He nucleotide sequence data reported within this paper will appear in the GenBank nucleotide database under the accession no. AF010416.VEB–LACTAMASE FROM E. COLITABLE two. MICs of -lactams for E. coli MG-1, E. coli JM109 harboring recombinant plasmids pRLT1 and pRLT50, and reference strain E. coli JMMIC ( g/ml) for: Antibiotic(s) E. coli MG-1a E. coli JM109 (pRLT1)b E. coli JM109 (pRLT50)c E. coli JMRESULTS Origin in the E. coli MG-1 isolate. E. coli MG-1 was isolated in 1996, at the Hopital Antoine Beclere, Clamart (a suburb of ^ ` Paris), France, from the pus of a 4-month-old Vietnamese boy hospitalized for serious respiratory problems. He was previously hospitalized in an intensive care unit in Vietnam. Antimicrobial regimens just before admission were not documented, and the patient didn’t get any antibiotic treatment before the isolation of the strain at the Hopital Antoine Beclere. A ^ ` routine antibiogram revealed high levels of resistance of E. coli MG-1 to amino, carboxy, and ureido-penicillins and to restricted and extended-spectrum cephalosporins (Table two). E. coli MG-1 was also resistant to kanamycin, chloramphenicol, tetracycline, gentamicin, tobramycin, netilmicin, amikacin, trimethoprim, and trimethoprim-sulfamethoxazole. In the course of a systematic multiresistant-bacteria rectal screening, the exact same E. coli MG-1 was isolated in conjunction with a K. pneumoniae MG-2 strain presenting a related resistance profile. Hybridizations and cloning on the ESBL gene. Preliminary hybridization experiments indicated that E. coli MG-1 harbored a TEM-derived resistance gene as indicated by a constructive hybridization signal detected by a blaTEM- probe (data not shown). blaSHV-3, oxa18, and blaPER-1 probes failed to provide constructive hybridization signals. PCR amplification applying TEM-1 intragenic primers and direct sequencing with the PCR product showed 100 identity with blaTEM-1. Considering the fact that TEM-1 will not be an ESBL, its presence could possibly not explain the uncommon resistance phenotype. DNA from E. coli MG-1 was partially digested with restriction endonuclease Sau3AI and ligated to BamHI-digested plasmid pBK-CMV.Hesperetin The ligation product was transformed into E.Mirogabalin coli JM109 by electroporation.PMID:24406011 Numerous recombinant colonies expressing among the following two phenotypes have been obtained: (i) a higher amount of resistance to amoxicillin, cephalothin, and ticarcillin, which was inhibited by clavulanic acid; or (ii) an extended-spectrum phenotype. The recombinant plasmids expressing each -lactamase resistance phenotype had been extracted and analyzed. The insert sizes ranged from 1.2 to 15 kb. Restriction maps had been generated for each plasmids pRLT1 and pRLT50 containing, respectively, a 1.2-kb insert expressing the ESBL in addition to a 1.3-kb insert expressing a penicillinase (pRLT50) (Fig. 1).Amoxicillin Amoxicillin-Clad Ticarcillin Ticarcillin-Cla Piperacillin Piperacillin-Cla Cephalothin Cephalothin-Cla Cefamandole Cefuroxime Cefuroxime-Cla Cefoxitin Cefoxitin-Cla Ceftazidime Ceftazidime-Cla Cefotaxime Cefotaxime-Cla Cefepime Cefepime-Cla Ceftriaxone Moxalactam Moxalactam-Cla Aztreonam Aztreonam-Cla Imipenema b512 128 512 256 256 four 128 4 32 64 4 four four 256 0.25 two 0.06 1 0.03 2 0.25 0.25 32 0.12 0.512 86 512 four 16 2 128 four 32 128 four 8 4 256 0.25 two 0.06 1 0.06 four 0.12 0.12 32 0.12 0.512 256 512 256 512 64 256 four 324 eight four four 2 1 1 0.12 0.06 0.12 0.03 0.12 0.12 0.12 0.25 0.25 0.2 1 two two 1 1 four four 0.five two two 4 two 0.25 0.25 0.06 0.06 0.06 0.06 0.06 0.12 0.25 0.12 0.06 0.E. coli MG-1 produces VEB-1 in conjunction with a TEM-1 pe.