DNA variation in a conifer,Cryptomeria japonica (Cupressaceae sensu lato)

We investigated the nucleotide variation of a conifer, Cryptomeria japonica, and the divergence between this species and its closest relative, Taxodium distichum, at seven nuclear loci (Acl5, Chi1, Ferr, GapC, HemA, Lcyb, and Pat). Samples of C. japonica were collected from three areas, Kantou-Toukai, Hokuriku, and Iwate. No apparent geographic differentiation was found among these samples. However, the frequency spectrum of the nucleotide polymorphism revealed excesses of intermediate-frequency variants, which suggests that the population was not panmictic and a constant size in the past. The average nucleotide diversity, pi, for silent sites was 0.00383. However, values of pi for silent sites vary among loci. Comparisons of polymorphism to divergence among loci (the HKA test) showed that the polymorphism at the Acl5 locus was significantly lower. We also observed a nearly significant excess of replacement polymorphisms at the Lcyb locus. These results suggested possibilities of natural selection acting at some of the loci. Intragenic recombination was detected only once at the Chi1 locus and was not detected at the other loci. The low level of population recombination rate, 4Nr, seemed to be due to both low level of recombination, r, and small population size, N.     

West–east contrast of phenology and climate in northern Asia revealed using a remotely sensed vegetation index

The phenology of the vegetation covering north Asia (mainly Siberia) and its spatial characterstics were investigated using remotely sensed normalized difference vegetation index (NDVI) data. The analysis used the weekly averaged NDVI over 5 years (1987-1991) using the second-generation weekly global vegetation index dataset (0.144 degrees x 0.144 degrees spatial resolution). In the seasonal NDVI cycle, three phenological events were defined for each pixel: green-up week (NDVI exceeds 0.2), maximum week, and senescence week (NDVI drops below 0.2). Generally there was a west-early/east-late gradient in the three events in north Asia. In the zonal transect between 45 degrees and 50 degrees N, the timing of green-up, maximum, and senescence near 60 degrees E (Kazakh) was about 3.4, 8.7, and 13.4 weeks earlier than near 110 degrees E (Mongolia) respectively. It has been suggested that vegetation near Kazakh only flourishes during a short period when water from snow melt is available from late spring to early summer. In Mongolia, abundant water is available for the vegetation, even in midsummer, because of precipitation. In the 50-60 degrees N zonal transect, the green-up and maximum near 40 degrees E were about 3.8 and 3.9 weeks earlier than near 115 degrees E, respectively. As for the week of senescence, there was no clear west-east trend. This west-to-east phenological gradient was related to the weekly cumulative temperature (over 0 degrees C). Weeks in which the cumalative temperature exceeded 40 degrees C and 140 degrees C had a similar west-east distribution to green-up and maximum NDVI.     

Lactoferrin stimulates A Staphylococcus aureus killing activity of bovine phagocytes in the mammary gland

Lactoferrin (Lf) may play a key role in the clearance of microorganisms from a host. To study in vitro the bactericidal mechanisms of Lf during nonlactating periods, we investigated whether the effects of Lf were influenced by bovine mammary gland secretory cells (MGSC) and fresh normal bovine serum (NBS) as a source of complement. Phagocytic killing tests demonstrated that a phagocytic mixture of unopsonized Staphylococcus aureus (S. aureus) and MGSC in the presence of Lf reduced bacterial growth, compared with that of unopsonized S. aureus and MGSC without Lf. The opsonization with Lf and fresh NBS together resulted in more than a 95% reduction in CFU. The activation of complement induced by Lf also resulted in increased deposition of C3 on S. aureus, and the phagocytic activity of MGSC was augmented by opsonization with Lf and fresh NBS. Inhibition of C3 deposition by Lf was not induced in the presence of Mg-EGTA, but was induced by the addition of bovine Lf antiserum. These results strongly suggest that Lf induces the activation of complement in fresh NBS mainly through an alternative pathway. The results demonstrated a Lf-dependent, antibody-independent and complement-mediated phagocytic killing of S. aureus, and implied that Lf was synergistically capable of activating both the alternative pathway of the bovine complement cascade and phagocytosis by phagocytes.     

Antibacterial effect of Kampo herbal formulation Hochu-ekki-to (Bu-Zhong-Yi-Qi-Tang) on Helicobacter pylori infection in mice

Because Helicobacter pylori (H. pylori) infection is a major cause of gastroduodenal diseases in humans, the eradication of H. pylori using antibiotics is very effective for the treatment of gastroduodenal diseases. However, it has recently been reported that resistance to these antibiotics is developing. In the present study, the antibacterial effect of a Kampo (traditional Japanese medicine) herbal formulation, Hochu-ekki-to (RET; Formula repletionis animalis et supletionis medii), against H. pylori was examined in vitro and in vivo. HET inhibited the growth of antibiotic-resistant strains of H. pylori as well as antibiotic-sensitive strains at a dose of 2.5 mg/ml in vitro. When 1,000 mg/kg of HET was administered orally to C57BL/6 mice for 7 days before or after inoculation with H. pylori, H. pylori in the stomach was significantly reduced in the HET-pre-treatment group compared with the control group. Furthermore, HET in combination with antibiotics completely eradicated the bacteria in mice. The expression of interferon (IFN)-gamma was induced in the gastric mucosa of the mice pre-treated with HET. There were no significant differences between the colonization of H. pylori in the control and HET treatment groups in IFN-gamma gene-deficient mice. These results suggest that the antibacterial effect of HET may be partly due to IFN-gamma induction, and that HET may be clinically useful for treatment of H. pylori infection.     

Effects of systemic and central nervous system localized inflammation on the contributions of metabolic precursors to the L-kynurenine and quinolinic acid pools in brain

L-Kynurenine and quinolinic acid are neuroactive L-tryptophan-kynurenine pathway metabolites of potential importance in pathogenesis and treatment of neurologic disease. To identify precursors of these metabolites in brain, [(2)H(3) ]-L-kynurenine was infused subcutaneously by osmotic pump into three groups of gerbils: controls, CNS-localized immune-activated, and systemically immune-activated. The specific activity of L-kynurenine and quinolinate in blood, brain and systemic tissues at equilibrium was then quantified by mass spectrometry and the results applied to a model of metabolism to differentiate the relative contributions of various metabolic precursors. In control gerbils, 22% of L-kynurenine in brain was derived via local synthesis from L-tryptophan/formylkynurenine versus 78% from L-kynurenine from blood. Quinolinate in brain was derived from several sources, including: local tissue L-tryptophan/formylkynurenine (10%), blood L-kynurenine (35%), blood 3-hydroxykynurenine/3-hydroxyanthranilate (7%), and blood quinolinate (48%). After systemic immune-activation, however, L-kynurenine in brain was derived exclusively from blood, whereas quinolinate in brain was derived from three sources: blood L-kynurenine (52%), blood 3-hydroxykynurenine or 3-hydroxyanthranilate (8%), and blood quinolinate (40%). During CNS-localized immune activation, > 98% of both L-kynurenine and quinolinate were derived via local synthesis in brain. Thus, immune activation and its site determine the sources from which L-kynurenine and quinolinate are synthesized in brain. Successful therapeutic modulation of their concentrations must take into account the metabolic and compartment sources.     

Characterization of a lysK gene as an argE homolog in Thermus thermophilus HB27

We conducted a chromosome walk to obtain a DNA fragment downstream of lysJ and found an argE homolog in a putative operon composed of lysJ-orfC-orfD-argE homologs. A knockout mutant of the argE homolog showed significantly slow growth on a minimal medium, and the growth was markedly improved by addition of lysine. We therefore termed this gene lysK. Purified LysK protein has deacetylating activities for both N(2)-acetyllysine and N(2)-acetylornithine at almost equal efficiency. These results suggest that lysK which may share an ancestor with argE functions not only for the lysine biosynthesis, but also for arginine biosynthesis in Thermus thermophilus.     

New convulsive compounds,brasiliamides A and B,from Penicillium brasilianum batista JV-379

New convulsive compounds, brasiliamides A (1) and B (2), were isolated by activity-guided fractionation from okara fermented with a soil isolate of Penicillium brasilianum Batista JV-379. Their structures were elucidated on the basis of spectral and chemical evidence and by X-ray crystallography of the hydrogenated product of 2. In the 1H- and 13C-NMR spectra of 2, the signals were complicated, all being doubled or broadened in several deuterated solvents at room temperature. The conformational change of 2 was clarified as the rotational isomerization of amide bonds in solution by NMR measurements at various temperatures. Four rotamers of 2 at two amide bonds were presented at -60 degrees C in CDCl3, whereas only two isomers were apparent at room temperature, owing to rapid rotation of one of the amide bonds. Brasiliamides A and B respectively showed convulsive activity against silkworms with ED50 values of 300 and 50 microg/g of diet.     

Two distinct high-affinity sulfate transporters with different inducibilities mediate uptake of sulfate in Arabidopsis roots

Sulfate transporters present at the root surface facilitate uptake of sulfate from the environment. Here we report that uptake of sulfate at the outermost cell layers of Arabidopsis root is associated with the functions of highly and low-inducible sulfate transporters, Sultr1;1 and Sultr1;2, respectively. We have previously reported that Sultr1;1 is a high-affinity sulfate transporter expressed in root hairs, epidermal and cortical cells of Arabidopsis roots, and its expression is strongly upregulated in plants deprived of external sulfate. A novel sulfate transporter gene, Sultr1;2, identified on the BAC clone F28K19 of Arabidopsis, encoded a polypeptide of 653 amino acids that is 72.6% identical to Sultr1;1 and was able to restore sulfate uptake capacity of a yeast mutant lacking sulfate transporter genes (K(m) for sulfate = 6.9 +/- 1.0 microm). Transgenic Arabidopsis plants expressing the fusion gene construct of the Sultr1;2 promoter and green fluorescent protein (GFP) showed specific localization of GFP in the root hairs, epidermal and cortical cells of roots, and in the guard cells of leaves, suggesting that Sultr1;2 may co-localize with Sultr1;1 in the same cell layers at the root surface. Sultr1;1 mRNA was abundantly expressed under low-sulfur conditions (50-100 microm sulfate), whereas Sultr1;2 mRNA accumulated constitutively at high levels under a wide range of sulfur conditions (50-1500 microm sulfate), indicating that Sultr1;2 is less responsive to changes in sulfur conditions. Addition of selenate to the medium increased the level of Sultr1;1 mRNA in parallel with a decrease in the internal sulfate pool in roots. The level of Sultr1;2 mRNA was not influenced under these conditions. Antisense plants of Sultr1;1 showed reduced accumulation of sulfate in roots, particularly in plants treated with selenate, suggesting that the inducible transporter Sultr1;1 contributes to the uptake of sulfate under stressed conditions.