MONOGRAPH OF THE GENUS PROTOSTELIUM

Three new species of Protostelium (Order Protostelida of the Mycetozoa) are described: P. irregularis, P. zonatura, and P. pyriformis, all with rather wide distribution and occurring on dead attached plant parts, less often in soil and humus. The latter two species differ from the first and from the other two known species, P. mycophaga Olive and Stoianovitch and P. arachisporum Olive, in the endogenous origin of the stalk. P. mycophaga var. crassipes, with vesicular stalk bases, is the third variety of that species to be described. A key to the genus is included.     

OLIGOAGARS ELICIT A PHYSIOLOGICAL RESPONSE IN GRACILARIA CONFERTA (RHODOPHYTA)

Gracilaria conferta (Schousboe ex Montagne) J. et G. Feldmann responded with an oxidative burst and rapid increases in respiration and halogenating activity when agar, agarose, or the agarose degradation products neoagarotetraose and neoagarohexaose were added to the growth medium. In contrast, carrageenan, oligocarrageenans, neoagarobiose, l-galactose, d-galactose, and several other mono- and oligosaccharides did not have any effect. Sixfold increases in respiration were observed 3 min after addition of neoagarohexaose. The response could only be induced in species of the genera Gracilaria and Gracilariopsis. Neoagarohexaose also elicited a release of hydrogen peroxide in less than 15 min, resulting in an immediate increase in algal brominating activity. Bleached thallus tips appeared a few hours after the addition of neoagarohexaose. This effect was dependent on the release of hydrogen peroxide and exposure to light. Exposure to light and oligosaccharide elicitors increased the production of reactive oxygen species, which reached destructive concentrations when both mechanisms were simultaneously active. Concentrations of 0.1 to 3.3 μM agarose or agars were sufficient to trigger an increase in respiration, an oxidative burst response, and tip bleaching. However, higher concentrations of neoagarohexaose and neoagarotetraose were necessary to elicit the responses, indicating that the alga is more sensitive to oligoagars with degrees of biose-polymerization > 3. The extremely short reaction time and high specificity indicate that intermediates of agar degradation are recognized by Gracilaria as messengers when microbial degradation of its cell wall occurs. The physiological responses may represent the early stages of algal defense mechanisms involved in repression of pathogen ingress.     

中国串珠藻属(红藻门)的两个新种

本文纪录了淡水红藻串珠藻属Batrachospermum的二个新种。它们采自中国的亚热地区,按其形态特征,它们属于旋转组Section Contorta。采自湖北省的弯形中珠藻B.curvatum与扭曲串珠藻B.tortuosum Kumano相似,但按其轮节、顶毛和受精丝等的形态与后者相区别。采自江西省的弯转串珠藻B.torsivum与赫罗西串珠藻B.hirosei Ratnasabapathy et Kumano相似,但轮节、中轴细胞、节丝和节间丝、果胞枝和受精丝等的大小和形态与后者相区别。     

SIMILAR UNSTABLE MUTATIONS IN THREE SPECIES OF GRACILARIA (RHODOPHYTA)1

Unstable mutants with similar variegated pigmentation were genetically characterized in the red algae. Gracilaria tikvahiae (McLachlan), G. foliifera (Forsk.) Børg. and. G. sjoestedtii (Kylin). All three mutants were green plants with flecks of red tissue where cells had reverted to wild type. The mutant green phenotypes were all recessive, and their genetic behavior in crosses indicated that each was the result of a single, unstable, nuclear gene. Wild-type revertant tissue was stable one it arose. Revertant plants obtained from spores and revertant fronds taken from variegated plants could not be distinguished from the normal wild type, either phenotypically or genetically. Reversion to wild type occurred during all phases of the life cycle. In crosses between the mutants and wild type, most of the F₁ tetrasporophytes were heterozygous wild-type plants, an observation consistent with the recessive nature of the mutations; however, a low frequency of homozygous unstable-green F₁ tetrasporophytes was also obttained from these crosses. The molecular basis of neither the mutant instability, i.e. the reversion to wild type, nor of the process producing the unstable green F₁ tetrasporophytes can yet be deduced, but the phenotype of the plants and genetic results suggest the involvement of transposable genetic elements.     

A MOLECULAR AND MORPHOLOGICAL ANALYSIS OF THE GYMNOGONGRUS DEVONIENSIS (RHODOPHYTA) COMPLEX IN THE NORTH ATLANTIC1

The Gymnogongrus devoniensis (Greville) Schotter complex in the North Atlantic Ocean was elucidated by comparative molecular, morphological, and culture studies. Restriction fragment length patterns and hybridization data on organellar DNA revealed two distinct taxa in samples from Europe and eastern Canada. Nucleotide sequences for the intergenic spacer between the large and small subunit genes of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), and the adjoining regions of both genes, differed by 12.5–13.4% between the two taxa. One of the taxa, which included material from the type locality of G. devoniensis at Torbay, Devon, England, was taken to represent authentic G. devoniensis. Within this taxon, samples from Ireland, England, northern France, northern Spain, and southern Portugal showed great morphological variation, particularly in habit, but their Rubisco spacer sequences were identical or differed by only a single nucleotide. Constant morphological features included the development, from a single auxiliary cell, of the spherical cystocarp with a thick mucilage sheath that appears to be typical of Gymnogongrus species with internal cystocarps. Two life-history types were found. Northern isolates underwent a direct-type life history, recycling apomictic females by carpospores, whereas the Portuguese isolate followed a heteromorphic life history in which carpospores gave rise to a crustose tetrasporophyte. The second group of samples, from Nova Scotia and Northern Ireland, provisionally referred to as Gymnogongrus sp., showed little morphological variation. The life history in both areas consists of apomictically reproducing diploid female gametophytes and diploid crustose bisporophytes and tetrasporophytes. Rubisco spacer sequences of the samples were identical, and the plasmid previously described in the Nova Scotian samples was also present in the Northern Ireland population. This species is widely distributed in the western Atlantic, from Newfoundland to Massachusetts. In Europe, gametophytes are known only at one site, but crusts are distributed from Denmark, Scotland (and probably Norway) to France. It is very likely that this species was introduced from one side of the North Atlantic to the other by shipping during the early nineteenth century. Several morphological features are unusual within the genus but are shared with G. leptophyllus J. Agardh from the eastern Pacific Ocean, and further work is necessary to determine whether Gymnogongrus sp. and G. leptophyllus are conspecific.     

DESICCATION TOLERANCE OF GRACILARIA TIKVAHIAE (RHODOPHYTA)1,2

The photosynthesis of Gracilaria tikvahiae McLachlan, normally a subtidal plant, returned to initial levels within a day following single desiccations to 50% and repeated desiccations to at least 15%. In contrast, growth rates were decreased by even very mild desiccation.     

GRACILARIA EDULIS (RHODOPHYTA) AS A BIOLOGICAL INDICATOR OF PULSED NUTRIENTS IN OLIGOTROPHIC WATERS

The response of the marine macroalga Gracilaria edulis (Gmelin) Silva to nutrient pulses of varying magnitude was investigated to test its applicability as a marine bioindicator at two oligotrophic locations. After exposure to nutrient pulses, algal amino acid, tissue nitrogen, and chlorophyll a content were assessed relative to algae incubated under control conditions (no nutrient enrichment). The smallest nutrient pulse involved a nutrient enrichment experiment conducted within a coral atoll, whereas two larger pulses resulted from sewage discharge to a tropical coastal bay. After exposure to the smallest nutrient pulse (10 × ambient), only changes in macroalgal amino acid concentration and composition were detected (mainly as increases in citrulline). At 100 × ambient concentrations, increases in tissue % nitrogen of the macroalgae were detected, in addition to responses in amino acids. Macroalgae exposed to the highest nutrient pulse (1000 × ambient) responded with increased chlorophyll a , tissue nitrogen, and amino acids within the three day incubation period. In contrast to these algal responses, analytical water sampling techniques failed to detect elevated nutrients when nutrient pulses were not occurring. The responses of this algal bioindicator to variable nutrient pulses may provide a useful tool for investigating the source and geographical extent of nutrients entering oligotrophic coastal waters.     

DISSECTION OF TWO DISTINCT DEFENSE‐RELATED RESPONSES TO AGAR OLIGOSACCHARIDES IN GRACILARIA CHILENSIS (RHODOPHYTA) AND GRACILARIA CONFERTA (RHODOPHYTA)1

The two agar‐producing red algae, Gracilaria chilensis C. J. Bird, McLachlan & E. C. Oliveira and Gracilaria conferta (Schousboe ex Montagne) Montagne, responded with hydrogen peroxide (H₂O₂) release when agar oligosaccharides were added to the medium. In G. conferta, a transient release was observed, followed by a refractory state of 6 h. This response was sensitive to chemical inhibitors of NADPH oxidase, protein kinases, protein phosphatases, and calcium translocation in the cell, whereas it was insensitive to inhibitors of metalloenzymes. Transmission electron microscopic observations of the H₂O₂‐dependent formation of cerium peroxide from cerium chloride indicated oxygen activation at the plasma membrane of G. conferta. A putative system, consisting of a receptor specific to agar oligosaccharides and a plasma membrane‐located NADPH oxidase, appears to be responsible for the release of H₂O₂ in G. conferta. Subcellular examination of G. chilensis showed that the H₂O₂ release was located in the cell wall. It was sensitive to inhibitors of metalloenzymes and flavoenzymes, and no refractory state was observed. The release was correlated with accumulation of an aldehyde in the algal medium, suggesting that an agar oligosaccharide oxidase is present in the apoplast of G. chilensis. The presence of this enzyme could also be demonstrated by polyacrylamide electrophoresis under nondenaturating conditions and proven to be variable. Cultivation of G. chilensis at 16 to 17°C resulted in significantly stronger expression of agar oligosaccharide oxidase than cultivation at 12°C, which indicates that the enzyme is used under conditions that generally favor microbial agar macerating activity.     

镰刀菌属分类进展

镰刀菌是真菌中最难鉴定和最具经济价值的属之一。自从1935年德国的Wollen-weber & Reinking发表了第一个以16组65种为内容的分类系统以来,人们对该属的分类展开了长期的研究和讨论,在国际上陆续出现了10种以此系统为基础的,而又各具特色的分类系统。它们是Snyder & Hansen(1940s,美国)的9种系统;Paйлло(1950,苏联)的55种系统;Gordon(1952,加拿大)的26种系统;Билай(1955,苏联)的26种系统;Messiaen & Cassini(1968,法国)的9种和9变种系统;Booth(1971,英国)的44种系统;松尾卓见(Matuo,1972,日本)的10种系统;Joffe(1974,以色列)的33种系统;Gerlach& Nirenberg(1982,德国)的90多种和变种的系统;Nelson等(1983,美国)的30种系统。本文重点论述了其中5种在国际上颇具影响的系统,并列表加以详细比较。     

PROTOPLAST ISOLATION AND CELL DIVISION IN THE AGAR-PRODUCING SEAWEED GRACILARIA (RHODOPHYTA)1

Methods were developed for the isolation of large numbers of healthy protoplasts from two species of the agarophyte Gracilaria; G. tikvahiae McLachlan and G. lemaneiformis (Bory) Weber-van Bosse. This is the first report of protoplast isolation and cell division in a commercially important, phycocolloid-producing red seaweed, as well as for a member of the Florideophycidae. The optimal enzyme composition for cell wall digestion and protoplast viability consisted of 3% Onozuka R-10, 3% Macerozyme R-10, 1% agarase and 0.5% Pectolyase Y- 23 dissolved in a 60% seawater osmoticum containing 1.0 M mannitol. The complete removal of the cell wall was confirmed by several different methods, including electron microscopic examination, and the absence of Calcofluor White (for cellulose) and TBO (for sulfated polysaccharide) staining. Spontaneous protoplast fusion was observed on several occasions. Protoplast viability was dependent upon the strain and age of the parent material, as well as the mannitol concentration of the enzyme osmoticum. Cell wall regeneration generally occurred in 2-6 days; cell division in 5-10 days. Protoplast-produced cell masses up to the 16-32 cell stage have been grown in culture. However, efforts to regenerate whole plants have been unsuccessful to date.     

DIURNAL FLUCTUATION OF NITRATE REDUCTASE ACTIVITY IN THE MARINE RED ALGA GRACILARIA TENUISTIPITATA (RHODOPHYTA)1

The biochemical characteristics and diurnal changes in activity of the enzyme nitrate reductase (NR; EC 1.6.6.1) from the marine red alga Gracilaria tenuistipitata var. liui Zhang et Xia are described. Different assay conditions were tested to determine the stability of NR. The crude extract of G. tenuistipitata has a NR specific activity of 10.2 U.mg⁻¹, which is higher than the NR activities found for other algae, plants, and fungi. This NR is highly active at a slightly alkaline pH and is stable over a wide range of temperature, with an optimal activity at 20° C. The apical portions of the thallus contain 64.9 ± 6.6% of the total NR specific activity. The apparent Michaelis-Menten (K_m) constant found for KNO₃ was 197 μM, and it was 95 μM for NADH. The NR from G. tenuistipitata can be included in the NADH-specific group, because no activity was found when NADPH was used as an electron donor. In extracts of algae grown under either continuously dim light or a light-dark cycle, the activity of NR exhibits a daily rhythm, peaking at the middle of the light phase, when activity is 30-fold higher than during the night phase.     

SYSTEMATIC EVALUATION OF THE GENUS HILDENBRANDIA (RHODOPHYTA): A SYNTHESIS OF TECHNIQUES

The evolutionary relationships among members of the red algal genus Hildenbrandia have not been well understood for several reasons. For example, the genus contains both marine and freshwater representatives, all of which are non-calcified and crustose, and few have definitive morphological characters for classification. Hildenbrandia is also assumed to be completely asexual (reproduction by tetrasporangia in marine forms and by gemmae in freshwater populations), and characters of the female gametangial system and post-fertilization structures are not available for comparative studies. Currently there are 14 marine and five freshwater species and infraspecific taxa recognized within the genus. We used phylogenetic analyses (parsimony, distance and maximum likelihood) of DNA sequences of commonly employed genes ( rbc L and 18S rRNA) to examine the evolutionary relationships among representatives of many of these taxa. In addition, we employed morphometrics (principal co-ordinates and cluster analyses) of several measured characters of these same representatives, as well as all available type specimens, to determine the number of morphologically-delimited entities within the genus. Thus far our results indicate that some characters traditionally used to distinguish species of Hildenbrandia , such as tetrasporangial division pattern, may not be useful in some cases, and a revision of the taxonomy of the genus will be necessary. Although the marine and freshwater species of Hildenbrandia appear to be well separated in our molecular analyses of European specimens, this trend was not observed for North American specimens. High sequence divergence values were calculated for both the rbc L and 18S rRNA genes of Hildenbrandia , compared to other red algal genera.     

DIFFUSION BOUNDARY LAYER TRANSPORT IN GRACILARIA CONFERTA (RHODOPHYTA)1

A theoretical framework on the combined effect of water velocity and solute concentration on the photosynthetic performance of the red alga Gracilaria conferta (Rhodophyta) is developed. This is based on the balance between the rate of transport through boundary layers and Michaelis-Menten-type equations for carbon consumption and for production of oxygen and hydroxyl ions. By comparing the theoretical models with experimental data, we found that the mechanism of enhancing photosynthetic rates by increasing water velocity cannot be attributed to enhanced bicarbonate and CO₂ transport, nor to CO₂ as a sole source of carbon. Velocity-facilitated photosynthesis may be due to the enhanced removal of OH^? ions, which inhibit photosynthesis when accumulated on the algal surface. Oxygen had no inhibitory effect on Gracilaria conferta.     

UPTAKE AND RELEASE OF NITROGEN BY THE MACROALGAE GRACILARIA VERMICULOPHYLLA (RHODOPHYTA)1

Macroalgae, often the dominant primary producers in shallow estuaries, can be important regulators of nitrogen (N) cycling. Like phytoplankton, actively growing macroalgae release N to the water column; yet little is known about the quantity or nature of this release. Using ¹⁵N labeling in laboratory and field experiments, we estimated the quantity of N released relative to assimilation and gross uptake by Gracilaria vermiculophylla (Ohmi) Papenfuss (Rhodophyta, Gracilariales), a non‐native macroalgae. Field experiments were carried out in Hog Island Bay, a shallow back‐barrier lagoon on the Virginia coast where G. vermiculophylla makes up 85%–90% of the biomass. There was good agreement between laboratory and field measurements of N uptake and release. Daily N assimilation in field experiments (32.3±7.2 μ mol N·g dw^?1·d^?1) was correlated with seasonal and local N availability. The average rate of N release across all sites and dates (65.8±11.6 μ mol N·g dw^?1·d^?1) was 67% of gross daily uptake, and also varied among sites and seasons (range=33%–99%). Release was highest when growth rates and nutrient availability were low, possibly due to senescence during these periods. During summer biomass peaks, estimated N release from macroalgal mats was as high as 17 mmol N·m^?2·d^?1. Our results suggest that most estimates of macroalgal N uptake severely underestimate gross N uptake and that N is taken up, transformed, and released to the water column on short time scales (minutes–hours).     

PROTEOMICS OF THE RED ALGA,GRACILARIA CHANGII (GRACILARIALES,RHODOPHYTA)1

The application of proteomics in alga research is still quite limited. The present report describes the establishment of the proteome of a red alga of economic importance, Gracilaria changii (Xia et Abbott) Abbott, Zhang et Xia. Initially, four protein extraction methods including direct precipitation by trichloroacetic acid/acetone, direct lysis using urea buffer, Tris buffer, and phenol/chloroform extraction were compared for their suitability to generate G. changii proteins for two‐dimensional gel electrophoresis (2‐DE). The phenol/chloroform protein extraction method gave the best 2‐DE resolution of the proteins. Using these 2‐DE gels and mass spectrometry, several proteins including pigment proteins, metabolic enzymes, and ion transporters were identified. These findings highlight the potential of using proteomic approaches for the investigation of G. changii protein function.     

TAXONOMY AND PHYLOGENY OF OSMUNDEA (RHODOMELACEAE, RHODOPHYTA) IN ATLANTIC EUROPE

Two species of Osmundea Stackhouse (Rhodomelaceae, Rhodophyta) that occur in Atlantic Europe have been confused under the names Osmundea ramosissima (Oeder) Athanasiadis and Osmundea truncata (Kützing) Nam et Maggs, regarded until now as a synonym of O. ramosissima. An epitype from its type locality (Stavanger, Norway) is selected for Osmundea ramosissima Athanasiadis, recognized here as a valid name for Fucus ramosissimus Oeder, nom. illeg. Details of vegetative and reproductive morphology of O. ramosissima are reported, based on material from France, the British Isles, and Helgoland. Osmundea ramosissima resembles other species of Osmundea in its vegetative axial segments with two pericentral cells and one trichoblast, spermatangial development from apical and epidermal cells (filament type), the formation of five pericentral cells in the procarp‐bearing segment of the female trichoblast, and tetrasporangial production from random epidermal cells. Among the species of Osmundea, O. ramosissima is most similar to O. truncata. Both species have discoid holdfasts, secondary pit connections between epidermal cells, and cup‐shaped spermatangial pits. They differ in that: (a) O. ramosissima lacks lenticular wall thickenings and refractive needle‐like inclusions in medullary cells, both of which are present in O. truncata; (b) O. ramosissima has branched spermatangial filaments that terminate in a cluster of several cells, whereas in O. truncata the unbranched spermatangial filaments have a single large terminal sterile cell; and (c) cystocarps of O. ramosissima lack protuberant ostioles but ostioles are remarkably protuberant in O. truncata. Phylogenetic analyses of rbcL sequences of Laurencia obtusa (Hudson) Lamouroux and all five Atlantic European species of Osmundea, including the type species, strongly support the generic status of Osmundea. Osmundea ramosissima and O. truncata are closely related (5.2% sequence divergence) and form a well‐supported clade sister to a clade consisting of O. pinnatifida (Hudson) Stackhouse, O. osmunda Stackhouse and O. hybrida (A. P. de Candolle) Nam. The formation of secondary pit connections between epidermal cells is a synapomorphy for the O. ramosissima+O. truncata clade. The close relationship between species with cup‐shaped spermatangial pits (Osmundea hybrida) and urn‐shaped pits (Osmundea pinnatifida and Osmundea osmunda) shows that spermatangial pit shape is not an important phylogenetic character. Parsimony analysis of a morphological data set also supports the genus Osmundea but conflicts with the molecular trees in infrageneric relationships, placing O. hybrida basal within the Osmundea clade and grouping O. osmunda and O. pinnatifida but not O. truncata and O. ramosissima. A key to Osmundea species is presented.     

QUANTIFICATION OF THE EFFECTS OF SPORELING COALESCENCE ON THE EARLY DEVELOPMENT OF GRACILARIA CHILENSIS (RHODOPHYTA)1

Sporeling coalescence in species of Gracilariales and Gigartinales is predicted to result in larger basal areas of growing disks as well as earlier initiation, increased abundance, and faster growth rates of erect shoots as compared to noncoalescent sporelings. These responses have been interpreted as providing mutual benefits for organisms living in aggregation, counterbalancing disadvantages associated with crowding. Quantitative evaluations of sporelings of Gracilaria chilensis failed to support several of these predictions. Sporelings were grown in the laboratory from a range of single sporelings to coalescent masses of 20 sporelings. Coalescent sporeling masses of G. chilensis exhibited larger basal areas than noncoalescent ones, but because the specific growth rates were inversely related to the original number of carpospores, no significant differences in actual area increments, during most of the experiment, were found among sporelings derived from one, two, or three to five coalescing sporelings. Initiation of erect shoots occurred at a similar time, regardless of their origin, i.e. coalescent or noncoalescent. Abundance of erect shoots was only loosely related to the number of coalescing sporelings. Even though by the end of the experiment (week 18), the total length of the longer erect shoots arising from coalescent sporeling masses was significantly greater than that of shoots arising from noncoalescent sporelings, total length was independent of the original number of coalescing sporelings. Furthermore, specific elongation rates between week 12 and week 18 were significantly greater for noncoalescent sporelings than for coalescent sporeling masses. Quantitative screening of other species seems necessary before generalizations on the ecological advantages of sporeling coalescence in seaweeds can be made.     

GENETIC DATA HINT AT A COMMON DONOR REGION FOR INVASIVE ATLANTIC AND PACIFIC POPULATIONS OF GRACILARIA VERMICULOPHYLLA (GRACILARIALES,RHODOPHYTA)1

Gracilaria vermiculophylla (Ohmi) Papenf., an agar‐producing red alga introduced from northeast Asia to Europe and North America, is often highly abundant in invaded areas. To assay its genetic diversity and identify the putative source of invasive populations, we analyzed the mitochondrial cytochrome c oxidase subunit I (cox1) gene from 312 individuals of G. vermiculophylla collected in 37 native and 32 introduced locations. A total of 19 haplotypes were detected: 17 in northeast Asia and three in Europe and eastern and western North America, with only one shared among all regions. The shared haplotype was present in all introduced populations and in ～99% of individuals in the introduced areas. This haplotype was also found at three native locations in east Korea, west Japan, and eastern Russia. Both haplotype and nucleotide diversities were extremely low in Europe and North America compared to northeast Asia. Our study indicates that the East Sea/Sea of Japan is a likely donor region of the invasive populations of G. vermiculophylla in the east and west Atlantic and the east Pacific.