Molecular typing of Clostridium difficile isolates cultured from patient stool samples and gastroenterological medical devices in a single Iranian hospital (original) (raw)
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African Journal of Microbiology Research, 2013
Detection of the source of Clostridium difficile for the control of nosocomial infections produced by these bacteria is very important. For this reason 84 C. difficile isolated from 250 stool samples of symptomatic patients, staff, asymptomatic patients at first day of admission, the same patients after seven days of hospitalization and 135 samples from their hospital environment were typed by AP-PCR. In addition to conventional standard tests, gas liquid chromatography was also used as complementary test to identify C. difficile isolates. All isolates were separated into 12 genotypes, with 31% falling into group I. Genotypes VIII, X and XII were found only in isolates of symptomatic patients while genotype I was observed in all C. difficile of patients and environment. Genotypes III was not detected in any C. difficile isolates except in one of the acquired isolates. C. difficile is frequently transmitted among hospitalized patients, staff, and their hospital environment.
Journal of Medical Microbiology, 2015
Clostridium difficile infection (CDI) leads to considerable morbidity and mortality among hospitalized patients. Faecal specimens from 1110 hospitalized patients suspected for CDI were cultured for isolation of C. difficile and characterization of virulence genes. PCR was carried out for toxigenic genes tcdA, tcdB, cdtA and cdtB and PCR-RFLP for fliC and slpA genes. Of 174 (15.7 %) C. difficile isolates, 121 (69.5 %) were toxigenic, amongst which 68 (56.2 %) also had both tcdA and tcdB genes. The remaining 53 (43.8 %) of the isolates also had at least one of the toxin genes. Binary toxin genes (cdtA and cdtB) with only one of the two components were present in 16 (9.2 %) of the 174 isolates. The other virulence genes-fliC and slpA-were present in 100 % of the isolates. The most frequent PCR-RFLP type of fliC gene was type I (n5101), followed by type VII (n549) and type III (n524). The slpA gene presented with three combinations of patterns. Characterization of virulence genes in C. difficile isolates is of extreme importance for epidemiological surveillance and control of outbreaks owing to the capacity of this bacterium to adapt to new environmental circumstances, leading to the emergence of new epidemic strains.
Journal of clinical microbiology, 2000
Toxigenic Clostridium difficile is the most common etiologic agent of hospital-acquired diarrhea in developed countries. The role of this pathogen in nosocomial diarrhea in Eastern Europe has not been clearly established. The goal of this study was to determine the prevalence of C. difficile in patients and the hospital environment in Belarus and to characterize these isolates as to the presence of toxin genes and their molecular type. C. difficile was isolated from 9 of 509 (1.8%) patients analyzed and recovered from 28 of 1,300 (2. 1%) environmental sites cultured. A multiplex PCR assay was used to analyze the pathogenicity locus (PaLoc) of all isolates, and strain identity was determined by an arbitrarily primed PCR (AP-PCR). The targeted sequences for all the genes in the PaLoc were amplified in all C. difficile strains examined. A predominantly homogeneous group of strains was found among these isolates, with five major AP-PCR groups being identified. Eighty-three percent of en...
Journal of Clinical Microbiology, 2009
Our laboratory has developed testing methods that use real-time PCR and pyrosequencing analysis to enable the rapid identification of potential hypervirulent Clostridium difficile strains. We describe a real-time PCR assay that detects four C. difficile genes encoding toxins A (tcdA) and B (tcdB) and the binary toxin genes (cdtA and cdtB), as well as a pyrosequencing assay that detects common deletions in the tcdC gene in less than 4 h. A subset of historical and recent C. difficile isolates (n ؍ 31) was also analyzed by pulsed-field gel electrophoresis to determine the circulating North American pulsed-field (NAP) types that have been isolated in New York State. Thirteen different NAP types were found among the 31 isolates tested, 13 of which were NAP type 1 strains. To further assess the best approach to utilizing our conventional and molecular methods, we studied the populations of C. difficile in patient stool specimens (n ؍ 23). Our results indicated that 13% of individual stool specimens had heterogeneous populations of C. difficile when we compared the molecular characterization results for multiple bacterial isolates (n ؍ 10). Direct molecular analysis of stool specimens gave results that correlated well with the results obtained with cultured stool specimens; the direct molecular analysis was rapid, informative, and less costly than the testing of multiple patient stool isolates.
Clinical Microbiology and Infection, 2001
Objectives To characterise genotypes of Clostridium difficile strains isolated from asymptomatic individuals and patients with diarrhea. Methods Fecal specimens from 235 asymptomatic infants <12 months, 76 asymptomatic children 1-11 years and 132 adult patients with antibiotic-associated and non-antibiotic-associated diarrhea obtained from Siriraj Hospital, Bangkok from October 1998 to April 1999 were examined for C. difficile by cycloserine-cefoxitin-fructose agar culture. The presence of the C. difficile toxin A gene was determined by specific PCR with the use of primers 5-(CCC AATAGA AGATTC AATATTAAG CTT)-3 and 5-(GGA AGA AAA GAA CTT CTG GCT CAC TCA GGT)-3. All C. difficile isolates were subsequently genotyped by pulsed-field gel electrophoresis (PFGE). Results The C. difficile strains were found in 28 (11.9%) asymptomatic infants, 16 (21.1%) asymptomatic children and 33 (25%) adult patients. In total, 14 PFGE types and eight subtypes designated as types A
PLOS ONE
In Colombia, the epidemiology and circulating genotypes of Clostridium difficile have not yet been described. Therefore, we molecularly characterized clinical isolates of C.difficile from patients with suspicion of C.difficile infection (CDI) in three tertiary care hospitals. C.difficile was isolated from stool samples by culture, the presence of A/B toxins were detected by enzyme immunoassay, cytotoxicity was tested by cell culture and the antimicrobial susceptibility determined. After DNA extraction, tcdA, tcdB and binary toxin (CDTa/CDTb) genes were detected by PCR, and PCR-ribotyping performed. From a total of 913 stool samples collected during 2013-2014, 775 were included in the study. The frequency of A/B toxinspositive samples was 9.7% (75/775). A total of 143 isolates of C.difficile were recovered from culture, 110 (76.9%) produced cytotoxic effect in cell culture, 100 (69.9%) were tcdA +/tcdB+, 11 (7.7%) tcdA-/tcdB+, 32 (22.4%) tcdA-/tcdB-and 25 (17.5%) CDTa+/CDTb+. From 37 ribotypes identified, ribotypes 591 (20%), 106 (9%) and 002 (7.9%) were the most prevalent; only one isolate corresponded to ribotype 027, four to ribotype 078 and four were new ribotypes (794,795, 804,805). All isolates were susceptible to vancomycin and metronidazole, while 85% and 7.7% were resistant to clindamycin and moxifloxacin, respectively. By multivariate analysis, significant risk factors associated to CDI were, staying in orthopedic service, exposure to third-generation cephalosporins and staying in an ICU before CDI symptoms; moreover, steroids showed to be a protector factor. These results revealed new C. difficile ribotypes and a high diversity profile circulating in Colombia different from those reported in America and European countries.
2014
Background and Aims: Clostridium difficile is an identified cause of antibioticassociated diarrhea, antibiotic-associated colitis, pseudomembranous colitis and nosocomial diarrhea. The objective of this survey was to determine molecular analysis of toxigenic Clostridium difficile isolates from hospital environment in Tehran tertiary medical centers. Materials and Methods: In this descriptive study, 100 hospital environmental specimens were collected. The specimens were cultured on a selective cycloserine cefoxitin fructose agar, and incubated in anaerobic conditions, at 37°C for 2 days. Clostridium difficile isolates were characterized by conventional biochemical tests. Bacterial cytotoxicity was assayed on tissue culture, and also all strains were typed by PCR ribotyping method. Results: Among toxigenic Clostridium difficile isolates, 6 isolates had the same PCR ribotyping patterns, and 11 isolates were typed in four different groups. Conclusion: Our findings showed that toxigenic ...
Molecular epidemiology of Clostridium difficile infection in Iranian hospitals
Antimicrobial Resistance & Infection Control
Background: Clostridium difficile infection (CDI) is known as one of the most important causes of nosocomial infections. The main objective of this study was to evaluate the presence of Clostridium difficile in the stool of hospitalized patients with diarrhea as well as in their environments. Methods: C. difficile isolates were characterized according to the presence of toxin genes and antibiotic resistance. Multilocus Sequence Typing Analysis (MLST) was applied for finding the genetic polymorphism and relationship among strain lineages. Results: A total of 821 samples (574 stools and 247 swabs) were collected between April 2015 and May 2017. The prevalence of C. difficile isolates was 28.6% (164/574) in patients and 19% (47/247) in swabs taken from medical devices, hands of healthcare workers and skin patient sites. Finally, 11.5% (66/574) toxigenic C. difficile strains isolated from stool samples of inpatients and 4.4% (11/247) from hands of healthcare workers and skin patient sites. All the toxigenic isolates were inhibited by a low concentration of vancomycin (MIC < 0.5 μg/ml). About 43% (33/77) and 39% of isolates were resistant to Clindamycin and moxifloxacin respectively. All isolates were susceptible to metronidazole. Toxigenic C. difficile strains were analyzed by MLST and were divided into 4 different STs. The detected types were ST-54 (57.9%), followed by ST-2 (31.6. %), ST-15 (5.3%) and ST-37 (5.3%), while none of the isolates were identified as ST-1 or ST-11. Significant risk factors for CDI appear to be advanced age, undergoing chemotherapy, previous surgery, and residence in the nursing home. Conclusions: CDI is common in Iran and further studies are recommended to monitor its epidemiological variations. Moreover, greater attempts must be made to encourage antibiotic stewardship by healthcare workers and the public.
Journal of medical microbiology, 2003
Ninety-five isolates of Clostridium difficile from symptomatic and asymptomatic patients and 18 from their environment in the intensive-therapy units (ITUs) of four teaching hospitals in Kuwait were typed by PCR amplification of rRNA intergenic spacer regions (PCR ribotyping). A total of 32 different ribotypes was detected among the clinical isolates. The predominant ribotypes from the clinical isolates were types 097 and 078, which accounted for approximately 40 % of all isolates in the ITUs in Kuwait. Ribotypes 097 (toxigenic), 078 (toxigenic) and 039 (non-toxigenic) were three distinct clones that were circulating in all four hospitals. Ribotypes 097, 078 and 076 (i.e. 50 % of isolates from symptomatic patients) were the predominant isolates associated with C. difficile-associated disease (CDAD). The environmental isolates belonged to a diverse range of ribotypes, with no particular types common to all the hospitals. Ribotype 078 was found only in the patient environment in Mubar...