The role of β-alanine in the biosynthesis of nitrate byAspergillus flavus

d-Serine (0.05m) inhibited nitrification byAspergillus flavus in media containing either peptone, aspartate,a-alanine or -alanine as the sole nitrogen source. A similar inhibition was observed in an aspartate + peptone medium, but nitrate was formed in a -alanine + peptone medium in the presence of the inhibitor. Exceptionally high yields of nitrate were obtained in the -alanine + peptone medium. In replacement cultures,d-serine inhibited nitrification of aspartate but not of -alanine. Manometric studies indicated that aspartate was decarboxylated byA. flavus and that the reaction was inhibited byd-serine. In view of these results, it is suggested that aspartate is a precursor and -alanine is an intermediate in nitrification by this fungus.     

The bactericidal effects of Lactobacillus acidophilus,garcinol and Protykin® compared to clarithromycin,on Helicobacter pylori

Chronic infection with Helicobacter pylori causes peptic ulcers, gastric cancer and lymphoma. We evaluated the inhibitory effects of the probiotic Lactobacillus acidophilus DDS-1J, the antibiotic clarithromycin and the natural antioxidants garcinol and Protykin^® (containing 50% trans-resveratrol) on Helicobacter pylori strain ATCC 49503. The findings of this study indicate that Lactobacillus acidophilus DDS-1J exerts a growth inhibitory effect on H. pylori at a ratio of 1:1 or higher in vitro. In the case of clarithromycin, garcinol and resveratrol, the bactericidal effect is time and concentration dependent. Clarithromycin completely inhibited growth at 62.5 g/ml at 6 h and at 31.5 g/ml at 12 h. For garcinol the highest concentration needed for complete inhibition was 31.5 g/ml at 6 h and 3.9 g/ml after 12 h incubation. For resveratrol, significant inhibition was noted at 1000 g/ml at 12 h only. The bactericidal effect of garcinol was reduced by the addition of resveratrol at all concentrations 125 g/ml at 6 and 12 h. We conclude from this study that Lactobacillus acidophilus DDS-1J inhibits H. pylori at 1:1 and higher ratios. Also, between the two antioxidants, garcinol is much more potent than resveratrol as a bactericidal agent against H. pylori, and that resveratrol may antagonize this effect. Finally, our study showed equivalent or better bactericidal activity of garcinol compared to clarithromycin against H. pylori at 6 and 12 h incubation, indicating a potential role for this antioxidant in treatment for H. pylori infection.     

Structure of root nodules formed by Rhizobium on the non-legume Trema cannabina var. scabra

The structure of the nodules formed by Rhizobium on the non-legume Trema cannabina var. scabra was studied using the light microscope. The overall features of the nodules showed greater resemblance to the non-legume rather than the legume nodule. Nodule squashes yielded bundles of infection threads and bacteroids with morphological differences from rhizobial cells grown on yeast-mannitol-glucose agar. Two types of cell infection occurred within the bacterial zone; plant cells were either, like legumes, filled with rhizobia released from the infection threads (less than one third of infected cells) or were filled with the extensive growth and development of the infection thread. The rate of nitrogen fixation in the Trema nodule was high. It seemed that host cells filled with threads were active in N fixation.     

Receptors for phorbol esters are primarily localized in neurons: Comparison of neuronal and glial cultures

Binding of [³H]PDB has been measured in the present study to determine the levels of protein kinase C in the neuronal and astrocytic glial cells in culture from rat brain. Binding of [³H]PDB to homogenates of cultured neuronal cells from the brains of normotensive and hypertensive rats was time-dependent and specific. The relative potency for competition by various phorbol esters to [³H]PDB binding was TPA > -PDD > POE > -PDD 4phorbol. Scatchard analysis showed that neuronal cultures from normotensive rat brains contained 2–3 fold more phorbol ester receptors compared with the glial cultures from the same brains. No differences in theK _d andB _max were observed between neuronal cultures from normotensive and spontaneously hypertensive rat brains. These studies suggest that the phorbol ester receptors are primarily localized in neuronal cells.     

Structure and floristic composition of the lowland rain forest of Los Tuxtlas,Mexico

Physiognomy, structure and floristic composition of one hectare of lowland tropical rain forest was studied in detail at Los Tuxtlas, Mexico. Physiognomically, the Los Tuxtlas forest should be classified as lowland tropical high evergreen rain forest. The forest showed a closed canopy at 30–35 m. Of all woody, non-climbing species with a DBH1.0 cm 89.4% (94.5% of all individuals) were evergreen, 25.4% (59.5% of the individuals) had compound leaves, and over 80% of species (and individuals) had leaves in the notophyll and mesophyll size classes. The forest structure was characterized by a low density (2976 individuals with a DBH1.0 cm, 346 individuals with a DBH10.0 cm, per ha, excluding vines) with an average basal area (38.1 m², DBH1.0 cm, 34.9 m², DBH10.0 cm, per ha, excluding vines). This was attributed to the relative maturity of the forest on the study plot. The study plot contained 234 species (11 208 individuals with a height 0.5 m), of which 55.1% (34.8% of individuals) were trees, 9.4% (6.8%) shrubs, 3.4% (44.3%) palms, 20.1% (5.2%) vines, 6.8% (8.7%) herbs and 5.1% (0.3%) of unknown lifeform. Furthermore, 58 species of epiphytes and hemi-epiphytes were found. Diversity of trees, shrubs and palms with a DBH1.0 cm was calculated as Shannon-Wiener index (4.65), Equitability index (0.65), and Simpson index (0.10). The dominance-diversity curve showed a lognormal form, characteristic for tropical rain forest. The community structure was characterized by a relative dominance of Astrocaryum mexicanum in the understorey, Pseudolmedia oxyphyllaria in the middle storeys, and Nectandra ambigens in the canopy. Species population structures of 31 species showed three characteristic patterns, differentiated by recruitment: continuously high, discontinuously high, and continuously low recruitment. Height/diameter and crown cover/diameter diagrams suggested a very gradual shift from height growth to crown growth during tree development. Forest turnover was calculated as 138 years. Compared to other tropical rain forests the Los Tuxtlas forest had 1. similar leaf physiognomical characteristics, 2. a lower diversity, 3. a lower density, 4. an average basal area, and 5. a slow canopy turnover.     

Comparative assessment of fermentation techniques useful in the processing of ogi

Ogiis processed traditionally by the use of uncontrolled spontaneous fermentation of maize, sorghum and millet. In this study, traditionally applied spontaneous fermentation was compared with accelerated batch fermentation (or back-slopping) and the use of starter cultures to initiate fermentation. Lactic acid bacteria populations comprised 95 of the total viable bacteria and remained prominent throughout the fermentations, while number of moulds and coliform bacteria declined as the fermentation progressed. The fermentation method involving the application of starter culture helps most to control the prevalence of coliforms and moulds. Lactic acid bacteria, such as Lactococcus raffinolactis, Pediococcussp.,Pediococcus pentosaceus, Lactobacillus plantarum, Lb. suebicus and Lb. brevis,were isolated at different processing stages of ogi using accelerated batch fermentation (back-slopping) technique. Highest increase in acidity was observed immediately after wet-milling and sieving fermenting maize grains at 28 and 48 h. Sharp increases in the reducing sugar levels were noted between 24 and 28 h of fermentations during wet-milling and sieving processes.     

The Hawaiian Archipelago: A Microbial Diversity Hotspot

The Hawaiian Archipelago is a biodiversity hotspot where significant endemism among eukaryotes has evolved through geographic isolation and local topography. To address the absence of corresponding region-wide data on Hawaiis microbiota, we compiled the first 16S SSU rDNA clone libraries and cultivated bacteria from five Hawaiian lakes, an anchialine pool, and the Lihi submarine volcano. These sites offer diverse niches over ~5000 m elevation and ~1150 nautical miles. Each site hosted a distinct prokaryotic community dominated by Bacteria. Cloned sequences fell into 158 groups from 18 Bacteria phyla, while seven were unassigned and two belonged in the Euryarchaeota. Only seven operational taxonomic units (each OTU comprised sequences that shared 97% sequence identity) occurred in more than one site. Pure bacterial cultures from all sites fell into 155 groups (each group comprised pure cultures that shared 97% 16S SSU rDNA sequence identity) from 10 Bacteria phyla; 15 Proteobacteria and Firmicutes were cultivated from more than one site. One hundred OTUs (60%) and 52 (33.3%) cultures shared <97% 16S SSU rDNA sequence identity with published sequences. Community structure reflected habitat chemistry; most -Proteobacteria occurred in anoxic and sulfidic waters of one lake, while -Proteobacteria were cultivated exclusively from fresh or brackish waters. Novel sequences that affiliate with an Antarctic-specific clade of Deinococci, and Candidate Divisions TM7 and BRC1, extend the geographic ranges of these phyla. Globally and locally remote, as well as physically and chemically diverse, Hawaiian aquatic habitats provide unique niches for the evolution of novel communities and microorganisms.     

Comparative Analysis of MGE 412 Patterns in 18 Isogenic Lines of Drosophila melanogaster

Comparative analysis of patterns of mobile genetic element 412 was conducted in 18 isogenic lines of Drosophila melanogaster isolated in three isogenic experiments in 1987 through 1999. Twelve extra-hot isogenization sites (in 15–18 lines) and 23 hot isogenization sites (10 lines) were found; of these, 19 occurred in the original heterogeneous line. These sites virtually do not overlap with hot induction sites of transposition. Sites of the latter group generally retain their positions during isogenization. It was shown that no more than 20% of the new sites were brought from the balancer by double recombination, while inbreeding and outbreeding caused 80% of them. Different factors were shown to have different hot isogenization sites. A similarity tree was constructed for the patterns of 18 isogenic lines. The maximum peak of the tree was very low (< 0.25), i.e., the isogenic lines are more similar to than different from one another. The tree was subdivided into subtrees. The division was in good agreement with the isogenization groups corresponding to individual isogenization experiments. Significant correlation was found between the total fragment length and the number of new sites per lines.     

Characterization of the effectors required for stable inheritance of Streptococcus pyogenes pSM19035-derived plasmids in Bacillus subtilis

The low-copy-number and broad-host-range pSM19035-derived plasmid pBT233 is stably inherited in Bacillus subtilis cells. Two distinct regions, segA and segB, enhance the segregational stability of the plasmid. Both regions function in a replicon-independent manner. The maximization of random plasmid segregation is accomplished by the recombination proficiency of the host or the presence of the pBT233 segA region. The segA region contains two open reading frames (or) [ and ]. Inactivation or deletion of or results in SegA^– plasmids. Better than random segregation requires an active segB region. The segB region contains two ors (or and or). Inactivation of either of the orfs does not lead to an increase in cell death, but or^– plasmids are randomly segregated. These results suggest that pBT233 stabilization relies on a complex system involving resolution of plasmid oligomers (segA) and on the function(s) encoded by the segB region.     

Biotransformation of 5βH-pregnan-3βol-20-one and cardenolides in cell suspension cultures of Nerium oleander L.

Summary In order to demonstrate enzyme activities playing a role in the biosynthesis of cardenolides and 2,6-dideoxysugars, 5H-pregnan-3ol-20-one and cardenolides (digitoxigenin, oleandrigenin/L-oleandrose, oleandrin, neriifolin, digitoxigeninmonodigitoxoside and strospeside) were fed to cell suspension cultures of Nerium oleander L.. It could be shown that cell suspension cultures of Nerium oleander L. are able to oxidize, isomerize and glucosylate 5H-steroidaglycones at C-3. The respective glucosides of the 5H-steroid-aglycones are the main biotransformation products. These cell cultures are an appropriate tool for the production of labelled 5H-steroidglucosides.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - IAA indole-3-acetic acid - EtOAc ethylacetate - MeOH methanol - MS Murashige & Skoog     

Cryptic growth in Klebsiella pneumoniae

Summary The ability of Klebsiella pneumoniae to grow on its own soluble lysis products is shown in a series of batch growth experiments. Maximum specific growth rate coefficients ranging from 0.69 to 1.46 h^-1 were obtained with experimental cryptic yield coefficients ranging between 0.42 to 0.52 (mg-cell-C/mg-substrate-C). These kinetic data are used to calibrate a model which demonstrates that depression of theoretical maximum yield coefficients relative to experimentally obtained values can be explained by cryptic growth phenomena without the need to resort to the use of physiologically undefined, mathematical constants. Growth of K. pneumoniae on sonicated cells derived from steady-state chemostat cultures was followed in batch culture and observed to occur with no lag phase. Batch growth curves did not indicate either diauxic or polyauxic growth, suggesting simultaneous utilization of the complex organic substrate mixture. These data suggest that cryptic growth is probably a real event occurring in growing chemostat cultures under ideal growth conditions and most probably also under starvation conditions.     

Micropropagation of flowering dogwood (Cornus florida) from seedlings

A method for regenerating whole plants from nodal (axillary bud) cultures of seedlings was developed for flowering dogwood (Cornus florida L.). The seed source significantly influenced the rate of proliferation, although cultures initiated from each of the seven mother trees produced some shoots. Woody plant medium (WPM) was superior to either Murashige and Skoog or Schenk and Hildebrandt basal medium. 6-Benzyladenine (BA) at 2.2 or 4.4 m stimulated the generation of significantly more useable shoots (1 cm) compared to all other concentrations (0.5–22.5 m tested. Thidiazuron (TDZ) at 0.6 and 1.1 m supported proliferation, but strongly inhibited shoot elongation. TDZ initiated cultures transferred to medium containing 4.4 m BA produced usable shoots after five additional subcultures. Shoots generated adventitious roots when exposed to either a 12-h pulse of relatively high concentrations (246–1230 m or continuous lower concentrations (0.5–49.0 m of indolebutyric acid (IBA) for longer periods. Microshoots produced the significantly greatest number of roots when subjected to 4.9 m IBA in WPM over a 4-week period. Whole plants were acclimatized to the laboratory conditions and subsequently to the greenhouse. The methodology described here should be useful in a breeding program by supplying multiple copies of unique, recombinant genotypes.     

Recombinant β-glucanase production and plasmid stability of Bacillus subtilis in cyclic fed batch culture

Recombinant -glucanase production and plasmid stability were higher in cyclic fed batch culture than in batch and chemostat culture. Plasmid stability was maintained indefinitely in cyclic fed batch cultures of Bacillus subtilis when subjected to cycle periods of 2 or 4 hours. Cycle periods greater than 6 hours resulted in loss of the plasmid. In batch and chemostat cultures plasmid loss was rapid. -1,3-glucanase productivity was highest in cyclic fed batch culture.     

Influence of phenolic acids on phosphorus mobilisation in acidic and calcareous soils

Phenolic acids and phenols are abundant in soils. However, little information is available on the role of these compounds in mobilisation of soil phosphorus (P). The present study examined the effects of three phenolics on P mobilisation in comparison with citric acid in three soils differing in chemical properties. The soils were incubated with organic compounds at concentrations of 0 to 100 mol g^–1 soil for 30 min. While the addition of phenolic acid anions and phenol decreased soil pH, citrate either increased or did not affect soil pH depending on the soil type. All the organic compounds increased the amounts of easily-mobilised P fractions in the order of catechol citric acid proto-catechuic acid caffeic acid for the acid soils and citric acid > catechol = proto-catechuic acid > caffeic acid for the calcareous soil. Phosphorus mobilisation did not correspond to the amounts of Ca, Fe or Al ions released from the soils. These results suggest that organic acids and phenols altered the P fractions from stable, sparingly-soluble forms to easily dissolvable forms, and that chelation or precipitation of cations with organic ligands occur before biodegradation of the ligands.     

Extraction and quantitative analysis of the blue-green pigment “marennine” synthesized by the diatom Haslea ostrearia

This study reports further information on mareninne, awater-soluble blue-green pigment synthesized by the diatom Hasleaostrearia, which is essential to the greening of maturing oysters inFrench production areas. The extraction process is reported, as well aspreliminary characterization of a partially purified marennine extract,including quantitative spectrophotometric analysis of intracellular pigment. Amean specific extinction coefficient, E ^1% _1cm = 17.2 at 669 nm, is proposed. Results forquantitative determination of marennine accumulated in cells during batchcultures are presented, and their importance for pigment production isconsidered in relation to potential industrial applications.     

MHC restriction of male-antigen-specific T helper cells collaborating in antibody responses

By priming female C57BL/6 mice with syngeneic male spleen cells and enriching inguinal and paraaortic lymph node cells in long-term culture (LTC) by repeated restimulations, H-Y-specific T helper cells can be produced. In response to male spleen cells carrying I-Ab antigens these cells activate antigenexpressing B cells to secrete polyclonal antibody. Before the end of the second week in LTC it was impossible to detect any helper activity. Induction of plaque-forming cells (PFC) also requires simultaneous recognition of antigen and I-A-encoded determinants in the stimulator-responder spleen-cell population. The testing of spleen cells fromH-2 recombinant strains as stimulator-responders to anti-H-Y helper T cells of C57BL/6 origin also revealed that other genes, telomeric toI-A, control the magnitude of both specific T-cell proliferation and helper-dependent B-cell activation.     

Further cytogenetic analyses of the Croatian triploid shallot “Ljutika” (Allium cepa var.viviparum, Alliaceae) and its comparison with the Indian triploid “Pran”

Feulgen and silver-stained karyotypes and meiosis of two triploid viviparous onion forms (Allium cepa var.viviparum), the Croatian Ljutika and the Indian Pran, were comparatively analyzed. The results of chromosome measurements show that Ljutika and Pran are karyologically not identical, although significant similarities were found in the morphology of their chromosomes. Five geographically distant clones of Ljutika showed good agreement in the number and gross morphology of the chromosomes and in the number and position of NORs and interphase nucleoli. Heterotrivalents were predominant in meiosis of Ljutika but a relatively high frequency of higher multivalents together with univalents and bivalents were also observed. The relationship between Ljutika and Pran and their possible origin are discussed.     

The function of mitochondria-rich cells (chloride cells) in teleost gills

Summary The CCs are the site of Cl^– transport in teleosts in sea water. The gills of freshwater teleosts contain at least two types of mitochondria-rich cell, the type and the type (Pisam and Rambourg, 1991). During seawater acclimation, the cells vanish and the cells are transformed and proliferate, and accessory cells appear in addition. This gives rise to the question of the function of cells in fish living in fresh water.According to the studies reviewed here, although they deal only with extrabranchial epithelia, the majority of evidence indicates that CCs (or MRCs) function as sites of active Ca²⁺ transport in freshwater teleosts. Moreover, some experimental results suggest that CCs are the Cl^– uptake site in freshwater teleosts. The main problem in characterizing the CC function is that they have not yet been adequately described from the biochemical standpoint. This applies particularly to their metabolic pattern and the composition of their apical and basolateral membranes, including their integrated proteins and cell-cell junctions.Experiments with organ tissue cultures such as gill organ cultures from Oncorhynchus mykiss (McCormick and Bern, 1989) and opercular membrane cultures from Oreochromis mossambicus (McCormick, 1990) will almost certainly yield important results. Primary cell cultures of CCs would be even better for characterizing CCs. Such a cell culture of rainbow trout respiratory cells has already been established (Pärt et al., 1993).     

Fromβ-lactams toα- andβ-amino acid derived peptides

Summary The potential of-lactams as intermediates for the access to- and-amino acid-derived peptides is shortly reviewed, with major focus on the technologies developed in our group. The two general strategies lie, on one side, in the oxidative ring expansion of 3-hydroxy-lactams toN-carboxy-amino acid anhydrides or Leuch's anhydrides and subsequent coupling with-amino acid esters and, on the other side, in the nucleophilic ring opening ofN-Boc--lactams. Both approaches have been successfully applied to the synthesis of,-diamino acid,-amino--hydroxy acid, polyhydroxylated-amino acid,,-disubstituted-amino acid,-amino acid,-amino--hydroxy acid and,-disubstituted-amino acid derived peptides. Because of the mild reaction conditions needed for the above transformations and the highly stereoselective procedures employed for the construction of the starting-lactam ring, the whole process allows the production of optically pure final products.     

Light and electron microscopic immunocytochemistry on the localization of 3β-hydroxysteroid dehydrogenase/isomerase in the bovine adrenal cortical cells

Summary The localization of 3-hydroxysteroid dehydrogenase/isomerase (3-HSD) was studied in bovine adrenal glands by light as well as electron microscopic immunocytochemistry, using anti-bovine adrenal 3-HSD antibody. With light microscopy the cytoplasm of the glomerulosa cells was weakly immunostained, while that of the fasciculata-reticularis cells was intensely immunostained though both the capsular connective tissue cells and the medullary cells were entirely negative for this reaction. Electron microscopic immunocytochemistry revealed that the positive reaction products for 3-HSD were present on the membrane of smooth endoplasmic reticulum of the cortical cells, especially that of the fasciculata and reticularis cells. Other cell organelles such as mitochondria and Golgi apparatus were entirely negative. The present results indicate that 3-HSD is present in the membrane of smooth endoplasmic reticulum of bovine adrenal cortical cells.Supported by grants from the Ministry of Education Science and Culture, Japan