Journal of Bacteriology & Parasitology

Abstract

Detection, Identification and Susceptibility Testing of Bacteria by Flow Cytometry

Sabine Nuding and Lutz T Zabel

Flow cytometry permits the simultaneous analysis of numerous parameters of eukaryotic cells as well as microorganisms making it a powerful tool for differentiation and functional analysis. However, application of flow cytometry in clinical microbiology is still infrequent. Here, we report on the development of flow cytometric assays to detect and identify bacteria, and determine antibiotic susceptibility. Susceptibility testing of a wide spectrum of bacterial culture isolates was based on the oxonol DiBAC4(3), and further modified to allow susceptibility testing using specimens such as blood cultures or swabs. Additionally an extended assay was developed for the rapid detection of methicillin
resistant Staphylococcus aureus (MRSA) within 6 hours from swabs. The results of 402 flow cytometric susceptibility tests were confirmed by Bauer-Kirby agar disk diffusion or automated systems with an average agreement of 94%. This assay brought essential time saving of one day particularly concerning extended beta lactamase producing bacteria (ESBL). Susceptibility testing performed directly from positive blood cultures or from swabs increased the rapidness furthermore. Susceptibility results of bacterial strains tested directly from blood culture achieved an accordance of 89% compared to Vitek® 2. Investigating 140 screening swabs of patients colonized with MRSA and of their contact persons
22 swabs were identified correctly to contain MRSA, 2 MRSA samples were not identified due to a lack of protein A expression of the MRSA-strains. False classifications as MRSA-positive swabs did not occur. Flow cytometry offers a suitable technique for the rapid detection, identification and susceptibility testing of bacteria with great potential.