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PubMed2001

Infection of hamsters with epidemiologically important strains of Clostridium difficile.

Five different toxigenic strains of Clostridium difficile of known human epidemiologic importance were tested for virulence in hamsters. Three strains-types B1, J9, and K14-have caused hospital outbreaks. Type Y2 is associated with a high rate of asymptomatic colonization in patients. The fifth strain, type CF2, is a toxin A-negative, toxin B-positive strain implicated in multiple human cases of C. difficile-associated diarrhea. Groups of 10 hamsters per strain were given 1 dose of clindamycin, followed 5 days later with gastric inoculation of 100 cfu of C. difficile. Hamsters given types B1, J9, K14, or Y2 showed 90%-100% colonization (albeit at a slower rate with type Y2) and 100% mortality of colonized animals. Hamsters challenged with type CF2 showed 60% (P= .01) colonization and 30% mortality (P= .0003). The hamster model demonstrated pathogenicity differences between a toxin variant strain and standard toxigenic strains but no significant differences among the standard strains.

Animals↗
PubMed2001

Molecular typing methods for the epidemiological identification of Clostridium difficile strains.

Toxigenic Clostridium difficile is the etiologic agent of C. difficile-associated diarrhea (CDAD), the most common cause of nosocomial diarrhea. Cross-infection between patients and transmission through the environment and medical personnel are important factors in the acquisition of CDAD. In order to understand differences in epidemiology and pathogenesis, a number of typing schemes have been developed. We will review the typing methods used to study the epidemiology of C. difficile infections and how they have evolved from a phenotypic identification to state of the art molecular methods, detecting genetic polymorphisms among strains. These molecular methods include PCR-based methods (arbitrarily primed-PCR [AP-PCR] and PCR ribotyping), restriction endonuclease analysis (REA) and pulse field gel electrophoresis (PFGE). The application, usefulness and feasibility of these methods are compared and discussed. Finally, the role of genomics as a tool to investigate CDAD is introduced.

Bacterial Typing Techniques↗
PubMed2001

Clostridium difficile-associated diarrhea and colitis.

Clostridium difficile is a spore-forming toxigenic bacterium that causes diarrhea and colitis, typically after the use of broad-spectrum antibiotics. The clinical presentation ranges from self-limited diarrhea to fulminant colitis and toxic megacolon. The incidence of this disease is increasing, resulting in major medical and economic consequences. Although most cases respond quickly to medical treatment, C difficile colitis may be serious, especially if diagnosis and treatment are delayed. Recurrent disease represents a particularly challenging problem. Prevention is best accomplished by limiting the use of broad-spectrum antibiotics and following good hygienic techniques and universal precautions to limit the transmission of bacteria. A high index of suspicion results in early diagnosis and treatment and potentially reduces the incidence of complications.

Adult↗
PubMed2006

Prophylactic Saccharomyces boulardii in the prevention of antibiotic-associated diarrhea: a prospective study.

BACKGROUND: Interest to probiotics for the prevention and treatment of antibiotic-associated diarrhea is increasing gradually. The most promising seems to be Saccharomyces boulardii . Using a double-blind controlled study, we investigated the preventive effect of S. boulardii on the development of antibiotic-associated diarrhea in patients under antibiotherapy but not requiring intensive care therapy. MATERIAL/METHODS: All the patients were hospitalized at the Gulhane Military Medical Academy, Department of Infectious Diseases and Clinical Microbiology. S. boulardii was given twice daily during the course of antibiotic therapy and application was initiated in all patients as late as after 48 hours of antibiotic therapy. A total of 151 patients completed the study. RESULTS: The antibiotic-associated diarrhea development ratio in placebo group was 9% (7/78) and in the study group 1.4% (1/73) (p < 0.05). Stool samples from the patients with antibiotic-associated diarrhea were stored at -70 degrees C and Clostiridium difficile toxin A assay was performed using Enzyme Immune Assay as late as in seven days. C. difficile toxin A assay yielded positive results in two (2/7) stool samples from the patients with antibiotic-associated diarrhea in the placebo group and a negative result in the only patient who developed antibiotic-associated diarrhea in the study group. CONCLUSIONS: The results implied that prophylactic use of Saccharomyces boulardii resulted in reduced, with no serious side effects, antibiotic-associated diarrhea in hospitalized patients.

Adult↗
PubMed1998

Review article: the use of biotherapeutic agents in the prevention and treatment of gastrointestinal disease.

There is presently a lack of well conducted clinical trials demonstrating any significant benefits of probiotics in humans. With the exception of diarrhoea due to rotavirus infection in children there is little evidence from randomized, double-blind, placebo-controlled studies that bacterial probiotics have a significant beneficial action in preventing diarrhoea of any cause. The yeast Saccharomyces boulardii has been shown to be of benefit in the prevention of antibiotic-associated diarrhoea but not in preventing infection with Clostridium difficile. S. boulardii may also be of benefit in preventing relapse of C. difficile infection. Because of the simplicity of in vitro systems and some animal models, beneficial characteristics of probiotics such as the ability of bacteria to bind to epithelial surfaces are not always transferable to humans. Thus any postulated benefit from consumption of probiotic bacteria should only be accepted as fact after testing in clinical studies. This review outlines our present knowledge of the mode of action of probiotics and presents the data from clinical trials on their use.

Anti-Bacterial Agents↗
PubMed2000

Intestinal colonization leading to fecal urobilinoid excretion may play a role in the pathogenesis of neonatal jaundice.

BACKGROUND: Neonatal hyperbilirubinemia remains of concern because of the potential danger for the central nervous system. Because urobilinogen is a nontoxic derivative of bilirubin, the current study was conducted to examine the fecal excretion of urobilinoids and bilirubin in healthy newborns and infants, as well as their intestinal bacteria capable of reducing bilirubin, to assess a possible relation to serum bilirubin levels during the first weeks of life. METHODS: Bilirubin pigments, urobilinoids, and porphyrins were measured in stools of infants during the first week (group A, n = 60) and between the second week and the first 6 months of life (group B, n = 64). Microbiologic analysis of stools was performed in selected cases and bilirubin-converting activity of isolated bacteria was determined in vitro. RESULTS: Urobilinoids were detectable in stools of 57% of the neonates at day 5, but not before. However, fecal urobilinoid production on that day was only a fraction of that observed in adults (0.07 vs. 0.7-3.6 mg/kg per day), whereas at week 6 it increased significantly to an average of 0.9 mg/kg per day. Microbiologic analysis of neonatal stools revealed two novel bacterial strains of Clostridium perfringens and Clostridium difficile capable of reducing bilirubin to urobilinoids. CONCLUSIONS: Urobilinoids can be detected in stools of 57% of newborns at day 5 after delivery. However, the urobilinoid production during the first week of life is quantitatively insufficient to contribute significantly to the removal of bilirubin. Enhancement of the microbial conversion of bilirubin could decrease the intestinal concentration of bilirubin and may decrease the degree or enhance the removal of neonatal hyperbilirubinemia.

Bacteria↗
PubMed2005

Alternative treatments for Clostridium difficile disease: what really works?

Vancomycin and metronidazole have been used for treating Clostridium difficile-associated disease (CDAD) for the past 25 years, but approximately 20 % of patients develop recurrent disease. The increasing incidence of nosocomial outbreaks, cases of recurrent CDAD and other complications (toxic megacolon, ileus, sepsis) has fuelled the search for different types of treatments. As the understanding of the pathogenesis of this disease has matured, newer treatment strategies that take advantage of these mechanisms have been developed. This review will describe such treatments and examine the evidence for each strategy.

Anti-Infective Agents↗
PubMed1994

Comparison of ribotyping, pulsed-field gel electrophoresis and random amplified polymorphic DNA for typing Clostridium difficile strains.

Clostridium difficile is a Gram-positive sporulating anaerobic bacillus which causes pseudomembranous colitis. Nosocomial acquisition of this bacteria has proved frequent, and epidemiological markers are needed to recognize and control common-source outbreaks. We therefore compared the results of pulsed-field gel electrophoresis (PFGE) after restriction with SmaI or NruI, random-amplified polymorphic DNA (RAPD) using 3 10-mer oligonucleotides, and ribotyping to differentiate between 30 unrelated strains of C. difficile belonging to 8 serotypes. The strains were separated into 26 different types by PFGE, 25 by RAPD, but into only 18 types by ribotyping. Median percentages of similarity between strains ranged from 27 in the PFGE assay to 90 in the ribotyping assay, but there was good agreement between the 3 methods for the clustering of strains. PFGE was more time-consuming than RAPD but its patterns were easier to analyze.

Bacterial Typing Techniques↗
PubMed1999

Clostridium difficile diarrhea and colitis: a clinical overview.

Infection with toxin-producing strains of Clostridium difficile is common and potentially life-threatening. It occurs mostly in patients in the hospital or nursing home who are taking or have recently taken antibiotics. Two toxins, A and B, damage the colonic mucosa, resulting in symptoms ranging from mild diarrhea to bloody diarrhea with fever and abdominal pain, colitis, or even pseudomembranous colitis. Severe cases may involve dehydration, toxic megacolon, or colonic perforation. This article reviews the microbiology, epidemiology, clinical manifestations, diagnosis, treatment, and prevention of this disease.

Adult↗
PubMed2002

[Clostridium difficile infection in a Department of Internal Medicine. A consecutive series of 45 patients].

OBJECTIVES AND METHODS: A retrospective study of 45 patients with Clostridium difficile infection over a 4-year period in a department of Internal Medicine. RESULTS: Mean age was 79 years; sex-ratio (F/M)=1.5; 38% of the patients had neurological or severe psychiatric disorders; 20% had a neoplastic disease. Ninety-three percent of cases had received one or more antibiotics before onset of diarrhea, prescribed mainly for a pulmonary infection. Amoxicillin clavulanic acid and cephalosporins were the most frequently used treatments, respectively in 48% and 40% of cases. For 25 patients (56%) Clostridium difficile-associated diarrhea was considered as a nosocomial infection, and as community-acquired diarrhea in 20 cases (44%). Treatment included isolation of the patient as soon as bacteriological diagnosis was known and specific therapy was instituted by metronidazole or vancomycin for a mean of 18 days. The addition of Saccharomyces boulardii was used in of cases. The clinical course was rapidly favorable for 80% of patients. Five patients died with complications of severe colitis in 2 cases. Mean hospital stay was 49 days (annual mean of the department=10 days). CONCLUSION: Clostridium difficile diarrhea concerns above all elderly patients with one or more underlying pathologies. Amoxicillin clavulanic acid and third-generation cephalosporins are the most frequently prescribed antibiotics in these cases and have the highest correlation with this infectious complication. This medical problem requires greater knowledge as it causes significant morbidity and increases the risk of prolonged hospital stays.

Aged↗
PubMed1991

Relapse versus reinfection with Clostridium difficile.

Relapse of Clostridium difficile-associated diarrhoea occurs in 15-20% of patients; however, whether relapse is due to an endogenous source of the organism or reinfection from the environment remains unclear. Restriction enzyme analysis (REA) of chromosomal DNA was used to type multiple isolates from ten patients who had experienced apparent relapses. More than half the relapses were due to infection with a new strain of C. difficile. The remaining patients were infected with the same strain, but whether this strain was acquired from the environment or from endogenous sources could not be determined. Relapses with a different strain of C. difficile could occur if an individual harboured more than one strain in their gastrointestinal tract. To investigate this possibility ten other patients were assessed for carriage of multiple strains. Ten colonies from a primary culture plate from each patient were typed by REA and tested for their ability to produce cytotoxin. All isolates from the same patient were identical by both methods, indicating that multiple carriage of strains may be a rare event.

Adult↗
PubMed2001

The effect of bacterial enterotoxins implicated in SIDS on the rabbit intestine.

The aim of this project was to characterise the type of damage caused to the intestine of the infant rabbit by bacterial enterotoxins implicated in sudden infant death syndrome (SIDS). Samples of the duodenum, jejunum, ileum, caecum and large intestine exposed to the toxins for up to 6 hours were examined by scanning (SEM) and transmission electron microscopy (TEM). The damage was quantitatively assessed (% villi damaged) by SEM and qualitatively by SEM and TEM. Clostridium perfringens enterotoxin, staphylococcal enterotoxin B and Clostridium difficile toxin A + toxin B combined all caused severe damage to the villi in the small intestine (80-90% damage). Clostridium difficile toxin B caused only slight damage (17% to the jejunum, 26% to the caecum). Clostridium perfringens alpha-toxin caused moderate damage to the small intestine (duodenum 34%, caecum 35%), and Escherichia coli STa caused significant damage to the small (53-70%) and large intestine (51%). The level of toxin damage increased with time, the small intestine being more susceptible generally to damage than the large intestine. Each toxin differed in its ability to damage the villi, microvilli, enterocytes and lamina propria.

Animals↗
PubMed1993

Enterotoxigenic bacteria in the sudden infant death syndrome.

Faecal samples from 123 infants who died with sudden infant death syndrome (SIDS) and from a comparative group of 52 age-matched babies were analysed for toxigenic bacteria and their toxins. Serum samples from the SIDS infants were also analysed for these toxins. A significantly higher proportion of toxigenic bacteria and their toxins were found in faecal samples of SIDS babies than in samples from the comparative group. These toxins were also found in serum from the SIDS babies. Clostridium perfringens was found in 54 (45.4%) of 119 SIDS cases compared with 10 (19.6%) of 51 healthy babies (chi 2 = 10.1, p < 0.01); C. difficile in 33 (27.7%) of 119 SIDS cases compared with 8 (14.8%) of 54 healthy babies (chi 2 = 3.43ns, p < 0.1); Staphylococcus aureus in 12 (27.3%; 66.7% enterotoxigenic) of 44 SIDS cases compared with 12 (85.7%; non-enterotoxigenic) of 14 healthy babies (chi 2 = 14.9, p < 0.001); C. botulinum in 6 (5.0%) of 120 SIDS cases compared with 0 of 53 healthy babies (chi 2 = 2.74, p < 0.1). Campylobacter jejuni, Yersinia enterocolitica, Vibrio parahaemolyticus, salmonellae and Bacillus cereus were not detected. Heat-labile toxin, lethal to mice (HLML) was found in 32 (27.1%) of 118 SIDS faecal samples compared with 5 (10.6%) of 47 healthy babies (chi 2 = 5.24, p < 0.05); cytotoxins in 38 (30.9%) of 123 SIDS faecal samples compared with 0 of 21 of healthy babies (chi 2 = 8.8, p < 0.01) and 24 (27.6%) of 87 SIDS serum samples. C. perfringens enterotoxin was detected in 33 (34.4%) of 96 SIDS faecal extracts compared with 0 of 23 of healthy babies (chi 2 = 10.94, p < 0.001), and in 27 (24.5%) of 110 SIDS serum samples. C. perfringens alpha-toxin (presumptive) was detected in 14 (17.5%) of 80 SIDS faecal extracts compared with 0 of 17 from healthy babies (chi 2 = 3.5ns, p congruent to 0.05) and in 2 (2.3%) of 87 SIDS serum samples. C. difficile toxin was detected in four SIDS faecal samples and two serum samples. C. botulinum toxin was detected in only one of 120 SIDS faecal samples compared with none of 49 from healthy babies. Staphylococcal enterotoxins were detected in 8 (19.5%) of 41 SIDS faecal samples compared with 0 of 19 from healthy babies (chi 2 = 4.278, p < 0.05), and in 4 (10.8%) of 37 SIDS serum samples.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed2006

In vitro activities of daptomycin, vancomycin, and penicillin against Clostridium difficile, C. perfringens, Finegoldia magna, and Propionibacterium acnes.

Daptomycin has in vitro activity against gram-positive anaerobic bacteria, although limited numbers of species have been tested. We studied the in vitro activities of daptomycin, vancomycin, and penicillin against more than 100 strains each of Clostridium difficile, C. perfringens, Finegoldia magna, and Propionibacterium acnes. Daptomycin Etest MICs and results from time-kill studies were determined for selected strains. For 392 of 421 strains (93%), daptomycin was inhibitory at < or =1 microg/ml, including 15 of 16 strains of C. difficile with elevated linezolid MICs of 8 and 16 microg/ml, all 32 strains with moxifloxacin MICs of > or =4 microg/ml, and all 16 strains resistant to clindamycin. Daptomycin MICs were also < or =1 microg/ml for all 16 F. magna strains resistant to clindamycin and all 32 strains resistant to tetracycline. Only one strain, a C. perfringens strain, had a MIC of >2 microg/ml to daptomycin. Eighty-five and 92.5% of the Etest MICs were within 1 dilution of the agar dilution method for all drugs at 24 and 48 h, respectively. In time-kill studies, a C. difficile strain was inhibited by both daptomycin and vancomycin at 1, 2, 4, 8, and 24 h; colony counts were decreased by 2.3 to 2.9 log at 24 h. Vancomycin was not bactericidal for C. perfringens; however, daptomycin showed bactericidal activity as early as 1 h at four and eight times the MIC and at 2 and 4 h at two and four times the MIC.

Anti-Bacterial Agents↗
PubMed1991

Molecular screening of Clostridium difficile toxins A and B genetic determinants and identification of mutant strains.

Three separate sets of polymerase chain reaction primers were designed to specifically detect the presence of a toxin A gene fragment, a toxin B gene fragment, and the entire toxin B gene. In addition toxin gene fragments that were amplified from well characterized toxic strains were tagged fluorescently and used as hybridization probes to screen C. difficile strains. A survey of 37 toxic strains and 10 non-toxic strains demonstrated that toxic strains normally contain the genetic composition for toxin A and toxin B simultaneously; whereas, non-toxic strains typically did not contain detectable toxin determinants. The only exception found was strain 39, which had the genetic composition for toxins A and B, but was not cytotoxic under the conditions tested.

Animals↗
PubMed2001

The role of antibiotics in the treatment of infectious diarrhea.

Infectious diarrhea is a significant cause of morbidity and mortality and a common complaint in clinical practice. Routine empirical use of antibiotics for infectious diarrhea should be avoided because of the self-limited nature of most cases, the cost of antibiotics, and the potential to worsen the already significant problem of antibiotic resistance of enteric pathogens. For patients with severe invasive or prolonged diarrhea or who are at high risk of complications, such as the elderly, diabetics, cirrhotics, and immunocompromised patients, empirical treatment with a quinolone antibiotic for 3 to 5 days can be considered. Antibiotic treatment can be highly effective for Shigella, ETEC, and V. cholerae infections, and metronidazole is indicated for C. difficile colitis. The impact of antibiotics for other specific pathogens is modest, and antibiotic therapy should be reserved for the same group of patients who would be considered for empirical treatment. The most significant problem in the antibiotic treatment of infectious diarrhea is the progressive increase in resistance among enteric pathogens; only the prudent use of antimicrobials in all areas of daily practice can limit or delay the impact of this serious problem.

Aged↗
PubMed1999

Comparative value of colonic biopsy and intraluminal fluid culture for diagnosis of bacterial acute colitis in immunocompetent patients. Infectious Colitis Study Group.

We compared the yield of intraluminal fluid culture to that of biopsy specimens obtained during colonoscopy for the diagnosis of bacterial colitis in 93 immunocompetent patients with a recent episode of diarrhea and macroscopic lesions of colitis. Stool culture findings were also available for 68 patients. At least one bacterial pathogen was isolated from the biopsy specimen, intraluminal fluid, or stool from 48 patients (51.6%). Salmonella species, Clostridium difficile, Klebsiella oxytoca, Shigella species, and Campylobacter species were recovered from 16 (17.2%), 15 (16.1%), 8 (8.6%), 7 (7.5%), and 4 (4.3%) of the patients, respectively. One Shigella species and one K. oxytoca strain were isolated from biopsy specimens but not from intraluminal fluid, and intraluminal fluid was the only positive specimen in 12 cases (yielding 1 Salmonella species, 2 Shigella species, 2 K. oxytoca, and 7 C. difficile isolates). In nine cases out of 10, toxin B was detected only in intraluminal fluid. A correlation of 91.2% was observed between stool and intraluminal fluid cultures for Salmonella, Shigella, and Campylobacter species isolations. Culture of biopsy specimens adds little to the diagnosis of infectious colitis, and stools and intraluminal fluids appear to have comparable value.

Acute Disease↗
PubMed1996

Inhibition of adhesion of Clostridium difficile to Caco-2 cells.

For many microorganisms, including Clostridium difficile, mucosal association is an important factor influencing intestinal colonisation and subsequent infection. Inhibition of adhesion of C. difficile to intestinal mucosa could be a new promising strategy for prevention and treatment of antibiotic-associated diarrhoea. We investigated the possibilities of influencing the adhesion of C. difficile by xylitol and bovine colostrum whey. Caco-2 cells and C. difficile cells were incubated with 1%, 5% and 10% solutions of xylitol and colostrum. Our study revealed that both xylitol and colostrum inhibited the adhesion of C. difficile to Caco-2 cells. Inhibition by xylitol was dose-dependent. When compared to the control, the count of adherent C. difficile decreased 3.4 times when treated with 1% xylitol, 12 times when 5% xylitol was applied, and 18.7 times when treated with 10% xylitol. The inhibition of adherence by colostrum was partially dose-dependent: 3.1 times in the case of 1%, and 5.5 times in the cases of 5% and 10% colostrum. Further experimental and clinical studies are needed for the application of xylitol and colostrum in the treatment and prophylaxis of pseudomembraneous colitis.

Animals↗