Emergence of a PER-1 Extended-Spectrum β-Lactamase-Producing Acinetobacter baumannii Clinical Isolate in Bulgaria

Abstract

Over the past decade, nosocomial outbreaks of Acinetobacter baumannii have been described with increasing frequency, mostly in intensive care units, burn units or surgical wards 1. This organism tends to be resistant to multiple antibiotics and to cause serious nosocomial infections. The main mechanisms of resistance to broad-spectrum β-lactams in A. baumannii are overexpression of chromosomal cephalosporinases, together with decreased outer membrane permeability 2. β-Lactamase production is an additional mechanism of β-lactam resistance. PER-1 is an extended-spectrum β-lactamase (ESBL) from Ambler class A, which was first detected in a Pseudomonas aeruginosa strain in France 3 and was then subsequently established in A. baumannii isolates from Turkey, France, Korea, Belgium and Romania 4-8. PER-1 production has been found to be an independent indicator of poor prognosis 9, but its detection by the double-disk synergy (DDS) test is difficult 5. In the course of routinely assessing all ceftazidimeresistant Acinetobacter spp. isolates by the DDS test, we identified a strain of A. baumannii (Aba 255-00) exhibiting a slight synergy between the ceftazidime (CAZ) disk and the amoxicillin/clavulanic acid (AMX) disk. The presence of an ESBL was suggested by this synergy image and its type was determined by polymerase chain reaction (PCR) amplification and sequencing. Aba 255-00 strain was isolated on February 11, 2000 from the wound of a male patient aged 65 years at a Surgery Clinic in Sofia, Bulgaria. Identification was performed by conventional tests and confirmed using the BBL Enteric/Nonfermenter ID System (Becton Dickinson, USA). Antimicrobial susceptibilities were determined by the disk diffusion and the agar dilution methods on Mueller–Hinton (MH) agar plates (Becton Dickinson, USA) according to the Clinical and Laboratory Standards Institute (CLSI)-2005 recommendations 10. Control strains included P. aeruginosa ATCC 27853 and E. coli ATCC 25922. The presence of an ESBL was investigated by the DDS test carried out with a CAZ disk (30 μg) placed next to an AMX (20/10 μg)containing disk at a distance of 20 mm (center to center) on a cloxacillin (200 mg/l)-containing MH agar plate, which partially inhibits the cephalosporinase activity 11. Aba 255-00 was resistant to all of the β-lactams tested, with the exception of imipenem and meropenem (Table 1). In the presence of clavulanic acid at a fixed concentration of 2 μg/ml, the minimum inhibitory concentration of CAZ was reduced from 512 μg/ml to 128 μg/ml. The studied strain of A. baumannii was also resistant to amikacin and gentamicin, but it was susceptible to ciprofloxacin.