The effect of thiaminase-induced subclinical thiamine deficiency on growth of weaner sheep

Three experiments were performed to examine for causes of poor growth of young Merino sheep. Weekly testing of animals 42 weeks of age for 10 weeks revealed that 90% of clinically poor animals were excreting high levels of thiaminase in their faeces; low levels of activity were present in 20% of clinically normal animals. There were significant differences in the mean erythrocyte transketolase activity of the thiaminase excreting poor animals and the thiaminase free normal animals. Other known causes of poor growth could not be demonstrated. Weekly monitoring of thiaminase activity in the faeces from 80 lambs 6 weeks of age showed 23% to be excreting significant levels of enzyme (>3mUg^–1 DM) throughtut a 10 week test period. Mean growth rates of these lambs were significantly below those of lambs not excreting thiaminase or excreting low levels intermittently. Supplementation of thiaminase excreting lambs with intra-muscular injections of thiamine HCl was associated with a statistically significant improved growth rate (P<0.01) compared to unsupplemented sheep excreting thiaminase. Mean growth rates of lambs not excreting thiaminase on a continuous basis (sampled weekly) were the same with or without thiamine HCl supplementation. High thiaminase levels were found in the ruminal fluids of trial animals excreting the enzyme in their faeces, confirming this previously established association.Bacillus thiaminolyticus was isolated from faeces and ruminal fluids from clinically poor animals and is the most likely source of the thiaminase. Subclinical thiamine deficiency was indicated by low erythrocyte transketolase activities and elevated TPP effects and is proposed as the cause of the poor growth by the young sheep.     

Natural establishment of thiaminase activity in the alimentary tract of newborn lambs and effects on thiamine status and growth rates

Thiaminase activity was detected in the faeces of lambs at 2 to 5 days of age. Levels of activity increased for 10 days and then declined over the next 3 to 4 weeks. Decreased erythrocyte transketolase activity indicated thiamine insufficiency in lambs with high thiaminase activity. Mean growth rates were 17% less in lambs with high thiaminase activity than in lambs with zero or low thiaminase activity. Bacillus thiaminolyticus was the only organism isolated which produced thiaminase. Treatment of newborn lambs with intramuscular injections of sulphadoxine did not prevent them from excreting thiaminase in their faeces. It is proposed that oral thiamine supplementation of lambs at 2 to 3 weeks of age may be the most appropriate prevention and treatment for subclinical thiamine deficiency of the cause described.     

Oral treatment of polioencephalomalacia and subclinical thiamine deficiency with thiamine propyl disulphide and thiamine hydrochloride

Thiaminase type I production by Bacillus thiaminolyticus and activity in vitro were repressed by the primary substate thiamine and by thiamine monophosphate and thiamine propyl disulphide. At thiamine concentrations of 300-3000 mumol/l production of active enzyme by B. thiaminolyticus, and activity of purified enzyme, were totally repressed. Growth of B. thiaminolyticus was inhibited by thiamine propyl disulphide at 3000 mumol/l. Activity of purified thiaminase was lost when incubated with ruminal fluid from healthy thiaminase free sheep. Enzyme activity was also lost when exposed to mildly alkaline conditions but was stimulated and stabilized against heat denaturation if treated with dithiothreitol. Thiaminase activities in the ruminal fluids of cases of ovine polioencephalomalacia were repressed within 2 h of oral administration of thiamine propyl disulphide. Blood pyruvate levels and transketolase activities were restored to normal following treatment. Treated animals recovered clinically and were returned to pasture.     

Thiamine deficiency in sheep exported live by sea

Three shipments of sheep being exported live by sea were examined to determine their thiamine status. Measurements were made of thiamine concentration in liver and ruminal contents, transketolase activity in erythrocytes and thiaminase activity in ruminal liquor. Sheep that died or were clinically ill and euthanased had significantly lower hepatic and ruminal thiamine concentrations than clinically healthy control sheep. A high proportion had thiamine concentrations comparable to those found in sheep that die with polioencephalomalacia. Thiamine concentrations decreased with increasing time that sheep were in pre-embarkation feedlots and on board ship. Destruction of thiamine in the rumen by thiaminase was not a significant factor. Erythrocyte transketolase activities indicated that many of the sheep that arrived in the Middle East without signs of clinical disease were also in a state of thiamine insufficiency.     

Faecal thiaminase, plasma lactate and pyruvate concentrations and erythrocyte transketolase activity changes in apparently normal replacement ewes after the initiation to the pasture

A study was made to investigate faecal thiaminase and the thiamine-related biochemical changes in apparently normal replacement ewes with a feed change, after the initiation without adaptation to the new pasture. Twenty-four female ewes were divided into two groups. Group A was managed in a system based on pasture and was compared with group B system based on a diet of concentrate and straw until moving to pasture 9 weeks after. Blood samples for lactate, pyruvate and erythrocyte transketolase activity determinations and faeces for thiaminase estimation were evaluated chronologically. At the end of a 126 days experimental period, live weights of groups were similar. We confirmed that clinically normal sheep may have thiaminase activity in the faeces and concluded that the thiaminase release increased during the diet changes, from concentrate to pasture, and that their continued excretion could develop some degree of thiamine deficiency.     

BIOCHEMISTRY OF NATURAL AND AMPROLIUM-INDUCED POLIOENCEPHALOMALACIA IN SHEEP

SUMMARY Polioencephalomalacia (PEM) induced in sheep was compared with the disease found in naturally occurring cases. Blood biochemical indicators measured were pyruvate, lactate, glucose, erythrocyte transketolase (TK) and stimulation of TK by addition of thiamine pyrophosphate (TPP effect). Faeces and rumen contents were assayed for thiaminase activity. The effect of treating affected sheep with thiamine was also noted. It was found that amprolium treatment could induce thrombocytopenia, but once the sheep became accustomed to amprolium in the diet they seemed to be resistant to this effect. In sheep receiving amprolium significant weight losses preceded the onset of clinical signs. Further weight loss continued throughout the recovery period despite removal of amprolium from the diet and treatment with thiamine. Blood glucose was variable, and was elevated only when marked clinical signs were present. Pyruvate and lactate levels showed marked variation throughout the trial. TK values were depressed and TPP effects increased well before the onset of clinical signs, although some naturally occurring cases had normal levels. Faecal thiaminase activity was negligible in all the sheep on the amprolium trial but most field cases had a high level. High faecal thiaminase was observed in about 5% of clinically normal animals from affected flocks. Depression of erythrocyte TK activity coupled with the presence of faecal thiaminase appeared to be the most reliable diagnostic biochemical parameters for PEM. Treatment of PEM affected sheep with thiamine rapidly brought the biochemical status of the animals to normal. However where advanced brain lesions were present the damage was permanent and such sheep treated with thiamine remained partially decorticate.     

Biochemical Changes in Apparently Normal Sheep from Flocks Affected by Polioencephalomalacia

Blood samples were obtained for determination of the activities of lactate, pyruvate and erythrocyte transketolase, and faeces as samples for estimation of thiaminase, from 190 apparently healthy sheep from 19 flocks in which at least one animal had been diagnosed with polioencephalomalacia (PEM), from 59 control animals and from 7 affected by PEM. Feed and pasture grass samples were collected for thiaminase analysis and thiaminase was assayed in ryegrass and fescue. Decreased erythrocyte transketolase activity, accompanied by a corresponding rise in the thiamin pyrophosphate effect, and estimation of the thiaminase content in faeces, may be useful diagnostic procedures by which to identify periods of greater risk, during which animals could develop PEM. The changes in the diet probably acted as a primary factor for microbial production of thiaminases in replacement ewes. In these, the prevalence was higher in the spring months.     

Role of thiamine status in sulphur induced polioencephalomalacia in sheep.

The effects of excess dietary sulphur were studied in sheep supplemented and unsupplemented with thiamine. The diets contained either 0.19 per cent sulphur (LS) or 0.63 per cent sulphur (HS) in combinations with 14 mg kg-1 thiamine (LB1) or 243 mg kg-1 thiamine (HB1). A total of 56 two-month-old lambs were used. Groups consisting of nine, nine, 22 and 16 lambs were fed LS-LB1, LS-HB1, HS-LB1 and HS-HB1 diets, respectively for 14 weeks. Out of 22 lambs fed the HS-LB1 diet, seven lambs developed neurological signs between the third and eighth week of the trial. Two of these lambs died, three that were in extremis were euthanased, and two recovered completely. All clinically affected animals had extensive malacic lesions in the cerebral cortex, midbrain and brainstem. None of the lambs from the LS groups or HS-HB1 group developed clinical signs. Several clinically normal lambs from the HS-LB1 group had necrotic lesions in their brains at gross and microscopic examination. Supplementation with dietary thiamine prevented development of clinical signs, but did not totally prevent development of microscopic brain lesions. Brain thiamine concentration, transketolase activity and thiamine pyrophosphate (TPP) effect were not different (P greater than 0.05) among groups. There was a strong effect (P less than 0.0001) of dietary thiamine supplementation on blood thiamine concentration and TPP effect. Blood thiamine concentration was higher whereas TPP effect was lower in the thiamine supplemented sheep. Blood and tissue thiamine concentrations in sheep exposed to high dietary sulphur did not indicate either systemic or local thiamine deficiency per se. Increased TPP effect in sheep fed the HS-LB1 diet indicated mild to moderate metabolic thiamine deficiency. Thiamine inadequacy may be an effect of an increased requirement for thiamine in animals exposed to excess dietary sulphur.     

Ruminant thiamine requirement in perspective

Thiaminases play an important role in the aetiology of CCN being responsible for the state of thiamine-deficiency which is an essential feature of the disease, evidence for which is presented here. These studies have led to a greater appreciation of the role of thiamine and thiaminases in ruminant nutrition especially as ruminal thiaminase activity is not confined to clinically affected animals but is of wider distribution. The importance of thiaminases in intensive beef production and the possibility of the need for thiamine supplementation in the form of a thiaminase resistant derivative is discussed.     

Severe thiamine deficiency in sheep with acute ruminal lactic acidosis

BACKGROUND: Thiamine status of ruminants is adversely affected in acidic rumen conditions. However, there have been limited published case study data related to thiamine deficiency of ruminants with acute ruminal lactic acidosis (ARLA). HYPOTHESIS: Thiamine deficiency would occur in sheep with ARLA. ANIMALS: Thirteen Ak-Karaman (white Karaman) sheep with ARLA, aged 1 year (ARLA group) and 10 healthy Ak-Karaman sheep, aged 1 year (control group) were used. METHODS: After clinical examination, rumen fluid samples of all sheep were obtained with a stomach tube and examined immediately. Blood samples were taken from a jugular vein of the sheep. Erythrocytic transketolase enzyme activity and hence thiamine pyrophosphate (TPP) effect were determined according to Clausen's method. RESULTS: History revealed that all sheep in the ARLA group had accidentally consumed excessive amounts of cracked barley. During clinical examination of the ARLA group, disturbed general condition, engorged scleral vessels, moderate to severe dehydration, and ruminal atony were recorded in the sheep. The results of the ruminal fluid analyses of the ARLA group demonstrated characteristics of ARLA. The results of clinical and ruminal fluid examination of control group were normal. The mean TPP effect (%) in the ARLA group (109 +/- 28) was significantly higher than in the control group (22.2 +/- 3.7) (P < .001). CONCLUSIONS AND CLINICAL IMPORTANCE: The present study revealed that severe thiamine deficiency occurred in sheep with ARLA. This result indicates that thiamine administration to sheep suffering from acute ruminal acidosis caused by overconsumption of readily fermentable carbohydrates could be beneficial in alleviating thiamine deficiency caused by ruminal acidosis.     

Polioencephalomalacia in adult sheep grazing pastures with prostrate pigweed

Polioencephalomalacia was diagnosed in 2 animals from different farms. In apparently healthy animals from same farms, fecal thiaminase and a significant reduction in erythrocyte trans-ketolase activity was observed. The presence of thiaminase in Amaranthus blitoides could have contributed to the development of polioencephalomalacia in sheep grazing on natural pastures.     

Thiamine status of feedlot cattle fed a high-concentrate diet

As thiamine status of ruminants is adversely affected by rumen acidity, this study investigated whether or not thiamine deficiency occurs in feedlot cattle fed a high concentrate diet. Fifty 1- to 2-year-old feedlot cattle fed a high concentrate diet (75% barley) for at least 3 mo (high concentrate diet group) and 15 healthy feedlot cattle of similar ages (control group) that were fed a low concentrate diet (30% barley) were used. Rumen fluid samples were obtained by rumenocentesis and their pH was determined with a portable pH meter. Blood samples taken from all animals from a jugular vein were used to determine erythrocyte transketolase enzyme activity, and hence thiamine pyrophosphate (TPP) effect. Odor and mean pH values of ruminal fluid samples from the high concentrate diet and control group were acidic (pH 5.3) and aromatic (pH 6.1), respectively. The mean TPP effect % in the high concentrate diet group (47.2 ± 3.2) was significantly higher than in the control group (19.53 ± 2.5) (P < 0.001). The study provides evidence of a TPP effect in feedlot cattle fed a high concentrate diet.     

Polioencephalomalacia in cattle consuming water with elevated sodium sulfate levels: A herd investigation

Polioencephalomalacia (PEM), hereafter used to refer to the specific lesion of cerebrocortical necrosis, developed in 11 of 110 mature cattle on pasture in central Saskatchewan. The primary water source contained a markedly elevated level of sodium sulfate (7200 ppm). The significant clinical findings of the herd investigation included depression, ataxia, cortical blindness, dysphagia, and death. Diagnosis of PEM was confirmed by histopathological evidence of cerebrocortical and subcortical necrosis with microvascular fibrinoid necrosis predominantly in the thalamic region of three affected cattle. The histopathology of sulfate-associated PEM observed in this herd appears to be unique and its features are presented and discussed. Mean levels for serum transketolase, copper, red blood cell transketolase activity, and thiamine (vitamin B₁) in all exposed young (n = 100) and mature (n = 99) animals did not reveal evidence of deficiencies. Although the blood thiamine status of the seven surviving, affected animals was not evaluated before treatment with exogenous thiamine, 199 members of the herd had blood thiamine levels within the reference range at the time of the outbreak. The outbreak resolved after cattle were moved to a water source containing acceptable levels of sodium sulfate.     

The response of Suffolk lambs to an escalating experimental infection with Dictyocaulus filaria.

Infection of Suffolk lambs with Dictyocaulus filaria larvae beginning at two months of age with one larva per day, six days per week, and doubling every four weeks to reach a final infection rate of 64 larvae per day gave rise to a pattern of larval excretion in the faeces which approximated to that seen in naturally infected Suffolk lambs in Midlothian in 1974. Several of the lambs were still infected with adult D filaria and were excreting larvae in their faeces 28 weeks after the infection began. However, when some of the lambs were challenged at this time, those which had received the previous escalating infection were resistant to the challenge infection.     

中草药复合添加剂对羊瘤胃消化酶活性的影响

为了研究自制中草药复合添加剂对绵羊瘤胃消化酶活性的影响,将60只4月龄健康杜寒杂一代绵羊随机分为4组进行试验,分别为公羊、母羊对照组(常规日粮),公羊、母羊试验组(常规日粮+中草药复合添加剂),试验期为45d。试验结束后取绵羊瘤胃液,分别测定瘤胃蛋白酶、羧甲基纤维素酶、α-淀粉酶活性。结果表明,该中草药复合添加剂对绵羊瘤胃蛋白酶、α-淀粉酶活性的影响均不显著(P>0.05),但显著(P<0.05)提高了绵羊瘤胃羧甲基纤维素酶的活性,且上述3种酶活性的变化均与绵羊性别无关。研究结果提示,该中草药复合添加剂能提高绵羊瘤胃羧甲基纤维素酶的活性,可促进饲草料中纤维素在羊瘤胃内的消化代谢。     

Effects of diets of high sulphur content and varied concentrations of copper, molybdenum and thiamine on in vitro phagocytic and candidacidal activity of neutrophils in sheep

Candidacidal and phagocytic tests were performed on 34 female sheep fed high sulphur diets containing varied concentrations of thiamine, copper, and molybdenum for 14 weeks. Tests were conducted at weekly intervals for five weeks during the latter half of the experiment. The ability of polymorphonuclear leucocytes from sheep on diets unsupplemented with thiamine to kill phagocytosed Candida albicans was lower than those on diets supplemented with thiamine. The ability of polymorphonuclear leucocytes to phagocytose C albicans was lower in animals on low copper diets than in animals on high copper diets. Factorial analysis showed that thiamine supplementation significantly increased (P less than 0.0001) candidacidal but not phagocytic activity. By contrast, copper supplementation significantly increased (P less than 0.0005) the phagocytic activity with no effect on candidacidal activity. The observed detrimental effect of high sulphur intake on in vitro polymorphonuclear leucocyte function can mean that ruminants in areas where large quantities of sulphur are taken in with water and feed have compromised immune function due to lower copper and thiamine status and hence are at risk of increased susceptibility to infections.     

Border disease: virus persistence, antibody response and transmission studies

Three groups of five and one group of four oestrus-synchronised sheep were inoculated with Border disease (BD) virus at 52 +/- 2 days after their first service. Transmission of virus to offspring as demonstrated by virus isolation, detection of viral antigen and, or antibody response occurred in 12 of 19 sheep and probably in four others which aborted or produced stillborn lambs. Both apparently normal and clinically affected animals excreted virus in saliva, urine and faeces, and excretion and contact transmission to sheep and pigs persisted for up to two and a half years. Most of the tissues of infected sheep contained virus titres between 10(3.5) and 10(5.5) TCID50 per g. The immune response in the lambs varied, in some it began before birth, in others a transient or low level response was observed in the first or second year, while others remained serologically negative for two and a half years.     

Clupeid Response to Stressors: The Influence of Environmental Factors on Thiaminase Expression

Abstract

Over the past five decades, a reproductive failure related to thiamine deficiency, referred to as thiamine deficiency complex (TDC), has been observed in valuable salmonine fishes in the Great Lakes and Finger Lakes in North America and the Baltic Sea in Europe. The cause of TDC has been linked to the consumption of clupeid fish, which contain high levels of a thiamine-destroying enzyme called thiaminase I (hereafter referred to as “thiaminase”). High activities of thiaminase have been reported from clupeids such as Alewife Alosa pseudoharengus, Gizzard Shad Dorosoma cepedianum and Atlantic (Baltic) Herring Clupea harengus, but no consistent explanation has accounted for the wide range of observed variation in levels of thiaminase in clupeids. Chronic stress can suppress the immune systems of Alewife and other fishes, thereby reducing the number of circulating white blood cells available to suppress bacteria. Because the presence of thiaminase has been associated with thiaminolytic bacteria isolated from Alewife viscera, we hypothesized that stressful conditions, which can potentially limit clupeid immune response or alter internal physiological conditions, could allow for thiaminase to be produced more efficiently by bacteria or thiaminolytic bacteria could proliferate, or both events could occur, resulting in a subsequent increase in thiaminolytic activity. In this study, Alewives and Gizzard Shad were exposed to severe winter temperatures and low food availability, respectively, in replicated pond experiments to evaluate the influence of stressful conditions on clupeid thiaminase activity. Though responses in circulating white blood cell counts and metrics of fish condition indicated that experimental treatments affected these clupeids, these effects were not related to increased thiaminase activity. The only significant treatment effect on clupeid thiaminase was an increase in mean thiaminase activity in Gizzard Shad from ponds where only high quality energy sources were available. These data indicate that variability in clupeid thiaminase may be related to diet composition.

Received June 19, 2012; accepted January 15, 2013     

Sulfur-induced polioencephalomalacia in sheep: some biochemical changes.

The effect of high dietary sulfur (S) supplementation on blood thiamine (B1) concentration, biochemical indices of liver, muscle and kidney damage and selected plasma electrolytes was studied in six sheep. Three of these sheep received an additional 230 mg thiamine/kg diet (Group 2). After approximately 2.5-3 weeks on this diet, all three sheep in the non-B1-supplemented group (Group 1) showed loss of appetite and developed mild neurological signs: depression, intermittent signs of excitation and head pressing. Increases in blood B1 concentration and plasma creatine kinase (CK) and aspartate aminotransferase (AST) were observed during this time in all affected animals. Clinical signs lasted only for two to five days. Sheep in group 2 were clinically normal throughout the experiment, but all of these animals also had elevated blood B1 concentrations and plasma CK activity at the 3 wk sampling. Plasma magnesium concentrations of group 1 sheep were elevated at the 2.5-3 wk and 6 wk samplings but they declined significantly (p less than 0.05) to low normal levels thereafter. Magnesium concentrations of group 2 sheep were low at the beginning but progressively increased during the course of the experiment. At necropsy, brain lesions suggestive of polioencephalomalacia (PEM) were observed in all sheep but were most marked in group 1. It is speculated that PEM may be caused by a direct toxic effect of S, S metabolites or B1 antimetabolites in the brain rather than by an in vivo B1 deficiency per se.     

Ruminal lactic acidosis in sheep and goats.

The clinical findings in 37 sheep and goats with acute ruminal lactic acidosis included a disturbed general condition characterised by anorexia, apathy, teeth grinding and muscle twitching, ruminal stasis, and the excretion of soupy or watery faeces. The ruminal fluid of affected animals was milky, had a sour odour and a low pH. There was a predominance of Gram-positive bacteria in smears of ruminal fluid. In comparison with 10 control animals, the rumen fluid of 23 sheep with ruminal lactic acidosis had higher lactic acid and lower volatile fatty acid concentrations. In addition, the affected animals often had haemoconcentration and metabolic acidosis. Treatment included single or repeated transfer of ruminal fluid from healthy cows and, depending on the severity, the administration of antacids, yeast and chlortetracycline, and the intravenous infusion of isotonic sodium chloride and 5 per cent sodium bicarbonate solutions. Of the 37 treated sheep and goats, four died within 24 hours, and three others were euthanased after one, two and three days because their condition rapidly deteriorated. Thirty animals were discharged one to nine days after treatment. Twenty-nine of them (78.4 per cent) recovered completely but one was euthanased later.