An efficient method for DNA transfer is essential for the genetic manipulation of any organism. Such a capacity will be required for the genetic analysis of Actinobacillus pleuropneumoniae as a swine pathogen, as well as for its manipulation for vaccination purposes. For this reason, the use of electroporation as a means of plasmid DNA introduction into this species was examined. The multiply antibiotic-resistant strain 80-8141 of Actinobacillus pleuropneumoniae harbors 3 plasmids: pYG10, pYG15, and pYG12 of 5.0, 2.7, and 2.5 kb, respectively. Electroporation of A pleuropneumoniae strain 4074 with a plasmid extract of strain 80-8141 showed that pYG10 encodes chloramphenicol resistance and that pYG12 encodes ampicillin resistance. Electrical pulse conditions for efficient electroporation of strain 4074 were examined by use of pYG10 DNA isolated from a 4074 transformant. Efficiency, expressed as transformants per microgram of plasmid DNA, increased directly with pulse amplitude. However, high efficiencies were only observed in a narrow window of pulse duration (gamma = 12 to 22 ms at 6.25 kV/cm). Longer pulse durations resulted in cell death. Electroporation efficiencies increased with cell density. Yield of transformants increased directly with DNA concentration. Results indicate that electroporation can be used to efficiently transform A pleuropneumoniae and that pYG10 and pYG12 are suitable plasmid vectors for use in the genetic manipulation of this organism. Actinobacillus (Haemophilus) pleuropneumoniae is a prominent cause of respiratory infections in swine. Clinical isolates of A pleuropneumoniae have been reported to be resistant to tetracycline, triple sulfonamides, ampicillin, and streptomycin. There has been particular concern over the increasing incidence of resistance to chloramphenicol, which may be related to the extensive use of this antibiotic for treatment of swine pleuropneumonia. In 1980, 95% of the strains of A pleuropneumoniae isolated from the St-Hyacinthe region of Quebec, Canada, were found to be sensitive to chloramphenicol; whereas in 1982, only 57% of surveyed strains were still sensitive to this antibiotic. Resistance to ampicillin, streptomycin, and sulfadiazine in A pl europneumoniae strains has been shown to be plasmid-mediated. The purpose of the study reported here was to use electroporation to analyze plasmids carried by a multiply antibiotic-resistant clinical isolate of A pleuropneumoniae. Electroporation involves the use of brief high-voltage electrical discharges to induce reversible permeability in both prokaryotic and eukaryotic membranes. Using a 5.0-kb A pleuropneumoniae plasmid encoding resistance to chloramphenicol, we have optimized electroporation as a means to transform this species. Conditions permitting an efficiency of over 10(5) transformants (Tfs)/microgram of plasmid DNA are described.

Somatosensory-evoked potentials (SEP) and spinal cord-evoked potentials (SCEP) were recorded in clinically normal adult cats in response to electrical stimulation of pudendal and tibial nerves to provide normative data that can be used in a clinical evaluation of pudendal nerve function in cats after sacral or sacrococcygeal luxations or fractures. Responses to tibial nerve stimulation were included in the study as an internal control because it is usually not involved in these types of injuries and because its SEP and SCEP are easily recorded. Evoked potentials were characterized by the latencies (ms) of positive (P or p) and negative (N or n) peaks. The SEP resulting from percutaneous pudendal nerve stimulation consisted of a prominent P-N-P potential in the 30- to 80-ms range. The pudendal SCEP was not successfully recorded because of large muscle artifacts evoked from the sacral area. The tibial SEP was similar to the pudendal SEP, except that the prominent P-N-P series in the 35- to 81-ms range was preceded by a smaller p-n-p-n sequence in the 7- to 23-ms range. The tibial SCEP consisted of a P-N-P series in the 2- to 4-ms range.

Eight bovine hearts with lesions of eosinophilic myositis (EM) and 2 bovine hearts without EM lesions were collected at slaughter. Blood samples from these 10 hearts, and the heart of a newborn calf also were collected. Histologically, Sarcocystis cruzi was identified in the 8 hearts with EM lesions and the 2 hearts without EM lesions, but not in the heart of the newborn calf. Serum was harvested from the 10 blood samples and was used in homologous, modified, passive cutaneous anaphylaxis test. Antigen was prepared from S cruzi bradyzoites isolated from the 2 hearts without EM lesions. Serum samples from the 8 cattle with EM lesions reacted positively to S cruzi antigen. When heat-inactivated IgE in serum (56 C for 4 hours) was used, all passive cutaneous anaphylaxis responses were considered negative. Using ELISA, serum IgE concentrations from the 10 cattle with and without EM lesions were 2.2 to 9 U/ml. As determined by radial immunodiffusion, IgM concentrations were 80 to 215 mg/dl. Immunoglobulin G concentrations were 420 to 2,050 mg/dl, but most were less than or equal to 1,700 mg/dl. Immunoglobin A concentrations were 0 to 62 mg/dl; 1 steer with EM lesions had 0 mg/dl. Double-gel immunodiffusion confirmed the presence of Sarcocystis-specific precipitating antibodies. Sera from the 10 cattle with and without EM lesions formed at least 1 precipitin band.

Cell proliferation kinetic values were established for the epidermis, hair follicle epithelium, and sebaceous glands of 10 Beagles and 4 Cocker Spaniels with healthy skin and coats. Values were established by intradermal pulse-labeling injections of [3H]thymidine, examination of cutaneous biopsied tissues, and autoradiography. The epidermal basal cell-labeling index was 1.41 +/- 0.46% for Beagles and 1.71 +/- 0.56% for Cocker Spaniels. The hair follicle basal cell-labeling index was 1.46 +/- 0.78 and 1.07 +/- 0.42%, respectively. Calculated epidermal cell-renewal time for viable layers of the epidermis was 23.38 +/- 5.93 days for Beagels and 20.97 +/- 4.92 days for Cocker Spaniels. Differences between cell kinetics data for the 2 breeds were not significant (P greater than 0.05). The basal cell-labeling index for the sebaceous gland was significantly (P = 0.009) lower for Cocker Spaniels (0.40 +/- 0.18%) than for Beagles (1.81 +/- 1.08%). Seemingly, epidermal and follicular cell proliferation kinetics in healthy dogs was similar between the 2 breeds, whereas sebaceous gland basal cells were less proliferative in healthy Cocker Spaniels than in healthy Beagles.

Serum concentrations of relaxin and progesterone were measured by specific radioimmunoassays in pregnant, pseudopregnant, or ovariectomized (between gestation weeks 4 and 5) pregnant Labrador Retriever bitches. Daily administration of 17 alpha-ethyl-19-nortestosterone was performed to maintain gestation in the ovariectomized pregnant bitches. This synthetic gestagen was selected because it did not interfere with the assay for endogenously secreted progesterone concentration in serum. Serum progesterone concentration was high in ovarian-intact pregnant or pseudopregnant bitches, but the mean progesterone concentration in pseudopregnant bitches (evaluated at 4 weeks after mating) was only 56% of the concentration in pregnant bitches. After ovariectomy, serum progesterone concentration decreased to undetectable values. Unlike progesterone, serum relaxin concentration increased during the latter half of pregnancy in the ovarian-intact and in the ovariectomized pregnant bitches, but relaxin was not detectable at any time in the pseudopregnant bitches. The amount of relaxin measured in the ovariectomized pregnant bitches was less (P less than 0.05) than that in ovarian-intact bitches, suggesting that the ovaries may have contributed to the total circulating relaxin concentration in the latter. Placental production of relaxin might have accounted for the serum relaxin concentration after ovariectomy; thus, the ovary and placenta each may secrete relaxin during gestation in bitches. Regardless of its source, measurement of serum relaxin concentration may offer a useful way of distinction between pregnancy and pseudopregnancy in dogs.

A series of coupled differential equations was used to model the temporal dynamics of rabies in raccoons in the mid-Atlantic region of the United States. The model takes explicit account of the development of natural immunity to rabies and was used to evaluate culling and vaccination elimination strategies. For habitats typical of the mid-Atlantic states, and given the assumptions of the model, it was estimated that elimination of rabies in raccoons by culling may involve the annual removal of over 32% of the raccoon population or the yearly vaccination of up to 99% of the susceptible fraction. Assuming a constant marginal cost for both culling and vaccination, the model suggests that, whatever the actual cost of each method, the cheapest strategy will always involve either culling or vaccination alone. A combined strategy of culling and vaccination will be cheaper than culling alone only when the per capita cost of vaccination is around one-fifth or less the per capita cost of culling.

A study on erythrocyte distribution was performed on 10 healthy, nonstressed adult white Pekin ducks. Results indicated 2 populations of erythrocytes, with average mean corpuscular volumes of 128.37 fl/cell and 308.50 fl/cell. Variations in erythrogram patterns were evident over time, when comparing specimens from different ducks or the same duck. There were 5 patterns of cell number/volume distribution observed between the 2 cell populations when all ducks were studied. Females had a greater change than did males when population density and volume percentage comparisons were made on erythrocyte compartments.

Data from slaughter plants (n = 3) and feedlots (n = 18) in eastern Washington were analyzed to characterize occurrence patterns of cysticercosis in Washington during 1984. Three concurrent peaks in cysticercosis rates (0.6/1,000 to 5/1,000 slaughtered cattle) were detected at 3 slaughter plants. Peaks were observed at 8 feedlots from December 1983 to March 1984, at 6 feedlots from April to July 1984, at 2 feedlots from August to October 1984, and at 3 feedlots from November 1984 to February 1985. Affected feedlots were not closely associated geographically and were feeding cattle from many, predominantely northwestern, origins. For 3 feedlots for which time in the feedlot was available for each slaughter shipment, an increase in cysticercosis rate with increasing time in the feedlot was noticed. Within these 3 feedlots, cases of cysticercosis were widely scattered spatially. The pattern of cysticercosis indicated human fecal contamination of a regionally available feed source. Of feedstuffs in use, potato waste, a byproduct of the processed potato industry, appeared to be the most likely source of Taenia saginata ova.

Exogenously administered vasopressors (sympathomimetics) were evaluated in halothane-anesthetized dogs to determine the effects of these drugs on cardiovascular function before and after hemorrhage. Six dogs were anesthetized with thiamylal sodium (20 mg/kg of body weight) and halothane (1.25 minimal alveolar concentration) in 100% oxygen. After instrumentation, cardiac output, systemic arterial blood pressure (SAP), heart rate (HR), left ventricular pressure, pulmonary arterial pressure, and an index of cardiac contractility (dP/dT) were measured. Stroke volume, cardiac index (CI), stroke index (SI), rate-pressure product, and systemic vascular resistance (SVR) were calculated. Epinephrine (0.1, 0.3, and 0.5 micrograms/kg/min [low, medium, and high doses, respectively]) and dobutamine (1, 5, and 10 micrograms/kg/min [low, medium, and high doses, respectively]) were infused. Methoxamine was given in a bolus of 0.22 mg/kg, IV. All measurements were taken at 2.5 minutes after infusion, and were repeated after removal of 40% of the estimated blood volume. Dobutamine administered at the low dose before hemorrhage increased SAP and dP/dT. At the high and medium dose, dobutamine significantly increased CI, dP/dT, and SAP with no significant change in HR or SVR. The medium dose of epinephrine was the most effective dose of epinephrine at increasing key variables (CI, SI, dP/dT). The response of CI and SI to this dose was not significantly different from the changes seen with high-dose administration of dobutamine. The dP/dT was significantly lower with epinephrine than with dobutamine, and SVR and HR were unchanged with epinephrine, except at the low dose, which decreased SVR. Methoxamine significantly decreased CI, SVR, and HR, whereas SVR and SAP were increased significantly. After hemorrhage, the only variables that had a significant change in the absolute magnitude of the response to a drug, relative to the response before hemorrhage, were a significantly reduced ability of dobutamine and methoxamine to increase SAP, and a significantly decreased ability of methoxamine to decrease CI. We concluded that dobutamine and epinephrine provide beneficial short-term support of the cardiovascular system in the halothane-anesthetized dog during acute hypovolemia.

Nucleic acid hybridization, bacteriologic culture, and a fluorescent antibody test were compared for detection of Leptospira interrogans serovar hardjo type hardjo-bovis in bovine urine. Seventy-five urine samples were collected from pregnant cows challenge exposed with type hardjo-bovis. Twenty samples were collected from steers not exposed to hardjo-bovis. Sediments from each sample were examined, using fluorescent antibodies and a repetitive sequence element nucleic acid probe, to detect the presence of leptospires. Urine samples were processed for bacteriologic culture, using standard techniques. Under laboratory conditions typically used for these techniques, leptospires were detected in 60 of 75 urine samples from challenge exposed cows by nucleic acid hybridization, in 24 samples by fluorescent antibody test, and in 13 samples by bacteriologic culture. Leptospires were not detected in the urine of steers not exposed to hardjo-bovis.