RESUMO
Several DNA-based and serological tests have been established for the detection of Theileria annulata infection, including polymerase chain reaction, reverse line blot and loop-mediated isothermal amplification, indirect enzyme-linked immunosorbent assay (ELISA), and competitive ELISA. In this study, we have applied knowledge from the development and application of a recombinant protein-based indirect ELISA and competitive ELISA to establish a rapid test for point-of-care diagnosis of T. annulata infection in the field to be used by the veterinarian. For the development of a lateral flow test, the recombinantly expressed T. annulata surface protein (TaSP) was applied as the test antigen and anti-TaSP antiserum as the control line. TaSP antigen conjugated to colloidal gold particles was used as the detection system for visualization at the test line for the binding of anti-TaSP antibody present in the serum of infected animals. The developed test specifically detected antibodies in the serum of animals experimentally infected with T. annulata and showed no cross-reactivity with serum from animals infected with other tested bovine pathogens (Trypanosoma brucei, Anaplasma marginale, Babesia bigemina, Babesia bovis, and Theileria parva). Testing of field samples was compared to results obtained by other serological tests, resulting in a sensitivity and specificity of 96.3% and 87.5% compared to indirect fluorescence antibody test, 98.7% and 81.8% compared to indirect ELISA, and 100% and 47.6% compared to competitive ELISA. In conclusion, a rapid test for the detection of T. annulata infection (T. annulata lateral flow device, Ta-LFD) has been developed, which is easy to perform, delivers results to be read by the naked eye within 10 min, and is suitable for the detection of infection in field samples.
Assuntos
Anticorpos Anti-Helmínticos/sangue , Doenças dos Bovinos/diagnóstico , Parasitologia/métodos , Theileria annulata/imunologia , Theileriose/diagnóstico , Animais , Antígenos de Protozoários , Bovinos , Doenças dos Bovinos/parasitologia , Parasitologia/normas , Sistemas Automatizados de Assistência Junto ao Leito/normas , Proteínas Recombinantes , Padrões de Referência , Sensibilidade e Especificidade , Testes Sorológicos/métodos , Testes Sorológicos/normas , Theileria annulata/isolamento & purificação , Theileriose/parasitologiaRESUMO
A lateral flow device (LFD) for the detection of all seven serotypes of foot-and-mouth disease virus (FMDV) was developed using a monoclonal antibody (Mab 1F10) shown to be pan-reactive to FMDV strains of each serotype by ELISA. The performance of the LFD was evaluated in the laboratory on suspensions of vesicular epithelia (304 positive and 1003 negative samples) from suspected cases of vesicular disease collected from 86 countries between 1965 and 2008 and negative samples collected from healthy animals. The diagnostic sensitivity of the LFD for FMDV was similar at 84% compared to 85% obtained by the reference method of antigen ELISA, and the diagnostic specificity of the LFD was approximately 99% compared to 99.9% for the ELISA. The device recognized FMDV strains of wide diversity of all seven serotypes but weaker reactions were often evident with those of type SAT 2, several viruses of which were not detected. Reactions with the viruses of swine vesicular disease and vesicular stomatitis that produce clinically indistinguishable syndromes in pigs and cattle, did not occur. The test procedure was simple and rapid, and typically provided a result within 1-10min of sample addition. Simple homogenizers that could be used in field conditions for preparing epithelial suspensions were demonstrated to be effective for LFD application. These data illustrate the potential for the LFD to be used next to the animal in the pen-side diagnosis of FMD and for providing rapid and objective support to veterinarians in their clinical judgment of the disease.
Assuntos
Anticorpos Monoclonais , Antígenos Virais/análise , Vírus da Febre Aftosa/isolamento & purificação , Febre Aftosa/diagnóstico , Kit de Reagentes para Diagnóstico/veterinária , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Antivirais/imunologia , Bovinos , Cromatografia/métodos , Cromatografia/veterinária , Colódio , Ensaio de Imunoadsorção Enzimática , Febre Aftosa/virologia , Cabras , Humanos , Filtros Microporos , Sensibilidade e Especificidade , Ovinos , SuínosRESUMO
OBJECTIVE: To explore the concurrent validity, responsiveness, and floor- and ceiling-effects of the 2 items of Action Research Arm Test (ARAT-2) in comparison with the original ARAT and the Fugl-Meyer Assessment for Upper Extremity (FMA-UE) during the first 4 weeks post-stroke. DESIGN: A prospective longitudinal cohort study. SUBJECTS: A non-selected cohort of 117 adults with first-ever stroke and impaired upper extremity function. METHODS: The activity capacity and motor function was assessed with ARAT and FMA-UE at 3 days, 10 days and 4 weeks post-stroke. RESULTS: Correlation between ARAT-2 and the other assessment scales was high (r=0.920.97) and ARAT-2 showed statistically significant changes between all time-points (effect size, r=0.310.48). The effect sizes for the change in ARAT and FMA-UE varied from 0.44 to 0.53. ARAT-2, similarly to ARAT, showed a floor effect at all time-points. The ceiling effect was reached earlier using ARAT-2 than with ARAT and FMA-UE. CONCLUSION: ARAT-2 appears to be valid and a responsive short assessment for upper extremity activity capacity, and suitable for use in the acute stage after stroke. However, when the highest score has been reached, the assessment needs to be complemented with other instruments.
Assuntos
Avaliação da Deficiência , Avaliação de Resultados em Cuidados de Saúde/métodos , Reabilitação do Acidente Vascular Cerebral/estatística & dados numéricos , Acidente Vascular Cerebral/fisiopatologia , Extremidade Superior/fisiopatologia , Adulto , Idoso , Feminino , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Recuperação de Função Fisiológica , Reprodutibilidade dos TestesRESUMO
Two lateral flow devices (LFD) for the detection of vesicular stomatitis (VS) virus (VSV), types Indiana (VSV-IND) and New Jersey (VSV-NJ) were developed using monoclonal antibodies C1 and F25VSVNJ-45 to the respective VSV serotypes. The performance of the LFDs was evaluated in the laboratory on suspensions of vesicular epithelia and cell culture passage derived supernatants of VSV. The collection of test samples included 105 positive for VSV-IND (92 vesicular epithelial suspensions and 13 cell culture antigens; encompassing 93 samples of subtype 1 [VSV-IND-1], 9 of subtype 2 [VSV-IND-2] and 3 of subtype 3 [VSV-IND-3]) and 189 positive for VSV-NJ (162 vesicular epithelial suspensions and 27 cell culture antigens) from suspected cases of vesicular disease in cattle and horses collected from 11 countries between 1937 and 2008 or else were derived from experimental infection and 777 samples that were either shown to be positive or negative for foot-and-mouth disease (FMD) virus (FMDV) and swine vesicular disease virus (SVDV) or else collected from healthy cattle or pigs and collected from 68 countries between 1965 and 2011. The diagnostic sensitivity of the VSV-IND (for reaction with VSV-IND-1) and VSV-NJ LFDs was either similar or identical at 94.6% (VSV-IND) and 97.4% (VSV-NJ) compared to 92.5% and 97.4% obtained by the reference method of antigen ELISA. The VSV-IND LFD failed to react with viruses of VSV-IND-2 and 3, while the VSV-NJ device recognized all VSV-NJ virus strains. The diagnostic specificities of the VSV-IND and VSV-NJ LFDs were 99.1% and 100, respectively, compared to 99.6% and 99.8% for the ELISA. Reactions with FMDV which can produce indistinguishable syndromes clinically in cattle, pigs and sheep and SVDV (vesicular disease in pigs) did not occur. These data illustrate the potential for the LFDs to be used next to the animal for providing rapid and objective support to veterinarians in their clinical judgment of vesicular disease and for the subtype (VSV-IND-1) and type-specific (VSV-NJ) pen-side diagnosis of VS and differential diagnosis from FMD.
Assuntos
Febre Aftosa/diagnóstico , Laboratórios/provisão & distribuição , Estomatite Vesicular/diagnóstico , Vírus da Estomatite Vesicular Indiana/isolamento & purificação , Vírus da Estomatite Vesicular New Jersey/isolamento & purificação , Animais , Bovinos , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/virologia , Diagnóstico Diferencial , Enterovirus Humano B/isolamento & purificação , Infecções por Enterovirus/diagnóstico , Infecções por Enterovirus/virologia , Ensaio de Imunoadsorção Enzimática , Vírus da Febre Aftosa , Sensibilidade e Especificidade , Ovinos , Doenças dos Ovinos/diagnóstico , Doenças dos Ovinos/virologia , Suínos , Estomatite Vesicular/virologiaRESUMO
A lateral flow device (LFD) for the detection of swine vesicular disease (SVD) virus (SVDV) and differential diagnosis from foot-and-mouth disease (FMD) was developed using a monoclonal antibody (Mab C70). The performance of the LFD was evaluated in the laboratory on suspensions of vesicular epithelia and cell culture passage derived supernatants of SVDV and porcine teschovirus (enterovirus; PEV). The collection of test samples included 157 which were positive for SVDV (84 vesicular epithelial suspensions and 73 cell culture antigens) from suspected cases of vesicular disease in pigs collected from 14 countries between 1966 and 2008 and 663 samples which were either shown to be negative for SVDV and FMD virus (FMDV) or else collected from healthy pigs or demonstrated to be positive for FMDV, PEV or vesicular exanthema (VEV) and collected from 16 countries between 1965 and 2008 or else were derived from experimental animals. Three further samples containing vesicular stomatitis virus (VSV) were also tested. The diagnostic sensitivity of the LFD for SVDV was similar at 82% compared to 86% obtained by the reference method of antigen ELISA, and the diagnostic specificity was 100% compared to 99.7% for the ELISA. The device recognized virus strains of each of the known genotypes of the sole SVDV serotype. Reactions with FMDV, VEV, VSV and PEV which can produce clinically indistinguishable syndromes in pigs, did not occur. These data illustrate the potential for the LFD to be used next to the animal for providing rapid and objective support to veterinarians in their clinical judgment of vesicular disease in pigs and for the specific pen-side diagnosis of SVD and differential diagnosis from FMD.
Assuntos
Técnicas de Laboratório Clínico/métodos , Enterovirus Humano B/isolamento & purificação , Doença Vesicular Suína/diagnóstico , Animais , Anticorpos Monoclonais , Anticorpos Antivirais , Diagnóstico Diferencial , Ensaio de Imunoadsorção Enzimática/métodos , Febre Aftosa/diagnóstico , Sensibilidade e Especificidade , Suínos/virologia , Doença Vesicular Suína/virologiaRESUMO
A lateral flow device (LFD) for the detection of foot-and-mouth disease virus (FMDV) of the SAT 2 serotype was developed using a monoclonal antibody (Mab 2H6). The performance of the LFD was evaluated in the laboratory on suspensions of vesicular epithelia: 305 positive for FMDV type SAT 2 from suspected cases of vesicular disease collected from 30 countries and 1002 samples shown to be negative for FMDV type SAT 2 collected from 67 countries between 1968 and 2008. The diagnostic sensitivity of the LFD for FMDV type SAT 2 was higher at 88% compared to 79% obtained by the reference method of antigen ELISA, and the diagnostic specificity of the LFD was approximately 99% compared to 100% for the ELISA. The device recognized FMDV strains of wide diversity within the FMDV SAT 2 serotype and gave a superior performance for their detection compared to the 1F10 LFD which had been developed previously and shown to perform less well for the detection of FMDVs of this particular serotype. Reactions in the SAT 2 2H6 LFD with the viruses of other FMDV serotypes and swine vesicular disease (which produces a clinically indistinguishable syndrome in pigs), did not occur. These data illustrate the potential for the LFD to be employed to complement the 1F10 device next to the animal in the pen-side diagnosis of FMD, for providing rapid and objective support to veterinarians in their clinical judgment of the disease and for specific confirmation of a FMDV type SAT 2 infection.