Development of polymerase chain reaction and comparison with in situ hybridization for the detection of Haemophilus parasuis in formalin-fixed, paraffin-embedded tissues.

DNA extraction and nested polymerase chain reaction (PCR) were developed for the detection of Haemophilus parasuis from formalin-fixed, paraffin-embedded tissues. The results for nested PCR were compared with those determined by in situ hybridization. The optimal results obtained show that use of xylene deparaffinization, digestion with proteinase K followed by nested PCR is a reliable detection method. A distinct positive signal was detected in 20 pigs naturally infected with H. parasuis by in situ hybridization. The rate of agreement between nested PCR and in situ hybridization for the detection of H. parasuis in formalin-fixed, paraffin-embedded tissues was 100%. The nested PCR could be applied successfully to formalin-fixed, paraffin-embedded tissues for the detection of H. parasuis with bacterial isolation.

Lectin histochemical study of lipopigments present in the cerebellum of Solanum fastigiatum var. fastigiatum intoxicated cattle.

This study was carried out to investigate the pattern of lectin binding in the cerebellum of calves poisoned with Solanum fastigiatum var. fastigiatum. For the experimental reproduction of the illness, S. fastigiatum var. fastigiatum was collected from farms where the intoxication occurs. The dried ground plant was administered to two 1-year-old cattle by a ruminal cannula. The animals received 5 g/kg b.w. daily, 5 days a week, during periods of 107 and 140 days. After these periods the animals were bled to death. For the histological study, transverse sections of the cerebellum were used. Paraffin-embedded sections were incubated with the following biotinylated lectins with different specificity: Concanavalia ensiformis (Con-A). Glycine max (SBA). Dolichos hiflorus (DBA), Ulex europeus-I (UEA-I). Triticum vulgaris (WGA), succynyl-WGA (sWGA). Arachis hypogaea (PNA), Ricinus communis-I (RCA-I) and Bandeirea simplicifolia-I (BS-I). Avidin-biotin-peroxidase complex was applied as a detection system. Purkinje cells showed vacuolation in the pericaryon. The stored material present in the cells reacted strongly with the following lectins: Con-A. sWGA, WGA and RCA-I. An irregular affinity was observed with PNA and DBA. The lectin-binding pattern was compatible with a glycolipid storage disease.

Immunohistochemical identification of caprine arthritis-encephalitis virus in paraffin-embedded specimens from naturally infected goats.

The expression of caprine arthritis-encephalitis virus capsid protein was studied in seropositive naturally infected asymptomatic goals (10< seropositive naturally infected encephalitic kids (12) and goats (4), and noninfected control goats (3). Rabbit antiserum to recombinant viral capsid and matrix proteins were used in a biotin-streptavidin-alkaline phosphatase complex immunohistochemical method on sections of formalin- and ethanol-fixed tissue specimens. Macrophages in inflamed areas of the lung (8/12), in the brain (5/16), and in the spinal cord (4/16) from encephalitic animals harbored viral antigens, as revealed by immunohistochemistry and use of a capsid protein-specific antiserum. Altogether 12/16 encephalitic animals tested positive for viral antigen. Viral antigens were found in 5/10 seropositive asymptomatic goals in macrophages located in the lung (3), the udder (1), and the medulla of lymph nodes (4). None of the control animals tested positive for viral antigen. Ethanol fixation showed highest sensitivity, and the lowest antigen concentration that revealed a positive signal discernible from background was twofold higher in ethanol-fixed specimens than in formalin-fixed specimens. The evaluation was performed on artificial antigen substrates embedded with defined concentrations of recombinant viral capsid protein. Immunohistochemistry is a valuable supplement to the methods presently available for diagnosis in cases suspicious of caprine arthritis-encephalitis.