Replication of bluetongue virus (BTV) in leukocytes from the blood of sheep, cattle, elk, and mule deer inoculated with BTV serotype 10 or 17 was assessed by immunocytochemical staining and dot blot northern hybridization to determine if differences in the prevalence of infection in this blood fraction might account for the differences in clinical disease among these species. Viremia was confirmed by virus isolation in all inoculated animals. Analysis of leukocytes with monoclonal antibodies specific for BTV proteins revealed low numbers of infected leukocytes in only 2 sheep 8 days after inoculation with BTV serotype 10. Most of the cells expressing BTV were identified morphologically as monocytes; approximately 10% of infected cells were lymphocytes. Bluetongue virus was not detected by use of dot-blot hybridization on samples of blood. Our results suggest that differential infection of leukocytes does not account for the pronounced differences in clinical signs and pathologic changes among ruminants.

Enzyme-linked immunosorbent assay and flow cytometric methods to screen hybridoma culture supernatants for antibodies to bovine neutrophils (surface antigen-specific) were optimized. Sensitivity of the 2 methods was compared. A panel of 14 murine monoclonal antibodies (MAB) to surface antigens of bovine polymorphonuclear neutrophilic leukocytes (neutrophils) was produced by hybridoma technology, and their isotypes were determined by whole-cell ELISA. Monoclonal antibody reactivity with neutrophils, eosinophils, and lymphocytes isolated on phosphate-buffered saline solution and on Ficoll-sodium diatrizoate were compared. Biochemical characterization of antigens recognized by MAB was performed by immunoblot analysis. Neutrophil plasma membranes were isolated on sucrose gradients (20, 32, and 50%) and purified for polypeptide characterization. Neutrophil surface proteins were characterized by external labeling with 125I. The flow cytometric method was proven to be more sensitive and rapid than ELISA to screen hybridoma supernatants. This method allowed light-scatter gating of live neutrophil populations for analysis, which eliminated nonspecific binding of antibodies to contaminating cells and dead neutrophils. The optimal conditions for flow cytometric analyses were 5 X 10(5) neutrophils and 1 microgram of fluorescein-labeled F(ab')2/assay as the second antibody. The optimal conditions for hybridoma screening by ELISA were neutrophil concentration of 2.5 X 10(5) well, using a 96-well polystyrene microtitration plate as solid support, and 2,2'-azino-di[3-ethyl-benzthiazoline sulfonate (6)] with H2O2 as the chromogenic substrate. Tissue culture plates as solid support and 3,3', 5,5'-tetramethyl benzidine, with H2O2 as the chromogenic substrate, were equally as sensitive. Panel MAB reacted differently with neutrophils, eosinophils, and lymphocytes. Isolation of these cells from blood on Ficoll-sodium diatrizoate generally did not alter MAB reactivity. Coomassie blue-stained gels of neutrophil plasma membrane proteins contained about 25 polypeptide bands, 13 of which were major bands. Autoradiography revealed about 11 surface proteins, 5 of which were heavily labeled with 125I. Monoclonal antibody S7G8 identified a 65-kd protein and MAB S8G10 identified 65- and 70-kd proteins. On the basis of molecular weight, MAB S7G8 and S8G10 are comparable to human CD15, CD16, and CD64 molecules. The MAB generated in this study are potential candidates to discern bovine neutrophil function and heterogeneity.

During the breeding season, the effect of constant administration of an agonist analog of gonadotropin-releasing hormone (GnRH; goserelin acetate) on reproductive activity of mares was determined. Twenty-four mares undergoing estrous cycles were allocated at random to 6 groups (n = 4/group) and, on May 29 (day 0), received no treatment (group 1, controls), 120 micrograms (group 2), 360 micrograms (group 3), 600 micrograms (group 4), or 1,200 micrograms (group 5) of GnRH agonist/d for 28 days via a depot implanted subcutaneously. The final group of mares (group 6) was treated with 120 miocrograms of GnRH agonist/d for 84 days (3 occasions at 28-day intervals). During a pretreatment period (April 19 to May 29) and for 90 days after initiation of GnRH agonist treatment, follicular development and ovulation were monitored by transrectal ultrasonography of the reproductive tract at 2- to 3-day intervals. On each occasion a blood sample was collected for determination of luteinizing hormone (LH) and progesterone. Estrous behavior was monitored by teasing of mares with a stallion. Initiation of agonist treatment was random, relative to the stage of the estrous cycle, and all mares ovulated within 11 days before or after implantation. in 3 of 4 nontreated control mares, estrous cycles were observed throughout the study, with interovulatory intervals ranging from 18 to 26 days. In the remaining mare, concentration of progesterone was high after asynchronous double ovulation during the pretreatment period, suggestive of persistent corpus luteum. In group-2 mares, ovulation occurred in all mares 7 days before and 2 days after initiation of treatment; however, the next anticipated ovulation was delayed in 3 of 4 mares (interovulatory interval, 33 to 70 days). Estrous cycles were not disrupted in the remaining mare. At higher doses (groups 3-5), 1 mare each from groups 3 and 5 ovulated between days 0 and 2 of treatment initiation, but faded to ovulate during the remainder of the study (anovulatory for > 88 days). Similarly, an additional 2 mares of groups 2 and 3 ovulated within 2 days of GnRH agonist treatment. A second ovulation occurred in these mares 32 to 35 days later, thereafter, both mares were anovulatory for the remainder of the study. In the remaining 8 mares, interovulatory intervals were either lengthened (n = 6 mares, range, 32 to 82 days) or were unaffected (n = 2) by treatment. One group-6 mare had a lengthened interovulatory interval, 1 was anovulatory for > 90 days, and the remaining 2 mares were unaffected by treatment. During the 28-day treatment period, serum concentration of LH decreased (P < 0.05) only in mares of groups 3-5. In group-6 mares, concentration of LH was unchanged during each 28-day period after depot GnRH agonist administration. Thus, constant administration of a GnRH agonist to mares during the breeding season disrupted their estrous cycles. Anovulation or lengthening of the interovulatory interval by GnRH agonist treatment was associated with persistence of a corpus luteum or an extended follicular phase.

Corynebacterium pseudotuberculosis phospholipase D (PLD) significantly affected viability of ovine neutrophils after 24 hours' exposure, This effect was more marked in cells that ingested PLD emulsified in oil. Treatment of neutrophils with PLD significantly (P < 0.05) reduced the ability of these cells to migrate toward activated sheep serum. The PLD was not chemotactic, but it activated normal sheep serum, producing factors that were chemotactic for neutrophils.

Streak retinoscopy was performed by 5 ophthalmologists on 256 eyes (191 dogs) to determine their postoperative refractive state after cataract extraction. Aphakic and pseudophakic eyes that had been implanted with 1 of 5 intraocular lenses (IOL) with dioptric powers ranging from +14.5 to +38 diopters (D) were studied. By use of ANOVA, breed and body type of dog and individual performing refraction were found to have no detectable effect on final refractive state. Mean refractive state of aphakic eyes was +14.4 +/- 2.10 D. Mean refractive state for different IOL powers was as follows: +14.5 D IOL = +11.54 +/- 1.18 D (n = 13); +30 D IOL = + 5.15 +/- 1.18 D (n = 105); +34.0 D IOL = +3.5 D (n = 1); +36 D IOL +2.34 +/- 0.73 D 9 (n = 61); and +38 D IOL = +1.41 +/- 0.56 D (n = 28). Residual hyperopia ranged from +0.5 D to +2.5 D with +38 D IOL, and no eyes were myopic (overcorrected) by use of any of the IOL studied. linear regression analysis of refractive state on IOL power for aU dogs predicted that dioptric strength of +41.53 D was necessary to best approximate emmetropia for the population as a whole. Body type of the dog had only slight effect (< 1.0 D) on predicted optimal IOL power. Further linear regression analysis of the 7 breeds studied predicted variations from +39.62 to +43.14 D in IOL powers necessary to approximate emmetropia. Results of the study support the routine use of canine IOL with dioptric strength of approximately +41.5 D in circumstances in which preoperative biometry and keratometry are not practical. The findings further suggest that, for the specific population of dogs studied, most of the dogs could be corrected to near emmetropia by use of a small range of IOL dioptric strengths, irrespective of body type or breed.

Ground reaction force (GRF) patterns from 20 clinically sound Dutch Warmbloods were recorded at the right fore-leading canter, and a standard horse was composed. These GRF data for the standard can be used for evaluation of jumping horses. The GRF patterns were asymmetric for all 4 limbs. The leading right forelimb decelerated the body. The trailing left forelimb propelled the body and decelerated it slightly. The trailing left hind limb propelled, and the leading right hind limb contributed to deceleration and propulsion. Referred to the maximal vertical load of the leading right forelimb, the load of the trailing left forelimb was 25% more; the load of the right hind limb was slightly less, whereas the load of the left hind limb was about 80% of that value.

Flow-volume loops generated from 6 Standardbreds at rest and during treadmill exercise were evaluated for their use in detecting upper airway obstruction. Tidal breathing flow-volume loops (TBFVL) were obtained from horses at rest and exercising at speeds corresponding to 75% of maximal heart rate and at maximal heart rate. The TBFVL were evaluated, using a pulmonary function computer; calculated indices describing airflow rate and expiratory-to-inspiratory airflow ratio for individual loops were determined. In addition to TBFVL indices, standard variables of upper airway function also were measured: peak airflow, peak pressure, and calculated inspiratory and expiratory impedances. Measurements were recorded before left recurrent laryngeal neurectomy (LRLN; baseline) and 14 days after surgically induced left laryngeal hemiplegia. When horses were at rest, TBFVL shape and indices describing the loop were highly variable. In contrast, in exercising horses, TBFVL shape was consistent and coefficients of variation of loop indices were less during exercise than at rest. After LRLN, TBFVL from exercising horses indicated marked inspiratory airflow limitation, while the expiratory airflow curve was preserved. Peak inspiratory flow rate and inspiratory flow at 50 and 25% of tidal volume decreased, and the ratio of peak expiratory to inspiratory airflow and that of midtidal volume expiratory and inspiratory airflow rates increased significantly (P < 0.05). Inspiratory impedance also increased after LRLN. Although in resting horses TBFVL were not a useful indicator of upper airway obstruction, examination of TBFVL from exercising horses allowed objective, specific, and repeatable detection of upper airway obstruction. The technique was noninvasive, rapid, and well tolerated by horses; thus, it is a potentially valuable clinical diagnostic test.

An intramammary device (IMD) was adapted for use in ewes; this device was made of abraded poly. ethylene material (1.7 mm in diameter, 47 mm long) and formed a 15-mm-diameter loop in the gland cistern. The IMD was inserted in 1 gland in each of 43 ewes. A significant (P < 0.0001) increase in milk somatic cell count (SCC) was observed in glands provided with an IMD. This increase was attributable to an increase in neutrophil numbers and was observed during the first 12 weeks after insertion. The IMD had a protective effect against experimentally induced staphylococcal mastitis (Staphylococcus aureus and S epidermidis), although different milk SCC were required for protection from each bacterial species in most ewes (10(6) and 2 X 10(5) cells/ml, respectively). Histologic studies revealed that the IMD induced local squamous metaplasia in the glandular part of the lactiferous sinus. Erythrocytes were found in milk from glands provided with an IMD throughout the studied period (35 days of the 45-day lactation) and, in some cases, blood clots were observed during the first 2 weeks of lactation. Glands with IMD also had lower milk production and quality at 30 and 32 days of lactation. Eight ewes with IMD were studied throughout a subsequent lactation. Milk from the IMD-containing glands had an increase in SCC, as in the previous lactation period; did not contain blood clots or erythrocytes; and had normal composition (similar to that in glands without the IMD).

To study behavioral and cardiopulmonary characteristics of horses recovering from inhalation anesthesia, 6 nonmedicated horses were anesthetized under laboratory conditions on 3 different days, with either halothane or isoflurane in O2. Anesthesia was maintained at constant dose (1.5 times the minimum alveolar concentration [MAC]) of halothane in O2 for 1 hour (H1), halothane in O2 for 3 hours (H3), or isoflurane in O2 for 3 hours (13). The order of exposure was set up as a pair of Latin squares to account for horse and trial effects. Circulatory (arterial blood pressure and heart rate) and respiratory (frequency, PaCO2, PaO, pHa) variables were monitored during anesthesia and for as long as possible during the recovery period. End-tidal percentage of the inhaled agent was measured every 15 seconds by automated mass spectrometry, then by hand-sampling after horses started moving. Times of recovery events, including movement of the eyelids, ears, head, and limbs, head lift, chewing, swallowing, first sternal posture and stand attempts, and the number of sternal posture and stand attempts, were recorded. The washout curve or the ET ratio (end-tidal percentage of the inhaled agent at time t to end-tidal percentage of the inhaled agent at the time the anesthesia circuit was disconnected from the tracheal tube) plotted against time was similar for HI and H3. The slower, then faster (compared with halothane groups) washout curve of isoflurane was explainable by changes in respiratory frequency as horses awakened and by lower blood/gas solubility of isoflurane. The respiratory depressant effects of isoflurane were marked and were more progressive than those for halothane at the same 1.5 MAC dose. During the first 15 minutes of recovery, respiratory frequency for group-13 horses increased significantly (P < 0.05), compared with that for the halothane groups. For all groups, arterial blood pressure increased throughout the early recovery period and heart rate remained constant. Preanesthesia temperament of horses and the inhalation agent used did not influence the time of the early recovery events (movement of eyelids, ears, head, and limbs), except for head lift. For events that occurred at anesthetic end-tidal percentage < 0.20, or when horses were awake, temperament was the only factor that significantly influenced the nature of the recovery (chewing P = 0.04, extubation P = 0.001, first stand attempt P = 0.008, and standing P = 0.005). The quality of the recoveries did not differ significantly among groups (H1, H3, I3) or horses; however 5 of 6 horses recovering from the H1 exposure had ideal recovery. During recovery, the anesthetic end-tidal percentage did not differ significantly among groups. However, when concentrations were compared on the basis of anesthetic potency (ie, MAC multiple) a significantly (P < 0.05) lower MAC multiple of isoflurane was measured for the events ear movement, limb movement, head lift, and first attempt to sternal posture, compared with that for horses given halothane, indicating that isoflurane may be a more-potent sedative than halothane in these horses.

Four-week-old chickens were inoculated orally with 1,000 or 100,000 oocysts of the ME-49 or GT-1 strain of Toxoplasma gondii, and their antibody responses were measured, using the direct modified agglutination test, latex agglutination test, indirect hemagglutination test, ELISA, and the Sabin-Feldman dye test. Antibodies against T gondii were detected by use of the modified agglutination test and ELISA within 2 weeks of oocyst inoculation, and antibodies persisted until termination of the study by postinoculation day 68. The latex agglutination test was insensitive in detecting T gondii antibodies, and antibodies were not detected by use of the dye and indirect hemagglutination tests. Of tissues bioassayed in mice for tissue cysts by pepsin digestion of individual organs of chickens on postinoculation day 68, tissue cysts were found in the brain of all 5, heart of 3, and leg muscles of 2, but not in the liver and breast muscles. None of the birds developed clinical toxoplasmosis.