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      Oral Administration of Electron-Beam Inactivated Rhodococcus equi Failed to Protect Foals against Intrabronchial Infection with Live, Virulent R. equi

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          Abstract

          There is currently no licensed vaccine that protects foals against Rhodococcus equi–induced pneumonia. Oral administration of live, virulent R. equi to neonatal foals has been demonstrated to protect against subsequent intrabronchial challenge with virulent R. equi. Electron beam (eBeam)-inactivated R. equi are structurally intact and have been demonstrated to be immunogenic when administered orally to neonatal foals. Thus, we investigated whether eBeam inactivated R. equi could protect foals against developing pneumonia after experimental infection with live, virulent R. equi. Foals (n = 8) were vaccinated by gavaging with eBeam-inactivated R. equi at ages 2, 7, and 14 days, or gavaged with equal volume of saline solution (n = 4), and subsequently infected intrabronchially with live, virulent R. equi at age 21 days. The proportion of vaccinated foals that developed pneumonia following challenge was similar among the vaccinated (7/8; 88%) and unvaccinated foals (3/4; 75%). This vaccination regimen did not appear to be strongly immunogenic in foals. Alternative dosing regimens or routes of administration need further investigation and may prove to be immunogenic and protective.

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          Most cited references43

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          Rhodococcus equi: an animal and human pathogen.

          Recent isolations of Rhodococcus equi from cavitatory pulmonary disease in patients with AIDS have aroused interest among medical microbiologists in this unusual organism. Earlier isolations from humans had also been in immunosuppressed patients following hemolymphatic tumors or renal transplantation. This organism has been recognized for many years as a cause of a serious pyogranulomatous pneumonia of young foals and is occasionally isolated from granulomatous lesions in several other species, in some cases following immunosuppression. The last decade has seen many advances in understanding of the epidemiology, pathogenesis, diagnosis, treatment, and immunity to infection in foals. The particular susceptibility of the foal is not understood but can be explained in part by a combination of heavy challenge through the respiratory route coinciding with declining maternally derived antibody in the absence of fully competent foal cellular immune mechanisms. R. equi is largely a soil organism but is widespread in the feces of herbivores. Its growth in soil is considerably improved by simple nutrients it obtains from herbivore manure. About one-third of human patients who have developed R. equi infections had contact in some way with herbivores or their manure. Others may have acquired infection from contact with soil or wild bird manure. R. equi is an intracellular parasite, which explains the typical pyogranulomatous nature of R. equi infections, the predisposition to infection in human patients with defective cell-mediated immune mechanisms, and the efficacy of antimicrobial drugs that penetrate phagocytic cells.
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            Epidemiology of Rhodococcus equi infections: a review.

            An overview of epidemiology of R. equi infection in foals is presented, emphasizing the importance of the virulence-associated antigens and plasmids as epidemiological markers. The monoclonal antibody-based colony blot test has been developed to identify rapidly and accurately virulent R. equi. Epidemiological studies conducted during the recent 5 years have revealed that: (1) avirulent R. equi are widespread in the feces of horses and their environment on every farm; (2) the feces of horses and the environment of the horse farms having endemic R. equi infections demonstrated heavy contamination with virulent R. equi, but the farms without the problem did not, thus suggesting that foals bred on a farm with endemic disease are exposed more frequently to virulent R. equi in their environment than those of a farm without the problem; (3) only virulent R. equi are isolated from lesions of naturally infected foals, showing that natural infections in foals are principally by virulent R. equi, but not avirulent organisms; (4) infected foals which constantly shed large quantities of virulent R. equi in their feces are the major source of virulent R. equi, which this may be the mechanism of progressive development of infection on farms with a history of the disease. At present, farms with a potential for endemic infection can be distinguished on the basis of the contamination with virulent R. equi, so regular examination of foals and their environment by virulence markers might be the most practical approach to control R. equi infection on endemic farms.
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              Role of the 85-kilobase plasmid and plasmid-encoded virulence-associated protein A in intracellular survival and virulence of Rhodococcus equi.

              Rhodococcus equi is a facultative intracellular pathogen of macrophages and a cause of pneumonia in young horses (foals) and immunocompromised people. Isolates of R. equi from pneumonic foals typically contain large, 85- or 90-kb plasmids encoding a highly immunogenic virulence-associated protein (VapA). The objective of this study was to determine the role of the 85-kb plasmid and VapA in the intracellular survival and virulence of R. equi. Clinical isolates containing the plasmid and expressing VapA efficiently replicated within mouse macrophages in vitro, while plasmid-cured derivatives of these organisms did not multiply intracellularly. An isolate harboring the large plasmid also replicated in the tissues of experimentally infected mice, whereas its plasmid-cured derivative was rapidly cleared. All foals experimentally infected with a plasmid-containing clinical isolate developed severe bronchopneumonia, whereas the foals infected with its plasmid-cured derivative remained asymptomatic and free of visible lung lesions. By day 14 postinfection, lung bacterial burdens had increased considerably in foals challenged with the plasmid-containing clinical isolate. In contrast, bacteria could no longer be cultured from the lungs of foals challenged with the isogenic plasmid-cured derivative. A recombinant, plasmid-cured derivative expressing wild-type levels of VapA failed to replicate in macrophages and remained avirulent for both mice and foals. These results show that the 85-kb plasmid of R. equi is essential for intracellular replication within macrophages and for development of disease in the native host, the foal. However, expression of VapA alone is not sufficient to restore the virulence phenotype.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                1 February 2016
                2016
                : 11
                : 2
                : e0148111
                Affiliations
                [1 ]Department of Large Animal Clinical Sciences, College of Veterinary Medicine & Biomedical Sciences, Texas A&M University, College Station, Texas, 77843–4475, United States of America
                [2 ]Department of Large Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, Georgia, 30602–7385, United States of America
                [3 ]Department of Microbial Pathogenesis and Immunology, Texas A&M Health Science Center, College Station, Texas, 77843, United States of America
                [4 ]Department of Veterinary Pathobiology, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, Texas, 77843–4467, United States of America
                [5 ]National Center for Electron Beam Research–IAEA Collaborative Centre for Electron Beam Technology, Texas A&M University, College Station, Texas, 77843, United States of America
                Institut National de la Santé et de la Recherche Médicale (INSERM), FRANCE
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Conceived and designed the experiments: JNR NDC AIB SG RCA WM SDP. Performed the experiments: JNR NDC AIB CNB MCC. Analyzed the data: JNR AIB NDC. Contributed reagents/materials/analysis tools: SG SDL WM SDP. Wrote the paper: JNR NDC AIB SG SDL RCA SDP.

                Article
                PONE-D-15-50358
                10.1371/journal.pone.0148111
                4735123
                26828865
                6a68f8bb-244f-4970-a5e9-a6b1addf1adb
                © 2016 Rocha et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 18 November 2015
                : 13 January 2016
                Page count
                Figures: 6, Tables: 2, Pages: 18
                Funding
                Funded by Grayson-Jockey Club Research Foundation ( https://www.grayson-jockeyclub.org/) (Grant to NDC) and Link Equine Research Endowment (Support to NDC). The open access publishing fees for this article have been covered by the Texas A&M University Online Access to Knowledge (OAK) Fund, supported by the University Libraries and the Office of the Vice President for Research.
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                Pulmonology
                Pneumonia
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                All 2 files are available from the Figshare ( https://figshare.com) database (DOIs: 10.6084/m9.figshare.2062566 and 10.6084/m9.figshare.2062575).

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