CONICET | Buscador de Institutos y Recursos Humanos

CMY-16 is a variant of the CMY/LAT AmpC -lactamase lineage differing from CMY-4 and CMY-12 in a single aminoacid substitution, and from CMY-2 in two. This marker has been described in P. mirabilis in Italy since 2006, and its emergence has been also reported in 2012 in Providencia stuartii in Tunisia. In this study we report the emergence of CMY-16 in P. mirabilis in Argentina, its kinetic parameters and the genetic context of its coding gene Methods: P. mirabilis 19 was isolated from an inpatient in Buenos Aires in 2012. Antimicrobial susceptibility was Determined according to CLSI. Phenotypic screening for AmpC--lactamases was performed using phenyl boronic acid (PBA) disks (300 μg). Specific primers were used to achieve the complete blaCMY amplicon. The genetic context of blaCMY-16 was analyzed by PCR mapping and sequencing. Conjugation assays were carried out using an E. coli J53 and E. coli HB101 as recipient strains. blaCMY-16 gene was cloned in pLB-II and transformed in E. coli DH5. CMY-16 was purified by ion exchange chromatography and the hydrolytic activity was characterized by standard kinetic measurements. Results: P. mirabilis 19 was resistant to ampicillin, cephalothin, cefoxitin, cefotaxime, ceftazidime, susceptible to amikacin and gentamicin and displayed reduced susceptibility to cefepime and imipenem. Synergy effect observedbetween the disk containing PBA and both oxyiminocephalosporins suggested the presence of AmpC -lactamases. The nucleotide sequence of the 1140 bp amplicon obtained using CMY specific primers corresponded to blaCMY-16. In good agreement with previously reported flanking regions, ISEcp1 was located upstream of blaCMY-16, and sugE and blc were identified downstream. The resistance marker could not be transferred by conjugation in the assayedconditions. CMY-16 hydrolyzes preferentially first generation cephalosporins. Non enzymatic resistance mechanisms may be responsible for the decreased susceptibility to cefepime and imipenem observed in P. mirabilis 19 Conclusions: Among AmpC -lactamases, CMY-2 and, to a lesser extend, DHA-1 constitute the most prevalent enzymes in Argentina; CMY-16 has not been previously reported. CMY-16 exhibit the typical properties attributed to broad spectrum CMY -lactamases