Utilization of Carbohydrates by Rhynchosporium secalis. I. Growth and Sporulation on Glucose, Galactose, and Galacturonic acid

The fungus Rhynchosporium secalis (Oud.) Davis grew and speculated in liquid nutrient media that contained glucose, galactose or galacturonic acid, or a pair of those substances, as the sole carbon source. Sporulation was inhibited by high concentrations of glucose and galacturonic acid. Growth and sporulation were greatest on glucose, and least on galactose. Growth was increased when glucose and galacturonic acid were mixed. Nitrogen concentration affected sporulation but not growth.     

Observations on the production and dispersal of spores, and infection by Rhynchosporium secalis

The numbers of spores of Rhynchosporium secalis washed from samples of barley plants taken weekly, varied markedly. No consistent association with amount of previous rainfall or length of the preceding dry period was detected. On potted seedlings exposed within a crop most infection occurred during long rain periods or when rain fell late in the evening; fewest lesions usually developed on seedlings prevented from touching adjacent plants. Rotorod traps fitted with a 13 mm diameter disc at the apex of each arm were operated under 24 cm diameter covers. Spores were collected on circular cover slips fixed to each disc with glycerine jelly. At all stages of crop growth more spores were trapped at ground level than at other heights tested up to 1 m. The number of spores trapped was not related to the quantity of or duration of rainfall but was related to the mean rate of fall during brief showers only. Efficiency of droplet retention was assessed in a wind tunnel. It declined rapidly when more than c. 5 pl was presented to the disc surface and was less at a wind speed of 1.0 than 0.5 m s-^l. Spore distribution on discs indicated that spores were washed from the surface during long rain periods.     

Transport and Metabolism of Lactose, Glucose, and Galactose in Homofermentative Lactobacilli

A number of species of lactobacilli were examined for their ability to ferment both the glucose and galactose moieties of lactose. Lactobacillus helveticus strains metabolized both the glucose and galactose moieties, whereas L. bulgaricus, L. lactis, and L. acidophilus strains metabolized only the glucose moiety and released galactose into the growth medium. All four species tested contained β-galactosidase activity, and no significant phospho-β-galactosidase activity was observed. L. bulgaricus and L. helveticus had a phosphoenolpyruvate (PEP):glucose phosphotransferase system for the uptake of glucose, but no evidence for a PEP:lactose phosphotransferase or PEP:galactose phosphotransferase system was obtained.     

Evolution of resistance to Rhynchosporium secalis (Oud.) Davis in barley composite cross II

Summary Changes in resistance to scald disease which occurred in barley composite Cross II over 45 generations were analyzed genetically. This population, which was synthesized in 1929 by pooling equal numbers of f₁ seeds from 378 pair wise crosses among 28 barley varieties, has subsequently been grown at Davis, California under standard agricultural conditions without conscious selection. Progenies derived from self-pollinated seeds from random plants taken from four generations (F₈, F₁₃, F₂₃, and F₄₅) were tested against four different races of scald (40, 61, 72, and 74), and rated as resistant, susceptible or segregating. Striking increases in the frequency of families resistant to races 40, 61, and 74 occurred in CC II. A test for randomness showed that quadruply susceptible and triply resistant families were more common than expected under the assumption that resistance to different races is independent. Positive correlations were found between resistance to races 40, 61, and 74, but resistance to race 72 was independent of resistance to all other races. Possible reasons for these correlations are discussed.     

The origin and colonization history of the barley scald pathogen Rhynchosporium secalis

The origins of pathogens and their past and present migration patterns are often unknown. We used phylogenetic haplotype clustering in conjunction with model-based coalescent approaches to reconstruct the genetic history of the barley leaf pathogen Rhynchosporium secalis using the avirulence gene NIP1 and its flanking regions. Our results falsify the hypothesis that R. secalis emerged in association with its host during the domestication of barley 10,000 to 15,000 years ago in the Fertile Crescent and was introduced into Europe through the migration of Neolithic farmers. Estimates of time since most recent common ancestor (2500-5000 BP) placed the emergence of R. secalis clearly after the domestication of barley. We propose that modern populations of R. secalis originated in northern Europe following a host switch, most probably from a wild grass onto cultivated barley shortly after barley was introduced into northern Europe. R. secalis subsequently spread southwards into already established European barley-growing areas.     

Metabolism of Carbohydrates by Pasteurella pseudotuberculosis

Cell-free extracts of Pasteurella pseudotuberculosis and P. pestis catalyzed a rapid and reversible exchange of electrons between pyridine nucleotides. Although the extent of this exchange approximated that promoted by the soluble nicotinamide adenine dinucleotide (phosphate) transhydrogenase of Pseudomonas fluorescens, the reaction in the pasteurellae was associated with a particulate fraction and was not influenced by adenosine-2'-monophosphate. The ability of P. pseudotuberculosis to utilize this system for the maintenance of a large pool of nicotinamide adenine dinucleotide phosphate could not be correlated with significant participation of the Entner-Doudoroff path or catabolic use of the hexose-monophosphate path during metabolism of glucose. As judged by the distribution of radioactivity in metabolic pyruvate, glucose and gluconate were fermented via the Embden-Meyerhof and Entner-Doudoroff paths, respectively. With the exception of hexosediphosphatase, all enzymes of the three paths were detected, although little or no gluconokinase or phosphogluconate dehydrase was present unless the organisms were cultivated with gluconate. The significance of these findings is discussed with respect to the regulation of carbohydrate metabolism in the pasteurellae, related enteric bacteria, and P. fluorescens.     

Cooperative Anti-Diabetic Effects of Deoxynojirimycin-Polysaccharide by Inhibiting Glucose Absorption and Modulating Glucose Metabolism in Streptozotocin-Induced Diabetic Mice

We had previously shown that deoxynojirimycin-polysaccharide mixture (DPM) not only decreased blood glucose but also reversed the damage to pancreatic β-cells in diabetic mice, and that the anti-hyperglycemic efficacy of this combination was better than that of 1-deoxynojirimycin (DNJ) or polysachharide alone. However, the mechanisms behind these effects were not fully understood. The present study aimed to evaluate the therapeutic effects of DPM on streptozotocin (STZ)-induced diabetic symptoms and their potential mechanisms. Diabetic mice were treated with DPM (150 mg/kg body weight) for 90 days and continued to be fed without DPM for an additional 30 days. Strikingly, decrease of blood glucose levels was observed in all DPM treated diabetic mice, which persisted 30 days after cessation of DPM administration. Significant decrease of glycosylated hemoglobin and hepatic pyruvate concentrations, along with marked increase of serum insulin and hepatic glycogen levels were detected in DPM treated diabetic mice. Results of a labeled ¹³C₆-glucose uptake assay indicated that DPM can restrain glucose absorption. Additionally, DPM down-regulated the mRNA and protein expression of jejunal Na⁺/glucose cotransporter, Na⁺/K⁺-ATPase and glucose transporter 2, and enhanced the activities as well as mRNA and protein levels of hepatic glycolysis enzymes (glucokinase, phosphofructokinase, private kinase and pyruvate decarboxylas E1). Activity and expression of hepatic gluconeogenesis enzymes (phosphoenolpyruvate carboxykinase and glucose-6-phosphatase) were also found to be attenuated in diabetic mice treated with DPM. Purified enzyme activity assays verified that the increased activities of glucose glycolysis enzymes resulted not from their direct activation, but from the relative increase in protein expression. Importantly, our histopathological observations support the results of our biochemical analyses and validate the protective effects of DPM on STZ-induced damage to the pancreas. Thus, DPM has significant potential as a therapeutic agent against diabetes.     

Remodeling of Oxidative Energy Metabolism by Galactose Improves Glucose Handling and Metabolic Switching in Human Skeletal Muscle Cells

Cultured human myotubes have a low mitochondrial oxidative potential. This study aims to remodel energy metabolism in myotubes by replacing glucose with galactose during growth and differentiation to ultimately examine the consequences for fatty acid and glucose metabolism. Exposure to galactose showed an increased [¹⁴C]oleic acid oxidation, whereas cellular uptake of oleic acid uptake was unchanged. On the other hand, both cellular uptake and oxidation of [¹⁴C]glucose increased in myotubes exposed to galactose. In the presence of the mitochondrial uncoupler carbonylcyanide p-trifluormethoxy-phenylhydrazone (FCCP) the reserve capacity for glucose oxidation was increased in cells grown with galactose. Staining and live imaging of the cells showed that myotubes exposed to galactose had a significant increase in mitochondrial and neutral lipid content. Suppressibility of fatty acid oxidation by acute addition of glucose was increased compared to cells grown in presence of glucose. In summary, we show that cells grown in galactose were more oxidative, had increased oxidative capacity and higher mitochondrial content, and showed an increased glucose handling. Interestingly, cells exposed to galactose showed an increased suppressibility of fatty acid metabolism. Thus, galactose improved glucose metabolism and metabolic switching of myotubes, representing a cell model that may be valuable for metabolic studies related to insulin resistance and disorders involving mitochondrial impairments.     

The relationship between leaf blotch caused by Rhynchosporium secalis and losses in grain yield of spring barley

Two methods were used to investigate the loss in grain yield associated with specific levels of leaf blotch. Yields from plots sprayed with fungicide were compared with those from unsprayed plots and yields of varieties of different susceptibility to the disease were compared with one another. A disease assessment key is presented, which was used to assess the percentage laminar area of the top two leaves affected by the disease. A linear relationship between disease on the upper two leaves and yield was established. Results from nine trials showed a consistent relationship between the disease level, at growth stage 11·1 (Feekes scale), and loss in yield. The loss in yield expressed as a percentage of the yield of an uninfected crop was equivalent to approximately two-thirds of the percentage of the flag-leaf area visibly infected, or one-half of the infected area on the second leaf. The predicted loss in yield is the average of these two estimates.     

Stimulation of Barley Plasmalemma H+-ATPase by Phytotoxic Peptides from the Fungal Pathogen Rhynchosporium secalis

A small family of necrosis-inducing peptides has been identified as virulence factors of Rhynchosporium secalis, a fungal pathogen of barley (Hordeum vulgare L.) Two members of this family, NIP1 and NIP3, were found to stimulate the phosphohydrolyzing activity of the Mg2+-dependent, K+-stimulated H+-ATPase of plasma membrane vesicles isolated from barley leaves by partitioning in an aqueous two-phase system. Stimulation of enzyme activity was saturated by 10 to 15 [mu]M fungal protein. Another member of the peptide family, NIP2, did not affect the enzyme, indicating that it has a different mode of action.     

Utilization of Lactose, Glucose, and Galactose by a Mixed Culture of Streptococcus thermophilus and Lactobacillus bulgaricus in Milk Treated with Lactase Enzyme

The mechanism responsible for an increased rate of acid production when yogurt starter cultures are grown in milk treated with lactase enzyme was investigated by studying carbohydrate utilization and acid development by a pure culture of Streptococcus thermophilus and a mixed yogurt starter culture consisting of S. thermophilus and Lactobacillus bulgaricus. In milk containing glucose, galactose, and lactose, glucose and lactose (but not free galactose) were fermented. Fermentation of lactose in control milk was accompanied by the release of free galactose, with the result that carbohydrate utilization was less efficient than in treated milk. This phenomenon also occurred when lactose was fermented by S. thermophilus in broth culture. Carbohydrate utilization by the mixed yogurt culture was more rapid when the lactose in milk was partially prehydrolyzed. Our results suggest that the more rapid acid development that took place when a mixed yogurt starter culture was grown in milk containing prehydrolyzed lactose was the result of a more rapid and efficient utilization of carbohydrate by S. thermophilus when free glucose in addition to lactose was available for fermentation. The evidence presented also suggests that uptake and utilization of glucose and lactose by S. thermophilus are different in broth and milk cultures.     

Uptake and Metabolism of Carbohydrates by Bradyrhizobium japonicum Bacteroids

Bradyrhizobium japonicum bacteroids were isolated anaerobically and were supplied with ¹⁴C-labeled trehalose, sucrose, UDP-glucose, glucose, or fructose under low O₂ (2% in the gas phase). Uptake and conversion of ¹⁴C to CO₂ were measured at intervals up to 90 minutes. Of the five compounds studied, UDP-glucose was most rapidly absorbed but it was very slowly metabolized. Trehalose was the sugar most rapidly converted to CO₂, and fructose was respired at a rate at least double that of glucose. Sucrose and glucose were converted to CO₂ at a very low but measurable rate (<0.1 nanomoles per milligram protein per hour). Carbon Number 1 of glucose appeared in CO₂ at a rate 30 times greater than the conversion of carbon Number 6 to CO₂, indicating high activity of the pentose phosphate pathway. Enzymes of the Entner-Doudoroff pathway were not detected in bacteroids, but very low activities of sucrose synthase and phosphofructokinase were demonstrated. Although metabolism of sugars by B. japonicum bacteroids was clearly demonstrated, the rate of sugar uptake was only 1/30 to 1/50 the rate of succinate uptake. The overall results support the view that, although bacteroids metabolize sugars, the rates are very low and are inadequate to support nitrogenase.     

Carbohydrate Metabolism in Leukocytes VII. Metabolism of Glucose, Acetate, and Propionate by Human Plasma Cells

Plasma cells obtained from the peripheral blood of a patient with multiple myeloma was incubated in serum and Krebs-Ringer bicarbonate buffer with (14)C-labeled glucose, acetate, and propionate. Glucose utilization by these cells amounted to 0.5 mumole per hr per 10(8) cells and was mainly via the Embden-Meyerhof pathway, and only 6% or less traversed the hexose monophosphate shunt. The presence of Krebs cycle activity was demonstrated by direct isolation of several labeled intermediates after incubation with either (14)C-acetate or (14)C-propionate. The distribution of (14)C in lactate, succinate, fumarate, malate, aspartate, and glutamate indicate a complete Krebs cycle. Acetate was metabolized via the Krebs cycle to the extent of 0.15 mumoles per hr per 10(8) cells, and the rate of propionate utilization was 0.17 mumoles per hr per 10(8) cells.     

Abnormal Behaviour of Stomata in Barley Leaves Infected with Rhynchosporium secalis (Oudem.) J. J. Davis

Infection of barley leaves by the fungus Rhynchosporium secalisincreases the degree to which stomata open in the light. Openingis enhanced at CO₂, levels between 0 and 4000 parts/10⁶. Theability of stomata to close in the dark, as normal, is retaineduntil an advanced stage of tissue necrosis is reached. Increased stomatal opening is confined to those areas of theleaf which have been colonized by the fungus. Abnormal stomatalbehaviour results from the loss of osmotically active substancesfrom the epidermis of diseased leaves with a consequent alterationof the turgor relations between guard cells and their surroundingepidermal cells.     

Role of galK and galM in Galactose Metabolism by Streptococcus thermophilus

Streptococcus thermophilus is unable to metabolize the galactose moiety of lactose. In this paper, we show that a transformant of S. thermophilus SMQ-301 expressing Streptococcus salivarius galK and galM was able to grow on galactose and expelled at least twofold less galactose into the medium during growth on lactose.