New plasmid-mediated aminoglycoside adenylyltransferase of broad substrate range that adenylylates amikacin

Abstract

The same aminoglycoside 2"-adenylyltransferase was isolated from four gram-negative species which were among a random group of gentamicin-resistant isolates from the same hospital. The enzyme was partially purified from a crude extract which also contained a second modifying enzyme identified as APH(3')-I. The substrate range of the new aminoglycoside 2"-adenylyltransferase included the newer aminoglycosides sisomicin and amikacin, but showed much-reduced activity against gentamicins C2 and Cla. The pH optimum was 7.8 to 8.0 for every substrate, and the molecular weight of the enzyme molecule was estimated at approximately 29,000. Genetic experiments clearly established that both enzymes were expressed by a conjugative plasmid.

Characterization of new resistance plasmids belonging to incompatibility group IncQ

“…This is the same type of resistance function as that observed in a number of different conjugative Gm plasmids from bacteria originating from similar sources in Bulgaria . From our results we cannot differentiate between the two ANT(2") determinants identified up to now (COOMBE andGEORGE 1981, LEE et al 1987) and the one present on pIE723. KmNm mediation by pIE639 is due to an APH(3') (5") enzyme function.…”

Construction of a gentamicin resistance gene probe for epidemiological studies

“…The clinical isolate E. coli VA292 and its transconjugant E. coli E292 were resistant to neomycin and were shown by bioassay (7) and the disk method (42) to contain a phosphotransferase. When total adenylyltransferase and phosphotransferase activity in pDGO114 was measured by bioassay, there was no increased inactivation of kanamycin or any inactivation of neomycin, confirming that there was no phosphotransferase activity.…”

“…On the basis of this substrate profile, the enzyme should be classified as AAD(2")-I. However, it did not completely rule out the possibility that the gene coded for AAD(2")-II, since the substrate affinity data for AAD(2")-II was determined with a highly purified enzyme preparation (7). An association of the AAD(2") gene with IncC (as we have found) and IncFII plasmids has been previously reported (11).…”

Decreased susceptibility to 4'-deoxy-6'-N-methylamikacin (BB-K311) conferred by a mutant plasmid in Escherichia coli

“…We have previously shown that a plasmid-borne mutation can increase the activity of aminoglycoside 3'-phosphotransferase II [APH(3')-II] against its aminoglycoside substrates, including minor ones such as amikacin, sufficiently to produce amikacin resistance in Escherichia coli (15). Recently, Coombe and George have reported on the activity of an aminoglycoside 2"-adenylyltransferase against amikacin in clinical isolates of four genera of Enterobacteriaceae (5). In the present study, we examined whether the alterations at Bacterial strais.…”