Project description:Listeria monocytogenes is a foodborne intracellular bacterial pathogen leading to human listeriosis. Despite a high mortality rate and increasing antibiotic resistance no clinically approved vaccine against Listeria is available. To identify antigens for this bacterial pathogen that can be encoded in mRNA vaccine formulations, we screened for Listeria epitopes presented on the surface of infected human cell lines by mass spectrometry-based immunopeptidomics. In between more than 15,000 human self-peptides, we detected 68 Listeria epitopes from 42 different bacterial proteins, including several known antigens. Peptide epitopes presented on different cell lines were often derived from the same bacterial surface proteins, classifying these antigens as potential vaccine candidates. Encoding these highly presented antigens in lipid nanoparticle mRNA vaccine formulations resulted in high levels of protection in vaccination challenge experiments in mice. Our results pave the way for the development of a clinical mRNA vaccine against Listeria and demonstrate the power of immunopeptidomics for next-generation bacterial vaccine development.
Project description:Mycoplasma hyopneumoniae, the causative agent of swine enzootic pneumonia, colonizes the cilia of swine lungs, causing ciliostasis and cell death. Mycoplasma hyopneumoniae is a component of the porcine respiratory disease complex (PRDC) and is especially problematic for the finishing swine industry, causing the loss of hundreds of millions of dollars in farm revenues worldwide. For successful infection, M. hyopneumoniae must effectively resist oxidative stresses due to the release of oxidative compounds from neutrophils and macrophages during the host’s immune response. However, the mechanism M. hyopneumoniae uses to avert the host response is still unclear. To gain a better understanding of the transcriptional responses of M. hyopneumoniae under oxidative stress, cultures were grown to early exponential phase and exposed to 0.5% percent hydrogen peroxide for 15 minutes. RNA samples from these cultures were collected and compared to RNA samples from control cultures using two-color PCR-based M. hyopneumoniae microarrays. This study revealed significant down-regulation of important glycolytic pathway genes and gene transcription proteins, as well as a protein known to activate oxidative stressor cascades in neutrophils. This study has also contained significantly differentially expressed genes common to other environmental stress responses, and merits further study of universal stress response genes of M. hyopneumoniae. Keywords: Mycoplasma hyopneumoniae, RNA microarray
Project description:This study investigated the ability of two novel adjuvant formulations, QCDC (Quil A/cholesterol/DDA/Carbopol) and QCDCR (QCDC/Bay R1005), in combination with a recombinant profilin vaccine, to modulate host protective immunity and to alter new gene expression during experimental avian coccidiosis.
Project description:This study investigated the ability of two novel adjuvant formulations, QCDC (Quil A/cholesterol/DDA/Carbopol) and QCDCR (QCDC/Bay R1005), in combination with a recombinant profilin vaccine, to modulate host protective immunity and to alter new gene expression during experimental avian coccidiosis. Four-condition experiment, Profilin only vs. Non-vaccination, Profilin only vs. Co-vaccination of QCDC plus profilin, Profilin only vs. Co-vaccination of QCDCR plus profilin, Biological replicates: 2 profilin only replicates, 2 Non-vaccination replicates, 2 QCDC plus profilin replicates, 2 QCDCR plus profilin replicates with dye-switching.
Project description:Mycoplasma hyopneumoniae is the causative agent of porcine enzootic pneumonia and a major factor in the porcine respiratory disease complex. A clear understanding of the mechanisms of pathogenesis does not exist. The virulence factors of M. hyopneumoniae are largely unknown and are most probably complex in nature. The transcriptional profile of intergenic regions is investigated using microarray PCR and oligonucleotide probes. It is hypothesized that intergenic regions of the Mhyo genome are being transcribed along with the ORFs.
Project description:Mycoplasma hyopneumoniae is the causative agent of porcine enzootic pneumonia and a major factor in the porcine respiratory disease complex. A clear understanding of the mechanisms of pathogenesis does not exist although it is clear that M. hyopneumoniae adheres to porcine ciliated epithelium by action of a protein called P97. Previous studies have shown variation in the gene encoding the P97cilium adhesin within different strains of M. hyopneumoniae, but the extent of genetic variation among field strains across the genome is not known. Since M. hyopneumoniae is a worldwide problem, it is reasonable to expect that a wide range of genetic variability may exist given all of the different breed and housing conditions. This variation may impact the overall virulence of a single strain. Using microarray technology, this study examined potential variation of fourteen field strains in comparison to strain 232 on which the array was based. Genomic DNA was obtained, amplified with TempliPhi™, and labeled indirectly with Alexa dyes. Post genomic hybridization, the arrays were scanned and data analyzed using a linear statistical model. Results indicate that genetic variation could be detected in all fourteen field strains but across different loci, suggesting that variation occurs throughout the genome. Fifty-nine percent of the variable loci were hypothetical genes. Twenty-two percent of the lipoprotein genes showed variation in at least one field strain. A permutation test identified a location in M. hyopneumoniae genome where spatial clustering of variability between the field strains and strain 232 exists. Keywords: CGH, Mycoplasma Hyopneumoniae
Project description:Mycoplasma hyopneumoniae pathogenic strains, like 7448, are the causative agents of porcine enzootic pneumonia. Non-pathogenic strains, like M. hyopneumoniae J, does not cause disease, although shares the entire repertoire of known virulence-related genes with M. hyopneumoniae 7448. In this context, the differential expression of ortholog genes is likely responsible, at least in part, for differences in pathogenicity or virulence level between these strains. Moreover, in the porcine lung, M. hyopneumoniae faces a hostile environment, with both oxidative and heat stresses. The performed comparative proteomics analyses provided evidence of differential stress responses between a pathogenic and a non-pathogenic M. hyopneumoniae strain, involving tens of proteins, including some known virulence factors. The results suggest that stress conditions trigger the expression of potential virulence factors in the pathogenic M. hyopneumoniae 7448, but not in the non-pathogenic M. hyopneumoniae J.
Project description:To development of our gene expression, we have employed whole rhesus monkey genome microarray expression profiling as a discovery platform to identify genes with the potential to induce immune response. The peripheral blood from rhesus macaques, who were immunized in groups of three with the capsular polysaccharide (CPS antigen), carrier protein tetanus toxoid (TT) or conjugate vaccine via intramuscular injection (i.m.) in the anterolateral thigh on days 0, 30 and 60 using the formulations, were obtained on days 0, 30, 60 and 90. PBMCs were collected for microarray assays. Dynamic expression variations of eight genes (KLRC1, LGALS13, LTB4DH, NUAK1, VNN2, GALNT3, LOC710050 and LOC716305) were maintained in Hib conjugate vaccine group throughout experiment comparing with the CPS antigen group and carrier protein TT group.
Project description:The COVID-19 pandemic has generated intense interest in the rapid development and evaluation of vaccine candidates for this disease and other emerging diseases. Several novel methods for preparing vaccine candidates are currently undergoing clinical evaluation in response to the urgent need to prevent the spread of COVID-19. In many cases, these methods rely on new approaches for vaccine production and immune stimulation. We report on the use of a novel method (SolaVAXTM) for production of an inactivated vaccine candidate and the testing of that candidate in a hamster animal model for its ability to prevent infection upon challenge with SARS-CoV-2 virus. The studies employed in this work included an evaluation of the levels of neutralizing antibody produced post-vaccination, levels of specific antibody sub-types to RBD and spike protein that were generated, evaluation of viral shedding post-challenge, flow cytometric and single cell sequencing data on cellular fractions and histopathological evaluation of tissues post-challenge. The results from this study provide insight into the immunological responses occurring as a result of vaccination with the proposed vaccine candidate and the impact that adjuvant formulations, specifically developed to promote Th1 type immune responses, have on vaccine efficacy and protection against infection following challenge with live SARS-CoV-2. This data may have utility in the development of effective vaccine candidates broadly. Furthermore, the results suggest that preparation of a whole virion vaccine for COVID-19 using this specific photochemical method may have utility in the preparation of one such vaccine candidate.