<p>It is known that antibiotics play a role in clearing bacterial infections, and sometimes yeast infections. Antibiotics can either kill or inhibit bacteria in many different ways, but they also effect the role white blood cells play in an infection. Some will enhance white blood cell effects, while other will inhibit. How antibiotics do this is not well understood. Azithromycin and daptomycin are two different antibiotics. Very little is known on how they affect the immune response, including phagocytosis, delayed type hypersensitivity (DTH), and cytokine release. Murine macrophages and neutrophils were treated with azithromycin or daptomycin, along with yeast, Candida lusitaniae, and plated to see the effects of the antibiotic on macrophage and neutrophil phagocytosis. Mice were also sensitized to dinitrofluorobenzene and given azithromycin or daptomycin to see how the antibiotics effect DTH. Lastly, an ELISA was done to see how much of the cytokine, IL-6, the macrophages treated with the antibiotics produced compared to the untreated macrophages. It was seen that both azithromycin and daptomycin have significant effects on phagocytosis and DTH. More research is needed to see the effects of the antibiotics on IL-6 release, as well as their effects on other aspects of the immune response.

<p>The complex interactions between diet and the microbiota that influence mucosal inflammation and inflammatory bowel disease (IBD) are poorly understood. Experimental colitis models provide the opportunity to control and systematically perturb diet and the microbiota in parallel to quantify the contributions between multiple dietary ingredients and the microbiota on host physiology and colitis. To examine the interplay of diet and the gut microbiota on host health and colitis, we fed over 40 different diets with varied macronutrient sources and concentrations to specific pathogen free or germ-free mice either in the context of healthy, unchallenged animals or the dextran sodium sulfate (DSS) colitis model with follow-up studies for 4 diets in the T cell transfer colitis model. Diet influenced physiology in both health and colitis across all models, with the concentration of protein and psyllium fiber having the most profound effects. Increasing dietary protein elevated gut microbial density and worsened DSS colitis severity. Depleting gut microbial density by using germ-free animals or antibiotics negated the effect of a high protein diet. Psyllium fiber influenced host physiology and attenuated colitis severity through microbiota-dependent and microbiota-independent mechanisms. Combinatorial perturbations to dietary protein and psyllium fiber in parallel explain most variation in gut microbial density, intestinal permeability, and DSS colitis severity, and changes in one ingredient can be offset by changes in the other. Our results demonstrate the importance of examining complex mixtures of nutrients to understand the role of diet in intestinal inflammation.

<p> Viral invasion of the central nervous system (CNS) is a significant cause of morbidity and mortlity worldwide, particularly in young children (1). The nervous system presents a challenging site for viruses to access, with multiple physical and immunological barriers that limit pathogen invasion. To invade the CNS, viruses must access cell-surface receptors for binding and entry events. Virus-receptor interactions also govern tropism and often control disease type and severity. For many viruses, the identities of receptors and other cellular determinants of viral tropism remain elusive. Understanding where and how viral capsid components engage neural receptors and the effect of these interactions on tropism and disease may illuminate targets to prevent viral neuroinvasion.</p><p> Mammalian orthoreoviruses (reoviruses) provide a highly tractable and well-established system to identify mechanisms of viral entry into the CNS. Reoviruses are non-enveloped particles containing a 10-segmented, double-stranded (ds) RNA genome that replicate well in culture and can be altered via a robust reverse-genetics system (2, 3). While reovirus causes similar age-restricted disease in many young mammals (4-6), most studies employ newborn mice. Following peroral or intracranial inoculation of newborn mice, reovirus displays serotype-specific patterns of tropism in the brain and concomitant disease (<b>Fig. I-1</b>). Serotype 1 (T1) strains infect ependymal cells lining the ventricles of the brain and cause a non-lethal hydrocephalus (7). In contrast, serotype 3 (T3) strains infect specific neuron populations in the CNS and produce a fulminant, and often lethal, encephalitis (8). These differences in tropism and disease have been genetically mapped to the reovirus S1 gene using single-gene reassortant viruses (9). However, viral and host gene sequences that mediate either T1 or T3 tropism have not been defined. </p><p> In Chapter I of my dissertation, I introduce key themes about mechanisms of neuroinvasion and the disease consequences of CNS infection. I describe fundamental knowledge and open areas of research pertaining to reovirus infection in the CNS and expand on reovirus-receptor interactions. I conclude Chapter I with a summary of viral oncolytic therapies and highlight strengths and opportunities for improvement of reovirus oncolytics. In Chapter II, I describe the design and implementation of &sigma;1- chimeric reoviruses to identify sequences in the S1 gene that dictate neurotropism and virulence in the CNS. In these studies, I found that homologous sequences at the viriondistal end of the viral attachment protein are responsible for neuron and ependymal cell targeting. In Chapter III, I identify sequences of the NgR1 reovirus receptor that are required for binding and post-binding functions and elucidate the viral ligand for NgR1, which is the &sigma;3 outer-capsid protein, using a combination of genetic, biochemical, and structural approaches. Finally, in Chapter IV, I review conclusions from results presented in Chapters II and III, examine new questions raised by these studies, and discuss future directions of this work. Collectively, my dissertation research has unveiled viral and host sequences that contribute to neural cell targeting and will improve strategies and knowledge to design targeted oncolytic therapies.</p><p>

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Giardia lamblia undergoes antigenic variation of variant-specific surface proteins (VSPs) that are encoded by a family of ~150 vsp genes only one of which is expressed at a time. The vsp gene promoters have not been previously studied. A comparison of the upstream non-coding region of vsp genes shows that they lack the AT-rich regions found in other Giardia gene promoters. We have determined that the core promoters of vsp A6 and vsp C5 genes extend from -57 to + 6 and -50 to + 6 respectively. Through linker scanning analysis, we have also identified regions within the vsp A6 core promoter important for promoter activity that span -7-3, -12-8, -17-13 and -42-38.There is no sequence similarity in the upstream regions of the previously characterized vsp genes that were analyzed, with the exception of a seven nucleotide region that encompasses the translation initiation site: Py A A T G T T. We have demonstrated that the four nucleotides flanking the start codon are essential for promoter activity. This result suggests that it may be an Inr element, which by definition determines the site of transcription initiation. In addition, this element loosely resembles the metazoan Inr consensus: Py Py A A/T Py Py. Using 5&#8242; RACE I have determined that for two vsp genes, the translation and transcription start sites are synonymous and reside within this conserved element. However, we were unable to identify protein factors that bind this region using electrophoretic mobility shift assays.A search for characteristic VSP motifs, such as CRGKA, amongst identified ORFs in the Giardia genome assembly in turn identified 180 ORFs which may be VSPs. Eighty one of these are found within contigs while 99 of these are found at contig and 80 ORFs have the Inr element identified in this study.This study supports the hypothesis that longer upstream non-coding regions of vsp genes play a role in regulating the expression of these genes and hence antigenic variation in G. lamblia.

Interest in the study of antiviral substances stems from the fact that classical chemotherapy as it is known in bacterial diseases has thus far not been successful in the field of viral diseases. However, any proven antiviral agent is a potential drug against virus infection. Although the ultimate goal of effective chemotherapy of virus infections may or may not be attained, information concerning such substances may provide clues to the nature of host-virus interactions and help to elucidate the patterns of the pathogenesis of virus diseases (Zimmermann and Schafer, 1960; Melnick and Rapp, 1965) and also to afford tools for the classification of viruses (Tamm and Eggers, 1962; Barry et al., 1962). [...]

The Citrobacter rodentium genome sequence contains a phoPQ operonhomologous (~79% identity) to that of S. typhimurium. We report that C. rodentiumPhoQ senses fluctuations in Mg2+ concentrations and acidic pH. Surprisingly, PhoQwas not activated by the presence of AMPs. However, activation by AMPs is observedwhen C. rodentium PhoP/PhoQ was expressed in as. typhimurium background. Weidentified an outer membrane protease of the omptin family that was responsible forinhibiting PhoQ activation by AMPs. In stark contrast to S. typhimurium, which relieson LPS modifications to resist AMPs, our results suggest that C. rodentium promotesresistance through a PhoP/PhoQ-dependent OM protease to inhibit disruption of theouter membrane by AMPs . / La séquence du génome de Citrobacter rodentium présente un opéron phoPQ(~79% identité) homologue à celui de S. typhimurium. Nous avons déterminé quePhoQ de C. rodentium perçoit les variations de pH et en Mg2+ du milieu environnant.De manière surprenante, les PAMs ne causent aucune augmentation d'activité dePhoQ. Néeanmoins, lorsque le système PhoP/PhoQ de C. rodentium est exprimé chezS. typhimurium les PAMs activent PhoQ. Nous avons identifié une protéine de lamembrane externe appartenant à la famille des omptin qui est responsable del'inactivité de PhoQ en présence des P AMs. Ces résultats suggèrent que le mécanismede résistance aux PAMs de C. rodentium serait régulé par le système PhoP/PhoQ et une protéase qui empêcherait la destruction de la membrane externe par les P AMs. Cemécanisme de défense est différent de celui du système PhoP/PhoQ de S. typhimuriumqui repose essentiellement sur des modification du LPS .