Immunohistochemical Detection ofHaemophilus ParasuisSerovar 5 in Formalin-Fixed, Paraffin-Embedded Tissues of Experimentally Infected Swine

Abstract

Abstract. An avidin-biotin complex immunohistochemistry technique was developed to detect Haemophilus parasuis serovar 5 in experimentally infected 18-21-day-old conventional pigs, using a rabbit polyclonal antiserum. Seven of 10 intratracheally inoculated animals developed a low to medium degree of fibrinous polyserositis; meninges and pleura were the most severely affected areas. Haemophilus parasuis was recovered from 9 of 10 pigs; in 2 of them H. parasuis was isolated from tracheal swabs only. Positive immunohistochemistry results, mainly observed as free bacteria or bacteria within inflammatory cell cytoplasm in the fibrinopurulent exudate, were observed in 8 of 10 animals. Cross-reactivity with Actinobacillus pleuropneumoniae was detected but not with other gram-positive and gram-negative bacteria tested. This immunohistochemistry technique seemed to be at least as sensitive as microbiologic cultures and could be useful in studies of pathogenesis and retrospective diagnosis. However, cross-reactivity with A. pleuropneumoniae means that positive immunohistochemistry results in lung tissue from field cases would be dubious.

Haemophilus parasuis and Glässer's disease in pigs: a review

“…Amano et al (1994) successfully isolated H. parasuis from nasal cavities and tonsils after intranasal inoculation of pigs with serovars 1, 4 and 5. Segales et al (1997) described common isolation of H. parasuis from tonsil and tracheal swabs from pigs after intratracheal inoculation and Kirkwood et al (2001) isolated H. parasuis from nasal cavity swabs from infected pigs.…”

“…IHC allows detection of non-living microorganisms in the cytoplasm of phagocytes (Amano et al, 1994;Segales et al, 1997). However, some polyclonal antibodies used for diagnosis of H. parasuis by the IHC method cross reacted with A. pleuropneumoniae (Segales et al, 1997).…”

“…parasuis isolation from clinical samples is performed on blood agar close to S. aureus culture as a source of NAD, on chocolate agar or on NAD supplemented PPLO media. Cultivation time between 24-48 h is recommended (Segales et al, 1997). H. parasuis is one of demanding microorganisms; its isolation from clinical samples is usually difficult and often complicated by contaminations.…”

“…H. parasuis endotoxin causes disseminated intravascular coagulation which results in microthrombus formation in various tissues (Peet et al, 1983;Amano et al, 1994). H. parasuis is commonly isolated from nasal cavities (Amano et al, 1994;Vahle et al, 1997), tonsils (Oliveira et al, 2001a) and the upper part of trachea (Segales et al, 1997).…”

Development of Polymerase Chain Reaction and Comparison with In situ Hybridization for the Detection of Haemophilus parasuis in Formalin-Fixed, Paraffin-Embedded Tissues

“…However, isolation of H. parasuis is difficult and often unsuccessful due to the fastidious growth of the bacteria [14]. Furthermore, culture was frequently unsuccessful because most pigs had been treated with antibiotics before they submitted into diagnostic laboratory.…”