Cloning of the class D beta-lactamase gene from Burkholderia pseudomallei and studies on its expression in ceftazidime-susceptible and -resistant strains

Ceftazidime is the antibiotic of choice for the treatment of melioidosis. Ceftazidime-resistant Burkholderia pseudomallei have been identified and beta-lactamase production implicated in resistance. In this study, 25 strains of B. pseudomallei (15 clinical and 10 environmental strains) were examined for their ability to yield mutants that overexpress beta-lactamase. Ceftazidime-resistant mutants were selected readily at high frequency and displayed four- to eight-fold increases in the MICs of ceftazidime. beta-Lactamase activities in both parent and mutant B. pseudomallei strains were examined by a spectrophotometric method. Twelve mutants (48%) showed approximately two- to 31-fold higher ceftazidimase activity compared with their parent strains and 10 (40%) demonstrated more than two-fold increases in imipenemase activity. A class D beta-lactamase gene from B. pseudomallei was cloned and sequenced. The encoded enzyme is an oxacillinase and is homologous to oxacillinases from Ralstonia pickettii and members of the genus Aeromonas. Reverse transcriptase PCR showed that transcription of the class D beta-lactamase gene is increased in ceftazidime-resistant mutants.