Bacteria detection by flow cytometry

Abstract

Since bacterial infection of the recipient has become the most frequent infection risk in transfusion medicine, suitable methods for bacteria detection in blood components are of great interest. Platelet concentrates are currently the focus of attention, as they are stored under temperature conditions, which enable the multiplication of most bacteria species contaminating blood donations. Rapid methods for bacteria detection allow testing immediately before transfusion in a bed-side like manner. This approach would overcome the sampling error observed in early sampling combined with culturing of bacteria and would, at least, prevent the transfusion of highly contaminated blood components leading to acute septic shock or even death of the patient. Flow cytometry has been demonstrated to be a rapid and feasible approach for detection of bacteria in platelet concentrates. The general aim of the current study was to develop protocols for the application of this technique under routine conditions. The effect of improved test reagents on practicability and sensitivity of the method is evaluated. Furthermore, the implementation of fluorescent absolute count beads as an internal standard is demonstrated. A simplified pre-incubation procedure has been undertaken to diminish the detection limit in a pragmatic manner. Additionally, the application of bacteria detection by flow cytometry as a culture method is shown, i.e., transfer of samples from platelet concentrates into a satellite bag, incubation of the latter at 37 degrees C, and measuring the contaminating bacteria in a flow cytometer.

Novel Flow Cytometry–Based Screening for Bacterial Contamination of Donor Platelet Preparations Compared with Other Rapid Screening Methods

“…Not only are NAT assays complex, but their time to result (approximately 4 h) is still relatively long for a rapid method. In contrast, flow cytometry for bacterial testing of PLTs has been demonstrated to be a rapid and feasible approach (17)(18)(19)(20). This method can be used either as a rapid test or, alternatively, in combination with a preincubation procedure (17,19 ).…”

“…Rapid methods for bacterial detection in PLTs should be easy to perform and not time-consuming to ensure the feasibility of performing them just before transfusion (13 ). These requirements can be met for bacterial detection by nucleic acid amplification techniques (NATs) (3,14,15 ), qualitative immunoassay [Pan Genera Detection (PGD)] (13,16 ), and fluorescence-activated cell sorting (FACS) (17)(18)(19)(20).…”

Ten Years of External Quality Control for Cellular Therapy Products in France

“…However, to improve this control, a revision of the 2.6.27 chapter is currently in progress with the development of rapid methods for sterility testing like PCR, flow cytometry or the detection of microcolonies. The Paul Erlich Institute (an Official Medicinal Laboratory Control for biological products) in Germany has already validated flow cytometry for the microbiological control of platelets (Karo et al, 2008) and it is in progress for the control of cell products. -Considering the rare and single character of some products and the benefit/risk, the challenge is to have the best suitable test to be sure to take the decision even if a contamination has been found.…”

Flow cytometry as a valuable tool to study cyanobacteria:A mini-review

“…A PubMed search query using the MeSH term "flow cytometry" returns nearly 180,000 papers published between 1979 and 2016 with over 50% published in only the last 15 years. FCM is also gaining popularity in environmental sciences and is being applied in various phytoplankton and bacterial analyses (Dubelaar and Jonker 2000;Vives-Rego et al 2000;Karo et al 2008;.…”

“…In particular cases an overlap in dye emission spectra and their interference with the autofluorescence of cell components can be estimated (Tashyreva et al 2013). Thus, attention should be paid to the choice of appropriate fluorescent dyes with respect to the particular phytoplankton and microorganism strain being analyzed and the available lasers with which the FCM instrument is equipped (Karo et al 2008).…”