Immuno-flow cytometry for the rapid identification of Staphylococcus aureus and the detection of methicillin resistance

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible S. aureus (MSSA) can be reliably differentiated by flow cytometry when labeled with nucleic acid dyes. The purpose of this study was to determine if this differentiation can be achieved while labeling with a S. aureus-specific anti-staphylococcal protein A antibody instead of nucleic acid dyes. A total of 103 S. aureus isolates were incubated for 4 h at 37°C in Mueller Hinton broth with and without oxacillin, then stained with anti-staphylococcal protein A antibody, and analyzed by flow cytometry using the Micro PRO™ instrument. Dot plots (side scatter vs. fluorescence intensity) of isolates exposed to oxacillin were examined to define two gates encompassing the majority of MSSA and MRSA signal events, respectively. The ratio of signal event counts in the two gates was called the gate signal count ratio (GSCR), and its performance was evaluated using receiver operating characteristic (ROC) curves. The GSCR could differentiate MRSA from MSSA with 98% sensitivity and 100% specificity using a cut-off of 0.6868 when the two gates were defined as follows: gate 1, fluorescence intensity 2-10, side scatter 5-70; gate 2, fluorescence intensity 7-700, side scatter 70-500. MRSA and MSSA can be accurately detected and differentiated by flow cytometry after 4 h of oxacillin exposure when labeled with anti-staphylococcal protein A antibody.

In vitro assessment of the antibacterial effect of watery date extracts (Mozafati, Zahedi and Piaroum) on Staphylococcus aureus and Escherichia coli

“…al 2012) investigated the use of flow cytometry in survivability estimates of S. aureus in combined culture method which were similar to the findings of the present study. Shrestha (et. al 2012) could identify Methicillin-Resistant S. aureus (MRSA) via flow cytometry.…”

A novel flow cytometry assay based on bacteriophage-derived proteins for Staphylococcus detection in blood

“…The cells are often labelled with fluorescent markers, therefore allowing the discrimination of different populations in a sample 42 . In the past years, this technique has been widely used for pathogen detection purposes, although the assays that have been described present some drawbacks, such as being laborious and expensive 43 or lack the ability to detect pathogens in biological samples 44,45 .…”