Contribution of Fluorescence Techniques in Determining the Efficiency of the Non-thermal Plasma Treatment

We have recently developed a non-thermal plasma (NTP) equipment intended to sterilize fragile medical devices and maintain the sterile state of items downstream the treatment. With traditional counts on agar plate a six log reduction of Staphylococcus aureus viability was obtained within 120 min of O2, Ar, or N2 NTP treatments. However to determine the best NTP process, we studied the different physiological states of S. aureus by flow cytometry (FC) and confocal laser scanning microscopy (CLSM) focusing on the esterasic activity and membrane integrity of the bacteria. Two fluorochromes, 5-(and-6)-carboxy-2′,7′-dichlorofluorescein diacetate and propidium iodide were used in order to distinguish three sub-populations: metabolically active, permeabilized, and damaged bacteria that can be in the viable but nonculturable state. FC and CLSM highlight that O2 and Ar NTP treatments were the most attractive processes. Indeed, a 5 min of Ar NTP generated a high destruction of the structure of bacteria and a 120 min of O2 NTP treatment led to the higher decrease of the total damaged bacteria population. SEM observations showed that in presence of clusters, bacteria of upper layers are easily altered compared to bacteria in the deeper layers. In conclusion, the plate counting method is not sufficient by itself to determine the best NTP treatment. FC and CLSM represent attractive indicator techniques to select the most efficient gas NTP treatment generating the lowest proportion of viable bacteria and the most debris

Microbiology of sugar-rich environments: diversity, ecology and system constraints

Microbial habitats that contain an excess of carbohydrate in the form of sugar are widespread in the microbial biosphere. Depending on the type of sugar, prevailing water activity and other substances present, sugar-rich environments can be highly dynamic or relatively stable, osmotically stressful, and/or destabilizing for macromolecular systems, and can thereby strongly impact the microbial ecology. Here, we review the microbiology of different high-sugar habitats, including their microbial diversity and physicochemical parameters, which act to impact microbial community assembly and constrain the ecosystem. Saturated sugar beet juice and floral nectar are used as case studies to explore the differences between the microbial ecologies of low and higher water-activity habitats respectively. Nectar is a paradigm of an open, dynamic and biodiverse habitat populated by many microbial taxa, often yeasts and bacteria such as, amongst many others, Metschnikowia spp. and Acinetobacter spp., respectively. By contrast, thick juice is a relatively stable, species-poor habitat and is typically dominated by a single, xerotolerant bacterium (Tetragenococcus halophilus). A number of high-sugar habitats contain chaotropic solutes (e.g. ethyl acetate, phenols, ethanol, fructose and glycerol) and hydrophobic stressors (e.g. ethyl octanoate, hexane, octanol and isoamyl acetate), all of which can induce chaotropicity-mediated stresses that inhibit or prevent multiplication of microbes. Additionally, temperature, pH, nutrition, microbial dispersion and habitat history can determine or constrain the microbiology of high-sugar milieux. Findings are discussed in relation to a number of unanswered scientific questions.status: publishe

Assessment of potential probiotic properties and multiple bacteriocin encoding-genes of the technological performing strain Enterococcus faecium MMRA

In the present study, Enterococcus faecium MMRA, a safety and technological performing strain, previously isolated from a well-known fermented dairy product of Tunisia, was evaluated for certain properties relevant to probiotic including acid and bile tolerance, gastrointestinal juice resistance and adhesive properties. Acidic pH (2.0-5.0) and bile salt concentrations of 0.3% were used as stress conditions. The adhesive properties were assessed by determination of bacterial hydrophobicity. The strain MMRA showed interesting features such as high tolerance to gastrointestinal tract transit, an extremely high survival rates under stress caused by acidic pHs or bile salts, and a high adhesive potential linked to the elevated percentage of hydrophobicity. Furthermore, the genome of this strain was examined for the occurrence of known enterocin (enterococcal bacteriocins) genes by means of specific PCR assays and this study revealed the genes coding for enterocins A, B, P and X. Although the joint production of these bacteriocins has not been verified, producers of multiple enterocins could have a great technological potential as protective and/or probiotic cultures in the food industry. The results suggested that E.faecium MMRA should be considered as a potential probiotic which meets the criteria to enhance healthy attributes of fermented dairy product besides both of hygienic and sensory aspects. © 2013 Elsevier Ltd.A part of the research presented in this paper was financially supported by the Spanish Agency for International Cooperation (AECI), Spain within the Inter-University Cooperation Programme Cooperation Programme and Scientific Research (PCI-Mediterranean) between Spain and Tunisia (project A/5664/06). Amel Rehaiem is the recipient of a MAEC-AECID fellowship (Spain).Peer Reviewe

Contribution of Fluorescence Techniques in Determining the Efficiency of the Non-thermal Plasma Treatment

International audienceWe have recently developed a non-thermal plasma (NTP) equipment intended to sterilize fragile medical devices and maintain the sterile state of items downstream the treatment. With traditional counts on agar plate a six log reduction of Staphylococcus aureus viability was obtained within 120 min of O2, Ar, or N2 NTP treatments. However to determine the best NTP process, we studied the different physiological states of S. aureus by flow cytometry (FC) and confocal laser scanning microscopy (CLSM) focusing on the esterasic activity and membrane integrity of the bacteria. Two fluorochromes, 5-(and-6)-carboxy-2′,7′-dichlorofluorescein diacetate and propidium iodide were used in order to distinguish three sub-populations: metabolically active, permeabilized, and damaged bacteria that can be in the viable but nonculturable state. FC and CLSM highlight that O2 and Ar NTP treatments were the most attractive processes. Indeed, a 5 min of Ar NTP generated a high destruction of the structure of bacteria and a 120 min of O2 NTP treatment led to the higher decrease of the total damaged bacteria population. SEM observations showed that in presence of clusters, bacteria of upper layers are easily altered compared to bacteria in the deeper layers. In conclusion, the plate counting method is not sufficient by itself to determine the best NTP treatment. FC and CLSM represent attractive indicator techniques to select the most efficient gas NTP treatment generating the lowest proportion of viable bacteria and the most debri

Biosurfactant production by Pseudomonas strains isolated from floral nectar

To screen and identify biosurfactant-producing Pseudomonas strains isolated from floral nectar; to characterize the produced biosurfactants; and to investigate the effect of different carbon sources on biosurfactant production.status: publishe

Innovative non-thermal plasma disinfection process inside sealed bags: Assessment of bactericidal and sporicidal effectiveness in regard to current sterilization norms.

In this work, we developed a device capable to generate a non-thermal plasma discharge inside a sealed bag. The aim of this study was to assess the effectiveness of the oxygen, nitrogen and argon plasma sterilization on Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis spores according to the NF EN 556 Norm. Moreover the bag integrity which is a critical key to maintain the sterile state of items after the end of the process was verified by Fourier Transform Infrared (FTIR) and X-ray Photoelectron Spectrometry (XPS) analyses. After plasma treatments, the bacterial counting showed a 6 log reduction of P. aeruginosa and S. aureus in 45 min and 120 min respectively whatever the gas used and a 4 log reduction of B. subtilis spores in 120 min with only oxygen plasma. These results were confirmed by Scanning Electron Microscopy (SEM) observations showing altered bacteria or spores and numerous debris. Taking into account the studied microorganisms, the oxygen plasma treatment showed the highest efficiency. FTIR and XPS analyses showed that this treatment induced no significant modification of the bags. To conclude this non-thermal plasma sterilization technique could be an opportunity to sterilize heat and chemical-sensitive medical devices and to preserve their sterile state after the end of the process

Scanning electron micrograph of <i>B</i>. <i>subtilis</i> spores before and after O₂ gas plasma treatments (100 W; 14 G and 1 sccm), (G x 7,500 for the left column and G x 20,0000 for the right column).

<p>(A-B) Untreated. (C-F) After O₂ plasma treatment.</p

Survival curves of <i>S</i>. <i>aureus</i> exposed to N₂, O₂, Ar non-thermal plasma (25 W; 14 G and 0.5 sccm).

<p>Each data represents the mean of data obtained from at least six independent experiments made in triplicate. Statistical analysis was performed by the Student's t test (*p<0.05).</p

Survival curves of <i>P</i>. <i>aeruginosa</i> exposed to N₂, O₂, Ar non-thermal plasma (25 W; 14 G and 0.5 sccm).

<p>Each data represents the mean of data obtained from at least six independent experiments made in triplicate. Statistical analysis was performed by the Student's t test (**p<0.01).</p