Fatal systemic phaeohyphomycosis caused by Ochroconis gallopavum in a dog (Canis familaris).

A 5-year-old Shetland Sheepdog was presented with a history of weakness, ataxia, anemia, thrombocytopenia, and occasional seizures. The dog had been treated for 6 months with prednisone for inflammatory bowel disease. A positive titer for Ehrlichia canis was detected 6 months before referral. The initial physical examination revealed a weak, laterally recumbent dog with pale mucous membranes. Neurologic examination revealed multiple neurologic deficits. A complete blood cell count (CBC) revealed normochromic, normocytic, nonregenerative anemia; lymphopenia; thrombocytopenia; and neutrophilic and monocytic leukocytosis. Urinalysis revealed proteinuria, with a specific gravity of 1.045. The dog was unresponsive to treatment and died. At necropsy, there was severe serofibrinous peritonitis and pleuritis, with randomly scattered dark brown necrotic foci present in multiple organs, including liver, spleen, kidney, and pancreatic lymph node. Histologically, there were extensive regions of parenchymal necrosis surrounded by neutrophils admixed with epithelioid macrophages, lymphocytes, and pigmented fungal organisms. Numerous brown, 2 to 6 microm in diameter, septate, branching hyphae, subsequently identified as Ochroconis gallopavum (formerly Dactylaria constricta var. gallopava), were observed.

Sequence variation of the ribosomal DNA second internal transcribed spacer region in two spatially-distinct populations of Amblyomma americanum (L.) (Acari: Ixodidae).

Sequence analysis of the ribosomal DNA second internal transcribed spacer (ITS 2) region in 2 spatially distinct populations of Amblyomma americanum (L.) revealed intraspecific variation. Nucleotide sequences from multiple DNA extractions and several polymerase chain reaction amplifications of eggs from mixed-parentage samples from both populations of ticks revealed that 12 of 1,145 (1.0%) sites varied. Three of the 12 sites of variation were distinct between the 2 A. americanum populations, which corresponded to a rate of 0.26%. Phylogenetic analysis based on ITS 2 sequences provided strong support (i.e., bootstrap value of 80%) that wild A. americanum clustered into a distinguishable group separate from those derived from colony ticks.

Purification and partial characterization of the OmpA family of proteins of Pasteurella haemolytica.

This study was conducted to partially characterize and identify the purity of two major outer membrane proteins (OMPs) (with molecular weights of 32,000 and 35,000 [32K and 35K, respectively]) of Pasteurella haemolytica. The 35K and 32K major OMPs, designated Pasteurella outer membrane proteins A and B (PomA and PomB, respectively), were extracted from P. haemolytica by solubilization in N-octyl polyoxyl ethylene. The P. haemolytica strain used was a mutant serotype A1 from which the genes expressing the 30-kDa lipoproteins had been deleted. PomA and PomB were separated and partially purified by anion-exchange chromatography. PomA but not PomB was heat modifiable. The N-terminal amino acid sequences of the two proteins were determined and compared with reported sequences of other known proteins. PomA had significant N-terminal sequence homology with the OmpA protein of Escherichia coli and related proteins from other gram-negative bacteria. Moreover, polyclonal antiserum raised against the E. coli OmpA protein reacted with this protein. PomA was surface exposed, was conserved among P. haemolytica biotype A serotypes, and had porin activity in planar bilayers. No homology between the N-terminal amino acid sequence of PomB and those of other known bacterial proteins was found. Cattle vaccinated with live P. haemolytica developed a significant increase in serum antibodies to partially purified PomA, as shown by enzyme-linked immunosorbent assays, and to purified PomA and PomB, as detected on Western blots and by densitometry.

Identification and comparison of macrophage-induced proteins and proteins induced under various stress conditions in Brucella abortus.

Brucella abortus is a facultative intracellular pathogen of cattle and humans that is capable of survival inside macrophages. In order to understand how B. abortus copes with the conditions during intracellular growth in macrophages, the protein synthesis pattern of the bacteria grown inside bovine macrophages has been compared with that of bacteria grown in the cell culture medium by two-dimensional polyacrylamide gel electrophoresis. Approximately 24 new proteins that are not detected in the bacteria grown in the cell culture medium have been induced during intracellular growth in macrophages. In contrast, approximately 50 proteins that were expressed during growth in cell culture medium were completely repressed during intracellular growth. The level of expression of 19 proteins increases while that of 54 proteins decreases during intracellular growth. To understand these results, the protein synthesis pattern of B. abortus during intracellular growth was compared with those during other stress conditions. Under each stress condition studied, several new proteins were induced that were not present during regular growth conditions. Comparison of the protein synthesis pattern of B. abortus during intracellular growth with those obtained under various stress conditions has indicated that the response to intracellular growth was not just a simple sum of stress conditions studied so far.

Has Medicaid growth crowded out state educational spending?

Researchers have argued that the dramatic increase in Medicaid spending during the late 1980s and early 1990s "crowded out" state spending on other activities, particularly education. Medicaid growth has, at least in part, been driven by increased federal eligibility and service mandates, federal court decisions on hospital and nursing home rates, and health care inflation; and the need to respond to these outside forces has placed increasing pressure on state finances. Other evidence, however, suggests that the adverse effect of Medicaid growth on state finances has been overstated. During the late 1980s and early 1990s, states shifted many human service programs from general fund to Medicaid financing and took advantage of Medicaid rules governing the use of provider donations and assessments, such as state matching and claiming payments to disproportionate-share hospitals to increase federal reimbursement without increasing the claims on their own revenues. But the increased burden of Medicaid growth on state finances may be more apparent than real. In this article, we test the crowding-out hypothesis using a two-stage, least-squares fixed-effects model of Medicaid's impact on educational spending from 1980 to 1990. Our results indicate that Medicaid growth has had no significant effect on educational spending. Rather, educational spending has responded more to changes in states' own-source revenues than to growth in Medicaid spending.

The role of eicosanoids in the chemotactic response to Pasteurella haemolytica infection.

The chemotactic role of eicosanoids in the pathogenesis of Pasteurella haemolytica infection was studied, using a tissue chamber infection model and pharmacological inhibitors of eicosanoid synthesis. Tissue chambers were implanted subcutaneously in 12 calves allotted to three treatment groups of equal size. At 45 days after implantation, calves received saline, dexamethasone, or phenylbutazone treatments, and tissue chambers in all animals were then inoculated with P. haemolytica. Chamber fluid samples were collected before inoculation and at 2, 6, 18, 40, and 90 h after inoculation. Bacterial counts, total leukocyte counts, pH and albumin concentrations in chamber fluids were determined using standard bacteriological and clinical pathological methods. Concentrations of eicosanoids and activity of interleukin-1 (IL-1) were measured by radioimmunoassay and a helper T cell bioassay, respectively. Concentrations of leukotriene B4 (LTB4), thromboxane B2 (TXB2), 6-keto-prostaglandin F1 alpha (PGF1 alpha) and prostaglandin E2 (PGE2) increased markedly after inoculation. An inhibitory effect of dexamethasone on both LTB4 production and neutrophil influx, together with the temporal relationship between these two events, suggested that LTB4 served as a chemo-attractant. Activity-time profiles for IL-1 in chamber fluids were similar to those of the eicosanoids. Phenylbutazone and dexamethasone reduced the severity of the inflammatory responses as measured by lower concentrations of albumin and higher pH in treated versus control chamber fluids. The results of this study suggest that eicosanoid inflammatory mediators play an important chemotactic role in the pathogenesis of P. haemolytica infection.

Biochemical and morphological alterations in xylazine-induced pulmonary edema.

Sprague-Dawley rats were given 42 mg/kg xylazine intramuscularly, and lungs were lavaged with phosphate-buffered saline 3, 6, and 12 hr later. Total protein, lactate dehydrogenase (LDH), xanthine oxidase (XO), tumor necrosis factor (TNF), and interleukin 1 (IL-1) were measured in bronchoalveolar lavage fluid (BALF). Protein concentration, LDH, XO, and TNF levels were increased (p < 0.05) in the BALF from xylazine-treated rats as compared to controls. IL-1 level was unchanged at 3 and 6 hr and was reduced (p < 0.05) at 12 hr. Another group of rats was given 42 mg/kg xylazine intramuscularly, and lungs were fixed 0.5 and 12 hr later. Histologically, severe pulmonary edema (PE) involving the alveoli and perivascular stroma was observed. Fibrin, increased numbers of eosinophils, and macrophages with foamy cytoplasm were present in the alveoli of all treated animals. Ultrastructurally, endothelial damage, characterized by thinning, detachment from basement membranes, or bleb formation, was observed. The lesions were similar in both xylazine groups, differing mainly in severity with the 12-hr group having more severe lesions than the 0.5-hr group. To determine whether endothelial injury is caused by direct toxicity of xylazine, bovine pulmonary artery endothelial cells (BPAECs) were incubated with xylazine (0.3, 3, and 30 micrograms) for 0.5 or 3 hr. Xylazine did not have any effects on BPAECs, as indicated by phase-contrast microscopy and dye-exclusion viability assay. These results indicate that xylazine-induced PE is due to increased permeability resulting from endothelial injury, which is not caused by direct effect of xylazine on pulmonary endothelium. While oxygen radicals and TNF are possibly involved, IL-1 does not appear to play a role in xylazine-induced PE.

Comparison of Brucella abortus antigen preparations for in vitro stimulation of immune bovine T-lymphocyte cell lines.

Three Brucella abortus antigen preparations were tested for stimulatory activity with immune bovine T-lymphocyte cell lines in vitro. A total of 32 polyclonal T-lymphocyte cell lines were derived from two steers each from four immunization groups: (1) B. abortus Strain 19 (S19) alone, (2) heat-killed B. abortus whole bacterial cells (HKC) alone, (3) S19 with recombinant human interleukin 2 (rHuIL-2), (4) HKC with rHuIL-2. Peripheral blood mononuclear cells were isolated at 2 and 9 weeks post immunization and cultured in vitro with either HKC antigen or B. abortus soluble antigen (BASA) with recombinant bovine interleukin 2 (rBoIL-2) to initiate four cell lines per steer. Sixteen of the resulting T-lymphocyte cell lines (from the S19 and S19+IL-2 groups) were tested through indirect immunofluorescence for expression of cell surface markers CD2, CD4, CD6, CD8, major histocompatibility complex (MHC) Class II molecules and a marker expressed on a subset of helper T-lymphocytes (Th) as well as sIgM, CD1 and a MHC Class II+ monocyte/macrophage marker. The T-lymphocyte cell lines were used to evaluate antigen-induced lymphoproliferative (LP) responses in a titration assay with HKC, BASA and gamma-irradiated B. abortus (gamma BA) antigens. The results indicate that most of the cells in many of the cell lines were typical activated T-lymphocytes as determined by surface marker expression and included cells positive for all T-lymphocyte markers tested. The cell lines contained no B-lymphocytes or mononuclear phagocytes. However, two cell lines contained significant populations (> 80%) of CD2-, CD4-, CD6-, CD8- cells that were both responsive to exogenous rBoIL-2 and were capable of exhibiting antigen-induced LP responses. In 22 of the 32 cell lines tested, gamma BA was superior to HKC at nearly every concentration tested in stimulating LP responses. This observation was independent of the immunization used to prime the T-lymphocytes in vivo. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed proteins with relative molecular masses common to all three antigen preparations as well as significant (P < 0.05) quantitative and qualitative differences in individual proteins between HKC and gamma BA. Together, the results suggest gamma BA may provide an in vitro antigenic stimulus which is deficient in HKC.

Serological survey for Lyme disease in domestic dogs and white-tailed deer from Oklahoma.

Sera from 223 randomly selected dogs and 489 white-tailed deer (Odocoileus virginianus) were tested for antibodies to Borrelia burgdorferi using an indirect kinetic ELISA. Dog samples were obtained in 1989 whereas deer samples were obtained between 1975 and 1990. Ten known negatives and two known positives from each group were run on each plate as controls. Samples showing mean mOD values above the mean of negatives + 3 SD were considered positive. Twenty-six dog (11.7%) and 22 deer (4.5%) samples were positive. Deer reactors were first detected among 1978 samples. Reactive deer were from central and eastern Oklahoma whereas reactive dogs were mostly from central Oklahoma. Confirmed human cases between 1986 and 1989 were distributed throughout the state, thus showing no correlation with either deer or dog results.

In vitro and in vivo 2',5'-oligoadenylate synthetase activity induced by recombinant DNA-derived bovine interferon alpha I1 in bovine alveolar macrophages and blood mononuclear cells.

Biological responses to recombinant DNA-derived bovine interferon alpha (rBoIFN-alpha I1) by bovine alveolar macrophages were examined by measuring viral yield reduction and 2',5'-oligoadenylate synthetase (2',5'-OAS) production by IFN-treated cells. In vitro IFN pretreatment of alveolar macrophages reduced viral yield in cultures challenged exposed with parainfluenza-3 virus, compared with control cultures. In vitro treatment of alveolar macrophages with IFN also resulted in increased 2',5'-OAS activity. The 2',5'-OAS activity was measured in alveolar macrophages and blood mononuclear leukocytes of calves injected IM with 3.6 x 10(6) U of rBoIFN-alpha I1/kg of body weight. The IFN action was monitored by measuring 2',5'-OAS activity of blood mononuclear leukocytes beginning 6 days before and ending 24 hours after IFN treatment. The 2',5'-OAS activity in the blood mononuclear leukocytes sharply increased 24 hours after IFN treatment, indicating response to IFN. The alveolar macrophages collected from the same calves 24 hours after IFN administration also had increased 2',5'-OAS activity, compared with alveolar macrophages from the same calves collected 6 days before treatment. Increased 2',5'-OAS activity indicates: a possible mechanism of IFN action in cattle that may be responsible for viral yield reduction; potential use of high enzyme activity as a marker for IFN induction; and potential use of 2',5'-OAS activity as a marker for determining effects of IFN on bovine macrophages and other cells of the bovine immune system.

The effects of adjuvants on immune responses in cattle injected with a Brucella abortus soluble antigen.

Five different adjuvants were examined for potentiation of humoral and cell-mediated immune (CMI) responses in cattle to a Brucella abortus soluble antigen (BASA). Two separate experiments were performed involving a total of 64 steers, divided among six groups (Experiment 1) and 9 groups (Experiment 2). The adjuvants used were: muramyl dipeptide, Freund's incomplete adjuvant, dimethyl-dioctadecyl ammonium bromide (DDA), Bordetella pertussis and Propionibacterium acnes. In each experiment, three groups received BASA (2 mg protein) subcutaneously with adjuvant, one group received a reduced dose of B. abortus Strain 19 (S19), one group served as unvaccinated controls, and another group received BASA alone. Primary responses were studied following a single immunization in comparison to the single inoculation with S19. For each experiment serum antibody responses and CMI responses were sequentially determined over a period of 56 days. Antibody responses to B. abortus were measured using the brucellosis card, rivanol precipitation-plate agglutination, complement fixation, and fluorometric immunoassay tests, and as well as with an enzyme-linked immunosorbent assay. The CMI response was measured using antigen-specific lymphoproliferation (LP) and skin testing for delayed-type hypersensitivity (DTH) to BASA (Experiment 2). Specific aspects of induced CMI responses investigated were macrophage activation (IL-1 production), helper T cell activation (IL-2 production), and release of soluble suppressor factor(s). In general, mean antibody responses were significantly higher (P less than 0.05) in immunized steers than in control steers and those receiving BASA alone. The LP responses to heat-killed B. abortus were generally higher in immunized groups than in the controls. The LP and DTH responses were greatest in the groups receiving S19 and BASA + DDA. Increased induction of IL-1 was largest in the group receiving BASA + DDA whereas IL-2 release was greatest in S19 vaccinated steers. Suppressor T cell responses were most obvious in the groups receiving S19, BASA + B. pertussis, and P. acnes. These studies demonstrated that DDA potentiates CMI responses to a soluble B. abortus antigen and may be useful as an adjuvant for future vaccines, particularly subunit vaccines.

Immunopotentiation of cattle vaccinated with a soluble Brucella abortus antigen with low LPS content: an analysis of cellular and humoral immune responses.

The adjuvant effects of dimethyl dioctadecyl ammonium bromide (DDA) alone or in combination with trehalose dimycolate (TDM) or muramyl dipeptide (MDP) on bovine humoral and cellular responses to a soluble protein extract of gamma irradiated Brucella abortus strain 19 (SPEBA) were investigated. Thirty-five beef steers were randomly allotted to nine groups. Three of these groups received SPEBA (2 mg protein per dose) subcutaneously in combination with adjuvants, one group received the reduced dose of B. abortus strain 19 (S19), and one group received SPEBA alone. Controls included groups receiving adjuvant preparations only or no vaccine. Immune responses to the various immunizations were assessed sequentially for 56 days using various in vitro and in vivo assays. The humoral response to B. abortus was measured using standard serologic tests, an enzyme-linked immunosorbent assay, and a quantitative fluorometric immunoassay. The cell-mediated immune (CMI) response was measured by antigen-specific lymphoproliferation (LP), interleukin 2 (IL 2) production, and soluble suppressor factor release. Skin testing at day 35 for delayed-type hypersensitivity (DTH) to SPEBA was also performed. Minimal humoral responses were induced with SPEBA alone. The highest and most sustained serum antibody responses to B. abortus antigens were elicited by the S19 vaccine. A combination of SPEBA with DDA + TDM induced higher antibody levels than SPEBA with DDA or SPEBA with DDA + MDP. Responses to DTH among the groups receiving SPEBA were most notable in the SPEBA with DDA + TDM groups. Increased IL 2 production was greatest in the S19 and SPEBA with DDA + TDM vaccinates. The results indicated that a combination of DDA + TDM best potentiated immune responses to a soluble B. abortus antigen preparation and may be useful as adjuvants for future vaccines.

Immunomodulation in cattle immunized with Brucella abortus strain 19.

Regulation of the bovine immune response to immunization with Brucella abortus Strain 19 (S19) was investigated through application of a modification of an assay to measure suppressor T lymphocyte activities in humans and through development and characterization of antigen-stimulated T lymphocyte lines in vitro. A total of nine of steers were alloted into two groups: control (n = 4) and S19-immunized (n = 5). Peripheral blood mononuclear cells (PBMC) from each animal were cultured in vitro with mitogens (concanavalin A (Con A) and pokeweed mitogen (PWM], B. abortus antigens (B. abortus soluble antigen (BASA) and whole heat-killed B. abortus cells (HKC)) and media alone periodically from days 4 through 49 of the experiment. Supernates from these cultures were assayed for immunomodulatory activity(s) by addition to indicator cultures stimulated with suboptimal concentrations of Con A. Supernates from PBMC of S19-immunized steers generated with B. abortus antigens significantly (P less than 0.05) suppressed indicator cell responses as compared to those from control steers on days 35 and 49 post-immunization. This suppressive activity from PBMC of immunized cattle with respect to that of control cattle could also be induced through mitogenic stimulation with Con A or PWM. On day 49 of the study, suppressive activity was spontaneously released from the PBMC of immunized cattle. T lymphocyte lines were initiated from two S19-immunized steers at 2 and 9 weeks post-immunization. These T cell lines were characterized with respect to proliferative responses to B. abortus antigens through in vitro assay and surface marker expression through indirect immunofluorescence with a limited panel of monoclonal antibodies. Results from the present study indicated that S19 immunization induces a subpopulation(s) of cells in the PBMC of cattle capable of regulating the in vitro response to B. abortus. This regulatory activity is detectable by in vitro assay as early as 7 weeks post-immunization. Furthermore, the regulatory cell(s) appear to involve BoCD8+ T, lymphocytes which are specific for B. abortus antigens.

Sepracell-MN for mononuclear cell separation from cattle: effect on mitogen-and antigen-induced lymphocyte blastogenesis.

Mononuclear leukocytes (MNC) were separated from heparinized and EDTA-treated whole bovine blood by centrifugation after mixing with a commercial colloidal silica preparation (Sepracell-MN (S-MN]. Cell yields and lymphocyte blast transformation (LBT) to pokeweed mitogen (PWM), phytohaemagglutinin (PHA), concanavalin A (Con A), and Brucella abortus antigens were tested against MNC obtained from heparinized whole blood using Ficoll-Hypaque (FH). Separation with S-MN was more rapid and less labor intensive than separation with FH. There was a higher average total yield of MNC but a lower percentage of monocytes in the FH- than in the S-MN-separated MNC. In mitogen-induced LBT assays, MNC responded comparably to each mitogen regardless of the separation technique or anticoagulant used, and a cell concentration effect was demonstrated. In general, FH-separated MNC responded greater to PWM than did S-MN/EDTA separated MNC, but S-MN/heparin separated MNC had the greatest LBT responses to PWM. Overall, S-MN/EDTA separated MNC had the greatest responses to PHA, and responses to Con A were variable among experiments with respect to the separation technique. In antigen-induced LBT assays, two B. abortus antigens were used: a heat-killed strain S1119 (HKA) and a gamma-irradiated strain 19 (gamma BA). The LBT responses of three steers vaccinated with live B. abortus strain 19 were compared with three nonvaccinated steers in three separate experiments. Using HKA, FH separation resulted in an overall greater LBT response for vaccinates than nonvaccinates and a greater differential between responses of vaccinates and nonvaccinates than did S-MN derived MNC regardless of the anticoagulant used. Using gamma BA, FH produced the most responsive MNC in one experiment and S-MN/heparin produced the most responsive MNC in the other. At the highest cell concentration tested, FH-separated MNC had the greatest LBT responses for vaccinated calves, but differences between S-MN- and FH-separated MNC responses were not significantly different (P greater than 0.05). In conclusion, S-MN is a rapid and simple technique for separation of MNC from bovine blood. The technique produces an adequate cell population for mitogen-induced LBT studies; however, FH-separated MNC were generally more responsive in the B. abortus-induced LBT assay.

Functional T cell recognition of synthetic peptides corresponding to continuous antibody epitopes of herpes simplex virus type 1 glycoprotein D.

Four synthetic peptides which correspond to continuous antibody epitopes of herpes simplex virus (HSV) type 1 glycoprotein D (gD) within amino acid residues 1-23 (8-23), 268-287 and 340-356 were evaluated for in vitro stimulating activity on HSV-primed murine T lymphocytes. All peptides stimulated lymphoproliferative responses and interleukin 2 (IL2) production from draining lymph node (LN) cell populations taken 5 days after footpad immunization with live HSV. Similar responses were elicited from splenic memory T cells only if these T cells were restimulated with HSV in vitro and rested prior to peptide stimulation. Furthermore, peptide stimulated memory T cell populations released soluble factor(s) into the culture supernates which modulated the induced lymphoproliferative and cytotoxic T lymphocyte (CTL) activities of HSV-stimulated, HSV-immune splenocytes (indicator cultures). Memory T cell supernates suppressed lymphoproliferation of indicator cultures, while CTL activity of indicator cultures was either enhanced or suppressed, depending on the peptide and concentration. In contrast, supernates generated by peptide stimulation of draining LN cells had no effect on CTL activity of indicator cultures. However, the lymphoproliferative responses were augmented with three of the four peptides at the highest concentration of peptides tested. Our experiments indicate T helper (Th) and T suppressor (Ts) lymphocyte recognition of four synthetic peptides which encompass continuous antibody epitopes of HSV gD. Immunization with one of these peptides (1-23) induces virus neutralizing antibodies and protection against lethal viral challenge. Th lymphocyte recognition of this peptide in particular, together with its observed function in the induction of protection against HSV infection, indicates that this peptide is a promising candidate as a synthetic vaccine against HSV infection.

An optimized enzyme-linked immunosorbent assay for quantitative diagnosis of bovine fascioliasis.

An optimized immunoassay for detection of antibody to Fasciola hepatica antigen in cattle was developed through the adaptation of a kinetics-dependent, enzyme-linked immunosorbent assay (k-ELISA) to a microplate format. Enhanced sensitivity and a strict quantitative nature were achieved with the utilization of enzyme kinetics. With this k-ELISA, significant (P less than 0.01) elevations in anti-F. hepatica antibody could be detected as early as 2 wk post-infection in experimentally infected calves. Furthermore, fluke-burden related differences in anti-F. hepatica antibody levels between 3 different levels of fluke infection were evident.

Regulation of herpes simplex virus-specific cell-mediated immunity by a specific suppressor factor.

Our study was designed to investigate the nature of an antigen-specific suppressor factor generated by antigen-stimulated herpes simplex virus (HSV)-immune splenocytes. Factor SF-200, a 90,000- to 100,000-dalton fraction obtained after Sephacryl gel filtration, suppressed the generation of HSV-specific cytotoxic T-lymphocyte and lymphoproliferative responses. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis of SF-200 indicated that it contained an I-J+, anti-idiotypic protein. It was possible to adsorb the suppressor activity of SF-200 to an anti-I-J immunoaffinity column. The suppressor activity could be eluted from the immunoaffinity column with a low-pH buffer. The acid-eluted material was determined to be both I-J+ and reactive with anti-HSV antiserum by Western blot analysis. Both SF-200 and the I-J+ suppressor activity suppressed only HSV-specific cell-mediated immunity responses. However, it was possible to generate nonspecific suppressor activity by incubating the I-J+ suppressor factor with Lyt 1+ splenocytes from HSV-immune mice. The implication of these results with respect to the model for a suppressor cell circuit regulating HSV-specific cell-mediated immunity responses is discussed.

Diagnosis of Fasciola hepatica infection in beef calves by plasma enzyme analysis.

Plasma analysis for albumin, total bilirubin, and total protein values and aspartate aminotransferase (AST), arginase, and gamma-glutamyl transpeptidase (GGT) activities was used for the early and quantitative diagnosis of experimental Fasciola hepatica infections in beef calves. Calves were infected on 3 occasions with 1,000 (n = 5), 100 (n = 5), or 10 (n = 4) metacercariae for a total infective dose of 3,000, 300, or 30, respectively. Albendazole (15 mg/kg of body weight) was administered to 7 infected calves on postinfection (initial) week (PIW) 13. All calves were euthanatized and necropsied on PIW 16 for the determination of fluke infections. Plasma constituents were determined weekly. Significant (P less than 0.05) increases in AST activity occurred as early as PIW 4 and GGT activity at PIW 9, as compared with that in noninfected controls. Fluke burden-related differences were observed in GGT activity from PIW 9 onward. Increases in AST activity reflected parenchymal liver damage, whereas increases in GGT reflected hepatobiliary damage; therefore, differentiation could be made between the migratory and ductal phases of the infection. There was no correlation between arginase activity and fluke infection. As compared with fecal examination results, plasma enzyme analysis gave an earlier and semiquantitative indication of F hepatica infection in experimentally infected calves. Although increases in these plasma constituents were not definitely diagnostic of fascioliasis, useful information on the size of the fluke burden and progress of the disease process could be obtained by these methods. Plasma enzyme analyses of AST and GGT were not indicative of chemotherapeutic success or failure when calves with mature F hepatica (14 weeks old) infections were treated.

Acquired spontaneous cytotoxicity of K562 in a subpopulation of monocytes.

Human peripheral blood monocytes were placed on a discontinuous density gradient of bovine serum albumin and fractionated into five subpopulations. Cells from each subpopulation were assayed for spontaneous cytotoxic activity against K562 tumor cells. Immediately following fractionation, monocytes were not cytotoxic. Following incubation for at least 48 hr, monocytes from two layers of the gradients clearly exhibited greater spontaneous cytotoxic activity than all others. The degree of cytotoxicity expressed by cells of these layers was enhanced by the addition of indomethacin and inhibited by prostaglandin E2 (PGE2). Monocytes acquiring spontaneous cytotoxicity did not secrete measurable levels of PGE2 and had increased levels of purine nucleoside phosphorylase after 72 hs of culture in vitro. Surface markers HNK-1 and Mac-1 normally associated with cytotoxic function, were detected on these cells by indirect immunofluorescence at isolation and after culture. The fraction with the greatest cytotoxic activity showed an increase in the proportion of cells displaying reactivity to HNK-1 after culture compared to initial isolation.

Efficacy of a benzenedisulfonamide against experimental Fasciola hepatica infections in calves.

The anthelmintic efficacy of a benzenedisulfonamide was evaluated by administering the drug parenterally at dosage levels of 2, 4, 8, and 16 mg/kg of body weight to crossbred Brahman calves with experimental Fasciola hepatica infections. In the 3-week period after treatment, fluke ova counts of treated calves were markedly reduced from counts obtained just before treatment. At necropsy, the mean fluke recovery for all 4 benzenedisulfonamide dosages were significantly (P less than 0.01) reduced. The efficacy of benzenedisulfonamide against F hepatica at dosage levels of 2, 4, 8, and 16 mg/kg was 97.5%, 99.5%, 100%, and 100%, respectively.