The prevalence of imipenem resistance among Pseudomonas aeruginosa isolates at a 195-bed tertiary care medical center in Cali, Colombia, rose from 2% in 1996 to 28% in 1997 and to over 40% in 2003. Many isolates showed high-level multiresistance, and phenotypic characterization suggested the spread of a predominant strain with minor variants. Sixty-six resistant isolates collected between February 1999 and July 2003 from hospitalized patients (n ‫؍‬ 54) and environmental samples (n ‫؍‬ 12) were subjected to a fuller analysis. Genetic fingerprints were compared by pulsed-field gel electrophoresis (PFGE) of SpeI-digested genomic DNA, and bla IMP and bla VIM genes were sought by PCR. PFGE and serotyping indicated that 52 of the 66 isolates belonged to a single strain, with 82% similarity; the PFGE pattern for this organism was designated pattern A. Two further pairs of isolates represented single strains; the remaining nine isolates were unique, and in the case of one isolate, no satisfactory PFGE profile could be obtained. The pattern A isolates were mostly of serotype O12 and were highly resistant to imipenem (MICs, 32 to >256 g/ml), with this resistance decreased eightfold or more in the presence of EDTA. They yielded amplicons with bla VIM -specific primers, and sequencing of DNA from a representative isolate revealed bla VIM-8 , a novel allele with three polymorphisms compared with the sequence of bla VIM-2 . Two of these nucleotide changes were silent, but the third determined a Thr139Ala substitution. Only 4 of 13 resistant isolates (2 clinical isolates and 2 environmental isolates) assigned to other PFGE types carried bla VIM alleles, whereas the others were less multiresistant and mostly had lower levels of imipenem resistance (MICs, <32 g/ml) which was not significantly reduced by EDTA. No bla IMP alleles were detected. During 2003, when the environmental study was undertaken, serotype O12 isolates with bla VIM were recovered from sinks and stethoscopes in the most-affected units, although not from the hands of staff; the problem declined once these reservoirs were disinfected and hygienic precautions were reinforced.

Pyrosequencing was used to detect rapidly and estimate the number of 23S rRNA genes with a G2576T mutation in 43 linezolid-resistant and -susceptible clinical isolates of enterococci. The method showed 100% concordance with PCR-restriction fragment length polymorphism for detecting isolates homozygous for either G2576 or T2576 or heterozygous for this mutation. A good correlation was found between linezolid MICs and the number of 23S rRNA gene copies carrying the mutation.

A strong association was observed between high-level resistance to ciprofloxacin and gentamicin for isolates of Enterococcus faecalis collected in the UK and Ireland as part of the BSAC Bacteraemia Resistance Surveillance Programme, 2001. Thus, 60 of 66 E. faecalis isolates with gentamicin MICs > or =512 mg/L were highly resistant to ciprofloxacin (MICs > or =32 mg/L), compared with only three of 83 E. faecalis isolates with normal gentamicin susceptibility (MICs < or =128 mg/L) (P<0.0001). Pulsed-field gel electrophoresis of SmaI-digested genomic DNA was used to investigate 38 representative E. faecalis isolates with the double resistance from 18 hospitals. Based on the criterion of > or =80% banding pattern similarity, two large clusters were observed: cluster 1 included 14 isolates, from seven hospitals, that were related at 84.8% similarity; cluster 2 included 10 isolates, from six hospitals, that were related at 83.3% similarity. Sporadic isolates and small clusters with the double resistance were also observed, but were not closely related to those in clusters 1 and 2. Further work is needed to characterize these two 'epidemic' E. faecalis strains and to investigate the presence of virulence genes.