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This study confirms that Taq DNA polymerase could be contaminated with the blaTEM-1(a) gene. It also proposes two different methods that could be used to overcome DNA contamination: (i) DNase I treatment prior to PCR amplification; and (ii) the use of a highly purified Taq DNA polymerase which was devoid of detectable contamination.

We investigated an outbreak of Acinetobacter baumannii in an intensive care unit and in the surgery, medicine, neurology, and urology wards of the Kosin University Gospel Hospital in Busan, Korea. The outbreak involved 36 cases of infection by A. baumannii producing the OXA-23 beta-lactamase over an 8-month period and was caused by a single pulsed-field gel electrophoresis clone. The epidemic isolates were characterized by a modified cloverleaf synergy test. Isoelectric focusing of crude bacterial extracts detected one nitrocefin-positive band with a pI value of 6.65. PCR amplification and characterization of the amplicons by direct sequencing indicated that the epidemic isolates carried a bla(OXA-23) determinant. The epidemic isolates were characterized by a multidrug resistance phenotype that remained unchanged over the outbreak, including penicillins, cephamycins, extended-spectrum cephalosporins, carbapenems, monobactams, and aminoglycosides. This study shows that the bla(OXA-23) resistance determinant may become an emerging therapeutic problem.

To determine the prevalence and genotypes of beta-lactamases among clones of a metagenomic library from the cold-seep sediments of Edison seamount (10,000 years old), we performed pulse-field gel electrophoresis, antibiotic susceptibility testing, pI determination, and DNA sequencing analysis. Among the 8,823 clones of the library, thirty clones produced beta-lactamases and had high levels of genetic diversity. Consistent with minimum inhibitory concentration patterns, we found that five (16.7%) of thirty clones produced an extended-spectrum beta-lactamase. 837- and 259-bp fragments specific to blaTEM genes were amplified, as determined by banding patterns of PCR amplification with designed primers. TEM-1 was the most prevalent beta-lactamase and conferred resistance to ampicillin, piperacillin, and cephalothin. TEM-116 had a spectrum that was extended to ceftazidime, cefotaxime, and aztreonam. The resistance levels conferred by the pre-antibiotic era alleles of TEM-type beta-lactamases were essentially the same as the resistance levels conferred by the TEM-type alleles which had been isolated from clinically resistant strains of bacteria of the antibiotic era. Our first report on TEM-type beta-lactamases of the pre-antibiotic era indicates that TEM-type beta-lactamases paint a picture in which most of the diversity of the enzymes may not be the result of recent evolution, but that of ancient evolution.

Among the 51 clinical isolates collected from a university hospital in Korea, nine isolates were resistant to cephamycins. Nine isolates were shown to produce CMY-11 and these also included three isolates producing TEM-1. The results from ERIC-PCR revealed that dissemination of CMY-11 was due to outbreaks of resistant species and to the intra-species spread of resistance to cephamycins in Korea. CMY-11 beta-lactamase genes from nine clinical isolates that were responsible for resistance to cephamycins (cefoxitin and cefotetan), amoxicillin, cephalothin and amoxicillin-clavulanic acid, were cloned and characterised. A sequence identical to the common regions in In6, In7 and a novel integron from pSAL-1 was found upstream from bla(CMY-11) gene at nucleotide 1-71. Eighteen nucleotides between position 71 and 72 were inserted into the bla(CMY-11) gene.