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

Saito, Takashi; Iinuma, Yoshitsugu; Takakura, Shunji; Fujihara, Noború; Inoue, Junya; Hamaguchi, Yukio; Ichiyama, Satoshi

doi:10.1007/s10156-005-0399-6

Cited by 15 publications

(5 citation statements)

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“…evaluated an experimental bacterial counter based on flow cytometry on different biological fluids. Applying a cut-off between 40–100 bacteria/μL according to the type of sample a 84.4% sensitivity and 86.0% specificity was obtained to predict culture positivity [21].…”

Section: Discussionmentioning

confidence: 99%

Evaluation of flow cytometry for the detection of bacteria in biological fluids

et al. 2019

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Objectives Conventional microbiological procedures for the isolation of bacteria from biological fluids consist of culture on solid media and enrichment broth. However, these methods can delay the microbiological identification for up to 4 days. The aim of this study was to evaluate the analytical performance of Sysmex UF500i (Sysmex, Kobe, Japan) as a screening method for the detection of bacteria in different biological fluids in comparison with direct Gram staining and the conventional culture on solid media and enrichment broth. Methods A total of 479 biological fluid samples were included in the study (180 ascitic, 131 amniotic, 56 synovial, 40 cerebrospinal, 36 pleural, 24 peritoneal, 9 bile and 3 pericardial fluids). All samples were processed by conventional culture methods and analyzed by flow cytometry. Direct Gram staining was performed in 339 samples. The amount of growth on culture was recorded for positive samples. Results Bacterial and white blood cell count by flow cytometry was significantly higher among culture positive samples and samples with a positive direct Gram stain compared to culture negative samples. Bacterial count directly correlated with the amount of growth on culture (Kruskall-Wallis H χ 2 (3) = 11.577, p = 0.009). The best specificity (95%) for bacterial count to predict culture positivity was achieved applying a cut-off value of 240 bacteria/μL. Conclusions Bacterial and white blood cell counts obtained with flow cytometry correlate with culture results in biological fluids. Bacterial count can be used as a complementary method along with the direct Gram stain to promptly detect positive samples and perform other diagnostic techniques in order to accelerate the bacterial detection and identification.

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Section: Discussionmentioning

confidence: 99%

Evaluation of flow cytometry for the detection of bacteria in biological fluids

et al. 2019

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“…In previous studies, the flow cytometry was compared with traditional techniques to validate this method as an alternative quantitative assay for cell viability assessment (Bunthof et al 1999;Malacrin o et al 2001;Holm et al 2004;Phe et al 2005;Saito et al 2005;Flint et al 2006;Pils et al 2006;Quiros et al 2007;Smigic et al 2009). All research works validated the flow cytometry against plate counts mainly to enumerate, assess and evaluate both bacteria and yeasts vitality, their optimal growth conditions and their response to different treatments.…”

Section: Discussionmentioning

confidence: 99%

Stress response assessment of Lactobacillus sakei strains selected as potential autochthonous starter cultures by flow cytometry and nucleic acid double-staining analyses

et al. 2013

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Aims: The aim of this study was to apply the flow cytometry to Lactobacillus sakei strains, selected as potential autochthonous starters, to investigate dynamics and physiological heterogeneity of microbial behaviour under different stress conditions. Methods and Results: A simultaneous nucleic acid double-staining assay was applied to discriminate cell populations in different physiological states after exposure to heat (50 and 55°C) and acid (pH 2Á5 and 3Á0) stresses. Alive cells with intact membranes, damaged cells still alive but with injured membranes, so with even a recovery ability, and dead cells with a permanent membrane damage were differentiated with a significant increase in damaged cells after stronger stress treatments. Conclusions: The existence and characteristics of subpopulations displaying heterogeneity in particular conditions are highly relevant, because specific subpopulations may show improved survival, changes and dynamics under stress conditions. Significance and Impact of Study: This assay has potential for physiological research on lactic acid bacteria and for application in the food industry. The assessment of intermediate physiological states in Lb. sakei strains with recovery possibility could be an important criterion for application of potential starter cultures. Application of flow cytometry and characterization of sorted subpopulations may contribute to further understanding of diversity and heterogeneity in physiology of bacterial populations.

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“…The second was the use of the automated urine particle analyzer UF-1000i as a flow cytometer for bacterial cell counting. The UF-1000i can count the numbers of bacteria in a urine specimen in 1 min with its separate flow channel in which the nucleic acids of the bacteria are stained with a specific fluorescent dye and discriminated from cell debris [13]. The results were shown in the main window of its analytical software and the operator can get the numbers of bacteria and the pattern of the bacterial counting channel at a glance (Figure S3).…”

Section: Discussionmentioning

confidence: 99%

“…In recent years, automated urine particle analyzers have been established as alternative screening methods for urine [13], [14]. The automated urine particle analyzer UF-1000i (Sysmex Corporation, Hyogo, Japan), which incorporates flow cytometry (FCM), has been proposed for bacterial detection at clinically relevant levels [15], [16].…”

Section: Introductionmentioning

confidence: 99%

Rapid Discrimination of Gram-Positive and Gram-Negative Bacteria in Liquid Samples by Using NaOH-Sodium Dodecyl Sulfate Solution and Flow Cytometry

Wada

Kono²,

Kawauchi³

et al. 2012

PLoS ONE

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BackgroundFor precise diagnosis of urinary tract infections (UTI), and selection of the appropriate prescriptions for their treatment, we explored a simple and rapid method of discriminating gram-positive and gram-negative bacteria in liquid samples.Methodology/Principal FindingsWe employed the NaOH-sodium dodecyl sulfate (SDS) solution conventionally used for plasmid extraction from Escherichia coli and the automated urine particle analyzer UF-1000i (Sysmex Corporation) for our novel method. The NaOH-SDS solution was used to determine differences in the cell wall structures between gram-positive and gram-negative bacteria, since the tolerance to such chemicals reflects the thickness and structural differences of bacterial cell walls. The UF-1000i instrument was used as a quantitative bacterial counter. We found that gram-negative bacteria, including E. coli, in liquid culture could easily be lysed by direct addition of equal volumes of NaOH-SDS solution. In contrast, Enterococcus faecalis, which is a gram-positive bacterium, could not be completely lysed by the solution. We then optimized the reaction time of the NaOH-SDS treatment at room temperature by using 3 gram-positive and 4 gram-negative bacterial strains and determined that the optimum reaction time was 5 min. Finally, in order to evaluate the generalizability of this method, we treated 8 gram-positive strains and 8 gram-negative strains, or 4 gram-positive and 4 gram-negative strains incubated in voluntary urine from healthy volunteers in the same way and demonstrated that all the gram-positive bacteria were discriminated quantitatively from gram negative bacteria using this method.Conclusions/SignificanceUsing our new method, we could easily discriminate gram-positive and gram-negative bacteria in liquid culture media within 10 min. This simple and rapid method may be useful for determining the treatment course of patients with UTIs, especially for those without a prior history of UTIs. The method may be easily applied in order to obtain additional information for clinical prescriptions from bacteriuria.

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

Cited by 15 publications

References 13 publications

Evaluation of flow cytometry for the detection of bacteria in biological fluids

Evaluation of flow cytometry for the detection of bacteria in biological fluids

Stress response assessment of Lactobacillus sakei strains selected as potential autochthonous starter cultures by flow cytometry and nucleic acid double-staining analyses

Rapid Discrimination of Gram-Positive and Gram-Negative Bacteria in Liquid Samples by Using NaOH-Sodium Dodecyl Sulfate Solution and Flow Cytometry

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