246
J. J. GIAMBRONE, W. I. ANDERSON, W. M. REID AND C. S. EIDSON
Hammond, D. M., and P. L. Long, Ed. University Park Press, Baltimore, Maryland, pp. 295-343. Winterfield, R. W., 1969. Immunity response to the infectious bursal agent. Avian Dis. 13: 548-557.
Wyeth, P. J., 1975. Effect of infectious bursal disease on the response of chickens to S. typhimurium and E. coli infections. Vet. Rec. 96: 238-243.
The Effect of Pokeberry Ingestion on Immune Response in Turkeys R. C. CATTLEY AND B. D. BARNETT
(Received for publication June 25, 1976)
ABSTRACT Two experiments were conducted to determine the effect of pokeberry ingestion on immune response in turkeys. In the first experiment, addition of pokeberries to a control diet at levels of 2.5% or 5.0% had no significant effect on feed consumption or production of sheep's red blood cell (SRBC) antibodies. In the second experiment, drenching with a pokeberry suspension had no significant effect on production of SRBC antibodies. POULTRY SCIENCE 56: 246-248, 1977
INTRODUCTION
MATERIALS AND METHODS
ORTON (1971) and Muenscher (1951) did not consider pokeberries to be toxic to the avian species. Barnett (1975), however, reported that young turkey poults exhibited mortality, reduced gains and lesions including abnormal hocks, ascites, and enlargement of the gall bladder with excess fluid when fed diets containing pokeberries. Ingestion of pokeberries by humans has caused transformation of lymphocytes into plasma cells (Barker et al., 1966, 1967). The appearance of dividing lymphocytes has been attributed to the mitogenic property of pokeweed. Hardin and Arena (1974) have interpreted hematologic abberation by pokeweed as being potentially immunosuppressive. The purpose of this study was to determine whether or not ingestion of pokeberries has any effect on the immune response in turkeys.
Two trials were conducted. In both trials SRBC (sheep's red blood cell) was the test antigen. The sheep's blood was collected in vacutainers with potassium oxalate for the anticoagulant. The cells were washed by centrifugation (1200 r.p.m. for 25 minutes) three times in 0.85% saline before suspending the cells at the desired concentration in 0.85% saline. Turkeys were inoculated with SRBC suspension. After allowing time for antibody production, serum was harvested and tested for SRBC antibodies. Determination of SRBC antibody was done by agglutination test. Frozen serum was thawed and heat inactivated at 56° C. to destroy complement. Dilutions of 1:5, 1:10, 1:20, 1:40, 1:80, 1:160, 1:320, 1:640, and 1:1280 were prepared in 0.5 ml. quantities with 0.85% saline, and 0.5 ml. of a 2% SRBC suspension was added to each. The final dilutions were twice those above (1:10, 1:20, etc.). Results were read as the highest dilution showing visible agglutination. Saline and
M
Published with the approval of the Director of the South Carolina Agricultural Experiment Station as Technical Contribution No. 1372.
Downloaded from http://ps.oxfordjournals.org/ at Carleton University on May 4, 2015
Poultry Science Department, Clemson University, Clemson, South Carolina 29631
247
POKEBERRIES AND IMMUNE RESPONSE
TABLE 1.—Control diet Percentage Ingredient
(%)
Ground yellow corn Soybean meal (49% protein) Coastal bermudagrass Dried whey Animal fat Ground limestone Defluorinated rock phosphate Turkey starter premix* Carbosep Salt
64.35 29.00 1.00 2.00 .50 .50 2.00 .25 .10 .30
consumption was determined. Two days after being placed on experimental rations, the turkeys were inoculated intravenously with 0.7 ml. of a 10% SRBC suspension. Blood was collected at 8 and 15 days post-inoculation. Serum was harvested and frozen at -15° C , until antibody titers were determined.
TABLE 2.—Effect
of adding pokeberries to a control diet on antibody titer to SRBC at 8 and 15 days post-inoculation and on feed consumption in young turkeys
Treatment* Control diet Control diet + 2.5% pokeberries Control diet + 5.0% pokeberries
Mean antibody titer (log 2 ) 15 days postinoculation
Mean feed consumption (grams/day)
8 days postinoculation
247.9 203.4 215.2
8.5477 7.2389 7.9219
*No significant effect (p < .05) due to treatment for any variable.
5.7220 5.5220 5.3599
Downloaded from http://ps.oxfordjournals.org/ at Carleton University on May 4, 2015
Trial 2. Twelve Broad Breasted White turkeys, 100 days of age, were placed in cages *Provides the following per kilogram of diet: Vitamin A, 8811 I.U.; vitamin D 3 , 3304 I.U.: and randomly assigned to two treatments, vitamin E, 9.9 I.U.; vitamin B 12 ,8.8 meg.; ribofla- six turkeys to a treatment. The experimental vin, 5.5 mg.; niacin, 55 mg.; pantothenic acid, 13.2 mg.; choline, 716 mg.; menadione sodium bisulfite, treatment was to drench the turkeys once 2.2 mg.; folacin, 1.1 mg.; ethoxyquin, 62.4 mg.; daily for eight consecutive days with 55 ml. biotin,. 11 mg.; pyridoxine HCL, 2.2 mg.; thiamine, of a pokeberry suspension, via esophageal 1.1 mg. cannula. The suspension was made by thawing frozen pokeberries, liquefing them in a non-inoculated serum were shown to exhibit blender and then mixing this material with no agglutination. an equal quantity of tap water. It is estimated All antibody titers were converted to base that the amount of pokeberries consumed was two logarithms, and all data were compiled over twice that consumed by birds on the and analyzed by the analysis of variance 5% diet of trial 1. The control turkeys were (Steel and Torrie, 1960). drenched with tap water at the same time and in the same manner. Feed and water Trial 1. Fifteen Broad Breasted White male were available ad libitum, except that feed turkeys, nine weeks of age, were placed in was withheld for one day prior to bleeding cages and randomly assigned to one of three the turkeys for immune serum. rations, five turkeys to a ration. These rations were: control diet (see Table 1), control diet On the third day of treatment, each turkey plus 2.5% pokeberries, and control diet plus was inoculated intravenously with 1.0 ml. of 5.0% pokeberries. Fresh pokeberries were a 10% SRBC suspension. Blood was collected picked and liquefied in a blender prior to at 7 and 14 days post-inoculation. Serum was adding to the control diet. Addition was at harvested and frozen at —15° C , until antithe expense of the total diet. The feed and body titers were determined. water were available ad libitum, except that RESULTS AND DISCUSSION feed was withheld for one day prior to bleeding the turkeys for immune serum. Feed The results for Trial 1 are summarized in
248
R. C. CATTLEY AND B. D. BARNETT
TABLE 3.—Effect of drenching with pokeberry suspension on antibody titer to SRBC at 7 and 14 days post-inoculation in young turkeys Mean antibody titer (log2) 7 days post- 14 days postinoculation inoculation 10.6552 8.0861
Treatment* Tap water drench Pokeberry suspension drench 10.2527 7.2527 *No significant effect (p < .05) due to treatment for any variable.
REFERENCES Barker, B. E., P. Fames and P. H. LaMarche, 1966. Peripheral blood plasmacytosis following systemic exposure to Phytolacca americana (pokeweed). Pediatrics, 38: 3: 390-493. Barker, B. E., P. Fames and P. H. LaMarche, 1967. Haematological effects of pokeweed. The Lancet, 1: 437. Barnett, B. D., 1975. Toxicity of pokeberries (fruit of Phytolacca americana large) for turkey poults. Poultry Sci. 54: 1215-1217. Hardin, J. W., and J. M. Arena, 1974. Human Poisoning from Native and Cultivated Plants, 2nd edition. Duke University Press, Durham, N.C., Pages 69-73. Morton, J. F., 1971. Plants Poisonous to People in Florida and Other Warm Areas. Hurricane House Publishers, Inc., Miami, Fla. Part I, pages 15-16. Muenscher, W. C. 1951. Poisonous Plants of the United States, 2nd edition. Macmillan Company, New York, N.Y. Part II, pages 73-74. Steel, R. G. D. and J. H. Torrie, 1960. Principles and Procedures of Statistics. McGraw-Hill Book Company, Inc., New York, N.Y. Chapter 7, pages 99-105.
Downloaded from http://ps.oxfordjournals.org/ at Carleton University on May 4, 2015
Table 2. Addition of pokeberries to the diet did not significantly affect (P < .05) feed consumption or antibody titer at 8 or 15 days post-inoculation. The results for Trial 2 are summarized in Table 3. Drenching with pokeberry suspension did not significantly affect (P < .05) antibody titer at 7 or 14 days post-inoculation. Based on feed consumption data from Trial 1, it was estimated that the turkeys receiving the drench in Trial 2 ingested about twice as much pokeberries as those receiving the control diet plus 5.0% pokeberries in Trial 1. None of the turkeys in Trial 1 or 2 reported here exhibited mortality or morbidity like that reported in younger poults by Barnett (1975). Presumably the difference in age is responsible for the lack of effect in the older birds used in these studies. These results do not rule out the possibility that higher levels of pokeberries might cause mortality, morbidity, or impair immune response in 56 or 100 day old turkeys. The lack of adverse effects in these trials suggest that consumption of a
limited quantity of pokeberries by range birds is not likely to interfere with the immune response from vaccines administered under commercial conditions. The marked hematological effects noted in humans by Barker et al. (1966, 1967) from very small quantities of pokeberries had suggested that immune response might be an unusually sensitive measure of pokeberry toxicity. Results reported here indicate this is not true in the turkey at least insofar as sheep's red blood cell antibodies are a measure of immune response.
J. J. GIAMBRONE, W. I. ANDERSON, W. M. REID AND C. S. EIDSON
Hammond, D. M., and P. L. Long, Ed. University Park Press, Baltimore, Maryland, pp. 295-343. Winterfield, R. W., 1969. Immunity response to the infectious bursal agent. Avian Dis. 13: 548-557.
Wyeth, P. J., 1975. Effect of infectious bursal disease on the response of chickens to S. typhimurium and E. coli infections. Vet. Rec. 96: 238-243.
The Effect of Pokeberry Ingestion on Immune Response in Turkeys R. C. CATTLEY AND B. D. BARNETT
(Received for publication June 25, 1976)
ABSTRACT Two experiments were conducted to determine the effect of pokeberry ingestion on immune response in turkeys. In the first experiment, addition of pokeberries to a control diet at levels of 2.5% or 5.0% had no significant effect on feed consumption or production of sheep's red blood cell (SRBC) antibodies. In the second experiment, drenching with a pokeberry suspension had no significant effect on production of SRBC antibodies. POULTRY SCIENCE 56: 246-248, 1977
INTRODUCTION
MATERIALS AND METHODS
ORTON (1971) and Muenscher (1951) did not consider pokeberries to be toxic to the avian species. Barnett (1975), however, reported that young turkey poults exhibited mortality, reduced gains and lesions including abnormal hocks, ascites, and enlargement of the gall bladder with excess fluid when fed diets containing pokeberries. Ingestion of pokeberries by humans has caused transformation of lymphocytes into plasma cells (Barker et al., 1966, 1967). The appearance of dividing lymphocytes has been attributed to the mitogenic property of pokeweed. Hardin and Arena (1974) have interpreted hematologic abberation by pokeweed as being potentially immunosuppressive. The purpose of this study was to determine whether or not ingestion of pokeberries has any effect on the immune response in turkeys.
Two trials were conducted. In both trials SRBC (sheep's red blood cell) was the test antigen. The sheep's blood was collected in vacutainers with potassium oxalate for the anticoagulant. The cells were washed by centrifugation (1200 r.p.m. for 25 minutes) three times in 0.85% saline before suspending the cells at the desired concentration in 0.85% saline. Turkeys were inoculated with SRBC suspension. After allowing time for antibody production, serum was harvested and tested for SRBC antibodies. Determination of SRBC antibody was done by agglutination test. Frozen serum was thawed and heat inactivated at 56° C. to destroy complement. Dilutions of 1:5, 1:10, 1:20, 1:40, 1:80, 1:160, 1:320, 1:640, and 1:1280 were prepared in 0.5 ml. quantities with 0.85% saline, and 0.5 ml. of a 2% SRBC suspension was added to each. The final dilutions were twice those above (1:10, 1:20, etc.). Results were read as the highest dilution showing visible agglutination. Saline and
M
Published with the approval of the Director of the South Carolina Agricultural Experiment Station as Technical Contribution No. 1372.
Downloaded from http://ps.oxfordjournals.org/ at Carleton University on May 4, 2015
Poultry Science Department, Clemson University, Clemson, South Carolina 29631
247
POKEBERRIES AND IMMUNE RESPONSE
TABLE 1.—Control diet Percentage Ingredient
(%)
Ground yellow corn Soybean meal (49% protein) Coastal bermudagrass Dried whey Animal fat Ground limestone Defluorinated rock phosphate Turkey starter premix* Carbosep Salt
64.35 29.00 1.00 2.00 .50 .50 2.00 .25 .10 .30
consumption was determined. Two days after being placed on experimental rations, the turkeys were inoculated intravenously with 0.7 ml. of a 10% SRBC suspension. Blood was collected at 8 and 15 days post-inoculation. Serum was harvested and frozen at -15° C , until antibody titers were determined.
TABLE 2.—Effect
of adding pokeberries to a control diet on antibody titer to SRBC at 8 and 15 days post-inoculation and on feed consumption in young turkeys
Treatment* Control diet Control diet + 2.5% pokeberries Control diet + 5.0% pokeberries
Mean antibody titer (log 2 ) 15 days postinoculation
Mean feed consumption (grams/day)
8 days postinoculation
247.9 203.4 215.2
8.5477 7.2389 7.9219
*No significant effect (p < .05) due to treatment for any variable.
5.7220 5.5220 5.3599
Downloaded from http://ps.oxfordjournals.org/ at Carleton University on May 4, 2015
Trial 2. Twelve Broad Breasted White turkeys, 100 days of age, were placed in cages *Provides the following per kilogram of diet: Vitamin A, 8811 I.U.; vitamin D 3 , 3304 I.U.: and randomly assigned to two treatments, vitamin E, 9.9 I.U.; vitamin B 12 ,8.8 meg.; ribofla- six turkeys to a treatment. The experimental vin, 5.5 mg.; niacin, 55 mg.; pantothenic acid, 13.2 mg.; choline, 716 mg.; menadione sodium bisulfite, treatment was to drench the turkeys once 2.2 mg.; folacin, 1.1 mg.; ethoxyquin, 62.4 mg.; daily for eight consecutive days with 55 ml. biotin,. 11 mg.; pyridoxine HCL, 2.2 mg.; thiamine, of a pokeberry suspension, via esophageal 1.1 mg. cannula. The suspension was made by thawing frozen pokeberries, liquefing them in a non-inoculated serum were shown to exhibit blender and then mixing this material with no agglutination. an equal quantity of tap water. It is estimated All antibody titers were converted to base that the amount of pokeberries consumed was two logarithms, and all data were compiled over twice that consumed by birds on the and analyzed by the analysis of variance 5% diet of trial 1. The control turkeys were (Steel and Torrie, 1960). drenched with tap water at the same time and in the same manner. Feed and water Trial 1. Fifteen Broad Breasted White male were available ad libitum, except that feed turkeys, nine weeks of age, were placed in was withheld for one day prior to bleeding cages and randomly assigned to one of three the turkeys for immune serum. rations, five turkeys to a ration. These rations were: control diet (see Table 1), control diet On the third day of treatment, each turkey plus 2.5% pokeberries, and control diet plus was inoculated intravenously with 1.0 ml. of 5.0% pokeberries. Fresh pokeberries were a 10% SRBC suspension. Blood was collected picked and liquefied in a blender prior to at 7 and 14 days post-inoculation. Serum was adding to the control diet. Addition was at harvested and frozen at —15° C , until antithe expense of the total diet. The feed and body titers were determined. water were available ad libitum, except that RESULTS AND DISCUSSION feed was withheld for one day prior to bleeding the turkeys for immune serum. Feed The results for Trial 1 are summarized in
248
R. C. CATTLEY AND B. D. BARNETT
TABLE 3.—Effect of drenching with pokeberry suspension on antibody titer to SRBC at 7 and 14 days post-inoculation in young turkeys Mean antibody titer (log2) 7 days post- 14 days postinoculation inoculation 10.6552 8.0861
Treatment* Tap water drench Pokeberry suspension drench 10.2527 7.2527 *No significant effect (p < .05) due to treatment for any variable.
REFERENCES Barker, B. E., P. Fames and P. H. LaMarche, 1966. Peripheral blood plasmacytosis following systemic exposure to Phytolacca americana (pokeweed). Pediatrics, 38: 3: 390-493. Barker, B. E., P. Fames and P. H. LaMarche, 1967. Haematological effects of pokeweed. The Lancet, 1: 437. Barnett, B. D., 1975. Toxicity of pokeberries (fruit of Phytolacca americana large) for turkey poults. Poultry Sci. 54: 1215-1217. Hardin, J. W., and J. M. Arena, 1974. Human Poisoning from Native and Cultivated Plants, 2nd edition. Duke University Press, Durham, N.C., Pages 69-73. Morton, J. F., 1971. Plants Poisonous to People in Florida and Other Warm Areas. Hurricane House Publishers, Inc., Miami, Fla. Part I, pages 15-16. Muenscher, W. C. 1951. Poisonous Plants of the United States, 2nd edition. Macmillan Company, New York, N.Y. Part II, pages 73-74. Steel, R. G. D. and J. H. Torrie, 1960. Principles and Procedures of Statistics. McGraw-Hill Book Company, Inc., New York, N.Y. Chapter 7, pages 99-105.
Downloaded from http://ps.oxfordjournals.org/ at Carleton University on May 4, 2015
Table 2. Addition of pokeberries to the diet did not significantly affect (P < .05) feed consumption or antibody titer at 8 or 15 days post-inoculation. The results for Trial 2 are summarized in Table 3. Drenching with pokeberry suspension did not significantly affect (P < .05) antibody titer at 7 or 14 days post-inoculation. Based on feed consumption data from Trial 1, it was estimated that the turkeys receiving the drench in Trial 2 ingested about twice as much pokeberries as those receiving the control diet plus 5.0% pokeberries in Trial 1. None of the turkeys in Trial 1 or 2 reported here exhibited mortality or morbidity like that reported in younger poults by Barnett (1975). Presumably the difference in age is responsible for the lack of effect in the older birds used in these studies. These results do not rule out the possibility that higher levels of pokeberries might cause mortality, morbidity, or impair immune response in 56 or 100 day old turkeys. The lack of adverse effects in these trials suggest that consumption of a
limited quantity of pokeberries by range birds is not likely to interfere with the immune response from vaccines administered under commercial conditions. The marked hematological effects noted in humans by Barker et al. (1966, 1967) from very small quantities of pokeberries had suggested that immune response might be an unusually sensitive measure of pokeberry toxicity. Results reported here indicate this is not true in the turkey at least insofar as sheep's red blood cell antibodies are a measure of immune response.
