Oral and parenteral vaccination of mice with protein-ergotamine conjugates and evaluation of protection against fescue toxicosis

Acremonium coenophialum produces ergopeptide alkaloids in tall fescue (Festuca arundinacea Schreb.). These ergot alkaloids decrease serum alkaline phosphatase (ALP) activity, serum cholesterol and prolactin concentrations, as well as average daily gains (ADG) in cattle. The objective of this study was to evaluate the protection of anti-ergotamine antibodies induced by either oral or parenteral vaccination with protein-ergotamine conjugates or passive vaccination with anti-ergovaline, monoclonal antibodies in a murine model of fescue toxicosis. Ergotamine (EG) was conjugated to bovine serum albumin (BSA) and cholera toxin subunit B (CTB) by the Mannich reaction. Mice were blocked based on weight and randomly allocated into five groups of 10 mice each. Treatment groups were as follows: (1) group vaccinated intraperitoneally (ip) with a BSA-EG conjugate and fed an endophyte-infected (EI) fescue diet (BSA-EG group); (2) group orally vaccinated with a CTB-EG conjugate mixed with free cholera toxin (CT) and fed an EI fescue diet (CTB-EG group); (3) nonvaccinated group fed an EI fescue diet (EI group); (4) group passively vaccinated with anti-ergovaline, monoclonal antibodies and fed an EI fescue diet (MoAB group); and (5) nonvaccinated group fed an endophyte-free (EF) fescue diet (EF group). The EI diet contained 1.5 ppm of Ergovaline (EV), whereas no EV was detected in the EF diet.Respective diets were similar upon nutritional analysis. Unvaccinated mice in the EI group exhibited features of fescue toxicosis as indicated by decreased serum ALP activity and cholesterol, and decreased weight gain as compared to mice in the EF group. Antibodies against EG and EV were present in sera of mice in the BSA-EG and MoAB groups, respectively. Mice orally vaccinated with the CTB-EG conjugate developed secretory IgA (sIgA) antibodies and short-lived, systemic IgG responses against EG. Weight gains were increased in the BSA-EG and CTB-EG groups and tended to be increased in the MoAB group vs. the unvaccinated EI group. Serum ALP activity was decreased in the BSA-EG and MoAB groups as compared to the EF group. Serum ALP activity was further decreased in the BSA-EG vaccinated group as compared to the EI group. Cholesterol concentrations were decreased in the EI, BSA-EG and MoAB groups as compared to the EF group. Prolactin concentrations were similar in all groups.     

Angiotensin-converting enzyme gene polymorphism is associated with coronary heart disease in non-insulin-dependent diabetic patients evaluated for 9 years

OBJECTIVES: To assess effects of vaccination against fescue toxicosis on weight gain, serum prolactin and cholesterol concentrations, and alkaline phosphatase (ALP) activity in mice fed an endophyte-infected (EI) or endophyte-free (EF) fescue diet. ANIMALS: 50 six-week-old male BALB/c mice. PROCEDURE: Mice were randomly allocated to the following 5 groups: 1, vaccinated intraperitoneally with a bovine serum albumin-ergotamine (EG) conjugate and fed an EI fescue diet; 2, orally vaccinated with cholera toxin (CT) subunit B-EG conjugate mixed with free CT and fed an EI fescue diet; 3, not vaccinated and fed an EI fescue diet; 4, passively vaccinated with monoclonal antibodies specific for ergovaline (EV) and fed an EI fescue diet; and 5, not vaccinated and fed an EF fescue diet. RESULTS: Antibodies against EG and EV were in serum of mice of groups 1 and 4, respectively. Secretory IgA and IgG coproantibodies against EG were induced in mice of group 2. Weight increased in groups 1 and 2 and tended to be increased in group 4 versus group 3. Prolactin concentration was similar in all groups; cholesterol concentration was decreased in groups 1, 3, and 4, compared with group 5. Compared with that in group 5, serum ALP activity decreased in groups 1 and 4 and was further decreased in group 1, compared with that in groups 2 and 3; it was negatively correlated with anti-EG titer. CONCLUSIONS AND CLINICAL RELEVANCE: Induction of anti-EG antibodies and administration of EV monoclonal antibodies tended to increase short-term weight gain in this murine model of fescue toxicosis. However, systemic IgG antibodies against EG or EV antibodies were not protective against decreases in serum ALP activity and cholesterol concentrations. Clinical significance of decreased ALP activity associated with vaccination is unknown, but represents a worsening of a response often associated with fescue toxicosis in cattle.     

Monocyte immune cell response and copper status in beef steers that grazed endophyte-infected tall fescue

A 3-yr study was conducted to evaluate immune response and Cu status of yearling beef steers as a consequence of grazing tall fescue (Festuca arundinacea Schreb.) infected (E+) with the endophyte fungus Neotyphodium coenophialum ([Morgan-Jones and Gams] Glenn, Bacon, and Hanlin). During a preliminary study in 1994, 24 weanling Angus and Angus x Hereford steers were blocked by breed and weight (initial BW 271 kg; SD 25) and were randomized to E+ and low endophyte (E-) fescue in pastures at Glade Spring, VA. Grazing began in April and was discontinued in July. In 1995 and 1996, 24 weanling Angus and Angus x Hereford steers (initial BW 249 kg, SD 20 and 240 kg, SD 15, respectively) were randomized to the E+ and E- pastures at Glade Spring during each year. Grazing began in April and continued until September in 1995 and October in 1996. In 1994, steers that grazed E+ fescue exhibited lower (P < .05) phagocytic activity, major histocompatibility complex (MHC) class II expression, ceruloplasmin, and serum Cu than steers that grazed E- tall fescue. During 1995, steers grazing E+ fescue had lower (P < .05) phagocytic activity and MHC class II expression than steers that grazed E- fescue. In 1996, one-half of the steers within each paddock received a Cu oxide bolus at the beginning of the grazing season. During 1996, phagocytic activity was lower (P < .01) and MHC class II expression tended (P < .07) to be lower in steers that grazed E+ tall fescue than in steers that grazed E- tall fescue. Copper supplementation increased (P < .05) MHC class II expression in July regardless of endophyte status over nonsupplemented steers. Steers that grazed E- tall fescue had higher (P < .05) plasma or serum Cu concentrations than steers that grazed E+ tall fescue in each year of the study. These data indicate that the endophyte compromised the immune function of grazing steers, and the data suggest a relationship with depressed Cu status.     

An evidence-based approach to pressure- and volume-limited ventilation strategies

To assess the efficacy of ammoniation in the detoxification of endophyte-infected tall fescue (Festuca arundinacea Schreb), 40 male Harlan Sprague-Dawley rats were randomly assigned to the following four treatments for 28 d: endophyte-free (E-), endophyte-infected (E+), ammoniated (2% dry matter basis, 7 d) endophyte-free (AE-), and ammoniated endophyte-infected (AE+) tall fescue seed. Total pyrrolizidine alkaloid (N-acetyl and N-formyl loline) and ergovaline contents of endophyte-infected fescue seed were reduced 24 and 54%, respectively, by ammoniation. Endophyte-infected treatment groups had lower (P < 0.01) daily feed intakes (DFI), daily weight gains (DWG), feed efficiencies, and primary serum hemagglutination titers to sheep red blood cell (SRBC) immunization than endophyte-free treatment groups. Performance parameters were higher (P < 0.01) for ammoniated diets in comparison to non-ammoniated diets; however, anti-SRBC titers were not significantly different. When compared to the E+ diet, the AE+ diet increased (P < 0.01) DFI (24%), DWG (41%) and feed efficiency (13%).     

Comparison of two ammoniation procedures to reduce the toxicity of endophyte-infected tall fescue seed fed to rats

To determine the effect of extending the duration of ammonia (2% dry matter basis) treatment ti'om 1 to 5 wk on the toxicity of endophyte-infected tall fescue seed, 60 male Harlan Sprague-Dawley rats were randomly assigned to the following six treatments during a 28-d trial: endophyte-free (E-), endophyte-infected (E+), 1 wk ammoniated endophyte-fee (1AE-), 1 wk ammoniated endophyte-infected (1AE+), 5 wk ammoniated endophyte-free (5AE-), and 5 wk ammoniated endophyte-infected (5AE+) tall fescue seed. The concentration of total pyrrolizidine alkaloids (N-acetyl and N-formyl loline) or E+ rescue was reduced from 4203 12 g/g to 3009 and 2533 I-tg/g by the 1AE+ and 5AE+ treatments, respectively. Ergovaline was lowered from 3.77 to 1.57 12 g/g by 1AE+ and eliminated by 5AE+. Endophyte-infected treatment groups had depressed (P < 0.0001) dally feed intakes (DFI), daily weight gains (DWG), feed efficiencies (G/F), primary antibody responses, and T cell and B cell mitogenic responses than endophyte-free treatment groups. Ammoniation of endophyte-infected rescue seed improved DFI and DWG (P < 0.0001) and G/F (P < 0.05); however, there was no difference in performance criteria between the 1-wk and 5-wk ammoniation treatments. Endophyte-induced depressions in immune function were not alleviated by ammoniation.     

Vaccination against ergot alkaloids and the effect of endophyte-infected fescue seed-based diets on rabbits

Three sequential experiments were conducted with rabbits to 1) determine the effect of endophyte-infected (E+) tall fescue seed on rabbit performance and examine the effect of anti-ergot alkaloid immunization on rabbit performance and protectiveness against fescue toxicosis, 2) compare immunogens designed to elicit systemic anti-ergot alkaloid antibodies, and 3) select a superior adjuvant. In Exp. 1, rabbits (n = 6/treatment) fed E+ fescue seed diets (20%, 340 ppb total ergot alkaloids) had reduced (P < .05) intake and weight gain compared with endophyte-free (E-) controls, whereas apparent diet digestibility was not different between E+ and E-. Rabbits immunized against ergot alkaloids (E+ vac) with lysergol conjugated to human serum albumin (Ly-HSA) had greater (P < .05) intake than E+ rabbits during the wk 1 of a 3-wk dietary challenge. In Exp. 2, rabbits (n = 4/treatment) were immunized with Ly-HSA, with H100-B (ergot alkaloid hapten, H100-different protein carrier, B conjugate), or combinations of both with alum as adjuvant. Greatest (P < .001) anti-ergot alkaloid antibody (Ab) titer developed in the group immunized with H100-B. In Exp. 3, rabbits (n = 4/treatment) were immunized with the immunogen H100-B in conjunction with six adjuvants. Freund's incomplete adjuvant (FIA) in combination with DEAE-dextran and FIA alone gave highest anti-ergot titers. In summary, rabbit weight gain and intake were reduced by feeding E+ fescue seed diets, immunization against ergot alkaloids provided temporary improvement in intake, and H100-B conjugate with FIA or FIA + DEAE-dextran as adjuvants elicited a superior anti-ergot immune response. We believe that rabbits may serve as a model animal for fescue toxicosis research.     

Plasma concentrations of prolactin, growth hormone, and luteinizing hormone in steers administered ergotamine or ergonovine

This research investigated whether ergot alkaloids associated with endophyte-infected tall fescue could alter plasma concentrations of pituitary hormones that regulate biological processes related to cattle performance. Seven Angus yearling steers received single i.v. injections of ergotamine tartrate, ergonovine maleate, or saline vehicle in a simple cross-over design. Each steer was given a different compound each week. Blood samples were collected at 15-min intervals for 45 min before and 240 min after treatments to assess plasma concentrations of prolactin, growth hormone, and LH. Respiratory rates were measured hourly to ascertain a systemic effect. Ambient temperature averaged 34 degrees C during data collection. Treatment x time was a significant source of variation for respiration rate and plasma concentrations of each hormone evaluated. Respiration rates were higher for ergonovine than for saline (P < .02) and ergotamine (P < .07) 30 min after treatment, but they were higher (P < .05) for ergotamine than for ergonovine and saline by 210 min after treatment. Both alkaloids transiently elevated (P < .01) plasma growth hormone concentrations compared with before alkaloid treatment and after saline treatment. Ergotamine reduced (P < .01) plasma concentrations of prolactin and LH throughout the 120-min period after treatment compared with concentrations before ergotamine treatment and after saline treatment. Ergonovine lowered (P < .01) prolactin concentrations for a shorter time than ergotamine and did not affect mean LH concentrations. Results indicated that ergot alkaloids implicated as contributing agents to fescue toxicosis can alter plasma concentrations of pituitary hormones important to cattle production.     

Growth and subsequent feedlot performance of estradiol-implanted vs nonimplanted steers grazing fall-accumulated endophyte-infested or low-endophyte tall fescue

A growing-finishing study using Angus steer calves was conducted in three phases: 1) grazing stockpiled 'Kentucky-31' tall fescue (Festuca arundinacea Schreb.) with high (65%; HE KY-31) and low (0%; LE KY-31) infestation rates of Acremonium coenophialum Morgan-Jones and Gams and 'Kenhy' and 'Johnstone' tall fescue with low (< 1%) infestation rate of Acremonium coenophialum from October 24 to December 19; 2) drylot feeding of Johnstone and HE KY-31 haylage (December 19 to April 10); and 3) feedlot finishing on a common high-concentrate diet (April 11 to August 1). In Phase 1, ADG was greatest (P < .05) for Kenhy, intermediate for Johnstone and LE KY-31, and lowest (P < .05) for HE KY-31. Implantation with estradiol 17-beta increased ADG (P < .01) by 23, 27, 7, and 2% for steers grazing Johnstone, HE KY-31, LE KY-31, and Kenhy, respectively. Dry matter digestibility and DMI of stockpiled Johnstone and HE KY-31 were not different (P > .10). During Phase 2, steers consuming Johnstone haylage had greater (P < .01) DMI, ADG, and gain:feed ratio (G:F) than steers consuming HE KY-31 haylage. During Phase 3, steers previously consuming Johnstone had greater DMI (P < .10); however, steers previously fed HE KY-31 had greater ADG (P < .05) and G:F (P < .01). By the end of the study, steer body weights were not different (P > .10) between treatments. These data indicate that growth-decreasing effects of endophyte-infested fescue were evident at hypothermal-ambient temperatures.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cottonseed meal or feather meal supplementation of ammoniated tropical grass hay for yearling cattle

Despite high N intake by cattle consuming ammoniated forages, improvements in ADG might be expected be feeding proteins that supply ruminally available and(or) undegraded intake protein. Growth studies with 220-kg steers were conducted to evaluate ammoniated hay plus a liquid cane molasses-based supplement containing urea or a combination of urea and .07, .14, or .21 kg of CP daily from cottonseed meal (CSM) or feather meal (FM). Steers grazed dormant pasture and were fed hay plus 1.36 kg of DM daily per steer of the supplement. In each of 3 yr, total DM intake was similar (P > .60) among treatments, averaging 3.0% of BW. Daily gain by steers supplemented with urea and maximum ADG were similar across years, ranging from .21 to .23 kg and .50 to .60 kg, respectively. In all years for CSM supplementation, and in two of the three years for FM supplementation, increasing level of CP supplementation resulted in linear (P < .01) increases in ADG and gain:feed ratio (GF), with no differences (P > .10) between the protein sources. In one year, ADG and GF by steers supplemented with FM were maximized at the .07 kg of CP level, suggesting greater efficiency of protein utilization in FM. Improved performance by cattle fed ammoniated tropical grass hay may be obtained by feeding a protein supplement that provides preformed protein that is either moderately or highly resistant to ruminal degradation. Ruminal degradation characteristics of protein sources may influence efficiency of protein utilization in cattle fed ammoniated tropical grass hay plus molasses-based supplements.     

Effect of ergotamine and ergonovine on plasma concentrations of thyroid hormones and cortisol in cattle

Plasma samples from two experiments were processed to determine whether ergot alkaloids associated with endophyte-infected tall fescue altered peripheral thyroxine (T4), triiodothyronine (T3), or cortisol concentrations in cattle. In Exp. 1, seven Angus steers (294 kg) received i.v. bolus injections of saline (SAL), ergonovine maleate (7 mg; EM), or ergotamine tartrate (7 mg; ET) at weekly intervals, and they received all treatments during the study. Blood was sampled every 15 min for 5 h, and treatments were given after h 1. Mean ambient temperature was 34 degrees C. Treatment x time affected plasma concentrations of T3 (P < .05) and of cortisol (P < .001) but not that of T4 (P > .2). Plasma T3 concentrations were not affected by SAL, whereas concentrations increased (P < .01) after either EM or ET treatment. Plasma cortisol concentrations were not altered by SAL or EM, but they were increased (P < .001) by ET treatment. In Exp. 2, six Holstein cows (499 kg) nursing calves received a bolus i.v. injection of SAL, EM (9.5 mg), or ET (9.5 mg) per estrous cycle, and all treatments were given over three cycles. Blood was sampled every 20 min for 5 h; treatments were given after h 1. Mean ambient temperature was 26 degrees C. Treatment x time affected T3 (P = .08) and cortisol (P < .001) and tended to influence (P = .16) T4 concentrations. Plasma T3, T4, and cortisol concentrations were not influenced by SAL treatment. Plasma T3 was higher (P < or = .01) after EM or ET treatment compared with pretreatment concentrations. Concentrations of T4 during the 4 h after EM and ET were increased (P < .001) compared with pretreatment. Plasma cortisol concentrations were not altered by EM but were increased (P < .001) by ET. Ergot alkaloids implicated as contributing agents to fescue toxicosis alter plasma concentrations of hormones important to metabolic and thermoregulatory functions in cattle.     

Modulation of endothelium-dependent hyperpolarization and relaxation to acetylcholine in rat mesenteric artery by cytochrome P450 enzyme activity

Twelve Hereford steers (average BW = 231 kg) that had previously grazed native rangeland (Range) or irrigated winter wheat pasture (Wheat) were allowed to graze locoweed-infested rangeland from April 1 to June 9, 1994 (six steers/previous grazing treatment). Relative consumption level of locoweed and other forage classes was measured as observed bites per steer. Liver biopsy and whole blood samples were obtained from each steer before and after grazing. Liver samples were analyzed for several minerals by inductively coupled plasma-atomic emission spectroscopy, and whole blood samples were analyzed for Se. Liver concentrations of Ba (P < .001), Cd (P < .001), Ca (P < .01), Cr (P < .01), Ni (P < .001), Na (P < .01), and V (P < .001) were greater and concentrations of Mn (P < .09), P (P < .01), and K (P < .07) were less in Wheat than in Range steers. Liver concentrations of Fe, Mg, S, and Zn and whole blood Se concentrations did not differ (P > .10) between the two groups. Liver concentrations of Cr (P < .04) and Mn (P < .001) were less, and Fe concentrations were greater (P < .01), in samples taken after grazing than in samples taken before grazing of locoweed-infested range. Whole blood Se concentrations decreased (P < .01) from the beginning to the end of the grazing period, but this effect was not related (P > .15) to locoweed consumption. Changes in liver concentrations of minerals were compared relative to consumption levels of all forage classes in the locoweed-infested range. Liver concentrations of Cu decreased (r2 = .45; P < .02) as the percentage of bites consumed as locoweed increased, but concentrations after grazing locoweed-infested range were still within normal ranges. Changes in liver concentrations of other minerals were not related (P > .15) to consumption of locoweed. These data indicate that previous grazing history can have significant effects on liver mineral stores and that, under our conditions, consumption of locoweed by grazing beef steers altered liver Cu concentrations. Toxic effects of locoweed consumption would likely occur before Cu deficiency would be induced by grazing locoweed-infested range; hence, supplementation of Cu would seem unlikely to alter the course of locoweed toxicosis.     

Influence of Neotyphodium coenophialum on copper concentration in tall fescue

Poor performance of livestock that graze tall fescue (Festuca arundinacea Schreb.) has been associated with the endophyte fungus Neotyphodium coenophialum [Morgan-Jones and Gams] Glenn, Bacon, and Hanlin). Recent evidence suggests lowered Cu status and a depression of Cu-related immune function in steers that graze endophyte-infected (E+) tall fescue. Greenhouse and field studies investigated relationships between the endophyte and Cu concentrations in tall fescue. Seventeen infected 'Kenhy' clones were divided, and one plant of each pair was treated three times with Benomyl to remove the endophyte (E-). Plants were watered with nutrient solution in a greenhouse for 6 mo before sampling. Copper concentrations were greater (P < .001) in E- than in E+ clones (3.4 vs 2.8 microg/g; SE, .06). In the second greenhouse experiment, genetically similar E+ and E- 'Kentucky'-31 (KY-31) and 'Georgia Jessup' were grown from seed and fertilized with nutrient solution to produce mature plants. Copper concentrations were higher (P < .05) in E- than in E+ tall fescue (8.6 vs 7.6 microg/g; SE, .3). In a field plot experiment in Texas, E+ and E- KY-31 were grown with 0, 50, and 100% replacement of potential evapotranspiration. By September, Cu concentrations were higher (P < .05) in E- than in E+ tall fescue (7.3 vs 6.6 microg/g; SE, .2). In pasture experiments, KY-31 E+ (> 70% infection level) and E- (< 5% infection level) tall fescue were grown in Virginia at two locations with three rates of N fertilizer. Copper concentrations were higher (P < .05) in E- than in E+ tall fescue (4.8 vs 4.5 microg/g; SE, .1) and increased (P < .01) linearly in response to N. Our data demonstrate that the presence of the endophyte is associated with lower Cu concentrations in tall fescue, which may contribute to lowered Cu status in animals and thus contribute to the etiology of fescue toxicity.     

Supplemental methionine and time of supplementation effects on ruminal fermentation, digesta kinetics, and in situ dry matter and neutral detergent fiber disappearance in cattle

Two experiments were conducted to study the effects of methionine supplementation on ruminal fermentation and digesta kinetics. In Exp. 1, nine ruminally cannulated beef heifers (average initial BW = 527 kg) in a crossover design were fed low-quality grass hay and cottonseed meal with or without 11.4 g of supplemental methionine (polysaccharide-coated). Particulate and fluid kinetics, rate of DM and NDF disappearance, ruminal VFA and NH3 N concentrations, and pH were not altered (P > .10) by supplemental methionine; however, ruminal purine concentration was greater (P < .05) in methionine-supplemented heifers than in unsupplemented heifers. In Exp. 2, 12 ruminally cannulated Holstein steers (average initial BW = 622 kg) grazing a fescue pasture were allotted to one of three groups: no supplemental methionine (CON) or 11.4 g of supplemental methionine fed at 0700 (AM) or at 1200 (PM). Forage intake, particulate kinetics, ruminal fluid kinetics, pH, VFA, and NH3 N concentrations were not altered (P > .10) by supplemental methionine or supplementation time. In situ rate of DM and NDF disappearance was greater (P < .05) in supplemented steers than in CON steers; AM steers exhibited faster (P < .05) rates than PM steers. Overall, methionine supplementation of low-quality forage increased ruminal purine concentration but did not alter in situ fermentation or digesta passage, whereas supplementation at 0700, but not at 1200, of steers grazing fescue forage increased rate of NDF fermentation.     

Alfalfa and escape protein supplements for grazed corn residues

Two calf corn-residue grazing trials were conducted to evaluate the effect of supplementing with two alfalfa hays (Medicago sativa), varying in their amount of CP and ADF, with and without escape protein (EP). In Trial 1, heifers (204 kg) grazed (1.36 heifers/ha) non-irrigated fields for 92 d. No difference (P > .05) in ADG was found when medium-protein (16.3% CP) and high-protein (18.0% CP) alfalfa hays were provided at 2.7 kg of DM.heifer-1.d-1. In Trial 2, 60 steers (257 kg) were supplemented individually with medium-protein (17.6% CP) alfalfa hay, 1.86 (MPL) and 2.27 kg DM (MPH), and one level of high-protein (18.7% CP) alfalfa hay, 1.86 kg DM (HPL). In a 2 x 3 factorial arrangement of treatments, steers were fed with or without EP (113 g.steer-1.d-1 from blood meal and corn gluten meal). Steers grazed a non-irrigated corn residue field (2.58 steers/ha) for 57 d and were watered and individually supplemented overnight. Diet quality and composition was estimated by collecting extrusa samples twice each week using four esophageally fistulated steers. No effects of alfalfa protein content or level (P > .05) were reflected in gain: 357, 309, and 326 g.steer-1.d-1 for steers receiving MPL, MPH, or HPL alfalfa treatments, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Group selection for adaptation to multiple-hen cages: humoral immune response

A selected line of White Leghorns, which has shown improved survivability and reduced feather loss in large multiple-hen cages, was evaluated for humoral immune response to SRBC under both stressed and unstressed conditions. Three lines of chickens (selected, control, and commercial) were housed in either single- (1 hen) or multiple-hen cages (12 hens, social competition) and subjected to a cold ambient temperature (0 C) at 33 wk of age and to two heating episodes (38 C) at 44 wk of age. Each hen was challenged intravenously with 1 mL of a 7% saline suspension of SRBC at the time that cold exposure was initiated. Hens subjected to high ambient temperatures had been exposed previously to a cold temperature, but were not challenged with SRBC until 16 to 18 h following the end of the second heating episode. Exposure to cold caused immunosuppression in single-caged hens, but not in hens in colony cages. Single- vs colony-caged hens of the control environment challenged with SRBC at 33 wk of age had similar primary hemagglutinin responses to SRBC. Hens subjected to heat experienced immunosuppression at 9 and 12 d following challenge to SRBC when compared to the controls. Hens of multiple-bird cages challenged with antigen at 44 wk of age had a significantly lower hemagglutinin response to SRBC than those reared in single-bird cages. The three lines of genetic stock had similar primary hemagglutinin responses to SRBC; the interactions of genetic stock with cage size or environmental temperature were not significant. It was concluded that genetically selecting hens for survival in multiple-hen cages did not affect their humoral immune response to SRBC.     

Effect of hemoglobin status on humoral immune response of weanling pigs differing in coping styles

The effects of hemoglobin (Hb) status and coping style of pigs on performance and humoral immune response were studied. Twenty-four, 4-wk-old crossbred barrows were assigned to groups of three pigs based on weight and litter origin. Groups were allotted according to a 2 x 2 factorial treatment arrangement: two blood Hb concentration classes (low vs high) and two immunization procedures (control vs immunized). Immunized pigs received an antigen cocktail containing keyhole limpet hemocyanin (KLH), ovalbumin (OA), and tetanus toxoid (TT) at weaning. Additionally, pigs were stratified according to behavioral coping style in response to exposure to a stressor. During 41 d after weaning (approximate time of immunization), blood Hb concentration, ADG, and ADFI were measured weekly and serum antibody titers to KLH, OA, and TT twice weekly. Average Hb concentration differed between low and high Hb pigs (P < .001; 10.0 vs 12.0 g/dL), but this difference declined with time after weaning. Neither immunization procedure nor coping style affected Hb concentrations. In addition, ADG and ADFI were unaffected by any of the treatments. However, ADG was slightly greater in high Hb status pigs (586 vs 633 g/d) and was paralleled by a slightly greater ADFI in high Hb status pigs (812 vs 899 g/d). Antibody responses were negatively or not related to Hb status at weaning. Antibody responses (depending on isotype and antigen) were or tended to be lower in pigs with high blood Hb concentrations. Behavioral coping style strongly affected humoral immune responsiveness; enhanced or accelerated antibody responses were found in pigs that had a passive coping style.     

Increased immune response elicited by DNA vaccination with a synthetic gp120 sequence with optimized codon usage

DNA vaccination elicits humoral and cellular immune responses and has been shown to confer protection against several viral, bacterial, and parasitic pathogens. Here we report that optimized codon usage of an injected DNA sequence considerably increases both humoral and cellular immune responses. We recently generated a synthetic human immunodeficiency virus type 1 gp120 sequence in which most wild-type codons were replaced with codons from highly expressed human genes (syngp120). In vitro expression of syngp120 is considerably increased in comparison to that of the respective wild-type sequence. In BALB/c mice, DNA immunization with syngp120 resulted in significantly increased antibody titers and cytotoxic T-lymphocyte reactivity, suggesting a direct correlation between expression levels and the immune response. Moreover, syngp120 is characterized by rev-independent expression and a low risk of recombination with viral sequences. Thus, synthetic genes with optimized codon usage represent a novel strategy to increase the efficacy and safety of DNA vaccination.     

Forage characteristics, steer performance, and water quality from bermudagrass pastures fertilized with two levels of nitrogen from swine lagoon effluent

Four .8-ha pastures of bermudagrass (Cynodon dactylon [L.] Pers.) were fertilized with either 456 or 873 kg/ha of nitrogen (N) from swine lagoon effluent (two replicates per treatment) and grazed by steers over two summers. Within each pasture, steers received forage only, an energy source (corn), a mixture of corn and soybean meal, or a mixture of corn and blood meal via electronic Calan feeders. All supplements were offered at a level of 1.36 kg/d, and the soybean meal and blood meal supplements provided similar among quantities of protein. Weight gains were similar among supplemented steers, but supplemented steers gained faster (P < .05) than controls. Nitrogen fertilization level had no effect on steer gains, steer grazing days per hectare, or in vitro dry matter disappearance, NDF, and ADF of clipped forage samples. Plant protein and nitrate ion concentrations were greater (P < .06) in clipped forage samples receiving the higher N application rate. Nitrate ion concentrations were greater in available forage samples from the pastures with the high N application rate. Mean total N and nitrate N concentrations were similar in water samples obtained from monitoring wells for the two N treatments over the 2 yr and there were no year x N interactions. Chloride concentrations were greater (P < .05) and pH and specific conductance were less in water samples collected from the 873 kg than from the 456 kg/ha N treatment. Long-term studies are needed to examine the possible cumulative effects of applying various levels of swine waste to the same land area.     

Intranasal immunization confers protection against murine Pneumocystis carinii lung infection

To evaluate the feasibility of mucosal immunization against Pneumocystis carinii (Pc) experimental infection, female BALB/c mice were intranasally immunized three times with soluble Pc antigens plus cholera toxin fraction B (Pc-CTB); control groups received either Pc antigen, CTB, or phosphate-buffered saline (PBS) alone. Two weeks after the last immunization, five animals from each group were sacrificed, and cellular and humoral immune responses were evaluated. The remaining five mice were CD4 depleted using a monoclonal antibody against mouse CD4 and inoculated with viable Pc. Significantly higher specific lymphoproliferative responses from tracheobronchial lymph node cells, immunoglobulin M (IgM) and IgG antibody levels in serum, and bronchoalveolar lavage (BAL)-derived IgA antibody concentrations were observed in the Pc-CTB group of mice relative to control groups (P < 0.01). Five weeks after challenge, no Pc organisms were observed in the lung smears of the Pc-CTB group, while the animals receiving antigen, adjuvant, or PBS had progressively higher numbers of Pc microorganisms. By Western blot analysis, a strongly reactive 55- to 60-kDa antigen was recognized by BAL IgA and by serum IgG. In summary, mucosal immunization elicited specific cellular and humoral immune responses and protected against Pc lung infection after immunosuppression.     

Cytokines as adjuvants for ruminant vaccines

Successful vaccination against any potential pathogen is critically dependent on inducing an appropriate immune response. The pivotal role of cytokines in the immune response to vaccination suggests that non-protective responses or responses that exacerbate disease subsequent to infectious challenge may be the result of limiting or preferential production of one or a number of these mediators. This suggests that the use of recombinant cytokines as vaccine adjuvants may offer a mechanism whereby the magnitude and phenotype of the immune response to vaccination can be specifically modified. In mice, recombinant cytokines have been used extensively as therapeutics, while studies describing vaccine adjuvant activity are more limited. Recombinant (r) interleukin (IL)-1, IL-2 and interferon (IFN) gamma have been used primarily to enhance humoral responses with enhanced protection assessed where appropriate. Cytokine adjuvant studies in ruminants have been restricted to recombinant ovine (rov) and bovine (rbov) IL-1 and IL-2. In sheep, their application has been optimised with respect to dose, route of delivery and formulation, for induction of humoral and cell mediated immunity (DTH and cytotoxicity) to the model protein antigen (Ag) avidin. The level of adjuvant activity of IL-1 in particular compares favourably to that of a variety of other traditional and new chemical adjuvants and detailed analysis has indicated no adverse local or systemic side-effects. Recent studies in our laboratory demonstrating the effectiveness of rovIL-1 as an adjuvant in single and multi-component bacterial toxoid vaccines, and studies from other laboratories demonstrating the application of rbovIL-1 as an adjuvant for the response in cattle to live attenuated viral vaccines, suggest that rIL-1 may become the adjuvant of choice for diseases where protection is mediated by high levels of circulating antibody (Ab). With respect to helminth parasites, IL-1 may prove useful as a component of vaccines based on "hidden Ags" which rely on induction of Ab. Based on analysis in mouse models of helminth infection, other cytokines such as IL-4 and IL-10 may be appropriate for vaccines based on induction of mechanisms involved in natural immunity.