Analytical validation of a novel point-of-care immunoassay for canine N-terminal pro-brain natriuretic peptide analysis.

BACKGROUND: N-terminal pro-brain natriuretic peptide (NT-proBNP) is a widely used point-of-care (POC) cardiac biomarker in human medicine. Canine NT-proBNP is used less in veterinary medicine, possibly due to the lack of a POC canine NT-proBNP assay resulting in temporal delay, increased degradation in transport, and high reported variability in the available assay. A new quantitative POC analyzer allows fast, onsite measurement of NT-proBNP, minimizing preanalytical error and reducing variability. OBJECTIVE: We aimed to analytically validate an NT-proBNP assay (Vcheck) according to American Society of Veterinary Clinical Pathology (ASVCP) and Clinical Laboratory Improvement Amendments (CLIA) specifications. METHODS: Archived and prospective plasma and serum samples were collected from male and female, client-owned dogs of various breeds with cardiac abnormalities (n = 81) and a healthy control population (n = 225). Precision, accuracy, analytical sensitivity, and specificity, and other statistical analyses were performed. RESULTS: Imprecision was considered acceptable with a coefficient of variation ranging from 9% at 4000 pmol/L to 20% at 600 pmol/L. The lower limit of quantitation was 650 pmol/L based on repetitive measures evaluation. Comparison of the Vcheck assay with the Cardiopet NT-proBNP assay revealed an excellent correlation with minimal bias when preanalytical factors were controlled. Significant degradation of NT-proBNP occurred when current methods were used at refrigerated and room temperatures, which could change diagnostic and prognostic decision-making. Age-partitioned reference intervals have high reference values of 750 pmol/L and 1280 pmol/L for juvenile and adult dogs, respectively. CONCLUSIONS: The Vcheck NT-ProBNP assay provides analytically acceptable results. Onsite testing can minimize variability related to preanalytical error and provide clinically useful contemporaneous results. Samples should be centrifuged immediately and analyzed within 2 hours of collection.

A review of the toxicology of oil in vertebrates: what we have learned following the Deepwater Horizon oil spill.

In the wake of the Deepwater Horizon (DWH) oil spill, a number of government agencies, academic institutions, consultants, and nonprofit organizations conducted lab- and field-based research to understand the toxic effects of the oil. Lab testing was performed with a variety of fish, birds, turtles, and vertebrate cell lines (as well as invertebrates); field biologists conducted observations on fish, birds, turtles, and marine mammals; and epidemiologists carried out observational studies in humans. Eight years after the spill, scientists and resource managers held a workshop to summarize the similarities and differences in the effects of DWH oil on vertebrate taxa and to identify remaining gaps in our understanding of oil toxicity in wildlife and humans, building upon the cross-taxonomic synthesis initiated during the Natural Resource Damage Assessment. Across the studies, consistency was found in the types of toxic response observed in the different organisms. Impairment of stress responses and adrenal gland function, cardiotoxicity, immune system dysfunction, disruption of blood cells and their function, effects on locomotion, and oxidative damage were observed across taxa. This consistency suggests conservation in the mechanisms of action and disease pathogenesis. From a toxicological perspective, a logical progression of impacts was noted: from molecular and cellular effects that manifest as organ dysfunction, to systemic effects that compromise fitness, growth, reproductive potential, and survival. From a clinical perspective, adverse health effects from DWH oil spill exposure formed a suite of signs/symptomatic responses that at the highest doses/concentrations resulted in multi-organ system failure.

Validation of an immunocytochemical assay for immunophenotyping of lymphoma in guinea pigs (Cavia porcellus).

BACKGROUND: Lymphoma is an important disease of pet guinea pigs, although validation of immunophenotyping techniques based on cytologic or hematologic samples has not been reported. OBJECTIVE: To describe an immunocytochemical method for immunophenotyping of lymphoma (as either T- or B-cell) in guinea pigs, and to validate antibodies for this purpose. METHODS: Blood and tissues were obtained at the time of necropsy from laboratory guinea pigs and a privately owned dog (control) euthanized for reasons unrelated to lymphoproliferative disease. Fine-needle aspirates of enlarged peripheral lymph nodes were obtained from a case of spontaneous lymphoma in a pet guinea pig. Anti-CD3 and anti-Pax5 antibodies were validated by a combination of western blotting performed on splenic lysates of both the dog and guinea pigs, immunohistochemical studies on normal guinea pig tissues, and immunocytochemistry on normal guinea pig peripheral blood and splenic impression smears. RESULTS: The antibodies bound to antigens of an appropriate size in both the dog and guinea pig splenic lysates by Western blot analysis. Immunohistochemistry and immunocytochemistry demonstrated the expected distribution of putative T- and B-lymphocytes in normal tissues, peripheral blood, and splenic impression smears. As a proof-of-principle for its clinical utility, this immunocytochemical assay was used to diagnose a B-cell phenotype in a spontaneous lymphoma case in a pet guinea pig. CONCLUSIONS: Here, we validated an immunocytochemical method for immunophenotyping of lymphoma in guinea pigs as either a T- or B-cell phenotype. This enables future research into the clinical attributes of these subtypes and may ultimately improve both prognostication and therapy of lymphoma in guinea pigs.

STRECK CELL PRESERVATIVE STABILIZES KOALA ( PHASCOLARCTOS CINEREUS) WHOLE BLOOD FOR COMPLETE BLOOD COUNTS.

Wildlife health assessments at remote sites may lead to delayed testing of whole blood for complete blood counts (CBC) resulting in artifacts affecting clinical interpretation. Streck Cell Preservative (SCP) is a proprietary liquid stabilization reagent designed to preserve human leukocytes and may be applicable to wildlife health assessments when immediate processing of blood is not possible. The purpose of this study was to determine if SCP adequately preserved EDTA-anticoagulated whole blood from koalas ( Phascolarctos cinereus) for up to 14 days. Blood from 12 captive adult koalas was collected and combined with SCP in a 1 : 1 ratio and refrigerated. Paired samples of SCP treated and untreated blood had CBCs performed at five time-points over 14 days. Streck Cell Preservative extended koala EDTA-anticoagulated whole blood viability to 14 days by decreasing cellular lysis. Species- and method-specific reference intervals for SCP should be generated to avoid clinical misinterpretation, especially when evaluating anemia.

Sample Collection.

Results may be changed with suboptimal sample collection and transport, which then result in incorrect diagnoses. Quality management of samples must start in the patient, extend through sampling itself, include appropriate short transport, and then be correctly accessioned at the referral laboratory or in-house station to ensure accurate diagnosis of disease. A quality assurance plan at the laboratory chosen is mandatory under American Society for Veterinary Clinical Pathology guidelines.

ASVCP guidelines: Allowable total error hematology.

The purpose of this document is to provide total allowable error (TEa ) recommendations for commonly analyzed hematology measurands for veterinary personnel. These guidelines define relevant terminology and highlight considerations specific to hematology measurands. They also provide reasons and guidelines for using TEa in instrument performance evaluation, including recommendations for when the total observed error exceeds the recommended TEa . Biological variation-based quality specifications are briefly discussed. The appendix describes the derivation of the hematology TEa recommendations and provides resources for external quality assurance/proficiency testing programs and a worksheet for implementation of the guidelines.

Generation of red drum (Sciaenops ocellatus) hematology Reference Intervals with a focus on identified outliers.

BACKGROUND: The Deepwater Horizon (DWH) oil spill in 2010 released millions of barrels of crude oil into the northern Gulf of Mexico, exposing numerous species of animals to the toxic components of oil. A comprehensive assessment of morbidity and mortality caused by DWH oil exposure was undertaken by the DWH Natural Resource Damage Trustees to characterize ecosystem damages. OBJECTIVES: This study aimed to characterize normal hematologic RIs in red drum fish with blood cell descriptions, and to demonstrate the importance of identifying and removing outliers when generating RI. METHODS: Two years after the oil spill, 57 adult, red drum fish of mixed sexes were caught along the eastern Louisiana coastline. Eight different sites were chosen to catch the fish; 6 sites were contaminated with oil, and 2 sites were not contaminated at the time of the oil spill. Hematologic RIs were generated from heparinized whole blood samples of healthy red drum as determined by gross examination and histopathologic examination. Two methods were used to detect hematologic effects likely caused by oil contamination. RESULTS: Red drum PCVs (RI 42-62%) were higher than previously reported in cold water and bottom-dwelling fish species, while absolute WBC counts (RI 2.9-8.7  × 109 /L) were comparable to WBC counts previously reported in other fish species with heterophil and lymphocyte absolute concentrations frequently being equivalent. Anemic animals (PCV<42%) were only identified in oil-contaminated sites. CONCLUSION: RIs in many wild fish species are lacking, and therefore, this study provides valuable baseline data on healthy red drum fish. The outliers assessed using ASVCP RI guidelines can provide valuable clinical information regarding individuals in population health assessments, which may be more sensitive for the detection of abnormalities than for population statistics comparing the mean. The importance of removing outliers and rerunning RI statistics is highlighted by this field example.

Changes in white cell estimates and plasma chemistry measurements following oral or external dosing of double-crested cormorants, Phalacocorax auritus, with artificially weathered MC252 oil.

Scoping studies were designed whereby double-crested cormorants (Phalacocorax auritus) were dosed with artificially weathered Deepwater Horizon (DWH) oil either daily through oil injected feeder fish, or by application of oil directly to feathers every three days. Preening results in oil ingestion, and may be an effective means of orally dosing birds with toxicant to improve our understanding of the full range of physiological effects of oral oil ingestion on birds. Blood samples collected every 5-6 days were analyzed for a number of clinical endpoints including white blood cell (WBC) estimates and differential cell counts. Plasma biochemical evaluations were performed for changes associated with oil toxicity. Oral dosing and application of oil to feathers resulted in clinical signs and statistically significant changes in a number of biochemical endpoints consistent with petroleum exposure. In orally dosed birds there were statistically significant decreases in aspartate amino transferase (AST) and gamma glutamyl transferase (GGT) activities, calcium, chloride, cholesterol, glucose, and total protein concentrations, and increases in plasma urea, uric acid, and phosphorus concentrations. Plasma electrophoresis endpoints (pre-albumin, albumin, alpha-2 globulin, beta globulin, and gamma globulin concentrations and albumin: globulin ratios) were decreased in orally dosed birds. Birds with external oil had increases in urea, creatinine, uric acid, creatine kinase (CK), glutamate dehydrogenase (GLDH), phosphorus, calcium, chloride, potassium, albumin, alpha-1 globulin and alpha-2 globulin. Decreases were observed in AST, beta globulin and glucose. WBC also differed between treatments; however, this was in part driven by monocytosis present in the externally oiled birds prior to oil treatment.

Weathered MC252 crude oil-induced anemia and abnormal erythroid morphology in double-crested cormorants (Phalacrocorax auritus) with light microscopic and ultrastructural description of Heinz bodies.

Injury assessment of birds following the Deepwater Horizon (DWH) oil spill in 2010 was part of the Natural Resource Damage Assessment. One reported effect was hemolytic anemia with the presence of Heinz bodies (HB) in birds, however, the role of route and magnitude of exposure to oil is unknown. The purpose of the present study was to determine if double-crested cormorants (Phalacocorax auritis; DCCO) exposed orally and dermally to artificially weathered crude oil would develop hemolytic anemia including HB and reticulocytosis. In the oral experiment, sub-adult, mixed-sex DCCOs were fed control (n = 8) or oil-injected fish with a daily target dose of 5 (n = 9) or 10 (n = 9) ml oil/kg for 21 days. Then, subadult control (n = 12) and treated (n = 13) cormorant groups of similar sex-ratio were dermally treated with approximately 13ml of water or weathered MC252 crude oil, respectively, every 3 days for 6 dosages approximating 20% surface coverage. Collected whole blood samples were analyzed by light (new methylene blue) and transmission electron microscopy. Both oral and dermal treatment with weathered DWH MC252 crude oil induced regenerative, but inadequately compensated, anemia due to hemolysis and hematochezia as indicated by decreased packed cell volume, relative increase in reticulocytes with lack of difference in corrected reticulocyte count, and morphologic evidence of oxidant damage at the ultrastructural level. Hemoglobin precipitation, HB formation, degenerate organelles, and systemic oxidant damage were documented. Heinz bodies were typically <2µm in length and smaller than in mammals. These oblong cytoplasmic inclusions were difficult to see upon routine blood smear evaluation and lacked the classic button appearance found in mammalian red blood cells. They could be found as light, homogeneous blue inclusions upon new methylene blue staining. Ultrastructurally, HB appeared as homogeneous, electron-dense structures within the cytosol and lacked membranous structure. Oxidant damage in avian red blood cells results in degenerate organelles and precipitated hemoglobin or HB with different morphology than that found in mammalian red blood cells. Ultrastructural evaluation is needed to definitively identify HB and damaged organelles to confirm oxidant damage. The best field technique based on the data in this study is assessment of PCV with storage of blood in glutaraldehyde for possible TEM analysis.

Organ weights and histopathology of double-crested cormorants (Phalacrocorax auritus) dosed orally or dermally with artificially weathered Mississippi Canyon 252 crude oil.

A series of toxicity tests were conducted to assess the effects of low to moderate exposure to artificially weathered Deepwater Horizon Mississippi Canyon 252 crude oil on representative avian species as part of the Natural Resource Damage Assessment. The present report summarizes effects of oral exposure (n=26) of double-crested cormorants (DCCO; Phalacrocorax auritus) to 5 or 10ml oil kg-1 day-1 for up to 21 days or dermal application (n=25) of 13ml oil to breast and back feathers every three days totaling 6 applications in 21 days on organ weights and histopathology. Absolute and relative kidney and liver weights were increased in birds exposed to oil. Additionally, gross and/or histopathologic lesions occurred in the kidney, heart, pancreas and thyroid. Clinically significant renal lesions in the orally dosed birds included squamous metaplasia and increased epithelial hypertrophy of the collecting ducts and renal tubules and mineralization in comparison to controls. Gross cardiac lesions including thin walls and flaccid musculature were documented in both orally and dermally dosed birds and myocardial fibrosis was found in low numbers of dermally dosed birds only. Cytoplasmic vacuolation of the exocrine pancreas was noted in orally dosed birds only. Thyroid follicular hyperplasia was increased in dermally dosed birds only possibly due to increased metabolism required to compensate damaged feather integrity and thermoregulate. Gastrointestinal ulceration was found in orally dosed birds only. There were no significant hepatic histopathologic lesions induced by either exposure route. Therefore, hepatic histopathology is likely not a good representation of oil-induced damage. Taken together, the results suggest that oral or dermal exposure of DCCOs to artificially weathered MC252 crude oil induced organ damage that could potentially affect survivability.

Reprint of: Assay Validation of the Cardiac Isoform of Troponin I in Double Crested Cormorant (Phalacrocorax auritus) Plasma for Diagnosis of Cardiac Damage.

Cardiac abnormalities, initially found in Deepwater Horizon weathered MC252 crude oil exposed Double Crested Cormorants (DCCOs) upon gross necropsy, were further investigated using echocardiography. Clinical and statistically significant changes including decreased ventricular myocardial contractility and arrhythmia were elucidated by echocardiography and interpreted by boarded cardiologists as potentially life threatening. The objective of this investigation was to initiate development of an antemortem, sensitive blood screening test for cardiac damage due to oil exposure of avian species. An assay for the cardiac isoform of troponin I (cTnI) which is known to be highly cross-reactive across mammalian species was chosen and analytically validated in DCCO. This is the first time this test has been analytically validated in avian species. All plasma samples from birds assessed as healthy had trace concentrations (<0.016ng/ml). The assays was precise and accurate revealing a coefficient of variation <3% and an R2>0.99. Diagnostic investigation revealed that the test appears to have diagnostic potential for the diagnosis of cardiomyocyte damage. Diagnostic sensitivity and specificity were 91% and 73% in this laboratory population. Due to an equivocal sample population in which health could not be proven, further investigation is needed to diagnostically validate troponin I in the assessment of oil exposure in DCCO.

Toxicological and thermoregulatory effects of feather contamination with artificially weathered MC 252 oil in western sandpipers (Calidris mauri).

The external contamination of bird feathers with crude oil might have effects on feather structure and thus on thermoregulation. We tested the thermoregulatory ability of western sandpipers (Calidris mauri) in a respirometry chamber with oil applied either immediately prior, or three days before the experiment. The birds were then exposed to a sliding cold temperature challenge between 27°C and -3°C to calculate thermal conductance. After the experiment, a large blood sample was taken and the liver extracted to measure a range of parameters linked to toxicology and oxidative stress. No differences in thermal conductance were observed among groups, but birds exposed to oil for three days had reduced body temperatures and lost more body mass during that period. At necropsy, oiled birds showed a decrease in plasma albumin and sodium, and an increase in urea. This is reflective of dysfunction in the kidney at the loop of Henle. Birds, especially when exposed to the oil for three days, showed signs of oxidative stress and oxidative damage. These results show that the ingestion of externally applied oil through preening or drinking can cause toxic effects even in low doses, while we did not detect a direct effect of the external oil on thermoregulation over the temperature range tested.

Effect of oral exposure to artificially weathered Deepwater Horizon crude oil on blood chemistries, hepatic antioxidant enzyme activities, organ weights and histopathology in western sandpipers (Calidris mauri).

Shorebirds were among birds exposed to Mississippi Canyon 252 (MC252) crude oil during the 2010 Deep Water Horizon (DWH) oil spill in the Gulf of Mexico. The western sandpiper (Calidris mauri) was chosen as one of four species for initial oral dosing studies conducted under Phase 2 of the avian toxicity studies for the DWH Natural Resource Damage Assessment (NRDA). Thirty western sandpipers were assigned to one of three treatment groups, 10 birds per group. The control group was sham gavaged and the treatment groups were gavaged with 1 or 5mL oil kg bw-1 daily for 20 days. Periodic blood samples for hemoglobin measurements were collected during the trial. A final blood sample used to determine hemoglobin concentration in addition to complete blood counts, plasma clinical chemistries, haptoglobin concentration and plasma electrophoresis was collected when birds were euthanized and necropsied on day 21. Tissues were removed, weighed and processed for subsequent histopathological evaluation. There were numerical decreases in hemoglobin concentrations in oil-dosed birds over the 21-day trial, but values were not significantly different compared to controls on day 21. There were no significant differences between controls and oiled birds in complete blood counts, plasma chemistries, haptoglobin concentration, and plasma electrophoresis endpoints. Of the hepatic oxidative stress endpoints assessed, the total antioxidant capacity assessment (Trolox equivalents) for the control group was lower compared to the 1mL oil kg bw-1 group. Absolute liver weights in the 5mL oil kg bw-1 group were significantly greater compared to controls. While not conclusive, the numerical decrease in hemoglobin concentration and significant increase in absolute liver weight are consistent with exposure to oil. Histological changes in the adrenal gland could be considered a non-specific indicator of stress resulting from exposure to oil. It is possible that the quantity of oil absorbed was not sufficient to induce clearly evident hemolytic anemia or that the western sandpiper is relatively insensitive to ingested oil.

Assay validation of the cardiac isoform of troponin I in double crested cormorant (Phalacrocorax auritus) plasma for diagnosis of cardiac damage.

Cardiac abnormalities, initially found in Deepwater Horizon weathered MC252 crude oil exposed Double Crested Cormorants (DCCOs) upon gross necropsy, were further investigated using echocardiography. Clinical and statistically significant changes including decreased ventricular myocardial contractility and arrhythmia were elucidated by echocardiography and interpreted by boarded cardiologists as potentially life threatening. The objective of this investigation was to initiate development of an antemortem, sensitive blood screening test for cardiac damage due to oil exposure of avian species. An assay for the cardiac isoform of troponin I (cTnI) which is known to be highly cross-reactive across mammalian species was chosen and analytically validated in DCCO. This is the first time this test has been analytically validated in avian species. All plasma samples from birds assessed as healthy had trace concentrations (<0.016ng/ml). The assays was precise and accurate revealing a coefficient of variation <3% and an R2>0.99. Diagnostic investigation revealed that the test appears to have diagnostic potential for the diagnosis of cardiomyocyte damage. Diagnostic sensitivity and specificity were 91% and 73% in this laboratory population. Due to an equivocal sample population in which health could not be proven, further investigation is needed to diagnostically validate troponin I in the assessment of oil exposure in DCCO.

Assessment of toxicity and coagulopathy of brodifacoum in Japanese quail and testing in wild owls.

Based on detection of hepatic residues, scavenging and predatory non-target raptors are widely exposed to second generation anticoagulant rodenticides (SGARs). A small proportion, generally <10%, of tested birds are diagnosed as acutely poisoned. Little is known, however, of sub-lethal effects of SGARs, such as interaction of clotting capacity with traumatic injury. Assessment of coagulation function of birds submitted live to wildlife rehabilitators or veterinarians may provide a means of establishing the proportion of animals suffering sub-lethal coagulopathies, as well as identifying individuals requiring treatment. As a first step in exploring the potential of this approach, we dosed Japanese quail (Coturnix japonica) with the SGAR, brodifacoum, at 0, 0.8, 1.4, 1.9, and 2.5 mg/kg and sampled birds at 1, 3, 5 and 7 days post-dosing. Prothrombin time (PT), which measures the extrinsic coagulation pathway, was significantly prolonged in 98% of brodifacoum-exposed quail in a dose- and time-dependent manner. 50-fold prolongation of PT occurred at higher brodifacoum dosages and correlated to hemorrhage found at necropsy. Activated clotting time (ACT), a measure of the intrinsic pathway also increased with dose and time. Hemoglobin (Hb) and hematocrit (Hct) decreased dose- and time-dependently at doses ≥1.4 mg/kg with no significant change at 0.8 mg/kg. Reference intervals for PT (10.0-16.2 s), ACT (30-180 s), Hb (9.6-18.4 g/dl), and Hct (34-55%) were established in Japanese quail. Species-specific reference intervals are required as barn owl PT (17-29 s) and quail PT were different. The proportion of brodifacoum-exposed quail with hemorrhage was not correlated with liver residues, but was correlated with PT, suggesting that this assay is a useful indicator of avian anticoagulant rodenticide exposure. PTs measured in free-living barn owls sampled between April 2009 and August 2010 in the lower Fraser Valley of BC do not suggest significant exposure to SGARs.

ASVCP guidelines: quality assurance for point-of-care testing in veterinary medicine.

Point-of-care testing (POCT) refers to any laboratory testing performed outside the conventional reference laboratory and implies close proximity to patients. Instrumental POCT systems consist of small, handheld or benchtop analyzers. These have potential utility in many veterinary settings, including private clinics, academic veterinary medical centers, the community (eg, remote area veterinary medical teams), and for research applications in academia, government, and industry. Concern about the quality of veterinary in-clinic testing has been expressed in published veterinary literature; however, little guidance focusing on POCT is available. Recognizing this void, the ASVCP formed a subcommittee in 2009 charged with developing quality assurance (QA) guidelines for veterinary POCT. Guidelines were developed through literature review and a consensus process. Major recommendations include (1) taking a formalized approach to POCT within the facility, (2) use of written policies, standard operating procedures, forms, and logs, (3) operator training, including periodic assessment of skills, (4) assessment of instrument analytical performance and use of both statistical quality control and external quality assessment programs, (5) use of properly established or validated reference intervals, (6) and ensuring accurate patient results reporting. Where possible, given instrument analytical performance, use of a validated 13s control rule for interpretation of control data is recommended. These guidelines are aimed at veterinarians and veterinary technicians seeking to improve management of POCT in their clinical or research setting, and address QA of small chemistry and hematology instruments. These guidelines are not intended to be all-inclusive; rather, they provide a minimum standard for maintenance of POCT instruments in the veterinary setting.

ASVCP guidelines: allowable total error guidelines for biochemistry.

As all laboratory equipment ages and contains components that may degrade with time, initial and periodically scheduled performance assessment is required to verify accurate and precise results over the life of the instrument. As veterinary patients may present to general practitioners and then to referral hospitals (both of which may each perform in-clinic laboratory analyses using different instruments), and given that general practitioners may send samples to reference laboratories, there is a need for comparability of results across instruments and methods. Allowable total error (TEa ) is a simple comparative quality concept used to define acceptable analytical performance. These guidelines are recommendations for determination and interpretation of TEa for commonly measured biochemical analytes in cats, dogs, and horses for equipment commonly used in veterinary diagnostic medicine. TEa values recommended herein are aimed at all veterinary settings, both private in-clinic laboratories using point-of-care analyzers and larger reference laboratories using more complex equipment. They represent the largest TEa possible without generating laboratory variation that would impact clinical decision making. TEa can be used for (1) assessment of an individual instrument's analytical performance, which is of benefit if one uses this information during instrument selection or assessment of in-clinic instrument performance, (2) Quality Control validation, and (3) as a measure of agreement or comparability of results from different laboratories (eg, between the in-clinic analyzer and the reference laboratory). These guidelines define a straightforward approach to assessment of instrument analytical performance.

Blood mineral concentrations in manatees (Trichechus manatus latirostris and Trichechus manatus manatus).

Limited information is available regarding the role of minerals and heavy metals in the morbidity and mortality of manatees. Whole-blood and serum mineral concentrations were evaluated in apparently healthy, free-ranging Florida (Trichechus manatus latirostris, n = 31) and Belize (Trichechus manatus manatus, n = 14) manatees. Toxicologic statuses of the animals and of their environment had not been previously determined. Mean mineral whole-blood (WB) and serum values in Florida (FL) and Belize (BZ) manatees were determined, and evaluated for differences with respect to geographic location, relative age, and sex. Mean WB and serum silver, boron, cobalt, magnesium, molybdenum, and WB cadmium concentrations were significantly higher in BZ versus FL manatees (P < 0.05). Mean WB aluminum, calcium, manganese, sodium, phosphorus, vanadium, and serum zinc concentrations were significantly lower in BZ versus FL manatees. Adult manatees had significant and higher mean WB aluminum, manganese, sodium, antimony, vanadium, and serum manganese and zinc concentrations compared to juvenile animals. Significant and lower mean WB and serum silver, boron, cobalt, and serum copper and strontium concentrations were present in adults compared to juveniles (P < or = 0.05). Females had significant and higher mean WB nickel and serum barium compared to males (P < or = 0.05). Mean WB arsenic and zinc, and mean serum iron, magnesium, and zinc concentrations fell within toxic ranges reported for domestic species. Results reveal manatee blood mineral concentrations differ with location, age, and sex. Influence from diet, sediment, water, and anthropogenic sources on manatee mineral concentration warrant further investigation.

ASVCP reference interval guidelines: determination of de novo reference intervals in veterinary species and other related topics.

Reference intervals (RI) are an integral component of laboratory diagnostic testing and clinical decision-making and represent estimated distributions of reference values (RV) from healthy populations of comparable individuals. Because decisions to pursue diagnoses or initiate treatment are often based on values falling outside RI, the collection and analysis of RV should be approached with diligence. This report is a condensation of the ASVCP 2011 consensus guidelines for determination of de novo RI in veterinary species, which mirror the 2008 Clinical Laboratory and Standards Institute (CLSI) recommendations, but with language and examples specific to veterinary species. Newer topics include robust methods for calculating RI from small sample sizes and procedures for outlier detection adapted to data quality. Because collecting sufficient reference samples is challenging, this document also provides recommendations for determining multicenter RI and for transference and validation of RI from other sources (eg, manufacturers). Advice for use and interpretation of subject-based RI is included, as these RI are an alternative to population-based RI when sample size or inter-individual variation is high. Finally, generation of decision limits, which distinguish between populations according to a predefined query (eg, diseased or non-diseased), is described. Adoption of these guidelines by the entire veterinary community will improve communication and dissemination of expected clinical laboratory values in a variety of animal species and will provide a template for publications on RI. This and other reports from the Quality Assurance and Laboratory Standards (QALS) committee are intended to promote quality laboratory practices in laboratories serving both clinical and research veterinarians.

ASVCP quality assurance guidelines: control of preanalytical and analytical factors for hematology for mammalian and nonmammalian species, hemostasis, and crossmatching in veterinary laboratories.

In December 2009, the American Society for Veterinary Clinical Pathology (ASVCP) Quality Assurance and Laboratory Standards committee published the updated and peer-reviewed ASVCP Quality Assurance Guidelines on the Society's website. These guidelines are intended for use by veterinary diagnostic laboratories and veterinary research laboratories that are not covered by the US Food and Drug Administration Good Laboratory Practice standards (Code of Federal Regulations Title 21, Chapter 58). The guidelines have been divided into 3 reports: (1) general analytical factors for veterinary laboratory performance and comparisons; (2) hematology, hemostasis, and crossmatching; and (3) clinical chemistry, cytology, and urinalysis. This particular report is one of 3 reports and provides recommendations for control of preanalytical and analytical factors related to hematology for mammalian and nonmammalian species, hemostasis testing, and crossmatching and is adapted from sections 1.1 and 2.3 (mammalian hematology), 1.2 and 2.4 (nonmammalian hematology), 1.5 and 2.7 (hemostasis testing), and 1.6 and 2.8 (crossmatching) of the complete guidelines. These guidelines are not intended to be all-inclusive; rather, they provide minimal guidelines for quality assurance and quality control for veterinary laboratory testing and a basis for laboratories to assess their current practices, determine areas for improvement, and guide continuing professional development and education efforts.