Laboratory culture and taxonomy ofHymenomonas coronata andOchrosphaera verrucosa (Class Prymnesiophyceae) from the Northwest Pacific

Hymenomonas coronata andOchrosphaera verrucosa, both members of the coccolithophorids, Class Prymnesiophyceae, have been studied by means of electron microscopy and with the aid of laboratory culture. Living specimens of these two species were collected in temperate and subtropical regions of Japan, including the Kii Peninsula and the Ryukyu Islands, and unialgal cultures were established in the laboratory. Their life histories are fundamentally identical, and consist of a non-motile vegetative stage that produces motile cells. The vegetative stage is either unicellular, or a packet consisting of a few cells. Both the non-motile cells and the motile cells are covered with two kinds of scales: these are thin scales of unmineralized nature and coccoliths. These two species differ from each other in the shape of the coccoliths and in the presence or absence of visible rudimentary haptonema, and they have been in separate families. The present study reveals that both species are fundamentally identical in the structure and the distribution of major organelles, especially with respect to two opposed pyrenoids which bulge from chloroplasts, each being traversed by two thylakoid bands, and a group of microtubules forming a flagellar root. On the basis of these characteristics, it would appear more logical to place these two species in the same family, namely the Hymenomonadaceae.     

Light and electron microscope observations of the type species of Syracosphaera,s. pulchra (Prymnesiophyceae)

Syracosphaera pulchra, the type species of the genus was isolated into unialgal culture and studied with both the light and electron microscope. A conspicuous coiling haptonema is present containing seven microtubules in the shaft and eight in the basal region; features shared with many taxa in the order Prymnesiales. The proximal and distal coccoliths differ in shape but resemble each other structurally: the outer elements alternate to make the rim. The proximal coccoliths possess an organic base-plate scale which is absent in the distal coccoliths. The uncalcified organic scales are ornamented by a radial, more or less concentric, fibrillar pattern and are arranged in several layers between the proximal coccoliths and the plasmalemma. The ultrastructure of the cell is typical of prymnesiophycean algae. The flagellar apparatus is characterized by the absence of secondary microtubular bundles which are usually well developed in other coccolithophorids with two microtubular roots. This feature is also rather similar to that found in members of the Prymnesiales.

This investigation has indicated that S. pulchra has, in some respects, a closer affinity with members of the Prymnesiales than with the coccolithophorids.     

Preliminary observations on differences of scale morphology at various stages in the life cycle of ‘Apistonema-Syracosphaera’ Sensu von Stosch

A preliminary study of the fine structure of various stages in the life cycle of von Stosch's solate of ‘Syracosphaera carterae’ has been made by means of sections and whole mounts. Protoplast structure appears to be similar throughout the cycle. However, the Apistonema thallus possesses a cell wall composed of numerous layers of closely adpressed rimless unmineralised scales. Two types of asexual swarmers are shown to be covered with one or two layers of somewhat similar scales. One type of swarmer possesses a short haptonema which is covered with small scales. In addition the coccolith-bearing stages are shown to be identical with Hymenomonas carterae sensu Manton and Peterfi (1969) and this isolate has therefore been transferred to H. carterae.     

Life history and taxonomy ofCricosphaera roscoffensis var.haptonemofera,var. nov. (class prymnesiophyceae) from the pacific

A new coccolithophoridCricosphaera roscoffensis var.haptonemofera is described by means of electron microscopy and with the aid of laboratory culture. The living specimens, which were in the motile phase, were collected in Okinawa, a subtropical region in Japan, and a unialgal culture was established in the laboratory. The life history was observed, starting from the motile cells. The life history consists of two phases: the motile phase and the benthic filamentous phase. The former is unicellular and presumably a diploid sporophyte, whereas the latter is similar to an alga previously known as “Apistonema submarinum” described by Dangeard (1934), and probably a haploid gametophyte. The motile cells bear two acronematic flagella and one short haptonema. The scales attached on the surface of the motile cells are of two kinds: one is an organic thin scale, and the other is a cricolith, a kind of coccolith, the latter being structurally almost identical with that ofCricosphaera roscoffensis studied by Gayral and Fresnel (1976). Because this species has neither haptonema nor a benthic filamentous phase, we propose, at present, to treat this alga as a new taxon at the rank of infraspecies, naming itCricosphaera roscoffensis var.haptonemofera Inouye et Chihara.     

Polyanion-mediated mineralization — assembly and reorganization of acidic polysaccharides in the Golgi system of a coccolithophorid alga during mineral deposition

Summary Immunolocalization of two highly acidic polysaccharides (PS-1 and PS-2) in a calcifying algaPleurochrysis carterae is described throughout the mineralization process, from before crystal nucleation through the cessation of crystal growth. This unicellular coccolithophorid alga is a useful model for mineralization because it produces calcified scales known as coccoliths in homogeneous cell culture. PS-1 and PS-2 were localized in the crystal coats of mature coccoliths and in electron dense Golgi particles. The polyanions are synthesized in medial Golgi cisternae and co-aggregate with calcium ions into discrete 25 nm particles. Particle-laden vesicles bud from cisternal margins and fuse with a coccolith-forming saccule containing an organic oval-shaped scale which forms the base of the future coccolith. The particles are localized on the base before the onset of mineral deposition and are present in the coccolith saccule throughout the period of crystal (CaCO₃) nucleation and growth. During the final phase of coccolith formation, the particles disappear, and the mature crystals acquire an amorphous coat containing PS-1 and PS-2 polysaccharides which remain with the mineral phase after the coccoliths are extruded from the cell. Postulated mechanisms of polyanion-mediated mineralization are reviewed and their relevance to the calcification of coccoliths is addressed.Abbreviations PS-1 polysaccharide one - PS-2 polysaccharide two - BSA bovine serum albumin - SDS sodium dodecyl sulfate - MES 2-(N-morpholino)-ethanesulfonic acid - EDTA ethylenediaminetetraacetic acid - DHA 3-deoxy-lyxo-2-heptulosaric acid - TCA trichloroacetic acid     

Intraspecific diversity and distribution of the cosmopolitan species Pseudo‐nitzschia pungens (Bacillariophyceae): morphology,genetics, and ecophysiology of the three clades

Three clades of Pseudo‐nitzschia pungens, determined by the internal transcribed space (ITS) region, are distributed throughout the world. We studied 15 P. pungens clones from various geographical locations and confirmed the existence of the three clades within P. pungens, based on ITS sequencing and described the three subgroups (IIIaa, IIIab, and IIIb) of clade III. Clade III (clade IIIaa) populations were reported for the first time in Korean coastal waters and the East China Sea. In morphometric analysis, we found the ultrastructural differences in the number of fibulae, striae, and poroids that separate the three clades. We carried out physiological tests on nine clones belonging to the three clades growing under various culture conditions. In temperature tests, only clade III clones could not grow at lower temperatures (10°C and 15°C), although clade I and II clones grew well. The estimated optimal growth range of clade I clones was wider than that of clades II and III. Clade II clones were considered to be adapted to lower temperatures and clade III to higher temperatures. In salinity tests, clade II and III clones did not grow well at a salinity of 40. Clade I clones were regarded as euryhaline and clade II and III clones were stenohaline. This supports the hypothesis that P. pungens clades have different ecophysiological characteristics based on their habitats. Our data show that physiological and morphological features are correlated with genetic intraspecific differentiation in P. pungens.     

Purification and Properties of Two Isozymes of γ-Glutamyltranspeptidase from Bacillus subtilis TAM-4

Two isozymes of γ-glutamyltranspeptidase, GGT-A and GGT-B, were purified to electrophoretic homogeneity from a culture broth of Bacillus subtilis TAM-4, which produces poly(γ-glutamic acid) (PGA) de novo. GGT-A was composed of three subunits with molecular weights of 23,000 (I), 39,000 (II), and 40,000 (III). GGT-B was composed of two subunits with molecular weights of 22,000 (I) and 39,000 (II). The N-terminal amino acid sequences of GGT-A subunit I and GGT-B subunit I were very similar. GGT-A subunit II and GGT-B subunit II had an identical N-terminal amino acid sequence. That of GGT-A subunit III showed no similarity to the other subunits. Both GGTs had similar enzymatic properties (optimum pH and temperature: pH 8.8 and 55°C) but showed a significantly different thermal stability at 55°C. Both GGT-A and -B used d-γ-glutamyl-p-nitroanilide as well as the l-isomer as the γ-glutamyl donor and used various amino acids and peptides as the acceptor. It was also found that the PGA produced by the strain was hydrolyzed to glutamic acid by its own GGTs.     

Proton nuclear magnetic resonance analysis of metabolic end products of the Bolivia strain of Trypanosoma cruzi and three of its clones

Proton nuclear magnetic resonance (¹H NMR) was used to study the in vivo metabolism of Trypanosoma cruzi, the pathogen causing American trypanosomiasis (Chagas' disease). Three clones were isolated from a strain of T. cruzi (Bolivia strain), The clones I, II and III and the original strain were characterized according to the spectra of their metabolic pathways to test the hypothesis that clonal evolution of T. cruzi has a major impact on biologically relevant properties of this parasite. T. cruzi (Bolivia strain) excreted acetate, alanine, glycerol, and succinate as major end products, in the proportion 6:4:2:2. Comparing the spectra of T. cruzi clones with the original Bolivia strain revealed both quantitative, as well as qualitative differences in the metabolites excreted: the clones I and II, as opposed to the Bolivia strain and clone III, excreted significant quantities of ethanol.     

COCCOLITH PRODUCTION AND DETACHMENT BY EMILIANIA HUXLEYI (PRYMNESIOPHYCEAE)1

Laboratory experiments were performed with the prymnesiophyte Emiliania huxleyi (Lohm.) Hay and Mohler, strain 88E, to quantify calcification per cell, coccolith detachment, and effects of coccolith production on optical scattering of individual cells. ¹⁴C incorporation into attached and detached coccoliths was measured using a bulk filtration technique. ¹⁴C-labeled cells also were sorted using a flow cytometer and analyzed for carbon incorporation into attached coccoliths. The difference between the bulk and flow cytometer analyses provided a ¹⁴C-based estimate of the rate of production of detached coccoliths. Coccolith production and detachment were separated in time in batch cultures, with most detachment happening well after calcification had stopped. Accumulation of coccoliths was maximum at the end of logarithmic growth with 50–80 coccoliths per cell (three to five complete layers of coccoliths around the cells). Net accretion rates of coccoliths were on the order of 7 coccoliths· cell^?1·d^?1 while net detachment rates were as high as 15 coccoliths· cell^?1·d^?1 for stationary phase cells. Equal numbers of coccoliths were attached and detached early in logarithmic growth, and as cells aged, the numbers of detached coccoliths exceeded the attached ones by a factor of 6. Our results demonstrate pronounced charges of forward angle light scatter and 90° light scatter of cells as they grow logarithmically and enter stationary phase. Counts of loose coccoliths in batch cultures are consistent with the detachment of coccoliths in layers rather than individual coccoliths.     

Light-dependent coccolith formation in the two forms of Coccolithus pelagicus

Summary Two methods were employed for measuring coccolith formation and photosynthesis in coccolithophorids. The first method was based on measurements of ¹⁴C radioactivity of cells on membrane filters before and after acid treatment. The second method involved a conversion of ¹⁴C in coccoliths or whole cells to BaCO₃ prior to counting. It was observed that in determinations of photosynthetic (or total) ¹⁴C by the first method, the count rate produced by a given amount of the isotope was 30–40% lower in the non-motile and motile forms of Coccolithus pelagicus than in C. huxleyi. There was no similarly great discrepancy in determinations of coccolith ¹⁴C.Light-dependent coccolith formation was demonstrated in both forms of C. pelagicus. The non-motile form may deposit several times more carbon in its coccoliths than it assimilates photosynthetically. In the motile form, coccolith carbon amounts to less than 2% of photosynthetic carbon.     

2-Keto-l-Gulonic Acid Fermentation

Fractionation of sorbitol metabolites in the culture liquid of Gluconobacter melanogenus IFO 3292 was examined by column chromatographic techniques. Ion exchange column chromatography of the culture supernatant allowed to divide the components of the metabolites into Fractions I, II, III and IV. Paperelectrophoretic and paperchromatographic analyses of these fractions revealed that Fractions I, II, III and IV contained neutral sugar, hexonic acids, 5-ketohexonic acid and 2-ketohexonic acids, respectively.

The neutral sugar in Fraction I, the 5-ketohexonic acid in Fraction III and the 2-ketohexonic acids in Fraction IV were isolated and determined to be l-sorbose, 5-keto-d- mannonic, 2-keto-d-gluconic and 2-keto-l-gulonic acids, respectively, from their physical properties. In Fraction II were contained two different hexonic acids, one of which was identified to be l-idonic acid by the aid of substrate specificity of a hexonic acid dehydrogenase of Pseudomonas aeruginosa, and the other was determined to be d-mannonic acid as the phenylhydrazide derivative.     

Adaptation to cycloheximide of macromolecular synthesis in Tetrahymena

Cycloheximide (CHI) at 10 ng/ml partially inhibited protein synthesis in exponential cultures of Tetrahymena Sp. At 20 ng/ml or greater, inhibition was complete. When protein synthesis was inhibited to any extent, cell division ceased immediately. In all instances where measured, synthesis of RNA and DNA also ceased. After a period of delay, cellular functions reinitiated in the order: (i) protein synthesis, (ii) DNA synthesis and, (iii) RNA synthesis and cell division. The delay in cell division was divided into three phases of: I, zero; II, low; and, III, fully recovered rates of exponential protein synthesis. The length of the three phases increased with increasing concentration of CHI Prior growth of cells for one generation in the presence of 7.5 ng/ml CHI (facilitation) eliminated phase I and slightly decreased phases II and III following subsequent challenge with an inhibitory concentration of CHI. Facilitation for six generations further decreased phases II and III. Protein synthesis and cell division were not inhibited during facilitation In the culture, succinate dehydrogenase activity did not increase during the delay but increased normally at the onset of division. In contrast, NADPH-cytochrome c reductase activity continued to increase for an hour after inhibition of protein synthesis, was constant for a period and did not increase again until an hour after reinitiatoin of cell division and RNA synthesis Inhibition of division of all cells was immediate and reinitiation of synthesis and cell division was non-synchronous.     

Morphologic variability of the coccolithophorid Calcidiscus leptoporus in the plankton, surface sediments and from the Early Pleistocene

On a global scale, morphological variability of the extant coccolithophorid Calcidiscus leptoporus (Murray and Blackman, 1898) Loeblich and Tappan was investigated in surface sediments and plankton samples and from an Early Pleistocene time-slice (1.8 Ma to 1.6 Ma). In the bivariate space coccolith diameter versus number of rays in the distal shield, Holocene samples follow a single, unimodal morphocline. Sample means of coccolith size and number of elements group in three clusters, I, II and III, which are of biogeographic significance. Clusters II and III coccoliths (mean coccolith size of 5.0 μm and 20.9 elements, and 6.6 μm and 25.6 elements, respectively) are found in a tropical belt extending from 11 °N to 17 °S with an annual minimum sea-surface temperature above 23.5 °C. Cluster I coccoliths (5.8 μm, 20.7 elements) are found in samples outside that belt. The distribution of coccoliths in the surface sediments is tentatively interpreted to be a result of mixing to a varying degree of at least three different morphotypes (‘small’, ‘intermediate’ and ‘large’), which were identified in the living plankton, and which are separated from each other at 5 μm and 8 μm mean coccolith diameter, respectively. A comparison of the surface sediments with the Early Pleistocene assemblages revealed that between 1.6 Ma and 1.8 Ma two morphoclines A and B existed, the first of which persisted until the Holocene in the form of C. leptoporus, while the second comprises only extinct morphotypes including Calcidiscus macintyrei as one end-member. During the Early Pleistocene morphocline A was more homogeneous and no clusters were evident.Morphocline B shows a clear bimodality with a separation of morphotypes at 9.5 μm. Our observations suggest that morphoclines are subsets within the total stratigraphical range of a single species, and represent the global variability of that species in a particular time interval. Morphotypes, which belong to a morphocline, represent the infra-specific variability of that species within the biogeographic and stratigraphic limits of that species.     

PLOIDAL CHANGES IN CLONAL CULTURES OF SPIROGYRA COMMUNIS AND IMPLICATIONS FOR SPECIES DEFINITION

A clonal culture of Spirogyra filaments of initially uniform width produced filaments of three additional significantly different widths. Group I filaments of the original clone were 30.9 ± 0.7 μm wide (mean ± SD, N = 50). Group I filaments produced Group II filaments (22.0 ± 1.1 μm) through vegetative growth and sexual reproduction. Zygospores from homothallic Group I filaments produced germlings representative of Groups I and II; zygospores from homothallic Group II filaments produced germlings representative of Group II only. Germlings of Groups III (27.7 ± 1.0 μm) and IV (44.9 ± 0.8 μm) were produced in the cross of I × II. Viable zygospores from homothallic Group III filaments were obtained. Cells of Group IV filaments were initially binucleate and did not conjugate. Of the six intergroup crosses possible, four resulted in conjugation-tube formation only; two crosses yielded zygospores (I × II and III × IV). Germlings from the successful cross of Groups III and IV produced filaments of all four groups. Chromosome counts were: Group I (24), Group II (12), Group III (18), and Group IV (24, one nucleus). Relative nuclear fluorescence values of mithramycin-stained DNA were (mean ± SD, N ≥ 30): Group I (11.1 ± 1.4), Group II (5.7 ± 0.7), Group III (8.8 ± 1.3), and Group IV (10.0 ± 0.9, one nucleus). Cytologically, Group II appears to be a diploid (2x), Group I a tetraploid (4x), and Group III a triploid (3x). Systematically, Groups I, II, and III key out to Spirogyra singularis, S. communis, and S. fragilis, respectively, using Transeau's mongraph of the family Zygnemataceae. These species are interpreted to represent a species complex of S. communis (whose name has priority) with the ancestral haploid (x = 6) missing.     

Continuous production of extracellular ultrafine calcite particles by the marine coccolithophorid alga Pleurochrysis carterae

A new procedure for the production of ultrafine calcite particles by the marine coccolithophorid alga Pleurochrysis carterae is reported. During continuous culture, calcite particles (coccoliths) were detached from the cell surface by optimized air-bubbling, which greatly reduced the damage associated with previous sonication methods. Detached calcite particles could be continuously recovered directly from the culture medium using a nylon mesh membrane filtration module. Cells remained viable and continued to produce coccoliths during culture. The optimum productivity of ultrafine calcite particles was 18 mg/l per day. These results demonstrate the potential for a continuous system for the photosynthetically driven removal of CO₂ and its fixation into ultrafine inorganic calcite particles. Correspondence to: T. Matsunaga     

Sex-ratio variation among Arisaema species with different patterns of gender diphasy

Arisaema species exhibit gender diphasy, or sex change, where individual plants produce either male, monoecious or female inflorescences depending on their size. Three basic sex-change patterns have been described in Arisaema. Type I species change between male and monoecious phases, type II species change between male, monoecious and female phases, while type III species change between male and female phases. Theoretical models suggest that sex ratios should be biased toward males, the sex with the lowest cost of reproduction. The goal of this study was to examine sex-ratio variation among Arisaema species that differ in sex-change patterns. Data from an extensive literature review, consisting of all available studies reporting Arisaema sex ratios, were combined with data from extensive field surveys of Arisaema dracontium and Arisaema triphyllum in southern Indiana, USA. This data set contains nearly 30 000 plants from 12 species. All species conformed to either the type I or type III pattern of sex change. There was little evidence for a distinct type II pattern of sex change, given that plants with monoecious inflorescences were rare relative to plants with pistillate inflorescences. The mean sex ratio in type I species (79.9% male) was significantly greater than in type III species (63.7% male). The data are consistent with the prediction that type I species are likely to have greater costs associated with female reproduction. We suggest that all Arisaema species have similar patterns of floral development, but differ in their ontogenetic patterns for male and female flowering.     

INCOMPLETENESS OF THE COCCOSPHERE AS A POSSIBLE STIMULUS FOR COCCOLITH FORMATION IN PLEUROCHRYSIS CARTERAE(PRYMNESIOPHYCEAE)1

Pleurochrysis carterae is a marine biflagellate that produces calcified structures called coccoliths. The coccoliths are formed inside the cells and released from the latter after formation. The light dependence of calcium incorporation in this species was studied using⁴⁵Ca as a tracer. Cells exposed to a repeating cycle of 16 h of light and 8 h of darkness incorporated calcium in extracellular coccoliths at a more or less constant rate throughout a cycle. The cells divided during the dark periods with a concomitant decrease in size. Their size increased during the light periods Coccolith formation in cells incubated in continuous darkness was greatly reduced and finally ceased. These cells did not divide and did not increase in size. Removal of extracellular coccoliths prior to the calcium incorporation experiments stimulated coccolith formation both in dark-incubated cells and in cells exposed to a repeating light-dark cycle. Cells in the stationary phase of growth ceased producing coccoliths. Calcification could be induced in these cells by removal of the extracellular coccoliths. Based on these findings we suggest that cells of Pleurochrysis carterae tend to produce a complete cover of coccoliths and that the available cell surface is a factor controlling coccolith formation.     

Novel detection of restriction fragment length polymorphisms in the human major histocompatibility complex

Cosmid genomic DNA clones have been used as hybridization probes in genomic Southern blot analysis to define restriction fragment length polymorphisms (RFLPs) in the major histocompatibility complex (MHC). Using 14 different enzymes and three overlapping cosmid clones we have detected six RFLPs in a 100 kilobase (kb) segment of DNA in the class III region extending centromeric of theTNFA gene towardHLA-DR. Four of the five RFLPs, defined using the enzymesTaqI,Rsa I,Hinc II, andHind III, and detected by the cosmid clone cosM7B, map to a 29 kb segment of DNA that includes all of the recently described G2 (BAT2) gene and a large portion of the 3 end of the G3 (BAT3) gene. The different RFLP variants were established by analyzing the DNA from three informative families and a panel of 51HLA-homozygous typing cell lines. CosM7B detectsTaq I variants of 4.3 kb, and 2.9 kb or 2.8 kb, Rsa I variants of 2.9 kb or 2.4 kb,Hinc II variants of 5.8 kb or 3.8 kb and 1.4 kb, and aHind III variant of 4.8 kb, while cosOT2 detects Taq I variants of 4.5 kb or 4 kb. The distribution of theRsa 1, Hinc II and Taq I RFLPs detected by cosM7B, and theTaq I RFLP detected with cosOT2, within the panel of cell line DNAs was assessed by Southern blotting. The 4.3 kbTaq I variant was observed in only one cell line with the extended haplotypeHLA-A29, C-, B44, SC30, DR4. The other RFLPs, however, occurred much more frequently. The 2.8 kb Taq I variant was observed in 20 % of haplotypes, the 2.9 kbRsa I variant was observed in 42% of haplotypes, and the 5.8 kbHinc I variant was observed in 12 % of haplotypes analyzed. The 4.5 kbTaq I variant detected by the overlapping cosmid cosOT2 was present in 21 % of haplotypes. Analysis of the RFLP variants with each other revealed seven different haplotypic combinations. Three of the haplotypic combinations were each subdivided into two subsets on the basis of the Nco I RFLP variant they carried at theTNF-B locus. These haplotypic combinations potentially allow differentiation among different extended haplotypes such asHLA-B8, SC01, DR3, HLA-B18, F1 C30, DR3, andHLA-B44, FC31, DR7. The RFLPs detected by the cosmid clones thus provide new tools which will be useful in the further genetic analysis of the MHC class III region.     

Effect of the MACS technique on rabbit sperm motility

Magnetic-activated cell sorting (MACS) separates apoptotic spermatozoa by the use of annexin V-conjugated nanoparticles which bind to phosphatidylserine that is externalized on the outer leaflet of the sperm plasma membrane. This technique yields two fractions: annexin V-negative (AnV⁻) and annexin V-positive (AnV⁺). The aim of the study was to evaluate the effect of MACS application on the motility parameters of rabbit spermatozoa. Rabbit semen samples collected separately from 4 bucks (I, II, III, and IV) were filtered and separated in a MACS system. The semen samples from a control (untreated) group, AnV⁻ and AnV⁺ fraction were evaluated using CASA system. The experiment was replicated 4 times for each buck. The AnV⁺ sperm had significantly lower concentration than the AnV⁻ fractions and the control samples (P<0.05 for bucks I, II, III, but not IV). We observed that the proportion of apoptotic spermatozoa in the semen of NZW bucks is about 20%. There was no significant difference in the percentage of motile and progressively motile spermatozoa between the AnVfractions and control samples. In conclusion, the MACS technique has no harmful effect on the rabbit sperm concentration and motility.     

Two new species of Isochrysis with remarks on the genus Ruttnera

Two marine strains of Chrysophyceae have been studied with the aid of light and electron microscopy. The benthic stages show definite similarities with two marine Gloeochrysis spp. (G. maritima and G. litoralis) described by Anand (1937), but the swarmers do not agree with the definition of this genus. The reason why they also cannot be assigned to the genus Ruttnera is discussed and their inclusion within the Isochrysidales is justified by the observation of haptophycean scales on the motile cells. The names Isochrysis maritima sp. nov. and Isochrysis litoralis sp. nov. are proposed in view of the fact that these strains might prove to be the species observed by Anand.