The role of Penicillia in ryegrass staggers

Tremorgenic strains of Penicillium verrucosum var cyclopium, P canescens, P janthinellum, P novaezeelandiae and P estinogenum were isolated from the faeces of 15 of 23 affected sheep and cattle in eight of nine field outbreaks of ryegrass staggers. One tremorgenic strain of P griseofulvum was isolated from the faeces of one of 25 sheep grazing in unaffected flocks. Tremorgenic strains of P verrucosum var cyclopium, P canescens, P janthinellum and P estinogenum were also isolated from the A horizon of New Zealand soils. Since a large proportion of experimentally dosed live P verrucosum var cyclopium died during passage through the gut, the faecal evidence from naturally staggering animals suggests that at least some outbreaks of ryegrass staggers are caused by tremorgenic Penicillia and that their source may be soil.     

The evaluation of endophyte toxin residues in sheep fat

Abstract

AIM: To monitor changes in concentrations of lolitrem B and epoxy-janthitrems in the fat of sheep grazing perennial ryegrass infected with wild-type- and AR37-endophyte, respectively, during the time of year when ryegrass staggers would be expected to be observed.

METHODS: Ten 5-month-old lambs with no previous exposure to endophytes were grazed on either wild-type (containing lolitrem B, n = 5) or AR37 (containing epoxy-janthitrems, n = 5) endophyte-infected perennial ryegrass (Lolium perenne L.) pastures between October 2008 and June 2009. Animals were regularly assessed for ryegrass staggers using the Keogh scale (0 = no signs, 5 = severe tremors). When a score of > 3.5 was observed animals were removed from the treatment pastures for 1 month. Fat biopsy samples were taken from each animal at approximately monthly intervals and analysed for endophyte metabolites using high performance liquid chromatography (HPLC) methods developed during this study. Regular herbage samples were also taken and concentrations of endophyte metabolites measured.

RESULTS: Efficient and reproducible methods to analyse both lolitrem B and epoxy-janthitrems in fat were developed. Concentrations of lolitrem B and epoxy-janthitrems in herbage and in sheep fat increased from late November to peak in mid-February. Ryegrass staggers was observed in both groups of sheep at this time. Following 1 month of grazing non-infected pasture mean concentrations in fat of lolitrem B decreased by 43% from 61.8 to 35.3 ppb, and of epoxy-janthitrems by 38% from 1032.0 to 639.5 ppb. Maximum concentrations in herbage of epoxy-janthitrems (35.7 ppm) were higher than of lolitrem B (3.4 ppm), but signs of staggers were less severe in sheep grazing pasture containing the former compared with the latter (median Keogh scores in late February were 2 and 3, respectively), consistent with epoxy-janthitrems being low potency toxins.

CONCLUSION: This study demonstrated that concentrations of epoxy-janthitrems and lolitrem B in sheep fat increased quickly during the initial phase of the study when concentrations in pasture increased, and decreased when animals were removed from pastures containing these compounds. These data will be used in the risk assessment of the endophyte metabolites.     

Selenium deficiency, the drug metabolising enzymes and mycotoxicoses in sheep

An examination was made of the relationship in Romney sheep between selenium deficiency and (i) the state of the liver drug metabolising enzyme system, (ii) subclinical liver damage, (iii) susceptibility to facial eczema, and (iv) susceptibility to ryegrasss staggers. The microsomal electron transport detoxifying enzymes based on cytochrome P-450 were unaffected by Se deficiency, suggesting that these enzymes are uninduced on ryegrass/clover pastures. Selenium deficiency had no effect on sporidesmin-induced liver damage, which makes it unlikely that peroxidative events countered by Se-dependent glutathione peroxidase (E.C.1.11.10) have a major role in the mechanism of sporidesmin toxicity. Selenium deficiency did appear to suppress the development of photosensitisation. There was no major effect on ryegrass staggers and no deficiency-induced subclinical liver damage.     

A simple treatment for “rye-grass staggers”?

Abstract

Extract

Sir,—A series of happy coincidences (serendipity?) has led us to a simple treatment which seems to have successfully alleviated symptoms of “rye-grass staggers” in a horse, a calf and two badly affected sheep. Although primarily engaged in a search for the causative agent(s) of rye-grass staggers, casual conversations with people having long experience of this disorder revealed many interesting observations. One of these was a racing-stable remedy for rye-grass staggers, which was the administration of “a couple of handfulls” of Epsom salts in a bran mash, with as much puha (Sonchus oleraceus) as the horse would eat. A suggestion that puha may be high in potassium led to the substitution of potassium chloride for puha, and on 9 March 1981, a thoroughbred mare was dosed with 100 g MgS0₄7H₂O and 6g KCl. This animal had marked symptoms of ryegrass staggers, such as swaying and head-weaving when standing, and a very irregular and unsteady gait at the walk. It is doubtful whether the animal could have trotted in this condition. The mare was greatly improved the following day, with no evidence of the above symptoms, and on the 13 March, the mare was able to trot freely around her paddock when chased by a motor bike. On the 16 March the mare again showed symptoms of rye-grass staggers, was treated as before and similarly improved. On 13 March a calf was reported to be staggering, and was drenched with 50 g MgSO₄7H₂O and 3 g K? 1. It too, recovered, and on 20 March showed only very slight symptoms, which had not worsened by 23 March and further treatment was not warranted.     

Ergovaline does not alter the severity of ryegrass staggers induced by lolitrem B

AIM: To investigate a possible interaction between lolitrem B and ergovaline by comparing the incidence and severity of ryegrass staggers in sheep grazing ryegrass (Lolium perenne) containing lolitrem B or ryegrass containing both lolitrem B and ergovaline.

METHODS: Ninety lambs, aged approximately 6 months, were grazed on plots of perennial ryegrass infected with either AR98 endophyte (containing lolitrem B), standard endophyte (containing lolitrem B and ergovaline) or no endophyte, for up to 42 days from 2 February 2010. Ten lambs were grazed on three replicate plots per cultivar. Herbage samples were collected for alkaloid analysis and lambs were scored for ryegrass staggers (scores from 0–5) weekly during the study. Any animal which was scored ≥4 was removed from the study.

RESULTS: Concentrations of lolitrem B did not differ between AR98 and standard endophyte-infected pastures during the study period (p=0.26), and ergovaline was present only in standard endophyte pastures. Ryegrass staggers was observed in sheep grazing both the AR98 and standard endophyte plots, with median scores increasing in the third week of the study. Prior to the end of the 42-day grazing period, 22 and 17 animals were removed from the standard endophyte and AR98 plots, respectively, because their staggers scores were ≥4. The cumulative probability of lambs having scores ≥4 did not differ between animals grazing the two pasture types (p=0.41).

CONCLUSIONS AND CLINICAL RELEVANCE: There was no evidence for ergovaline increasing the severity of ryegrass staggers induced by lolitrem B. In situations where the severity of ryegrass staggers appears to be greater than that predicted on the basis of concentrations of lolitrem B, the presence of other tremorgenic alkaloids should be investigated.     

Lolitrem B residue in fat tissues of cattle consuming endophyte-infected perennial ryegrass straw.

Lolitrems are neurotoxins found in endophyte-infected perennial ryegrass. Lolitrems, primarily lolitrem B, are the causative agents of ryegrass staggers in livestock. To guarantee the safety of meat produced from cattle consuming endophyte-infected perennial ryegrass, lolitrem B concentrations in tissues of Japanese Black cattle were determined using high-performance liquid chromatography. Lolitrem B was not detected in muscle, liver, kidney, or cerebrum of a Japanese Black cow with signs of ryegrass staggers. In contrast, perirenal fat contained 210 ppb lolitrem B. Three cows that received half as much perennial ryegrass straw as the cow with ryegrass staggers showed no clinical signs of ryegrass staggers. However, low concentrations of lolitrem B (less than 150 ppb) were detected in their fat tissue. These observations indicate that human exposure to the neurotoxic effect of lolitrem B through beef is unlikely. The amount of lolitrem B consumed by cattle can be estimated by the determination of lolitrem B in fat tissue.     

The effect of glyphosate, paraquat and paclobutrazol on lolitrem B levels in endophyte-infected perennial ryegrass

Two herbicides (glyphosate and paraquat) and a plant growth regulator (paclobutrazol) were applied to endophyteinfected (Acremonium lolii) perennial ryegrass swards. Subsamples of these swards were then chemically analysed at intervals up to 28 days later for lolitrem B, the compound responsible for perennial ryegrass staggers in domestic livestock. Glyphosate and paclobutrazol had no effect on lolitrem B concentrations. Paraquat applications decreased lolitrem B concentrations in the herbage. Because none of the chemicals tested increased the concentration of lolitrem B in the herbage, they are unlikely to be directly implicated in perennial ryegrass staggers in grazing animals.     

A review of the Neotyphodium lolii / Lolium perenne symbiosis and its associated effects on animal and plant health, with particular emphasis on ryegrass staggers

Abstract Ryegrass staggers is a seasonal mycotoxicosis of grazing livestock characterised by tremors, in coordination and a staggering gait almost unaccompanied by physical lesions. Deaths occur only as a consequence of accident or starvation. Outbreaks, in summer and autumn, occur only on pasture in which endophyte (Neotyphodium lolii)-infected perennial ryegrass (Lolium perenne) predominates and usually on which animals are grazed intensively. Animals recover when moved to a different type of grazing or after rain has promoted pasture growth. The disease was recognised for 80 years before its cause was discovered as a consequence of a grazing trial of sheep on three ryegrass cultivars which happened to have three different levels of endophyte infection. The endophyte was first formally described as Acremonium loliae, later corrected to Acremonium lolii, and was finally placed in the genus Neotyphodium. It produces a number of secondary metabolites of which lolitrem B is the principal one causing ryegrass staggers symptoms. Ergopeptides are also produced which cause heat stress and lack of productivity. N. lolii is symptomless in the plant, seed borne and grows intercellularly in the aerial parts, mainly in reproductive tillers and leaf sheaths but sparsely in leaf blades. It dies in stored seed and infection rates of different ryegrass cultivars have depended on seed storage times during their production. In addition, N. Lolii produces insect feeding deterrents, among them peramine, which protects infected plants from pest predation. Because of this, control of ryegrass staggers by elimination of endophyte-infected ryegrass is not feasible in areas in which insect predation is a serious pasture problem. However, N. lolii strains vary in the secondary metabolites they produce allowing the selection of strains that produce desirable metabolites. By inoculating such strains into uninfected ryegrass plants it is possible to produce cultivars which do not cause ryegrass staggers but resist insect predation. This review aims to provide a comprehensive summary of the current understanding of the N. lolii / L. perenne symbiosis, the toxins it is known to produce, their effects on animals and plants and the strategies used to control their ill effects while maximising their beneficial ones.     

Young perennial ryegrass staggers in a dairy herd

Ryegrass staggers was diagnosed in a group of young stock on a dairy farm in the Netherlands. The cattle were fed on ryegrass hay originating from a farm in the region. The typical clinical signs observed in this herd were difficulty in rising, hypermetria of especially the front legs, incoordination, and tremors of the head and neck muscles. The morbidity of the illness was about 80 percent and there was no mortality. Four weeks after withdrawal of the hay, the animals recovered completely. The diagnosis was confirmed by demonstrating high concentrations of Lolitrem-B (1.7 ppm) in the hay. This neurotoxin is usually produced by the endophyte Neotyphodium lolii with which ryegrass may be infected. This is the first case of ryegrass staggers in cows reported in the Netherlands since 1992.     

Ryegrass staggers in Canadian wapiti deer

Extract

Sir:- We would like to report an outbreak of ryegrass staggers in Canadian wapiti (Cervus elaphus mannitobensis) which occurred at the Invermay Agricultural Research Centre, Mosgiel. As far as we are aware this is the first recorded case of ryegrass staggers in deer associated with grazing perennial rye grass (Lolium perenne L.).     

The clinical differentiation of nervous and muscular locomotor disorders of sheep in Australia

SUMMARY Many of the nervous and muscular locomotor disorders that affect sheep throughout Australia are commonly referred to as ‘staggers’ syndromes. The range of clinical signs displayed by sheep suffering these disorders is sufficiently diverse to enable each syndrome to be graded into one of 5 progressive clinical groups. The first group, the limb paresis syndromes, includes the primary myopathies associated with the ingestion of Ixiolaena brevicompta, Malva parviflora, and Trachymene ochracea, as well as selenium and vitamin E disorders, Paroo virus staggers, congenital progressive muscular dystrophy, humpy back, hypocalcaemic muscle weakness, Tribulus terrestris staggers and tetanus . The second group is characterised by limb paresis with knuckling of the fetlocks, and includes the plant-associated toxicities of Romulea rosea, Stachys arvensis, Trachyandra divaricata, and Tribulus micrococcus, together with haloxon toxicity, enzootic ataxia (copper deficiency), and the probably genetic disorders of segmental axonopathy, neuroaxonal dystrophy, and degenerative thoracic myelopathy. Other locomotor disorders that fit more loosely into this group are listerial myelitis (post-dipping staggers), vitamin A deficiency, cervico-thoracic vertebral subluxation, Stypandra glauca toxicity, Ipomoea spp toxicity, ivermectin toxicity, and botulism . The third group, the falling syndromes, includes the probably genetic disorders of thalamic cerebellar neuropathy, cerebellar abiotrophy, and globoid cell leucodystrophy, together with Swainsona spp toxicity . The fourth group, the falling-with-tremors syndromes, includes the plant-associated toxicities of phalaris staggers, perennial rye grass staggers and nervous ergotism (Claviceps paspali) . The fifth group, the convulsive syndromes, includes the polioencephalomalacic entity nardoo fern (Marsilea drummondii) staggers, outbreaks of focal symmetrical encephalomalacia and the tunicaminyluracil toxicities known as annual ryegrass (Lolium rigidum) toxicity, annual beard grass (Polypogon monspeliensis) toxicity, blow away grass (Agrostis avenaceae) toxicity, and water damaged wheat (Triticum aestivum) toxicity . A dichotomous system is presented for the differential diagnosis of these groups of conditions .     

Detection of endophyte toxins in the imported perennial ryegrass straw

From 1997 to 1999, 29 cases of disorders were detected in cattle and horses that had been fed ryegrass straw imported from the U.S.A. These animals showed symptoms resembling ryegrass staggers and the clinical signs disappeared after removal of the straw. Endophytic hyphae were detected in the seeds of all straw samples that were responsible for the clinical cases. Lolitrem B concentrations in the straw ranged between 972 and 3740 ppb. Ergovaline concentrations were between 355 and 1300 ppb. Even though the concentrations of lolitrem B were lower than the toxic threshold proposed by Oregon State University in better part of the cases, our observations suggest the possibility that lolitrem B lower than the proposed threshold can bring disorders to sensitive individuals.     

Evaluation of a screening programme for identification of mannosidosis heterozygotes in angus cattle

Sir, — In recent years, much research effort has been directed towards identifying the cause of Ryegrass staggers (RGS). Many promising leads have been investigated, including studies of fungi in soil, ryegrass leaves, and dead leaf litter that could produce tremorgenic mycotoxins⁽³⁾. Several groups of fungi produce tremorgens⁽⁴⁾ which, when injected into sheep, produce symptoms similar to those in field outbreaks of RGS. However, under field conditions, populations of these fungi have not been shown to differ significantly between toxic and non-toxic pasture⁽⁷⁾ (McKenzie &; Byford, unpublished data).     

Tryptamine alkaloid toxicosis in feedlot sheep

Tryptamine alkaloid toxicosis (Phalaris staggers) was diagnosed in feedlot sheep. Clinical signs of toxicosis, which were exacerbated by excitement, included gait abnormalities, muscular tremors, nystagmus, and convulsions. An estimated 8% of the most severely affected lambs had clinical signs of toxicosis. Gross lesions detected in the brain of affected lambs consisted of focal gray-green discoloration in the brain stem and thalamus; these areas had microscopic evidence of intraneuronal pigment accumulation. Brain specimens obtained at slaughter indicated that 60% of the lambs had lesions consistent with tryptamine alkaloid toxicosis. Tryptamine alkaloids were found in low concentrations in the feed. Lambs exposed to these feeds had higher death losses than those that were not exposed to the feeds. Cobalt concentration in the feed was higher than that previously reported to be associated with Phalaris staggers.     

Molds, mycotoxins, and mycotoxicosis

Interest in mycotoxins and mycotoxicosis in humans and animals has greatly increased in recent years. Horses have long been considered very susceptible to molds. The signs, treatment, and prevention of several conditions, such as leukoencephalomalacia, aflatoxicosis, ergotism, fescue toxicity, slobbering disease, ryegrass staggers, and moldy sweet clover disease, are discussed.     

不同类型黑麦草营养价值评估

文章旨在通过体内外试验评估不同类型黑麦草的营养价值。试验选择平均体重为（66.87±2.34）kg的绵羊12头,随机分成3组,每组4个重复,每个重复1头。3组绵羊分别饲喂新鲜、青贮和晒干黑麦草。经过14?d的饲养试验后收集粪便和牧草样品进行后续分析。结果：晒干黑麦草的干物质、有机物和无氮浸出物含量均表现最高（P＜0.05）。新鲜黑麦草粗蛋白质和半纤维素含量较青贮黑麦草分别显著提高42.10%和10.82%（P＜0.05）。青贮黑麦草粗脂肪、粗纤维、中性洗涤纤维和酸性洗涤纤维含量显著高于其他两种类型的黑麦草（P＜0.05）。新鲜、青贮和晒干黑麦草的粗蛋白质、粗脂肪和粗纤维表观消化率无显著差异（P＞0.05）。晒干黑麦草干物质表观消化率较新鲜和青贮黑麦草显著提高6.34%和6.21%（P＜0.05）,同时晒干黑麦草有机物表观消化率较新鲜黑麦草显著提高6.65%（P＜0.05）,无氮浸出物表观消化率较青贮黑麦草显著提高15.44%（P＜0.05）。当体外代谢能值以MJ/kg有机物表示时,新鲜和青贮黑麦草代谢能值较晒干黑麦草分别显著提高21.34%和22.41%（P＜0.05）,而当体外代谢能值以MJ/kg干物质表示时,新鲜黑麦草能值最高,青贮黑麦草能值次之,晒干黑麦草能值最低（P＜0.05）。结论：新鲜、晒干和青贮黑麦草在不同营养成分上各有优势,但3种牧草主要营养物质的表观消化率和代谢能值无显著差异。因此,无论是新鲜、晒干还是青贮黑麦草都可以作为反刍动物粗饲料的良好来源。 [关键词]黑麦草;青贮;营养价值;消化     

Increased tolerance to annual ryegrass toxicity in sheep given a supplement of cobalt

SUMMARY To determine whether oral cobalt supplements could modify the clinical onset of annual ryegrass toxicity, groups (n = 5) of sheep were dosed orally with 0, 4 or 16 mg cobalt/day. After 3 weeks on this treatment, toxic ryegrass seed was added to their feed to provide 0, 0.15 and 0.30 mg corynetoxins/kg body weight, daily. Sheep receiving cobalt ingested 30% more toxin than did unsupplemented sheep before clinical signs developed (P = 0.03). There was no significant difference between groups receiving 4 and 16 mg cobalt. The results showed that cobalt delayed, but did not prevent, the onset of clinical signs of annual ryegrass toxicity.     

Mycotoxic tremorgens of Claviceps paspali and Penicillium cyclopium: a comparative study of effects on sheep and cattle in relation to natural staggers syndromes

Claviceps paspali sclerotia were dosed orally at various levels to sheep and cattle and the neurotoxic effects are described. Evidence is presented which indicates that the tremors are produced by a neutral tremorgen fraction. The ergot alkaloids, principally D-lysergic acid alpha-hydroxyethylamide and its isolysergic acid isomer, are not involved. The mixture of C paspali tremorgens has not been separated and characterised but its components are chemically related to other known tremorgens, such as penitrem A, produced by saprophytic moulds, notably Penicillium cyclopium. P cyclopium mycelium and C paspali sclerotia were dosed to produce comparable effects in sheep and in cattle. Similarly, effects of penitrem A and tremorgen concentrate of C paspali were compared after intravenous dosage to sheep. Observed enurotoxic effects of both moulds were identical in most respects. A low level of tremorgenic activity was detected at the honeydew stage of C paspali. These findings are discussed in relation to staggers syndromes occurring in grazing livestock.     

The effect of grain species, processing and time of feeding on the efficiency of feed utilization and microbial protein synthesis in sheep

This study investigated the effect of cereal grain species (sorghum, wheat, oats and barley), extent of processing (cracked barley, finely ground barley, and wet whole barley) and time of feeding (barley grain mixed with ryegrass hay or fed 2 h before hay was fed), on whole tract dry matter digestibility (WTDMD), and microbial protein synthesis (MPS), as a supplement to ryegrass hay when fed to rumen-cannulated sheep.Expected dry matter digestibility (EDMD) in mixtures of cereal grain and ryegrass hay was calculated by interpolation between in vitro dry matter digestibility (DMD) of each grain and the ryegrass hay. These were compared with measured actual WTDMD to detect positive or negative associative effects. Among grain species, the percentage difference in digestibility between actual WTDMD and EDMD was negative at − 6.6% units for wheat but positive at + 2.3%, + 4.3% and + 5.7% units for sorghum, oats, cracked or finely ground barley, respectively.As expected, the supplementation of sheep fed ryegrass hay with different sources of carbohydrates increased urinary allantoin output (as an indicator of MPS) when compared to sheep fed ryegrass hay alone. The concentration of urinary allantoin was significantly higher in sheep supplemented with sorghum (1916 mg/sheep/day) than wheat, oats or cracked barley ([mean ± S.E.M.] 1451 ± 24 mg/sheep/day) grain. There was a significantly higher urinary allantoin concentration in sheep fed cracked barley compared to finely ground barley or wet whole barley (1479 vs. 1095 vs. 1031 mg/sheep/day, respectively). There was no significant (P > 0.05) difference in urinary allantoin output, expressed as output/kg DM intake, when cracked barley was mixed and fed with the hay or fed 2 h before hay. However, in terms of total output of allantoin this was significantly higher (P < 0.05) (1479 vs. 1209 mg/sheep/day).In sacco degradability characteristics of organic matter and nitrogen for sorghum, oats, wheat, barley at different levels of processing and for ryegrass hay were also measured in the rumen of cannulated sheep.Among grain species, wheat had the highest effective organic matter degradability in the rumen (78.1%) while sorghum had the lowest. The effective degradability of protein of finely ground barley in the rumen was found to be higher than cracked barley or wet whole barley.Wheat grain, being highly degradable in the rumen, had a negative effect on WTDMD. In contrast, sorghum grain, being more slowly degradable in the rumen, would be expected to provide a substantially increased supply of energy to microbes over time in the rumen for MPS.Both cracked barley and finely ground barley also had a positive associative effect on WTDMD when fed with ryegrass increasing it by 5.7% units. The MPS was significantly higher (P < 0.05) in sheep fed cracked barley compared to finely ground barley or wet whole barley. This supports the hypothesis that slowly degrading carbohydrate sources synchronise more closely with available N from degradation of forage in the rumen.