Genetic variability among ampC genes from acinetobacter genomic species 3.
As a part of a nationwide study in Spain, 15 clinical isolates of Acinetobacter genomic species 3 (AG3) were analyzed. The main objective of the study was to characterize the ampC genes from these isolates and to determine their involvement in beta-lactam resistance in AG3. The 15 AG3 isolates showed different profiles of resistance to ampicillin (range of MICs, 12 to >256 microg/ml). Nucleotide sequencing of the 15 ampC genes yielded 12 new AmpC enzymes (ADC-12 to ADC-23). The 12 AG3 enzymes showed 93.7 to 96.1% amino acid identity with respect to the AmpC enzyme from Acinetobacter baumannii (ADC-1 enzyme). Eight out of fifteen ampC genes were expressed in Escherichia coli cells under the control of a common promoter, and with the exception of one isolate (isolate 65, which showed lower beta-lactam MICs), significant differences in overall beta-lactam MICs for E. coli cells expressing AG3 ampC genes were not revealed. No significant differences in ampC gene expression in AG3 clinical isolates were revealed by reverse transcription-PCR analysis. A detailed analysis of the 12 AmpC protein sequences revealed that amino acid replacements (in comparison with those of ADC-1) occurred mainly in the same positions, although none were located in important functional domains such as the Omega- loop or conserved beta-lactamase motifs. Kinetic experiments performed with three representative AmpC enzymes (ADC-14, -16, and -18) in some cases revealed dramatic changes in K(m) and k(cat) values for beta-lactams. No ISAba1 was detected upstream of the ampC genes. Our results reveal 12 new ampC genes in AG3. The enzymes showed a moderate degree of variability, and they are tentatively named ADC-12 to ADC-23.
Beceiro
Alejandro
A
Servicio de Microbiología, Complejo Hospitalario Universitario Juan Canalejo-INIBIC, La Coruña, Spain.
Pérez
Astrid
A
Fernández-Cuenca
Felipe
F
Martínez-Martínez
Luis
L
Pascual
Alvaro
A
Vila
Jordi
J
Rodríguez-Baño
Jesús
J
Cisneros
Jose Miguel
JM
Pachón
Jerónimo
J
Bou
Germán
G
D000150:Acinetobacter / Q000145:classification / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D040981:Acinetobacter baumannii / Q000201:enzymology; D000595:Amino Acid Sequence; D000668:Ampicillin Resistance / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D014644:Genetic Variation; D006801:Humans; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D013030:Spain; D018440:beta-Lactam Resistance / Q000235:genetics
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins