Transcriptome analysis of signal transduction pathway involved in light inducing astaxanthin accumulation in Haematococcus pluvialis / 生物工程学报

The unicellular green alga Haematococcus pluvialis is the best source of natural astaxanthin (AST) in the world due to its high content under stress conditions. Although high light (HL) can effectively induce AST biosynthesis, the specific mechanisms of light signal perception and transduction are unclear. In the current study, we used transcriptomic data of normal (N), high white light (W), and high blue light (B) to study the mechanisms of light inducing AST accumulation from the point of photoreceptors. The original data of 4.0 G, 3.8 G, and 3.6 G for N, W, and B were obtained, respectively, by the Illumina Hi-seq 2000 sequencing technology. Totally, 51 954 unigenes (at least 200 bp in length) were generated, of which, 20 537 unigenes were annotated into at least one database (NR, NT, KO, SwissProt, Pfam, GO, or KOG). There were 1 255 DEGs in the W vs N, 1 494 DEGs in the B vs N, and 1 008 DEGs in the both W vs N and B vs N. KEGG enrichment analysis revealed that photosynthesis, oxidative phosphorylation, carotenoid biosynthesis, fatty acids biosynthesis, DNA replication, nitrogen metabolism, and carbon metabolism were the significantly enriched pathways. Moreover, a large number of genes encoding photoreceptors and predicted interacting proteins were predicted in Haematococcus transcriptome data. These genes showed significant differences at transcriptional expression levels. In addition, 15 related DEGs were selected and tested by qRT-PCR and the results were significantly correlated with the transcriptome data. The above results indicate that the signal transduction pathway of "light signal - photoreceptors - interaction proteins - (interaction proteins - transcription factor/transcriptional regulator) - gene expression - AST accumulation" might play important roles in the regulation process, and provide reference for further understanding the transcriptional regulation mechanisms of AST accumulation under HL stress.

Advances in astaxanthin biosynthesis in Haematococcus pluvialis / 生物工程学报

Astaxanthin is widely applied as a nutraceutical, pharmaceutical, and aquaculture feed additive because of its high antioxidant activity. Haematococcus pluvialis is a microalgal species that can largely accumulate astaxanthin under adverse environmental conditions. Here we review the research progress of astaxanthin biosynthesis in H. pluvialis, including the induction and regulation of massive astaxanthin, the relationship between astaxanthin synthesis, photosynthesis and lipid metabolism.

Aquaculture biological waste as culture medium to cultivation of Ankistrodesmus gracilis (Reinsch) Korshikov / Resíduos biológicos de aquicultura como meio de cultura para o cultivo de Ankistrodesmus gracilis (Reinsch) Korshikov

Abstract Current study investigated the effectiveness of different macrophytes as culture media for Ankistrodesmus gracilis in laboratory conditions. Significant difference (p < 0.05) was reported in cell density with regard to conventional culture medium (CHU12) and macrophytes culture media. Mean cell density in NPK, Eichhornia crassipes and E. azurea media was higher (p < 0.05) than in conventional culture medium. Chlorophyll-a was higher than 1 g.L-1, except in CHU12 (0.7 ± 0.4 g.L-1) and T. domingensis (0.8 ± 0.3 g.L-1) media. Nitrate decreased sharply as from the 7th-day of the experiment. Ammonium and total phosphorus were highest in culture media and ranged between 0.4 g.L-1 (P. cordata medium) and 1.7 g.L-1 (CHU12 medium) for ammonium, and between 0.8 g.L-1 (CHU12 medium) and 1.9 g.L-1 (T. domingensis medium) for total phosphorus. Results revealed inorganic fertilizer and macrophytes combined with vitamins may be effective as culture media and strongly supports the growth of Ankistrodesmus gracilis, since cell density and biochemical composition are similar to or higher than conventional culture medium (CHU12). Macrophyte is a tool for aquaculture since biological wastes may be used with nutrients to improve the cultivation of microalgae.

Resumo O presente estudo investigou a eficácia na utilização de diferentes macrófitas como meio de cultura para Ankistrodesmus gracilis em condições laboratoriais. Foi observada diferença significativa (p < 0,05) entre a densidade celular em relação aos meios de cultura convencional (CHU12) e de macrófitas. Os meios de cultura com Eichhornia crassipes, E. azurea e NPK apresentaram densidade celular maiores (p < 0,05) que o meio de cultura convencional. Os teores de clorofila-a foram superiores a 1 g.L-1, exceto nos meios de cultura CHU12 (0.7 ± 0.4 g.L-1) e de T. domingensis (0.7 ± 0.3 g.L-1). O nitrato diminuiu acentuadamente a partir do 7º dia de experimento. Dentre os nutrientes, amônia e o fósforo total foram os mais elevados nos meios de cultura, variando entre 0.4 g.L-1 (meio de P. cordata) a 1,7 g.L-1 (meio CHU12) para amônia e, entre 0.8 g.L-1 (meio CHU12) e 1,9 gL-1 (meio de T. domingensis) para o fósforo total. Os resultados do presente estudo mostraram que o uso de fertilizantes inorgânicos e macrófitas, combinados com vitaminas, pode ser eficaz como meio de cultura no crescimento de Ankistrodesmus gracilis, uma vez que a densidade celular e a composição bioquímica foram semelhantes ou superiores ao meio de cultura convencional (CHU12). As macrófitas são ferramentas a serem adotadas na aquicultura, desde que os resíduos biológicos podem ser usados como nutrientes para melhorar o cultivo de microalgas.

In vitro inactivation of thrombin generation by polysulfated fractions isolated from the tropical coenocytic green seaweed Caulerpa racemosa (Caulerpaceae, Bryopsidales) / Inativação in vitro da geração de trombina por frações polissulfatadas isoladas da macroalga verde cenocítica tropical Caulerpa racemosa (Caulerpaceae; Bryopsidales)

Pharmacological efficacy of Caulerpa racemosa (Chlorophyta) sulfated polysaccharidic (SPs) fractions (F I→III) on models of coagulation and inflammation has been demonstrated, but not their effects on thrombin generation (TG). This study examined fractions for composition and physical-chemical characteristics and in vitro inactivation of TG by F I and F II in 60-fold diluted human plasma using continuous method. Papain-extraction yield of 0.7% revealed F I→III by DEAE-cellulose chromatography, with differences among the relative proportions of sulfate (17.37-24.00%), total sugars (30.03-48.34%) and absence of proteins. Charge density patterns and molecular sizes > 100 kDa of the fractions were verified by both agarose/polyacrylamide analyses, respectively. These electrophoreses combined with toluidine blue/Stains-All also indicated nonSPs. Anticoagulant effects of 4.76 (F I), 12.00 (F II) and 2.32 (F II) IU mg-1 by activated partial thromboplastin time test were recorded against heparin (193 IU mg-1), without changes in prothrombin time. Diluted plasma treated with F I and F II reduced concentration-dependent and sulfation pattern TG by both intrinsic and extrinsic pathways, with 50% inactivation by intrinsic pathway of F II even at 4.1 µg. Heparin abolished TG at least 4-fold lower. Therefore, C. racemosa produces SPs with TG inhibition.

Eficácia farmacológica de frações (F I→III) polissacarídicas sulfatadas (PSs) da Chlorophyta Caulerpa racemosa sobre modelos de coagulação e inflamação tem sido demonstrada, exceto seus efeitos sobre geração de trombina (GT). Examinaram-se frações quanto à composição, características físico-químicas e inativação in vitro de GT por F I e F II, em plasma humano diluído 60 vezes usando método contínuo. Rendimento de extração-papaína (0,7%) revelou, por cromatografia de DEAE -celulose, F I→III com diferenças entre as proporções relativas de sulfato (17,37-24,00%), açúcares totais (30,03-48,34%) e ausência de proteínas. Foram verificados, por ambas as análises agarose/poliacrilamida, graus de densidade de carga e tamanhos moleculares > 100 kDa das frações, respectivamente. Também essas eletroforeses, combinadas com azul de toluidina/Stains-All, indicaram polissacarídeos não sulfatados. Foram registrados, pelo teste do tempo de tromboplastina parcial ativada, efeitos anticoagulantes de 4,76 (F I), 12,00 (F II) e 2,32 (F II) UI mg-1 contra heparina (193 UI mg- 1), porém não modificando tempo de protrombina. Plasma diluído tratado com F I e F II reduziu GT por ambas as vias intrinsíca/extrínsica, dependente de concentração e grau de sulfatação, com F II em 4,1 µg apresentando eficácia de 50% pela via intrínsica. Heparina, quatro vezes menos, aboliu GT. Portanto, C. racemosa produz PSs com inibição de GT.

An anti-dengue and anti-herpetic polysulfated fraction isolated from the coenocytic green seaweed Caulerpa cupressoides inhibits thrombin generation in vitro / Uma fração polissulfatada anti- dengue e anti- herpética isolada da alga marinha verde cenocítica Caulerpa cupressoides inibe geração de trombina in vitro

The antiviral potency of Caulerpa cupressoides (Chlorophyta) sulfated polysaccharidic fractions (Cc -SP1→3) associated with thrombin generation (TG) assay is unknown. This study analyzed the structure of Cc-SP1 and its effects against herpes viruses (HSV-1 and HSV-2), dengue-virus (DENV-1) , human-metapneumovirus (HMPV) and adenovirus (AdV) in Vero and mosquito C6/36 cell-lines and as inhibitor of TG in 60- fold diluted human plasma using continuous system. Infrared analysis indicated ulvan containing 11% sulfate, 40.16% total sugars and 6% uronic acid. Procedures of agarose/polyacrylamide gels electrophoresis revealed two major components presenting sulfate and non SPs, with molecular dispersion ranging from 8 to > 100 kDa, respectively, as confirmed by gel permeation chromatography. Cc-SP1 did not cause cytotoxicity up to 1000 µg mL-1 and was more effective inhibitor of DENV-1 (96%, 0.35 µg mL-1) compared with HSV-1 (90%, 27 µg mL- 1) and HSV-2 (99.9%, 0.9 µg mL-1) in Vero cell-line, whose selectivity indexes were > 714, > 9.2 and > 131, respectively, but was inactive against HMPV, AdV and C6/36 cell-line using plate reduction assay. Cc-SP1 required concentration 20.8-fold higher of SPs than heparin for abolishes intrinsically TG, but at concentrations so far the anti-HSV-1. Therefore, TG assay provides data to guide studies of Cc-SP1 on anticoagulant/antiviral effects.

A potência antiviral de frações polissacarídicas sulfatadas de Caulerpa cupressoides (Chlorophyta) (Cc-PS1→3) é desconhecida e associada com ensaio de geração de trombina (GT). Analisaram-se de Cc-PS1 estrutura e efeitos contra herpesviroses (HSV-1 e HSV-2), vírus da dengue (DENV-1), metapneumovírus-humano (MPVH) e adenovírus (AdV) em linhagens de células Vero e de mosquito C6/36 e como inibidora de GT em plasma humano diluído 60 vezes usando sistema contínuo. Análise de infravermelho indicou 'ulvana' contendo sulfato (11%), açúcares totais (40,16%) e ácido urônico (6%). Procedimentos de electroforese em géis de agarose/poliacrilamida revelaram dois componentes majoritários apresentando sulfato e polissacarídeos não sulfatados, com dispersão molecular variando de 8 a > 100kDa, respectivamente, confirmada por cromatografia de permeação em gel. Cc-PS1 não causou citotoxicidade até 1000 µg mL-1 e se mostrou, em linhagem de célula Vero usando ensaio de redução de placas, inibidora efetiva de DENV-1 (96%; 0,35 µg mL-1) comparada com HSV-1 (90%; 27 µg mL-1) e HSV-2 (99,9%; 0,9 µg mL-1), cujos índices de seletividade foram > 714; > 9,2 e > 278, respectivamente, enquanto, demonstrou-se ineficaz sobre HMPV, AdV e linhagem de célula C6/36. Foi requerida concentração 20,8 vezes maior de Cc-PS1 que heparina para abolir GT intrinsicamente, mas em concentrações diferentes da anti-HSV-1. Portanto, ensaio de GT fornece dados para estudos direcionados sobre efeitos anticoagulante/antiviral de Cc-PS1.

Degradation of textile dyes by cyanobacteria

Abstract Dyes are recalcitrant compounds that resist conventional biological treatments. The degradation of three textile dyes (Indigo, RBBR and Sulphur Black), and the dye-containing liquid effluent and solid waste from the Municipal Treatment Station, Americana, São Paulo, Brazil, by the cyanobacteria Anabaena flos-aquae UTCC64, Phormidium autumnale UTEX1580 and Synechococcus sp. PCC7942 was evaluated. The dye degradation efficiency of the cyanobacteria was compared with anaerobic and anaerobic-aerobic systems in terms of discolouration and toxicity evaluations. The discoloration was evaluated by absorption spectroscopy. Toxicity was measured using the organisms Hydra attenuata, the alga Selenastrum capricornutum and lettuce seeds. The three cyanobacteria showed the potential to remediate textile effluent by removing the colour and reducing the toxicity. However, the growth of cyanobacteria on sludge was slow and discoloration was not efficient. The cyanobacteria P. autumnale UTEX1580 was the only strain that completely degraded the indigo dye. An evaluation of the mutagenicity potential was performed by use of the micronucleus assay using Allium sp. No mutagenicity was observed after the treatment. Two metabolites were produced during the degradation, anthranilic acid and isatin, but toxicity did not increase after the treatment. The cyanobacteria showed the ability to degrade the dyes present in a textile effluent; therefore, they can be used in a tertiary treatment of effluents with recalcitrant compounds.

Response of phytoplankton to an experimental fish culture in net cages in a subtropical reservoir / Resposta do fitoplâncton ao cultivo experimental de peixes em tanques-rede em um reservatório subtropical

Abstract This study aimed to evaluate nutrients concentration and spatial-temporal changes in phytoplankton biovolume during an experimental fish culture in net cages in a lateral arm of Salto Caxias reservoir, Brazil. Two sampling stations were placed in the affected lateral arm and other two in a cageless lateral arm. Neither abiotic variables nor phytoplankton biovolume presented significant differences between the treatments. Only temporal changes were confirmed by the analysis performed. Both lateral arms were classified as oligotrophic, reflecting low influence of the net cages. Phytoplankton growth seems to be limited by nitrogen. Biovolume values were, in general, low and five major functional groups were recognized (E, F, G, K and P). In summer higher biovolume values were observed and representatives of Chlorophyceae and Cyanobacteria belonging to the functional groups F and K, respectively, were the most important. In winter phytoplankton was mainly composed by Bacillariophyceae taxa from P group. G group was also restricted to winter and E group occurred in winter and summer. The variations recorded in phytoplankton structure appear to have been mainly influenced by seasonal changes in temperature, precipitation and nutrients availability. The effects of net cages on the abiotic variables and phytoplankton biovolume appear to have been small, probably due to the small number of net cages employed and the system dilution capacity. However, a permanent monitoring of phytoplankton is recommended, since this environment has a carrying capacity, from which the trophic state may increase.

Resumo Este estudo objetivou avaliar alterações nas variáveis abióticas e no biovolume fitoplanctônico durante o cultivo experimental de peixes em tanques-rede em um braço lateral do reservatório de Salto Caxias, Brasil. Foram selecionadas duas estações de amostragem no braço com tanques-rede e outras duas em um braço sem tanques. As variáveis abióticas e o biovolume fitoplanctônico não apresentaram diferenças significativas entre os locais estudados. Apenas mudanças temporais foram confirmadas pelas análises utilizadas. Os dois braços laterais foram classificados como oligotróficos, refletindo a baixa influência dos tanques-rede. O crescimento do fitoplâncton parece ter sido limitado principalmente por nitrogênio. Os valores de biovolume foram, em geral, baixos e cinco principais grupos funcionais foram observados (E, F, G, K e P). No verão, os maiores valores de biovolume foram observados e representantes de Chlorophyceae e Cyanobacteria dos grupos funcionais F e K, respectivamente, se destacaram. No inverno, o fitoplâncton foi composto principalmente por táxons de Bacillariophyceae do grupo P. O grupo G também foi restrito ao inverno e o grupo E ocorreu no inverno e verão. As variações registradas na estrutura do fitoplâncton parecem ter sido principalmente influenciadas pelas mudanças sazonais de temperatura, precipitação e disponibilidade de nutrientes. Os efeitos dos tanques-rede sobre as variáveis abióticas e biovolume fitoplanctônico parecem ter sido pequenos, provavelmente devido ao pequeno número de tanques utilizados e a capacidade de diluição do sistema. Entretanto, o monitoramento permanente do fitoplâncton é recomendado, uma vez que este ambiente possui uma capacidade de suporte, a partir da qual o estado trófico pode aumentar.

Phytoplankton composition of the water and gastrointestinal tract of the mussel Diplodon enno (Ortmann, 1921) from São Francisco river (Bahia, Brazil) / Composição fitoplanctônica da água e do trato gastrointestinal do molusco Diplodon enno (Ortmann) do rio São Francisco (BA - Brasil)

Abstract The knowledge on diet composition of the freshwater mussel Diplodon enno (Ortmann) would aid in its culture and propagation allowing, this way, the replacement of natural endangered populations in Brazil. Microalgae are the main food source for captive mussels and unionids have displayed an ability to sort algae based on the cellular characteristics prior to ingestion. The main objective of the present work is to analyze the phytoplankton composition of the water from and of the gastrointestinal contents of the mussel D. enno, as an initial step for development of a suitable rearing diet. Therefore, water samples and bivalve specimens were collected from S. Francisco River, city of Paulo Afonso, Bahia, Brazil. The microalgal composition found in water and stomach/gut content samples was very diverse being represented by the following divisions: Cyanophyta, Chlorophyta, Dinophyta and Heterokontophyta (Diatoms). Concerning the relative abundance of microalgae divisions, it is possible to state, for the water and gastrointestinal contents, that Cyanophyta represents 15% and 14%, Chlorophyta 54% in both, Heterokontophyta 31% and 27% and Dinophyta 0% and 5%, respectively. According to the Brazilian CETESB criteria for phytoplankton species classification, 50% of Cyanophyta and 15% of Chlorophyta species observed in the water samples were classified as “very frequent”, as were 68% of Heterokontophyta and 33% of Chlorophyta species in the gut/stomach tract samples. Focusing at a species level, although in the water only Coelastrum sp. and Chroococcus sp. were observed in 100% and 75% of the samples, respectively, in the gastrointestinal tract the species Staurastrum sp., Aulacoseira sp., Scenedesmus sp. and Fragilaria crotonensis occurred in 80% to 100% of the samples. The present results showed that D. enno feeds not only on small chlorophytes microalgae, due to their convenient size that facilitates higher feeding rates, but also on large size diatoms, due to a possible nutritional advantage for the bivalves. Thus, a diet composed by large diatoms and small chlorophytes microalgae may be considered as the most reasonable for the maintenance of D. enno populations.

Resumo O conhecimento da composição da dieta do molusco de água doce Diplodon enno é de extrema importância para a sua cultura e propagação, permitindo desta forma a recuperação de populações em perigo de extinção no Brasil. As microalgas são a principal fonte de alimento para moluscos filtradores e os uniónidos sendo selecionadas por estes com base nas suas características celulares. O principal objectivo deste trabalho é analisar a composição fitoplanctônica da água e do conteúdo gastrointestinal do molusco D. enno, de forma a desenvolver uma dieta apropriada ao seu cultivo. Para isso, amostras de água e espécimes de bivalves foram recolhidos do rio São Francisco, cidade de Paulo Afonso, Bahia, Brasil. A composição de microalgas encontrada na água e no conteúdo do estômago/intestino foi muito diversa, sendo representada pelas seguintes divisões: Cyanophyta, Chlorophyta, Dinophyta e Heterokontophyta (diatomáceas). Atendendo à abundância relativa de cada divisão de microalgas, podemos afirmar que na água e no trato gastrointestinal Cyanophyta representa 15% e 14%, Chlorophyta 54% em ambos, Heterokontophyta 31% e 27% e Dinophyta 0% and 5%, respectivamente. De acordo com o critério CETESB de classificação de espécies de fitoplâncton, 50% das espécies de Cyanophyta e 15% das de Chlorophyta observadas nas amostras de água foram classificadas como “muito frequentes”, tal como aconteceu para 68% das espécies de Heterokontophyta e 33% das de Chlorophyta nas amostras do conteúdo do trato gastrointestinal. Realçando as espécies presentes, apesar de apenas Coelastrum sp. e Chroococcus sp. terem sido observadas em 100% e 75% das amostras de água, respectivamente, nas amostras de trato gastrointestinal Staurastrum sp., Aulacoseira sp., Scenedesmus sp. e Fragilaria crotonensis foram identificadas em entre 80% e 100% das amostras. Os resultados deste estudo mostram que D. enno se alimenta não apenas de pequenas microalgas clorófitas, devido ao seu pequeno tamanho que possibilita maiores taxas de filtração, como também de diatomáceas maiores, devido a uma possível vantagem nutricional para os bivalves. Assim sendo, uma dieta composta por diatomáceas e pequenas clorófitas poderá ser considerada a mais indicada para a manutenção de populações de D. enno.

Enhancement of pigment extraction from B. braunii pretreated using CO2 rapid depressurization

Abstract Extraction of compounds from microalgae requires cell disruption as a pretreatment to increase extraction yield. Botryococcus braunii is a microalga with a significant content of carotenoids and other antioxidant compounds, such as chlorophylls. Cell disruption of B. braunii using CO2 rapid depressurization was studied as a pretreatment for the extraction of carotenoid and chlorophyll pigments. We studied the effect of temperature (21–49 °C) and pressure (6–13 MPa) during static compression on pigment recovery with supercritical CO2 at 40 °C, 30 MPa and solvent flow of 4.7 L NPT/min. Within the experimental region, the extraction yield of carotenoids and chlorophylls increased by 2.4- and 2.2-fold respectively. Static compression conditions of high pressure and low temperature increased the extraction of carotenoids and especially chlorophylls. We selected 21 °C and 13 MPa as the cell disruption condition, which produced 1.91 g/kg d.s. of carotenoids and 14.03 mg/kg d.s. of chlorophylls. Pretreated microalga gave a 10-fold higher chlorophyll extraction yield compared to the untreated sample. While for carotenoids and tocopherols were 1.25 and 1.14-fold higher, respectively. Additionally, antioxidant activity of pretreated microalga (33.22 mmol TE/kg oil) was significantly higher than the value for the untreated samples (29.11 mmol TE/kg oil) (p ≤ 0.05). Confocal microscopy images showed morphological differences between micro-colonies with and without disruption treatment, suggesting that partial cell disruption by rapid depressurization improved the extraction of microalga compounds.

Structure and dynamics of the phytoplankton community within a maturation pond in a semiarid region / Estrutura e dinâmica da comunidade fitoplanctônica de uma lagoa de maturação em região semiárida

Abstract In northeastern Brazil, stabilization ponds are very suitable for wastewater treatment because of the relative great land availability and environmental conditions (e.g., high temperature) favorable for microorganism optimal development. However, blooms of potentially toxic cyanobacteria may affect the use of these treatment ponds due to resulting effluent poor quality. The objective of this study was to evaluate the dynamics of phytoplankton communities and the occurrence of cyanobacteria in a maturation pond located immediately after a series of two ponds. Temperature, dissolved oxygen, pH, BOD, N, and P were measured during a period of four months when samples were collected from the surface and the bottom of 7 sampling points distributed inside the pond. The phytoplankton of collected samples was also identified and classified using a conventional optical microscopy. Analysis of variance and Tukey test were used to evaluate the results. The three phytoplankton divisions found (Cyanophyta, Chlorophyta, and Euglenophyta) did not change considerably through surface and bottom. However, they changed greatly over the sampled months; great dominance of Cyanophyta was found at April and October, while Chlorophyta dominated the lagoon in September. Low superficial organic loads (between 78 and 109 kg BOD.ha–1.d–1) and N:P ≤ 10 were the determinant factors that favored the predominance of Cyanophyta. The presence of two potentially toxic species of Cyanophyta, Oscillatoria sp. and Microcystis aeruginosa, indicates that caution is required when considering the final destination of treated effluent and suggests a need to assess the risks and benefits associated with the use of the treatment technology.

Resumo No nordeste do Brasil, as lagoas de estabilização são muito adequadas para o tratamento de águas residuárias por causa da disponibilidade relativamente grande de terra e das condições ambientais (por exemplo, altas temperaturas) favoráveis ao melhor desenvolvimento dos microorganismos. Entretanto, florações de cianobactérias potencialmente tóxicas podem afetar o uso dessas lagoas de tratamento, devido à consequente qualidade inferior do efluente. O objetivo deste estudo foi avaliar a dinâmica das comunidades de fitoplâncton e a ocorrência de cianobactérias em uma lagoa de maturação situada após duas lagoas em série. Temperatura, oxigênio dissolvido, pH, DBO, N e P foram medidos durante um período de quatro meses, durante o qual amostras foram coletadas na superfície e fundo em sete pontos de amostragem da lagoa. As comunidades de fitoplâncton das amostras coletadas foram também identificadas e classificadas utilizando-se um microscópio óptico convencional. Para avaliar os resultados utilizou-se a análise de variância e o teste de Tukey. Para as três divisões de fitoplâncton encontradas (Cyanophyta, Chlorophyta e Euglenophyta), não houve diferença significativa para as amostras de superfície e de fundo de um mesmo mês. Entretanto, ocorreu grande variação para as amostras dos diferentes meses; nos meses de abril e outubro houve uma predominância de Cyanophyta, ao passo que em setembro o predomínio na lagoa foi de Chlorophyta. Os fatores determinantes que favoreceram o predomínio de Cyanophyta foram a baixa carga orgânica superficial aplicada (entre 78 e 109 kg DBO.ha–1.d–1) e N:P ≤ 10. A presença de duas das espécies de Cyanophyta, Oscillatoria sp. e Microcystisaeruginosa, consideradas potencialmente tóxicas, indica que é necessária precaução quando se considera o destino final do efluente tratado e sugere a necessidade de avaliar os riscos e benefícios associados ao uso da tecnologia de tratamento.

Low field nuclear magnetic resonance for rapid quantitation of microalgae lipid and its application in high throughput screening / 生物工程学报

A rapid and accurate determination method of lipids in microalgae plays a significant role in an efficient breeding process for high-lipid production of microalgae. Using low field nuclear magnetic resonance (LF-NMR), we developed a direct quantitative method for cellular lipids in Chlorella protothecoides cells. The LF-NMR signal had a linear relationship with the lipid content in the microalgae cells for both dry cell samples and algal broth samples (R2 > 0.99). These results indicated that we could use this method for accurate determination of microalgal lipids. Although LF-NMR is a rapid and easy lipid determination method in comparison to conventional methods, low efficiency would limit its application in high throughput screening. Therefore, we developed a novel combined high throughput screening method for high-lipid content mutants of C. protothecoides. Namely, we initially applied Nile red staining method for semi-quantification of lipid in the pre-screening process, and following with LF-NMR method for accurate lipid determination in re-screening process. Finally, we adopted this novel screening method in the breeding process of high-lipid content heterotrophic cells of C. protothecoides. From 3 098 mutated strains 108 high-lipid content strains were selected through pre-screening process, and then 9 mutants with high-lipid production were obtained in the re-screening process. In a consequence, with heterotrophical cultivation of 168 h, the lipid concentration could reach 5 g/L, and the highest lipid content exceeded 20% (W/W), which was almost two-fold to that of the wild strain. All these results demonstrated that the novel breeding process was reliable and feasible for improving the screening efficiency.

Growth of Haematococcus pluvialis Flotow in alternative media / Crescimento de Haematococcus pluvialis Flotow em meios alternativos

Abstract Current study investigates the effect of two alternative media NPK (20-5-20) fertilizer and NPK plus macrophyte (M+NPK) compared to the commercial medium (WC) under growth rate and physiological parameters in batch culture mode (2-L), and verifies whether the use of fertilizer (NPK) and macrophyte (Eichhornia crassipes) would be a good tool for Haematococcus pluvialis culture in the laboratory. The highest number of cells of H. pluvialis has been reported in NPK medium (5.4 × 105cells.mL–1) on the 28th day, and in the M+NPK and WC media (4.1 × 105 cells.mL–1 and 2.1 × 105 cells.mL–1) on the 26th day, respectively. Chlorophyll-a contents were significantly higher (p<0.05) in NPK medium (41-102 µg.L–1) and lower in WC and M+NPK media (14-61 µg.L–1). The astaxanthin content was less than 0.04 mg.L–1. Production cost of 10-L of H. pluvialis was low in all media, and NPK and M+NPK media had a cost reduction of 65% and 82%, respectively when compared with commercial medium (WC). The use of a medium based on commercial fertilizer and macrophyte (E. crassipes) produced a new medium formulation that proved to be efficient, at least in batch culture mode, in promoting high density culture of H. pluvialis. NPK and macrophyte (E. crassipes) medium seems to be an adequate alternative to replace the conventional one (WC).

Resumo O presente estudo investigou o efeito de dois meios alternativos, NPK (20-5-20) e NPK mais macrófita (M+NPK), em relação ao meio comercial (WC) para avaliar a taxa de crescimento e parâmetros fisiológicos em cultivo estático (2-L), a fim de verificar se o fertilizante (NPK) e a macrófita (Eichhornia crassipes) podem ser utilizados no cultivo de Haematococcus pluvialis em laboratório. O maior número de células de H. pluvialis foi observado no meio NPK (5,4 × 105 células.mL–1) no vigésimo oitavo dia, e nos meios M+NPK e WC foi de 4,1 × 105 células.mL–1 e 2,1 × 105.celulas.mL–1 no vigésimo sexto dia, respectivamente. Os teores de clorofila-a foram significativamente maiores (p<0,05) em meio NPK (41-102 g.L–1) quando comparado aos meios WC e M+NPK (14-61 g.L–1). O teor de astaxantina foi menor que 0,04 mg.L–1. O custo de produção de 10-L de H. pluvialis foi baixo em todos os meios sendo que os meios NPK e M+NPK apresentaram uma redução de custos de 65% e 82%, respectivamente, quando comparados ao meio comercial. O meio contendo fertilizante e o de macrófita (E. crassipes) obtiveram resultados eficientes em cultivo estático, com alta densidade celular de H. pluvialis. O meio NPK e o de macrófita (E. crassipes) demonstraram ser uma alternativa adequada para substituir o meio comercial (WC).

Toxicity effects of nickel electroplating effluents treated by photoelectrooxidation in the industries of the Sinos River Basin / Avaliação do efeito tóxico de efluentes oriundos da indústria galvânica na região da Bacia do Rio dos Sinos após o tratamento pelo processo de fotoeletrooxidação

The Sinos river Basin is an industrial region with many tanneries and electroplating plants in southern Brazil. The wastewater generated by electroplating contains high loads of salts and metals that have to be treated before discharge. After conventional treatment, this study applied an advanced oxidative process to degrade organic additives in the electroplating bright nickel baths effluent. Synthetic rinsing water was submitted to physical-chemical coagulation for nickel removal. The sample was submitted to ecotoxicity tests, and the effluent was treated by photoelectrooxidation (PEO). The effects of current density and treatment time were evaluated. The concentration of total organic carbon (TOC) was 38% lower. The toxicity tests of the effluent treated using PEO revealed that the organic additives were partially degraded and the concentration that is toxic for test organisms was reduced.

A Bacia do Rio dos Sinos é uma região do Sul do Brasil que sedia um importante pólo da indústria coureiro-calçadista incluindo, desde o beneficiamento das peles, em curtumes, à fabricação dos acessórios, em indústrias metalúrgicas e de galvanoplastia. O efluente gerado por processos galvânicos contém carga elevada de sais e metais que devem ser tratados antes do descarte. Neste artigo foi avaliada a aplicação de um processo oxidativo avançado após o tratamento convencional do efluente, visando degradar os aditivos orgânicos presentes nos efluentes dos banhos de eletrodeposição de níquel brilhante. Um efluente sintético foi tratado pelo processo físico-químico de coagulação para a remoção do níquel. A amostra foi, então, submetida a testes de ecotoxicidade e, em seguida, o efluente foi tratado pelo processo de fotoeletrooxidação (FEO). A influência da densidade de corrente e o tempo de tratamento foram avaliados. Foi verificada uma redução de carbono orgânico total superior a 38%. Os ensaios de ecotoxicidade para o efluente tratado por fotoeletroxidação indicaram que a FEO degradou parcialmente os aditivos orgânicos e reduziu a concentração que provoca efeitos tóxicos para os organismos teste.

Antibacterial, cytotoxicity and anticoagulant activities from Hypnea esperi and Caulerpa prolifera marine algae

Extracts from 2 algal species [Hypnea esperi and Caulerpa prolifera] from Suez Canal region, Egypt were screened for the production of antibacterial compounds against some pathogenic bacteria. The bacteria tested included Escherichia coli, Pseudomonas aeruginosa, Salmonella typhimurium, Aeromonas hydrophila, Bacillus subtilis and Staphylococcus aureus. Algal species displayed antibacterial activity. The methanolic extracts showed variable response by producing various zones of inhibition against studied bacteria. The tested Gram-negative bacteria were less affected by studied algal extracts than Gram-positive bacteria. We determined some biopotentials properties such as cytotoxicity and anticoagulant activity of most potent algal active extracts. The secondary metabolites of only Hypnea esperi algal extract effectively prevented the blood clotting to the extent of 120 seconds. Minimum inhibitory concentration [MIC] indicated that all potent tested algal extract C inhibits Bacillus subtilis and Staphylococcus aureus. Minimum bactericidal concentration [MBC] was between 1 and 1.4mg/ml. The algal isolates from Egypt have been found showing promising results against infectious bacteria instead of some synthetic antibiotics

Growth and development time of subtropical Cladocera Diaphanosoma birgei Korinek, 1981 fed with different microalgal diets / Crescimento e tempo de desenvolvimento de uma espécie de Cladocera subtropical Diaphanosoma birgei Korinek, 1981 alimentada com dietas de microalgas.

The aim of this work was to investigate the growth performance of Diaphanosoma birgei fed with two Chlorophyceae algae, Ankistrodesmus gracilis and Haematococcus pluvialis using monoalgal diets and simpler mixed diets. D. birgei was daily fed on four treatments: 1) 100% Ankistrodesmus gracilis (Ag); 2) 100% Haematoccocus pluvialis (Hp); 3) 25% A. gracilis + 75% H. pluvialis (Ag-25+Hp-75) and 4) 75% A. gracilis + 25% H. pluvialis (Ag-75+Hp-25). The fecundity curve of D. birgei showed that the mixed feed Ag-25+Hp-75 and temperature 24±2°C triggered fast fecundity at approximately two days. The fecundity was low when based only on H. pluvialis (Hp), albeit with greater longevity (19 days) and a higher number of broods (8). D. birgei fed on Ag and Ag-75+Hp-25 diets in this experiment sustained higher growth rate and higher lipid content in these treatments. The present study showed that A. gracilis diet and mixed microalgae diets tested were able to support the egg production and development of D. birgei.

O objetivo deste estudo foi avaliar o crescimento de uma espécie de Cladocera Diaphanosoma birgei alimentada com dois tipos de microalgas Chlorophyceae, Ankistrodesmus gracilis e Haematococcus pluvialis cultivadas em monocultura e cultura mista. Quatro dietas foram utilizadas: 1) 100% Ankistrodesmus gracilis (Ag); 2) 100% Haematoccocus pluvialis (Hp); 3) cultura mista com 25% A. gracilis + 75% H. pluvialis (Ag-25+Hp-75) e 4) cultura mista com 75% A. gracilis + 25% H. pluvialis (Ag-75+Hp-25). A curva de fecundidade de D. birgei mostrou que a dieta mista Ag-25+Hp-75 e temperatura de 24±2°C promoveu rápida fecundidade em aproximadamente dois dias. A fecundidade foi baixa quando D. birgei foi alimentada somente com H. pluvialis (Hp), porém foi observada maior longevidade (19 dias) e consequentemente, maior número de descendentes (8). As dietas Ag and Ag-75+Hp-25 apresentaram efeito direto na taxa de crescimento de D. birgei com elevados teores de lipídios nestes tratamentos. O presente estudo mostrou que as dietas contendo somente A. gracilis e as dietas mistas de microalgas foram capazes de manter níveis adequados na produção de ovos e no desenvolvimento de D. birgei.

Structural features and assessment of zymosan-induced arthritis in rat temporomandibular joint model using sulfated polysaccharide / Características estruturais e ensaio de artrite induzida por zimosam em modelo de articulação temporomandibular de ratos usando polissacarídeo sulfatado

The green seaweed Caulerpa cupressoides var. lycopodium contains three SPs fractions (Cc-SP1, Cc-SP2 and Cc-SP3). Cc-SP1 and Cc-SP2 had anticoagulant (in vitro), pro- and antithrombotic, antinociceptive and/or anti-inflammatory (in vivo ) effects. This study analyzed structural features and the antinociceptive and anti-inflammatory effects of Cc-SP1 on zymosan-induced acute arthritis of the rat temporomandibular joint (TMJ). Cc -SP1 was investigated by infrared technique. Male Wistar rats (200-240 g) received subcutaneously (s.c.) Cc-SP1 1h prior to intra-articular (i.art.) injection of zymosan (2 mg joint -1) or saline (0.9%) into the left TMJ. Mechanical hypernociception was measured by the electronic Von Frey method in the basal and 4h after zymosan injection. Animals were euthanized 6h after zymosan injection and the TMJ cavity was removed for total leukocyte counts from the synovial fluid and myeloperoxidase (MPO) activity assessment. Cc-SP1 (1, 3 or 9 mg kg-1) containing sulfate ester, galactose-6-sulfate, uronic acid and glycosidic linkages reduced zymosan-induced hypernociception (78.12, 81.13 and 87.43%, respectively, p < 0.01), and inhibited the total leukocyte influx (85, 88.14 and 89.95%, respectively, p < 0.01), being confirmed by MPO activity (p < 0.05). Therefore, Cc-SP1 reveals a pharmacological tool for treating inflammatory arthropathies.

A alga marinha verde Caulerpa cupressoides var. lycopodium contém três frações de PSs (Cc-PS1; Cc-PS2 e Cc-PS3). Cc-PS1 e Cc-PS2 apresentaram efeitos anticoagulante (in vitro), pró- e antitrombótico, antinociceptivo e/ou anti-inflammatório (in vivo). Analisaram-se as características estruturais e os efeitos antinociceptivo e anti-inflamatório de Cc-PS1 sobre artrite aguda induzida por zimosam na articulação temporomandibular (ATM) de ratos. Foi investigada a Cc-PS1 por técnica de infravermelho. Ratos machos Wistar (200-240 g) receberam subcutaneamente (s.c.) Cc-PS1 1h antes de injeção intra-articular (i.art.) de zimosam (2 mg articulação-1) ou salina (0,9%) na ATM esquerda. A hipernocicepção mecânica foi mensurada por método Von Frey elétrico em zero e 4h após injeção de zimosam. Os animais foram entanasiados após 6h de injeção de zimosam e a cavidade da ATM foi removida para contagem de leucócitos totais do fluído sinovial e ensaio da atividade de mieloperoxidase (MPO). A Cc-PS1 (1; 3 ou 9 mg kg-1) contendo éster sulfato, galactose-6-sulfato, ácido urônico e ligações glicosídicas reduziu a hipernocicepção induzida por zimosam (78,12; 81,13 e 87,4%, respectivamente; p < 0,01), além de inibir o influxo de leucócitos totais (85; 88,14 e 89,95%, respectivamente; p < 0,01), sendo, ainda, confirmado pela atividade de MPO (p < 0,05). Portanto, a Cc-PS1 revela como uma ferramenta farmacológica para tratar de artropatias inflamatórias.

Algas marinas bentónicas de la Isla Gorgona, costa pacífica colombiana / Benthic marine algae of Gorgona island, Colombian Pacific coast

The knowledge of the biodiversity of Gorgona, a continental island in Colombia, is very limited in the case of algae. We present an updated list of the benthic marine algae of Gorgona Island, associated with different marine environments and type of substrates, such as coral reefs, rocks, and soft bottoms. Field samplings were taken between October 2010 and June 2011. We estimated algal cover, species composition and algal diversity for each reef environment using 0.25 m² quadrats. Ecological and environmental aspects are also included. A total of 43 species of algae for the three types of habitats are documented: 24 Rhodophyta, 12 Clorophyta, six Ochrophyta and one species of Cyanobacteria, and 55% of the algal composition corresponded to red algae. Rev. Biol. Trop. 62 (Suppl. 1): 27-41. Epub 2014 February 01.

Se presenta el listado de las algas marinas bénticas del Parque Nacional Natural Isla Gorgona, el inventario corresponde a la flora algal asociada a los diferentes ambientes marinos de la isla, a saber: arrecifes coralinos y fondos blandos. El monitoreo de las comunidades de algas se llevó a cabo entre octubre de 2010 y mayo de 2011, se realizó una estimación de abundancia y cobertura para lo cual se establecieron transeptos paralelos a la línea de costa y se evaluó el porcentaje de cobertura en cuadrantes de 0.25m². Se reportó un total de 43 especies para los dos ambientes estudiados (coralino y fondos blandos), 24 Rhodophyta, 12 Clorophyta, 6 Ochrophyta y una Cyanobacteria. El grupo más representativo fue el perteneciente a la clase Rhodophyceae, que correspondió al 55.8% de la flora inventariada. Los resultados de nuevos registros en la isla ratifican su importancia fitogeográfica, por cuanto la composición florística insular contribuye significativamente a la biodiversidad de la costa pacífica colombiana, destacándose la presencia de arrecifes coralinos como los principales ambientes marinos para el desarrollo de la flora algal de la región.

Effects of some parameters in upscale culture of Haematococcus pluvialis Flotow / Efeito de alguns parâmetros no cultivo em larga escala de Haematococcus pluvialis Flotow

Growth rate and medium parameters between two bench scale volumes (13-L and 250-L) were compared. Experiments were maintained batch mode and culture parameters were periodically measured during a 13-day period. Culture growth during the cultivation of algae Haematococcus pluvialis was determined qualitatively by cell counting, optical density, dry weight, ash content, amount of chlorophyll-a, total organic carbon content and by direct measuring of medium nutrients and some abiotic aspects. Vegetative cell growth was higher when cultured in 13-L with 1.33 x 105 cells.mL−1 on the 12th day than when cultured in 250-L. Significant difference (p < 0.05) in the biology and water culture of H. pluvialis, with the exception of dry weight, ash, nitrite and ammonia, was reported between the volumes. Data obtained in current study for the upscale culture maintenance of H. pluvialis in laboratory conditions shows that it should be undertaken in a 13-L volume due to a greater time span of cells in a vegetative state, greater cell density, lipids and chlorophyll-a contents. Light was of paramount importance on the direct performance of H. pluvialis on the algal biological conditions.

Foram avaliadas a taxa de crescimento e condições do meio de cultura em volumes de 13-L e 250-L em sistema estático, durante o período de 13 dias para a microalga Haematococcus pluvialis. Foi determinada qualitativamente a contagem de células, densidade, peso seco, teor de cinzas, clorofila-a, teor de carbono orgânico total e avaliação de nutrientes e fatores abióticos do meio de cultura. O crescimento foi mais elevado em volume de 13-L com 1,33 x 105 células.mL−1 no décimo segundo dia, do que em volume de 250-L. Em relação ao meio de cultivo e aspectos biológicos de H. pluvialis, foram observadas diferenças significativas (p < 0,05) entre os dois volumes com exceção do peso seco, cinzas, nitrito e amônia. Para cultivo em larga escale de H. pluvialis é recomendado nestas condições laboratoriais, o volume de 13-L devido ao maior tempo das células em estado vegetativo, maior densidade celular e elevados teores de lipídios e clorofila-a. A luz interferiu diretamente nas condições biológicas de H. pluvialis.

Analysis of mRNA expression profiles of carotenogenesis and astaxanthin production of Haematococcus pluvialis under exogenous 2, 4-epibrassinolide (EBR)

The fresh-water green unicellular alga Haematococcus pluvialis is known to accumulate astaxanthin under stress conditions. In the present study, transcriptional expression of eight genes involved in astaxanthin biosynthesis exposed to EBR (25 and 50 mg/L) was analyzed using qRT-PCR. The results demonstrated that both 25 and 50 mg/L EBR could increase astaxanthin productivity and the eight carotenogenic genes were up-regulated by EBR with different expression profiles. Moreover, EBR25 induction had a greater influence on the transcriptional expression of ipi-1, ipi-2, crtR-B, lyc and crtO (> 5- fold up-regulation) than on psy, pds, bkt; EBR50 treatment had a greater effect on the transcriptional expression of ipi-2, pds, lyc, crtR-B, bkt and crtO than on ipi-1 and psy. Furthermore, astaxanthin biosynthesis under EBR was up-regulated mainly by ipi1־ and psy at the post-transcriptional level, pds, lyc, crtR-B, bkt and crtO at the transcriptional level and ipi-2 at both levels.

Adaptability of oleaginous microalgae Chlorococcum alkaliphilus MC-1 cultivated with flue gas / 生物工程学报

In order to lower the cost of lipid production of microalgae and reduce greenhouse gas emissions, microalgae Chlorococcum alkaliphilus MC-1 with the characteristics of rapid pH drift and high pH adaptability, was cultivated with bubbling of flue gas. The experiment was first performed in the photobioreactor (15 L) in three groups (control group, CO2 group and flue gas group), then, in the open raceway pond (24 m2). The adaptability of microalgae MC-1 to the cultivation with flue gas was studied. The results showed that the maximum biomass concentration, growth rate, total lipid content and CO2 fixation rate were (1.02+/-0.07) g/L, (0.12+/-0.02) g/(L.d), (37.84+/-0.58)% and (0.20+/-0.02) g/(L.d) in the photobioreactor treated with flue gas, 36%, 33.33%, 15.34% and 33.33% higher than those of the CO2 group, respectively. In the open raceway pond with aeration of flue gas, the maximum biomass concentration, growth rate, total lipid content and CO2 fixation rate were 147.40 g/m2, 14.73 g/(m2.d), 35.72% and 24.01 g/(m2.d), respectively, which were similar to the cultivation with pure CO2. The toxic heavy metal contents (Pb, As, Cd and Cr) in the biomass of MC-1 treated with flue gas were all below the legal limits. Additionally, the absorptive effect of CO2, NO and SO2 were determined. In the photobioreactor and open raceway pond, the average absorption ratios of these gases were all higher than previous studies. Therefore, our study showed that MC-1 can adapt to the cultivation with flue gas, and it is feasible to enlarge the outdoor cultivation of MC-1 for lipid production coupling with emissions reduction of flue gas.