SUMMARY: Resistance of strain HB251 to the newer β-lactam antibiotics, in particular ceftazidime and aztreonam, results from production of the extended-spectrum β-lactamase TEM-6. The corresponding structural gene, , and its promoter region were amplified by the polymerase chain reaction. Analysis of the sequence of the amplification product showed that differed by two nucleotide substitutions from , the gene encoding TEM-1 penicillinase in plasmid pBR322. The mutations led to the substitution of a lysine for a glutamic acid at position 102 and of a histidine for an arginine at position 162 of the unprocessed TEM-1 protein. The presence of a 116 bp DNA insert upstream from resulted in the creation of hybrid promoter P in which the —10 region was that of TEM-1 promoter P whereas the — 35 canonical sequence TTGACA was provided by the right end of the insert. P was found to be 10 times more active than P and to confer higher levels of antibiotic resistance upon the host. Analysis of the sequence of the insert indicated that the 116 bp fragment is related to insertion sequence IS but differs from it by three internal deletions that removed regions encoding the transposase. The distribution of the IS-like element in clinical isolates of was studied by the polymerase chain reaction and by DNA-DNA hybridization. The element appeared to be widespread and was detected in strains producing TEM-6 or other TEM variants.


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