Plasma endotoxin concentration in horses: a methods study

Background: Accurate determination of plasma endotoxin concentration is critical for ex vivo and in vitro cellular and molecular studies of endotoxemia in horses. However, reports are conflicting with respect to anticoagulant, handling, and sample preparation.

Objective:

Objective: The purpose of this study was to determine the effect of blood sample fraction and handling time on measurement of endotoxin concentration in horses.

Methods:

Methods: Whole blood, anticoagulated with 3.8% (0.12 M) sodium citrate (9:1), was collected from 5 healthy horses. Whole blood (WB), platelet-rich plasma (PRP), and platelet-poor plasma (PPP) were spiked with endotoxin (2 EU/mL). Endotoxin-spiked WB samples were centrifuged immediately to generate PRP for measurement. Endotoxin concentration was subsequently measured by Limulus amebocyte assay at 0, 15, 30, 45, and 60 minutes. Assays were performed in triplicate and results were analyzed using Student's t -test, with significance set at P < .05.

Results:

Results: Mean endotoxin concentrations in 2 EU/mL-spiked WB were significantly different from those in PPP at all time points tested. Recovery of endotoxin in PRP generated from WB was significantly diminished after just 15 minutes.

Conclusion:

Conclusion: PRP generated from WB is significantly more reliable than PPP in determining endotoxin concentration ex vivo. Measurement of endotoxin in PRP generated from WB was significantly diminished after 15 min, identifying a time frame within which to process blood samples for endotoxin analysis.     

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.     

Mortierella wolfii keratomycosis in a horse

Purpose To describe a case of superficial keratomycosis caused by Mortierella wolfii (M. wolfii) in a horse. Methods A thoroughbred filly was presented with painful right eye of 2 days’ duration. A superficial corneal ulcer was observed ventrally together with multifocal punctuate opacities axially. Samples were collected by swabbing and scraping the ulcerated lesion and submitted for microbiologic and cytologic examination. Results Microscopic evaluation of debrided corneal tissue revealed the presence of nonseptate fungal hyphae, and culture of a corneal swab yielded fungal growth. Medical treatment with topical antifungal, antibiotic and autogenous serum and systemic anti‐inflammatory resolved the problem within 2 weeks. Conclusions Cytologic evaluation of a corneal scraping was useful to make a clinical diagnosis of keratomycosis. Based on the mycological characteristics, the fungus isolated from the corneal lesion was identified as M. wolfii. To the authors’ knowledge, this is the first case report of equine keratomycosis associated with this fungus, although the organism is known to infect various organs of cattle.     

Influence of ketamine or xylazine supplementation on isoflurane anaesthetized horses‐ a controlled clinical trial

ObjectivesTo determine the influence of ketamine or xylazine constant rate infusions on isoflurane requirements, cardiovascular parameters and quality of anaesthesia in horses undergoing elective surgery.Study designProspective, matched paired clinical trial.AnimalsFifty four adult Warmblood horses.MethodsAfter premedication with acepromazine, xylazine and butorphanol, anaesthesia was induced with ketamine-midazolam and maintained with isoflurane alone (I), isoflurane with either 1 mg kg⁻¹ hour⁻¹ ketamine (IK) or same dose of xylazine (IX). End tidal concentration of isoflurane (Fe’Iso) was adjusted by the same anaesthetist in all horses according to a scoring system. Dobutamine was infused to maintain mean arterial pressure (MAP) =70 mmHg. Arterial blood gases, heart rate (HR), respiratory rate, MAP and cardiac output (lithium dilution) were measured. Groups I and IK received xylazine before recovery. Recovery quality was scored.ResultsMean ± SD averaged Fe’Iso (volume%) was significantly lower in IX (0.95 ± 0.07) and IK (0.97 ± 0.08) than in I (1.16 ± 0.13). In group IX, HR was significantly lower and averaged MAP (90 ± 13 mmHg) significantly higher than in groups I (71 ± 7 mmHg) and IK (76 ± 7 mm Hg). Differences in other cardiopulmonary variables did not reach statistical significance. All horses recovered well with best score in group IX.ConclusionsBoth CRIs of xylazine and of ketamine resulted in pronounced reduction of isoflurane requirements and blood pressure support based on routinely monitored parameters. Cardiac output appeared well maintained in all three protocols, but lithium dilution induced errors mean the results are untrustworthy. The work requires repetition with another mode of measurement of cardiac output.Clinical relevanceAll three protocols provided good clinical anaesthesia with clinically acceptable cardiovascular effects.     

马疱疹病毒1型LAMP检测方法的建立

本试验旨在建立一种检测马疱疹病毒1型（EHV-1）的快速、灵敏、特异的环介导等温扩增技术（LAMP）,同时评价该方法的可靠性。根据马鼻肺炎糖蛋白B（gB）基因特异保守序列设计多对LAMP引物,利用LAMP Real Time Turbidimeter LA-320仪监测反应进程,进行引物筛选和反应条件的优化,建立能特异性扩增EHV-1 DNA的LAMP检测方法,并加入SYBR GreenⅠ通过肉眼判断结果。该方法在65 ℃恒温下作用50 min,使EHV-1 DNA获得了高效率的特异性扩增,与其他马易感病毒如马疱疹病毒4型（EHV-4）等无交叉反应;且具有极高的灵敏性,可检测到10^-4稀释的目标病毒,比普通PCR的灵敏度高10倍;反应结束后加入SYBR GreenⅠ肉眼观察的结果与LAMP Real Time Turbidimeter LA-320仪监测结果一致。通过将4份临床样品的LAMP检测结果与已得到验证的PCR结果进行比对,结果显示符合率为100%。本研究建立的LAMP检测方法具有快速、特异、灵敏、简单易操作且设备要求低等特点,具有实地检测EHV-1的前景。     

Pharmacokinetics and pharmacodynamics of intravenous romifidine and propranolol administered alone or in combination for equine sedation

Objective

Propranolol has been suggested for anxiolysis in horses, but its sedation efficacy and side effects, both when administered alone and in combination with α₂-adrenoceptor agonists, remain undetermined. This study aimed to document the pharmacokinetics and pharmacodynamics of propranolol, romifidine and their combination.

Two successful embryo transfers of mini-donkey embryos in Brazilian Northeastern jennies using an alternative method: Case report

Pregnancy rates after embryo transfer (ET) are disappointing in donkey species. This study aims to report two successful ET of mini-donkey embryos using Brazilian Northeastern jennies as recipients. Eighteen embryo flushes were performed 9 days post-ovulation in two non-pregnant mini-donkeys jennies (11 and 7 cycles per jenny). Eleven embryos (61%, 11/18) were collected and transferred to Brazilian Northeastern jennies 4–6 days post-ovulation by conventional (n = 6) or an alternative (n = 5) technique. The alternative method consisted of inserting a Polansky equine vaginal speculum smeared with lubricant in the vagina of the recipient jenny. The arms of the speculum were extended to allow the visualization of the cervix. Then, using an adapted crafted, elongated, toothed tissue grasping forceps, the external cervical os was held, and the cervix was gently pulled backward, aiming to straight the cervical canal. The ET gun was inserted through the vagina and cervix by visual inspection, and the embryo was released into the uterine lumen. All embryos collected were Grade 1 and classified as Expanded Blastocysts. No jennies become pregnant after conventional ET (0/6), whereas two recipient jennies (40%, 2/5) become pregnant and delivered offspring in the following year after ET using the alternative technique. In conclusion, Brazilian Northeastern jennies can be used as embryo recipients using the alternative method proposed in the present study. However, further investigations are needed to improve the knowledge and results of ET in donkey species.     

Demonstration of Heterogeneous Genotypes of Taylorella equigenitalis Isolated from Horses in Six European Countries by Pulsed-Field Gel Electrophoresis

Forty-six isolates of Taylorella equigenitalis were analysed by pulsed-field gel electrophoresis (PFGE) after separate digestion of the genomic DNA with ApaI and with NotI. The isolates had been obtained from horses in six European countries and were classified into 18 genotypes. In Belgium, 2 genotypes were detected in 2 isolates, in England 9 among 15, in Finland 2 in 2, in France 2 among 10, in Sweden 3 among 5, and in Switzerland 3 among 12. Two English isolates and 4 French isolates gave identical PFGE profiles to those of Kentucky 188 from the United States. A common genotype was found in 5 isolates from Belgium and England and also in 10 isolates from France and Switzerland. The analysis of genomic DNA from 12 isolates of T. equigenitalis obtained from male horses in France, Sweden and Switzerland gave no evidence of a sex-related difference in the genomic DNA. Genomic DNA from 11 streptomycin (STM)-susceptible isolates obtained in Sweden and Switzerland were classified into four genotypes by PFGE. Each of the six genotypes determined among the 17 isolates from these two countries had single phenotypes for resistance or susceptibility to STM.