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.
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An 11‐year‐old female spayed German Wirehaired Pointer with a 1‐week history of lethargy, hyporexia, diarrhea, and coughing presented with pericardial effusion causing cardiac tamponade. An ...echocardiogram revealed no structural cause for pericardial effusion. The pericardial effusion was an exudate with mixed macrophagic and neutrophilic inflammation. Morulae occasionally were found within neutrophils. The pericardial fluid and blood were qPCR and cPCR positive for Anaplasma phagocytophilum (NC State University, Vector‐borne Disease Diagnostic Laboratory, Raleigh, NC). The dog's blood was negative by ELISA (Vetscan Flex4 Rapid Test, Zoetis, Parsippany, NJ) for A. phagocytophilum antibodies at initial presentation and subsequently positive (SNAP4DxPlus, IDEXX, Westbrook, ME) 7 days later. After pericardiocentesis and administration of doxycycline (5 mg/kg PO q12h for 14 days), a repeat echocardiogram performed 1 month later showed no recurrence of pericardial effusion.
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Prior to introduction of a new method to the diagnostic laboratory, analytical performance must be validated to ensure operation within the manufacturer's specifications and/or within predetermined ...quality requirements. In addition, the new method may require diagnostic performance assessment to ensure it differentiates between diseased and nondiseased individuals as intended. These 2 phases of assessment, while complementary, are not equivalent and require a different set of experiments, statistical analyses, and interpretation. Studies of analytical performance typically include a method comparison experiment, the purpose of which is to identify bias (inaccuracy) of the “test” (or “index”) method (new method) relative to a “comparative method” (established method). Analysis of method comparison data is facilitated by commercial software programs that present the statistical significance of identified bias; however, the clinical relevance of any bias also should be considered. Studies of diagnostic performance should not be pursued until analytical performance is fully characterized and may not be required for well‐established tests or for those for which results are nonspecific (ie, not referable to a specific disease or condition). Diagnostic performance assessment may include assessment of sensitivity, specificity, predictive values, odds ratios, and/or likelihood ratios. The purpose of this review is to clarify differences between the assessment of analytical and diagnostic performance, and to explore the method comparison study and bias assessment from a perspective not addressed in prior veterinary articles.
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Climate change is influencing polar bear (Ursus maritimus) habitat, diet, and behavior but the effects of these changes on their physiology is not well understood. Blood-based biomarkers are used to ...assess the physiologic health of individuals but their usefulness for evaluating population health, especially as it relates to changing environmental conditions, has rarely been explored. We describe links between environmental conditions and physiologic functions of southern Beaufort Sea polar bears using data from blood samples collected from 1984 to 2018, a period marked by extensive environmental change. We evaluated associations between 13 physiologic biomarkers and circumpolar (Arctic oscillation index) and regional (wind patterns and ice-free days) environmental metrics and seasonal and demographic co-variates (age, sex, season, and year) known to affect polar bear ecology. We observed signs of dysregulation of water balance in polar bears following years with a lower annual Arctic oscillation index. In addition, liver enzyme values increased over time, which is suggestive of potential hepatocyte damage as the Arctic has warmed. Biomarkers of immune function increased with regional-scale wind patterns and the number of ice-free days over the Beaufort Sea continental shelf and were lower in years with a lower winter Arctic oscillation index, suggesting an increased allocation of energetic resources for immune processes under these conditions. We propose that the variation in polar bear immune and metabolic function is likely indicative of physiologic plasticity, a response that allows polar bears to remain in homeostasis even as they experience changes in nutrition and habitat in response to changing environments.
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Captive‐bred rhesus macaques of Indian origin represent one of the most important large animal models for infectious disease, solid organ transplantation, and stem cell research. There is a dearth of ...information defining hematopoietic development, including neutrophil leukocyte differentiation in this species using multicolor flow cytometry. In the current study, we sought to identify cell surface markers that delineate neutrophil progenitor populations with characteristic immunophenotypes. We defined four different postmitotic populations based on their CD11b and CD87 expression pattern, and further refined their immunophenotypes using CD32, CD64, lactoferrin, and myeloperoxidase as antigenic markers. The four subsets contained myelocyte, metamyelocyte, band, and segmented neutrophil populations. We compared our flow cytometry‐based classification with the classical nuclear morphology‐based classification. We found overlap of immunological phenotype between populations of different nuclear morphology and identified phenotypically different subsets within populations of similar nuclear morphology. We assessed the responsiveness of these populations to stimulatory signals, such as LPS, fMLP, or PMA, and demonstrated significant differences between human and rhesus macaque neutrophil progenitors. In this study, we provided evidence for species‐specific features of granulopoiesis that ultimately manifested in the divergent immunophenotypes of the fully differentiated segmented neutrophils of humans and rhesus macaques. Additionally, we found functional markers that can be used to accurately quantify neutrophil progenitors by flow cytometry. Although these markers do not coincide with the classical nuclear‐morphology‐based grading, they enable us to perform functional studies monitoring immunophenotypic markers.
A flow cytometric protocol using species‐specific characteristics to define rhesus macaque neutrophil progenitors in the bone marrow.
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Disorders of the avian reproductive tract are common, yet monitoring their resolution presents a diagnostic dilemma. Reproductive hormones such as luteinizing hormone or estrogen are the best ...reflection of reproductive status, but the required sample volumes and lack of reference intervals limit their clinical utility. An alternative analyte is blood triglyceride, the concentration of which rises markedly during sustained estrogen release from the ovary. Portable meters for measuring human blood triglyceride concentration offer the advantage of using minimal sample volumes, but these have not been validated for use in birds. We assessed the precision and accuracy of 2 portable meters for measuring blood triglyceride concentration in pooled whole blood and plasma from chickens (n = 42), and performed method comparison using a reference analyzer and determined total error. Within-run repeatability was fair-to-excellent using whole blood and plasma (range: 2.5–11.5%), and between-run repeatability using plasma was similar (3.1–12.2%). The meters performed well in recovery and dilution studies in which almost all readings fell within the preset requirement of 75–125%. Correlations between each meter, using whole blood and plasma, and the reference analyzer, using plasma only, were high to very high (0.86–0.98). Bias determined by Bland–Altman analysis was similar between whole blood and plasma for each meter, yet markedly different between the meters. The calculated total observed error was consequently within our pre-set total allowable error of 25% for one meter but not the other, indicating the requirement for a meter-specific reference interval.
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Letter to the Editor and Authors' Response Nemetz, Larry; Irvine, Katherine L.; Mans, Christoph ...
Journal of avian medicine and surgery,
12/2019, Volume:
33, Issue:
4
Journal Article
Peer reviewed
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Female reproductive disorders, such as chronic egg laying, are common in captive psittacine birds. While a disease diagnosis related to reproductive disorders can often be accomplished by physical ...examination and diagnostic imaging, monitoring of the response to environmental modification and medical treatment is more challenging. Monitoring ideally would involve measurement of luteinizing hormone or estrogen to assess ovarian activity. However, the amount of blood required for hormone analysis is greater than the small sample size that one can collect from these birds. Additionally, the lack of reference intervals limits their use as a diagnostic tool. Because plasma triglyceride increases during sustained estrogen release from the ovary, it may be used as an alternative method for assessing ovarian activity in birds. Point-of-care (POC) analyzers for measuring lipids in human plasma use very small sample volumes and have been used for measuring triglycerides in animals, including chickens. The authors therefore performed a method comparison study with 2 POC analyzers and a reference analyzer and plasma and whole blood from psittacine birds to determine whether these meters are suitable for triglyceride measurement in a known population of psittacine birds. Correlation, Deming regression, and Bland-Altman analyses were used to assess performance, and the total observed error for each meter relative to the reference analyzer was calculated. One of the meters exhibited fair performance and, with species-specific reference intervals, is likely to be clinically useful for triglyceride measurement in psittacine birds. The other meter demonstrated poor performance with unacceptable error, and its use for this purpose is strongly discouraged.
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A 10‐year‐old female spayed Miniature Schnauzer was presented for investigation of an intra‐nasal mass. The mass was diagnosed by histopathologic examination as an undifferentiated round cell ...neoplasm with an infiltrate of segmented leukocytes, interpreted as neutrophilic inflammation. The mass was treated with palliative radiation and systemic chemotherapy due to the presence of regional lymph node metastasis. During subsequent monitoring over several months, the peripheral leukocyte concentration was repeatedly within reference intervals to slightly increased with low numbers of toxic neutrophils. Four months after the initial diagnosis, there was a significant leukocytosis of 66 100 cells/μL, and 39 700 cells/μL of the leukocytes had variably mature, lobulated, and hypolobulated nuclei, and grey cytoplasm with clear vacuoles, resembling grey eosinophils. To further characterize these cells, peripheral blood smears from the patient and a canine control with eosinophilia were stained for alkaline phosphatase (AP), peroxidase, and esterase activities, and with Luxol fast blue (LFB). Histopathologic sections of the nasal mass were stained with LFB and immunohistochemically for tryptase. On blood smears, the cytoplasm of the suspected grey eosinophils stained for AP and granules stained with LFB confirmed that there was an eosinophilic lineage. Peroxidase staining was weak, and esterase staining was absent. On histopathologic sections from the nasal mass, the segmented leukocytes contained LFB‐staining granules, indicating an eosinophilic infiltrate was present. Neoplastic cells expressed tryptase, which confirms a mast cell lineage. Our findings suggest that grey eosinophils might be under‐recognized and interpreted incorrectly as toxic neutrophils. This report expands the canine breeds in which these eosinophils have been identified.
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