Applications of Flow Cytometry to Clinical Microbiology

Abstract

SUMMARY Classical microbiology techniques are relatively slow in comparison to other analytical techniques, in many cases due to the need to culture the microorganisms. Furthermore, classical approaches are difficult with unculturable microorganisms. More recently, the emergence of molecular biology techniques, particularly those on antibodies and nucleic acid probes combined with amplification techniques, has provided speediness and specificity to microbiological diagnosis. Flow cytometry (FCM) allows single- or multiple-microbe detection in clinical samples in an easy, reliable, and fast way. Microbes can be identified on the basis of their peculiar cytometric parameters or by means of certain fluorochromes that can be used either independently or bound to specific antibodies or oligonucleotides. FCM has permitted the development of quantitative procedures to assess antimicrobial susceptibility and drug cytotoxicity in a rapid, accurate, and highly reproducible way. Furthermore, this technique allows the monitoring of in vitro antimicrobial activity and of antimicrobial treatments ex vivo. The most outstanding contribution of FCM is the possibility of detecting the presence of heterogeneous populations with different responses to antimicrobial treatments. Despite these advantages, the application of FCM in clinical microbiology is not yet widespread, probably due to the lack of access to flow cytometers or the lack of knowledge about the potential of this technique. One of the goals of this review is to attempt to mitigate this latter circumstance. We are convinced that in the near future, the availability of commercial kits should increase the use of this technique in the clinical microbiology laboratory.

Feasibility of flow cytometry for the detection of bacteria from body fluid samples

“…4 We evaluated the feasibility of using the BF-FCM to detect bacteria in body fluid samples. Until now, only few reports have documented bacterial detection using FCM.…”

“…FCM has already been used in clinical microbiology for urinalysis, analysis of cerebrospinal fluid (CSF), and bile analysis, and is able to provide accurate and precise quantification of microscopic elements in urine and bile samples. [4][5][6][7][8][9] However, in CSF samples, FCM bacterial counts have been unreliable in hemorrhagic samples and in samples collected by ventricular drainage, where interference by platelets and cells debris was observed. 7,8 In the present study, we evaluated the feasibility of using an FCM-based experimental automated bacteria counter (BF-FCM; Sysmex, Kobe, Japan) to quickly detect bacteria in body fluid samples.…”

Application of flow cytometry to industrial microbial bioprocesses

“…Over the last three decades, due to its versatility, and combined with automated sample handling for an increased sample throughput, flow cytometry (FC) technique has impacted biotechnology significantly [47,48]. Concurrent with the development of fluorescent dyes technology, this technique has evolved as one of the most powerful and versatile tools available to study microbial suspensions, giving rise to an increasing number of biotechnological applications, books and reviews [1,[49][50][51][52][53][54][55]. Some current FC applications in biotechnology range from cell count and identification to its use in bioprocess control and prediction for the development of more accurate kinetic models.…”

Flow cytometry applications in the food industry

“…It can be used for food samples, but requires prior isolation of the target organism to generate antibodies. The use of speciWc antibodies directed against pathogens such as certain strains of Escherichia coli, Salmonella, Pseudomonas and other cell types allowed their cytometric detection with sensitivity and speciWcity [1]. Magnetic separation can also be used to make the analysis faster and more speciWc, and to overcome the presence of noncellular particles [85].…”