Dans les produits laitiers, les cultures bioprotectrices et leurs métabolites représentent une alternative d’intérêt aux conservateurs chimiques pour lutter contre les contaminations fongiques. L’objectif de cette thèse était de développer des ingrédients antifongiques, issus de la fermentation d’un substrat laitier par différents microorganismes, bactéries ou champignons, utilisables dans des produits laitiers variés. Pour ce faire, nous avons dans un premier temps caractérisé la diversité des contaminants fongiques des produits laitiers et de leur environnement afin de sélectionner les cibles fongiques les plus pertinentes pour le reste de notre étude. Ensuite, nous avons criblé, in vitro, l’activité antifongique de fermentats issus de la fermentation de 2 substrats laitiers par 698 souches de bactéries lactiques, propioniques et de champignons grâce à une nouvelle méthode de criblage haut-débit, dans une matrice mimant le fromage. Après optimisation des conditions de fermentation pour améliorer l’activité antifongique de ces fermentats, les plus actifs ont été testés à l’échelle du laboratoire sur des fromages blancs et des fromages à pâte pressée à croûte morgée (PPCM) avant d’être testés à l’échelle pilote dans des crèmes fraiches et des fromages à PPCM. Cette étape nous a permis (i) de valider l’activité antifongique des fermentats en produits réels en mettant en place des challengetests et des tests d’usage et (ii) d’évaluer leur impact sur les qualités organoleptiques des produits grâce à des analyses sensorielles. Les molécules impliquées dans l’activité antifongique ont ensuite été identifiées grâce à différentes méthodes (chromatographie en phase gazeuse ou liquide couplée ou non à la spectrométrie de masse) et l’impact de ces composés sur la croissance des cibles fongiques a été évalué. Le criblage in vitro a permis la sélection de 3 ingrédients antifongiques qui se sont tous révélés actifs in situ. Les tests de durabilité ont révélé que le fermentat issu de Lactobacillus rhamnosus CIRM-BIA 1952 avait une activité prometteuse dans les crèmes fraiches. Pour le fromage à PPCM, le fermentat issu de Propionibacterium jensenii CIRM-BIA 1774, qui retarde jusqu’à 16 jours la croissance de Mucor racemosus et Penicillium commune pourrait être utilisé à la place de la natamycine. La caractérisation des composés antifongiques dans les fermentats a mis en évidence certains acides organiques, des acides gras, des composés volatils et un peptide qui, ensemble, jouent très probablement un rôle dans l’activité antifongique de ces fermentats. Nous avons enfin montré l’effet fongistatique du fermentat le plus actif sur Rhodotorula mucilaginosa UBOCC-A-216004 et Mucor racemosus UBOCC-A-109155 ainsi que son impact sur la physiologie des spores de M. racemosus. L’ensemble de ces résultats devrait donc conduire au développement de nouveaux ingrédients laitiers antifongiques qui pourraient remplacer avantageusement les conservateurs dans les produits laitiers, ils apportent en outre des éléments d’information sur les molécules et les mécanismes mis en œuvre dans l’inhibition antifongique. / In dairy products, bioprotective cultures and their metabolites represent an interesting alternative to chemical preservatives. The aim of this PhD thesis was to develop antifungal dairy ingredients, derived from the fermentation of dairy substrates by different microorganisms, bacteria and fungi, which could be used in various dairy products. To do so, we first studied the diversity of spoilage fungi in a large variety of dairy products and their environment in order to select the most appropriate fungal targets. Then, we screened the in vitro antifungal activity of fermentates obtained from the fermentation of 2 dairy substrates by 698 lactic acid bacteria, propionibacteria and fungal strains using a novel high-throughput method in a cheese mimicking model. After optimizing fermentation conditions to enhance antifungal activity, the most active fermentates were tested at a lab scale in semi-hard and fresh cheese before evaluation at a pilot scale in semi-hard cheese and sour cream. This step allowed us (i) to validate the fermentate antifungal activities in real products using challenge- and durability-tests and (ii) to evaluate their impact on the products’ organoleptic properties using sensorial analysis. Antifungal molecules were then identified using different analytical methods (high performance liquid chromatography, gas chromatography coupled or not with mass spectrometry) and impact of these compounds on fungal growth was studied. The in vitro screening allowed selecting 3 antifungal ingredients that were also active in situ. Durability tests revealed that Lactobacillus rhamnosus CIRMBIA 1952 fermentate had a promising activity in sour cream. In semi-hard cheese, Propionibacterium jensenii CIRMBIA 1774 fermentate, which delayed Mucor racemosus and Penicillium commune growth for up to 16 days, could be used instead of natamycin. Antifungal compounds present in fermentates consisted of organic acids, free fatty acids, volatile compounds and peptides which altogether might play a role in the antifungal activity of these fermentates. Finally, the fungistatic effect of P. jensenii CIRM-BIA 1774 fermentate against R. mucilaginosa UBOCC-A-216004 and M. racemosus UBOC-A-109155 was demonstrated, as well as its impact on the physiology of M. racemosus spores. Together, these results should lead to the development of new antifungal dairy ingredients which could replace preservatives in dairy products. Finally, these results gave us new information concerning antifungal molecules and their action mechanisms.

no / One of the major roles lichens play in shaping the natural world, both physically and biologically, is as agents in soil development: formerly considered only in a geological context, recent research has shown that they are capable of biodeteriorating stone substrata within a relatively short timescale. Chemical alteration of the substratum is achieved by the disruptive action of many species, particularly those capable of producing an oxalate at the thallus–substratum interface . The oxalate contributes significantly to the bulk and composition of the thallus itself and persists as an obvious encrustation after the lichen’s death. In the past, these disfiguring oxalate residues on ancient monuments have been variously interpreted as resulting from former mechanical/chemical renovation treatments, atmospheric pollution , and climatic weathering. The origin and nature of oxalate accretions, the factors involved in pedogenesis , and the development of lichen mosaics are reviewed. Aesthetic disfigurement versus physical damage to stonework is considered, and various aspects of stonework conservation are discussed.

La contamination fongique des produits laitiers est à l’origine de pertes économiques conséquentes et de gaspillage alimentaire. Dans un contexte de demande pour plus de « naturalité », les cultures de bioprotection et leurs métabolites représentent une alternative d’intérêt aux conservateurs chimiques pour lutter contre ces contaminants.Les objectifs de cette thèse étaient i) de sélectionner des micro-organismes présentant une activité antifongique, pour élaborer des cultures de bioprotection applicables dans des produits laitiers, et ii) d’étudier les composés potentiellement supports de l’activité antifongique observée. Dans un premier temps, l’activité antifongique de 32 souches de bactéries lactiques et propioniques a été étudiée en modèles « fromage » et « yaourt ». L’étude de combinaisons de souches et de leur innocuité a conduit à sélectionner 2 combinaisons binaires de lactobacilles (A1 et A3). Leur efficacité et applicabilité a été évaluée à l’échelle pilote en fabrication de crème fraîche et de fromage.Les challenges tests et tests d’usages ont montré que selon le produit laitier, A1 et A3 ont une activité antifongique similaire ou supérieure que les cultures bioprotectrices commerciales. Selon l’inoculum ajouté, ces cultures n’impactent pas les caractéristiques technologiques et organoleptiques des produits laitiers. Des méthodes chromatographiques des composés antifongiques suivies d’analyses statistiques ont permis de mettre en évidence des « cocktails » de 2 à 17 composés, selon la matrice et la culture considérée, qui sont probablement supports de l’activité antifongique.Ces travaux contribuent à une meilleure compréhension des mécanismes d’action de l’activité antifongique et devraient conduire au développement de cultures antifongique pour remplacer les conservateurs dans les produits laitiers. / Fungal contamination of dairy products is responsible for economic losses and food waste. In a context of “preservative-free” product demand, bioprotective cultures and their metabolites represe,t an alternative of interest of chemical preservatives to control these spoilers.The objective of this study was i) to select microorganisms exhibiting an antifungal activity, in order to elaborate bioprotectivecultures applicable in dairy products, and ii) to study the compounds potentially supporting the observed activity. Firstly, the antifungal activity of 32 strains of lactic acid and propionic bacteria screened in cheese model and yogurt. Strain combinaison study and safety assessment led to the selection of 2 binary lactobacilli combinations (A1 and A3). Their efficiency and applicability were then evaluated in pilot-scale productions of sour cream and cheese.Challenge and shelf life tests showed that depending on the dairy product, A1 and A3 have a similar or higher antifungal activity than the commercial bioprotective cultures. In addition, depending of inoculum, A1 and A3 did not impact the technological and organoleptic characteristics. Chromatographic methods and statistical analyses allowed identifying cocktails of 2 to 17 compounds, according to the considered dairy product and culture that probably support the antifungal activity.The obtained results contribute to a better understanding of the antifungal activity action mechanisms and should lead to the development of antifungal cultures to replace preservatives in dairy products.

Trichoderma spp. is one of the most researched fungi as biocontrole agent of diseases, being antagonistic to several phytopathogens in different crops. The soilborne pathogen Fusarium oxysporum causes wilt in several crops, including chrysanthemum, is of difficult control, due mainly to its survival capacity in the soil for long periods, even without the presence of the host. Studies about the population dynamics of Trichoderma spp. and Fusarium spp. and of the associated native microbiota they necessary, especially, to observe the impact of the biocontrols addition in the soil. Ornamental plants, such chrysanthemum, are cultivated in Rio Grande do Sul, but they are susceptible to several diseases, among them, the wilt of Fusarium oxysporum, mainly in protected environment. Aiming at the biocontrol of the wilt caused by F. oxysporum in chrysanthemum, as well as the understanding of the population dynamics of the microbiota in this patossystem, this work had for objectives: to study the dynamics of the fungic population present in soil used in the chrysanthemum cultivation in greenhouse in presence and ausence of wilt symptoms; to select and identify isolates of Fusarium pathogenic to chrysanthemum; to isolate and select antagonists, of the gender Trichoderma, effective in the biocontrol of Fusarium oxysporum in vitro; to verify, in vivo, the effectiveness of the antagonists tested in vitro in control of F. oxysporum; to evaluate the survival of Trichoderma sp. in substract with the incorporation of biological products of the fungus; to analyze the population dynamics of Fusarium sp. and Trichoderma sp. in sterilized soil and with addition of bioprotector; to identify the isolates of Trichoderma sp. used in the biocontrol of the pathogen; to evaluate the effect of administrations of Trichoderma sp. in non-target organisms. The soil sampling cultivated with chrysanthemum in greenhouse showed variation in the fungic populations of Trichoderma sp. and Fusarium sp. as a function of the occurrence of wilt symptoms in plants. Of the isolates of Fusarium sp. inoculated, 25,3% were pathogenic to chrysanthemum, which were found at points with visible symptoms of the disease on the plants. The biological products of Trichoderma sp. used varied in their effectiveness in the control of the wilt of chrysanthemum, being able to reach 100% of biocontrol as in the case of isolate UFSMT15.1. The desinfestation of the soil with methyl bromide reduced the population of Trichoderma, Fusarium and other fungi. The incorporation of the biological products in the soil promoted population growth of Trichoderma sp. and inhibited the growth of Fusarium sp. The molecular characterization of the isolates of Trichoderma indicated that the region of ITS of isolates UFSMT15.1, UFSMT16 and UFSMT17 of Trichoderma sp. it presents a simple band with a fragment of approximately 600 bp and the isolate UFSMT17 present high phylogenetic similarity with the specie Trichoderma aureoviride. In the rats treated with biological product of active Trichoderma sp., no viable spores of the fungus werw found in the sampled tissues after a hour of the administration of the bioproduct. / Trichoderma spp. Persson é um dos fungos mais pesquisados como agente de biocontrole de doenças, sendo antagonista a vários fitopatógenos em diferentes culturas. O patógeno de solo Fusarium oxysporum Link causador de murcha vascular em várias culturas, inclusive em crisântemo, é de difícil controle, devido principalmente à sua capacidade de sobrevivência no solo por longos períodos, mesmo sem a presença do hospedeiro. Estudos sobre a dinâmica populacional de Trichoderma spp. e de Fusarium spp. e da microbiota nativa associada são necessários, em especial, para observar-se o impacto da adição de biocontroladores no solo. Plantas ornamentais, como o crisântemo, são cultivadas no Rio Grande do Sul, mas são suscetíveis a várias doenças, dentre as quais, a murcha por Fusarium tem se destacado, principalmente, em ambiente protegido. Visando o biocontrole da murcha vascular causada por F. oxysporum em crisântemo, bem como, o entendimento da dinâmica populacional da microbiota nesse patossistema, este trabalho teve por objetivos: estudar a dinâmica da população fúngica presente em solo utilizado no cultivo de crisântemo em estufa na presença e ausência de sintomas de murcha; selecionar e identificar isolados de Fusarium patogênicos ao crisântemo; isolar e selecionar antagonistas, do gênero Trichoderma, eficazes no biocontrole de Fusarium oxysporum em teste in vitro; verificar a eficácia in vivo dos antagonistas testados no controle in vitro de F. oxysporum; avaliar a sobrevivência de Trichoderma sp. em substrato com incorporação de biopreparados desse fungo; analisar a dinâmica populacional de Fusarium sp. e Trichoderma sp. presentes em solo esterilizado e povoado com bioprotetores utilizados no cultivo de crisântemo em ambiente protegido; caracterizar molecularmente os isolados de Trichoderma sp. utilizados no biocontrole do patógeno; avaliar o efeito de administrações de Trichoderma sp. em organismos não-alvo. A amostragem de solo cultivado com crisântemo em estufa mostrou variação nas populações fúngicas de Trichoderma sp. e de Fusarium sp., em função da ocorrência de sintomas de murcha nas plantas. Dos isolados de Fusarium sp. inoculados, 25,3% foram patogênicos ao crisântemo, os quais foram amostrados em pontos com sintomas visíveis da doença nas plantas. Os biopreparados de Trichoderma sp. utilizados variaram em sua eficácia no controle da murcha do crisântemo, podendo atingir 100% de biocontrole como no caso do isolado UFSMT15.1. A desinfestação do solo com brometo de metila reduziu a população de Trichoderma, Fusarium e outros gêneros fúngicos. A incorporação do biopreparado no solo promoveu crescimento populacional de Trichoderma sp. e inibiu o crescimento de Fusarium sp. A caracterização molecular dos isolados de Trichoderma indicou que a região do ITS dos isolados UFSMT15.1, UFSMT16 e UFSMT17 de Trichoderma sp. apresenta uma banda simples com um fragmento de aproximadamente 600 pares de bases e o isolado UFSMT17 possui alta similaridade filogenética com a espécie Trichoderma aureoviride. Nos ratos tratados com o biopreparado de Trichoderma sp. ativo, não foram encontrados conídios viáveis do fungo nos tecidos amostrados após uma hora da administração do bioproduto.

L'industrie alimentaire cherche constamment à améliorer les processus de conservation des aliments. La biopréservation s’impose comme une alternative à l’utilisation de conservateurs dits «chimiques» et fait appel à des micro-organismes sélectionnés en tant que cultures protectrices pour leurs propriétés antimicrobiennes naturels. L'objectif de cette étude était de tirer profit de la diversité de Carnobacterium maltaromaticum afin d’identifier des souches capables d'inhiber la croissance du pathogène alimentaire Listeria monocytogenes. Pour cela, une méthode de typage moléculaire permettant la reconnaissance fine de souches a été établie, puis une méthode de criblage d’activités antibactériennes à robustesse élevée a été mise au point. La méthode de typage moléculaire est une méthode MLVA (Multiple-locus variable-number tandem repeat (VNTR) analysis) avec 3 loci de VNTR (VNTR-A, VNTR-B, VNTR-C) permet de discriminer 15 génotypes différents au sein d’une collection de 24 souches. Cela confirme la grande diversité génotypique et phénotypique présente au sein de la population de C. maltaromaticum et la performance de la méthode. La méthode de criblage est basée sur la réalisation d’essais de compétition à haut débit où chaque membre d’une collection de C. maltaromaticum a été co-cultivé avec une souche bioluminescente de L. monocytogenes. Il a été montré que la production de luminescence est le reflet de la croissance de L. monocytogenes en micro-culture et qu’il est ainsi possible de tester la stabilité du pouvoir antagoniste de chaque membre de la collection dans de multiples conditions de cultures différentes. Cette méthode a ainsi permis d’identifier des souches de C. maltaromaticum dont la propriété anti-L. monocytogenes est peu influencée par les conditions de cultures. Outre l’intérêt fondamental que présente cette méthode pour l’étude des interactions compétitives dans l’aliment, elle a permis d’identifier des souches présentant un fort potentiel de valorisation dans le domaine de la biopréservation alimentaire / The food industry is constantly seeking to improve food preservation processes. Biopreservation imposes itself as an alternative to the use of chemical preservatives and involves micro-organisms selected as protective cultures for their natural antimicrobial properties. The aim of this study was to benefit from the diversity of Carnobacterium maltaromaticum in order to identify strains capable of inhibiting the growth of Listeria monocytogenes, a food-borne pathogen. For this purpose, a molecular typing method that enables fine strain identification has been established, then a method of screening of antibacterial activities with high robustness has been developed. The molecular typing method is a MLVA (Multiple-locus variable-number tandem repeat (VNTR) analysis) with 3 VNTR loci (VNTR-A, VNTR-B, VNTR-C). It allowed to discriminate 15 different genotypes among a collection of 24 C. maltaromaticum strains. These results confirmed the high strain diversity within this species and showed the high performance of this method. The screening method is based on high throughput competition assays where each member of a collection of C. maltaromaticum strains was co-cultivated with a bioluminescent strain of L. monocytogenes. It was established that the bioluminescence of this strains is highly correlated to bacterial growth in micro-cultures, indicating that it is possible to investigate the stability of antagonistic properties of each collection member under multiple different culture conditions. The use of this method allowed to identify C. maltaromaticum strains with bioprotection properties that are poorly influenced by culture conditions. Besides the fundamental interest of this method for investigations of competitive interactions in food, it allowed to identify strains with high potential for bioprotection applications

Orientador: Marcelo de Almeida Silva / Resumo: A soja possui grande importância no mercado mundial, e para atender a sua crescente demanda é preciso aumentar a produtividade, que tem se tornado um desafio em condições de ausência de chuva. Para contornar os efeitos negativos do déficit hídrico, tem-se adotado práticas agrícolas com a aplicação de bioestimulantes como os compostos por ácidos fúlvicos e extrato da alga marinha Ascophyllum nodosum (L.), que atuam na melhoria do desenvolvimento das plantas em condições estressantes. Neste estudo, objetivou-se compreender como ácidos fúlvicos e extrato da alga marinha Ascophyllum nodosum (L.) atuam em conjunto nos mecanismos fisiológicos e bioquímicos das plantas e possibilitando a mitigação dos efeitos negativos gerados pelo estresse, e ainda determinar qual formulação e dose do bioestimulante tem melhor efeito sobre o desenvolvimento de plantas de soja submetidas ao déficit hídrico. O experimento foi conduzido em casa de vegetação, em vasos contendo 10 kg de solo sob delineamento experimental de blocos casualizados em fatorial duplo com dois fatores adicionais, com cinco repetições e 11 tratamentos constituídos por três formulações do bioestimulante e três doses (0,25; 0,50 e 1,0 kg ha-1), controle com déficit hídrico e controle sem déficit hídrico. As plantas foram submetidas a 50% da capacidade de campo por três dias, em seguida foram reidratadas e receberam a aplicação do bioestimulante, com exceção dos controles. Foram avaliados parâmetros de crescimento, fisiológicos, b... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Soy has great importance in the world market, and to meet its growing demand it is necessary to increase productivity, which has become a challenge in conditions of absence of rain. To circumvent the negative effects of the water deficit, agricultural practices have been adopted with the application of biostimulants such as those composed of fulvic acids and extract from the seaweed Ascophyllum nodosum (L.), which act in improving the development of plants in stressful conditions. In this study, the objective was to understand how fulvic acids and extract of the seaweed Ascophyllum nodosum (L.) act together in the physiological and biochemical mechanisms of plants and enabling the mitigation of the negative effects generated by stress, and also determine which formulation and dose of biostimulant has a better effect on the development of soybean plants submitted to water deficit. The experiment was carried out in a greenhouse, in pots containing 10 kg of soil under a randomized double-block design with two additional factors, with five replications and 11 treatments consisting of three biostimulant formulations and three doses (0.25; 0.50 and 1.0 kg ha-1), control with water deficit and control without water deficit. The plants were submitted to 50% of the field capacity for three days, then they were rehydrated and received the application of the biostimulant, with the exception of the controls. Growth, physiological, biochemical parameters and productivity components were e... (Complete abstract click electronic access below) / Doutor

La compréhension de la plus grande stabilisation des molécules biologiques par un disaccharide<br />tel que le tréhalose par rapport à d'autres excipients fait l'objet de recherches accrues. Nous<br />avons étudié l'influence du tréhalose et de deux autres stéréoisomères, le maltose et le sucrose,<br />sur la structure et sur la dynamique de l'eau et d'une protéine globulaire modèle, le lysozyme. Nous<br />avons montré que le tréhalose induit une plus grande déstructuration du réseau de liaisons hydrogène (LHs)<br />de l'eau que le maltose et le sucrose, au-delà d'une concentration de 40-50 %, à laquelle le réseau<br />de LHs des sucres percole. En outre, le nombre d'hydratation des sucres et le<br />nombre de LHs sucre-sucre suggèrent que les solutions aqueuses de tréhalose sont plus "homogènes",<br />pour des concentrations entre 33 et 66 % pds. Nous avons également localisé les zones<br />d'interaction des sucres avec le lysozyme et montré par spectroscopie Raman que le tréhalose le stabilise<br />davantage.