Is sperm cryopreservation in absence of permeable cryoprotectants suitable for subfertile donkeys?

Sperm from fertile donkeys have been successfully frozen in absence of permeable cryoprotectants. The aim of this study was to determine whether this cryopreservation method is suitable for subfertile donkeys in comparison to conventional sperm freezing with glycerol. Ejaculates were collected from four Andalusian Donkeys: three fertile and one subfertile. Semen was frozen with an extender containing glycerol (GLY), or adding instead sucrose 0.25 molar and 1% bovine serum albumin (SUC) as non‐permeable cryoprotectants. After thawing, samples were assessed for total (TM, %) and progressive (PM, %) sperm motility by CASA, plasma membrane integrity (PMI, %) by epifluorescence microscopy and DNA integrity (DFI, %) by SCSA. Results (mean ± SD) were compared between extenders in fertile and subfertile donkeys using the Student's t test. No differences between GLY and SUC treatments were found in the fertile group for the sperm parameters assessed. In subfertile donkey ejaculates, GLY resulted in significantly higher values than SUC for TM (25.5 ± 3.1 vs. 19.6 ± 1.9) and PM (13.3 ± 5.1 vs. 4.0 ± 1.2), respectively. In conclusion, considering all the sperm parameters assessed, sperm freezing in absence of permeable cryoprotectants may not be still an option for cryopreservation of subfertile donkey sperm.     

藏野驴与家驴血清同工酶的比较研究

采用聚丙烯酰胺凝胶电泳对藏野驴与家驴４种血清同工酶进行比较研究。结果发现，除ＬＤＨ同工酶以外，藏野驴与家驴的酯酶、淀粉酶和碱性磷酸酶酶谱都有颇大的差异。     

Seasonal Variations in the Abundance of Gasterophilus spp. Larvae in Donkeys in Northern Jordan

One hundred and five donkeys were examined over a 12-month period for the presence of Gasterophilus spp. larvae in their stomachs. G. intestinalis and G. nasalis were found, but there was a distinct seasonal variation in their abundance. The numbers of second-stage larvae (L₂) peaked during the early winter and early summer, but they were absent or at low levels during the early spring and early autumn. This suggests that there are two periods of adult fly activity (late autumn and late spring) for both G. nasalis and G. intestinalis. Third-stage larvae (L₃) were found throughout the year, but the lowest levels were seen in the late summer and autumn. The parasites also had an aggregated distribution, which may be due to host selection by the fly. The numbers of larvae did not vary significantly with age, which may be due to a lack of acquired protective immunity or to tolerance after repeated exposure.     

Seminal parameters and field fertility of cryopreserved donkey jack semen after insemination of horse mares

Reasons for performing study: As mule production is often concentrated in remote areas of the world, a simplified semen cryopreservation protocol is required. Aim: To compare the seminal parameters of cryopreserved donkey semen in lactose‐EDTA and lactose‐yolk extenders and the fertility rates on horse mares. Methods: Trial 1: Sperm total and progressive motility, vigour (scale 0–5), morphology (major and minor defects) and plasma membrane integrity (HOST) were evaluated in 25 ejaculates from 5 donkey jacks immediately after collection (raw), after chilling to 5°C (chilled) and after freezing/thawing. The semen was mixed with skimmed‐milk extender, centrifuged, and then re‐suspended in lactose‐EDTA or lactose‐yolk extender. Semen was loaded into 0.5 ml straws and chilled to 5°C for 1 h, after which samples were either evaluated (chilled semen) or placed above liquid nitrogen for 20 min prior to immersion. Seminal parameters were evaluated by ANOVA and Tukey's test. Trial 2: Cryopreserved semen from 3 males was used to inseminate 53 mares at 60 oestrous cycles randomly assigned to lactose‐yolk (n = 30 cycles) or lactose‐EDTA (n = 30 cycles) extenders. Pregnancy diagnosis was performed 15 and 25 days post ovulation. The pregnancy rates were compared using Chi‐squared tests. Results: Trial 1: No significant differences were evident in any seminal parameters between extenders after either chilling or cryopreservation. Total and progressive motility were significantly (P<0.05) lower in cryopreserved semen than raw and chilled semen for both extenders. Trial 2: Pregnancy rates did not significantly differ between extenders (lactose‐EDTA extender 53.33 and 43.33%; lactose‐yolk 50.0 and 46.66% for Days 15 and 25 post ovulation, respectively). Conclusions: Cryopreservation of donkey semen using the simplified lactose‐yolk extender resulted in similar seminal parameters and fertility rates when compared to lactose‐EDTA extender. Potential relevance: Lactose‐yolk extender may be advocated as a simple, easy to prepare extender, for use in geographically isolated enterprises producing mules throughout the world.     

Conjunctival fungal flora in healthy donkeys

OBJECTIVE: To identify and quantify ocular fungi from healthy donkeys living in the center of Italy. ANIMALS STUDIED: One hundred and two Amiata donkeys were examined. PROCEDURES: Conjunctival swabs from both eyes were seeded onto Sabouraud dextrose agar (SDA) and malt extract agar (MEA), and incubated at 25 degrees C over a 10-day period. Filamentous fungi identification was achieved to the genus level; yeast colonies were identified for macro-micromorphologic and physiological characteristics. RESULTS: Eighty-one donkeys out of 102 (79.4%) were positive for fungi; 47/102 (46.1%) had positive cultures from both eyes. Most frequently recovered fungal genera were Aspergillus spp., Penicillium spp., Cladosporium spp., Acremonium spp. Different fungal genera and/or species were recovered from the same donkey in 43 cases (42.1%). Yeasts were isolated from five subjects; the yeasts were never associated with molds. The number of colony forming units (CFU) ranged from 1 to 100. CONCLUSIONS: Aspergillus was the most commonly isolated fungal genus (33%). This result agrees with the findings of similar surveys carried out in horses. There was a remarkable presence of fungi and perfect forms. These observations may be explained by the optimal conditions for presence and development of fungi in the conjunctival fornix microenvironment in Amiata donkeys.     

The influence of various confounding factors on mechanical nociceptive thresholds in the donkey

ObjectiveTo evaluate a mechanical nociceptive threshold (MNT) testing device in the donkey, and to investigate the influence of potential confounders on MNTs generated.Study designProspective, randomised.AnimalsSixteen castrated male donkeys aged 4–9 years, weighing 105–170 kg.MethodsMechanical nociceptive thresholds were measured using an actuator with three pins placed on the dorsal aspect of the distal limb, connected to a force meter. The pins (surface area 15 mm²) were extruded onto the limb by pressurising an air-filled syringe, until the MNT force (when foot-lift was observed) or 25 N (cut-off force) was reached. Effect on MNT of presence of a companion donkey, the limb tested, rate of application of force, testing location, level of distraction, ambient temperature and hair cover at the test site was evaluated. Long and short-term repeatability of MNT was assessed. Data were analysed using general linear models and Mann–Whitney U tests, p < 0.05 was considered significant.ResultsIncreasing the rate of force application significantly increased the mean ± SD MNT from 9.2 ± 2.0 N when applied at 0.4 N sec^?1 to 10.6 ± 2.1 N when applied at 1.2 N sec^?1 (p = 0.001). No other factors significantly influenced MNT. Mean MNT remained stable over a 3 week period, however MNTs were significantly (p = 0.006) higher (12.8 ± 3.0 N cf 10.3 ± 1.9 N) after a 12 month interval.Conclusions and clinical relevanceWhen designing studies measuring MNT in donkeys, rate of application of force must be standardised. Donkeys’ MNTs have good short-term stability suggesting this technique is appropriate for short-term analgesiometry studies; however variability of MNTs over the long-term is greater.     

新疆驴和青海驴MSTN基因多态性分析

 为揭示中国家驴 MSTN 基因的遗传多样性,以新疆驴和青海驴为研究对象,根据GenBank上公布的马的 MSTN 序列,设计9对特异性引物,利用驴混合DNA池为模板对该 MSTN 基因部分序列进行PCR扩增和测序分析。结果显示,扩增出的驴 MSTN 基因部分序列长度为3242 bp,包括5'-UTR区(671 bp)、第一外显子(373 bp)、第一内含子(1825 bp)和第二外显子(373 bp)。在该基因中共发现4个SNPs,分别为65 bp处T→Ｃ（5'-UTR）,872 bp处A→G（第一外显子）,2017 bp处G→A (第一内含子),2395处C→G突变（第一内含子）。利用PCR-RFLP技术对新疆驴和青海驴共计80个样本进行基因分型,4个突变位点共检测到6种单倍型（H1-H6）,其中H1是最主要的单倍型。两个家驴品种 MSTN 基因的遗传多样性很丰富（H=0.6044）,这对中国驴的遗传资源保护具有重要作用。     

补饲高蛋白质、高脂肪精饲料对驴产乳量和乳成分的影响

本试验旨在研究补饲高蛋白质、高脂肪精饲料对驴产乳量和乳成分的影响。选择 ３～７岁的健康泌乳母驴 ３４头,设 Ａ（１２头）、Ｂ（１０头）和 Ｃ（１２头）３个区,共 ５个处理。所有驴饲喂基础饲粮。每个区均设对照组,各试验组驴分别补饲粗蛋白质水平为 ３１．４４％（试验 １组,Ａ区,４头）、粗脂肪水平为 ２５．７７％［２０％棉籽油（试验 ２组,Ａ区,４头）、２０％葵花籽油（试验 ３组,Ｂ区,５头）、２０％菜籽油（试验 ４组,Ｃ区,６头）］的试验精饲料 １．５ｋｇ／ｄ。预试期 １４ｄ,正试期１４ｄ。结果表明：１）除含 ２０％葵花籽油的试验精饲料外,其余试验精饲料采食量（＞１３００ｇ／ｄ）均达到预期的采食量,且未出现消化异常,表明所用精饲料适口性较好,驴对它们有较好的适应性;２）补饲含 ２０％棉籽油和含 ２０％菜籽油的试验精饲料均显著提高了驴日均产乳量（Ｐ＜０．０５）,驴日均产乳量可达 １．５ｋｇ／ｄ,而日均总泌乳量可达 ４ｋｇ／ｄ;３）补饲不同试验精饲料对乳成分无显著影响（Ｐ＞０．０５）,驴乳中乳蛋白、乳糖、粗灰分含量较稳定,而乳脂含量变化非常大。总之,补饲高蛋白质、高脂肪的精饲料可提高驴产乳量,但对乳成分的影响不明显。