Molecular epidemiology of endemic Clostridium difficile infection

This is the first study to provide a comprehensive insight into the molecular epidemiology of endemic Clostridium difficile and particularly that associated with a recently recognized epidemic strain. We DNA fingerprinted all C. difficile isolates from the stools of patients with symptomatic antibiotic-associated diarrhoea and from repeated samples of the inanimate ward environment on two elderly medicine hospital wards over a 22-month period. Notably, C. difficile was not recoverable from either ward immediately before opening, but was found on both wards within 1-3 weeks of opening, and the level of environmental contamination rose markedly during the first 6 months of the study period. C. difficile infection (CDI) incidence data correlated significantly with the prevalence of environmental C. difficile on ward B (r = 0.76, P < 0.05) but not on ward A (r = 0.26, P > 0.05). We found that RAPD and RS-PCR typing had similar discriminatory power, although, despite fingerprinting over 200 C. difficile isolates, we identified only six distinct types. Only two distinct C. difficile strains were identified as causing both patient infection and ward contamination. Attempts to determine whether infected patients or contaminated environments are the prime source for cross-infection by C. difficile had limited success, as over 90% of C. difficile isolates were the UK epidemic clone. However, a non-epidemic strain caused a cluster of six cases of CDI, but was only isolated from the environment after the sixth patient became symptomatic. The initial absence of this strain from the environment implies patient-to-patient and/or staff-to-patient spread. In general, routine cleaning with detergent was unsuccessful at removing C. difficile from the environment. Understanding the epidemiology and virulence of prevalent strains is important if CDI is to be successfully controlled.     

Acquisition of Clostridium difficile from the hospital environment

An outbreak of antibiotic-associated colitis that occurred on a ward of a Michigan hospital during February-April, 1984, was studied by bacteriophage-bacteriocin typing. Stools from the seven involved patients yielded Clostridium difficile isolates of types B1537 or Cld7;B1537. C. difficile was recovered from 31.4% of environmental cultures obtained on the ward, and the majority of isolates were types B1537 or Cld7;B1537. When the ward was disinfected with unbuffered hypochlorite (500 parts per million (ppm) available chlorine), surface contamination decreased to 21% of initial levels and the outbreak subsequently ended. Phosphate buffered hypochlorite (1,600 ppm available chlorine, pH 7.6) was even more effective; its use resulted in a 98% reduction in surface contamination. These findings suggest that environmental contamination with C. difficile is important in the epidemiology of antibiotic-associated colitis, and that hypochlorite is effective in eliminating C. difficile from the hospital environment.     

Prospective evaluation of environmental contamination by Clostridium difficile in isolation side rooms.

We determined prospectively the frequency, persistence and molecular epidemiology of Clostridium difficile environmental contamination after detergent-based cleaning in side rooms used to isolate patients with C. difficile diarrhoea. Approximately one-quarter of all environmental sites in side rooms sampled over four-week periods were contaminated with C. difficile. The overall side room prevalence of environmental C. difficile declined from 35% initially, to 24% in week 2, 18% in week 3, and 16% in week 4. The bed frame was the most common site from which C. difficile was recovered, although the floor was the most contaminated site in terms of total numbers of colonies. C. difficile was recovered significantly more frequently from swabs plated directly on to C. difficile selective media containing lysozyme than from enrichment broth (P< 0.001), emphasizing the benefit of lysozyme supplementation. The great majority of C. difficile isolates (87% of all isolates, 84% of patient isolates) was indistinguishable from the UK epidemic strain (PCR ribotype 1). It thus could not be determined whether environmental contamination was a cause or a consequence of diarrhoea. Our findings highlight the need for improved approaches to hospital environmental hygiene, and call into question current UK guidelines that recommend detergent-based cleaning to remove environmental C. difficile. In particular, improved cleaning of frequently touched sites in the immediate bed space area is required.     

Environmental survey to assess viral contamination of air and surfaces in hospital settings

This report describes an outbreak of Clostridium difficile infection (CDI) in a vascular surgery ward in 2009 caused by a high-level clindamycin-resistant ribotype 106. A case of CDI was defined as a patient with diarrhoea, positive for C. difficile toxin and negative for other enteric pathogens. Cultures were sent to the Scottish Salmonella Shigella and Clostridium difficile Reference Laboratory (SSSCDRL) for PCR ribotyping, antibiotic susceptibility testing and PCR detection of ermB. The mean age of the nine patients was 73 years (range: 38–90 years). All had received clindamycin and ciprofloxacin. All cases were typed as PCR ribotype 106 and they showed high-level resistance to clindamycin. Five of these isolates were tested by PCR for the presence of the ermB gene and no amplification was detected. This strain has rarely been isolated from patients on this ward. The outbreak was controlled successfully by closure of the ward with terminal cleaning, reinforcement of infection control precautions and the introduction of a new antibiotic policy. It is notable that this outbreak was caused by a strain with high-level clindamycin resistance not mediated by ermB. It also re-emphasizes that outbreaks of CDI can be caused by C. difficile PCR ribotypes other than 027. The outbreak was most likely associated with the use of clindamycin and ciprofloxacin cross-infection with spores in this environment. Implementation of strict infection control precautions, antimicrobial stewardship and enhanced environmental cleaning are key components in managing such an outbreak successfully. The number of meticillin-resistant Staphylococcus aureus acquisitions also fell substantially after these interventions.     

A targeted strategy to wipe out Clostridium difficile

This study evaluated daily cleaning with germicidal bleach wipes on wards with a high incidence of hospital-acquired Clostridium difficile infection (CDI). The intervention reduced hospital-acquired CDI incidence by 85%, from 24.2 to 3.6 cases per 10,000 patient-days, and prolonged the median time between hospital-acquired CDI cases from 8 to 80 days.     

An evaluation of hospital cleaning regimes and standards

A four-part study assessing cleanliness in up to 113 environmental surfaces in an operating theatre and a hospital ward is reported. Surfaces were assessed visually, using microbiological methods and ATP bioluminescence. Results from a preliminary random survey indicated variability in cleanliness. These results were then used to select sites for monitoring before and after routine cleaning, over a 14-day period. Using published microbiological and ATP specifications 70 and 76% of these sites were unacceptable after cleaning. Visual assessment was a poor indicator of cleaning efficacy with only 18% considered unacceptable. Sites most likely to fail in the ward were in the toilet and kitchen, areas which are frequently implicated in the spread of infectious intestinal disease. Operating theatre sites had lower ATP results but 61% of sites would be considered unacceptable. There was no significant difference in general microbiological or ATP results overall before and after routine cleaning. Although some important hand contact sites showed no significant difference, overall there was a significant decrease in staphylococcal and enterobacteria counts in the ward but not in the operating theatre after cleaning. The routine cleaning programmes used did not include a biocide and cleaning using a hypochlorite based sanitizer gave much lower values. The results are discussed in relation to infection control, cleaning audits and cleaning schedules: an integrated cleaning monitoring programme using ATP bioluminescence in conjunction with visual and microbiological assessments is recommended.     

Clostridium difficile colitis: a review

Clostridium difficile has become an increasingly important nosocomial pathogen and is one of the most common causes of hospital-acquired diarrhea. The incidence of C difficile infection (CDI) is increasing worldwide. Overuse of antibiotics is felt to be a major contributing factor leading to the increased incidence of CDI. The clinical manifestations of CDI vary from a mild form of the disease to fulminant diarrhea, leading to significant patient morbidity and mortality. The increasing incidence of CDI has a major impact on increasing health care costs. This article will summarize the epidemiology, pathogenesis, clinical manifestations, laboratory diagnosis, and treatment options for CDI, as well as infection-control measures for the prevention of CDI.     

A clinical risk index for Clostridium difficile infection in hospitalised patients receiving broad-spectrum antibiotics

Identification of a population at high risk for Clostridium difficile infection (CDI) would enable CDI prevention strategies to be designed. The purpose of this study was to create a clinical risk index that would predict those at risk for CDI. A CDI risk index was therefore developed, based on a cohort of hospital patients given broad-spectrum antibiotics, and divided into a development and validation cohort. Logistic regression equations helped identify significant predictors of CDI. A scoring algorithm for CDI risk was created using identified risk factors and collapsed to create four categories of CDI risk. The area under the receiver operating characteristic (aROC) curve was used to measure goodness-of-fit. Among 54 226 patients, 392 tested positive for C. difficile. Age 50-80 years [odds ratio (OR: 0.5; P<0.0116)], age >80 years (OR: 2.5; P<0.0001), haemodialysis (OR: 1.5; P=0.0227), non-surgical admission (OR: 2.2; P<0.0001) and increasing length of stay in the intensive care unit (OR: 2.1; P<0.0001) were significantly associated with CDI. A simple risk index using presence of significant variables was significantly associated with increasing risk for CDI in both development (OR: 3.57; P<0.001; aROC: 0.733) and validation (OR: 3.31; P<0.001; aROC: 0.712) cohorts. An OR-derived risk index did not perform as well as the simple risk index. This easily implemented risk index should allow stratification of patients into risk group categories for development of CDI and help fashion preventive strategies.     

Value of lysozyme agar incorporation and alkaline thioglycollate exposure for the environmental recovery of Clostridium difficile

Clostridium difficile is an increasingly prevalent nosocomial pathogen. Environmental contamination by spores is believed to be a major factor propagating the spread of C. difficile. Various approaches including the use of bile salts have been described to enhance the recovery of C. difficile from clinical and environmental specimens. We found that lysozyme (5 mg/L) incorporated into a selective medium containing bile salts significantly increased the recovery of C. difficile from swabs of 197 environmental sites (11% versus 24% samples positive, P< 0.01). Furthermore, in a separate series of experiments additional use of cooked meat broth enrichment significantly enhanced the recovery of C. difficile (35% versus 45%, P = 0.009). Conversely, we found that pre-exposure to alkaline thioglycollate did not improve the yield of C. difficile. Lysozyme incorporation markedly increases the recovery of C. difficile from environmental samples probably by stimulation of spore germination. Our findings suggest that previous attempts to determine the level of environmental C. difficile contamination have markedly underestimated the true prevalence of this pathogen.     

Efficacy of hospital cleaning agents and germicides against epidemic Clostridium difficile strains.

OBJECTIVE: To compare the effects of hospital cleaning agents and germicides on the survival of epidemic Clostridium difficile strains. METHODS: We compared the activity of and effects of exposure to 5 cleaning agents and/or germicides (3 containing chlorine, 1 containing only detergent, and 1 containing hydrogen peroxide) on vegetative and spore forms of epidemic and non-epidemic C. difficile strains (3 of each). We carried out in vitro exposure experiments using a human fecal emulsion to mimic conditions found in situ. RESULTS: Cleaning agent and germicide exposure experiments yielded very different results for C. difficile vegetative cells, compared with those for spores. Working-strength concentrations of all of the agents inhibited the growth of C. difficile in culture. However, when used at recommended working concentrations, only chlorine-based germicides were able to inactivate C. difficile spores. C. difficile epidemic strains had a greater sporulation rate than nonepidemic strains. The mean sporulation rate, expressed as the proportion of a cell population that is in spore form, was 13% for all strains not exposed to any cleaning agent or germicide, and it was significantly increased by exposure to cleaning agents or germicides containing detergent alone (34%), a combination of detergent and hypochlorite (24%), or hydrogen peroxide (33%). By contrast, the mean sporulation rate did not change substantially after exposure to germicides containing either a combination of detergent and dichloroisocyanurate (9%) or dichloroisocyanurate alone (15%). CONCLUSIONS: These results highlight differences in the activity of cleaning agents and germicides against C. difficile spores and the potential for some of these products to promote sporulation.     

Activity of a dry mist hydrogen peroxide system against environmental Clostridium difficile contamination in elderly care wards

Clostridium difficile causes serious healthcare-associated infections. Infection control is difficult, due in part to environmental contamination with C. difficile spores. These spores are relatively resistant to cleaning and disinfection. The activity of a dry mist hydrogen peroxide decontamination system (Sterinis((R))) against environmental C. difficile contamination was assessed in three elderly care wards. Initial sampling for C. difficile was performed in 16 rooms across a variety of wards and specialties, using Brazier's CCEY (cycloserine-cefoxitin-egg yolk) agar. Ten rooms for elderly patients (eight isolation and two sluice rooms) were then resampled following dry mist hydrogen peroxide decontamination. Representative isolates of C. difficile were typed by polymerase chain reaction ribotyping. C. difficile was recovered from 3%, 11% and 26% of samples from low, medium and high risk rooms, respectively. In 10 high risk elderly care rooms, 24% (48/203) of samples were positive for C. difficile, with a mean of 6.8 colony-forming units (cfu) per 10 samples prior to hydrogen peroxide decontamination. Ribotyping identified the presence of the three main UK epidemic strains (ribotypes 001, 027 and 106) and four rooms contained mixed strains. After a single cycle of hydrogen peroxide decontamination, only 3% (7/203) of samples were positive (P<0.001), with a mean of 0.4 cfu per 10 samples ( approximately 94% reduction). The Sterinis((R)) hydrogen peroxide system significantly reduced the extent of environmental contamination with C. difficile in these elderly care rooms. This relatively quick and user-friendly technology might be a more reliable method of terminally disinfecting isolation rooms, following detergent cleaning, compared to the manual application of other disinfectants.     

Gaseous and air decontamination technologies for Clostridium difficile in the healthcare environment

The recent data for hospital-acquired infections suggest that infection rates for meticillin-resistant Staphylococcus aureus (MRSA) and Clostridium difficile are beginning to decrease. However, while there is still pressure to maintain this trend, the resistance of C. difficile spores to standard detergents continues to present a problem for many UK hospitals trying to prevent its spread or control outbreaks. Alternative disinfection technologies such as gaseous decontamination are currently being marketed to the healthcare sector as an alternative/supplement to manual disinfection, and have been shown to be effective in reducing environmental contamination. When used correctly, they offer a complementary technology to manual cleaning that increases the probability of an effective reduction in viability and provides a comparatively uniform distribution of disinfectant. Three gaseous decontamination technologies are examined for their suitability in reducing environmental contamination with C. difficile: gaseous hydrogen peroxide, chlorine dioxide and ozone. Air decontamination and UV-based technologies are also briefly described. We conclude that while there is a role to play for these new technologies in the decontamination of ward surfaces contaminated with C. difficile, the requirement for both a preclean before use and the limited 'in vivo' evidence means that extensive field trials are necessary to determine their cost-effectiveness in a healthcare setting.     

Emergence and control of fluoroquinolone-resistant, toxin A-negative, toxin B-positive Clostridium difficile.

BACKGROUND: Clostridium difficile is a major cause of infectious diarrhea in hospitalized patients. Between August 2003 and January 2004, we experienced an increase in the incidence of C. difficile-associated disease. We describe the investigation into and management of the outbreak in this article. METHODS: A total of 73 consecutive patients with nosocomial C. difficile-associated diarrhea were identified. C. difficile isolates were characterized using toxin-specific enzyme immunoassays, a tissue-culture fibroblast cytotoxicity assay, polymerase chain reaction (PCR), and antimicrobial susceptibility tests. Rates of recurrence and of C. difficile colitis were recorded. Changes in antibiotic use and infection control policies were documented. RESULTS: The incidence of C. difficile-associated diarrhea peaked at 21 cases per 1,000 patient admissions. Of the C. difficile isolates recovered, 85 (95%) were identical toxin A-negative and toxin B-positive strains, corresponding to toxinotype VIII and PCR ribotype 017. All clonal isolates were resistant to multiple antibiotics, including ofloxacin, ciprofloxacin, levofloxacin, moxifloxacin, and gatifloxacin (minimum inhibitory concentrations [MICs] of greater than 32 micro g/mL) and erythromycin, clarithromycin, and clindamycin (MICs of greater than 256 micro g/mL). Recurrent C. difficile-associated disease occurred in 26 (36%) of the patients. At least 10 (14%) of the patients developed C. difficile colitis. Additional infection control measures introduced included the use of ward memos, a hand-hygiene awareness campaign, increased environmental cleaning, attention to prescribing practices for antibiotics, increased awareness of diarrheal illness, and early isolation of affected patients. Total use of fluoroquinolones did not change throughout the study period. Despite persistence of this toxin-variant strain, the incidence of C. difficile-associated disease in our institution decreased to fewer than 5 cases per 1,000 admissions. CONCLUSIONS: We report on the emergence of a fluoroquinolone- and clindamycin-resistant, toxin A-negative, and toxin B-positive strain of C. difficile associated with an outbreak of C. difficile-associated disease in our institution during a 6-month period. We found that careful attention to improvement of infection control interventions was the most important means of controlling this nosocomial pathogen.     

Lack of enhanced effect of a chlorine dioxide-based cleaning regimen on environmental contamination with Clostridium difficile spores

Spores of Clostridium difficile may play a significant role in transmission of disease within the healthcare environment and are resistant to a variety of detergents and cleaning fluids. A range of environmental cleaning agents has recently become available, many of which claim to be sporicidal. We investigated the effect of changing to a chlorine dioxide-based cleaning regimen on C.?difficile environmental contamination and patient infection rates. The prevalence of environmental contamination was unaffected with a rate of 8% (9/120) before and 8% (17/212) following the change. Rates of patient infection were also unchanged during these periods.     

Effectiveness and cost-effectiveness of a Clostridium difficile vaccine candidate in a hospital setting

Toxoid vaccines against Clostridium difficile infections (CDI) appear promising in reducing the risk of developing toxin-mediated symptoms. We sought to evaluate the effectiveness and cost-effectiveness of a vaccine candidate in a hospital setting. We developed an agent-based simulation model of nosocomial CDI in a 300-bed hospital. Targeting high-risk patients for vaccination, we estimated the reduction of symptomatic CDI. Using the net reduction of CDI-associated isolation days, we evaluated the vaccine’s cost-effectiveness from a healthcare provider perspective over a 2-year period with an average monthly incidence of 5 cases per 10,000 patient-days pre-vaccination.Assuming a vaccine efficacy in the range 60–90%, vaccinating 40% of high-risk patients pre-admission reduced symptomatic CDI by 16.6% (95% CI: 15.2, 17.9). When the vaccine coverage increased to 80%, the reduction of symptomatic CDI was 34.6% (95% CI: 33.7, 35.9). For a willingness to pay (WTP) of CDN$1000 (corresponding to the average costs of case isolation per day), vaccine was cost-effective for vaccination costs per individual (VCPI) up to CDN$111 in the scenario of 40% vaccine coverage. With the same WTP, vaccine was cost-effective for VCPI up to CDN$121 when the vaccine coverage increased to 80%. A significant portion (~80%) of hospital colonization is caused by environmental transmission of C. difficile, which markedly reduced the effectiveness of vaccine below its assumed efficacy. However, due to the number of CDI-associated isolation days averted, vaccination of high-risk patients can be cost-effective depending on the WTP and the VCPI.     

Airborne hydrogen peroxide for disinfection of the hospital environment and infection control: a systematic review

We reviewed the effectiveness of airborne hydrogen peroxide as an environmental disinfectant and infection control measure in clinical settings. Systematic review identified ten studies as eligible for inclusion. Hydrogen peroxide was delivered in the form of vapour and dry mist in seven and three studies, respectively. Pathogens evaluated included meticillin-resistant Staphylococcus aureus (MRSA), Clostridium difficile and multiple bacterial types, in five, three, and two studies, respectively. Before the application of any cleaning intervention, 187/480 (39.0%; range: 18.9-81.0%) of all sampled environmental sites were found to be contaminated by the studied pathogens in nine studies that reported specific relevant data. After application of terminal cleaning and airborne hydrogen peroxide, 178/630 (28.3%; range: 11.9-66.1%) of the sampled sites in six studies and 15/682 (2.2%; range: 0-4.0%) of the sampled sites in ten studies, respectively, remained contaminated. Four studies evaluated the use of hydrogen peroxide vapour for infection control. This was associated with control of a nosocomial outbreak in two studies, eradication of persistent environmental contamination with MRSA and decrease in C. difficile infection in each of the remaining two studies.     

Gastrointestinal carriage of Clostridium difficile in cats and dogs attending veterinary clinics.

Cats and dogs being treated at two veterinary clinics were investigated for gastrointestinal carriage of Clostridium difficile using selective solid and enrichment media. Thirty-two (39.5%) of 81 stool samples yielded C. difficile. There were significant differences in isolation rates between clinics, 61.0% of animals being positive at one clinic compared to 17.5% at the other (Chi-square, P less than 0.005). Of 29 animals receiving antibiotics, 15 (52.0%) harboured C. difficile while 11 (23.9%) of 46 animals not receiving antibiotics were positive (Chi-square, P less than 0.01). There was no difference in carriage rate between cats (38.1%) and dogs (40.0%). The environment at both veterinary clinics was surveyed for the presence of C. difficile. Fifteen of 20 sites at one clinic were positive compared to 6 of 14 sites at the other clinic. Both cytotoxigenic and noncytotoxigenic isolates of C. difficile were recovered from animals and environmental sites. These findings suggest that household pets may be a potentially significant reservoir of infection with C. difficile.     

Eclipse系统中通量平滑对单发脑转移瘤调强放疗的剂量学影响研究

目的:探讨Eclipse计划系统中不同的通量平滑值在单发脑转移瘤调强放疗(intensity modulated radiotherapy,IMRT)中的剂量学差异。方法:随机选取20例单发脑转移瘤患者的临床资料,基于Eclipse v13.6计划系统和Varian Trilogy加速器为每个患者制订6组逆向IMRT计划。使用默认的X、Y方向通量平滑值X=40、Y=30作为对照组,其他5组在相同的优化条件下,改变X、Y方向的通量平滑值分别为X=0、Y=0,X=20、Y=15,X=80、Y=60,X=100、Y=80,X=200、Y=150重新进行计划设计和评估。比较各计划靶区的D2、D98、Dmean和适形性指数(conformity index,CI)、均匀性指数(homogeneity index,HI),危及器官(organs at risk,OAR)的剂量学参数脑干Dmax、视神经Dmax、视交叉Dmax、晶体Dmax、眼球Dmax及各计划的机器跳数和Gamma通过率。采用SPSS 22.0进行统计学分析。结果:与对照组相比,X=0、Y=0和X=20、Y=15计划组的D2有统计学差异(P<0.05),所有组的D98均有统计学差异(P<0.05),X=0、Y=0和X=200、Y=150计划组的Dmean有统计学差异(P<0.05),所有计划组的CI均有统计学差异(P<0.05),X=100、Y=80和X=200、Y=150计划组的HI有统计学差异(P<0.05)。与对照组相比,所有计划组脑干Dmax、视神经Dmax、眼球Dmax均无统计学差异(P>0.05);对照组的晶体Dmax低于X=80、Y=60,X=100、Y=80,X=200、Y=150计划组(P<0.05);对照组的视交叉Dmax低于X=100、Y=80和X=200、Y=150计划组(P<0.05)。随着通量平滑值的增加,机器跳数减少,Gamma通过率增加。结论:综合考虑单发脑转移瘤IMRT的临床需求和计划执行效率,X、Y方向通量平滑值建议采用默认的X=40、Y=30为佳。     

Laboratory diagnosis of Clostridium difficile infection in Spain: a population-based survey

A survey of laboratory diagnosis of Clostridium difficile infection (CDI) was conducted in 103 Spanish hospitals. A mean of 23.2 stool specimens/1000 admissions were processed to detect CDI. Overall, 35.9% of the laboratories specifically selected stool specimens for diagnostic C.?difficile toxin testing. The most commonly used criteria were loose or watery stools, previous antibiotic therapy and nosocomial diarrhoea. Most laboratories (95.1%) processed the stool specimens in house, mainly five to seven days/week. All laboratories except one detected toxins directly in stool specimens, and 94.9% used enzyme immunoassays (EIAs). Only 16.3% of the laboratories used toxigenic culture as a diagnostic tool, and most of them used EIAs to detect toxins in isolates. The most common diagnostic strategy was toxin detection in stool specimens using EIA alone (79.6%), and the second most common strategy was the combination of toxin detection in stool specimens and isolates (10.2%). The estimated annual incidence of CDI was 1.71 cases/1000 admissions, and this was significantly higher in large hospitals. CDI is underdiagnosed in Spain because most laboratories use EIAs performed directly on stool specimens as the only diagnostic procedure. A national laboratory survey of diagnostic methods and testing protocols for the diagnosis of CDI is simple and inexpensive, and could act as the first step towards implementing national standardized criteria for optimal diagnosis of CDI.     

The incidence of nosocomial toxigenic clostridium difficile associated diarrhea in Tehran tertiary medical centers

Clostridium difficile is the most common cause of nosocomial diarrhea. It is usually a consequence of antibiotic treatment, But sporadic cases can occur. This study was aimed to determine the frequency of the nosocomial Clostridium difficile (C. difficile) associated diarrhea in Tehran University of Medical Sciences hospitals and study of antibacterial susceptibility of isolates. In this study a total of 942 stool samples from patients with nosocomial diarrhea that were hospitalized in Imam Khomeini hospital, Shariati hospital and Children clinical center were collected. The samples were cultured on a selective cycloserine cefoxitin fructose agar (CCFA) and incubated in anaerobic conditions, at 37°C for 5 days. Isolates were characterized to species level by conventional biochemical tests. Bacterial cytotoxicity was assayed on tissue culture (vero). Antimicrobial sensitivity of isolated toxigenic C. difficile were investigated by kirby Beuer method (disk diffusion). Our findings show that, of the total patients, 57 toxigenic C. difficile (6.1%) were isolated. Results of statistical analysis show significant differences between the rate of isolated toxigenic C. difficile and age group of patients (P<0.05). Among the wards of selected hospitals, in gastroenterology of Children clinical center, Toxigenic C. difficile was isolated from patients most frequently. The sensitivity of isolates to vancomycin, Chloramphenicol and ceftriaxone were higher than other antibiotics. Toxigenic C. difficile is a common hospital-acquired infection. The organism was found in 6.1% hospitalized patients. Further studies to evaluate the rate and role of toxigenic C. difficile in nosocomial diarrheal processes, ecological and pathogenic terms are suggested.