Use of dimethylformamide to cryopreserve alpaca semen previously incubated with collagenase

The objective of this study was to evaluate the effect of collagenase and two final dimethylformamide (DMF) concentrations (4% and 7%) on alpaca frozen-thawed sperm quality. A total of 25 ejaculates from 5 alpaca were obtained using electroejaculation. Each individual ejaculate was evaluated and then diluted 4:1 in a solution of 1 mg/ml collagenase in HEPES-TALP medium and incubated for 4 min at 37°C. Subsequently, samples were diluted in TRIS-fructose-citric acid-egg yolk and cooled to 5°C. Then, each sample was divided in two aliquots and DMF at final concentration of 4% or 7% was added, equilibrated for 1 hr at 5°C and frozen over liquid nitrogen vapours. A Kruskal–Wallis test was used to evaluate the sperm morphometry, and Completely Random Block designs were used to analyse sperm motility, viability, membrane function and acrosome status. After collagenase incubation, none of the samples showed thread formation, and sperm parameters were preserved. Non-progressive motile sperm were higher (p < .05) in equilibrated samples (4% DMF: 31.8 ± 8.3% and 7% DMF: 36.3 ± 11.8%) compared to raw (10.1 ± 4.3%) and frozen-thawed semen (4% DMF: 9.7 ± 1.8% and 7% DMF: 7.5 ± 3.2%). Sperm membrane function, membrane integrity and intact acrosomes were higher (p < .05) in raw semen (40.1 ± 12.2%, 94.6 ± 3.2% and 91.3 ± 8.1%) compared to frozen-thawed samples (4% DMF: 19.8 ± 4.7%, 53.2 ± 2.7%, 65.7 ± 8.7% and 7% DMF: 20.4 ± 4.5%, 54.1 ± 1.4%, 64.6 ± 9.1%). Length of the sperm head was lower in frozen-thawed samples, being statistically different with 4% DMF compared to pre-freezing samples. The ratio between acrosome and head areas was greater (p < .05) in frozen-thawed samples. Incubation of raw alpaca semen with collagenase decreased the thread formation without affecting sperm quality. Frozen of collagenase treated alpaca semen with 4% or 7% DMF did not preserve the sperm parameters in thawed samples.     

Diagnosis and treatment of mastitis in mares

Infection and inflammation of the udder (mastitis) is a common condition affecting all domestic mammals, but it appears to be less prevalent in mares than in dairy cows and dairy goats. The seemingly reduced incidence of mastitis in mares can be partially explained by the smaller size and relatively concealed location of the mare’s udder, coupled with a smaller storage capacity than cows and goats. Mastitis can affect lactating, peripartum, dry mares, mares at dry-off or prepubertal foals. Common clinical signs include swollen mammary tissue, abnormal mammary gland secretion, fever and anorexia; less common signs are hindlimb lameness and a swollen mammary vein. On rare occasions, mastitis pathogens can severely affect the nursing foal and mares may develop fibrotic tissue and consequent agalactia in the side(s) or quarter(s) affected. Based on the clinical presentation, mastitis can be classified as acute or chronic, and clinical or subclinical. Diagnosis is based on the clinical signs aided with aerobic culture and cytological evaluation of the gland secretion. In addition, these ancillary tests can also be used to assess prognosis and duration of treatment. Mares suffering from mastitis may present neutrophilia and hyperfibrinogenaemia. Treatment for mastitis includes antimicrobial therapy (systemic and/or locally), nonsteroidal anti-inflammatory drugs, frequent milking and cold hosing with/without hot-packing applied on the gland. While the frequent monitoring of mares after weaning and reducing food intake should be part of common practices at weaning, cleaning of the udder, control of insect populations and frequent milking of mares with a foal unable to nurse can also aid in preventing mastitis.     

Validation of a multiplex PCR assay to detect Babesia spp. and Anaplasma marginale in cattle in Uruguay in the absence of a gold standard test

Detection of bovine Babesia spp. and Anaplasma marginale is based on the reading of Giemsa-stained blood or organ smears, which can have low sensitivity. Our aim was to improve the detection of bovine Babesia spp. and A. marginale by validating a multiplex PCR (mPCR). We used 466 samples of blood and/or organs of animals with signs and presumptive autopsy findings of babesiosis or anaplasmosis. The primers in our mPCR amplified the rap-1a gene region of Babesia bovis and B. bigemina, and the msp-5 region of A. marginale. We used a Bayesian model with a non-informative priori distribution for the prevalence estimate and informative priori distribution for estimation of sensitivity and specificity. The sensitivity and specificity for smear detection of Babesia spp. were 68.6% and 99.1%, and for A. marginale 85.6% and 98.8%, respectively. Sensitivity and specificity for mPCR detection for Babesia spp. were 94.2% and 97.1%, and for A. marginale 95.2% and 92.7%, respectively. Our mPCR had good accuracy in detecting Babesia spp. and A. marginale, and would be a reliable test for veterinarians to choose the correct treatment for each agent.     

Hypertrophic gastropathy associated with gastric sarcoma in a dog

A 14-y-old spayed female Labrador Retriever was presented with an 8-mo history of chronic vomiting. Abdominal ultrasound and gastrointestinal endoscopy revealed a mass protruding into the gastric lumen, with cytologic features suggestive of sarcoma. A partial gastrectomy was performed; the gastric body and antrum were thickened, with a cerebriform appearance of the mucosal surface. Histologic examination revealed a submucosal neoplastic proliferation of fusiform cells variably arranged in irregular bundles and scattered whorls. Fusiform cells strongly reacted to antibodies against vimentin, S100, and neuron-specific enolase; glial fibrillary acidic protein was moderately and multifocally expressed. Pancytokeratin, KIT, α–smooth muscle actin, and desmin were nonreactive. Histologic and immunohistochemical findings suggested a diagnosis of gastric sarcoma with features referable to a non-GIST (gastrointestinal stromal tumor), non–smooth muscle NIMT (non-angiogenic, non-lymphogenic intestinal mesenchymal tumor). The overlying gastric mucosa was thickened by elongated and dilated gastric glands, predominantly lined by intensely periodic acid-Schiff–stained mucous cells. This altered mucosal architecture was suggestive of Ménétrier-like disease. Although this disease has been hypothesized to predispose to gastric adenocarcinoma in dogs, an association with gastric sarcoma has not been documented previously in the veterinary literature, to our knowledge.     

Free-range system and supplementation of 25-hydroxicholecalciferol increases the performance and serum vitamin levels in mixed-parity sows

Improvements in sow productivity have raised questions regarding dietary vitamin D recommendations. The present study aimed to evaluate the effects of the housing system with access to sunlight exposure and supplementation of 25-hydroxicholecalciferol on performance and serum levels of 25(OH)D₃ in sows during gestation and lactation. Sows were distributed in an experimental design with two housing systems: gestation crates or gestation free-range system with external area for sunlight exposure; and two diets: 0 or 50 μg of 25-hydroxicholecalciferol kg⁻¹. The use of 25-hydroxicholecalciferol tended (P = 0.052) to improve total born and influenced (P = 0.046) on number of born alive. Litter weight at birth was also increased (P = 0.01) by 25-hydroxicholecalciferol supplementation; 25-hydroxicholecalciferol supplementation and housing system (free-range with sunlight exposure) tended to increase weaning weight (P = 0.07) and litter daily gain (P = 0.051) during lactation. Exposure to sunlight and 25-hydroxicholecalciferol supplementation increased 25(OH)D₃ serum levels when compared with control treatment during gestation (136.95 vs. 113.92 ng mL⁻¹; P = 0.035) and lactation (120.29 vs. 88.93 ng mL⁻¹; P = 0.026). In conclusion, the association of 25-hydroxicholecalciferol supplementation with exposure to sunlight during gestation improved significantly 25(OH)D₃ serum levels and consequently performance traits in gestation and lactation.     

Definition and clinical evaluation of a recruiting airway pressure based on the specific lung elastance in anesthetized dogs

ObjectiveTo determine the specific lung elastance (SE_L) in anesthetized dogs and to evaluate the efficacy of a SE_L-based recruiting airway pressure (RP_aw) at improving global and regional lung aeration.Study designRetrospective and prospective clinical study.AnimalsA total of 28 adult dogs were included in the retrospective study and six adult dogs in the prospective study.MethodsRetrospective study: SE_L and SE_L-based RP_aw were determined using previously published data. In mechanically ventilated dogs undergoing thoracic computed tomography (CT), SE_L was calculated as ΔP_L/(V_T/EELV), where ΔP_L is the driving transpulmonary pressure, V_T is the tidal volume and EELV is the end-expiratory lung volume. The ratio of lung to respiratory system elastance (E_L/E_rs) was determined. SE_L and E_L/E_rs were used to calculate the SE_L-based RP_aw. Prospective study: dogs underwent thoracic CT at end-expiration and at end-inspiration using the SE_L-based RP_aw, and global and regional aeration was determined. For analysis of regional aeration, lungs were divided into cranial, intermediate and caudal regions. Regional compliance was also calculated. A p value <0.05 was considered significant.ResultsThe SE_L and E_L/E_rs were 12.7 ± 3.1 cmH₂O and 0.54 ± 0.07, respectively. The SE_L-based RP_aw was 29.1 ± 7.6 cmH₂O. In the prospective study, the RP_aw was 28.2 ± 1.3 cmH₂O. During RP_aw, hyperinflation increased (p = 0.0003) whereas poorly aerated (p < 0.0001) and nonaerated (p = 0.01) tissue decreased. Normally aerated tissue did not change (p = 0.265). Regional compliance was higher in the intermediate (p = 0.0003) and caudal (p = 0.034) regions compared with the cranial region. Aeration did not differ between regions (p > 0.05).Conclusions and clinical relevanceAn SE_L-based RP_aw reduces poorly and nonaerated lung tissue in anesthetized dogs. In nonsurgical anesthetized dogs, an RP_aw near 30 cmH₂O is effective at improving lung aeration.     

Plasma histamine concentrations in horses administered sodium penicillin,guaifenesin–xylazine–ketamine and isoflurane with morphine or butorphanol

ObjectiveVarious drugs administered to horses undergoing surgical procedures can release histamine. Histamine concentrations were evaluated in horses prepared for surgery and administered butorphanol or morphine intraoperative infusions.Study designProspective studies with one randomized.AnimalsA total of 44 client-owned horses.MethodsIn one study, anesthesia was induced with xylazine followed by ketamine–diazepam. Anesthesia was maintained with guaifenesin–xylazine–ketamine (GXK) during surgical preparation. For surgery, isoflurane was administered with intravenous (IV) morphine (group M: 0.15 mg kg^–1 and 0.1 mg kg^–1 hour^–1; 15 horses) or butorphanol (group B: 0.05 mg kg^–1 and 0.01 mg kg^–1 hour^–1; 15 horses). Histamine and morphine concentrations were measured using enzyme-linked immunoassay before opioid injection (time 0), and after 1, 2, 5, 30, 60 and 90 minutes. In a subsequent study, plasma histamine concentrations were measured in 14 horses before drug administration (baseline), 15 minutes after IV sodium penicillin and 15 minutes after starting GXK IV infusion. Statistical comparison was performed using anova for repeated measures. Pearson correlation compared morphine and histamine concentrations. Data are presented as mean ± standard deviation. Significance was assumed when p ≤ 0.05.ResultsWith histamine, differences occurred between baseline (3.2 ± 2.4 ng mL^–1) and GXK (5.2 ± 7.1 ng mL^–1) and between baseline and time 0 in group B (11.9 ± 13.4 ng mL^–1) and group M (11.1 ± 12.4 ng mL^–1). No differences occurred between baseline and after penicillin or between groups M and B. Morphine concentrations were higher at 1 minute following injection (8.1 ± 5.1 ng mL^–1) than at 30 minutes (4.9 ± 3.1 ng mL^–1) and 60 minutes (4.0 ± 2.5 ng mL^–1). Histamine correlated with morphine at 2, 30 and 60 minutes.Conclusions and clinical relevanceGXK increased histamine concentration, but concentrations were similar with morphine and butorphanol.     

芩术安胎散的毒性和临床安全性试验研究

试验通过对芩术安胎散的急性毒性、亚慢性毒性和临床安全性进行研究,为芩术安胎散对家猫先兆性流产的预防与治疗提供参考。急性毒性试验：制备芩术安胎散药液,取6周龄健康昆明小白鼠60只,随机分为5组,每组12只,第1~4组的给药剂量分别为6 000、4 800、3 840和3 072 mg/kg,对照组给予等量纯净水,10 d内观察有无中毒和死亡,计算半数致死量（LD₅₀）;另取6周龄健康昆明小白鼠20只,随机分为2组：试验组给予2.0 g/mL芩术安胎散药液,18 h内灌服3次,每次0.8 mL,对照组给予等量纯净水,给药后饲养7 d,计算最大耐受量（MTD）。亚慢性毒性试验：24只7周龄SD雌性大鼠随机均分为高、中、低剂量组和对照组,在30 d内,每日分别给予4 800、2 400和1 200 mg/kg芩术安胎散,对照组以等量纯净水进行灌胃,每日观察和记录各组大鼠的精神状态、有无中毒症状和死亡;第31天对各组大鼠称重、采血,进行血液学检测,剖检各组大鼠,观察主要脏器有无病变并制作病理切片。临床安全性试验：选取2~5岁健康雌性家猫20只,适应性饲养10 d,随机分组,每组5只,分别为低剂量组（1倍临床推荐剂量：1.15 g/kg）、中剂量组（3倍临床推荐剂量：3.45 g/kg）、高剂量组（5倍临床推荐剂量：5.75 g/kg）及空白对照组,将药物置于胶囊内,口服给药,空白对照组给予空胶囊,每日1次,连续给药7 d,每日观察各组家猫食欲、精神状态及排便情况,于第8天对各组家猫进行静脉采血,检测血常规和血液生化指标。结果显示,急性毒性试验无小鼠死亡,LD₅₀>6 000 mg/kg;小鼠对芩术安胎散的最大耐受量为240 g/kg,表明该受试药物无明显毒性。在亚慢性毒性试验中,各组大鼠的生长发育情况、血常规指标、脏器系数与对照组相比均无显著差异（P>0.05）;高、中剂量组与低剂量组、对照组相比血清总胆固醇含量显著下调（P<0.05）,除此之外的生化指标均无显著差异（P>0.05）。病理剖检和组织切片观察结果显示,高剂量组大鼠主要组织器官与对照组相比无明显异常。在临床安全性试验中,不同剂量组家猫精神状态、被毛光泽度、粪便情况均正常,血液学指标与对照组相比差异均不显著（P>0.05）。本试验结果表明,中药芩术安胎散无明显毒性,家猫按临床推荐剂量使用是安全的。