Experimental Infection of Calves with Escherichia coli O104:H4 outbreak strain.

In 2011, a severe outbreak of hemolytic-uremic syndrome was caused by an unusual, highly virulent enterohemorrhagic E. coli (EHEC) O104:H4 strain, which possessed EHEC virulence traits in the genetic background of human-adapted enteroaggregative E. coli. To determine magnitude of fecal shedding and site of colonization of EHEC O104:H4 in a livestock host, 30 (ten/strain) weaned calves were inoculated with 10(10) CFU of EHEC O104:H4, EHEC O157:H7 (positive control) or E. coli strain 123 (negative control) and necropsied (4 or 28 d.p.i.). E. coli O157:H7 was recovered until 28 d.p.i. and O104:H4 until 24 d.p.i. At 4 d.p.i., EHEC O104:H4 was isolated from intestinal content and detected associated with the intestinal mucosa. These results are the first evidence that cattle, the most important EHEC reservoir, can also carry unusual EHEC strains at least transiently, questioning our current understanding of the molecular basis of host adaptation of this important E. coli pathovar.

[Evolution and infection biology of hemolytic-uremic syndrome (HUS) associated E. coli (HUSEC)]. / Evolution und Infektionsbiologie der mit dem hämolytisch-urämischen Syndrom (HUS) assoziierten E. coli (HUSEC).

Shiga toxin (Stx)-producing Escherichia coli (STEC), which cause hemolytic-uremic syndrome (HUS), are designated as HUSEC. Their exceptional genome variability driven by evolutionary diversification permits fast adaptation to changed environmental conditions. The HUSEC collection (http://www.ehec.org), which has been established at the Institute for Hygiene in Münster, contains 42 EHEC reference strains (HUSEC001-HUSEC042). It represents a unique repository collection of pathogens and is extremely helpful for the analysis of evolutionary changes and fixed properties in the STEC that cause the most severe host injury. Such genomic attributes include slowly evolving loci, mobile genetic elements that often encode virulence factors and are assimilated via horizontal gene transfer. Current evolutionary models indicate that numerous outbreak strains evolved recently and that highly pathogenic HUSEC descend from less pathogenic progenitors. However, additional data suggest that HUSEC have small effective population sizes. The HUSEC collection is also a valuable resource with which to study important non-Shiga toxin virulence factors.

[Granular-cell tumor (Abrikossoff's tumor): precise preoperative diagnosis is crucial when for minimally-invasive resection]. / Abrikossoff-Granularzelltumor des Ösophagus. Exakte präoperative Diagnostik ist entscheidendfür minimal-invasives Vorgehen.

HISTORY AND ADMISSION FINDINGS: A 38-year-old man was referred to a gastroenterologist because of thoracic pain and dysphagia of uncertain cause. INVESTIGATIONS: Endoscopy revealed a centrally depressed, coarse submucosal tumor, 1.0 cm in diameter, in the anterior wall of the esophagus at about 35 cm from the teeth. Button-hole biopsy revealed histologically and immunohistochemically a granular cell tumor (Abrikossoff's tumor), positive for S-100 protein. CT-staging was unremarkable, except for a thickening of the distal esophageal wall. TREATMENT AND COURSE: Minimally invasive submucosal resection was undertaken using the videoscopic/endoscopic rendezvous technique. Histological examination confirmed complete resection of the tumor. The clinical course was uneventful and endoscopy 6 and 12 weeks after the operation showed complete resection of the tumor. CONCLUSION: An exact preoperative diagnosis beyond a mere description of site and morphology, but also providing the histopathological data creates favorable conditions for planning and performing a minimally invasive resection. Submucosal resection with the videoscopic/endoscopic in rendezvous technique offers the possibility of complete resection, which can often not be achieved in a submucosal tumor by only endoscopic resection.

Large and ongoing outbreak of haemolytic uraemic syndrome, Germany, May 2011.

Since early May 2011, an increased incidence of haemolytic uraemic syndrome (HUS) and bloody diarrhoea related to infections with Shiga toxin-producing Escherichia coli (STEC) has been observed in Germany, with most cases in the north of the country. Cases reported from other European countries had travelled to this area. First results of a case­control study conducted in Hamburg suggest an association between the occurrence of disease and the consumption of raw tomatoes, cucumber and leaf salad.

Do complement factor H 402Y and C7 M allotypes predispose to (typical) haemolytic uraemic syndrome?

Typical haemolytic uraemic syndrome (HUS) is mainly caused by infections with enterohaemorrhagic Escherichia coli, whereas in atypical, nonbacteria-associated HUS, complement plays a dominant role. Recently, complement has also been shown to be involved in typical HUS. In this study, mostly weakly significant associations with homozygosities of complement allotype C7 M and inversely with factor H 402H were found, suggesting that 402Y and C7 M allotypes predispose to (typical) haemolytic uraemic syndrome.

Incidence and risk factors for community-acquired acute gastroenteritis in north-west Germany in 2004.

In developed countries, acute gastroenteritis (AGE) is a major source of morbidity. However, only a few studies have estimated its incidence and the associated medical burden. This population-based study determined the incidence of community-acquired AGE patients seeking medical care and the relative role of various pathogens. Stool samples from patients with AGE presenting to a general practitioner (GP), pediatrician, or specialist in internal medicine for that reason were screened for various bacterial and viral enteropathogens. A control group was established as well. Incidences were calculated by the number of positive patients divided by the general population. The study was performed in north-west Germany in 2004. The incidence of AGE patients requiring medical consultation was 4,020/100,000 inhabitants. Children (<5 years of age) were at the highest risk (13,810/100,000 inhabitants). Of the patients, 6.6% were tested positive for an enteropathogenic bacteria and 17.7% for a viral agent. The predominant pathogens were norovirus (626/100,000) and rotavirus (270/100,000). Salmonella was the most frequently detected bacteria (162/100,000). The results presented confirm AGE and, specifically, AGE of viral origin as a major public health burden in developed countries.

Sorbitol-fermenting enterohaemorrhagic Escherichia coli O157:H- causes another outbreak of haemolytic uraemic syndrome in children.

An outbreak of haemolytic uraemic syndrome (HUS) among children caused by infection with sorbitol-fermenting enterohaemorrhagic Escherichia coli O157:H- (SF EHEC O157:H-) occurred in Germany in 2002. This pathogen has caused several outbreaks so far, yet its reservoir and routes of transmission remain unknown. SF EHEC O157:H- is easily missed as most laboratory protocols target the more common sorbitol non-fermenting strains. We performed active case-finding, extensive exploratory interviews and a case-control study. Clinical and environmental samples were screened for SF EHEC O157:H- and the isolates were subtyped by pulsed-field gel electrophoresis. We identified 38 case-patients in 11 federal states. Four case-patients died during the acute phase (case-fatality ratio 11%). The case-control study could not identify a single vehicle or source. Further studies are necessary to identify the pathogen's reservoir(s). Stool samples of patients with HUS should be tested with an adequate microbiological set-up to quickly identify SF EHEC O157:H-.

[Surveillance of infectious foodborne diseases after enactment of the German Protection against Infection Act in 2001. Potentials and requirements]. / Surveillance Lebensmittel-übertragener Infektionserkrankungen durch das Infektionsschutzgesetz: Möglichkeiten und Anforderungen.

The epidemiology of infectious foodborne diseases has changed. Outbreaks more frequently occur geographically dispersed or protractedly over longer periods of time, and they often appear as a scatter of seemingly sporadic cases. This hampers and delays the identification of their epidemiological link. The surveillance of infectious foodborne diseases has to be refined accordingly to be able to detect these diffuse outbreaks. The German Protection against Infection Act, enacted in 2001, offers the potential of increased sensitivity due to timely electronic reporting of individual cases and detailed data accompanying each report. In addition to a timely and comprehensive reporting system, subtyping of pathogens has become an invaluable tool in identifying epidemiologically linked cases, i.e. outbreaks. Still, the sensitivity of foodborne disease surveillance still hinges on the willingness of physicians to order stool testing for enteric pathogens (and to report suspected outbreaks to local health departments). Without the active participation of physicians, the chance of detecting outbreaks and successfully investigating them is markedly reduced. Consequently, the general preventive strategy would be jeopardised, namely to understand the (often new) mechanisms by which contamination and disease transmission occur well enough to interrupt them.

Molecular and phenotypic profiling of sorbitol-fermenting Escherichia coli O157:H- human isolates from Finland.

This study investigated the occurrence of virulence-associated genes, including stx1, stx2, stx2c, stx2d, stx2e, eae and its subtypes (alpha, beta, gamma, epsilon), efa1, cdt-V cluster, enterohaemorrhagic Escherichia coli (EHEC)-hlyA, katP, espP, etpD, sfpA and the flagellar fliC gene, in nine sorbitol-fermenting (SF), beta-glucuronidase-positive E. coli O157:H- (non-motile) isolates obtained from humans in Finland between 1997 and 2001. In addition, the production of Shiga toxin (Stx), cytolethal distending toxin (CDT)-V and EHEC haemolysin (EHEC-Hly) was studied, and the phage type (PT) and pulsed-field gel electrophoresis (PFGE) types were determined. All nine isolates carried eae-gamma, efa1, EHEC-hlyA, etpD, sfpA and fliC; eight also harboured the cdt-V gene cluster and five were positive for stx2. None of the isolates harboured stx1, stx2c, stx2d, stx2e, katP or espP. All isolates harbouring the corresponding genes also produced Stx2 and CDT-V in titres ranging from 1:32 to 1:128 and from 1:2 to 1:4, respectively. None of the isolates expressed EHEC-Hly on enterohaemolysin agar. Seven isolates belonged to PT88 and two had a PT88 variant pattern. Seven isolates showed a close genetic relationship, with a PFGE similarity index (SI) of 92-98%. Two isolates, temporally the first and last, obtained 5 years apart, were the most divergent (SI of 71% and 85%, respectively). The study demonstrated that SF E. coli O157:H- isolates from Finland are closely related and show a close relationship with SF E. coli O157 strains isolated in Germany. This finding suggests a clonality of SF E. coli O157:H- isolates from different geographical regions.

Urease genes in non-O157 Shiga toxin-producing Escherichia coli: mostly silent but valuable markers for pathogenicity.

The distribution of ureC was investigated among 294 Escherichia coli isolates, comprising 72 strains from the E. coli standard reference collection (ECOR), 62 strains from the diarrhoeagenic E. coli (DEC) collection, and 160 clinical isolates of Shiga toxin-producing E. coli (STEC). The ureC gene was more frequent among STEC isolates harbouring eae than among those lacking eae (p < 0.0001). All clinical STEC isolates of serogroups O111 and O145 contained ureC, but only two of 294 isolates expressed urease activity. The silencing of urease expression could not be linked to a stop codon in ureD. The frequent occurrence of ure genes in eae-positive STEC isolates makes them valuable markers for virulence.

[Surface disinfection in nursing homes--what is really happening? Study of control success in three Duisburg nursing homes]. / Flächendesinfektion in Pflegeheimen, was geschieht wirklich? Studie zur Erfolgskontrolle in drei Duisburger Pflegeheimen.

The effectiveness of cleaning and disinfection of environmental surfaces was evaluated in three nursing homes using bacteriological monitoring. Samples from inmates (nose, throat and wounds) and surface cleaning equipment were also taken. Cleaning solutions, disinfectants and cleaning clothes were found to be highly contaminated in two of three institutions. Referring to the surfaces in some cases disinfection didn't reduce bacterial colony counts and seeded MRSA as a potential pathogen in one nursing home. Six MRSA-positive inmates and identical strains were registered in the environment. MRSA can be used as a marker organism to demonstrate effectiveness of cleaning. To achieve further improvement bacteriological monitoring can help in focussing special cleaning and disinfection related problems.

Serum antibody reactivity against recombinant PrtC of Porphyromonas gingivalis following periodontal therapy.

OBJECTIVES: In 34 patients with chronic periodontitis, the presence of IgA, IgG, and IgG subclass serum antibodies against recombinant PrtC (rPrtC) of Porphyromonas gingivalis was assessed by immunoblot analysis 24 months after therapy. METHODS: rPrtC was produced from P. gingivalis ATTC 33277 using the plasmid pGEX-2T. In addition, intraoral colonization with P. gingivalis was detected by PCR in subgingival plaque and swab samples from buccal mucosae, tonsils and tongue at baseline, 10 d, and 3, 6, 9, 12, 18, and 24 months. RESULTS: All patients were found to harbor P. gingivalis in the oral cavity at least once during the observation period. The identified antibody responses against the rPrtC of P. gingivalis were IgA (97%, i.e. 33/34 patients) and IgG (100%, i.e. 34/34), with an IgG subclass distribution of IgG2 (65%, i.e. 22/34 patients) > IgG3 (47%, i.e. 16/34) > IgG1 (38%, i.e. 13/34) > IgG4 (29%, i.e. 10/34). Anti-rPrtC IgA and IgG antibody reactivity was found in all but one patients (anti-rPrtC IgA negative), who tested negative for P. gingivalis at all of the assessed intraoral sites for at least 6 months before sera collection. There was no association between IgG subclass reactivity against the rPrtC of P. gingivalis and progression of periodontal attachment loss. CONCLUSION: The results indicated that anti-rPrtC IgA and IgG antibodies may serve as an indicator for past or present intraoral colonization with P. gingivalis.

Guided tissue regeneration using a polylactic acid barrier.

OBJECTIVES: The purpose of this study was to determine the relative impact of various predictors responsible for the variability in treatment outcome after guided tissue regeneration (GTR) in intraosseous periodontal defects. PATIENTS AND METHODS: 30 patients with chronic periodontitis and at least one intraosseous periodontal lesion (> or =4 mm) were enrolled. Following full-mouth scaling, GTR using polylactic acid membranes was performed at one site in each patient. Main periodontal pathogens, defect morphology, membrane exposure and smoking habit were assessed as predictor variables. Alveolar bone level change served as the primary outcome variable in a multiple regression analysis. RESULTS: After 12 months, the 29 patients completing the study showed alveolar bone changes ranging from 4 mm bone gain to 1 mm bone loss (mean: 1.6+/-0.4 mm gain). Active smoking (beta-weight:-0.49, P=0.003) and persistence of subgingival infection with P. gingivalis (P.g.) (beta-weight:-0.25, P=0.11) were associated with poor treatment outcome. Deep initial intraosseous defects (beta-weight: 0.32, P=0.045) were associated with favorable treatment outcome, and membrane exposure had no impact on bone gain. CONCLUSION: Active smoking was the strongest predictor variable negatively affecting alveolar bone gain following GTR in the treatment of periodontal defects. It was followed by a positive influence of a deeper intraosseous defect and by a negative effect by persistent subgingival infection of P. gingivalis. The relative impact of these factors may be useful in assessing the prognosis of GTR in intraosseous periodontal defects.

Guided tissue regeneration using a polylactic acid barrier. Part I: Environmental effects on bacterial colonization.

OBJECTIVES: The purpose of this study was to assess the dynamics of bacterial colonization in intra-osseous defects following guided tissue regeneration (GTR) therapy using a resorbable barrier. PATIENTS AND METHODS: In each of 30 patients, one intra-osseous defect was treated with GTR using a polylactic acid membrane (Guidor). Plaque samples were taken from the defect site, other teeth and mucous membranes following initial therapy (baseline), and at 3, 6 and 12 months after periodontal surgery. Additionally, samples were taken from the defect sites at 1, 2 and 4 weeks. Actinobacillus actinomycetemcomitans (A.a.), Porphyromonas gingivalis (P.g.), and Bacteroides forsythus (B.f.) were detected by polymerase chain reaction (PCR). Supportive periodontal therapy was performed at 3-month intervals. RESULTS: In the 29 patients completing the study, the assessed microflora was detected in 3 (A.a.), 13 (P.g.) and 14 (B.f.) defect sites at baseline, in 2 (A.a.), 2 (P.g.) and 2 (B.f.) following surgical debridement, and in 6 (A.a.), 10 (P.g.) and 22 (B.f.) at 12 months. Defect site colonization following GTR therapy was significantly correlated with presurgical colonization at other assessed teeth (A.a. and P.g.: tau = 0.45 and 0.66, respectively; P < 0.001), or on mucous membranes (B.f.: tau = 0.44, P < 0.001). CONCLUSION: The colonization of periodontal pathogens at sites treated by GTR may correlate with the intra-oral presence of these pathogens before surgery. If colonization of GTR sites by periodontal pathogens is to be prevented, intra-oral suppression/eradication of these pathogens may be required before surgery.

The high-pathogenicity island is absent in human pathogens of Salmonella enterica subspecies I but present in isolates of subspecies III and VI.

In this study we tested 74 Salmonella strains of all eight Salmonella groups and were able to demonstrate the presence of two high-pathogenicity island types in strains of Salmonella groups IIIa, IIIb, and VI. Most high-pathogenicity island-positive isolates produced yersiniabactin under iron-limited conditions and were positive for the high-molecular-weight proteins HMWP1 and HMWP2.

Genetic diversity of intimin genes of attaching and effacing Escherichia coli strains.

In this study, we determined the sequences of four intimin variant genes detected in attaching and effacing Escherichia coli isolates of human origin. Three of them were novel and were designated eae-eta (eta), eae-iota (iota), and eae-kappa (kappa). The fourth was identical to the recently described eae-zeta (zeta), isolated from a bovine E. coli O84:NM isolate. We compared these sequences with those of published intimin-alpha, intimin-beta, intimin-gamma1, intimin-gamma2, intimin- epsilon, and intimin-theta alleles. Sequence analysis of these 10 intimin alleles confirmed extensive genetic diversity within the intimin gene family in E. coli. The genetic diversity was more prominent in the 3' region (starting at bp 2,112), which encodes the binding domain of intimin. Phylogenetic analyses revealed four groups of closely related intimin genes: alpha and zeta; beta and kappa; gamma1 and gamma2/theta; and epsilon and eta. Calculation of homoplasy ratios of sequences of the 5' region of eae (positions 1 to 2,111) revealed evidence for intragenic recombination. Split decomposition analysis also indicates that recombination events have played a role in the evolutionary history of eae. In conclusion, we recommend an eae nomenclature system based on the Greek alphabet and provide an updated PCR scheme for amplification and typing of E. coli eae.

A long-term study on the prevalence of shiga toxin-producing Escherichia coli (STEC) on four German cattle farms.

The occurrence of Shiga toxin-producing Escherichia coli (STEC) was studied on four cattle farms. STEC were detected in 29-82% of the cattle. STEC with additional EHEC markers were detected on all farms. The occurrence of the complete virulence marker pattern (stx1 and/or stx2, eae, EHEC(hlyA), katP, espP) was correlated with the presence of known STEC serotypes. STEC O26:H11 and O165:H25 with the complete pattern of virulence markers were the most prevalent. STEC O157 (H7/H-) STEC O103:H2 and STEC O145:H- were found sporadically. Five clonal subgroups of the STEC O26:H11 isolates were identified by pulsed-field gel electrophoresis. STEC O26:H11 were present in three groups of cattle. This serotype was detected in a single group over the entire fattening period. Most STEC O26:H11 with the complete pattern of potential virulence markers were found in clinically healthy cattle. These animals may represent a risk factor for farmers and consumers.

[Not Available]. / Infektionen des Menschen durch enterohämorrhagische Escherichia coli (EHEC) in der Bundesrepublik Deutschland von 1998 bis 2001 Prävalenz und Typenspektrum : Prävalenz und Typenspektrum.

Intestinal infections in Germany due to enterohemorrhagic E. coli bacteria (EHEC) between 1998 and 2001 reveal a large scale of biological diversity of their pathogens. However, no dramatic increase of their clinical importance and public health implications has been observed. As strains of serovar O157:H7 have continuously declined as causative agents, other serovars such as O26:H11 and O103:H2 have replaced them. The great diversity of the EHEC pathogens might point to a great number of various infection routes and sources. Since recently new pathogenic factors of EHEC bacteria have been detected (especially by the sequencing of the genome of EHEC), it is currently not possible to define a clear-cut difference between human pathogens and nonhuman pathogens. The enhanced surveillance of EHEC pathogens with respect to their biological diversity and dynamics, their epidemic spread, and their infection routes and sources remain an essential task of the public health authorities.

Identification and characterization of a novel genomic island integrated at selC in locus of enterocyte effacement-negative, Shiga toxin-producing Escherichia coli.

The selC tRNA gene is a common site for the insertion of pathogenicity islands in a variety of bacterial enteric pathogens. We demonstrate here that Escherichia coli that produces Shiga toxin 2d and does not harbor the locus of enterocyte effacement (LEE) contains, instead, a novel genomic island. In one representative strain (E. coli O91:H(-) strain 4797/97), this island is 33,014 bp long and, like LEE in E. coli O157:H7, is integrated 15 bp downstream of selC. This E. coli O91:H(-) island contains genes encoding a novel serine protease, termed EspI; an adherence-associated locus, similar to iha of E. coli O157:H7; an E. coli vitamin B12 receptor (BtuB); an AraC-type regulatory module; and four homologues of E. coli phosphotransferase proteins. The remaining sequence consists largely of complete and incomplete insertion sequences, prophage sequences, and an intact phage integrase gene that is located directly downstream of the chromosomal selC. Recombinant EspI demonstrates serine protease activity using pepsin A and human apolipoprotein A-I as substrates. We also detected Iha-reactive protein in outer membranes of a recombinant clone and 10 LEE-negative, Shiga toxin-producing E. coli (STEC) strains by immunoblot analysis. Using PCR analysis of various STEC, enteropathogenic E. coli, enterotoxigenic E. coli, enteroaggregative E. coli, uropathogenic E. coli, and enteroinvasive E. coli strains, we detected the iha homologue in 59 (62%) of 95 strains tested. In contrast, espI and btuB were present in only two (2%) and none of these strains, respectively. We conclude that the newly described island occurs exclusively in a subgroup of STEC strains that are eae negative and contain the variant stx(2d )gene.