In vitro activity of ceftaroline against Burkholderia pseudomallei

6 derive mainly from wild-type cepha-losporinases by structural alterations in the R2 binding site that accommodates the R2 lateral side chain of b-lactams. Taken together, these findings suggest that acquired resistance to avi-bactam and extension of the hydrolysis spectrum of class C b-lactamases might result from the same structural changes. It prompted us to investigate the inhibitory activity of avibactam against extended-spectrum AmpC b-lactamases containing structural changes in the R2 binding site. Six isogenic Escherichia coli recombinant clones—E. coli TOP10 pEC14, E. coli TOP10 pEC16, E. coli TOP10 pEC17, E. coli TOP10 pEC18, E. coli TOP10 pMEV and E. coli TOP10 pBER—that have been characterized in previous work 6 – 8 were tested in this study. They expressed extended-spectrum AmpC enzymes with representative structural alterations in the R2 binding site, such as the Val-298Leu and His-296Pro replacements, 7 the Ser-287Asn and Ser-287Cys substitutions, 7 the Ser-282 duplication 6 and the tandem duplication of the alanine residues at positions 294 and 295. 8 All these modifications occurred in the R2 loop 7 or the H-9 and H-10 helices, 6 – 8 which are secondary structures surrounding the R2 binding site. 5 The MICs of ceftazidime, cefepime and imipenem were determined using an agar dilution technique on Mueller – Hinton agar (Sanofi Diagnostics Pasteur, Paris, France) plates and Mueller – Hinton agar supplemented with 4 mg/L avibactam with an inocu-lum of 10 4 cfu per spot, and were interpreted according to the guidelines of the CLSI. 9 Results are shown in Table 1. Avibactam restored the susceptibility to ceftazidime and cefepime of all recombinant clones, thus confirming its potency against these variant enzymes. These results are in agreement with the crystallographic study carried out by Lahiri et al., 10 which showed the weak interaction between avibactam and the R2 binding site of the AmpC b-lactamase of Pseudomonas aeruginosa. According to that study, the carboxamide and sulphate groups of avibactam, and the carbonyl oxygen of the acylated compound, did not interact with the R2 binding site of the AmpC enzyme. By contrast, the hydrophobic side chain of the Ala-293 residue, which is located in the R2 loop, interacted with the piperidine ring of the inhibitor. 10 In conclusion, the present study demonstrates that avibactam, which is a novel covalent non-b-lactam b-lactamase inhibitor, remains active against extended-spectrum AmpC b-lactamases, suggesting that this molecule could be used in clinical practice with limited risk of selecting …