An efficient, single-step method for purification of the 110-kilodalton (kDa) hemolysin of Actinobacillus pleuropneumoniae was developed. An immunoaffinity column was made by cross-linking murine monoclonal antibody 8C2 to the 110-kDa hemolysin of A pleuropneumoniae strain J45 serotype 5 to protein A-agarose beads. Purified hemolysin with high hemolytic activity was obtained after washing the column with phosphate-buffered saline solution, and eluting the hemolysin with 50 mM diethylamine, pH 11.0. The same column was also used to purify the hemolysin from A pleuropneumoniae strain 4074 serotype 1. The purification procedure could be completed within 5 hours, and almost 50% of the total hemolytic activity and hemolysin protein was recovered in pure form.

Postoperative abdominal fluid changes were compared in 2 groups of horses; those undergoing double small-colon resection and anastomosis (n = 10) and those undergoing exploratory celiotomy alone (n = 5). Peritoneal fluid was collected before surgery and on postoperative days 1, 3, 5, and 7. Total and differential nucleated cell counts, RBC numbers, and total protein and fibrinogen concentrations were evaluated. In both groups, all values were significantly higher than normal on the first postoperative day (after small-colon resection and anastomoses, WBC = 130,350 +/- 23,310 cells/microliter, RBC 7,389,000 +/- 6,234,000 cells/microliter, total protein = 3.63 +/- 0.16 g/dl; after exploratory celiotomy alone, WBC = 166,620 +/- 34,340 cells/microliter, RBC = 295,000 +/- 86,070 cells/microliter, total protein 4.38 +/- 0.54 g/dl). The number of total peritoneal nucleated cells and RBC significantly decreased after the first postoperative day, whereas total protein and fibrinogen concentrations, percent neutrophils, and percent mononuclear cells remained unchanged. None of the values had returned to normal by postoperative day 7 (after small-colon resection and anastomoses, WBC = 45,600 +/- 8,765 cells/microliter, RBC = 95,390 +/- 53,380 cells/microliter, total protein = 4.39 +/- 0.23 g/dl; after exploratory celiotomy alone, WBC = 43,340 +/- 7,746 cells/microliter, RBC = 12,860 +/- 11,790 cells/microliter, total protein = 3.92 +/- 2.20 g/dl.) The resection and anastomosis group had a significantly lower total protein concentration on the first postoperative day and a significantly higher mean total RBC count over the entire 7-day postoperative evaluation than did horses that underwent celiotomy alone. Other values in the 2 groups of horses did not differ significantly. As a result, there was insufficient evidence to conclude that resection and anastomosis of the small colon in healthy horses causes a different inflammatory response than does manipulation of the intestine alone.

When Salmonella typhimurium and Clostridium perfringens were tested in conventional chickens, larger numbers of S typhimurium and C perfringens adhered to Eimeria tenella-infected ceca than to uninfected ceca. In germ-free chickens, S typhimurium and C perfringens adhered to the E tenella-infected cecal mucosa more than to the uninfected cecal mucosa, but fewer Bacteroides vulgatus and Bifidobacterium thermophilum adhered to the E tenella-infected ceca than to the uninfected ceca. Many bacteria adhered to the lesions caused by E tenella as observed by scanning electron microscopy. On the basis of our findings, we suggest that infection with E tenella upsets the balance of competitive adherence of bacteria, allowing more colonization of S typhimurium and C perfringens.

Three-week-old weaned and colostrum-deprived neonatal (< 1 day old) pigs were inoculated to determine the pathogenicity of 2 enterotoxigenic Escherichia coli isolates that do not express K88, K99, F41, or 987P adhesins (strains 2134 and 2171). Strains 2134 and 2171 were isolated from pigs that had diarrhea after weaning attributable to enterotoxigenic E coli infection. We found that both strains of E coli adhered in the ileum and caused diarrhea in pigs of both age groups. In control experiments, adherent bacteria were not seen in the ileum of pigs < 1 day old or 3 weeks old that were noninoculated or inoculated with a nonpathogenic strain of E coli. These control pigs did not develop diarrhea. Antisera raised against strains 2134 and 2171 and absorbed with the autologous strain, grown at 18 C, were used for bacterial-agglutination and colony-immunoblot assays. Both absorbed antisera reacted with strains 2134 and 2171, but not with strains that express K99, F41, or 987P adhesins. A cross-reaction was observed with 2 wild-type K88 strains, but not with a K12 strain that expresses K88 pili. Indirect immunofluorescence with these absorbed antisera revealed adherent bacteria in frozen sections of ileum from pigs infected with either strain. We concluded that these strains are pathogenic and express a common surface antigen that may be a novel adhesin in E coli strains that cause diarrhea in weaned pigs.

The role of silage feeding in the origin of an epizootic of encephalitic listeriosis in a sheep flock was investigated by use of a new direct Listeria-selective isolation and enumeration medium, in combination with serotyping and phage typing. The silage contained high numbers (about 10(6) cells/g) of a L monocytogenes strain indistinguishable with respect to serovar and phagovar from that isolated from the brains of sick sheep. These results provided unambiguous bacteriologic evidence of the epidemiologic link between silage consumption and listeriosis in ruminants.

Pharmacokinetics, CSF penetration, and hematologic effects of oral administration of pyrimethamine were studied after multiple dosing. Pyrimethamine (1 mg/kg of body weight) was administered orally once a day for 10 days to 5 adult horses, and blood samples were collected frequently after the first, fifth, and tenth doses. The CSF Samples were obtained by cisternal puncture 4 to 6 hours after administration of the first, third, seventh, and tenth doses. Pyrimethamine concentration in plasma and CSF Was quantified by gas chromatography, and plasma concentration-time data were analyzed, using a pharmacokinetic computer program. Repeated daily dosing resulted in accumulation of pyrimethamine in plasma, with steady state being achieved within 5 days, when the mean peak plasma concentration was more than twice that measured after the first dose. Pyrimethamine concentration in CSF was 25 to 50% of corresponding plasma concentration and did not appear to accumulate with successive administration of doses. Blood samples collected during and after the dosing regimen were submitted for hematologic analysis; neutrophil numbers decreased slightly, but remained within normal range for adult horses.

It has been established that L-gamma-glutamylated derivatives of alpha-amino acids are delivered more efficiently to the kidneys than are the parent alpha-amino acids. Therefore, we synthesized L-gamma-glutamyl-S-(1,2-dichlorovinyl)-L-cysteine (L-gamma-glutamyl-L-DCVC), the simplest L-gamma-glutamylated derivative of the nephrotoxic alpha-amino acid S-(1,2-dichlorovinyl)-L-cysteine (L-DCVC), and investigated its effects on renal function and ultrastructure in pentobarbital-anesthetized dogs. Intravenous doses of 23.15 and 92.60 micromoles of L-gamma-glutamyl-L-DCVC/kg of body weight induced significant increases in urinary protein output and significant decreases in the clearance of inulin during the 6-hour post-injection period. Changes were not observed in any of the other 13 renal function variables or in the 11 plasma and blood variables that were monitored throughout the same period. Both doses of L-gamma-glutamyl-L-DCVC induced renal ultrastructural lesions in the S1 and S2 cells of the canine proximal tubule; the remaining 8 cell types downstream and the glomeruli were not damaged. The onset and magnitude of renal function changes and the cell types affected by L-gamma-glutamyl-L-DCVC were virtually identical to those observed previously following IV administration of equivalent doses of L-DCVC to pentobarbital-anesthetized dogs. Rapid removal of the L-gamma-glutamyl group from L-gamma-glutamyl-L-DCVC (ie, deglutamylation) resulting in formation of the parent alpha-amino acid, L-DCVC, can best explain the extreme similarity in the nephrotoxic profiles of these 2 toxicants.

The B1 strain of Newcastle disease virus (NDV-B1), which is nonpathogenic for newly hatched chickens, killed embryos when it was used to inoculate chicken eggs at embryonation day 18. Treatment of NDV-B1 with an alkylating agent, ethylmethane sulfonate (EMS) markedly reduced the pathogenicity of the virus for 18-day-old chicken embryos. Eggs inoculated with the modified virus (NDV-B1-EMS) hatched, and the virus was isolated from lungs and spleen of 1-day-old chickens. The hatched chickens developed antibody to NDV and were protected against challenge exposure (at 4 weeks of age) with a highly virulent GB-Texas strain of NDV. Presence of maternal antibody to NDV in embryonating eggs did not influence the protective ability of NDV-B1-EMS, which also induced protective immunity when administered to 4-week-old chickens. The 50% protective dose of NDV-B1-EMS in maternal antibody-negative and -positive embryos was calculated to be 10.77 and 17.70 embryo 50% lethal doses, respectively. Results of the study indicated that NDV-B1-EMS may be used as an embryo vaccine to protect chickens against Newcastle disease.

Reference strains for Haemophilus parasuis serovars 1 to 7 were examined for virulence by inoculation of guinea pigs. Guinea pig response to intraperitoneal inoculation was similar for the 7 reference strains. However, apparent differences in virulence were detected after intratracheal inoculation. Cells of the reference strains for serovars 1 and 5 were most invasive, causing moribundity or death at higher doses and a persistent septicemia at lower doses. Haemophilus parasuis could be isolated from respiratory and systemic sites; purulent bronchopneumonia, pericarditis, and pleuritis were apparent in infected guinea pigs. Inoculation of cells of the reference strains for serovars 2 and 6 also resulted in bronchopneumonia and moribundity or death in some guinea pigs; however, reisolation of H parasuis and microscopic lesions at necropsy were less pronounced than those observed with serovars 1 and 5. Inoculation of cells of serovars 3, 4, and 7 induced only transient clinical signs and minimal evidence of H parasuis infection at necropsy. The data from intratracheal inoculation of guinea pigs are similar to data from other investigations in swine, indicating differences in the pathogenic potential of H parasuis strains. Thus, guinea pigs may be useful as a laboratory animal model for examining cellular factors associated with virulence and immunogenicity of H parasuis.

A study for about a 30-month period was done to compare strongyle control programs, using per os treatments of ivermectin (IVE) paste exclusively or alternation of 4 antiparasitic paste compounds: IVE, oxfendazole (OFZ), oxibendazole (OBZ), or pyrantel pamoate (PRT). Every 8 weeks, 1 group of horses (barn C; n = 14 to 16) was given IVE paste exclusively, and a second group (barn E; n = 16) was given the 4 antiparasitic pastes on an alternating schedule. Worm eggs and larvae per gram of feces (epg and lpg, respectively) values were determined every 2 weeks during the investigation. This study in grazing horses (mares and fillies), naturally infected with internal parasites, was conducted during the period between Oct 22, 1987 and Feb 8, 1990, with an additional observation on Mar 28, 1990. For barn-C horses, treated exclusively with IVE (200 micrograms/kg of body weight) 14 times, 2-week posttreatment mean strongyle epg and lpg (small strongyle) values were reduced 99 to 100%. Mean strongyle epg and lpg (small strongyle) values for each 2-week sample period remained low (< 20) throughout the study period, except for 1 moderate transient increase in July 1988. For the entire study period, the aggregate mean strongyle epg value was 12 and the lpg value was 6. Two-week posttreatment mean strongyle epg and lpg (small strongyle) values for barn-E horses, treated alternately with therapeutic (approx) dosage of IVE (200 micrograms/kg, 4 times), OFZ (10 mg/kg; 5 times), OBZ (10 mg/kg; 4 times), or PRT (6.6 mg base/kg; 2 times), varied within and between compounds. Posttreatment (2-week) mean epg values were reduced 100% by IVE, 0 to 100% by OFZ, 74 to 100% by OBZ, and 92 to 100% by PRT. Mean small strongyle lpg values at 2 weeks after treatment indicated reduction of: 100% for IVE, 0 to 76% for OFZ, 43 to 100% for OBZ, and 97 to 100% for PRT. For the entire study period, the aggregate mean strongyle epg value was 54 and the lpg value was 64. The epg and lpg reduction values for the 2 benzimidazoles indicated an increase in the benzimidazole-resistant segment of small strongyles. Fecal cultures for horses in both groups contained larvae of large strongyles (Strongylus vulgaris and S edentatus) only at the time of initial treatment. Treatment program evaluations included necropsy of 9 foals (56 to 203 days old), 7 born to barn-C mares and 2 born to barn-E mares. Generally, low numbers of bots (Gasterophilus intestinalis) and Habronema muscae were found in foals of both groups. Intestinal stages of immature and mature ascarids (Parascaris equorum) were also found in foals born to both groups of mares. Mature pinworms (Oxyuris equi) were found in 1 barn-C foal. Only barn-E foals had Strongyloides westeri. Small strongyles were detected in foals of both groups, up to several thousand in some. The latter finding indicates, in particular, that although barn-C mares had low small strongyle epg and lpg values throughout the study, eggs and larvae built up on pasture in sufficiently high numbers for major transmission to foals born there. Small strongyles in foals were composed of 3 genera and 10 species for barn-C foals and of 3 genera and 8 species for barn-E foals. Seasonal transmission of parasites also was observed.