Abstract.
The molecular diversity of inhibitor-resistant TEM (IRT) enzymes was explored using a strategy which involved DNA amplification by polymerase chain reaction (PCR), analysis of restriction fragment length polymorphism (RFLP), and direct nucleotide sequencing. The study of plasmid-borne genes from 27 strains, resistant to amoxicillin and β-lactamase-inhibitor combinations, identified mutations resulting in amino acid change at positions 69, 244, 275, and 276 known to be associated with the IRT phenotype and a mutation at nucleotide position 162 in the promoter region. These mutations were found to lie on two different gene sequences, described here as ``TEM-1B like'' and ``TEM-2 like'' restriction linkage groups. Further analysis, of nucleotide sequences of promoter and coding regions of the β-lactamases, confirmed that a given mutation causing IRT phenotype could be associated with two different gene sequence frameworks and two different causal mutations could lie on identical gene sequence framework. These data argue in favor of convergent phenotypic evolution of IRT enzymes under the selective pressure imposed by the intensive clinical use of β-lactam–β-lactamase inhibitor combinations.
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References
Ambler RP (1980) The structure of β-lactamases. Philos Trans R Soc Lond Biol 289:321–331
Ambler RP, Coulson AFW, Frere J-M, Ghuysen J-M, Joris B, Forsman M, Levesque RC, Tiraby G, Waley SG (1991) A standard numbering scheme for the Class A β-lactamases. Biochem J 276:269–272
Belaaouaj A, Lapoumeroulie C, Canna MM, Vedel G, Névot P, Krishnamoorthy R, Paul G (1994) Nucleotide sequences of the genes coding for the TEM-like β-lactamases IRT-1 and IRT-2 (formerly called TRI-1 and TRI-2). FEMS Microbiol Lett 120:75–80
Bémer-Melchior P, Gilly L, Jugroot-Klotz K, Brun T, Névot P, Paul G (1995) Etude de la resistance a l’amoxicilline et a Fassociation amoxicillin-acide clavulanique Bans 231 souches cliniques d’Escherichia coli isolées en 1992 a I’Hôpital Cochin. Pathol Biol (Paris) 43:760–765
Bijvoet SM, Hayden MR (1992) Mismatch PCR: a rapid method to screen for the Pro207 → Leu mutation in the lipoprotein lipase (LPL) gene. Hum Mol Genet 1:541
Blazquez J, Baquero M-R, Canton R, Alos I, Baquero F (1993) Characterization of a new TEM-type β-lactamase resistant to clavulanate, sulbactam, and tazobactam in a clinical isolate of Escherichia coli. Antimicrob Agents Chemother 37:2059–2063
Bonomo RA, Currie-McCumber C, Shlaes D (1992) OHIO-1 β-lacta-mase resistant to mechanism-based inactivators. FEMS Microbiol Lett 92:79–82
Brun T, Peduzzi J, Caniça MM, Paul G, Névot P, Barthélémy M, Labia R (1994) Characterization and amino acid sequence of IRT-4, a novel TEM-type enzyme with a decreased susceptibility to β-lac-tamase inhibitors. FEMS Microbiol Lett 120:111–118
Bush K, Jacoby GA, Medeiros AA (1995) A functional classification scheme for β-lactamases and its correlation with molecular structure. Antimicrob Agents Chemother 39:1211–1233
Canna MM, Ly CY, Brun T, Névot P, Paul G, Krishnamoorthy R (1994) Molecular diversity of genes encoding TEM-type β-lacta-mases resistant to suicide inhibitors (IRT). 14th Interdisciplinary Meeting on Anti-Infectious Chemotherapy, Paris, Abstract 229/C13, p 174
Datta N, Kontomichalou P (1965) Penicillinase synthesis controlled by infectious R factors in Enterobacteriaceae. Nature (London) 208: 239–241
Delaire M, Labia R, Samama J-P, Masson J-M (1992) Site-directed mutagenesis at the active site of Escherichia coli TEM-1 β-lacta-mase. Suicide inhibitor-resistant mutants reveal the role of arginine 244 and methionine 69 in catalysis. J Biol Chem 267:20600–20606.
Goussard S, Courvalin P (1991) Sequence of the genes bla T-1B and bla T-2. Gene 102:71–73
Hawley DK, McClure WR (1983) Compilation and analysis of Escherichia coli promoter DNA sequences. Nucleic Acids Res 11:22372255
Henquell C, Chanal C, Sirot D, Labia R, Sirot J (1995) Molecular characterization of nine different types of mutants among 107 inhibitor-resistant TEM β-lactamases from clinical isolates of Escherichia coli. Antimicrob Agents Chemother 39:427–430
Heritage J, Hawkey PM, Todd N, Lewis IJ (1992) Transposition of the gene encoding a TEM-12 extended-spectrum β-lactamase. Antimicrob Agents Chemother 36:1981–1986
Hibbert-Rogers L, Heritage J, Todd N, Hawkey P (1994) Convergent evolution of TEM-26, a β-lactamase with extended-spectrum activity. J Antimicrob Chemother 33:707–720
Ish-Horowicz D, Burke JF (1981) Rapid and efficient cosmid cloning. Nucleic Acids Res 9:2989–2998
Jacoby GA, Medeiros AA (1991) More extended-spectrum β-lactama-ses. Antimicrob Agents Chemother 35:1697–1704
Jacoby GA, Sutton L (1985) β-lactamases and β-lactam resistance in Escherichia coli. Antimicrob Agents Chemother 28:703–705
Joris B, Ghuysen J-M, Dive G, Renard A, Dideberg O, Charlier P, Frere J-M, Kelly JA, Boyington JC, Moews PC, Knox JR (1988) The active -site-serine penicillin-recognizing enzymes as members of the Streptomyces R61 DD-peptidase family. Biochem J 250:313324
Knox JR (1995) Extended-spectrum and inhibitor-resistant TEM-Type β-lactamases: mutations, specificity, and three-dimensional structure. Antimicrob Agents Chemother 39:2593–2601
Labia R, Barthelemy M, Fabre C, Guionie M, Peduzzi J (1979) Kinetic studies of three R-Factor mediated β-lactamases In: Hamilton-Miller JMT, Smith JT (eds) Beta-lactamases. Academic Press, London, pp 429–442
Lemozy J, Sirot D, Chanal C, Huc C, Labia R, Dabernat H, Sirot J (1995) First characterization of inhibitor-resistant TEM (IRT) β-lactamases in Klebsiella pneumoniae strains. Antimicrob Agents Chemother 33:2580–2582
Liu PYF, Gur D, Hall LMC, Livermore DM (1992) Survey of the prevalence of β-lactamases amongst 1000 Gram-negative bacilli isolated consecutively at the Royal London Hospital. J Antimicrob Chemother 30:429–447
Martinez JL, Vicente MF, Delgado-Iribarren A, Perez-Diaz JC, Baquero F (1989) Small plasmids are involved in amoxicillinclavulanate resistance in Escherichia coli. Antimicrob Agents Chemother 33:595 (Letter)
Matthew M, Hedges RW (1976) Analytical isoelectric focusing of R factor-determined β-lactamases: correlation with plasmid compatibility. J Bacteriol 125:713–718
Reguera JA, Baquero F, Perez-Diaz JC, Martinez JL (1991) Factors determining resistance to β-lactam combined with β-lactamase inhibitors in Escherichia coli. J Antimicrob Chemother 27:569–575
Rychlick W, Rhoads RE (1989) A computer program for choosing optimal oligonucleotides for filter hybridization, sequencing and in vitro amplification of DNA. Nucleic Acids Res 17:8543–8551
Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning. A laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY
Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chainterminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467
Stapleton P, Wu P-J, King A, Shannon K, French G, Phillips I (1995) Incidence and mechanisms of resistance to the combination of amoxicillin and clavulanic acid in Escherichia coli. Antimicrob Agents Chemother 39:2478–2483
Sutcliffe JG (1978) Nucleotide sequence of the ampicillin gene of Escherichia coli plasmid pBR322. Proc Natl Acad Sci USA 75: 3737–3741
Thomson CJ, Amyes SGB (1992) TRC-1: emergence of a clavulanic acid-resistant TEM β-lactamase in a clinical strain. FEMS Microbiol Lett 91:113–118
Vedel G, Belaaouaj A, Gilly L, Labia R, Philippon A, Névot P, Paul G (1992) Clinical isolates of Escherichia coli producing TRI β-lac-tamases: novel TEM-enzymes conferring resistance to β-lactamase inhibitors. J Antimicrob Chemother 30:449–462
Zhou XY, Bordon F, Sirot D, Kitzis M-D, Gutmann L (1994) Emergence of clinical isolates of Escherichia coli producing TEM-1 derivatives or an OXA-1 β-lactamase conferring resistance to β-lactamase inhibitors. Antimicrob Agents Chemother 38:10851089
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Received: 18 March 1996 / Accepted: 15 July 1996
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Caniça, M., Lu, C., Krishnamoorthy, R. et al. Molecular Diversity and Evolution of bla TEM Genes Encoding β-Lactamases Resistant to Clavulanic Acid in Clinical E. coli . J Mol Evol 44, 57–65 (1997). https://doi.org/10.1007/PL00006121
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DOI: https://doi.org/10.1007/PL00006121