Effect of Environmental Factors on Cyanobacterial Abundance and Cyanotoxins Production in Natural and Drinking Water, Bangladesh.

Cyanobacterial blooms commonly appear during the summer months in ponds, lakes and reservoirs in Bangladesh. In these areas, fish mortality, odorous water and fish and human skin irritation and eye inflammation have been reported. The influence of physicochemical factors on the occurrence of cyanobacteria and its toxin levels were evaluated in natural and drinking water in Bangladesh. A highly sensitive immunosorbent assay was used to detect microcystins (MCs). Cyanobacteria were found in 22 of 23 samples and the dominant species were Microcystis aeruginosa, followed by Microcystisflosaquae, Anabeana crassa and Aphanizomenon flosaquae. Cyanobacterial abundance varied from 39 to 1315 x 10(3) cells mL(-1) in natural water and 31 to 49 x 10(3) cells mL(-1) in tap water. MC concentrations were 25-82300 pg mL(-1) with the highest value measured in the fish research pond, followed by Ishakha Lake. In tap water, MC concentrations ranged from 30-32 pg mL(-1). The correlation between nitrate-nitrogen (NO3-N) concentration and cyanobacterial cell abundance was R2 = 0.62 while that between cyanobacterial abundance and MC concentration was R2 = 0.98. The increased NO3-N from fish feed, organic manure, poultry and dairy farm waste and fertilizer from agricultural land eutrophicated the water bodies and triggered cyanobacterial bloom formation. The increased amount of cyanobacteria produced MCs, subsequently reducing the water quality.

Morphological, molecular, and biochemical characterization of astaxanthin-producing green microalga Haematococcus sp. KORDI03 (Haematococcaceae, Chlorophyta) isolated from Korea.

A unicellular red microalga was isolated from environmental freshwater in Korea, and its morphological, molecular, and biochemical properties were characterized. Morphological analysis revealed that the isolate was a unicellular biflagellated green microalga that formed a non-motile, thick-walled palmelloid or red aplanospore. To determine the taxonomical position of the isolate, its 18S rRNA and rbcL genes were sequenced and phylogenetic analysis was performed. We found that the isolate was clustered together with other related Haematococcus strains showing differences in the rbcL gene. Therefore, the isolated microalga was classified into the genus Haematococcus, and finally designated Haematococcus sp. KORDI03. The microalga could be cultivated in various culture media under a broad range of pH and temperature conditions. Compositions of the microalgal cellular components were analyzed, and its protein, carbohydrate, and lipid compositions were estimated to be 21.1 ± 0.2%, 48.8 ± 1.8%, and 22.2 ± 0.9%, respectively. In addition, D-glucose and D-mannose were the dominant monosaccharides in the isolate, and its amino acids were composed mainly of aspartic acid, glutamic acid, alanine, and leucine. Moreover, several polyunsaturated fatty acids accounted for about 80% of the total fatty acids in Haematococcus sp. KORDI03, and the astaxanthin content in the red aplanospores was estimated to be 1.8% of the dry cell weight. To the best of our knowledge, this is the first report of an Haematococcus sp. isolated from Korea, which may be used for bioresource production in the microalgal industry.

Photo-oxidative stress by ultraviolet-B radiation and antioxidative defense of eckstolonol in human keratinocytes.

Ultraviolet-B (UV-B) irradiation has been known to generate oxidative stress by increasing reactive oxygen species (ROS) in skin cells. Several naturally occurring antioxidant compounds isolated from marine algae are believed to protect against ROS. In this study, we assessed the antioxidative effect of eckstolonol isolated from Ecklonia cava against UV-B-induced ROS in human keratinocytes (HaCaTs). We investigated the effects of photo-oxidative stress by UV-B (50 mJ/cm(2)) and the antioxidative effects of eckstolonol using fluorometry, flow cytometry, microscopy, and cell viability and comet assays. UV-B irradiation decreased cell viability, which was restored in a dose-dependent manner with eckstolonol treatment (0, 5, 50, 100, and 200 µM). Moreover, eckstolonol reduced UV-B-induced ROS, lipid peroxidation, damaged DNA levels, and cell death. These antioxidative effects seem to be due to the enzymatic activities of catalase (CAT) and superoxide dismutase (SOD). Collectively, these results indicate that eckstolonol is capable of protecting keratinocytes from photo-oxidative stress.

Draft genome sequence of Marinobacterium stanieri S30, a strain isolated from a coastal lagoon in Chuuk state in Micronesia.

In this study, we isolated xylan-degrading bacteria from a coastal lagoon of Micronesia and identified the bacteria as Marinobacterium stanieri S30. GSFLX 454 pyrosequencing and sequence analysis of the M. stanieri S30 genome generated 4,007 predicted open reading frames (ORFs) that could be candidate genes for producing enzymes with different catalytic functions.

Neuroprotective effect of phlorotannin isolated from Ishige okamurae against H2O2 -induced oxidative stress in murine hippocampal neuronal cells, HT22.

The present study is designed to investigate the neuroprotective effect of a kind of phlorotannins, diphlorethohydroxycarmalol (DPHC) isolated from Ishige okamurae against hydrogen peroxide (H(2)O(2))-induced oxidative stress in murine hippocampal neuronal cells, HT22. H(2)O(2) treatment induced neurotoxicity, whereas DPHC prevented cells from H(2)O(2)-induced damage then restoring cell viability was significantly increased. DPHC slightly reduced the expression of Bax induced by H(2)O(2) but recovered the expression of Bcl-xL as well as caspase-9 and -3 mediated PARP cleavage by H(2)O(2). Intracellular reactive oxygen species (ROS) and lipid peroxidation was overproduced as the result of the addition of H(2)O(2); however, these ROS generations and lipid peroxidation were effectively inhibited by addition of DPHC in a dose-dependent manner. Moreover, DPHC suppressed the elevation of H(2)O(2)-induced Ca(2+) release. These findings indicate that DPHC has neuroprotective effects against H(2)O(2)-induced damage in neuronal cells, and that an inhibitory effect on ROS production may contribute to the underlying mechanisms.

Complete genome sequence of strain s85, a novel member of the family Flavobacteriaceae.

An agar-degrading marine bacterium identified as a novel member of the family Flavobacteriaceae (strain S85) was isolated from seawater in Micronesia. The sequenced strain S85 genome is composed of 3,384,629 bp in a circular chromosome, which includes 2,883 complete open reading frames.

Isolation, purification, and enzymatic characterization of extracellular chitosanase from marine bacterium Bacillus subtilis CH2.

A Bacillus subtilis strain was isolated from the intestine of Sebastiscus marmoratus (scorpion fish) that was identified as Bacillus subtilis CH2 by morphological, biochemical, and genetic analyses. The chitosanase of Bacillus subtilis CH2 was best induced by fructose and not induced with chitosan, unlike other chitosanases. The strain was incubated in LB broth, and the chitosanase secreted into the medium was concentrated with ammonium sulfate precipitation and purified by gel permeation chromatography. The molecular mass of the purified chitosanase was detected as 29 kDa. The optimum pH and temperature of the purified chitosanase were 5.5 and 60°C, respectively. The purified chitosanase was continuously thermostable at 40°C. The specific acitivity of the purified chitosanase was 161 units/mg. The N-terminal amino acid sequence was analyzed for future study.

Complete genome sequence of the agarase-producing marine bacterium strain s89, representing a novel species of the genus Alteromonas.

We report here the annotated genome sequence of the marine bacterium Alteromonas sp. S89 and the identification of six genes coding for agar-degrading enzymes. The sequenced Alteromonas sp. S89 genome is composed of a 3,864,871-bp circular chromosome that includes 3,236 complete open reading frames.

Whole-genome sequence of the xylanase-producing Mesoflavibacter zeaxanthinifaciens strain S86.

We isolated Mesoflavibacter zeaxanthinifaciens S86 as xylanase-producing bacteria from seawater sampled in Micronesia. Analysis of the M. zeaxanthinifaciens genome revealed that it contains a single circular chromosome of 3,704,661 bp with 3,249 putative open reading frames.

Chromene induces apoptosis via caspase-3 activation in human leukemia HL-60 cells.

In this study, the potent anti-tumor effects of brown algae on human leukemia HL-60 cells were investigated. The Sargassum siliquastrum extract among the 14 species of brown algae exhibited profound growth inhibitory effect on HL-60 cells in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, therefore, S. siliquastrum was selected for use in further experiments. The highest inhibitory activity of S. siliquastrum on HL-60 cells was detected in the chloroform fraction, and the active compound was identified as a kind of chromene, sargachromanol E (SE). SE treatment showed significant growth inhibitory effects on HL-60 cells in a dose-dependent manner by inducing apoptosis, as evidenced by the formation of apoptotic bodies, fragmented DNA ladder, and the accumulation of DNA in the sub-G(1) phase of cell cycle. SE induced apoptosis was accompanied by downregulation of Bcl-xL, upregulation of Bax, activation of caspase-3, and cleavage of poly (ADP-ribose) polymerase (PARP). Moreover, z-DEVD-fmk, a caspase-3 inhibitor, significantly inhibited cell cytotoxicity, apoptotic characteristics such as apoptotic bodies, sub-G(1) DNA content, and cleavage of PARP induced by SE. These results suggest that SE exerts its growth inhibitory effects on HL-60 cells through caspase-3-mediated induction of apoptosis. Therefore, SE offers promising chemotherapeuric potential to prevent cancers such as human leukemia.

Evaluation of anti-inflammatory effect of fucoxanthin isolated from brown algae in lipopolysaccharide-stimulated RAW 264.7 macrophages.

In this study, potential anti-inflammatory effect of fucoxanthin isolated from brown algae was assessed via inhibitory effect of nitric oxide (NO) production in lipopolysaccharide (LPS) induced RAW 264.7 macrophage cells. The Myagropsis myagroides was selected for further experiments due to its profound NO inhibitory effect, and was partitioned with different organic solvents. Highest NO inhibitory effect was detected in the chloroform fraction, and the active compound was identified as fucoxanthin, a kind of carotenoid available in brown algae evidenced high correlation with the inhibitory effect of NO production (r(2)=0.9511). Though, fucoxanthin significantly inhibited the NO production, it slightly reduced the prostaglandin E(2) (PGE(2)) production. The inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2) protein expressions were inhibited by fucoxanthin. Further, RT-PCR analysis indicated that the iNOS and COX-2 mRNA expressions were suppressed by fucoxanthin. Moreover, the release of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and interleukin-6 (IL-6), and the mRNA expression levels of those cytokines were reduced by the addition of fucoxanthin in a dose-dependent manner. Hence, these results suggest that the use of fucoxanthin may be a useful therapeutic approach for the various inflammatory diseases.

Inhibitory effect of diphlorethohydroxycarmalol on melanogenesis and its protective effect against UV-B radiation-induced cell damage.

In this study, potential inhibitory effect of 21 species of marine algae on melanogenesis was assessed via tyrosinase inhibitory effect. The Ishige okamurae extract tested herein evidenced profound tyrosinase inhibitory effect, compared to that exhibited by other marine algae extracts. Thus, I. okamurae was selected for use in further experiments, and was partitioned with different organic solvents. Profound tyrosinase inhibitory effect was detected in the ethyl acetate fraction, and the active compound was identified as the carmalol derivative, diphlorethohydroxycarmalol (DPHC), which evidenced higher levels of activity than that of commercial whitening agent. Intracellular reactive oxygen species (ROS) induced by ultraviolet (UV)-B radiation was reduced by the addition of DPHC and cell viability was dose-dependently increased. Moreover, DPHC demonstrated strong protective properties against UV-B radiation via damaged DNA tail length and morphological changes in fibroblast. Hence, these results indicate that DPHC isolated from I. okamurae has potential whitening effects and prominent protective effects on UV-B radiation-induced cell damages which might be used in pharmaceutical and cosmeceutical industries.

OPTIMAL GROWTH CONDITIONS AND ANTIOXIDATIVE ACTIVITIES OF CYLINDROTHECA CLOSTERIUM (BACILLARIOPHYCEAE)(1).

We isolated the unialgal strain of Cylindotheca closterium (Ehrenb.) Reimann et J. C. Lewin and produced an axenic strain using an antibiotic cocktail of enriched f/2 artificial seawater medium. The optimal growth conditions were estimated under 27 different combinations of temperature, salinity, and nutrients, and mass culture was performed based on the best specific growth conditions. Its antioxidant activities were determined from the extracts of methanol, water, and enzymes (proteases and carbohydrases). The maximum specific growth rate (µmax ) varied from 0.63 to 0.97 · d(-1) . The maximum cell density was 7.20 × 10(4) cells · mL(-1) , while the µmax was 0.82 · d(-1) in culture conditions of 20°C, 30 psu (practical salinity unit), and "F" nutrient concentrations on day 10 of the culture period. The scavenging rates for 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical were 72.5% and 69.4% from Viscozyme and methanol extracts, respectively. The enzymatic extracts of C. closterium prepared by the hydrolyses of Amyloglucosidase (AMG) and Viscozyme showed 45.8% and 45.5% nitric-oxide-scavenging rates, slightly lower than the activity of alpha-tocopherol (α-tocopherol) but similar to butylated hydroxytoluene (BHT). The extract from methanol and water showed 44.8% and 44.4% scavenging rates, statistically similar with BHT. The metal-chelating activities of the Kojizyme, Alcalase, methanol, Viscozyme, and Neutrase extracts were 67.1, 53.9, 53.2, 52.1, and 50.2 %, respectively, five to six times higher than the commercial antioxidants. The AMG, Viscozyme, and Neutrase extracts showed a remarkable linoleic acid peroxidation inhibition, which was higher than BHT and statistically similar with α-tocopherol.

Protective effect of enzymatic extracts from microalgae against DNA damage induced by H2O2.

The enzymatic extracts from seven species of microalgae (Pediastrum duplex, Dactylococcopsis fascicularis, Halochlorococcum porphyrae, Oltmannsiellopsis unicellularis, Achnanthes longipes, Navicula sp. and Amphora coffeaeformis) collected from three habitats (freshwater, tidal pool, and coastal benthic) at Jeju Island in Korea were investigated for their antioxidant activity. Of the extracts tested, the AMG 300 L (an exo 1, 4-alpha-D-glucosidase) extract of P. duplex, the Viscozyme extract of Navicula sp., and the Celluclast extract of A. longipes provided the most potential as antioxidants. Meanwhile, the Termamyl extract of P. duplex in an H(2)O(2) scavenging assay exhibited an approximate 60% scavenging effect. In this study, we report that the DNA damage inhibitory effects of P. duplex (Termamyl extract) and D. fascicularis (Kojizyme extract) were nearly 80% and 69% respectively at a concentration of 100 microg/ml. Thus, it is suggested that the microalgae tested in this study yield promising DNA damage inhibitory properties on mouse lymphoma L 5178 cells that are treated with H(2)O(2). Therefore, microalgae such as P. duplex may be an excellent source of naturally occurring antioxidant compounds with potent DNA damage inhibition potential.