M. Kuswandi


Isolation of Klebsiella pneumoniae resistant mutants against BRL 41897A antibiotics using TnphoA has yielded 30 mutants which have been characterized. In order to identify whether the mutants are as a result of the insertion (transposition) of TnphoA, and to count how many copies of TnphoA are inserted in the each mutant chromosome, several chromosomal DNA of the mutants have been cut with restriction enzymes and analised using Southern blot method and DIG conjugation. The results showed that at least one mutant (KSL19) carried one copy of transposon, whereas at least 2 mutants (KSL38 and KSL52, ) have 2 copies. It is obvious that TnphoA mutagenesis provides an efficient method of selecting mutants which are defective in genes controlling expression of the outer membrane and the periplasmic membrane proteins.

Key words: Resistant K. pneumoniae mutants- TnphoA mutagenesis – BRL 41897A- Southern Blot.

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Basker , M.J., C.L. Branch, S.C. Finch, A.W. Guest, P.H. Milner, M.J. Pearson, R.J. Ponsford and T.C. Smak. 1986. Studies on Semi-synthetic 7–Formamido-cephalosporins. J.Antibiotics. 39(12):1788-1791.

Basker, M.J., R.A. Edmonson, SJ. Knott, RJ. Ponsford, B. Slocombe and SS. White. 1984. In vitro Antibacterial Properties of BRL36650, a novel 6–Substituted Penicillin. Antimicrob. Ag. Chemother., 26:734-740.

Critchley,I.A., M.J.Baker, R.A.Edmondson and S.J.Knott.1991. Uptake of a Catecholic Cephalosporin by the Iron Transport System of E.coli. J.Antimicrob.Chemother., 28:377-388.

Curtis, N.A. C., R.L. Eisenstandt, S.J. East, R.J. Conford, L.A. Walker and A.J. White. 1988. Iron Regulated Outer Membrane Proteins of E. coli K-12 and Mechanism of Action of Catechol-substitutecephalosporins. Antimicrob. Ag. Chemother., 32:1879-1886.

Hill, H.R., C.E., Hunt and J.M. Matsen. 1974. Nosocomial Colonisation with Klebsiella type 26 in Neonatal Intensive-care Unit Associated with an Outbreak of Sepsis, Meningitis and Necrotizing Endocolitis. J. Pediatrics, 85:415-419.

Jacoby. 1996. In vivo Selection of Porin-deficient Mutants of Klebsiella pneumoniae with Increased Resistance to Cefoxitin and Expanded-spectrum Cephalosporin. Antimicrob. Ag. Chemother., 40(2):342-347.

Kuswandi, M dan Lambert, P.L. 1997. Mekanisme Uptake dari BRL 41897A, Suatu Derivat Sefalosporin yang Mengandung Katekol oleh Klebsiella pneumoniae., Majalah Farmasi Indonesia 8(1).

Kuswandi, M. 1997. Isolation of Klebsiella Pneumoniae Mutants Resistant to the Catecholic Cephalosporin BRL 41897. Proceeding Indonesian Biotechnology Conference, Jakarta.

Manoil, C. and J. Beckwith. 1985. TnphoA: A Transposon Probe for Protein Export Signals. Proc.Natl.Acad.Sci. USA. 82:8129-8133.

Martinez-martinez, L., S. Hernandez-Alex, S. Alberti, J.M. Tomas, V.J. Benedi and G.A.

McGowan,J.E.(1985). Changing Etiology of Nosocomial Bacteremia and Fungemia and other Hospital Required Infections. Rev.Inf.Dis., 7(suppl) :s357-s370.

Nikaido, H. and E.Y. Rosenberg. 1990. Cir and Fiu Proteins in the Outer Membrane of E. coli Catlyse Transport of Monomeric Catechols: Study with -lactam Antibiotics Containing Catechol and Analogous Groups. J. Bacteriol., 172:1361-1367.

Pornull,K.J., E. Goranson, A.S. Rytting and K. Dornbusch. 1993. ESBL in E. coli and Klebsiella spp. in European Septisemia Isolates. J. Antimicrob. Chemother., 32:559-570.

Reish, O., S. Ashkenazi, N. Naor, Z. Samra and P. Merlob. 1993. An Outbreak of Multiresistant Klebsiella in a Neonatal Intensive-care unit. J. Hosp. Infect., 25:287-291.

Sambrook,J.T. and E.F.Fritsch.1989. Molecular Cloning: a Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York.

Southern, E.M. 1975. Detection of Specific Sequences Among DNA Fragments Separated by Gel Electrophoresis. J. Mol. Biol., 98:503-517.

Taylor, R.K., C. Manoil and J.J. Mikalinos. 1989. Broad-Host range Vectors for delivery of TnphoA: Use in Genetic Analysis of Secreted Determinats of Vibrio cholerae. J. Bacteriol., 171:1870-1878.

Ullman,U. 1978. The Patterns of Multiple Resistance in Enterobacteriaceae of Hospitalized Patients. Infection, 6:S219-S223.

Watanabe, N., T. Nagasu, K.Katsu, and K. Kitoh. 1987. E 0702, A New Cephalosporin, is Incorporated Into E. coli Cells via TonB Dependent Iron transport system. Antimicrob. Ag. Chemother., 31:497-504.

Williams, P. 1983. The Role of the O and K Antigens in Determining the Resistance of K. aerogenes to Serum Killing and Phagocytosis. J. Ge. Microbiol., 129, 2181-2191.

Young, S.E.J. 1982. Bacteremia: A Survey of Cases Reported to the PHLS Communicable Disease Surveillance Centre. J. Infection, 5, 19-26



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