Adhesion of equine spermatozoa to homologous oviduct epithelial cells (OEC) in vitro results in specific changes in spermatozoa and OEC function. To test the hypothesis that adhesion of spermatozoa affects protein synthesis and secretion by OEC, the following treatment groups were established in culture: OEC with culture medium only; control spermatozoa in culture medium only; OEC in coculture with spermatozoa; and OEC and spermatozoa in coculture, but physically separated by a microporous membrane. The experiment was replicated within each of 4 ejaculates from 3 stallions. De novo protein secretion by OEC was measured and compared by incorporation of [35S]methionine, and evaluated, using two-dimensional polyacrylamide gel electrophoresis and fluorography. Monolayers of OEC secreted a large number of proteins of molecular mass ranging from 14 to 205 kd. Adhesion of spermatozoa consistently caused reduced synthesis of 2 OEC secretory proteins and new or increased synthesis of 6 proteins. When spermatozoa and OEC were separated by a microporous membrane, some but not all of these changes were duplicated. Synthesis of 3 OEC secretory proteins, unaffected by binding of spermatozoa, was reduced when spermatozoa were prevented from contact with OEC by a microporous membrane. Adhesion of equine spermatozoa to homologous OEC monolayers and presence of equine spermatozoa resulted in qualitative and quantitative changes in synthesis and secretion of proteins by OEC. These changes have implications for storage, longevity, and maturation of spermatozoa.

Objective--To test the hypothesis that endothelium-derived nitric oxide modulates vasomotor reactivity in equine digital arteries. Design--Digital arteries were isolated from adult horses, and their vasodilator properties were examined in an in vitro controlled environment. Animals--Five adult horses (1 gelding, 4 mares) without evidence of hoof or vascular disease were studied. Procedure--Arterial rings with or without endothelium were exposed to endothelium-dependent vasodilator drugs in the presence or absence of a pharmacologic inhibitor of the enzyme nitric oxide synthase. Results--Vasodilator effects of 3 endothelium-dependent vasorelaxant agents were significantly greater in endothelium-intact vessels than in endothelium-denuded vessels. Moreover a nitric oxide synthase inhibitor reduced vasodilator responses to endothelium-dependent vasodilators in endothelium-intact arteries, but had no discernable effects in endothelium-denuded arteries. Conclusions--These findings indicate the presence of endothelium-derived relaxing factor/nitric oxide in blood vessels of horses, and identify vascular endothelium as an endogenous modulator of vasomotor tone in the digital arteries of this species.

An avirulent live Salmonella choleraesuis culture (SC-54) was evaluated for use as an effective vaccine in preventing salmonellosis caused by S choleraesuis in pigs. Eighty-two pigs, 3 to 4 weeks old, were randomly assigned to 1 of 2 treatment groups, which were designated as either vaccinates or controls. After vaccination, all pigs were examined for fecal shedding of S choleraesuis, rectal temperature, and 10 clinical variables. Significant difference was not detected between vaccinated and nonvaccinated pigs for 14 days (phase I) after intranasal administration of the vaccine. Efficacy and duration of immunity were examined by intranasally challenge exposing respective pigs from either treatment group with a virulent field isolate of S choleraesuis at 2, 8, or 20 weeks after vaccination (phases II-IV). Pigs were again evaluated for 14 days after challenge exposure, and 10 clinical variables and rectal temperature were monitored. Surviving pigs were euthanatized and evaluated for gross lesions, and samples of 7 organs were collected. These organ samples were homogenized, and level of S choleraesuis infection was determined. After virulent challenge exposure during phases II-IV, the clinical status of the SC-54 vaccinates was significantly (P < 0.05) superior to that of nonvaccinates for rectal temperature, feces consistency, behavior, appetite, body condition, and mean score for the 10 clinical variables. Quantitative bacteriologic culture of the tonsil, lung, liver, spleen, mesenteric lymph nodes, ileum, and colon samples indicated consistent reduction of organ colonization in vaccinates; bacteria numbers in the mesenteric lymph nodes, lungs, and ileum were significantly (P < 0.05) reduced. Gross lesions in pigs indicated reduction of pneumonia in vaccinates. Pigs also had consistent weight gain throughout all phases of the study after challenge exposure, although the differences were not significant. In conclusion, a single intranasally administered dose of SC-54 given to 3- to 4-week-old pigs proved to be safe and efficacious and to provide protection to pigs at least 20 weeks after initial vaccination.

Twenty-four, adult, female Beagles were arranged by body weight from greatest to least and allocated to 2 groups of 12 dogs, using random numbers. Dogs were housed collectively in 2 adjacent metal buildings, each divided into 4 rooms measuring 2.1 X3.7 m. Each room was paneled and carpeted and had an access door to the outside with a connecting run that measured 2.1 X 9.1 m. Each run had a surface consisting of 5 cm of pea gravel overlayng 5 cm of sand, and was partially covered by an awning that provided shade at its proximal end. For placement in room/run units, dogs in each of the treated and control groups were allotted to 4 subgroups of 3 dogs each. Each subgroup of dogs was placed in a separate room/run unit. Units containing treatment or control subgroups were alternated to avoid placing identically treated subgroups adjacent to each other. Dogs of subgroups A, C, E, and G were treated with lufenuron monthly at a minimal target dosage of 10 mg/kg of body weight; those of subgroups B, D, F, and H were treated with excipient tablets. Dogs were treated on study days 7, 37, 68, and 98. Each dog was infested with 100 newly emerged, unfed, insectary-reared adult Ctenocephalides felis on each of study days 0 and 2. Thereafter, infestations on all dogs were dependent on continued development of fleas either in the indoor or outdoor environment. Numbers of fleas on each of the treated and control dogs were determined, using a nondestructive counting technique on days 6, 14, 21, 28, 35, 56, 70, 84, 98, 112, and 119. On study day 21 and on each collection day thereafter, numbers of adult fleas recovered from treated dogs were significantly (P < 0.05) fewer than those recovered from control dogs. Proportion reduction of fleas on treated vs control dogs exceeded 90% by study day 35 and 95% by study day 56. Efficacies exceeded 95% on all remaining study days except days 98 (94.4% and 119 (90%). Results of this study indicate that control of flea populations can be achieved in treated dogs approximately 4 to 5 weeks after initial treatment with lufenuron, and that continued monthly treatments will maintain effective control of flea infestations. Adverse reactions or side effects to treatment with lufenuron were not observed in dogs after treatment at any time throughout the study.

A surgical technique was developed for implanting a flexible polyurethane cannula in a lateral ventricle in the brain of calves. Initially, measurements were made on 25 calves at necropsy to develop equations for calculating coordinates for cannula placement. The distance (cm) caudal, in the sagittal plane, from the coronal suture line to the center of a hole to be drilled in the parietal bone of the skull was: 0.73 + (0.00925 X body weight [kg]). The distance (cm) lateral from the midline to the center of the hole to be drilled was: 0.018 + (0.6464 X distance caudal). The depth (cm) from the surface of the skull to the dorsal surface of the lateral ventricle was: 2.29 + (0.0159 X body weight [kg]). Surgery was subsequently performed on 17 calves. A 5-mm-diameter hole was drilled through the skull with a hand trephine at coordinates derived from the aforementioned regression equations. A polyurethane cannula (total length, 30 cm; 1 mm ID; 2 mm OD) covering a stainless-steel 20-gauge blunt-tipped needle (stylet) was lowered through the brain and into a lateral ventricle at an angle of 20.5 degrees relative to the frontal bones of the skull. The blunt-tipped needle was then removed, and CSF was allowed to drip from the cannula to verify placement. One stainless-steel screw was inserted 0.6 cm medial, and another was inserted 0.6 cm caudal to the hole in the skull. The area around the cannula, bone screws, and hole in the skull was covered with dental acrylic (approx 2 cm in diameter) to stabilize the cannula. With minimal restraint of calves, injection of substances into and withdrawal of CSF from a lateral ventricle of the brain were possible in most calves for at least 6 weeks after surgery was performed.

A total of 22 clinical streptococcal isolates, predominantly Streptococcus zooepidemicus, associated with endometritis in horses were tested for their ability to withstand the natural bactericidal properties of freshly obtained blood. During a 3-hour incubation in blood from a single horse, 8 of these isolates survived and grew; the remainder were killed. To determine whether this ability to grow extended to blood of other horses, 5 of these growing isolates were tested for their ability to grow in the blood of 5 additional horses. The same 5 horses were used for each isolate. The isolates grew in blood of some of the horses, but were killed in blood of the others. However, the horse's blood that mediated killing was different for each isolate. Killing required leukocytes, but the specificity for killing appeared to reside in plasma, although plasma by itself was not bactericidal. Heat-stable and heat-labile components in plasma, interpreted as antibody and complement, respectively, appeared necessary for killing. Isolates that could grow in fresh blood lost this ability after 10 passages in artificial media. Results of these experiments of phagocytosis in fresh blood may provide helpful insights into the phagocytosis of S zooepidemicus in equine uterine f1uid.

The stability of ionized calcium (CaI) concentration and pH in sera (n = 14) stored at 23 or 4 C for 6, 9, 12, 24, 48, or 72 hours, or -10 C for 1, 3, 7, 14, or 30 days was evaluated. Also studied were the effects of oxygen exposure, cold handling, and feeding on CaI and pH values. Results indicated that serum CaI concentration was stable throughout 72 hours of storage at 23 or 4 C, and for 7 days at -10 C. Serum CaI concentration significantly (P < 0.05) decreased by 14 days of storage at -10 C. Serum pH was stable for 6 hours at 23 or 4 C, and for 24 hours at -10 C, but significantly (P < 0.05) increased by 9 hours of storage at 23 or 4 C and by 3 days at -10 C. Exposure of the surface of the serum to air immediately before measurement had no effect on CaI or pH values, but mixing serum with air resulted in significantly (P < 0.05) decreased CaI concentration and increased pH. Handling of blood on ice resulted in significantly (P < 0.05) higher serum pH, compared with blood handled at 23 C, but serum CaI concentration was unaffected. Serum obtained at 2 hours after feeding did not have any significant changes in CaI, total calcium, or pH values. It appears that if canine serum is obtained, handled, and stored anaerobically, CaI concentration can be accurately measured after 72 hours at 23 or 4 C, or after 7 days at -10 C.

Milk antimicrobial residues are a serious concern for the dairy industry. Residues of the tetracycline family of antimicrobials have been reported in market milk by investigators, using radioimmunoassay and microbial receptor technology (hereafter referred to as the Charm II test). In response to these reports, an investigation was conducted to determine the potential of 3 extra-label routes of oxytetracycline (OTC) administration to cause milk residues above the Food and Drug Administration safe value of 30 parts per billion (ppb). Lactating Holstein cows were administered OTC once by use of 1 of 3 routes: IV at 16.5 mg/kg of body weight (n = 6); IM at 11 mg/kg (n = 6); and intrauterine (IU) at 2 g in 500 ml of saline solution/cow (n = 6). Duplicate milk samples were collected at the milking prior to drug administration and for the next 13 milkings at 12-hour intervals. Concentrations of OTC in milk samples were analyzed by use of the Charm II test for tetracyclines (limit of OTC detection, approx 5 ppb) and were compared with concentrations determined by use of a high-performance liquid chromatography (HPLC) method (lower limit of OTC quantitation, approx 2 ppb). The potential for milk OTC residues above the Food and Drug Administration safe value of 30 ppb after treatment was considerably greater for the IV and IM routes, compared with the IU route. Mean peak OTC concentrations in milk at the first milking after treatment for the HPLC and Charm II tests were approximately 3,700 to 4,200 ppb for the IV route, 2,200 to 2,600 ppb for the IM route, and 186 to 192 ppb for the IU route, respectively. Pharmacokinetic analysis, based on milk OTC concentrations, indicated that the area under the curve (AUC) and milk maximal concentration (Cmax) differed significantly (P < 0.001) among routes of administration. The AUC was similar for IV and IM administrations; values for both were greater than the AUC for IU administration. The Cmax was greatest for IV, intermediate for IM, and least for IU administration. There were significant (P less than or equal to 0.01) differences in AUC between assay methods (Charm II vs HPLC) for the IV route. Concentrations of OTC in milk determined by the Charm II test were often greater than those determined by HPLC. Administration of OTC to lactating cows via these routes is extra-label drug use. Failure to withhold the product from early milkings of cows administered OTC by the IV or IM route should be considered a potential cause of OTC residues in market milk. Milk from nearly all cows contained OTC (< 30 ppb), the Food and Drug Administration safe level, by 120 hours after OTC administration. Use of appropriate withholding times and antibiotic residue testing is indicated to avoid OTC residues.

Intraocular production of Toxoplasma gondii-specific antibody in cats has been estimated by comparing the ratio of T gondii-specific antibody in aqueous humor and serum with the ratio of total immunoglobulins in serum and aqueous humor (Goldmann-Witmer coefficient; aqueous antibody coefficient; C value). It has been proposed that in human beings, comparison of the ratio of T gondii-specific antibody in aqueous humor and serum with the ratio of antibodies against a nonocular pathogen in serum and aqueous humor is more accurate than methods using total immunoglobulin quantification. We developed an ELISA for detection of calicivirus-specific antibodies in the serum and aqueous humor of cats. By evaluating calicivirus-specific antibody concentrations in the aqueous humor of healthy and diseased cats, calicivirus was assessed as a nonintraocular pathogen. The ratio of T gondii-specific antibodies in the aqueous humor and serum and the ratio of calicivirus-specific antibodies in serum and aqueous humor were evaluated as a means of estimating intraocular T gondii-specific antibody production. A field strain of feline calicivirus was isolated, cultured, and purified. A calicivirus-specific IgG ELISA was developed for detection of feline calicivirus-specific IgG in serum and aqueous humor. Calicivirus-specific IgG was measured in the serum and aqueous humor from 3 groups of control cats. Results suggested that calicivirus is a nonintraocular pathogen in cats and that calicivirus IgG detected in aqueous humor is attributable to leakage across a damaged blood-ocular barrier. Intraocular production of T gondii-specific antibodies was estimated, using 2 formulas. The C value was calculated by multiplying the ratio of T gondii-specific IgM or IgG in aqueous humor and serum by the ratio of total immunoglobulins (using the corresponding IgM or IgG class) in serum and aqueous humor. The Ctc value (Toxoplasma-calicivirus Goldmann-Witmer coefficient) was calculated by multiplying the ratio of T gondii-specific IgM or IgG in aqueous humor and serum by the ratio of calicivirus-specific IgG in serum and aqueous humor. Serum and aqueous humor samples were obtained from 41 client-owned cats with uveitis, and T gondii-specific C values and Ctc values were calculated. Toxoplasma gondii-specific IgM or IgG C values of 10 or greater or T gondii-specific IgM or IgG Ctc values of 1 or greater were considered to be suggestive of intraocular T gondii-specific antibody production. Of the 41 cats, 20 (48.7%) had evidence of intraocular production of T gondii-specific antibody on the basis of either an IgM or IgG C value of 10 or greater. A Ctc value could not be calculated in 3 cats because calicivirus-specific IgG was not present in aqueous humor. Of the 38 cats for which Ctc values could be calculated, 25 (65.8%) had evidence of intraocular production of T gondii-specific antibody on the basis of either an IgM or IgG Ctc value of 1 or greater. The C values and Ctc values were in agreement for 75.9% of IgM containing samples and 75% of IgG containing samples. Sensitivity, specificity, predictive value of a positive test result, and predictive value of a negative test result for an IgM or IgG C value, when compared with the corresponding IgM or IgG Ctc value were determined. The results indicate that use of the C value for estimation of intraocular T gondii-specific antibody production will result in 28.6 (IgM) to 50% IgG) false-negative results and 12.5% (IgM and IgG) false-positive results, when compared with the Ctc value.

A virologic survey was conducted on calves with diarrhea associated with bovine rotavirus (BRV) on a closed dairy farm. The BRV was detected from 32 of 219 (14.6%) fecal specimens repeatedly collected from 56 calves born during the years 1992-1993, regardless of whether they had diarrhea. Most of the 32 strains were isolated from fecal specimens obtained from 2-to 6-week-old calves. After electrophoresis of doublestranded viral RNA from the 32 strains, genomic RNA migration patterns were similar to those of the predominant BRV strains isolated at the same farm during the years 1990-1991. All representative strains were identified as G serotype 6 (G6) and P type 5 (P5) by results of the virus-neutralization test and polymerase chain reaction procedure. Thus, BRV had no change in genomic RNA electropherotypes and serologic antigenicities in a closed dairy herd over a period of several years.