Haihui Huang

Antimicrobial resistance in Clostridium difficile.

Clostridium difficile is the leading cause of hospital-acquired diarrhoea and the number of outbreaks has risen markedly since 2003. The emergence and spread of resistance in C. difficile is complicating treatment and prevention. Most isolates are still susceptible to vancomycin and metronidazole (MTZ), however transient and heteroresistance to MTZ have been reported. The prevalence of resistance to other antimicrobial agents is highly variable in different populations and in different countries, ranging from 0% to 100%. Isolates of common polymerase chain reaction (PCR) ribotypes are more resistant than uncommon ribotypes. Most of the resistance mechanisms that have been identified in C. difficile are similar to those in other Gram-positive bacteria, including mutation, selection and acquisition of the genetic information that encodes resistance. Better antibiotic stewardship and infection control are needed to prevent further spread of resistance in C. difficile.

Comparison of a Commercial Multiplex Real-Time PCR to the Cell Cytotoxicity Neutralization Assay for Diagnosis of Clostridium difficile Infections

ABSTRACT A commercial multiplex real-time PCR assay (Cepheid Xpert C. difficile assay) for the diagnosis of Clostridium difficile infection was evaluated. The sensitivity and specificity of the Cepheid assay were 97.1% and 93.0% for fresh stools, using the cell cytotoxicity neutralization assay as the reference. Using PCR ribotyping as the reference for ribotype 027 strains, the corresponding figures were 100% and 98.1%, respectively.

Clostridium difficile infections in a Shanghai hospital: antimicrobial resistance, toxin profiles and ribotypes

The incidence of Clostridium difficile infection (CDI) has risen markedly since 2003, however data from China are limited. A 1-year study was conducted at the University Hospital Huashan to characterise clinical isolates of C. difficile. Of 74 isolates, 56 were from the first episode of CDI (43 A(+)B(+) and 13 A(-)B(+)), 5 were from recurrences and 13 were toxin-negative. No binary toxin or TcdC deletion was detected. All strains were susceptible to metronidazole, vancomycin, meropenem and piperacillin/tazobactam. Resistance to moxifloxacin, ciprofloxacin, levofloxacin, erythromycin, clindamycin, tetracycline, rifampicin and fusidic acid was found in 46.4%, 100%, 60.7%, 71.4%, 71.4%, 35.7%, 25.0% and 17.9% of the isolates, respectively. All moxifloxacin-resistant isolates carried a mutation in either gyrA, gyrB or both. Fourteen different polymerase chain reaction ribotypes were identified, with a specific clone (SH II) accounting for 25% of isolates. No isolates belonged to ribotype 027. The present study is the first systematic survey of clinical C. difficile isolates in China. Further surveillance is required to detect clustering of cases and to monitor the emergence of specific highly virulent clones and resistance.

Distinct ribotypes and rates of antimicrobial drug resistance in Clostridium difficile from Shanghai and Stockholm

Seventy-five clinical isolates of Clostridium difficile from Shanghai and 80 from Stockholm were investigated. The prevalence of toxin A-negative, toxin B-positive isolates of C. difficile among isolates from Shanghai (33.3%) was significantly higher than among isolates from Stockholm (0%). Both sets of isolates were fully susceptible to metronidazole and vancomycin. However, the MICs of fluoroquinolones, erythromycin-clindamycin, tetracycline, rifampin and fusidic acid were significantly higher for the Shanghai isolates than for the Stockholm isolates. Thirty-three PCR ribotypes were identified; a dominant clone, 017, accounted for 18.7% of Shanghai isolates, whereas clone 005 dominated among Stockholm isolates, accounting for 11.3%. Strains 027 and 078 were not detected. No outbreak occurred during the study period.

Antimicrobial susceptibility and heteroresistance in Chinese Clostridium difficile strains

One hundred and ten toxigenic Clostridium difficile isolates collected between December 2008 and May 2009 at Fudan University Hospital Huashan were analyzed for their antibiotic susceptibility patterns and resistance molecular basis. The heteroresistance to metronidazole in fresh isolates were detected as well. Sixteen different PCR ribotypes were identified with a dominant clone 017 accounting for 37.3% of the isolates, followed by 001 and H. Ribotype 027 was not found but one isolate belonged to ribotype 078. All the isolates were susceptible to vancomycin and piperacillin/tazobactam. Seventy-eight fresh isolates were tested for heteroresistance to metronidazole, 18 (23.1%) of them were found to be positive. Resistance to moxifloxacin, ciprofloxacin, levofloxacin, erythromycin, clindamycin, tetracycline, rifampin, rifaximin and fusidic acid was found in 61.8%, 100%, 66.4%, 85.3%, 88.1%, 62.7%, 29.1%, 29.1% and 8.2% of the isolates, respectively. The isolates of common PCR ribotypes were more frequently resistant than the uncommon ribotypes. The prevalence of resistance genes and mutations among the resistant isolates was as follows: ermB, 69.1%; tetM, 97.1%; gyrA mutation, 63.2%; gyrB mutation, 4.4%; gyrA and gyrB mutation, 32.4%; rpoB mutation, 100%, respectively. The resistance related fusA mutation was only found in one isolate with minimum inhibitory concentration of 4 mg/L.

Risk factors of Clostridium difficile infections among patients in a university hospital in Shanghai, China

Clostridium difficile infection (CDI) is an increasing concern in China. However, the risk factors of CDI are rarely reported in the Chinese population. A prospective observational study was therefore conducted among patients with hospital-acquired C. difficile diarrhoea and the risk factors of CDI in a retrospective case-control study. The CDI patients were compared with the non-CDI diarrhoeal patients and those without diarrhoea, respectively. The recurrent CDI patients were compared with the corresponding non-recurrent CDI patients and those without diarrhoea, respectively. Overall, of the 240 patients with hospital-acquired diarrhoea 90 (37.5%) were diagnosed as CDI, and 12 (13.3%) of the 90 CDI patients experienced recurrence. Multivariate analysis indicated that renal disease, malignancy, hypoalbuminemia, prior antibiotic treatment, chemotherapy, nasogastric tube use, length of stay>14 days and intra-abdominal surgery, defined daily dose of antimicrobial agents≥19, prior use of more than three antimicrobial agents, and use of carbapenems were independent risk factors for the first episode of CDI. Use of laxatives, the first- and second-generation narrow-spectrum cephalosporins or metronidazole was identified as protective factors. It is necessary to make testing of C. difficile available as a routine practice and control these risk factors in Chinese hospitals to avoid CDI outbreaks.

Toxin profiles, PCR ribotypes and resistance patterns of Clostridium difficile: a multicentre study in China, 2012–2013

A total of 178 clinical isolates of Clostridium difficile were collected from five major teaching hospitals representing northern, eastern and southern China from August 2012 to July 2013. Among the 178 isolates, 162 (91.0%) were toxigenic, including 66 (40.7%) toxin A-negative, toxin B-positive, 95 (58.6%) toxin A-positive, toxin B-positive and only 1 (0.6%) toxin A-, toxin B- and binary toxin-positive. Twenty-nine different PCR ribotypes were identified, of which 017 (21.0%), 012 (17.3%) and novel type H (16.7%) were the most prevalent. PCR ribotypes 027 and 078 were not found. All toxigenic strains were susceptible to metronidazole, vancomycin and piperacillin/tazobactam. Resistance to moxifloxacin, clindamycin, erythromycin, tetracycline, chloramphenicol, fusidic acid, imipenem, linezolid and rifampicin was observed in 45.1%, 79.6%, 75.3%, 46.9%, 3.7%, 29.6%, 4.9%, 2.5% and 12.3% of the isolates, respectively. Ribotype 017 showed resistance to more antimicrobial agents tested than other ribotype strains.

Evaluation of the in vitro activity of levornidazole, its metabolites and comparators against clinical anaerobic bacteria

This study evaluated the in vitro anti-anaerobic activity and spectrum of levornidazole, its metabolites and comparators against 375 clinical isolates of anaerobic bacteria, including Gram-negative bacilli (181 strains), Gram-negative cocci (11 strains), Gram-positive bacilli (139 strains) and Gram-positive cocci (44 strains), covering 34 species. Minimum inhibitory concentrations (MICs) of levornidazole, its five metabolites and three comparators against these anaerobic isolates were determined by the agar dilution method. Minimum bactericidal concentrations (MBCs) of levornidazole and metronidazole were measured against 22 strains of Bacteroides fragilis. Levornidazole showed good activity against B. fragilis, other Bacteroides spp., Clostridium difficile, Clostridium perfringens and Peptostreptococcus magnus, evidenced by MIC90 values of 0.5, 1, 0.25, 2 and 1mg/L, respectively. The activity of levornidazole and the comparators was poor for Veillonella spp. Generally, levornidazole displayed activity similar to or slightly higher than that of metronidazole, ornidazole and dextrornidazole against anaerobic Gram-negative bacilli, Gram-positive bacilli and Gram-positive cocci, especially B. fragilis. Favourable anti-anaerobic activity was also seen with levornidazole metabolites M1 and M4 but not M2, M3 or M5. For the 22 clinical B. fragilis strains, MBC50 and MBC90 values of levornidazole were 2mg/L and 4mg/L, respectively. Both MBC50/MIC50 and MBC90/MIC90 ratios of levornidazole were 4, similar to those of metronidazole. Levornidazole is an important anti-anaerobic option in clinical settings in terms of its potent and broad-spectrum in vitro activity, bactericidal property, and the anti-anaerobic activity of its metabolites M1 and M4.