A pseudomolecule assembly of the Rocky Mountain elk genome

Rocky Mountain elk (Cervus canadensis) populations have significant economic implications to the cattle industry, as they are a major reservoir for Brucella abortus in the Greater Yellowstone area. Vaccination attempts against intracellular bacterial diseases in elk populations have not been successful due to a negligible adaptive cellular immune response. A lack of genomic resources has impeded attempts to better understand why vaccination does not induce protective immunity. To overcome this limitation, PacBio, Illumina, and Hi-C sequencing with a total of 686-fold coverage was used to assemble the elk genome into 35 pseudomolecules. A robust gene annotation was generated resulting in 18,013 gene models and 33,422 mRNAs. The accuracy of the assembly was assessed using synteny to the red deer and cattle genomes identifying several chromosomal rearrangements, fusions and fissions. Because this genome assembly and annotation provide a foundation for genome-enabled exploration of Cervus species, we demonstrate its utility by exploring the conservation of immune system-related genes. We conclude by comparing cattle immune system-related genes to the elk genome, revealing eight putative gene losses in elk

In vitro and in vivo evaluation of novel biodegradable polymer adjuvants for vaccine delivery

Infectious disease remains a constant threat to the health of man and his animals. Vaccination has been declared one of the medical triumphs of the twentieth century. For man or animal, vaccination remains the best and most cost effective means for the prevention of disease. Many novel vaccine antigens are rationally designed peptides and recombinant proteins which require the use of adjuvants or other immune enhancers to increase efficacy. Currently, there is a need not only for single dose vaccines (to improve patient compliance and improve animal welfare by reducing livestock handling) but also adjuvants that preserve the immunogenicity of the protein during encapsulation, storage and release and enhance the host's immune response to the antigen. Biodegradable polyanhydrides have shown many characteristics that fulfill these ideals but further study is needed. The studies presented in this dissertation were undertaken with the intent to define the interaction(s) between novel biodegradable polyanhydride microspheres and the host immune system. In order to address the role of polyanhydride chemistry on murine dendritic cells (DCs) in vitro, DC activation by polyanhydride microspheres was evaluated by surface marker expression and cytokine secretion. Several murine models, including a transgenic T cell transfer model, were used to evaluate the induction of antigen-specific immune response by immunizing mice with microsphere adjuvanted ovalbumin. The in vivo studies using ovalbumin encapsulated microspheres were carried out in three mouse strains to evaluate the memory or recall response induced by a single microsphere vaccination and to evaluate strain differences in response to the polyanhydride microspheres. Finally, microspheres loaded with the protease digested vaccine antigen derived from Brachyspira hyodysenteriae was used to vaccinate mice and pigs prior to disease challenge studies designed to evaluate the induction of protective immunity. Taken together, this body of work further adds to our knowledge of polyanhydride microspheres and their potential use as vaccine carriers.</p

Digital Dermatitis in Cattle: Current Bacterial and Immunological Findings

Globally; digital dermatitis is a leading form of lameness observed in production dairy cattle. While the precise etiology remains to be determined; the disease is clearly associated with infection by numerous species of treponemes; in addition to other anaerobic bacteria. The goal of this review article is to provide an overview of the current literature; focusing on discussion of the polybacterial nature of the digital dermatitis disease complex and host immune response. Several phylotypes of treponemes have been identified; some of which correlate with location in the lesion and some with stages of lesion development. Local innate immune responses may contribute to the proliferative, inflammatory conditions that perpetuate digital dermatitis lesions. While serum antibody is produced to bacterial antigens in the lesions, little is known about cellular-based immunity. Studies are still required to delineate the pathogenic traits of treponemes associated with digital dermatitis; and other host factors that mediate pathology and protection of digital dermatitis lesions

Digital Dermatitis in Cattle: Current Bacterial and Immunological Findings

Globally; digital dermatitis is a leading form of lameness observed in production dairy cattle. While the precise etiology remains to be determined; the disease is clearly associated with infection by numerous species of treponemes; in addition to other anaerobic bacteria. The goal of this review article is to provide an overview of the current literature; focusing on discussion of the polybacterial nature of the digital dermatitis disease complex and host immune response. Several phylotypes of treponemes have been identified; some of which correlate with location in the lesion and some with stages of lesion development. Local innate immune responses may contribute to the proliferative, inflammatory conditions that perpetuate digital dermatitis lesions. While serum antibody is produced to bacterial antigens in the lesions, little is known about cellular-based immunity. Studies are still required to delineate the pathogenic traits of treponemes associated with digital dermatitis; and other host factors that mediate pathology and protection of digital dermatitis lesions

First Molecular Confirmation of Treponema spp. in Lesions Consistent with Digital Dermatitis in Chilean Dairy Cattle

Digital dermatitis (DD) is a highly contagious and infectious disease in cattle which has a considerable negative economic impact worldwide, and adversely affects animal welfare. Members of the genus Treponema are the only bacterial agents for which there is consistent evidence of participation in DD lesions. In Chile, DD has been described since the 1990s, but only under a clinical approach. To date, the presence of the pathogenic agent has not been confirmed in Chile by any type of confirmatory microbiological diagnosis. The aim of the present study was to detect the presence of Treponema spp. DNA in lesions consistent with DD, in Chilean dairy cattle for the first time. We provide PCR confirmation of Treponema spp. in Chilean dairy cattle affected by DD. The high rate of positive results, as well as the proportion of the main Treponema species involved, is in line with what have been described in published studies elsewhere. Future herd control plans should benefit from the molecular detection of these pathogenic bacteria associated with DD

Inbred Rats as a Model to Study Persistent Renal Leptospirosis and Associated Cellular Immune Responsiveness

Pathogenic species of Leptospira cause leptospirosis, a bacterial zoonotic disease with a global distribution affecting over one million people annually. Rats are regarded as one of the most significant reservoir hosts of infection for human disease, and in the absence of clinical signs of infection, excrete large numbers of organisms in their urine. A unique biological equilibrium exists between pathogenic leptospires and reservoir hosts of infection, but surprisingly, little is known concerning the host's cellular immune response that facilitates persistent renal colonization. To address this deficiency, we established and applied an immunocompetent inbred rat model of persistent renal colonization; leptospires were detected in urine of experimentally infected rats by 3 weeks post-infection and remained positive until 8 weeks post-infection. However, there was little, if any, evidence of inflammation in colonized renal tubules. At 8 weeks post-infection, a robust antibody response was detected against lipopolysaccharide and protein outer membrane (OM) components. Purified B and T cells derived from the spleen of infected and non-infected rats proliferated in response to stimulation with 0.5 μg of OM fractions of Leptospira, including CD4+ T cells, which comprised 40% of proliferating cells, compared to 25% in non-infected controls. However, analysis of gene expression did not determine which immunoregulatory pathways were activated. Lymphocytes purified from the lymph node draining the site of colonization, the renal lymph node, also showed an increase in percentage of proliferating B and T cells. However, in contrast to a phenotype of 40% CD4+ T cells in the spleen, the phenotype of proliferating T cells in the renal lymph node comprised 65% CD4+ T cells. These results confirm that the renal lymph node, the local lymphoid organ, is a dominant site containing Leptospira reactive CD4+ T cells and highlight the need to consider the local, vs. systemic, immune responses during renal colonization infection. The use of inbred immunocompetent rats provides a novel tool to further elucidate those pathophysiological pathways that facilitate the unique biological equilibrium observed in reservoir hosts of leptospirosis

Interaction of Bovine Peripheral Blood Polymorphonuclear Cells and Leptospira Species; Innate Responses in the Natural Bovine Reservoir Host

Cattle are the reservoir hosts of Leptospira borgpetersenii serovar Hardjo, and can also be reservoir hosts of other Leptospira species such as L. kirschneri, and L. interrogans. As a reservoir host, cattle shed Leptospira, infecting other animals, including humans. Previous studies with human and murine neutrophils have shown activation of neutrophil extracellular trap or NET formation, and upregulation of inflammatory mediators by neutrophils in the presence of Leptospira. Humans, companion animals and most widely studied models of Leptospirosis are of acute infection, hallmarked by systemic inflammatory response, neutrophilia and septicemia. In contrast, cattle exhibit chronic infection with few outward clinical signs aside from reproductive failure. Taking into consideration that there is host species variation in innate immunity, especially in pathogen recognition and response, the interaction of bovine peripheral blood polymorphonuclear cells (PMNs) and several Leptospira strains was evaluated. Studies including bovine-adapted strains, human pathogen strains, a saprophyte and inactivated organisms. Incubation of PMNs with Leptospira did induce slight activation of neutrophil NETs, greater than unstimulated cells but less than the quantity from E. coli P4 stimulated PMNs. Very low but significant from non-stimulated, levels of reactive oxygen peroxides were produced in the presence of all Leptospira strains and E. coli P4. Similarly, significant levels of reactive nitrogen intermediaries (NO2) was produced from PMNs when incubated with the Leptospira strains and greater quantities in the presence of E. coli P4. PMNs incubated with Leptospira induced RNA transcripts of IL-1β, MIP-1α, and TNF-α, with greater amounts induced by live organisms when compared to heat-inactivated leptospires. Transcript for inflammatory cytokine IL-8 was also induced, at similar levels regardless of Leptospira strain or viability. However, incubation of Leptospira strains with bovine PMNs did not affect Leptospira viability as measured by limiting dilution culture. This is in contrast to previously reported results of innate inflammatory activation by Leptospira in human and other animal models, or the activation and interaction of bovine PMNs with Escherichia coli and other bacterial pathogens. While it could be hypothesized that variations in innate receptor recognition, specifically variance in toll-like receptor 2, could underlie the observed reduction of activation in bovin

Vaccine Adjuvants: Current Challenges and Future Approaches

For humans, companion animals, and food producing animals, vaccination has been touted as the most successful medical intervention for the prevention of disease in the twentieth century. However, vaccination is not without problems. With the development of new and less reactogenic vaccine antigens, which take advantage of molecular recombinant technologies, also comes the need for more effective adjuvants that will facilitate the induction of adaptive immune responses. Furthermore, current vaccine adjuvants are successful at generating humoral or antibody mediated protection but many diseases currently plaguing humans and animals, such as tuberculosis and malaria, require cell mediated immunity for adequate protection. A comprehensive discussion is presented of current vaccine adjuvants, their effects on the induction of immune responses, and vaccine adjuvants that have shown promise in recent literature.This is a manuscript of an article published as Wilson-Welder, Jennifer H., Maria P. Torres, Matt J. Kipper, Surya K. Mallapragada, Michael J. Wannemuehler, and Balaji Narasimhan. "Vaccine adjuvants: current challenges and future approaches." Journal of Pharmaceutical Sciences 98, no. 4 (2009): 1278-1316. DOI: 10.1002/jps.21523. Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0). Copyright 2008 Wiley-Liss, Inc. and the American Pharmacists Association. Posted with permission

Polyanhydride microparticles enhance dendritic cell antigen presentation and activation

The present studies were designed to evaluate the adjuvant activity of polyanhydride microparticles prepared in the absence of additional stabilizers, excipients, or immune modulators. Microparticles composed of varying ratios of either 1,6-bis(p-carboxyphenoxy)hexane (CPH) and sebacic acid (SA) or 1,8-bis(p-carboxyphenoxy)-3,6-dioxaoctane (CPTEG) and CPH were added to in vitro cultures of bone marrow-derived dendritic cells (DCs). Microparticles were efficiently and rapidly phagocytosed by DCs in the absence of opsonization and without centrifugation or agitation. Within 2 h, internalized particles were rapidly localized to an acidic, phagolysosomal compartment. By 48 h, only a minor reduction in microparticle size was observed in the phagolysosomal compartment, indicating minimal particle erosion consistent with being localized within an intracellular microenvironment favoring particle stability. Polyanhydride microparticles increased DC surface expression of MHC II, the co-stimulatory molecules CD86 and CD40, and the C-type lectin CIRE (murine DC-SIGN; CD209). In addition, microparticle stimulation of DCs also enhanced secretion of the cytokines IL-12p40 and IL-6, a phenomenon found to be dependent on polymer chemistry. DCs cultured with polyanhydride microparticles and ovalbumin induced polymer chemistry-dependent antigen-specific proliferation of both CD4+ OT-II and CD8+ OT-I T cells. These data indicate that polyanhydride particles can be tailored to take advantage of the potential plasticity of the immune response, resulting in the ability to induce immune protection against many types of pathogens