PHENOTYPIC PLASTICITY IN SCENEDESMUS COMMUNIS (CHLOROPHYCEAE). I. RELATIONSHIP OF S. COMMUNIS TO S. KOMAREKII1

When scenedesmus communis Hegew. (UTEX 76) was transferred daily in dilute media and a low cell density was maintained (ca. 1000 cells · mL^?1), up to 30% unicells were produced in that population. Unlike previously described uncell-coenobium-unicell transformation with other species, these unicells never produced S. communis coenobia (large coenobium type, LCT) but rather small coenobium type (SCT) resembling S. komarekii Hegew. Growth and morphological development of the paratype strain of S. komarekii (UTEX 1236) was compared with an isolated SCT strain (SCT 76–8). SCT 76–8 never produced LCTs and grew significantly faster than UTEX 1236. Both SCT 76–8 and UTEX 1236 produced uncells at low cell densities. Coenobia formed when cell densities increased over time in batch cultures. SCT 76–8 and UTEX 1236 did not differ morphologically when viewed with the light microscope. Under scanning electron microscopy, an outer opaque layer covered an inner warty layer on unicells. The outer layer was reduced or absent in coenobia from batch cultures in stationary growth. In addition, long spikelets, not present on the walls of unicells, were prominent on coenobial walls. The spikelets of UTEX 1236 appeared smaller and more uniformly distributed than in strain 76–8. In contrast, the surface wall morphology of LCT S. communis was composed of an outer reticulate layer supported by spikelets and appeared as a pentagonal meshwork covering the cell walls. This phenotypic plasticity, as demonstrated by SEM and light microscopy, provides further evidence needed for an understanding of Scenedesmus evolution.     

LAGERHEIMIA HINDAKII IS THE UNICELLULAR STAGE OF A SCENEDESMUS1

Extensive variability in spine number and position was emphasized in the protologue for the spiny unicell Lagerheimia hindakii Hegew. et A. Schmidt. Using an axenic clone established from culture Hindk 1975/95, which provided the type material of that organism, we initiated a morphogenetic study. While in the unicellular stage, three short spines were usually formed at each cell pole. A colonial morph was first observed during the initial isolation procedure after unicells were dispersed on firm agar and allowed to grow. We propose that L. hindakii is a Scenedesmus, close to S. subspicatus R. Chod. This organism is unlike most representatives of the genus Scenedesmus because colonies do not readily appear in dilute media; they also fail to form in Bristol's liquid medium unless there is an organic supplement, e.g. glucose. Hindk 1975/95 may be more closely related to truly unicellular taxa than to other Scenedesmus.     

Influence of barley straw (Hordeum vulgare L.) extract on phytoplankton dominated by Scenedesmus species in laboratory conditions: the importance of the extraction duration

The response of a natural phytoplankton assemblage dominated by algae of the genus Scenedesmus to the addition of barley straw extract was studied in a laboratory experiment. The aim of the study was to compare the inhibiting effect of water extracts obtained by soaking the straw for 1, 2 and 3 months. We analysed the response of four species, Scenedesmus subspicatus, Scenedesmus ecornis, Scenedesmus quadricauda and Scenedesmus acuminatus, during 14 days of their exposure to different types of barley straw extract. S. subspicatus and S. ecornis responded with decreasing numbers only to the addition of the 3-month solution (ANOVA; F?=?290.1, p <0.001; and F?=?11.8, p <0.01, respectively); the two other species were inhibited by all types of extracts. The results indicate the need for more research on the importance of extraction duration to the inhibitory abilities of barley straw which can be applied in the management of water quality in water bodies.     

No difference in colony formation of Scenedesmus obliquus exposed to lakes with different nutrient levels

The algal genus Scenedesmus is famous for its highly phenotypic plasticity in response to various environmental factors. In laboratory, axenic cultures of Scenedesmus often fail to form colonies and remain in unicellular morph. To examine whether unicellular Scenedesmus can form colonies after exposure to natural lakes, dialysis bags and plastic bottles which contained the precultured Scenedesmus obliquus were exposed in two lakes with different nutrient levels for four weeks. Results showed S. obliquus grew well in dialysis bags but not in plastic bottles; S. obliquus can form colonies when incubated in dialysis bags which allow exchange with the aquatic environment of the substances, regardless of the nutrient levels of the two lakes. However, no colonies were observed in S. obliquus incubated in plastic bottles exposed in both lakes. This suggested that active growth and zooplankton infochemicals contributed together to the colony formation of S. obliquus exposed in situ environment.     

Physiological,Biochemical, and Molecular Characters for the Taxonomy of the Subgenera of Scenedesmus (Chlorococcales,Chlorophyta)

Biochemical and physiological properties of 16 Scenedesmus species representing the three subgenera Scenedesmus, Acutodesmus, and Desmodesmus are not suitable for species differentiation. All Scenedesmus species studied thus far produce secondary carotenoids, e.g. astaxanthin and canthaxanthin, under nitrogen-deficient conditions. In addition, with the exception of one strain, hydrogenase activity under anaerobic conditions is generally present. Sequence analyses of ribosomal 18S RNAs indicate that the subgenus Desmodesmus is phylogenetically well separated from the other subgenera, whereas the separation of Scenedesmus and Acutodesmus appears less convincing and is dismissed in favour of a single subgenus, Scenedesmus. Three taxa formerly assigned to the genus Chlorella are shown to be unicellular species of the genus Scenedesmus. “Chlorella” fusca var. vacuolata and “C.” fusca var. rubescens, which is closely related to S. obliquus, belong to the Scenedesmus/‘Acutodesmus group. “C.” fusca var. fusca is closely related to S. communis and thus belongs to the subgenus Desmodesmus. Inclusion of Kermatia pupukensis into the genus Scenedesmus is also strongly supported by the RNA data which furthermore indicate a relationship with the subgenus Desmodesmus.     

CRYPTIC SPECIES OF SCENEDESMUS (CHLOROPHYTA) FROM DESERT SOIL COMMUNITIES OF WESTERN NORTH AMERICA1

Nine isolates of unicellular green algae were obtained from six geographically separate desert microbiotic crust communities in western North America. Microscopically, eight isolates strongly resembled unicellular forms of Scenedesmus obliquus (Turpin) Kützing. They are oval or crescent shaped, often flattened on one side, with knobby cell apices. SEM indicated a lack of wall ornamentation. Fine filaments connecting cells pole to pole were observed in some isolates, as previously documented in Scenedesmus (Dactylococcus) dissociatus and S. obliquus. The ninth isolate was spherical, without knobby apices or connections between cells, and was similar to unicellular forms that were originally classified as species of Chlorella (Scenedesmus vacuolatus and S. rubescens). None of the isolates formed coenobia in liquid culture. Phylogenetic analysis of the 18S rRNA gene placed all desert isolates in the genus Scenedesmus, separating them into two or three weakly resolved groups along with published sequences of other Scenedesmus isolates. Phylogenetic analyses of the internal transcribed spacer region revealed well‐supported lineages of desert algae that were unsupported with 18S data alone. The eight S. obliquus‐like desert strains formed two distinct clades that excluded the S. obliquus strains from geographically widespread nondesert habitats. The ninth strain was outside of the S. obliquus group, associated with S. raciborskii and S. pectinatus. These results demonstrate three lineages of Scenedesmus from desert soils and provide robust support for the presence of cryptic species in S. obliquus, a morphospecies that is said to have a cosmopolitan distribution. Three new species of Scenedesmus are described.     

THE ROLE OF UNICELLS IN THE POLYMORPHIC SCENEDESMUS ARMATUS (CHLOROPHYCEAE)1

The UTEX 2193 strain of Scenedesmus armatus (Chod.) Chod, when cultured in any of several media (whether natural or artificial, concentrated or dilute) produced a variety of colonial morphologies as well as a unicell population. Morphological expression was related to culture ape. When the initial cell density was just a feu1 hundred cells per mL. the culture first produced a unicell population, then spiny colonies, and as stationary phase was approached, spine-less colonies. Two classes of spiny colonies were detected. Type I colonies had elongate cells with the terminal cells shorter than median cells. Spines were longer than cell length. The wider, oval, grainy cells of Type II colonies were uniform m length. Spines were shorter and thicker than those on Type I colonies. Only Type I colonies produced unicells: the latter appeared as two morphs. The smaller unicell was obovoid with four delicate spines: the larger had ovate cells bearing four thicker spines. Control of unicell development in all media was achieved by carefully monitoring colony type and cell number used for the inoculum. A unicellular population developed in batch culture in defined media, both concentrated and dilute, when the initial cell density (either Type I or Type II colonies) was low (below 1000 cells-mL^?1), as well as in synchronous cultures. With higher initial cell densities, e.g. 2 × 10⁴ cells·mL^?1, the inoculum had to contain Type I colonies to produce unicells. Unicells were also produced in water from Agronomy Pond, where the strain originated. We discuss the role of unicell populations in the distribution of Scenedesmus.     

SCENEDESMUS WALL ORNAMENTATION. I. SCENEDESMUS PARISIENSIS CULTURES1

Four strains of Scenedesmus parisiensis Chodat were studied in xenic and axenic culture in 3 media as well as in cultures incubated in sterile vessels in nature. Organized coenobia are usually produced but these may have merely short spines, spines and serrate edges, or lack wall ornamentation. Because the serrate edge is either not formed or cannot be readily detected in most cases, it is not a satisfactory morphological feature for delimiting this species. In laboratory studies it is noted that S. parisiensis might be confused with S. denticulatus rather than S. brasilien-sis. Inasmuch as both xenic and axenic cultures of the -f strains produced similar results, S. parisiensis can be readily characterized.     

Activities of superoxide dismutase (SOD) isoforms during growth of <Emphasis Type="Italic">Scenedesmus</Emphasis> (chlorophyta) species and strains grown in batch-cultures

Growth of Scenedesmus species and strains, grown for 28 days in mineral BBM medium in batch-cultures, displayed sigmoidal kinetics that comprised a lag, exponential and declining growth phases. Total SOD activity in these autotrophically cultured organisms, which oscillated within 0.6 – 1.4 Umg protein⁻¹, was rather species-specific and only to some extent depended on the growth phase. Contrary, three S. obliquus strains: wild type 276-6, mutant with blocked PS I (strain 56.80) and mutant with blocked PS II (strain 57.80), cultured for 7 days on BBM medium supplemented with bacto-tryptone and yeast extract (BBM+) turned out to be time-dependent and to have several times higher total SOD activity than one obtained for Scenedesmus grown autotrophically. Regardless of the media composition, the phase of growth and studied organism, dominant isoforms of total SOD were together determined Fe- and Mn-SOD. Profiles of SOD isoforms, obtained after PAGE analysis of all autotrophically and exponentially growing organisms, revealed that one Mn-SOD and one Cu/Zn-SOD bands located on gels at the same position whereas location of three bands of Fe-SOD depended on the strain. This suggests the presence of two different groups of Fe-SODs in analyzed organisms. Identical SOD profiles found in two S. armatus strains (276-4a and 276-4d) and S. subspicatus correspond well with their taxonomic position. The SOD profile of S. armatus B1-76 distinctly differed from two other S. armatus strains but was identical to S. microspinal B1-76 and S. quadricauda G-15 despite the fact that there were significant growth rate differences between these three species. SODs profiles of S. acutus 437 and S. obliguus 453 were species-specific. In S. obliquus strains cultured on BBM+ medium, there are four SOD bands: one slightly visible band of Mn-SOD, two intensive bands of Fe-SOD and one band of Cu/Zn-SOD. The above finding suggests that antioxidant response of algae kept in batch-cultures differs according to medium composition and the SOD activity mainly restricted to chloroplasts.     

<Emphasis Type="Italic">Scenedesmus</Emphasis> sp. cultivation using commercial-grade ammonium sources

The use of commercial-grade nutrients such as agricultural fertilizers is important for commercial microalgae cultivation, and this is particularly the case for biofuel production which is associated with low added value. Nitrogen is a very important macronutrient in microalgae cultivation, and ammonium sources are cheaper than nitrate sources. However, the growth response and cellular composition can be altered by the different nutrient sources. In the study reported here, we investigated the effects of different ammonium doses and commercial-grade macronutrients from agricultural fertilizers on the growth of Scenedesmus sp. BR003, a promising genus for biofuel production. Five growth media were developed using fertilizers and evaluated during Scenedesmus sp. cultivation under autotrophic conditions. The growth media differed in terms of their composition and concentration of macronutrients. We found that all commercial-grade media supported equal or higher cell concentrations, dry weight, water-soluble proteins, neutral carbohydrates, and total lipid production compared to the conventional BG11 medium. However, the commercial-grade growth medium with the highest ammonium content affected the coenobium pattern of Scenedesmus sp. BR003. Commercial-grade nutrient sources were a low-cost alternative to improve the growth of Scenedesmus sp. BR003. The different fertilizers also allowed for manipulation of microalgae chemical composition and phenotypic plasticity to target traits of commercial interest. Our results demonstrate the potential of using ammonium from agricultural fertilizers as a nitrogen source in combination with other commercial-grade macronutrients sources. In addition, this work demonstrates the ability of a robust Scenedesmus strain to grow in media of different compositions, even when a high dosage of ammonium was used.     

AFLP analysis shows high incongruence between genetic differentiation and morphology‐based taxonomy in a widely distributed tortoise

Morphology has traditionally been used to diagnose the taxa of various taxonomic ranks. However, there is growing evidence that morphology is not always able to reveal cryptic taxa, and that pronounced morphological variation could reflect phenotypic plasticity rather than evolutionary divergence. Spur‐thighed tortoises (the Testudo graeca complex), distributed in the western Palaearctic region, are characterized by high morphological variability and complicated taxonomy, which are under debate. Previous molecular studies using mainly mitochondrial DNA (mtDNA) sequences have revealed incongruence between genetic differentiation and morphology‐based taxonomy, suggesting that morphological variability is the result of phenotypic plasticity and stabilizing selection, which masks the true genealogies. In the present study, we used a range‐wide sampling and nuclear Amplified fragment length polymorphism (AFLP) markers to investigate genetic differentiation within the T. graeca complex. We found that spur‐thighed tortoises are differentiated into four geographically well‐defined AFLP groups: Balkans–Middle Eastern, western Mediterranean, Caucasian and central‐eastern Iranian. Compared with the distribution of mtDNA lineages, the groups are largely concordant, although the AFLP markers are less sensitive and distinguish fewer groups than do mtDNA sequences. The AFLP groups show an allopatric or parapatric distribution. The AFLP differentiation conflicts with the previously proposed morphology‐based taxonomy of the complex, suggesting that local adaptation to different environmental conditions may have led to the great extent of morphological variation within the same lineages. We propose a re‐evaluation of the taxa that were confirmed genetically using a thorough morphological analysis corrected for phenotypic plasticity. © 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, ●● , ●●–●●.     

PATTERNS OF VARIATION IN THE STELLARIA LONGIPES COMPLEX: EFFECTS OF POLYPLOIDY AND NATURAL SELECTION

Patterns of morphological variation were studied in herbarium specimens of Stellaria longipes, an herbaceous perennial and subsequently in a growth chamber experiment using three cytotypes (4x, 6x, 8x) of S. longipes and diploids of its proposed progenitor S. longifolia. Despite extensive phenotypic plasticity in many traits, patterns of variation resulting from ecotypic differentiation within S. longipes could be detected in the field. A distinct form of S. longipes, which is restricted primarily to arctic and alpine tundra locations, shows genetic differentiation for the following traits: few flowers per ramet, a low proportion of flowering ramets, and ovate leaves. The three cytotypes of S. longipes could be distinguished by their mean genotypic value for leaf length and number of flowers per ramet. The extent of phenotypic plasticity in these traits makes it unlikely that the cytotypes could be distinguished in the field. The direction and extent of morphological divergence between S. longifolia and S. longipes suggest an alloploid origin for S. longipes. Variational trends (among-habitat types and cytotypes) in trait means are similar to those reported previously for the pattern of plasticity. This supports the argument that similar forces guide evolution of the mean and pattern of plasticity of a trait.     

Grazing resistance in nutrient-stressed phytoplankton

Grazing experiments were performed with the zooplankters Daphnia pulex and Daphnia magna, feeding on phosphorus-saturated and phosphorus-limited cells of two green algae (Scenedesmus subspicatus and Selenastrum capricornutum). P-limited algal cells passed largely intact through the gut and were thus spared from heavy grazing pressure. P-saturated algal cells, in contrast, were efficiently assimilated. Structural and morphological changes in the P-limited cells most probably reduced their digestibility. This phenomenon may be an important factor in zooplankton production and competition, and may serve as an example of a highly efficient strategy of P-limited algae to resist heavy grazing pressure.     

Isolation, culture and characterization of a new strain of Euglena gracilis

A new strain of Euglena gracilis Klebs has been isolated from a highly polluted river; it was named MAT. Strain growth in different culture media was evaluated under heterotrophic and autotrophic conditions. Total lipid, sugar, protein and chlorophyll a production were studied. Results obtained for MAT were compared with data obtained for a UTEX Culture Collection strain. Likewise, cells from both strains were bleached using streptomycin, and grown in the same media used for green samples. Both MAT and UTEX showed clear differences in their biochemical composition and growth rate depending on the media used. They also exhibited different growth patterns. E. gracilis medium proved to be the best culture environment for both strains either in autotrophic or heterotrophic conditions. Results show that basal contents of lipids, sugars, proteins and chlorophyll a vary depending on the strain, and thus values obtained for one strain do not apply to another. Moreover, strain origin may have an influence on the mechanisms of adaptation or defense developed by each sample.     

Phenotypic plasticity inPotamogeton (Potamogetonaceae)

Sources of the extensive morphological variation of the species and hybrids ofPotamogeton were studied, especially from the viewpoint of the stability of the morphological characters used inPotamogeton taxonomy. Transplant experiments, the cultivation of clones under different values of environmental factors, and the cultivation of different clones under uniform conditions were performed to assess the proportion of phenotypic plasticity in the total morphological variation. Samples from 184 populations of 41Potamogeton taxa were grown. The immense range of phenotypic plasticity, which is possible for a single clone, is documented in detail in 14 well-described examples. The differences among distinct populations of a single species observed in the field were mostly not maintained when grown together under the same environmental conditions. Clonal material cultivated under different values of environmental factors produced distinct phenotypes, and in a few cases a single genotype was able to demonstrate almost the entire range of morphological variation in an observed trait known for that species. Several characters by recent literature claimed to be suitable for distinguishing varieties or even species were proven to be dependent on environmental conditions and to be highly unreliable markers for the delimiation of taxa. The unsatisfactory taxonomy that results when such classification of phenotypes is adopted is illustrated by three examples from recent literature. Phenotypic plasticity was found to be the main source of morphological variation within the species ofPotamogeton, having much more influence than morphological differences caused by different genotypes.     

The effect of nitrogen and temperature changes on Solidago canadensis phenotypic plasticity and fitness

Phenotypic plasticity is commonly considered to contribute to the invasion success of invasive species. However, the importance of phenotypic plasticity, nitrogen (N) levels and warming to the invasion of invasive species is unclear. The effects of warming and N addition on the morphology, biomass allocation and biochemistry traits of Solidago canadensis and their plasticity were investigated by conducting a pot experiment. The results showed that the effect of N addition on biomass was improved for S. canadensis; whereas warming displayed no significant effect, their positive synergistic interact effect resulted in the overall significant increase in plant performance. The mean phenotypic plasticity indices (MPPI) of biochemistry and total parameters demonstrated a difference between operations, and the higher value was observed in N interaction with temperature treatments than N addition or warming alone. The observed MPPI indicated the biochemistry parameters > morphological parameters > allocation parameters. The MPPI of biochemistry parameters, morphological parameters and total parameters exhibited significant and positive correlations with N level and MPPI of morphological parameters was also significantly positively correlated with the fitness of S. canadensis. These results indicated that the global warming and N addition would make the invaded habitat more suitable for the growth of invasive S. canadensis, and even may effectively increase the invasion risk of S. canadensis through the enhanced phenotypic plasticity, which is a crucial factor to help species deal with the changing environment.     

Strukturveränderungen am PlasmalemmaAphelidium-infizierter Scenedesmus-Zellen

Zusammenfassung Es werden Strukturveränderungen am Plasmalemma vonScenedesmus acutus-Zellen beschrieben, die vonAphelidium spec. infiziert waren. Der Parasit entwickelt imScenedesmus-Cytoplasma ein Plasmodium, dessen Plasmalemma von dem durch den Parasiten eingestülpten Teil desScenedesmus-Plasmalemmas umgeben ist. Beide Membranen verlaufen mit geringem Abstand parallel und bilden manchmal eine compound membrane. Der verlagerte Teil desScenedesmus-Plasmalemmas hat eine mehr oder weniger deutlich asymmetrische Struktur, während der nicht verlagerte Teil (an der Zellwand) symmetrisch ist. Das Bild dieser Membranen ist von der Fixierung abhängig. Reste des nichtverlagerten Teils desScenedesmus-Plasmalemmas kommen sogar dann noch vor, wenn das Cytoplasma völlig aufgelöst ist; sie treten in Form Lamellen-ähnlicher, trilaminarer, symmetrischer Strukturen auf. Gelegentlich fusionieren die Plasmalemmen des Wirtes und des Parasiten artifiziell miteinander und bilden Vesikel. Unterschiede zwischen dieser parasitischen Assoziation und der vonSc. armatus mit einem anderen Typ vonAphelidium (Schnepf et al. 1971) werden erwähnt.

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Structural modifications in the plasmalemma ofAphelidium-infectedScenedesmus cells

Summary Modifications in the structure of the plasmalemma of cells ofScenedesmus acutus on infection by a strain ofAphelidium are described. The parasite develops a plasmodium inside theScenedesmus cytoplasm, its plasmalemma is surrounded by a dislocated portion of theScenedesmus plasmalemma. Both membranes run parallel, a short distance apart, and occasionally form a compound membrane. The dislocated portion of theScenedesmus plasmalemma has a more or less asymmetrical structure whereas the undislocated portion (at the cell wall) is nearly symmetrical. The appearance of these modifications is dependent on the fixation method. Remnants of the undislocated part of theScenedesmus plasmalemma are found even when the host cytoplasm has lysed completely; they occur in the form of lamella-like, trilaminar symmetrical structures. Occasionally the plasmalemma of the host and of the parasite fuse artificially to form vesicles. Differences between this parasitic association and that ofSc. armatus with another strain ofAphelidium (Schnepf et al. 1971) are described.

     

Gordonia sinesedis sp. nov., a novel soil isolate

The taxonomic position of an actinomycete isolated from soil was evaluated using a polyphasic approach. The organism, strain J72, was found to have chemical and morphological properties consistent with its assignment to the genus Gordonia. A nearly complete 16S rDNA sequence of the strain was determined by direct sequencing of the amplified gene. The tested strain formed a distinct phylogenetic line within the evolutionary radiation occupied by the genus Gordonia and was most closely related to G. polyisoprenivorans DSM 44302^T. The phenotypic profile of strain 372 readily distinguishes it from representatives of the validly described species of Gordonia. The combined genotypic and phenotypic data show that strain J72 merits recognition as a new species of Gordonia. The name proposed for the new species is Gordonia sinesedis; the type strain is J72^T (= DSM 44455^T = NCIMB 13802^T). This revised version was published online in June 2006 with corrections to the Cover Date.     

“DNA signaturing” database construction for Tetradesmus species identification and phylogenetic relationships of Scenedesmus-like green microalgae (Scenedesmaceae,Chlorophyta)

Species identification of Scenedesmus-like microalgae, comprising Desmodesmus, Tetradesmus, and Scenedesmus, has been challenging due to their high morphological and genetic similarity. After developing a DNA signaturing tool for Desmodesmus identification, we built a DNA signaturing database for Tetradesmus. The DNA signaturing tool contained species-specific nucleotide sequences of Tetradesmus species or strain groups with high similarity in ITS2 sequences. To construct DNA signaturing, we collected data on ITS2 sequences, aligned the sequences, organized the data by ITS2 sequence homology, and determined signature sequences according to hemi-compensatory base changes (hCBC)/CBC data from previous studies. Four Tetradesmus species and 11 strain groups had DNA signatures. The signature sequence of the genus Tetradesmus, TTA GAG GCT TAA GCA AGG ACCC, recognized 86% (157/183) of the collected Tetradesmus strains. Phylogenetic analysis of Scenedesmus-like species revealed that the Tetradesmus species were monophyletic and closely related to each other based on branch lengths. Desmodesmus was suggested to split into two subgenera due to their genetic and morphological distinction. Scenedesmus must be analyzed along with other genera of the Scenedesmaceae family to determine their genetic relationships. Importantly, DNA signaturing was integrated into a database for identifying Scenedesmus-like species through BLAST.