An outbreak of Mycobacterium abscessus infection in a Pediatric Intensive Care Unit in Kuwait.

BACKGROUND: Mycobacterium abscessus has been associated with respiratory tract infections, localized skin and soft tissue infections and sepsis. However, outbreaks of M. abscessus are rare. AIM: to report an outbreak of M. abscessus causing respiratory tract infections in a Pediatric Intensive Care Unit (PICU) in Kuwait, its investigation and control measures. METHODS: Respiratory secretions were obtained from ventilator-dependent patients showing signs of sepsis, including fever, malaise and weight loss. The specimens were cultured on appropriate routine media. After the results of the sample taken from the index case as acid-fast bacilli positive, all patients were screened for M. abscessus carriage. Isolates were identified by INNO-LiPA Mycobacteria v2 line probe assay and DNA sequencing. Molecular fingerprinting DiversiLab strain typing was performed on the isolates. Epidemiologic investigation was conducted during the outbreak. FINDINGS: the outbreak affected 5 patients, 4 of whom had severe infections including 1 patient with septicemia. Asymptomatic carriage of outbreak strain was found in 1 patient. All environmental samples were negative for M. abscessus but some were positive for M. gordonae and M. fortuitum. The source could not be identified. Stringent infection control measures were put in place, including reemphasizing hand hygiene and closure of the Pediatric Intensive Care Unit to new admissions. A year later, no further case has occurred after the last case. CONCLUSION: To our knowledge, this is the first report of a hospital-acquired outbreak of respiratory tract infection caused by M. abscessus in a Pediatric Intensive Care Unit. In the absence of definite source identification, reinforcement of standard infection control guidelines was successful in containing the outbreak.

An outbreak of CTX-M-15-producing Klebsiella pneumoniae isolates in an intensive care unit of a teaching hospital in Kuwait.

OBJECTIVE: This study reports an outbreak of Klebsiella pneumoniae infections in 14 patients during a 2-month period (August-September, 2008) in the intensive care unit (ICU) of a teaching hospital in Kuwait. MATERIALS AND METHODS: The clinical sources were blood (9), urine (3) and respiratory secretions (2) identified by the automated VITEK-2 ID System. Susceptibility testing was performed by the E-test method. Extended-spectrum ß-lactamase (ESBL) production was assessed using the ESBL E-test and confirmed by PCR. Carriage of bla genes was determined by PCR and sequence analysis. The transferability of resistance phenotypes was demonstrated by conjugation experiments and clonal relatedness was determined by PFGE. RESULTS: The isolates were susceptible to imipenem, meropenem, and tigecycline and produced ESBL. All isolates yielded an amplicon of 499 bp with universal consensus primers (MA primers). DNA sequence analysis showed that they all harboured bla CTX-M-15 and bla TEM-1 genes. The environmental isolate obtained from a suction machine was also CTX-M-15/TEM-1 producer. The resistance phenotypes were transferrable to the Escherichia coli J53 r strain. PFGE, revealed two clones, A and B, related with a Dice coefficient of >94.1%. A mortality rate of 21.4% was recorded. CONCLUSION: The outbreak was contained by robust and aggressive infection control measures. This study highlights the first outbreak of CTX-M-15-producing K. pneumoniae associated with high mortality in an adult medical ICU in Kuwait.

Sequence analysis of bla(CTX-M) genes carried by clinically significant Escherichia coli isolates in Kuwait hospitals.

OBJECTIVE: To investigate the extent, distribution and sequence analysis of bla(CTX-M) genes carried by Escherichia coli isolated from patients admitted to all government hospitals in Kuwait. METHODS: Extended-spectrum ß-lactamase (ESBL)-producing E. coli isolates were collected from the 8 major hospitals in Kuwait. CTX-M ESBLs were analyzed by PCR and sequenced. Clonality of the positive isolates was determined for genetic relatedness using pulsed-field gel electrophoresis (PFGE) with XbaI digestion of the genomic DNA. RESULTS: Of the 136 ESBL-positive isolates, 106 (77.9%) harbored bla(CTX-M) genes. Among these, bla(CTX-M-15) was the most frequent with a prevalence rate of 84.1%, followed by bla(CTX-M-14) (6.8%), bla(CTX-M-14b) (5.7%) and bla(TOHO-1) (3.4%). Ninety-three (87.7%) were isolated from Kuwaiti (35.9%), Egyptian (31.1%) and Indian (20.8%) nationals; the majority of isolates positive for bla(CTX-M-15) were mainly from these 3 nationalities. PFGE analysis did not demonstrate any clustering of positive isolates in any particular hospital. CONCLUSION: This study confirms an explosive emergence of CTX-M-15 ß-lactamase among E. coli isolates in Kuwait and shows that the strains were clonally heterogeneous with no evidence of inter- or intra-hospital spread. Thus Kuwait may represent an important source of CTX-M-15-positive E. coli.

National surveillance of antimicrobial susceptibility of CTX-M-positive and -negative clinical isolates of Escherichia coli from Kuwait government hospitals.

Antibiotic resistance in Escherichia coli is becoming a complex therapeutic problem. Surveillance programs are valuable tools and offer important information on bacterial resistance trends. This study was undertaken to determine the susceptibility of clinically significant isolates of E. coli obtained from patients admitted to 8 Kuwait government hospitals and to examine how this was influenced by the production of CTX-M extended-spectrum beta-lactamases (ESBLs). The susceptibility of 876 consecutive clinically significant strains of E. coli to 13 antibiotics was determined by Etest. ESBL production was assessed by ESBL-Etest method and the presence of CTX-M beta-lactamases was confirmed by PCR technique. Of the 876 isolates, 604 (69%) were highly non-susceptible to ampicillin with MIC(90 )of >256 microg/ml. Resistance to the 3(rd)-generation cephalosporins ranged from 7.5% in the Maternity hospital to 29% in the Ibn Sina hospital; ciprofloxacin resistance rates ranged from 14% and 40%, respectively. Carbapenems and amikacin demonstrated excellent activity. The minimum inhibitory concentrations (MIC(90)) of cefotaxime, ceftazidime, cefepime and ciprofloxacin were >256, 64, >256 and >32 microg/ml, respectively for CTX-M-positive isolates versus 0.5, 1, 025 and 0.125 microg/ml for CTX-M-negative strains. Frequencies of CTX-M-positive isolates within the cefotaxime MIC ranges of 1-2, 3-8, 9-16 and >16 microg/ml were 0, 4, 15 and 81%, respectively. In conclusion, the susceptibility of E. coli to the 3(rd )generation cephalosporins and ciprofloxacin was influenced by the presence of CTX-M ESBL and a high proportion of the CTX-M-producing isolates were in the susceptibility ranges of cefotaxime.

Comparative in vitro activity of tigecycline and nine other antibiotics against gram-negative bacterial isolates, including ESBL-producing strains.

The enterobacteriaceae, especially Escherichia coli and Klebsiella spp., as well as Acinetobacter spp., are important agents of nosocomial infections in hospitalized patients. A total of 460 Gram-negative bacteria (GNb), were investigated for their susceptibility to tigecycline and 9 other antibiotics by the etest. ESBL production was inferred from ESBL etest phenotypes. All the GNb, including the ESBL-producers, were susceptible to tigecycline with MIC(90 )ranges of 0.25 to 2 microg/ml. Imipenem and meropenem were very active against ESBL and non-ESBL producers; mean MIC(90)s of 0.19 and 0.09 microg/ml and 0.05 microg/ml and 0.02 microg/ml, respectively. The MIC(90)s of imipenem and meropenem for the Acinetobacter spp. were 16 and >32 microg/ml, respectively with resistance rates of 64.3 and 66.1%. ESBL production was detected in 62% and 82.1% of the E. coli and K. pneumoniae isolates, respectively. Resistance to ciprofloxacin was higher among the ESBL-producing strains of E. coli and K. pneumoniae than the non-ESBL producers. Comparatively, tigecycline had excellent in vitro activities against ESBL-producing enterobacteriaceae and demonstrated superior activity against Acinetobacter spp. Increasing ESBL production and resistance to ciprofloxacin and gentamicin in enterobacteriaceae require careful selection of empirical therapy. Tigecycline holds promise as an alternative choice of therapy for infections caused by ESBL-producing isolates and multi-drug resistant Acinetobacter spp.

Role of tigecycline in the control of a carbapenem-resistant Acinetobacter baumannii outbreak in an intensive care unit.

The incidence of Acinetobacter baumannii infection has greatly increased over recent decades with infections occurring more in critically ill hospitalised patients. Hospital outbreaks of multiple antibiotic-resistant strains are posing an increasing threat to public health. Three different outbreaks of multidrug-resistant A. baumannii (MRAB) infections involving 24 patients, aged 16-75 years occurred in the intensive care unit in the course of one year. The isolates were cultured from clinical samples and identified using automated Vitek II ID system and the API 20NE system. Susceptibility testing was done by the E-test method. Molecular typing of the isolates was determined by pulsed-field electrophoresis. Screening of both patients and the environment was carried out. The acquisition time, i.e. the time of admission to time of acquiring infection, ranged from 3 to 31 days. All isolates were multiply resistant (MRAB), including resistance to carbapenems (MRAB-C) in the majority of cases but susceptible to tigecycline, with a minimum inhibitory concentration (MIC(90)) of 2 microg/mL. The overall mortality rate was 16.7%. Time-to-clearance of the MRAB-C was 8.3 days in the first outbreak, when tigecycline was not used, and 2.8 and 3.1 days during the second and third outbreaks, respectively, when tigecycline was used, and all but one patient survived. Environmental screening revealed gross contamination of many surfaces and equipment within the unit. The outbreak strains belonged to two distinct clones (D and E) whereas the 14 environmental strains belonged to three distinct groups (A-C). The outbreak of infections treated with tigecycline was successfully eliminated in conjunction with an aggressive infection control strategy.

Evidence from CD spectra and melting temperatures for stable Hoogsteen-paired oligomer duplexes derived from DNA and hybrid triplexes.

The pyr*pur.pyr type of nucleic acid triplex has a purine strand that is Hoogsteen-paired with a parallel pyrimidine strand (pyr*pur pair) and that is Watson-Crick-paired with an antiparallel pyrimidine strand (pur.pyr pair). In most cases, the Watson-Crick pair is more stable than the Hoogsteen pair, although stable formation of DNA Hoogsteen-paired duplexes has been reported. Using oligomer triplexes of repeating d(AG)12 and d(CT)12 or r(CU)12 sequences that were 24 nt long, we found that hybrid RNA*DNA as well as DNA*DNA Hoogsteen-paired strands of triplexes can be more stable than the Watson-Crick-paired strands at low pH. The structures and relative stabilities of these duplexes and triplexes were evaluated by circular dichroism (CD) spectroscopy and UV absorption melting studies of triplexes as a function of pH. The CD contributions of Hoogsteen-paired RNA*DNA and DNA*DNA duplexes were found to dominate the CD spectra of the corresponding pyr*pur.pyr triplexes.

Hybrid oligomer duplexes formed with phosphorothioate DNAs: CD spectra and melting temperatures of S-DNA.RNA hybrids are sequence-dependent but consistent with similar heteronomous conformations.

Knowledge of the relative stabilities of S-DNA.RNA hybrids of different sequences is important for choosing RNA targets for hybridization with antisense phosphorothioate oligodeoxyribonucleotides (S-DNAs). It is also important to know how hybrid secondary structure varies with sequence, since different structures could influence thermal stability and the activity of RNase H. Our approach has been to study relatively simple sequences consisting of repeating di-, tri-, and tetranucleotides, which allow the maximum resolution of nearest-neighbor effects. Circular dichroism (CD) spectra and melting temperatures were acquired for 16 hybrid sequences that could be formed by mixing S-DNA and RNA oligomers of 24 nucleotides in length. CD spectra of S-DNA.RNA hybrids were sequence-dependent and were similar to those of analogous unmodified hybrids. From singular value decomposition, the major CD spectral component was like that of the A-conformation. Three nearest-neighbor relationships among the hybrid CD spectra were in as good agreement as are such relationships among spectra of duplex RNAs. Tm values ranged from 44.1 degrees C for S-d(ACT)8. r(AGU)8 to 66.6 degrees C for S-d(CCT)8.r(AGG)8 (in 0.15 M K+, phosphate buffer, pH 7). The S-DNA.RNA hybrids had a sequence-dependence of melting temperatures that was approximately the same as that calculated using published data for normal DNA.RNA hybrids [Sugimoto, N., Nakano, S., Katoh, M., Matsumura, A., Nakamuta, H., Ohmichi, T.,Yoneyama, M., & Sasaki, M. (1995) Biochemistry 34, 11211-11216]. In general, sequence-dependent CD spectra and Tm values of S-DNA.RNA hybrids appear to reflect the unique nearest-neighbor interactions of adjacent base pairs, where the S-DNA and RNA strands are in different, but relatively uniform, conformations.

Evaluation of the Baxter-MicroScan 4-hour enzyme-based yeast identification system.

A new 4-h Yeast Identification Panel (YIP; Baxter-MicroScan, W. Sacramento, Calif.) was compared with the API 20C Yeast Identification System (Analytab Products, Inc., Plainview, N.Y.) in the identification of recent clinical yeast isolates. The YIP had a 94% correlation (288 of 306) in identifying 22 species within the genera Candida, Hansenula, Pichia, Rhodotorula, Saccharomyces, and Torulopsis. Correlation dropped to 65% for those species within the genera of slower growing yeasts, i.e., Blastoschizomyces spp., Crpytococcus spp., Geotrichum spp., Hyphopichia spp., Phaeococcomyces spp., Prototheca spp., and Trichosporon spp. Overall correlation with the API 20C was 92% (365 of 401) for those taxa included in the data base and 85% (373 of 437) for all yeasts encountered in the study. There were 36 (8.2%) discrepant identifications, which were due in part to the limited data base. Expansion of the data base plus the easy inoculation, reading, and rapid results of the YIP should make it an excellent method for yeast identification.