Equine neonatal isoerythrolysis: evidence for prevention by maternal antibodies to the Ca blood group antigen

Foals with the Ca blood group antigen on their RBC were given colostrum with anti-Ca antibodies (6 foals) or colostrum without anti-Ca antibodies (6 foals). The PCV were determined at birth and 2, 4, and 6 days after birth for the foals in each group. Significant differences were not observed for the PCV between the 2 groups, indicating that foals were not adversely affected by ingesting colostrum with the anti-Ca antibody. Standardbred mares without the Aa blood group antigen were evaluated to determine whether production of anti-Ca antibodies influenced production of anti-Aa antibodies. Of 266 mares without the Aa antigen, 3 of 61 (5%) mares without the Ca blood group antigen produced anti-Aa antibodies and 43 of 205 (21%) with the Ca blood group antigen produced anti-Aa antibodies. These 2 groups of mares were significantly (P = 0.006) different; Ca-negative mares were less likely to produce antibodies to Aa than were mares with the Ca blood group antigen. This observation was consistent with a hypothesis of antibody-mediated immunosuppression of immune response to the Aa blood group antigen by antibodies to the Ca blood group antigen, ie, when a mare is exposed to her foal's RBC and already has antibodies to the Ca blood group antigen on the foal's RBC, then she is less likely to initiate an immune response to the Aa blood group antigen also on the foal's RBC.     

Clinical survey of antibodies against red blood cells in horses after homologous blood transfusion

Serum samples of 20 horses were evaluated for antibodies against RBC after homologous blood transfusion. Transfusion-associated antibodies against RBC were detected in 10 horses. Antibodies recognizing horse blood group antigens Aa, Ae, Db, and Dc were identified. Antibodies against Aa were found in all samples from Aa-negative horses that were transfused with Aa-positive RBC. Antibodies against Aa persisted for at least 1 year after transfusion. Antibodies against Ae were detected in 7 of 8 horses transfused with Ae-positive RBC. Initial appearance and persistence of antibodies against Ae differed among the horses; antibodies were initially detected 1 week to 154 weeks after transfusion and disappeared as early as 4 weeks after transfusion. Antibodies against Db or Dc were detected in less than or equal to 33% of the horses that lacked Db or Dc antigens and were transfused with Db- or Dc-positive RBC. Antibodies against Db and Dc were initially detected in sera later than were the A-system antibodies. Three mares with transfusion-associated antibodies subsequently produced healthy offspring. Two foals had RBC antigens corresponding to their dam's alloantibodies; maternal colostrum with antibodies against Aa was withheld from the Aa-positive foal. The Db-positive foal remained healthy after nursing the mare with serum antibodies against Db.     

Effects of blood contamination of cerebrospinal fluid on results of indirect fluorescent antibody tests for detection of antibodies against Sarcocystis neurona and Neospora hughesi.

The purpose of this study was to determine the effect of blood contamination of cerebrospinal fluid (CSF) on the results of indirect fluorescent antibody tests (IFATs) for Sarcocystis neurona and Neospora hughesi. The in vitro study used antibody-negative CSF collected from non-neurologic horses immediately after euthanasia and blood samples from 40 healthy horses that had a range of IFAT antibody titers against S. neurona and N. hughesi. Serial dilutions of whole blood were made in seronegative CSF to generate blood-contaminated CSF with red blood cell (RBC) concentrations ranging from 10 to 100,000 RBCs/microl. The blood-contaminated CSF samples were then tested for antibodies against both pathogens using IFAT. Blood contamination of CSF had no detectable effect on IFAT results for S. neurona or N. hughesi at any serologic titer when the RBC concentration in CSF was <10,000 RBCs/microl. At concentrations of 10,000-100,000 RBCs/microl of CSF, positive CSF results (IFAT titer >or=5) for S. neurona and N. hughesi were detected only when the corresponding serum titers were >or=160 and >or=80, respectively. The IFAT performed on CSF is reliable for testing horses for equine protozoal myeloencephalitis caused by S. neurona or N. hughesi, even when blood contamination causes the RBC concentration in CSF to be up to 10,000 RBCs/microl.     

Susceptibility of equine erythrocytes to oxidant-induced rheologic alterations

OBJECTIVE: To evaluate the rheologic responses of equine versus human RBC to oxidant stress induced by superoxide anions. SAMPLE POPULATION: Equine blood samples were obtained from 8 healthy, 3- to 6-year-old various breed horses of either sex; human blood samples were obtained from 8 healthy adults. PROCEDURE: Washed RBC were exposed to superoxide anions generated by the xanthine oxidase (XO)-hypoxanthine system (XO activity of 0 to 0.1 U/ml). Deformability of RBC was assessed by ektacytometry, and RBC aggregation was measured in autologous plasma or 3% solution of dextran 70 via a defined-shear photometric technique. RESULTS: Equine RBC had XO dose-dependent increases in methemoglobin concentration that were greater by 60 to 110% than in human RBC and an enhanced tendency for echinocyte formation (ie, 40% echinocyte formation at highest activity of XO). Oxidant stress reduced deformability (ie, increased rigidity) for equine and human RBC with the effect more prominent for equine RBC. Equine RBC aggregation had a biphasic response with a significant increase in plasma and dextran 70 at low XO activities and inhibition at high activities; echinocytes were incorporated into equine, but not human, RBC aggregates. CONCLUSIONS AND CLINICAL RELEVANCE: Compared with human RBC, equine RBC are more sensitive to oxidant damage as judged by the extent of methemoglobin formation, alteration of aggregation, and reduction of cellular deformability. The high susceptibility of equine RBC to oxidant damage, and the resulting hemorheologic alterations, may have important consequences for tissue perfusion and cardiovascular adequacy in horses; they may be of particular relevance in physiologic or pathophysiologic changes associated with increased oxidant stress.     

Effects of blood contamination of cerebrospinal fluid on western blot analysis for detection of antibodies against Sarcocystis neurona and on albumin quotient and immunoglobulin G index in horses.

OBJECTIVE: To determine effects of blood contamination on western blot (WB) analysis of CSF samples for detection of anti-Sarcocystis neurona antibodies, and on CSF albumin and IgG concentrations, albumin quotient (AQ), and IgG index in horses. DESIGN: Prospective in vitro study. SAMPLES: Blood with various degrees of immunoreactivity against S neurona was collected from 12 healthy horses. Cerebrospinal fluid without immunoreactivity against S neurona was harvested from 4 recently euthanatized horses. PROCEDURE: Blood was serially diluted with pooled nonimmunoreactive CSF so that final dilutions corresponded to 10(-3) to 100 microliters of blood/ml CSF, and WB analysis was performed on contaminated CSF samples. Number of RBC, albumin and IgG concentrations, AQ, and IgG index were also determined. RESULTS: Antibodies against S neurona were detected in CSF contaminated with 10(-3) microliters of strongly immunoreactive blood/ml. In CSF samples contaminated with 10 microliters of blood/ml, AQ remained within reference range. Volume of blood required to increase IgG index varied among blood samples and was primarily influenced by serum IgG concentrations. Number of RBC in contaminated samples was correlated with volume of blood added, but not with degree of immunoreactivity detected in contaminated CSF samples. CONCLUSIONS AND CLINICAL RELEVANCE: During collection of CSF from horses, contamination with blood may introduce serum antibodies against S neurona at concentrations sufficient for detection by WB analysis, thus yielding false-positive results. When blood is moderately or strongly immunoreactive, the amount of contaminating albumin may be small enough as to not increase AQ above reference range. In these cases, AQ and IgG index should be interpreted with caution.     

Clinical, immunophenotypic and functional characterisation of T-cell leukaemia in six horses

REASON FOR PERFORMING STUDY: Lymphoid leukaemia (LL) is rare in equids. In man, immunophenotypic classification identifies distinct leukaemic types with different treatment strategies. Improved understanding and classification of equine LL may allow similar advances. OBJECTIVES: To document the clinical, immunophenotypic and functional characteristics in 6 cases of equine LL of T-cell origin. METHODS: The clinical records and pathological findings from 6 cases of equine LL were analysed. Immunohistochemistry to identify T or B lymphocytes was performed on paraffin embedded tissues in 4 cases. Peripheral blood mononuclear cells (PBMC) were phenotyped for expression of CD4, CD8, MHC class I and II and B-cell antigens in 4 cases using monoclonal antibodies (mAbs) and flow cytometry. Neoplastic lymphocytes from 4 horses were stimulated with mitogens. RESULTS AND CONCLUSIONS: Six horses of various breeds were identified with LL of T-cell origin. The clinical course and presenting signs varied. Neoplastic lymphocytes were identified in peripheral blood samples from all horses and tissue invasion was confirmed at examination post mortem in 4 horses. Immunophenotyping identified a predominance of CD3+ T-cells in lymphoid tissues and CD4+ T-cells in circulating peripheral blood mononuclear cells (PBMC) in the affected horses. Neoplastic lymphocytes from the 4 cases that were tested failed to proliferate in response to mitogens. POTENTIAL RELEVANCE: Characterisation of the clinical, pathological and immunological findings in 6 horses with LL has added to reports of this rare condition, characterised it in greater detail and therefore provides a starting point for further investigations.     

Adsorptive effects of di-tri-octahedral smectite on Clostridium perfringens alpha, beta, and beta-2 exotoxins and equine colostral antibodies

OBJECTIVE: To determine the adsorptive capability of di-tri-octahedral smectite (DTOS) on Clostridium perfringens alpha, beta, and beta-2 exotoxins and equine colostral antibodies. SAMPLE POPULATION: 3 C perfringens exotoxins and 9 colostral samples. PROCEDURES: Alpha, beta, and beta-2 exotoxins were individually co-incubated with serial dilutions of DTOS or bismuth subsalicylate, and the amount of toxin remaining after incubation was determined via toxin-specific ELISAs. Colostral samples from healthy mares were individually co-incubated with serial dilutions of DTOS, and colostral IgG concentrations were determined via single radial immunodiffusion assay. RESULTS: Di-tri-octahedral smectite decreased the amount of each C perfringens exotoxin in co-incubated samples in a dose-dependent manner and was more effective than bismuth subsalicylate at reducing exotoxins in vitro. Decreases in the concentration of IgG were detected in samples of colostrum that were combined with DTOS at 1:4 through 1:16 dilutions, whereas no significant decrease was evident with DTOS at the 1:32 dilution. CONCLUSIONS AND CLINICAL RELEVANCE: Di-tri-octahedral smectite effectively adsorbed C perfringens exotoxins in vitro and had a dose-dependent effect on the availability of equine colostral antibodies. Results suggested that DTOS may be an appropriate adjunctive treatment in the management of neonatal clostridiosis in horses. In vivo studies are necessary to fully assess the clinical efficacy of DTOS treatment.     

Modified in vitro method to label equine red blood cells with technetium 99m in concentrated whole blood

An in vitro method to label equine RBC with technetium 99m was modified to achieve quantitative labeling of cells in concentrated whole blood. After a blood sample was incubated with a reducing agent (stannous citrate), an oxidizing reagent (NaOCl) and a chelating agent (EDTA) were added to inactivate residual Sn2+ in the plasma. This step prevented premature reduction of pertechnetate in plasma. Labeling of RBC from 9 healthy horses, using a standard whole blood protocol, resulted in only moderate labeling efficiency (44 to 85%) and indicated a linear relationship between labeling efficiency and PCV. Effects of increased incubation time, increased incubation temperature, prelabeling sedimentation, and double addition of NaOCl/EDTA were investigated in whole blood from 10 healthy horses. Labeling efficiency was improved by each independent factor and by combination of factors. Highest labeling efficiencies (96 to 97%) were achieved when blood samples were sedimented for 20 minutes before being labeled, regardless of incubation time or incubation temperature. Morphologic features of RBC were unaffected by labeling procedures. In vivo whole blood clearance time for labeled cells was determined in 5 healthy horses. Sedimented blood samples were labeled, using a standard 15-minute incubation time at 20 to 22 C. Mean clearance half-time for 5 horses was approximately 20 hours. More than 95% of 99mTc activity was associated with the cells during the 24 hours after reinjection.     

Detection of equine antiplatelet and antineutrophil antibodies by enzyme-linked immunosorbent assay

An enzyme-linked immunosorbent assay (elisa) was standardised and applied for the detection of anti-platelet and antineutrophil antibodies using a heterologous system consisting of equine platelets or neutrophils and antisera raised in rabbits. The standardised technique consisted of using Immulon type 3 plate, I per cent gelatine as a blocking solution, poly-L-lysine buffer as a coating solution, unfixed antigen, 90 μl test serum, horseradish peroxidase conjugated antibody and o-phenylenediamine dihydrochloride as a substrate. The number of unfixed platelets or neutrophils required for optimum detection of antibodies was 250,000 per well. Unfixed cellular antigens were as good as their extracts and superior to paraformaldehyde-fixed antigens in detecting specific antibodies. Microtitre plates coated with platelet or neutrophil antigens could be stored at 4° and −70°C for four to five weeks without significant loss of antigenicity. The ELISA was very sensitive in that antiplatelet antibody was detected up to a titre of 1:204,800 and antineutrophil antibody to a titre of 1:51,200. Some cross-reactivity (1:1600) was detected in antiplatelet and antineutrophil sera for neutrophil and platelet antigens, respectively. Platelet-associated antibody was also detected in extracts from platelets pretreated with 1:2 and 1:8 dilutions of antiplatelet serum. Standardised elisa detected antiplatelet antibodies in nine and antineutrophil antibodies in three of 100 isologous equine blood typing sera.     

Comparison of various canine blood-typing methods

OBJECTIVE: To compare canine blood-typing results determined by use of the card (CARD), gel (GEL), Michigan State University (MSU), and tube (TUBE) tests. SAMPLE POPULATION: Blood samples from 23 healthy dogs. PROCEDURES: Blood samples anticoagulated with EDTA were screened by use of each blood-typing method according to manufacturers' protocols. RESULTS: Strong RBC agglutination reactions were observed with dog erythrocyte antigen (DEA) 1.1 reagents of the CARD and GEL tests as well as MSU test (only after adding Coombs' reagent) in 9 blood samples. By use of the CARD test, RBCs from 4 additional dogs agglutinated weakly; on the basis of MSU test results, these 4 dogs were classified as DEA 1.2 positive. All blood samples agglutinated with the B antigen reagent of the TUBE test. All but 2 blood samples had strong positive reactions with the DEA 4 reagent of the MSU test. All but 3 blood samples reacted with the E antigen reagent of the TUBE test. Three blood samples agglutinated with the DEA 3 reagent of the MSU test and A antigen reagent of the TUBE test. Five blood samples had strong agglutination reactions with the DEA 5 reagent of the MSU test. CONCLUSIONS AND CLINICAL RELEVANCE: Use of the CARD test allows for rapid identification of DEA 1.1 but may produce weak reactions with blood from DEA 1.2-positive dogs. The GEL test is a reliable and rapid clinical laboratory method for identification of DEA 1.1. The MSU test requires Coombs' reagent for identification of DEA 1.1 and 1.2.     

Validation of thromboelastometry in horses

Background: Thromboelastometry is used for identifying or monitoring coagulation abnormalities. It has been validated in several species but not in horses and the characteristics of the equine thromboelastogram have not yet been detailed. Objectives: The purpose of this study was to validate a thromboelastometer to be used with equine blood and to define the normal equine thromboelastogram. Methods: A Rotem-gamma thromboelastometer (Pentapharm GmbH, Munich, Germany) was used on 38 citrated blood samples to investigate native coagulation, the intrinsic and extrinsic pathways, the function of fibrinogen (largely dependent on its concentration), and the presence of fibrinolysis. Using classic validation approaches, we evaluated the imprecision of the method and the influence of hemolysis and storage time and temperature. The normal thromboelastogram was defined in both saddle and racing horses (the latter sampled before and after the race). Results: For imprecision tests, the analytical variations were <10%. The equine thromboelastogram had a pattern similar to those of other species, but the intrinsic and extrinsic pathways were less and more efficient, respectively. Reference intervals in racing horses, especially after exercise, were different from those of saddle horses, most likely due to a higher RBC mass. Coagulability decreased in hemolyzed samples and significant changes were found between nonrefrigerated and refrigerated blood samples stored for 20 hours. Conclusions: The Rotem-gamma thromboelastometer is a precise instrument for use with equine blood samples. The equine thromboelastogram is similar to that of other species, but reference intervals vary with aptitude and exercise. Hemolysis and refrigeration alter thromboelastometric results.     

Tears and aqueous humor from horses inoculated with Leptospira contain antibodies which bind to cornea

An antigenic relationship between Leptospira interrogans and equine cornea was previously described by us. An enzyme-linked immunosorbent assay was employed in the present work to investigate the existence of anti-leptospira and anti-cornea antibodies in tears, aqueous humor and serum from horses inoculated i.m. with those antigens. Ten days after a booster by the same route, antibodies that bind to microtiter plates, coated with an homogenate of either equine cornea or leptospira, were detected in those fluids and in the sera. At the same time, the corneas of the horses began to develop a diffuse opacity. This finding of anti-leptospira antibodies in equine tears and aqueous humor shows the pathway along which they arrive at the cornea and bind to it.     

Comparison of 4 blood storage methods in a protocol for equine pre-operative autologous donation

OBJECTIVE: To compare viability of equine whole blood stored by 4 different methods, and to establish optimal storage protocols for an equine autologous blood donation program. STUDY DESIGN: In vitro study of stored equine whole blood. Animals- Six healthy adult horses. METHODS: Blood from each horse was collected into 4 different containers: glass bottles containing acid-citrate-dextrose solution (ACD), plastic bags containing ACD, citrate-phosphate-dextrose (CPD), and CPD with supplemental adenine (CPDA-1). Blood was stored for 5 weeks and sampled at 2-day intervals. Standard hematologic and biochemical variables were evaluated, and adenosine-5-triphosphate (ATP) and 2,3-diphosphoglycerate (2,3-DPG) concentrations were measured and normalized to total hemoglobin content. RESULTS: Plasma hemoglobin, % hemolysis, lactate, potassium, ammonia, and lactate dehydrogenase (LDH) increased, whereas glucose concentration and pH decreased in all stored blood over 5 weeks. There was a temporal increase in hemolysis with all storage methods, but the increase was greatest in glass bottles. Lactate and ammonia were highest in CPD and CPDA-1 samples, indicating more active red blood cell (RBC) metabolism. 2,3-DPG concentrations decreased during storage, but were optimally preserved with CPDA-1. ATP concentrations were significantly higher for blood stored in CPDA-1, and were lowest in glass bottles. CONCLUSIONS: Hematologic and biochemical values measured for blood stored in CPDA-1 are suggestive of improved RBC viability compared with other storage methods. With the exception of ATP, results from stored equine blood were similar to those reported for other species. CLINICAL RELEVANCE: Commercial CPDA-1 bags appear to be the optimal storage method for equine whole blood.     

Differential Expression of Monocarboxylate Transporter 1 and Ancillary Protein CD147 in Red Blood Cells of Show Jumping Horses

We compare the expression levels of the lactate transporter complex consisting of the lactate transporter, monocarboxylate transporter 1 (MCT1), and its ancillary protein, cluster of differentiation 147 (CD147), in the membranes of red blood cells (RBCs) from two breeds of jumping horses and associate the expression levels of these proteins with their jumping ability. The expression levels of MCT1 and CD147 proteins on the membranes of RBCs collected from 30 show jumping horses of two different breeds were quantified: the Brazilian Sport Horses (n = 17) and the European Warmbloods (n = 13). The levels of MCT1 and CD147 in the RBC membranes were measured by western blot using horse-specific antibodies. Statistical analyses included unpaired Student t test and chi-squared test. According to the expression levels of MCT1 and CD147 proteins, 88% of the Brazilian Sport Horses were categorized as high lactate transporters (HTs) and the remaining 12% as low lactate transporters (LTs). The opposite was found for the European Warmbloods, where most animals (77%) were classified as LTs and the remaining animals (23%) were classified as HTs. Brazilian Sport Horses express statistically significantly higher levels of CD147 and MCT1 than European Warmbloods. The classification of horses considering the expression of proteins involved in the ability to transport lactate through the complex MCT1-CD147 seems to be breed dependent, with horses that are able to jump higher obstacles showing lower expression of the MCT1-CD147 complex in their RBCs.     

Artifactual changes in equine blood following storage,detected using the Advia 120 hematology analyzer

Background — Delayed analysis of blood samples may be caused by restricted access to laboratories. Artifactual changes may occur in the measured analytes as a consequence of delayed analysis and may complicate interpretation of the data.
Objective — The purpose of this study was to characterize artifactual changes in equine blood, due to storage, using the Advia 120 hematology analyzer.
Methods — Samples of blood from 5 horses were analyzed using the Advia 120 soon after collection and again after 24 and 48 hours of storage at either 4°C or ambient laboratory temperature (∼24°C).
Results — Delayed analysis of equine blood samples resulted in increased numbers of normocytic hypochromic RBCs, increased numbers of macrocytic hypochromic RBCs, misclassification of granulocytes as mononuclear cells using the basophil reagent method, and pseudothrombocytosis, due to misclassification of ghost RBCs as platelets. The latter artifact was corrected by an amended version of the software. Many of the artifactual changes were identified by morphology flags.
Conclusion — Characteristic changes in cytograms produced by the Advia 120 allowed recognition of artifactual changes in stored equine blood samples. These changes were less pronounced in samples stored at 24°C than at 4°C.     

Studies on Blood and Serum Types of the Icelandic Horses

By means of isoimmunizations and heteroimmunizations 10 equine blood typing reagents were isolated. The specific antibodies were complete agglutinins, which were used in the direct agglutination test in saline medium. The reagents were designated A₂, C, D, E, G, H, I, K, Da₁, and Da₂ reagent. Da₁ and Da₂ are preliminary designations.The data obtained from blood typing of a family material and a population material of Icelandic horses showed that the occurrence of each blood type factor is controlled by a single, dominant gene. The family data tended to show that the blood factors under investigation belonged to 8 blood type systems. The A system contained the antigens A₂ and Da₂. These antigens are related to each other through a linear subgroup relationship. The D system had the factors D and J. The G, E, G, I, K, and Da₁ systems are one-factor, two-allele blood type systems. The H factor was not observed in Icelandic horses. In connection with the establishment of the 8 blood type systems it must be emphasized that the problem of allelism or nonallelism of 2 genes can only be solved by means of relevant family data. Because of the rare occurrence of some of the blood factors in the Icelandic horse such data were in some cases not available. Thus some conclusions were based on results from two-by-two contingency tables with the use of population data. This was used particularly for the D and G systems, and additional family data are necessary for a definite establishment of these systems.Exceptions to the genetic theory, apparently caused by erroneous registration, were presented.Finally, estimates were given of gene frequencies of the causative genes among Icelandic horses.Starch gel electrophoresis of sera from Icelandic horses revealed the existence of 21 transferrin phenotypes. The data obtained supported the theory advanced, that transferrin polymorphism in horses is controlled by 6 autosomal codominant alleles: Tf^D, Tf^F, Tf^H, Tf^M, Tf^O, and Tf^H.925 randomly selected Icelandic horses were typed for serum transferrin and the gene frequencies were estimated.Starch gel electrophoresis of about 100 horse serum samples did hot reveal individual variation of the equine haptoglobin and ceruloplasmin. Studies on approximately 300 sera showed an identical serum amylase pattern.     

Clinical evaluation of the CA530-VET hematology analyzer for use in veterinary practice

BACKGROUND: The CA530-VET is a completely automated impedance cell hematology analyzer, which yields a 16-parameter blood count including a 3-part leukocyte differential. OBJECTIVES: The aim of this study was to examine the operational potential of the CA530-VET and its value for use in veterinary practice. METHODS: The analyzer was tested for blood carry-over, precision, and accuracy. Comparison methods included the CELL-DYN 3500, microhematocrit centrifugation, manual platelet (PLT) counting for feline and equine species, and a 100-cell manual WBC differential. Blood samples for comparison of the methods were obtained from 242 dogs, 166 cats, and 144 horses. RESULTS: The carry-over ratio (K) was 0.28% for RBC, 0.59% for PLT, 0.32% for WBC, and 0.18% for hemoglobin (HGB) concentration. Coefficients of variation (CVs) for within-batch precision and duplicate measurement of blood samples were clearly within the required limits, except for duplicate platelet counts in cats (8.7%) and horses (9.5%). The WBC count was in excellent agreement for dogs and horses and RBC count was in excellent agreement for horses. The accuracy of feline WBC counts was not acceptable, with the exception of values at the high end of the range. RBC counts in dogs and cats, and HGB concentration and MCV in all 3 species were sufficiently accurate. The CA530-VET HCT results were in excellent agreement with microhematocrit results in horses but exceeded the maximum allowed inaccuracy for cats and dogs. In all species, PLT counts established mechanically and manually were not in adequate agreement. Large differences were found between the CA530-VET and the manual differential percentage for lymphocytes and "mid-sized cells" (monocytes and basophilic granulocytes). CONCLUSIONS: The CA530-VET can be considered useful for routine canine, feline, and equine blood cell analyses. It should not be considered accurate, however, for PLT counts, feline total WBC counts in the subnormal and normal range, and leukocyte differentials, except for granulocytes.     

Evaluation of the opsonic capacity of core lipopolysaccharide antiserum of equine origin against smooth Escherichia coli 0111:B4, using macrophage chemiluminescence

A study was performed to determine whether equine antiserum to core lipopolysaccharide (LPS) would enhance phagocytosis of smooth gram-negative (GN) organisms by equine macrophages. Five healthy adult horses (group A) were immunized with a bacterin prepared from the J-5 mutant of Escherichia coli 0111:B4 and Salmonella minnesota R595 to produce antibodies to core LPS. Five horses (group B) served as nonimmunized controls and were given physiologic saline solution instead of the rough mutant bacterin. Serum antibody titers to core LPS and to smooth E coli 0111:B4 were determined by indirect ELISA. Four serum pools were prepared: pool 1 = sera from horses in group B prior to immunization; pool 2 = sera from horses in group A prior to immunization (preimmune serum); pool 3 = sera from horses in group B, 7 days after the last saline injection; pool 4 = sera from horses in group A, 7 days after the last immunization (core LPS antiserum). The serum pools, either unheated or heated 30 minutes at 56 C, in 3 dilutions (1/50, 1/100, 1/500) were used to opsonize smooth E coli 0111:B4 in an assay of equine peritoneal macrophage chemiluminescence (CL). Peritoneal fluid was collected from clinically normal horses and the macrophages were purified by adherence to borosilicate glass scintillation vials. Each serum type and dilution was added to triplicate vials containing 10(7) colony-forming units of E coli 0111:B4. Luminol-dependent CL was measured with a liquid scintillation counter in the out-of-coincidence mode. Each serum dilution was tested in duplicate vials without bacteria to asses serum-induced nonspecific CL.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pitfalls in immunofluorescence testing in dermatology. III. Pemphigus-like antibodies in the horse and direct immunofluorescence testing in equine dermatophilosis

Indirect immunofluorescence testing for pemphigus-like antibodies was performed on 79 horses: 28 horses with various nonpemphigus dermatologic diseases, 21 horses with various nondermatologic diseases, and 30 normal horses. Pemphigus-like antibodies were detected in 6 horses: 3 normal horses with titers of 1:40, 2 horses with dermatophilosis at titers of 1:10 and 1:80, and 1 horse with lymphosarcoma at a titer of 1:320. It was concluded that equine pemphigus-like antibodies are a potential source of misinterpretation and misdiagnosis in indirect immunofluorescence testing. Direct immunofluorescence testing for whole immunoglobulin, IgG, IgM, and IgA was performed on skin lesions from 2 horses with dermatophilosis. Diffuse intercellular deposition of whole immunoglobulin and IgG was found in both horses. It was concluded that equine dermatophilosis is a potential source of misinterpretation and misdiagnosis in direct immunofluorescence testing.     

Viruses associated with outbreaks of equine respiratory disease in New Zealand

AIM: To identify viruses associated with respiratory disease in young horses in New Zealand.

METHODS: Nasal swabs and blood samples were collected from 45 foals or horses from five separate outbreaks of respiratory disease that occurred in New Zealand in 1996, and from 37 yearlings at the time of the annual yearling sales in January that same year. Virus isolation from nasal swabs and peripheral blood leukocytes (PBL) was undertaken and serum samples were tested for antibodies against equine herpesviruses (EHV-1, EHV-2, EHV-4 and EHV-5), equine rhinitis-A virus (ERAV), equine rhinitis-B virus (ERBV), equine adenovirus 1 (EAdV-1), equine arteritis virus (EAV), reovirus 3 and parainfluenza virus type 3 (PIV3).

RESULTS: Viruses were isolated from 24/94 (26%) nasal swab samples and from 77/80 (96%) PBL samples collected from both healthy horses and horses showing clinical signs of respiratory disease. All isolates were identified as EHV-2, EHV-4, EHV-5 or untyped EHV. Of the horses and foals tested, 59/82 (72%) were positive for EHV-1 and/or EHV-4 serum neutralising (SN) antibody on at least one sampling occasion, 52/82 (63%) for EHV-1-specific antibody tested by enzyme-linked immunosorbent assay (ELISA), 10/80 (13%) for ERAV SN antibody, 60/80 (75%) for ERBV SN antibody, and 42/80 (53%) for haemagglutination inhibition (HI) antibody to EAdV-1. None of the 64 serum samples tested were positive for antibodies to EAV, reovirus 3 or PIV3. Evidence of infection with all viruses tested was detected in both healthy horses and in horses showing clinical signs of respiratory disease. Recent EHV-2 infection was associated with the development of signs of respiratory disease among yearlings [relative risk (RR)=2.67, 95% CI=1.59-4.47, p=0.017].

CONCLUSIONS: Of the equine respiratory viruses detected in horses in New Zealand during this study, EHV-2 was most likely to be associated with respiratory disease. However, factors other than viral infection are probably important in the development of clinical signs of disease.