69. In STEC O157:H7 strain EDL933, only small (ca. 3 kb) initial and terminal fragments of this ORF are present. PCR analysis with primers complementary to the efa1/lifA sequence of strain 493/89 indicated that the complete sequence is present in each of 10 SF STEC O157:H- isolates but in none of 10 STEC O157:H7 strains investigated. The presence of the complete efa1/lifA also in both tested E. coli O55:H7 strains supports the hypothesis that SF STEC O157:H- are phylogenetically closer to the proposed E. coli O55:H7 ancestor than STEC O157:H7. Our data demonstrate the presence of a potential virulence gene in SF STEC O157:H- that is only rudimentarily present in STEC O157:H7.

Bacterial Proteins↗
PubMed2002

Virulence factors of uropathogens.

Urinary tract infections are among the most frequent infections encountered in developed countries. The vast majority of community-acquired urinary tract infections are caused by Escherichia coli. However, other bacterial species play an important role in nosocomial urinary tract infections. All these species are equipped with a variety of virulence factors. The best characterized are those from Escherichia coli. Among the first virulence factors that come into play during establishment of a urinary tract infection are adhesins. Besides their primary function as adhesin molecules several other additional functions can now be attributed to these organelles. Adhesins may also function as invasins, promote biofilm formation and transmit signals to epithelial cells resulting in inflammation. Furthermore, subunit proteins of adhesins seem to be promising vaccines. Later in infection, toxins seem to enhance virulence. However, for cytotoxic necrotizing factor type 1 this is controversial. Many virulence factors of uropathogenic bacteria are encoded by foreign DNA stretches inserted into the core genome. These pathogenicity islands or islets were obviously acquired via horizontal gene transfer creating new pathotypes more efficient in establishing infection. The role of new virulence factors and the new functions of already known virulence factors will be discussed as well as the concept of the composite genome of uropathogenic Escherichia coli.

Adhesins, Escherichia coli↗
PubMed2002

Genetic structure and distribution of four pathogenicity islands (PAI I(536) to PAI IV(536)) of uropathogenic Escherichia coli strain 536.

For the uropathogenic Escherichia coli strain 536 (O6:K15:H31), the DNA sequences of three pathogenicity islands (PAIs) (PAI I(536) to PAI III(536)) and their flanking regions (about 270 kb) were determined to further characterize the virulence potential of this strain. PAI I(536) to PAI III(536) exhibit features typical of PAIs, such as (i) association with tRNA-encoding genes; (ii) G+C content differing from that of the host genome; (iii) flanking repeat structures; (iv) a mosaic-like structure comprising a multitude of functional, truncated, and nonfunctional putative open reading frames (ORFs) with known or unknown functions; and (v) the presence of many fragments of mobile genetic elements. PAI I(536) to PAI III(536) range between 68 and 102 kb in size. Although these islands contain several ORFs and known virulence determinants described for PAIs of other extraintestinal pathogenic E. coli (ExPEC) isolates, they also consist of as-yet-unidentified ORFs encoding putative virulence factors. The genetic structure of PAI IV(536), which represents the core element of the so-called high-pathogenicity island encoding a siderophore system initially identified in pathogenic yersiniae, was further characterized by sample sequencing. For the first time, multiple PAI sequences (PAI I(536) to PAI IV(536)) in uropathogenic E. coli were studied and their presence in several wild-type E. coli isolates was extensively investigated. The results obtained suggest that these PAIs or at least large fragments thereof are detectable in other pathogenic E. coli isolates. These results support our view that the acquisition of large DNA regions, such as PAIs, by horizontal gene transfer is an important factor for the evolution of bacterial pathogens.

Codon↗
PubMed2002

Loss of regulatory protein RfaH attenuates virulence of uropathogenic Escherichia coli.

RfaH is a regulatory protein in Escherichia coli and Salmonella enterica serovar Typhimurium. Although it enhances expression of different factors that are proposed to play a role in bacterial virulence, a direct effect of RfaH on virulence has not been investigated so far. We report that inactivation of rfaH dramatically decreases the virulence of uropathogenic E. coli strain 536 in an ascending mouse model of urinary tract infection. The mortality rate caused by the wild-type strain in this assay is 100%, whereas that of its isogenic rfaH mutant does not exceed 18%. In the case of coinfection, the wild-type strain 536 shows higher potential to colonize the urinary tract even when it is outnumbered 100-fold by its rfaH mutant in the inoculum. In contrast to the wild-type strain, serum resistance of strain 536rfaH::cat is fully abolished. Furthermore, we give evidence that, besides a major decrease in the amount of hemin receptor ChuA (G. Nagy, U. Dobrindt, M. Kupfer, L. Emody, H. Karch, and J. Hacker, Infect. Immun. 69:1924-1928, 2001), loss of the RfaH protein results in an altered lipopolysaccharide phenotype as well as decreased expression of K15 capsule and alpha-hemolysin, whereas levels of other pathogenicity factors such as siderophores, flagella, Prf, and S fimbriae appear to be unaltered in strain 536rfaH::cat in comparison to the wild-type strain. trans complementation of the mutant strain with the rfaH gene restores wild-type levels of the affected virulence factors and consequently restitutes virulence in the mouse model of ascending urinary tract infection.

Animals↗
PubMed2002

A single nucleotide exchange in the wzy gene is responsible for the semirough O6 lipopolysaccharide phenotype and serum sensitivity of Escherichia coli strain Nissle 1917.

Structural analysis of lipopolysaccharide (LPS) isolated from semirough, serum-sensitive Escherichia coli strain Nissle 1917 (DSM 6601, serotype O6:K5:H1) revealed that this strain's LPS contains a bisphosphorylated hexaacyl lipid A and a tetradecasaccharide consisting of one E. coli O6 antigen repeating unit attached to the R1-type core. Configuration of the GlcNAc glycosidic linkage between O-antigen oligosaccharide and core (beta) differs from that interlinking the repeating units in the E. coli O6 antigen polysaccharide (alpha). The wa(*) and wb(*) gene clusters of strain Nissle 1917, required for LPS core and O6 repeating unit biosyntheses, were subcloned and sequenced. The DNA sequence of the wa(*) determinant (11.8 kb) shows 97% identity to other R1 core type-specific wa(*) gene clusters. The DNA sequence of the wb(*) gene cluster (11 kb) exhibits no homology to known DNA sequences except manC and manB. Comparison of the genetic structures of the wb(*)(O6) (wb(*) from serotype O6) determinants of strain Nissle 1917 and of smooth and serum-resistant uropathogenic E. coli O6 strain 536 demonstrated that the putative open reading frame encoding the O-antigen polymerase Wzy of strain Nissle 1917 was truncated due to a point mutation. Complementation with a functional wzy copy of E. coli strain 536 confirmed that the semirough phenotype of strain Nissle 1917 is due to the nonfunctional wzy gene. Expression of a functional wzy gene in E. coli strain Nissle 1917 increased its ability to withstand antibacterial defense mechanisms of blood serum. These results underline the importance of LPS for serum resistance or sensitivity of E. coli.

Alleles↗
PubMed2002

Influence of the leuX-encoded tRNA5(Leu) on the regulation of gene expression in pathogenic Escherichia coli.

The leuX gene encoding the minor tRNA5(Leu) is important for the expression of several virulence factors of pathogenic Escherichia coli strains. The differential usage of minor codons to control the expression of specialized genes has been proposed to be a general mechanism of bacteria to regulate gene expression at the posttranscriptional level. The minor codon usage theory foots on the biased codon usage of bacterial genes and the selective availability of tRNA isoacceptors. We aimed at the further investigation of the regulatory role of the tRNA5(Leu) for gene expression in pathogenic E. coli. For this purpose, the molecular mechanism underlying the tRNA5(Leu)-dependent regulation of different virulence-associated genes of pathogenic E. coli as well as the regulation of leuX transcription under various growth conditions were investigated in detail. The global impact of the presence or absence of the leuX-encoded tRNA on gene expression of the uropathogenic E. coli strain 536 was studied by proteome analysis. The obtained results argue for a general importance of the tRNA5(Leu) for gene expression of E. coli and the involvement of this tRNA in global regulatory networks.

Animals↗