The acid protease Acp1 is produced by Sclerotinia sclerotiorum during plant infection. We explored the mechanism involved in the triggering of that production and found that cyclic AMP played a positive role. Acp1 could be produced in the sole presence of exogenous cyclic AMP. The use of molecules known to increase or decrease the intracellular cyclic AMP levels confirmed the impact of this nucleotide on the protease production and suggested its endogenous site of action. Further pharmacological studies showed the specific effect of cyclic AMP on Acp1 production and suggested that protein kinase A would be its likely target. Together, these results provide the first indication that the production of a pathogenesis-related fungal protease could depend on a cyclic AMP/Protein kinase A signalling pathway.

Sclerotinia sclerotiorum, a necrotrophic fungus, produces many different lytic enzymes. This g which are plant contact, pH, nitrogen and production responds to several input signals among carbon sources. In order to understand the mechanisms that underlie the set up of the plant degrading machinery, we used molecular and biochemical tools to study the regulation of polygalacturonase and protease production during plant infection.

Holstein cows (n = 24) averaging 42 d in milk were used in a randomized complete block design during a 4-wk trial. A control total mixed ration (TMR) was compared with TMR supplemented with Ca salts of fatty acids from canola oil, soybean oil, or linseed oil. The three vegetable oils were progressively more unsaturated; the dominant fatty acids were, respectively, cis-delta-9-C18:1, C18:2, and C18:3. Apparent total tract digestibility of dry matter, crude protein, ether extract, and neutral detergent fiber was higher for rations containing Ca salts than for the control ration. Milk yield increased linearly as the unsaturation of the dominant fatty acid in the Ca salts increased. Milk fat percentage was reduced when Ca salts were added to the rations. The addition of Ca salts to the ration decreased the proportions of saturated fatty acids that contained C6 to C16 and increased the proportions of C18:0, cis-delta-C18:1, and trans-delta-11-C18:1 in milk fat. Proportions of C18:2 and C18:3 increased linearly, and cis-delta-9-C18:1 decreased linearly, as the unsaturation of the dominant fatty acid in the Ca salts increased. The proportion of fat that was liquid at 5 degrees C was higher for butter from cows fed diets containing Ca salts, but the proportion of liquid fat at 20 degrees C was not affected. Calcium salts of unsaturated fatty acids added to the diets of dairy cows improved the thermal properties of milk fat.

Ten Holstein cows in early lactation were used in a replicated 5 × 5 Latin square design to study the effects of MgO and three buffers added to diets containing Ca salts of canola oil fatty acids. Treatments were 1) control (basal diet; no buffer), 2) 1.1% NaHCO3 plus 1.1% KHCO3, 3) 1.9% NaHCO3,4) 0.5% MgO, and 5) 2.0% Na sesquicarbonate (percentage of dry matter). The control diet contained 53% grass silage, 43% concentrate, and 4% Ca salts. Body weight, intake, milk yield, and percentages of milk fat, protein, and lactose were unaffected by treatments. Buffers and MgO tended to increase triacylglycerol extraction by the mammary gland and changed the proportions of some fatty acids in milk. Arterial concentrations of acetate and triacylglycerol were correlated with their respective arteriovenous differences. Extraction by the mammary gland was high for acetate (≈58.2%), triacylglycerol (≈47.3%), propionate (≈34.6%), and glucose (≈24.3%). Extraction of free fatty acids, phospholipids, or cholesterol was negligible. Mammary triacylglycerol arteriovenous difference tended to be higher when MgO was fed than when NaHCO3 was fed. Sodium sesquicarbonate, NaHCO3, and the blend of bicarbonate buffers increased C18:2 in milk fat when compared with the control treatment. The concentration of C18:2 in milk fat decreased when MgO was fed, but the ratio of cis-C18:1 to trans-C18:1 increased compared with effects of dietary NaHCO3. Medium-chain fatty acids in milk fat tended to be higher with Na sesquicarbonate than with NaHCO3. Buffers and MgO modified the profiles of fatty acids in milk.

Holstein cows (n = 24) averaging 39 d of lactation were used in a randomized complete block design during an 8-wk trial. From wk 1 to 4, diets contained 62% alfalfa silage and 38% concentrates (dry matter basis), and, from wk 5 to 8, diets contained 47% forage and 53% concentrates. The concentrates were increased for the second phase so that the effect of bicarbonates could be expressed more fully. Diets 1, 2, and 3 contained 2% of a blend of Na and K bicarbonates and 0, 2, or 4% of Ca salts of canola oil fatty acids (percentage of dry matter), respectively. Diet 4 contained the same percentage of Ca salts as did diet 3 but without bicarbonates. Dry matter intake decreased linearly (wk 4), and milk yield was altered quadratically (wk 4), as the percentage of Ca salts in the diet increased. Milk fat percentage (wk 8) and yield (wk 4 and 8), as well as milk protein percentage (wk 4 and 8) and yield (wk 4), decreased linearly as the percentage of Ca salts in the diet increased. Short- and medium-chain saturated fatty acids decreased linearly, and C18:0, trans-delta-11-C18:1, cis-delta-9-C18:1, cis-delta-11-C18:1, and C18:2 increased linearly, as Ca salts in the diet increased. Addition of Na and K bicarbonates to the diets that contained Ca salts increased milk and milk protein yields and increased the proportions of C18:2 in milk fat at wk 8. Dietary bicarbonates had no effect on the responses of other milk fatty acids to supplementation of 4% Ca salts of canola oil fatty acids.

Twenty-four multiparous Holstein cows (48 d in lactation) were used in an 8-wk trial employing a completely randomized block design to determine the effect of heat treatment of full fat soybeans on the fatty acid composition of milk fat. Treatments consisted of a total mixed ration supplemented with 1) ground raw, 2) extruded, 3) micronized, or 4) roasted soybeans included at 17.5% of the dry matter of the total mixed ration. Dietary treatment had no effect on body weight, yields of milk or solids-corrected milk, fat percentage, or yield of milk fat or protein. Cows fed ground raw soybeans had a higher dry matter intake and a higher percentage of protein in milk than those fed heat-treated soybeans. The concentrations of saturated fatty acids from C8:0 to C16:0 were lower, and cis-delta-9-C18:1 tended to be higher, in the milk fat of cows fed heat-treated soybeans than in the milk fat of cows fed ground raw soybeans. The concentration of trans-delta-11-C18:1 was lowest for cows fed ground raw soybeans, intermediate for cows fed micronized and roasted soybeans, and highest for cows fed extruded soybeans. Cows fed extruded soybeans had lower concentrations of C18:2 and C18:3 than did cows fed micronized or roasted soybeans; cows fed ground raw soybeans had intermediate concentrations of C18:2 and C18:3.

An apparently novel 1,3-β-glucan synthase from the oomycete Saprolegnia monoica has been characterized. The enzyme exhibits properties that differ markedly from those of the enzyme previously described [Fèvre, M. & Dumas, C. (1977). J Gen Microbiol 103, 297-306] as it is active at alkaline pH, stimulated by the divalent cations Ca2+, Mg2+ and Mn2+, and appears to be located mainly in the apical part of the hypha. Taking into consideration the differences in pH optimum and effect of divalent ions, each enzyme activity could be assayed in the presence of the other. The insoluble polymeric product of the enzyme with alkaline pH optimum was characterized as a linear 1,3-β-glucan. Comparisons of the general properties of 1,3-β-glucan synthases suggest that enzymes from the oomycetes are more closely related to enzymes from higher plants than to those of true fungi, reflecting the fact that the oomycetes are highly divergent from chitinous fungi.

Twenty-four multiparous Holstein cows, 4 of which were ruminally fistulated, were assigned to one of four diets containing full-fat soybeans, either raw or extruded at 120, 130, or 140 degrees C. Our hypothesis was that the extrusion of full-fat soybeans, as well as the extrusion temperature, would affect the bypass of fatty acids in the rumen and, thus, would modify the fatty acid profile of milk fat. Total mixed diets containing 23.7% soybeans (percentage of DM) were fed for 8 wk. Milk yield was lower, and the proportion of milk CP was higher, for cows fed raw soybeans than for cows fed extruded soybeans. Compared with raw soybeans, extruded soybeans increased the concentration of delta-11-trans-C18:1 from 2.72 to 11.41% in milk fat but had no effect on yield or percentage of milk fat. Polyunsaturated fatty acids of raw soybeans disappeared more rapidly than did those of extruded soybeans from bags incubated in the rumen of fistulated cows. However, more delta-11-trans-C18:1 and C18:0 appeared in bags containing extruded soybeans than in bags containing raw soybeans. Extrusion of full-fat soybeans influenced the metabolism of fatty acids in the rumen and the fatty acid profile of milk fat, but the temperature of extrusion had only minor effects on these parameters.

Twenty-four multiparous Holstein cows averaging 104 d of lactation were used in a trial with a split-plot design to evaluate the nutritive value of two silages, timothy grass or alfalfa, both treated with formic acid and stored in plastic bag silos. Silages were offered for ad libitum intake either alone or with 17 or 34% (dry matter basis) dry-rolled barley. Both silages contained similar amounts of acid detergent fiber (ADF) (27.5 and 26.7% for timothy grass and alfalfa, respectively). After 110 d of storage, alfalfa silage contained higher amounts of organic acids and NH3 N but had lower soluble N. Total dry matter intake (DMI) and silage DMI were similar between cows fed both silages. Increased barley proportion decreased silage DMI (19.2 to 14.2 kg/d). Apparent total tract digestibility of dry matter was unaffected by treatment. The digestibility of neutral detergent fiber and ADF was higher for the timothy grass silage than for alfalfa silage and was unaffected by the barley percentage added to either silage. Milk yield was lower (23.9 to 22.6 kg/d) for cows fed the highest proportion of barley. The 4% fat-corrected milk yield was unaffected by treatment. Percentages of fat, protein, and total solids in milk were higher for cows fed diets with the higher barley content. Milk fat and protein yields were similar among treatments. Urea in blood was lower for cows fed timothy grass silage than for cows fed alfalfa silage (4.68 vs. 6.23 mg/100 ml). These results suggest that timothy grass silage and alfalfa silage, when stored at a similar ADF content, have comparable nutritive value for midlactation cows.

Eighteen multiparous Holstein cows in midlactation were used in a switchback design trial to evaluate the effect of chop length (3 or 30 mm) of timothy grass silage, containing less than 30% ADF and treated with formic acid, on DMI and cow performance. Within chop length, addition of NaHCO3 (2% of DMI) or the replacement of 30% of silage DM with juice-extracted grass pellets was also evaluated. Cows were fed a TMR composed of 90% silage and 10% concentrate. Silage preservation characteristics were not different between chop lengths. The DMI and apparent digestibility were similar among treatments. Yields of 4% FCM (24.9 vs. 22.7 kg/d), fat (1.03 vs. .93 kg/d), and protein (.83 vs .77 kg/d) were higher with the short chopped silage. Milk protein, milk NPN content, and serum urea were higher for cows fed long chopped silage. Yields of milk and milk constituents were not affected by the addition of juice-extracted grass pellets or NaHCO3. Fat percentage and fatty acid composition of milk remained unchanged by treatments. Reduction of particle size, from 30 to 3 mm, of timothy grass, treated with formic acid at harvest and using compaction at ensiling in bag silos, did not affect silage conservation characteristics but did improve milk, fat, and protein yields when cows were fed high silage diets.

Eighteen midlactation Holstein cows, averaging 80 to 125 d of lactation, were used in a trial of switchback design to evaluate two timothy silages, wilted or direct-cut and treated with formic acid, and three treatments, control, NaHCO3 added at 2% of DM, or juice-extracted grass pellets replacing 30% silage DM. Cows fed direct-cut silage that had been treated with formic acid consumed more DM (19.8 vs. 18.6 kg/d) than those fed wilted silage. The DMI was also increased with NaHCO3 and grass pellet treatments. However, NaHCO3 reduced digestibility of most nutrients without affecting pH of rumen fluid or degradation of DM and NDF. Compared with wilted silage, direct-cut silage that had been treated with formic acid contained more degradable NDF (86 vs. 84.5%). Milk yield (24.9 vs. 23.6 kg/d) was higher for cows fed direct-cut than wilted silage, but 4% FCM yield remained unchanged. Addition of NaHCO3 tended to increase yields of milk and 4% FCM. Treatments did not affect milk composition. Serum urea N was higher for cows fed the direct-cut silage than for cows fed wilted silage. Silage type had more impact on feed intake and performance than did NaHCO3 or juice-extracted grass pellets.

Theophylline has been reported to inhibit excitatory noncholinergic but not cholinergic-neurotransmission in guinea-pig bronchi. As theophylline might exert this effect through an inhibition of phosphodiesterases (PDE), and since many types of PDE have now been described, the aim of this study was to investigate the effects of three specific inhibitors of PDE on the electrical field stimulation (EFS) of the guinea-pig isolated main bronchus in vitro. The drugs used were siguazodan, rolipram and zaprinast, which specifically inhibit PDE types, III, IV and V, respectively. Guinea-pig bronchi were stimulated transmurally with biphasic pulses (16 Hz, 1 ms, 320 mA for 10 s) in the presence of indomethacin 10(-6) M and propranolol 10(-6) M. Two successive contractile responses were observed: a rapid cholinergic contraction, followed by a long-lasting contraction due to a local release of neuropeptides from C-fibre endings. Rolipram (10(-9) to 10(-6) M) but not siguazodan or zaprinast, inhibited the peptidergic contraction in a concentration-dependent manner. Conversely, the cholinergic response was unaffected. Contractile responses induced by exogenous acetylcholine (10(-8) to 10(-3) M) or [Nle10]NKA(4-10) (10(-10) to 10(-6) M) were also unaffected by rolipram, siguazodan and zaprinast (10(-7) M). These results demonstrate that concentrations of rolipram, similar to those which inhibit PDE, reduce the release of sensory neuropeptides from C-fibre endings, and suggest that the cyclic adenosine monophosphate (AMP) PDE type IV is specifically involved in this effect, as in other anti-inflammatory effects.

La famille forme le cadre nécessire au développement psychologique de l'enfant.

Enriched preparations of chitin synthase were obtained from cell homogenates from Saprolegnia monoica. Chitin synthase was solubilized from a mixed membrane fraction by two successive digitonin treatments. Glycerol gradient centrifugation of the solubilized proteins separated the chitin synthase activity from the majority of proteins and from beta-1,3 and beta-1,4 glucan synthases. The properties of chitin synthase from this Oomycete fungus are similar to those reported for the enzymes of chitinous fungi. The solubilized enzyme catalysed the synthesis in vitro of spindle-like crystals of chitin. Biophysical analysis (by electron and X-ray diffractometry and infrared spectroscopy) demonstrated that the polymer synthesized in vitro was alpha-chitin. These results and those previously reported demonstrate unambiguously that chitin synthase and chitin are normal components of the cell wall of the cellulosic fungus S. monoica.

Bull (n = 30) and steer (n = 30) carcasses were measured for meat quality in the longissimus muscle after 48 hr on electrically stimulated (ES) sides or after 6 days on their unstimulated counterparts. Electrical stimulation and 48 hr aging had the same tenderizing effect as a 6 day aging period on both bulls and steers. Compared to meat from steers, that from bulls was tougher and had higher ultimate pH values and collagen contents. On an overall quality basis however, ES meat aged for 48 hrs was equivalent to nonstimulated meat aged for 6 days.

The presence of α chitin in hyphal walls and regenerating protoplast walls of the cellulosic fungus Saprolegnia monoica has been demonstrated by biophysical analyses. In hyphal walls, chitin appeared as globular particles while in protoplast walls chitin had a microfibrillar structure. Chitin synthesized in vitro by partially purified soluble chitin synthase appeared as spindle-like particles. This variation of morphology may reflect the structural role played by crystalline chitin during morphogenesis.

The membrane-localized 1,3-β-glucan synthase (EC 2.4.1.34; UDPglucose: 1,3-β-d-glucan 3-β-glucosyltransferase) from Saprolegnia monoica was greatly enriched by a two-step purification procedure. Starting with a microsomal preparation, the enzyme was solubilized with a cholamidopropyl-dimethyl-ammonio-propanesulfonate (Chaps)/octylglucoside mixture and further purified by ultracentrifugation on a linear glycerol density gradient. The most purified enzyme preparation showed enrichment in 34, 48 and 50 kDa polypeptides. The true involvement of these three bands in the 1,3-β-glucan synthase structure was demonstrated by three experimental procedures. After enzyme purification, entrapment gave selective synthase sedimentation with its reaction product and subsequent SDS-PAGE electrophoresis showed the presence of these three polypeptides. Lectin binding on solubilized preparation provoked a shift in the density gradient of the synthase activity correlating with the distribution of these three bands. Western analysis also showed that the doublet 48, 50 kDa was glycosylated. Finally, the three bands were used to raise polyspecific antibodies which precipitated 1,3-β-glucan synthase activity.

Sixty Canadian Holstein male calves (mean weight 261 kg), previously fed all-concentrate diets, were allocated to one of six different sequences of barley supplementation. Rolled barley was provided in the diet at three levels (0, 17 and 34% on a DM basis) with either grass or corn silages, and fed as a total mixed ration for 91, 77 and 70 d during periods 1,2, and 3, respectively, to ensure consumption of 330 kg of barley per animal for the entire experiment. Barley supplementation in the last period improved average daily gain over the whole experiment by 0.223 kg d−1 (P = 0.0154), and feed efficiency by 55% (P = 0.0062) compared to inclusion of barley in the silage mixtures during the first period. Feed efficiency improved with concentrate supplementation of both silages (P ≤ 0.0001), but the improvement with grass silage was greater than with corn silage (P = 0.0041). Dry matter and energy digestibilities were not affected by feed sequences (P = 0.1025, P = 0.0833, respectively), but they were higher for mixed diets than for pure silage (P = 0.0186, P = 0.0218, respectively). Dry matter intake was 0.81 kg d−1 higher (P ≤ 0.0001) when concentrate was added to the diet. These results suggest that energy feeds such as barley are more efficiently used in the last part of growth. This experiment demonstrates the feasibility of switching grain-fed calves to roughage diets. Key words: Barley, dairy bulls, energy distribution, corn silage, grass silage

The presence of chitin in hyphal cell walls and regenerating protoplast walls ofSaprolegnia monoi¨ca was demonstrated by biochemical and biophysical analyses. α-Chitin was characterized by X-ray diffraction, electron diffraction, and infrared spectroscopy. In hyphal cell walls, chitin appeared as small globular particles while cellulose, the other crystalline cell wall component, had a microfibrillar structure. Chitin synthesis was demonstrated in regenerating protoplasts by the incorporation of radioactiveN-acetylglucosamine into a KOH-insoluble product. Chitin synthase activity of cell-free extracts was particulate. This activity was stimulated by trypsin and inhibited by the competitive inhibitor polyoxin D (Ki 20 μM). The reaction product was insoluble in 1M KOH or 1M acetic acid and was hydrolyzed by chitinase into diacetylchitobiose. Fungal growth and cell wall chitin content were reduced when mycelia were grown in the presence of polyoxin D. However, hyphal morphology was not altered by the presence of the antibiotic indicating that chitin does not seem to play an important role in the morphogenesis ofSaprolegnia.