CNRS Laboratory for Microbiology, Adaptation & Pathogenesis

Lyon, France

CNRS Laboratory for Microbiology, Adaptation & Pathogenesis

Lyon, France

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Pigeot-Remy S.,CNRS Research on Catalysis and Environment in Lyon | Pigeot-Remy S.,Ahlstrom | Simonet F.,CNRS Research on Catalysis and Environment in Lyon | Errazuriz-Cerda E.,Center Communications Dimagerie Of Laennec Cecil | And 3 more authors.
Applied Catalysis B: Environmental | Year: 2011

In order to identify some of the potential bacterial targets, the effects of TiO2 nanoparticles on bacteria in aqueous suspension were assessed in the dark and under UV-A (λ>340nm) radiation exposure, using the microorganism model Escherichia coli K-12. Illumination was produced with a HPK 125W lamp and suspended TiO2 Degussa P-25 was used as the photocatalyst, absorbing all the incident UV-A radiations.The impact of the photocatalyst on the bacteria was investigated by monitoring cell cultivability, cell wall integrity and nucleic acid stability. The contact of TiO2 particles with bacteria in the dark increased the bacterial sensitivity to membrane-perturbing agents, suggesting an increase in outer membrane permeability. In contrast, the contact between SiO2 particles, with an average particle size similar to that of TiO2 P-25, and bacteria did not induce any alteration of the cell permeability. The TiO2 deleterious action on the envelope integrity continued during the UV-A radiations exposure. Impacts on bacterial permeability precede the total loss of cultivability. After 2.5h of photocatalytic treatment at 3.45mW/cm2, bacteria were no longer cultivable on their standard growth medium. However, some of them could become cultivable again under specific environmental conditions appropriate to their survival. These resilient bacteria exposed again to UV-A photocatalysis were more resistant to the treatment.Some chemical by-products released during photocatalytic inactivation of the bacteria were also monitored. The appearance of oxamic and oxalic acids as well as ammonium cations, sulfate and nitrate anions were observed. Transmission electron microscopy (TEM) was used to study the morphological damages to E. coli structure during the photocatalytic inactivation of the microorganisms. After 1.5. h of treatment, bacteria showed disorganized membrane structures, while bacteria were still visible although they were no longer cultivable after a longer exposure time. These results were correlated with damages of nucleic acids at in vivo level. An analysis by electrophoresis revealed that bacterial DNA and RNA molecules completely disappeared after 7 h of photocatalytic treatment. © 2011.


Pigeot-Remy S.,CNRS Research on Catalysis and Environment in Lyon | Pigeot-Remy S.,Ahlstrom | Pigeot-Remy S.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Simonet F.,CNRS Research on Catalysis and Environment in Lyon | And 3 more authors.
Water Research | Year: 2012

In order to compare the disinfection potential of photocatalysis and photochemistry, the effects of these two processes on bacteria in water were investigated under exposure to UV-A and UV-C. The well-known bacterial model Escherichia coli (E. coli) was used as the experimental organism. Radiation exposure was produced with an HPK 125 W lamp and the standard TiO 2 Degussa P-25 was used as the photocatalyst.Firstly, the impact of photocatalysis and photochemistry on the cultivability of bacterial cells was investigated. UV-A radiation resulted in low deleterious effects on bacterial cultivability but generated colonies of size smaller than average. UV-C photocatalysis demonstrated a greater efficiency than UV-A photocatalysis in altering bacterial cultivability. From a cultivability point of view only, UV-C radiation appeared to be the most deleterious treatment.A rapid epifluorescence staining method using the LIVE/DEAD Bacterial Viability Kit was then used to assess the modifications in bacterial membrane permeability. UV-A radiation did not induce any alterations in bacterial permeability for 420 min of exposure whereas only a few minutes of exposure to UV-C radiation, with the same total radiance intensity, induced total loss of permeability. Moreover, after 20 and 60 min of exposure to UV-C and UV-A photocatalysis respectively, all bacteria lost their membrane integrity, suggesting that the bacterial envelope is the primary target of reactive oxygen species (ROS) generated at the surface of TiO 2 photocatalyst. These results were further confirmed by the formation of malondialdehyde (MDA) during the photocatalytic inactivation of bacterial cells and suggest that destruction of the cell envelope is a key step in the bactericidal action of photocatalysis. The oxidation of bacterial membrane lipids was also correlated with the monitoring of carboxylic acids, which can be considered as representatives of lipid peroxidation by-products.Finally, damages to bacterial morphology induced by UV-C photocatalysis and photochemistry were investigated through Scanning electron microscopy (SEM). Bacterial cells were observed on microscopy pictures at exposure durations corresponding to a loss of cultivability. After 90 min of exposure to UV-C radiation, bacterial cells showed little alteration of their outer membrane whereas they suffered deep deleterious damages under UV-C photocatalysis exposure. © 2012 Elsevier Ltd.


Thabet S.,CNRS Research on Catalysis and Environment in Lyon | Thabet S.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Weiss-Gayet M.,University Claude Bernard Lyon 1 | Dappozze F.,CNRS Research on Catalysis and Environment in Lyon | And 2 more authors.
Applied Catalysis B: Environmental | Year: 2013

We have investigated the antimicrobial effects of photocatalysis on yeast (Saccharomyces cerevisiae), an essential eukaryotic unicellular model of living cells. As compared to UV-A irradiation, photocatalytic inactivation kinetics revealed a faster microbial cell cultivability inactivation. Optimal experimental conditions required a semiconductor concentration of 0.1g/l with 3.8mW/cm2 UV-A radiance intensity. Cell viability was monitored by plasma membrane permeability and loss of enzymatic activity using double fluorescent dye staining and flow cytometry. Plasma membrane permeability and enzymatic activity were almost simultaneously targeted. Esterase enzymatic activity decreased progressively, suggesting that the intracellular protein pool was sequentially damaged by the treatment. In yeast cells, the presence of a thick cell wall did not prevent the plasma membrane from being a prime photocatalytic target. Monitoring of chemical byproducts confirmed the loss of membrane integrity. A massive loss of potassium, major cation in yeast, that was released first and in large amounts, remained constant beyond 5h of treatment and was correlated to the number of damaged membrane-cells. On the contrary, ammonium ions concentration gradually increased, suggesting their generation through the photocatalytic process, probably via amino acid and protein degradation. Oxamic and oxalic acid that could also arise from damages to amino acids were detected. Amino acid analysis revealed a main component, increasing over time, corresponding to glycine. Glycine could be produced via the transformation of other amino acids, released from cell wall, membrane and intracellular proteins. © 2013 Elsevier B.V.


Billon-Grand G.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Rascle C.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Droux M.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Rollins J.A.,University of Florida | Poussereau N.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis
Molecular Plant Pathology | Year: 2012

During pathogenesis on sunflower cotyledons, Botrytis cinerea and Sclerotinia sclerotiorum show a striking resemblance in symptom development. Based on pH change profiles, the colonization process of both fungi can be divided into two stages. The first stage is associated with a pH decrease, resulting from an accumulation of citric and succinic acids. The second stage is correlated with a pH increase, resulting from an accumulation of ammonia. In this article, we also report that oxalic acid is produced at the late stage of the colonization process and that ammonia accumulation is concomitant with a decrease in free amino acids in decaying tissues. Sclerotinia sclerotiorum produces eight-fold more oxalic acid and two-fold less ammonia than B.cinerea. Consequently, during sunflower cotyledon colonization by B.cinerea, pH dynamics differ significantly from those of S. sclerotiorum. Invitro assays support the inplanta results and show that decreases in pH are linked to glucose consumption. At different stages of the colonization process, expression profiles of genes encoding secreted proteases were investigated. This analysis highlights that the expression levels of the B. cinerea protease genes are higher than those of S. sclerotiorum. This work suggests that the overt similarities of S. sclerotiorum and B. cinerea symptom development have probably masked our recognition of the dynamic and potentially different metabolic pathways active during host colonization by these two necrotrophic fungi. © 2011 Bayer Crop Science. Molecular Plant Pathology © 2011 BSPP and Blackwell Publishing Ltd.


Vacher S.,CONIDIA | Hernandez C.,CONIDIA | Bartschi C.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Poussereau N.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis
Building and Environment | Year: 2010

Biocide-free and biocide-treated plasterboards as well as aluminum plate as a reference material normally considered as being insensitive to mould growth have been used as substrate to check the influence of different common wall coverings, i.e. paints and wall papers, on fungal growth. The results described in this paper show that any non-biodegradable material (such as aluminum) can become a substrate to fungal infestation once painted or wall paper applied, depending on the type of paint or wall paper used. Moreover, a biodegradable material treated with a biocide (biocide-treated plasterboard) offers partial resistance to fungal growth at a biodegradable surface covering. The main conclusion of this study is that composition of the surface covering applied on building materials is as important as the substrate itself when considering the bioreceptivity of this material to potential fungal infestation. Accordingly, any discussion on the ability of a given building material to resist or not to fungal infestation must refer to the exact composition of the surface covering (paint, varnish, wall paper, etc). This has not often been the case in many of the previous studies published on the topic. © 2009 Elsevier Ltd. All rights reserved.


Hervet E.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Charpentier X.,Columbia University | Charpentier X.,Joseph Fourier University | Vianney A.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | And 4 more authors.
Infection and Immunity | Year: 2011

Legionella pneumophila is the etiological agent of Legionnaires' disease. Crucial to the pathogenesis of this intracellular pathogen is its ability to subvert host cell defenses, permitting intracellular replication in specialized vacuoles within host cells. The Dot/Icm type IV secretion system (T4SS), which translocates a large number of bacterial effectors into host cell, is absolutely required for rerouting the Legionella phagosome. Many Legionella effectors display distinctive eukaryotic domains, among which are protein kinase domains. In silico analysis and in vitro phosphorylation assays identified five functional protein kinases, LegK1 to LegK5, encoded by the epidemic L. pneumophila Lens strain. Except for LegK5, the Legionella protein kinases are all T4SS effectors. LegK2 plays a key role in bacterial virulence, as demonstrated by gene inactivation. The legK2 mutant containing vacuoles displays less-efficient recruitment of endoplasmic reticulum markers, which results in delayed intracellular replication. Considering that a kinase-dead substitution mutant of legK2 exhibits the same virulence defects, we highlight here a new molecular mechanism, namely, protein phosphorylation, developed by L. pneumophila to establish a replicative niche and evade host cell defenses. © 2011, American Society for Microbiology.


Landraud P.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Chuzeville S.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Billon-Grande G.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Poussereau N.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Bruel C.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis
PLoS ONE | Year: 2013

Fungi are known to adapt to pH partly via specific activation of the Pal signaling pathway and subsequent gene regulation through the transcription factor PacC. The role of PacC in pathogenic fungi has been explored in few species, and each time its partaking in virulence has been found. We studied the impact of pH and the role of PacC in the biology of the rice pathogen Magnaporthe oryzae. Conidia formation and germination were affected by pH whereas fungal growth and appressorium formation were not. Growth in vitro and in planta was characterized by alkalinization and ammonia accumulation in the surrounding medium. Expression of the MoPACC gene increased when the fungus was placed under alkaline conditions. Except for MoPALF, expression of the MoPAL genes encoding the pH-signaling components was not influenced by pH. Deletion of PACC caused a progressive loss in growth rate from pH 5 to pH 8, a loss in conidia production at pH 8 in vitro, a loss in regulation of the MoPALF gene, a decreased production of secreted lytic enzymes and a partial loss in virulence towards barley and rice. PacC therefore plays a significant role in M. oryzae's biology, and pH is revealed as one component at work during interaction between the fungus and its host plants. © 2013 Landraud et al.


Thabet S.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Thabet S.,CNRS Research on Catalysis and Environment in Lyon | Simonet F.,CNRS Research on Catalysis and Environment in Lyon | Lemaire M.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | And 2 more authors.
Applied and Environmental Microbiology | Year: 2014

We have investigated the antimicrobial effects of photocatalysis on the yeast model Saccharomyces cerevisiae. To accurately study the antimicrobial mechanisms of the photocatalytic process, we focused our investigations on two questions: the entry of the nanoparticles in treated cells and the fate of the intracellular environment. Transmission electronic microscopy did not reveal any entry of nanoparticles within the cells, even for long exposure times, despite degradation of the cell wall space and deconstruction of cellular compartments. In contrast to proteins located at the periphery of the cells, intracellular proteins did not disappear uniformly. Disappearance or persistence of proteins from the pool of oxidized intracellular isoforms was not correlated to their functions. Altogether, our data suggested that photocatalysis induces the establishment of an intracellular oxidative environment. This hypothesis was sustained by the detection of an increased level of superoxide ions (O2 °-) in treated cells and by greater cell cultivability for cells expressing oxidant stress response genes during photocatalytic exposure. The increase in intracellular ROS, which was not connected to the entry of nanoparticles within the cells or to a direct contact with the plasma membrane, could be the result of an imbalance in redox status amplified by chain reactions. Moreover, we expanded our study to other yeast and filamentous fungi and pointed out that, in contrast to the laboratory model S. cerevisiae, some environmental strains are very resistant to photocatalysis. This could be related to the cell wall composition and structure. © 2014, American Society for Microbiology.


Regoudis E.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Pelandakis M.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis
Experimental Parasitology | Year: 2016

The amoeba-flagellate Naegleria fowleri is a causative agent of primary amoebic meningoencephalitis (PAM). This thermophilic species occurs worldwide and tends to proliferate in warm aquatic environment. The PAM cases remain rare but this infection is mostly fatal. Here, we describe a single copy region which has been cloned and sequenced, and was used for both conventional and real-time PCR. Targeting a single-copy DNA sequence allows to directly quantify the N. fowleri cells. The real-time PCR results give a detection limit of 1 copy per reaction with high reproducibility without the need of a Taqman probe. This procedure is of interest as compared to other procedures which are mostly based on the detection of multi-copy DNA associated with a Taqman probe. © 2015 Elsevier Inc.


Risler A.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Coupat-Goutaland B.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis | Pelandakis M.,CNRS Laboratory for Microbiology, Adaptation & Pathogenesis
Parasitology Research | Year: 2013

We examined a partial SSU-rDNA sequence from 20 Acanthamoeba isolates associated with keratitis infections. The phylogenetic tree inferred from this partial sequence allowed to assign isolates to genotypes. Among the 20 isolates examined, 16 were found to be of the T4 genotype, 2 were T3, 1 was a T5, and 1 was a T2, confirming the predominance of T4 in infections. However, the study highlighted other genotypes more rarely associated with infections, particularly the T2 genotype. Our study is the second one to detect that this genotype is associated with keratitis. Additionally, the phylogenetic analyses showed five main emerging clusters, T4/T3/T11, T2/T6, T10/T12/T14, T13/T16, and T7/T8/T9/T17, regularly obtained whichever method was used. A similar branching pattern was found when the full rDNA sequence was investigated. © 2013 Springer-Verlag Berlin Heidelberg.

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