Introduction
Many equids are vaccinated against equine influenza annually as it causes a highly contagious respiratory infection. In horses, both age and adiposity contribute to increased levels of inflammatory markers, which could affect the response to vaccination. In humans, a chronic inflammatory state associated with obesity can impair response to vaccination or infection (Sheridan et al., 2012). The objective of this study was to determine which factors would influence the response to the equine influenza vaccination.
Material and methods
Surplus to diagnostic requirement serum samples were obtained from 55 donkeys that had recently received a booster dose of a commercially-available inactivated virus equine influenza vaccine. Antibody levels against a component strain of the vaccine (influenza A/equine/Newmarket/2/93 [H3N8]) were measured using the single radial haemolysis assay. An equine-specific ELISA (R&D Systems) was used to measure serum tumour necrosis factor alpha (TNF-α). High molecular weight (HMW) adiponectin was measured using a human ELISA kit (Millipore) previously shown to be relevant for the horse (Woolridge et al., 2012). Non-esterified fatty acid (NEFA) levels were measured using a kit from Randox with some modifications to the manufacturer’s instructions. Additional data including age, weight, body condition score (BCS), and total cholesterol and triglyceride serum levels were kindly provided by the Donkey Sanctuary. Univariate analysis was conducted using Pearson correlation for normally distributed variables and Spearman’s rank correlation for variables that were not normally distributed. To evaluate the effect of gender, comparisons were made using a t-test for normally-distributed variables or a Mann-Whitney test for variables not normally distributed. Significant differences were determined at the level of p<0.05.
Results
In this study, the factor with the greatest influence on the response to vaccination was gender, with females having significantly higher antibody levels than males (Table 1). In addition, the mean age of the female donkeys was significantly greater than that of the male donkeys. There was no correlation between BCS or weight and antibody levels, but NEFA levels were negatively correlated with antibody levels (p=0.044). Associations between NEFA and age, weight and days since vaccination also reached statistical significance. Triglyceride levels were also positively correlated with days since vaccination although antibody levels were not. Positive correlations were seen between serum adiponectin and age, and triglyceride and cholesterol levels.
Table 1. Influence of gender on variables measured
Variable Mean ±SD (Range) p-value
Males (n=28) Females (n=27)
Age (years) 19.2 ±11.8 25.0 ±6.4 .028
Antibody (area of lysis - mm2) 178.9 ±39.3 208.9 ±42.3 .009
Conclusions
Negative correlations were expected between antibody levels and both age and BCS, but were not seen. This may have been confounded by the narrow distribution of BCS in the study population (the majority had a BCS score of 2.5–3.5 on a scale of 1–5) and the overwhelming influence of gender with female donkeys having a higher antibody response despite a greater mean age. It has been demonstrated in human subjects that influenza vaccination can cause alterations to the lipid profile (Tsai et al., 2005). The correlation between NEFA and serum antibody levels warrants further investigation as does the finding that gender has a significant impact on response to equine influenza vaccination in donkeys.
Acknowledgments The authors gratefully acknowledge The Donkey Sanctuary for providing surplus to diagnostic requirement serum samples and data, Waltham Centre for Pet Nutrition for funding the study and Dr Marnie Brennan for assistance with the statistical analysis.
References
P. A. Sheridan, H. A. Paich, J. Handy, E. A. Karlsson, M. G. Hudgens, A. B. Sammon, L. A. Holland, S. Weir, T. L. Noah, M. A. Beck. 2012. Obesity is associated with impaired immune response to influenza vaccination in humans. International Journal of Obesity, 36, 1072-1077.
M. Y. Tsai, N. Q. Hanson, R. J. Straka, T. R. Hoke, J. M. Ordovas, J. M. Peacock, V. L. Arends, D. K. Arnett. 2005. Effect of influenza vaccine on markers of inflammation and lipid profile. The Journal of Laboratory and Clinical Medicine, 145, 323-327.
A. A. Wooldridge, H. G. Edwards, E. P. Plaisance, R. Applegate, D. R. Taylor, J. Taintor, Q. Zhong, R. L. Judd. 2012. Evaluation of high-molecular weight adiponectin in horses. American Journal of Veterinary Research, 73, 1230-40.
In the last one hundred years, tractors with much more power replaced animals all over the world, except in developing countries. Besides the advantage in power, the use of tractors increases agriculture productivity and time efficiency of field operations. Despite the unquestionable advantage of tractors for agriculture performance, in recent years the interest by animal traction is growing, even in developed countries. It is important to understand why. This interest usually is linked to small scale farming, but not only. Environmental concern is one of the most significant reasons, in different aspects: use of working animals as a renewable source of energy; forest management in protected areas to protect soil physical properties related to compaction affected by tillage treatments; field operations in environment protected areas, among others. In old vineyards, but producing a high value output, plant spacing turn animal traction without alternative. The maintenance of the gene pool in domestic animals; the creation of jobs in rural areas, such as farrier, harness maker and implement manufacturer, are also important issues. New challenges: new implements must be designed and new mechanical solutions achieved.
In Ethiopia the Donkey Sanctuary started its health and welfare activities in 1994. This was the time when the International Donkey Protection Trust and Addis Ababa University concluded an agreement and signed a memorandum of understanding. Since that time and up to 2003, activities were centred in the districts of Bereh, Boset, Ada and Akaki and Addis Ababa Grain Market located at a radius of 100-150 kms from Debre Zeit, the centre with a stationery clinic. The facilities of the latter include an open-air clinic, surgical theatre, laboratory, store for drugs, boxes for mules and donkeys, offices and a training room. As of 2004, three districts namely: Dugda Bora, Lume and Sebata have been added as sites of operation. Resistance at the very beginning by donkey owners to bring their animals for treatment was a problem but over the years the service has obtained wide acceptance, the trend exhibiting exponential growth: Ada (R2=0.87), Akaki (R2=0.88), Bereh (R2=0.70), Boset (R2=0.70), and for mules (R2=0.84). The clinic has so far provided treatments to 289,999 donkeys and 14,665 mules, and 217,609 owners have benefited. In 2003 two satellite projects have been set up in the northern part of Ethiopia: Amhara and Tigray and have so far treated more than 72,000 donkeys and mules and more than 29,000 owners have benefited. The centre at Debre Zeit was also designed to cater for local and international training, to date it has provided training to 217 veterinary professionals (28 foreign vets, 41 local vets, 17 foreign and 101 local animal health assistants, 30 local animal health technicians) in donkey medicine, surgery, farriery and general welfare. Since 2002, 297 clinical year students enrolled for the DVM degree in the Faculty of Veterinary Medicine of Addis Ababa University have been trained in donkey medicine and surgery. Three to four clinical year students accompany mobile teams in field operations on a weekly rotation basis. Recently, the project has embarked on an education and extension programme in conjunction with its treatment activities and so far 51,513 owners have benefited. In the recently launched school education programme 1014 students in 7 primary schools are attending sessions in the basics of animal welfare. Baseline data development for the understanding of the health and welfare problems of donkeys in Ethiopia is being pursued by the project staff and through the supervision of DVM thesis involving final year students.
This paper proposes an approach to use pain-relevant pathologies to enhance our understanding of the clinical and behavioural signs of pain in donkeys and outlines initial results of this ongoing investigation.
The methodology is summarized as follows. Trained veterinary clinicians examined live donkeys under two situations: A) before being euthanized due to a terminal illness or reduced quality of life (n= 347 sedentary donkeys in UK; DU) or B) when about to be slaughtered in an abattoir (n=164 working donkeys in Mexico; DM). The animals that represent populations in terms of age (years) for DU and DM respectively, were: <5 = 0.5%, 31.7%; 5-15 = 4%; 67.7%; 16-20 = 6%, 0.6%; >20 = 89.5% and 0%. For sex were: stallions 0.5%, 44%; geldings 52.2%, 8%; females 47.3%, 48% for DU and DM respectively. The body condition was: <2 =18.5%, 44.7%; 2.5 – 3 = 56.5%, 54%; >3.5 = 25%, 1% for DU and DM respectively and the girth was: 115 cm ±SD 9.6, 112cm ±SD 7.3 for DU and DM respectively.
The clinical examination (CE) included the oral mucosa, heart & respiratory rate, rectal temperature, plus an evaluation of 6 demeanours and 47 behaviours/signs that could relate to pain. At this point an overall pain visual analogue score was derived (VAS 0cm = no pain to 10cm = the worst pain). At post-mortem (PM), lesions/pathologies were noted, grouped by system-organ/tissue, ranked (mild, moderate, severe) and classified according to the following potentially painful pathologies: 1) trauma, 2) inflammation, 3) over-distension 4) perforation/ rupture, 5) stripping/ ulceration, 6) adhesions, 7) swelling, 8) exposure of sub-chondral bone. From these observations a second VAS was produced.
Raw correlations from the two populations showed that donkeys given a higher VAS at CE and PM presented with a greater severity of lesions in more systems as well as a higher heart rate at CE than those donkeys with a lower PM VAS. Moderate to severe pain identified CE was often recognized as severe pain at the PM stage. These initial observations show promise, and so further analysis will be done to test the relationships between pain indicators and pain pathologies.
Retrospective analysis of the clinic database between 2003-2005 indicated that tetanus (27 cases), colic (134), rabies (7), strangles (17), hyena bite (429), dystocia (39), hernia (27) and car accidents are the major causes of direct mortality in donkeys of Central Ethiopia. The case fatality rates of donkeys affected by these diseases were: tetanus 22%, colic 27%, dystocia 15%, strangles 12%, hernia 11%, hyena bite 7% and rabies 100%, in clinical intervention sites of the Donkey Sanctuary. In the non-intervention areas the mortality rates are expected to be higher as there would be no treatment intervention. In this paper the two major causes of mortality: tetanus and colic are discussed. In the project in Ethiopia, treatments of advanced cases of tetanus following standard procedures have been successful. Of 27 cases treated 21 have been cured. The treatment has, however, been expensive, costing the project an average of 1400 Eth Birr per donkey (mature donkey costs 300-400 Eth Birr) and requiring a long period of hospitalisation. In an ideal situation, prophylactic immunisation is the best option. Economic realities however prohibit such an option from being feasible in countries like Ethiopia. An extension system and strategy that focus on the prevention of wounds using proper harnesses/saddles and institution of a sound hoof care programme is the option in mind. Colic characteristics are: gastric impaction 8%, obstruction of small intestines 7%, colonic impactions 19%, flatulent colic 23%, enterolithiasis/foreign bodies 24%, throboembolism due to strongylosis and undiagnosed cases 19%. Free access to mouldy or coarse feed, ingestion of polythene bags, fertiliser sacks and used fabrics of nylon clothes were the major causes. Common sites of lodgement for enteroliths were the proximal portion of the transverse colon, transverse and small colons. Enterolith/foreign body was often diagnosed in pregnant donkeys with colic. Few cases were relieved by surgery. A considerable number of the enteroliths were removed by manual traction. Veterinary intervention alone will do little to alleviate such problems. The use of school children to collect polyethylene bags and fertiliser sacks, improve farmers' awareness of the risk being posed by these materials and that of braided nylon ropes. Encouraging farmers to prevent donkeys from grazing at waste disposal sites is also another preventive measure that can be practised against colic. Extension and education programmes for owners should pay particular attention to these issues.