immunohistochemical detection of clostridia species in paraffin-embedded tissues of experimentally inoculated guinea pigs detecção imunohistoquímica de várias espécies de clostridia em tecidos parafinizados de cobaias inoculadas experimentalmente

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ID: 254854
2005
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Abstract
Blackleg is caused by Clostridium chauvoei, whereas malignant oedema is caused by C. chauvoei, C. septicum, C. sordellii, C. perfringens type A, and/or C. novyi type A. Anti-C. chauvoei, anti-C. septicum, anti-C. sordellii and anti-C. novyi type A polyclonal antibodies were produced in rabbits and purified in a column of DEAE-cellulose. Aliquots of the antisera were conjugated with fluorescein isothiocyanate and the remaining was used for the streptavidin biotin peroxidase technique (SBPT). SBPT was standardized to detect C. chauvoei, C. septicum, C. sordellii and C. novyi type A in formalin-fixed, paraffin-embedded tissues of guinea pigs. SBPT was compared to a fluorescent antibody technique (FAT). Sections and smears of muscle from inoculation area (MIA), heart, liver, spleen and kidney, were obtained for both SBPT and FAT. Cross-reactions between the different Clostridial species were not observed. C. chauvoei and C. septicum were detected in all specimens from the animals inoculated with these microorganisms, while only sections of muscle obtained from all the animals inoculated with C. sordellii and C. novyi type A were positive. The same results observed by the SBPT, were obtained on tissue smears of these microorganisms stained by the FAT. The results indicate that SBPT is suitable for detection of C. chauvoei, C. septicum, C. sordellii and C. novyi type A in formalin-fixed, paraffin-embedded tissues of guinea pigs.
O carbúnculo sintomático é causado pelo Clostridium chauvoei, enquanto o edema maligno é causado pelo C. chauvoei, C. septicum, C. sordellii, C. perfringens tipo A, e/ou C. novyi tipo A. Corpos policlonais anti-C. chauvoei, anti-C. septicum, anti-C. sordellii and anti-C.novyi tipo A foram produzidos em coelhos e purificados em uma coluna de DEAE-celulose. Alíquotas das imunoglobulinas foram conjugadas com isotiocianato de fluoresceína e o restante foi usado na técnica de streptavidina biotina peroxidase (SBP). SBP foi padronizada para detectar C. chauvoei, C. septicum, C. sordellii e C. novyi tipo A em tecidos de cobaias fixados em formol e incluídos em parafina. A mesma foi comparada com a técnica de imunofluorescência direta (IFD). Secções e impressões do músculo da área de inoculação, coração, fígado, baço e rim, foram obtidas para ambas as técnicas. Nenhuma reação cruzada foi observada quando tecidos inoculados com outras espécies de Clostridia foram tratadas com esses quatro anticorpos primários. C. chauvoei e C. septicum foram detectados em todos os espécimes provenientes dos animais inoculados com esses microrganismos, enquanto somente secções de músculo obtidas de todos os animais inoculados com C. sordellii e C. novyi tipo A foram positivas. Os mesmos resultados observados pela técnica de SBP, foram obtidos em impressões de tecidos desses microrganismos corados pela IFD. Os resultados indicam que a técnica de SBP permite a detecção de C. chauvoei, C. septicum, C. sordellii e C. novyi tipo A em tecidos de cobaias fixados em formol e incluídos em parafina.
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Authors Ronnie A. Assis, Francisco C.F. Lobato, Rogéria Serakides, Renato L. Santos, Guilherme R.C. Dias, Ricardo. A.P. Nascimento, Vera. L.V. Abreu, Patrícia M. Parreiras, Francisco A. Uzal
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Journal AIP Conference Proceedings
Year 2005
DOI
10.1590/S0100-736X2005000100002
URL
http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-736X2005000100002
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