J. Comp.
Path.
1992 Vol.
Neosporosis
107, 231-237
in the Aborted
Fetus
and Newborn
Calf
H. Ogino, E. Watanabe *, S. Watanabe?, H. Agawat, M. NaritaS, M. Haritani$ and K. Kawashimat Chuo Livestock Hygiene Service Center, Nishikanbara, Niigata, *Chuo Livestock Hygiene Service Center, Jyoyo, Kyoto, THigashihiroshima Livestock Hygiene Service Center, Higashihiroshima, Hiroshima and iNational Institute of Animal Health, Kannondai, Tsukuba, Ibaraki, 305 Japan
Summary Protozoa1 encephalitic lesions were found in four aborted fetuses and one dead newborn calf. The organism was identified as Neospora caninum by immunoperoxidase. The brain lesions were of two forms. One was observed in three fetuses of 5 months gestation and was characterized by multifocal necrosis. The other was found in a 7-month fetus and in a newborn calf, and showed severe infiltration with macrophages and plasma cells containing IgG. This association, between the age of fetus and inflammation, may reflect development of the immune system in bovine fetuses.
Introduction
Numerous sporadic cases of bovine abortion owing to congenital disease are reported in the literature. Bovine abortion caused by protozoa1 fetal infections such as Sarcocystis, Hammondia and Toxoplasma is almost unrecorded, perhaps because of the unavailability of freshly collected fetal or maternal tissue or a paucity
of parasite
stages
in tissues
examined.
Neosporacaninumis a newly described
protozoan parasite that can cause fetal disease in dogs (Dubey, Carpenter, Speer, Topper and Uggla, 1988a; Dubey, Hattel, Lindsay and Topper, 198813; Dubey and Lindsay, 1989; Umemura, Shiraki, Morita, Shimada, Haritani, Kobayashi and Yamagata, 1992) and other animals (Dubey, Lindsay and Lipscomb, 1990a), including cattle (Barr, Anderson, Bianchard, Daft, Kinde and Conrad, 1990; Barr, Anderson, Dubey and Conrad, 1991a; Barr, Conrad, Dubey and Anderson, 1991 b; Dubey, Leathers and Lindsay, 1989; Dubey, Miller, Lindsay and Topper, 1990b; Shivaprasad, Ely and Dubey, 1989). The organism causes myositis and encephalomyelitis, but neither its life cycle nor the source of infection is known. The purpose of the present report is to describe the brain lesions in four aborted bovine fetuses of different gestational ages and in one newborn dead calf in which N. caninumwas probably the direct cause of death. In addition, the relationship between the N. caninumantigen and immunoglobulin-containing cells was investigated by an immunoperoxidase technique. 0021-9975/92/060231+07
008.00/O
0
1992 Academic
Press Limited
232
H. Ogino
Materials
et al.
and Methods
A total of 163 brain tissue samples, 115 collected from aborted fetuses and 48 from a slaughter house, and one calf which died 1 day after birth, were fixed in 10 per cent neutral buffered formalin, stained with haematoxylin and eosin (HE) and the periodic acid-Schiff (PAS) method. N. cuninum antigen and immunoglobulin (IgG and IgM)-containing cells were demonstrated by an avidin-biotin-complex (ABC) immunoperoxidase method with a kit as described previously (Lindsay and Dubey, 1989). Anti-N caninum rabbit serum, supplied by Dr J. P. Dubey, University of California, U.S.A., anti-Toxoplasma go&ii and anti-Hammondia hammondi rabbit sera, supplied by Dr K. Shimura, National Institute of Animal Health, Japan, anti-bovine IgG (Fc) (EY Lab.) and anti-bovine IgM (p) (Miles Lab.) rabbit sera were used at a dilution of 1 in 1000, 1 in 2048, 1 in 8192, 1 in 1000 and 1 in 2000, respectively. Sections were counterstained with methyl green. Serum from a non-immunized rabbit was used as the control.
Results Brain tissue from three fetuses, Nos. 1, 2 and 3, aborted at 5 months of gestation, from fetus No. 4, collected from a slaughter house at 7 months of gestation and from one dead newborn calf all had brain lesions associated with protozoa1 infection. Organisms resembling N. caninum were detected in all except fetus 1 (Table 1). Three fetuses, Nos. 1, 2 and 3, had multifocal areas of necrosis ranging from 100 pm to 1 mm in diameter. These foci were characterized, histologically, by disruption and pallor of the neuropil and showed pyknotic and karyorrhectic nuclear debris (Fig. 1). They were distributed in the white and grey matter of the brain and spinal cord. A small number of protozoa1 tachyzoites were found in association with these foci, except in fetus 1. In fetus 4 and the newborn calf (5)) the encephalitic lesion was characterized by moderate to severe perivascular infiltration by lymphocytes, macrophages, plasma cells and glial cells (Figs 2 and 3). Tachyzoites were commonly found in the peripheral areas of blood vessels (Fig. 4). In the newborn calf and in fetus 4, tissue cysts were seen primarily in the neurones and were oval or round in shape (Fig. 5). The tissue
Detection
Animal
of histological
lesion,
Gestational months
no.
Histological
Necrosis Fetus Fetus Fetus Fetus Calf
1 2 3 4
5 5 5 7 1 day
Table 1 antigen and immuaoglobulin-containing CNS
N. caninum
++* ++ ++ -
lesion*
Gliosis
N. caninumt ~ PVC
++ ++
Ig-ce11st
.-
__Tachyzoite
-
+++ +++
cells in the
+ ++ ++ ++
PVC = Perivascular cuffing. *: - , no lesion; + , slight; + + , moderate; + + + , severe. t: - , none; + , small number; + + , moderate number; + + + , large number.
Cyst + +
I&
W ++ +++
-
Neosporosis
Large
area of central
necrosis
in the cerebrum
Fig. 2.
Fig.
1.
Severe
inflammatory
reaction
in the spinal
Fig. 3.
Perivascular
cuffing
Fig. 4.
Small
of tachyzoites
group
composed
of plasma
in newborn
in Cattle
in fetus 2. HE cord of newborn
cells in the cerebrum calf. HE x 1000.
233
x
200.
calf. HE
X 160.
of fetus 4. HE x 400.
234
H. Ogino
et al.
cyst wall was up to 2.5 pm thick, homogeneous and enclosed slender bradyzoites (Fig. 6). Tissue cysts were separate from the inflammatory foci and showed no surrounding cellular reaction. Immunohistologically, the protozoa1 structures reacted positively with antiN. caninum serum, but not with anti-Toxoplasma gondii or anti-Hammondia hammondisera. The tachyzoites stained more intensely than did the bradyzoites. The cyst wall was also stained. Free tachyzoites and clusters of tachyzoites were associated with neural degeneration and some were found in blood vessels (Fig. 7). IgG-containing cells were present in the perivascular cuffs and were diffuse in the nodular foci (Fig. 8). They were detected in fetus 4 and in the newborn calf, but not in fetuses 1, 2 and 3. The number of positive cells in the newborn calf was much greater than that in fetus 4. IgM-containing cells were not found. Discussion The results of the present study revealed four aborted fetuses (Nos 1 to 4) and one dead newborn calf that had protozoa1 lesions in the central nervous system. The lesions were of two forms. One form, observed in fetuses 1, 2 and 3, was characterized by focal necrosis (necrotizing encephalitis) and the other, observed in fetus 4 and in a newborn calf, was characterized by moderate to severe infiltration of macrophages and plasma cells. Four of the brain lesions contained the protozoa1 organisms. These findings resembled those described for encephalitic protozoa1 infection in bovine fetuses associated with 3v. caninum-like organisms (Barr et al., 1990, 1991a,b; Dubey et al., 1990b). It is well known that protozoan parasites such as Sarcocystis (Hong, Giles, Newman and Fayer, 1982; Jolley, Jensen, Hancock and Swift, 1983; McCausland, Badman, Hides and Slee, 1984) and T. gondii (Dubey, 1981; Dubey and Schmitz, 1981) cause abortion in cows and produce encephalitic lesions. Sarcocystisis identified by its specific endothelial cell tropism and the presence of protozoa1 clusters in endothelial cells of various tissues. Degenerating ‘7: gondii is difficult to differentiate from degenerating host cells and diagnosis of toxoplasma1 abortion may depend more on recognition of lesions in the placenta than on identification of the parasite in tissue (Dubey, 1981; Dubey and Schmitz, 1981). The encephalitic lesions and structure of the tachyzoites of TT gondii are similar in appearance to those of N. caninum. Therefore, the identification of .N. caninumin the present four cases was by an immunoperoxidase technique and a specific antiserum, as in previous reports (Lindsay and Dubey, 1989; Barr et al., 1991b). The ability of fetuses to develop specific immunoglobulin to a variety of non-pathogenic antigens and congenitally acquired agents has been investigated in several species (Osburn, MacLachlan and Terrell, 1982). In the present study, fetuses at 5 months gestation (Nos 1, 2 and 3) had multifocal necrosis and tachyzoites, but no inflammatory cell reaction, suggesting that central nervous system cells were being destroyed by active multiplication of tachyzoites. However, fetus 4 (7 months) and the newborn calf had a
Neosporosis
Fig. 5.
Tissue cyst in a neurone
Fig. 6.
Tissue cyst with
Fig. 7.
Three
tachyzoites
Fig. 8.
Many
IgG-containing
of newborn
in Cattle
calf. Immunoperoxidase
a thick cyst wall in newborn (arrows)
235
in and around
(IP)
x 1000.
calf. IP x 1000. a small vessel in newborn
cells in the perivascular
cuffing
calf. IP x 1000
of fetus 4. IP X 400.
H. Ogino et al.
236
prominent inflammatory response with many IgG-containing cells and tissue cysts. This apparent association between gestational age and the degree of tissue inflammation may reflect the progressive development of the immune system in bovine fetuses. Acknowledgment The authors
thank
Mrs Y. Ando and T. Fujisawa
for preparing
the photographs.
References Barr, B. C., Anderson, M. L., Bianchard, P. C., Daft, B. M., Kinde, H. and Conrad, P. A. (1990). Bovine fetal encephalitis and myocarditis associated with protozoa1 infection. Veterinary Pathology, 27, 354-36 1. Barr, B. C., Anderson, M. L., Dubey, J. P. and Conrad, P. A. (1991a). Neosporu-like protozoa1 infections associated with bovine abortions. Veterinary Pathology, 28, 110-l 16. Barr, B. C., Conrad, P. A., Dubey, J. P. and Anderson, M. L. (1991 b). JVeosporu-like encephalomyelitis in a calf: Pathology, ultrastructure, and immunoreactivity. Journal of Veterinary Diagnostic Investigation, 3, 3946. Dubey, J. P. (1981). Epizootic Toxoplasmosis associated with abortion in dairy goats in Montana. 3ournal of the American Veterinary Medical Association, 178, 661-670. Dubey, J. P., Carpenter, J. L., Speer, C. A., Topper, M. J. and Uggla, A. (1988a). Newly recognized fatal protozoan disease of dogs. Journal of the American Veterinary Medical Association, 192, 1269-l 285. Dubey, J. P., Hattel, A. L., Lindsay, D. S. and Topper, M. J. (1988b). Neonatal Neospora caninum infection in dogs: Isolation of the causative agent and experimental transmission. Journal of the American Veterinary Medical Association, 193, 1259-1263. Dubey, J. P., Leathers, C. W. and Lindsay, D. S. (1989). Jveosporu cuninum-like protozoon associated with fetal myelitis in newborn calves. Journal of Parasitology, 75, 146-148. Dubey, J. P. and Lindsay, D. S. (1989). Fatal Jveosporu caninum infection in kittens. Journal of Parasitology, 75, 148-l 5 1. Dubey, J. P., Lindsay, D. S. and Lipscomb, T. P. (1990a). Neosporosis in cats. Veterinary Pathology, 27, 335-339. Dubey, J. P., Miller, S., Lindsay, D. S. and Topper, M. J. (1990b). ~eospora caninum-associated myocarditis and encephalitis in an aborted calf. Journal of Veterinary Diagnostic Investigation, 2, 66-69. Dubey, J. P. and Schmitz, J. A. (1981). Abortion associated with Toxoplasmosis in sheep in Oregon, Journal of the American Veterinary Medical Association, 178, 675-678. Hong, C. B., Giles, R. C., Newman, I. E. and Fayer, R. ( 1982). Surcocystosis in an aborted bovine fetus. Journal of the American Veterinary Medical Association, 181, 585-588. Jolley, W. R., Jensen, R., Hancock, H. A. and Swift, B. L. (1983). Encephalitic Sarcocystosis in a newborn calf. American Journal of Veterinary Research, 44, 1908-1911. Lindsay, D. S. and Dubey, J. P. ( 1989). I mmunohistochemical diagnosis of Neospora caninum in tissue section. American Journal of Veterinary Research, 50, 1981-I 983. McCausland, I. P., Badman, R. T., Hides, S. and Slee, K. J. (1984). Multiple apparent Sarcocystis abortion in four bovine herds. Cornell Veterinarian, 74, 146154. Osburn, B. I., MacLachlan, N. J. and Terrell, T. G. (1982). Ontogeny of the immune system. Journal of the American Veterinary Medical Association, 181, 1049-1052. Shivaprasad, H. L., Ely, R. and Dubey, J. P. (1989). A Neosporu-like protozoon found in an aborted bovine placenta. Veterinary Parasitology, 34, 145-148.
Neosporosis
in Cattle
237
Umemura, T., Shiraki, K., Morita, T., Shimada, A., Haritani, M., Kobayashi, M. and Yamagata, S. (1992). N eos p orosis in a dog: The first casereport in Japan. Journal of Veterinary Medical Science, 54, 157-I 59.
1
Received, March 3Oth, 1992 Accepted, June lst, 1992
Path.
1992 Vol.
Neosporosis
107, 231-237
in the Aborted
Fetus
and Newborn
Calf
H. Ogino, E. Watanabe *, S. Watanabe?, H. Agawat, M. NaritaS, M. Haritani$ and K. Kawashimat Chuo Livestock Hygiene Service Center, Nishikanbara, Niigata, *Chuo Livestock Hygiene Service Center, Jyoyo, Kyoto, THigashihiroshima Livestock Hygiene Service Center, Higashihiroshima, Hiroshima and iNational Institute of Animal Health, Kannondai, Tsukuba, Ibaraki, 305 Japan
Summary Protozoa1 encephalitic lesions were found in four aborted fetuses and one dead newborn calf. The organism was identified as Neospora caninum by immunoperoxidase. The brain lesions were of two forms. One was observed in three fetuses of 5 months gestation and was characterized by multifocal necrosis. The other was found in a 7-month fetus and in a newborn calf, and showed severe infiltration with macrophages and plasma cells containing IgG. This association, between the age of fetus and inflammation, may reflect development of the immune system in bovine fetuses.
Introduction
Numerous sporadic cases of bovine abortion owing to congenital disease are reported in the literature. Bovine abortion caused by protozoa1 fetal infections such as Sarcocystis, Hammondia and Toxoplasma is almost unrecorded, perhaps because of the unavailability of freshly collected fetal or maternal tissue or a paucity
of parasite
stages
in tissues
examined.
Neosporacaninumis a newly described
protozoan parasite that can cause fetal disease in dogs (Dubey, Carpenter, Speer, Topper and Uggla, 1988a; Dubey, Hattel, Lindsay and Topper, 198813; Dubey and Lindsay, 1989; Umemura, Shiraki, Morita, Shimada, Haritani, Kobayashi and Yamagata, 1992) and other animals (Dubey, Lindsay and Lipscomb, 1990a), including cattle (Barr, Anderson, Bianchard, Daft, Kinde and Conrad, 1990; Barr, Anderson, Dubey and Conrad, 1991a; Barr, Conrad, Dubey and Anderson, 1991 b; Dubey, Leathers and Lindsay, 1989; Dubey, Miller, Lindsay and Topper, 1990b; Shivaprasad, Ely and Dubey, 1989). The organism causes myositis and encephalomyelitis, but neither its life cycle nor the source of infection is known. The purpose of the present report is to describe the brain lesions in four aborted bovine fetuses of different gestational ages and in one newborn dead calf in which N. caninumwas probably the direct cause of death. In addition, the relationship between the N. caninumantigen and immunoglobulin-containing cells was investigated by an immunoperoxidase technique. 0021-9975/92/060231+07
008.00/O
0
1992 Academic
Press Limited
232
H. Ogino
Materials
et al.
and Methods
A total of 163 brain tissue samples, 115 collected from aborted fetuses and 48 from a slaughter house, and one calf which died 1 day after birth, were fixed in 10 per cent neutral buffered formalin, stained with haematoxylin and eosin (HE) and the periodic acid-Schiff (PAS) method. N. cuninum antigen and immunoglobulin (IgG and IgM)-containing cells were demonstrated by an avidin-biotin-complex (ABC) immunoperoxidase method with a kit as described previously (Lindsay and Dubey, 1989). Anti-N caninum rabbit serum, supplied by Dr J. P. Dubey, University of California, U.S.A., anti-Toxoplasma go&ii and anti-Hammondia hammondi rabbit sera, supplied by Dr K. Shimura, National Institute of Animal Health, Japan, anti-bovine IgG (Fc) (EY Lab.) and anti-bovine IgM (p) (Miles Lab.) rabbit sera were used at a dilution of 1 in 1000, 1 in 2048, 1 in 8192, 1 in 1000 and 1 in 2000, respectively. Sections were counterstained with methyl green. Serum from a non-immunized rabbit was used as the control.
Results Brain tissue from three fetuses, Nos. 1, 2 and 3, aborted at 5 months of gestation, from fetus No. 4, collected from a slaughter house at 7 months of gestation and from one dead newborn calf all had brain lesions associated with protozoa1 infection. Organisms resembling N. caninum were detected in all except fetus 1 (Table 1). Three fetuses, Nos. 1, 2 and 3, had multifocal areas of necrosis ranging from 100 pm to 1 mm in diameter. These foci were characterized, histologically, by disruption and pallor of the neuropil and showed pyknotic and karyorrhectic nuclear debris (Fig. 1). They were distributed in the white and grey matter of the brain and spinal cord. A small number of protozoa1 tachyzoites were found in association with these foci, except in fetus 1. In fetus 4 and the newborn calf (5)) the encephalitic lesion was characterized by moderate to severe perivascular infiltration by lymphocytes, macrophages, plasma cells and glial cells (Figs 2 and 3). Tachyzoites were commonly found in the peripheral areas of blood vessels (Fig. 4). In the newborn calf and in fetus 4, tissue cysts were seen primarily in the neurones and were oval or round in shape (Fig. 5). The tissue
Detection
Animal
of histological
lesion,
Gestational months
no.
Histological
Necrosis Fetus Fetus Fetus Fetus Calf
1 2 3 4
5 5 5 7 1 day
Table 1 antigen and immuaoglobulin-containing CNS
N. caninum
++* ++ ++ -
lesion*
Gliosis
N. caninumt ~ PVC
++ ++
Ig-ce11st
.-
__Tachyzoite
-
+++ +++
cells in the
+ ++ ++ ++
PVC = Perivascular cuffing. *: - , no lesion; + , slight; + + , moderate; + + + , severe. t: - , none; + , small number; + + , moderate number; + + + , large number.
Cyst + +
I&
W ++ +++
-
Neosporosis
Large
area of central
necrosis
in the cerebrum
Fig. 2.
Fig.
1.
Severe
inflammatory
reaction
in the spinal
Fig. 3.
Perivascular
cuffing
Fig. 4.
Small
of tachyzoites
group
composed
of plasma
in newborn
in Cattle
in fetus 2. HE cord of newborn
cells in the cerebrum calf. HE x 1000.
233
x
200.
calf. HE
X 160.
of fetus 4. HE x 400.
234
H. Ogino
et al.
cyst wall was up to 2.5 pm thick, homogeneous and enclosed slender bradyzoites (Fig. 6). Tissue cysts were separate from the inflammatory foci and showed no surrounding cellular reaction. Immunohistologically, the protozoa1 structures reacted positively with antiN. caninum serum, but not with anti-Toxoplasma gondii or anti-Hammondia hammondisera. The tachyzoites stained more intensely than did the bradyzoites. The cyst wall was also stained. Free tachyzoites and clusters of tachyzoites were associated with neural degeneration and some were found in blood vessels (Fig. 7). IgG-containing cells were present in the perivascular cuffs and were diffuse in the nodular foci (Fig. 8). They were detected in fetus 4 and in the newborn calf, but not in fetuses 1, 2 and 3. The number of positive cells in the newborn calf was much greater than that in fetus 4. IgM-containing cells were not found. Discussion The results of the present study revealed four aborted fetuses (Nos 1 to 4) and one dead newborn calf that had protozoa1 lesions in the central nervous system. The lesions were of two forms. One form, observed in fetuses 1, 2 and 3, was characterized by focal necrosis (necrotizing encephalitis) and the other, observed in fetus 4 and in a newborn calf, was characterized by moderate to severe infiltration of macrophages and plasma cells. Four of the brain lesions contained the protozoa1 organisms. These findings resembled those described for encephalitic protozoa1 infection in bovine fetuses associated with 3v. caninum-like organisms (Barr et al., 1990, 1991a,b; Dubey et al., 1990b). It is well known that protozoan parasites such as Sarcocystis (Hong, Giles, Newman and Fayer, 1982; Jolley, Jensen, Hancock and Swift, 1983; McCausland, Badman, Hides and Slee, 1984) and T. gondii (Dubey, 1981; Dubey and Schmitz, 1981) cause abortion in cows and produce encephalitic lesions. Sarcocystisis identified by its specific endothelial cell tropism and the presence of protozoa1 clusters in endothelial cells of various tissues. Degenerating ‘7: gondii is difficult to differentiate from degenerating host cells and diagnosis of toxoplasma1 abortion may depend more on recognition of lesions in the placenta than on identification of the parasite in tissue (Dubey, 1981; Dubey and Schmitz, 1981). The encephalitic lesions and structure of the tachyzoites of TT gondii are similar in appearance to those of N. caninum. Therefore, the identification of .N. caninumin the present four cases was by an immunoperoxidase technique and a specific antiserum, as in previous reports (Lindsay and Dubey, 1989; Barr et al., 1991b). The ability of fetuses to develop specific immunoglobulin to a variety of non-pathogenic antigens and congenitally acquired agents has been investigated in several species (Osburn, MacLachlan and Terrell, 1982). In the present study, fetuses at 5 months gestation (Nos 1, 2 and 3) had multifocal necrosis and tachyzoites, but no inflammatory cell reaction, suggesting that central nervous system cells were being destroyed by active multiplication of tachyzoites. However, fetus 4 (7 months) and the newborn calf had a
Neosporosis
Fig. 5.
Tissue cyst in a neurone
Fig. 6.
Tissue cyst with
Fig. 7.
Three
tachyzoites
Fig. 8.
Many
IgG-containing
of newborn
in Cattle
calf. Immunoperoxidase
a thick cyst wall in newborn (arrows)
235
in and around
(IP)
x 1000.
calf. IP x 1000. a small vessel in newborn
cells in the perivascular
cuffing
calf. IP x 1000
of fetus 4. IP X 400.
H. Ogino et al.
236
prominent inflammatory response with many IgG-containing cells and tissue cysts. This apparent association between gestational age and the degree of tissue inflammation may reflect the progressive development of the immune system in bovine fetuses. Acknowledgment The authors
thank
Mrs Y. Ando and T. Fujisawa
for preparing
the photographs.
References Barr, B. C., Anderson, M. L., Bianchard, P. C., Daft, B. M., Kinde, H. and Conrad, P. A. (1990). Bovine fetal encephalitis and myocarditis associated with protozoa1 infection. Veterinary Pathology, 27, 354-36 1. Barr, B. C., Anderson, M. L., Dubey, J. P. and Conrad, P. A. (1991a). Neosporu-like protozoa1 infections associated with bovine abortions. Veterinary Pathology, 28, 110-l 16. Barr, B. C., Conrad, P. A., Dubey, J. P. and Anderson, M. L. (1991 b). JVeosporu-like encephalomyelitis in a calf: Pathology, ultrastructure, and immunoreactivity. Journal of Veterinary Diagnostic Investigation, 3, 3946. Dubey, J. P. (1981). Epizootic Toxoplasmosis associated with abortion in dairy goats in Montana. 3ournal of the American Veterinary Medical Association, 178, 661-670. Dubey, J. P., Carpenter, J. L., Speer, C. A., Topper, M. J. and Uggla, A. (1988a). Newly recognized fatal protozoan disease of dogs. Journal of the American Veterinary Medical Association, 192, 1269-l 285. Dubey, J. P., Hattel, A. L., Lindsay, D. S. and Topper, M. J. (1988b). Neonatal Neospora caninum infection in dogs: Isolation of the causative agent and experimental transmission. Journal of the American Veterinary Medical Association, 193, 1259-1263. Dubey, J. P., Leathers, C. W. and Lindsay, D. S. (1989). Jveosporu cuninum-like protozoon associated with fetal myelitis in newborn calves. Journal of Parasitology, 75, 146-148. Dubey, J. P. and Lindsay, D. S. (1989). Fatal Jveosporu caninum infection in kittens. Journal of Parasitology, 75, 148-l 5 1. Dubey, J. P., Lindsay, D. S. and Lipscomb, T. P. (1990a). Neosporosis in cats. Veterinary Pathology, 27, 335-339. Dubey, J. P., Miller, S., Lindsay, D. S. and Topper, M. J. (1990b). ~eospora caninum-associated myocarditis and encephalitis in an aborted calf. Journal of Veterinary Diagnostic Investigation, 2, 66-69. Dubey, J. P. and Schmitz, J. A. (1981). Abortion associated with Toxoplasmosis in sheep in Oregon, Journal of the American Veterinary Medical Association, 178, 675-678. Hong, C. B., Giles, R. C., Newman, I. E. and Fayer, R. ( 1982). Surcocystosis in an aborted bovine fetus. Journal of the American Veterinary Medical Association, 181, 585-588. Jolley, W. R., Jensen, R., Hancock, H. A. and Swift, B. L. (1983). Encephalitic Sarcocystosis in a newborn calf. American Journal of Veterinary Research, 44, 1908-1911. Lindsay, D. S. and Dubey, J. P. ( 1989). I mmunohistochemical diagnosis of Neospora caninum in tissue section. American Journal of Veterinary Research, 50, 1981-I 983. McCausland, I. P., Badman, R. T., Hides, S. and Slee, K. J. (1984). Multiple apparent Sarcocystis abortion in four bovine herds. Cornell Veterinarian, 74, 146154. Osburn, B. I., MacLachlan, N. J. and Terrell, T. G. (1982). Ontogeny of the immune system. Journal of the American Veterinary Medical Association, 181, 1049-1052. Shivaprasad, H. L., Ely, R. and Dubey, J. P. (1989). A Neosporu-like protozoon found in an aborted bovine placenta. Veterinary Parasitology, 34, 145-148.
Neosporosis
in Cattle
237
Umemura, T., Shiraki, K., Morita, T., Shimada, A., Haritani, M., Kobayashi, M. and Yamagata, S. (1992). N eos p orosis in a dog: The first casereport in Japan. Journal of Veterinary Medical Science, 54, 157-I 59.
1
Received, March 3Oth, 1992 Accepted, June lst, 1992
