Streptomyces Pepsin Inhibitor-Insensitive Carboxyl Proteinase from Lentinus edodes

A Streptomyces-pepsin inhibitor (S-PI or Pepstatin Ac), and DAN-insensitive carboxyl proteinase was found in a still culture filtrate of Lentinus edodes. The new carboxyl proteinase was purified, and about 9 mg purified enzyme was obtained from 19 liters of culture filtrate, with 11% recovery. The enzyme showed a single band on polyacrylamide gel electrophoresis. The molecular weight and isoelectric point were 40,000 and pH 4.2, respectively. The enzyme did not contain histidine and was composed of 387 amino acid residues. The enzyme was most active between pH 2.7 ~ 2.9, and stable over a pH of 3.2 ~ 5.2 for 3 hr at 37°C. The enzyme was not inhibited by S-PI or synthetic pepsin inhibitors such as DAN and EPNP. The physicochemical and enzymological properties were very similar to those of Scytalidium lignicolum carboxyl proteinase A, which was reported to be an S-PI-, and DAN-insensitive carboxyl proteinase.     

Absolute Configuration of (+)-1-Acetoxy-p-menth-3-one Isolated from Essential Oil of Mentha gentilis (L.)

The most effective electro-energizing fermentation (E-E F) conditions for l-glutamate (l-Glu) production by Brevibacterium flavum No. 2247 were determined. The adding of 0.01 mm neutral red at the beginning of cultivation was found most effective. A 1.5 V direct current was applied to the culture broth at 6~8 hr after inoculation in the cathode compartment, l-Glu was produced at 51.0 mg per ml, and this is about a 15 % increase in yield compared to the yield of the not electro-energizing (E-E) control (44.3 mg/ml).     

Fermentation of plant cell walls by ruminal bacteria,protozoa and fungi and their interaction with fibre particle size

Abstract

The objective of the experiment was to evaluate the contribution of various ruminal microbial groups to the fermentation of cell walls of corn stover with different particle sizes based on ruminal gas production in vitro. Physical, chemical, and antibiotical methods were used to differentiate groups of bacteria, protozoa and fungi in rumen fluid, offering following rumen microbial groups: whole rumen fluid (WRF), bacterial (B), protozoal (P), fungal (F), bacterial plus protozoal (B + P), bacterial plus fungal (B + F), protozoal plus fungal (P + F), and negative control (CON). Cell walls from corn stover were ground and ball milled to produce two different particle sizes. The results showed that digestion of the cell walls was undertaken by the interaction among ruminal bacteria, protozoa and fungi, and such co-actions seemed to fail alternation by one of three microbial groups or any combinations. However, B + P group showed a significant contribution to the degradation of milled cell walls, and B + F group revealed a great synergy effect on the ground cell walls degradation. Particle size of cell walls also had a considerable influence on their fermentation extent instead of the fermentative patterns by various rumen microbial groups.     

Effects of nitrate adaptation by rumen inocula donors and substrate fiber proportion on in vitro nitrate disappearance,methanogenesis, and rumen fermentation acid

A study was conducted to evaluate the main effects of dietary nitrate adaptation by cattle and alfalfa cell wall to starch ratio in in vitro substrates on nitrate disappearance and nitrite and volatile fatty acid (VFA) concentrations, as well as hydrogen (H₂) and methane (CH₄) accumulations. Rumen fluid from steers fed diets containing urea or nitrate was added into in vitro incubations containing sodium nitrate as the sole nitrogen source and 20 cell wall : 80 starch or 80 cell wall : 20 starch as the carbohydrate source. The results showed that during 24 h incubation, rumen fluid inoculums from steers adapted to dietary nitrate resulted in more rapid nitrate disappearance by 6 h of incubation (P < 0.01), no significant effect on nitrite concentration and diminished CH₄ accumulation (P < 0.05). Cell wall to starch ratio did not affect nitrate disappearance, CH₄ accumulation and total VFA concentration. The higher cell wall ratio had the lower total gas production and H₂ concentration (P < 0.05). Ammonia-N (NH₃-N) concentration increased because of adaptation of donors to nitrate feeding (P < 0.05). Nitrate adaptation did not alter total VFA concentration, but increased acetate, and decreased propionate and butyrate molar proportions (P < 0.01).     

Effects of rumen-escape starch and coarseness of ingredients in pelleted concentrates on performance and rumen wall characteristics of rosé veal calves

The objective was to study the effect of rumen-escape starch and coarseness of ingredients in pelleted concentrates on performance, carcass quality and rumen wall characteristics in rosé veal calf production. Two alternative concentrates (Coarse and Slow) were compared with a traditional (Control) concentrate. Control was based on finely ground ingredients, whereas in Coarse, the same ingredients were coarsely ground resulting in a mean particle size before pelleting of 1.5 in Coarse and 0.6 mm in Control. Slow compared with Control and Coarse contained finely ground sorghum and corn instead of barley and wheat which increased the amount of rumen-escape starch to 59 compared with 22 g/kg in Control and Coarse. All concentrates had the same total starch (362 g/kg), NDF (168 g/kg), CP (154 g/kg) and DE (15.5 MJ/kg DM) content and a pellet diameter of 3.5 to 4 mm. Use of an ‘indicator of starch digestibility’ method gave a value of 98.6% for Control and Coarse and 91.1% for Slow (P < 0.001). A total of 57 Holstein bull calves (n = 19 per treatment) were offered one of the three concentrates ad libitum from weaning 2 1/2 months of age) to slaughter (<10 months of age). Concentrate intake was recorded individually. Barley straw was available ad libitum but intake was not recorded. Average daily gain (1.43 kg/day), concentrate conversion efficiency (3.7 kg DM concentrate/kg gain), LW at slaughter (386 kg), carcass weight (194 kg) and EUROP conformation (3.9) were not affected by type of concentrate (P > 0.05). Papillae length and shape evaluated in atrium ruminis and the cranial part of the ventral rumen sac at slaughter were not affected by type of concentrate (P > 0.05). Rumen wall characteristics showed degrees of plaque formation (i.e., papillary aggregation), hyperaemia and necrotic areas in all treatment groups, but with no general difference between type of concentrate (P > 0.05). Incidence of liver abscesses (LAs, 16%) was not affected by type of concentrate (P > 0.05). There were no differences in performance or rumen wall characteristics between liver-abscessed and non-abscessed calves. The results show a high level of production performance with the three types of pelleted concentrates and indicates that neither the more coarse ingredients nor the additional rumen-escape starch tested, when fed ad libitum, could improve rumen wall characteristics or reduce LAs of rosé veal calves.     

Contrasting geographic patterns of genetic variation for molecular markers vs. phenotypic traits in the energy grass Miscanthus sinensis

Species and hybrids of Miscanthus are a promising energy crop, but their outcrossing mating systems and perennial life cycles are serious challenges for breeding programs. One approach to accelerating the domestication of Miscanthus is to harness the tremendous genetic variation that is present within this genus using phenotypic data from extensive field trials, high‐density genotyping and sequencing technologies, and rapidly developing statistical methods of relating phenotype to genotype. The success of this approach, however, hinges on detailed knowledge about the population genetic structure of the germplasm used in the breeding program. We therefore used data for 120 single‐nucleotide polymorphism and 52 simple sequence repeat markers to depict patterns of putatively neutral population structure among 244 Miscanthus genotypes grown in a field trial near Aberystwyth (UK) and delineate a population of 145 M . sinensis genotypes that will be used for association mapping and genomic selection. Comparative multivariate analyses of molecular marker and phenotypic data for 17 traits related to phenology, morphology/biomass, and cell wall composition revealed significant geographic patterns in this population. A longitudinal cline accounted for a substantial proportion of molecular marker variation (R² = 0.60, P = 3.4 × 10^?15). In contrast, genetic variation for phenotypic traits tended to follow latitudinal and altitudinal gradients, with several traits appearing to have been affected by divergent selection (i.e., Q_ST >> F_ST). These contrasting geographic trends are unusual relative to other plants and provide opportunities for powerful studies of phenotype–genotype associations and the evolutionary history of M. sinensis.     

Melanin is crucial for growth of the black yeast Hortaea werneckii in its natural hypersaline environment

Melanin has an important role in the ability of fungi to survive extreme conditions, like the high NaCl concentrations that are typical of hypersaline environments. The black fungus Hortaea werneckii that has been isolated from such environments has 1,8-dihydroxynaphthalene-melanin incorporated into the cell wall, which minimises the loss of glycerol at low NaCl concentrations. To further explore the role of melanin in the extremely halotolerant character of H. werneckii, we studied the effects of several melanin biosynthesis inhibitors on its growth, pigmentation and cell morphology. The most potent inhibitors were a 2,3-dihydrobenzofuran derivative and tricyclazole, which restricted the growth of H. werneckii on high-salinity media, as shown by growth curves and plate-drop assays. These inhibitors promoted release of the pigments from the H. werneckii cell surface and changed the medium colour. Inhibitor-treated H. werneckii cells exposed to high salinity showed both decreased and increased cell lengths. We speculate that this absence of melanin perturbs the integrity of the cell wall in H. werneckii, which affects its cell division and exposes it to the harmful effects of high NaCl concentrations. Surprisingly, melanin had no effect on H. werneckii survival under H₂O₂ oxidative stress.