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Abstract Vaccines are the most effective and sustainable intervention to control ticks and tick-borne diseases (TBD). Using a personalized vaccine design based on regional tick genotypes, a Rhipicephalus appendiculatus Subolesin protective antigen was used in a field trial evaluating tick vaccine efficacy, effectiveness, and safety in cattle infested with multiple tick species in different Ugandan agro-ecological zones. Vaccination with SUB was safe with a protective capacity against anemia and infection, and reduced the number of infested cattle, tick fitness (feeding and reproduction) with vaccine effectiveness against multiple tick species between 93.2% at 167-196 days post-vaccination (dpv) and 61.4% at 251–327 dpv. Total integrated vaccine efficacy/effectiveness was estimated as 98.8%. The Subolesin-based vaccine is protective against multiple cattle tick infestations under field conditions in Uganda. These results support registration and commercialization of the vaccine to reduce tick populations and associated risks for human and animal TBD and chemical acaracides in Uganda.

Abstract Vector or reservoir species of three diseases of crustaceans listed in the Animal Health Law were identified based on evidence generated through an extensive literature review, to support a possible updating of Regulation (EU) 2018/1882. Crustacean species on or in which Taura syndrome virus (TSV), Yellow head virus (YHV) or White spot syndrome virus (WSSV) were identified, in the field or during experiments, were classified as reservoir species with different levels of certainty depending on the diagnostic tests used. Where experimental evidence indicated transmission of the pathogen from a studied species to another known susceptible species, the studied species was classified as vector species. Although the quantification of the risk of spread of the pathogens by the vectors or reservoir species was not part of the terms of reference, such risks do exist for the vector species, since transmission from infected vector species to susceptible species was proven. Where evidence for transmission from infected crustaceans was not found, these were defined as reservoirs. Nonetheless, the risk of the spread of the pathogens from infected reservoir species cannot be excluded. Evidence identifying conditions that may prevent transmission by vectors during transport was collected from scientific literature. It was concluded that it is very likely to almost certain (90–100%) that WSSV, TSV and YHV will remain infective at any possible transport condition. Therefore, vector or reservoir species that may have been exposed to these pathogens in an affected area in the wild or aquaculture establishments or by water supply can possibly transmit WSSV, TSV and YHV.

Abstract Vector or reservoir species of five mollusc diseases listed in the Animal Health Law were identified, based on evidence generated through an extensive literature review, to support a possible updating of Regulation (EU) 2018/1882. Mollusc species on or in which Mikrocytos mackini, Perkinsus marinus, Bonamia exitiosa, Bonamia ostreae and Marteilia refringens were detected, in the field or during experiments, were classified as reservoir species with different levels of certainty depending on the diagnostic tests used. Where experimental evidence indicated transmission of the pathogen from a studied species to another known susceptible species, this studied species was classified as a vector species. Although the quantification of the risk of spread of the pathogens by the vectors or reservoir species was not part of the terms of reference, such risks do exist for the vector species, since transmission from infected vector species to susceptible species was proven. Where evidence for transmission from infected molluscs was not found, these were defined as reservoir. Nonetheless, the risk of the spread of the pathogens from infected reservoir species cannot be excluded. Evidence identifying conditions that may prevent transmission by vectors or reservoir mollusc species during transport was collected from scientific literature. It was concluded that it is very likely to almost certain (90–100%) that M. mackini, P. marinus, B. exitiosa B. ostreae and M. refringens will remain infective at any possible transport condition. Therefore, vector or reservoir species that may have been exposed to these pathogens in an affected area in the wild or at aquaculture establishments or through contaminated water supply can possibly transmit these pathogens. For transmission of M. refringens, the presence of an intermediate host, a copepod, is necessary.

Abstract Vector or reservoir species of five fish diseases listed in the Animal Health Law were identified, based on evidence generated through an extensive literature review (ELR), to support a possible updating of Regulation (EU) 2018/1882. Fish species on or in which highly polymorphic region‐deleted infectious salmon anaemia virus (HPR∆ ISAV), Koi herpes virus (KHV), epizootic haematopoietic necrosis virus (EHNV), infectious haematopoietic necrosis virus (IHNV) or viral haemorrhagic septicaemia virus (VHSV) were detected, in the field or during experiments, were classified as reservoir species with different levels of certainty depending on the diagnostic tests used. Where experimental evidence indicated transmission of the pathogen from a studied species to another known susceptible species, the studied species was classified as a vector species. Although the quantification of the risk of spread of the pathogens by the vectors or reservoir species was not part of the terms or reference, such risks do exist for the vector species, since transmission from infected vector species to susceptible species was proven. Where evidence for transmission from infected fish was not found, these were defined as reservoirs. Nonetheless, the risk of the spread of the pathogens from infected reservoir species cannot be excluded. Evidence identifying conditions that may prevent transmission by vectors or reservoir fish species during transport was collected from scientific literature. For VHSV, IHNV or HPR∆ ISAV, it was concluded that under transport conditions at temperatures below 25°C, it is likely (66–90%) they will remain infective. Therefore, vector or reservoir species that may have been exposed to these pathogens in an affected area in the wild, aquaculture establishments or through water supply can possibly transmit VHSV, IHNV or HPR∆ ISAV into a non‐affected area when transported at a temperature below 25°C. The conclusion was the same for EHN and KHV; however, they are likely to remain infective under all transport temperatures.

Abstract This Scientific Opinion addresses a European Commission request on the welfare of calves as part of the Farm to Fork strategy. EFSA was asked to provide a description of common husbandry systems and related welfare consequences, as well as measures to prevent or mitigate the hazards leading to them. In addition, recommendations on three specific issues were requested: welfare of calves reared for white veal (space, group housing, requirements of iron and fibre); risk of limited cow–calf contact; and animal‐based measures (ABMs) to monitor on‐farm welfare in slaughterhouses. The methodology developed by EFSA to address similar requests was followed. Fifteen highly relevant welfare consequences were identified, with respiratory disorders, inability to perform exploratory or foraging behaviour, gastroenteric disorders and group stress being the most frequent across husbandry systems. Recommendations to improve the welfare of calves include increasing space allowance, keeping calves in stable groups from an early age, ensuring good colostrum management and increasing the amounts of milk fed to dairy calves. In addition, calves should be provided with deformable lying surfaces, water via an open surface and long‐cut roughage in racks. Regarding specific recommendations for veal systems, calves should be kept in small groups (2–7 animals) within the first week of life, provided with ~ 20 m2/calf and fed on average 1 kg neutral detergent fibre (NDF) per day, preferably using long‐cut hay. Recommendations on cow–calf contact include keeping the calf with the dam for a minimum of 1 day post‐partum. Longer contact should progressively be implemented, but research is needed to guide this implementation in practice. The ABMs body condition, carcass condemnations, abomasal lesions, lung lesions, carcass colour and bursa swelling may be collected in slaughterhouses to monitor on‐farm welfare but should be complemented with behavioural ABMs collected on farm.

Abstract Trained immunity is the capacity of innate immune cells to produce an improved response against a secondary infection after a previous unrelated infection. Salmonellosis represents a public health issue and affects the pig farming industry. In general, vaccination against salmonellosis is still facing problems regarding the control of distinct serovars. Therefore, we hypothesized that an immunostimulant based on heat inactivated Mycobacterium bovis (HIMB) could have an immune training effect in pigs challenged with Salmonella enterica serovar Choleraesuis (S. Choleraesuis) and decided to explore the amplitude of this non-specific immune response. For this purpose, twenty-four 10 days-old female piglets were randomly separated in three groups: immunized group (n = 10) received orally two doses of HIMB prior to the intratracheal S. Choleraesuis-challenge, positive control group (n = 9) that was only challenged with S. Choleraesuis, and negative control group (n = 5) that was neither immunized nor infected. All individuals were necropsied 21 days post-challenge. HIMB improved weight gain and reduced respiratory symptoms and pulmonary lesions caused by S. Choleraesuis in pigs. Pigs immunized with HIMB showed higher cytokine production, especially of serum TNFα and lung CCL28, an important mediator of mucosal trained immunity. Moreover, immunized pigs showed lower levels of the biomarker of lipid oxidation malondialdehyde and higher activity of the antioxidant enzyme superoxide dismutase than untreated challenged pigs. However, the excretion and tissue colonization of S. Choleraesuis remained unaffected. This proof-of-concept study suggests beneficial clinical, pathological, and heterologous immunological effects against bacterial pathogens within the concept of trained immunity, opening avenues for further research.

BackgroundExcessive and irrational use of antibiotics as growth promoters in poultry has been one of key factors contributing to increased emergence of antibiotics resistant bacteria. Several alternatives for antibiotic growth promoters are being sought, and the search for effective probiotics to be used as feed additives is amongst the promising ones. Our study aimed to isolate and test potential probiotics bacteria from cloacal swabs of various indigenous chicken (Gallus domesticus) breeds from rural outskirts of the Kathmandu valley (Nepal).MethodsSelective isolation of probiotics was conducted by micro-aerophilic enrichment of sample in MRS Broth at 37°C, followed by culturing on MRS agar supplemented with 5 g/L of CaCO3. Isolated bacterial colonies producing transparent halo were selected as potential lactic acid bacteria (LAB), and tested for their antibacterial activity, phenotypic and biochemical characteristics, acidic yield, and tolerance to acid and bile.ResultsA total of 90 potential LAB were isolated from cloacal samples collected from 41 free-ranging chickens of indigenous breeds. Of these, 52 LAB isolates (57%) showed variable antibacterial activity to at least one bacterial pathogen. Of 52 LAB, 46 isolates fulfilled phenotypic and biochemical criteria of Lactobacillus spp. Of these, 37 isolates produced varying percentage yields of lactic acid, 27 isolates showed survival at pH 3.0, and 17 isolates showed survival tolerances in the presence of 0.3% and 0.5% bile salts for 24 hours. Phylogenetic analysis of 16S rDNA sequencing of LAB isolates fulfilling in vitro probiotics properties showed that 3 isolates had genetic identity of 99.38% with Lactobacillus plantarum, while one isolate was genetically similar (99.85%) with the clade of L. reuteri, L. antri and L. panis.ConclusionOur study identified four Lactobacillus spp. strains having potential probiotics properties. Further investigations are needed to evaluate these isolates to be used as poultry probiotics feed supplement.

Bagaza virus emerged in Spain in 2010 and was not reported in other countries in Europe until 2021, when the virus was detected by molecular methods in a corn bunting and several red-legged partridges in Portugal. Sequencing revealed high similarity between the 2021 strains from Portugal and the 2010 strains from Spain.

Abstract In this Opinion, the antimicrobial‐resistant bacteria responsible for transmissible diseases that constitute a threat to the health of certain kept fish species have been assessed. Atlantic salmon (Salmo salar), carp (Cyprinus spp.), rainbow trout (Oncorhynchus mykiss), sea bream (Sparus aurata) and tilapia (Oreochromis spp.), selected as representative of the most important fish species and production systems that are commercially reared in fresh and saltwater farms, were the focus of this assessment. The assessment was performed following a methodology based on information collected by an extensive literature review and expert judgement. Details of the methodology used for this assessment are explained in a separate Opinion. The global state of play of antimicrobial resistance in Aeromonas hydrophila, Aeromonas salmonicida, Flavobacterium psychrophilum and Flavobacterium columnare is provided. Among these bacteria, none was identified as being among the most relevant antimicrobial‐resistant bacteria in the assessed kept fish species in the EU due to the very limited scientific evidence available.

Abstract In this opinion, the antimicrobial‐resistant bacteria responsible for transmissible diseases that constitute a threat to the health of pigs have been assessed. The assessment has been performed following a methodology based on information collected by an extensive literature review and expert judgement. Details of the methodology used for this assessment are explained in a separate opinion. A global state of play of antimicrobial resistant Escherichia coli, Streptococcus suis, Actinobacillus pleuropneumoniae, Pasteurella multocida, Glaeserella parasuis, Bordetella bronchiseptica, Staphylococcus aureus, Staphylococcus hyicus, Brachyspira hyodysenteriae, Trueperella pyogenes, Erysipelothrix rhusiopathiae, Streptococcus dysgalactiae, Mycoplasma hyosynoviae, Mycoplasma hyorhinis, Mycoplasma hyopneumoniae and Brachyspira pilosicoli has been provided. Among those bacteria, EFSA identified E. coli and B. hyodysenteriae with > 66% certainty as being the most relevant antimicrobial resistant bacteria in the EU based on the available evidence. The animal health impact of these most relevant bacteria, as well as their eligibility for being listed and categorised within the animal health law framework will be assessed in separate scientific opinions.

Abstract In this opinion, the antimicrobial‐resistant bacteria responsible for transmissible diseases that constitute a threat to poultry health have been assessed. The assessment has been performed following a methodology based on information collected by an extensive literature review and expert judgement. Details of the methodology used for this assessment are explained in a separate opinion. A global state of play is provided for: Avibacterium (Haemophilus) paragallinarum, Bordetella avium, Clostridium perfringens, Enterococcus faecalis and Enterococcus cecorum, Erysipelothrix rhusiopathiae, Escherichia coli, Gallibacterium spp., Mycoplasma synoviae, Ornithobacterium rhinotracheale, Pasteurella multocida, Riemerella anatipestifer and Staphylococcus aureus. Among those bacteria, EFSA identified Escherichia coli, Enterococcus faecalis and Enterococcus cecorum with ≥ 66% certainty as being the most relevant antimicrobial resistant bacteria in the EU based on the available evidence. The animal health impact of these most relevant bacteria, and their eligibility for being listed and categorised within the Animal Health Law Framework, will be assessed in separate scientific opinions.

Abstract In this opinion, the antimicrobial‐resistant bacteria responsible for transmissible diseases that constitute a threat to the health of farmed rabbits have been assessed. The assessment has been performed following a methodology based on information collected through an extensive literature review and expert judgement. Details of the methodology used for this assessment are explained in a separate opinion. A global state of play on antimicrobial resistance in clinical isolates of Pasteurella multocida, Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Bordetella bronchiseptica, Clostridium difficile, Clostridium perfringens and Clostridium spiroforme is provided. Among these bacteria, none were identified as being the most relevant antimicrobial‐resistant bacteria in rabbits in the EU due to the very limited scientific evidence available.

Abstract In this opinion, the antimicrobial resistant bacteria responsible for transmissible diseases that constitute a threat to the health of cattle have been assessed. The assessment has been performed following a methodology based on information collected by an extensive literature review and expert judgement. Details of the methodology used for this assessment are explained in a separate opinion. A global state of play on antimicrobial resistance in clinical isolates of Escherichia coli (non‐VTEC), Klebsiella pneumoniae, Staphylococcus aureus, Streptococcus uberis, Streptococcus dysgalactiae, Pasteurella multocida, Mannheimia haemolytica, Histophilus somni, Mycoplasma bovis, Moraxella bovis, Fusobacterium necrophorum and Trueperella pyogenes is provided. Among those bacteria, EFSA identified E. coli and S. aureus with ≥ 66% certainty as being the most relevant antimicrobial resistant bacteria in cattle in the EU based on the available evidence. The animal health impact of these most relevant bacteria, as well as their eligibility for being listed and categorised within the animal health law framework will be assessed in separate scientific opinions.

Abstract In this opinion, the antimicrobial‐resistant bacteria responsible for transmissible diseases that constitute a threat to the health of horses have been assessed. The assessment has been performed following a methodology composed of information collected via an extensive literature review and expert judgement. Details on the methodology used for this assessment are explained in a separate opinion. A global state of play of antimicrobial‐resistant Actinobacillus equuli, Dermatophilus congolensis, Enterococcus spp., Escherichia coli, Klebsiella pneumoniae, Pasteurella spp., Pseudomonas aeruginosa, Rhodococcus equi, Staphylococcus aureus and Streptococcus dysgalactiae subsp. dysgalactiae/equisimilis and Streptococcus equi subsp. equi and subsp. zooepidemicus has been provided. Among those bacteria, EFSA identified E. coli, Staphylococcus aureus and R. equi with more than 66% certainty as the most relevant antimicrobial‐resistant bacteria in the EU, given their importance as causative agents of clinical disease in horses and the significant levels of resistance to clinically relevant antimicrobials. The animal health impact of these ‘most relevant’ bacteria as well as their eligibility of being listed and categorised within the animal health law framework will be assessed in separate scientific opinions.

A collaboration program was established between the group of Health and Biotechnology (SaBio) of the IREC Institute of Game and Wildlife Research (CSIC-UCLM-JCCM, Spain) and the National Agricultural Research Organization of Uganda (NARO) for the development of vaccines for the control of cattle ticks in Uganda. Controlled pen trials identified a tick protective antigen, Rhipicephalus appendiculatus Subolesin, and a cross-species-effective vaccine formulation. As the next step, a controlled vaccine field trial has been approved by Ugandan state regulatory authorities, the Uganda National Council for Science and Technology (UNCST) and the National Drug Authority (NDA), to evaluate the efficacy and effectiveness of the vaccine formulation for the control of cattle tick infestations under field conditions. The results of this trial may lead to the approval of the vaccine for application in Uganda to improve cattle health and production while reducing the use of acaricides.

Abstract The European Commission requested EFSA assess antimicrobial‐resistant bacteria responsible for animal transmissible diseases, with a view to listing such pathogens for European Union action. This Scientific Opinion addresses the ad hoc method developed: (i) to give a global state of play as regards resistant animal pathogens that cause transmissible animal diseases, (ii) to identify the most relevant bacteria in the EU and (iii) to summarise their actual or potential animal health impact, and to perform their assessment for being listed and categorised according to the criteria of Articles 7, 5, 9 and 8 within the Animal Health Law (AHL) framework. An extensive literature review is carried out to give the global state of play of selected resistant bacteria that constitute a threat to animal health (i). An expert judgement procedure, based on the outcome of the literature review, is applied to identify which among those bacteria subjected to the literature review are the ‘most relevant’ in the European Union (ii). Their animal health impact in the European Union and their assessment for being listed and categorised according to the AHL framework will follow the ‘ad hoc method for the assessment on listing and categorisation of animal diseases within the framework of the Animal Health Law’ that EFSA has developed in the past (iii). The assessment of (i) and (ii) is addressed in distinct scientific opinions that are published separately by animal species of interest (dogs and cats, horses, pigs, poultry, cattle, small ruminants, rabbits and aquatic animal species). The assessment of (iii) is addressed in distinct scientific opinions and published separately by the animal pathogen.

Abstract In this opinion the antimicrobial‐resistant bacteria responsible for transmissible diseases that constitute a threat to dog and cat health have been assessed. The assessment has been performed following a methodology based on information collected via an extensive literature review and expert judgement. Details of the methodology used for this assessment are explained in a separate opinion. A global state of play of antimicrobial resistant Staphylococcus pseudintermedius, Staphylococcus aureus, Staphylococcus schleiferi, Escherichia coli, Proteus mirabilis, Klebsiella spp., Enterobacter spp., Pseudomonas aeruginosa, Clostridium perfringens, Clostridioides difficile, Enterococcus faecalis and Enterococcus faecium has been provided. Among those bacteria, EFSA identified S. pseudintermedius, E. coli and P. aeruginosa with > 90% certainty as the most relevant antimicrobial resistant bacteria in the EU based on the available evidence. The animal health impact of these most relevant bacteria, as well as their eligibility for being listed and categorised within the animal health law framework will be assessed in separate scientific opinions.

Wildlife management and conservation requires monitoring of species distribution and population indicators, especially when the unbalanced demographic changes of some species can affect the whole ecosystem functioning. The populations of wild boar (Sus scrofa) have, over the past few decades, undergone an expansion around the world, reaching situations of overabundance that can cause serious economic, ecological and health problems. This numerical increase of wild boar and its new spatial invasion can affect certain vulnerable species in sensitive ecological zones, such as the main inland wetland complex in SW Europe.In this context, we aim to (i) examine the association of wild boar abundance and that of lagomorphs, and waterbird productivity, controlling the possible effects of wild carnivores and other environmental predictors, and (ii) applying a survey method useful for managers to evaluate expected effect of wild boar abundance.Overall, the presence of wild boar has been detected in 80.8% of the monitored wetlands (n = 26), but a high variation of abundance rates was found. Wild boar abundance negatively associated with the productivity of the entire community of waterbirds, a priori highly vulnerable, breeding on shores or islands, both colonially (genus Gelochelidon, Himantopus, Recurvirostra, Sternula, Glareola, Tringa and Phoenicopterus) or solitary (genus Anas, Spatula, Mareca, Charadrius, Vanellus, Anser, Ardea, Aythya, Fulica, Netta, Oxyura and Tadorna).In addition, we evidenced a clear negative association of wild boar and wild rabbit (Oryctolagus cuniculus) population abundance in the monitored wetlands. Rabbits are a keystone species in the study area and represent stable prey for many endangered predators.Our results suggest that potential impacts caused by high wild boar abundance may be already reaching unsustainable levels for some wetlands. Inter-species integrated monitoring is key to address the population management plans of wild boar populations in wetlands and to preserve the most vulnerable species.

Abstract EFSA assessed the role of seropositive wild boar in African swine fever (ASF) persistence. Surveillance data from Estonia and Latvia investigated with a generalised equation method demonstrated a significantly slower decline in seroprevalence in adult animals compared with subadults. The seroprevalence in adults, taking more than 24 months to approach zero after the last detection of ASFV circulation, would be a poor indicator to demonstrate the absence of virus circulation. A narrative literature review updated the knowledge on the mortality rate, the duration of protective immunity and maternal antibodies and transmission parameters. In addition, parameters potentially leading to prolonged virus circulation (persistence) in wild boar populations were reviewed. A stochastic explicit model was used to evaluate the dynamics of virus prevalence, seroprevalence and the number of carcasses attributed to ASF. Secondly, the impact of four scenarios on the duration of ASF virus (ASFV) persistence was evaluated with the model, namely a: (1) prolonged, lifelong infectious period, (2) reduction in the case‐fatality rate and prolonged transient infectiousness; (3) change in duration of protective immunity and (4) change in the duration of protection from maternal antibodies. Only the lifelong infectious period scenario had an important prolonging effect on the persistence of ASF. Finally, the model tested the performance of different proposed surveillance strategies to provide evidence of the absence of virus circulation (Exit Strategy). A two‐phase approach (Screening Phase, Confirmation Phase) was suggested for the Exit Strategy. The accuracy of the Exit Strategy increases with increasing numbers of carcasses collected and tested. The inclusion of active surveillance based on hunting has limited impact on the performance of the Exit Strategy compared with lengthening of the monitoring period. This performance improvement should be reasonably balanced against an unnecessary prolonged ‘time free’ with only a marginal gain in performance. Recommendations are provided for minimum monitoring periods leading to minimal failure rates of the Exit Strategy. The proposed Exit Strategy would fail with the presence of lifelong infectious wild boar. That said, it should be emphasised that the existence of such animals is speculative, based on current knowledge.

Understanding the dynamics of a wildlife population in relation to hunting strategies is essential to achieve sustainable management. We used monitoring data over 25 years from two red deer (Cervus elaphus) populations with different management (with and without supplemental feeding) in South Central Spain to: (i) characterise the density dependence of population dynamics under contrasted management, and (ii) provide the basis for sustainable extraction by considering the theoretical maximum sustainable yield (MSYt) as the reference. The red deer population displayed a typical management reactive culling approach (‘saw-tooth-like’ curves), with occasional strong annual harvests but not occurring on a regular basis. Interestingly, we found reduced population growth at high densities in both populations, indicating that density-mediated factors determined population growth even when artificial feeding was provided. However, no effects of sex not age class of the extracted population on the population growth rate were determined. The total number of animals hunted was only slightly above those predicted by MSYt (i.e. K50%) in both populations, despite high densities close to theoretical K, being consistent throughout the study period. The extraction rates (30.3 and 34.0%, for supplemented and unsupplemented populations, respectively) were 13.3% and 10.2% lower compared to the MSYt situation in the unsupplemented and supplemented populations, respectively. Long term population monitoring data provided feasible and suitable baseline values to optimise the sustainable exploitation of red deer populations in the Mediterranean ecosystem under these contrasting management scenarios. Adaptive management, involving objective-driven decision making informed by data on red deer population dynamic, can contribute (i) to maximising the total extraction over the long term while (ii) reducing the ecological impact of high population densities.

Abstract Animal tuberculosis (TB) remains a major problem in some countries despite the existence of control programmes focused mainly on cattle. In this species, aerogenous transmission is accepted as the most frequent infection route, affecting mainly the respiratory system. Under the hypothesis that the oral route could be playing a more relevant role in transmission, diagnosis and disease persistence than previously thought, this study was performed to assess the course of TB infection in cattle and its effects on diagnosis depending on the route of entry of Mycobacterium bovis. Two groups of five calves each were either endotracheally (EC) or orally (OC) challenged. Necropsies were carried out 12 weeks after challenge except for three OC calves slaughtered 8 weeks later. All animals reacted to the tuberculin skin test and the entire EC group was positive to the interferon-gamma release assay (IGRA) 2 weeks after challenge and thereafter. The first positive IGRA results for OC calves (3/5) were recorded 4 weeks after challenge. Group comparison revealed significant differences in lesion and positive culture location and scoring. TB-compatible gross lesions and positive cultures were more frequently found in the thorax (p

Vaccines are an environmentally friendly alternative to acaracides for the control of tick infestations, to reduce the risk for tick-borne diseases affecting human and animal health worldwide, and to improve animal welfare and production. Subolesin (SUB, also known as 4D8) is the functional homolog of Akirin2 involved in the regulation of development and innate immune response, and a proven protective antigen for the control of ectoparasite infestations and pathogen infection. Oral vaccination combining protein antigens with immunostimulants has proven efficacy with increased host welfare and safety, but has not been used for the control of tick infestations. Here we describe the efficacy of oral vaccination with a formulation combining Rhipicephalus microplus SUB and heat inactivated Mycobacterium bovis (IV) on cattle tick infestations and fertility. The levels of IgG antibody titers against SUB and M. bovis P22, and the expression of selected immune response genes were determined and analyzed as possible correlates of protection. We demonstrated that oral immunization with the SUB+IV formulation resulted in 51% reduction in the number of female ticks and 30% reduction in fertility with an overall efficacy of 65% in the control of R. microplus infestations by considering the cumulative effect on reducing tick survival and fertility in cattle. The akr2, IL-1β, and C3 mRNA levels together with antibody levels against SUB correlated with vaccine efficacy. The effect of the oral immunization with SUB+IV in cattle on tick survival and fertility is essential to reduce tick infestations, and extended previous results on the effect of R. microplus SUB for the control of cattle tick infestations. These results support the development of oral vaccines formulations for the control of tick infestations and the incidence of tick-borne diseases.

We investigated adult Eurasian wild boar (Sus scrofa) survival and death in 2 tuberculosis-endemic populations with different harvest pressure in Spain. Overall, tuberculosis accounted for 30% of total deaths. Increased survival in protected areas has direct implications for wild boar management and tuberculosis control.

Abstract Council Regulation (EC) No 1099/2009 on the protection of animals at the time of killing lists in Annex I the stunning interventions currently allowed in the EU, together with the related conditions under which those interventions can be implemented. The regulation allows the Commission to amend Annex I, listing additional stunning interventions, provided they ensure a level of animal welare at least equivalent to that ensured by the one already approved. EFSA was requested to perform such assessment with regard to the implementation of the low atmospheric pressure stunning (LAPS) system on broiler chickens. The ad hoc Working Group (WG) set up by EFSA performed the assessment in three main steps, i.e. checking the data provided against the criteria laid down in the EFSA Guidance (EFSA AHAW Panel, 2013); running an extensive literature search, followed by data extraction and performing a judgemental ranking exercise based on expert opinion. As main outcome, the LAPS intervention was found to be able to provide a level of animal welfare not lower than that provided by at least one of the currently allowed methods. The overall assessment of EFSA is valid ONLY under the technical conditions described in the submission and for broiler chickens, intended for human consumption, weighting less than 4 kg. Deviations from these conditions might have different consequences for animal welfare which were not assessed in this exercise. The LAPS method may, in addition to commercial slaughter, be suitable for depopulation, respecting the technical conditions defined in the present conclusions. The WG considers that a revision of the present version of the EFSA Guidance could be beneficial.

Anaplasma phagocytophilum transmembrane and surface proteins play a role during infection and multiplication in host neutrophils and tick vector cells. Recently, A. phagocytophilum Major surface protein 4 (MSP4) and Heat shock protein 70 (HSP70) were shown to be localized on the bacterial membrane, with a possible role during pathogen infection in ticks. In this study, we hypothesized that A. phagocytophilum MSP4 and HSP70 have similar functions in tick-pathogen and host-pathogen interactions. To address this hypothesis, herein we characterized the role of these bacterial proteins in interaction and infection of vertebrate host cells. The results showed that A. phagocytophilum MSP4 and HSP70 are involved in host-pathogen interactions, with a role for HSP70 during pathogen infection. The analysis of the potential protective capacity of MSP4 and MSP4-HSP70 antigens in immunized sheep showed that MSP4-HSP70 was only partially protective against pathogen infection. This limited protection may be associated with several factors, including the recognition of non-protective epitopes by IgG in immunized lambs. Nevertheless, these antigens may be combined with other candidate protective antigens for the development of vaccines for the control of human and animal granulocytic anaplasmosis. Focusing on the characterization of host protective immune mechanisms and protein-protein interactions at the host-pathogen interface may lead to the discovery and design of new effective protective antigens.

Abstract After a request from the European Commission, EFSA's Panel on Animal Health and Welfare summarised the main characteristics of 36 vector‐borne diseases (VBDs) in 36 Disease Profiles. The risk of introduction in the EU through movement of livestock or pets was assessed for each of the 36 VBDs individually, using a semiquantitative Method to INTegrate all relevant RISK aspects (MINTRISK model), which was further modified to a European scale into the EFSA-VBD-RISK-model. Only eight of the 36 VBD‐agents had an overall rate of introduction in the EU (being the combination of the rate of entry, vector transmission and establishment) which was estimated to be above 0.001 introductions per year. These were Crimean‐Congo haemorrhagic fever virus, bluetongue virus, West Nile virus, Schmallenberg virus, Hepatozoon canis, Leishmania infantum, Bunyamwera virus and Highlands J. virus. For these eight diseases, the annual extent of spread was assessed, assuming the implementation of available, authorised prevention and control measures in the EU. Further, the probability of overwintering was assessed, as well as the possible impact of the VBDs on public health, animal health and farm production. For the other 28 VBD‐agents for which the rate of introduction was estimated to be very low, no further assessments were made. Due to the uncertainty related to some parameters used for the risk assessment or the instable or unpredictability disease situation in some of the source regions, it is recommended to update the assessment when new information becomes available. Since this risk assessment was carried out for large regions in the EU for many VBD‐agents, it should be considered as a first screening. If a more detailed risk assessment for a specific VBD is wished for on a national or subnational level, the EFSA‐VBD‐RISK‐model is freely available for this purpose.

Abstract This scientific opinion addresses animal welfare aspects of slaughtering of livestock pregnant animals. Term of Reference (ToR) 1 requested assessment of the prevalence of animals slaughtered in a critical developmental stage of gestation when the livestock fetuses might experience negative affect. Limited data on European prevalence and related uncertainties necessitated a structured expert knowledge elicitation (EKE) exercise. Estimated median percentages of animals slaughtered in the last third of gestation are 3%, 1.5%, 0.5%, 0.8% and 0.2% (dairy cows, beef cattle, pigs, sheep and goats, respectively). Pregnant animals may be sent for slaughter for health, welfare, management and economic reasons (ToR2); there are also reasons for farmers not knowing that animals sent for slaughter are pregnant. Measures to reduce the incidence are listed. ToR3 asked whether livestock fetuses can experience pain and other negative affect. The available literature was reviewed and, at a second multidisciplinary EKE meeting, judgements and uncertainty were elicited. It is concluded that livestock fetuses in the last third of gestation have the anatomical and neurophysiological structures required to experience negative affect (with 90–100% likelihood). However, there are two different possibilities whether they perceive negative affect. It is more probable that the neurophysiological situation does not allow for conscious perception (with 66–99% likelihood) because of brain inhibitory mechanisms. There is also a less probable situation that livestock fetuses can experience negative affect (with 1–33% likelihood) arising from differences in the interpretation of the fetal electroencephalogram, observed responses to external stimuli and the possibility of fetal learning. Regarding methods to stun and kill livestock fetuses at slaughter (ToR4), sets of scenarios and respective actions take account of both the probable and less probable situation regarding fetal ability for conscious perception. Finally, information was collated on methods to establish the dam's gestational stage based on physical features of livestock fetuses (ToR5).

Abstract Background Piroplasmosis are among the most relevant diseases of domestic animals. Babesia is emerging as cause of tick-borne zoonosis worldwide and free-living animals are reservoir hosts of several zoonotic Babesia species. We investigated the epidemiology of Babesia spp. and Theileria spp. in wild ungulates and carnivores from Northern Italy to determine which of these apicomplexan species circulate in wildlife and their prevalence of infection. Methods PCR amplification of the V4 hyper-variable region of the 18S rDNA of Babesia sp./Theileria sp was carried out on spleen samples of 1036 wild animals: Roe deer Capreolus capreolus (n = 462), Red deer Cervus elaphus (n = 52), Alpine Chamois Rupicapra rupicapra (n = 36), Fallow deer Dama dama (n = 17), Wild boar Sus scrofa (n = 257), Red fox Vulpes vulpes (n = 205) and Wolf Canis lupus (n = 7). Selected positive samples were sequenced to determine the species of amplified Babesia/Theileria DNA. Results Babesia/Theileria DNA was found with a mean prevalence of 9.94% (IC95% 8.27-11.91). The only piroplasms found in carnivores was Theileria annae, which was detected in two foxes (0.98%; IC95% 0.27-3.49). Red deer showed the highest prevalence of infection (44.23%; IC95% 31.6-57.66), followed by Alpine chamois (22.22%; IC95% 11.71-38.08), Roe deer (12.55%; IC95% 9.84-15.89), and Wild boar (4.67%; IC95% 2.69-7.98). Genetic analysis identified Babesia capreoli as the most prevalent piroplasmid found in Alpine chamois, Roe deer and Red deer, followed by Babesia bigemina (found in Roe deer, Red deer and Wild boar), and the zoonotic Babesia venatorum (formerly Babesia sp. EU1) isolated from 2 Roe deer. Piroplasmids of the genus Theileria were identified in Wild boar and Red deer. Conclusions The present study offers novel insights into the role of wildlife in Babesia/Theileria epidemiology, as well as relevant information on genetic variability of piroplasmids infecting wild ungulates and carnivores.

A meeting and course supported by the Vice-Presidency for International Affairs of the Spanish National Research Council (CSIC) and the National Agricultural Research Organization of Uganda (NARO) were held at the National Livestock Resources Research Institute (NaLIRRI) in Nakyesasa, Wakiso, Uganda on September 2−9, 2019. The activities were conducted within the collaboration program between the Institute of Game and Wildlife Research (IREC, CSIC-UCLM-JCCM, Spain) and NARO for the development of vaccines and other interventions for the control of cattle ticks in Uganda.

Innate immunity is evolutionarily conserved in multicellular organisms and was considered to lack memory until very recently. One of its more characteristic mechanisms is phagocytosis, the ability of cells to engulf, process and eventually destroy any injuring agent. We report the results of an ex vivo experiment in bovine macrophages in which improved clearance of Mycobacterium bovis (M. bovis) was induced by pre-exposure to a heat killed M. bovis preparation. The effects were independent of humoral and cellular adaptive immune responses and lasted up to six months. Specifically, our results demonstrate the existence of a training effect in the lytic phase of phagocytosis that can be activated by killed mycobacteria, thus suggesting a new mechanism of vaccine protection. These findings are compatible with the recently proposed concept of trained immunity, which was developed to explain the observation that innate immune responses provide unspecific protection against pathogens including other than those that originally triggered the immune response.

Mycobacteria of the Mycobacterium tuberculosis complex (MTBC) greatly affect humans and animals worldwide. The life cycle of mycobacteria is complex and the mechanisms resulting in pathogen infection and survival in host cells are not fully understood. Recently, comparative genomics analyses have provided new insights into the evolution and adaptation of the MTBC to survive inside the host. However, most of this information has been obtained using M. tuberculosis but not other members of the MTBC such as M. bovis and M. caprae. In this study, the genome of three M. bovis (MB1, MB3, MB4) and one M. caprae (MB2) field isolates with different lesion score, prevalence and host distribution phenotypes were sequenced. Genome sequence information was used for whole-genome and protein-targeted comparative genomics analysis with the aim of finding correlates with phenotypic variation with potential implications for tuberculosis (TB) disease risk assessment and control. At the whole-genome level the results of the first comparative genomics study of field isolates of M. bovis including M. caprae showed that as previously reported for M. tuberculosis, sequential chromosomal nucleotide substitutions were the main driver of the M. bovis genome evolution. The phylogenetic analysis provided a strong support for the M. bovis/M. caprae clade, but supported M. caprae as a separate species. The comparison of the MB1 and MB4 isolates revealed differences in genome sequence, including gene families that are important for bacterial infection and transmission, thus highlighting differences with functional implications between isolates otherwise classified with the same spoligotype. Strategic protein-targeted analysis using the ESX or type VII secretion system, proteins linking stress response with lipid metabolism, host T cell epitopes of mycobacteria, antigens and peptidoglycan assembly protein identified new genetic markers and candidate vaccine antigens that warrant further study to develop tools to evaluate risks for TB disease caused by M. bovis/M.caprae and for TB control in humans and animals.

Tuberculosis (TB) remains a pandemic affecting billions of people worldwide, thus stressing the need for new vaccines. Defining the correlates of vaccine protection is essential to achieve this goal. In this study, we used the wild boar model for mycobacterial infection and TB to characterize the protective mechanisms elicited by a new heat inactivated Mycobacterium bovis vaccine (IV). Oral vaccination with the IV resulted in significantly lower culture and lesion scores, particularly in the thorax, suggesting that the IV might provide a novel vaccine for TB control with special impact on the prevention of pulmonary disease, which is one of the limitations of current vaccines. Oral vaccination with the IV induced an adaptive antibody response and activation of the innate immune response including the complement component C3 and inflammasome. Mycobacterial DNA/RNA was not involved in inflammasome activation but increased C3 production by a still unknown mechanism. The results also suggested a protective mechanism mediated by the activation of IFN-γ producing CD8+ T cells by MHC I antigen presenting dendritic cells (DCs) in response to vaccination with the IV, without a clear role for Th1 CD4+ T cells. These results support a role for DCs in triggering the immune response to the IV through a mechanism similar to the phagocyte response to PAMPs with a central role for C3 in protection against mycobacterial infection. Higher C3 levels may allow increased opsonophagocytosis and effective bacterial clearance, while interfering with CR3-mediated opsonic and nonopsonic phagocytosis of mycobacteria, a process that could be enhanced by specific antibodies against mycobacterial proteins induced by vaccination with the IV. These results suggest that the IV acts through novel mechanisms to protect against TB in wild boar.

Because of their relative simplicity and the barriers to gene flow, islands are ideal systems to study the distribution of biodiversity. However, the knowledge that can be extracted from this peculiar ecosystem regarding epidemiology of economically relevant diseases has not been widely addressed. We used information available in the scientific literature for 10 old world islands or archipelagos and original data on Sicily to gain new insights into the epidemiology of the Mycobacterium tuberculosis complex (MTC). We explored three nonexclusive working hypotheses on the processes modulating bovine tuberculosis (bTB) herd prevalence in cattle and MTC strain diversity: insularity, hosts and trade. Results suggest that bTB herd prevalence was positively correlated with island size, the presence of wild hosts, and the number of imported cattle, but neither with isolation nor with cattle density. MTC strain diversity was positively related with cattle bTB prevalence, presence of wild hosts and the number of imported cattle, but not with island size, isolation, and cattle density. The three most common spoligotype patterns coincided between Sicily and mainland Italy. However in Sicily, these common patterns showed a clearer dominance than on the Italian mainland, and seven of 19 patterns (37%) found in Sicily had not been reported from continental Italy. Strain patterns were not spatially clustered in Sicily. We were able to infer several aspects of MTC epidemiology and control in islands and thus in fragmented host and pathogen populations. Our results point out the relevance of the intensity of the cattle commercial networks in the epidemiology of MTC, and suggest that eradication will prove more difficult with increasing size of the island and its environmental complexity, mainly in terms of the diversity of suitable domestic and wild MTC hosts.

Vector or reservoir species of five mollusc diseases listed in the Animal Health Law were identified, based on evidence generated through an extensive literature review, to support a possible updating of Regulation (EU) 2018/1882. Mollusc species on or in which Mikrocytos mackini, Perkinsus marinus, Bonamia exitiosa, Bonamia ostreae and Marteilia refringens were detected, in the field or during experiments, were classified as reservoir species with different levels of certainty depending on the diagnostic tests used. Where experimental evidence indicated transmission of the pathogen from a studied species to another known susceptible species, this studied species was classified as a vector species. Although the quantification of the risk of spread of the pathogens by the vectors or reservoir species was not part of the terms of reference, such risks do exist for the vector species, since transmission from infected vector species to susceptible species was proven. Where evidence for transmission from infected molluscs was not found, these were defined as reservoir. Nonetheless, the risk of the spread of the pathogens from infected reservoir species cannot be excluded. Evidence identifying conditions that may prevent transmission by vectors or reservoir mollusc species during transport was collected from scientific literature. It was concluded that it is very likely to almost certain (90–100%) that M. mackini, P. marinus, B. exitiosaB. ostreae and M. refringens will remain infective at any possible transport condition. Therefore, vector or reservoir species that may have been exposed to these pathogens in an affected area in the wild or at aquaculture establishments or through contaminated water supply can possibly transmit these pathogens. For transmission of M. refringens, the presence of an intermediate host, a copepod, is necessary. This publication is linked to the following EFSA Journal articles: http://onlinelibrary.wiley.com/doi/10.2903/j.efsa.2023.8172/full, http://onlinelibrary.wiley.com/doi/10.2903/j.efsa.2023.8174/full This publication is linked to the following EFSA Supporting Publications articles: http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8122/full, http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8123/full, http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8124/full [ABSTRACT FROM AUTHOR]

Vector or reservoir species of five fish diseases listed in the Animal Health Law were identified, based on evidence generated through an extensive literature review (ELR), to support a possible updating of Regulation (EU) 2018/1882. Fish species on or in which highly polymorphic region‐deleted infectious salmon anaemia virus (HPR∆ ISAV), Koi herpes virus (KHV), epizootic haematopoietic necrosis virus (EHNV), infectious haematopoietic necrosis virus (IHNV) or viral haemorrhagic septicaemia virus (VHSV) were detected, in the field or during experiments, were classified as reservoir species with different levels of certainty depending on the diagnostic tests used. Where experimental evidence indicated transmission of the pathogen from a studied species to another known susceptible species, the studied species was classified as a vector species. Although the quantification of the risk of spread of the pathogens by the vectors or reservoir species was not part of the terms or reference, such risks do exist for the vector species, since transmission from infected vector species to susceptible species was proven. Where evidence for transmission from infected fish was not found, these were defined as reservoirs. Nonetheless, the risk of the spread of the pathogens from infected reservoir species cannot be excluded. Evidence identifying conditions that may prevent transmission by vectors or reservoir fish species during transport was collected from scientific literature. For VHSV, IHNV or HPR∆ ISAV, it was concluded that under transport conditions at temperatures below 25°C, it is likely (66–90%) they will remain infective. Therefore, vector or reservoir species that may have been exposed to these pathogens in an affected area in the wild, aquaculture establishments or through water supply can possibly transmit VHSV, IHNV or HPR∆ ISAV into a non‐affected area when transported at a temperature below 25°C. The conclusion was the same for EHN and KHV; however, they are likely to remain infective under all transport temperatures. This publication is linked to the following EFSA Journal articles: http://onlinelibrary.wiley.com/doi/10.2903/j.efsa.2023.8172/full, http://onlinelibrary.wiley.com/doi/10.2903/j.efsa.2023.8173/full This publication is linked to the following EFSA Supporting Publications articles: http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8122/full, http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8123/full, http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8124/full [ABSTRACT FROM AUTHOR]

Vector or reservoir species of three diseases of crustaceans listed in the Animal Health Law were identified based on evidence generated through an extensive literature review, to support a possible updating of Regulation (EU) 2018/1882. Crustacean species on or in which Taura syndrome virus (TSV), Yellow head virus (YHV) or White spot syndrome virus (WSSV) were identified, in the field or during experiments, were classified as reservoir species with different levels of certainty depending on the diagnostic tests used. Where experimental evidence indicated transmission of the pathogen from a studied species to another known susceptible species, the studied species was classified as vector species. Although the quantification of the risk of spread of the pathogens by the vectors or reservoir species was not part of the terms of reference, such risks do exist for the vector species, since transmission from infected vector species to susceptible species was proven. Where evidence for transmission from infected crustaceans was not found, these were defined as reservoirs. Nonetheless, the risk of the spread of the pathogens from infected reservoir species cannot be excluded. Evidence identifying conditions that may prevent transmission by vectors during transport was collected from scientific literature. It was concluded that it is very likely to almost certain (90–100%) that WSSV, TSV and YHV will remain infective at any possible transport condition. Therefore, vector or reservoir species that may have been exposed to these pathogens in an affected area in the wild or aquaculture establishments or by water supply can possibly transmit WSSV, TSV and YHV. This publication is linked to the following EFSA Journal articles: http://onlinelibrary.wiley.com/doi/10.2903/j.efsa.2023.8173/full, http://onlinelibrary.wiley.com/doi/10.2903/j.efsa.2023.8174/full This publication is linked to the following EFSA Supporting Publications articles: http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8122/full, http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8123/full, http://onlinelibrary.wiley.com/doi/10.2903/sp.efsa.2023.EN-8124/full [ABSTRACT FROM AUTHOR]

Animal tuberculosis (TB) is often maintained by multi-host communities, including livestock and wildlife. Quantitative studies of such communities require estimating the true prevalence of TB, correcting the apparent prevalence by the diagnostic sensitivity (Se) and specificity (Sp) of the test. The goal of this study was to lay the foundations for estimating the true prevalence of TB in wild ungulate populations (wild boar and two cervids: red deer and fallow deer). We used Bayesian latent class models to assess the Se and Sp of gross pathology, IS6110 real-time PCR in tissues, bacteriological culture, and P22 indirect ELISA. We analyzed 308 harvested wild ungulates (211 wild boar and 97 cervids: 92 red deer and 5 fallow deer). The Se of bacteriological culture (80.4%, CI95 61.0–96.3%) and gross pathology (87.9%, CI95 69.5–99.9%) was reasonably good in wild boar. These tests showed lower Se in cervids: 60.2% (CI95 38.3–82.3%) for bacteriological culture and 81.5% (CI95 63.6–96.2%) for gross pathology. The Se of the real-time PCR was low (50.7% in wild boar and 53.0% in cervids). These tests showed Sp between 95.2 and 99.1% in both taxa. The P22 ELISA performed reasonably well in wild boar (Se = 71.9%, CI95 59.2–83.4%; Sp = 98.8%, CI95 96.9–99.9%) but lacked Sp in cervids (Se = 77.1%, CI95 62.9–89.7%; Sp = 74.5%, CI95 65.7–83.3%). The real-time PCR in wild boar and cervids and bacteriological culture in cervids tended to show higher Se in low-prevalence populations, possibly due to a higher proportion of early-stage TB lesions. In cervids, the parallel interpretation of gross pathology and bacteriological culture significantly improved the diagnostic performance (Se = 93.1%, CI95 84.7–98.9%; Sp = 92.9%, CI95 86.0–98.3%). Our results allow the estimation of true prevalence from the results of a single diagnostic test applied to harvested wild boar, red deer, and fallow deer, paving the way for more precise quantitative ecological studies of the multi-host TB maintenance community.