Repeat patient testing-quality control compared to commercial quality control material for the Sysmex XT-2000iV hematology analyzer in a multi-site veterinary laboratory.

BACKGROUND: Quality control material (QCM) for hematology in veterinary laboratories is limited, and repeat patient testing quality control (RPT-QC) is an alternative method using excess matrix-specific samples. OBJECTIVES: This study aimed to determine if median differences between RPT-QC analyses for each time interval for RBC, HGB, HCT, and WBC were the same, determine if unified RPT-QC limits can be applied to a network of veterinary laboratories, compare the performance of RPT-QC to commercial QCM for the reference analyzer and evaluate the experience over a 4 month period and design, improve and implement an automated spreadsheet for RPT-QC data management. METHODS: The potential to unify individual analyzer RPT-QC limits for red blood cells (RBC), hematocrit (HCT), hemoglobin (HGB), and white blood cells (WBC) on multi-site Sysmex XT-2000-iV analyzers was explored by a difference of means test and confidence interval determination for the median difference for each network analyzer in comparison to the network reference analyzer. User experience of an automated RPT-QC data management Excel spreadsheet was collected by user feedback during monthly meetings. Numbers of out-of-control results and the root causes for these for RPT-QC were compared against those of a commercial QCM over a 4-month period. RESULTS: Differences between individual analyzer RPT-QC limits were too large to allow for unification of network limits. The automated spreadsheet successfully highlighted out-of-control events for RPT-QC. Trends or shifts were more frequent for commercial QCM based on observed performance and a 1-2.5 s QC rule than for RPT-QC. Following routine troubleshooting, RPT-QC out-of-control events were resolved with an alternative RPT-QC sample indicating random error associated with excessive deterioration. Use of an automated spreadsheet for recording RPT-QC, documentation and troubleshooting of out-of-control events, and collating monthly summary calculations were considered an asset in laboratory quality management. CONCLUSIONS: RPT-QC can be successfully implemented and integrated into a multi-site veterinary laboratory. Individual analyzer RPT-QC limit generation is recommended. The deterioration of commercial QCM caused shifts or trends in QC results, which initiated more repeat analyses and investigations than did RPT-QC.

Biological variation of routine haematology and biochemistry measurands in the horse.

BACKGROUND: Clinical pathology results are typically interpreted by referring to population-based reference intervals. The use of individualised (subject-based) reference intervals is more appropriate for measurands with a high degree of variation between individuals. OBJECTIVES: To determine the biological variation of routinely analysed equine haematology and biochemistry measurands and calculate indices of individuality and reference change values which enable production of individualised reference intervals, in a group of healthy, privately owned horses. STUDY DESIGN: In a prospective cohort study, thirty-nine privately owned horses were sampled by jugular venipuncture for analysis of haematology and biochemistry measurands at weekly intervals for 6 weeks. METHODS: Haematology was analysed on the day of collection. Serum was frozen and biochemistry analyses performed on thawed samples. Duplicate results were obtained and the coefficient of variation was calculated for analytical variation, within-subject variation and between-subject variation. The index of individuality and reference change value were derived for each measurand. RESULTS: Haematology (red blood cell count, mean corpuscular haemoglobin and mean cell volume) and biochemistry measurands (total protein, globulins, albumin, gamma-glutamyl transferase, aspartate aminotransferase) demonstrated high individuality, indicating that individualised reference intervals are more appropriate for evaluation of these measurands. Two haematology (mean corpuscular haemoglobin concentration and platelets) and three biochemistry measurands (chloride, glucose and sodium) had low individuality, indicating that the use of traditional population-based reference intervals is appropriate for these measurands. Remaining measurands had intermediate individuality suggesting interpretation of the reference change value should occur with consideration of the population-based reference interval. MAIN LIMITATIONS: The use of privately owned horses, variable management and environmental factors. CONCLUSIONS: The use of individualised reference intervals is justified for many measurands in horses, supporting the use of serial sampling, consideration of biological variation and application of reference change values for improved clinical decision making and patient management in equine practice.

Clinical application of biological variation data to facilitate interpretation of canine and feline laboratory results.

Interpretation of laboratory results is based on comparison of the patient's own results against established decision thresholds or reference intervals in the context of the clinical presentation and history. Blood measurand analysis has pre-analytical, analytical and physiological sources of variation, which may complicate interpretation of results. Biological variation describes the physiological random fluctuation of blood measurands around a homeostatic set point, which varies within and between individuals. This article reviews the practical applications of biological variation in the everyday clinical setting. Examples are offered to highlight how biological variation can be used to: (1) assess the usefulness of subject-based reference intervals, (2) determine measurand homeostatic set points, (3) interpret single or serial results for diagnosis of disease and (4) evaluate changes in serial results during monitoring.

Determination of haematological reference intervals in healthy adult greyhounds.

OBJECTIVE: The purpose of this study was to establish breed-specific reference intervals for haematological measurands in non-racing greyhounds. Suitability of the data for partitioning according to sex was also examined. METHODS: Haematological data were collected from 304 healthy non-racing greyhounds and analysed using non-parametric methods. Results were compared with non-breed-specific canine reference intervals and also with greyhound reference intervals obtained by other investigators. RESULTS: Compared with non-breed-specific reference intervals, the results showed comparable mean and upper limit and higher lower limit for erythrocyte count; higher values for haemoglobin, haematocrit and mean corpuscular volume; and lower values for total leucocyte count and absolute concentration of neutrophils, lymphocytes, monocytes, eosinophils and platelets. Partitioning according to sex was recommended by the statistical analysis for all analytes except haematocrit and total leucocyte count. CLINICAL SIGNIFICANCE: In this study the reference intervals were derived from a large sample size. The results are in general agreement with previous reports, although higher values for low reference limits have been noted for the erythroid parameters, and lower values for upper reference limits have been observed for the total and different leucocyte counts. Breed-specific reference intervals provide a useful clinical tool for haematological evaluations.

Determination of serum biochemistry reference intervals in a large sample of adult greyhounds.

BACKGROUND: As large breed, highly muscled dogs, greyhounds are regarded as physiologically different to other breeds. Biochemistry reference intervals have previously been determined using small numbers of greyhounds or based on the observations from racing dogs. OBJECTIVES: The purpose of this study was to develop statistically defined reference intervals for biochemical analytes in healthy non-racing greyhounds. Partitioning according to gender was also examined. METHODS: Biochemical analytes from a population of non-racing healthy greyhounds, including 269 males, 202 females and 28 dogs where gender had not been recorded, were examined using a non-parametric statistical approach. The dogs were aged between one and nine years old. RESULTS: Total protein, albumin, globulin and creatinine reference intervals differed from generic reference intervals used for dogs. The reference intervals for greyhounds in this study are similar to those obtained by other studies of greyhounds, but often had a narrower range of values, likely reflecting more accurate estimation associated with larger numbers of reference individuals. Recommended methods for assessment of partitioning do not indicate a need to partition according to gender.

Canine paediatric oncology: retrospective assessment of 9522 tumours in dogs up to 12 months (1993-2008).

Little information is available on the occurrence of neoplasms in dogs up to the age of 12 months. This is a retrospective review of histopathological diagnoses of neoplasia in dogs up to the age of 12 months based on biopsy specimens submitted to a commercial veterinary diagnostic laboratory in the United Kingdom between 1993 and 2008. In 20 280 histological submissions, 9522 neoplasms were identified. Canine cutaneous histiocytoma (n = 8465; 89%) was the most common histological type. Neoplasms other than histiocytoma (n = 1057; 11%) were grouped as benign epithelial (n = 375; 4%), haematopoietic (n = 229; 2%), benign mesenchymal (n = 145; 2%), miscellaneous (n = 118; 1%), non-hematopoietic malignant mesenchymal (n = 118; 1%) or malignant epithelial tumours (n = 72; <1%). Excluding canine cutaneous histiocytoma, 52% of tumours (n = 547) were benign, and 66% were from the skin or soft tissues. These data provide valuable epidemiological information on neoplasms occurring in juvenile dogs in the United Kingdom.

Feline paediatric oncology: retrospective assessment of 233 tumours from cats up to one year (1993 to 2008).

OBJECTIVES: To determine which types of tumour occur in cats up to the age of 12 months based on biopsies submitted to Idexx Laboratories, Wetherby, UK. METHODS: Retrospective review of histopathological diagnoses of tumours in cats up to the age of 12 months from biopsies received between September 1993 and March 2008. RESULTS: A total of 4196 submissions from cats 12 months old or younger were identified; 233 biopsies (6%) were neoplastic and fulfilled the search criteria. Tumours were categorised as haematopoietic (n=73, 31%), malignant epithelial (n=44; 19%), malignant mesenchymal (n=38; 16%), benign epithelial (n=37; 16%), benign mesenchymal (n=30, 13%) and miscellaneous (n=11; 5%). The most frequent tumours were lymphoma (n=51; 22%), soft-tissue sarcoma (n=34; 15%), mast cell tumour (n=22; 9%) and squamous cell carcinoma (n=16; 7%). The most common tumour site was the skin and soft tissues (41% of tumours). In all, 164 neoplasms (70%) were malignant or had malignant potential. CLINICAL SIGNIFICANCE: These data provide unique epidemiological information on a poorly characterised subgroup of feline cancer patients in the UK.

Establishment of the European College of Veterinary Clinical Pathology (ECVCP) and the current status of veterinary clinical pathology in Europe.

After 5 years of development, the European College of Veterinary Clinical Pathology (ECVCP) was formally recognized and approved on July 4, 2007 by the European Board of Veterinary Specialisation (EBVS), the European regulatory body that oversees specialization in veterinary medicine and which has approved 23 colleges. The objectives, committees, basis for membership, constitution, bylaws, information brochure and certifying examination of the ECVCP have remained unchanged during this time except as directed by EBVS. The ECVCP declared full functionality based on the following criteria: 1) a critical mass of 65 members: 15 original diplomates approved by the EBVS to establish the ECVCP, 37 de facto diplomates, 7 diplomates certified by examination, and 5 elected honorary members; 2) the development and certification of training programs, laboratories, and qualified supervisors for residents; currently there are 18 resident training programs in Europe; 3) administration of 3 annual board-certifying examinations thus far, with an overall pass rate of 70%; 4) European consensus criteria for assessing the continuing education of specialists every 5 years; 5) organization of 8 annual scientific congresses and a joint journal (with the American Society for Veterinary Clinical Pathology) for communication of scientific research and information; the College also maintains a website, a joint listserv, and a newsletter; 6) collaboration in training and continuing education with relevant colleges in medicine and pathology; 7) development and strict adherence to a constitution and bylaws compliant with the EBVS; and 8) demonstration of compelling rationale, supporting data, and the support of members and other colleges for independence as a specialty college. Formal EBVS recognition of ECVCP as the regulatory body for the science and practice of veterinary clinical pathology in Europe will facilitate growth and development of the discipline and compliance of academic, commercial diagnostic, and industry laboratories in veterinary clinical pathology. Future needs are in developing sponsorship for resident positions, increasing employment opportunities, increasing compliance with laboratory, training, and continuing education standards, and advancing relevant science and technology.

Inactivation of the p16 cyclin-dependent kinase inhibitor in high-grade canine non-Hodgkin's T-cell lymphoma.

The significance of p16/Rb tumor suppressor pathway inactivation in T-cell non-Hodgkin's lymphoma (NHL) remains incompletely understood. We used naturally occurring canine NHL to test the hypothesis that p16 inactivation has specific pathologic correlates. Forty-eight samples (22 T-cell NHL and 26 B-cell NHL) were included. As applicable, metaphase- or array-based comparative genomic hybridization, Southern blotting, promoter methylation, and Rb phosphorylation were used to determine the presence, expression, and activity of p16. Fisher's exact test was used to test for significance. Deletion of p16 (or loss of dog chromosome 11) was restricted to high-grade T-cell NHL (lymphoblastic T-cell lymphoma and peripheral T-cell lymphoma, not otherwise specified). These were characterized by a concomitant increase of tumor cells with Rb phosphorylation at canonical CDK4 sites. Rb phosphorylation also was seen in high-grade B-cell NHL (diffuse large B-cell lymphoma and Burkitt-type lymphoma), but in those cases, it appeared to be associated with c-Myc overexpression. The data show that p16 deletion or inactivation occurs almost exclusively in high-grade T-cell NHL; however, alternative pathways can generate functional phenotypes of Rb deficiency in low-grade T-cell NHL and in high-grade B-cell NHL. Both morphologic classification according to World Health Organization criteria and assessment of Rb phosphorylation are prognostically valuable parameters for canine NHL.

Multilobulated "flower" cells in a subcutaneous mass aspirate from a cat.

An 8-year-old intact male cat was presented with a subcutaneous mass in the region of the right jugular vein. Cytologic and histopathologic examinations revealed cells with multilobulated nuclei (flower cells). Immunochemistry using a panel of markers showed vimentin-positivity on cytologic specimens, and postive staining for CD79a and BLA36 on histologic specimens. The final diagnosis was lymphoma of B-cell origin. We have observed similar multilobulated cells in ascites fluid, thoracic fluid, and peripheral blood from dogs and cats with a variety of lymphoid and myeloid neoplasms. Cells with multilobulated nuclei that resemble flower petals also have been described in humans. These cells are infrequently observed in canine and feline cytology specimens and require immunochemistry to determine their cell of origin.

Prognostic factors affecting survival of 507 horses with joint disease: (1983 to 1990).

Between July 1, 1983 and December 31, 1990, risk factors were determined for all horses with joint disease presented to a referral center, of being discharged, of ever becoming sound, or of being alive at 3 mo follow-up. Logistic multiple-regression models were done separately for foals (< or = 4 mo), yearlings (> 4-24 mo) and racing or nonracing adult horses (> 24 mo). The breakdown in this study was 53 foals, 87 yearlings, 141 nonracing adults, and 226 racing adults. Thirty-one foals (58%), 68 yearlings (78%), 119 non-racing adults (84%), and 213 racing adults (94%) were discharged. Foals with a less severe lameness, duration of illness of > 1 d, and infectious arthritis had increased odds of discharge. At follow-up, 12 of 18 (67%) were alive, 10 (56%) of which were sound. Yearlings with osteochondrosis had higher odds of discharge; at follow-up, 38 of 49 (78%) were alive, 32 (65%) of which were sound. For non-racing adults, horses with less severe lameness, without a miscellaneous diagnosis, or intended for pleasure use had increased odds of discharge. At follow-up, 55 of 78 (70%) were alive and 33 of 58 (57%) with soundness data became sound. Risk factors for higher odds of being alive at follow-up were carpal lameness, arthroscopic surgery, a prognosis other than poor, became sound, above-median hospitalization costs, and duration of follow-up. The 161 racing adults (76% of discharges), with follow-up, were more likely to have had osteoarthritis, higher hospital costs, hospitalization > 1 d, and arthroscopy. Sixty-four (60%) of these became sound; the odds increased if the horse was not severely lame at admission or was hospitalized for > 1 d. Risk factors and prognosis differed by age-use group among horses seen at our hospital.

Detection and clinical significance of plasma glutathione-S-transferases in dogs with lymphoma.

BACKGROUND: The objective of this study was to determine if glutathione-S-transferases were detectable in the plasma of dogs and to determine if concentrations of the a- and pi-subtypes were related with tumor response to single agent anthracycline (e.g., doxorubicin) chemotherapy in dogs with lymphoma. MATERIALS AND METHODS: Plasma was obtained from 10 healthy, normal dogs and from 11 dogs with lymphoma before treatment, 3 weeks after 1 dose of doxorubicin and every 3 weeks thereafter until relapse (the physical detection of recurrent and enlarged peripheral lymph nodes). Plasma concentration of alpha and pi-GST was determined by use of an ELISA technique with well plates pre-coated with IgG[anti-Canine alpha-GST or anti-Human pi-GST]. RESULTS: Mean plasma alpha-GST concentrations did not significantly decline after 1 dose of doxorubicin chemotherapy; however, mean plasma alpha-GST concentrations were markedly increased (p < 0.05) at the time of relapse (the physical detection of recurrent and enlarged peripheral lymph nodes). CONCLUSIONS: In this study we show that a relationship exists between the plasma alpha-GST concentration and the clinical response of dogs with lymphoma to doxorubicin chemotherapy.

Right leg muscle atrophy and osteopenia caused by renal adenocarcinoma in a cockatiel (Melopsittacus, undulatus).

A 7-year-old cockatiel presented with a 1 week history of right leg lameness. A renal adenocarcinoma invading and constricting the right ischiatic nerve resulted in disuse atrophy of the affected leg and radiographic evidence of osteopenia. This report illustrates the natural behavior, radiographic, and pathologic appearance of malignant renal tumors in birds.

Serum alpha 1-acid glycoprotein concentrations before and after relapse in dogs with lymphoma treated with doxorubicin.

OBJECTIVE: To determine whether serum alpha 1-acid glycoprotein (AGP) concentration was a useful marker of relapse in dogs with lymphoma that were in clinical remission following treatment with doxorubicin. DESIGN: Cohort study. ANIMALS: 12 dogs with lymphoma and 10 healthy dogs. PROCEDURE: Serum AGP concentration was measured in the healthy dogs and in the dogs with lymphoma before treatment, 3 weeks after the first dose of doxorubicin was administered, and every 3 weeks thereafter until relapse (i.e., recurrence of clinically detectable disease such as palpable enlargement of peripheral lymph nodes). Serum AGP concentrations were determined by use of a radial immunodiffusion kit. RESULTS: Mean serum AGP concentration in healthy dogs was significantly less than concentration in dogs with lymphoma prior to treatment. Mean serum AGP concentrations after the first and each subsequent dose of doxorubicin were not significantly different from concentration in healthy dogs. However, mean serum AGP concentrations 3 weeks prior to and at the time of relapse were significantly higher than concentration measured after the first dose of doxorubicin, and were not significantly different from concentration measured before treatment. CLINICAL IMPLICATIONS: Results suggest that measuring serum AGP concentration may be a useful method of predicting relapse before recurrence of clinically detectable disease in dogs with lymphoma undergoing treatment with doxorubicin.

Unusual presentation of an Amazon parrot (Amazona species) with hepatocellular carcinoma.

Non-haematopoietic hepatic malignancies are uncommon in birds. The clinical presentation (i.e. chronic buphthalmos)and non-specific radiographic findings observed in this adult Amazon parrot (Amazona spp.) were not consistent with previous reports describing the natural behaviour of metastatic hepatocellular carcinoma in birds.

Acute synovitis and intra-articular methylprednisolone acetate in ponies.

OBJECTIVE: To determine how acute synovitis, with and without intra-articular methylprednisolone acetate (MPA), affect synthesis of proteoglycan, total protein, and collagen in articular cartilage and total protein synthesis in synovial membrane. DESIGN: Synovitis was induced in 10 ponies by the injection of 0.5 ng lipopolysaccharide (LPS) into the left radiocarpal and midcarpal joints every 2 days for a total of four treatments. Synovitis was documented by clinical examination and synovial fluid analyses. Two days before euthanasia, MPA (0.1 mg/kg) was injected with the last dose of LPS into both the left and right radiocarpal and midcarpal joints of five of these ponies. Proteoglycan synthesis in articular cartilage explants from these joints was measured by incorporation of sodium [35S]sulfate. The size of the proteoglycan monomers and their aggregation with hyaluronan was assessed by size-exclusion chromatography. Protein synthesis in articular cartilage was measured by incorporation of [3H]proline and collagen synthesis by conversion of [3H]proline into [3H]hydroxyproline. Protein synthesis was measured in synovial membrane explants by incorporation of [35S]methionine. RESULTS: Ponies developed carpal effusion and mild lameness accompanied by increased total nucleated cell count and total solids in synovial fluid in response to the LPS injections. Moderate to severe synovial membrane proliferation and inflammation were observed histopathologically in joints injected with LPS but no consistent light-microscopical changes were observed in the articular cartilage from these joints. Intra-articular MPA alone was associated with decreased proteoglycan synthesis and increased protein and collagen synthesis in the cartilage explants. Total protein synthesis by synovial membrane was also increased by MPA alone. In contrast, no differences in protein or proteoglycan synthesis were observed in explants from the joints with synovitis, with or without intra-articular MPA. Treatment with MPA, LPS, and LPS/MPA did not alter proteoglycan aggregate size, but LPS-induced synovitis resulted in an increase in the second largest population of monomers. MPA increased the synthesis of small proteoglycan monomers. CONCLUSION: Based on the methods used, acute synovitis prevented changes induced by intra-articular MPA alone. Results suggested that the effect of intra-articular MPA on joint metabolism was different between inflamed and normal joints. Experimental studies must consider the effect of inflammation, as well as the potential to introduce in vitro culture artifacts when investigating the effect of intra-articular corticosteroids on chondrocyte function.

Concentrations of keratan sulfate in plasma and synovial fluid from clinically normal horses and horses with joint disease.

OBJECTIVE: To determine whether keratan sulfate concentrations in plasma or synovial fluid from clinically normal horses were different from concentrations in horses with joint disease and whether concentrations varied with type of joint disease. DESIGN: Case-control study. ANIMALS: 67 clinically normal horses, 10 clinically normal foals, and 160 horses with joint disease. PROCEDURE: ELISA was used to measure keratan sulfate concentrations. RESULTS: Mean plasma keratan sulfate concentration (mean +/- SEM, 580 +/- 124 ng/ml) in foals peaked at 10 weeks of age. Mean plasma keratan sulfate concentration in clinically normal horses was 200 ng/ml (95% confidence interval, 157 to 251 ng/ml). Horses with osteochondral (chip) fractures, other closed intraarticular fractures, inflammatory arthritis (synovitis), infectious arthritis, or osteochondrosis had significantly higher plasma keratan sulfate concentrations than did clinically normal horses, but horses with osteoarthritis did not. Breed, gender, and type of joint disease affected keratan sulfate concentration in synovial fluid. Standard-breds with chip fractures of the metacarpophalangeal/ metatarsophalangeal joints had significantly higher keratan sulfate concentrations in synovial fluid than did Thoroughbreds. Keratan sulfate concentrations in synovial fluid from osteoarthritic carpal joints were lower than concentrations in normal carpal joints and tarsocrural joints with inflammatory joint disease. CLINICAL IMPLICATIONS: Keratan sulfate concentration alone was not a specific marker of joint disease but was affected by various joint diseases.