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.

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.

Erysipelothrix rhusiopathiae-specific T-cell responses after experimental infection of chickens selectively bred for high and low serum levels of mannose-binding lectin.

Erysipelas, caused by infection with Erysipelothrix rhusiopathiae (ER) is an important emerging disease in laying hens. We have earlier observed prominent mannose-binding lectin (MBL) acute phase responses in experimentally ER infected chickens. The present study aimed to further examine immune responses to ER by using chickens selectively bred for high (L10H) and low (L10L) serum MBL levels. Chickens were infected with ER at 3 weeks of age and immune parameters and bacterial load were monitored in blood until day 18 after infection. Blood and spleen leukocytes collected on day 18 were stimulated in vitro with ER antigens and blast transformation of different T-cell populations was assessed. The ER infection gave a very varied outcome and no clear differences were observed between L10H and L10L chickens with respect to leukocyte counts, bacterial load or clinical outcome. Nonetheless, rapid innate responses, e.g., heterophilia and increased serum MBL levels were noted in bacteraemic chickens. All ER infected chickens also showed transient increased expression of mannose receptor MRC1L-B and decreased expression of major histocompatibility complex II on monocytes day 1 after infection indicating monocyte activation or relocation. In vitro ER stimulation showed antigen specific blast transformation of CD4+, TCRγ/δ-CD8αß+ and TCRγ/δ+CD8αß+ spleen cells from all infected chickens. For CD4+ and TCRγ/δ-CD8αß+ cells the proportions of blast transformed cells were significantly higher for samples from L10L chickens than those for samples from L10H chickens. This is the first observation of ER-specific T-cells in chickens and interestingly a Th1-type response comprising cytotoxic T-cells was indicated.

Immune cell composition of the bronchoalveolar lavage fluid in healthy and respiratory diseased dromedary camels.

BACKGROUND: Respiratory diseases are among the most common and expensive to treat diseases in camels with a great economic impact on camel health, welfare, and production. Bronchoalveolar lavage fluid (BALF) has been proven as a valuable sample for investigating the leukocyte populations in the respiratory tract of several species. In the present study, fluorescent antibody labeling and flow cytometry were used to study the immune cell composition of BALF in dromedary camels. Animals with clinical respiratory diseases (n = seven) were compared with apparently healthy animals (n = 10). In addition, blood leukocytes from the same animals were stained in parallel with the same antibodies and analyzed by flow cytometry. RESULTS: Camel BALF macrophages, granulocytes, monocytes, and lymphocytes were identified based on their forward and side scatter properties. The expression pattern of the cell markers CD172a, CD14, CD163, and MHCII molecules on BALF cells indicates a similar phenotype for camel, bovine, and porcine BALF myeloid cells. The comparison between camels with respiratory disease and healthy camels regarding cellular composition in their BALF revealed a higher total cell count, a higher fraction of granulocytes, and a lower fraction of macrophages in diseased than healthy camels. Within the lymphocyte population, the percentages of helper T cells and B cells were also higher in diseased than healthy camels. The elevated expression of the activation marker CD11a on helper T cells of diseased camels is an indication of the expansion of helper T cells population due to infection and exposure to respiratory pathogens. The higher abundance of MHCII molecules on BALF macrophages from diseased camels indicates a polarization toward an inflammatory macrophage phenotype (M1) in respiratory diseased camels. No significant differences were observed in the systemic leukogram between healthy and diseased animals. CONCLUSIONS: Collectively, the current study represents the first report on flow cytometric analysis of immune cell composition of bronchoalveolar lavage fluid (BALF) in dromedary camels.

BASELINE HEMATOLOGIC AND BIOCHEMICAL VALUES AND CORRELATIONS TO ENVIRONMENTAL PARAMETERS IN MANAGED JAPANESE SPIDER CRABS (MACROCHEIRA KAEMPFERI).

Although the evaluation of hematologic and biochemical parameters is a well-established diagnostic tool in vertebrate medicine, comprehensive understanding of these parameters in invertebrate species is lacking. This study provides baseline hemocyte concentrations and biochemistry values for a population of managed Japanese spider crabs (JSC; Macrocheira kaempferi) housed at six different public aquariums. The methodology for obtaining diagnostic hemolymph samples is described. Distinct hemocyte types were identified, including hyaline cells, semigranulocytes, and granulocytes, with hyaline cells as the predominant type. Correlates to exam findings and environmental parameters were evaluated and included higher absolute semigranulocyte counts (r = 0.65, P = 0.020) and triglyceride levels (r = 0.44, P = 0.014) in JSC with exoskeletal lesions; higher total protein (mean = 5.93 g/dl, P = 0.028), cholesterol (median = 18.5 mg/dl, P = 0.018), triglyceride (median = 15.5 mg/dl, P = 0.002), and amylase (median = 243 U/L, P = 0.013) in nonmolting JSC compared with JSC that have previously molted since acquisition (total protein mean = 4.83 g/dl, cholesterol median = 14 mg/dl, triglyceride median = 6.4 mg/dl, and amylase median = 131 U/L); and lower relative and absolute granulocyte counts (mean = 8.83% P = 0.030, median = 1,162 cells/µl P = 0.006, respectively) and higher albumin (median = 1.35 g/dl, P = 0.031) in JSC housed at facilities that used ozone sterilization. The data presented serve as a foundation for understanding basic clinical parameters in JSC hemolymph, as well as the potential influence of environmental stressors on those parameters.

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.

Inflammation and its association with oxidative stress in dogs with heart failure.

BACKGROUND: Inflammation and oxidative stress can contribute to the development and progression of heart failure. This study aimed to investigate the association between inflammatory and oxidative stress markers in dogs with congestive heart failure (CHF). Associations between the disease severity marker N-terminal pro-B-type natriuretic peptide (NT-proBNP) and markers of inflammation and oxidative stress were also determined. RESULTS: Thirty-seven dogs with cardiovascular diseases (dilated cardiomyopathy, DCM (16 dogs), myxomatous mitral valve disease, MMVD (21 dogs)) and ten healthy dogs were included in this prospective study. The patients were further divided into groups with (26) and without CHF (11). We found a significantly higher serum concentration of C-reactive protein (P = 0.012), white blood cell (P = 0.001), neutrophil (P = 0.001) and monocyte counts (P = 0.001) in patients with CHF compared to control dogs. The concentration of tumor necrosis factor-alpha (TNF-α) was significantly higher in patients with CHF compared to patients without CHF (P = 0.030). No significant difference was found in most of the measured parameters between MMVD and DCM patients, except for glutathione peroxidase (GPX) and NT-proBNP. In patients with CHF, TNF-α correlated positively with malondialdehyde (P = 0.014, r = 0.474) and negatively with GPX (P = 0.026, r = - 0.453), and interleukin-6 correlated negatively with GPX (P = 0.046, r = - 0.412). NT-proBNP correlated positively with malondialdehyde (P = 0.011, r = 0.493). In patients without CHF none of the inflammatory and oxidative stress markers correlated significantly. Furthermore, in the group of all cardiac patients, GPX activity significantly negatively correlated with NT-proBNP (P = 0.050, r = - 0.339) and several markers of inflammation, including TNF-α (P = 0.010, r = - 0.436), interleukin-6 (P = 0.026, r = - 0.382), white blood cell (P = 0.032, r = - 0.369), neutrophil (P = 0.027, r = - 0.379) and monocyte counts (P = 0.024, r = - 0.386). CONCLUSION: Inflammatory and oxidative stress markers are linked in canine CHF patients, but not in patients without CHF. These results suggest complex cross communication between the two biological pathways in advanced stages of CHF.

Evaluation of hematological alteration of vector-borne pathogens in cats from Bangkok, Thailand.

BACKGROUND: Cats can be carriers of infected arthropods and be infected with several vector-borne pathogens (VBPs) but there is limited knowledge about their pathogenic role in cats. This study aimed to assess the prevalence of some feline vector-borne agents by molecular technique and to characterize the hematological findings associated with these infections in a cat population from Bangkok Thailand. RESULTS: PCR was positive with at least one pathogen in 237 out of 372 subjects (63.7%), with prevalence of 39.5% (147/372) for Babesia spp., 36.9% (137/372) for hemoplasmas and 3.2% (12/372) for Hepatozoon spp. The cats older than 1 year were at significantly greater risk for VBPs infection (P = 0.001; OR = 1.43; 95% CI: 1.12 - 1.81) and hemoplasmas infection (χ2 = 10.8, df = 1; P < 0.0001; OR = 2.45; 95% CI: 1.49 - 4.01). A significant association between hematological findings and hemoplasma infection were identified in the present study. Besides, VBPs infection revealed more frequent in male cats (χ2= 6.38, df = 1, P = 0.01). Macrocytic hypochromic type of anemia was observed in cats infested with blood-sucking arthropods compared to the non-infested cats presented. CONCLUSIONS: The current study confirmed that Babesia, Hepatozoon and hemoplasmas had infected semi-domesticated cats in Bangkok, Thailand, with Babesia and hemoplasmas being dominant in prevalence. Some hematological findings were significantly associated with cats infected with vector-borne pathogens in this study including leukocyte count and platelets count that may help support veterinary technicians in diagnosis and appropriate treatment. Campaigns of VBPs monitoring in Bangkok emphasizing on the investigation of vectors and possible routes of the infection in animals should be conducted to prevent the transmission of the pathogens.

Canine Parvovirus Infections in Taiwanese Pangolins (Manis pentadactyla pentadactyla).

Canine parvovirus type 2 (CPV-2) is among the most important and highly contagious pathogens that cause enteric or systemic infections in domestic and nondomestic carnivores. However, the spillover of CPV-2 to noncarnivores is rarely mentioned. Taiwanese pangolins (Manis pentadactyla pentadactyla) are threatened due to habitat fragmentation and prevalent animal trafficking. Interactions between Taiwanese pangolins, humans, and domestic animals have become more frequent in recent years. However, information about the susceptibility of pangolins to common infectious agents of domestic animals has been lacking. From October 2017 to June 2019, 4 pangolins that were rescued and treated in wildlife rescue centers in central and northern Taiwan presented with gastrointestinal signs. Gross and histopathological examination revealed the main pathologic changes to be necrotic enteritis with involvement of the crypts in all intestinal segments in 2 pangolins. By immunohistochemistry for CPV-2, there was positive labeling of cryptal epithelium throughout the intestine, and immunolabeling was also present in epidermal cells adjacent to a surgical amputation site, and in mononuclear cells in lymphoid tissue. The other 2 pangolins had mild enteritis without crypt involvement, and no immunolabeling was detected. The nucleic acid sequences of polymerase chain reaction (PCR) amplicons from these 4 pangolins were identical to a Chinese CPV-2c strain from domestic dogs. Quantitative PCR revealed a higher ratio of CPV-2 nucleic acid to internal control gene in the 2 pangolins with severe intestinal lesions and positive immunoreactivity. Herein, we present evidence of CPV-2 infections in pangolins.

Expression-based analysis of genes related to single nucleotide polymorphism hits associated with bovine leukemia virus proviral load in Argentinean dairy cattle.

In dairy cattle infected with bovine leukemia virus (BLV), the proviral load (PVL) level is directly related to the viral transmission from infected animals to their healthy herdmates. Two contrasting phenotypic groups can be identified when assessing PVL in peripheral blood of infected cows. A large number of reports point to bovine genetic variants (single nucleotide polymorphisms) as one of the key determinants underlying PVL level. However, biological mechanisms driving BLV PVL profiles and infection progression in cattle have not yet been elucidated. In this study, we evaluated whether a set of candidate genes affecting BLV PVL level according to whole genome association studies are differentially expressed in peripheral blood mononuclear cells derived from phenotypically contrasting groups of BLV-infected cows. During a 10-mo-long sampling scheme, 129 Holstein cows were phenotyped measuring anti-BLV antibody levels, PVL quantification, and white blood cell subpopulation counts. Finally, the expression of 8 genes (BOLA-DRB3, PRRC2A, ABT1, TNF, BAG6, BOLA-A, LY6G5B, and IER3) located within the bovine major histocompatibility complex region harboring whole genome association SNP hits was evaluated in 2 phenotypic groups: high PVL (n = 7) and low PVL (n = 8). The log2 initial fluorescence value (N0) transformed mean expression values for the ABT1 transcription factor were statistically different in high- and low-PVL groups, showing a higher expression of the ABT1 gene in low-PVL cows. The PRRC2A and IER3 genes had a significant positive (correlation coefficient = 0.61) and negative (correlation coefficient = -0.45) correlation with the lymphocyte counts, respectively. Additionally, the relationships between gene expression values and lymphocyte counts were modeled using linear regressions. Lymphocyte levels in infected cows were better explained (coefficient of determination = 0.56) when fitted a multiple linear regression model using both PRRC2A and IER3 expression values as independent variables. The present study showed evidence of differential gene expression between contrasting BLV infection phenotypes. These genes have not been previously related to BLV pathobiology. This valuable information represents a step forward in understanding the BLV biology and the immune response of naturally infected cows under a commercial milk production system. Efforts to elucidate biological mechanisms leading to BLV infection progression in cows are valuable for BLV control programs. Further studies integrating genotypic data, global transcriptome analysis, and BLV progression phenotypes are needed to better understand the BLV-host interaction.

The relationship of bovine milk somatic cell count to neutrophil level in samples of cow's milk assessed by an automatic cell counter.

This research communication describes the application of a fluorescent automatic cell counter Lactoscan SCC for simultaneous determination of somatic cell count and neutrophils in bovine milk. The obtained results were compared with results obtained by a flow cytometer and a light microscope. The Pearson correlations between the methods were calculated. A comparison between the main characteristics of the three kinds of analysis was made - the assay duration and the intra-assay precision. A relation between the SCC and neutrophil cells was observed in 55 milk samples. The obtained results confirm that the simultaneous determination of SCC and neutrophil analysis are necessary and support the early diagnosis of mastitis, the timely treatment of the animal and the avoidance of major economic losses.

Neutrophil and CD4+ milk cell count related to natural incidence of mastitis in Jersey cattle.

This Research Communication describes the relation between somatic cells and microbial content in milk from Jersey cattle. Milk samples were classified in groups: healthy, dirty and mastitic (from Staphylococcus spp., Escherichia coli, Coliforms). The somatic cells in each of those groups were analysed by two methods - flow cytometric and automatic fluorescent cell counting. Those methods were compared. Total somatic cell count (SCC), neutrophil count, and lymphocytes with cluster of differentiation 4 (CD4+cells) were determined. There was a positive relationship between microbes and somatic cells. It was noticed that the neutrophil count was generally increased together with SCC, whilst the CD4+ cell count was higher in healthy milk samples (about 8%) compared to mastitic ones (about 3%). Lower number of CD4+ cells (from 1 to 4%) was determined in samples positive for Staphylococcus spp. but with lower SCC (from 2.7 to 4.0 × 105 cells/ml). Also, the number of CD4+ cells in Staphylococcus spp.-positive samples increased (to 4.8%) together with higher SCC, something that was not observed in the other mastitic samples. Knowledge of those relations could be useful for veterinary medical tests in the initial phase of inflammation.

EFFICACY OF HUMAN RECOMBINANT GRANULOCYTE COLONY-STIMULATING FACTOR (G-CSF, FILGRASTIM; NEUPOGEN®) IN NEUTROPENIC CETACEANS.

Neutrophils are one of the initial cell lines of protection against pathogens, and when their concentrations in the blood are low, animals are highly susceptible to infections. Neutropenia has been reported in cetaceans secondary to administration of systemic sulfa antibiotics or antifungal medications and severe, overwhelming infection. Filgrastim was administered to treat neutropenia over a 15-y period in 11 cetaceans comprising four species-beluga (Delphinapterus leucas, n = 1), bottlenose dolphin (Tursiops truncatus, n = 4), killer whale (Orcinus orca, n = 5), and short-finned pilot whale (Globicephala macrorhynchus, n = 1)] ranging in age from 1 wk to >24 y. Seven study animals received multiple doses (2-6). All animals responded to at least one dose (1-7 µg/kg) of parenteral filgrastim characterized by an increase in peripheral immature (band) neutrophils, segmented neutrophils, or both. In most cases (9/11), neutrophil counts increased within 48 h of a single dose. Duration of response varied but was at least 2 wk in eight of the 11 animals and 5-9 d in the remaining animals. No adverse reactions were observed in any cases.

HEMATOLOGIC VALUES OF CAPTIVE HEALTHY RED-AND-GREEN MACAWS (ARA CHLOROPTERUS).

Hematologic reference values can be used to help monitor the health of birds kept in captivity for conservation and management purposes. The reference intervals for hematologic analysis in clinically healthy, captive red-and-green macaws (n = 42) were determined and evaluated as to whether they varied with sex. Males showed higher hemoglobin values than females. This report defines hematologic reference intervals of captive, healthy red-and-green macaws kept under similar conditions.

BASELINE PHYSIOLOGIC AND HEMATOLOGIC HEALTH IN WILD-CAUGHT MUSKRATS (ONDATRA ZIBETHICUS) FROM A NEAR-PRISTINE ECOSYSTEM IN NORTHERN MINNESOTA.

Muskrat (Ondatra zibethicus) populations show long-term and widespread declines across North America, necessitating research into potential mechanistic explanations, including population health. Previous research established reference hematology values, a proxy of individual health, of muskrats occurring in highly modified ecosystems. However, our knowledge of hematology metrics in muskrat populations occurring in more natural ecosystems is limited. We measured several hematological parameters of wild-caught muskrats (n = 73) in the Greater Voyageurs Ecosystem in northern Minnesota in 2018-2019 to establish baseline muskrat health in a relatively intact, near-pristine ecosystem. Additionally, we measured rectal temperature and heart and respiratory rates and collected whole blood for complete blood cell count assessment. We established baseline physiologic and hematologic reference ranges for the population and describe variations between total white blood cells, nucleated cell differentials, and basic erythron and platelet estimates and demonstrate methods of estimation to be poor proxies for more standardized counting methods. Our results establish a baseline to compare muskrat health assessments for populations affected by landscape change or in decline.

PARTIAL HEMATOLOGY, BIOCHEMISTRY, AND PROTEIN ELECTROPHORESIS REFERENCE INTERVALS FOR CAPTIVE SPOTTED TURTLES (CLEMMYS GUTTATA).

Spotted turtles (Clemmys guttata) are an endangered species and are commonly encountered in the pet trade and in many zoological collections across the United States, yet peer-reviewed published reference intervals (RI) for common clinicopathologic tests are unavailable for this species. The objectives of this study were to calculate partial RI for routine hematology, biochemistry, and electrophoretic analyses, as well as to perform an initial comparison of capillary zone electrophoresis (CZE) and agarose gel electrophoresis (AGE) in this species. A single blood sample was obtained from a single collection of 32 apparently healthy captive spotted turtles weighing at least 100 g and was submitted for standard hematologic and biochemistry analyses, as well as electrophoresis via CZE and AGE methods. Partial RI were calculated for corresponding analytes for each type of testing. While CZE and AGE protein fractions were found to have good correlation, some significant differences were observed, reinforcing that RI should be reported with the specific method used for their determination. The spotted turtle electrophoretograms were distinctly different from those previously reported from turtles in the same taxonomic family, including differences in the number and relative prominence of protein fractions.

COMPARISON OF THREE ANALYZERS FOR ASSESSING COMPLETE BLOOD COUNTS IN NONHUMAN PRIMATES.

Diagnostic hematology can prove challenging to the exotic animal practitioner presented with a nonhuman primate patient. Few point-of-care automated cell counters are calibrated for primate samples. Twenty-one samples from 17 nonhuman primates presented to an exotic animal practice were analyzed. Samples were run on both canine and feline settings on each of two veterinary point-of-care analyzers: one that assays by impedance technology, and one that assays by laser flow cytometry. Samples were also sent to a reference laboratory to be assayed on an analyzer that performs simultaneous impedance and laser measurements of blood cells and has been calibrated for use in nonhuman primates. Fourteen analytes were assessed for each sample on each machine. Manual hematocrits and total white blood cell counts were also performed on 16 of the samples. Statistical analysis indicated some variance between individual parameters, but overall correlation was acceptable.

HEMATOLOGY, PLASMA BIOCHEMISTRY, AND PLASMA PROTEIN ELECTROPHORESIS REFERENCE INTERVALS FOR BLUE IGUANAS (CYCLURA LEWISI) FROM GRAND CAYMAN ISLAND.

The blue iguana (Cyclura lewisi) is an endangered rock iguana species native to Grand Cayman, in the Cayman Islands. Health assessments were conducted on captive and free-roaming iguanas in 2001 and 2003-2014 and were performed in the summer wet season (June-July) of 2003-2004 and 2010-2014 and in the winter dry season (November-December) of 2001 and 2005-2009. Morphometric data were recorded from iguanas when blood samples were collected: 903 samples were collected and data from 890 samples from 775 iguanas were included. Samples were analyzed for hematology, plasma biochemistry, protein electrophoresis, mineral panels, 25-hydroxyvitamin D, and testosterone. Reference intervals were created for captive subadults, captive adults, and free-roaming adults when data were sufficient. Significant differences among these groups were described, as were differences on the basis of sex, season, and origin (captive vs free-roaming). In captive iguanas, most analytes were significantly different between subadults and adults, mature heterophils and copper were significantly higher in the dry season, zinc levels were significantly higher in the wet season, and cholesterol and triglycerides were significantly higher in adult females than adult males. Testosterone in adult males was significantly higher in the dry season. These results will aid in future health assessments and disease investigations in wild and captive populations of blue iguanas and are of comparative value for other Cyclura species that are free-roaming, captive, and, especially, in similar conservation release programs.

Effects of Season, Location, Species, and Sex on Hematologic and Plasma Biochemical Values and Body Mass in Free-ranging Grebes (Aechmophorus species).

The effects of season, location, species, and sex on body weight and a comprehensive array of blood chemistry and hematology analytes were compared for free-ranging western (Aechmophorus occidentalis) and Clark's (Aechmophorus clarkii) grebes. Birds (n = 56) were collected from Puget Sound, WA, and Monterey Bay and San Francisco Bay, CA, from February 2007 to March 2011. The data supported generalization of observed ranges for most analytes across Aechmophorus grebe metapopulations wintering on the Pacific coast. Notable seasonal and location effects were observed for packed cell volume (winter 6% greater than fall; winter California [CA] 5% greater than Washington [WA]), total white blood cell count (CA 3.57 × 103 cells/µL greater than WA), heterophils (WA 10% greater than CA), lymphocytes (winter 19% greater than fall), heterophil to lymphocyte ratio (fall 5.7 greater than winter), basophils (CA greater than WA), plasma protein (WA about 10 g/L [1.0 g/dL] greater than CA), plasma protein to fibrinogen ratio (winter about 15 greater than fall), potassium (CA 2 mmol/L greater than WA), and liver enzymes (alanine aminotransferase, aspartate aminotransferase, lactate dehydrogenase: WA greater than CA). Within California, season had a greater effect on body mass than sex (mean winter weights about 200 g greater than fall), whereas within a season, males weighed only about 80 g more than females, on average. These data give biologists and veterinarians quantitative reference values to better assess health at the individual and metapopulation level.

Immune responses upon experimental Erysipelothrix rhusiopathiae infection of naïve and vaccinated chickens.

Erysipelas, a disease caused by Erysipelothrix rhusiopathiae (ER), is an increasing problem in laying hens housed in cage-free systems. This study aimed to monitor immune responses during ER infection of naïve chickens and chickens vaccinated intra muscularly with a commercial inactivated ER vaccine. Chickens were infected intra muscularly with ER at 30 days of age and blood leukocyte counts, serum levels of mannose binding lectin (MBL) and ER-specific IgY were monitored until the experiment was terminated at day 15 after infection. ER was detected in blood from more chickens and at higher bacterial counts in the naïve group (day 1: 1 of 7 chickens; day 3: 6 of 6 chickens) than in the vaccinated group (day 1: 0 of 7 chickens; day 3: 1 of 6 chickens). During the acute phase of infection transient increases in circulating heterophil numbers and serum MBL levels were detected in all ER infected chickens but these responses were prolonged in chickens from the naïve group compared to vaccinated chickens. Before infection IgY titers to ER in vaccinated chickens did not differ significantly from those of naïve chickens but vaccinated chickens showed significantly increased IgY titers to ER earlier after infection compared to chickens in the naïve group. In conclusion, the ER infection elicited prompt acute innate responses in all chickens. Vaccinated chickens did not have high IgY titers to ER prior to infection but did however show lower levels of bacteraemia and their acute immune responses were of shorter duration.