The Detection of Transposable ampicillin-resistance in Klebsiella pneumonia


Genetically i stable cliiical straii of K|ebsieqa pnelnania Nal0 was isolated fion respirarofy rractinfection. The isolaLe harboN large singie non-conjugative plasmid p,(1 (120kbp) which encoding multiresistance forantibiotics.Successive subcuhuring of tbe isolate for about 480 generations yielded several plasmid derivarive. Allmaintained ampicillin fesistance. while lhey appealed va able deleted or abolished to the othef antibiotic resistanceOne of these derived plasmkls (,KPr) encoding arnpicillin (Ap') and bimethoprim (Tp) .esistance wastransfomred into E.coli I,/1t4294 which aheady hafborc a non-conjugatlve p1^snid pACyCISl (Cn, rd.'fhehansfornred cells (pKP2 ah.l p,1CYC 184) were subjected to successive subculturing for about 480 genefations inampicilljn medium at 37'C with selecrion for ampiciuin and tetracycline and chloramphenicol. Subsequentlyscrcening lbr plasrnid content rcvealed that the prcseoce (on agarose) ofappfoximately severr bands ofplasnid DNArangifg beiween p,(P? d,d pACyCl3l in |i.olec|Jaf size. Each ofthese bands was eluied ffom the gel, purified aDdre-transfomed inlfo r.coli liM2 94 co]mpelent ce|.|.-The hansformed colonies resisiani io ampicillin, chloramphenicol and tetmcycline were selected. The plasmidconient of these cells revealed the presence of extra size (approximately 5kb) to the pfe transfonned DNA band@'1CyCl8'l) ^s;ndicaled by gel electrophoresis. The added DNA could represent a rmnsposable DNA fiagrnenrc,cod,q IDLi lir "e,5ta ce