Acute inhibition of glucose-6-phosphate translocator activity leads to increased de novo lipogenesis and development of hepatic steatosis without affecting VLDL production in rats.

Glucose-6-phosphatase (G6Pase) is a key enzyme in hepatic glucose metabolism. Altered G6Pase activity in glycogen storage disease and diabetic states is associated with disturbances in lipid metabolism. We studied the effects of acute inhibition of G6Pase activity on hepatic lipid metabolism in nonanesthetized rats. Rats were infused with an inhibitor of the glucose-6-phosphate (G6P) translocator (S4048, 30 mg. kg(-1). h(-1)) for 8 h. Simultaneously, [1-(13)C]acetate was administered for determination of de novo lipogenesis and fractional cholesterol synthesis rates by mass isotopomer distribution analysis. In a separate group of rats, Triton WR 1339 was injected for determination of hepatic VLDL-triglyceride production. S4048 infusion significantly decreased plasma glucose (-11%) and insulin (-48%) levels and increased hepatic G6P (201%) and glycogen (182%) contents. Hepatic triglyceride contents increased from 5.8 +/- 1.4 micromol/g liver in controls to 20.6 +/- 5.5 micromol/g liver in S4048-treated animals. De novo lipogenesis was increased >10-fold in S4048-treated rats, without changes in cholesterol synthesis rates. Hepatic mRNA levels of acetyl-CoA carboxylase and fatty acid synthase were markedly induced. Plasma triglyceride levels increased fourfold, but no differences in plasma cholesterol levels were seen. Surprisingly, hepatic VLDL-triglyceride secretion was not increased in S4048-treated rats. These studies demonstrate that inhibition of the G6Pase system leads to acute stimulation of fat synthesis and development of hepatic steatosis, without affecting hepatic cholesterol synthesis and VLDL secretion. The results emphasize the strong interactions that exist between hepatic carbohydrate and fat metabolism.

Chlorogenic acid and synthetic chlorogenic acid derivatives: novel inhibitors of hepatic glucose-6-phosphate translocase.

The enzyme system glucose-6-phosphatase (EC 3.1.3.9) plays a major role in the homeostatic regulation of blood glucose. It is responsible for the formation of endogenous glucose originating from gluconeogenesis and glycogenolysis. Recently, chlorogenic acid was identified as a specific inhibitor of the glucose-6-phosphate translocase component (Gl-6-P translocase) of this enzyme system in microsomes of rat liver. Glucose 6-phosphate hydrolysis was determined in the presence of chlorogenic acid or of new synthesized derivatives in intact rat liver microsomes in order to assess the inhibitory potency of the compounds on the translocase component. Variation in the 3-position of chlorogenic acid had only poor effects on inhibitory potency. Introduction of lipophilic side chain in the 1-position led to 100-fold more potent inhibitors. Functional assays on isolated perfused rat liver with compound 29i, a representative of the more potent derivatives, showed a dose-dependent inhibition of gluconeogenesis and glycogenolyosis, suggesting glucose-6-phosphatase as the locus of interference of the compound for inhibition of hepatic glucose production also in the isolated organ model. Gl-6-P translocase inhibitors may be useful for the reduction of inappropriately high rates of hepatic glucose output often found in non-insulin-dependent diabetes.

Kodaistatins, novel inhibitors of glucose-6-phosphate translocase T1 from Aspergillus terreus thom DSM 11247. Isolation and structural elucidation.

Two novel compounds, kodaistatin A, C35H34O11, molecular weight 630, and kodaistatin C, C35H34O12, molecular weight 646, have been isolated from cultures of Aspergillus terreus Thom DSM 11247 by solid-phase extraction, size-exclusion chromatography, and various preparative HPLC steps. The use of a range of 2D NMR measurements, in particular 13C-13C correlation measurements, has led to the clarification of the structure of kodaistatin A. Kodaistatin C is a hydroxylated derivative of kodaistatin A. Both natural products contain hydroxylated aspulvinones and identical highly substituted polyketide units. An X-ray single crystal structure analysis of aspulvinon E demonstrated the z-configuration at the central double bond. The kodaistatins are effective inhibitors of the glucose-6-phosphate translocase component of the glucose-6-phosphatase system (EC 3.1.3.9), an enzyme system which is important for the control of blood glucose levels. The IC50 is 80 nM for kodaistatin A and 130 nM for kodaistatin C.

Large-scale management systems and parasite populations prevalence and resistance of parasitic agents in animal effluents and their potential hygienic hazard.

Keeping large numbers of animals in large-scale, pseudo-industrial units poses similar problems to the overstocking of small farms within any one area, as far as the disposal of animal effluents is concerned. The prevalence of parasites in pigs, poultry and cattle on large-scale farms is listed, and examples of the resistance of their exogenic stages are given. The possibilities of transmission to individuals of the same species or to other species including man are evaluated. Guidelines for safe or restricted disposal of the effluents as fertilizers for different plant crops are given. On recycling effluents as animal food, no problems should arise from the parasitologist's point of view if poultry manure is fed to pigs or cattle or if swine or cattle effluents are fed to poultry. Other combinations are hazardous.

The use of a morantel sustained release bolus in the seasonal control of parasitic gastroenteritis in second-season cattle.

The efficacy of a morantel sustained release bolus was evaluated during the 1980 summer grazing season in 216 yearling, second-season cattle in nine trials in West Germany. In five trials individual farms were used, in which medicated and control groups could be grazed on adjacent pastures. In four additional trials medicated and control groups were maintained on separate farms where the sites and management practice were matched as far as possible. Faecal worm egg counts, herbage larval counts and weight gain were monitored at regular intervals throughout the season. The faecal output of gastrointestinal nematode eggs was reduced by 55.8 percent in the medicated animals during the period from May to early August, while worm egg counts were equal to that of the control animals during the later part of the season. Herbage larval counts were similar on pastures grazed by medicated and nonmedicated cattle. In the five trials where individual farms were used the average daily weight gains over the entire grazing period were 900 (+/- 222) g/animal/day in the medicated group compared with 826 (+/- 263) g/animal in the nonmedicated cattle, a non-significant advantage. No advantage for the treated over the control groups was detected in the additional 4 trials but this was attributed largely to the difficulty in obtaining comparable groups of animals in comparable grazing conditions in "matching" farms, leading to inter-farm variation.

Apparent digestibility of nutrients and nitrogen balance during experimental infection of calves with Eimeria bovis.

The apparent digestibilities (AD) of dry matter (DM), organic matter (OM), crude ash (CA), crude fiber (CFi), crude fat (CFa) crude protein (CP) and nitrogen-free extracts (NFE) and the nitrogen balance were investigated during experimental Eimeria bovis coccidiosis in calves. noninfected pair-fed controls and controls fed on a normal plan of nutrition were included in the study to allow differentiation between the effects of infection and of changes in feed intake. Primary infection with 5 x 10(4) oocysts (n = 4, group A) caused mild diarrhea and calves infected primarily with 1 X 10(5) oocysts (n = 5, group B) suffered from mild (three calves) to severe hemorrhagic (two calves) diarrhea. No clinical disease was seen after reinfection of the group A calves with 1 X 10(5) oocysts. The primary infection with 5 X 10(4) oocysts or reinfection with twice the primary inoculum did not affect AD of nutrients or the overall nitrogen balance (RT). AD of DM, NFE or OM were higher in group B during patency and in the pair-fed group C calves (n = 5) than in the reinfected but healthy group A calves. AD of CFi of the group B calves even exceeded the values of the pair-fed controls. The two calves of group B that suffered from hemorrhagic diarrhea and anorexia had low values of AD of CP during the acute phase of the disease and the plasma nitrogen levels were reduced in this group. Severe clinical coccidiosis transiently reduced the nitrogen balance. It is concluded that the transient increase of AD of nutrients, especially of CFi, during clinical coccidiosis reflect hypomotility and that anorexia and intestinal leakage impair the nitrogen balance and cause weight depression.

T cell responses in calves to a primary Eimeria bovis infection: phenotypical and functional changes.

The study aimed to characterize T cell responses in calves to a primary E. bovis infection. For this purpose, peripheral blood lymphocytes (PBL) were isolated from six infected calves and three controls during prepatency (Day 12 post infection (p.i.), patency (Day 25 p.i.) and postpatency (Day 35 p.i.). In addition, lymphocytes were isolated from various lymphatic organs (lnn. cervicales superficiales, lnn. jejunales craniales, lnn. jejunales caudales, lnn. caecales, lnn. colici, Peyer's patches (PP) and spleen) at necropsy (Day 35 p.i.). FACS analyses determined the proportions of CD4+-, CD8+-, CD2+-, and gammadelta+-T cells. Proliferative responses of the cells after stimulation with Concanavalin A (Con A) and an E. bovis-merozoite I antigen (EbAg) were measured. Furthermore, in situ hybridization experiments were performed for the detection of IL-2 and IL-4 mRNA in histological sections of lymphatic organs. Proportions of CD4+-, CD8+- and CD2+-expressing PBL were significantly increased 12 days p.i. in infected calves. While the proportions of CD4+- and CD8+-PBL declined until day 25 p.i. and finally reached control values, proportions of activated PBL (CD2+-T cells) remained at a high level throughout the observation period. Those of gammadelta+-PBL, in contrast, remained unaffected. The proportions of CD4+-, gammadelta+- and CD2+-T cells in lymphatic organs were significantly increased in comparison to uninfected controls, when determined 35 days p.i. Concerning the proportions of CD8+-T cells of the organs, however, there were no differences between the groups. PBL and cells from lymphatic organs except those from the PP showed strong proliferative response to the mitogen Con A, without a significant difference between the groups. Reactions to EbAg in contrast differed significantly between controls and E. bovis infected calves. Proliferation responses of PBL of infected animals were highest 12 days p.i.; subsequently they decreased and 35 days p.i. they were found within the ranges of controls. Lymphocytes isolated from lymphatic organs of infected animals reacted significantly stronger than lymphocytes from control animals, whereby most marked differences occured with cells from lymph nodes draining E. bovis infested parts of the intestine and from the spleen. These reactions were accompained by an increased transcription of the IL-2 gene but not of the IL-4 gene in gut associated lymphnodes of infected calves when compared with infected controls. The data suggest strong antigenic stimuli by developing first generation schizonts, and of predominant involvement of (CD4+) Th1 cells in the course of a primary E. bovis infection of calves.

Efficacy of two formulations ('injectable' and 'pour on') of moxidectin against gastrointestinal nematode infections in grazing cattle.

The efficacy of moxidectin, 'injectable' and 'pour on', against gastrointestinal nematodes was determined in cattle in two separate field trials (Trial I in 1990 and Trial II in 1991) with respectively 88 and 94 young grazing cattle of either sex. The efficacy was measured on the basis of the reduction of the egg output and of the evaluation of the results from larval differentiation. Animals in Group MI received 0.2 mg kg-1 body weight (b.w.) moxidectin injectable solution in Trial I on Day 0. Group CI was not given any medication on Day 0, but 0.2 mg kg-1 b.w. ivermectin injectable solution (Ivomec) was applied after 2 weeks to prevent clinical disease. In Trial II, animals in Group MP were treated with pour on moxidectin (0.5 mg kg-1 b.w.) on Day 0. Animals in Group CP serving as controls for Group MP during the first part of the trial received the same formulation at the same dose 2 weeks after treatment of Group MP. When the egg output was compared within treated groups, the egg count reduction was very similar post treatment (p.t.) with both formulations being 96.3% and 96.6% on Day 7 after the application of injectable moxidectin or pour on moxidectin, respectively, and 90.7% and 92.5% on Day 28 p.t. When egg counts of treated and control animals were compared (corrected for the e.p.g. values before treatment) the egg count reduction was 95.4% and 91.5% on Day 7 and 92.9% and 84.8% on Day 14 p.t. with either the injectable or pour on formulation. Pour on moxidectin seemed to be more effective against Ostertagia spp. than against Cooperia spp. Animals treated with injectable moxidectin gained significantly more body weight (4.2 kg per animal) than the controls from Day -7 to Day +14, while no significant difference in weight gain was achieved within 2 weeks after treatment with pour on moxidectin.

Prevalence and control of benzimidazole-resistant small strongyles on German thoroughbred studs.

The prevalence of benzimidazole-resistant small strongyles was determined in a survey, conducted on 14 thoroughbred studs, which compared the faecal egg counts of groups of horses before and after treatment with the recommended doses of cambendazole (20 mg kg-1 b.w.) or febantel (6 mg kg-1 b.w.). Benzimidazole-resistant cyathostomes were found on all farms examined. Pyrantel pamoate (19 mg kg-1 b.w.), oxibendazole (10 mg kg-1 b.w.) and ivermectin (0.2 mg kg-1 b.w.) reduced the strongyle egg counts on these studs by 97-100% at 2 weeks post-treatment. However, 6 weeks after dosing the reduction of the strongyle egg output had decreased to an average of 67.8% (8.7-97.1%) with pyrantel pamoate and 51.2% (0-95.8%) with oxibendazole, whereas ivermectin still suppressed the egg counts by 98.2% (95-100%).

Detection of Trypanosoma congolense antibodies with indirect ELISAs using antigen-precoated microtitre plates.

The study reports the performance of four indirect enzyme-linked immunosorbent assays (ELISAs) for antibody (AB) detection using microtitre plates which were precoated with native or heat/detergent denatured antigens (AGs) from Trypanosoma congolense (T.c.) and T. vivax (T.v.), and stored for between 1 to 206 days at +37 degrees C. Bovine serum samples were obtained by sequential bleeding of 3-months old T.c.-infected bulls and their uninfected cohorts, as well as by a single bleeding of uninfected adult cattle. The first day of AB detection, and observations on samples after this (defined as estimated ELISA sensitivity), depended on the cut-off value in the specific ELISAs. Cut-off values from pre- and early post-infection samples of individual animals demonstrated a seroconversion in all ELISAs on average after 10-15 days post-infection (dpi). The AB detection was delayed in the T.c. native and denatured AG-based ELISAs using cut-off points from uninfected cohort cattle (16.5 dpi, 19.3 dpi) and the adult cattle population (22.1 dpi, 25.0 dpi). The T.v. AG-based ELISAs however lacked crossreactiviy to T.c. ABs. The estimated sensitivity of each T.c. AG-based ELISA was above 96% throughout, but significantly lower for the T.c. native AG-based ELISA (91.1%) when the adult cattle derived cut-off point was used (p<0.01). The sensitivity of the phase contrast buffy coat technique was similar to the T.c. AG-based ELISAs, but significantly lower when the T.c. denatured AG-based ELISA was used at the adult cattle derived cut-off point (p<0.05). The implications of the results and future research aspects on ELISAs to detect trypanosomal ABs and AGs are discussed.

Efficacy of febantel in sheep experimentally infected with five species of gastrointestinal nematodes.

The anthelmintic activity of the guanidine derivative febantel was tested in a total of 76 experimentally infected lambs in two control tests. Doses of 5.0 and 7.5 mg per kg body-weight were administered orally against fourth and pre-adult fifth or adult stages of Haemonchus contortus, Ostertagia circumcincta, Trichostrongylus colubriformis, B unostomum trigonocephalum and Oesophagostomum columbianum. The efficacy was between 98.5 and 100 per cent. The drug was well tolerated.

Efficacy of four anthelmintics against benzimidazole-resistant cyathostomes of horses.

In order to confirm benzimidazole resistance as recommended at a workshop of the Commission of the European Communities the isolate 'E' of cyathostome strongyles originating from a stud where benzimidazole resistance had been demonstrated by egg hatch tests and by egg count reduction tests was investigated in two series of critical tests. Each of 11 foals reared strongyle-free was infected with 130,000 third stage cyathostome larvae. One animal remained untreated, two pairs of foals were treated with paste formulations of the (pro)benzimidazoles cambendazole (20 mg/kg bodyweight) or febantel (6 mg/kg bodyweight) and two groups of three foals were given pastes containing the non-benzimidazole drugs pyrantel pamoate (19 mg/kg bodyweight) or ivermectin (0.2 mg/kg bodyweight) either at 101 days (trial 1) or at 59 to 62 days (trial 2) after infection. Strongyles were counted in faecal samples collected daily between treatment and post mortem examination five or seven days later and worm burdens were counted in the intestinal contents and mucosal digests. Nine species of the cyathostome subfamily were found in the infected foals. The numbers of luminal stages were reduced by only 3.1 and 20.2 (mean 7.9) per cent by cambendazole and by 13.6 and 32.8 (mean 21.3) per cent by febantel in the individual animals. However, pyrantel pamoate (93.6 to 98.2, mean 96.3 per cent reduction) and ivermectin (100 per cent reduction) were highly effective. These trials provide the first report of benzimidazole resistant Cylicostephanus poculatus anywhere in the world and demonstrate (pro)benzimidazole resistance in seven other species for the first time in Europe.

Efficacy of an oxibendazole-trichlorfon paste formulation against third stage larvae of Gasterophilus intestinalis and its safety in horses.

A paste formulation containing 14.3 per cent of oxibendazole and 44 per cent of trichlorfon was administered to 33 ponies and horses. The dose rate used was equivalent to 10 mg and 30 mg/kg bodyweight, of oxibendazole and trichlorfon respectively. After treatment 25 animals passed between one and 82 third stage larvae of Gasterophilus intestinalis in their faeces. Dosing with 0.2 mg ivermectin/kg bodyweight three weeks later resulted in six animals expelling between one and four bots. The efficacy of the oxibendazole-trichlorfon paste was on average 96.2 per cent. This drug combination given to 52 ponies and horses at the indicated dose rate and to six ponies at twice that dose was tolerated without side effects except transient softening of the faeces in several animals and mild symptoms of colic in two horses.

[The helminth fauna of the gastrointestinal tract of young cattle in northwest Germany]. / Ein Beitrag zur Helminthenfauna des Magen-Darm-Traktes von Jungrindern in Nordwestdeutschland.

Ten unmedicated animals of a controlled anthelmintic trial were used to analyse the present gastrointestinal helminth fauna of young cattle from the northwest of Germany. This survey done in August revealed Ostertagia ostertagi to represent the most prevalent and abundant species followed by Cooperia oncophora and Nematodirus helvetianus. O. lyrata, O. leptospicularis, C. zurnabada, C. punctata, Trichostrongylus axei, Haemonchus contortus, Bunostomum phlebotomum, Capillaria bovis and Trichuris discolor were less prevalent and abundant. These findings are closely similar to findings from investigations more than 20 years ago although since that time anthelmintics have been used increasingly.