A New OXA Variant from Achromobacter sp.: a Species Specific Marker

PAPALIA M., ; ALMUZARA M., ; TRAGLIA G., ; RAMÍREZ MS.,; BELMONTE A., ; FAMIGLIETTI A., ; VAY C., ; GUTKIND G., ; RADICE M

San Francisco

Congreso; 52 nd Interscience Conference on Antimicrobial Agents and Chemotherapy; 2012

ASM

Background: Achromobacter spp are opportunistic bacteria that cause infections in patients with underlying disorders and are frequently misidentified by conventional phenotypic methods. In this study phenotype-based techniques and molecular tools were conducted to characterize 5 ceftazidime and/or imipenem resistant isolates recovered from fibrocystic patientsMethods: Phenotypic biochemical approaches included denitrification and carbohydratemetabolic reactions. Total protein profiles were resolved by SDS-PAGE. Genotypic identification was performed by 16S rRNA gene amplification and sequencing. The presence of the A. xylosoxidans species marker blaOXA-114 (Turton et al) was investigated by PCR amplification and sequencing. A RFLP-PCR using ApaI enzyme was designed in order to characterize the amplified fragment. A TAIL-PCR approach was used to identify and sequence a new blaOXA gene Results: Biochemical tests strongly suggested that the isolates correspond to A. ruhlandii. Electrophoretic protein profiles were identical between them but different from those of A. xylosoxidans. The 16S rRNA gene sequence displayed more than 99% similarity to the sequences of the type strains of all species of Achromobacter. Positive PCR amplifications were obtained using the primers described by Turton et al in the 5 isolates. Amplicons had the expected size but differed from blaOXA-114 in 56-58 nucleotides (90% identity, translated in 15 amino acid changes) and displayed 99% identity between them. ApaI digestions were positive for amplicons corresponding to the new variant but not for blaOXA-114 amplicons. The new OXA variant was deposited as EMBL HE614014.1. Neither ESBL (CTX-M, SHV, PER, TEM) nor MBL (IMP, VIM, SPM, GIM, SIM) or KPC coding genes were detectedConclusions: Although A. ruhlandii is considered a soil inhabitant, these isolates were recovered from fibrocystic patient specimens. Here we report a new OXA variant seemingly ubiquitous in one species of Achromobacter. ApaI RFLP-PCR would be a practical tool to discern between this variant and blaOXA-114