Marine planktonic and benthic organisms: an ocean of diversity in the collections of the State of São Paulo / Organismos planctônicos e bentônicos marinhos: um oceano de diversidade nas coleções do Estado de São Paulo

Abstract Brazil is characterized as a megadiverse country, and one of the factors that guarantees the knowledge and preservation of this biodiversity is an improvement in the biological collections. They represent a country's biological, scientific, cultural, and genetic heritage and their preservation depends on a joint effort of researchers, institutions, and public authorities. Marine biological collections of invertebrates and algae of the state of São Paulo represent a high percentage of the national collections, with five biological collections deposited in state institutions. Currently, these collections safeguard the vast majority of the state's marine collection and make up one of the largest in the country. Therefore, the objective of this research is to present information on marine biological collections in the state of São Paulo, their current status, funding, and future perspectives, creating communication opportunities and considering the factors that impact their development. Support from the São Paulo Research Foundation (FAPESP) surpasses state limits, since many studies address broader areas and/or establish diverse partnerships with other institutions from outside São Paulo. This is reflected in the taxonomic and geographic scope of the biological collections in the state, since these host specimens/species from different regions of the country, or even from other countries. However, it is worth mentioning that it is not enough to invest in expeditions to enrich the collections without valuing the training of qualified personnel, both for the collections maintenance and for their taxonomic refinement.

Resumo O Brasil é caracterizado como um país megadiverso e uma das prioridades no conhecimento e conservação dessa biodiversidade é seguramente a melhoria das coleções biológicas. Estas representam patrimônio biológico, científico, cultural e genético de um país, e sua preservação depende de um esforço coletivo de pesquisadores, instituições e do poder público. Com relação às coleções biológicas marinhas de invertebrados e algas, o estado de São Paulo representa uma alta porcentagem do acervo nacional, com cinco principais coleções depositadas em instituições estaduais. Atualmente estas coleções salvaguardam a grande maioria do acervo marinho estadual e compõem um dos maiores do país. Assim sendo, o objetivo desta pesquisa é o de apresentar informações sobre as coleções biológicas marinhas do estado de São Paulo, estado atual, financiamentos e perspectivas futuras, criando oportunidades de comunicação e considerando os fatores que impactam o desenvolvimento das mesmas. O apoio financeiro da Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) às instituições paulistas extravasa os limites do estado, visto que muitas pesquisas abordam áreas mais abrangentes e/ou estabelecem parcerias diversas com outras instituições não paulistas. Isso se reflete inclusive na abrangência taxonômica e geográfica das coleções biológicas do Estado, uma vez que estas mantêm espécimes/espécies de diferentes regiões do país, ou mesmo de outros países. No entanto, vale ressaltar que não basta investir em expedições de coleta para enriquecimento das coleções sem valorizar a formação de pessoal qualificado, tanto para a curadoria dos acervos quanto para o refinamento taxonômico.

Environmental Stress Tolerance and Antioxidant Response of Palisada perforata (Rhodophyta) from a Tropical Reef1.

In this study, we analyzed the antioxidant activity and total phenolic content of the intertidal seaweed Palisada perforata collected from different reef microhabitats (sheltered site, tide pool, plateau, and exposed site) along the coast of Pernambuco (Brazil). Both parameters were compared with the same parameters of this species grown in the laboratory under two experiments simulating temperature, salinity, and desiccation conditions found in the reef. After both experiments (temperature x salinity and desiccation), the algal photosynthetic performance was measured through chlorophyll fluorescence parameters using a pulse-amplitude modulation fluorometer to test their stress response. Palisada perforata likely underwent stress by desiccation due to tidal fluctuations rather than to temperature or salinity changes. This conclusion agrees with our observations of the plateau site´s specimens, which were exposed to both air and UV radiation during low tides and exhibited higher antioxidant activity to avoid oxidative damage. However, despite the environmental stress, the antioxidant activity remained low, suggesting that photoinhibition is a crucial protection mechanism against oxidative damage.

Chemical structure and snake antivenom properties of sulfated agarans obtained from Laurencia dendroidea (Ceramiales, Rhodophyta).

Aqueous and KCl-soluble polysaccharides were extracted from Laurencia dendroidea (Rhodomelaceae, Ceramiales) and their chemical profile was accessed by anion-exchange chromatography, chemical and spectroscopic analyses. The homogeneous agaran DHS-4 (181.3 × 103 g. mol-1, 21.3% of NaSO3) presents A units mostly 2-sulfated (18.9 mol%), nonsubstituted (15.3 mol%) and 6-O-methylated (10.1 mol%), while B units are l-sugars composed predominantly by galactose 6-sulfate precursor units (19.2 mol%) and 3,6-anhydrogalactose (13.8 mol%), besides non-precursor galactose 6-sulfate units bearing d-xylose substituents on C-3 (8.1 mol%). The crude KCl-soluble DHS agaran (20.5% of NaSO3) inhibited proteolysis and hemolysis induced by Lachesis muta and Bothrops jararaca venoms. DHS was able to inhibit up to 75% the L. muta venom hemorrhagic effect and to reduce the lethality displayed by B. jararaca venom, increasing the mice survival time up to 3 times. Therefore, this agaran has high potential to be used as an additional tool to treat snakebite envenomation.

Biohydroxylation of (-)-ambrox®, (-)-sclareol, and (+)-sclareolide by whole cells of Brazilian marine-derived fungi.

A screening was performed using nine marine-derived fungi as biocatalysts and the natural products (-)-ambrox® (1), (-)-sclareol (2), and (+)-sclareolide (3) in order to select the microorganisms able to catalyze the biooxidation of these compounds. It was observed that only the Aspergillus sydowii CBMAI 934, Botryosphaeria sp., Eutypella sp., and Xylaria sp. presented active oxidoreductases and catalyzed the regioselective hydroxylation in the natural products. The hydroxylated metabolites obtained were 1ß-hydroxy-ambrox (1a) (14%, A. sydowii CBMAI 934); 3ß-hydroxy-ambrox (1b) (17%, Botryosphaeria sp.; 11%, Eutypella sp.); 3ß-hydroxy-sclareol (2a) (31%, Xylaria sp.; 69%, Botryosphaeria sp.; 55%, Eutypella sp.); 18-hydroxy-sclareol (2b) (10%, Xylaria sp.); and 3ß-hydroxy-sclareolide (3a) (34%, Botryosphaeria sp.; 7%, Eutypella sp.). This is the first report of biohydroxylation of (-)-ambrox® (1), (-)-sclareol (2), and (+)-sclareolide (3) by whole mycelia of marine-derived fungi.

Chemical diversity and antileishmanial activity of crude extracts of Laurencia complex (Ceramiales, Rhodophyta) from Brazil

Chemical profiles of extracts of four species from Laurencia complex (Ceramiales, Rhodophyta) from different populations collected along Southeast Brazilian coast were assessed by High Performance Liquid Chromatography coupled with a Diode Array Detector in order to observe geographic chemical variability. Aiming to evaluate the impact of chemical diversity on potential pharmaceutical uses, the extracts were tested against the promastigote form of Leishmania amazonensis. The most active extracts were submitted to anti-amastigote and cytotoxicity assays. Principal Component Analysis of the chromatograms resulted in four major groups of chemical profiles according to the presence of leishmanicidal chamigranes (-)-elatol and obtusol. The existence of chemotypes, displaying variable pharmacological action, is proposed for the differences observed in L. dendroidea samples. Although all extracts were found active against promastigote form of L. amazonensis, their efficacy was remarkably different and not related to the variation of (-)-elatol and obtusol, which indicates the presence of additional compounds with antileishmanial activity. Moreover, the active extracts also displayed anti-amastigote activity and none of them were considered cytotoxic. The results highlight that the knowledge of chemical geographic variability can be valuable in the search of new antileishmanial compounds from marine sources.

Characterization of volatile composition of Laurencia dendroidea by gas chromatography-mass spectrometry

In this study we report the characterization of the volatile compounds of Laurencia dendroidea. Solvent extracts (dichloromethane and methanol), hydrodistillation extracts and headspace solid-phase microextraction samples were obtained and analyzed by GC-MS. Forty-six volatile components were identified in L. dendroidea, among them hydrocarbons, alcohols, phenols, aldehydes, ketones, acids, esters and terpenes.

Comparative analysis of the corps en cerise in several species of Laurencia (Ceramiales, Rhodophyta) from the Atlantic Ocean

Different species of Laurencia have proven to be a rich source of natural products yielding interesting bioactive halogenated secondary metabolites, such as terpenoids and acetogenins. It is shown that such compounds are accumulated in the spherical, reniform to claviform refractive inclusions called corps en cerise (CC), which are intensively osmiophilic and located mainly in the cortical cells of the thalli and also in trichoblast cells. Up to now, it was believed that CC were present only in these two kinds of cells. Recently, however, a species of Laurencia, L. marilzae, with CC in all cells of the thallus, i.e., cortical, medullary, including the pericentral and axial cells, as well as in the trichoblasts, was described from the Canary Islands, and subsequently also reported to Brazil and Mexico. Within the Laurencia complex, only Laurencia species produce CC. Since the species of Laurencia are targets of interest for the prospection of bioactive substances due to their potential antibacterial, antifungal, anticholinesterasic, antileishmanial, cytotoxic, and antioxidant activities, the present paper carries out a comparative analysis of the corps en cerise in several species of Laurencia from the Atlantic Ocean to obtain basic information that can support natural product bioprospection projects. Our results show that the number and size of the CC are constant within a species, independent of the geographical distribution, corroborating their use for taxonomical purposes to differentiate groups of species that present a lower number from those that have a higher number. In this regard, there was a tendency for the number of CC to be higher in some species of Laurencia from the Canary Islands. The presence of CC can also be used to distinguish species in which these organelles are present in all cells of the thallus from those in which CC are restricted to the cortical cells. Among the species analyzed, L. viridis displayed the most varied secondary metabolites composition, such as sesquiterpenes, diterpenes, triterpenes, all of which showed potent antiviral, cytotoxic, and antitumoral activities, including protein phosphatase type 2A (PP2A) inhibitory effects.

Morphological and molecular studies on the Brazilian native red seaweed Laurencia oliveirana (Rhodomelaceae, Ceramiales)

Morphological and molecular studies were carried out on Laurencia oliveirana from the type locality (Arraial do Cabo, Rio de Janeiro, Brazil). This species is easily recognized by its small size, sub-erect habit forming intricate cushion-like tufts and unilateral pectinate branching. The species displays all the typical characters of the genus Laurencia, such as the production of the first pericentral cell underneath the basal cell of the trichoblast, tetrasporangia produced from particular pericentral cells, with the third and fourth pericentral cells becoming fertile, without production of additional pericentral cells, spermatangial branches produced from one of two laterals on the suprabasal cell of trichoblasts, and procarp-bearing segment with five pericentral cells. Details of tetrasporangial plants and development of procarp and male plants are described for the first time for the species. The phylogenetic position of L. oliveirana was inferred by analysis of the chloroplast-encoded rbcL gene sequences from 57 taxa. In all phylogenetic analyses, L. oliveirana grouped with L. caraibica, L. caduciramulosa, L. venusta and L. natalensis, forming a monophyletic clade within the Laurencia sensu stricto. The genetic divergence between L. oliveirana and the molecularly closest species, L. caraiba collected in Brazil, was 2.3%.

Influence of tourist activity on the diversity of seaweed from reefs in Maracajaú, Atlantic Ocean, Northeast Brazil

The worldwide increase in recreational activities and marine tourism is reason for concern due to its impact on reef environments. In order to assess the effect of tourism on the reef area in Maracajaú (Northeast Brazil), a study was conducted based on the presence of different seaweed species. A region of intense tourist activity was chosen, and another where tourism is prohibited. Result comparison used richness (S), biomass, diversity (H), and dominance (D), as well as an analysis of similarity between samples. Both areas exhibited differences in specific composition, biomass, diversity, richness and seaweed dominance. The highest values for biomass, richness and diversity were recorded in the tourism-free region, while the greatest dominance rates were found in the area of intense tourist activity. The latter was characterized by the dominant presence of Caulerpa racemosa (Forsskål) J. Agardh (Chlorophyta) and turf algae. These characteristics are a clear indication that the reef area subject to heavy tourist activity is undergoing degradation.

Effects of temperature, salinity and irradiance on carposporeling development of Hidropuntia caudata (Gracilariales, Rhodophyta)

The success of seaweed cultivation depends on the scientific control of the tolerance limits and the optimal physiological conditions that affect the spore germination and the early development of algal species. In order to establish cultivation techniques for spores of Hidropuntia caudata (J. Agardh) Gurgel & Fredericq, the effects of irradiance, salinity, and temperature on the carpospore germination and carposporeling development were evaluated under laboratory conditions. Five photon flux densities (PFD, from 18 to 200 µmol photons m-2s-1), six salinity values (from 7 to 55 psu), and four temperatures (from 20 ºC to 35 ºC) were investigated. The level of irradiance caused significant differences in the growth, in the following order: 200±5 > 100±5 ~= 62.5±2.5 > 30±1.5 > 18±1 µmol of photons m-2s-1, but they did not inhibit the carposporeling development. Maximum growth occurred under 35 psu, while at 15 psu the formation of carposporeling erect axis was limited. The optimal temperature for growth was 25 ºC, while at 35 ºC the spores died. These results show the importance of previous knowledge on the tolerance limits and optimal conditions for sporeling development of H. caudata for the implementation of an aquaculture program.

Chemical structure of the complex pyruvylated and sulfated agaran from the red seaweed Palisada flagellifera (Ceramiales, Rhodophyta).

A homogeneous agaran fraction from Palisada flagellifera (Laurencia complex, Rhodomelaceae, Ceramiales) was obtained by aqueous room-temperature extraction, followed by ion-exchange chromatography. This galactan presents a highly complex structure with at least 18 different types of derivatives. The A units were found mostly pyruvylated, 2-sulfated (∼34%), and 6-methylated (∼34%), with the latter partially 2- and 2,4-sulfated. Minor amounts of ß-D-galactopyranosyl units 2-, 6- and 2,6-sulfated, 6-glycosylated, and non-substituted are also present. The B-units are L-sugars composed predominantly of their cyclized derivatives, 3,6-anhydrogalactose and 3,6-anhydro-2-O-methylgalactose (∼56%). The former are linked to ß-D-galactosyl (6-methyl) (6-glycosylated) units, as well as to 4,6-O-(1-carboxyethylidene)-ß-D-galactose 2-sulfate in the proportion of 3:1.8, respectively. A significant amount (∼18%) of the α-L-galactopyranosyl units are linked to pyruvylated ß-D-galactose 2-sulfate residues. An important part of the B-units (20%) is represented by α-L-galactose 6-sulfate substituted on C-3 by xylosyl, galactosyl and/or 2,3-di-O-methylgalactose units or sulfate groups that preclude their cyclization to 3,6-anhydrogalactosyl derivative. The precursor units are present in relatively low percentages. Kinetic studies suggest that in P. flagellifera agaran the cyclizable units are linked to 6-O-methyl-ß-D-galactosyl and/or ß-D-galactosyl units (6-glycosylated). The structural complexity of this polysaccharide is increased by the presence of 2- and 3,6-sulfated α-L-galactoses, with the latter additionally 2-O-methylated. Therefore, the major subfraction obtained from the cold extract contains structurally complex sulfated, methylated, and pyruvylated agaran.

Biochemical composition of two red seaweed species grown on the Brazilian coast.

BACKGROUND: Algae species have been used as an important source of food because they are highly nutritive considering their vitamin, protein, mineral, fiber, essential fatty acid and carbohydrate contents. However, a large number of seaweeds have been poorly studied, especially Brazilian species. Two red macroalgae species from the Brazilian coast (Plocamium brasiliense and Ochtodes secundiramea) were assessed with respect to their total lipid, fatty acid, total nitrogen, protein, amino acid and total carbohydrate contents. RESULTS: The total lipid contents (dry weight) were 36.3 and 35.4 g kg(-1); fatty acid contents were 9.3 and 12.1 g kg(-1); total nitrogen contents were 37.4 and 24.9 g kg(-1); protein contents were 157.2 and 101.0 g kg(-1); amino acid contents were 127.5 and 91.4 g kg(-1); and total carbohydrate contents were 520.3 and 450.7 g kg(-1) for P. brasiliense and O. secundiramea, respectively. CONCLUSION: Considering these compositions, both algae species were determined to have sources of protein, essential amino acids and carbohydrates similar to the edible seaweeds Laminaria japonica and Palmaria palmata.

Sesquiterpenes from the essential oil of Laurencia dendroidea (Ceramiales, Rhodophyta): isolation, biological activities and distribution among seaweeds

Two known sesquiterpenes (1R*,2S*,3R*,5S*,8S*,9R*)-2,3,5,9-tetramethyltricyclo[6.3.0.0(1,5)]undecan-2-ol and (1S*,2S*,3S*,5S*,8S*,9S*)-2,3,5,9-tetramethyltricyclo-[6.3.0.0(1,5)]undecan-2-ol were isolated for the first time from the essential oil of the red seaweed Laurencia dendroidea collected in the Brazilian coast. These compounds were not active against eight bacteria strains and the yeast Candida albicans, but showed some antioxidant activity. Both compounds were also found in other seaweed species showing that they are not exclusive taxonomic markers to the genus Laurencia.

Overview of the taxonomy and of the major secondary metabolites and their biological activities related to human health of the Laurencia complex (Ceramiales, Rhodophyta) from Brazil

In Brazil, the Laurencia complex is represented by twenty taxa: Laurencia s.s. with twelve species, Palisada with four species (including Chondrophycus furcatus now that the proposal of its transference to Palisada is in process), and Osmundea and Yuzurua with two species each. The majority of the Brazilian species of the Laurencia complex have been phylogenetically analyzed by 54 rbcL sequences, including five other Rhodomelacean species as outgroups. The analysis showed that the Laurencia complex is monophyletic with high posterior probability value. The complex was separated into five clades, corresponding to the genera: Chondrophycus, Laurencia, Osmundea, Palisada, and Yuzurua. A bibliographical survey of the terpenoids produced by Brazilian species showed that only six species of Laurencia and five of Palisada (including C. furcatcus) have been submitted to chemical analysis with 48 terpenoids (47 sesquiterpenes and one triterpene) isolated. No diterpenes were found. Of the total, 23 sesquiterpenes belong to the bisabolane class and eighteen to the chamigrene type, whose biochemical precursor is bisabolane, two are derived from lauranes and four are triquinols. Despite the considerable number of known terpenes and their ecological and pharmacological importance, few experimental biological studies have been performed. In this review, only bioactivities related to human health were considered.

Search for cytotoxic agents in multiple Laurencia complex seaweed species (Ceramiales, Rhodophyta) harvested from the Atlantic Ocean with emphasis on the Brazilian State of Espírito Santo

The development of new anti-cancer drugs of algal origin represents one of the least explored frontiers in medicinal chemistry. In this regard, the diversity of micro- and macroalgae found in Brazilian coastal waters can be viewed as a largely untapped natural resource. In this report, we describe a comparative study on the cytotoxic properties of extracts obtained from the Laurencia complex: Laurencia aldingensis, L. catarinensis, L. dendroidea, L. intricata, L. translucida, L. sp, and Palisada flagellifera. All of these species were collected in the coastal waters of the State of Espírito Santo, Brazil. Four out of the twelve samples initially investigated were found to show significant levels of toxicity towards a model tumor cell line (human uterine sarcoma, MES-SA). The highest levels of cytotoxicity were typically associated with non-polar (hexane) algal extracts, while the lowest levels of cytotoxicity were found with the corresponding polar (methanol) extracts. In this report, we also describe a biological model currently in development that will not only facilitate the search for new anti-cancer drug candidates of algal origin, but also permit the identification of compounds capable of inducing the destruction of multi-drug resistant tumors with greater efficiency than the pharmaceuticals currently in clinical use.

Screening for antifungal activities of extracts of the brazilian seaweed genus Laurencia (Ceramiales, Rhodophyta)

The resistance of pathogens to commonly used antibiotics has enhanced morbidity and mortality and has triggered the search for new drugs. Several species of the red alga genus Laurencia are very interesting candidates as potential sources of natural products with pharmaceutical activity because they are known to produce a wide range of chemically interesting halogenated secondary metabolites. This is an initial report of the antifungal activities of the secondary metabolites of five species of Laurencia, collected in the state of Espírito Santo, against three strains of pathogenic fungi: Candida albicans (CA), Candida parapsilosis (CP), and Cryptococcus neoformans (CN). Minimum inhibitory concentrations (MIC) of the algal extracts were determined by serial dilution method in RPMI 1640 Medium in 96-well plates according to the NCCLS and microbial growth was determined by absorbance at 492nm. A result showing maintenance or reduction of the inoculum was defined as fungistatic, while fungicidal action was no observed growth in the 10 µL fungistatic samples subcultured in Sabouraud Agar. Our results indicate that apolar extracts of Laurencia species possess antifungal properties and encourage continued research to find new drugs for therapy of infectious diseases in these algae.

Antileishmanial sesquiterpenes from the Brazilian red alga Laurencia dendroidea.

Investigation of the bioactive crude extracts from two populations of the red alga Laurencia dendroidea from the southeastern Brazilian coast led to the identification of five sesquiterpenes: (+)-obtusane (1), a triquinane derivative (2), (-)-elatol (3), obtusol (4), and cartilagineol (5). An antileishmanial bioassay against Leishmania amazonensis was conducted for crude lipophilic extracts and for sesquiterpenes 2, 3, and 4. Compounds 3 and 4 displayed in vitro and in vivo leishmanicidal activity and very low cytotoxicity.

Induction of halogenated vesicle transport in cells of the red seaweed Laurencia obtusa.

In clones of the red alga Laurencia obtusa, the frequency of vesicle transport from corps en cerise (CC) to the cell wall region was evaluated in response to differences in temperature, irradiance, desiccation, bacterial fouling, and bromine (Br) availability. In addition, the morphology of the corps en cerise was analyzed. Traffic of vesicles was induced by exposing L. obtusa to low temperatures and variations in irradiance. It was also verified that bacterial fouling induced vesicle traffic. Under high temperatures and desiccation, the membranous tubular connections were lost and transport of vesicles was not seen. The morphology of the corps en cerise varied according to the availability of Br in seawater. Exocytosis of secondary metabolites by L. obtusa was shown to vary in relation to temperature, irradiance, desiccation and bacterial fouling. The data suggest that the transport of vesicles in L. obtusa may be related to the inhibition of the microfouling community on the algal surface.

Sulfated xylomannans isolated from red seaweeds Chondrophycus papillosus and C. flagelliferus (Ceramiales) from Brazil.

Sulfated xylomannans were isolated from two species of genus Chondrophycus by aqueous extraction followed by KCl fractionation. Structural determination of the native, desulfated and Smith-degraded KCl-precipitated polysaccharides carried out by composition and methylation analysis and NMR spectroscopy (1D and 2D experiments) showed the following general structure: [see text] These xylomannans present different degrees of branching (15-25%) by beta-D-Xylp (70-80%) and beta-D-Manp-2-S (20-30%) and molecular weights (33-222kDa). This is the first report of the presence of a sulfated xylomannan in species of order Ceramiales.

Bisabolanes from the red alga Laurencia scoparia.

Three novel halogenated beta-bisabolene sesquiterpenoids (1-3), together with two know triquinane alcohol sesquiterpenes (6 and 7), were isolated from the red alga Laurencia scoparia and their structures elucidated by spectroscopic methods. Single-crystal X-ray crystallography allowed us to confirm the structure of 1 as well as to determine the absolute configuration of all stereocenters. To the best of our knowledge, the isolation of beta-bisabolenes from the genus Laurencia has no precedent in the literature. Compound 1 showed weak in vitro anthelmintic activity against parasitant stage (L4) Nippostrongilus brasiliensis.