OBJECTIVE: To clinically evaluate previously developed pain scales [Donkey Chronic Pain Composite Pain Scale (DCP-CPS), Donkey Chronic Pain Facial Assessment of Pain (DCP-FAP) and combined Donkey Chronic Pain Scale (DCPS)], including behavioural and facial expression-based variables, for the assessment of chronic pain in donkeys.
METHODS: Prospective, blinded clinical study.
METHODS: A group of 77 donkeys (34 patients and 43 healthy control animals).
METHODS: Animals were assessed by two observers that were blinded to the condition of the animals.
RESULTS: Both DCP-CPS and DCP-FAP, and resulting combined DCPS scores, showed good interobserver reliability [intraclass correlation coefficient (ICC) = 0.91, 95% confidence interval (CI) = 0.86-0.95, p < 0.001; ICC = 0.71, CI = 0.50-0.83, p < 0.001 and ICC = 0.84, CI = 0.72-0.91, p < 0.001, respectively]. All scores (DCP-CPS, DCP-FAP and the resulting combined DCPS) were significantly higher for patients than for controls at all time points (p < 0.001 for all three scales). Sensitivity and specificity for identification of pain (cut-off value >3) was 73.0% and 65.1% for DCP-CPS, and 60.9% and 83.3% for DCP-FAP, respectively. For the combined DCPS, sensitivity was 87.0% and specificity 90.9% (cut-off value >6).
CONCLUSIONS: Based on behavioural and facial expression-based variables, DCPS proved a promising and reproducible tool to assess different types of chronic pain in donkeys. The combination of behavioural and facial expression-based variables showed the best discriminatory characteristics in the current study. Further studies are needed for refinement of these tools.

The reproductive performance of horse sperm and donkey sperm has been reported to differ. Sperm proteins play a crucial role in sperm viability and fertility. Although differences between species are known, no prior study has investigated disparities in the sperm proteome between horses and donkeys. Therefore, this study characterized and compared the sperm proteomes of horses and donkeys using 4D-DIA mass spectrometry technology. We identified 3436 proteins in horse sperm and 3404 proteins in donkey sperm. Of these, 3363 proteins were expressed in both horse and donkey sperm, with 73 proteins being specifically expressed in horse sperm, and 41 in donkey sperm. According to data analysis, donkeys exhibited a greater percentage of motility and progressive movement in straight-line sperm than horses, as well as lower percentages of static and slow sperm than horses. Joint analysis of the results from the horse and donkey sperm proteomes and their CEROS II-read parameters demonstrated a possible association between sperm proteins and their sperm viability patterns. These findings suggest that there are discrepancies in the expression levels and protein compositions of horse and donkey sperm and that certain specific proteins may be responsible for the differences in performance between these two species.

Previous hematologic and serum biochemistry reference interval (RI) values have been established for donkeys in various geographic regions, life-stages, or for specific donkey breeds. The last extensive investigation establishing RIs for adult donkeys in the United States (U.S.) was published over three decades ago. We aimed to establish updated robust RIs using a reference population of apparently healthy adult donkeys from across the U.S. Standard sized (n = 102), miniature (n = 17), and mammoth (n = 1) donkeys from four different states were enrolled, with 20% of the study population including donkeys captured directly from the wild in Death Valley National Park, CA. RIs were established in accordance with the American Society for Veterinary Clinical Pathology and Clinical Laboratory Standards Institute guidelines. The findings will assist practitioners with the interpretation of their complete blood count and biochemistry panel results in U.S. donkeys. This study also highlights a comparison of results for some important analytes in U.S. donkeys compared to U.S. horses and previously established donkey RIs.

The donkey has always been a notable working animal and its importance as a companion animal has been growing over the last few years. However, there are only a few studies about cardiology in this species. Therefore, this study aimed to evaluate the influence of age, sex, training, and bodyweight on cardiac dimension in mixed-breed donkeys. Forty-five clinically and para-clinically healthy mixed-breed donkeys were included, and B-mode and M-mode echocardiographic measurements were recorded. Multivariate linear regression analysis was performed to identify the effect of age, sex, bodyweight, and training on cardiac measurements. Most of the echocardiographic measurements showed a strong statistically significant correlation with bodyweight. Linear regression analysis between echocardiographic measurements and bodyweight was performed to obtain the values of the intercept and slope of the linear equation to calculate the echocardiographic measurements as a function of bodyweight. This is the first study reporting a strong linear correlation between echocardiographic measurements and bodyweight and reporting a correlation between training and echocardiographic parameters in donkeys, suggesting that this variable should be considered when assessing cardiac dimensions in these animals.

An adult jenny (5-years-old, non-pregnant) was presented to the Veterinary Teaching Hospital (VTH) of the University of Sassari, with a recent history of appetite loss, extreme underweight condition and reluctance to move. On physical inspection, emaciation [body condition score, BCS: 3/9], muscular waste [muscular condition score, MCS: 1/5], loose/running faeces [faecal score, FS: 2/8], and a general state of mild dehydration were found. Blood analyses outlined a general undernourishment condition [circulating albumins, ALB: 17.6 g/L (21.6-31.6 g/L)] with underlying systemic inflammatory profile and moderate increase in circulating enzymes to explore liver function [aspartate amino-transferase, AST: 657 u/L (279-430 u/L); alanine amino-transferase ALT: 60 u/L (5-14 u/L); gamma-glutamyl-transferase, γ-GT: 87 IU/L (14-69 IU/L); total bilirubin close to the upper limit, TB: 0.20 mg/dL(0.07-0.21 mg/dL)]and hyperlipaemia [TG: 8.70 mmol/L (0.60-2.87 mmol/L)], following fat depots mobilisation, with total cholesterol closed to the lower limit of the physiological range. Hyper-phosphataemia was linked to haemolytic anaemia [P:1.81 mmol/L (0.77-1.39 mmol/L) and red blood cells, RBC: 4.14 1012/L (4.40-7.10 1012)] aligned with the TB to the upper limit. On ultrasound abdominal imaging, enlarged and hyper-echogenic liver was observed. Based on the clinical evaluation, a condition of hepatic lipidosis was diagnosed, requiring dedicated nutritional treatment to solve the extreme emaciation along with the metabolic disorder in support of medical therapy. A two-step feeding protocol was planned to support treatments aiming at immediate re-hydration (Ringer lactate solution 2 ml/kg/8 h). The nutritional objectives were meant at first to restart the voluntary feed intake. Gradual increasing energy provision through a palatable hay-based diet was planned to cover one fourth of daily metabolizable energy requirement calculated on the expected metabolic weight, adjusted according to the daily intake of feed and clinical condition. At the conclusion of this first 7-day phase, circulating blood parameters were closer to the reference values and the BCS moved from 3 to 4 out of 9. Bowel motility was restored, and faecal score improved (4/8). In the second phase, allowance to pasture and a combination diet with compound mixed feed were designed. Within four weeks of starting the nutritional plan, blood parameters were re-established to reference values. The gradual feed provision calculated in this two-phase approach proved successful in support of the overall clinical improvement observed after four weeks of treatment, in a severely undernourished jenny with compromised liver functions.

This study investigated the description and feasibility of a surgical procedure for enucleation-exenteration of the equine eye via the supraorbital fossa. A preliminary study was performed on both eyes of four cadaveric heads of native breed donkeys (Equus asinus) to describe the surgical anatomy and demonstrate a new supraorbital enucleation surgical approach. For the clinical study, eight donkeys were admitted for unilateral enucleation. All procedures were performed in a lateral recumbent position under the influence of inhalation anesthesia in combination with a retrobulbar nerve block. A semi-circular incision was made in the skin and fascia of the supraorbital fossa to gain access into the orbital cavity, after which the periorbital fat was dissected and removed. Bleeding was controlled by electrocautery and large blood vessels were ligated, then the eyeball was dissected sharply and freed from its bony attachment. The procedure was successfully accomplished in all clinical cases, and no significant complications occurred during or throughout the postoperative follow-up period. The initial results suggest the feasibility and safety of the supraorbital enucleation technique described in this study for equine eye enucleation. This new technique seems promising due to its feasibility, safety, and positive outcomes observed in both cadaveric and clinical studies.

With more than 150 recognized breeds, donkeys assume relevant economic importance, especially in developing countries. Even if the estimated number of heads worldwide is 53M, this species received less attention than other livestock species. Italy has traditionally been considered one of the cradles of European donkey breeding, and despite a considerable loss of biodiversity, today still counts nine autochthonous populations. A total of 220 animals belonging to nine different populations were genotyped using the double-digest restriction site associated DNA (ddRAD) sequencing to investigate the pattern of diversity using a multi-technique approach. A total of 418,602,730 reads were generated and successfully demultiplexed to obtain a medium-density SNP genotypes panel with about 27K markers. The diversity indices showed moderate levels of variability. The genetic distances and relationships, largely agree with the breeding history of the donkey populations under investigation. The results highlighted the separation of populations based on their genetic origin or geographical proximity between breeding areas, showed low to moderate levels of admixture, and indicated a clear genetic difference in some cases. For some breeds, the results also validate the success of proper management conservation plans. Identified runs of homozygosity islands, mapped within genomic regions related to immune response and local adaptation, are consistent with the characteristics of the species known for its rusticity and adaptability. This study is the first exhaustive genome-wide analysis of the diversity of Italian donkey populations. The results emphasized the high informativeness of genome-wide markers retrieved through the ddRAD approach. The findings take on great significance in designing and implementing conservation strategies. Standardized genotype arrays for donkey species would make it possible to combine worldwide datasets to provide further insights into the evolution of the genomic structure and origin of this important genetic resource.
Donkeys assume relevant economic importance in several countries worldwide. However, the genetic structure of these populations is less investigated compared to other species. The aim of this study was to investigate the genetic background of nine different Italian donkey populations. A total of 220 animals were genotyped with about 27K markers extracted by the double-digest restriction site associated DNA sequencing. The consistency of the results across different approaches agreed with the demographic history, the origin, and previous results on the nine donkey populations, suggesting that our conclusions are robust. Moreover, the results of the present study highlighted low to moderate levels of admixture and, for some breeds, confirmed the success of proper management conservation plans.

BACKGROUND: Equine herpesvirus type 1 (EHV-1) is commonly associated with horse abortion. Currently, there are no reported cases of abortion resulting from EHV-1 infection in donkeys.
RESULTS: This was the first survey-based study of Chinese donkeys. The presence of EHV-1 was identified by PCR. This survey was conducted in Chabuchar County, North Xinjiang, China, in 2020. A donkey EHV-1 strain (Chabuchar/2020) was successfully isolated in MDBK cells. Seventy-two of 100 donkey sera were able to neutralize the isolated EHV-1. Moreover, the ORF33 sequence of the donkey-origin EHV-1 Chabuchar/2020 strain showed high levels of similarity in both its nucleotide (99.7‒100%) and amino acid (99.5‒100%) sequences, with those of horse EHV-1 strains. EHV-1 Chabuchar/2020 showed significant consistency and was classified within cluster 1 of horse EHV-1 strains. Further, analysis of the expected ORF30 nucleotide sequence revealed that donkey EHV-1 strains contained guanine at position 2254, resulting in a change to aspartic acid at position 752 of the viral DNA polymerase. Therefore, these strains were classified as horse neuropathogenic strains. Lastly, a phylogenetic tree was constructed using the partial ORF68 nucleotide sequences, showing that the identified donkey EHV-1 strain and the EHV-1 strain found in aborted Yili horses in China comprised a novel independent VIII group.
CONCLUSIONS: This study showed the first isolation and identification of EHV-1 as an etiological agent of abortions in donkeys. Further analysis of the ORF33, ORF30, and ORF68 sequences indicated that the donkey EHV-1 contained the neuropathogenic genotype of strains in the VIII group. It is thus important to be aware of EHV-1 infection in the donkey population, even though the virus has only been identified in donkey abortions in China.

This paper discusses the discoveries of early donkey and the earliest mule remains in Central Anatolia from the site Derekutuğun. This site represents the remains of a village dating back to the Early Bronze Age and Assyrian Trade Colonies period, associated with mining. The archaeofaunal assemblage was studied by the author and his team using classical archaeozoological methods. The dental remains of the Equidae found at Derekutuğun have been re-examined and are described in this article. The dental evidence indicates that donkeys, and possibly the earliest mules ever found in Central Anatolia, were kept at this site. Although the paper is based on the archaeozoological remains, written sources from the period also support the faunal identification. Derekutuğun was a small settlement that specialized in processing copper ore, and which was an important hub for a trade network because of its extensive mining and extraction operations.

BACKGROUND: Metabolic changes in donkey meat during the early postmortem period have not been previously reported.
METHODS: The LC-MS-based metabolomics technique was conducted to understand the metabolic profiles and identify the key metabolites of donkey meat in the first 48 h postmortem.
RESULTS: The pH values showed a decreasing trend followed by an increasing trend. Shear force was the lowest at 4 h and the highest at 24 h (p < 0.05). For the metabolome, some candidate biomarker metabolites were identified, such as adenine, inosine, n-acetylhistidine, citric acid, isocitrate, and malic acid. Predominant metabolic pathways, such as citrate cycle (TCA cycle), alanine, aspartate and glutamate metabolism, and purine metabolism, were affected by aging time. Overabundant n-acetylhistidine was identified in LT, declined at 12 h postmortem aging, and then increased. This may explain the significantly lower pH at 12 h postmortem. Adenine was higher at 4 h postmortem, then declined. Decreased ADP may indicate a fast consumption of ATP and subsequent purine metabolism in donkey meat.
CONCLUSIONS: The results of this study provided new insights into early postmortem aging of donkey meat quality.