Hyperbaric oxygen effects on brown recluse spider (Loxosceles reclusa) envenomation in rabbits.

Human loxoscelism was modeled in albino rabbits by injection of brown recluse spider (Loxosceles reclusa) venom, and the effects of daily or twice-daily hyperbaric oxygen treatment on wound healing were investigated. Lesions similar to those seen in humans were produced in rabbits by intradermal injection of 200 microliters of a venom extract (0.21 microgram protein per microliter), including edema and erythema, ischemia and cyanosis in the first 12 hr, extensive purpura by 24 hr, and crateriform ulcer formation by day four, with induration and eschar formation. Hyperbaric oxygen treatments, consisting of two atmospheres absolute (2 ATA) for 60 min, were applied daily (n = 8) or twice daily (n = 8), while control animals (n = 8) received no treatment. Treatments were initiated 72 hr after venom injection (day 3) to duplicate typical clinical treatment delays, and were administered for seven consecutive days. No significant effects of hyperbaric oxygen treatment on lesion healing were seen as measured by lesion area. However, histologic evaluation of wound tissue collected at euthanasia on day 24 showed clear differences between rabbits receiving twice-daily treatments and those receiving daily or no treatment. The former showed complete re-epithelization or slight ulceration, while the latter usually had necrotic cavities extending into the dermis, with myonecrosis and inflammatory cell accumulation. Thus, no superficial differences were seen between groups, but twice-daily treatments resulted in enhanced recovery at the histologic level.

Comparison of the resistance of C57BL/6 and C3H/He mice to infection with Mycobacterium paratuberculosis.

Susceptibility of C57BL/6 (Bcgs) and C3H/HeN (Bcgr) mice to an intraperitoneal infection with Mycobacterium paratuberculosis strain 19698 was compared (by histopathology and the number of mycobacteria isolated from the spleen). Mycobacterial counts from the spleen of Bcgr mice progressively decreased over the course of infection but remained unchanged in Bcgs mice. Granulomatous lesions and acid-fast bacteria were consistently present in the liver and lymph nodes of Bcgs mice, whereas lesions were transient or absent in Bcgr mice. These results indicate that Bcgr mice are inherently resistant to M. paratuberculosis, whereas Bcgs mice are inherently susceptible. These differences may prove useful in elucidating the mechanisms of resistance and susceptibility to paratuberculosis and other mycobacterial infections.

Natural bovine lentiviral type 1 infection in Holstein dairy cattle. I. Clinical, serological, and pathological observations.

Clinical, serological, and pathological abnormalities observed in Holstein cows naturally infected with bovine lentivirus 1 bovine immunodeficiency virus (BIV) and other infections were progressive and most commonly associated with weight loss, lymphoid system deficiency, and behavioral changes. Clinical evidence of meningoencephalitis was dullness, stupor, and occasional head or nose pressing postures. The polymerase chain reactions associated the BIV provirus with the lesions in the central nervous system and lymphoid tissues. Multiple concurrent infections developed in retrovirally infected cows undergoing normal stresses associated with parturition and lactation. A major functional correlate of the lymphoreticular alterations was the development of multiple secondary infections which failed to resolve after appropriate antibacterial therapy. The chronic disease syndrome in dairy cows associated with BIV may be useful as a model system for investigation of the pathogenesis of the nervous system lesions and lymphoid organ changes that occur in humans with lentiviral infection.

Differences in the kinetics of T cell accumulations in C3H/HeN (Bcg-resistant) and C57BL/6 (Bcg-susceptible) mice infected with Mycobacterium paratuberculosis.

The accumulation of various T cell subsets in Bcg-susceptible (C57BL/6) and- resistant (C3H/HeN) strains of mice were compared following an intraperitoneal infection with Mycobacterium paratuberculosis. Groups of mice from both strains were killed at 3, 5, 10, 15, 30, and 150 days after infection and lymphocytes were harvested from the peritoneal exudate cells (PEC), spleen, intestinal epithelial lymphocytes (IEL), lamina propria lymphocytes (LPL), Peyer's patches, and mesenteric lymph node (MLN) and labelled with monoclonal antibodies to CD3, CD4, CD8, gamma delta TCR, CD25, and CD44 for flow cytometric analysis. Uninfected C3H/HeN mice had higher proportions of CD4+ cells in the spleen, MLN, LPL, IEL, and Peyer's patches, while uninfected C57BL/6 mice had higher proportions of CD8+ and/or gamma delta T cells. Significant increases in accumulation of CD8+ and gamma delta T cells were detected in the peritoneum and other tissues in both strains of mice after infection. Higher CD4/CD8 ratios were observed in most lymphoid tissues of C3H/HeN mice, while increased proportions of CD8+ and/or gamma delta T cells were present in C57BL/6 mice. These results indicate that significant differences in T cell profiles exist between these two strains of mice, both inherently and in response to infection with M. paratuberculosis. Innately lower levels of CD4+ cells and/or higher percentages of CD8+ and gamma delta T cells may play a role in the increased suspectibility of C57BL/6 mice to infection with M. paratuberculosis.

Natural bovine lentivirus type 1 infection in Holstein dairy cattle. II. Lymphoid tissue lesions.

Bovine immunodeficiency virus (BIV) in Holstein cows was associated with morphologic evidence of lymphoid organ deficiency. Cows were subjected to normal management practices including parturition and lactation without adverse environmental stresses. During the clinical disease process there was marked weight loss and wasting with frequent and severe concurrent infections. Lymphoid follicular hyperplasia and dysplasia in lymph nodes, and hypertrophy and hyperplasia in hemal lymph nodes were characteristics of the lymphoid tissues. Atrophy of lymphoid cell compartments with depletion of lymphocytes and a lymphocytic lymphoid folliculitis were components of the lymphoid system pathology. The nodal tissue lesions resembled those observed in feline, simian, and human lentiviral disease. A functional correlation with immune system deficiency was the development of multiple bacterial infections which failed to resolve after appropriate therapy. The BIV-associated disease syndrome in dairy cows may be useful as a model system for investigation of the pathogenesis of the lymphoid organ changes that occur in humans and animals with lentiviral infection.

Encephalitis, lymphoid tissue depletion and secondary diseases associated with bovine immunodeficiency virus in a dairy herd.

Encephalitis, lymphoid tissue depletion and secondary infections occurred over a 5-yr-period in Holstein cows infected with bovine immunodeficiency virus (BIV). There were 59 cattle studied, the majority during 1991, when a severe environmental stress occurred, each with one or more primary causes of death, natural or by euthanasia, and most with several secondary diseases. The encephalitis was characterized by meningeal, perivascular and parenchymal infiltration with lymphocytes, occasional plasma cells and macrophages with perivascular edema in some cows. Affected areas included the cerebrum, cerebellum, and spinal cord with no particular distribution pattern recognized. The lymphoid depletion was primarily an absence of follicular development in nodes draining regions with secondary infections such as chronic mastitis and chronic suppurative pododermatitis. Paucity of lymphocytes in thymic-dependent regions of lymph nodes and the spleen suggested a primary depletion of T cells. Secondary infections were often multiple with each cow having several minor conditions, usually considered short-term and treatable. These included mastitis and pododermatitis, with many cows having non-responding abscesses, cellulitis and myositis attributed to injection site infections. A large number of the cattle had parturition difficulties such as dystocia, obturator paralysis, and metritis. Pulmonary, cardiovascular, and intestinal disease were recognized as both primary and secondary disease conditions. There was a high level of infection with bovine leukemia virus with 4 of the 59 cattle having lymphosarcoma. Under practical conditions, the infection with BIV has a different effect on the host than has been observed under experimental conditions. The presence of BIV combined with the stresses associated with parturition and a modern dairy production system were considered causal for the development of untreatable secondary diseases in immunocompromised cattle. The peak incidence in 1991 was attributed to increased environmental stress during renovation of the barn facility. During this time the cattle were kept on open pasture, exposed to an extremely wet winter, and spring weather conditions. The effect of co-infection with bovine leukemia virus, the influence of immunocompromise on the chronicity of mastitis, the relationship with laminitis and pododermatitis, and several questions related to viral transmission, complementarism with bovine leukemia virus, viral reactivation and immunoprophylaxis all remain as viable avenues for future investigations.

Spontaneous murine thymocyte comitogenic activity consistent with interleukin-1 in cattle naturally infected with Mycobacterium paratuberculosis.

The production of comitogenic activity consistent with interleukin-1 (IL-1) activity by blood monocytes from cattle with naturally acquired paratuberculosis was examined by murine thymocyte proliferation. In addition, IL-1-like activity in response to homologous and heterologous antigens was determined. Activity was determined in nine cattle naturally infected with Mycobacterium paratuberculosis and six non-infected cattle. Comitogenic properties were measured in response to M. paratuberculosis antigen (johnin), bacterial lipopolysaccharide (LPS) as a positive control, and culture media as a negative control. Monocytes from infected cattle spontaneously released high levels of IL-1-like activity in the absence of stimuli and significantly (P less than 0.05) increased activity in response to LPS. With johnin, M. bovis PPD and KLH stimulation, comitogenic activity was similar to spontaneous levels. Non-infected cattle had significantly (P less than 0.05) increased comitogenic activity when blood monocytes were stimulated with KLH, M. bovis PPD, johnin, and lipopolysaccharide when compared with non-stimulated cells in that group. Johnin produced the greatest response in non-infected animals. The data suggest that blood monocytes in infected cattle are non-specifically activated with respect to IL-1 production. Alternatively, a defective regulatory mechanism for IL-1 may be operative in infected cattle. In addition, the previous observation that mycobacterial antigens are potent inducers of IL-1 was also verified.

Effect of neonatal thymectomy and antilymphocyte globulin on non-specific mitogen stimulation in Holstein-Friesian calves.

Nine of seventeen neonatal Holstein-Friesian calves were thymectomized, treated with antilymphocyte globulin, and monitored for immunologic functional ability for 4 to 6 months. The thymus weights for 4 to 10-day-old calves and 4 to 6-month-old calves indicated a continued increase in total weight. This indicated significant thymic involution had not occurred at 4 to 6 months. Following thymectomy a wasting syndrome was not observed although an increased incidence of a lowly virulent virus infection did occur. A significant decrease in circulating lymphocytes was observed. Peripheral blood lymphocytes were stimulated in vitro by non-specific mitogens, phytohemagglutinin, bacterial lipopolysaccharide and pokeweed mitogen using the whole blood culture method. Observations included a greater response to phytohemagglutinin and pokeweed mitogen in summer months and variable age related response to all mitogens. Lipopolysaccharide stimulation results were inconclusive. It was concluded that neonatal thymectomy was not a satisfactory experimental procedure for the production of selective immunosuppression in the bovine species.

Immunosuppression of lymphocyte blastogenesis in cattle infected with Ostertagia ostertagi and/or Trichostrongylus axei.

Twenty 4-month-old calves were infected with O ostertagi and/or T axei and the responses to phytolectins were evaluated. Whole blood cultures were incubated with phytohemagglutinin (PHA), concanavalin A (Con A) and pokeweed mitogen (PWM). The blastogenic response was determined by tritiated thymidine uptake with results presented as counts per minute (cpm), stimulation indices (SI) and a mononuclear cell responsive index determined by dividing the phytomitogen induced cpm by the absolute mononuclear cell number per ul. The control group results were adjusted to 100 percent and changes in the percentage difference by the parasitized calves was determined. There was a decline in lymphocyte responsiveness to PHA beginning at the time of infection. Significant depression of responses to PHA was observed in all parasitized calves 8 weeks after infection although clinical signs of parasitism did not occur. Lymphocyte responses to PW, were not different in infected calves from the control, although the O ostertagi group had significantly higher PWM mean upon than the calves infected with T. axei. A slight depression in response to Con A was also observed at 8 weeks after infection followed by a significant increase after 10 weeks. The immunosuppression appeared to be a feature of gastrointestinal parasitism and related to infections with O. ostertagi and/or T. axei.

Visualization of eosinophil chemotactic factor in abomasal tissue of cattle by immunoperoxidase staining during Ostertagia ostertagi infection.

Eosinophil chemotactic factor (ECF) was localized predominantly in the intestinal cells and lateral hypodermal cords of developing fifth stage larvae (L5) of Ostertagia ostertagi within abomasal tissue cross-sections by peroxidase in an antibody sandwich technique using monoclonal antibody to ECF. Cooperia oncophora larvae in tissue cross-sections did not stain using this technique. These experiments demonstrate that ECF is localized in Ostertagia ostertagi organelles and is probably released by the developing L5 into the abomasal tissue surrounding the parasitized gland. The presence of ECF within O. ostertagi larvae in situ and the results of previous experiments demonstrated in vitro and in vivo ECF chemotactic activity help to explain why eosinophils are observed histologically in abomasal tissues from cattle with ostertagiasis.

Sequential histopathologic changes of type I, pre-type II and type II ostertagiasis in cattle.

Yearling cattle in Louisiana were examined at monthly intervals for abomasal nematode burdens and histological lesions over a year. Tracer calves were grazed for 3 to 4 weeks and removed from pasture for 2 to 3 weeks, then slaughtered; a few animals were killed in extremis shortly after removal from pasture. Histological changes were correlated with worm burdens and characterized according to the type of Ostertagia ostertagi infection present. In cattle with acute Type I ostertagiasis, changes varied from eosinophil infiltration to glandular dilation and slight mucous cell hyperplasia with submucosal edema. During the summer months the cattle had worm burdens that were primarily early 4th stage larvae (EL4), with changes characterized by minimal glandular dilation and mucous cell metaplasia and moderate lymphoid cell proliferation and with intramucosal migration of EL4. In the autumn, the maturation of EL4 produced the Type II syndrome with severe glandular changes, prominent mucosal hyperplasia and marked lymphoid cell accumulation. With increased duration of the pre-Type II interval, there was greater development of the subepithelial lymphoid tissue and increased frequency of epithelial globule leukocytes. The lymphoproliferation which occurred during the prolonged pre-Type II interval appeared to be related to the increased severity and mortality seen with the Type II ostertagiasis syndrome.

Encephalitis attributed to dactylariosis in Japanese quail chicks (Coturnix coturnix japonica).

Dactylaria gallopava was isolated from brain tissue of 1-to-3-week-old quail chicks. Successive batches demonstrated elevated (15-20%) mortality preceded by incoordination and lateral recumbency. Chicks exhibited cerebellar and cerebral encephalitis characterized by brown-red discoloration of affected brain tissue. Decontamination of setters and hatchers resulted in abrupt cessation of mortality in subsequent placements, implicating incubators as the source of infection.

Mucinous hyperplasia in the kidney and ureter of a horse.

A case of mucinous hyperplasia in the right renal pelvis and ureter is reported in a 23-year-old, male, Quarter horse. The affected right renal pelvis was markedly distended and filled with inspissated yellowish-green mucous material compressing the adjacent cortical and medullary parenchyma. Multiple, similar masses were present protruding from the right urethral mucosa. Histologically, the masses were characterized by a cystic glandular and papillary hyperplasia of the mucosa of the renal pelvis and ureter. This is believed to be the first reported case in an equine species.

Histopathology of C57BL/6 mice inoculated orally with Mycobacterium paratuberculosis.

The susceptibility of C57BL/6 mice to oral inoculation with Myobacterium paratuberculosis was evaluated histopathologically. Granulomatous lesions containing acid-fast bacteria developed in the mesenteric lymph nodes in over 50% of the mice by 11 months after inoculation. The results suggest that C57BL/6 mice may be useful for studying infection, pathogenesis, and other aspects of paratuberculosis.

Effect of strategic anthelmintic treatment and pasture management on productivity and control of nematode parasites in weaner-yearling beef cattle.

Three groups of 17 beef calves were used to evaluate effects of strategic anthelmintic treatment on safe (group 1) and contaminated (group 2) pasture in comparison with minimal treatment at weaning and contaminated pasture (group 3). The investigation extended from weaning in November 1982 to the following August. Results of faecal egg counts, herbage larval counts, plasma pepsinogen and tracer calf worm counts in autumn and spring indicated minimal levels of infection on safe pastures provided in November and April (group 1). A decided weight gain advantage for group 1 was achieved from November to April, but the rate of gain was not consistent after April and transfer to the second safe pasture. Final average weights in late August were: group 1, 368 kg; group 2, 336 kg; group 3, 262 kg. All were significantly different (P less than 0.05). Worm counts from representative yearlings in September revealed low to moderate levels of Ostertagia ostertagi in group 1. In contrast group 2 cattle had large, almost exclusively O ostertagi infections; group 3 cattle had exceedingly high levels of Trichostrongylus axei infection and moderate to high levels of O ostertagi. Marginal evidence of type 2 ostertagiasis was observed in individual animals of group 2 and group 3.

Measurement of lymphoblast proliferative capacity of stimulated blood mononuclear cells from cattle with chronic paratuberculosis.

Concanavalin A (conA) blast proliferation as a quantitative measure of lymphoblast proliferative capacity by blood mononuclear cell supernatants was measured in cattle naturally infected with Mycobacterium paratuberculosis and in healthy control cattle. Blast cell proliferation was significantly reduced in infected animals, compared with control cattle when blood mononuclear cells were stimulated with conA. Proliferation was significantly greater than media control when M bovis purified protein derivative and johnin were used to stimulate cells from the infected group. After sensitizing control and affected cattle with M paratuberculosis bacterin (live M bovis and keyhole limpet hemocyanin in Freund's incomplete adjuvant), infected animals had no difference in blast cell proliferative capacity with the mycobacterial antigens and conA stimulation, whereas healthy animals had significantly increased blast proliferation in response to all the sensitizing antigens. The blast cell proliferative capacity in infected animals with keyhole limpet hemocyanin stimulation was increased significantly after sensitization; however, it remained significantly less than that in the sensitized control group. These data indicate that cattle naturally infected with M paratuberculosis probably produce suboptimal interleukin-2 (IL-2) activity in response to a potent IL-2 inducer (conA) and fail to optimize IL-2 activity when sensitized with a potent immunogen (keyhole limpet hemocyanin).

Gross and light microscopic lesions in neonatal gnotobiotic dogs inoculated with a canine rotavirus.

Gross, light microscopic, and morphometric changes were observed in the small intestines of 11 neonatal gnotobiotic dogs after oral inoculation with a canine rotavirus. Starting in pups killed at postinoculation hour (PIH) 24, gross changes consisted of moderate dilatation and thinning of the walls of the small intestine, hyperemia, and a moderate amount of semiliquid-to-liquid greenish-yellow intestinal contents. In the jejunum and ileum of inoculated pups killed from PIH 18 to 48, columnar villus epithelial cells on the upper one-third of the villus were necrotic, and foci denude of epithelium were seen on the upper regions of villi. Inoculated pups killed from PIH 24 to 72 had mild-to-moderate villus atrophy in the jejunum and ileum, villi covered with cuboidal-to-flat squamous-like epithelial cells, and absence of large, clear vacuoles in the jejunal and ileal villus epithelial cells similar to those seen in villus epithelial cells of 9 control pups. Microscopic sections of the jejunum and ileum from inoculated pups killed from PIH 72 to 154 showed slight-to-moderate villus atrophy, and the villi were covered with cuboidal-to-low columnar epithelial cells. Morphometric results indicated lower mean villus-crypt ratios in the duodenum, jejunum, and ileum of inoculated pups. In addition, the morphometry of the small intestine indicated that crypt cell hyperplasia occurred in the duodenum early during infection without obvious villus atrophy, and hyperplasia of the crypt cells in the jejunum and ileum was observed later during the infection.

Dermal responses to Ostertagia ostertagi in Ostertagia ostertagi- and Cooperia punctata-inoculated calves.

Calves harboring patent Ostertagia ostertagi or Cooperia punctata were given intradermal injections of O ostertagi 3rd-stage larval antigen. The initial injections were followed 30 days later by a 2nd series of injections. Skin thickness was measured at injection sites for 72 hours after injection. Selected injection sites including saline solution control sites were biopsied at 30 minutes, at 3, 24, 48, and 72 hours, and at 30 days after injection. After the 1st series of injections, there was a clear distinction in dermal reactions between O ostertagi-inoculated calves and C punctata-inoculated calves; after 24 hours, reactions were not seen in the C punctata-inoculated calves. Marked dermal reactions occurred in the O ostertagi-inoculated calves. The reactions at 30 minutes and 3 hours were characterized by slight-to-extensive infiltration of neutrophils and dermal edema. The 24-hour cellular reaction was principally due to neutrophil and eosinophil infiltration with edema and necrosis. Reactions at 48 to 72 hours were due to eosinophils and perivascular accumulations of macrophages and lymphocytes. Necrosis, neutrophils, and edema were present in foci where fragments of nematodes were located. On reinjection, a clear distinction in dermal reactions between calves was not seen based on the type of nematode infection. Thirty days after dermal inoculation, large nodules developed at the site of the initial antigen injection. The nodules were characterized by marked intradermal proliferation of lymphocytes in a follicular pattern with occasional macrophages and rare multinucleated giant cells.

Effects of dialyzable lymph node extracts on lymphoblast proliferative capacity of blood mononuclear cells in cattle with chronic paratuberculosis.

Dialyzable lymph node extracts (DLE) containing transfer factor prepared from calves sensitized to Mycobacterium paratuberculosis and keyhole-limpet hemocyanin (KLH) were administered to 4 adult cows with chronic paratuberculosis. Cutaneous delayed hypersensitivity, lymphocyte blastogenesis, monocyte migration-inhibition, and lymphoblast proliferative capacity as a reflection of interleukin-2 (IL-2) activity were measured in response to M bovis purified protein derivative, johnin, and KLH before and after treatment with DLE. Change in cutaneous delayed hypersensitivity was not evident after DLE treatment. Alterations in histologic features of pre- and posttreatment sections of ileum and mesenteric lymph nodes were not detected. Lymph node extract treatment significantly (P less than 0.05) increased IL-2 activity and migration-inhibition in response to johnin and KLH in vitro. Treatment had no effect on lymphocyte blastogenesis. The data indicate that cattle with chronic paratuberculosis may benefit from DLE treatment, by virtue of increased IL-2 activity, and that effects of DLE are at least partially mediated by an increase in IL-2 activity.

Celiac trunk cannulation for obtaining abomasal lymph from cattle.

Cannulation of the celiac trunk was surgically performed in 26 Holstein steers. The procedure was successful in 23 (88.5%) of the steers. Twenty-two of the steers were infected either naturally or experimentally with the abomasal nematode, Ostertagia ostertagi and/or other gastrointestinal parasites. The remaining 4 steers were not infected. Lymph obtained after surgery was used in various immunologic and biochemical assays. Daily lymph flow rate and total and differential WBC counts were determined after surgery in 4 of the infected and 3 of the noninfected steers. Steers were euthanatized for tissue specimen collection 7 days after surgery. At the time of euthanasia, lymph was still flowing from the cannula of 13 (56.5%) of the steers in which surgery was successful. This surgical procedure represents a valuable technique for studying at the local level, immunologic and physiologic responses of cattle to infection with O ostertagi.