Regulation of the bacterial second messenger cyclic-di-GMP in Salmonella Typhimurium allows it to delicately alter phenotypes to optimize invasion and survive intracellularly in epithelial cells and macrophages to become virulent and cause infection. The concentration of cyclic-di-GMP is regulated by the presence of external stimuli, sensor CMEs (diguanylate cyclases, DGCs, and phosphodiesterases, PDEs), and cyclic-di-GMP binding effectors. Previous studies established that maintenance of low cyclic-di-GMP concentrations is required for survival in macrophages and that the deletion of 3 active PDEs reduces this survival. This study aimed to further investigate the regulation of cyclic-di-GMP for survival in macrophages and epithelial cells. Salmonella Typhimurium mutants were generated and used for an infection assay with RAW 264.7 macrophage and HeLa epithelial cell lines to determine active CMEs via intracellular survival. Intracellular survival was quantified by plate counting of cell lysates at 1-, 4-, and 24-hours post-infection. Our result showed that the previously identified 3 PDEs also influenced the infection of epithelial cells. We re-established the decreased survival in the RAW 264.7 macrophage cell line and determined that the cyclic-di-GMP-binding cellulose synthase BcsA was responsible for decreased survival in macrophages. Finally, we identified an active DGC whose deletion within the 3xKO PDEs restores survival levels, suggesting that this enzyme is responsible for the synthesis of cyclic-di-GMP during macrophage infection. Further studies on how cyclic-di-GMP regulates Salmonella Typhimurium intracellular survival could lead to identifying a potential alternative drug target for treating its infections.

Regulation of the bacterial second messenger cyclic-di-GMP in Salmonella Typhimurium allows it to delicately alter phenotypes to optimize invasion and survive intracellularly in epithelial cells and macrophages to become virulent and cause infection. Cyclic-di-GMP concentration is regulated by the presence of external stimuli, sensory diguanylate cyclases (DGCs) and phosphodiesterases (PDEs), and cyclic-di-GMP binding effectors. Previous studies established that maintenance of low cyclic-di-GMP concentrations is required for survival in macrophages, and that deletion of 3 active PDEs reduces this survival. Here I showed that these 3 PDEs also influenced the infection of epithelial cells. Further studies re-established the decreased survival in an immortalized macrophage cell line and determined that cyclic-di-GMP-binding cellulose synthase BcsA was responsible for the decreased survival in macrophages. Finally, I also identified an active DGC whose deletion within the 3xKO restores survival levels, suggesting that this enzyme is responsible for the synthesis of cyclic-di-GMP during macrophage infection.

Bacteria-based cancer therapy (BBCT) holds immense promise in addressing the limitations in treatment of solid tumors. Bacterial strains used for BBCT are engineered to express therapeutics, facilitate precise navigation within the tumor microenvironment by enhancing bacteria's motility, chemotaxis (movement toward or away from specific chemicals), or other mechanisms that aid in reaching and infiltrating the tumor tissue effectively, and complementing traditional chemotherapy and immunotherapies while minimizing side effects. Bacterial motility not only influences the ability of bacteria to navigate within the tumor but also plays a pivotal role in optimizing drug delivery, treatment efficacy, and minimizing potential obstacles associated with the complex microenvironment of human tissues. However, the current understanding of bacterial motility remains limited. In this thesis, we use a reductionist approach and study bacteria motile behavior within human tissue phantoms (collagen and agar) and the bacteria-hydrogel interactions. Apart from motility, it is important to analyze the mechanical properties of the hydrogels the bacteria interact with as they play a vital role in overall behavior and physics of bacteria movement. To that extent, there exists a gap in our understanding of the viscoelastic properties of hydrogels. Lastly, systematic and comprehensive investigation of bacteria behavior in hydrogels requires tracking of thousands of individual cells. Thus, there is an unmet need to develop new automated techniques to reduce the labor-intensive manual tracking of bacteria in low-contrast hydrogel environments, with feature sizes comparable to that of bacteria. To address these gaps, this thesis proposes a trident approach towards mechanistic understanding of bacteria motility in time-invariant agar and temporally evolving collagen hydrogels to bridge critical gaps in understanding bacterial motile behavior in these media, non-destructive microelastography-based mechanical characterization of hydrogels with less than 4.7% error compared with rheology, and the development of deep learning-enabled automated bacteria tracking tools with 77% precision. / Master of Science / There exists a huge scope for improvement in cancer therapies. The gold standard chemotherapy and immunotherapies are responsible for a lot of side effects. Over a century ago, bacteria-based cancer therapy started to develop and over the period, it was discovered that they can be effective when used with traditional therapies improving precision and reducing side effects. The motility of bacteria is shown to improve bacterial distribution in solid tumors. However, the physical underpinnings of bacteria motility in the tumor environment remains understudied. This thesis proposes a trident approach, investigating bacteria motility in tissue-like environments (hydrogels), characterizing the mechanical properties of hydrogels using acoustic waves to capture bacteria-induced alterations, and developing deep-learning-enabled automated bacterial tracking approach for high throughput analysis of experimental data. We report bacteria behavior and motility patterns in hydrogels, the mechanics of these hydrogels with less than 4.7% error compared with standard characterization methods, and automated bacteria tracking with 77% precision to inform the development and advancement of bacteria-based drug delivery systems. In summary, these tools can help improve our understanding of bacteria-hydrogel interactions, allowing us to develop innovative bacteria-based cancer therapies in the long term.

Opioids have been shown to modulate immune function in a variety of assays and animal models. In a more limited number of studies, opioids have been shown to sensitize to infection. Heroin, the prototypical opioid drug of abuse, is rapidly metabolized to morphine in the body. Morphine has been used as an analgesic for hundreds of years, and continues to be a drug of choice for treating pain in ICU and trauma patients. The continued use of these opioid compounds in humans warrants further investigation of their effect on immune responses against, and progression of, common bacterial infections. Two infections were investigated in this thesis using murine models, Acinetobacter baumannii and Salmonella typhimurium. A recent increase in the prevalence of A. baumannii infections among healthy, but wounded, military personnel, lead to the hypothesis that analgesic morphine might sensitize to infection with this multiply-drug resistant bacterium. A systemic, intraperitoneal A. baumannii infection model was established in mice that resulted in rapid, disseminated disease where animals became septic as organisms replicated in the blood, lungs, and other organs. This model was used to investigate the role of various parameters of innate immune defenses to Acinetobacter. Neutralization of neutrophils by antibody depletion greatly sensitized to this infection. Infection resulted in a rapid, biphasic induction of both IL-17 and the chemokine, KC/CXCL1, a major chemotactic factor for neutrophils, that continued to rise through 18h after bacterial inoculation. However, depletion of either IL-17 or KC/CXCL1 using monoclonal antibodies failed to sensitize to Acinetobacter infection. Further, IL-17 receptor KO mice were not sensitized to this infection. Collectively, these results suggest that there must be other chemotactic factors for neutrophils that can compensate for the absence of IL-17 and KC. Morphine, delivered by extended release pellet, sensitized two strains of mice to two strains of Acinetobacter, as measured by mortality to a sublethal challenge dose, and this effect was blocked by administration of the opioid-receptor antagonist, naltrexone. . Morphine increased Acinetobacter burdens in the organs and blood of infected mice, and increased the levels of pro-inflammatory cytokines. Evidence for an effect of morphine on neutrophil infiltration was obtained. Morphine decreased the total numbers of cells, as well as the total numbers of neutrophils and macrophages infiltrating into the peritoneal cavity. This inhibition of neutrophil accumulation correlated with suppression of levels of both IL-17 and KC/CXCL1. The evidence supports the conclusion that morphine sensitizes to Acinetobacter infection by suppressing the response of neutrophils, potentially via depression of neutrophil chemotactic factors IL-17 and KC. However, taken together with the data above there are probably additional factors in addition to IL-17 and KC that are sensitizing the animals to infection in the presence of morphine. In addition to these studies, the opioid-receptor dependency of morphine-mediated sensitization to Salmonella enteric serovar Typhimurium was examined. Previous experiments had determined that extended release morphine pellets sensitized mice to a sublethal dose of Salmonella, as determined by survival and bacterial burdens in the organs of infected mice, but naltrexone resulted in only incomplete reversal of the morphine-mediated effects. To further characterize the receptor dependency of the observed phenomenon, mu-opioid receptor knockout (MORKO) mice were used. MORKO mice were found to be completely resistant to the lethal effects of morphine plus infection observed in wild-type (WT) mice. In addition, MORKO mice showed greatly reduced bacterial burdens and pro-inflammatory cytokine levels when treated with morphine and challenged with a sublethal challenge dose of Salmonella, in comparison to WT mice. In summary, the studies presented in this thesis explored basic mechanisms of innate immunity to A. baumannii using a systemic model of infection. The work provides additional evidence that morphine sensitizes to infection, using models of Acinetobacter and Salmonella in mice. An implication of this work is use of caution in the administration of opioids in patients that are susceptible to opportunistic infections. / Microbiology and Immunology

The first recorded observation of bacterial biofilms dates back to the 17th century by Antoine Van Leeuwenhoek. Today, biofilms are known as bacteria encapsulated within a self-produced extracellular matrix adherent to biotic or abiotic surfaces. Since the initial discovery of biofilms, research has explored the structure and function of biofilms. Only until recently has the role of biofilms within the medical setting become apparent. Here, we investigate the role of curli-extracellular DNA (eDNA) complexes in disease pathogenicity and explore the ability to target bacterial amyloid curli as a novel anti-biofilm therapeutic target. Biofilms of enteric bacteria, such as Escherichia coli and Salmonella enterica serovar Typhimurium, are composed of various components that act in consortium to fortify the extracellular matrix. One of the main components of enteric biofilms is amyloid curli. Curli, one of the best characterized bacterial amyloids, is a protein with a conserved cross beta sheet structure that forms basket like structures encapsulating the bacteria. Within the biofilm, curli serves to fortify the extracellular matrix, aids in bacterial attachment and protects bacteria from harsh environmental conditions. Extracellular DNA (eDNA) is another integral component of enteric biofilms. Recent reports from our lab has suggested that curli forms irreversible complexes with eDNA. Even with exposure to DNases, co-localized curli and eDNA can be observed. Other components of enteric biofilms include cellulose and Biofilm Associated Protein A. Biofilms of S. Typhimurium have been associated with significant disease pathologies. In addition to identifying the existence of curli-eDNA complexes within S. Typhimurium biofilms, our lab has also reported that curli-eDNA complexes of S. Typhimurium potentiate the autoimmune disease Systemic Lupus Erythematosus (SLE). SLE is an autoimmune disease characterized by the production of type I interferons and autoantibodies, although the etiology remains unknown. Systemically, curli binds to and activates the Toll like Receptor (TLR)1/2 complex leading to a pro-inflammatory response. In these studies we aimed to identify the innate immune mechanisms leading to the autoimmune phenotype following stimulation with curli-eDNA complexes. As TLR9 is activated by unmethylated bacterial DNA CpG DNA sequences leading to the production of type I interferons we hypothesized a potential role for TLR9 in recognizing eDNA of the curli-eDNA complex leading to the generation of the hallmarks of SLE. To investigate this hypothesis, we stimulated wild-type, TLR2 knockout, TLR9 knockout and TLR2-9 double knockout immortalized macrophages with curli-eDNA complexes purified from S. Typhimurium biofilms. We observed a significant reduction in the transcript level of type I interferons (IFN), Ifnβ, Isg15 and Cxcl10, upon stimulation of TLR2 knockout, TLR9 knockout and TLR2-9 double knockout immortalized macrophages implicating a role in TLR9 recognition of the curli-eDNA complex. As there was a significant reduction of type I interferon levels upon stimulation of TLR2 knockout macrophages, we hypothesized that TLR2 may serve as a carrier to bring the curli-eDNA complex into the endosome containing TLR9. To inhibit phagocytosis, we pretreated cells with endocytosis inhibitors and stimulated wild-type macrophages with curli-eDNA complexes. We observed a reduction in the transcript level of Ifnβ suggesting that curli-eDNA complexes gain access to endosomal TLR9 via TLR2 engagement. Finally, to explore the role of TLR2 and TLR9 in the production of autoantibodies, curli-eDNA complexes were intraperitoneal injected twice weekly for six weeks into C57BL/6 wild-type, TLR2 knockout, TLR9 mutant and TLR2 knockout-TLR9 mutant mice. We observed a robust generation of anti-double stranded autoantibodies within the first three weeks, however the production of autoantibodies was significantly decreased and delayed in the TLR2 knockout, TLR9 mutant and TLR2 knockout-TLR9 mutant mice. Overall, these data suggest that curli acts as a carrier for DNA to elicit an autoimmune response via TLR2 and TLR9. Within biofilms of S. Typhimurium, curli is the main proteinaceous component. Biofilms lacking curli destabilize and fail to form mature biofilms. Recent research has shown that in response to the production of host amyloids, the body will generate anti-amyloid antibodies in the serum. Monoclonal antibodies (mAb) generated from serum antibodies have been shown to have pan anti-amyloid properties in vitro and in vivo due to the β-sheet conformational epitope. As amyloids from both human and bacterial origin share a β-sheet conformational structure, we hypothesized as to if the anti-amyloid mAbs can eradicate S. Typhimurium biofilms by targeting curli. We incubated S. Typhimurium biofilms in the presence of various mAbs (ALZ.4A6, ALZ.4GI, ALZ.2C10 and ALZ.3H3) and observed a significant reduction of biofilm thickness and curli content within the biofilm. We deduced that ALZ.3H3 conferred the greatest anti-biofilm response. When we visualized the three-dimensional architecture of biofilms incubated with ALZ.3H3, we observed that ALZ.3H3 induced the formation of a loose architecture compared to untreated biofilms that were dense and compact. The resulting loose biofilm architecture induced by incubation with ALZ.3H3 enhanced the susceptibility of the biofilms to antibiotic exposure and macrophage clearance. We also observed enhanced biofilm eradication in vivo when catheters precoated with S. Typhimurium biofilms were inserted into the back flanks of mice that were percutaneously injected with ALZ.3H3. Both in vitro and in vivo, combination therapy of ALZ.3H3 and antibiotic enhanced biofilm clearance. In summary, we propose a novel anti-biofilm strategy by targeting the amyloid component of the biofilm, thus satisfying an unmet need in the art of biofilm prevention. Overall, these data in summation significantly broadens our understanding of disease pathogenicity and the role of curli-eDNA complexes in S. Typhimurium biofilms. As amyloid-eDNA complexes may be found in other biofilms, these results may extend beyond enteric bacteria proving novel insight into host-microbe interactions and the generation of novel anti-biofilm therapeutics. / Microbiology and Immunology

Attenuated bacterial strains have been investigated on the premise of selective tumor colonization and drug delivery potential for decades. Salmonella Typhimurium VNP20009 was derived from the parental strain 14028 through genetic modification and tumor targeting ability, being well studied for anticancer effects in mice. In 2001 Phase 1 Clinical Trials, patients diagnosed with melanoma were introduced with VNP20009, resulting in safe delivery of the strain and targeting to the tumor, however no anticancer effects were observed. Recently, it was discovered that VNP20009 contains a SNP in cheY, which encodes the chemotaxis response regulator of flagellar motor function, rendering the strain deficient in chemotaxis. Replacement of cheY with the 14028 wild-type copy resulted in a 70% restoration of phenotype in traditional chemotaxis capillary assays compared to the parental strain. We attempted to optimize the chemotactic potential of VNP20009 but were unable without reversing the attenuated state of VNP20009. Due to the role of chemotaxis in bacterial tumor colonization and eradication remaining unclear, we aimed to compare VNP20009 and VNP20009 cheY+ primary tumor colonization and impact on metastasis in an aggressive 4T1 mouse mammary carcinoma model. Bacterial tumor colonization and metastatic potential of the cancerous cells to the lungs appear bacterial chemotaxis independent. Moreover, mice bearing tumors exposed to Salmonella exhibited increased morbidity that was associated with significant liver disease. Our results suggest that in our timeline VNP20009 may not be safe or efficacious when used in the context of immunocompetent animals with aggressive, metastatic breast cancer. In a novel approach, we aimed to understand the bacterial-cancer cell relationship within the tumor microenvironment, with an emphasis on gene expression changes occurring within the eukaryotic transcriptome. We employed the B16-F10 mouse melanoma model because VNP20009 is known to colonize and eradicate these tumors in mice. First, we optimized a timeline for Salmonella treatment of mouse melanoma, finding a dramatic delay in tumor growth between 2 and 7 days due to the presence of Salmonella. Additionally, we observed upregulation of the IFN-gamma signaling pathway within tumor tissue upon exposure to Salmonella after 7 days. In future studies, we aim to analyze the bacterial transcriptome in the tumor microenvironment to gain unique understanding and contribute to knowledge supporting bacterial-mediated cancer therapies. / Ph. D. / Bacteria have become our allies in the fight against cancer. Strains of Salmonella, normally thought of as a cause of gastrointestinal discomfort, are able to target cancer in the body and effectively shrink tumors in several animal models. Specifically, a strain of Salmonella Typhimurium called VNP20009, has shown great promise as an anticancer agent. Research on VNP20009 culminated in a Phase 1 Clinical Trial in which safe delivery of the strain and targeting to the tumor were achieved, however no anticancer effects were observed. We hypothesized further targeting of Salmonella could be achieved using chemotaxis, the coordination of flagellar driven movement with sensing environmental chemical gradients, akin to the nose of the bacterium. We discovered strain VNP20009 to be defective in chemotaxis, due to a genetic mutation that occurred during the strain’s construction. We were able to restore chemotaxis of the strain, at least partially, and discovered we could not further optimize chemotaxis without compromising the safety profile of VNP20009. We tested the effect of chemotaxis on tumor colonization in a mouse breast cancer model and found that the bacteria had an additive effect in causing liver disease and morbidity of the mice. We finally examined genome-wide gene expression changes occurring in the tumor microenvironment, as a response to anticancer agent VNP20009 colonization in a mouse melanoma model of cancer. Overall, this work contributes significantly to the understanding of VNP20009 chemotaxis and its tumor targeting abilities.

In the twenty-first century, advances in synthetic biology and molecular tools to implement programmable behavior into microbes have fueled significant efforts to develop microbial-based therapeutics. Bacteria and viruses have been explored independently for their ability to replicate and induce cytotoxic effects in cancer cells selectively. This dissertation aims to co-opt the anti-tumor capabilities of gram-negative Salmonella enterica subspecies enterica serotype Typhimurium (referred to as Salmonella typhimurium moving forward) and picornaviruses (small RNA viruses with positive sense genomes) to develop a potent, single bacterial-viral consortium- based system to treat solid tumors.I first describe our efforts to co-opt S. typhimurium’s natural internalization into hosts and intracellular space-sensing to deliver self-amplifying picornaviral RNA. Protein effectors that promote intracellular survival of S. typhimurium within the Salmonella-Containing-Vacuole (SCV) are transcribed by Salmonella Pathogenicity Island-2 (SPI-2) promoters, which turn on after sensing the intracellular pH, ion concentrations, and oxidative stressors. These effectors are then translocated into the host’s cytoplasm by a needle apparatus that connects the SCV and cytoplasm, which is also transcribed by SPI-2 promoters. By using the SPI-2 promoter PsseA to drive the expression of fluorescent reporters and membrane-disrupting proteins (eukaryotic and prokaryotic), efficient escape of Salmonella-produced proteins into tumor-host cells was established. RNA delivery into host cells was also made possible by a secondary SPI-2 promoter, PsseJ, which transcribes RNA polymerase T7 (T7), which then transcribes a T7-promoter-driven Poliovirus replicon or full-length Senecavirus A (SVA). Inoculation of this engineered S. typhimurium strain on a panel of cancer cell lines identified the system’s ability to deliver viral replicons and full-length viruses in a small cell cancer cell line, H446. In a murine model, S. typhimurium delivery of SVA was then shown to clear xenografted H446 tumors. Motivated by the possibility of delivering other picornaviral species with similar anti-tumor properties, but documented healthy tissue cytotoxicity, S. typhimurium was further engineered to control SVA viral spread. By driving Tobacco Etch Virus (TEV) protease expression via a second SseA promoter, and replacing a natural cleavage site on SVA with the TEV-cleavage domain, we demonstrate TEV-dependent SVA spread in H446 cells. I conclude with efforts on engineering TEV-dependent-SVA transgene expression to confer greater antitumor properties. Interferon-gamma and granulocyte-macrophage colony-stimulating factor (GM-CSF) have been reported to attenuate H446 growth in vitro. Expression of interferon-gamma off SVA would produce a direct selective pressure against viral replication and virion production. However, a fusion of human GM-CSF to Nano-Luciferase protein on the TEV-dependent SVA genome maintained luminescent signals, GM-CSF activity, and TEV-dependent spread, providing a framework to survey anti-tumor properties of SVA-transgenes. Furthermore, I address our development of syngeneic models for Salmonella-mediated delivery of SVA, an important step towards clinical applications of the system as immunocompetent models more closely correlate to immunocompetent patient populations. SVA’s efficient entry and replication in neuroendocrine-derived tissue identified murine neuroblastoma N1E-115 cells as a suitable cell line for SVA cytotoxicity studies. However, the ability of these cells to support bacterial-viral superinfections is unknown. Here, we show that Salmonella-mediated launch of SVA leads to viral spread that can attenuate heterologous hind flank tumor growth and improve their survival along with mice engrafted with orthotopic intracranial brain tumors.

Les séquences d'insertion sont des parasites moléculaires d'ADN codant uniquement pour leur machinerie de transposition et sont retrouvées majoritairement dans les génomes et les plasmides des procaryotes. Le séquençage génomique massif de la dernière décennie nous a permis de détecter chez des souches bactériennes cliniques plusieurs gènes de résistance aux antibiotiques associés aux séquences d'insertion sous forme de transposons composés. Il a été suggéré que les séquences d'insertion jouent un rôle prépondérant dans l'évolution bactérienne et qu'elles possèdent un pouvoir recruteur permettant de sélectionner, réorganiser et propager des gènes conférant un avantage sélectif à l'hôte. Nous avons partiellement caractérisé le potentiel recruteur de deux séquences d'insertion associées à des gènes de résistance, la séquence IS26 de Proteus vulgaris et la séquence ISJO de Salmonella typhimurium, en observant les étapes de recrutement d'un gène chromosomique et leurs distributions épidémiologiques en consultant les banques de données. / Insertion sequences are DNA molecular parasites encoding exclusively their transposition function, and are mainly found within the genomes and plasmids of prokaryotic organisms. The massive genomic sequencing we have witnessed in the last decade has allowed us to detect in clinical bacterial isolates many antibiotic resistance genes associated with insertion sequences in the form of composite transposons. It has been suggested that insertion sequences may act as a primary modulator of bacterial evolution, and that they possess a recruitment capacity allowing them to select, reorganize and spread genes encoding adaptation functions for their hosts. We have partially characterized the recruitment potential of two insertion sequences well known for their association with antibiotic resistance genes, IS10 from Salmonella typhimurium and 1S26 from Proteus vulgaris, by observing the steps involved in the recruitment process of a chromosomal gene and their epidemiological distribution by consulting various databanks.

La pollution engendrée par les pratiques d'élevage constitue une préoccupation majeure dans beaucoup de pays. Il a été notamment démontré que les fèces animales logent des pathogènes potentiellement nocifs, dont le Escherichia coli 0157:H7 et des espèces du genre Salmonella. Ainsi, le recyclage des résidus organiques qui n'ont pas été préalablement soumis à un traitement adéquat pose un risque élevé de contamination de l'environnement, du sol et des eaux souterraines. L'élimination des pathogènes par les techniques de biopasteurisation s'avère un moyen prometteur pour la gestion des résidus animaux et en particulier du fumier solide. Bien que chacune des techniques existantes possèdent des avantages, il y a toutefois toujours place à l'amélioration. Dans cette étude, nous avons présenté un nouveau système de traitement en continu par biopasteurisation appelé SHOCMD (Système d'Hygiénisation en Oxygénation Contrôlée), et son efficacité à éliminer les bactéries pathogènes. Les résultats ont été comparés à ceux d'un système conventionnel, soit la cellule de compostage à aération forcée. Afin de vérifier l'efficacité du SHOCMD , deux pathogènes majeurs, soit E, coli 0157:H7 et Salmonella enterica subsp. enterica serovar Typhimurium, ont été introduits dans le fumier solide de porc. À la fin du traitement, les échantillons ont été analysés en utilisant des techniques microbiologiques et moléculaires. De plus, la diversité bactérienne, un paramètre important pour une gestion efficace de la biopasteurisation, a été déterminée à l'aide de techniques moléculaire. Les résultats ont démontré que le SHOCMD est un système efficace pour l'hygiénisation du fumier solide et que le produit final répond aux normes de sécurité concernant les pathogènes. / Pollution by livestock wastes has become a great concern in many countries. It is well documented that potentially harmful pathogens including Escherichia coli 0157:117 and Salmonella spp. are shed in animal feces. Recycling of these wastes without treatment poses a risk of contamination of the environment, soil and groundwater. Bio-pasteurization is a promising technique for management of solid animal wastes especially manure. However, even if each method has its own advantages there is still room for improvement. In this study, we present a new bio-pasteurization system called SHOCIM (Système d'Hygiénisation en Oxygénation Contrôlée) and the efficiency of the SHOCIM to eliminate pathogens was compared to the Cell, which is one of the conventional Systems. To evaluate the efficiency of SHOC™, two majors pathogens, E. coli 0157:H7 and Salmonella enterica subsp. enterica serovar Typhimurium were introduced in manure. At the end of treatment, the samples were analyzed with both microbiological and molecular techniques. Bacterial diversity which is important in the effective management of biopasteurization process was monitored by molecular techniques as well. Results demonstrated that SHOCIM is an efficient facility for manure treatment and the final product meet the standards for pathogens safety.

Au cours de ce travail, différents glucosylamines et aminodésoxyglucoses ont été synthétisés et caractérisés par différentes méthodes spectroscopiques dont l’IRTF, la RMN 1H, 13C et MALDI-Tof MS. L’étude des propriétés biologiques de ces molécules réalisée, d’une part, avec deux champignons du bois, Coriolus versicolor et Poria placenta, et d’autre part, avec trois microorganismes potentiellement rencontrés dans des aliments, Listeria innocua, Salmonella typhimurium et Fusarium proliferatum ont indiqué une contribution positive de la N-alkylation, du degré de N-substitution et de la quaternisation sur l’inhibition de leur croissance. Par ailleurs, l’impact sur la bioactivité, de la position du groupe amine sur le sucre, a été étudié. Il a été montré que la position du groupe amine sur le C-1 du glucose conduisait à une activité antifongique contre C. versicolor et P. placenta plus prononcée alors que la position C-3 du glucose était favorable à une activité antimicrobienne contre L. innocua et S. typhimurium. / In this study different glucosylamines and amino desoxyglucoses were synthesized and characterised using various spectroscopic methods including IRFT, both 1H and 13C NMR spectroscopy and MALDI-Tof MS. Biological assessment of these compounds realised with two wood decay fungi, Coriolus versicolor and Poria placenta on one hand, and with three food microorganisms Listeria innocua, Salmonella typhimurium and Fusarium proliferatum on other hand, indicated a positive impact of both N-alkylation and degree of N-substitution and quaternisation on their growth inhibition. Furthermore, a biological impact of the amine position on sugar was studied. It was found that amine function attached to the C-1 of glucose conducted to the best antifungal activity against both C. versicolor and P. placenta while that fixed on the C-3 of glucose was indicated for antibacterial activity against L. innocua and S. typhimurium.