Reduction of Cr(VI) by a Bacillus sp.

A Bacillus sp. RE was resistant to chromium and reduced Cr(VI) without accumulating chromium inside the cell. When Cr(VI) was 10 and 40 μg ml⁻¹, >95% of the total Cr(VI) was reduced in 24 and 72 h of growth, respectively, whereas at 80 μg Cr(VI) ml⁻¹ only 50% of Cr(VI) was reduced. However growth was not affected; the cell mass was 0.7–0.8 mg ml⁻¹ in all cases. The cell-free extract showed Cr(VI) reducing enzyme activity which was enhanced (>5 fold) by NADH and NADPH. Like whole cells the enzyme also reduced Cr(VI) with decreasing efficiency on increasing Cr(VI) concentration. The enzyme activity was optimal at pH 6.0 and 30 °C. The enzyme was stable up to 30 °C and from pH 5.5 to 8, but from pH 4 to 5 the enzyme was severely destabilized. Its K_m and V_max were 14 μm and 3.8 nmol min⁻¹ mg⁻¹ respectively. The enzyme activity was enhanced by Cu²⁺ and Ni²⁺ and inhibited by Hg²⁺. Received 21 September 2005; Revisions requested 5 October 2005; Revisions received 16 November 2005; Accepted 16 November 2005     

Chromium Reduction,Plant Growth–Promoting Potentials,and Metal Solubilizatrion by <Emphasis Type="Italic">Bacillus sp</Emphasis>. Isolated from Alluvial Soil

The plant growth–promoting potentials, production of siderophore and solubilization of insoluble phosphorus (P) and zinc and lead by the chromium (vi) -reducing Bacillus species, PSB 1, PSB 7, and PSB 10, was assessed both in the presence and absence of chromium under in vitro conditions. The Bacillus strains tolerated chromium up to the concentration of 500 (PSB1), 400 (PSB7), and 550 μg ml⁻¹ (PSB10), respectively, on nutrient agar plates. Bacillus sp. PSB 10 reduced Cr (vi) by 87% at pH 7, which was followed by Bacillus sp. PSB 1 (83%) and PSB 7 (74%) in nutrient broth after 120 h of incubation. A concentration of 50 μg ml⁻¹ of Cr (vi) was completely reduced by Bacillus sp. PSB 1 and PSB 10 (after 100 h) and PSB 7 (after 120 h). The Bacillus strains PSB 1, PSB 7, and PSB 10 produced 19.3, 17.7, and 17.4 μg ml⁻¹ of indole acetic acid, respectively, in luria bertani broth at 100 μg ml⁻¹ of tryptophan, which consistently decreased with an increase in chromium concentration. The Bacillus strains were positive for siderophore, HCN, and ammonia both in the absence and presence of chromium. The Bacillus strains solubilized 375 (PSB 1), 340 (PSB 7), and 379 (PSB 10) μg ml⁻¹ P, respectively, in Pikovskaya broth devoid of chromium. In contrast, chromium at 150 μg ml⁻¹ reduced the amount of P solubilized by 17 (PSB 1), 15 (PSB 7), and 9% (PSB 10) compared to control. The tested bacterial strains solubilized a considerable amount of zinc and lead in nutrient broth both in the absence and presence of chromium. Generally, the chromium reduction and the plant growth–promoting potentials of chromium-reducing Bacillus were strongly correlated at the tested concentration of chromium. The present observations demonstrated that the chromium-reducing, metal-solubilizing, and plant growth–promoting potentials of the Bacillus strains PSB1, PSB 7, and PSB10 were not adversely affected by the chromium application and, hence, may be applied for raising the productivity of crops under metal-contaminated soils.     

Induction of axenic culture of Arthrospira (Spirulina) platensis based on antibiotic sensitivity of contaminating bacteria

Arthrospira platensis SAG 21.99 and the isolated bacteria (Halomonas spp., Staphylococcus sp., etc.) from the culture of A. platensis SAG 21.99 were treated with five antibiotics to determine the minimal lethal concentrations. The combination of a washing step and a consecutive treatment with antibiotics, imipenem (100 μg ml⁻¹), neomycin (100 μg ml⁻¹) and cycloheximide (20 μg ml⁻¹), treatment step was highly effective in eliminating bacteria. An axenic culture of A. platensis SAG 21.99 could be induced within 3 days using this method. This technique is a simple and rapid method for obtaining axenic cultures of filamentous cyanobacteria.     

Studies on inositol-mediated expression of MAL gene encoding maltase and phospholipid biosynthesis in Schizosaccharomyces pombe

In this study, the effects of inositol addition on expression of the MAL gene encoding maltase and phosphatidylinositol (PI) biosynthesis in Schizosaccharomyces pombe (a naturally inositol-requiring strain) were examined. We found that specific maltase activity was at its maximum when the concentration of added inositol reached 6 μg ml⁻¹ in a synthetic medium containing 2.0% (w/v) glucose. When the concentration of added inositol was 1 μg ml⁻¹ in the medium, repression of MAL gene expression occurred at glucose concentration higher than 0.2% (w/v). However, when S. pombe was cultured in the synthetic medium containing 6 μg ml⁻¹, repression of maltase gene expression occurred only at initial glucose concentration above 1.0% (w/v). More mRNA encoding maltase was detected in the cells grown in the medium with 6 μg ml⁻¹ inositol than in those grown in the same medium with 1 μg ml⁻¹ inositol. These results demonstrate that higher inositol concentrations in the synthetic medium could derepress MAL gene expression in S. pombe. PI content of the yeast cells grown in the synthetic medium with 6 μg ml⁻¹ of inositol was higher than that of the yeast cells grown in the same medium with 1 μg ml⁻¹ of inositol. This means that PI may be involved in the derepression of MAL gene expression in S. pombe.     

Bioremoval of hexavalent chromium from water by a salt tolerant bacterium, Exiguobacterium sp. GS1

Pollution of terrestrial surfaces and aquatic systems by hexavalent chromium, Cr(VI), is a worldwide public health problem. A chromium resistant bacterial isolate identified as Exiguobacterium sp. GS1 by 16S rRNA gene sequencing displayed high rate of removal of Cr(VI) from water. Exiguobacterium sp. GS1 is 99% identical to Exiguobacterium acetylicum. The isolate significantly removed Cr(VI) at both high and low concentrations (1–200 μg mL⁻¹) within 12 h. The Michaelis–Menten K _m and V _max for Cr(VI) bioremoval were calculated to be 141.92 μg mL⁻¹ and 13.22 μg mL⁻¹ h⁻¹, respectively. Growth of Exiguobacterium sp. GS1 was indifferent at 1–75 μg mL⁻¹ Cr(VI) in 12 h. At initial concentration of 8,000 μg L⁻¹, Exiguobacterium sp. GS1 displayed rapid bioremoval of Cr(VI) with over 50% bioremoval in 3 h and 91% bioremoval in 8 h. Kinetic analysis of Cr(VI) bioremoval rate revealed zero-order in 8 h. Exiguobacterium sp. GS1 grew and significantly reduced Cr(VI) in cultures containing 1–9% salt indicating high salt tolerance. Similarly the isolate substantially reduced Cr(VI) over a wide range of temperature (18–45  °C) and initial pH (6.0–9.0). The T _opt and initial pH_opt were 35–40  °C and 7–8, respectively. Exiguobacterium sp. GS1 displayed a great potential for bioremediation of Cr(VI) in diverse complex environments.     

High regenerative nature ofMentha arvensis internodes

Media and incubation conditions have been defined for highly efficient regeneration of shoots from internode explants of slow and fast growing cultivars ofMentha arvensis. Internodal segments excised from thein vitro raised shoots were inoculated on the MS medium supplemented with combinations of 5 concentrations of l-napthalene acetic acid (NAA) and 3 concentrations of 6-benzyl amino purine (BAP). The media containing 2 μg ml⁻¹ NAA, 10 Μg ml⁻¹ BAP and 1 μg ml⁻¹ NAA, 5 μg ml⁻¹ BAP proved best for shoot regeneration and growth responses on cv Himalaya and cv Kalka explants, respectively. In 12 weeks time, on average one explant of cv Himalaya produced about 200 shoots and that of cv Kalka produced about 180 shoots. The Himalaya explants required higher concentrations of NAA and BAP for high efficiency proliferation as compared to the Kalka explants. The experiments demonstrated that internodal tissue inMentha arvensis can be induced to obtain direct shoot regenerants with high efficiency. The analysis of the RAPD profiles of 100 regenerated plantlets each of cv Himalaya and Kalka showed more than 99.9% homogeneity in bands with respect to the parents.     

Antimicrobial activity of an endophytic <Emphasis Type="Italic">Xylaria</Emphasis> sp.YX-28 and identification of its antimicrobial compound 7-amino-4-methylcoumarin

An endophytic Xylaria sp., having broad antimicrobial activity, was isolated and characterized from Ginkgo biloba L. From the culture extracts of this fungus, a bioactive compound P3 was isolated by bioactivity-guided fractionation and identified as 7-amino-4-methylcoumarin by nuclear magnetic resonance, infrared, and mass spectrometry spectral data. The compound showed strong antibacterial and antifungal activities in vitro against Staphylococcus aureus [minimal inhibitory concentrations (MIC) 16 μg·ml⁻¹], Escherichia coli (MIC, 10 μg·ml⁻¹), Salmonella typhia (MIC, 20 μg·ml⁻¹), Salmonella typhimurium (MIC, 15 μg·ml⁻¹), Salmonella enteritidis (MIC, 8.5 μg·ml⁻¹), Aeromonas hydrophila (MIC, 4 μg·ml⁻¹), Yersinia sp. (MIC, 12.5 μg·ml⁻¹), Vibrio anguillarum (MIC, 25 μg·ml⁻¹), Shigella sp. (MIC, 6.3 μg·ml⁻¹), Vibrio parahaemolyticus (MIC, 12.5 μg·ml⁻¹), Candida albicans (MIC, 15 μg·ml⁻¹), Penicillium expansum (MIC, 40 μg·ml⁻¹), and Aspergillus niger (MIC, 25 μg·ml⁻¹). This is the first report of 7-amino-4-methylcoumarin in fungus and of the antimicrobial activity of this metabolite. The obtained results provide promising baseline information for the potential use of this unusual endophytic fungus and its components in the control of food spoilage and food-borne diseases.     

Polysaccharides isolated from <Emphasis Type="Italic">Phellinus baumii</Emphasis> stimulate murine splenocyte proliferation and inhibit the lipopolysaccharide-induced nitric oxide production in RAW264.7 murine macrophages

Polysaccharides isolated from Phellinus baumii (PBP) significantly enhanced both lipopolysaccharide (LPS)-induced B lymphocyte proliferation and concanavalin A (Con A)-induced T lymphocyte proliferation. However, PBP (12.5–100 μg ml⁻¹) significantly suppressed the LPS-induced nitric oxide (NO) production in RAW264.7 cells in a concentration-dependent manner. The maximal inhibition of PBP on NO production was 37.5% at 100 μg ml⁻¹. These results provide useful in vitro information to explain the immunostimulating activity and anti-inflammatory activity of PBP.     

Novel Antifouling and Antimicrobial Compound from a Marine-Derived Fungus Ampelomyces sp.

In this study, using a bioassay-guided isolation and purification procedure, we obtained 3-chloro-2,5-dihydroxybenzyl alcohol from a marine-derived Ampelomyces species that effectively inhibited larval settlement of the tubeworm Hydroides elegans and of cyprids of the barnacle Balanus amphitrite. The inhibitive effect on larval settlement was nontoxic and the EC₅₀ of 3-chloro-2,5-dihydroxybenzyl alcohol ranged from 3.19 μg ml⁻¹ to 3.81 μg ml⁻¹ while the LC₅₀ was 266.68 μg ml⁻¹ for B. amphitrite cyprids; EC₅₀ ranged from 0.67 μg ml⁻¹ to 0.78 μg ml⁻¹, and LC₅₀ was 2.64 μg ml⁻¹ for competent larvae of H. elegans, indicating that inhibitive effect of this compound was nontoxic. At a concentration of 50 μg per disc, this compound showed strong inhibitive effects on the growth of 13 out of 15 marine bacterial species tested in disc diffusion bioassay. Overall, the high inhibitory activities against bacteria and larval settlement as well as the non- or low-toxic nature of this compound to the barnacle and polychaete larvae suggest this compound could be a potent antifoulant and/or antibiotic.     

Effects of Thymus kotschyanus var. glabrescens Boiss. extract on mitomycin-C induced chromosomal aberrations and sister chromatid exchanges in human lymphocytes

In this investigation, clastogenic effects of Thymus kotschyanus var. glabrescens Boiss. extract (TE) and anticlastogenic effects of this extract against Mitomycin C (MMC) induced chromosome damage have been evaluated in human peripheral blood lymphocytes in vitro. Two series of experiments were conducted. In the first, only 10⁻⁵, 10⁻⁴, 10⁻³ and 10⁻² μl ml⁻¹ concentrations of TE were used for 48 h to detect potential clastogenicity. In the second, MMC (0.38 μg ml⁻¹) plus 10⁻⁵, 10⁻⁴, 10⁻³ and 10⁻² μl ml⁻¹ concentrations of TE were used for 48 h to determine anticlastogenic effects. TE did not increase sister chromatid exchanges (SCEs) (except 10⁻² μl ml concentration) and chromosome aberrations (CAs) significantly compared with negative and solvent controls. However, it decreased the frequency of MMC induced chromosome aberrations. Decreasing was significant at 10⁻⁴, 10⁻³ and 10⁻² μl ml⁻¹ concentrations. On the other hand, TE significantly increased MMC-induced SCEs for all treatment groups compared with positive control.     

Inhibition of galectin-3 mediated cellular interactions by pectic polysaccharides from dietary sources

Pectic polysaccharides from dietary sources such as Decalepis hamiltonii—swallow root (SRPP), Hemidesmus indicus (HPP), Nigella sativa—black cumin (BCPP), Andrographis serpyllifolia—(APP), Zingiber officinale—ginger (GRPP) and, citrus pectin (CPP) were examined for galectin inhibitory activity. Inhibition of (a) galectin-3 of MDA-MB-231 cells induced hemagglutination of red blood cells; (b) galectin-3 mediated interaction between normal/metastatic human buccal cells (NBC)/(MBC) and; (c) invasion of MDA-MB-231 and MBC in the invasive chamber was assessed. Results indicated that SRPP inhibited hemagglutination at Minimum Inhibitory Concentration (MIC) of 1.86 μg ml⁻¹ equivalent of carbohydrate as apposed to those of BCPP (130 μg ml⁻¹), APP (40 μg ml⁻¹), HPP (40 μg ml⁻¹) and CPP (25 μg ml⁻¹). GRPP even at concentration >1–6 mg ml⁻¹ did not inhibit agglutination. Also SRPP showed ∼15 and 2 fold potent anti hemagglutination activity relative to that of galectin-3 specific sugars—galactose (MIC-27.1 μg ml⁻¹) and lactose (MIC-4.16 μg ml⁻¹) respectively. Further, SRPP at 10 μg ml⁻¹ inhibited agglutination of NBC by galectin-3 of MDA-MB-231 cells. Modified swallow root pectic polysaccharide (MSRPP) of 50 kDa retained anti hemagglutination activity (MIC of 1.03 μg ml⁻¹) and inhibited MDA-MB-231 and MBC invasion by 73 and 50% with an IC₅₀ of 136 and 200 μg ml⁻¹ respectively. Both SRPP and MSRPP induced apoptosis up to 80% at 100 μg ml⁻¹ concentration by activating ∼2 and 8 folds of Caspase-3 activity. Sugar composition analysis and its correlation with the galectin inhibitory property indicated that pectic polysaccharides with higher arabinose and galactose content—arabinogalactan inhibited hemagglutination significantly.     

Production of indole-3-acetic acid, aromatic amino acid aminotransferase activities and plant growth promotion by Pantoea agglomerans rhizosphere isolates

The production of auxins, such as indole-3-acetic acid (IAA), by rhizobacteria has been associated with plant growth promotion, especially root initiation and elongation. Six indole-producing bacteria isolated from the rhizosphere of legumes grown in Saskatchewan soils and identified as Pantoea agglomerans spp. were examined for their ability to promote the growth of canola, lentil and pea under gnotobiotic conditions and for tryptophan (Trp)-dependent IAA production. Five of the isolates enhanced root length, root weight or shoot weight by 15–37% in at least one of the plant species, but isolates 3–117 and 5–51 were most consistent in enhancing plant growth across the three species. Indole concentrations in the rhizosphere of plants grown under gnotobiotic conditions increased in the presence of the rhizosphere isolates and when Trp was added 3 days prior to plant harvest. Isolates 3–117, 5–51 and 5–105 were most effective in increasing rhizosphere indole concentrations. Colony hybridization confirmed that all of the isolates possessed the ipdC gene which codes for a key enzyme in the Trp-dependent IAA synthetic pathway. The activity of amino acid aminotransferase (AAT), catalyzing the first step in the Trp-dependent synthetic pathway, was examined in the presence of Trp and other aromatic amino acids. All of the isolates accumulated Trp internally and released different amounts of IAA. The production of IAA from the isolates was greatest in the presence of Trp, ranging from 2.78 to 16.34 μg mg protein⁻¹ in the presence of 250 μg of Trp ml⁻¹. The specific activity of AAT was correlated with the concentration of IAA produced in the presence of Trp but not when tyrosine (Tyr), phenylalanine (Phe) or aspartate (Asp) was used as a sole nitrogen source. Isolate 3–117, which produced significant concentrations of IAA in the presence and absence of Trp, was able to use aromatic amino acids as sole sources of nitrogen and was most consistent in enhancing the growth of canola, lentil and pea may have potential for development as a plant growth-promoting inoculant. Responsible Editor: Peter A. H. Bakker.     

Carbon-limited fed-batch production of medium-chain-length polyhydroxyalkanoates from nonanoic acid by <Emphasis Type="Italic">Pseudomonas putida</Emphasis> KT2440

Pseudomonas putida KT2440 grew on glucose at a specific rate of 0.48 h⁻¹ but accumulated almost no poly-3-hydroxyalkanoates (PHA). Subsequent nitrogen limitation on nonanoic acid resulted in the accumulation of only 27% medium-chain-length PHA (MCL-PHA). In contrast, exponential nonanoic acid-limited growth (μ = 0.15 h⁻¹) produced 70 g l⁻¹ biomass containing 75% PHA. At a higher exponential feed rate (μ = 0.25 h⁻¹), the overall productivity was increased but less biomass (56 g l⁻¹) was produced due to higher oxygen demand, and the biomass contained less PHA (67%). It was concluded that carbon-limited exponential feeding of nonanoic acid or related substrates to cultures of P. putida KT2440 is a simple and highly effective method of producing MCL-PHA. Nitrogen limitation is unnecessary.     

Salicylic Acid Enhances Biocontrol Efficacy of the Antagonist Cryptococcus laurentii in Apple Fruit

Biological control and induced resistance are two of the promising approaches to the control of postharvest diseases. This study was conducted to evaluate the efficacy of salicylic acid (SA) alone or in combination with an antagonistic yeast, Cryptococcus laurentii, in controlling the blue mold disease caused by Penicillium expansum on apple fruit wounds. SA alone significantly inhibited the spore germination of P. expansum in vitro when its concentration was increased to 1000 μg ml⁻¹, but it was not effective in controlling the disease in vivo. Simultaneous application of SA and C. laurentii to the wounds on the apple fruit surface showed that SA could improve the efficacy of C. laurentii against P. expansum in a concentration-dependent manner, being most effective at 10 μg ml⁻¹ but less effective at a higher or lower concentrations. Besides reducing the blue mold incidence in the local wound sites, the combination of C. laurentii with SA at 10 μg ml⁻¹ also had a synergistic effect on the induction of fruit resistance to the disease, which might be associated with a rapid increase in peroxidase, phenylalanineamonialyase and lipoxygenase activities. In addition, SA at 100 μg ml⁻¹ or above showed an adverse effect on the growth of C. laurentii in vitro and in vivo, whereas it had no effect when its concentration was decreased to 10 μg ml⁻¹ or lower. This suggested that SA could enhance the biological activity of C. laurentii in apple fruit by inducing resistance to pathogens based on the antagonistic activity of C. laurentii.     

Endophytic fungi with anti-microbial,anti-cancer and anti-malarial activities isolated from Thai medicinal plants

A total of 81 Thai medicinal plant species collected from forests in four geographical regions of Thailand were examined for the presence of endophytic fungi with biological activity. Of 582 pure isolates obtained, 360 morphologically distinct fungi were selected for cultivation on malt Czapek broth and yeast extract sucrose broth, from which extracts were tested for biological activity. Extracts of 92 isolates could inhibit Mycobacterium tuberculosis (MIC 0.0625–200 μg ml⁻¹) when tested by the microplate Alamar blue assay, while extracts of six inhibited Plasmodium falciparum (IC₅₀ of 1.2–9.1 μg ml⁻¹) as determined by the [³H]hypoxanthine incorporation method. Strong anti-viral activity against Herpes simplex virus type 1 was observed in 40 isolates (IC₅₀ of 0.28–50 μg ml⁻¹). The sulphorhodamine B assay for activity against cancer cell lines revealed that 60 were active against human oral epidermoid carcinoma cells (EC₅₀ 0.42–20 μg ml⁻¹) and 48 against breast cancer cells (EC₅₀ 0.18–20 μg ml⁻¹). Bioactivity profile was affected by the type of culture medium. Given the high incidence of bioactive extracts and the fact that most of the isolated fungi could not be identified due to lack of spore formation, the results suggested that Thai medicinal plants can provide a wide variety of endophytes that might be a potential source of novel bioactive compounds. This revised version was published online in August 2006 with corrections to the Cover Date.     

Antimicrobial activity of crude extracts from mangrove fungal endophytes

The aim of this work was to select endophytic fungi from mangrove plants that produced antimicrobial substances. Minimal inhibitory concentrations (MIC) and minimal bactericidal concentrations (MBC) or minimal fungicidal concentrations (MFC) of crude extracts from 150 isolates were determined against potential human pathogens by a colorimetric microdilution method. Ninety-two isolates (61.3%) produced inhibitory compounds. Most of the extracts (28–32%) inhibited Staphylococcus aureus (MIC/MBC 4–200/64–200 μg ml⁻¹). Only two extracts inhibited Pseudomonas aeruginosa (MIC/MBC 200/>200 μg ml⁻¹). 25.5 and 11.7% inhibited Microsporum gypseum and Cryptococcus neoformans (MIC/MFC 4–200/8–200 μg ml⁻¹ and 8–200/8–200 μg ml⁻¹, respectively), while 7.5% were active against Candida albicans (MIC/MFC 32–200/32–200 μg ml⁻¹). None of the extracts inhibited Escherichia coli. The most active fungal extracts were from six genera, Acremonium, Diaporthe, Hypoxylon, Pestalotiopsis, Phomopsis, and Xylaria as identified using morphological and molecular methods. Phomopsis sp. MA194 (GU592007, GU592018) isolated from Rhizophora apiculata showed the broadest antimicrobial spectrum with low MIC values of 8–32 μg ml⁻¹against Gram-positive bacteria, yeasts and M. gypseum. It was concluded that endophytic fungi from mangrove plants are diverse, many produce compounds with antimicrobial activity and could be suitable sources of new antimicrobial natural products.     

Use of clove oil to anaesthetize freshwater amphipods

The use of clove oil as a potential anaesthetic for freshwater amphipods was examined at 20 °C. Individuals of Gammarus minus, a common species in southern Illinois, USA, spanning the entire body size range (4.3–14.3 mm), were used to test four anaesthetic concentrations varying from 1.48 × 10⁻⁴ ml ml⁻¹ to 5.9 × 10⁻⁴ ml ml⁻¹. Small-bodied individuals (mean size = 5.4 mm ± 0.27SE) were used to test additional concentrations, up to 14.7 × 10⁻⁴ ml ml⁻¹, a 10-fold span, to identify potential lethal concentrations. At the lowest concentration, time to anaesthesia and recovery was constant at all body sizes. For the three next higher concentrations, time to anaesthesia decreased with increasing concentration while recovery time increased. Activity of amphipods was not affected by the ethanol carrier. In addition, activity did not differ between amphipods that had recovered from anaesthesia and unexposed amphipods. At clove oil concentrations of 8.84 × 10⁻⁴ ml ml⁻¹ and 14.7 × 10⁻⁴ ml ml⁻¹, mortality was 7 and 40%, respectively, indicating, that 5.9 × 10⁻⁴ ml ml⁻¹ was a safe working concentration. No mortality was observed with Gammarus acherondytes, a federally endangered cave amphipod on which the protocol with 80 μl of stock was used in the field. The method enabled us to obtain information on the endangered amphipod which normally would have required the sacrifice of individuals. Thus, research can continue on species for which population numbers are low and for which basic information is needed to formulate meaningful recovery plans.     

Heterotrophic high-cell-density fed-batch and continuous-flow cultures of <Emphasis Type="Italic">Galdieria sulphuraria</Emphasis> and production of phycocyanin

Production of biomass and phycocyanin (PC) were investigated in highly pigmented variants of the unicellular rhodophyte Galdieria sulphuraria, which maintained high specific pigment concentrations when grown heterotrophically in darkness. The parental culture, G. sulphuraria 074G was grown on solidified growth media, and intensely coloured colonies were isolated and grown in high-cell-density fed-batch and continuous-flow cultures. These cultures contained 80–110 g L⁻¹ biomass and 1.4–2.9 g L⁻¹ PC. The volumetric PC production rates were 0.5–0.9 g L⁻¹ day⁻¹. The PC production rates were 11–21 times higher than previously reported for heterotrophic G. sulphuraria 074G grown on glucose and 20–287 times higher than found in phototrophic cultures of Spirulina platensis, the organism presently used for commercial production of PC.     

Cytotoxic and non-cytotoxic exometabolites of the cyanobacterium Nostoc insulare

The isolation, identification and quantification of exometabolites from culture media of the cyanobacterium Nostoc insulare are described. Besides the known exometabolite 4,4′-dihydroxybiphenyl (I), two more compounds, the β-carboline 9H-pyrido(3,4-b)indole (norharmane, II) and N,N′-(4,5-dimethyl-1,2-phenylene)bis-acetamide (III), were discovered. Concentrations of all three compounds in media and biomass of five 250 L cultures of N. insulare were determined. Culture medium values for I ranged between 200 and 1,250 μg L⁻¹ (1.1–6.7 μmol L⁻¹), for III between 115 and 390 μg L⁻¹ (0.5–1.8 μmol L⁻¹), whereas concentrations of II were conspicuously lower (2–16 μg L⁻¹ or 0.014–0.094 μmol L⁻¹). Amounts of III per volume of culture medium were about tenfold higher than in the biomass contained in an equal culture volume. This difference is an indication for an active excretion of III. Amounts of I and II in biomass and media were of no significant difference. In the neutral red uptake assay, I and II were found to be toxic against eukaryotic cells as follows: I was of considerable cytotoxicity in concentrations from 1,000 to 10 mg L⁻¹ and of lower cytotoxicity (causing a 27 % decrease of cell viability) in a concentration of 1,000 μg L⁻¹, whereas II was merely of considerable cytotoxicity in concentrations from 1,000 to 100 mg L⁻¹. Because of the cytotoxicity of I and because of the many known pharmacological effects of II there is a possibility that a certain amount of risk to humans and livestock comes from cultures or even from biomass- free culture media of N. insulare. The natural function of the examined exometabolites is discussed.     

Endocrinological investigation into the reproductive cycles of two sympatric skate species, Malacoraja senta and Amblyraja radiata, in the western Gulf of Maine

The smooth skate, Malacoraja senta, and thorny skate, Amblyraja radiata, are two commercially exploited batoids found within the Gulf of Maine. During the past five years, we conducted a large study to accurately describe important biological life history parameters previously lacking for these species. As part of that project, the current study reports our findings on the hormonal profiles associated with the reproductive cycles of M. senta and A. radiata. Blood samples were obtained from mature M. senta and A. radiata of both sexes from all months of the year, and plasma testosterone (T), estradiol (E₂) and progesterone (P₄) concentrations were determined using radioimmunoassay (RIA). In female M. senta and A. radiata, monthly T concentrations ranged from 4,522 pg ml⁻¹ to 1,373 pg ml⁻¹ and 31,940 pg ml⁻¹ to 14,428 pg ml⁻¹, E₂ concentrations from 831 pg ml⁻¹ to 60 pg ml⁻¹ and 8,515 pg ml⁻¹ to 2,902 pg ml⁻¹, and P₄ concentrations from 3,027 pg ml⁻¹ to 20 pg ml⁻¹ and 3,264 pg ml⁻¹ to 331 pg ml⁻¹, respectively. No statistical differences were detected between any months for any hormone. Estradiol concentrations were not correlated with ovary weight, shell gland weight, or diameter of the largest follicles in either species. Monthly T concentrations in male M. senta and A. radiata ranged from 23,146 to 12,660 pg ml⁻¹ and from 57,500pg ml⁻¹ to 24,737 pg ml⁻¹, while E₂ concentrations ranged from 7.5 pg ml⁻¹ to undetectable and 103 to 30 pg ml⁻¹, respectively. No statistical differences were observed between months for either steroid. Testosterone concentrations were weakly correlated with testes weight and percent of stage VI spermatocysts in A. radiata, however, no correlation was detected between T and stage VI spermatocysts in M. senta. Collectively, these data support the previous conclusion that M. senta and A. radiata of both sexes are capable of reproducing year round in the western Gulf of Maine.