The eradication of tuberculosis, brucellosis and leucosis in cattle has not yet been achieved in the entire Italian territory. The region of Lazio, Central Italy, represents an interesting case study to evaluate the evolution of costs for these eradication programmes, as in some provinces the eradication has been officially achieved, in some others the prevalence has been close to zero for years, and in still others disease outbreaks have been continuously reported. The objectives of this study were i) to describe the costs for the eradication programmes for tuberculosis, brucellosis and leucosis in cattle carried out in Lazio between 2007 and 2011, ii) to calculate the ratio between the financial contribution of the European Union (EU) for the eradication programmes and the estimated total costs and iii) to estimate the potential savings that can be made when a province gains the certification of freedom from disease. For the i) and ii) objectives, data were collected from official sources and a costing procedure was applied from the perspective of the Regional Health Service. For the iii) objective, a Bayesian AR(1) regression was used to evaluate the average percentage reduction in costs for a province that gained the certification. The total cost for the eradication programmes adjusted for inflation to 1 January 2016 was estimated at 18 919 797 euro (5th and 95th percentiles of the distribution: 18 325 050-19 552 080 euro). When a province gained the certification of freedom from disease, costs decreased on average by (median of the posterior distribution) 47.5%, 54.5% and 54.9% for the eradication programmes of tuberculosis, brucellosis and leucosis, respectively. Information on possible savings from the reduction of control costs can help policy makers operating under budget constraints to justify the use of additional resources for the final phase of eradication.

The aim of this study was to develop and validate different innovative DNA extraction methods to detect Mycobacterium avium subsp. paratuberculosis (MAP) DNA from bovine and buffalo colostrum. Paratuberculosis is a chronic inflammatory infection of domestic and wild animals, especially ruminants, caused by MAP. The primary route of disease transmission is feces, but MAP can also be excreted in milk and colostrum. In 2015, the Italian Ministry of Health has issued a voluntary control plan of MAP in order to allow risk‐based certification of bovine and buffaloes farms. In addition to the annual diagnostic screening and to the clinical surveillance of animals the plan includes the adoption of biosecurity and management measures to progressively mitigate the incidence of MAP. To achieve this goal it is crucial to ensure the accuracy of the methods used to detect the presence of MAP in bovine and buffaloes milk and colostrum, in order to: (1) support a "safe colostrum farm‐bank" set‐up and thus prevent the main within‐farm MAP transmission route and (2) to allow the MAP‐free certification of milk products for export purposes. To achieve these goals, seven different DNA extraction protocols were identified from bibliography, out of which three methods were finally selected after the adoption of an evaluation procedure aimed at assessing the efficiency of extraction of DNA, the purity of DNA and the adaptability of the DNA amplification: NucleoSpin® Food Kit (Macherey‐Nagel), NucleoSpin® Food Kit (Macherey‐Nagel) combined with the magnetic beads, and QIAamp Cador Pathogen Mini kit (QIAGEN). In particular, the NucleoSpin® Food Kit (Macherey‐Nagel) and the QIAamp Cador Pathogen Mini kit (QIAGEN) were tested on bovine and buffalo colostrum, showing a LOD between 4 × 104 (2.6 × 106 cfu/ml) and 4.08 (26.7 cfu/ml) IS900 target copies and a LOD between 5.3 × 105 (4.1 × 106 cfu/ml) and 53 (4.1 × 103 cfu/ml) IS900 target copies, respectively.

Bovine tuberculosis (bTB) is a worldwide zoonosis that affects many species of domestic and wild animals. Mycobaterium bovis is the main cause of infection in water buffalo (Bubalus bubalis) and bovines and is of great concern for human health and for buffalo producers in Italy. The bTB eradication programme is based on slaughterhouse surveillance and intradermal skin tests. Other in vivo diagnostic methods such as the interferon-gamma (IFN-γ) assay have been developed and are widely used in cattle to accelerate the elimination of bTB positive animals. The present study is the first to assess the use and performance of IFN-γ assays, which is used as an ancillary test for bTB diagnosis in water buffalo, and presents the results of a field-evaluation of the assay from 2012 to 2019 during the buffalo bTB eradication programme in Italy. The study involved 489 buffaloes with a positive result to the single intradermal tuberculin test (SITT). The IFN-γ assays and single intradermal comparative tuberculin test were used as confirmation tests. Then, a total of 458 buffaloes, reared on officially tuberculosis-free (OTF) herds, that were confirmed bTB-free for at least the last 6 years were subjected to IFN-γ testing. Furthermore, to evaluate the IFN-γ test in an OTF herd with Paratuberculosis (PTB) infection, 103 buffaloes were subjected to SITT and IFN-γ test simultaneously. Four interpretative criteria were used, and the IFN-γ test showed high levels of accuracy, with sensitivity levels between 75.3% (CI 95% 71.2–79.0%) and 98.4% (CI 95% 96.7–99.4%) and specificity levels between 94.3% (CI 95% 91.2–96.50%) and 98.5% (CI 95% 96.9–99.4%), depending on the criterion used. Finally, in the OTF herd with PTB infection, in buffalo, the IFN-γ test displayed high specificity values according to all 4 interpretative criteria, with specificity levels between 96.7% (CI 95% 88.4–99.5%) and 100% (CI 95% 96.2–100%), while SITT specificity proved unsatisfactory, with a level of 45.3% (CI 95% 35.0–55.7%). Our results showed that the IFN-γ test in the buffalo species could reach high Sensitivity and Specificity values, and that the level of Sensitivity and Specificity could be chosen based on the interpretative criterion and the antigens used depending on the health status of the herd and the epidemiological context of the territory. The IFN-γ test and the use of different interpretative criteria proved to be useful to implement bTB diagnostic strategies in buffalo herds, with the possibility of a flexible use of the assay.