Sensitivity and specificity of a competitive enzyme immunoassay in the serodiagnosis of bovine brucellosis

O trabalho teve por objetivo avaliar a sensibilidade e a especificidade de um teste imunoenzimático competitivo, empregando como conjugado os anticorpos monoclonais BM-38 e BM-40, no diagnóstico sorológico da brucelose bovina. Foram examinados 74 soros de bovinos dos quais havia sido isolada Brucella abortus e 2.118 soros de bovinos procedentes de rebanhos livres de brucelose e que apresentaram resultado negativo quando submetidos ao teste Rosa Bengala. O teste imunoenzimático competitivo, usando qualquer dos dois conjugados, foi capaz de revelar a presença de anticorpos contra o lipopolissacáride bacteriano em todos os soros de bovinos infectados, o que resulta em uma sensibilidade de 100%. A especificidade do teste usando o conjugado BM-38 foi de 98.82% e usando o conjugado BM-40 foi de 99,95%. Estes resultados indicam que o teste imunoenzimático competitivo, principalmente ao se empregar o conjugado BM-40, consiste em um método bastante útil para ser usado como teste confirmatório no diagnóstico sorológico da brucelose bovina.The purpose of this work was to evaluate the sensitivity and the specificity of a competitive enzyme immunoassay, using as conjugate the monoclonal antibodies BM-38 and BM-40, in the serodiagnosis of bovine brucellosis. Seventy-four sera from culture-positive cattle and 2,118 cattle sera from herds free from brucellosis and negative to the Rose Bengal plate test were examined. The competitive enzyme immunoassay, using any of the two conjugates, was able to reveal the presence of antibodies to Brucella lipopolysaccharide in all of the 74 sera of the infected cattle, resulting in a sensitivity of 100%. The specificity of the test using the conjugate BM-38 was 98.82% and using the conjugate BM-40 was 99.95%. These results indicated that the competitive enzyme immunoassay, mainly when using the conjugate BM-40, consists in a technique very useful in the confirmation of the serological diagnosis of bovine brucellosis

Comparative studies on the pathogenicity and tissue distribution of three virulence variants of classical swine fever virus, two field isolates and one vaccine strain, with special regard to immunohistochemical investigations

<p>Abstract</p> <p>Background</p> <p>The aim of this study was to compare the tissue distribution and pathogenicity of three virulence variants of classical swine fever virus (CSFV) and to investigate the applicability of various conventional diagnostic procedures.</p> <p>Methods</p> <p>64 pigs were divided into three groups and infected with the highly virulent isolate ISS/60, the moderately virulent isolate Wingene'93 and the live attenuated vaccine strain Riems, respectively. Clinical signs, gross and histopathological changes were compared in relation to time elapsed post infection. Virus spread in various organs was followed by virus isolation, by immunohistochemistry, applying monoclonal antibodies in a two-step method and by <it>in situ </it>hybridisation using a digoxigenin-labelled riboprobe.</p> <p>Results</p> <p>The tissue distribution data are discussed in details, analyzing the results of the various diagnostic approaches. The comparative studies revealed remarkable differences in the onset of clinical signs as well as in the development of the macro- and microscopical changes, and in the tissue distribution of CSFV in the three experimental groups.</p> <p>Conclusion</p> <p>The present study demonstrates that in the case of highly and moderately virulent virus variants the virulence does not affect the pattern of the viral spread, however, it influences the outcome, the duration and the intensity of the disease. Immunohistochemistry has the advantage to allow the rapid detection and localisation of the virus, especially in cases of early infection, when clinical signs are still absent. Compared to virus isolation, the advantage of this method is that no cell culture facilities are required. Thus, immunohistochemistry provides simple and sensitive tools for the prompt detection of newly emerging variants of CSFV, including the viruses of very mild virulence.</p

Clustering of classical swine fever virus isolates by codon pair bias

<p>Abstract</p> <p>Background</p> <p>The genetic code consists of non-random usage of synonymous codons for the same amino acids, termed codon bias or codon usage. Codon juxtaposition is also non-random, referred to as codon context bias or codon pair bias. The codon and codon pair bias vary among different organisms, as well as with viruses. Reasons for these differences are not completely understood. For classical swine fever virus (CSFV), it was suggested that the synonymous codon usage does not significantly influence virulence, but the relationship between variations in codon pair usage and CSFV virulence is unknown. Virulence can be related to the fitness of a virus: Differences in codon pair usage influence genome translation efficiency, which may in turn relate to the fitness of a virus. Accordingly, the potential of the codon pair bias for clustering CSFV isolates into classes of different virulence was investigated.</p> <p>Results</p> <p>The complete genomic sequences encoding the viral polyprotein of 52 different CSFV isolates were analyzed. This included 49 sequences from the GenBank database (NCBI) and three newly sequenced genomes. The codon usage did not differ among isolates of different virulence or genotype. In contrast, a clustering of isolates based on their codon pair bias was observed, clearly discriminating highly virulent isolates and vaccine strains on one side from moderately virulent strains on the other side. However, phylogenetic trees based on the codon pair bias and on the primary nucleotide sequence resulted in a very similar genotype distribution.</p> <p>Conclusion</p> <p>Clustering of CSFV genomes based on their codon pair bias correlate with the genotype rather than with the virulence of the isolates.</p

Challenge of Pigs with Classical Swine Fever Viruses after C-Strain Vaccination Reveals Remarkably Rapid Protection and Insights into Early Immunity

Pre-emptive culling is becoming increasingly questioned as a means of controlling animal diseases, including classical swine fever (CSF). This has prompted discussions on the use of emergency vaccination to control future CSF outbreaks in domestic pigs. Despite a long history of safe use in endemic areas, there is a paucity of data on aspects important to emergency strategies, such as how rapidly CSFV vaccines would protect against transmission, and if this protection is equivalent for all viral genotypes, including highly divergent genotype 3 strains. To evaluate these questions, pigs were vaccinated with the Riemser® C-strain vaccine at 1, 3 and 5 days prior to challenge with genotype 2.1 and 3.3 challenge strains. The vaccine provided equivalent protection against clinical disease caused by for the two challenge strains and, as expected, protection was complete at 5 days post-vaccination. Substantial protection was achieved after 3 days, which was sufficient to prevent transmission of the 3.3 strain to animals in direct contact. Even by one day post-vaccination approximately half the animals were partially protected, and were able to control the infection, indicating that a reduction of the infectious potential is achieved very rapidly after vaccination. There was a close temporal correlation between T cell IFN-γ responses and protection. Interestingly, compared to responses of animals challenged 5 days after vaccination, challenge of animals 3 or 1 days post-vaccination resulted in impaired vaccine-induced T cell responses. This, together with the failure to detect a T cell IFN-γ response in unprotected and unvaccinated animals, indicates that virulent CSFV can inhibit the potent antiviral host defences primed by C-strain in the early period post vaccination

Human Muscle Satellite Cells as Targets of Chikungunya Virus Infection

BACKGROUND: Chikungunya (CHIK) virus is a mosquito-transmitted alphavirus that causes in humans an acute infection characterised by fever, polyarthralgia, head-ache, and myalgia. Since 2005, the emergence of CHIK virus was associated with an unprecedented magnitude outbreak of CHIK disease in the Indian Ocean. Clinically, this outbreak was characterized by invalidating poly-arthralgia, with myalgia being reported in 97.7% of cases. Since the cellular targets of CHIK virus in humans are unknown, we studied the pathogenic events and targets of CHIK infection in skeletal muscle. METHODOLOGY/PRINCIPAL FINDINGS: Immunohistology on muscle biopsies from two CHIK virus-infected patients with myositic syndrome showed that viral antigens were found exclusively inside skeletal muscle progenitor cells (designed as satelllite cells), and not in muscle fibers. To evaluate the ability of CHIK virus to replicate in human satellite cells, we assessed virus infection on primary human muscle cells; viral growth was observed in CHIK virus-infected satellite cells with a cytopathic effect, whereas myotubes were essentially refractory to infection. CONCLUSIONS/SIGNIFICANCE: This report provides new insights into CHIK virus pathogenesis, since it is the first to identify a cellular target of CHIK virus in humans and to report a selective infection of muscle satellite cells by a viral agent in humans

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