Bovine purified protein derivative (bPPD) and avian purified protein derivative (aPPD) are widely used for bovine tuberculosis diagnosis. However, little is known about their qualitative and quantitative characteristics, which makes their standardisation difficult. In addition, bPPD can give false-positive tuberculosis results because of sequence homology between Mycobacterium bovis (M. bovis) and M. avium proteins. Thus, the objective of this study was to carry out a proteomic characterisation of bPPD, aPPD and an immunopurified subcomplex from bPPD called P22 in order to identify proteins contributing to cross-reactivity among these three products in tuberculosis diagnosis. Trypsin digests of bPPD, aPPD and P22 were analysed by nanoscale liquid chromatography-electrospray ionization tandem mass spectrometry. Mice were immunised with bPPD or aPPD, and their serum was tested by indirect ELISA for reactivity against these preparations as well as against P22. A total of 456 proteins were identified in bPPD, 1019 in aPPD and 118 in P22; 146 of these proteins were shared by bPPD and aPPD, and 43 were present in all three preparations. Candidate proteins that may cause cross-reactivity between bPPD and aPPD were identified based on protein abundance and antigenic propensity. Serum reactivity experiments indicated that P22 may provide greater specificity than bPPD with similar sensitivity for ELISA-type detection of antibodies against M. tuberculosis complex. The subpreparation from bPPD called P22 may be an alternative to bPPD for serodiagnosis of bovine tuberculosis, since it shares fewer proteins with aPPD than bPPD does, reducing risk of cross-reactivity with anti-M. avium antibodies.

Background: To control the transmission of relevant shared diseases, such as animal tuberculosis (TB) and African swine fever (ASF), it is essential to reduce the risk of interaction between livestock and wild ungulates. In Eastern and Central Europe, the current spread of ASF virus affecting wild boar and domestic pigs (especially those raised outdoors and/or in backyards) has devastated the pig sector in affected regions and is seriously threatening other exporting countries. Here, we evaluated the risk of wildlife-livestock interactions on 45 outdoor pig farms in Spain, the second largest pork producer in the EU and then proposed biosecurity-related actions. An integrated, systematic wildlife risk mitigation protocol based on interviews, questionnaires and field audits was developed and applied on each farm. Results: Most of the interaction risk points were associated with water sources (84.2%; 701/832), mainly springs and ponds, which accounted for almost all the specific points with high or very high risk scores. The risk of interaction at feeding points (6.9%; 57/832) and those associated with facilities for livestock and/or game management (8.9%; 74/832) were rated as low and very low risk, respectively. Wild boar were present and hunted on 69% of the farms. Supplementary feeding for wild ungulate species (mainly wild boar) was provided on almost half (48.9%; 22/45) the surveyed farms. Risk mitigation actions were categorised to target water access, waterers, food, other livestock species, grazing, wildlife, and offal disposal. Of the total number of actions (n = 2016), 82.7% were identified as priority actions while 17.3% represented alternative options which were identified less cost-effective. On average, 37.1 (median: 32; range 14–113) action proposals per study farm were made and 2.0 (median: 1; range 0–4) per risk point. The mean estimated cost of implementing the proposed priority actions was 14,780 €/farm (25.7 €/hectare and 799.4 €/risk point). Conclusions: This study expands the knowledge of interaction risks between domestic pigs and wild ungulates in outdoor pig farming systems and highlights the importance of considering local risks and management practices when designing and prioritising adapted wildlife risk mitigation and biosecurity actions. This practical and feasible protocol developed for Mediterranean ecosystems is easily transferable to professionals and can be adapted to extensive (outdoor) production or epidemiological systems in other European regions.