A Multi-Laboratory Comparison of Methods for Detection and Quantification of African Swine Fever Virus.

African swine fever is a viral disease of the family Suidae. Methods to detect and quantify African swine fever virus (ASFV) include qPCR and virus infectivity assays. Individual laboratories often use in-house procedures for these assays, which can hamper the comparison of results. The objective of this study was to estimate the probability of ASFV detection using these assays, and to determine the inter-test correlations between results. This was achieved by testing a panel of 80 samples at three reference laboratories. Samples were analysed using nucleic acid extraction and qPCR, as well as virus infectivity assays. For qPCR, a very high probability (ranging from 0.96 to 1.0) of detecting ASFV DNA was observed for all tested systems. For virus infectivity assays in cells, the probability of detecting infectious ASFV varied from 0.68 to 0.90 and was highest using pulmonary alveolar macrophages, followed by MARC145 cells, peripheral blood monocytes, and finally wild boar lung cells. Intraclass correlation coefficient estimates of 0.97 (0.96-0.98) between qPCR methods, 0.80 (0.74-0.85) to 0.94 (0.92-0.96) between virus infectivity assays, and 0.77 (0.68-0.83) to 0.95 (0.93-0.96) between qPCR methods and virus infectivity assays were obtained. These findings show that qPCR gives the highest probability for the detection of ASFV.

Increase in infected corneal ulcerations in dogs during the northern Colorado's 2020 wildfire season.

OBJECTIVE: In the fall of 2020, Colorado experienced the two largest wildfires in state history. The smoke blanketed the college town of Fort Collins, Colorado, the location of the Veterinary Teaching Hospital at Colorado State University (CSU-VTH). The objective for this cross-sectional observational study was to evaluate how these wildfires and the corresponding elevated air quality index (AQI)) was associated with infected corneal ulcerations in dogs when compared to the two previous years. ANIMALS: Seventeen dogs were included in this study. PROCEDURES: Medical records from dogs presented to the CSU-VTH ophthalmology service with infected corneal ulcerations in August, September, and October of 2020, 2019, and 2018 were evaluated. Only corneal ulcerations with growth on their microbial cultures were included in this study. RESULTS: The study revealed a significant increase in prevalence of infected corneal ulcerations in dogs presented to the CSU-VTH during the three wildfire months of 2020 that is, 3.5% (9/255) when compared with the two previous years, 2019: 1.0% (4/383, p = 0.04), and 2018: 0.9% (4/457) (p = .01). The AQI (mean ± standard error) was also significantly elevated for dogs that presented with infected corneal ulcerations in 2020 (70.2 ± 5.8) compared with 2019 (19.7 ± 8.7) and 2018 (45.6 ± 8.7) (p < .01). CONCLUSIONS AND CLINICAL RELEVANCE: Elevation of AQI from wildfires seems to be correlated with an increased prevalence of infected corneal ulceration in dogs. As the duration and frequency of wildfires continues to rise globally, the effects of these wildfires on animal health should be investigated further.

Cataracts and phacoemulsification in the Siberian Husky: A retrospective and multicentric study (2008-2018).

PURPOSE: To compare cataract characteristics and complications related to cataracts and phacoemulsification in the Siberian Husky (Huskies) versus other canine breeds (non-Huskies). ANIMALS: A total of 50 Huskies (92 eyes) and 96 non-Huskies (182 eyes) were evaluated. METHODS: Medical records of Huskies (at four university veterinary hospitals, 2008-2018) and non-Huskies (Colorado State University, 2017-2018) diagnosed with cataracts were reviewed. Age of dog, cataract stage at presentation, and pre- and post-operative complications were recorded and analyzed. RESULTS: Mean (±standard deviation) age at presentation was significantly lower in Huskies (3.5 ± 3.3 years) compared to non-Huskies (9.5 ± 2.9 years) (p < .0001). Huskies more commonly presented with hereditary cataracts than non-Huskies (84% versus 52%) and a significantly higher percentage of non-Huskies presented with diabetic cataracts than Huskies (48% versus 16%; p = .0001). Cataract stage at presentation did not differ between Huskies and non-Huskies. Phacoemulsification was performed in 40% (20 out of 50 dogs, 39 out of 92 eyes) of Huskies and 42% (40 out of 96 dogs, 74 out of 182 eyes) of non-Huskies. Pre-operative and post-operative retinal detachment were more common in Huskies than non-Huskies (13% versus 2% and 10% versus 1%, respectively) but the difference was not significant. Other post-operative complications occurred with similar frequency in both groups (p ≥ .17). CONCLUSIONS: Huskies evaluated for cataracts were younger and less likely to present with diabetic cataracts than other canine breeds and, although not statistically significant, had a clinically important increased risk of retinal detachment pre- and post-phacoemulsification.

Individual and herd-level milk ELISA test status for Johne's disease in Ireland after correcting for non-disease-associated variables.

Antibody-detecting tests for Mycobacterium avium ssp. paratuberculosis (MAP) have low sensitivity and imperfect specificity for detection of infection. Sensitivity increases as the disease progresses. Aside from infection status and stage of disease, several factors affect test performance. These factors have not yet been studied in dairy cows producing lower volumes of milk with higher solids concentration, such as those managed in low-input, pasture-based production systems. Furthermore, the effect of correcting for these associations on individual and herd test status is also unknown. The first objective of this study was to examine the relationship between MAP antibody response in milk and milk yield, somatic cell count (SCC), fat and protein contents, and stage of lactation in dairy cows enrolled in the national Johne's Disease Control Programme (JDCP) in Ireland. The second objective was to examine the effect of correcting the antibody response for these associations on the test status of individual cows and herds, given that individual tests are often used to define a herd's status. Data were extracted for herds in the JDCP from January 2014 to December 2015 inclusive, consisting of 42,657 milk recordings from 18,569 cows across 187 dairy herds. Two linear regression models were constructed to investigate the association between log-transformed MAP sample-to-positive ratio and milk recording data and in primi- and multiparous cows. Days in milk was modeled as a B-spline in each model, and cow and herd were included as random effects. Across both models, natural log-transformed MAP antibody response was negatively associated with milk yield, positively associated with protein and fat production, and had a curvilinear association with log-transformed SCC. The association between MAP antibody response and days in milk varied over the course of the lactation. However, when combined, these variables explained only 5.1% of the variation in the antibody response of the population. After correcting for these associations, 93 multiparous cows and 20 primiparous cows changed category (negative, suspect, or positive). When considered at the herd-test level, out of a total of 531 herd tests, 1 herd changed from negative to positive, and 5 herds changed from positive to negative. This study provides useful information to aid in the interpretation of antibody results for herds testing animals for the presence of MAP infection. At an overall population level, correction of the serological response for non-disease-associated factors has the potential to change the status of only a small number of cows. At the herd level, the proportion of herds changing status was minimal. However, depending on the implications of a herd-level serological diagnosis, consideration should be given to correcting for these non-disease-associated variables within the context of national JD control programs.

Welfare of pigs during killing for purposes other than slaughter.

Pigs at different stages of the production cycle may have to be killed on-farm for purposes other than slaughter (where slaughter is defined as killing for human consumption) either individually (e.g. severely injured pigs) or on a large scale (e.g. unproductive animals or for disease control reasons). This opinion assessed the risks associated with the on-farm killing of pigs and included two phases: 1) handling and moving of pigs and 2) killing methods (including restraint). The killing methods were subdivided into four categories: electrical methods, mechanical methods, gas mixture methods and lethal injection. Four welfare consequences to which pigs can be exposed to during on-farm killing were identified: pain, fear, impeded movement and respiratory distress. Welfare consequences and relevant animal-based measures were described. In total, 28 hazards were associated with the welfare consequences; majority of the hazards (24) are related to Phase 2 (killing). The main hazards are associated with lack of staff skills and training, and poor-designed and constructed facilities. Staff was identified as an origin of all hazards, either due to lack of skills needed to perform appropriate killing or due to fatigue. Corrective measures were identified for 25 hazards. Outcome tables linking hazards, welfare consequences, animal-based measures, hazard origins, preventive and corrective measures were developed and mitigation measures proposed.

Stunning methods and slaughter of rabbits for human consumption.

This opinion on the killing of rabbits for human consumption ('slaughtering') responds to two mandates: one from the European Parliament (EP) and the other from the European Commission. The opinion describes stunning methods for rabbits known to the experts in the EFSA working group, which can be used in commercial practice, and which are sufficiently described in scientific and technical literature for the development of an opinion. These are electrical stunning, mechanical stunning with a penetrative and non-penetrative captive bolt and gas stunning. The latter method is not allowed in the EU anymore following Council Regulation (EC) No 1099/2009, but may still be practiced elsewhere in the world. Related hazards and welfare consequences are also evaluated. To monitor stunning effectiveness as requested by the EP mandate, the opinion suggests the use of indicators for the state of consciousness, selected on the basis of their sensitivity, specificity and ease of use. Similarly, it suggests indicators to confirm animals are dead before dressing. For the European Commission mandate, slaughter processes were assessed from the arrival of rabbits in containers until their death, and grouped in three main phases: pre-stunning (including arrival, unloading of containers from the truck, lairage, handling/removing of rabbits from containers), stunning (including restraint) and bleeding (including bleeding following stunning and bleeding during slaughter without stunning). Ten welfare consequences resulting from the hazards that rabbits can be exposed to during slaughter are identified: consciousness, animal not dead, thermal stress (heat or cold stress), prolonged thirst, prolonged hunger, restriction of movements, pain, fear, distress and respiratory distress. Welfare consequences and relevant animal-based measures (indicators) are described. Outcome tables linking hazards, welfare consequences, indicators, origins, preventive and corrective measures are developed for each process. Mitigation measures to minimise welfare consequences are also proposed.

Scientific opinion concerning the killing of rabbits for purposes other than slaughter.

Rabbits of different ages may have to be killed on-farm for purposes other than slaughter (where slaughter is defined as killing for human consumption) either individually or on a large scale (e.g. for production reasons or for disease control). The purpose of this opinion was to assess the risks associated to the on-farm killing of rabbits. The processes during on-farm killing that were assessed included handling, stunning and/or killing methods (including restraint). The latter were grouped into four categories: electrical methods, mechanical methods, controlled atmosphere method and lethal injection. In total, 14 hazards were identified and characterised, most of these related to stunning and/or killing. The staff was identified as the origin for all hazards, either due to lack of the appropriate skill sets needed to perform tasks or due to fatigue. Possible corrective and preventive measures were assessed: measures to correct hazards were identified for five hazards and the staff was shown to have a crucial role in prevention. Five welfare consequences of the welfare hazards to which rabbits can be exposed to during on-farm killing were identified: not being dead, consciousness, pain, fear and distress. Welfare consequences and relevant animal-based measures were described. Outcome tables linking hazards, welfare consequences, animal-based measures, origins, preventive and corrective measures were developed for each process. Mitigation measures to minimise welfare consequences are proposed.

Health and welfare of rabbits farmed in different production systems.

The AGRI committee of the European Parliament requested EFSA to assess the welfare of rabbits farmed in different production systems, including organic production, and to update its 2005 scientific opinion about the health and welfare of rabbits kept for meat production. Considering reproducing does, kits and growing rabbits, this scientific opinion focusses on six different housing systems, namely conventional cages, structurally enriched cages, elevated pens, floor pens, outdoor/partially outdoor systems and organic systems. To compare the level of welfare in the different housing systems and rabbit categories, welfare impact scores for 20 welfare consequences identified from the literature were calculated, taking their occurrence, duration and severity into account. Based on the overall welfare impact score (sum of scores for the single welfare consequences), obtained via a 2-step expert knowledge elicitation process, the welfare of reproducing does is likely (certainty 66-90%) to be lower in conventional cages compared to the five other housing systems. In addition, it is likely to extremely likely (certainty 66-99%) that the welfare of kits is lower in outdoor systems compared to the other systems and that the welfare is higher in elevated pens than in the other systems. Finally, it is likely to extremely likely (certainty 66-99%) that the welfare of growing rabbits is lower in conventional cages compared to the other systems and that the welfare is higher in elevated pens than in the other systems. Ranking of the welfare consequences allowed an analysis of the main welfare consequences within each system and rabbit category. It was concluded that for reproducing does, as well as growing rabbits, welfare consequences related to behavioural restrictions were more prominent in conventional cages, elevated pens and enriched cages, whereas those related to health problems were more important in floor pens, outdoor and organic systems. Housing in organic rabbit farming is diverse, which can result in different welfare consequences, but the overall welfare impact scores suggest that welfare in organic systems is generally good.

INVESTIGATION INTO CARDIOVASCULAR ASSESSMENT OF CAPTIVE ADULT SCARLET IBIS (EUDOCIMUS RUBER).

Cardiovascular lesions are commonly diagnosed postmortem in scarlet ibis (Eudocimus ruber), but antemortem diagnosis is rare. The aim of this study was to evaluate the cardiovascular health of a zoo population (n = 44) of apparently healthy, adult, scarlet ibis. A cross-sectional study design was employed whereby each animal was manually restrained for physical examination, phlebotomy, and echocardiographic examination performed with a 12-MHz transducer and a ventromedial approach, and observed intervals were calculated for 12 parameters. Seven individuals from the study population had high left-sided mid-ventricular velocities (2.59-5.89 m/sec) compared with values in other species. Follow-up examination suggested that these mid-ventricular obstructive lesions were dynamic and transient in nature rather than caused by fixed lesions within the outflow tract and may therefore be associated with stress. Conscious echocardiography proved to be feasible, although, unsurprisingly, the stress response in nonhabituated birds appeared to increase blood flow velocities. Handling protocols likely have a significant effect on echocardiographic parameters and should be taken into consideration when interpreting findings. Serum cholesterol concentrations were generally high (7.4-13.0 mmol/L), and further work is required to investigate the relationship between circulating cholesterol and the development of atherosclerosis in scarlet ibis. Serum cardiac troponin I concentrations were measured, and four animals were identified with suspected elevated levels, likely indicative of myocardial damage.

SIGMA Animal Disease Data Model: A comprehensive approach for the collection of standardised data on animal diseases.

The European Commission is routinely asking EFSA for scientific and technical support in the epidemiological analysis of animal disease outbreaks (i.e. African swine fever, lumpy skin disease and avian influenza) and to report or assess surveillance data (i.e. Echinococcus multilocularis and avian influenza). For this purpose, EFSA has over the last years carried out several data collections and gathered specific information on outbreaks, surveillance activities and concerned animal populations (i.e. poultry, domestic pigs, cattle and wildlife such as wild boar). EFSA aims to work together closely with Member States in order to (i) reduce the Member States' manual input of the data to be submitted to EFSA; (ii) avoid double reporting to EFSA; (iii) provide the Member States with tools to produce automatically their own draft national reports on animal health and surveillance in a protected environment to ensure data protection; (iv) increase the quality of the data received from the Member States; and (v) shorten the time to retrieve up-to-date data, relevant for risk assessment purposes. With this purpose, EFSA launched a project called SIGMA. It is important to highlight that the SIGMA - Animal Disease Data Model (σ-ADM) focuses on data which are known to be already collected by several Member States under different legal frameworks and for different purposes. The version presented in this report, will be subject to modifications and updates derived from the feedback during the implementation phase.

Risk of survival, establishment and spread of Batrachochytrium salamandrivorans (Bsal) in the EU.

Batrachochytrium salamandrivorans (Bsal) is an emerging fungal pathogen of salamanders. Despite limited surveillance, Bsal was detected in kept salamanders populations in Belgium, Germany, Spain, the Netherlands and the United Kingdom, and in wild populations in some regions of Belgium, Germany and the Netherlands. According to niche modelling, at least part of the distribution range of every salamander species in Europe overlaps with the climate conditions predicted to be suitable for Bsal. Passive surveillance is considered the most suitable approach for detection of Bsal emergence in wild populations. Demonstration of Bsal absence is considered feasible only in closed populations of kept susceptible species. In the wild, Bsal can spread by both active (e.g. salamanders, anurans) and passive (e.g. birds, water) carriers; it is most likely maintained/spread in infected areas by contacts of salamanders or by interactions with anurans, whereas human activities most likely cause Bsal entry into new areas and populations. In kept amphibians, Bsal contamination via live silent carriers (wild birds and anurans) is considered extremely unlikely. The risk-mitigation measures that were considered the most feasible and effective: (i) for ensuring safer international or intra-EU trade of live salamanders, are: ban or restrictions on salamander imports, hygiene procedures and good practice manuals; (ii) for protecting kept salamanders from Bsal, are: identification and treatment of positive collections; (iii) for on-site protection of wild salamanders, are: preventing translocation of wild amphibians and release/return to the wild of kept/temporarily housed wild salamanders, and setting up contact points/emergency teams for passive surveillance. Combining several risk-mitigation measures improve the overall effectiveness. It is recommended to: introduce a harmonised protocol for Bsal detection throughout the EU; improve data acquisition on salamander abundance and distribution; enhance passive surveillance activities; increase public and professionals' awareness; condition any movement of captive salamanders on Bsal known health status.

Guidance on the assessment criteria for applications for new or modified stunning methods regarding animal protection at the time of killing.

This guidance defines the process for handling applications on new or modified stunning methods and the parameters that will be assessed by the EFSA Animal Health and Welfare (AHAW) Panel. The applications, received through the European Commission, should contain administrative information, a checklist of data to be submitted and a technical dossier. The dossier should include two or more studies (in laboratory and slaughterhouse conditions) reporting all parameters and methodological aspects that are indicated in the guidance. The applications will first be scrutinised by the EFSA's Applications Desk (APDESK) Unit for verification of the completeness of the data submitted for the risk assessment of the stunning method. If the application is considered not valid, additional information may be requested from the applicant. If considered valid, it will be subjected to assessment phase 1 where the data related to parameters for the scientific evaluation of the stunning method will be examined by the AHAW Panel. Such parameters focus on the stunning method and the outcomes of interest, i.e. immediate onset of unconsciousness or the absence of avoidable pain, distress and suffering until the loss of consciousness and duration of the unconsciousness (until death). The applicant should also propose methodologies and results to assess the equivalence with existing stunning methods in terms of welfare outcomes. Applications passing assessment phase 1 will be subjected to the following phase 2 which will be carried out by the AHAW Panel and focuses on the animal welfare risk assessment. In this phase, the Panel will assess the outcomes, conclusions and discussion proposed by the applicant. The results of the assessment will be published in a scientific opinion.

Animal welfare aspects in respect of the slaughter or killing of pregnant livestock animals (cattle, pigs, sheep, goats, horses).

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).