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Tribulus Ataxia or 'Cathead Staggers'

C.A. Bourke, B.V.Sc. (HONS), H.D.A. Veterinary Research Officer, Department of Agriculture, Regional Veterinary Laboratory, ORANGE

INTRODUCTION

Prior to September of 1981 the state of knowledge regarding this disease was roughly as follows:

In 1938, a curious hindquarter ataxia was observed in some mobs of sheep in the Coonabarabran district. Since that time several minor outbreaks of this disorder have been seen in the Coonabarabran and Dubbo areas. These outbreaks seem to have followed years of prolonged drought and seem to have been associated, in many instances, with the grazing of a plant identified as Tribulus terrestris and locally referred to as Cathead. The sheep involved tend to have been Border Leicester and their crosses and were generally in older rather than younger age groups. The disease state was observed to be slowly progressive but essentially irreversible, with eventual death due to thirst or misadventure. Post-mortem findings and laboratory results, including histopathology, were negative or non specific. The causal agent and its effects in the animal were generally regarded as unknown. Overall, the disease, even 42 years after its initial occurrence, was still considered a mystery.

In the face of a fresh outbreak in 1981, we at the Orange Veterinary Laboratory decided it was time this disease was more thoroughly and persistently investigated, so that the mystery might finally be solved. Having had previous field experience working in the Coonabarabran District, the task of researching the problem fell to me. This paper represents the results of these investigations to date (March, 1982).

DISTRIBUTION AND INCIDENCE OF THE DISEASE

The following lists are a compilation of all officially recorded cases of this disease, useful flock incidence figures are only available for the 1981 outbreak.

Recorded cases for 1981 outbreak

Note: first cases started to appear in March, 1981,

Owner	Ewes run	No. lost	No. still affected	Breed	Occurrence of last new case
WR Merrygoen	780	350	30	Merino	February, 1982
	850	760	90	Border cross
HB Mendooran	550	12	10	Border cross	February 1982
GHV Gilgandra	700	280	30	Border L.	February, 1982
	600	10	-	Merino
SK Merrygoen	350	160	20	Border L.	February, 1982
	500	-	-	Merino
SS Merrygoen	350	200	9	Border cross	-
	250	-	-	Merino
HK Gilgandra	1300	14	10	Border Cross	January, 1982
TL Gilgandra	3000	32	45	Merino & Border cross	-
MD Mendooran	450	15	-	Corriedale	September, 1981
HB Mendooran	900	100	15	Border C. Merino	February, 1982
	700	-	-	Merino
GG New Mollyan	2500	70	20	Border C.	February, 1982
YG Merrygoen	1500	25	5	Border C.	February, 1982
HF Mendooran	470	40	6	Border C.	September, 1981
QF Mendooran	250	70	-	Border C.	-
PL Neilrex	-	few	-	Merino	-
WG Weetalibah	700	100	-	Border C.	-
	160	-	-	Merino
LM Neilrex	700	30	few	Border C.	-
KB Mendooran	850	110	20	Border C.	February, 1982
MG Mendooran	800	20	-	Merino	-
KB Binnaway	-	16	-	Border C.	-
RB Mendooran	480	300	-	Border C.	-
BB Binnaway	-	few	-	Border C.	-
TH Bugaldie	105	10+	-	Border L.	-
WU Weetalibah	450	20+	-	Border C.	-
PP Binnaway	850	30+	-	Border C.	-
HR Curban	700	25+	-	Border C.	-
FW Obley (Yeoval)	150	67	-	Border C.	-
MB Ballimore	-	12+	-	Border C.	-
OW Trangie	-	few rams	-	Border L.	-
Lyons, 'Geenobby', Wellington	346	35	15	Border C.	-
	30	7	-	Southdown
	130	5	-	Border L.
Olsen, 'Inverness', - Wellington	-	few	-	Border L.	-

Previous recorded cases of the disease

Year	Owner	Breed
1938/39	NG New Molyan	Border Leicesters & Border Crosses
	GW Mendooran	Merinos
1944/45/46	Park of Narrabri	Dorset Horns
	MC Binnaway	Border Leicesters & Border Crosses
	GW Mendooran	Corriedales
1947	KB Mendooran	Border Crosses & Comebacks
	RB Binnaway	Border Crosses
	TG Binnaway	Corriedales
	MC Binnaway	Border Leicesters & Border Crosses
	JH Birnaway	Merinos
	WD Tooraweenah	Merinos
	NG New Molyan	Border Leicesters & Border Crosses
	AB Gilgandra	Border Leicesters & Merinos
	MI Gilgandra	Border Crosses
	SK Gilgandra	Merinos
	GW Mendooran	Corriedales & Merinos
	GB Mendooran	Corriedale cross
	BS Merrygoen	Merino
	GB Dubbo	Border Leicesters
	DB Dunnedoo	Border Leicesters
	DX Bylong (Mudgee)	Merinos
1954/55	TB Mendooran and other holdings not on official records.	Border Crosses
1965/66/67	JB Tooraweenah	Breed not recorded
	LR Binnaway	Border Leicesters & Border Crosses
	MS Warrumbungle	Border Crosses
	RG New Molyan	Border Crosses
	SG Binnaway and other holdings for which the official records have been lost.	Breed not recorded

Official records are only a reflection of those holdings that experienced significant problems with this disease, many owners with only small losses did not report their problem. Further official records tend to reflect the number of animals affected in the middle of an outbreak period rather than the total for the end of an outbreak period, i.e. they understate the incidence.

After reviewing all available records pertaining to each affected property listed, the following facts emerge.

1. Every affected mob of sheep grazed good growths of Cathead for a period of 6 weeks or more during the summer/autumn period of the year in question.

2. Clinical cases may occur while the Cathead is being grazed but in general they do not appear until several months after sheep are removed from it.

3. Clinical cases may develop slowly over weeks or months or they may develop rapidly over days or even hours.

4. Many new cases of the disease seem to appear following a period of stress, for example, the stress of shearing, lambing or droving. Frequently, clinically normal animals become obviously ataxic within a matter of hours of mustering and droving.

5. Although sheep of any sex may be affected, the age of animals seems important, since most reports involve sheep that are four tooth or older. However, cases as young as 10 months old have been seen.

6. The breed of sheep involved is very significant. Cases have occurred in Dorset Horns, Southdowns, Merinos, Corriedales, Border Leicesters, Comebacks and Crossbreds, but there would appear to be a real 'Resistance' on the part of Merinos and a real 'susceptibility' on the part of British Breeds and their crosses. The Border Leicester would appear to be particularly susceptible and this involves more than just a grazing preference for Tribulus plants.

7. Affected properties in general experience an incidence of less than 5%, however, a number of holdings experience losses of 30% to 60%.

8. An endemic area for this disease exists along a corridor of land that roughly follows the Castlereagh river and runs from Binnaway to Mendooran and then on to Gilgandra. Approximately 75% of all recorded affected properties lie within this zone.

The total area of distribution of affected properties is much broader than first imagined and includes the following locations outside the endemic area - Narrabri, Curban, Trangie, Yeoval, Bugaldie, Tooraweenah, Dubbo, Mudgee, Warrumbungle, Dunedoo & Wellington.

10.The causal agent of this disease does not appear to be a microorganism, a helminth, an arthropod or a genetic factor. Consequently, it seems most likely that this disorder results from a toxicity or a deficiency state.

DISTRIBUTION AND INCIDENCE OF THE PLANT TRIBULUS TERRESTRIS

In looking for a factor that was common to all affected mobs regardless of their location, year of occurrence, etc., it became increasingly clear that the only consistent common factor was the grazing of Tribulus. I should stress, not just grazing some Tribulus for some brief period, but more specifically, the grazing of pastures which consisted largely or even solely of the plant Tribulus for periods of six weeks or longer. With this fact in mind, a detailed study of the plant was made to see how it might correlate with the distribution and incidence of the disease. The findings from this study were very encouraging.

The broad description Tribulus terrestris is applied by botanists to all plants of the Cathead type when in fact there is good evidence to suggest that, in fact, several species of this genus are involved, and even stronger evidence to support the concept of more than 50 types or varieties within the species terrestris itself. Consequently, even though any plant type of this group will be consistently identified by the Government Botanist as Tribulus terrestris, this identification, though correct in itself, is very misleading because, in fact, it is a generalisation.

In Australia alone there are 19 different types of Tribulus terrestris, and of great significance to this disease, 18 of these types are in fact native to Australia and only one of them has been introduced from overseas. Fortunately, at least one plant ecologist has gone to the trouble of mapping out the limit of distribution of this introduced form, and also a rough history of its introduction and spread can be compiled from various other sources. In brief, the existence of the introduced form was first recorded in N.S.W. in 1895 and again in 1910; it was incorrectly thought to be a native to this State and farmers were warned that it was starting to spread. By 1920, it had reached the south east corner of South Australia and the hotter northern districts of Victoria. Between 1920 and 1935 it seemed to be making great strides in some parts of the North West Slopes of N.S.W. and, in fact, it became more successful in its establishment in that district than in any other part of Australia. Concurrently to all this spread in the Eastern States, the plant seems to have gained entry to the south western districts of Western Australia, radiating out, it would seem, from the port of Fremantle. I feel certain it was more than pure chance that the first cases of Cathead Staggers were seen in the North West slopes of N.S.w. in 1938, and I attribute them directly to the very successful establishment of the introduced type of Tribulus terrestris in that region. In fact, regardless of the original country of origin of this type of Tribulus (and that could have been any one of five continents where Tribulus is known to occur), it is hard to imagine that the plant could have been more successful than it has been on the south eastern side of the Warrumbungle Mountains. Many districts may report growths of Cathead but very few can boast of thousands of acres of almost pure Catheads of the introduced type for two to three months of the year, following periods of drought. This is in fact what happens in the endemic area of the Cathead Staggers disease.

Further north and west of that district, large growths of Tribulus can occur, but the type of plant involved is the Australian native form and in fact the locals even change the common name, referring to it as Yellow Vine. Why was the introduced type of Tribulus so successful in the Coonabarabran district and why did its 'golden period' for establishment occur from 1930 onwards to the present day? To answer these questions, one needs to understand the growth habit and requirements of the plant itself. In brief, they are as follows:

1. Tribulus is a long growing plant that does not compete very well with other taller growing vegetation; for large scale establishment it requires areas denuded of other vegetation, i.e. bare areas in summer. Cultivation in general and winter cereal cropping in particular assist greatly in this regard, as does overgrazing of areas by sheep. The distribution of the plant correlates well with the sheep-wheat zone of N.S.W.

2. Tribulus requires prolonged high light intensity and prolonged elevated temperatures for its optimum growth, consequently, it is a summer growing species and is particularly suited to the sunlight and temperature conditions of North Western N.S.W.

3. For optimum growth, Tribulus has a higher water requirement than one might imagine. However, it responds very poorly to prolonged damp conditions; what it does require are short heavy downfalls of rain several weeks apart. Typically, it responds well to the reliable summer storm pattern in the vicinity of the Warrumbungle uplift. In southern N.S.W., it does not experience this weather pattern and as a consequence its growth is sub-optimal.

4. Contrary to popular belief, Tribulus requires good fertility levels for optimum growth. It has a relatively high nitrogen and phosphorus requirement and consequently responds well on cultivation areas where leguminous leys have been used and superphosphate applications are regular. The plant also prefers deep soils of good texture that facilitate the rapid establishment of its large tap root.

5. Tribulus is basically an annual (although some perennial habit is displayed), with a prolific seeding habit and very rapid growth. The plant can progress from germination to flowering to seed set within a period of 21 days. Tribulus is in effect in all stages of growth at the one time. However, sheep tend to selectively graze the leaves first, the stems second and the seeds last. Individual animals can be observed to eat all three parts of the plant at the same time.

6. Optimum establishment of Tribulus on a particular area is dependant on high germination rates, so that the Tribulus plants can better compete with other weed species by the very rapid establishment of high plant densities of themselves. These high germination rates are strongly favoured by the preconditioning effect of prolonged hot dry weather, i.e. by the preconditioning effect of drought. In normal years, germination is more patchy.

7. The distribution of the seeds of the introduced form of Tribulus by the wool of sheep, or in the dung of animals, or on the feet of animals is only a minor means of spread. The most successful means of distributing the seeds of this plant is by the rubber tyre. In this respect, the tyres of tractors, trucks, cars and motor bikes have been particularly beneficial to this plant's establishment.

CLINICAL SIGNS OF THE DISEASE

Sheep affected by Cathead Staggers generally develop a progressive hindquarter ataxia over a period of weeks or months. In some individuals, the progression is quite rapid, particularly if they are stressed. In other individuals, the initial onset of obvious clinical signs occurs quite suddenly, e.g. in a matter of hours following mustering and droving. However, in the more slowly developing cases the following clinical progression will be seen.

1. A slight stiffness or irregularity in the gait of the hind legs, the occasional dragging of the toe instead of lifting the foot up and putting it down cleanly.

2. Gradually the hind legs tend to bow outwards and a noticeable flaccidity develops in the hindquarter.

3. Toe dragging becomes more frequent and drag trails can be seen in the dust. The hock joints tend to be held in more constant flexion.

4. Affected animals start to 'crouch' and 'sway' in the hindquarter. That is, the flexion of the hocks causes the rump to drop, i.e. crouch down, and possibly impaired joint proprioception results in the back legs leaning to one side, i.e. sway over.

5. By this stage, animals may at times bow in at the hocks or assume irregular postures with their hind legs.

6. Forward progression can be achieved in one of two ways, either by 'bounding' or 'crabbing'. Bounding is often favoured by Border Leicesters; here the animal arches its back and moves both hind legs simultaneously, such that the hind feet are carried well forward and hit the ground together. Crabbing is generally favoured by Merinos or Crossbred types; here the animal moves its front legs in a forward direction and its hind legs in a sideways direction, the backbone curves to one side, and the overall motion is that of a crab.

7. Affected animals will support themselves by leaning against fences, stumps, posts or even against each other. They spend progressively longer periods lying down, and the difficulty in raising themselves up again, increases.

8. Weeks, or more typically months, after the first clinical signs, affected cases are found in lateral recumbency, unable to raise themselves at all, and unable to remain standing even if manually lifted up. The animal will continue to eat any feed that is within reach, but within a matter of days will die of thirst, crow-pick, flystrike etc.

It is important to realise that even though this disease is in essence a flaccid paresis of the hind legs with an incoordination component, some few individuals do at times display a degree of spasticity or a degree of foreleg involvement. Any foreleg involvement appears as a quick jerky action of the front legs, the legs appearing to be flicked forward in a mild form of 'goose stepping'. Bounding actions should not be misinterpreted as spasticity nor should the lying down in lateral recumbency with all legs extended.

Urination and defaecation appear to remain normal throughout the course of the disease. However, the musculature of the hindquarters gradually undergoes atrophy, and various areas of the body may show bruising or rubbing due to mechanical trauma.

PATHOGENESIS OF THE DISEASE

An accurate assessment of just exactly what is happening to sheep affected by this disease is at this stage somewhat hypothetical. However, my current understanding of the disease, based on all the facts available at present, favours the following assessment.

Prolonged ingestion by sheep of large quantities of an introduced form of the plant Tribulus terrestris appears to result either directly or indirectly in a neurotoxicity state. The cycle of events in any single outbreak year is as follows:

a. Feb., Mar., April, May - during this period summer storms and bare paddocks result in the growth of large quantities of Tribulus, such that a minimum of 4 to 8 weeks of solid grazing by sheep takes place and a few ataxic cases develop.

b. June, July - although some clinical cases are already apparent, the majority of them are yet to appear; a few more develop during this period.

c. Aug., Sept. - the development of fresh clinical cases reaches a peak during this period, probably in association with lambing and shearing stresses.

d. Oct., Nov., Dec., Jan. - small numbers of fresh clinical cases will appear at any time during this period, i.e. even up to 9 months after removal from the toxic plant.

Allowing for the fact that some new toxicities are slow to develop clinically, there is still a very protracted period between the removal of some animals from the plant and the appearance of first clinical signs in them i.e., up to nine months. Allied with this is the phenomenon that clinical cases can be precipitated rather suddenly, for example the stress of mustering and droving apparently clinically normal sheep can produce very obvious ataxia cases in a matter of hours. These phenomena could perhaps indicate an indirect toxicity. For example, does the plant toxin interfere with say liver function, resulting in the liver cells themselves producing a neurotoxin which is accumulated first and then suddenly released at a later period in response to stress? Alternatively, is the neurotoxin in the plant itself, but does it need to accumulate in say the fat depots of the body first, until sufficient quantity is available for later sudden release and effective neurotoxicosis? Here again, stress may be a factor in instigating the fat mobilisation.

If in fact, the neurotoxicity is actually direct, then the nine month delay period possible is still reasonably consistent with other recorded plant toxin ataxias similar to Cathead Staggers. Coyotillo Ataxia in goats, for example, has a maximum delay period of four months. In Cycad Leaf Ataxia in cattle, the recognised period possible for maximum development of spinal cord lesions is six months. This disease incidentally on histopathology, is remarkably similar to Cathead Staggers. In Neurolathyrism in humans it can take up to six months for clinical signs to develop and again the histopathology of that disorder is similar to Cathead Staggers.

One might well ask why is this disease assumed to be a primary nervous disorder rather than say a primary muscle problem. Here we must rely on histopathological findings and observation of the progressive development of clinical signs. Admittedly, pathology of the musculature is a feature of this disease in advanced cases, however, histopathological studies of early cases are strongly suggestive of problems in the C.N.S. before there are any problems in the musculature.

When one considers carefully the clinical signs of typical cases of Cathead Staggers the following conclusions can be drawn -

1. The animal has lost the use of some groups of Extensor Muscles in the hind legs and as a consequence, Flexor Muscle activity becomes dominant. Hence, the classical 'crouched' stance.

2. The animal has lost the use of some muscle groups in the hind legs that act specifically in joint fixation and stabilisation. Hence the classical 'swaying' stance.

3. The animal typically demonstrates a paresis that is flaccid not spastic, but very occasionally spasticity does occur.

4. Foreleg involvement is an unusual complication but it does occasionally occur.

5. Affected animals never adopt the 'dog sitting' posture so typical of many nervous disorders of the hindquarters.

Electromyography carried out on only two typical cases has demonstrated a partial denervation of the Semitendinosus and Semimembranosus muscle groups. Considering that the action of both these muscle groups in the sheep involves extension of the hip, stifle and hock joints, this finding is very consistent with the clinical signs. It is worthy of note that the Semitendinosus also functions as a flexor of the stifle joint when the limb is in the free non weight bearing position, possibly this reflects one reason for affected animals resting in lateral recumbency rather than the more normal sternal recumbency.

Nerve innervation for the Semitendinosus and Semimembranosus comes from the Tibial nerve, a branch of the Sciatic, which has its origins at the level of L6, L7 and S1. Degenerative changes at the level of the sciatic nerve are a reasonably common finding in this disease and this is consistent with the partial denervation concept. Marked atrophy of some hind leg muscle groups, another common finding, is also consistent with denervation.

The specificity of a flaccid paresis is such that a lower motor neurone (spinal cord) disorder would seem at first extremely likely; degenerative changes observed in some ventral horn neurons in the cord grey matter would be consistent with this concept, as would an apparent lack of degenerative changes in the descending motor pathways in the cords of many affected cases. However, mild brain lesions are a regular finding, mild degenerative changes in the ventral columns of the cord white matter (i.e., an area specific for some descending pyramidal tracts) occur in some cases, foreleg involvement can occur, and a degree of spasticity is also possible. In other words, findings of this nature are very suggestive of an upper motor neurone (brain) problem. However, for a brain lesion to result in a flaccid paresis of the hind legs would seem to require a problem specifically involving the pyramidal (corticospiral) motor system, and more precisely, the hind limb sub area of that system.

Since mild degenerative changes in the fibre tracts of the dorso-medial columns of the cord are the most consistent cord findings, then the fact that they are specific for the ascending sensory pathways from joint proprioceptors to the nuclei of the medulla, seems rather significant. Further, because the fibre tract degenerations seen are frequently describable as Wallerian in type, then presumably the fibre degenerations are secondary to a primary degenerative state at the level of the cell bodies (i.e. nuclei). With this in mind, fairly regular findings of scattered degenerate neurones in the medulla and brain stem may well prove significant. Because a sensory input deficit can give rise to a motor output deficit, the significance of ascending sensory pathway pathology should not be overlooked.

When one considers the histopathology of the brain in more detail, the finding of a moderate sub acute choroiditis in some affected animals is strange, and may imply that the choroid plexus of the ventricles is responding to the passage across the blood/CSF barrier of an irritant substance. It would also presumably suggest increased production of CSF. Further to this concept, examination of the brains of 3 animals autopsied after being fed a 100% Tribulus diet for 30 days revealed a curious marked focal oedema specifically in sections taken at the level of the Mammillary Bodies, and at no other level. The location of this oedema was essentially the white matter of the lateral brain stem but it involved portion of the choroid plexus of the lateral ventricle, the posterior aspect of the caudate nucleus, the lateral aspect of the thalamus and aspects of the lateral geniculate body. Significant numbers of fibre tracts in the region were degenerate and there was an overall appearance of microscopic 'holes'. However, concurrent jaundice in these animals makes accurate interpretation of the findings difficult.

The histopathology of the nervous system in affected individuals varies significantly from case to case; this may simply reflect stages of progression of the disease. Some cases show pathology at the level of the brain, spinal cord and peripheral nerves, whilst others show changes at only one or two of these levels. The changes seen are generally mild but in some cases moderately severe. As a consequence, the investigating pathologist must examine all three levels of the nervous system in great detail and look very carefully or significant changes will be missed. Cases that are the most advanced clinically tend to show the least pathology microscopically. Histopathological studies into this disease are far from complete at present, and the following description is a composite one that draws on the findings from several cases rather than being typical of any one individual case. The histopathology of this disease is not pathognomonic for it alone.

BRAIN

Focal areas of micro-gliosis in the cerebral cortex and brain stem. Scattered degenerative neurons in the caudate nucleus, cerebal cortex, brain stem (including caudate nucleus and thalamus) and the medulla. Mild degenerative changes in the white matter of the brain stem and medulla. The exact locations of all these changes is requiring of further more detailed studies.

CORD

Degenerative changes involving the dorsal columns, in particular the dorso-medial component, mild cervically progressing to moderately severe in the lumbar segments. There is also ventral column involvement in some individuals. These changes may involve segmented demyelination and/or Wallerian degeneration. Scattered degenerate neurons in the ventral horns of the grey matter, more typically at the thoracic level but also cervically, and occasionally at the lumbar level.

PERIPHERAL NERVES

Mild to moderately severe degenerative changes involving more typically the sciatic and femoral nerves. These changes generally involve a segmental demyelination and vary in degree from bundle to bundle. In some individuals these changes also appear in the brachial nerves.

Although this disease appears to be a primary nervous disorder an appreciation of the secondary muscle effects are paramount to an understanding of how the clinical signs progress and why the C.N.S. pathology appears milder as the clinical signs become more advanced. The result of initial degenerative changes in the C.N.S. and peripheral nerves is partial denervation of some muscle groups, in particular those of the hind legs, possibly specifically the semimembranosus and the semitendinosus. The ability to use these muscles is impaired and they gradually atrophy. The normal bias in favour of extensor muscles subsides, the joint ligaments and tendons as a consequence assume greater control, and an ever increasing flexion of the hock and stifle joints results. Atrophy of some muscle groups, excessive use of others, and a general inability to forage, result in an ever increasing generalised muscular weakness. The nervous component of this disease is only limited and terminates relatively early in the course of the disease. However, it is the permanent nature of these nerve tract degenerations that results in the slowly progressive muscular disorder that follows.

TOXICOLOGICAL ASPECTS OF THE DISEASE

Members of the plant group Tribulus terrestris are known to be capable of causing at least two disease states in sheep.

1. Nitrate/Nitrite poisoning. This results in fairly rapid deaths when hungry mobs of sheep are suddenly given access to reasonable quantities of Tribulus. The Nitrate content of Tribulus plants in terms of KNO₃(%) of dry weight ranges from 0.12 to 7.12% depending on growing conditions.

2. Geeldikkop. This condition is of particular concern in South Africa but it can occur under Australian conditions as well. It involves a primary hepatotoxicity with secondary jaundice and photosensitisation. It is currently regarded as a mycotoxicosis that involves ingestion of Tribulus and at the same time ingestion of the saprophytic fungus Pithomyces chartarum. There would seem to be a specific toxic principle in the plant itself, quite apart from the sporidesmin toxin produced by the fungus. The disease does not occur unless both toxins are operative. There would appear to be good evidence to suggest that the plant is only toxic when it is growing vigorously.

Neither of these two diseases have a hindquarter ataxia component at any stage.

Whilst investigating Geeldikkop, South African workers screened Tribulus plants for possible toxic factors. The only potential toxins uncovered, other than Nitrite, were a group of steroidal saponins that occurred at a level of 0.5% to 2.0% of the dry weight of the plant. These saponins were shown to give the following sapogenins on hydrolysis, DIOSGENIN, RUSCOGENIN, GITOGENIN and 25D - SPIROSTA-3; 5-DIENE. No other potential toxins were uncovered, however, this analytical work was done prior to 1960 and it is reasonable to expect that substances from less obvious chemical groups may have been missed. Some of these saponins were demonstrated to be hepatotoxic but there appears to be no suggestion that they might be neurotoxic. In fact, no saponin containing plant material of any description has ever been proven to be neurotoxic to herbivorous mammals when given by the oral route. It would appear that ruminants are particularly good at breaking down saponins during the digestive process. Inflammation of the gut wall would seem to be an essential prerequisite to facilitate appreciable transport of saponins across the gut wall and into the blood stream. Because at least one type of saponin has been proven capable of prolonged cholinesterase inhibition, this aspect was followed up by subjecting the bloods of 6 normal and 6 ataxic sheep to plasma and R.B.C. cholinesterase activity tests. No significant difference could be seen between the two groups. If saponins are the toxic principle in Cathead Staggers, then the assumption at this stage would be that the C.N.S. effect was secondary to a primary liver effect.

A review of the available literature on all recorded plant toxicities capable of causing chronic irreversible hindquarter ataxias similar to Cathead Staggers, has been undertaken. The intention being to obtain an indication of what toxic chemical groups might be active in diseases of this nature. In addition, it was hoped that more light might be cast on the histopathology and pathogenesis of this type of disease. The following is a summary of that review.

1. 'Guajillo Wobbles', seen in sheep in Texas during prolonged droughts, an irreversible hindquarter ataxia due to the ingestion of the shrub Acacia berlandieri. Toxic principle is the aliphatic amine N-methyl A-phenylethylamin.

2. 'Coyotillo Ataxia' seen in goats in parts of the United States during drought situations. It results from the ingestion of the shrub Karwinskia huobolditara.

3. 'Cycad and Macrozamia ataxia', seen in Australia, New Guinea and the Caribbean in cattle generally, but occasionally in sheep. An irreversible chronic hindquarter ataxia due to the ingestion of Cycad and Macrozamia palms. Toxic principle is believed to be a compound with a molecular weight exceeding 1,000.

4. 'Neurolathyrism', seen mainly in India in humans. This is a chronic irreversible paresis of the lower limbs due to the ingestion of chick pea seeds (Lathyrus sp.) and certain vetches (Vicia sp.). Toxic principles are believed to be the non protein amino acids β-N-oxalyl-α,β-diaminopropionic acid and α-N-oxalyl-^∝βdiaminobutyric acict as well as the compound β-cyano-L-alanine.

5. 'Chronic Milkvetch Paresis', seen in sheep and cattle in Colorado, Utah and Wyoming (U.S.A.), it is a hindquarter ataxia that results from the ingestion of aliphatic nitro compounds. The toxic principle in Miserotoxin, i.e. a glycoside of 3-nitro-1-propanol which is split during ruminant digestion and the toxic portion liberated. Milkvetch is a species of Astragalus.

6. 'Tropical Ataxic Neuropathy', seen in humans in Nigeria, it is an irreversible progressive ataxia of the lower limbs that results from the ingestion of Cassava (Manihot esculenta). The toxic principle is the cyano-genetic glycoside Linamarin. This is essentially a Nitrile compound that has been synthesised from an amino acid.

Thus the potential toxic groups involved in diseases of this type include:

1. Aliphatic amines, i.e. Methyl Phenylethylamine.

2. Compounds with molecular weights of the order of 1,000.

3. Non protein amino acids, i.e. oxalyl diaminopropionic acid and oxalyl diaminobutyric acid.

4. Aliphatic nitro compounds, i.e. nitropropanol.

5. Cyano substances, i.e. the cyanogenetic glycoside Linamarin and Cyanoalanine.

Even though the histopathological changes in the C.N.S. described for each of the above ataxias demonstrate close similarities between one another, and even though they correlate reasonably well with those of Cathead Staggers, it would appear that the actual toxins themselves demonstrate distinct differences in their methods of action. That is the nett effect on cell bodies and axons is the same, i.e. a 'dying back' degeneration process. The means of achieving it can vary from:

a. interference with methionine and cysteine pathways in the case of Linamarin and Cyanoalanine.

b. primary interference with urea synthesis in the liver followed by a secondary C.N.S. effect presumably involving a chronic ammonia toxicity, in the case of Diaminobutyric acid.

c. accumulation in brain tissue followed by lysosome damage, enhanced protein degradation and consequent ammonia production, in the case of Diaminopropionic acid, or alternatively, this toxin can interfere with the glutamate transport system resulting in an accumulation of glutamate in the environment of nerve synapses, glutamate being potentially neurotoxic.

d. in the case of phenylethylamine, speculation exists as to its involvement in the production in the animal's body of a methylated derivative of serotonin, a substance known to be neurotoxic, or alternatively to the involvement of phenylethylamine itself in the precipitation of a dopamine deficiency, in certain groups of brain nuclei, of the type seen in human Idiopathic Parkinsonism.

In an effort to gain more insight into the mechanics of Cathead Staggers at the physiological level, a routine screening was carried out on blood samples from 42 ataxic and 42 'normal' sheep from four of the affected properties. These sheep were a balanced mixture of Border Leicesters, Merinos and Border Merino cross breeds. The sample involved roughly equal numbers of early and advanced ataxia cases. The following table indicates parameters examined and group mean results obtained.

Parameter	Normal Value	Ataxic Group	Normal Group
Serum Calcium	>2.2m mol/L	2.7	2.7
Serum Magnesium	>0.7 mol/L	0.9	1.0
Serum Phosphorus	>1.3 mol/L	1.8	1.9
Serum OCT	<20 i.u./L	24	25
Serum CPK	<30 i.u./L	548	87
Serum GOT	<100 i.u./L	97	74
R.B.C. x 106	8.0 - 16.0	12.1	11.7
P.C.V.	24 - 50	39	39
P.P.	6.0 - 7.5	8.8	8.3
W.B.C. x 103	4.0 - 12.0	11.7	9.1
Neut (%)	10 - 50	64	40
Lymph (%)	40 - 75	38	56
Eosino (%)	0 - 10	3	5
Glutathione Peroxidase (Selenium indicator)	>150 i.u./gHb	741	744

There were no significant differences observed between both groups, or any sub groups within each group, for any of the parameters listed, except for CPK levels and the Neutrophil/Lymphocyte ratios. The ataxic group had high CPK values and elevated neutrophil counts. Both of these findings are probably directly attributable to secondary muscular pathology. The other parameters scored values that were essentially within the normal range in both groups.

Feeding trials with Tribulus, in an effort to demonstrate its potential toxicity to sheep have mainly been concerned with the disease Geeldikkop. In the early days of researching this disease (Circa, 1920), South African workers demonstrated the plant's Nitrate potential but had difficulty in experimentally reproducing Geeldikkop on a consistent basis. By 1980, the ability to experimentally produce Geeldikkop on a repeatable basis was achieved by concurrently feeding the plant and the fungus. At no stage in the more than sixty years of Geeldikkop research in South Africa has a hindquarter ataxia, associated with the ingestion of Tribulus, been demonstrated. This fact should emphasise the point that the following criteria are essential in the design of a feeding trial aimed at demonstrating the potential neurotoxicity of Tribulus terrestris.

1. The variety of Tribulus being fed should be that which, in Australia, is loosely referred to as Tribulus Introduced.

2. The plant must be fed in the green, well grown state. Any limited storage period should involve refrigeration at all times, after the plant is harvested, in containers sealed against moisture loss.

3. The plant must constitute the whole diet of the animals being fed and be fed at a level in excess of the maintenance requirement.

4. The plant must be fed for a period of at least 8 weeks.

5. The breed of sheep selected for the trial should preferably be Border Leicesters or at least a British (Mutton) Breed rather than Merinos.

6. The sheep used should preferably be adult and certainly not at the lamb or weaner age.

7. The trial sheep should be kept under observation for the development of clinical signs for a period of at least 9 months after the period of experimental feeding is completed.

In Australia, very few feeding trials have been carried out with Tribulus terrestris. With the exception of the very recent ones, all of the trials were attempting to reproduce Geeldikkop and the possibility of a Hindquarter Ataxia disease potential was not even thought of. The following is a list of all known feeding trials with this plant in Australia.

1. Location - Glenfield.

(A) Period of feeding - February 15, 1924 to March 7, 1924, total 22 days. 2 Merino ewes with 2 lambs at foot; feed used was a mixture of Tribulus Introduced and Tribulus Australian Native; the feed was harvested at Merriwa (a distance of 6 hours by train), held in chaff bags for a day or so and then railed to Glenfield, where it was further held and gradually fed out. Heat stressing and 'Sweating' of the plant material would have been inevitable. Findings were negative (Seddon).

(B) Period of feeding - February 27, 1924 to March 31, 1924, total 33 days. One Merino ewe with lamb at foot; feed used was Tribulus Introduced; feed was harvested and handled as for A. Findings were negative (Seddon).

(C) Period of feeding - February 27, 1924 to April 4, 1924, total 37 days. One Merino ewe with lamb at foot; feed used was Tribulus Australian Native; feed was harvested and handled as for A. Findings were negative (Seddon).

2. Location - Merriwa.

Period of feeding - March 5, 1924 to March 31, 1924, total 26 days, 4 ewes and lambs probably of the Merino breed; half were fed Tribulus Introduced and half were fed Tribulus Australian Native; feed was harvested twice a week and stored in chaff bags (not refrigerated). Plant material would have been heat stressed and 'sweated' during storage. Findings were negative (Seddon).

3. Location - Nyngan.

Period of feeding - January 26, 1926 to February 9, 1926, total 15 days, 2 Merino lambs fed fresh cut green Tribulus of an uncertain varietal type. Findings were negative (Seddon).

4. Location - Yanco.

Period of feeding - February 25, 1929 to April 20, 1929, total 54 days. 1 aged Merino ewe and 3 Merino hoggets; fed well grown green fresh Tribulus cut daily from the irrigation channel overflows; varietal type uncertain but Tribulus Introduced known to have arrived in this district in 1892 and spread since that time. No evidence to suggest the presence of Tribulus Australian Native. Findings were very significant, i.e. no evidence of Geeldikkop was seen but an incidental finding was the development of 'a weakness in the hind legs' of one of the hoggets (Rose).

5. Location - Orange.

Period of feeding - January 18, 1982 to February 18, 1982, total 32 days. 3 Border Leicester weaners; fed well grown green fresh Tribulus Introduced, cut once a week, refrigerated at all times, held in sealed containers and fed out daily ad lib. Findings very significant. By day 17, two animals had started intermittent toe dragging in the hind legs; by day 27, all three animals appeared to be showing a vague hind leg weakness, by day 29 appetite was depressed; by day 32 all three animals were inappetent, depressed and clinically jaundiced. The animals were returned to normal feed (lucerne hay) but they failed to recover. One was sacrificed and the other two had died within 7 days. Autopsy of all three revealed severe jaundice and histopathology demonstrated lesions specific for Geeldikkop. The animals were fully housed at all times, hence photosensitisation was not a feature. Culture of Tribulus plant material for fungi present revealed Cladosporium cladosporioides, Stemphyllium botryosum, Curvularia affinis, Alternaria alternata, Phoma sp., and a single colony of Pithomyces chartarum. The finding of a specific marked focal lesion in the brain stem on histopathology was suggestive of a possible primary C.N.S. lesion for Cathead Staggers (Bourke).

6. Location - Orange.

Period of feeding - February 20, 1982 and still continuing. Anticipated total period 60 days. 2 adult Border Leicester rams; feeding regime as for previous trial (i.e. 5). No significant findings as yet (Bourke).

ACKNOWLEDGEMENTS

The assistance of the following people in this investigation is gratefully acknowledged. Geoff Wise and Frank Nottle for various field aspects, Brian Farrow for the electromyography, Alex Nickandrow for fungal cultures, Bill Hartley for his histopathology during the 1960's outbreak, and various members of staff at the Regional Veterinary Laboratory, Orange.

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