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1.
Front Immunol ; 13: 901372, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35651616

RESUMEN

T cell-mediated immunity plays a central role in the control and clearance of intracellular Coxiella burnetii infection, which can cause Q fever. Therefore, we aimed to develop a novel T cell-targeted vaccine that induces pathogen-specific cell-mediated immunity to protect against Q fever in humans while avoiding the reactogenicity of the current inactivated whole cell vaccine. Human HLA class II T cell epitopes from C. burnetii were previously identified and selected by immunoinformatic predictions of HLA binding, conservation in multiple C. burnetii isolates, and low potential for cross-reactivity with the human proteome or microbiome. Epitopes were selected for vaccine inclusion based on long-lived human T cell recall responses to corresponding peptides in individuals that had been naturally exposed to the bacterium during a 2007-2010 Q fever outbreak in the Netherlands. Multiple viral vector-based candidate vaccines were generated that express concatemers of selected epitope sequences arranged to minimize potential junctional neo-epitopes. The vaccine candidates caused no antigen-specific reactogenicity in a sensitized guinea pig model. A subset of the vaccine epitope peptides elicited antigenic recall responses in splenocytes from C57BL/6 mice previously infected with C. burnetii. However, immunogenicity of the vaccine candidates in C57BL/6 mice was dominated by a single epitope and this was insufficient to confer protection against an infection challenge, highlighting the limitations of assessing human-targeted vaccine candidates in murine models. The viral vector-based vaccine candidates induced antigen-specific T cell responses to a broader array of epitopes in cynomolgus macaques, establishing a foundation for future vaccine efficacy studies in this large animal model of C. burnetii infection.


Asunto(s)
Coxiella burnetii , Fiebre Q , Animales , Anticuerpos Antibacterianos , Vacunas Bacterianas , Modelos Animales de Enfermedad , Epítopos de Linfocito T , Cobayas , Humanos , Ratones , Ratones Endogámicos C57BL , Péptidos , Fiebre Q/prevención & control , Linfocitos T
2.
Pathogens ; 9(3)2020 Mar 07.
Artículo en Inglés | MEDLINE | ID: mdl-32156028

RESUMEN

The wild pig population on Molokai, Hawaii, USA is a possible reservoir for bovine tuberculosis, caused by Mycobacterium bovis, and has been implicated in decades past as the source of disease for the island's domestic cattle. Heat-inactivated vaccines have been effective for reducing disease prevalence in wild boar in Spain and could prove useful for managing M. bovis in Molokai wild pigs. We designed an experiment to test this vaccine in wild pigs of Molokai genetics. Fifteen 3-4-month-old pigs were orally administered 106-107 colony forming units (cfu) of heat-inactivated M. bovis (Vaccinates; n = 8; 0.2 mL) or phosphate buffered saline (Controls; n = 7; 0.2 mL). Each dose was administered in a 0.5 mL tube embedded in a fruit candy/cracked corn mix. Boosters were given seven weeks post-prime in the same manner and dose. Nineteen weeks post-prime, pigs were orally challenged with 1 × 106 cfu of virulent M. bovis. Twelve weeks post-challenge, pigs were euthanized and necropsied, at which time 23 different tissues from the head, thorax, and abdomen were collected and examined. Each tissue was assigned a lesion score. Ordinal lesion score data were analyzed using non-parametric Wilcoxon Signed Rank test. Effect size was calculated using Cohen's d. Four of eight Vaccinates and four of seven Controls had gross and microscopic lesions, as well as culture-positive tissues. Vaccinates had statistically lower lesion scores than Controls in the following areas: gross thoracic lesion scores (p = 0.013 Cohen's d = 0.33) and microscopic thoracic lesion scores (p = 0.002, Cohen's d = 0.39). There were no differences in head lesion scores alone, both gross and microscopic, nor were there differences when comparing combined gross and microscopic head and thoracic lesion scores. These results are indicative that this vaccination protocol affords a modest degree of infection containment with this vaccine in Molokai wild pigs.

3.
Front Immunol ; 10: 207, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-30828331

RESUMEN

Coxiella burnetii, the causative agent of Q fever, is a Gram-negative intracellular bacterium transmitted via aerosol. Regulatory approval of the Australian whole-cell vaccine Q-VAX® in the US and Europe is hindered by reactogenicity in previously exposed individuals. The aim of this study was to identify and rationally select C. burnetii epitopes for design of a safe, effective, and less reactogenic T-cell targeted human Q fever vaccine. Immunoinformatic methods were used to predict 65 HLA class I epitopes and 50 promiscuous HLA class II C. burnetii epitope clusters, which are conserved across strains of C. burnetii. HLA binding assays confirmed 89% of class I and 75% of class II predictions, and 11 HLA class II epitopes elicited IFNγ responses following heterologous DNA/DNA/peptide/peptide prime-boost immunizations of HLA-DR3 transgenic mice. Human immune responses to the predicted epitopes were characterized in individuals naturally exposed to C. burnetii during the 2007-2010 Dutch Q fever outbreak. Subjects were divided into three groups: controls with no immunological evidence of previous infection and individuals with responses to heat-killed C. burnetii in a whole blood IFNγ release assay (IGRA) who remained asymptomatic or who experienced clinical Q fever during the outbreak. Recall responses to C. burnetii epitopes were assessed by cultured IFNγ ELISpot. While HLA class I epitope responses were sparse in this cohort, we identified 21 HLA class II epitopes that recalled T-cell IFNγ responses in 10-28% of IGRA+ subjects. IGRA+ individuals with past asymptomatic and symptomatic C. burnetii infection showed a comparable response pattern and cumulative peptide response which correlated with IGRA responses. None of the peptides elicited reactogenicity in a C. burnetii exposure-primed guinea pig model. These data demonstrate that a substantial proportion of immunoinformatically identified HLA class II epitopes show long-lived immunoreactivity in naturally infected individuals, making them desirable candidates for a novel human multi-epitope Q fever vaccine.


Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Coxiella burnetii/inmunología , Epítopos de Linfocito T/inmunología , Memoria Inmunológica , Fiebre Q/inmunología , Animales , Vacunas Bacterianas/inmunología , Biomarcadores , Linfocitos T CD4-Positivos/metabolismo , Citocinas/metabolismo , Ensayo de Immunospot Ligado a Enzimas , Cobayas , Antígenos HLA/inmunología , Antígenos HLA/metabolismo , Humanos , Inmunización , Inmunogenicidad Vacunal , Interferón gamma/biosíntesis , Fiebre Q/metabolismo , Fiebre Q/prevención & control
4.
PLoS One ; 13(10): e0205882, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30312355

RESUMEN

Historically, vaccination with Coxiella burnetii whole cell vaccines has induced hypersensitivity reactions in humans and animals that have had prior exposure to the pathogen as a result of infection or vaccination. Intradermal skin testing is routinely used to evaluate exposure in humans, and guinea pig hypersensitivity models have been developed to characterize the potential for reactogenicity in vaccine candidates. Here we describe a refinement of the guinea pig model using an alternate vaccine for positive controls. An initial comparative study used viable C. burnetii to compare the routes of sensitizing exposure of guinea pigs (intranasal vs intraperitoneal), evaluation of two time points for antigen challenge (21 and 42 days) and an assessment of two routes (intradermal and subcutaneous) of challenge using the ruminant vaccine Coxevac as the antigenic control. Animals sensitized by intraperitoneal exposure exhibited slightly larger gross reactions than did those sensitized by intranasal exposure, and reactions were more pronounced when skin challenge was performed at 42 days compared to 21 days post-sensitization. The intradermal route proved to be the optimal route of reactogenicity challenge. Histopathological changes at injection sites were similar to those previously reported and a scoring system was developed to compare reactions between groups receiving vaccine by intradermal versus subcutaneous routes. Based on the comparative study, a standardized protocol for assessment of vaccine reactogenicity in intranasally-sensitized animals was tested in a larger confirmatory study. Results suggest that screens utilizing a group size of n = 3 would achieve 90% power for detecting exposure-related reactogenic responses of the magnitude induced by Coxevac using either of two outcome measures.


Asunto(s)
Vacunas Bacterianas/inmunología , Coxiella burnetii , Fiebre Q/prevención & control , Administración Intranasal , Animales , Antígenos/química , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Cobayas , Inyecciones Intradérmicas , Inyecciones Subcutáneas , Piel , Resultado del Tratamiento , Vacunación
5.
J Clin Microbiol ; 56(7)2018 07.
Artículo en Inglés | MEDLINE | ID: mdl-29695520

RESUMEN

Yersinia pestis is the causative agent of plague and is considered a category A priority pathogen due to its potential for high transmissibility and the significant morbidity and mortality it causes in humans. Y. pestis is endemic to the western United States and much of the world, necessitating programs to monitor for this pathogen on the landscape. Elevated human risk of plague infection has been spatially correlated with spikes in seropositive wildlife numbers, particularly rodent-eating carnivores, which are frequently in contact with the enzootic hosts and the associated arthropod vectors of Y. pestis In this study, we describe a semiautomated bead-based flow cytometric assay developed for plague monitoring in wildlife called the F1 Luminex plague assay (F1-LPA). Based upon Luminex/Bio-Plex technology, the F1-LPA targets serological responses to the F1 capsular antigen of Y. pestis and was optimized to analyze antibodies eluted from wildlife blood samples preserved on Nobuto filter paper strips. In comparative evaluations with passive hemagglutination, the gold standard tool for wildlife plague serodiagnosis, the F1-LPA demonstrated as much as 64× improvement in analytical sensitivity for F1-specific IgG detection and allowed for unambiguous classification of IgG status. The functionality of the F1-LPA was demonstrated for coyotes and other canids, which are the primary sentinels in wildlife plague monitoring, as well as felids and raccoons. Additionally, assay formats that do not require species-specific immunological reagents, which are not routinely available for several wildlife species used in plague monitoring, were determined to be functional in the F1-LPA.


Asunto(s)
Animales Salvajes , Monitoreo Epidemiológico/veterinaria , Citometría de Flujo/métodos , Peste/veterinaria , Yersinia pestis , Animales , Anticuerpos Antibacterianos/sangre , Anticuerpos Antibacterianos/inmunología , Antígenos Bacterianos/inmunología , Proteínas Bacterianas/inmunología , Pruebas de Inhibición de Hemaglutinación , Pruebas de Hemaglutinación , Inmunoensayo , Peste/sangre , Peste/diagnóstico , Peste/microbiología , Reproducibilidad de los Resultados , Yersinia pestis/inmunología
6.
J Wildl Dis ; 50(2): 384-8, 2014 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-24484483

RESUMEN

Plague surveillance is routinely conducted to predict future epizootics in wildlife and exposure risk for humans. The most common surveillance method for sylvatic plague is detection of antibodies to Yersinia pestis F1 capsular antigen in sentinel animals, such as coyotes (Canis latrans). Current serologic tests for Y. pestis, hemagglutination (HA) test and enzyme-linked immunosorbent assay (ELISA), are expensive and labor intensive. To address this need, we developed a complete lateral flow device for the detection of specific antibodies to Y. pestis F1 and V antigens. Our test detected anti-F1 and anti-V antibodies in serum and Nobuto filter paper samples from coyotes, and in serum samples from prairie dogs (Cynomys ludovicianus), lynx (Lynx canadensis), and black-footed ferrets (Mustela nigripes). Comparison of cassette results for anti-F1 and anti-V antibodies with results of ELISA or HA tests showed correlations ranging from 0.68 to 0.98. This device provides an affordable, user-friendly tool that may be useful in plague surveillance programs and as a research tool.


Asunto(s)
Animales Salvajes , Peste/veterinaria , Yersinia pestis , Animales , Peste/diagnóstico , Peste/microbiología
7.
J Wildl Dis ; 49(3): 738-40, 2013 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-23778634

RESUMEN

During 2007-09, we necropsied striped skunks (Mephitis mephitis) from Colorado, USA. Eight of 51 (16%) had severe infections with the subcutaneous filarid nematode Filaria taxideae, and four of the infected skunks (50%) had dermatitis that was histologically associated with parasite ova in the skin.


Asunto(s)
Filariasis/veterinaria , Mephitidae/parasitología , Animales , Colorado/epidemiología , Filariasis/diagnóstico , Filariasis/epidemiología
8.
J Wildl Dis ; 49(4): 932-9, 2013 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-24502720

RESUMEN

Multiple publications have reported the use of coyotes (Canis latrans) in animal-based surveillance efforts for the detection of Yersinia pestis. Coyotes are likely exposed via flea bite or oral routes and are presumed to be resistant to the development of clinical disease. These historic data have only been useful for the evaluation of the geographic distribution of Y. pestis in the landscape. Because the canid immunologic response to Y. pestis has not been thoroughly characterized, we conducted experimental inoculation of captive-reared, juvenile coyotes (n = 8) with Y. pestis CO92 via oral or intradermal routes. We measured the humoral response to Y. pestis fraction 1 capsular protein (anti-F1) and found a significant difference between inoculation groups in magnitude and duration of antibody production. The anti-F1 titers in animals exposed intradermally peaked at day 10 postinoculation (PI; range = 1∶32 to 1∶128) with titers remaining stable at 1∶32 through week 12. In contrast, orally inoculated animals developed higher titers (range = 1∶256 to 1∶1,024) that remained stable at 1∶256 to 1∶512 through week 6. No clinical signs of disease were observed, and minimal changes were noted in body temperature, white blood cell counts, and acute phase proteins during the 7 days PI. Gross pathology was unremarkable, and minimal changes were noted in histopathology at days 3 and 7 PI. Rechallenge at 14 wk PI via similar dosage and routes resulted in marked differences in antibody response between groups. Animals in the orally inoculated group produced a striking increase in anti-F1 titers (up to 1∶4,096) within 3 days, whereas there was minimal to no increase in antibody response in the intradermal group. Information gathered from this experimental trial may provide additional insight into the spatial and temporal evaluation of coyote plague serology.


Asunto(s)
Coyotes/sangre , Peste/veterinaria , Yersinia pestis/clasificación , Animales , Anticuerpos Antibacterianos , Coyotes/inmunología , Pruebas de Hemaglutinación/veterinaria , Inmunidad Humoral , Peste/inmunología , Peste/microbiología , Peste/patología , Factores de Tiempo
9.
Curr Pharm Biotechnol ; 13(6): 1061-9, 2012 May.
Artículo en Inglés | MEDLINE | ID: mdl-22039796

RESUMEN

The use of macrocyclic lactones has become the main stay for the treatment of endo- and ectoparasites in the cattle industry. Here we review those drugs that are currently approved for use in cattle in the United States. The general efficacy, tissue distribution and toxicity of each drug formulation are discussed. Included is a discussion regarding the current status for nematode anthelmintic resistance in cattle populations within the United States. Also provided is a current summary of ecological effects of macrocyclic lactones residues in manure.


Asunto(s)
Antiparasitarios/uso terapéutico , Enfermedades de los Bovinos/tratamiento farmacológico , Macrólidos/uso terapéutico , Enfermedades Parasitarias en Animales/tratamiento farmacológico , Animales , Antiparasitarios/farmacología , Bovinos , Residuos de Medicamentos/toxicidad , Resistencia a Medicamentos , Heces/química , Macrólidos/farmacología , Parásitos/efectos de los fármacos , Estados Unidos
10.
J Wildl Dis ; 46(2): 585-90, 2010 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-20688655

RESUMEN

Individuals of three species of cervids, with varying degrees of alopecia, were examined for ectoparasites: Rocky Mountain elk (Cervus elaphus nelsoni) and mule deer (Odocoileus hemionus hemionus) in Colorado and white-tailed deer (Odocoileus virginianus) in South Dakota. Hair follicle mites were recovered and identified as Demodex kutzeri, a species originally described from the European red deer (Cervus elaphus, from Austria) and the sika deer (Cervus nippon pseudaxis, captive in Germany). These findings expand the geographic range of D. kutzeri to North America and extend its host range to include the genus Odocoileus. Thus, the host range for D. kutzeri spans two subfamilies of cervids. Additionally, D. kutzeri was identified in material from a white-tailed deer collected in South Carolina in 1971, indicating this parasite has been present, but unrecognized, on US cervids for some time.


Asunto(s)
Alopecia/veterinaria , Antílopes/parasitología , Ciervos/parasitología , Infestaciones por Ácaros/veterinaria , Alopecia/epidemiología , Alopecia/parasitología , Animales , Animales Salvajes/parasitología , Colorado/epidemiología , Femenino , Folículo Piloso/parasitología , Interacciones Huésped-Parásitos , Masculino , Infestaciones por Ácaros/epidemiología , Infestaciones por Ácaros/parasitología , South Dakota/epidemiología , Especificidad de la Especie
11.
J Vet Diagn Invest ; 20(5): 650-3, 2008 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-18776103

RESUMEN

Bovine viral diarrhea virus (BVDV) is a significant viral pathogen of domestic cattle. Worldwide, there is evidence of BVDV exposure and infection in wild ungulates; however, the frequency and significance of such events are unknown. To determine the prevalence and distribution of Colorado deer, elk, and moose persistently infected (PI) with BVDV, a cross-sectional study was conducted using full-thickness ear tissue samples collected from animals presented to the Colorado Division of Wildlife for chronic wasting disease surveillance in the 2005-2006 hunting season. Tissue from 5,597 harvested animals (2,934 mule deer, 2,516 elk, 141 white-tailed deer, and 6 moose) was paraffin-embedded and stained for BVDV using immunohistochemistry. A single adult male mule deer had BVDV antigen in the skin; staining distribution was consistent with that seen in PI cattle. Skin and lymph node were also positive for viral RNA by polymerase chain reaction, and the virus was determined to be a type 1. The prevalence of BVDV PI cervids in Colorado is very low. However, the identification of a naturally infected adult PI animal in the wild suggests that the virus infects free-ranging populations. The source of the BVDV is unknown and is assumed to be spillover from cattle or maintenance within wildlife populations. Consideration of a potential wild animal reservoir is important in the design and implementation of BVDV management practices in cattle.


Asunto(s)
Animales Salvajes/virología , Virus de la Diarrea Viral Bovina/aislamiento & purificación , Infecciones por Pestivirus/veterinaria , Animales , Colorado/epidemiología , Estudios Transversales , Ciervos/virología , Incidencia , Infecciones por Pestivirus/epidemiología , Rumiantes/virología
12.
J Wildl Dis ; 43(2): 309-14, 2007 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-17495319

RESUMEN

Chronic wasting disease (CWD) was diagnosed in a free-ranging moose (Alces alces shirasi) killed by a hunter in Jackson County, Colorado, USA, in September 2005. The diagnosis was based upon immunohistochemistry (IHC) demonstrating the presence of accumulations of CWD-associated prion protein (PrP(CWD)) in tissue sections of medulla oblongata at the level of the obex (dorsal motor nucleus of the vagus) and in retropharyngeal lymph node (RPLN); additional testing by IHC revealed deposits of PrP(CWD) in multiple sections of medulla oblongata and cervical spinal cord as well as palatine tonsil and submandibular lymph node tissues. Western blot confirmed the presence of PrP(CWD) in RPLN and tonsil tissue. The PrP(CWD) also was detected via enzyme-linked immunosorbent assay of RPLN tissue. Spongiform encephalopathy was observed in sections of the brainstem and cervical spinal cord, although no clinical signs were noted by the hunter who killed the animal. The affected moose was homozygous for methionine at codon 209 of the prion protein coding region. In October 2006, two additional free-ranging moose were diagnosed with CWD. Epidemiology and implications of CWD in moose remain to be determined.


Asunto(s)
Ciervos , Priones/aislamiento & purificación , Enfermedad Debilitante Crónica/diagnóstico , Animales , Animales Salvajes , Colorado , Inmunohistoquímica/veterinaria , Masculino , Enfermedad Debilitante Crónica/patología , Enfermedad Debilitante Crónica/transmisión
13.
J Zoo Wildl Med ; 35(2): 142-6, 2004 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-15305507

RESUMEN

Black-footed ferrets (Mustela nigripes) are highly susceptible to sylvatic plague, caused by the bacterium Yersinia pestis, and this disease has severely hampered efforts to restore ferrets to their historic range. A study was conducted to assess the efficacy of vaccination of black-footed ferrets against plague using a recombinant protein vaccine, designated F1-V, developed by personnel at the U.S. Army Medical Research Institute of Infectious Diseases. Seven postreproductive black-footed ferrets were immunized with the vaccine, followed by two booster immunizations on days 23 and 154; three control black-footed ferrets received a placebo. After the second immunization, antibody titers to both F1 and V antigen were found to be significantly higher in vaccinates than controls. On challenge with 7,800 colony-forming units of virulent plague by s.c. injection, the three control animals died within 3 days, but six of seven vaccinates survived with no ill effects. The seventh vaccinate died on day 8. These results indicate that black-footed ferrets can be immunized against plague induced by the s.c. route, similar to fleabite injection.


Asunto(s)
Vacunas Bacterianas , Hurones , Peste/veterinaria , Proteínas Recombinantes de Fusión/inmunología , Yersinia pestis/inmunología , Animales , Femenino , Inyecciones Subcutáneas/veterinaria , Masculino , Peste/prevención & control , Distribución Aleatoria , Proteínas Recombinantes de Fusión/administración & dosificación , Análisis de Supervivencia , Factores de Tiempo , Vacunación/veterinaria , Vacunas Sintéticas , Yersinia pestis/patogenicidad
14.
J Vet Diagn Invest ; 15(4): 311-9, 2003 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-12918810

RESUMEN

Tissue samples (n = 25,050 total) from 23,256 mule deer (Odocoileus hemionus), Rocky Mountain elk (Cervus elaphus nelsoni), and white-tailed deer (Odocoileus virginianus) collected statewide in Colorado were examined for chronic wasting disease (CWD) using an enzyme-linked immunosorbent assay developed by Bio-Rad Laboratories, Inc. (brELISA), in a 2-phase study. In the validation phase of this study, a total of 4,175 retropharyngeal lymph nodes (RLN) or obex (OB) tissue samples were examined independently by brELISA and immunohistochemistry (IHC). There were 137 IHC-positive samples and 4,038 IHC-negative samples. Optical density (OD) values from brELISA were classified as "not detected" or "suspect" based on recommended cutoff values during the validation phase. Using IHC-positive cases as known CWD-infected individuals and assuming IHC-negative cases as uninfected, the relative sensitivity of brELISA depending on species ranged from 98.3% to 100% for RLN samples and 92.1% to 93.3% for OB samples; the relative specificity of brELISA depending on species ranged from 99.9% to 100% for RLN samples and was 100% for OB samples. Overall agreement between brELISA and IHC was > or = 97.6% in RLN samples and > or = 95.7% in OB samples of all species where values could be calculated; moreover, mean brELISA OD values were > or = 46X higher in IHC-positive samples than in IHC-negative samples. Discrepancies were observed only in early-stage cases of CWD. Based on the validation phase data, only RLN samples were collected for the field application phase of this study and only samples with brELISA OD values > 0.1 were examined by IHC. Among 20,875 RLN samples screened with brELISA during this second testing phase, 155 of 8,877 mule deer, 33 of 11,731 elk, and 9 of 267 white-tailed deer samples (197 total) had OD values > 0.1 and were further evaluated by IHC to confirm evidence of CWD infection. Of cases flagged for IHC follow-up, 143 of 155 mule deer, 29 of 33 elk, and all 9 white-tailed deer were confirmed positive. Mean (+/- SE) OD values for IHC-positive cases detected during the field application phase were comparable with those measured in RLN tissues during the validation phase. Based on these data, brELISA was determined to be an excellent rapid test for screening large numbers of samples in surveys designed to detect CWD infections in deer and elk populations.


Asunto(s)
Ciervos , Ensayo de Inmunoadsorción Enzimática/veterinaria , Enfermedades por Prión/veterinaria , Priones/análisis , Síndrome Debilitante/veterinaria , Enfermedades de los Animales/diagnóstico , Animales , Animales Salvajes , Anticuerpos Monoclonales , Diagnóstico Diferencial , Ensayo de Inmunoadsorción Enzimática/normas , Inmunohistoquímica , Vigilancia de la Población , Enfermedades por Prión/diagnóstico , Priones/inmunología , Sensibilidad y Especificidad , Síndrome Debilitante/diagnóstico
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