P and N deficiency change the relative abundance and function of rhizosphere microorganisms during cluster root development of white lupin (Lupinus albus L.)

We studied microbe-plant interactions of white lupin, a cluster root-forming plant, under low P and N conditions to examine increased nutrient acquisition by plants either by a shift to a more specialized microbial community or changes in microbial enzyme production. White lupin plants were grown in rhizoboxes filled with either P- or N-deficient soil; fertilized soil was used as control. After cultivation of plants in a glasshouse for 41 d, plant growth (shoot and roots) and P and N accumulation in shoots were measured. Microbial functions were analyzed by P- and N-cycling enzymes. The microbial community structure was estimated by fingerprinting (denaturing gradient gel electrophoresis) and sequencing techniques. P deficiency induced the released citrate and acid phosphomonoesterases from cluster roots and stimulated the production of microbe-derived alkaline phosphomonoesterase in the rhizosphere. P deficiency decreased microbial diversity in the cluster root rhizosphere. Increased relative abundance of Burkholderiales in the rhizosphere of P deficient plants might be responsible for the degradation of different organic P fractions such as phytates. N deficiency induced an increase of the number of nodules and P concentration in shoot as well as roots of white lupin. We clarified that high release of citrate from cluster roots might be the preferred mechanisms to meet the P demand of nodulated plants under N deficiency. In addition, the high abundance of Rhizobiales and Rhodospirillales in the rhizosphere of cluster roots showed that the importance of N-fixing microorganisms under N deficiency. The contribution of rhizosphere microorganisms due to similar activities of N-cycling enzymes under the two different N treatments is less important for N nutrition of plants. Further understanding of the regulation of cluster roots under N-deficiency will provide new information on the interactions between P and N nutrition.     

Analysis of acrylamide in green tea by gas chromatography-mass spectrometry

Optimization of the solid-phase extraction cleanup procedure enabled the GC-MS analysis of acrylamide in tea samples without the interference of bromination by tea catechins. Although polyvinylpolypyrrolidone (PVPP) is available for removing tea catechins from tea extract, the peaks derived from PVPP had the same retention time as brominated acrylamide in mass chromatograms obtained by GC-MS. A considerable amount of acrylamide was formed at roasting temperatures of > or =120 degrees C; the highest acrylamide level was observed when tea samples were roasted at 180 degrees C for 10 min. Higher temperatures and longer processing times caused a decrease in the acrylamide content. Furthermore, an analysis of 82 tea samples showed that rather than the reducing sugar content, the asparagine content in tea leaves was a significant factor related to acrylamide formation in roasted products. The acrylamide level in roasted tea products was controlled by asparagine in the presence of reducing sugars.     

Effects of feeding level of milk replacer on body growth,plasma metabolite and insulin concentrations,and visceral organ growth of suckling calves

The objective was to evaluate effects of feeding level of milk replacer on body growth, plasma metabolite and insulin concentrations, and allometric growth of visceral organs in suckling calves. Holstein bull calves (n = 8; 3–4 days of age) were fed either a low amount (average 0.63 kgDM/day, LM) or high amount (average 1.15 kgDM/day, HM) of high protein milk replacer until they were slaughtered at 6 weeks of age. Body weight (BW) at 4, 5, and 6 weeks of age, feed intake, average daily gain, and feed efficiency were higher in the HM than LM calves. The HM group had higher plasma glucose at 3 and 4 weeks of age and insulin levels after the age of 4 weeks compared with LM calves whereas no effect was detected on plasma nonesterified fatty acid or urea nitrogen concentrations. The HM calves had greater empty body weight (EBW), viscera‐free BW and most of the organs dissected than LM calves. Relative weights (% of EBW) of liver, spleen, kidneys, and internal fat were higher, whereas head and large intestine was lower in HM than LM calves. The results suggest that increased milk feeding levels would accelerate the growth of the body and specific organs.     

Epigallocatechin gallate promotes the vasorelaxation power of the antiatherosclerotic dipeptide trp-his in contracted rat aorta

The aim of this study was to demonstrate the enhancement of the vasorelaxation power of the antiatherosclerotic voltage-dependent L-type Ca(2+) channel (VDCC)-blocking peptide Trp-His by epigallocatechin gallate (EGCg). We found that 300 μM EGCg dramatically enhanced the magnitude of Trp-His-induced vasorelaxation by a factor of >6 (EC(50) of Trp-His: EGCg(-), 2.80 ± 0.05 mM; EGCg(+), 0.45 ± 0.04 mM) in phenylephrine-contracted rat aorta. The enhancing effect of EGCg was completely abolished in endothelium-removed aorta and high K(+)-contracted aorta. The enhancement of Trp-His-induced vasorelaxation by EGCg was significantly diminished by either N(G)-monomethyl-l-arginine acetate (NO synthase (NOS) inhibitor) or 1-H-[1,2,4]oxadiazolo[4,3]quinoxalin-1-one (soluble guanylyl cyclase inhibitor), together with the enhancement of NOS activity by EGCg. These results indicate that the enhancing effect of EGCg in Trp-His-induced vasorelaxation may be involved in the activation of NO/cGMP pathway.     

Analysis of bacterial communities in <Emphasis Type="Italic">Nannochloropsis</Emphasis> sp. cultures used for larval fish production

ABSTRACT:   Phytoplankton used in fish hatcheries is mass-cultured in the open air and usually contains large numbers of bacteria. In commercial fish production, the phytoplankton cultures are usually added into the larval rearing tanks; however, the numbers and types of bacteria introduced into the rearing tanks simultaneously are unknown. In this study, the bacterial community structures in Nannochloropsis sp. cultures were analyzed by using fluorescence in situ hybridization (FISH). A direct viable count (DVC)-FISH analysis was also performed as DVC is useful for the detection of actively growing cells. Total numbers of bacteria in Nannochloropsis sp. cultures ranged from 7.72 × 10⁵−2.39 × 10⁶ cells/mL. High proportions of the total bacteria (31.6–53.6%) in the Nannochloropsis sp. cultures showed growth potential. DVC-FISH analysis revealed that α-proteobacteria and the Cytophaga – Flavobacterium cluster were abundant in the bacterial community of actively growing cells. Thus, the high growth potentials of the distinct bacterial communities in Nannochloropsis sp. culture must influence the bacterial communities in larval rearing tanks.     

Development of avian embryo manipulation techniques and their application to germ cell manipulation

The development of chicken embryo culture techniques, from single‐cell stage to hatching, makes it possible to manipulate developing embryos at any developmental stage. Production of germline chimeric chickens by the transfer of stage X blastodermal cells or primordial germ cells enables the manipulation of germline cells in vitro. Production of transgenic chickens has been attempted by the retroviral vector method, microinjection of DNA into a fertilized ovum at the single‐cell stage, use of chimeric intermediates produced by the transfer of stage X blastodermal cells or primordial germ cells, manipulation of spermatozoa, and in vivo manipulation of gonads. So far, the only non‐viral method that has successfully produced transgenic chickens is microinjection of DNA into a fertilized ovum. Manipulation of primordial germ cells could become an efficient system for producing transgenic chickens by combining it with the highly efficient transfection method or the in vitro culture system for primordial germ cells. Preservation of avian genetic resources has now become possible by cryopreservation of stage X blastodermal cells or primordial germ cells as well as spermatozoa. The development of nuclear transfer techniques for avian species is necessary.     

Allele‐specific polymerase chain reaction typing and sequencing of mitochondrial D‐loop region in broiler chickens in Japan

This study aimed to comprehend a feature of single nucleotide polymorphism (SNP) in mitochondrial DNA (mtDNA) mainly of general broiler chickens in Japan. We typed two SNP sites (199C/T and 792A/G) of the D‐loop region in mtDNA by allele‐specific PCR (AS‐PCR) in 359 broiler (182 chunky and 177 cobb) and 506 layer (233 White Leghorn, 140 Barred Plymouth Rock and 133 Rhode Island Red) chickens. The SNP of 199C or 792A by AS‐PCR was observed in the chunky and cobb chickens, and not in the layers. The haplotype 199T/792G was observed in a part of cobb and all layers. By the result of AS‐PCR haplotyping and the broiler brands, the D‐loop region was sequenced in 44 broiler chickens (20 chunky and 24 cobb) and compared with the layers’ sequence data. Among the broiler and layer chickens, 21 SNP sites (including one insertion) and 11 sequence haplotypes were observed. Haplotype variation or correspondence was observed in and between the broiler brands. This study provides important information to establish a chicken meat traceability system by SNP haplotyping of mtDNA in Japan.     

Irradiation history of Itokawa regolith material deduced from noble gases in the Hayabusa samples

Noble gas isotopes were measured in three rocky grains from asteroid Itokawa to elucidate a history of irradiation from cosmic rays and solar wind on its surface. Large amounts of solar helium (He), neon (Ne), and argon (Ar) trapped in various depths in the grains were observed, which can be explained by multiple implantations of solar wind particles into the grains, combined with preferential He loss caused by frictional wear of space-weathered rims on the grains. Short residence time of less than 8 million years was implied for the grains by an estimate on cosmic-ray-produced (21)Ne. Our results suggest that Itokawa is continuously losing its surface materials into space at a rate of tens of centimeters per million years. The lifetime of Itokawa should be much shorter than the age of our solar system.     

Purification and characterization of phytase induced in tomato roots under phosphorus-deficient conditions

Phytase (myo-inositol hexakisphosphate phosphohydrolase; EC 3.1.3.8) was purified from roots of tomato plants grown under phosphorus-deficient conditions using five purification schemes. The phytase was successfully separated from the major acid phosphatase to an electrophoretic homogeneity. The native molecular weight of this enzyme was estimated to be about 164 kD by Bio-Gel P-200 gel filtration. The molecular weight of the subunit on SDS-PAGE was approximately 82 kD, indicating that the native form of the enzyme was a homodimer. The isoelectric point of tomato phytase was about 5.5. The enzyme exhibited a high affinity for phytic acid (K _m = 38 μM), and was strongly inhibited by phosphate, molybdate and fluoride. Among other characteristics of tomato phytase, the pH and temperature optima were 4.3 and 45°C, respectively. Tomato phytase contained a fairly high concentration of aspartic, glutamic acid and glycine residues.     

Isolation and characterization of a cDNA from Catharanthus roseus which is highly homologous with phosphate transporter

The PIT1 gene which is highly homologous with phosphate transporter was isolated from Catharanthus roseus and analyzed. The cBNA PIT1 contained an open reading frame of 542 amino acids and its sequence showed a 31, 30, and 34% identity with the phosphate transporter of Saccharomyces cerevisiae (PBO84), Neurospora crassa (PHO-5), and Glomus versiforme (GvPT), respectively. Furthermore, the cDNA PIT1 encoded a highly hydrophobic protein with 12 putative membrane-spanning regions and contained a conserved amino acid sequence reported in the human glucose transporter super-family* S. cerevisiae strain DpU (pho84 knockout strain) was unable to grow on low phosphate (55 μM) medium (LP medium). Expression of the PIT1 cDNA enabled DpU to grow on LP medium. Northern hybridization analysis revealed that the PIT1 gene was expressed in roots, stems, and young whole plant of C. roseus, but not in leaves.