DEVELOPING A THROMBOELASTOGRAPHY ASSAY IN ELASMOBRANCHS.

Thromboelastography (TEG) is a hemostatic assay evaluating clot initiation time, kinetics, strength, and extent of fibrinolysis. Hemostatic assays in nonmammalian species have been less extensively studied because of lack of taxon-specific reagents and unique physiology. Hemostatic or hemorrhagic disease has been described postmortem in elasmobranchs, but antemortem detection of coagulopathies is limited in this taxon. The study aimed to establish an elasmobranch TEG protocol to improve hemostatic evaluation and facilitate advanced treatment options for animals under human care. Multiple clotting initiators were assessed for efficacy with frozen-thawed citrated plasma, fresh citrated plasma, and fresh whole citrated blood: RapidTEGTM, citrated kaolin, Reptilase®, and species brain-derived thromboplastin prepared by two different methods. Initial evaluation found plasma samples clot inconsistently, but TEG analyses using fresh whole blood consistently led to measurable TEG reactions using multiple clotting initiators. The most reliable elasmobranch TEG results were observed using citrated fresh whole blood and the RapidTEG clot initiation reagent.

INVESTIGATION OF A POINT-OF-CARE VISCOELASTIC COAGULATION MONITOR AND ITS COMPARISON TO THROMBOELASTOGRAPHY IN CLINICALLY HEALTHY AFRICAN ELEPHANTS (LOXODONTA AFRICANA).

Elephant endotheliotropic herpesvirus (EEHV) can induce fatal hemorrhagic disease (HD) in African elephants (Loxodonta africana). Once clinical signs develop, progression is rapid, even with aggressive treatment. There is a critical need to develop point-of-care diagnostic tests to aid in identification of EEHV-HD prior to the onset of overt clinical signs. Study objectives were to investigate a novel, point-of-care viscoelastic coagulation monitor (VCM Vet), compare the results to thromboelastography (TEG), and report traditional hemostatic analytes in adult African elephants. Whole blood was collected from seven clinically healthy elephants (four females and three males, 18-47 yr) and analyzed in duplicate via VCM Vet and kaolin-activated TEG 1-3 and 30 min following collection, respectively. Separated plasma was frozen for ancillary coagulation testing. Both analyses generated quantifiable clotting reactions with variables (median [range]) describing clot formation rate (VCM Vet, clot time = 682 s [530-987 s], clot formation time = 244 s [186-744 s], Alpha = 40° [14-47°]; TEG, reaction time = 6.2 min [3.7-11.8 min], kinetic time = 1.3 min [0.9-2.6 min], Alpha = 70° [57-77°]), clot strength (VCM Vet, maximum clot formation = 34 units [20-45 units]; TEG, maximum amplitude = 75 mm [69-80 mm], shear elastic modulus strength = 14.7 Kdynes/s [11.3-19.5 Kdynes/s]), and clot lysis (VCM Vet, lysis index at 30 min = 100% [100-99%], lysis index at 45 min = 98% [95-100%]; TEG, lysis index at 30 min = 0% [0-0.4%], lysis index at 60 min = 1.4% [0-2.6%]) recorded. Additional testing (median [range]) included D-dimer concentration (33 ng/ml [28-94 ng/ml]), prothrombin time (12.4 s [12.2-13.2 s]), activated partial thromboplastin time (17.2 s [14.2-18.8 s]), and fibrinogen concentration (297 [282-383] mg/dL). Tracings generated by VCM Vet and TEG were clinically similar, and there was visual agreement and minimal difference between quantitative variables for duplicate tests. VCM Vet is a promising, user-friendly tool for use in identification and management of coagulopathies in African elephants.

Dose-dependent effect of canine lyophilized platelet on an in vitro hemodilution model.

The transfusion of stored platelets has emerged as an efficient method for treating dogs with thrombocytopenia. However, the availability of fresh platelets is limited in veterinary medicine due to demanding storage conditions. Lyophilized platelets have long shelf lives and can be easily stored, increasing their accessibility for thrombocytopenic dogs. Due to the lack of research and information on the dose effect, canine lyophilized platelets are used at a clinical dosage without research-based evidence. This study was to evaluate the dose effect of lyophilized canine platelets on blood coagulability. Three different concentrations of lyophilized canine platelets were added to in vitro hemodilution blood model, increasing the platelet count by 25, 50, and 100 × 106/ml and coagulation profiles were analyzed. The coagulability was evaluated via the plasma fibrinogen concentration, coagulation time, thromboelastography (TEG), and platelet function analyzer (PFA). Higher concentrations of lyophilized platelets showed dose-dependent association with decreased aPTT and R-time of TEG and increased alpha angle and MA of TEG. These results showed the potential that the higher dose of canine lyophilized platelets better improve blood coagulability than the standard dose and provided the basis for further safety and clinical studies.

Point-of-care viscoelastic coagulation assessment in healthy dogs during the perianesthetic period.

BACKGROUND: The viscoelastic coagulation monitor (VCM Vet) is a novel, portable device that provides a global assessment of hemostasis. The study aims were to evaluate serial viscoelastic analysis during the perianesthetic period in healthy dogs and to compare the agreement between two VCM Vet devices. Twenty healthy dogs undergoing orthopedic surgery were enrolled. Whole blood samples were collected from an intravenous catheter at four time points: baseline, 15 min after premedication, 60 min after inhalant initiation, and 60 min after inhalant termination. Viscoelastic tests were performed in duplicate on different devices, providing: clot time (CT; seconds), clot formation time (CFT; seconds), alpha angle (α; degrees), amplitude (units) at 10 (A10) and 20 (A20) minutes post clot time, maximum clot firmness (MCF; units), and lysis index (%) at 30 (Li30) and 45 (Li45) minutes post maximum clot formation. RESULTS: One hundred sixty samples were analyzed. The speed of CT and CFT significantly decreased an average of 25.5 s (95% confidence interval [CI]15.9-35.0) and 6.9 s (95% CI 3.1-10.7) per time point, respectively. There were no significant changes in clot strength or lysis variables. The Bland-Altman style plot shows an acceptable rate of agreement for all variables with intra-class correlation ranging from 0.64-0.94. CONCLUSION: The rate of clot formation (CT and CFT) decreased over the perianesthetic period in healthy dogs undergoing surgery. These changes were small and occurred without changes in clot strength or fibrinolysis rate, thus were not clinically relevant. There was clinically acceptable consistency between devices.

WHOLE BLOOD THROMBOELASTOGRAPHY IN HEALTHY ADULT CAMELIDS (VICUGNA PACOS AND CAMELUS DROMEDARIUS).

Thromboelastography (TEG) provides a global assessment of hemostasis and fibrinolysis and has broad applications to identify and monitor coagulation dysfunction in veterinary patients. Although alpacas are susceptible to a wide variety of coagulopathies, the assessment of TEG has not been reported in clinically healthy alpacas to date. The purpose of this study was to evaluate the analytical performance of recombinant human tissue factor (rhTF)- and kaolin-activated TEG and to establish reference intervals for TEG parameters (reaction [R] and clotting [K] times, angle [α], maximum amplitude [MA], and shear elastic modulus [G]) in healthy, adult alpacas. Kaolin and rhTF-activated TEG were performed using citrated whole blood samples from 20 clinically healthy, nonpregnant, adult Huacaya alpacas each after 30 min of sample storage at room temperature. Six individuals of a related species, dromedary camels, were also sampled for comparative purposes. All data were presented descriptively, assessed for normality, and compared using either independent-sample t tests or Mann-Whitney U tests, with P ≤ 0.05 considered significant. Reference intervals were calculated using a robust method and Box-Cox-transformed data. Mean TEG values (reference intervals) were determined for rhTF-activated TEG as follows: R 6.99 min (3.41-12.71), K 3.43 min (1.61-6.42), α 48.51° (27.21-67.38), MA 52.05 mm (21.53-65.92), and G 5.71 kdyn/cm2 (1.87-9.60), while mean values (reference intervals) for kaolin-activated TEG included R 7.72 min (4.48-11.43), K 4.24 min (2.03-9.20), α 45.06° (23.66-64.20), MA 52.18 mm (33.49-66.63), and G 5.78 kdyn/cm2 (NR-9.66). None of the measured TEG values differed significantly between activators, suggesting that activator choice may have a limited effect on TEG parameters in healthy alpacas. TEG results in alpacas were comparable to those of dromedary camels. These results will thus provide a useful starting point in the evaluation of hemostasis in adult camelids.

A pilot study evaluating the Calibrated Automated Thrombogram assay and application of plasma-thromboelastography for detection of hemostatic aberrations in horses with gastrointestinal disease.

BACKGROUND: Critically ill horses, such as horses with gastrointestinal (GI) disease, often suffer from hemostatic aberrations. Global hemostatic tests examining the initiation of coagulation, clot strength and fibrinolysis, such as the Calibrated Automated Thrombogram (CAT) and plasma-thromboelastography (TEG) have not been evaluated in horses. This study aimed to evaluate CAT and apply plasma-TEG in horses. Test performance of CAT was evaluated on equine platelet poor plasma with intra- and inter-assay variability (CV) and a heparin dilution curve. To examine clinical performance of both tests, group comparisons were assessed comparing healthy horses, horses with mild and severe GI disease with both CAT and plasma-TEG. RESULTS: For CAT, intra- and inter-assay CVs were established for lag-time (1.7, 4.7%), endogenous thrombin potential (1.6, 4.6%), peak (2.6, 3.9%) and time to peak (ttPeak) (1.9, 3.4%). Increasing heparin concentrations led to the expected decrease in thrombin generation. In the group comparison analysis, CAT showed significant higher peak (p = 0.04) and ttPeak (p = 0.008) in the severe GI disease group compared to horses with mild GI disease and healthy horses, respectively. Plasma-TEG showed an increased angle (p = 0.032), maximum amplitude (p = 0.017) and shear elastic force (G) (p = 0.017) in the severe GI disease group compared to healthy horses. CONCLUSIONS: CAT performed well in horses. Both CAT and plasma-TEG identified hemostatic aberrations in horses with severe GI disease compared to healthy horses. Further studies including more horses, are needed to fully appreciate the use of CAT and plasma-TEG in this species.

BASELINE PLASMA THROMBOELASTOGRAPHY IN KEMP'S RIDLEY (LEPIDOCHELYS KEMPII), GREEN (CHELONIA MYDAS) AND LOGGERHEAD (CARETTA CARETTA) SEA TURTLES AND ITS USE TO DIAGNOSE COAGULOPATHIES IN COLD-STUNNED KEMP'S RIDLEY AND GREEN SEA TURTLES.

Cold-stunning in sea turtles is a frequent natural cause of mortality and is defined as a hypothermic state due to exposure to water temperatures <12°C. Derangements of biochemistry and hematology data by cold stunning have been well documented, although the effects on coagulation have not yet been investigated. The objectives of this study were to characterize the hemostatic state of non-cold-stunned sea turtles and to compare cold-stunned sea turtles at admission and after successful rehabilitation via a sea turtle-specific thromboelastography (TEG) protocol. TEG enables evaluation of the entire coagulation process, and the methodology has recently been established in sea turtles. Initially, 30 wild and apparently healthy sea turtles were sampled as controls: loggerhead sea turtles (Caretta caretta), n =17; Kemp's ridley sea turtles (Lepidochelys kempii), n = 8; and green turtles (Chelonia mydas), n = 5. In addition, paired TEG samples were performed on 32 Ch. mydas and 14 L. kempii at admission and prerelease after successful rehabilitation from cold stunning. Statistically significant differences in reaction time, kinetics, angle, and maximum amplitude parameters in L. kempii and Ch. mydas species demonstrated that the time taken for blood clot formation was prolonged and the strength of the clot formed was reduced by cold stunning. These findings indicate that cold stunning may cause disorders in hemostasis that can contribute to the severity of the condition. Early diagnosis of coagulopathies in the clinical assessment of a cold-stunned sea turtle may influence the treatment approach and clinical outcome of the case.

Thromboelastometric assessment of hemostasis following hydroxyethyl starch (130/0.4) administration as a constant rate infusion in hypoalbuminemic dogs.

BACKGROUND: The primary aim was to evaluate by means of thromboelastometry (ROTEM) the effects of hydroxyethyl starch (HES) 130/0.4 administered as a constant rate infusion (CRI) on hemostasis in hypoalbuminemic dogs. The second aim was to use ROTEM analysis to detect whether all hypoalbuminemic dogs of our population were hypercoagulable. RESULTS: The study sample was 20 hypoalbuminemic dogs (albumin < 2 g/dl) with normal perfusion parameters and requiring intravenous fluid therapy. In order to support plasma colloid osmotic pressure, in addition to crystalloid, HES 130/0.4 was administered as a constant rate infusion at 1 ml/kg/h (group 1, n = 11) or 2 ml/kg/h for 24 h (group 2, n = 9). Blood samples were collected at baseline (T0) and 24 h postinfusion (T1); coagulation was assessed by standard coagulation profile (prothrombin time, activated partial thromboplastin time, and fibrinogen), and ROTEM analysis (in-TEM®, ex-TEM® and fib- TEM® profile). No statistically significant differences in ROTEM values in group 1 were observed (P > 0.05), whereas in group 2 statistically significant differences (P < 0.05) were found at T1 in the in-TEM® profile [decrease in clot formation time (P = 0.04) and increase in α angle (P = 0.02)] and in the ex-TEM® profile [increase in maximum clot firmness (P = 0.008) and α angle (P = 0.01)]; no changes were identified in the fib-TEM® profile. In both groups, a statistically significant decrease (P = 0.007) in hematocrit was noted, whereas no statistically significant differences in platelet count and standard coagulation profile were found. In group 2, a statistically significant increase in TS values (P = 0.03) was noted at T1. ROTEM tracings indicating a hypercoagulable state were observed in 7/20 dogs at T0 (5/11 in group 1 and 2/9 in the group 2). CONCLUSION: Our findings suggest that HES 130/0.4 administered as CRI does not cause hypocoagulability in hypoalbuminemic dogs. A trend toward hypercoagulability, probably related to the underlying diseases, was observed in group 2 at T1. Although all dogs were hyoalbuminemic, only 7/20 were hypercoagulable at T0, confirming the lack of correlation between albumin level and prothrombotic state.

Hyperfibrinolysis diagnosed with rotational thromboelastometry and treated with tranexamic acid in a dog with acute traumatic coagulopathy.

INTRODUCTION: A 4-year old male Australian Cattle Dog involved in a road traffic accident was presented with severe polytrauma to the Small Animal Clinic, University of Zurich. He was presented in hemorrhagic shock, with an initial lactate of 10.3mmol/l and ongoing bleeding from multiple injury sites. Acute traumatic coagulopathy diagnosed with ROTEM within one hour after accident showed marked hypocoagulation and hyperfibrinolysis. Treatment with a total dose of 40mg/kg of tranexamic acid intravenously resulted in successful elimination of hyperfibrinolysis in the following, serially measured ROTEM tracings.

THROMBOELASTOGRAPHY IN THE HEALTHY ASIAN ELEPHANT ( ELEPHAS MAXIMUS): REFERENCE INTERVALS AND EFFECTS OF STORAGE.

Hemorrhagic disease associated with elephant endotheliotropic herpesvirus infection is the most-frequent cause of mortality in captive Asian elephants ( Elephas maximus). Survival relies on intensive monitoring of hemostatic status. Thromboelastography (TEG) utilizes whole blood samples containing all the blood components of hemostasis and is therefore a sensitive indicator of the clinical status in the patient. This study was performed to assess the practicability of TEG in Asian elephants in a zoo environment. Citrated stabilized whole blood samples were obtained from 44 healthy Asian elephants. Kaolin-activated TEG was performed on whole blood at 60 min and 24 hr postsampling (to replicate shipment to an external laboratory) as well as on freeze-thawed plasma samples, 12-14 mo postsampling. Reference intervals were calculated for fresh whole blood and freeze-thawed plasma samples. In the 24-hr analysis, storage artifacts, likely due to cellular degeneration, resulted in a hypercoagulable thromboelastogram and thus reduced sensitivity for detecting coagulopathies. Therefore, delayed analysis of whole blood samples is not recommended.

Preliminary reference intervals and the impact of citrate storage time for thrombelastography in cats including delta and the velocity curve.

BACKGROUND: Thrombelastography is a useful tool in assessment of hemostasis. Beside the traditional variables, the velocity curve and the variable delta have lately earned attention. The velocity curve provides knowledge about the speed of clot formation including information about thrombin generation. Delta, which only reflects enzymatic coagulation, allows the determination of the origin of hypercoagulability when compared to clot rigidity, a variable that reflects both platelet and enzymatic activity. The aim was to establish preliminary reference intervals for feline thrombelastography including the velocity curve variables and delta obtained after 60 min of storage including the assessment of coefficients of variation. Furthermore, the effect of citrate storage time (30 versus 60 min) on feline thrombelastography will be determined. RESULTS: Prolonged storage times significantly reduced reaction (R) (P = 0.019) and clotting (K) (P = 0.008) times, split point (SP) (P = 0.019) and time to maximum rate of thrombus generation (TMRTG) (P = 0.023) values whereas maximum rate of thrombus generation (MRTG) significantly increased (P = 0.040). Preliminary reference intervals: R (min): 2.7-18.1; K (min): 0.8-3.9; alpha (°): 27.6-75.2; maximum amplitude (mm): 18.5-62.5; clot rigidity (dyn/cm2): 1.2-8.2; coagulation index: -4.6 - 2.6; SP (min): 2.4-15.4; delta (min): 0.3-3.1; thrombus generation (mm/min): 255.3-751.2; MRTG (mm/min): 4.0-19.3; TMRTG (min): 3.5-22.0; maximum rate of lysis (mm/min): 0.0-4.7 and time to maximum rate of lysis (min): 0.4-55.8. CONCLUSION: Storage for 60 versus 30 min induces hypercoagulable tracings including the velocity curve, some of which variables (MRTG, TMRTG) might function as sensitive markers for changes in the coagulation activity. Because of the impact of citrate storage time on thrombelastography, reference intervals have to be established using a specific and constant storage time in each laboratory.

Thromboelastographic profile of goat blood after the experimental injury of the femoral artery and use of QuikClot gauze and Celox gauze dressings.

The aim of this study was to evaluate the suitability of thromboelastometry for the analysis of blood test results in goats after the use of hemostatic dressings to control massive bleeding. The study was carried out on 12 goats, 6 animals in each of two subgroups. In all experimental animals incision of the femoral artery was performed, and bleeding was controlled with QuikClot gauze in the first group and Celox gauze in the second group. Dressings were applied for 60 minutes. Blood samples for thromboelastometry were collected from the jugular vein before the incision and 60 min after the application of a dressing. Clotting time (CT), clot formation time (CFT), maximum clot firmness (MCF) and α angle (°) were measured in three standard ROTEM assays (system with generation of reaction curve, numerical parameters and size of the blood clot): intrinsic coagulation pathway (INTEM), extrinsic coagulation pathway (EXTEM) and functional fibrinogen (FIBTEM). Complete hemostasis of the injured femoral artery was found in all goats. No significant differences between pre- and post-incision thromboelastometric parameters were found in any tests in any of the groups, which indicates that the use of dressings was not associated with blood coagulation disorders. This study is the first to describe the use of thromboelastometry in goats for the assessment of clot formation and hemostatic disorders.

In vitro effects of 6 % hydroxyethyl starch 130/0.42 solution on feline whole blood coagulation measured by rotational thromboelastometry.

BACKGROUND: The artificial colloid, hydroxyethyl starch (HES), is recommended for intravascular volume expansion and colloid-osmotic pressure enhancement in dogs and cats. A well-known side effect of HES solutions in humans and dogs is coagulopathy. However, HES-associated coagulopathy has thus far not been investigated in cats. The goal of this study was to assess the in vitro effects of 6 % HES 130/0.42 on feline whole blood samples using rotational thromboelastometry (ROTEM). A further goal was to develop feline reference intervals for ROTEM at our institution. In this in vitro experimental study, blood samples of 24 adult healthy cats were collected by atraumatic jugular phlebotomy following intramuscular sedation. Baseline ROTEM analyses (using ex-tem, in-tem and fib-tem assays) were performed in duplicate. Additionally, ROTEM analyses were performed on blood samples after dilution with either Ringer's acetate (RA) or 6 % HES 130/0.42 (HES) in a 1:6 dilution (i.e. 1 part solution and 6 parts blood). RESULTS: Coefficients of variation of duplicate measures were below 12 % in all ex-tem assays, 3 of 4 in-tem assays but only 1 of 3 fib-tem assays. Reference intervals were similar albeit somewhat narrower than those previously published. Dilution with both solutions lead to significantly prolonged CT (in-tem), CFT (ex-tem and in-tem), and reduced MCF (ex-tem, in-tem, and fib-tem) and alpha (ex-tem and in-tem). Compared to RA, dilution with HES caused a significant prolongation of CT in fib-tem (P = 0.016), CFT in ex-tem (P = 0.017) and in-tem (P = 0.019), as well as a reduction in MCF in in-tem (P = 0.032) and fib-tem (P = 0.020), and alpha in ex-tem (P = 0.014). However, only a single parameter (CFT in ex-tem) was outside of the established reference interval after dilution with HES. CONCLUSIONS: In vitro hemodilution of feline blood with RA and HES causes a small but significant impairment of whole blood coagulation, with HES leading to a significantly greater effect on coagulation than RA. Further studies are necessary to evaluate the in vivo effects and the clinical significance of these findings.

EFFECT OF ε-AMINOCAPROIC ACID ON FIBRINOLYSIS IN PLASMA OF ASIAN ELEPHANTS (ELEPHAS MAXIMUS).

ε-Aminocaproic acid (EACA) is a lysine analogue antifibrinolytic drug used to treat bleeding disorders in humans and domestic animals. Use in zoological medicine is rare and dose recommendations are anecdotal, but EACA may be a valuable therapeutic option for bleeding disorders in exotic species, including Asian elephants ( Elephas maximus ). This study used an in vitro model of hyperfibrinolysis and a thromboelastograph-based assay to estimate the therapeutic plasma concentration of EACA in Asian elephants (61.5 µg/ml, 95% CI = 34.6-88.5 µg/ml). Substantial but incomplete inhibition of lysis was seen at relatively low concentrations of EACA (40 µg/ml). Asian elephants appear sensitive to EACA-mediated inhibition of hyperfibrinolysis. Doses published for domestic animals, targeting higher plasma concentrations, may be inappropriate in this species.

Thromboelastographic Clot Characteristics of Autologous Equine Blood Products After Activation by Autologous Thrombin, Bovine Thrombin, or Calcium Chloride.

OBJECTIVE: To compare clotting efficiency of platelet-rich plasma (PRP) and concentrated platelet-poor plasma (cPPP) to citrated whole blood after activation by autologous thrombin, bovine thrombin, or calcium chloride (CaCl2 ). STUDY DESIGN: Experimental study. ANIMALS: Healthy adult horses (n = 6). METHODS: PRP and cPPP were prepared by commercial devices. Using thromboelastography, clotting variables were compared after activation of citrated autologous blood, PRP, and cPPP by autologous thrombin, bovine thrombin, or CaCl2 , respectively. RESULTS: PRP had the greatest clot strength and quickest clot rate, whereas cPPP had the weakest clot strength, slowest clot rate, and longest clot initiation time. Bovine thrombin resulted in the shortest clot initiation time, quickest clot rate, and was similar to CaCl2 for greatest clot strength. CaCl2 also resulted in the longest clot initiation time and time to reach maximum clot strength. Autologous thrombin resulted in the lowest clot strength. CONCLUSION: When combined with either bovine thrombin or CaCl2 , PRP provided the best combinations for clinical use. Autologous thrombin was suboptimal, but could be an autologous alternative for clinical application. As prepared here, cPPP had inefficient clotting, but may be sufficient for plasma spray indications.

Evaluation of coagulation via thromboelastography in healthy horses administered dexamethasone.

Dexamethasone was administered to healthy horses daily for 7 days. Blood samples were collected at 3 time points from both treatment and non-treatment groups, and analyzed via thromboelastography (TEG). There were no significant differences in TEG parameters between treated and untreated horses, or within treatment groups over time.

Évaluation de la coagulation par une thrombo-élastographie chez des chevaux en santé ayant reçu de de la dexaméthasone. La dexaméthasone a été administrée à des chevaux en santé pendant 7 jours. Des échantillons de sang ont été prélevés à trois moments auprès des groupes de traitement et des groupes sans traitement et ensuite analysés par thrombo-élastographie (TEG). Il n'y avait aucune différence significative dans le temps pour les paramètres TEG entre les chevaux traités et non traités ou à l'intérieur des groupes de traitement.(Traduit par Isabelle Vallières).

Thromboelastography Values in Hispaniolan Amazon Parrots ( Amazona ventralis ): A Pilot Study.

Thromboelastography (TEG) provides a global assessment of coagulation, including the rate of clot initiation, clot kinetics, achievement of maximum clot strength, and fibrinolysis. Thromboelastography (TEG) is used with increasing frequency in the field of veterinary medicine, although its usefulness in avian species has not been adequately explored. The purpose of this preliminary study was to assess the applicability of TEG in psittacine birds. Kaolin-activated TEG was used to analyze citrated whole blood collected routinely from 8 healthy adult Hispaniolan Amazon parrots ( Amazona ventralis ). The minimum and maximum TEG values obtained included time to clot initiation (2.6-15 minutes), clot formation time (4.3-20.8 minutes), α angle (12.7°-47.9°), maximum amplitude of clot strength (26.3-46.2 mm), and percentage of lysis 30 minutes after achievement of maximum amplitude (0%-5.3%). The TEG values demonstrated comparative hypocoagulability relative to published values in canine and feline species. Differences may be explained by either the in vitro temperature at which TEG is standardly performed or the method of activation used in this study. Although TEG may have significant advantages over traditional coagulation tests, including lack of need for species-specific reagents, further evaluation is required in a variety of avian species and while exploring various TEG methodologies before this technology can be recommended for use in clinical cases.

Thromboelastography in Selected Avian Species.

Currently available assay methods and reagents are not optimized for evaluating avian hemostasis; therefore, assessing avian coagulopathies is challenging. Recently, thromboelastography (TEG), which measures the viscoelastic properties of blood, has been used clinically in mammalian species to diagnose and characterize hemostatic disorders. To evaluate TEG in healthy individuals of 6 avian species, we modified existing mammalian TEG protocols to allow analysis of citrated, avian whole-blood samples collected from scarlet ibis (Eudocimus ruber) (n = 13), American flamingos ( Phoenicopterus ruber ) (n = 13), helmeted Guinea fowl ( Numida meleagris ) (n = 12), Amazon parrots (Amazona species) (n = 9), Humboldt penguins ( Spheniscus humboldti ) (n = 6), and domestic chickens (n = 16). Activated partial thromboplastin time, prothrombin time, and fibrinogen were measured as a means of comparison. Regardless of the mode of activation, clot formation in the species studied was markedly delayed compared with mammals. Because of prolonged reaction time (14.7-52.7 minutes) with kaolin and diluted tissue factor, undiluted human tissue factor was used in all avian samples because it provided the shortest reaction time. Species differed significantly in reaction time (P = .007), clotting rate (P < .001), rate of clot formation (α angle; P < .001), and maximum amplitude (P < .001) values, indicating that species-specific reference intervals are necessary. Based on these results, TEG with specific reference intervals could prove useful in evaluating avian hemostatic disorders.

Inflammation-induced haemostatic response in layer chickens infected with Streptococcus equi subsp. zooepidemicus as evaluated by fibrinogen, prothrombin time and thromboelastography.

Streptococcus zooepidemicus has recently been shown to be a severe pathogen in layer chickens, where it is able to cause serious lesions in the vascular system. To evaluate the haemostatic response, 10 layer chickens were inoculated intravenously with S. zooepidemicus. Four hypotheses were tested: that the infection-induced inflammation would increase the plasma fibrinogen (Fbg) concentration, would prolong the prothrombin time (PT) and would prompt hypercoagulability or hypocoagulability as assessed by whole-blood thromboelastography (TEG), and that a possible correlation would exist between one of the TEG values and Fbg/PT. Each parameter was measured at days 1, 3 and 6 post inoculation (p.i.), and compared with the values at day 0 from each individual bird and with values obtained from non-infected control chickens (n = 10). In the infected chickens, the mean (± standard error) of Fbg was higher at day 3 p.i. (9.4 ± 1.4 g/l) and day 6 p.i. (8.0 ± 0.7 g/l) and the PT was prolonged at day 6 p.i. (168.1 ± 21.0 sec) compared with the day 0 standards (2.6 ± 0.2 g/l and 104.6 ± 2.0 sec, respectively) (P < 0.05). The majority of infected chickens demonstrated a hypercoagulable TEG result with increased mean values of the clot formation rate (α-angle) and maximal amplitude (MA) of TEG tracing at day 3 p.i. (83.1 ± 0.7°, 83.8 ± 1.4 mm) and day 6 p.i. (84.0 ± 0.4°, 89.8 ± 1.0 mm) compared with the day 0 values (75.8 ± 2.2° and 66.9 ± 1.4 mm, respectively) (P < 0.05). In control birds, the means of Fbg (1.5 ± 0.1 g/l), PT (79.4 ± 6.4 sec), TEG-α (76.7 ± 1.5°) and TEG-MA (64.0 ± 2.3 mm) were lower at day 6 compared with values observed for the infected chickens (P < 0.05). A negative correlation coefficient (-0.71) was found between the clot formation time (TEG-K) and Fbg at day 1 in the control group (P = 0.02). In conclusion, infection with S. zooepidemicus following intravenous injection in layer chickens induced haemostatic alterations including hyperfibrinogenaemia, prolonged PT, and hypercoagulability as measured by increased TEG-α and TEG-MA.

Thromboelastography in healthy dairy cows.

Thromboelastography is a whole blood-based coagulation assay that can be used to investigate hypocoagulability and hypercoagulability, as seen with thromboembolic diseases and disseminated intravascular coagulation. Numerous coagulopathies due to different causes are reported in cows. The objective was to establish reference intervals for thromboelastography using the TEG 5000 (Haemonetics GmbH, Munich, Germany) with citrated whole blood samples and kaolin activation in dairy cows and to investigate possible thromboelastographic changes between cows in different lactation periods. An additional objective was to test the stability of samples for up to 100h. Sixty blood samples from healthy Holstein-Friesian cows were examined. The samples were allocated to 3 different lactation groups (≤30 d postcalving, 31-99 d postcalving, ≥100 d postcalving). Thromboelastography was performed by using the TEG 5000 analyzer with citrated whole blood samples with kaolin activation. The calculated reference intervals were as follows: reaction time=2.2 to 6.2min, coagulation time=0.8 to 2.0min, angle α=58.2 to 81.8°, maximum amplitude=64.3 to 89.2mm, and clot rigidity=9.2 to 41.2 dyn/cm(2). The 3 different lactation groups showed no significant differences in TEG parameters. No significant difference was seen in samples stored for up to 48h at room temperature, which indicates that delays in processing samples, such as those arising during transit, are not an issue.