- Email: [email protected]
Incorporation of H3-Glucose into the Parasite Cells of Eimeria tenella
Incorporation of H3 -Glucose into the Parasite Cells of Eimeria tenella TOSH1AKI MATSUZAWA1 Sankyo Company Limited, The Central Research Laboratories, 1-2-58, Iliromachi, Shinagawa, Tokyo, Japan (Received for publication January 2, 1979)
1979 Poultry Science 58:1007-1008
Ryley et al. (1969) employing biochemical and electron microscopic techniques, demonstrated the presence of amylopectin in sporozoites, schizont, macrogametes, and oocysts of Eimeria tenella and E. brunetti. The source of amylopectin in the parasite cells is not yet known. However, the glucose reductions in blood and tissues have been reported in chicks infested with E. tenella (Pratt, 1940, 1941; Edgar et al, 1944; Daugherty and Herrick, 1952; Gill and Ray, 1954; and Freeman, 1970). It is suggested that glucose is required for growth by E. tenella. The purpose of the present study was to incorporate tritiated-glucose (H -glucose) into the parasite cells of E. tenella. Two-week-old White Leghorn male chicks were inoculated with 500,000 sporulated oocysts of E. tenella. Those chicks ceca were ligated under ether anaethesia on 48 and 96 hr after inoculation and .5 ml of a solution containing .8 Mg H 3 -glucose (specific activity .8 juCi/mg) in .1 M phosphate buffer pH 6.0 was injected into each cecum. Thirty minutes later, chicks were killed and ligated ceca were removed. The cecal mucous tissue was sectioned. Tissues were fixed for 2 hr in a 6.25% glutaraldehyde solution with .1 M phosphate buffer at pH 7.4. The tissues were washed with .1 M phosphate and post-fixed in .1 M phos-
phate buffered osmium at pH 7.4 for an hour. The tissues were dehydrated by increasing concentrations of ethanol and were embedded in epon. Sections were cut with glass knives on a Porter-Blum MT I microtome and mounted on naked copper grids. Electron microscopic autoradiography was performed according to the method of Mizuhira (1971). The ultrathin section was stained with lead citrate for 5 min followed by saturation in uranyl acetate for 30 min. The sections were examined in a Hitachi HU-11 DS electron microscope. The results are shown in Figures 1 and 2. Silver grains of H -glucose were located over the nucleus (N), nucleolus (NC), mitochondria (M), and endoplasmic reticulum (E) of the first schizont, but were not observed over the
4$f -
M ) %
'Present address: Yamanouchi Pharmaceutical Company Limited, Institute of Development and Research, Toxicology and Safety Research, 1-1-8, Azusawa, Itabashi, Tokyo 174, Japan.
-'i*>
•* '/.
:•
t£*Sa0* •* '^*'-J^^':.'''^S^N«i FIG. 1. First generation schizont of/;', tenella. The silver grains (t) of H3-glucose are located over the nucleous (N), nucleolus (Nu), mitochondria (M), and endoplasmic reticulum (E) of first schizont.
1007
Downloaded from http://ps.oxfordjournals.org/ at Univ of Iowa-Law Library on May 26, 2015
ABSTRACT The incorporation of H3-glucose into the parasite cells of Eimeria tenella was observed with electron microscopic autoradiography. H3-glucose was incorporated into the nucleus, nucleolus, mitochondria, and endoplasmic reticulum of the first schizont, but was not incorporated into the nucleus and nucleolus of young macrogametocyte.
1008
MATSUZAWA clear division at this stage ceases and does not require exogenous glucose. REFERENCES
nucleus and nucleolus of young macrogametcytes. It is assumed that no silver grains of H -glucose in the nucleus and nucleolus of macrogametocytes were present because nu-
Downloaded from http://ps.oxfordjournals.org/ at Univ of Iowa-Law Library on May 26, 2015
FIG. 2. Macrogametocyte of E. tenella. The silver grains (t) of H3-glucose are located over the nucleous (N), nucleolus (Nu), mitochondria (M), and endoplasmic reticulum (E) of young macrogametocyte.
Daugherty, J. W., and Herrick, C. A., 1952. Cecal coccidiosis and carbohydrate metabolism in chicks. J.Parasitol. 38:299-304. Edgar, S. A., C. A. Herrick, and L. A. Fraser, 1944. Glycogen in the life cycle of the coccidium, Eimeria tenella. Trans. Amer. Microscopical Soc. 63:199-202. Freeman, B. M., 1970. Carbohydrate stores in chickens infected with Eimeria tenella. Parasitology 61:245-251. Gill, B. S., and H. N. Ray, 1954. Glycogen and it's probable significance in Eimeria tenella. Railliet and Lucet, 1891. Indian J. Vet. Sci. 26:223-228. Mizuhira, B., 1971. Page 71—80 in Electron microscope (Japanese). Ishiyaku Press, Tokyo. Pratt, I., 1940. The effect of Eimeria tenella (coccidia) upon the blood sugar of the chicken. Trans. Amer. Microscopical Soc. 59:31 — 37. Pratt, I., 1941. The effect of Eimeria tenella (coccidia) upon the glycogen stores of the chicken. Amer. J. Hyg. 3 4 : 5 4 - 6 1 . Ryley, J. F., M. Bentley, D. J. Manners, and J. R. Stark, 1969. Amylopectin, the storage polysaccharide of the coccidia Eimeria brunetti and E. tenella. J. Parasitol. 55:839-845.
1979 Poultry Science 58:1007-1008
Ryley et al. (1969) employing biochemical and electron microscopic techniques, demonstrated the presence of amylopectin in sporozoites, schizont, macrogametes, and oocysts of Eimeria tenella and E. brunetti. The source of amylopectin in the parasite cells is not yet known. However, the glucose reductions in blood and tissues have been reported in chicks infested with E. tenella (Pratt, 1940, 1941; Edgar et al, 1944; Daugherty and Herrick, 1952; Gill and Ray, 1954; and Freeman, 1970). It is suggested that glucose is required for growth by E. tenella. The purpose of the present study was to incorporate tritiated-glucose (H -glucose) into the parasite cells of E. tenella. Two-week-old White Leghorn male chicks were inoculated with 500,000 sporulated oocysts of E. tenella. Those chicks ceca were ligated under ether anaethesia on 48 and 96 hr after inoculation and .5 ml of a solution containing .8 Mg H 3 -glucose (specific activity .8 juCi/mg) in .1 M phosphate buffer pH 6.0 was injected into each cecum. Thirty minutes later, chicks were killed and ligated ceca were removed. The cecal mucous tissue was sectioned. Tissues were fixed for 2 hr in a 6.25% glutaraldehyde solution with .1 M phosphate buffer at pH 7.4. The tissues were washed with .1 M phosphate and post-fixed in .1 M phos-
phate buffered osmium at pH 7.4 for an hour. The tissues were dehydrated by increasing concentrations of ethanol and were embedded in epon. Sections were cut with glass knives on a Porter-Blum MT I microtome and mounted on naked copper grids. Electron microscopic autoradiography was performed according to the method of Mizuhira (1971). The ultrathin section was stained with lead citrate for 5 min followed by saturation in uranyl acetate for 30 min. The sections were examined in a Hitachi HU-11 DS electron microscope. The results are shown in Figures 1 and 2. Silver grains of H -glucose were located over the nucleus (N), nucleolus (NC), mitochondria (M), and endoplasmic reticulum (E) of the first schizont, but were not observed over the
4$f -
M ) %
'Present address: Yamanouchi Pharmaceutical Company Limited, Institute of Development and Research, Toxicology and Safety Research, 1-1-8, Azusawa, Itabashi, Tokyo 174, Japan.
-'i*>
•* '/.
:•
t£*Sa0* •* '^*'-J^^':.'''^S^N«i FIG. 1. First generation schizont of/;', tenella. The silver grains (t) of H3-glucose are located over the nucleous (N), nucleolus (Nu), mitochondria (M), and endoplasmic reticulum (E) of first schizont.
1007
Downloaded from http://ps.oxfordjournals.org/ at Univ of Iowa-Law Library on May 26, 2015
ABSTRACT The incorporation of H3-glucose into the parasite cells of Eimeria tenella was observed with electron microscopic autoradiography. H3-glucose was incorporated into the nucleus, nucleolus, mitochondria, and endoplasmic reticulum of the first schizont, but was not incorporated into the nucleus and nucleolus of young macrogametocyte.
1008
MATSUZAWA clear division at this stage ceases and does not require exogenous glucose. REFERENCES
nucleus and nucleolus of young macrogametcytes. It is assumed that no silver grains of H -glucose in the nucleus and nucleolus of macrogametocytes were present because nu-
Downloaded from http://ps.oxfordjournals.org/ at Univ of Iowa-Law Library on May 26, 2015
FIG. 2. Macrogametocyte of E. tenella. The silver grains (t) of H3-glucose are located over the nucleous (N), nucleolus (Nu), mitochondria (M), and endoplasmic reticulum (E) of young macrogametocyte.
Daugherty, J. W., and Herrick, C. A., 1952. Cecal coccidiosis and carbohydrate metabolism in chicks. J.Parasitol. 38:299-304. Edgar, S. A., C. A. Herrick, and L. A. Fraser, 1944. Glycogen in the life cycle of the coccidium, Eimeria tenella. Trans. Amer. Microscopical Soc. 63:199-202. Freeman, B. M., 1970. Carbohydrate stores in chickens infected with Eimeria tenella. Parasitology 61:245-251. Gill, B. S., and H. N. Ray, 1954. Glycogen and it's probable significance in Eimeria tenella. Railliet and Lucet, 1891. Indian J. Vet. Sci. 26:223-228. Mizuhira, B., 1971. Page 71—80 in Electron microscope (Japanese). Ishiyaku Press, Tokyo. Pratt, I., 1940. The effect of Eimeria tenella (coccidia) upon the blood sugar of the chicken. Trans. Amer. Microscopical Soc. 59:31 — 37. Pratt, I., 1941. The effect of Eimeria tenella (coccidia) upon the glycogen stores of the chicken. Amer. J. Hyg. 3 4 : 5 4 - 6 1 . Ryley, J. F., M. Bentley, D. J. Manners, and J. R. Stark, 1969. Amylopectin, the storage polysaccharide of the coccidia Eimeria brunetti and E. tenella. J. Parasitol. 55:839-845.