Applications and challenges for single-bacteria analysis by flow cytometry

Wu, Lina; Wang, Shuo; Song, Yu; Wang, Xu; Yan, Xiaomei

doi:10.1007/s11426-015-5518-3

Cited by 37 publications

(20 citation statements)

References 109 publications

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“…In addition, non-viable bacteria cannot be identified with this method, since they must first be cultured to obtain material for MALDI-TOF MS. In order to detect these bacteria as well, researchers are looking for alternative methods such as Flow Cytometry ( Gunasekera, Attfield & Veal, 2000 ; Wu, Wang, Song, Wang & Yan, 2016 ) or 16S sequencing of extracted DNA which can identify very low bacterial loads, and non-viable or damaged bacteria as well as live bacteria ( Ben-Amor et al., 2005 ; Kuczynski et al., 2012 ; Xu, 2014 ). The 16S sequencing method identifies all DNA associated with a particular sequence unique to bacteria, and therefore can be used to identify DNA even from non-viable bacteria.…”

Section: Discussionmentioning

confidence: 99%

Microbiota of semen from stallions in Sweden identified by MALDI-TOF

Al-Kass

Eriksson

Bagge

et al. 2020

Veterinary and Animal Science

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Highlights Bacteria isolated from stallion semen vary in number and species found. Differences may depend on culture conditions and method of identification. 64% bacteria in stallion semen were cultured and identified by MALDI-TOF. The number of bacteria varied considerably among samples, even from the same stud. More information is needed on their effect on sperm quality.

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Section: Discussionmentioning

confidence: 99%

Microbiota of semen from stallions in Sweden identified by MALDI-TOF

Al-Kass

Eriksson

Bagge

et al. 2020

Veterinary and Animal Science

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“…the Cytoflex system of Beckman Coulter) have been developed with a very high sensitivity using adopted strategies for single molecule fluorescence detection in sheathed flow. Using label-free side scatter detection, a single bacterial cell can be well discriminated from the instrumentation background and total bacterial counts can be made based on a volumetric control that enable a direct quantification of the bacterial concentration [43,44].…”

Section: Discussionmentioning

confidence: 99%

Flow cytometry, a powerful novel tool to rapidly assess bacterial viability in metal working fluids: Proof-of-principle

et al. 2019

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Metalworking fluids (MWF) are water- or oil-based liquids to cool and lubricate tools, work pieces and machines, inhibit corrosion and remove swarf. One of the major problems in the MWF industry is bacterial growth as bacterial enzymes can cause MWF degradation. In addition, bacteria can form biofilms which hamper the functioning of machines. Last but not least, some bacterial by-products are toxic (e.g. endotoxins) and present potential health risks for metalworking machine operators via the formation of aerosols. Therefore, a novel fast yet accurate analytical method to evaluate and predict the antibacterial capacity of MWF would be an important asset. As such a tool is currently lacking, the present study aimed to develop a protocol based on flow cytometry (FCM) to assess the antibacterial potential of newly developed MWF independent of bacterial growth. Results of this novel method were compared to a biochallenge test currently used in MWF industry and also to traditional plate counts. Our results represent a proof-of-principle that FCM can reliably predict the antibacterial capacity of MWF already within one day of incubation with Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Proteus mirabilis, being substantially faster than the current growth-based methods.

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“…Flow cytometry is a widely used technique to investigate all kind of particles with respect to their shape, granularity and fluorescence. The size of the particles, which can be analyzed, ranges from mammalian cells to much smaller bacteria cells [37,38]. This technology has been successfully used in the eukaryotic and prokaryotic sensors mentioned above and it was our first aim to establish the routine application of flow cytometry for the quantitative readout of our epigenetic memory systems, which previously was mainly based on fluorescence spectroscopy in whole-cell lysates [9].…”

Section: Establishment Of Flow Cytometry Detection For the Arabinose mentioning

confidence: 99%

Development of an epigenetic tetracycline sensor system based on DNA methylation

2020

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Bacterial live cell sensors are potentially powerful tools for the detection of environmental toxins. In this work, we have established and validated a flow cytometry readout for an existing bacterial arabinose sensor system with DNA methylation based memory function (Maier et al., 2017, Nat. Comm., 8:15336). Flow cytometry readout is convenient and enables a multiparameter analysis providing information about single-cell variability, which is beneficial for further development of sensor systems of this type in the future. We then designed a tetracycline sensor system, because of the importance of antibiotics pollution in the light of multi-resistant pathogens. To this end, a tetracycline trigger plasmid was constructed by replacing the araC repressor gene and the ara operator of the arabinose trigger plasmid with the tetR gene coding for the tetracycline repressor and the tet operon. After combination with the memory plasmid, the tetracycline sensor system was shown to be functional in E. coli allowing to detect and memorize the presence of tetracycline. Due to a positive feedback between the trigger and memory systems, the combined whole-cell biosensor showed a very high sensitivity for tetracycline with a detection threshold at 0.1 ng/ml tetracycline, which may be a general property of sensors of this type. Moreover, acute presence of tetracycline and past exposure can be detected by this sensor using the dual readout of two reporter fluorophores.

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Applications and challenges for single-bacteria analysis by flow cytometry

Cited by 37 publications

References 109 publications

Microbiota of semen from stallions in Sweden identified by MALDI-TOF

Microbiota of semen from stallions in Sweden identified by MALDI-TOF

Flow cytometry, a powerful novel tool to rapidly assess bacterial viability in metal working fluids: Proof-of-principle

Development of an epigenetic tetracycline sensor system based on DNA methylation

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