Using registry data to identify individual dairy cows with abnormal patterns in routinely recorded somatic cell counts.

Data from the Danish milk recording system routinely enter the Danish Cattle Database, including somatic cell counts (SCC) for individual animals. Elevated SCC can signal intramammary inflammation, suggesting subclinical mastitis. Detecting mastitis is pivotal to limit severity, prevent pathogen spread, and target treatment or culling. This study aimed to differentiate normal and abnormal SCC patterns using recorded registry data. We used registry data from 2010 to 2020 for dairy cows in herds with 11 annual milk recordings. To create consistency across herds, we used data from 13,996 unique animals and eight different herds, selected based on the amount of data available, only selecting Holstein animals and conventional herds. We fitted log10-transformed SCC to days in milk (DIM) using the Wilmink and Wood's curve functions, originally developed for milk yield over the lactation. We used Nonlinear Least Square and Nonlinear Mixed Effect models to fit the log10-transformed SCC observations to DIM at animal level. Using mean squared residuals (MSR), we found a consistently better fit using a Wood's style function. Detection of MSR outliers in the model fitting process was used to identify animals with log10(SCC) curves deviating from the expected "normal" curve for that same animal. With this study, we propose a method to identify single animals with SCC patterns that indicate abnormalities, such as mastitis, based on registry data. This method could potentially lead to a registry data-based detection of mastitis cases in larger dairy herds.

Perceptions of the rabbit as a low investment 'starter pet' lead to negative impacts on its welfare: Results of two Danish surveys.

Concerns over compromised companion rabbit (Oryctolagus cuniculus domesticus) welfare are widespread. The welfare problems have been linked to the perception of rabbits as low investment 'children's pets.' To test this hypothesis and investigate the current conditions for rabbits, data were gathered from two surveys in 2021: a nationally representative survey of Danish companion animal owners (Survey I) and a detailed social media-based survey of Danish rabbit owners (Survey II). Using logistic regression, three owner-related variables (whether a child/adult was responsible for care of the rabbit, owner-opinion on rabbits' suitability as 'starter pets' and willingness-to-pay [WTP] for veterinary treatment) were employed to investigate the effect of rabbit status on owner-provision of selected husbandry conditions. The 76 (Survey I) and 4,335 (Survey II) responses suggested that most rabbits are acquired for children and are solitarily housed, and that many are kept in cages of an unsuitable size and not checked daily. Owners who perceived rabbits as 'starter pets' and with lower WTP were more likely to house rabbits in restricted space and to not provide continuous gnawing opportunities, ad libitum hay or routine healthcare. A child fulfilling the role of the rabbit's main caretaker was also associated with inadequate housing type and fewer gnawing opportunities. Thus, many rabbits live in unsuitable conditions, and owners who perceive rabbits as low investment 'children's pets' are more likely to not provide recommended resources. Changing owners' perceptions of rabbits and promoting suitable husbandry through official education programmes and minimum requirements is important if there are to be improvements made to rabbit welfare.

BIOLOGICAL VARIATION OF HEMATOLOGY AND BIOCHEMISTRY PARAMETERS FOR THE ASIAN ELEPHANT (ELEPHAS MAXIMUS), AND APPLICABILITY OF POPULATION-DERIVED REFERENCE INTERVALS.

The aim of this study was to objectively evaluate the biological variation of healthy Asian elephant (Elephas maximus) hematology and biochemistry parameters, therefore enabling evidence-based clinical decision-making to improve patient management. Ten clinically healthy elephants had blood samples collected weekly for 5 wk under standardized conditions. The analytical, between- and within-individual variation, index of individuality, and reference change values were calculated using previously reported methods. Large between-individual variation and small within-individual variation for almost all parameters indicated that individual normal values should be used for interpreting blood results from Asian elephants.

Control of paratuberculosis: who, why and how. A review of 48 countries.

Paratuberculosis, a chronic disease affecting ruminant livestock, is caused by Mycobacterium avium subsp. paratuberculosis (MAP). It has direct and indirect economic costs, impacts animal welfare and arouses public health concerns. In a survey of 48 countries we found paratuberculosis to be very common in livestock. In about half the countries more than 20% of herds and flocks were infected with MAP. Most countries had large ruminant populations (millions), several types of farmed ruminants, multiple husbandry systems and tens of thousands of individual farms, creating challenges for disease control. In addition, numerous species of free-living wildlife were infected. Paratuberculosis was notifiable in most countries, but formal control programs were present in only 22 countries. Generally, these were the more highly developed countries with advanced veterinary services. Of the countries without a formal control program for paratuberculosis, 76% were in South and Central America, Asia and Africa while 20% were in Europe. Control programs were justified most commonly on animal health grounds, but protecting market access and public health were other factors. Prevalence reduction was the major objective in most countries, but Norway and Sweden aimed to eradicate the disease, so surveillance and response were their major objectives. Government funding was involved in about two thirds of countries, but operations tended to be funded by farmers and their organizations and not by government alone. The majority of countries (60%) had voluntary control programs. Generally, programs were supported by incentives for joining, financial compensation and/or penalties for non-participation. Performance indicators, structure, leadership, practices and tools used in control programs are also presented. Securing funding for long-term control activities was a widespread problem. Control programs were reported to be successful in 16 (73%) of the 22 countries. Recommendations are made for future control programs, including a primary goal of establishing an international code for paratuberculosis, leading to universal acknowledgment of the principles and methods of control in relation to endemic and transboundary disease. An holistic approach across all ruminant livestock industries and long-term commitment is required for control of paratuberculosis.

Epidemiological and economic consequences of purchasing livestock infected with Mycobacterium avium subsp. paratuberculosis.

BACKGROUND: Paratuberculosis (PTB) is a chronic disease which may lead to reduced milk yield, lower animal welfare and death in cattle. The causative agent is Mycobacterium avium subsp. paratuberculosis (MAP). The economic consequences are particularly important incentives in the control and eradication of the infection. One strategy to control PTB in a herd is to purchase animals from farms with a low risk of MAP infection. We wanted to investigate the epidemiological and economic consequences of buying livestock from different supplier farms of low, medium or high risk, as well as farms with unknown status. We also wanted to estimate the probability of spontaneous fadeout if the farmer of an initially MAP-free herd bought a specified number of infected animals in a single year, or continually bought infected animals. This was achieved through simulation modeling, and the effects of consistently introducing one, five or ten infected animals annually into an initially infection-free herd was also modeled. RESULTS: Our findings show that once infected, a farm can relatively safely purchase animals from other low and medium-risk farms without experiencing an increase in the prevalence, highlighting the importance of certification programmes. Furthermore, farms free of MAP are highly susceptible and cannot purchase more than a small number of animals per year without having a high risk of being infected. The probability of spontaneous fadeout after 10 years was 82% when introducing a single infected animal into an initially MAP-free herd. When purchasing ten infected animals, this probability was 46%. The continual purchase of infected animals resulted in very low probabilities of spontaneous fadeout. CONCLUSIONS: We demonstrated that MAP-free farms can purchase a small number of animals, preferably from certified farms, each year and still remain free of MAP. Already infected farms have little risk of increasing the prevalence on a farm when purchasing animals from other farms.

Changes in concentrations of haemostatic and inflammatory biomarkers in synovial fluid after intra-articular injection of lipopolysaccharide in horses.

BACKGROUND: Septic arthritis is a common and potentially devastating disease characterized by severe intra-articular (IA) inflammation and fibrin deposition. Research into equine joint pathologies has focused on inflammation, but recent research in humans suggests that both haemostatic and inflammatory pathways are activated in the joint compartment in arthritic conditions. The aim of this study was to characterize the IA haemostatic and inflammatory responses in horses with experimental lipopolysaccharide (LPS)-induced joint inflammation. Inflammation was induced by IA injection of LPS into one antebrachiocarpal joint of six horses. Horses were evaluated clinically with subjective grading of lameness, and blood and synovial fluid (SF) samples were collected at post injection hours (PIH) -120, -96, -24, 0, 2, 4, 8, 16, 24, 36, 48, 72 and 144. Total protein (TP), white blood cell counts (WBC), serum amyloid A (SAA), haptoglobin, iron, fibrinogen, thrombin-antithrombin (TAT) and d-dimer concentrations were assessed in blood and SF. RESULTS: Intra-articular injection of LPS caused local and systemic signs of inflammation including increased rectal temperature, lameness and increased joint circumference and skin temperature. Most of the biomarkers (TP, WBC, haptoglobin, fibrinogen and TAT) measured in SF increased quickly after LPS injection (at PIH 2-4), whereas SAA and d-dimer levels increased more slowly (at PIH 16 and 144, respectively). SF iron concentrations did not change statistically significantly. Blood WBC, SAA, haptoglobin and fibrinogen increased and iron decreased significantly in response to the IA LPS injection, while TAT and d-dimer concentrations did not change. Repeated pre-injection arthrocenteses caused significant changes in SF concentrations of TP, WBC and haptoglobin. CONCLUSION: Similar to inflammatory joint disease in humans, joint inflammation in horses was accompanied by an IA haemostatic response with changes in fibrinogen, TAT and d-dimer concentrations. Inflammatory and haemostatic responses were induced simultaneously and may likely interact. Further studies of interactions between the two responses are needed for a better understanding of pathogenesis of joint disease in horses. Knowledge of effects of repeated arthrocenteses on levels of SF biomarkers may be of value when markers are used for diagnostic purposes.

Multistrain models predict sequential multidrug treatment strategies to result in less antimicrobial resistance than combination treatment.

BACKGROUND: Combination treatment is increasingly used to fight infections caused by bacteria resistant to two or more antimicrobials. While multiple studies have evaluated treatment strategies to minimize the emergence of resistant strains for single antimicrobial treatment, fewer studies have considered combination treatments. The current study modeled bacterial growth in the intestine of pigs after intramuscular combination treatment (i.e. using two antibiotics simultaneously) and sequential treatments (i.e. alternating between two antibiotics) in order to identify the factors that favor the sensitive fraction of the commensal flora. Growth parameters for competing bacterial strains were estimated from the combined in vitro pharmacodynamic effect of two antimicrobials using the relationship between concentration and net bacterial growth rate. Predictions of in vivo bacterial growth were generated by a mathematical model of the competitive growth of multiple strains of Escherichia coli. RESULTS: Simulation studies showed that sequential use of tetracycline and ampicillin reduced the level of double resistance, when compared to the combination treatment. The effect of the cycling frequency (how frequently antibiotics are alternated in a sequential treatment) of the two drugs was dependent upon the order in which the two drugs were used. CONCLUSION: Sequential treatment was more effective in preventing the growth of resistant strains when compared to the combination treatment. The cycling frequency did not play a role in suppressing the growth of resistant strains, but the specific order of the two antimicrobials did. Predictions made from the study could be used to redesign multidrug treatment strategies not only for intramuscular treatment in pigs, but also for other dosing routes.

Modeling the growth dynamics of multiple Escherichia coli strains in the pig intestine following intramuscular ampicillin treatment.

BACKGROUND: This study evaluated how dosing regimen for intramuscularly-administered ampicillin, composition of Escherichia coli strains with regard to ampicillin susceptibility, and excretion of bacteria from the intestine affected the level of resistance among Escherichia coli strains in the intestine of nursery pigs. It also examined the dynamics of the composition of bacterial strains during and after the treatment. The growth responses of strains to ampicillin concentrations were determined using in vitro growth curves. Using these results as input data, growth predictions were generated using a mathematical model to simulate the competitive growth of E. coli strains in a pig intestine under specified plasma concentration profiles of ampicillin. RESULTS: In vitro growth results demonstrated that the resistant strains did not carry a fitness cost for their resistance, and that the most susceptible strains were more affected by increasing concentrations of antibiotics that the rest of the strains. The modeling revealed that short treatment duration resulted in lower levels of resistance and that dosing frequency did not substantially influence the growth of resistant strains. Resistance levels were found to be sensitive to the number of competing strains, and this effect was enhanced by longer duration of treatment. High excretion of bacteria from the intestine favored resistant strains over sensitive strains, but at the same time it resulted in a faster return to pre-treatment levels after the treatment ended. When the duration of high excretion was set to be limited to the treatment time (i.e. the treatment was assumed to result in a cure of diarrhea) resistant strains required longer time to reach the previous level. CONCLUSION: No fitness cost was found to be associated with ampicillin resistance in E. coli. Besides dosing factors, epidemiological factors (such as number of competing strains and bacterial excretion) influenced resistance development and need to be considered further in relation to optimal treatment strategies. The modeling approach used in the study is generic, and could be used for prediction of the effect of treatment with other drugs and other administration routes for effect on resistance development in the intestine of pigs.

Apramycin treatment affects selection and spread of a multidrug-resistant Escherichia coli strain able to colonize the human gut in the intestinal microbiota of pigs.

The effect of apramycin treatment on transfer and selection of an Escherichia coli strain (E. coli 912) in the intestine of pigs was analyzed through an in vivo experiment. The strain was sequenced and assigned to the sequence type ST101 and serotype O11. It carried resistance genes to apramycin/gentamicin, sulphonamide, tetracycline, hygromycin B, ß-lactams and streptomycin [aac(3)-IV, sul2, tet(X), aph(4), bla TEM-1 and strA/B], with all but tet(X) located on the same conjugative plasmid. Nineteen pigs were randomly allocated into two inoculation groups, one treated with apramycin (pen 2) and one non-treated (pen 3), along with a non-inoculated control group (pen 1). Two pigs of pen 2 and 3 were inoculated intragastrically with a rifampicin resistant variant of the strain. Apramycin treatment in pen 2 was initiated immediately after inoculation. Strain colonization was assessed in the feces from all pigs. E. coli 912 was shown to spread to non-inoculated pigs in both groups. The selective effect did not persist beyond 3 days post-treatment, and the strain was not detected from this time point in pen 2. We demonstrated that E. coli 912 was able to spread between pigs in the same pen irrespective of treatment, and apramycin treatment resulted in significantly higher counts compared to the non-treated group. This represents the first demonstration of how antimicrobial treatment affects spread of resistant bacteria in pig production. The use of apramycin may lead to enhanced spread of gentamicin-resistant E. coli. Since gentamicin is a first-choice drug for human bacteremia, this is of concern.

Gross and histopathological evaluation of human inflicted bruises in Danish slaughter pigs.

BACKGROUND: Human inflicted bruises in slaughter pigs are hampering animal welfare, are an infringement of the animal protection act, and are a focus of public attention. The aim of the present study was to evaluate the gross appearance of human inflicted bruises in slaughter pigs and to compare the inflammatory changes in two lesions as a basis for estimating the age of lesions in the same pig. Pigs with human inflicted bruises slaughtered at two major slaughterhouses in Denmark from November 2013 to May 2014 were evaluated. After slaughter, the bruises were examined grossly and skin and underlying muscle tissue from two similar but separate bruises (a and b) on each pig were sampled for histology. RESULTS: Skin and muscle tissue from 101 slaughter pigs were subjected to gross evaluation. Eighty-one of these were also subjected to histological evaluation. Most frequently (51 out of 101 pigs, 50 %), bruises had a tram-line pattern due to blunt trauma inflicted with long objects such as sticks. Other bruises reflected the use of tattoo-hammers, plastic paddles, double U profiles and chains. Histological evaluation of two bruises from a pig with multiple lesions was found insufficient to assess the overall age of the lesions as substantial variation in the inflammatory response between bruises was present. CONCLUSIONS: Grossly, the pattern of bruises often reflected the shape of the object used for inflicting the lesions. When determining the age of multiple bruises on a pig more than two lesions should be evaluated histologically.

Student Preparation and the Power of Visual Input in Veterinary Surgical Education: An Empirical Study.

In recent years, veterinary educational institutions have implemented alternative teaching methods, including video demonstrations of surgical procedures. However, the power of the dynamic visual input from videos in relation to recollection of a surgical procedure has never been evaluated. The aim of this study was to investigate how veterinary surgical students perceived the influence of different educational materials on recollection of a surgical procedure. Furthermore, we investigated if surgical technique was associated with a certain method of recollection or use of educational material. During a basic surgical skills course, 112 fourth-year veterinary students participated in the study by completing a questionnaire regarding method of recollection, influence of individual types of educational input, and homework preparation. Furthermore, we observed students performing an orchiectomy in a terminal pig lab. Preparation for the pig lab consisted of homework (textbook, online material, including videos), lecture, cadaver lab, and toy animal models in a skills lab. In the instructional video, a detail was used that was not described elsewhere. Results show that 60% of the students used a visual dynamic method as their main method of recollection and that video was considered the most influential educational input with respect to recollection of a specific procedure. Observation of students' performance during the orchiectomy showed no clear association with students' method of recollection but a significant association (p=.002) with educational input. Our results illustrate the power of a visual input and support prior findings that knowledge is constructed from multiple sources of information.

Pharmacokinetic-pharmacodynamic model to evaluate intramuscular tetracycline treatment protocols to prevent antimicrobial resistance in pigs.

High instances of antimicrobial resistance are linked to both routine and excessive antimicrobial use, but excessive or inappropriate use represents an unnecessary risk. The competitive growth advantages of resistant bacteria may be amplified by the strain dynamics; in particular, the extent to which resistant strains outcompete susceptible strains under antimicrobial pressure may depend not only on the antimicrobial treatment strategies but also on the epidemiological parameters, such as the composition of the bacterial strains in a pig. This study evaluated how variation in the dosing protocol for intramuscular administration of tetracycline and the composition of bacterial strains in a pig affect the level of resistance in the intestine of a pig. Predictions were generated by a mathematical model of competitive growth of Escherichia coli strains in pigs under specified plasma concentration profiles of tetracycline. All dosing regimens result in a clear growth advantage for resistant strains. Short treatment duration was found to be preferable, since it allowed less time for resistant strains to outcompete the susceptible ones. Dosing frequency appeared to be ineffective at reducing the resistance levels. The number of competing strains had no apparent effect on the resistance level during treatment, but possession of fewer strains reduced the time to reach equilibrium after the end of treatment. To sum up, epidemiological parameters may have more profound influence on growth dynamics than dosing regimens and should be considered when designing improved treatment protocols.

Mean effective sensitivity for Mycobacterium avium subsp. paratuberculosis infection in cattle herds.

BACKGROUND: Mycobacterium avium subsp. paratuberculosis (MAP) infections in cattle are generally challenging to detect and cost-effective test strategies are consequently difficult to identify. MAP-specific antibody ELISAs for milk and serum are relatively inexpensive, but their utility is influenced by a number of factors such as herd size, herd composition and diagnostic sensitivity. The sensitivity of the test increases with the age of the tested animal, and therefore the general, or "mean effective sensitivity" (defined as the mean of the sensitivities for all animals within a population, MES), for detecting MAP within a herd is dependent upon the age distribution of the herd. For this study we used a dataset of cattle from 4,259 dairy herds and 4,078 non-dairy herds. The aim was to investigate the MES for groups of cattle considered to be reasonable entities for MAP surveillance and control, in order to assist the decision-makers in planning and optimizing these programs economically. We compared six different groups of cattle (three dairy and three non-dairy) in Denmark by calculating the MES for each herd in each group. RESULTS: The distribution of MES showed a large variation within and between groups, and in some groups we found a bimodal distribution of MES. Dairy herds generally showed higher MES than non-dairy herds. Dairy herds in a control programme for paratuberculosis showed a MES similar to all other dairy herds from which animals > 2.0 years were tested (both groups had a median MES = 0.60). For the non-dairy groups, the sensitivity became much higher when animals < 2.0 years and herds with less than 25 cattle were excluded, resulting in a median MES of 0.65. CONCLUSION: The results showed that MES could indicate the effectivity of testing different cattle groups for MAP, given that the data used are unbiased.

mRNA expression of genes involved in inflammation and haemostasis in equine fibroblast-like synoviocytes following exposure to lipopolysaccharide, fibrinogen and thrombin.

BACKGROUND: Studies in humans have shown that haemostatic and inflammatory pathways both play important roles in the pathogenesis of joint disease. The aim of this study was to assess mRNA expression of haemostatic and inflammatory factors in cultured equine fibroblast-like synoviocytes exposed to lipopolysaccharide (LPS), fibrinogen and thrombin. Synovial membranes were collected from metacarpo-phalangeal joints of 6 skeletally mature horses euthanized for non-orthopaedic reasons. Passage 4 fibroblast-like synoviocytes were left non-treated or treated with either 0.1 µg/ml LPS, 5 mg/ml fibrinogen or 5 U/ml thrombin and harvested at time points 0, 6, 24 and 48 h. mRNA expression of serum amyloid A (SAA), interleukin-6 (IL-6), monocyte chemotactic protein 1 (MCP-1), tissue factor (TF), plasminogen activator inhibitor 1 (PAI-1), urokinase plasminogen activator (uPA), vascular endothelial growth factor (VEGF) and protease activator receptor 1 (PAR-1) was assessed using quantitative real time reverse transcriptase PCR. RESULTS: LPS caused a significant increase in mRNA expression of SAA, IL-6, MCP-1 and uPA, and a decrease in TF, PAI-1 and PAR-1 when compared to non-treated cells. Treatment with thrombin resulted in increased mRNA expression of SAA, IL-6, MCP-1 and PAI-1, and a decreased PAR-1 expression compared to non-treated cells. The fibrinogen-treated synoviocytes showed significantly increased mRNA expression of IL-6, MCP-1, TF and PAI-1, and decreased PAR-1 expression compared to non-treated cells. CONCLUSION: LPS, fibrinogen and thrombin induced an increased gene expression of inflammatory markers in isolated equine fibroblast-like synoviocytes. LPS caused changes in gene expression promoting increased fibrinolysis, while fibrinogen and thrombin changed the gene expression resulting potentially in reduced fibrinolysis. Overall, it appeared that both inflammatory and haemostatic stimuli affected expression of genes involved in inflammatory and haemostatic pathways, supporting their importance in equine joint diseases.

Enhanced adherence of methicillin-resistant Staphylococcus pseudintermedius sequence type 71 to canine and human corneocytes.

The recent worldwide spread of methicillin-resistant Staphylococcus pseudintermedius (MRSP) in dogs is a reason for concern due to the typical multidrug resistance patterns displayed by some MRSP lineages such as sequence type (ST) 71. The objective of this study was to compare the in vitro adherence properties between MRSP and methicillin-susceptible (MSSP) strains. Four MRSP, including a human and a canine strain belonging to ST71 and two canine non-ST71 strains, and three genetically unrelated MSSP were tested on corneocytes collected from five dogs and six humans. All strains were fully characterized with respect to genetic background and cell wall-anchored protein (CWAP) gene content. Seventy-seven strain-corneocyte combinations were tested using both exponential- and stationary-phase cultures. Negative binomial regression analysis of counts of bacterial cells adhering to corneocytes revealed that adherence was significantly influenced by host and strain genotype regardless of bacterial growth phase. The two MRSP ST71 strains showed greater adherence than MRSP non-ST71 (p < 0.0001) and MSSP (p < 0.0001). This phenotypic trait was not associated to any specific CWAP gene. In general, S. pseudintermedius adherence to canine corneocytes was significantly higher compared to human corneocytes (p < 0.0001), but the MRSP ST71 strain of human origin adhered equally well to canine and human corneocytes, suggesting that MRSP ST71 may be able to adapt to human skin. The genetic basis of the enhanced in vitro adherence of ST71 needs to be elucidated as this phenotypic trait may be associated to the epidemiological success and zoonotic potential of this epidemic MRSP clone.

Haematological and biochemical reference intervals for free-ranging brown bears (Ursus arctos) in Sweden.

BACKGROUND: Establishment of haematological and biochemical reference intervals is important to assess health of animals on individual and population level. Reference intervals for 13 haematological and 34 biochemical variables were established based on 88 apparently healthy free-ranging brown bears (39 males and 49 females) in Sweden. The animals were chemically immobilised by darting from a helicopter with a combination of medetomidine, tiletamine and zolazepam in April and May 2006-2012 in the county of Dalarna, Sweden. Venous blood samples were collected during anaesthesia for radio collaring and marking for ecological studies. For each of the variables, the reference interval was described based on the 95% confidence interval, and differences due to host characteristics sex and age were included if detected. To our knowledge, this is the first report of reference intervals for free-ranging brown bears in Sweden. RESULTS: The following variables were not affected by host characteristics: red blood cell, white blood cell, monocyte and platelet count, alanine transaminase, amylase, bilirubin, free fatty acids, glucose, calcium, chloride, potassium, and cortisol. Age differences were seen for the majority of the haematological variables, whereas sex influenced only mean corpuscular haemoglobin concentration, aspartate aminotransferase, lipase, lactate dehydrogenase, ß-globulin, bile acids, triglycerides and sodium. CONCLUSIONS: The biochemical and haematological reference intervals provided and the differences due to host factors age and gender can be useful for evaluation of health status in free-ranging European brown bears.

Prevalence and characterisation of band-shaped tail lesions in Holstein cows.

The aim of the study was to characterise and determine the prevalence of band-shaped tail lesions in Holstein cows. Lesions were present either as wounds or by epithelised granulation/connective tissue formations. Both types were characterised by a median localisation 7 cm from the tip of the tail, and they occurred on the dorsal aspect of the tail. From here they encircled the tail either completely or in varying degrees, and they were often present as isolated lesions (93%). The prevalence of band-shaped tail lesions was found to be 25% among 2099 cows examined in 16 Danish Holstein herds with a variation from 18 to 40% between herds. In the herds, the wound lesions and the connective tissue formations accounted for 22% and 78% of all band-shaped tail lesions, respectively. Among 458 Holstein cows examined at an abattoir the prevalence of band-shaped tail lesions was 23%, i.e. similar to the prevalence within the herds. At the abattoir the share of band-shaped wound lesions was 67% and the band-shaped connective tissue formation 33%. Associations between the occurrence of band-shaped tail lesions and parity and lack of the tail tip were observed.

Vaccination of poultry against highly pathogenic avian influenza - Part 2. Surveillance and mitigation measures.

Selecting appropriate diagnostic methods that take account of the type of vaccine used is important when implementing a vaccination programme against highly pathogenic avian influenza (HPAI). If vaccination is effective, a decreased viral load is expected in the samples used for diagnosis, making molecular methods with high sensitivity the best choice. Although serological methods can be reasonably sensitive, they may produce results that are difficult to interpret. In addition to routine molecular monitoring, it is recommended to conduct viral isolation, genetic sequencing and phenotypic characterisation of any HPAI virus detected in vaccinated flocks to detect escape mutants early. Following emergency vaccination, various surveillance options based on virological testing of dead birds ('bucket sampling') at defined intervals were assessed to be effective for early detection of HPAIV and prove disease freedom in vaccinated populations. For ducks, virological or serological testing of live birds was assessed as an effective strategy. This surveillance could be also applied in the peri-vaccination zone on vaccinated establishments, while maintaining passive surveillance in unvaccinated chicken layers and turkeys, and weekly bucket sampling in unvaccinated ducks. To demonstrate disease freedom with > 99% confidence and to detect HPAI virus sufficiently early following preventive vaccination, monthly virological testing of all dead birds up to 15 per flock, coupled with passive surveillance in both vaccinated and unvaccinated flocks, is recommended. Reducing the sampling intervals increases the sensitivity of early detection up to 100%. To enable the safe movement of vaccinated poultry during emergency vaccination, laboratory examinations in the 72 h prior to the movement can be considered as a risk mitigation measure, in addition to clinical inspection; sampling results from existing surveillance activities carried out in these 72 h could be used. In this Opinion, several schemes are recommended to enable the safe movement of vaccinated poultry following preventive vaccination.

Scientific Committee guidance on appraising and integrating evidence from epidemiological studies for use in EFSA's scientific assessments.

EFSA requested its Scientific Committee to prepare a guidance document on appraising and integrating evidence from epidemiological studies for use in EFSA's scientific assessments. The guidance document provides an introduction to epidemiological studies and illustrates the typical biases, which may be present in different epidemiological study designs. It then describes key epidemiological concepts relevant for evidence appraisal. This includes brief explanations for measures of association, exposure assessment, statistical inference, systematic error and effect modification. The guidance then describes the concept of external validity and the principles of appraising epidemiological studies. The customisation of the study appraisal process is explained including tailoring of tools for assessing the risk of bias (RoB). Several examples of appraising experimental and observational studies using a RoB tool are annexed to the document to illustrate the application of the approach. The latter part of this guidance focuses on different steps of evidence integration, first within and then across different streams of evidence. With respect to risk characterisation, the guidance considers how evidence from human epidemiological studies can be used in dose-response modelling with several different options being presented. Finally, the guidance addresses the application of uncertainty factors in risk characterisation when using evidence from human epidemiological studies.

Welfare of sheep and goats during killing for purposes other than slaughter.

Sheep and goats 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 (i.e. on-farm killing of unproductive, injured or terminally ill animals) or on a large scale (i.e. depopulation for disease control purposes and for other situations, such as environmental contamination and disaster management) outside the slaughterhouses. The purpose of this opinion was to assess the hazards and welfare consequences associated with the on-farm killing of sheep and goats. The whole killing procedure was divided into Phase 1 (pre-killing) - that included the processes (i) handling and moving the animals to the killing place and (ii) restraint of the animals before application of the killing methods and Phase 2 - that included stunning and killing of the animals. The killing methods for sheep and goats were grouped into three categories: (1) mechanical, (2) electrical and (3) lethal injection. Welfare consequences that sheep and goats may experience during each process were identified (e.g. handling stress, restriction of movements and tissue lesions during restraint) and animal-based measures (ABMs) to assess them were proposed. During application of the killing method, sheep and goats will experience pain and fear if they are ineffectively stunned or if they recover consciousness. ABMs related to the state of consciousness can be used to indirectly assess pain and fear. Flowcharts including ABMs for consciousness specific to each killing method were included in the opinion. Possible welfare hazards were identified for each process, together with their origin and related preventive and corrective measures. Outcome tables linking hazards, welfare consequences, ABMs, origins, preventive and corrective measures were developed for each process. Mitigation measures to minimise welfare consequences were proposed.