| Antibiotic-associated diarrhoea (AAD) is described as diarrhoea that occurs in conjunction with antibiotic administration and cannot be explained by another diagnosis. AAD can range from a mild, self-limiting illness to more serious and progressive disease such as pseudomembranous colitis (PMC). AAD results from overgrowth of the intestinal mucosa by pathogenic microorganisms after antibiotic treatment, however, it may also occur in response to a reduction in the concentration of fecal flora.Clostridium difficile is the most important infectious cause of AAD. Since 2002, the incidence and severity of C. difficile infection (CDI) have increased dramatically in North America and Europe. Almost 10-30% of AAD is due to C. difficile; when considering severe cases of AAD such as documented antibiotic-associated PMC, 90%-100% are attributed to CDI.However, there is still a paucity of data in China. A prospective, observational study was conducted to evaluate the clinical and molecular characteristics of CDI in patients with AAD at University Hospital Huashan from August 2012 to July 2013.There are two parts in this study:Part one. Clinical characteristics of Clostridium difficile infection in hospitalized patients with antibiotic-associated diarrhoeaPart two. Molecular characteristics of Clostridium difficile clinical Isolates: antimicrobial resistance, toxin profiles and PCR ribotyping.Part one. Clinical Characteristics of Clostridium difficile Infection in Hospitalized Patients with Antibiotic-associated DiarrhoeaAll fecal specimens from patients who hospitalized in Huashan Hospital and developed AAD were sent to microbiological laboratory for the detection of toxigenic C. difficile from August 1,2012 to July 31,2013. Eligibility criteria included being a hospitalized patient aged ≥18 years with acute diarrhoea, and receiving antibiotic treatment ≤4 weeks before the onset of diarrhoea. Patients with chronic diarrhoea or who had history of using a laxative within 3 days preceding diarrhoea onset were excluded. A case of AAD was defined as otherwise unexplained diarrhoea in hospitalized patient occuring in association with the administration of antibiotics. A case-patient with CDI was defined as a positive toxigenic C. difficile culture from a diarrhea hospitalized patient, or PMC diagnosed during enteroscopy. A questionnaire was completed for each patient. The following data were collected:demographic data; presumed risk factors in the 4 weeks before the onset of diarrhea; biological parameters; and clinical course and outcomes. SPSS, version 16.0 was used for statistical analyses.AAD developed in 206 hospitalized patients and toxigenic C. difficile was isolated from 30.6%(63/206) of diarrhoeal stool samples. The frequency of AAD was highest in the Intensive care unit (10.7%,56/265) while the proportion of CDI in AAD was highest in the Geriatric unit (38%,11/29). The mean age of patients with CDI and non-C. difficile AAD were 66±17 and 62±20 years, respectively. Most of the patients were male. Diabetes mellitus and malignancy were the most common underlying diseases in both groups. Carbapenems were prescribed significantly more frequently in patients with CDI than those with non-C. difficile AAD (42.9% vs.28.0%, p= 0.036). After univariate analysis and using a multivariate logistic regression model, only the use of carbapenems was found to be significantly associated with increased risk of CDI (RR,2.31; 95% CI,1.22 to 4.38; P= 0.011). Neither fever nor abdominal pain showed significant differences between CDI and non-C. difficile AAD patients. Most of AAD ranged in severity from mild to moderate; only one case with pseudomembranous colitis was identified.36.5% (23/63) and 5.6% (8/143) of patients with CDI or with non-C. difficile AAD received metronidazole and/or vancomycin as empirical treatment, respectively. Almost 90% of patients with CDI or non-C. difficile AAD were cured; 2 patients had CDI recurrence.In conclusion, C. difficile plays an important role in AAD in China. It is difficult for clinicians to identify CDI cases based on clinical presentation of AAD patients since the severity ranges of disease ranges from mild to moderate diarrhoea. Therefore, C. difficile infections should be included in the routine differential diagnoses for hospitalized patients with diarrhoea, especially AAD.Part Two. Molecular Characteristics of Clostridium difficile Clinical Isolates:PCR ribotyping, Toxin profiles and Antimicrobial resistanceAll C. difficile strains isolated from patients who hospitalized in Huashan Hospital and developed AAD were collected from August 1,2012 to July 31,2013. C. difficile isolates were analyzed for their antibiotic susceptibility patterns using agar dilution method. PCR ribotyping and Real-time PCR to detect tcdA, tcdB, ctdA, and cdtB genes were performed. Toxin B was reconfirmed by the cytotoxicity neutralization assay.Totally 64 toxigenic C. difficile isolates were collected,67.2% of the isolates were toxin A-positive and toxin B-positive (A+B+),32.8% were toxin A-negative but toxin B-positive (A’B+). No binary toxin was detected in any isolates. Eighteen different PCR ribotypes were identified with a specific clone (H) accounting for 18.8%, followed by 012 (14.1%) and 017 (12.5%). None of them belonged to ribotype 027. All isolates were susceptible to metronidazole, vancomycin. Resistance to erythromycin, clindamycin, tetracycline, moxifloxacin, levofloxacin, fusidic acid, rifampin and imipenem was found in 73.4%,84.4%,48.4%,42.2%,47.4%,35.9%, 7.8% and 6.2% of the isolates, respectively. Coresistance to erythromycin, clindamycin and fluroquinolones were found in 32.8% of C. difficile isolates. All the ribotype H isolates and 62.5% of ribotype 017 isolates showed multiresistant to the antimicrobial agents tested.In conclusion, most of the clinical isolates of C. difficile isolated in Huashan Hospital are toxin A-positive and toxin B-positive. Ribotype H is the dominant clone. All the isolates are still fully susceptible to metronidazole and vancomycin, however, the resistant rates to other antimicrobial agents tested such as clindamycin are high.Summary:1. Clostridium difficile was the most important infectious cause of AAD, accounting for 30.6%.2. Use of carbapenems was found to significantly increase the risk of CDI; patient demographics, presumed risk factors, clinical manifestations and laboratory findings revealed no significant difference between patients with CDI and non-C. difficile AAD.3. Most of the clinical isolates of C. difficile were toxin A-positive and toxin B-positive. Ribotype H was the dominant clone. All the isolates were still susceptible to metronidazole and vancomycin, however, the resistant rates to other antimicrobial agents tested such as clindamycin were very high.The present study is the first systematic survey of clinical and molecular C. difficile infections among patients with antibiotic-associated diarrhoea in China. The results suggest CDI should be included in the routine differential diagnoses for hospitalized patients, especially who presenting with AAD. A limitation of this study is that only one hospital is enrolled therefore the data could not represent China. Further national surveillance is crucial to monitor the incidence, identify populations at risk, and characterize the molecular epidemiology of strains causing CDI. |
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