Ester Clavero

Morphological, genetic and mating diversity within the widespread bioindicator Nitzschia palea (Bacillariophyceae)

Trobajo R., Clavero E., Chepurnov V.A., Sabbe K., Mann D.G., Ishihara S. and Cox E.J. 2009. Morphological, genetic and mating diversity within the widespread bioindicator Nitzschia palea (Bacillariophyceae). Phycologia 48: 443–459. DOI: 10.2216/08-69.1. Nitzschia palea (Kützing) W. Smith is believed to be a widely distributed diatom and is common in various lotic and lentic freshwater habitats. However, it is also taxonomically problematic. As part of a multidisciplinary study of this diatom, 25 clones identified morphologically as N. palea were isolated from different freshwater habitats around the world (Belgium, Brazil, Egypt, India, Japan, Paraguay, Spain, Sri Lanka and the United Kingdom). Morphological and genetic diversity (using the hypervariable D1–D2 domains of LSU rDNA) were investigated, and an almost complete set of interclonal crossing experiments was carried out. Results indicate that N. palea is not a simple, homogeneous taxon and that N. palea will probably have to be split into three or more species. Molecular and mating groups do not separate along the traditional morphological boundaries among N. palea varieties, in particular between vars palea and debilis, two taxa that are commonly used to discriminate between different degrees of water pollution. At least two of the putative species within the N. palea complex appear to be geographically widespread. Because of the complexity of variation revealed by the LSU, mating and morphometric data, it is clear that further work, using extra genetic markers and new isolates, will be needed to determine the full extent of cryptic and pseudocryptic speciation in N. palea and to investigate whether the segregate species are ecologically differentiated and have value as indicators.

SALINITY TOLERANCE OF DIATOMS FROM THALASSIC HYPERSALINE ENVIRONMENTS

Thirty‐four benthic diatom strains were isolated from thalassic hypersaline marine environments and their salinity tolerance characterized in growth experiments conducted at salinities ranging from 0.5% to 17.5% (weight of total salts per volume, g·100 mL−1). The results were compared with the patterns of diatom species distribution and abundance in hypersaline evaporation ponds and tidal channels of Guerrero Negro, Baja California Sur, Mexico. The isolated strains were representative of the diatom assemblages present in the saltern ponds but were less so of natural assemblages in tidal channels. In general, we found a clear decreasing trend of diatom diversity in the field and in the isolated strains with increasing salinity. With some exceptions, the upper limit of salinity tolerance in cultivated strains corresponded to their distribution in field samples. However, the relative abundance of species in the field was not correlated with growth rates achieved in culture for the same salinities. Most cultured strains exhibited extreme euryhalinity growing well from brackish to hypersaline conditions, but the particulars of salt tolerance were quite diverse among strains. The most halotolerant taxa, two Amphora species, Amphora cf. subacutiuscula Schoeman, Nitzschia fusiformis Grunow, and Entomoneis sp., grew well in salinities ranging from 0.5% to 15%. Three strains of Pleurosigma strigosum W. Smith that were unable to grow in salinities less than 5% total salts represent the only true halophilic diatoms ever reported. The fact that many strains displayed a remarkable halotolerance, with optimal or near‐optimal growth rates at salinities as high as three times that of seawater, implies that diatoms from hypersaline environments are evolutionarily highly adapted to such environments.

The use of partial cox1, rbcL and LSU rDNA sequences for phylogenetics and species identification within the Nitzschia palea species complex (Bacillariophyceae)

Cox1 has been suggested as a barcode marker for diatoms but it has not been tested intensively in any group of closely related species outside Sellaphora. We evaluated the use of cox1, LSU and rbcL for phylogeny and identification in the taxonomically problematic but ecologically important freshwater diatom Nitzschia palea, for which LSU rDNA sequences, mating and morphological data had already been published, and for which DNA and/or clonal isolates were still available; some new isolates were added. The previous and new information concur in suggesting that N. palea is a complex of several or many species. Where cox1 sequences could be obtained, they were more variable than LSU and rbcL and discriminated between lineages that differed in their morphologies, mating compatibilities, LSU sequences or rbcL sequences. Repeated failures for some strains to recover cox1 sequences from DNA that yielded LSU and rbcL suggest that cox1 will be impractical as a universal barcode marker in diatoms until better primers are designed. LSU and rbcL, though less variable than cox1, can be sequenced reliably in N. palea and, together, seem to show sufficient discrimination to be worth further consideration for species recognition.

Vibrating membrane filtration as improved technology for microalgae dewatering.

The effect of shear-enhanced filtration by vibratory process in microalgae dewatering is presented in this paper. The aim of this research was to investigate the technical performance and improvement of vibrating membrane filtration compared with conventional tangential cross-flow filtration in microalgae concentration. An industrial-scale available commercial set-up was used. Several membrane materials as polyethersulfone, polyacrylonitrile, etc., and mean pore sizes (from 7000Da to 0.2μm) were tested and compared in both filtration set-ups. Experiments were carried-out with Nannochloropsis gaditana and Phaeodactylum tricornutum microalgae. It has been demonstrated that, even if the choice of the membrane depends on its cut-off, its material and the type of microalgae filtrated, dynamic filtration is always the best technology over a conventional one. If with conventional filtration permeability values were in the vicinity of 10L/h/m(2)/bar in steady state phase, with dynamic filtration these values increased to 30L/h/m(2)/bar or more.

Why there is such luxurious growth in the hypogean environments

Organisms building biofilms are of considerable interest in the context of degradation of cultural heritage. Particularly, hypogean environments exposed to artificial light are colonized by microbial communities, which damage walls and frescoes. In order to ascertain the mechanisms by which phototophic biofilms thrive under the particular conditions of hypogea, the organism composition and three-dimensional structure of biofilms from the Roman catacombs St. Callistus and Domitilla were studied. The main phototrophic organisms forming the biofilms were filamentous sheathed cyanobacteria and mosses. Biofilms were spatially very heterogeneous in thickness, density and organism composition but could be classified as regards their main organisms. There was a trend of decreasing diversity in the phototrophic composition of the biofilms under lower irradiances, the one at the lowest irradiance being uniquely built by erected filaments of Leptolynghya sp. Except for this biofilm, the main organism composition was not clearly related with decreasing irradiance. However, biofilms from dim light samples were porous and the filamentous cyanobacteria in them were erected. Leplolyngbya sp., the most ubiquitous species, displayed a high number of phycobilisomes and its hormogonia a gliding movement that allowed colonization of substrata. Such mechanisms may have an important role for thriving under the low light conditions of the catacombs.

Biorefinery concept in a microalgae pilot plant. Culturing, dynamic filtration and steam explosion fractionation.

A 53 m(3) microalgae pilot plant with semi-closed photobioreactors has been operated in batch mode at Tarragona, where proper climatic conditions exist. Operations carried-out were microalgae culturing, concentration, cell disruption and lipid extraction. Culturing was performed with and without CO2 fertilization. pH was used to control CO2 dosing. With CO2 fertilization best microalgae concentrations were obtained. Productivities up to 19.9 g/m(2)/day were reached. Although sedimentation and centrifugation was performed, studies focused on dynamic membrane filtration as improved technique compared with conventional one. Significant fouling reduction was obtained by using a vibrational membrane setup. Cell disruption including product pre-extraction was performed by using steam explosion at moderate conditions, as novel technique for this application. Lipid extraction was performed by using conventional techniques with and without exploded material. The amount of lipids obtained from exploded material was higher than with non-exploded material, which shows that steam explosion provides a clear enhancement.

Potential pre-concentration methods for Nannochloropsis gaditana and a comparative study of pre-concentrated sample properties.

We compared potential pre-concentration techniques for Nannochloropsis gaditana (Nng) by testing natural sedimentation; flocculation with aluminium sulphate, polyaluminium chloride and chitosan; and induced pH. Promising flocculation efficiencies and concentration factors were obtained in a short time with alkalinity-induced flocculation at an adjusted pH of 9.7 and with chitosan at an adjusted pH of 9.9 using a concentration of 30mgL(-1). The sedimentation rates of alkalinity-induced flocculation were also evaluated. Additionally, viscosity, particle size distribution and Ca/Mg ions were analysed for pre-concentrated samples of N. gaditana (Nng) and the previously studied Phaeodactylum tricornutum (Pht) which were obtained by various different harvesting methods under optimal conditions. The rheological properties of the concentrated algae suspensions of two microalgal species showed Newtonian behaviour. The mean diameters of the flocs were between 39 and 48μm. The Ca/Mg analysis showed that Mg(+2) is the triggering ion for alkalinity-induced flocculation in the conditions studied.

THE FINE STRUCTURE OF TWO SMALL AMPHORA SPECIES. A. TENERRIMA ALEEM & HUSTEDT AND A. TENUISSIMA HUSTEDT

The type materials of Amphora tenerrima Aleem & Hustedt and Amphora tenuissima Hustedt have been examined with light and electron microscopy in order to amplify the data on these two small species apparently lacking ventral striae. Both species, though well characterised by light microscopy in their original descriptions, appear problematic in view of information from use of the SEM. A. tenerrima from the type material has biseriate dorsal striae and ventral striae formed by one row of cuneate shallow depressions of the basal siliceous layer, a striae shape that has not been recorded for any species of the subgenus Halamphora Cleve. On the other hand, the conspicuous puncta of the connecting bands allow a differentiation of this species from very similar taxa. A. tenuissima has uniseriate dorsal striae formed by rows of small round puncta. Ventral striae are thin slits arranged in a higher density than dorsal striae. These and other fine structural features combined with morphometrical data make them distinguishable from apparently similar taxa even under the light microscope. The features revealed by SEM for both species complete the original descriptions.

Microalgae fractionation using steam explosion, dynamic and tangential cross-flow membrane filtration

In this study, the microalga Nannochloropsis gaditana was subjected to acid catalysed steam explosion treatment and the resulting exploded material was subsequently fractionated to separate the different fractions (lipids, sugars and solids). Conventional and vibrational membrane setups were used with several polymeric commercial membranes. Two different routes were followed: 1) filtration+lipid solvent extraction and 2) lipid solvent extraction+filtration. Route 1 revealed to be much better since the used membrane for filtration was able to permeate the sugar aqueous phase and retained the fraction containing lipids; after this, an extraction required a much lower amount of solvent and a better recovering yield. Filtration allowed complete lipid rejection. Dynamic filtration improved permeability compared to the tangential cross-flow filtration. Best membrane performance was achieved using a 5000Da membrane with the dynamic system, obtaining a permeability of 6L/h/m2/bar.

Efficient harvesting of Chaetoceros calcitrans for biodiesel production

Harvesting is one of the key challenges to determine the feasibility of producing biodiesel from algae. This paper presents experimental results for a cost-effective system to harvest Chaetoceros calcitrans, using natural sedimentation, flocculation, and inducing pH. No efficient sedimentation of microalgal cells was observed only by gravity. By alkalinity-induced flocculation, at a pH value of 9.51, 86% recovery of the cells was achieved with a sedimentation rate of 125 cm/h and a concentration factor (CF) of 4 (volume/volume (v/v)) in 10 min. The maximum photochemical quantum yield of photosystem II (Fv/Fm) of concentrated cells was almost the same as fresh culture (0.621). Commercial flocculants, aluminium sulphate and poly-aluminium chloride (PAC), were also successful in harvesting the studied algal cells. Optimum concentration of aluminium sulphate (AS) could be concluded as 10 ppm with 87.6% recovery and 7.10 CF (v/v) in 30 min for cost-efficient harvesting, whereas for PAC it was 20 ppm with 74% recovery and 6.6 CF (v/v). Fv/Fm yields of concentrated cells with AS and PAC showed a 1% reduction compared to fresh culture. Mg+2 was the triggering ion for alkalinity-induced flocculation in the conditions studied. The rheology behaviour of the concentrated cells was Newtonian with values between 2.2 × 10−3 and 2.3 × 10−3 Pa s at 30°C.