Michel Gautier

Agrocampus Ouest, Roazhon, Brittany, France

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Publications (74)144.37 Total impact

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    ABSTRACT: Le comité d’organisation : Véronique Broussolle, UMR 408 SQPOV, INRA PACA Avignon Frédéric Carlin, UMR 408 SQPOV, INRA PACA Avignon Didier Lereclus, UMR 319 Micalis, INRA Jouy-en-Josas Christina Nielsen-Leroux, UMR 1319 Micalis, INRA Jouy-en-Josas Vincent Sanchis, UMR 1319 Micalis, INRA Jouy-en-Josas
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    ABSTRACT: Lysozyme is mainly described active against Gram-positive bacteria, but is also efficient against some Gram-negative species. Especially, it was recently demonstrated that lysozyme disrupts Escherichia coli membranes. Moreover, dry-heating changes the physicochemical properties of the protein and increases the membrane activity of lysozyme. In order to elucidate the mode of insertion of lysozyme into the bacterial membrane, the interaction between lysozyme and a LPS monolayer mimicking the E. coli outer membrane has been investigated by tensiometry, ellipsometry, Brewster angle microscopy and atomic force microscopy. It was thus established that lysozyme has a high affinity for the LPS monolayer, and is able to insert into the latter as long as polysaccharide moieties are present, causing reorganization of the LPS monolayer. Dry-heating increases the lysozyme affinity for the LPS monolayer and its insertion capacity; the resulting reorganization of the LPS monolayer is different and more drastic than with the native protein.
    Biochimica et Biophysica Acta (BBA) - Biomembranes 01/2015; 1848(1). DOI:10.1016/j.bbamem.2014.10.026 · 3.43 Impact Factor
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    ABSTRACT: Antimicrobial resistance is currently an important public health issue. The need for innovative antimicrobials is therefore growing. The ideal antimicrobial compound should limit antimicrobial resistance. Antimicrobial peptides or proteins such as hen egg white lysozymes are promising molecules that act on bacterial membranes. Hen egg white lysozymes have recently been identified as active on Gram-negative bacteria due to disruption of the outer and cytoplasmic membrane integrity. Furthermore, dry-heating (7days and 80°C) improves the membrane activity of lysozymes, resulting in higher antimicrobial activity. These in vivo findings suggest interactions between lysozymes and membrane lipids. This is consistent with the findings of several other authors who have shown lysozyme interaction with bacterial phospholipids such as phosphatidylglycerol and cardiolipin. However, until now, the interaction between lysozyme and bacterial cytoplasmic phospholipids has been in need of clarification. This study proposes the use of monolayer models with a realistic bacterial phospholipid composition in physiological conditions. The lysozyme/phospholipid interactions have been studied by surface pressure measurements, ellipsometry and atomic force microscopy. Native lysozyme has proved able to absorb and insert into a bacterial phospholipid monolayer, resulting in lipid packing reorganization, which in turn has lead to lateral cohesion modifications between phospholipids. Dry-heating of lysozymes has increased insertion capacity and ability to induce lipid packing modifications. These in vitro findings are then consistent with the increased membrane disruption potential of dry heated lysozyme in vivo compared to native lysozyme. Moreover, an eggPC monolayer study suggested that lysozyme/phospholipid interactions are specific to bacterial cytoplasmic membranes. Copyright © 2015. Published by Elsevier B.V.
    Biochimica et Biophysica Acta (BBA) - Biomembranes 01/2015; 1848(4). DOI:10.1016/j.bbamem.2015.01.008 · 3.43 Impact Factor
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    ABSTRACT: As a 1st step, this study aimed at investigating the microbial quality of liquid egg white in a French egg processing company. Thirty raw and 33 pasteurized liquid egg white samples were analyzed. Pasteurization was globally found efficient on mesophilic contaminants (1.7 ± 1.6 and 0.8 ± 0.9 log CFU/mL in raw and pasteurized samples, respectively), including for the control of Salmonella. However,Gram-positive enterococci were still detected in the pasteurized samples. As a 2nd step, a representative bacterial collection was built for exploring the spoilage issue in egg-based chilled desserts. Custard cream was chosen as growth medium since this food is widely used for the production of French chilled desserts. All of the 166 isolates of the bacterial collection were shown to be able to grow and to induce spoilage of the custard cream at refrigeration temperature (10 °C). Several spoilage types were highlighted in the custard cream, on the basis of changes regarding pH, consistency, production of holes or gas. As a 3rd step, bacterial enzymatic activities were explored on custard cream-based agar media. The bacterial collection was reduced to 43 isolates, based on further selection regarding the genera and the spoilage types previously highlighted. Albeit to different degrees, all these isolates were able to produce proteases. A large part of these isolates also expressed lipolytic and amylolytic activities. This study emphasizes the need to control egg white contamination and especially with Gram-positive heat-resistant Enterococi, in order to guarantee the shelf life of egg-based chilled desserts.
    Journal of Food Science 01/2015; 80(2). DOI:10.1111/1750-3841.12764 · 1.79 Impact Factor
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    ABSTRACT: The objective of this study was to develop and evaluate a SYBR Green real-time PCR method for the specific detection of Salmonella spp using a novel target, the siiA gene. Primers specificity testing was done on a panel of 76 Salmonella strains and 32 non-Salmonella strains. The primers directed against the siiA gene amplified all Salmonella strains tested, while non-Salmonella strains were not amplified. The melting temperatures of the 107bp amplicons were consistently specific as they gave melting peaks around 75.5°C. The precision of the assay, based on intra and inter-run variations, was shown to be widely acceptable. In the second part of this study, 45 Salmonella strains were screened for the presence of 6 virulence-associated genes (sopB, cat2, safC, sefB and SC1248) located in several Salmonella Pathogenicity Islands (SPIs) and the spvC gene from the Salmonella virulence plasmid. The prevalence of these genes ranged from 51% to 100%. Variable virulence gene profiles were obtained even within the same serotype. Copyright © 2014. Published by Elsevier B.V.
    Journal of Microbiological Methods 12/2014; 109. DOI:10.1016/j.mimet.2014.11.018 · 2.10 Impact Factor
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    ABSTRACT: Although egg white is known as expressing efficient antimicrobial properties under physiological conditions, little is known on the microbial quality of industrial egg white. This issue is of great importance when egg white enters into the composition of highly perishable products, such as chilled egg-based desserts. Due to the high thermo-sensitivity of its constitutive proteins, liquid egg white is stabilized at low heat-treating times and temperatures (2 to 6 min at 55°C to 57°C in France). The remaining of heat-resisting bacteria may shorten the shelf-life of the desserts, with damaging health and economical consequences. The objective of this study was to investigate the microbial quality of raw and pasteurized liquid egg white products collected in a French company at two consecutive warm and cold seasons. A total of 63 samples were analyzed, including 30 raw and 33 pasteurized (at 57°C for 6 min) liquid egg white products. The level and type of bacterial contamination was evaluated by pour-plating in PCA (incubation for 24h at 30°C under aerobic conditions) and 16SrDNA sequencing of the colonies. The average counts were fairly low and a broad range of population was highlighted from one sample to another, namely 1.7 ± 1.6 log CFU/mL
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    ABSTRACT: Bacillus cereus group bacteria are opportunistically pathogenic spore-forming microorganisms well known in the sector of pasteurized food products because of their involvement in spoilage events. In the sector of egg product processing, these bacteria may lead to important economic losses. It seemed then relevant to study their behavior in egg white, a widely used egg product usually recognized as developing different levels of antimicrobial activities depending on the environmental conditions. A strong bactericidal effect (decrease in the bacterial population of 6.1 ± 0.2 log CFU/ml) was observed for 68 B. cereus group isolates, independently incubated at 30°C in egg white at pH 9.3 (natural egg white pH). To determine which components could explain such a strong bactericidal effect, an experimental strategy was carried out, based on egg white fractionation by ultrafiltration and by anion-exchange liquid chromatography. The role of the protein fraction was thus demonstrated, and subsequent nano-liquid chromatography-tandem mass spectrometry analyses allowed identification of ovotransferrin as a major protein involved. The strong bactericidal effect was confirmed in the presence of commercial ovotransferrin. Such a bactericidal effect (i.e., a decrease in the bacterial population through cell death) had never been described because ovotransferrin is known for its bacteriostatic effect (i.e., inhibition of growth) due to its ability to chelate iron. Surprisingly, the addition of iron did not reverse the bactericidal effect of ovotransferrin under alkaline conditions (pH 9.3), whereas it completely reversed this effect at pH 7.3. Ovotransferrin was shown to provoke a perturbation of the electrochemical potential of the cytoplasmic membrane. A membrane disturbance mechanism could, hence, be involved, leading to the lysis of B. cereus group bacteria incubated in egg white.
    Journal of food protection 06/2014; 77(6):955-62. DOI:10.4315/0362-028X.JFP-13-473 · 1.80 Impact Factor
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    ABSTRACT: Bovine abortion of unknown infectious etiology still remains a major economic problem. Thus, we investigated whether Brucella spp., Listeria monocytogenes, Salmonella spp., Campylobacter spp. and Coxiella burnetii are associated with abortion and/or stillbirth in Tunisian dairy cattle. Using a pan-Chlamydiales PCR, we also investigated the role of Chlamydiaceae, Waddlia chondrophila, Parachlamydia acanthamoebae and other members of the Chlamydiales order in this setting. Veterinary samples taken from mid to late-term abortions from twenty dairy herds were tested. From a total of 150 abortion cases collected, infectious agents were detected by PCR in 73 (48.66%) cases, 13 (8.66%) of which represented co-infections with two infectious agents. Detected pathogens include Brucella spp (31.3%), Chlamydiaceae (4.66%), Waddlia chondrophila (8%), Parachlamydia acanthamoebae (5.33%), Listeria monocytogenes (4.66%) and Salmonella spp. (3.33%). In contrast, Campylobacter spp. and Coxiella burnetii DNA were not detected among the investigated veterinary samples. This demonstrates that different bacterial agents may cause bovine abortion in Tunisia. This is the first report suggesting the role of Parachlamydia acanthamoebae in bovine abortion in Africa. Further studies with a larger number of samples are necessary to confirm whether this emerging pathogen is directly linked to abortion in cattle.
    PLoS ONE 03/2014; 9(3):e91549. DOI:10.1371/journal.pone.0091549 · 3.23 Impact Factor
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    ABSTRACT: To evaluate the food safety and spoilage risks associated with psychrotrophic Bacillus cereus group bacteria for the egg product industry, and to search for relevant risk markers. A collection of 68 psychrotrophic B. cereus group isolates, coming from pasteurized liquid whole egg products, was analyzed through a principal component analysis (PCA) regarding their spoilage and food safety risk potentials. The principal component analysis showed a clear differentiation between two groups within the collection, one half of the isolates representing a safety risk and the other half a spoilage risk. Relevant risk markers were highlighted by PCA, i.e. (i) for the food safety risk, the presence of the specific 16S rDNA-1m genetic signature and the ability to grow at 43°C on solid medium; and (ii) for the spoilage risk, the presence of the cspA genetic signature. This work represents a first step in the development of new diagnostic technologies for the assessment of the microbiological quality of foods likely to be contaminated with psychrotrophic B. cereus group bacteria. This article is protected by copyright. All rights reserved.
    Journal of Applied Microbiology 02/2014; 116. DOI:10.1111/jam.12462 · 2.39 Impact Factor
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    ABSTRACT: For food as well as for medical applications, there is a growing interest for novel and natural antimicrobial molecules. Lysozyme is a promising candidate for the development of such molecules. This protein is largely studied and known for its muramidase activity against Gram-positive bacteria, but it also shows antimicrobial activity against Gram-negative bacteria, especially when previously modified. In this study, the activity of dry-heated lysozyme (DH-L) against E. coli has been investigated and compared to that of native lysozyme (N-L). Whereas N-L only delays bacterial growth, DH-L causes an early stage population decrease. The accompanying membrane permeabilization suggests that DH-L induces either larger pores or more pores in the outer membrane as compared to N-L, as well as more ion channels in the inner membrane. The strong morphological modifications observed by optical microscopy and AFM when E. coli cells are treated with DH-L are consistent with the suggested disturbances of membrane integrity. The higher hydrophobicity, surface-activity and positive charge induced by dry-heating would be responsible of the increased activity of DH-L on the E. coli membranes.
    Journal of Agricultural and Food Chemistry 01/2014; 62(7). DOI:10.1021/jf405155p · 3.11 Impact Factor
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    ABSTRACT: All over the world, the incidence of Salmonella spp contamination on different food sources like broilers, clams and cow milk has increased rapidly in recent years. The multifaceted properties of Salomnella serovars allow the microorganism to grow and multiply in various food matrices, even under adverse conditions. Therefore, methods are needed to detect and trace this pathogen along the entire food supply network. In the present work, PFGE and ERIC-PCR were used to subtype 45 Salmonella isolates belonging to different serovars and derived from different food origins. Among these isolates, S. Enteritidis and S. Kentucky were found to be the most predominant serovars. The Discrimination Index obtained by ERIC-PCR (0.85) was slightly below the acceptable confidence value. The best discriminatory ability was observed when PFGE typing method was used alone (DI = 0.94) or combined with ERIC-PCR (DI = 0.93). A wide variety of profiles was observed between the different serovars using PFGE or/and ERIC-PCR. This diversity is particularly important when the sample origins are varied and even within the same sampling origin.
    PLoS ONE 12/2013; 8(12):e81315. DOI:10.1371/journal.pone.0081315 · 3.23 Impact Factor
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    ABSTRACT: Abstract This article reports the prevalence and antibiotic resistance of the Bacillus cereus group isolated from different foods (milk and dairy products, spices, and rice salad) in Morocco. In total, 402 different food samples collected from 2008 to 2010 were analyzed by microbiological methods to isolate B. cereus. The strains were subjected to a polymerase chain reaction test in order to verify whether they belonged to the B. cereus group. Sixty-four of all isolates (15.9%) were found to be positive. Among the sources, B. cereus strains from milk and dairy products constituted the largest proportion of isolates (33/64; 51.6%) followed by spices (22/64; 34.4%) and salad with rice (9/64; 14.1%). The genetic diversity of the strains of B. cereus group was examined by pulsed-field gel electrophoresis (PFGE) of chromosomal DNA digested with SmaI. The enzyme restriction profiles showed a high degree of polymorphism among the strains. The results showed that PFGE analysis could reveal the genetic differences among B. cereus strains. Investigation of antibiotic-resistance profiles showed that isolates were resistant to ampicillin (98.4%), tetracycline (90.6%), oxacillin (100%), cefepime (100%), and penicillin (100%), and were susceptible to chloramphenicol (67.2%), erythromycin (84.4%), and gentamicin (100%). The results of this study indicated that B. cereus could be a significant etiological agent of food poisoning in Morocco because of its high prevalence. Also, we demonstrated that the majority of strains came from milk and dairy products. However, additional research involving cytotoxicity tests is needed to more evaluate this sanitary risk.
    Foodborne Pathogens and Disease 11/2013; 11. DOI:10.1089/fpd.2013.1615 · 2.09 Impact Factor
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    ABSTRACT: Natural preservatives answer the consumer demand for long shelf-life foods, considering synthetic molecules are perceived as a health risk. Lysozyme is already used because of its muramidase activity against Gram-positive bacteria. It is also described active against some Gram-negative bacteria; membrane disruption would be involved, but the mechanism remains unknown. In the presently reported study, a spectrophotometric method using the mutant Escherichia coli ML-35p, has been adapted to investigate membrane disruption by lysozyme for long durations. Lysozyme rapidly increases the permeability of the outer membrane of E. coli due to large size pore formation. A direct delayed activity of lysozyme against the inner membrane is also demonstrated, but without evidence of perforations.
    Journal of Agricultural and Food Chemistry 09/2013; 61(41). DOI:10.1021/jf4029199 · 3.11 Impact Factor
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    ABSTRACT: Bacterial membranes are often thought to be the main targets of the antimicrobial activity of egg white. In order to test this hypothesis, the state of the membranes of Escherichia coli K-12 cells during either bactericidal (45°C) or bacteriostatic (30°C) incubation in egg white at natural alkaline pH was studied by biochemical methods. Namely, the permeability of the outer membrane was evaluated through its ability to incorporate a hydrophobic fluorescent probe (1-N-phenylnaphthylamine), and the permeability of the cytoplasmic membrane was evaluated through the release of a specific intracellular enzyme (β-galactosidase). The bacteria were observed by atomic force microscopy in order to support the biochemical results. At 45°C, the outer membrane of E. coli K-12 incorporated the hydrophobic probe, suggesting that it was disrupted. In addition, the cytoplasmic β-galactosidase was released at this temperature. The atomic force microscopy analysis revealed the formation of spheroplasts, which provided further evidence of the cell wall disruption and a progressive release of cellular contents. At 30°C, biochemical and micrographic experiments confirmed that membrane integrity was preserved. These techniques provide a useful approach for studying the mechanisms of bacterial cell death in egg white.
    Journal of food protection 09/2013; 76(9):1523-9. DOI:10.4315/0362-028X.JFP-12-418 · 1.80 Impact Factor
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    ABSTRACT: Rennes – France) Quelques mois après l’autorisation de BACTOCELL® DRINK en eau de boisson, Lallemand Animal Nutrition a organisé sa deuxième journée technique BACTOCELL® pondeuses. Cet évènement a eu lieu le 26 Juin à Rennes où plus d’une trentaine de professionnels du monde avicole étaient présents. Le temps d’une demi-journée, différents sujets autour de la microflore des pondeuses ont été abordés. Plusieurs intervenants se sont succédés, apportant des présentations variées et complémentaires : Sophie Jan ( INRA-Agrocampus) Rennes, Hervé Morvan (LDA 22), Adeline Huneau (ANSES).
  • S. Merzougui · M. Lkhider · N. Grosset · M. Gautier · N. Cohen
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    ABSTRACT: Bacillus cereus is widespread pathogen. It is widely distributed in several environments such as soil and plants and is commonly isolated from food and additives. In this study we analyzed 18 foodborne B. cereus strains isolated in Morocco from food samples (milk and dairy products, rice salade and spices), in order to investigate the genetic diversity (assessed by PFGE and ERIC-PCR). The food samples were collected from hotels (n=38), restaurants (n=32) and private companies (n=26) in several cities in Morocco. The results obtained in this study confirmed the diversity of B. cereus strains. The results showed that PFGE analysis has good discriminatory power but it is long to implement, which promotes the use of ERIC-PCR as a complementary tool for molecular typing. However, additional studies involving
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    ABSTRACT: The species Bacillus cereus, known for its ability to cause food borne disease, consists of a large variety of strains. An important property for discrimination of strains is their growth temperature range. Fifty-two strains of Bacillus cereus isolated from different sources of food (milk, dairy product, spices and rice salad) for two years were determined to be either mesophilic or psychrotrophic by growth at 6 °C and at 43° C on optimal agar medium. The strains were also screened by real time polymerase chain reaction to discriminate between mesophilic and psychrotrophic types. The result obtained allowed highlighting eight profiles. Thirty seven of the 52 strains were able to grow at 6°C, but only thirteen conformed to the new psychrotolerant species Bacillus weihenstephanensis. The presence of the gene components encoding production of enterotoxins Nhe, Hbl, EntT and a recently described cytotoxin K was determined by PCR. All the strains possessed genes for at least one of these toxins. The nhe genes were detected in a higher proportion than hbl genes. Haemolytic enterotoxin was detected in 71.1 per cent of the isolates. Results of this study indicate that there are intermediate forms between B. cereus and B. weihenstephanensis, these results might be of importance for gaining further understanding of the growth properties of B. weihenstephanensis and psychrotolerant B. cereus as well as their contribution to food poisoning. However, no relationship among haemolysis test, enterotoxin genes and growth temperatures of the strains was found.
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    ABSTRACT: Abstract 1. The growth of Bacillus cereus group bacteria often limits the shelf-life of pasteurised liquid egg products and is also a putative toxin producer. This study was performed to better understand the route of contamination by B. cereus in egg products by studying the factors affecting eggshell contamination on-farm. 2. Eggs were collected in warm and cold seasons in 50 conventional laying farms in Western France. Egg surfaces were analysed for the presence of B. cereus group bacteria, environmental measurements were recorded and production practices were identified through a questionnaire filled out by the farmers. 3. A total of 44% of the farms were contaminated by mesophilic and 10% by psychrotrophic B. cereus group bacteria. No significant effect of the season was observed, whatever the thermal type. Several procedures were associated with reduced eggshell contamination by mesophilic bacteria, including the limitation of dust formation from manure and feeding and efficient disinfection of the silo, houses and the sanitary wall between houses. 4. The research highlights the need to promote prevention strategies, from farm to fork, for the control of eggshell and putative subsequent egg product contamination by B. cereus group bacteria.
    British Poultry Science 05/2013; 54(3). DOI:10.1080/00071668.2013.783900 · 0.78 Impact Factor
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    ABSTRACT: The species Bacillus cereus, known for its ability to cause food borne disease, consists of a large variety of strains. An important property for discrimination of strains is their growth temperature range. Fifty-two strains of Bacillus cereus isolated from different sources of food (milk, dairy product, spices and rice salad) for two years were determined to be either mesophilic or psychrotrophic by growth at 6 °C and at 43° C on optimal agar medium. The strains were also screened by real time polymerase chain reaction to discriminate between mesophilic and psychrotrophic types. The result obtained allowed highlighting eight profiles. Thirty seven of the 52 strains were able to grow at 6°C, but only thirteen conformed to the new psychrotolerant species Bacillus weihenstephanensis. The presence of the gene components encoding production of enterotoxins Nhe, Hbl, EntT and a recently described cytotoxin K was determined by PCR. All the strains possessed genes for at least one of these toxins. The nhe genes were detected in a higher proportion than hbl genes. Haemolytic enterotoxin was detected in 71.1 per cent of the isolates. Results of this study indicate that there are intermediate forms between B. cereus and B. weihenstephanensis, these results might be of importance for gaining further understanding of the growth properties of B. weihenstephanensis and psychrotolerant B. cereus as well as their contribution to food poisoning. However, no relationship among haemolysis test, enterotoxin genes and growth temperatures of the strains was found.