Immunogenicity of Potomac horse fever vaccine when simultaneously co-administered with rabies vaccine in a multivalent vaccine or as two monovalent vaccines at separate sites.

BACKGROUND: Potomac horse fever (PHF) is a potentially fatal enterocolitis of horses caused by Neorickettsia risticii. The disease was originally recognised almost 40 years ago in the state of Maryland in the US. It is now known to occur in many areas of North America, as well as having been described in South America and Europe. Monocomponent PHF vaccines are available, but clinical protection with vaccination has been reported to be inconsistent. OBJECTIVES: This study was designed to assess the immunogenicity of a commercially available Potomac Horse Fever (PHF) vaccine when administered as either a monovalent PHF vaccine simultaneously co-administered with a separate monovalent Rabies vaccine or as a multivalent PHF/Rabies vaccine in horses. STUDY DESIGN: Randomised parallel group trial. METHODS: Ninety-one client or University owned horses participated in this open-label randomised study, with 45 horses receiving the monovalent vaccines at separate sites and 46 receiving the multivalent vaccine at a single site. Serum PHF IFA titres were determined twice prior to vaccination and at 1, 2 and 3 months after vaccination. RESULTS: Both vaccination protocols exhibited poor immunogenicity, with only one-third of all the animals demonstrating seroconversion, defined as an increase in titre of greater than 400 over baseline, at any time point after vaccination. The monovalent PHF vaccine exhibited significantly greater immunogenicity in terms of the number of horses exhibiting seroconversion, as compared to the multivalent vaccine, at one (20 vs. 11, P = 0.03) and two (18 vs. 9, p = 0.02) months post vaccination. The monovalent PHF vaccine also exhibited significantly greater immunogenicity in terms of the median (interquartile range) IFA titres, as compared to the multivalent vaccine, at one (800 [200-1600] vs. 400 [200-800], P = 0.009) and 2 months (400 [200-1600] vs. 400 [100-800], P = 0.02) post vaccination. There was no significant difference between groups at 3 months in either seroconversion rate or median IFA titers. MAIN LIMITATIONS: This study did not assess the actual protective effects of PHF vaccination but rather used the serologic response to vaccination as a surrogate biomarker of immunity. CONCLUSIONS: The multivalent PHF/Rabies vaccine exhibited lower immunogenicity as compared to the monovalent PHF vaccine co-administered with a separate Rabies vaccine.

Rickettsia rickettsii Whole-Cell Antigens Offer Protection against Rocky Mountain Spotted Fever in the Canine Host.

Rocky Mountain spotted fever (RMSF) is a potentially fatal tick-borne disease in people and dogs. RMSF is reported in the United States and several countries in North, Central, and South America. The causative agent of this disease, Rickettsia rickettsii, is transmitted by several species of ticks, including Dermacentor andersoni, Rhipicephalus sanguineus, and Amblyomma americanum RMSF clinical signs generally include fever, headache, nausea, vomiting, muscle pain, lack of appetite, and rash. If untreated, it can quickly progress into a life-threatening illness in people and dogs, with high fatality rates ranging from 30 to 80%. While RMSF has been known for over a century, recent epidemiological data suggest that the numbers of documented cases and the fatality rates remain high in people, particularly during the last two decades in parts of North America. Currently, there are no vaccines available to prevent RMSF in either dogs or people. In this study, we investigated the efficacies of two experimental vaccines, a subunit vaccine containing two recombinant outer membrane proteins as recombinant antigens (RCA) and a whole-cell inactivated antigen vaccine (WCA), in conferring protection against virulent R. rickettsii infection challenge in a newly established canine model for RMSF. Dogs vaccinated with WCA were protected from RMSF, whereas those receiving RCA developed disease similar to that of nonvaccinated R. rickettsii-infected dogs. WCA also reduced the pathogen loads to nearly undetected levels in the blood, lungs, liver, spleen, and brain and induced bacterial antigen-specific immune responses. This study provides the first evidence of the protective ability of WCA against RMSF in dogs.

Targeting prohibitins at the cell surface prevents Th17-mediated autoimmunity.

T helper (Th)17 cells represent a unique subset of CD4+ T cells and are vital for clearance of extracellular pathogens including bacteria and fungi. However, Th17 cells are also involved in orchestrating autoimmunity. By employing quantitative surface proteomics, we found that the evolutionarily conserved prohibitins (PHB1/2) are highly expressed on the surface of both murine and human Th17 cells. Increased expression of PHBs at the cell surface contributed to enhanced CRAF/MAPK activation in Th17 cells. Targeting surface-expressed PHBs on Th17 cells with ligands such as Vi polysaccharide (Typhim vaccine) inhibited CRAF-MAPK pathway, reduced interleukin (IL)-17 expression and ameliorated disease pathology with an increase in FOXP3+-expressing Tregs in an animal model for multiple sclerosis (MS). Interestingly, we detected a CD4+ T cell population with high PHB1 surface expression in blood samples from MS patients in comparison with age- and sex-matched healthy subjects. Our observations suggest a pivotal role for the PHB-CRAF-MAPK signalling axis in regulating the polarization and pathogenicity of Th17 cells and unveil druggable targets in autoimmune disorders such as MS.

Comparative Study of Two Droplet-Based Dissolving Microneedle Fabrication Methods for Skin Vaccination.

Dissolving microneedles (DMNs) have been widely studied in medical applications due to their pain-free administration, superior efficiency, and safe drug delivery. In skin vaccination, preserving the activity of the encapsulated antigen is an important consideration, as antigen activity is lost during DMN fabrication because of various stress factors. These stress factors vary between fabrication methods and each method affects the antigen's activity to different degrees. In this study, the activity of encapsulated antigens delivered by DMNs is compared between two recently developed DMN fabrication methods; droplet-born air blowing (DAB) and centrifugal lithography (CL) for a model scrub typhus vaccine antigen, ScaA. Although the in vitro analysis of ScaA-loaded DMNs (ScaA-DMNs) does not show any differences in physical properties depending on the fabrication methods, the immunogenicity of the CL-produced ScaA-DMN is significantly higher based on cytokine measurement and humoral immunity. DAB and CL differ in their solidification conditions, suggesting that solidification factors critically affect the encapsulated antigen's activity. ScaA-DMNs may also be stably stored for 4 weeks at room temperature. In conclusion, CL is a superior DMN fabrication method compared with DAB, and this study proves that DMN is feasible and practical for skin vaccination.

Comparative genomic analysis of Rickettsia rickettsii for identification of drug and vaccine targets: tolC as a proposed candidate for case study.

BACKGROUND: Antibiotic resistance is increasing rapidly in pathogenic organisms, creating more complications for treatment of diseases. Rocky Mountain spotted fever (RMSF) is a neglected tropical disease in humans caused by Rickettsia rickettsii for which no effective therapeutic is available. Subtractive genomics methods facilitate the characterization of non-homologous essential proteins that could be targeted for the discovery of potential therapeutic compounds against R. rickettsii to combat RMSF. Present study followed an in-silico based methodology, involving scanning and filtering the complete proteome of Rickettsia rickettsii by using several prioritization parameters in the search of potential candidates for drug development. Further the putative targets were subjected to series of molecular dockings with ligands obtained from PDB ligand database to identify suitable potential inhibitors. The comparative genomic analysis revealed 606 non-homologous proteins and 233 essential non-homologous proteins of R. rickettsii. The metabolic pathway analysis predicted 120 proteins as putative drug targets, out of which 56 proteins were found to be associated with metabolic pathways unique to the bacteria and further subcellular localization analysis revealed that 9 proteins as potential drug targets which are secretion proteins, involved in peptidoglycan biosynthesis, folate biosynthesis and bacterial secretion system. As secretion proteins are more feasible as vaccine candidates, we have selected a most potential target i.e. tolC, an outer membrane efflux protein that belongs to type I secretion system and has major role in pathogen survival as well as MDR persistence. So for case study, we have modelled the three dimensional structure of tolC (tunnel protein). The model was further subjected to virtual screening and in-silico docking. The study identified three potential inhibitors having PDB Id 19V, 6Q8 and 39H. Further we have suggested that the above study would be most important while considering the selection of candidate targets and drug or vaccine designing against R. rickettsii.

Tifus y laboratorio en la España de posguerra / No disponible

El tifus exantemático padecido de forma epidémica en los años consecutivos a la Guerra Civil fue momento de una peculiar movilización científica en el Instituto y Escuela Nacional de Sanidad que comenzaban su andadura bajo el franquismo, contando con una significativa participación multinacional, desde la Fundación Rockefeller al Instituto Pasteur, que convirtió España en nudo donde se cruzaban las teorías y las prácticas de las escuelas más relevantes en torno a la consecución de una vacuna eficaz. El artículo reconstruye los elementos en dicha movilización y analiza sus resultados. Las carencias informativas del momento, cierta tensión entre los participantes y la rápida evolución de la situación privaron de resultados prácticos inmediatos a la Sanidad española, lo que no puede decirse en lo tocante al recorrido profesional de varios de los científicos funcionarios implicados. La auténtica explotación de resultados de esta experiencia se llevó a cabo en los Estados Unidos años después (AU)

No disponible

Vaccination with an Attenuated Mutant of Ehrlichia chaffeensis Induces Pathogen-Specific CD4+ T Cell Immunity and Protection from Tick-Transmitted Wild-Type Challenge in the Canine Host.

Ehrlichia chaffeensis is a tick-borne rickettsial pathogen and the causative agent of human monocytic ehrlichiosis. Transmitted by the Amblyomma americanum tick, E. chaffeensis also causes disease in several other vertebrate species including white-tailed deer and dogs. We have recently described the generation of an attenuated mutant strain of E. chaffeensis, with a mutation in the Ech_0660 gene, which is able to confer protection from secondary, intravenous-administered, wild-type E. chaffeensis infection in dogs. Here, we extend our previous results, demonstrating that vaccination with the Ech_0660 mutant protects dogs from physiologic, tick-transmitted, secondary challenge with wild-type E. chaffeensis; and describing, for the first time, the cellular and humoral immune responses induced by Ech_0660 mutant vaccination and wild-type E. chaffeensis infection in the canine host. Both vaccination and infection induced a rise in E. chaffeensis-specific antibody titers and a significant Th1 response in peripheral blood as measured by E. chaffeensis antigen-dependent CD4+ T cell proliferation and IFNγ production. Further, we describe for the first time significant IL-17 production by peripheral blood leukocytes from both Ech_0660 mutant vaccinated animals and control animals infected with wild-type E. chaffeensis, suggesting a previously unrecognized role for IL-17 and Th17 cells in the immune response to rickettsial pathogens. Our results are a critical first step towards defining the role of the immune system in vaccine-induced protection from E. chaffeensis infection in an incidental host; and confirm the potential of the attenuated mutant clone, Ech_0660, to be used as a vaccine candidate for protection against tick-transmitted E. chaffeensis infection.

Lesser known aspects of Ludwik Fleck's (1896-1961) heroic life during World War II.

Professor Ludwik Fleck was a famous scientist and a prominent philosopher. Although his life and work were studied extensively, the Second World War period was a subject of some discussion and controversy. On account of his Jewish origin, he was first arrested and moved from the Lwów ghetto to the 'Laokoon' factory and then imprisoned in KL Auschwitz-Birkenau and in KL Buchenwald. Fleck produced the anti-typhus vaccine in the chemo-bacteriological laboratory in the Jewish Hospital at Kuszewicza Street and in the 'Laokoon' factory in Lwów. During his incarceration in KL Auschwitz-Birkenau, Fleck worked in the camp laboratory in Block 10 carrying out bacteriological studies for the inmates and then was assigned to work in the Wasserman station in Rajsko. From January 1944 Fleck performed routine laboratory tests in Block 50 in KL Buchenwald. Though Fleck had a privileged life in the camp, he participated in the sabotage activities organized by the camp resistance.

Enhanced protection against Rickettsia rickettsii infection in C3H/HeN mice by immunization with a combination of a recombinant adhesin rAdr2 and a protein fragment rOmpB-4 derived from outer membrane protein B.

BACKGROUND: Two surface proteins of Rickettsia rickettsii, outer membrane protein B (OmpB) and adhesion 2 (Adr2), have been recognized as protective antigens. Herein, the immunization with both OmpB and Adr2 was performed in mice so as to explore whether their combination could induce an enhanced immunoprotection against R. rickettsii infection. METHODS: C3H/HeN mice were immunized with recombinant protein rAdr2 or/and rOmp-4, a fragment derived from OmpB, and then mice were challenged with R. rickettsii. After which rickettsial loads in mice were measured by quantitative PCR. The specific antibodies in mouse sera were determined by ELISA and antigen-specific cytokines secretion by mouse T cells were analyzed in vitro. RESULTS: After challenge with R. rickettsii, the mice immunized with rAdr2 or/and rOmpB-4 had significant lower rickettsial load in livers, spleens, or lungs compared to PBS mock-immunized mice. Particularly, the load in lungs of mice immunized with both rAdr2 and rOmpB-4 was significantly lower than that with either of them. High levels of specific antibodies were detected in sera from mice immunized with rAdr2 or/and rOmpB-4, but the ratios of specific IgG2a to IgG1 induced by their combination were significantly higher than that by either rAdr2 or rOmpB-4. Following stimulation with rAdr2 or/and rOmpB-4, the INF-γ secreted by CD4(+) T cells from infected mice was significantly higher than that by cognate cells from uninfected mice. And the TNF-α secreted by CD4(+) or CD8(+) T cells from infected mice was markedly greater than that by cognate cells from uninfected mice after stimulation by their combination but not either of them. CONCLUSION: The combination of rAdr2 and rOmpB-4 conferred an enhanced protection against R. rickettsii infection in mice, which was mainly dependent on a stronger Th1-oriented immunoresponse with greater INF-γ and TNF-α secretion by antigen-specific T cells and specific IgG2a elicited by the combination.

[Spotted fever and the invention of its serodiagnosis and vaccination in the Austro-Hungarian army in World War I]. / Das Fleckfieber und die Erfindung seiner Serodiagnose und Impfung bei der k. u. k. Armee im Ersten Weltkrieg.

After description of the medical institutions and epidemiological situations of the Austro-Hungarian army in World War I the provisions against spotted fever focused on louse control are discussed. The letter specified for the army had to be adjusted for the local populations. 1915 in the k.u.k. military service in Galicia Edmund Weil and Arthur Felix cultivated Proteus strains from urine of soldiers with spotted fever. As sera of such patients agglutinated these bacteria in considerable titers the investigators developed the reliable diagnostic "Weil-Felix-Test" used still today. In the same military area and time Rudolf Weigl invented the anal infection of lice. This enabled him to harvest a great amount of louse intestines containing the spotted fever Rickettsiae in their epithelial cells. Lots with defined numbers of intestines were homogenized, sterilized and used with success as vaccine for medical staff. This sort of vaccine still was used in World War II.

Discovery of novel cross-protective Rickettsia prowazekii T-cell antigens using a combined reverse vaccinology and in vivo screening approach.

Rickettsial agents are some of the most lethal pathogens known to man. Among them, Rickettsia prowazekii is a select agent with potential use for bioterrorism; yet, there is no anti-Rickettsia vaccine commercially available. Owing to the obligate intracellular lifestyle of rickettsiae, CD8(+) T cells are indispensable for protective cellular immunity. Furthermore, T cells can mediate cross-protective immunity between different pathogenic Rickettsia, a finding consistent with the remarkable similarity among rickettsial genomes. However, Rickettsia T cell antigens remain unidentified. In the present study, we report an algorithm that allowed us to identify and validate four novel R. prowazekii vaccine antigen candidates recognized by CD8(+) T cells from a set of twelve in silico-defined protein targets. Our results highlight the importance of combining proteasome-processing as well as MHC class-I-binding predictions. The novel rickettsial vaccine candidate antigens, RP778, RP739, RP598, and RP403, protected mice against a lethal challenge with Rickettsia typhi, which is indicative of cross-protective immunity within the typhus group rickettsiae. Together, our findings validate a reverse vaccinology approach as a viable strategy to identify protective rickettsial antigens and highlight the feasibility of a subunit vaccine that triggers T-cell-mediated cross-protection among diverse rickettsiae.

Discovery of a protective Rickettsia prowazekii antigen recognized by CD8+ T cells, RP884, using an in vivo screening platform.

Rickettsia prowazekii has been tested for biological warfare due to the high mortality that it produces after aerosol transmission of very low numbers of rickettsiae. Epidemic typhus, the infection caused by these obligately intracellular bacteria, continues to be a threat because it is difficult to diagnose due to initial non-specific symptoms and the lack of commercial diagnostic tests that are sensitive and specific during the initial clinical presentation. A vaccine to prevent epidemic typhus would constitute an effective deterrent to the weaponization of R. prowazekii; however, an effective and safe vaccine is not currently available. Due to the cytoplasmic niche of Rickettsia, CD8(+) T-cells are critical effectors of immunity; however, the identification of antigens recognized by these cells has not been systematically addressed. To help close this gap, we designed an antigen discovery strategy that uses cell-based vaccination with antigen presenting cells expressing microbe's proteins targeted to the MHC class I presentation pathway. We report the use of this method to discover a protective T-cell rickettsial antigen, RP884, among a test subset of rickettsial proteins.

Protective immunity induced by immunization with a live, cultured Anaplasma marginale strain.

Despite significant economic losses resulting from infection with Anaplasma marginale, a tick-transmitted rickettsial pathogen of cattle, available vaccines provide, at best, only partial protection against clinical disease. The green-fluorescent protein expressing mutant of the A. marginale St. Maries strain is a live, marked vaccine candidate (AmStM-GFP). To test whether AmStM-GFP is safe and provides clinical protection, a group of calves was vaccinated, and clinical parameters, including percent parasitized erythrocytes (PPE), packed cell volume (PCV) and days required to reach peak bacteremia, were measured following inoculation and following tick challenge with wild type St. Maries strain (AmStM). These clinical parameters were compared to those obtained during infection with the A. marginale subsp. centrale vaccine strain (A. centrale) or wild type AmStM. AmStM-GFP resulted in similar clinical parameters to A. centrale, but had a lower maximum PPE, smaller drop in PCV and took longer to reach peak bacteremia than wild type AmStM. AmStM-GFP provided clinical protection, yielding a stable PCV and low bacteremia following challenge, whereas A. centrale only afforded partial clinical protection.

Genome-wide screening and identification of antigens for rickettsial vaccine development.

The capacity to identify immunogens for vaccine development by genome-wide screening has been markedly enhanced by the availability of microbial genome sequences coupled to proteomic and bioinformatic analysis. Critical to this approach is in vivo testing in the context of a natural host­pathogen relationship, one that includes genetic diversity in the host as well as among pathogen strains. We aggregate the results of three independent genome-wide screens using in vivo immunization and protection against Anaplasma marginale as a model for discovery of vaccine antigens for rickettsial pathogens. In silico analysis identified 62 outer membrane proteins (Omp) from the 949 predicted proteins in the A. marginale genome. These 62 Omps were reduced to 10 vaccine candidates by two independent genome-wide screens using IgG2 from vaccinates protected from challenge following vaccination with outer membranes (screen 1) or bacterial surface complexes (screen 2). Omps with broadly conserved epitopes were identified by immunization with a live heterologous vaccine, A. marginale ssp. centrale (screen 3), reducing the candidates to three. The genome-wide screens identified Omps that have orthologs broadly conserved among rickettsial pathogens, highlighted the importance of identifying immunologically subdominant antigens, and supported the use of reverse vaccinology approaches in vaccine development for rickettsial diseases.

Chlamydia trachomatis vaccine research through the years.

Chlamydia trachomatis is a Gram-negative obligate intracellular bacterium. It is the leading cause of bacterial sexual transmitted infections (STIs). World Health Organization figures estimated that over 90 million new cases of genital C. trachomatis infections occur worldwide each year. A vaccination program is considered to be the best approach to reduce the prevalence of C. trachomatis infections, as it would be much cheaper and have a greater impact on controlling C. trachomatis infections worldwide rather than a screening program or treating infections with antibiotics. Currently, there are no vaccines available which effectively protect against a C. trachomatis genital infection despite the many efforts that have been made throughout the years. In this paper, the many attempts to develop a protective vaccine against a genital C. trachomatis infection will be reviewed.

Molecular basis of immunity to rickettsial infection conferred through outer membrane protein B.

Pathogenic rickettsiae are the causative agents of Rocky Mountain spotted fever, typhus, and other human diseases with high mortality and an important impact on society. Although survivors of rickettsial infections are considered immune to disease, the molecular basis of this immunity or the identification of protective antigens that enable vaccine development was hitherto not known. By exploring the molecular pathogenesis of Rickettsia conorii, the agent of Mediterranean spotted fever, we report here that the autotransporter protein, rickettsial outer membrane protein B (rOmpB), constitutes a protective antigen for this group of pathogens. A recombinant, purified rOmpB passenger domain fragment comprised of amino acids 36 to 1334 is sufficient to elicit humoral immune responses that protect animals against lethal disease. Protective immunity requires folded antigen and production of antibodies that recognize conformational epitopes on the rickettsial surface. Monoclonal antibodies (MAbs) 5C7.27 and 5C7.31, which specifically recognize a conformation present in the folded, intact rOmpB passenger domain, are sufficient to confer immunity in vivo. Analyses in vitro indicate this protection involves a mechanism of complement-mediated killing in mammalian blood, a means of rickettsial clearance that has not been previously described. Considering the evolutionary conservation of rOmpB and its crucial contribution to bacterial invasion of host cells, we propose that rOmpB antibody-mediated killing confers immunity to rickettsial infection.

Vi-CRM 197 as a new conjugate vaccine against Salmonella Typhi.

An efficacious, low cost vaccine against typhoid fever, especially for young children, would make a major impact on disease burden in developing countries. The virulence capsular polysaccharide of Salmonella Typhi (Vi) coupled to recombinant mutant Pseudomonas aeruginosa exoprotein A (Vi-rEPA) has been shown to be highly efficacious. We investigated the use of carrier proteins included in infant vaccines, standardized the conjugation process and developed key assays required for routine lot release at production scale. Vi from a BSL1 organism, Citrobacter freundii, strain WR7011, was used as an alternative to Vi from S. Typhi. We showed that Vi conjugated to CRM(197), a non-toxic mutant of diphtheria toxin, widely used in commercial vaccines, was produced at high yield. Vi-CRM(197) proved immunogenic in animal studies, even without adjuvant. Thus, Vi-CRM(197) appears to be a suitable candidate for the development of a commercially viable, effective typhoid vaccine for developing countries.