Resident bacteria are incriminated in the pathogenesis of experimental colitis and inflammatory bowel diseases. We investigated the relative roles of various enteric bacteria populations in the induction and perpetuation of experimental colitis. HLA-B27 transgenic rats received antibiotics (ciprofloxacin, metronidazole, or vancomycin-imipenem) in drinking water or water alone in either prevention or treatment protocols. Mice were treated similarly with metronidazole or vancomycin-imipenem before or after receiving 5% dextran sodium sulfate (DSS). Germfree transgenic rats were colonized with specific-pathogen-free enteric bacteria grown overnight either in anaerobic or aerobic atmospheres. Nontransgenic rats colonized with anaerobic bacteria served as negative controls. Although preventive metronidazole significantly attenuated colitis in transgenic rats and DSS-treated mice, it had no therapeutic benefit once colitis was established. Ciprofloxacin also partially prevented but did not treat colitis in B27 transgenic rats. In both animal models vancomycinimipenem most effectively prevented and treated colitis. Germfree transgenic rats reconstituted with enteric bacteria grown under anaerobic conditions had more aggressive colitis than those associated with aerobic bacteria. These results suggest that a subset of resident luminal bacteria induces colitis, but that a complex interaction of commensal aerobic and anaerobic bacteria provides the constant antigenic drive for chronic immune-mediated colonic inflammation.Rapidly growing evidence supports the influence of normal enteric bacteria on the pathogenic process of intestinal inflammation and extraintestinal manifestations in experimental colitis and human inflammatory bowel diseases (IBD) (40-43). Both spontaneous and induced inflammation in multiple widely diverse rodent models have been associated with commensal luminal bacteria (1, 11-13, 16, 22, 31, 44, 45, 52, 54). The influence of resident bacteria on the induction and perpetuation of spontaneous colitis and gastritis has been thoroughly studied in HLA-B27/␤ 2 -microglobulin transgenic (B27 TG) rats. Colitis, gastritis, and joint inflammation fail to develop in B27 TG rats raised under germfree (sterile) conditions (36, 49). Moreover, when transferred into a specific-pathogenfree (SPF) environment, B27 TG rats universally develop immune-mediated colitis and gastritis within 1 month of bacterial colonization (36).However, not all luminal bacteria have equal abilities to cause inflammation. Antibiotics with narrow specificities, such as metronidazole, which is selectively active against anaerobic bacteria, are effective in Crohn's colitis and ileocolitis (47) and also attenuate chronic experimental intestinal inflammation induced by indomethacin or carageenan in rats and guinea pigs, respectively (32, 54). In addition, overgrowth of predominantly anaerobic bacteria in bypassed small intestinal segments can lead to systemic inflammation. A jejunal self-filling blind loop induces hepatobiliary inflammation rese...

Within the last few years, methicillin-resistant Staphylococcus aureus (MRSA) strains encoding Panton-Valentine leukocidin (PVL) have emerged and spread worldwide. This epidemic can be attributed to a small number of distinct clones. The present study used a novel assay, based on multiplex linear DNA amplification and subsequent microarray hybridisation, to simultaneously detect all relevant exotoxins, antimicrobial resistance determinants and the allelic variants of agr. The genes of the staphylococcal exotoxin-like (set) locus were also included for typing purposes. This assay, together with multilocus sequence typing (MLST) and spa typing, was applied to 56 clinical isolates and reference strains representing all major pandemic PVL-MRSA lineages, as well as to phylogenetically-related strains and putative ancestors. Array hybridisation results allowed the assignment of isolates to clonal groups, which were in accordance with MLST and spa typing data. Ten distinct clonal groups of PVL-MRSA (ST1, ST5, ST8, ST22, ST30, ST59/359, ST80/583, ST88, ST93 and ST152), including 12 MLST types, were identified and analysed with regard to resistance determinants and genes coding for exotoxins. The array hybridisation data confirmed that pandemic PVL-positive strains originate from very diverse genetic backgrounds, and provided insights into the evolution of some lineages. The DNA microarray technique provides a valuable epidemiological tool for the detailed characterisation of clinical isolates and comparison of strains at a global level.

BackgroundTimely identification of pathogens is crucial to minimize mortality in patients with severe infections. Detection of bacterial and fungal pathogens in blood by nucleic acid amplification promises to yield results faster than blood cultures (BC). We analyzed the clinical impact of a commercially available multiplex PCR system in patients with suspected sepsis.MethodsBlood samples from patients with presumed sepsis were cultured with the Bactec 9240™ system (Becton Dickinson, Heidelberg, Germany) and aliquots subjected to analysis with the LightCycler® SeptiFast® (SF) Test (Roche Diagnostics, Mannheim, Germany) at a tertiary care centre. For samples with PCR-detected pathogens, the actual impact on clinical management was determined by chart review. Furthermore a comparison between the time to a positive blood culture result and the SF result, based on a fictive assumption that it was done either on a once or twice daily basis, was made.ResultsOf 101 blood samples from 77 patients, 63 (62%) yielded concordant negative results, 14 (13%) concordant positive and 9 (9%) were BC positive only. In 14 (13%) samples pathogens were detected by SF only, resulting in adjustment of antibiotic therapy in 5 patients (7,7% of patients). In 3 samples a treatment adjustment would have been made earlier resulting in a total of 8 adjustments in all 101 samples (8%).ConclusionThe addition of multiplex PCR to conventional blood cultures had a relevant impact on clinical management for a subset of patients with presumed sepsis.

Acute graft-versus-host disease (aGVHD) often limits feasibility and outcome of allogeneic bone marrow transplantation. Current pathophysiologic concepts of aGVHD involve conditioning regimens, donor-derived T cells, proinflammatory cytokines, and bacterial lipopolysaccharide (LPS) as a major trigger for aGVHD. LPS derives mostly from gram-negative bacteria and can enter circulation through the impaired mucosal barrier after the conditioning regimen. Probiotic microorganisms have been shown to alter the composition of the intestinal microflora and thereby mediate anti-inflammatory effects. We hypothesized that modifying the enteric flora using the probiotic microorganism Lactobacillus rhamnosus GG, would ameliorate aGVHD. Here we show that oral administration of Lactobacillus rhamnosus GG before and after transplantation results in improved survival and reduced aGVHD. Furthermore, subculturing of mesenteric lymph node tissue revealed a reduced translocation of enteric bacteria. Our findings suggest that alteration of the intestinal microflora plays an important role in the initiation of experimental aGVHD. IntroductionAcute graft-versus-host disease (aGVHD) remains one of the major obstacles in allogeneic bone marrow transplantation (BMT). Despite the development of potent immunosuppressive drugs and reduction of conditioning regimens, a high percentage of patients develop aGVHD, resulting in a high mortality after transplantation. Bacterial lipopolysaccharide (LPS) is considered a major player in the development of aGVHD. 1-3 Hence, bowel decontamination using broad-spectrum antibiotics prior to transplantation has been introduced as standard practice to date. [4][5][6] In experimental studies, the proinflammatory potency of LPS varies from bacterial species to species. However, the clinical impact of different intestinal microorganisms on aGVHD is still unclear.Chronic inflammatory bowel disease (IBD) appears to be the result of a genetically determined unbalanced immune response to ubiquitous luminal bacterial antigens. 7 Probiotic bacteria, mainly lactobacilli and bifidobacteria, are defined as living microbes, which confer benefits to the host. There is increasing evidence that probiotic therapy is effective in the treatment of IBD. 8 However, the mechanisms by which these bacteria mediate their effects are still unclear. It has been shown that probiotic bacteria have a temporary impact on the composition of the intestinal flora, but a direct influence on the immune system has also been suggested. [9][10][11] To address the impact of modulation of the bowel flora on aGVHD, we used a well-described murine transplantation model in which aGVHD is induced across a haploidentical major histocompatibility complex (MHC) mismatch. The model is characterized by severe damage of the bowel mucosa, high-serum LPS levels after transplantation, and strong release of proinflammatory cytokines. 12-14 Study designMice, BMT, assessment of GVHD, and treatment protocols C57BL/6 and B6D2F1 mice (8 to 12 weeks old) were purchas...

The aim of the present study was to investigate strains of methicillin-resistant Staphylococcus aureus (MRSA) for the presence of the lukS-lukF determinant of Panton-Valentine leukocidin and to further characterize strains found to contain the genes. During the past 2 years, MRSA containing the lukS-lukF genes for Panton-Valentine leukocidin, particularly those emerging outside of hospitals, have become of interest. MRSA strains sent to the national reference center in Germany were investigated for lukS-lukF by polymerase chain reaction (PCR). If the presence of lukS-lukF was demonstrated, strains were further characterized by molecular typing (determination of SmaI pattern, spa sequence, and multilocus sequence type), PCR demonstration of resistance genes, and characterization of the SCCmec element. Since the end of 2002, MRSA containing Panton-Valentine leukocidin genes have been demonstrated as the causative agent of 28 cases of infection (9 community-acquired cases, 19 sporadic nosocomial cases) in different areas of Germany. Twenty-seven of these 28 isolates exhibited a unique pattern of genomic typing: all exhibited multilocus sequence type 80, spa sequence type 44, and a SmaI macrorestriction pattern that corresponds to a community-acquired strain of MRSA from France and Switzerland. In addition to resistance to oxacillin, the strains exhibited resistance to ciprofloxacin, tetracycline (tetM), and fusidic acid, the last of which is encoded by the far-1 gene. The far-1 gene was shown to be located on the plasmid. One isolate corresponded to community MRSA (cMRSA) of multilocus sequence type 1 from the USA.

The luminal bacterial load and composition determines the activity of cecal inflammation in genetically susceptible hosts. Lowering cecal bacterial concentrations can diminish inflammation in remote organs.

Escherichia coli strain Nissle 1917 (EcN) is as effective in maintaining remission in ulcerative colitis as is treatment with mesalazine. This study aims to evaluate murine models of acute and chronic intestinal inflammation to study the antiinflammatory effect of EcN in vivo. Acute colitis was induced in mice with 2% dextran-sodium sulfate (DSS) in drinking water. EcN was administered from day ؊2 to day ؉7.

A duplex LightCycler PCR assay targeting the mecA gene and a Staphylococcus aureus-specific marker was used to test 165 S. aureus strains and 80 strains of other bacterial species. Within an assay time of 60 min plus 10 min for sample preparation, S. aureus as well as the presence or absence of the mecA gene was correctly identified.