Bobbi J. Conner

Systematic evaluation of evidence on veterinary viscoelastic testing part 4: Definitions and data reporting.

Objective To systematically examine evidence surrounding definitions and reporting of data for viscoelastic testing in veterinary medicine. Design Standardized, systematic evaluation of the literature, categorization of relevant articles according to level of evidence and quality, and development of consensus on conclusions for application of the concepts to clinical practice. Setting Academic and referral veterinary medical centers. Results Databases searched included Medline, CAB abstracts, and Google Scholar. Conclusions All 4 standard thromboelastography (TEG) and rotational thromboelastometry (ROTEM) variables should be universally reported, and the reporting of shear elastic modulus in addition to maximum amplitude (MA) is encouraged. There is insufficient evidence to support universal usage of the coagulation index at this time. The K value and clot formation time are the most variable of the 4 parameters, with alpha angle, MA, and maximum clot firmness generally the least variable. Individual studies should report sufficient data on patients and institutional controls to enable definitions of hypo- and hypercoagulability to be evaluated post-hoc, and it is recommended that all studies specifically report how these conditions were defined. In reporting data relating to fibrinolysis, the TEG variables LY30, LY60, CL30, CL60, and the ROTEM variables LI30, LI60, ML, LOT, and LT should be documented. Studies should report sufficient data on patients and controls to enable definitions of hyper- and hypofibrinolysis to be evaluated post-hoc, and we suggest that standard TEG/ROTEM assays may be unable to detect hypofibrinolysis in companion animals. We recommend that every center establish reference intervals, which are specific to either TEG or ROTEM. These reference intervals should be established using veterinary clinical pathology guidelines, standardized protocols, and a minimum of 40 healthy animals. There are currently insufficient data in companion animals to suggest a utility for Vcurve variables beyond that of standard TEG variables.OBJECTIVE To systematically examine evidence surrounding definitions and reporting of data for viscoelastic testing in veterinary medicine. DESIGN Standardized, systematic evaluation of the literature, categorization of relevant articles according to level of evidence and quality, and development of consensus on conclusions for application of the concepts to clinical practice. SETTING Academic and referral veterinary medical centers. RESULTS Databases searched included Medline, CAB abstracts, and Google Scholar. CONCLUSIONS All 4 standard thromboelastography (TEG) and rotational thromboelastometry (ROTEM) variables should be universally reported, and the reporting of shear elastic modulus in addition to maximum amplitude (MA) is encouraged. There is insufficient evidence to support universal usage of the coagulation index at this time. The K value and clot formation time are the most variable of the 4 parameters, with alpha angle, MA, and maximum clot firmness generally the least variable. Individual studies should report sufficient data on patients and institutional controls to enable definitions of hypo- and hypercoagulability to be evaluated post-hoc, and it is recommended that all studies specifically report how these conditions were defined. In reporting data relating to fibrinolysis, the TEG variables LY30, LY60, CL30, CL60, and the ROTEM variables LI30, LI60, ML, LOT, and LT should be documented. Studies should report sufficient data on patients and controls to enable definitions of hyper- and hypofibrinolysis to be evaluated post-hoc, and we suggest that standard TEG/ROTEM assays may be unable to detect hypofibrinolysis in companion animals. We recommend that every center establish reference intervals, which are specific to either TEG or ROTEM. These reference intervals should be established using veterinary clinical pathology guidelines, standardized protocols, and a minimum of 40 healthy animals. There are currently insufficient data in companion animals to suggest a utility for Vcurve variables beyond that of standard TEG variables.

Effects of acepromazine maleate on platelet function assessed by use of adenosine diphosphate activated– and arachidonic acid– activated modified thromboelastography in healthy dogs

OBJECTIVE To evaluate the effect of acepromazine maleate administered IV on platelet function assessed in healthy dogs by use of a modified thromboelastography assay. ANIMALS 6 healthy adult mixed-breed dogs. PROCEDURES Dogs received each of 3 treatments (saline [0.9% NaCl] solution [1 to 2 mL, IV] and acepromazine maleate [0.05 and 0.1 mg/kg, IV]) in a randomized crossover study with a minimum 3-day washout period between treatments. From each dog, blood samples were collected via jugular venipuncture immediately before and 30 and 240 minutes after administration of each treatment. A modified thromboelastography assay, consisting of citrated kaolin-activated (baseline assessment), reptilase-ADP-activated (ADP-activated), and reptilase-arachidonic acid (AA)-activated (AA-activated) thromboelastography, was performed for each sample. Platelet inhibition was evaluated by assessing the percentage change in maximum amplitude for ADP-activated or AA-activated samples, compared with baseline values. Percentage change in maximum amplitude was analyzed by use of Skillings-Mack tests with significance accepted at a family-wise error rate of P < 0.05 by use of Bonferroni corrections for multiple comparisons. RESULTS No significant differences were found in the percentage change of maximum amplitude from baseline for ADP-activated or AA-activated samples among treatments at any time. CONCLUSIONS AND CLINICAL RELEVANCE Platelet function in dogs, as assessed by use of a modified thromboelastography assay, was not inhibited by acepromazine at doses of 0.05 or 0.1 mg/kg, IV. This was in contrast to previous reports in which it was suggested that acepromazine may alter platelet function via inhibition of ADP and AA.

Establishment of reference values for various coagulation tests in healthy Florida manatees (Trichechus manatus latirostris) and evaluation of coagulation in debilitated manatees during rehabilitation

OBJECTIVE To establish reference ranges for coagulation parameters in healthy Florida manatees (Trichechus manatus latirostris) and compare results with those for debilitated manatees undergoing treatment at a rehabilitation facility. DESIGN Prospective study. ANIMALS 29 healthy manatees and 45 debilitated manatees with various diseases. PROCEDURES Manatees considered healthy on the basis of results of physical examination, CBC, and serum biochemical analysis underwent coagulation testing including measurement of prothrombin time, partial thromboplastin time, D-dimer concentration, platelet count, and fibrinogen concentration to establish reference ranges. For comparison, a group of manatees undergoing rehabilitation was also tested, and the results were compared. Thromboelastography was also performed on some animals. RESULTS Values for D-dimer concentration were significantly higher in debilitated versus healthy animals. There was no significant difference for prothrombin time, partial thromboplastin time, platelet count, or fibrinogen concentration between groups. Thromboelastography was performed on 8 healthy animals. CONCLUSIONS AND CLINICAL RELEVANCE Reference ranges were established for various tests of coagulation that may assist clinicians during the initial evaluation and rehabilitation of Florida manatees. Future research to evaluate the effect of specific disease processes on the coagulation cascade is recommended.

Coagulation abnormalities in 5 cats with naturally occurring cytauxzoonosis

OBJECTIVE To characterize hemostasis and determine if disseminated intravascular coagulation (DIC) is present in cats with cytauxzoonosis. DESIGN Cross-sectional study. SETTING University teaching hospital. ANIMALS Five client-owned cats with cytologic and PCR-confirmed cytauxzoonosis. INTERVENTIONS None. MEASUREMENTS AND MAIN RESULTS Admission samples were collected for hemostasis testing including platelet count, activated partial thromboplastin time, prothrombin time, fibrinogen, antithrombin (AT), d-dimer, protein C, plasminogen, antiplasmin, factors VII, VIII, IX, X, and XI, von Willebrand factor, and thromboelastography. Results were compiled for combined criteria used to define DIC, and all 5 cats satisfied criteria using a previously described modified scoring system for DIC in cats. The abnormalities found in all 5 cats included thrombocytopenia, low protein C activity, and prolonged prothrombin time; however, none of the cats had low AT activity. None of the cats had clinical signs of hemorrhage despite thrombocytopenia, coagulation factor deficiency (5/5 cats), and thromboelastographic evidence of hypocoagulability (2/5 cats). Three of 5 cats survived to hospital discharge. The nonsurvivors had disseminated cytauxzoonosis with schizont-laden macrophages in vessels of various organs. CONCLUSIONS This is the first report that comprehensively describes the hemostastic status of cats with naturally occurring infection with Cytauxzoon felis. All 5 cats had laboratory evidence of overt DIC. Unlike human and canine models of sepsis-induced DIC, AT deficiency was not found in this series of cats. Further research is warranted to investigate therapeutic strategies targeting thrombotic DIC to improve survival in cats with cytauxzoonosis.Objective To characterize hemostasis and determine if disseminated intravascular coagulation (DIC) is present in cats with cytauxzoonosis. Design Cross-sectional study. Setting University teaching hospital. Animals Five client-owned cats with cytologic and PCR-confirmed cytauxzoonosis. Interventions None. Measurements and Main Results Admission samples were collected for hemostasis testing including platelet count, activated partial thromboplastin time, prothrombin time, fibrinogen, antithrombin (AT), d-dimer, protein C, plasminogen, antiplasmin, factors VII, VIII, IX, X, and XI, von Willebrand factor, and thromboelastography. Results were compiled for combined criteria used to define DIC, and all 5 cats satisfied criteria using a previously described modified scoring system for DIC in cats. The abnormalities found in all 5 cats included thrombocytopenia, low protein C activity, and prolonged prothrombin time; however, none of the cats had low AT activity. None of the cats had clinical signs of hemorrhage despite thrombocytopenia, coagulation factor deficiency (5/5 cats), and thromboelastographic evidence of hypocoagulability (2/5 cats). Three of 5 cats survived to hospital discharge. The nonsurvivors had disseminated cytauxzoonosis with schizont-laden macrophages in vessels of various organs. Conclusions This is the first report that comprehensively describes the hemostastic status of cats with naturally occurring infection with Cytauxzoon felis. All 5 cats had laboratory evidence of overt DIC. Unlike human and canine models of sepsis-induced DIC, AT deficiency was not found in this series of cats. Further research is warranted to investigate therapeutic strategies targeting thrombotic DIC to improve survival in cats with cytauxzoonosis.

Normal Hemostatic Profiles and Coagulation Factors in Healthy Free-Living Florida Manatees (Trichechus manatus latirostris)

Abstract Hemostatic disorders presumptively play an important role in the pathophysiology of several important disease conditions in the Florida manatee (Trichechus manatus latirostris). Prior to pursuing such clinical implications, it is essential to establish normal hemostatic profiles in clinically healthy animals. During annual health assessments of free-living manatees organized by the US Geological Survey, blood samples were collected from 12 healthy animals from the Atlantic coast and 28 from the Gulf of Mexico coast of Florida, with body lengths of 210–324 cm. The following analyses were performed on citrated plasma: prothrombin (PT), partial thromboplastin time (PTT), and concentrations of fibrinogen, D-dimers, and coagulation factors VII, VIII, IX, X, XI, and XII. Compared to other mammalian species, manatees had short PT (9.2±1.5 s) and PTT (10.7±0.5 s), fibrinogen was 369±78.7 mg/dL, antithrombin III was 132±11%, and D-dimer was 142±122 ng/mL. Baseline concentrations for the listed coagulation factors were established. When comparing coagulation factors between locations, Atlantic coast manatees had significantly higher factors VIII, IX, and X than did Gulf Coast manatees. This finding may reflect differences in water salinity, diet, or genetics. There were no differences in coagulation factors when among sexes and sizes. These baselines for hemostatic profiles and coagulation factors in healthy free-living manatees lay the foundation for diagnosis and future research of hemostatic disorders and contribute to understanding their role in the pathophysiology of manatees affected by various diseases.

Effects of dipyrone, meloxicam, or the combination on hemostasis in conscious dogs

OBJECTIVE To compare the effects of dipyrone, meloxicam, and of the combination of these drugs on hemostasis in dogs. DESIGN Prospective, blinded, randomized crossover study. SETTING Research laboratory at a veterinary teaching hospital. ANIMALS Six adult dogs. INTERVENTIONS Animals received 4 intravenous treatments with 15-day washout intervals: control (physiological saline, 0.1 mL/kg), meloxicam (0.2 mg/kg), dipyrone (25 mg/kg), and dipyrone-meloxicam (25 and 0.2 mg/kg, respectively). A jugular catheter was placed for drug injection and for collecting samples for whole blood platelet aggregation (WBPA) and thromboelastometry assays at baseline, 1, 2, 3, 5, and 8 hours after treatment administration. The percent change from baseline of lag time and of the area under the curve (AUC) of impedance changes in response to collagen-induced platelet activation were recorded during WBPA. Thromboelastometry-derived parameters included clotting time, clot formation time, alpha-angle, and maximum clot firmness. The buccal mucosal bleeding time was evaluated by a blinded observer at baseline, 1, 3, and 5 hours after treatment injection. MEASUREMENTS AND MAIN RESULTS No significant changes in WBPA and thromboelastometry were recorded in the control treatment. Dipyrone significantly (P < 0.05) increased the lag time for 2 hours and decreased the AUC for 3 hours after injection. Meloxicam did not alter WBPA. Dipyrone-meloxicam significantly increased lag time for 2 hours and decreased the AUC for 5 hours after treatment injection. Experimental treatments did not differ from the control treatment for thromboelastometry and buccal mucosal bleeding time. CONCLUSIONS While meloxicam does not alter hemostasis by the methods evaluated, dipyrone inhibits platelet aggregation for up to 3 hours. Meloxicam-dipyrone combination causes more prolonged inhibition of platelet function than dipyrone alone. Decreased platelet aggregation induced by dipyrone and dipyrone-meloxicam does not appear to impact the viscoelastic properties of the blood clot nor increase the risk of bleeding in dogs without preexisting hemostatic disorders.Objective To compare the effects of dipyrone, meloxicam, and of the combination of these drugs on hemostasis in dogs. Design Prospective, blinded, randomized crossover study. Setting Research laboratory at a veterinary teaching hospital. Animals Six adult dogs. Interventions Animals received 4 intravenous treatments with 15-day washout intervals: control (physiological saline, 0.1 mL/kg), meloxicam (0.2 mg/kg), dipyrone (25 mg/kg), and dipyrone-meloxicam (25 and 0.2 mg/kg, respectively). A jugular catheter was placed for drug injection and for collecting samples for whole blood platelet aggregation (WBPA) and thromboelastometry assays at baseline, 1, 2, 3, 5, and 8 hours after treatment administration. The percent change from baseline of lag time and of the area under the curve (AUC) of impedance changes in response to collagen-induced platelet activation were recorded during WBPA. Thromboelastometry-derived parameters included clotting time, clot formation time, alpha-angle, and maximum clot firmness. The buccal mucosal bleeding time was evaluated by a blinded observer at baseline, 1, 3, and 5 hours after treatment injection. Measurements and Main Results No significant changes in WBPA and thromboelastometry were recorded in the control treatment. Dipyrone significantly (P < 0.05) increased the lag time for 2 hours and decreased the AUC for 3 hours after injection. Meloxicam did not alter WBPA. Dipyrone-meloxicam significantly increased lag time for 2 hours and decreased the AUC for 5 hours after treatment injection. Experimental treatments did not differ from the control treatment for thromboelastometry and buccal mucosal bleeding time. Conclusions While meloxicam does not alter hemostasis by the methods evaluated, dipyrone inhibits platelet aggregation for up to 3 hours. Meloxicam-dipyrone combination causes more prolonged inhibition of platelet function than dipyrone alone. Decreased platelet aggregation induced by dipyrone and dipyrone-meloxicam does not appear to impact the viscoelastic properties of the blood clot nor increase the risk of bleeding in dogs without preexisting hemostatic disorders.

PROPOSAL AND APPLICATION OF A NOVEL DISSEMINATED INTRAVASCULAR COAGULATION SCORING SYSTEM IN THE FLORIDA MANATEE (TRICHECHUS MANATUS LATIROSTRIS)

Abstract:  Disseminated intravascular coagulopathy (DIC) is an acquired disorder of hemostasis resulting in activation of the coagulation and fibrinolytic pathways. It is reported secondarily to multiple disease processes and can be associated with increased mortality. Previous research at Tampas Lowry Park Zoo (LPZ) demonstrated that Florida manatees (Trichechus manatus latirostris) with cold stress syndrome (CSS) demonstrated thromboembolic disease. The object of this retrospective study was to establish the presence and clinical relevance of DIC in Florida manatees admitted to LPZ for rehabilitation from 07 March 2010 to 15 August 2015. A coagulation panel, including prothrombin time, partial thromboplastin time, platelet count, fibrinogen level, and D-dimer level was used to diagnose DIC. There were 100 cases identified in the study period: 35 trauma, 43 CSS, 17 secondary to harmful algae blooms (HAB), and five miscellaneous. Manatees with CSS had the highest incidence of DIC with 24 of 43 cases (56%) affected, followed by trauma with 18 of 35 cases (52%) affected. None of the manatees with HAB were found to have DIC. Manatees that developed DIC during rehabilitation or when DIC progressed did not survive. Due to the clinical implications of DIC, identifying its presence and recognizing its severity could improve clinical outcomes by enabling more intensive treatment protocols.

Identifying coagulopathies in the pathophysiology of cold stress syndrome in the Florida manatee Trichechus manatus latirostris

Cold stress syndrome (CSS) in the Florida manatee Trichechus manatus latirostris has been defined as morbidity and mortality resulting from prolonged exposure to water temperatures <20°C. The pathophysiology is described as multifactorial, involving nutritional, immunological and metabolic disturbances; however, the exact mechanisms are unknown. We hypothesized that thromboembolic complications contribute to the pathophysiology of CSS in addition to the previously described factors. During the winter of 2014-2015, 10 Florida manatees with clinical signs of CSS were presented to Lowry Park Zoo, Tampa, FL, USA. Thromboelastography (TEG) and coagulation panels were performed at admission. In addition, coagulation panel data from 23 retrospective CSS cases were included in the analyses. There were numerous differences between mean values of TEG and coagulation parameters for healthy manatees and those for CSS cases. Among TEG parameters, reaction time (R), clot formation time (K) and percentage of clot lysed after 30 min (LY30) values were significantly different (p < 0.05) between the 2 groups. CSS cases also had significantly higher mean D-dimer concentration and coagulation factor XI activity, prolonged mean activated partial thromboplastin time (aPTT) and significantly decreased mean antithrombin activity. These combined abnormalities include clinicopathologic criteria of disseminated intravascular coagulation, indicating an increased risk of thromboembolic disease associated with manatee CSS.

COAGULATION ASSESSMENT: UNDERUTILIZED DIAGNOSTIC TOOLS IN ZOO AND AQUATIC ANIMAL MEDICINE

Abstract Veterinarians specializing in nondomestic species are faced with unique challenges regarding research and diagnostic capabilities given the wild and frequently dangerous nature of their patients. Standard diagnostic techniques used in small or large animal practice are not always possible due to anatomical constraints, size, tractability, or the inherent risk of anesthesia in highly valued, rare species. Diagnostic modalities that utilize simple, relatively noninvasive techniques show promise in evaluating nondomestic species and elucidating the pathophysiology behind poorly characterized disease processes in both wild and captive populations. Coagulation profiles, which may include prothrombin time (PT), partial thromboplastin time (PTT), D-dimer concentration, platelet count, and thromboelastography (TEG) are frequently used in domestic species but often overlooked in exotic medicine due to lack of normal reference values and/or availability. Whenever possible, coagulation profiles should be utilized in the evaluation of various disease processes including neoplasia, sepsis, trauma, inflammation, toxin exposure, and envenomation. There are several reports of coagulopathies in both wild and captive species; however, few studies on coagulation profiles have been published on nondomestic species. Clinicians should consider coagulation testing as part of the diagnostic work-up in nondomestic species. A review of available coagulation diagnostic tests is provided here in addition to summarizing the pertinent coagulation disorders currently established in the literature.