Characterization and molecular epidemiology of extended spectrum beta-lactamase producing Enterobacter cloacae isolated from a Tunisian hospital.

In 2009, out of the 66 nonrepetitive Enterobacter cloacae collected at Charles Nicolle hospital in Tunisia, 44 were extended spectrum ß-lactamase (ESBL) producers. The aim of the current study was to detect and characterize the genes encoding the ESBLs including blaTEM, blaSHV, and blaCTX-M groups by polymerase chain reaction and sequencing. Pulsed-field gel electrophoresis (PFGE) analysis was used to determine the genetic relatedness between isolates. All strains were susceptible to carbapenems. They were resistant to fluoroquinolones, gentamicin, tobramycin, and trimethoprim+sulfamethoxazole but variably resistant to netilmicin, amikacin, and tetracyclines. Sequence analysis of the polymerase chain reaction products revealed the presence of blaCTX-M-15 (39 strains), blaSHV-12 (6 strains), and blaSHV-27 (1 strain). The coexistence of two ESBLs was observed in two isolates harboring, respectively, SHV-12+CTX-M-15 and SHV-27+CTX-M-15. PFGE revealed 36 unrelated profiles. Diffusion of E. cloacae producing CTX-M-15 ESBL in our hospital is the consequence of dissemination of identical or related plasmids harboring the CTX-M-15 gene.

Carbapenem-resistant Acinetobacter baumannii producing the carbapenemase OXA-23 in Tunisia.

AIM: To analyze the mechanisms of resistance to carbapenems among imipenem resistant A. baumannii recovered from different wards at Charles Nicolle Hospital. METHODS: From January to December 2007, 50 carbapenem-resistant A. baumannii isolates were recovered from hospitalized patients. MICs were performed by agar dilution method and interpreted according to CLSI guidelines. Metallo-ß-lactamase production was evaluated using imipenem-EDTA disk synergy test. PCR and DNA sequencing targeting blaOXA genes were performed and pulsed field gel electrophoresis was used for epidemiologic study. RESULTS: Most of the isolates were obtained from patients hospitalized in surgery (62%) and Intensive Care Units (22%). All strains showed high level of resistance to ticarcillin (MIC50 > 2048µg/ml), ticarcillin-clavulanic acid (MIC50 >1024µg/ml), aztreonam (MIC50 = 512µg/ml), ceftazidim (MIC50 = 512µg/ml), imipenem (MIC50 = 512µg/ml), meropenem (MIC50 =128µg/ml) and cefepime (MIC50 = 256µg/ml). Metallo-ß-lactamase production was negative for all isolates. The co-existence of blaOXA-51-like/ blaOXA-23-like was detected in 82% (n= 41). The genes blaOXA- 24-like and blaOXA-58-like were not found in any isolate. All isolates harboured a blaOXA-51-like gene. Sequencing confirmed the presence of blaOXA-23 and blaOXA-69 genes. Eight distinct patterns were observed (A: 41 isolates, B: 1 isolate, C: 1 isolate, D: 1 isolate, E: 1 isolate, F: 2 isolates, G: 1 isolate, H: 2 isolates). CONCLUSION: Production of OXA-23 was the important mechanism of resistance to carbapenem among A. baumannii. Strengthening of prevention measures are required to control further spread of carbapenemases in Tunisia.

Clonal spread of carbapenem resistant Pseudomonas aeruginosa in an university hospital.

AIM: We examined 14 Pseudomonas aeruginosa clinical isolates collected in 2000 from patients hospitalised in different wards at Charle Nicolle hospital from Tunisia. METHODS: Analysis includes serotyping, antimicrobial susceptibility profile, beta-lactamase detection, randomly amplified polymorphic DNA and pulsed field gel electrophoresis. All Pseudomonas aeruginosa strains belonged to serotype O12 and they demonstrated a high level of resistance to all antibiotioc tested. RESULTS: Beta-lactamase detection revealed that 9 of these strains had ceftazidime activity restored by cloxacillin and none of the 14 strains were metallo-beta-lactamase producing. Randomly amplified polymorphic DNA analysis and pulsed field gel electrophoresis were able to discriminate isolates and gave concordant results which showed epidemiologically related strains. CONCLUSION: These results confirm a clonal spread of multiresistant Pseudomonas aeruginosa O12 throughout the hospital.

Spread of multidrug resistant Acinetobacter baumannii in a teaching hospital.

BACKGROUND: A. baumannii is an important opportunistic pathogen widely distributed in the hospital environment and responsible for a variety of nosocomial infections especially in patients from intensive care units. AIM: We describe an outbreak of Acinetobacter baumannii (16 stains) in 3 intensive care units (I, II, III) at Charles Nicolle hospital of Tunis over a 5 month period (March to July 2005). METHODS: The antimicrobial susceptibility was determined by disc diffusion test and the genetic relatedness of isolates was done by Random Amplified Polymorphic DNA (RAPD) analysis. Two strains not related to the outbreak were used for the discrimination of the technique. RESULTS: Samples were collected from blood (44%), materials (31%), pus (6.5%), urines (6.5%) and respiratory tract (12.5%). Antibiotic resistance pattern showed 2 different profiles. However, molecular typing of isolates revealed 3 distinct profiles (A, B, C) represented respectively by 8, 7 and one isolates. The major profile was the profile A found in 5 patients and in materials. It was appeared firstly in intensive care unit I, then in the 2 other units (II and III). The profile B was observed also in the 3 units. However, the profile C was found in one patient in unit I. CONCLUSION: These data emphasize the need for active surveillance for multidrug-resistant Acinetobacter baumannii, and the value of molecular typing of strains in hospital settings to investigate spread of infection.

[Pseudomonas aeruginosa: a multicentric study of antibiotic resistance (1999-2000)]. / Pseudomonas aeruginosa: étude multicentrique de la résistance aux antibiotiques (1999-2000).

A retrospective multicentric study was carried out over a period of 2 years (1999-2000). 2659 strains of Pseudomonas aeruginosa were collected from 4 university hospitals (Charles Nicolle Hospital, Pediatric Hospital and National Centre of Bone Marrow Transplantation in Tunis, Habib Bourguiba Hospital in Sfax). Epidemiological profile and antibiotic susceptibility were analysed. All bacteria were identified by conventional methods and antibiotic susceptibility tests were performed according to CA-SFM guidelines. The strains were recovered essentially from surgical wards (33%) and intensive care units (22%). Pseudomonas aeruginosa was isolated mainly from pus (36%), urine (32%) and respiratory samples (18%). 25% of strains were resistant to ticarcilline, 18% to cefsulodine, 9% to ceftazidime, 14% to imipenem and amikacin and 25% to ciprofloxacin. Moreover, the resistance rates varied from hospital to hospital and from unit to another. The resistant strains were isolated particularly from urology and intensive care units: respectively 62% and 39% for ticarcilline; 26% and 13% for ceftazidime. The acquired resistance to b-lactams seems largely due to penicillinase production. The frequency of resistance to ceftazidime was the lowest and seems associated to chromosomal cephalosporinase over production.

[Multidrug resistance in Klebsiella pneumoniae: multicenter study]. / Multiresistance aux antibiotiques de Klebsiella pneumoniae: etude multicentrique.

The extensive use of broad spectrum antibiotics, especially the third generation cephalosporins (C3G), was followed by the emergence of newer plasmid mediated betalactamases called extended spectrum betalactamases (ESBLs). To assess the impact of K. pneumoniae resistant to 3GC in Tunisia, this study was conducted in 3 teaching hospitals. A total of 1110 strains of K pneumoniae was collected. The antibiotics susceptibilities were tested by diffusion method using Mueller-Hinton agar. The quality control was regularly performed. I ESBLs producing solates were detected using the double-disc synergy test. Data analysis was done using the Whonet 4 software. 23.6% K. pneumoniae isolates showed phenotype pattern of ESBLs producers. The double-disc synergy test was positive in 75% of the cases. These isolates were recovered from hospitalized patients in different wards but mainly from pediatrics (23.6%), medicine (23.2%), surgery (22.9%), intensive care units (11%) and neonatology (11%). 54% were isolated from urines, 22% from blood cultures. These isolates remained susceptible to imipenem (100%) and most of them to cefoxitin (96.4%) but all had associated resistance to aminoglycosides, quinolones and trimethoprim-sulfamethoxazole. The prevalence of multidrug resistant K. pneumoniae is high. This resistance can be minimized by the implementation of infection control measures including handwashing and isolation procedures.