Development of a peroxidase‐antiperoxidase (PAP) technique for the identification of Haemophilus somnus in pneumonic calf lungs in Denmark

A peroxidase-antiperoxidase (PAP) technique was developed for the identification of Haemophilus somnus bacteria in lung tissues of calves. Antisera raised against somatic and wall antigens of a Danish and American strain of H. somnus were produced. Experimentally infected murine tissues were used for the determination of the sensitivity and specificity of antiserum that had been heterologously absorbed with antigens of cross-reacting bacteria, i.e. Pasteurella haemolytica and Pasteurella multocida. None of the antisera reacted with Actinomyces pyogenes. An antiserum raised against somatic antigens of the Danish strain of H. somnus revealed the highest sensitivity in the PAP technique and became specific following absorption. Heterologous absorption also rendered this antiserum specific in crossed immunoelectrophoresis. Subsequently, the PAP technique was applied on formalin-fixed pneumonic lung tissues of 86 calves. An immunodiagnosis of H. somnus pneumonia was obtained in 15 of 17 lungs from which the bacterium had been isolated. Moreover, immunostained bacteria were also demonstrated in 20 lungs from which H. somnus had not been isolated. Thus, application of immunohistochemistry significantly enhanced the diagnostic sensitivity of H. somnus pneumonia of calves and should be used as a potent supplementary tool for the routine screening of suspected lung tissues of calves from which bacterial isolation is negative.