Br. J. exp. Path. (1979) 60, 417
PATHOGENICITY OF DIFFERENT STRAINS OF RIFT VALLEY FEVER VIRUS IN SWISS ALBINO MICE 0. TOMORI* AND 0. KASALI From the Virus Research Laboratory* and the Department of Veterinary Pathology, University of Ibadan, Ibadan, Nigeria Received for publication March 6, 1979
Summary. Laboratory mice were inoculated with 3 different strains of Rift Valley fever virus (RVFV): namely, the prototype, the Nigerian, and the Lunyo variant strains of RVFV. Animals were inoculated with either infective mouse brain or serum by the i.c. or i.p. route and organs of inoculated animals examined for virus content and histopathological changes. Animals inoculated with either the classical orthe Nigerian strain showed identical histopathological changes. These changes varied from mild encephalitis only, to a ditropism involving mild lesions in the brain and liver or the typically severe RVFV hepatitis. The type of lesion shown was dependent on the source of inoculum and the route of inoculation. On the other hand, animals inoculated by the i.c. route with the infective brain material of the Lunyo variant virus showed a mild encephalitis, while the use of the i.p. route or infective blood source of the Lunyo virus resulted in the development of typical RVFV hepatitis. Typical intranuclear eosinophilic inclusion bodies were found in both the brain and liver of animals inoculated with the Lunyo variant virus. These inclusion bodies were found only in the livers of mice inoculated with the infective blood material of either the prototype or the Nigerian strain. There was no correlation between the virus titres in the different organs and the severity of histopathological lesions.
WEINBREN, WILLIAMS AND HADDOW (1957) described the Lunyo virus, a variant of the prototype pantropic Rift Valley fever virus (RVFV), originally reported by Daubney, Hudson and Garnham ( 1931). The Lunyo variant virus differs from the prototype strain by its lack of haemagglutinating properties, and also by the histological lesions produced in laboratory mice. According to Weinbren et al. (1957), mice infected with the early passage of the Lunyo strain showed a viral encephalitis with the presence in the cornu ammmonis of intranuclear eosinophilic inclusion bodies, indistinguishable from those seen in the liver of mice infected with the classical or prototype RVFV. Virus strains indistinguishable in complement-fixation (CF) tests from pantropic RVFV have been isolated in Nigeria from a Merino sheep imported from South
Africa (Ferguson, 1959); and from pools of Culicoides and Culex antennatus mosquitoes (Lee, 1970). The absence of reports of clinical RVFV disease in any indigenous stock, despite widespread RVFV immunity in Nigerian livestock population (Fagbami, Tomori and Kemp, 1973), and the reports of a subelinical RVFV disease in the West African dwarf sheep experimentally infected with the Nigerian strain of RVFV (Fagbami et al., 1975), led to the speculation that the Nigerian strain may be another RVFV variant. A series of studies to determine the relationships between the Nigerian, the prototype and the Lunyo variant strains of RVFV were carried out. These studies employed the 3 different RVFV strains in experimental infection of the West African dwarf sheep, a comparison of the immunological reactions of the antigen-antibody
418
0. TOMORI AND 0. KASALI
systems of the 3 strains, as well as a study of the pathogenicity of the different RVFV strains for laboratory animals following experimental infection. The results of the experimental infection studies in the West African dwarf sheep and the immunological reactions between the strains have been reported elsewhere (Tomori, 1979a, b). This paper reports the findings of the pathogenic changes induced in laboratory mice following infection with these virus strains. MATERIALS AND METHODS
Viruses. Details of the virus strains used have been previously described (Tomori, 1979a). Briefly, the Nigerian strain of RVFV designated lb Ar55171 was used at the 4th mouse brain passage. It was originally isolated by Lee (1970) from a pool of Culex antennatus mosquitoes. The pantropic RVFV strain, designated lb Ar92620, was isolated by Smithburn, Haddow and Gillet (1948) from a mixed pool of Eretmapodites and Aedes spp. mosquitoes. It had undergone over 100 mouse intracerebral (i.e.) passages. The Lunyo variant, designated Ib Ar92621, was the 4th mouse brain passage of the original strain isolated from a pool of Aedes spp. mosquitoes by Weinbren et al. (1957). Both the lb Ar92620 and Ib Ar92621 virus strains were kindly supplied by the Director, East African Virus Research Institute, Entebbe, Uganda. Virus strains were passaged i.c. in suckling 2-3-day-old mice, and brain and serum material harvested from sick mice for titration. When infective brain and serum titres were 3 0 dex LD5o/0-02 ml or above (dex = logio; Haldane, 1960), seed viruses for experimental infection studies were prepared. This was in the form of pooled brain or serum material harvested from sick mice. Initial brain-virus titres for the 3 RVFV strains were between 5 0 and 6-5 dex LD5o/0 02 ml; however, virus strains lb Ar55171 and lb Ar92621 were passaged twice and lb Ar92620 three times to achieve the desired serum-virus titres. Experimental animals. Two- to 3-day-old laboratory bred Swiss albino mice certified free of any intercurrent disease were used. Experimental procedure. Experimental animals in groups of 30 animals each were inoculated with either infective brain or serum material of the different strains of RVFV, using the i.c. or intraperitoneal (i.p.) routes. Each animal was inoculated with 2 dex LD50 of virus. When mice became sick or moribund, they were anaesthesized and examined by necropsy, brain and liver being removed. Blood for viraemia
studies was also collected from each set of animals. Histopathology. Detailed necropsy examinations of both infected and control mice were carried out. The organs were cut in halves; one set was titrated for virus content, and the other halves were fixed in Bouin's fluid for 24 h, followed by 70% alcohol. After fixation, the tissues were embedded in paraffin, sectioned at a thickness of 5-7 ,um and stained with haematoxylin and eosin. Virus assay.- Viraemia, as well as the virus content in the brain and liver of infected animnals, was determined by the i.c. inoculation of 2-3-day-old suckling mice. Blood or organs from 4 animals per group were pooled, and 10% suspensions prepared in a diluent made up of 0-02 ml phosphate-buffered physiological saline (pH 7-2), to which was added 0-75% bovine plasma albumen (Armour Fraction V), supplemented with 100 u/ml of penicillin and 100 ,ug/ml streptomycin. Samples yielding virtus were further titrated for end-points, which were calculated by the method of Reed and Muench (1938), and expressed in dex (Haldane, 1960). RESULTS
The types of histopathological changes shown by albino mice infected with the 3 different strains of RVFV can be classified into 4 broad groupings (Table I). Animals inoculated by the i.p. route with the infective brain material of both the Nigerian strain (lb Ar55171) and the prototype (lb Ar92620) strain of RVFV showed no significant lesions in either the brain or liver. A mild encephalitis characterized by scanty necrosis of glial cells and mild polymorphonuclear infiltration was the only significant lesion seen in the mice infected by the i.c. route with brain material of either the Nigerian or the prototype RVFV strain. In addition to the mild encephalitis, the presence of intranuclear eosinophilic inclusion bodies was demonstrated in the brain sections of mice inoculated by the i.c. route with infective brain material of the Lunyo variant virus. Animals infected by the i.c. route with infective blood material of either the Nigerian or prototype RVFV strains exhibited identical histopathological
PATHOGENICITY OF RIFT VALLEY FEVER VIRUS
419
TABLE I. Cla8ssification of Histopathological Changes induced in Swiss Albino Mice infected by the Intracerebral and Intraperitoneal Routes with Different Strains of RVFV Virus
Sour ce of inoculum Brain
Bloodl
Route of inoculation Intracerebral Intraperitoneal Intracerebral Intraperitoneal
lb Ar 55171 (Nigerian) A B C D
lb Ar 92620 (Prototype) A B C D)
lb Ar 92621 (Lunyo) Al D D 1)
A= Mild encephalitis. Al - Aild encephalitis writh intranuclear eosinophilic inclusion bodies (IEIB). B =No significant lesion. C = Ditropism; mil(d encephalitis, and hepatitis with IEIB in hepatocytes only. D = Typical RVFV hepatitis with IEIB in hepatocytes.
lesions in the brain and liver. The lesions in the brain were characterized by a mild encephalitis with mild and scattered polymorphonuclear cell infiltration in the cerebrum and brainstem. In the liver, there were focal areas of necrosis of hepatocytes and the presence of intranuclear eosinoplhilic inclusion bodies in some necrotic hepatocytes. The fourth tvpe of lesion was the typical RVFV hepatitis, characterized by massive coagulative necrosis of the hepatocytes. The affected hepatocytes have eosinophilic, hyaline cytoplasm and pyknotic nuclei. The periportal areas were infiltrated by mononuclear cells composed chiefly of lymphocytes and plasma cells (Fig. 1). Small intranuclear inclusion bodies (Fig. 2) were present in some of the necrotic cells. These lesions were common to animals inoculated by the i.p. route with infective brain material of the Lunyo RVFV variant virus or those inoculated with infective blood material of the same virus by either the i.c. or i.p. route. Animals iinoculated i.c. with the infective blood material of either the Nigerian or prototype RY'FV strain also showed similar lesions. Table II shows the virus titres in the brain, liver and blood of mice inoculated with the different strains of RVFV. There was no correlation between the virus titres in the different organs (brain and liver) 28
and the severity of histopathological lesions. The virus titre in the brain of mice inoculated with the infective brain material of the Nigerian strain of RVFV' was significantly lower than those of mice inoculated with the same material of the other 2 strains. It is also of note that no virus was detected in organs of mice inoculated with any of the 3 RVFV strain when the infective brain material was inoculated by the i.p. route. DISCUSSION
The inoculation of laboratory mice with either the Nigerian or the prototype strain of RVFV resulted in identical histopathological changes. The major distinguishing feature between the histopathological changes induced by either of these 2 virus strains and the Lunyo strain is the presence of intranuclear eosinophilic inclusion bodies in the liver and brain samples of mice infected with the Lunyo variant strain irrespective of source of inoculum or route of inoculation. These inclusion bodies were found only in the liver samples of mice inoculated with infective blood material of either the Nigerian or prototype RVFV strain. Weinbren et al. (1957) reported the presence of inclusion bodies in the brains of mice inoculated with the neural source of Lunyo variant virus. Although the Nigerian strain was
420
0. TOMORI AND 0. KASALI
FIG. 1. Massive coagulative necrosis of hepatocytes with periportal infiltration by mononuclear cells, composed mainly of lymphocytes and plasma cells, in a mouse inoculated i.p. with infective brain material of the Lunyo variant of Rift Valley fever virus. H. and E. x 45. FIG. 2.-Presence of eosinophilic intranuclear inclusion body in the hepatocytes of a mouse inoculated i.p. with infective brain material of the Lunyo variant of Rift Valley fever virus. H. and E. x 1,000.
421
PATHOGENICITY OF RIFT VALLEY FEVER VIRUS
TABLE II.-Virus Titres in the Brain, Liver and Blood of Swiss Albino Mice inoculated with Different Strains of Rift Valley Fever Virus Virus strain lb Ar55171 (Nigerian)
Source of inoculum Brain
Bloo(d lb Ar92620 (T'antropic)
Brain
Bloodl Ib Ar92621 (Ltinyo)
Brain
Blood *
Route of inoculation i.e. i.p. i.e. i.p. i.e. i.p. i.e. i.p. i.e. i.p. i.c. i.p.
Average* survival time 1-0 10-0 2-0 2-6
1-0 > 14-0
1-2 2-0 1-0 10-0 1.0 3.5
Virus titrest (dex LD50/ml) in 'A
Brain 6-9 0.0 4-6 3-2 95 0 39 3-1 9.5 0-0
4.3 3' 1
Liver 3-3 00 3-2 3-5 2-7 0 1-5 5-3 4-2 0-0 3-2 5-3
Blood 1-6 0.0 1-2 1.1 2-2 0 2-6 2-4 3-8 0 2-0 24
Average survival time in days.
t Viruis titres represent the titres of pooled samples from 4 sick animals.
used at a inuch lower passage than the classical prototype RVFV, mice were not susceptible to i.p. inoculation of the infective brain material of the Nigerian strain. Smithburn (1949) also reported that the pathogenicity of the prototype virus for abdominal visceral tissues was so easily reduced by continued brain-to-brain passage that i.p. introduction of the agent seldom resulted in death. Not only did all the mice inoculated i.p. with the infective braini material of the Nigerian RVFV strain survive, but virus was not isolated nor was any significant lesion found in any of the organs examined. This, however, was not the case with the Lunyo virus; while no virus was detected in organs of mice inoculated i.p. with infective brain material of Lunyo variant virus, typical RVFV hepatic lesions were demonstrated in the livers of killed animals. From these experiments, there was no correlation between the virus titre of the various organs and the severity of histopathological changes. The rapid multiplication of RVFV in these organs and short average survival time might have been responsible for the paucity of histopathological lesions, particularly in animals infected with the brain material by the neural route. In previous studies employing sero28*
logical tests (Tomori, 1979b), it was shown that the Nigerian strain was immunologically closely related to, if not identical with, the prototype RVFV. The present studies confirm the previous observations and further strengthens the conclusions that the Nigerian strain of RVFV is a strain of the classical pantropic RVFV and not a variant like the Lunyo virus. REFERENCES DAIJBNEY, R., HUDSON, J. R. & GARNHAM, P. C. (1931) An Undescribed Virus Disease of Sheep, Cattle and Man from East Africa. J. Path. Bact., 34, 545. FAGBAMI, A. H., TOMORI, 0. & KEMP, G. E. (1973) A Survey of Nigerian Domestic Animals for Serum Neutralising Antibody to Indigenous Rift Valley Fever Virus. Nigerian Vet. J., 2, 45. FAGBAMI, A. H., ToMoRI, O., FABIYI, A. & IsouN, T. T. (1975) Experimental Rift Valley Fever in West African Dwarf Sheep. Res. Vet. Sci., 18, 334. FERGUSON, W. (1959) Identification of Rift Valley Fever in Nigeria. Bull. Epizootic Dis. Africa, 7, 317HALDANE, J. B. S. (1960) Dex or Order of Magnitude? Nature (Lond.), 187, 879. KITCHEN, S. F. (1950) The Development of Neurotropism in Rift Valley Fever Virus. Ann. Trop. Med. Parasitol., 44, 132. LEE, V. H. (1970) University of Ibadan Arbovirus Research Project, 1970 Annual Report. p. 30. REED, L. J. & MUENCH, H. (1938) A Simple Method of Estimating Fifty Per Cent End Points. Amer. J. trop. Med. Hyg., 27, 493. SMITHBITRN, K. C., HADDOW, A. J. & GILLET, J. D. (1948) Rift Valley Fever. Isolation of the Virus
422
0. TOMORI AND 0. KASALI
from Wild Mosquitoes. Br. J. exp. Path., 29, 107. SMITHBURN, K. C. (1949) Rift Valley Fever; the Neurotropic Adaptation of the Virus and the Experimental Use of this Modified Virus as a Vaccine. Br. J. exp. Path., 30, 1. TOMORI, 0. (1979a) Clinical, Virological and Serological Response of West African Dwarf Sheep to Experimental Infection with Different Strains
of Rift Valley Fever Virus. Res. Vet. Sci., 26, 152. TOMORI, 0. (1979b) Immunological Reactions of Rift Valley Fever Virus Strains from East and West Africa. Res. Vet. Sci., 26, 160. WEINBREN, M. P., WILLIAMS, M. C. & HADDOW, A. J. (1957) A Variant of Rift Valley Fever Virus. S. Afr. Med. J., 31, 951.
PATHOGENICITY OF DIFFERENT STRAINS OF RIFT VALLEY FEVER VIRUS IN SWISS ALBINO MICE 0. TOMORI* AND 0. KASALI From the Virus Research Laboratory* and the Department of Veterinary Pathology, University of Ibadan, Ibadan, Nigeria Received for publication March 6, 1979
Summary. Laboratory mice were inoculated with 3 different strains of Rift Valley fever virus (RVFV): namely, the prototype, the Nigerian, and the Lunyo variant strains of RVFV. Animals were inoculated with either infective mouse brain or serum by the i.c. or i.p. route and organs of inoculated animals examined for virus content and histopathological changes. Animals inoculated with either the classical orthe Nigerian strain showed identical histopathological changes. These changes varied from mild encephalitis only, to a ditropism involving mild lesions in the brain and liver or the typically severe RVFV hepatitis. The type of lesion shown was dependent on the source of inoculum and the route of inoculation. On the other hand, animals inoculated by the i.c. route with the infective brain material of the Lunyo variant virus showed a mild encephalitis, while the use of the i.p. route or infective blood source of the Lunyo virus resulted in the development of typical RVFV hepatitis. Typical intranuclear eosinophilic inclusion bodies were found in both the brain and liver of animals inoculated with the Lunyo variant virus. These inclusion bodies were found only in the livers of mice inoculated with the infective blood material of either the prototype or the Nigerian strain. There was no correlation between the virus titres in the different organs and the severity of histopathological lesions.
WEINBREN, WILLIAMS AND HADDOW (1957) described the Lunyo virus, a variant of the prototype pantropic Rift Valley fever virus (RVFV), originally reported by Daubney, Hudson and Garnham ( 1931). The Lunyo variant virus differs from the prototype strain by its lack of haemagglutinating properties, and also by the histological lesions produced in laboratory mice. According to Weinbren et al. (1957), mice infected with the early passage of the Lunyo strain showed a viral encephalitis with the presence in the cornu ammmonis of intranuclear eosinophilic inclusion bodies, indistinguishable from those seen in the liver of mice infected with the classical or prototype RVFV. Virus strains indistinguishable in complement-fixation (CF) tests from pantropic RVFV have been isolated in Nigeria from a Merino sheep imported from South
Africa (Ferguson, 1959); and from pools of Culicoides and Culex antennatus mosquitoes (Lee, 1970). The absence of reports of clinical RVFV disease in any indigenous stock, despite widespread RVFV immunity in Nigerian livestock population (Fagbami, Tomori and Kemp, 1973), and the reports of a subelinical RVFV disease in the West African dwarf sheep experimentally infected with the Nigerian strain of RVFV (Fagbami et al., 1975), led to the speculation that the Nigerian strain may be another RVFV variant. A series of studies to determine the relationships between the Nigerian, the prototype and the Lunyo variant strains of RVFV were carried out. These studies employed the 3 different RVFV strains in experimental infection of the West African dwarf sheep, a comparison of the immunological reactions of the antigen-antibody
418
0. TOMORI AND 0. KASALI
systems of the 3 strains, as well as a study of the pathogenicity of the different RVFV strains for laboratory animals following experimental infection. The results of the experimental infection studies in the West African dwarf sheep and the immunological reactions between the strains have been reported elsewhere (Tomori, 1979a, b). This paper reports the findings of the pathogenic changes induced in laboratory mice following infection with these virus strains. MATERIALS AND METHODS
Viruses. Details of the virus strains used have been previously described (Tomori, 1979a). Briefly, the Nigerian strain of RVFV designated lb Ar55171 was used at the 4th mouse brain passage. It was originally isolated by Lee (1970) from a pool of Culex antennatus mosquitoes. The pantropic RVFV strain, designated lb Ar92620, was isolated by Smithburn, Haddow and Gillet (1948) from a mixed pool of Eretmapodites and Aedes spp. mosquitoes. It had undergone over 100 mouse intracerebral (i.e.) passages. The Lunyo variant, designated Ib Ar92621, was the 4th mouse brain passage of the original strain isolated from a pool of Aedes spp. mosquitoes by Weinbren et al. (1957). Both the lb Ar92620 and Ib Ar92621 virus strains were kindly supplied by the Director, East African Virus Research Institute, Entebbe, Uganda. Virus strains were passaged i.c. in suckling 2-3-day-old mice, and brain and serum material harvested from sick mice for titration. When infective brain and serum titres were 3 0 dex LD5o/0-02 ml or above (dex = logio; Haldane, 1960), seed viruses for experimental infection studies were prepared. This was in the form of pooled brain or serum material harvested from sick mice. Initial brain-virus titres for the 3 RVFV strains were between 5 0 and 6-5 dex LD5o/0 02 ml; however, virus strains lb Ar55171 and lb Ar92621 were passaged twice and lb Ar92620 three times to achieve the desired serum-virus titres. Experimental animals. Two- to 3-day-old laboratory bred Swiss albino mice certified free of any intercurrent disease were used. Experimental procedure. Experimental animals in groups of 30 animals each were inoculated with either infective brain or serum material of the different strains of RVFV, using the i.c. or intraperitoneal (i.p.) routes. Each animal was inoculated with 2 dex LD50 of virus. When mice became sick or moribund, they were anaesthesized and examined by necropsy, brain and liver being removed. Blood for viraemia
studies was also collected from each set of animals. Histopathology. Detailed necropsy examinations of both infected and control mice were carried out. The organs were cut in halves; one set was titrated for virus content, and the other halves were fixed in Bouin's fluid for 24 h, followed by 70% alcohol. After fixation, the tissues were embedded in paraffin, sectioned at a thickness of 5-7 ,um and stained with haematoxylin and eosin. Virus assay.- Viraemia, as well as the virus content in the brain and liver of infected animnals, was determined by the i.c. inoculation of 2-3-day-old suckling mice. Blood or organs from 4 animals per group were pooled, and 10% suspensions prepared in a diluent made up of 0-02 ml phosphate-buffered physiological saline (pH 7-2), to which was added 0-75% bovine plasma albumen (Armour Fraction V), supplemented with 100 u/ml of penicillin and 100 ,ug/ml streptomycin. Samples yielding virtus were further titrated for end-points, which were calculated by the method of Reed and Muench (1938), and expressed in dex (Haldane, 1960). RESULTS
The types of histopathological changes shown by albino mice infected with the 3 different strains of RVFV can be classified into 4 broad groupings (Table I). Animals inoculated by the i.p. route with the infective brain material of both the Nigerian strain (lb Ar55171) and the prototype (lb Ar92620) strain of RVFV showed no significant lesions in either the brain or liver. A mild encephalitis characterized by scanty necrosis of glial cells and mild polymorphonuclear infiltration was the only significant lesion seen in the mice infected by the i.c. route with brain material of either the Nigerian or the prototype RVFV strain. In addition to the mild encephalitis, the presence of intranuclear eosinophilic inclusion bodies was demonstrated in the brain sections of mice inoculated by the i.c. route with infective brain material of the Lunyo variant virus. Animals infected by the i.c. route with infective blood material of either the Nigerian or prototype RVFV strains exhibited identical histopathological
PATHOGENICITY OF RIFT VALLEY FEVER VIRUS
419
TABLE I. Cla8ssification of Histopathological Changes induced in Swiss Albino Mice infected by the Intracerebral and Intraperitoneal Routes with Different Strains of RVFV Virus
Sour ce of inoculum Brain
Bloodl
Route of inoculation Intracerebral Intraperitoneal Intracerebral Intraperitoneal
lb Ar 55171 (Nigerian) A B C D
lb Ar 92620 (Prototype) A B C D)
lb Ar 92621 (Lunyo) Al D D 1)
A= Mild encephalitis. Al - Aild encephalitis writh intranuclear eosinophilic inclusion bodies (IEIB). B =No significant lesion. C = Ditropism; mil(d encephalitis, and hepatitis with IEIB in hepatocytes only. D = Typical RVFV hepatitis with IEIB in hepatocytes.
lesions in the brain and liver. The lesions in the brain were characterized by a mild encephalitis with mild and scattered polymorphonuclear cell infiltration in the cerebrum and brainstem. In the liver, there were focal areas of necrosis of hepatocytes and the presence of intranuclear eosinoplhilic inclusion bodies in some necrotic hepatocytes. The fourth tvpe of lesion was the typical RVFV hepatitis, characterized by massive coagulative necrosis of the hepatocytes. The affected hepatocytes have eosinophilic, hyaline cytoplasm and pyknotic nuclei. The periportal areas were infiltrated by mononuclear cells composed chiefly of lymphocytes and plasma cells (Fig. 1). Small intranuclear inclusion bodies (Fig. 2) were present in some of the necrotic cells. These lesions were common to animals inoculated by the i.p. route with infective brain material of the Lunyo RVFV variant virus or those inoculated with infective blood material of the same virus by either the i.c. or i.p. route. Animals iinoculated i.c. with the infective blood material of either the Nigerian or prototype RY'FV strain also showed similar lesions. Table II shows the virus titres in the brain, liver and blood of mice inoculated with the different strains of RVFV. There was no correlation between the virus titres in the different organs (brain and liver) 28
and the severity of histopathological lesions. The virus titre in the brain of mice inoculated with the infective brain material of the Nigerian strain of RVFV' was significantly lower than those of mice inoculated with the same material of the other 2 strains. It is also of note that no virus was detected in organs of mice inoculated with any of the 3 RVFV strain when the infective brain material was inoculated by the i.p. route. DISCUSSION
The inoculation of laboratory mice with either the Nigerian or the prototype strain of RVFV resulted in identical histopathological changes. The major distinguishing feature between the histopathological changes induced by either of these 2 virus strains and the Lunyo strain is the presence of intranuclear eosinophilic inclusion bodies in the liver and brain samples of mice infected with the Lunyo variant strain irrespective of source of inoculum or route of inoculation. These inclusion bodies were found only in the liver samples of mice inoculated with infective blood material of either the Nigerian or prototype RVFV strain. Weinbren et al. (1957) reported the presence of inclusion bodies in the brains of mice inoculated with the neural source of Lunyo variant virus. Although the Nigerian strain was
420
0. TOMORI AND 0. KASALI
FIG. 1. Massive coagulative necrosis of hepatocytes with periportal infiltration by mononuclear cells, composed mainly of lymphocytes and plasma cells, in a mouse inoculated i.p. with infective brain material of the Lunyo variant of Rift Valley fever virus. H. and E. x 45. FIG. 2.-Presence of eosinophilic intranuclear inclusion body in the hepatocytes of a mouse inoculated i.p. with infective brain material of the Lunyo variant of Rift Valley fever virus. H. and E. x 1,000.
421
PATHOGENICITY OF RIFT VALLEY FEVER VIRUS
TABLE II.-Virus Titres in the Brain, Liver and Blood of Swiss Albino Mice inoculated with Different Strains of Rift Valley Fever Virus Virus strain lb Ar55171 (Nigerian)
Source of inoculum Brain
Bloo(d lb Ar92620 (T'antropic)
Brain
Bloodl Ib Ar92621 (Ltinyo)
Brain
Blood *
Route of inoculation i.e. i.p. i.e. i.p. i.e. i.p. i.e. i.p. i.e. i.p. i.c. i.p.
Average* survival time 1-0 10-0 2-0 2-6
1-0 > 14-0
1-2 2-0 1-0 10-0 1.0 3.5
Virus titrest (dex LD50/ml) in 'A
Brain 6-9 0.0 4-6 3-2 95 0 39 3-1 9.5 0-0
4.3 3' 1
Liver 3-3 00 3-2 3-5 2-7 0 1-5 5-3 4-2 0-0 3-2 5-3
Blood 1-6 0.0 1-2 1.1 2-2 0 2-6 2-4 3-8 0 2-0 24
Average survival time in days.
t Viruis titres represent the titres of pooled samples from 4 sick animals.
used at a inuch lower passage than the classical prototype RVFV, mice were not susceptible to i.p. inoculation of the infective brain material of the Nigerian strain. Smithburn (1949) also reported that the pathogenicity of the prototype virus for abdominal visceral tissues was so easily reduced by continued brain-to-brain passage that i.p. introduction of the agent seldom resulted in death. Not only did all the mice inoculated i.p. with the infective braini material of the Nigerian RVFV strain survive, but virus was not isolated nor was any significant lesion found in any of the organs examined. This, however, was not the case with the Lunyo virus; while no virus was detected in organs of mice inoculated i.p. with infective brain material of Lunyo variant virus, typical RVFV hepatic lesions were demonstrated in the livers of killed animals. From these experiments, there was no correlation between the virus titre of the various organs and the severity of histopathological changes. The rapid multiplication of RVFV in these organs and short average survival time might have been responsible for the paucity of histopathological lesions, particularly in animals infected with the brain material by the neural route. In previous studies employing sero28*
logical tests (Tomori, 1979b), it was shown that the Nigerian strain was immunologically closely related to, if not identical with, the prototype RVFV. The present studies confirm the previous observations and further strengthens the conclusions that the Nigerian strain of RVFV is a strain of the classical pantropic RVFV and not a variant like the Lunyo virus. REFERENCES DAIJBNEY, R., HUDSON, J. R. & GARNHAM, P. C. (1931) An Undescribed Virus Disease of Sheep, Cattle and Man from East Africa. J. Path. Bact., 34, 545. FAGBAMI, A. H., TOMORI, 0. & KEMP, G. E. (1973) A Survey of Nigerian Domestic Animals for Serum Neutralising Antibody to Indigenous Rift Valley Fever Virus. Nigerian Vet. J., 2, 45. FAGBAMI, A. H., ToMoRI, O., FABIYI, A. & IsouN, T. T. (1975) Experimental Rift Valley Fever in West African Dwarf Sheep. Res. Vet. Sci., 18, 334. FERGUSON, W. (1959) Identification of Rift Valley Fever in Nigeria. Bull. Epizootic Dis. Africa, 7, 317HALDANE, J. B. S. (1960) Dex or Order of Magnitude? Nature (Lond.), 187, 879. KITCHEN, S. F. (1950) The Development of Neurotropism in Rift Valley Fever Virus. Ann. Trop. Med. Parasitol., 44, 132. LEE, V. H. (1970) University of Ibadan Arbovirus Research Project, 1970 Annual Report. p. 30. REED, L. J. & MUENCH, H. (1938) A Simple Method of Estimating Fifty Per Cent End Points. Amer. J. trop. Med. Hyg., 27, 493. SMITHBITRN, K. C., HADDOW, A. J. & GILLET, J. D. (1948) Rift Valley Fever. Isolation of the Virus
422
0. TOMORI AND 0. KASALI
from Wild Mosquitoes. Br. J. exp. Path., 29, 107. SMITHBURN, K. C. (1949) Rift Valley Fever; the Neurotropic Adaptation of the Virus and the Experimental Use of this Modified Virus as a Vaccine. Br. J. exp. Path., 30, 1. TOMORI, 0. (1979a) Clinical, Virological and Serological Response of West African Dwarf Sheep to Experimental Infection with Different Strains
of Rift Valley Fever Virus. Res. Vet. Sci., 26, 152. TOMORI, 0. (1979b) Immunological Reactions of Rift Valley Fever Virus Strains from East and West Africa. Res. Vet. Sci., 26, 160. WEINBREN, M. P., WILLIAMS, M. C. & HADDOW, A. J. (1957) A Variant of Rift Valley Fever Virus. S. Afr. Med. J., 31, 951.
