Thermostable phycocyanin from the red microalga <Emphasis Type="Italic">Cyanidioschyzon merolae</Emphasis>, a new natural blue food colorant

The demand for natural food colorants is growing as consumers question the use of artificial colorants more and more. The phycobiliprotein C-phycocyanin of Arthospira platensis is used as a natural blue colorant in certain food products. The thermoacidophilic red microalga Cyanidioschyzon merolae might provide an alternative source of phycocyanin. Cyanidioschyzon merolae belongs to the order Cyanidiophyceae of the phylum Rhodophyta. Its natural habitat are sulfuric hot springs and geysers found near volcanic areas in, e.g., Yellowstone National Park in the USA and in Java, Indonesia. It grows optimally at a pH between 0.5 and 3.0 and at temperatures up to 56 °C. The low pH at which C. merolae grows minimizes the risk of microbial contamination and could limit production loss. As C. merolae lacks a cell wall, phycocyanin with a high purity number of 9.9 could be extracted by an osmotic shock using a simple ultrapure water extraction followed by centrifugation. The denaturation midpoint at pH 5 was 83 °C, being considerably higher than the A. platensis phycocyanin (65 °C). The C. merolae phycocyanin was relatively stable at pH 4 and 5 up to 80 °C. The high thermostability at slightly acidic pH makes the C. merolae phycocyanin an interesting alternative to A. platensis phycocyanin as a natural blue food colorant.     

Potential risk of biologic pollution associated to the introduction of <i>Pinus radiata</i> in grassland areas

Afforestation is a recommended practice to mitigate global warming. However, their implementation may generate undesirable impacts, mostly if exotic species are used. Plantations of Pinus radiata D Don in Ventania (Bs. As., Argentina) soils showed notorious increments of extractable P (Pe), which could affect the dynamic of this element as well as the degree of phosphorus saturation (GSP_Bray). The objectives of this study were: i) to quantify the GSP_Bray in Mollisols afforested with P. radiata comparing the results with those coming from adjacent, natural grassland areas (base line); ii) to evaluate the potential environmental risk induced by afforestation through the identification of a change point (PC) in the GSP_Bray indicative of a phosphate leaching increment. Treatments included mature stands of P. radiata (TB) and adjacent areas with natural grassland vegetation (TP). Samples were taken at 0-15; 15-30 and 30-45 cm soil depth, and texture, pH, total organic carbon (COT), Pe, soluble reactive phosphorus (PSR), phosphorus sorption index (ISP) and GSP_Bray were determined. The results showed a significant acidification in TB and an increase in the COT stock, indicating an additional atmospheric CO₂ sequestration by the trees. The Pe and PSR values were notoriously higher in TB, and they were reflected in a significant increment in the GSP_Bray with respect to TP. The detection of a significant PC in the GSP_Bray-PSR regression indicates higher chances of phosphate leaching in the forest stands, which could reach water courses, lakes and artificial reservoirs promoting their eutrophication. Because of the potential environmental pollution risk of biologic origin derived from the afforestation with P. radiata in Mollisols areas, their inclusion in clean development practices must be reconsidered.     

Identification and Characterization of Pathogenic Aeromonas veronii Biovar Sobria Associated with Epizootic Ulcerative Syndrome in Fish in Bangladesh

Sparse information is available on the virulence factors of Aeromonas strains isolated from diseased fish, from the environment, and from humans. In the present study, 52 Aeromonas isolates obtained from epizootic ulcerative syndrome (EUS) lesions in fish, from the aquatic environment, and from children with diarrhea in Bangladesh were identified by biochemical phenotyping (i.e., PhenePlate [PhP] typing) and DNA fingerprinting and then characterized with respect to certain putative virulence factors. The isolates from the fish exhibiting EUS symptoms were identified to be Aeromonas veronii biovar sobria by fatty acid methyl ester analysis and amplified fragment length polymorphism fingerprinting. Biochemical phenotyping revealed that all EUS-associated isolates belonged to a unique phenotype which was not identified among more than 1,600 environmental and diarrheal isolates in a previously collected database of PhP types of Bangladeshi Aeromonas isolates. The 52 Aeromonas isolates were investigated for the production of hemolysin and cytotoxin; for hemagglutination with erythrocytes from fish, human, and rabbit sources; for the presence of a cytolytic enterotoxin gene; and for adhesion to and invasion into fish cell lines. All of the EUS isolates produced all of the virulence factors investigated, as did also some of the environmental isolates, but the isolates from EUS were unique in their ability to agglutinate fish erythrocytes. Our results suggest that a clonal group of A. veronii biovar sobria is associated with, and may be a causative agent of, EUS in fish in Bangladesh.     

Bacoside production by suspension cultures of Bacopa monnieri (L.) Pennell

Cell suspension cultures of Bacopa monnieri (L.) Pennell, grown in modified MS medium, grew some 5–6 fold over 40 days. Selected cell lines produced the important saponin, bacoside A, up to 1 g/100 g dry wt after this time.     

Comparative analyses of the lysine binding site properties of apolipoprotein(a) kringle IV types 7 and 10.

Apolipoprotein(a) [apo(a)] shares extensive sequence similarity with plasminogen and consists of multiple tandem repeats of domains similar to plasminogen kringle IV (KIV), followed by domains homologous to the plasminogen KV and protease domains. The apo(a) KIV domains can be classified into 10 types on the basis of amino acid sequence (KIV(1)-KIV(10)) of which KIV(10) contains a canonical lysine binding site (LBS); KIV(10) mediates the lysine-dependent interaction of Lp(a) with certain biological substrates. Molecular modeling studies indicated the presence of weak LBS in each of KIV(5)-KIV(8), and subsequent biochemical studies have revealed contributions of these kringles to lysine-mediated interactions involving apo(a). The present study describes the direct demonstration of a weak LBS within KIV(7), as well as the first characterization of the ligand specificity of an LBS outside that of KIV(10). We have expressed both KIV(7) and KIV(10) from bacterial cells and purified them to homogeneity from cell lysates. Equilibrium binding analyses of the KIV(7) LBS using intrinsic fluorescence revealed an affinity for L-lysine and its analogues approximately 10-fold weaker (K(D) = 230 +/- 42 microM for epsilon-aminocaproic acid) than that of KIV(10) (K(D) = 33 +/- 4 microM for epsilon-aminocaproic acid). Moreover, we demonstrated differences in specificity of the LBS of KIV(7) in comparison with KIV(10) in that KIV(7) preferentially bound L-proline. Both kringles bind 4-aminobutyric acid with similar affinities albeit with apparently different mechanisms. Key Phe(62) --> Tyr and Asp(56) --> Glu substitutions in the KIV(7) LBS result in alterations in the size of the LBS and in the spatial relationship between the cationic and anionic centers in the LBS and thus account for the differences in the binding properties of KIV(7) and KIV(10).     

Rhythmic changes in testicular activity with lunar cycle in the forktail rabbitfish

Testosterone, 11-ketotestosterone and 17α ,20β-dihydroxy-4-pregnen-3-one in the forktail rabbitfish Siganus argenteus peaked around the full moon and the last quarter moon, respectively. Since changes in these hormones were correlated with histological change in the testis, the present study provided conclusive evidence for importance of the lunar periodicity in the reproductive activity of the forktail rabbitfish.     

Ameliorating effect of β-carotene on ethylmethane sulphonate-induced genotoxicity in the fish Oreochromis mossambicus

Genotoxic effects have been assessed in the fish Oreochromis mossambicus treated separately and conjointly with 0.2% ethylmethane sulphonate (EMS) and 0.05% β-carotene (BC) during five different time periods (6, 24, 48, 72 and 96 h) by analysis of endpoints such as chromosome aberrations, abnormal red blood cell nuclei, abnormal sperm morphology and protein contents (both qualitative and quantitative) of selected tissues, viz. muscle, heart, eye, brain, gill, liver, spleen and kidney. In addition, the relative efficacy of three doses of BC 0.02, 0.05 and 0.1%, in ameliorating genotoxic effects of 0.2% EMS was also tested after a treatment period of 48 h. EMS caused chromosomal aberrations, nuclear anomalies in red blood cells, abnormal sperm morphology and an apparent alteration of protein synthesis in various tissues. Some of these genotoxic effects of EMS appeared to be ameliorated by all three doses of BC, of which the 0.02% dose showed a marginally better efficacy.     

Neurons are protected from excitotoxic death by p53 antisense oligonucleotides delivered in anionic liposomes.

The potential of anionic liposomes for oligonucleotide delivery was explored because the requirement for a net-positive charge on transfection-competent cationic liposome-DNA complexes is ambiguous. Liposomes composed of phosphatidylglycerol and phosphatidylcholine were monodisperse and encapsulated oligonucleotides with 40-60% efficiency. Ionic strength, bilayer charge density, and oligonucleotide chemistry influenced encapsulation. To demonstrate the biological efficacy of this vector, antisense oligonucleotides to p53 delivered in anionic liposomes were tested in an in vitro model of excitotoxicity. Exposure of hippocampal neurons to glutamate increased p53 protein expression 4-fold and decreased neuronal survival to approximately 35%. Treatment with 1 microm p53 antisense oligonucleotides in anionic liposomes prevented glutamate-induced up-regulation of p53 and increased neuronal survival to approximately 75%. Encapsulated phosphorothioate p53 antisense oligonucleotides were neuroprotective at 5-10-fold lower concentrations than when unencapsulated. Replacing the anionic lipid with phosphatidylserine significantly decreased neuroprotection. p53 antisense oligonucleotides complexed with cationic liposomes were ineffective. Neuroprotection by p53 antisense oligonucleotides in anionic liposomes was comparable with that by glutamate receptor antagonists and a chemical inhibitor of p53. Anionic liposomes were also capable of delivering plasmids and inducing transgene expression in neurons. Anionic liposome-mediated internalization of Cy3-labeled oligonucleotides by neurons and several other cell lines demonstrated the universal applicability of this vector.