We studied the uptake and sequential transneuronal passage of pseudorabies virus (PRV) in rat CNS by use of a combination of immunohistochemistry and in situ hybridization. Protocols for rapid detection of PRV by immunohistochemistry and in situ hybridization in rats with PRV infection of the CNS after intranasal instillation of a wild-type strain of PRV were optimized in vitro, using porcine kidney-15 cells. Pseudorabies virus-specific hybridization signals appeared in the cytoplasm and nucleus of PRV-infected porcine kidney-15 cells by postinoculation (PI) hour 6. In tissue sections of PRV-infected rats, PRV nucleic acids were detected in areas of the rat brain in close proximity to the areas in which PRV antigens were evident. The PRV was initially found in the nucleus of trigeminal ganglion neurons at PI hour 24. At PI hour 72, PRV antigens were observed in the mid-brain, and 24 hours later, in the telencephalon. We also found evidence of specific progressive transsynaptic transmission of the virus, and, on the basis of that, we have constructed a map of the synaptic contacts and pathways in the brain. Therefore, combined use of immunohistochemistry and in situ hybridization was useful for characterizing the pathogenesis of PRV in the CNS of rats after intranasal inoculation, following a pattern that mimics PRV infection of the natural host.

Systemic administration of dexamethasone led to a significant reduction in the size of the tuberculin reaction in response to intradermal injection of bovine purified protein derivative in 18 cattle experimentally sensitized to Mycobacterium bovis (P < 0.01) and 8 cattle naturally infected with M bovis (P < 0.001). The reaction in 6 of the 7 M bovis-infected cattle that received dexamethasone was classified as negative for the standard interpretation of the single intradermal comparative tuberculin test. Significantly fewer BoCD2+ (P < 0.05) and BoCD4+ T cells (P < 0.001) were present at the reaction site and in blood of dexamethasone-treated cattle, compared with untreated control cattle. Significantly fewer cells expressing the interleukin-2 receptor and WC1+ gamma delta T cells (P < 0.001), and a significantly greater number of cells expressing the ACT2 antigen (P < 0.05) were found at the reaction site in dexamethasone-treated cattle than in controls. The number of BoCD8+ T cells at the reaction site and in blood was not significantly affected by administration of dexamethasone. In vitro production of interferon-gamma by lymphocytes incubated with bovine purified protein derivative also was significantly lower (P < 0.01) in the dexamethasone-treated cattle.

Aflatoxin (AF)-contaminated and fumonisin B1 (FB1)-contaminated (culture material from Fusarium moniliforme) diets were fed singly and in combination to growing cross-bred barrows. Six barrows (3 replicates of 2 each; mean body weight, 17.5 kg) per group were fed: 0 mg of AF and 0 mg of FB1/kg of feed (control); 2.5 mg of AF/kg of feed; 100 mg of FB1/kg of feed; or 2.5 mg of AF plus 100 mg of FB1/kg of feed for 35 days. The effects on production performance, serum biochemical, hematologic, immunologic, and pathologic measurements were evaluated. Body weight, gain, and feed consumption were significantly (P < 0.05) decreased by AF and AF plus FB1 diets. The FB1 diet decreased feed consumption, and although body weight was numerically decreased, it was not statistically significant. Aflatoxin increased serum gamma-glutamyltransferase (GGT) activity and total iron concentration and decreased urea nitrogen concentration and unsaturated iron-binding capacity. The FB1-alone diet increased serum GGT activity, whereas the AF plus FB1 diet increased serum aspartate transaminase, cholinesterase, alkaline phosphatase, and GGT activities, increased RBC count, triglycerides, and total iron concentrations, and decreased unsaturated iron-binding capacity and urea nitrogen concentration. For the most part, the effects of the AF plus FB1 diet on body weight and hematologic measurements could be considered additive. However, the effect of the AF plus FB1 diet on cholinesterase and alkaline phosphatase activities was greater than additive and was a synergistic response. One pig in the FB1-diet group and 2 pigs in the combination-diet group died. Postmortem lesions in pigs of the FB1-diet group consisted of ascites and increased liver weight. Observations at necropsy for pigs of the AF plus FB1-diet group consisted of hydrothorax, ascites, pulmonary edema, gastric erosions and ulceration, and increased liver and spleen weights. The AF diet increased relative liver weight and resulted in liver that was pale, rubbery, and resistant to cutting. Histologic lesions consisted of hepatic necrosis or degeneration, or both, with variable degrees of bile duct proliferation in barrows of the AF-diet groups. Renal tubular nephrosis was observed in barrows of the FB1 diet group, but this was not consistent in the AF plus FB1-diet group. Cell-mediated immunity, as measured by mitogen-induced lymphoblastogenic stimulation index, was decreased in barrows of the AF and FB1-diet groups, and values in barrows given the combination diet were significantly decreased from those in barrows given the single toxin diets. It was concluded that AF and FB1 (from culture material), singly or in combination, can adversely affect clinical performance, serum biochemical, hematologic, and immunologic values and induce lesions in growing barrows. For most of the variables we evaluated under our study conditions and dosages of toxins, measurements were affected more by the combination diet than by either single toxin diet, and the toxic responses could be described as additive or more than additive, particularly for induction of liver disease.

Objective-To develop a system for analysis of immune response variables in the lymph draining the lung and to establish baseline data for clinically normal calves. Design-Surgery was performed on 6 calves to insert a cannula into the efferent lymphatic duct of the caudal mediastinal lymph node to create a long-term thoracic lymph fistula draining to the exterior. Lymph was collected daily, and on the fifth postoperative day, calves were exposed to an aerosol of cell culture medium (mock infection). For the next 10 days, lymph was collected for analysis and, on the tenth day, necropsy was performed. Animals-Six 6- to 8-week-old Holstein bull calves. Procedure-Daily lymph samples were evaluated for: flow rate; total and differential cell counts; and IgG, IgM, IgA, IgE, and protein concentrations. On days -4, -1, 1, 4, 7, and 10, cells were stained and quantitated by fluorescence-activated cell sorter analysis for T, B, CD4+, and CD8+ cells. Blood lymphocytes were evaluated on days -1 and 10 for comparison. Results-Flow was established for up to 25 days, with a mean rate between 11 and 22 ml/h. Protein concentrations in lymph and plasma did not indicate a protein drain. Although mean lymphocyte counts reflected a slight gradual decrease in lymph lymphocytes, this effect was not apparent in every calf, nor was the effect seen in blood lymphocytes. There were no significant changes in IgG, IgM, IgA, or IgE concentration, with the exception of IgA concentration in 1 calf that developed an abscess at the cannulation site. The T-cell subset absolute numbers of CD4+ and CD8+ cells decreased slightly over time, but the CD4+-to-CD8+ cell ratio remained almost constant at near 2. Conclusion-Creation of a thoracic lymphatic fistula appears to be a useful technique for studying effects of lung infection on immunologic variables, with potential application to bacterial and viral respiratory tract diseases.

Objective: To examine the effects of giardiasis on production and carcass quality, using growing lambs as a domestic ruminant model. Design: Randomized block. Animals: Giardia-free lambs: 23 in infected group, 24 in control group. Procedure: Six-week-old, specific-pathogen-free lambs were infected with Giardia trophozoites; control lambs received saline solution. Clinical signs of infection, body weight, and feed intake were determined for 10 weeks. Carcass weight and quality were determined at slaughter weight of 45 kg. Results: Giardia infection persisted from weeks 7 to 16. For 5 weeks after challenge exposure, abnormal feces were more frequently observed in infected lambs. Giardia infection was associated with a decrease in rate of weight gain and impairment in feed efficiency. Time to reach slaughter weight was extended in infected lambs, and the carcass weight of Giardia-infected lambs was lower than that of control lambs. Conclusion: Giardiasis has a negative effect on domestic ruminant production. Clinical Relevance: Giardiasis in domestic ruminants is an economically important disease, thus necessitating control or elimination of the infection.

Microcytosis is a common laboratory finding in dogs with congenital portosystemic shunt (PSS), although its pathogenesis is not yet understood. Because the most common cause of microcytosis in dogs is absolute or relative iron deficiency, iron status was evaluated in 12 young dogs with PSS. Complete blood counting was done before surgical correction in all dogs, and in 5 dogs after surgery, by use of an automated hematology analyzer. Serum iron concentration and total iron-binding capacity (TIBC) were determined colorimetrically, and percentage of transferrin saturation was calculated. Erythrocyte protoporphyrin content was quantified by use of front-face fluorometry. Serum ferritin concentration was measured by use of ELISA. Serum ceruloplasmin content was determined colorimetrically (with p-phenylenediamine dihydrochloride as substrate) as an indirect indicator of subclinical inflammation, which may result in impaired iron utilization. Special stains were applied to liver (10 dogs; Gomori's) and bone marrow aspiration biopsy (7 dogs; Prussian blue) specimens for qualitative assessment of tissue iron content. Nonpaired Student's t-tests were used to compare serum iron concentration, TIBC, percentage of transferrin saturation, and erythrocyte protoporphyrin, ferritin, and ceruloplasmin concentrations in dogs with PSS with those in clinically normal dogs. All dogs had microcytosis before surgery; microcytosis resolved in 3 dogs after surgical correction. Serum iron concentration and TIBC were significantly lower, in PSS-affected dogs than in clinically normal dogs. Erythrocyte protoporphyrin, ferritin, and ceruloplasmin concentrations in PSS-affected dogs were not significantly different from those in healthy dogs. Excess iron was not detected consistently in liver or bone marrow samples. These results suggest that relative iron deficiency, perhaps associated with altered iron transport and not absolute iron deficiency, is related to microcytosis in dogs with PSS.

The effects of dexamethasone (0.2 mg/kg of body weight; IV, IM, and PO) and methylprednisolone acetate (120 mg, given intra-articularly) on serum osteocalcin and cortisol concentrations were studied in 6 horses. Serum osteocalcin and cortisol concentrations were serially monitored after each treatment. A significant (P < 0.05) decrease in serum osteocalcin and cortisol concentrations was observed from 12 to 24 and 2 to 48 hours, respectively, after IV and IM administrations of dexamethasone. Serum osteocalcin and cortisol concentrations were significantly decreased from 6 to 48 and 3 to 72 hours, respectively, after oral administration. In contrast, a change in serum osteocalcin concentration was not detected after intra-articular administration of methylprednisolone. Oral, IV, or IM treatment with 0.2 mg of dexamethasone/kg caused a decrease in serum osteocalcin concentration in horses.

Objective--To determine the relative role of endogenous prostaglandin F2 alpha (PGF2 alpha) secretion in cloprostenol-induced abortion in mares that no longer require luteal progesterone secretion for maintenance of pregnancy, and to evaluate the ability of a prostaglandin cyclooxygenase inhibitor (flunixin meglumine) to prevent cloprostenol-induced abortion. Design--The effect of flunixin meglumine on PGF2 alpha secretion and outcome of pregnancy was compared between mares treated with cloprostenol only and mares treated with cloprostenol plus flunixin meglumine. Animals--Five pregnant mares, aged 4 to 15 years, of light-horse type. Procedure--Cloprostenol (250 micrograms) was administered at 24-hour intervals to 5 pregnant mares. Flunixin meglumine (500 mg, IV) was administered at 8-hour intervals starting 15 minutes before the first cloprostenol administration. Hourly blood samples were analyzed for 15-ketodihydro-PGF2 alpha, progesterone, and estrogen concentrations. Previously reported data on cloprostenol-induced abortion in 6 pregnant mares treated daily with cloprostenol only were used as historic controls. Results--The mean (+/- SEM) interval from first cloprostenol administration to fetal expulsion 56.4 (+/- 13.7) hours and number of cloprostenol administrations 3.2 (+/- 0.6) in the 5 flunixin meglumine-treated mares were not significantly different, compared with values for 6 pregnant mares treated daily with cloprostenol only, 48.6 (+/- 5.6) hours and 2.8 (+/- 0.2) cloprostenol administrations. Flunixin meglumine did not inhibit endogenous PGF2 alpha secretion. Prostaglandin F2 alpha secretion rates on the day before and day of fetal expulsion were similar in both groups. Conclusion--Flunixin meglumine at a dosage of 500 mg/animal, administered IV every 8 hours, is ineffective in modulating uterine PGF2 alpha secretion during cloprostenol-induced abortion. Clinical Relevance--Flunixin meglumine is ineffective in the modulation of prostaglandin-induced uterine PGF2 alpha secretion and, therefore, does not offer a viable alternative for the prevention of abortion in mares at risk of abortion because of systemic illness.

Cytologic examination of bronchoalveolar lavage fluid (BALF), including phenotypic analysis of lymphocytes, was performed on 32 Standardbreds with poor race performance and endoscopic examination findings characteristic of inflammatory airway disease (IAD). Nucleated cell counts in BALF from IAD-affected horses were higher than those in control horses; the cytologic profile of BALF in affected horses included mixed inflammation, characterized by mild neutrophilia, lymphocytosis, and monocytosis. Eosinophil and mast cell counts were not higher in the IAD-affected group, compared with those in the control group; however, 4 IAD-affected horses had marked eosinophilia (24.7 +/- 4.8% SEM) in BALF. Phenotypic analysis of lymphocytes in BALF obtained from IAD-affected horses revealed a low proportion of CD4-positive cells and B cells, compared with those in the control group; these findings may have been representative of a greater proportion of non-B, non-T cells (null cells) in horses with IAD. The cytologic profile of BALF obtained from horses with IAD differed from that in horses affected with chronic obstructive pulmonary disease, suggesting that the pathogenesis of inflammation in horses with IAD may differ from that of chronic obstructive pulmonary disease.

Uptake of ferritin by M cells in follicle-associated epithelium at various sites in the small and large intestines was examined in 4 healthy 5-week-old pigs by use of electron microscopy. A 2.5% solution of ferritin in saline was injected into ligated loops of the jejunum and ileum containing aggregations of lymphoid follicles (Peyer's patches), as well as into intestinal loops containing lymphoglandular complexes at the ileocecal junction, in the central colonic flexure, and in the rectum. As negative control, saline solution was injected into loops at identical localizations. After an exposure period of 2 hours, uptake of ferritin by M cells, but not by enteroabsorptive cells of the small and large intestines, was observed. Numbers of M cells with ferritin and total M cells were counted and the percentage was calculated. Total number of M cells was highest in lymphoglandular complexes in the rectum and lowest on domes of the ileal Peyer's patch. High numbers of M cells with ferritin were found on domes of the jejunal Peyer's patch, and in lymphoglandular complexes at the ileoceral entrance and in the rectum. Only a few M cells on domes of the ileal Peyer's patch and in lymphoglandular complexes in the central colonic flexure contained ferritin. The percentage of M cells with internalized ferritin was similar on domes of the ileal Peyer's patch, and in lymphoglandular complexes at the ileocecal junction and in the rectum. It was higher on domes of the jejunal Peyer's patches and lower in lymphoglandular complexes of the central colonic flexure. Ferritin was found in the apical tubulovesicular system, multivesicular bodies, and a few vacuoles in the central area of M cells. Ferritin was exocytosed into the lateral intercellular spaces next to M cells. Uptake of ferritin by intraepithelial cells in the follicle-associated epithelium could not be documented, but ferritin was present in vesicles of subepithelial macrophages.