Comparison of Sysmex XN-V body fluid mode and deep-learning-based quantification with manual techniques for total nucleated cell count and differential count for equine bronchoalveolar lavage samples.

BACKGROUND: Bronchoalveolar lavage (BAL) is a diagnostic method for the assessment of the lower respiratory airway health status in horses. Differential cell count and sometimes also total nucleated cell count (TNCC) are routinely measured by time-consuming manual methods, while faster automated methods exist. The aims of this study were to compare: 1) the Sysmex XN-V body fluid (BF) mode with the manual techniques for TNCC and two-part differential into mononuclear and polymorphonuclear cells; 2) the Olympus VS200 slide scanner and software generated deep-learning-based algorithm with manual techniques for four-part differential cell count into alveolar macrophages, lymphocytes, neutrophils, and mast cells. The methods were compared in 69 clinical BAL samples. RESULTS: Incorrect gating by the Sysmex BF mode was observed on many scattergrams, therefore all samples were reanalyzed with manually set gates. For the TNCC, a proportional and systematic bias with a correlation of r = 0.79 was seen when comparing the Sysmex BF mode with manual methods. For the two-part differential count, a mild constant and proportional bias and a very small mean difference with moderate limits of agreement with a correlation of r = 0.84 and 0.83 were seen when comparing the Sysmex BF mode with manual methods. The Sysmex BF mode classified significantly more samples as abnormal based on the TNCC and the two-part differential compared to the manual method. When comparing the Olympus VS200 deep-learning-based algorithm with manual methods for the four-part differential cell count, a very small bias in the regression analysis and a very small mean difference in the difference plot, as well as a correlation of r = 0.85 to 0.92 were observed for all four cell categories. The Olympus VS200 deep-learning-based algorithm also showed better precision than manual methods for the four-part differential cell count, especially with an increasing number of analyzed cells. CONCLUSIONS: The Sysmex XN-V BF mode can be used for TNCC and two-part differential count measurements after reanalyzing the samples with manually set gates. The Olympus VS200 deep-learning-based algorithm correlates well with the manual methods, while showing better precision and can be used for a four-part differential cell count.

Automated hematological cell count using sysmex XN-1000V in Testudo hermanni: Agreement with manual count.

Mediterranean area represents the main habitat of Testudo hermanni. Clinical signs of disease of these tortoises are non-specific, making the hematology results crucial in revealing underlying pathological conditions. However, accurate automated identification of blood cell populations is hampered by the presence of nucleated erythrocytes (NRBC) and thrombocytes (Thr), necessitating manual methods such as counting chambers. The aim of the study was to assess the performance of the novel automated hematology analyzer Sysmex XN-1000 V, which includes a a specific channel (WNR) for counting NRBC, in accurately identify and quantify the different blood cell populations of Testudo hermanni. Additionally, its agreement with manual counts was evaluated. Fifty heparinized blood samples were initially counted using the Neubauer improved chamber and then analysed twice with Sysmex XN-1000 V. Thirteen out of 50 samples were instrumentally counted again after 48 h to assess the inter-assay precision. All WNR scattergrams were re-analysed using an ad hoc gate panel to differentiate two populations: NRBCs (weak fluorescence signal) and WBC + Thr (high fluorescence signal). Sysmex XN-1000 V demonstrated optimal intra- and inter-assay precision for NRBCs (CV 0.98% ± 1.96; 1.31% ± 2.98) and moderate precision for WBC + Thr (CV 9.24% ± 16.61; 12.69% ± 10.35). No proportional nor constant errors were observed between the methods for both the populations. The instrumental NRBC counts were consistently slightly lower, while WBC + Thr counts were slightly higher compared to manual counts. These findings suggest that Sysmex XN-1000 V can be used for analyzing cell populations in heparinized blood of Testudo hermanni. However, specific instrumental reference intervals are suggested.

A comparison study between the Siemens ADVIA 120 and manual method for the differential white blood cell count in goats.

BACKGROUND: Although widely used, the ADVIA 120 hematology analyzer has not been previously validated for determining the differential leukocyte count in goats. OBJECTIVES: The aim of this study was to compare the differential leukocyte counts provided by the ADVIA 120 (A-diff) and the manual method (M-Diff) in goats. METHODS: EDTA blood samples that were analyzed within 4 h of collection were used in the study. The following exclusion criteria were applied: inappropriately filled tubes or tubes containing clots, erroneous ADVIA peroxidase cytograms, and blood smears of poor quality. The A-Diff was compared with the M-Diff performed by two independent observers on 200 leukocytes. RESULTS: Forty samples were included after previously excluding eight samples. The correlation between the A-Diff and M-Diff was very strong for eosinophils (r = .870, p < .001) and strong for lymphocytes (r = .796, p < .001) and neutrophils (r = .730, p < .001), while no significant correlation was observed for monocytes (r = .026, p = .872). The Passing-Bablok regression analyses revealed statistically significant constant errors for neutrophils (5.83%; 95% confidence interval [CI]: 0.41%, 12.18%) and eosinophils (1.89%; 95% CI: 1.17%, 2.71%). Bland-Altman analyses showed a statistically significant negative bias for lymphocytes (-5.0%) and a statistically significant positive bias for eosinophils (2.2%). The very low basophil percentages precluded a meaningful method comparison. CONCLUSIONS: The ADVIA 120 overall demonstrated good performance for the differential WBC count in goats under the conditions of this study. Therefore, it can be considered suitable for routine hematologic screening in goats. Nonetheless, it should be emphasized that any abnormal result should be confirmed with a blood smear evaluation.

Diagnostic utility of the total nucleated cell count for differentiation of septic and sterile peritoneal effusions in dogs.

BACKGROUND: Rapid and accurate diagnosis of septic peritonitis is critical for initiating appropriate medical and surgical management. OBJECTIVES: The aim of this study was to determine the diagnostic utility of the total nucleated cell count (TNCC), absolute neutrophil count, neutrophil percentage, and total protein (TP) to distinguish septic versus non-septic peritoneal effusions in dogs. METHODS: Electronic medical records were retrospectively searched for peritoneal fluid samples from 2008 to 2018 and classified as septic or non-septic based on bacterial culture and/or cytology results. Receiver operator characteristic curves (ROCs) were used to describe the overall diagnostic utility of each test, with optimal cutpoints analyzed to dichotomize continuous variables. Positive and negative likelihood ratios were calculated at these cutpoints. RESULTS: A total of 166 unique samples, including 87 septic and 79 non-septic peritoneal effusions, were included. There were no significant differences in dog sex, age, or days hospitalized between groups. Septic effusions had significantly higher TP, TNCC, absolute neutrophil count, and neutrophil percentage compared with non-septic effusions. The area under the curve of the ROC curves was TNCC (0.80), absolute neutrophil count (0.80), neutrophil percentage (0.64), and TP (0.63). For TNCC and absolute neutrophil count, optimal cutoffs were 17.13 × 103 cells/µL and 19.88 × 103 cells/µL, resulting in positive and negative likelihood ratios of 2.39 and 0.28 and 2.85 and 0.28, respectively. CONCLUSIONS: Total nucleated cell counts and absolute neutrophil counts aid in the differentiation of septic and non-septic peritoneal effusions with similar diagnostic utility but are not sufficiently sensitive or specific to use without concurrent microscopic evaluation.

Performance evaluation of the Sysmex XN-1000V in side-by-side comparison with the Siemens ADVIA 120 and manual methods for healthy CD Sprague-Dawley rats and CD-1 mice.

BACKGROUND: The Sysmex XN-1000V automated hematology analyzer with multispecies software was released in June 2017 for use in research laboratories. Laser light, impedance, fluorescent staining, and fluorescent flow cytometry are used to analyze whole blood for CBC, reticulocyte counts, and WBC counts, including a 5-part differential leukocyte analysis. OBJECTIVES: A side-by-side comparison of the Sysmex XN-1000V with the Siemens ADVIA 120 in analyzing blood from healthy mice and rats will provide insight into the performance of the new analyzer and its capabilities for use in drug development studies. Method correlation analyses on normal mouse and rat hematology data collected with both analyzers and manual reference methods will help determine the reliability of the data produced using the Sysmex XN-1000V analyzer. METHODS: Whole blood samples collected in K2 EDTA from healthy CD-1 mice and CD Sprague-Dawley rats were analyzed in parallel with the XN-1000V and ADVIA 120 analyzers. Male and female mice, approximately 6-9 weeks old, and male and female rats, approximately 7-9 weeks old, were included in this study. Manual reference methods for WBC differential leukocyte analysis and packed cell volume (PCV) measurements were also performed. EP Evaluator version 11.2 (Data Innovations LLC, South Burlington, VT, USA) was used for method comparison statistical analysis. RESULTS: Most hematologic parameters for naïve mice and rats achieved correlation in the fair to excellent range, with the majority showing very good to excellent correlation with low biases (<11.0%) for cohorts analyzed separately and when cohort data were combined. CONCLUSIONS: The Sysmex XN-1000V Hematology Analyzer provided comparable results to those obtained from the Siemens ADVIA 120. We found the Sysmex XN-1000V Hematology Analyzer to be acceptable for use in drug development studies for rats and mice.

Repeat patient testing-quality control with canine samples shows promise as an alternative to commercial quality control material for a network of four Sysmex XT-2000iV hematology analyzers.

BACKGROUND: Repeat patient testing-quality control (RPT-QC) uses retained patient samples as an alternative to commercial quality control material (QCM). We elected to calculate and validate RPT-QC limits for red blood cell count (RBC), hemoglobin (HBG), hematocrit (HCT), and white blood cell count (WBC). OBJECTIVES: (1) To validate RPT-QC across a network of four harmonized Sysmex XT-2000iV hematology analyzers and determine the total error that can be controlled with RPT-QC. (2) To generate quality control (QC) limits using the standard deviation (SD) of the duplicate measurement differences and determine a suitable simple QC rule with a probability of error detection >0.85 and probability of false rejection <0.05. (3) Monitor RPT-QC using sigma metrics as a performance indicator and (4) to challenge RPT-QC to ensure acceptable sensitivity. METHODS: Fresh adult canine EDTA samples with results within reference intervals were selected and run again on days 2, 3, and 4. QC limits were generated from the SD of the duplicate measurement differences. The QC limits were challenged using interventions designed to promote unstable system performance. The total error detectable by RPT-QC was determined using EZRULES 3 software. RESULTS: In all, 20-40 data points were needed for RPT-QC calculations and validated using 20 additional data points. The calculated limits differed among the network of analyzers. The total error that could be controlled was the same or better than that of the manufacturer's commercially available quality control material using the same analyzer for all measurands except hematocrit, which required a higher total error goal than that proposed by ASVCP guidelines to achieve an acceptable probability of error detection. The challenges designed to mimic unstable system performance were successfully detected as out-of-control QC. CONCLUSIONS: The challenges for RPT-QC resulted in acceptable detection of potential unstable system performance. This initial study demonstrates that RPT-QC limits differ among the network of Sysmex XT-2000iV analyzers, indicating a requirement to customize for the individual analyzer and laboratory conditions. RPT-QC could achieve ASVCP total allowable error goals for RBC, HGB, and WBC, but not for HCT. Sigma metrics were consistently >5.5 for RBC, HGB, and WBC, but not for HCT.

Pathogen and severity-dependent immune responses in bovine mastitis: highlight the dynamics of differential somatic cell count.

Immune responses in bovine clinical mastitis (CM) probably differ depending on the causative pathogen and disease severity. The observational study aimed to investigate whether both factors are associated with the dynamics of immune indicators, including somatic cell score (SCS), white blood cell count (WBC), serum albumin/globulin (A/G) ratio, and differential somatic cell count (DSCC). We collected blood and milk samples 0, 3, 5, 7, 14, and 21 days after CM occurred in 38 cows, and grouped the cases (n=49) by disease severity and pathogen. We analyzed data using a linear mixed model considering the effects of pathogens and severity, calculated estimated-marginal means for indicators at each time point, and compared the means between groups. The dynamics of WBC varied depending on both pathogen and severity. WBC changed drastically in either severe or coliform-caused CM, slightly elevated in streptococcal mastitis, but unchanged in staphylococcal mastitis. This possibly relates to the deficiency in innate immune response toward staphylococci. The A/G ratio also changed depending on severity, as it dropped sharply only in severe CM. We observed a non-linear relationship between DSCC and SCS, possibly due to mammary epithelial cells shedding in milk when CM occurred. When cows recovering from Streptococcus dysgalatiae mastitis, DSCC decreased while SCS remained high, suggesting a healing process requiring more macrophages. Our results demonstrate that both the severity and pathogen are associated with immune responses in CM, providing insights into mastitis pathogenesis.

Validation of a hematology analyzer in donkey medicine.

Asinina de Miranda is a protected donkey sub-species from the Mirandês plateau in northeastern of Portugal. Donkeys are animals that have substantially lost their place as working animals in modern society, this had led to a decrease in their population numbers. A need to preserve native species has led to the foundation of organizations like Associação para o Estudo e Proteção do Gado Asinino (AEPGA) and the development of studies regarding breed welfare, such as hematology. The IDEXX ProCyte Dx is a veterinary hematology analyzer validated for several species, but not for donkeys. The aim of this study was to validate the ProCyte Dx for Asinina de Miranda donkeys. The validation requires a controlled study of precision, carryover, linearity and comparison between the equipment and the manually obtained values for the leukocyte differential count and hematocrit. Results indicated coefficient of variation was good (below 5 %) for both the intra-assay and the inter-assay precision, except for basophils. Carryover was 0 % for all the parameters except platelets (5.88 %). Linearity showed a very high Pearson correlation coefficient, above 0.99, for erythrocytes, hematocrit, hemoglobin, leucocytes, neutrophils, lymphocytes, monocytes, eosinophils, platelets and plateletcrit. Comparison demonstrated excellent agreement for hematocrit (rs=0.96) and good Spearman rank correlation for neutrophils (rs=0.84) and lymphocytes (rs=0.90). Accuracy for total leukocyte count and platelets could not be determined. In conclusion, the ProCyte Dx seems appropriate to be used in Asinina de Miranda hematology.

Retrospective evaluation of hematological ratios in canine parvovirosis: 401 cases.

BACKGROUND: The utility of neutrophil-to-lymphocyte ratio (NLR), platelet to-lymphocyte ratio (PLR) and monocyte-to-lymphocyte ratio (MLR) as prognostic indicators has not been investigated in canine parvovirosis (CPV). HYPOTHESIS: To evaluate whether these hematological ratios obtained at hospital admission in CPV are associated with outcome or duration of hospitalization. ANIMALS: Four hundred one client-owned dogs presented with CPV. Methods-Retrospective multicenter cohort study. Medical records were reviewed to identify dogs with CPV. Data regarding signalment, complete blood count at admission, duration of hospitalization and outcome were collected. RESULTS: Of the 401 dogs included in the study, 336 (83.8%) survived to discharge. The median (25th and 75th percentiles) PLR in nonsurvivors (336.56 [159.84-635.77]) was significantly higher than in survivors (217.65 [117.67-389.65]) (P = .003). The area under the receiver-operating characteristic curve for nonsurvival was 0.615 (95% CI [0.593-0.691], P = .003). A cut off of 700 showed a 21.5% sensitivity and 90% specificity for nonsurvival. No association was observed between hospitalization duration and either hematological ratios or total WBC counts. The median (25th and 75th percentiles) lymphocyte count was below reference interval in all dogs and was significantly lower in the dogs which died (0.82 × 109 /L [0.5-1.87]) than in survivors (1.27 × 109 /L [0.73-2.22]) (P = .005). The median (25th and 75th percentiles) monocyte count however was lower in survivors (0.38 × 109 /L [0.29-1.59]), than in nonsurvivors (0.73 × 109 /L [0.1-2]) (P = .002). CONCLUSIONS: Evaluation of PLR at hospital admission might be a useful marker of disease severity and could have prognostic value in dogs with CPV.

Haematological reference intervals for pregnant Icelandic mares on pasture.

BACKGROUND: Few studies have been conducted on haematological reference intervals (RIs) in Icelandic horses. Reference intervals have been published for Icelandic horses in Austria and a preliminary study in Iceland compared haematological values in riding horses to published RIs for other breeds as well as Icelandic horses abroad. Haematological parameters can vary greatly due to factors such as breed, gender, age, reproductive status, and training, as well as feeding, prior exercise and management method. Icelandic broodmares are kept on pasture under supervision throughout the year, with haylage provided during the winter, and it is therefore of interest to establish haematological reference intervals for pregnant broodmares in Iceland. The purpose of this study was to establish haematological RIs specific to Icelandic broodmares in the first months of pregnancy, kept on pasture. Blood samples from 183 mares, stabilised in EDTA were analysed using IDEXX ProCyte Dx and total protein was analysed in serum samples from 157 of the 183 mares, using IDEXX Catalyst One analyser. The RIs were established using the guidelines of the American Society for Veterinary Clinical Pathology. RESULTS: The RIs for red blood cell count, haematocrit and haemoglobin were higher in pasture-kept Icelandic mares in early pregnancy, most of which were lactating, than in pregnant mares of other breeds. This was also true for white blood cell count, as well as numbers of monocytes, eosinophils, and basophils, which in some instances might illustrate problems in the automated categorisation of some leukocytes. CONCLUSIONS: As no RIs have been published for other pasture-kept Icelandic horses, future investigations should include other groups of pasture-kept Icelandic horses. Such an analysis might elucidate the effect of breed, management, and pregnancy on haematological values in pasture-kept Icelandic horses.

Evaluation of canine and feline leukocyte differential counts obtained with the scil vCell 5 compared to the Advia 2120 hematology analyzer and a manual method.

The vCell 5 (scil Animal Care), a point-of-care hematology analyzer (POCA), was recently introduced to veterinary laboratories. This laser- and impedance-based analyzer is capable of providing a CBC with 5-part WBC differential count (Diff) along with WBC cytograms and flags serving as interpretation aids for numerical results. We compared the scil POCA-Diff to reference methods (i.e., manual differential count, Advia 2120 hematology analyzer [Siemens]) for canine and feline blood samples and considered WBC cytograms and flags. Total observed error (TEo), calculated from CV and bias%, was compared to total allowable error (TEa). Data were analyzed before and after a review process (exclusion of flagged and samples with invalid cytograms). For both species, correlation was good-to-excellent (rs = 0.81-0.97) between both analyzers for all variables, except for feline monocytes (rs = 0.21-0.63) and canine monocyte% (rs = 0.50). Smallest biases were seen for neutrophils (dog: -5.7 to 0.8%; cat: 1.5-9.4%) with both reference methods. Quality requirements (TEo < TEa) were fulfilled for canine and feline neutrophils (TEo = 5.3-10.6%, TEa = 15%) and eosinophils (TEo = 67.1-83%, TEa = (90)-50%) considering at least one reference method. Our review process led to mildly higher rs-values for most variables. Although not completely satisfactory, the scil POCA provides reliable results in compliance with ASVCP quality goals for canine and feline neutrophils and eosinophils. Analyzer flag and cytogram analysis served as useful tools for QA, indicating the necessity for manual review of blood smears, and contributed to improvement of scil POCA performance.

Effect of storage on the nitro blue tetrazolium reduction test in dog blood samples.

INTRODUCTION: The nitro blue tetrazolium (NBT) reduction test (NBTT) has been used for measuring the metabolic activity of phagocytes of mammals. Activated neutrophils transform NBT into formazan in the cytoplasm. The NBTT can detect the activation of neutrophils in peripheral blood and is used to assess neutrophil function in dogs. However, the NBTT is not used frequently in the clinical setting, as samples should be processed after blood collection. OBJECTIVE: The aim of this study was to evaluate the effect of storage on NBTT in dog blood samples. MATERIALS AND METHODS: Residual EDTA blood samples from 22 dogs were included of different ages, breeds, and sex. The buffy coat layer was separated from the blood and incubated with 0.1% NBT. The NBTT was performed at 0, 24, 48, and 72 h after the collection of blood. Blood samples were stored at 4°C until the tests were performed. Blood smears were evaluated by light microscopy, and the NBT reduction rate was reported, which represents the percentage of activated neutrophils. The NBT reduction rate was calculated after counting 300 neutrophils in each slide. RESULTS: The means of NBTT in dog blood samples at 0, 24, 48, and 72 h were 8.3%, 8.5%, 8.7%, and 7.8%, respectively. No significant differences were observed between time points. CONCLUSIONS: This study showed that the NBTT can be performed up to 72 h after the collection of canine blood if correctly refrigerated at 4°C. This finding supports the performance of the NBTT in the clinical setting.

Haematological values in cattle reared in humid and subhumid tropics of Mexico.

The objective of this study was to determine some factors that influence the haematological values of cattle reared in the humid and subhumid tropics of Mexico. Whole blood samples were taken from 1355 crossbred cattle in the years 2017 to 2019. Haematocrit (HTC, %), total plasma protein (TPP, g/dL) and peripheral eosinophils count (PEOS, × 103/µL) were determined manually, and the main haematological variables were recorded with an automatic analyser. The statistical analysis considered as classification variables age, sex, season (cold, dry and rainy), year (2017, 2018 and 2019) and origin of the cattle. The mean of the haematological parameters was determined along with the confidence limits (CL) of the different categories of animals according to age. Calves younger than 1-year-old presented higher levels of HTC, red blood cell count (RBC), haemoglobin (HGB), red blood cell distribution width (RDW), platelet number (PLT), white blood cell count (WBC) and lymphocyte count (LYMF) than animals older than 2 years of age. However, they showed the lowest mean cell volume (MCV) and TPP values. In cows, the highest levels of PEOS, granulocytes (GRAN), MCV and medium cells (MID) were observed and the lowest HTC, RBC, RDW and WBC levels. Intervals were determined with the 1st quartile (Q1) or lower confidence interval (90% CI) as the minimum values and the 3rd quartile (Q3) or upper confidence interval (90% CI) as the maximum values. The haematological parameters of cattle reared in the Southeast of Mexico are significantly affected by age, sex and environmental conditions.

Agreement Study between Total Leukocyte Count Methodologies in South American Sea Lions (Otaria byronia) and Peruvian Fur Seals (Arctocephalus australis).

South American sea lions (Otaria byronia) and Peruvian fur seals (Arctocephalus australis) are sympatric species inhabiting the coastal Peruvian marine ecosystem. Declining abundance has prompted population health monitoring programs, including temporal monitoring of blood parameters. Several methods exist to determine total leukocyte count, yet no studies have evaluated agreement between methods in pinnipeds. We assessed agreement between total leukocyte counts determined by blood film estimate, Leuko-TIC, HemoCue, and UNOPETTE methods by using archival results from pinnipeds at Punta San Juan, Peru. Blood film estimates were prospectively performed, and resulting data were compared with retrospective leukocyte counts obtained from both species between 2009 and 2019 by using the other methodologies. Agreement in hematologic counts between methods was evaluated using Passing-Bablok regression and Bland-Altman plots (α=0.05). In total, 295 individuals (201 A. australis and 94 O. byronia) were included in the analysis. The blood film estimate method resulted in the highest leukocyte values (P<0.0001). Leuko-TIC counts were significantly higher than HemoCue counts (P<0.0001). Constant and proportional error was present in the agreement between the blood film estimate method and the other methods. Given the variation demonstrated between the different methodologies, additional research is needed to further evaluate agreement between these methodologies. The results underscore the importance of maintaining consistency in leukocyte count methodology for monitoring trends in population health over time. Method consistency may be the more important clinical consideration for assessing changes in leucocyte count over time and avoiding apparent changes depending on the methodology used.

Case of mastocytemia and systemic mastocytosis in a dog: ProCyte Dx and Sysmex XT-2000i scattergram findings, morphologic features, and c-kit somatic mutation analysis.

A 10-year-old female Golden Retriever was presented for a recheck after the complete removal of low-grade complex mammary carcinoma. The in-house ProCyte Dx automated counts revealed moderate regenerative anemia and moderate eosinophilia. The ProCyte Dx WBC scattergram showed a cloud in an unusual place parallel and to the right of the monocyte dot plot location. Cells were classified as either monocytes or neutrophils with no clear separation. Complete blood count analysis performed in the laboratory on a Sysmex XT-2000iV analyzer showed moderate regenerative anemia and WBC count within RI; a differential count was not provided by the instrument. On the Sysmex XT-2000iV DIFF scattergram, neutrophil and eosinophil dot plots were present at the respective locations and appeared separated, but the instrument did not provide numerical results. In addition to the normal lymphocyte dot plot location, the second cloud of cells classified as lymphocytes was displayed to the right of the monocyte dot plot area. On the WBC/BASO scattergram, the second population of cells was present above and to the right of the leukocyte cluster. Morphologic assessment of the blood smear detected mastocytemia with 16% poorly granulated and degranulated mast cells. FNAs from the liver and spleen contained large aggregates of poorly granulated mast cells. C-kit somatic mutation screening detected the presence of point mutation S479I in exon 9 of the canine c-KIT gene. This is the first description of abnormal scattergrams from ProCyte Dx and Sysmex XT-2000iV analyzers in a dog with concurrent mastocytemia and systemic mastocytosis, and where cytologic assessments of a blood smear, liver, and spleen, and c-kit somatic mutation analysis were performed.

Myeloperoxidase enzyme and Ferric-reducing antioxidant power concentrations in lean and obese dogs.

The study aimed to assess plasma Myeloperoxidase enzyme (MPO) and Ferric-reducing antioxidant power (FRAP) in obese dogs and compare them with ideal body weight dogs. Thirty-two dogs were distributed into two groups according to a 9-point body condition score (BCS), as follows: Control group (n = 16), dogs with a BCS of 4 or 5; Obese group (n = 16), dogs with a BCS of 8 or 9. Plasma MPO and FRAP assays, neutrophil count, lipid profile (cholesterol and triglycerides), and systolic blood pressure (SBP) were evaluated in both groups. The duration of obesity was defined based on history. The obese group showed higher values for body weight, BCS, SBP, neutrophil count, triglycerides, and MPO than the Control group. A positive correlation was observed between MPO concentrations and BCS and body weight. FRAP concentrations showed a positive correlation with the duration of obesity. The results suggested that an inflammatory state caused by obesity may promote increased neutrophil count and MPO concentrations, besides the positive correlation between MPO with BCS and body weight. The obesity in dogs promoted slight active MPO elevation, influenced by body weight, BCS, and neutrophil count. The FRAP assay did not show the expected reduction and, therefore, needs further investigation.

Comparison of six microscopic methods and two operators for estimation of white and red blood cells in canine urine sediment.

BACKGROUND: Little information is currently available about the analytical variability of urinalysis. OBJECTIVE: We aimed to compare results obtained by two operators using six microscopic methods in the quantification of urinary leukocytes (WBC) and erythrocytes (RBC). METHODS: Forty urine samples (10 mL) were centrifuged (450g, 5 minutes) and resuspended in 0.5 mL of supernatant. Two operators with different expertise in urinalysis interpreted sediment results using the six methods, obtained by combining the use of microscope slides (Slide) or counting chambers (Chamber) with three different techniques: bright-field (BF) microscopy, phase-contrast (PC) microscopy, and stained sediment (SS) evaluations. The mean WBC and RBC counts from 10 fields (Slide) or squares (Chamber) observed at 400× were used to calculate the difference and agreement between operators and methods. We also estimated the concordance between methods in classifying microhematuric or pyuric samples. RESULTS: Operator 2 counted significantly lower WBC counts using Slide+BF (P = 0.009) and Slide+PC (P = 0.001) than Operator 1, whereas no inter-operator differences were recorded for RBC counts. The concordance between the operators ranged from "good" to "very good." No differences or biases were found for WBC counts among the methods, and concordances were "good" to "very good"; proportional biases were found for RBC counts between Slide+BF vs Slide+SS and Slide+PC vs Slide+SS. Concordance measurements for RBC counts ranged from "good" to "very good." CONCLUSIONS: All methods yielded good reproducibility among operators, although stained SS evaluations allowed better identification of WBC by the inexperienced operator. However, we suspected that the SS preparations affected RBC counts. All other methods yielded reproducible WBC and RBC counts.

Establishment of preliminary reference intervals and cytochemical staining of blood cells in big-bellied seahorses (Hippocampus abdominalis).

BACKGROUND: Despite their popularity, hematology reference intervals (RIs) have not been established in big-bellied seahorses (Hippocampus abdominalis). OBJECTIVE: The objectives of this study were to establish hematologic RIs to compare values between sex in regard to cytochemical staining of blood cells. We also sought to compare white blood cell concentrations using the Natt and Herrick technique vs blood smear estimates. METHODS: Forty-three healthy individuals from the Aquarium du Québec (22 females and 21 males) were included. Normal health status was confirmed by an unremarkable physical examination in five individuals and by necropsy of five other individuals, of which all were excluded from further analyses. Venipuncture was performed from the ventral coccygeal vein in the remaining 33 individuals without anesthesia using heparinized insulin syringes. A blood volume of 0.05 to 0.1 ml was collected to prepare Wright Giemsa-stained blood smears and hematocrits immediately after venipuncture. Whole blood was stored in heparinized Eppendorf tubes to determine red and white blood cell concentrations using the Natt and Herrick technique with a hemocytometer in 10 individuals; these results were compared with blood smear estimates. Additional blood smears were stained with alkaline phosphatase substrate, periodic acid Schiff, and toluidine blue stains. RESULTS: The reference intervals included the packed cell volume (27.4-67.5%), thrombocyte count (19.5-197.7 × 109 /L), and white blood cell (WBC) count (2-54.8 × 109 /L), including neutrophils (1.1-21.3 × 109 /L), lymphocytes (2.7-45.5 × 109 /L), and monocytes (0-2.2 × 109 /L). The WBC hemocytometer counts showed no correlation with blood smear estimates (Spearman's rho = 0.2). There was also no significant difference between the sexes. CONCLUSIONS: These preliminary reference intervals will help assess the health of seahorses.

Use of serum amyloid A in equine medicine and surgery.

Serum amyloid A (SAA) has become an indispensable part of the management of equine patients in general practice and specialized hospital settings. Although several proteins possess acute phase properties in horses, the usefulness of SAA exceeds that of other acute phase proteins. This is due to the highly desirable kinetics of the equine SAA response. SAA concentrations exhibit a rapid and pronounced increase in response to inflammation and a rapid decline after the resolution of inflammation. This facilitates the detection of inflammatory disease and real-time monitoring of inflammatory activity. SAA may be used in all stages of patient management: (1) before diagnosis (to rule in/rule out inflammatory disease), (2) at the time of diagnosis (to assess the severity of inflammation and assist in prognostication), and (3) after diagnosis (to monitor changes in inflammatory activity in response to therapy, with relapse of disease, or with infectious/inflammatory complications). By assessing other acute phase reactants in addition to SAA, clinicians can succinctly stage inflammation. White blood cell counts and serum iron concentration change within hours of an inflammatory insult, SAA within a day, and fibrinogen within 2-3 days; the interrelationship of these markers thus indicates the duration and activity of the inflammatory condition. Much research on the equine SAA response and clinical use has been conducted in the last decade. This is the prerequisite for the evidence-based use of this analyte. However, still today, most published studies involve a fairly low number of horses. To obtain solid evidence for use of SAA, future studies should be designed with larger sample sizes.

Hematologic Reference Intervals and Comparison of Natt-Herrick Technique and Smear-Based Leukocyte Estimation in Cockatiels (Nymphicus hollandicus).

Although cockatiels are among the most common avian species maintained as companion animals in the United States, information on standard hematologic reference values for this species is limited. The objectives of this study were to establish hematologic reference intervals (RI) for cockatiels, compare methods using both the Natt-Herrick technique (NHT) and the smear-based estimation technique (SBT), explore age and sex differences in the hematologic findings for this species, and produce the first cockatiel RI for fibrinogen concentration and thrombocyte estimate. Healthy cockatiels (60 males and 60 females, 2-11 years old) from a research colony were included in this study. Blood samples were placed in dipotassium ethylenediaminetetraacetic acid tubes, and erythrocyte counts and thrombocyte estimates were determined via automated analyzer (ADVIA 120) and SBT, respectively. Moreover, leukocyte concentrations were determined using both NHT and SBT to compare these common methods for measuring a complete blood count in cockatiels. Data were analyzed for outliers, distributions, descriptive statistics, and RI via Reference Value Adviser, a set of macroinstructions for Microsoft Excel (Microsoft, Redmond, WA, USA). Lymphocytes were the predominant leukocyte across both methods. According to the NHT, females had significantly higher concentrations of total leukocytes, heterophils, bands, lymphocytes, basophils, and total plasma protein compared with males. Significant inverse polynomial relationships were noted between total leukocyte count and age and lymphocyte counts and age for NHT. Total leukocyte count produced via NHT and SBT were compared using Passing-Bablok and Bland-Altman plots, and no significant constant or proportional biases were found. However, these methods showed wide limits of agreement. While the RI were interchangeable between methods from a clinical standpoint, the same method should be used to assess changes in an individual. The reported RI are uniquely robust given the sample size, balanced sex and age distributions, inclusion criteria, and control over sample collection.