Identification of species in the Acinetobacter calcoaceticusA. baumannii complex isolated from blood cultures by an oligonucleotide assay
Abstract number: P1987
Chang T-C., Su S-C., Lee N-Y., Dijkshoorn L., Ko W-C.
Objectives:Acinetobacter calcoaceticus (genomic species 1), A. baumannii (genomic species 2), and unnamed genomic species 3 and 13TU are phenotypically similar and were referred to as a group, the A. calcoaceticusA. baumannii (ACB) complex. Members of the complex are the most common acinetobacters among clinical isolates. The aim of this study was to construct an oligonucleotide array with probes based on the ITS sequence to differentiate different species in the ACB complex and to determine the species distribution of 291 blood isolates of the complex.
Methods: Four oligonucleotide probes (18- to 30-mers), based on the intergenic spacer (ITS) sequences, were designed to identify the four genomic species in the ACB complex. An Acinetobacter-specific and an ACB complex-specific probe were also designed. In addition, a positive control probe was designed from a highly conserved region in the 16S rRNA gene. The array (4 by 4 mm) contained 12 dots (4 by 3 dots), including one Acinetobacter-specific probe, one ACB complex-specific probe, four probes for identification of each of the four genomic species in the ACB complex, one positive control probe, two negative control dots (tracking dye only), and three dots of the position marker probe. One pair of bacteria-specific universal primers 2F and 10R was used to amplify the ITS regions, with each of the two primers labeled with a digoxigenin molecule at its 5' end. The amplified PCR products were hybridised with the arrays at 50°C for 90 min. After hybridisation, alkaline phosphatase-conjugated anti-digoxigenin antibodies were used to reveal the hybridisation signals.
Results: After testing a collection of 52 strains of the ACB complex and 137 strains not belonging to the complex, both the identification sensitivity and specificity of the array were 100%. By using the array, the species distribution of 291 bacteremic isolates of the ACB complex were determined to be A. baumannii (221 strains, 75.9%), genomic species 3 (67 strains, 23.0%), genomic species 13 TU (2 strains, 0.7%), and unidentified Acinetobacter sp. (1 strain, 0.3%).
Conclusion: In conclusion, clinical isolates of the ACB complex can be quickly and accurately identified to species level by the current array. Acinetobacter baumannii (75.9%) was the predominant species among monomicrobial bacteraemia caused by the complex. The current method provides a rapid and accurate way to species identification of the ACB complex.
|Session name:||18th European Congress of Clinical Microbiology and Infectious Diseases|
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