Accuracy and inter-rater reliability of lung auscultation by bovine practitioners when compared with ultrasonographic findings.

In practice, veterinary surgeons frequently rely on lung auscultation as a confirmation test for pneumonia. To what extent diagnostic accuracy of lung auscultation varies between different practitioners is currently unknown. In this diagnostic test study, 49 Dutch veterinarians each auscultated between 8 and 10 calves, and communicated whether they would decide to treat the animal with antimicrobials or not. They were not allowed to perform any other aspect of the clinical examination. Their decisions were compared with lung ultrasonography findings. The average sensitivity and specificity of lung auscultation were 0.63 (sd=0.2; range=0.2-1.0) and 0.46 (sd=0.3; range=0.0-1.0), respectively. Of the participants, 8.2 per cent were 100 per cent sensitive, 16.3 per cent were 100 per cent specific, and only 4.0 per cent were perfect. The Krippendorff's alpha was 0.18 (95 per cent confidence interval: -0.01 to 0.38), signifying poor reliability between multiple raters. Regardless of the poor diagnostic accuracy in this study, especially the large variation in a confirmation test between different practitioners could potentially cause professional damage as well as misuse of antimicrobials. This study could be seen as a gentle stimulus to regularly evaluate one's diagnostic skills. Both complementary training and the use of more accurate techniques with less inter-rater variation could improve the situation.

Rethinking the role of alpha toxin in Clostridium perfringens-associated enteric diseases: a review on bovine necro-haemorrhagic enteritis.

Bovine necro-haemorrhagic enteritis is an economically important disease caused by Clostridium perfringens type A strains. The disease mainly affects calves under intensive rearing conditions and is characterized by sudden death associated with small intestinal haemorrhage, necrosis and mucosal neutrophil infiltration. The common assumption that, when causing intestinal disease, C. perfringens relies upon specific, plasmid-encoded toxins, was recently challenged by the finding that alpha toxin, which is produced by all C. perfringens strains, is essential for necro-haemorrhagic enteritis. In addition to alpha toxin, other C. perfringens toxins and/or enzymes might contribute to the pathogenesis of necro-haemorrhagic enteritis. These additional virulence factors might contribute to breakdown of the protective mucus layer during initial stage of pathogenesis, after which alpha toxin, either or not in synergy with other toxins such as perfringolysin O, can act on the mucosal tissue. Furthermore, alpha toxin alone does not cause intestinal necrosis, indicating that other virulence factors might be needed to cause the extensive tissue necrosis observed in necro-haemorrhagic enteritis. This review summarizes recent research that has increased our understanding of the pathogenesis of bovine necro-haemorrhagic enteritis and provides information that is indispensable for the development of novel control strategies, including vaccines.

Toxin-neutralizing antibodies protect against Clostridium perfringens-induced necrosis in an intestinal loop model for bovine necrohemorrhagic enteritis.

BACKGROUND: Bovine necrohemorrhagic enteritis is caused by Clostridium perfringens type A. Due to the rapid progress and fatal outcome of the disease, vaccination would be of high value. In this study, C. perfringens toxins, either as native toxins or after formaldehyde inactivation, were evaluated as possible vaccine antigens. We determined whether antisera raised in calves against these toxins were able to protect against C. perfringens challenge in an intestinal loop model for bovine necrohemorrhagic enteritis. RESULTS: Alpha toxin and perfringolysin O were identified as the most immunogenic proteins in the vaccine preparations. All vaccines evoked a high antibody response against the causative toxins, alpha toxin and perfringolysin O, as detected by ELISA. All antibodies were able to inhibit the activity of alpha toxin and perfringolysin O in vitro. However, the antibodies raised against the native toxins were more inhibitory to the C. perfringens-induced cytotoxicity (as tested on bovine endothelial cells) and only these antibodies protected against C. perfringens challenge in the intestinal loop model. CONCLUSION: Although immunization of calves with both native and formaldehyde inactivated toxins resulted in high antibody titers against alpha toxin and perfringolysin O, only antibodies raised against native toxins protect against C. perfringens challenge in an intestinal loop model for bovine necrohemorrhagic enteritis.

The C-terminal domain of Clostridium perfringens alpha toxin as a vaccine candidate against bovine necrohemorrhagic enteritis.

Bovine necrohemorrhagic enteritis is caused by Clostridium perfringens and leads to sudden death. Alpha toxin, together with perfringolysin O, has been identified as the principal toxin involved in the pathogenesis. We assessed the potential of alpha toxin as a vaccine antigen. Using an intestinal loop model in calves, we investigated the protection afforded by antisera raised against native alpha toxin or its non-toxic C-terminal fragment against C. perfringens-induced intestinal necrosis. Immunization of calves with either of the vaccine preparations induced a strong antibody response. The resulting antisera were able to neutralize the alpha toxin activity and the C. perfringens-induced endothelial cytotoxicity in vitro. The antisera raised against the native toxin had a stronger neutralizing activity than those against the C-terminal fragment. However, antibodies against alpha toxin alone were not sufficient to completely neutralize the C. perfringens-induced necrosis in the intestinal loop model. The development of a multivalent vaccine combining the C-terminal fragment of alpha toxin with other C. perfringens virulence factors might be necessary for complete protection against bovine necrohemorrhagic enteritis.

Clostridium perfringens strains from bovine enterotoxemia cases are not superior in in vitro production of alpha toxin, perfringolysin O and proteolytic enzymes.

BACKGROUND: Bovine enterotoxemia is a major cause of mortality in veal calves. Predominantly veal calves of beef cattle breeds are affected and losses due to enterotoxemia may account for up to 20% of total mortality. Clostridium perfringens type A is considered to be the causative agent. Recently, alpha toxin and perfringolysin O have been proposed to play an essential role in the development of disease. However, other potential virulence factors also may play a role in the pathogenesis of bovine enterotoxemia. The aim of this study was to evaluate whether strains originating from bovine enterotoxemia cases were superior in in vitro production of virulence factors (alpha toxin, perfringolysin O, mucinase, collagenase) that are potentially involved in enterotoxemia. To approach this, a collection of strains originating from enterotoxemia cases was compared to bovine strains isolated from healthy animals and to strains isolated from other animal species. RESULTS: Strains originating from bovine enterotoxemia cases produced variable levels of alpha toxin and perfringolysin O that were not significantly different from levels produced by strains isolated from healthy calves and other animal species. All tested strains exhibited similar mucinolytic activity independent of the isolation source. A high variability in collagenase activity between strains could be observed, and no higher collagenase levels were produced in vitro by strains isolated from enterotoxemia cases. CONCLUSIONS: Bovine enterotoxemia strains do not produce higher levels of alpha toxin, perfringolysin O, mucinase and collagenase, as compared to strains derived from healthy calves and other animal species in vitro.