Comparison between Flow Cytometry and Traditional Culture Methods for Efficacy Assessment of Six Disinfectant Agents against Nosocomial Bacterial Species

Massicotte, Richard; Mafu, Akier Assanta; Ahmad, Darakhshan; Deshaies, Francis; Pichette, Gilbert; Belhumeur, Pierre

doi:10.3389/fmicb.2017.00112

Cited by 21 publications

(13 citation statements)

References 17 publications

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“…Indeed, although it might be difficult, if not impossible, to directly correlate both approaches, each one leads to the conclusion that revivifiable members were least affected in the new diluents. That flow cytometry can advantageously replace traditional microbiological methods was proved in more simple systems 31,32 . Since viability in the defrosted, raw and purified transplants was nearly as high or even indistinguishable from that in the fresh preparations, we can reasonably propose the new diluents as an alternative solution for viable and ready-to-use transplants that circumvents the use of glycerol.…”

Section: Discussionmentioning

confidence: 99%

A Guide for Ex Vivo Handling and Storage of Stool Samples Intended for Fecal Microbiota Transplantation

Burz

Abraham

Fonseca

et al. 2019

Sci Rep

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Owing to the growing recognition of the gut microbiota as a main partner of human health, we are expecting that the number of indications for fecal microbiota transplantation (FMT) will increase. Thus, there is an urgent need for standardization of the entire process of fecal transplant production. This study provides a complete standardized procedure to prepare and store live and ready-to-use transplants that meet the standard requirements of good practices to applied use in pharmaceutical industry. We show that, if time before transformation to transplants would exceed 24 hours, fresh samples should not be exposed to temperatures above 20 °C, and refrigeration at 4 °C can be a safe solution. Oxygen-free atmosphere was not necessary and simply removing air above collected samples was sufficient to preserve viability. Transplants prepared in maltodextrin-trehalose solutions, stored in a -80 °C standard freezer and then rapidly thawed at 37 °C, retained the best revivification potential as proven by 16S rRNA profiles, metabolomic fingerprints, and flow cytometry assays over a 3-month observation period. Maltodextrin-trehalose containing cryoprotectants were also efficient in preserving viability of lyophilized transplants, either in their crude or purified form, an option that can be attractive for fecal transplant biobanking and oral formulation.

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Section: Discussionmentioning

confidence: 99%

A Guide for Ex Vivo Handling and Storage of Stool Samples Intended for Fecal Microbiota Transplantation

Burz

Abraham

Fonseca

et al. 2019

Sci Rep

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“…However, there is no documentation on the use of PTB to control post-harvest diseases caused by bacteria. Flow cytometry is a rapid sensitive method for quantifying of live and dead cells in suspension ( O’Brien-Simpson et al, 2016 ; Massicotte et al, 2017 ). Propidium monoazide (PMA) is high-affinity photoreactive DNA binding dye that has recently been used to distinguish intact from membrane- damage of bacterial cells and other microorganisms.…”

Section: Introductionmentioning

confidence: 99%

Antibacterial Effect of Potassium Tetraborate Tetrahydrate against Soft Rot Disease Agent Pectobacterium carotovorum in Tomato

Ahmed

Arif

2017

Front. Microbiol.

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Soft rot caused by Pectobacterium carotovorum is one of most common bacterial diseases occurring in fruits and vegetables worldwide, yet consumer-acceptable options for post-harvest disease management are still insufficient. We evaluated the effect of potassium tetraborate tetrahydrate (B4K2O7.4H2O) (PTB) on the growth of P. carotovorum using strain BA17 as a representative of high virulence. Complete inhibition of bacterial growth was achieved by treatment with PTB at 100 mM both at pH 9.2 and after adjustment to pH 7.0. Bactericidal activity was quantified and validated by counting fluorescently labeled live and dead bacterial cells using flow cytometry, and reconfirmed using qPCR with high-affinity photoreactive DNA binding dye propidium monoazide (PMA). The results of flow cytometry, qPCR, and culturing confirmed that bacterial cells were killed following exposure to PTB at 100 mM. Bacterial cell membranes were damaged following a 5-min treatment and extrusion of cytoplasmic material from bacterial cells was observed using electronic transmission microscopy. Soft rot incidence on inoculated tomato fruit was significantly reduced by dipping infected fruits in PTB at 100 mM for 5 min and no lesions developed following a 10-min treatment. PTB does not pose a hazard to human health and is an effective alternative to other bactericides and antibiotics for controlling soft rot disease of tomato caused by P. carotovorum.

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“…Furthermore, this difference points out the presence of stressed subpopulations, which are not able to grow on agar plates [16]. In turn, Massicotte et al [26] explained this discrepancy, mainly, with the chemical interaction/interference of hypochlorite ions and cell components. According to these authors, chlorine can react with nucleic acids causing a reduction of fluorescence.…”

Section: Resultsmentioning

confidence: 99%

Evaluation by Flow Cytometry of Escherichia coli Viability in Lettuce after Disinfection

et al. 2019

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Foodborne outbreaks due to the consumption of ready-to-eat vegetables have increased worldwide, with Escherichia coli (E. coli) being one of the main sources responsible. Viable but nonculturable bacteria (VBNC) retain virulence even after some disinfection procedures and constitute a huge problem to public health due to their non-detectability through conventional microbiological techniques. Flow cytometry (FCM) is a promising tool in food microbiology as it enables the distinction of the different physiological states of bacteria after disinfection procedures within a short time. In this study, samples of lettuce inoculated with E. coli were subject to disinfection with sodium hypochlorite at free chlorine concentrations of 5, 10, 25, 50, and 100 mg·L−1 or with 35% peracetic acid at concentrations of 5, 10, 25, and 50 mg·L−1. The efficiency of these disinfectants on the viability of E. coli in lettuce was evaluated by flow cytometry with LIVE/DEAD stains. Results from this study suggest that FCM can effectively monitor cell viability. However, peracetic acid is more effective than sodium hypochlorite as, at half the concentration, it is enough to kill 100% of bacteria and always induces a lower percentage of VBNC. Finally, we can conclude that the recommended levels of chemical disinfectants for fresh fruit and vegetables are adequate when applied in lettuce. More importantly, it is possible to ensure that all cells of E. coli are dead and that there are no VBNC cells even with lower concentrations of those chemicals. These results can serve as guidance for lettuce disinfection, improving quality and the safety of consumption.

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Comparison between Flow Cytometry and Traditional Culture Methods for Efficacy Assessment of Six Disinfectant Agents against Nosocomial Bacterial Species

Cited by 21 publications

References 17 publications

A Guide for Ex Vivo Handling and Storage of Stool Samples Intended for Fecal Microbiota Transplantation

A Guide for Ex Vivo Handling and Storage of Stool Samples Intended for Fecal Microbiota Transplantation

Antibacterial Effect of Potassium Tetraborate Tetrahydrate against Soft Rot Disease Agent Pectobacterium carotovorum in Tomato

Evaluation by Flow Cytometry of Escherichia coli Viability in Lettuce after Disinfection

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