1973 Onderstepoort Journal of Veterinary Research, Volume 40, 1973

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Contentes Pages: Onderstepoort Journal of Veterinary Research, Vol ??, 19??

CONTENTS

*Volume 40: Number 1*

Serological titres following vaccination of sheep and goats with Brucella melitensis Rev 1 vaccine	Worthington, RW, Mulders, MSG, McFarlane, IS & Becker, D	1

A serological investigation on adult cattle vaccinated with Brucella abortus strain 19	Worthington, RW, Mulders, MSG, McFarlane, IS & Becker, D	7

Margaropus wileyi sp. nov. (Ixodoidea, Ixodidae), a new species of tick from the reticulated giraffe	Walker, JB & Laurence BR	13

Studies on schistosomiasis. 4. Differential staining of live and dead cercariae after immobilization with physostigmin	Van Wyk, JA	23

The toxicology and pathology of dieldrin and photodieldrin poisoning in two antelope species	Wiese, IH, Basson, NCJ, Basson, PA, Naude, TW & Maartens, BP	31


*Volume 40: Number 2*

Foot-and-mouth disease in the African elephant (Loxodonta africana)	Howell, PG, Young, E & Hedger, RS	41

Electron microscopic studies on equine encephalosis virus	Lecatsas, G, Erasmus, BJ & Els, HJ	53

The diagnosis of bovine mastitis : a critical evaluation of a polyvalent radial immunodiffusion test and other methods	Giesecke, WH, Van den Heever, LW, Du Toit, IJ & Beyer, MCE	59

Helminths from the mountain reedbuck, Redunca fulvorufula (Afzelius, 1815)	Baker, MK & Boomker, J	69

Research notes:

Mechanism of envelopment of herpesvirus by the nucelar envelope	Lecatsas, G & Poste, G	71

Electron microscopy of bluetongue virus RNA	Els, HJ	73


*Volume 40: Number 3*

Studies on the development of a vaccine against bovine ephemeral fever	Theodoridis, A, Boshoff, SET & Botha, MJ	77

Effects of ephemeral fever on milk production and reproduction of dairy cattle	Theodoridis, A, Giesecke, WH & Du Toit, IJ	83

Molecular hybridization studies on the relationships between different serotypes of bluetongue virus and on the difference between the virulent and attenuated strains of the same serotype	Huismans, H & Howell, PG	93

Studies on the enhancement of immunity to Corynebacterium pseudotuberculosis	Cameron, CM & Fuls, WJ	105

Photosensitivity in South Africa. 1. A comparative study of Asaemia Axillaris (Thunb.) Harv. ex Jackson and Lasiospermum bipinnatum (Thunb.) Druce poisoning in sheep	Kellerman, TS, Basson, PA, Naude, TW, Van Rensburg, IBJ & Welman, WG	115

Lesions in sheep skeletal and oesophageal muscle in vermeersiekte (Geigeria ornativa O. Hoffm. poisoning)	Pienaar, JG, Kriek, NPJ, Naude, TW, Adelaar, TF & Ellis, SD	127


*Volume 40: Number 4*

Variation in morphology of ephemeral fever virus	Theodoridis, A & Lecatsas, G	139

Clostridium perfringens type D epsilon prototoxin. Some chemical, immunological and biological properties of a highly purified prototoxin	Worthington, RW, Mulders, MSG & Van Rensburg, JJ	143

Enzymatic activation of Clostridium perfringens epsilon prototoxin and some biological properties of activated toxin	Worthington, RW, Mulders, MSG & Van Rensburg, JJ	151

Studies on schistosomiasis. 5. Sampling methods for estimating the numbers of cercariae in suspension with special reference to the infestation of experimental animals	Van Wyk, JA & Groeneveld, HT	155

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Now showing 1 - 20 of 21

Lesions in sheep skeletal and oesophageal muscle in vermeersiekte (Geigeria ornativa O. Hoffm. poisoning)
(Published by the Government Printer, Pretoria, 1973) Pienaar, J.G.; Kriek, N.P.J.; Naude, T.W.; Ellis, Shirley D.; Adelaar, T.F.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Vermeersiekte in sheep, which is characterized clinically by vomition of ruminal contents, often accompanied by stiffness or paralysis, is caused by various plant species of the genus Geigeria. A histopathological study of the skeletal and oesophageal muscles of three experimental and three natural ovine cases revealed lesions in every case. In paraffin sections examined by light microscopy, vacuolation of focal groups of muscle fibres was seen. The sarcoplasm in the vicinity of the vacuoles was hyalinized and single or multiple centrally displaced sarcolemmal nuclei occurred in the vacuoles. A variation in the size of muscle fibres in these foci was seen in the more chronic experimental cases. Small "atrophic" hyalinized fibres with centralization and proliferation of sarcolemmal nuclei were encountered. Electron micrographs revealed that the vacuoles in the sarcoplasm resulted from focal degeneration of myofibrils in otherwise intact muscle fibres. The thick myofilaments disappeared first, causing dissolution of the A-band in affected myofibrils. Shredding of the remaining thin filaments eventually lead to the complete destruction of myofibrils and the appearance in the fibre of irregular areas of fine granular material, containing remnants of myofilaments, Z-band material and swollen vacuolated mitochondria. Due to excessive loss of myofibrils the diameter of some muscle fibres was reduced.
Photosensitivity in South Africa. 1. A comparative study of Asaemia axillaris (Thunb.) Harv. Ex Jackson and Lasiospermum bipinnatum (Thunb.) Druce poisoning in sheep
(Pretoria : The Government Printer, 1973) Basson, P.A.; Kellerman, T. Stephanus; Naude, T.W.; Van Rensburg, I.B.J.; Welman, Wilhelmina G.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Experimental evidence is presented to confirm field observations that Asaemia axillaris (Thunb.) Harv. ex Jackson and Lasiospermum bipinnatum (Thunb.) Druce cause ovine hepatogenous photosensitization. The two plants have fairly distinct distributions. While Lasiospermum is most prevalent in theeastern Karoo, Asaemia is commonest in the west and, except perhaps in the southern central Karoo,they seldom occur together. The locality of an outbreak should therefore be taken into account when making a diagnosis. Peripheral fatty changes and midzonal necrosis of liver lobules were seen in both Asaemia and Lasiospermum poisoning. Asaemia poisoning could, however, be distinguished from Lasiospermum poisoning by the presence of centrilobular fatty degeneration and the absence of haemorrhages and bile pigmentation. Although it may be difficult to distinguish between the two types of plant poisoning they are easily differentiated from geeldikkop (a major photosensitizing disease of the Karoo), which lacks a zonal pattern of hepatocytic degeneration and necrosis. Moreover, the crystalloid material characteristic of geeldikkop is not encountered in Asaemia or Lasiospermum poisoning.
Molecular hybridization studies on the relationships between different serotypes of bluetongue virus and on the difference between the virulent and attenuated strains of the same serotype
(Pretoria : The Government Printer, 1973) Howell, P.G.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.; Huismans, H. (Henk), 1942-
Isolates of ³H-labelled messenger RNA of a number of different bluetongue virus serotypes were hybridized with saturating amounts of denatured ³²P-labelled double-stranded RNA of different serotypes. These cross-hybridization products were then analysed by polyacrylamide gel electrophoresis. The results indicate relatively large differences between the various serotypes. Only a few of the genome segments in the different serotypes were completely homologous. Each of the cross- hybridization patterns obtained using the genome of Serotype 10 and any one of the other serotypes was unique and characteristic for the strain under investigation. The patterns furthermore clearly indicated different degrees of homology between the genomes of the different serotypes. The immunological specificity of the serotypes appears to be determined mainly by the second genome segment of the virus while genome segment six could be of secondary importance. These results were supported by a study of the cross-hybridization patterns between different isolates of Serotype 4. Cross-hybridization experiments between virulent and attenuated strains of the same serotype also indicated small differences. In all the serotypes investigated the process of attenuation involved changes in genome segments two and six. This result would tend to implicate the same genome segments in the determination of both the immunological specificity and the virulence of the virus.
Effects of ephemeral fever on milk production and reproduction of dairy cattle
(Pretoria : The Government Printer, 1973) Theodoridis, A.; Giesecke, W.H.; Du Toit, I.J.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
The effect of ephemeral fever (EF) on the lactation, pregnancy and fertility of 19 dairy cows was studied. During the febrile reaction milk yields were reduced by an average of 58,7 ± 22 %, depending on the stage of lactation at which the infection was contracted. Ephemeral fever predisposes lactating udders to mastitogenic bacterial infections by deleteriously affecting the leucocytic udder barrier. Delayed oestrus was associated with EF in five out of 14 cows examined.
Studies on the enhancement of immunity to Corynebacterium pseudotuberculosis
(Pretoria : The Government Printer, 1973) Cameron, Colin McKenzie; Fuls, W.J.P.; De Lange, M.; Howell, P.G.; Bigalke, R.D.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Experiments were undertaken to enhance the immune response of mice, guinea pigs and sheep to C. pseudotuberculosis vaccine. The antibody response of guinea pigs was not increased either by the simultaneous administration of formalin inactivated vaccine and heterologous live vaccine, or by vaccines grown on special media and extracted with ether : ethanol or sodium lauryl sulphate, or by endotoxin. In mice and sheep the immunity was not enhanced either by the intravenous administration of inactivated vaccines or by the subcutaneous injection of live vaccines prepared from avirulent strains. In sheep an inactivated vaccine, concentrated tenfold, caused a moderate increase in the antibody titre and there was a marked improvement in their immunity. Such sheep were markedly more resistant to the lethal effects of a subacute infection of living bacteria than those that received standard inactivated vaccine. It is concluded that concentrated C. pseudotuberculosis vaccine does not confer an absolute immunity but it would be of value when used shortly before possible exposure to infection, e.g. shearing.
Studies on the development of a vaccine against bovine ephemeral fever
(Pretoria : The Government Printer, 1973) Theodoridis, A.; Boshoff, S.E.T.; Botha, M.J.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Bovine ephemeral fever virus of low passage level (5th to 8th) in tissue culture has proved to be more antigenic for cattle than higher passage level (17th to 19th) or mouse-brain-adapted virus. However, the neutralizing antibody response appears to be short-lived and therefore probably provides inadequate protection. Similar results were obtained when ephemeral fever virus of both low and high passage level was adsorbed onto aluminium hydroxide and inoculated into cattle. Emulsions of ephemeral fever virus of both low and high passage level in Freund's incomplete mineral oil adjuvant induced high neutralizing antibody titres in vaccinated cattle. These antibodies were detectable for at least a year and the animals were found to be immune when infected with virulent bovine ephemeral fever virus.
Electron microscopy of bluetongue virus RNA
(Published by the Government Printer, Pretoria, 1973) Els, H.J.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
RNA fragments were released from bluetongue virus particles using the Kleinschmidt spreading technique. Electron microscopy of viruses in protein monolayers spread on urea showed that each virion liberated an average of about 6 fragments although up to 10 fragments were occasionally observed. A large variation of filament lengths was found with an average of 3,5 µm total composite length and with a maximum of 6,5 µm leading to an estimated RNA molecular mass of 13,6 X 10⁶ . In addition, a background of loose dispersed fragments was invariably found.
Mechanism of envelopment of herpesvirus by the nuclear envelope
(Published by the Government Printer, Pretoria, 1973) Lecatsas, G.; Poste, G.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Studies on schistosomiasis. 5. Sampling methods for estimating the numbers of cercariae in suspension with special reference to the infestation of experimental animals
(Published by the Government Printer, Pretoria, 1973) Van Wyk, Jan Aucamp; De Lange, M; Bigalke, R.D.; Cameron, Colin McKenzie; Walker, Jane B.; Verster, Anna J.M.; De Kock, V.E.; Groeneveld, Hendrik T.
Various sampling methods for Schistosoma mattheei cercariae, including sampling from different concentrations, were tested. The most satisfactory method consisted of mixing the cercarial suspension by pouring it to and fro between two cylinders and then sampling it by oral suction on a bulb pipette with a large aperture. The aliquots withdrawn were stored in bottles containing formol-saline for later microscopic examination. This method of sampling was tested extensively. The results showed that the variation between aliquots was larger than that expected for either a binomial or a Poisson distribution, indicating that the cercariae were not randomly distributed in the suspension. Moreover, significant variations occurred between the totals of series of aliquots, possibly indicating uncontrolled factors, e.g. in mixing. From these results, methods of making up doses of carcariae for infesting sheep and cattle were developed: The Sheep Method is used for doses of up to 15 000 cercariae and is therefore applicable to infestation of primates as well as sheep. Several series of aliquots are made up, some of which (one or more series per animal) are used for infesting the animals concerned (Infestation Series), while others are retained for estimating the infestation doses (Estimation and Additional Estimation Series). One Estimation and one Additional Estimation Series suffice for estimating the numbers of cercariae in up to eight Infestation Series drawn from the same cercarial suspension, which is mixed immediately before each series of aliquots is withdrawn. In the Bovine Method, which is used for doses of over 15000 cercariae, an Estimation Series and an Additional Estimation Series of aliquots are withdrawn from every dose of cercariae to be used for infesting an animal. Thus the number of cercariae is estimated separately for each animal. Formulae were compiled for these methods to determine the upper levels (with a specified probability) of the percentage errors in the estimated doses. These formulae were applied to the results and some expected percentage errors in the numbers of cercariae estimated by the two methods, calculated. A result for the probability distribution of aliquot counts under the assumption of randomness is proved for the first time.
Helminths from the mountain reedbuck, Redunca fulvorufula (Afzelius, 1815)
(Published by the Government Printer, Pretoria, 1973) Boomker, Jacob Diederik Frederik; Baker, Maureen K.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Helminth parasites recovered from the mountain reedbuck in the Loskop Dam Nature Reserve and the Mountain Zebra National Park are recorded. The following species are new host records: Moniezia expansa, Cooperia hungi, C. oncophora, C. pectinata, C.yoshidai, Gongylonema sp, Haemonchus krugeri, Impalaia tuberculata, Nematodirus spathiger, Oesophagostomum columbianum, Skrjabinema sp.
The diagnosis of bovine mastitis : a critical evaluation of a polyvalent radial immunodiffusion test and other methods
(Published by the Government Printer, Pretoria, 1973) Giesecke, W.H.; Van den Heever, Louwrens Wepener; Du Toit, I.J.; Beyer, Maria C. E.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
A critically comparative investigation was performed on 228 quarter samples from 26 Friesland-type dairy cows respectively to evaluate the diagnostic value of conventional direct and indirect cytological methods and a single radial immunodiffusion test based on a polyvalent antiserum. Data obtained suggest considerable diagnostic inaccuracies for all the methods tested. The lack of an adequate "mastitis standard" prevented a precise determination of the magnitude of the diagnostic inaccuracies.
Electron microscopic studies on equine encephalosis virus
(Published by the Government Printer, Pretoria, 1973) Lecatsas, G.; Erasmus, B.J.; Els, H.J.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Electron microscopic studies carried out on thin sections of BHK 21 cells infected with equine encephalosis virus (EEV) have revealed a fine cytopathology closely resembling that shown by bluetongue and African horsesickness viruses. The characteristics include the presence of granular inclusion bodies from which mature virus particles appear to arise, swelling of the rough endoplasmic reticulum and the presence of inclusion bodies in the mitochondrial matrix. The diameter of the sectioned virus particle is approximately 73 nm, and 60 nm when negatively stained. The experimental observations suggest that the capsid is composed of 32 morphological subunits.
Foot-and-mouth disease in the African elephant (Loxodonta africana)
(Published by the Government Printer, Pretoria, 1973) Howell, P.G.; Young, E.; Hedger, R.S.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
A strain of SAT 2 foot-and-mouth disease virus which was experimentally inoculated into the epidermis of the tongues of captive African elephants produced vesicular lesions at the site of inoculation. After a short period of viraemia, secondary lesions developed in the mouth and on the feet giving rise to extensive tissue damage and the separation of the soles. In spite of close contact there was no spread of the disease to other elephants and by conventional sampling techniques no carrier virus could be demonstrated. The neutralizing antibody response was of a low order and this finding together with the observations made during the course of the experimental disease are discussed in relation to the possible role of the elephant in the epizootiology of foot-and-mouth disease in Africa.
The toxicology and pathology of dieldrin and photodieldrin poisoning in two antelope species
(Published by the Government Printer, Pretoria, 1973) Wiese, I.H.; Basson, P.A.; Naude, T.W.; Maartens, B.P.; Basson, N.C.J.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Blesbuck (Damaliscus dorcas phillipsi) and springbuck (Antidorcas marsupialis) were found to be more susceptible to dieldrin and its photoisomer than any vertebrates reported on previously in the literature. In blesbuck the mean cumulative lethal dose of dieldrin at various dietary levels was 9,07 mg/kg and of photodieldrin 1,90 mg/kg. Under field conditions the calculated median lethal dose of photodieldrin to blesbuck was 1,21 mg/kg; to springbuck it was 3,97 mg/kg. In both species rams were more susceptible to photodieldrin than ewes. It was proved that antelope mortalities on veld sprayed with dieldrin for harvester termite (Hodotermes mossambicus) control was principally due to photodieldrin intoxication. In addition to the usual violent nervous signs encountered in chlorinated hydrocarbon intoxication a dumb syndrome, in which blindness was rather characteristic, was seen, especially in the field cases. The pathological changes in the blesbuck and springbuck were similar. Degenerative lesions typical of various conditions, including chlorinated hydrocarbon poisoning, were encountered in the skeletal muscles, myocardium, kidneys and liver of cases of both dieldrin and photodieldrin poisoning. The severity and widespread nature of the lesions in the skeletal muscles and myocardium of natural cases were reminiscent of white muscle disease and capture myopathy. Congestion, oedema and glial swelling of the brain were constant features.
Enzymatic activation of Clostridium perfringens epsilon prototoxin and some biological properties of activated toxin
(Published by the Government Printer, Pretoria, 1973) Worthington, R.W.; Mulders, M.S.G.; Van Rensburg, J.J.; De Lange, M.; Bigalke, R.D.; Cameron, Colin McKenzie; Walker, Jane B.; Verster, Anna J.M.; De Kock, V.E.
Maximal activation of Clostridium perfringens epsilon toxin was only achieved by the combined action of trypsin and chymotrypsin. Impure preparations of trypsin, presumably containing small amounts of chymotrypsin were more efficient in activating prototoxin than pure trypsin. Activated toxin was readily absorbed by brain tissue and smaller amounts were possibly absorbed by kidney tissue. Other tissues absorbed only very small amounts of toxin. Injection of mice with toxoid 3 h prior to challenge with toxin increased their resistance 32 times.
Studies on schistosomiasis. 4. Differential staining of live and dead cercariae after immobilization with physostigmin
(Published by the Government Printer, Pretoria, 1973) Van Wyk, Jan Aucamp; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
A modified eosin technique similar to that used for differential staining of live and dead spermatozoa stained dead cercariae of Schistosoma mattheei red while those that were alive remained colourless and transparent. Prior to exposure to the dye live cercariae were immobilized with physostigmin, which was better for that purpose than five other substances tested. The lowest concentration of physostigmin to immobilize cercariae quickly and effectively was found to be 3,3 x 10-⁶ M.
Clostridium perfringens type D epsilon prototoxin. Some chemical, immunological and biological properties of a highly purified prototoxin
(Published by the Government Printer, Pretoria, 1973) Worthington, R.W.; Mulders, M.S.G.; Van Rensburg, J.J.; De Lange, M.; Bigalke, R.D.; Cameron, Colin McKenzie; Walker, Jane B.; Verster, Anna J.M.; De Kock, V.E.
Highly purified Cl. perfringens type D epsilon prototoxin was prepared by ammonium sulphate precipitation and DEAE cellulose chromatography of culture filtrate of cultures of Cl. perfringens type D (Strain ET 468). Preparations of prototoxin were electrophoretically heterogeneous. The protein bands demonstrable in polyacrylamide gel electrophoresis were, however, all immunologically identical and toxic. The faster moving bands were shown to be degradation products of the main prototoxin band which was the slowest moving of the major bands. There was an inverse relationship between electrophoretic mobility and the activation ratio of these degradation products. The unde- graded prototoxin could be separated from its degradation products by CM cellulose chromatography but degradation appears to be a continual process and isolation of an absolutely pure product was not achieved.
Margaropus wileyi sp. nov. (Ixodoidea, Ixodidae), a new species of tick from the reticulated giraffe
(Published by the Government Printer, Pretoria, 1973) Walker, Jane B.; Laurence, B.R.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Descriptions are given of the male and female of Margaropus wileyi sp. nov. collected from the reticulated giraffe in Kenya. A key is given to the three species now known in the genus Margaropus and their differential diagnosis is discussed.
A serological investigation on adult cattle vaccinated with Brucella abortus strain 19
(Published by the Government Printer, Pretoria, 1973) Worthington, R.W.; Mulders, M.S.G.; McFarlane, I.S.; Becker, D.; De Lange, M.; Bigalke, R.D.; Howell, P.G.; Reinecke, R.K.; Walker, Jane B.; De Kock, V.E.
Serum antibody titres were followed for a period of two years in 99 beef and 29 dairy cows which had been vaccinated as adults with B. abortus strain 19. Agglutination titres remained above diagnostic levels throughout but complement fixation, mercaptoethanol agglutination and rivanol agglutination titres generally returned to negative levels within 6 months. Coombs tests and agglutination at 56° proved to be of limited value for recognising antibody titres caused by vaccination.
Variation in morphology of ephemeral fever virus
(Published by the Government Printer, Pretoria, 1973) Theodoridis, A.; Lecatsas, G.; De Lange, M.; Bigalke, R.D.; Cameron, Colin McKenzie; Walker, Jane B.; Verster, Anna J.M.; De Kock, V.E.
Electron microscope studies on the three ephemeral fever virus (EFV) strains EFVl, EFV13 (South Africa) and Japanese bovine epizootic fever (BEF) virus (Strain YHK) showed that the normally cone-shaped South African EFV, may, under certain circumstances, appear bullet-shaped while the Japanese EFV which is usually bullet-shaped may appear as a cone. These findings suggest that ephemeral fever virus is pleomorphic.

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