PHYLOGENETIC AFFINITIES OF “CHANTRANSIA” STAGES IN MEMBERS OF THE BATRACHOSPERMALES AND THOREALES (RHODOPHYTA)1

This study evaluated the phylogenetic relationship among samples of “Chantransia” stage of the Batrachospermales and Thoreales from several regions of the world based on sequences of two genes—the plastid‐encoded RUBISCO LSU gene (rbcL) and the nuclear SSU ribosomal DNA gene (SSU rDNA). All sequences of “Chantransia macrospora” were shown to belong to Batrachospermum macrosporum based on both molecular markers, confirming evidence from previous studies. In contrast, nine species are now associated with “Chantransia pygmaea,” including seven species of the Batrachospermales and two of the Thoreales. Therefore, the presence of “C. macrospora” in a stream can be considered reliable evidence that it belongs to B. macrosporum, whereas the occurrence of “C. pygmaea” does not allow the recognition of any particular species, since it is associated with at least nine species. Affinities of “Chantransia” stages to particular taxa were congruent for 70.5% of the samples comparing the rbcL and SSU analyses, which were associated with the same or closely related species for both markers. Sequence divergences have been reported in the “Chantransia” stage in comparison to the respective gametophyte, and this matter deserves further attention.     

CONSPECIFICITY AND INDO-PACIFIC DISTRIBUTION OF SYMBIODINIUM GENOTYPES (DINOPHYCEAE) FROM GIANT CLAMS

We previously reported the occurrence of genetically‐diverse symbiotic dinoflagellates (zooxanthellae) within and between 7 giant clam species (Tridacnidae) from the Philippines based on the algal isolates' allozyme and random amplified polymorphic DNA (RAPD) patterns. We also reported that these isolates all belong to clade A of the Symbiodinium phylogeny with identical 18S rDNA sequences. Here we extend the genetic characterization of Symbiodinium isolates from giant clams and propose that they are conspecific. We used the combined DNA sequences of the internal transcribed spacer (ITS)1, 5.8S rDNA, and ITS2 regions (rDNA‐ITS region) because the ITS1 and ITS2 regions evolve faster than 18S rDNA and have been shown to be useful in distinguishing strains of other dinoflagellates. DGGE of the most variable segment of the rDNA‐ITS region, ITS1, from clonal representatives of clades A, B, and C showed minimal intragenomic variation. The rDNA‐ITS region shows similar phylogenetic relationships between Symbiodinium isolates from symbiotic bivalves and some cnidarians as does 18S rDNA, and that there are not many different clade A species or strains among cultured zooxanthellae (CZ) from giant clams. The CZ from giant clams had virtually identical sequences, with only a single nucleotide difference in the ITS2 region separating two groups of isolates. These data suggest that there is one CZ species and perhaps two CZ strains, each CZ strain containing individuals that have diverse allozyme and RAPD genotypes. The CZ isolated from giant clams from different areas in the Philippines (21 isolates, 7 clam species), the Australian Great Barrier Reef (1 isolate, 1 clam species), Palau (8 isolates, 7 clam species), and Okinawa, Japan (1 isolate, 1 clam species) shared the same rDNA‐ITS sequences. Furthermore, analysis of fresh isolates from giant clams collected from these geographical areas shows that these bivalves also host indistinguishable clade C symbionts. These data demonstrate that conspecific Symbiodinium genotypes, particularly clade A symbionts, are distributed in giant clams throughout the Indo‐Pacific.     

Variation of cassiicolin genes among Chinese isolates of <Emphasis Type="Italic">Corynespora cassiicola</Emphasis>

Corynespora cassiicola is a species of fungus that is a plant pathogen of many agricultural crop plants, including severe target spot disease on cucumber. Cassiicolin is an important effector of pathogenicity of this fungus. In this study, we collected 141 Corynespora isolates from eighteen hosts, and the casscolin gene was detected in 82 C. cassiicola strains. The deduced protein sequences revealed that 72 isolates contained the Cas2 gene, two strains from Gynura bicolor harboured the Cas2.2 gene, and 59 isolates without a cassiicolin gene were classified as Cas0. Phylogenetic analyses was performed for the 141 isolates using four loci (ITS, ga4, caa5, and act1) and revealed two genetic clusters. Cluster A is composed of four subclades: subcluster A1 includes all Cas2 isolates plus 18 Cas0 strains, subcluster A2 includes the eight Cas5 isolates and one Cas0 isolate, and subclusters A3 and A4 contain Cas0 strains. Cluster B consists of 21 Cas0 isolates. Twenty-two C. cassiicola strains from different toxin classes showed varying degrees of virulence against cucumber. Cas0 or Cas2 strains induced diverse responses on cucumber, from no symptoms to symptoms of moderate or severe infection, but all Cas5 isolates exhibited avirulence on cucumber.     

Phylogenetic Analysis of the Genus Bifidobacterium and Related Genera Based on 16S rDNA Sequences

The 16S rRNA gene sequences were determined for type strains of 21 Bifidobacterium species. A phylogenetic tree was constructed using the determined sequences and sequences from DNA databases, which contain the sequences of 11 type strains of Bifidobacterium species and 11 strains of related genera. All species of the genus Bifidobacterium and Gardnerella vaginalis ATCC 14018 belonged to a cluster phylogenetically distinct from the other genera. The cluster was divided into two subclusters: subcluster 1 composed of most species of Bifidobacterium and G. vaginalis, and subcluster 2 consisting of two species, B. denticolens and B. inopinatum; both of which were isolated from human dental caries. In the genus Bifidobacterium, four groups of species are known to be moderately to highly related by DNA-DNA hybridization. The four groups of species exhibited more than 99% similarity among their 16S rDNA sequences within each group. These results indicated that species with around 99% or more similarity in their 16S rDNA sequences should be confirmed for species identities.     

Emergence of carbapenem non‐susceptible multidrug resistant Acinetobacter baumannii strains of clonal complexes 103B and 92B harboring OXA‐type carbapenemases and metallo‐β‐lactamases in Southern India

The molecular epidemiology and carbapenem resistance mechanisms of clinical isolates of Acinetobacter baumannii obtained from a south Indian tertiary care hospital were investigated by repetitive extragenic palindromic sequence PCR (REP‐PCR) and multi‐locus sequence typing (MLST). Analysis of resistant determinants was achieved by PCR screening for the presence of genes encoding OXA‐carbapenemases, metallo‐β‐lactamases (MBLs) and efflux pumps. REP‐PCR generated around eight clusters of high heterogeneity; of these, two major clusters (I and V) appeared to be clonal in origin. Analysis of representative isolates from different clusters by MLST revealed that most of the isolates belonged to sequence type 103 of CC103^B. Second most prevalent ST belonged to clonal complex (CC) 92^B which is also referred to as international clone II. Most of the isolates were multi‐drug resistant, being susceptible only to polymyxin‐B and newer quinolones. Class D β‐lactamases such as bla_{OXA‐51‐like} (100%), bla_{OXA‐23‐like} (56.8%) and bla_{OXA‐24‐like} (14.8%) were found to be predominant, followed by a class B β‐lactamase, namely bla_IMP‐1 (40.7%); none of the isolates had bla_OXA‐58 like, bla_NDM‐1 or bla_SIM‐1. Genes of efflux‐pump adeABC were predominant, most of isolates being biofilm producers that were PCR‐positive for autoinducer synthase gene (>94%). Carbapenem non‐susceptible isolates were highly diverse and present throughout the hospital irrespective of type of ward or intensive care unit. Although previous reports have documented diverse resistant mechanisms in A. baumannii, production of MBL and OXA‐type of carbapenamases were found to be the predominant mechanism(s) of carbapenem resistance identified in strains isolated from Southern India.     

Differentiation of species in the Bacillus brevis group and the Bacillus aneurinolyticus group based on the electrophoretic whole-cell protein pattern

Ninety strains of eleven Bacillus species in the Bacillus brevis group and the Bacillus aneurinolyticus group were compared with the electrophoretic whole-cell protein pattern. The strains were separated into two clusters at the similarity of 55%. One cluster (cluster 1) was consisted of strains from the B. brevis group, and another cluster (cluster 2) was composed of strains from the B. aneurinolyticus group. The cluster 1 was separated into eight subclusters. Out of eight subclusters, seven subclusters contain strains from B. brevis, B. laterosporus, B. agri, B. reuszeri, B. choshinensis, B. formosus, and B. borstelensis. Another subcluster was further separated into two related clusters, which corresponded to B. centrosporus and B. parabrevis, and they were fused at the similarity of 76%. Cluster 2 was separated into two subclusters, which corresponded to B. aneurinolyticus and B. migulanus. The above eleven species showed characteristic patterns distinct from one another, and this correlated well with the published DNA relatedness data. The comparison of the electrophoretic whole-cell protein pattern proved to be useful for evaluation of taxonomic relationships among these taxa.     

MOLECULAR AND PHYLOGENETIC CHARACTERIZATION OF PHORMIDIUM SPECIES (CYANOPROKARYOTA) USING THE CPCB‐IGS‐CPCA LOCUS1

The accurate determination of species of Cyanoprokaryota/Cyanophyceae has many important applications. These include the assessment of risk with regard to blooms in water reservoirs as well as the identification of species capable of producing valuable bioactive compounds. Commonly, Cyanoprokaryota are classified based on their morphology. However, morphological criteria are not always reliable because they may change, for example, due to environmental factors. Thus, genetic and molecular analyses are a promising additional approach, but their application has so far been limited to relatively few genera. In light of this, we present here the first characterization of species and strains of the genus Phormidium Kütz. based on the cpcB‐IGS‐cpcA locus of the phycocyanin operon. In phylogenetic analyses using deduced amino acid sequences of the cpcB‐cpcA regions, Phormidium was found to be polyphyletic. This analysis appeared to be dominated by the cpcB region, which is characterized by a relatively high percentage of informative substitutions. The percentage of variable positions within the cpcB‐IGS‐cpcA locus overall was 16.5%, thereby indicating a level of divergence remarkably higher than that reported for Nodularia and Arthrospira in previous studies relying on cpcB‐IGS‐cpcA. Further, alignment of informative nucleotide substitutions in the cpcB‐IGS‐cpcA sequences revealed a mosaic distribution, which may be indicative of genetic recombination events. Finally, the length and sequences of the IGS region alone proved useful as markers to differentiate the cyanobacterial genus Phormidium. However, whether the IGS region per se is sufficiently discriminatory to differentiate between Phormidium species or even strains requires further investigation using newly identified Phormidium sequence data.     

REPORT OF A NEW INVASIVE ALGA IN THE ATLANTIC UNITED STATES: “HETEROSIPHONIA” JAPONICA IN RHODE ISLAND1

Recent collections of tetrasporangiate “Heterosiphonia” japonica Yendo from Watch Hill to Point Judith, Rhode Island, represent the first report of this nonnative alga in the western Atlantic. Native to the Pacific Ocean, this species was unintentionally introduced into European waters by 1984 and has subsequently invaded the eastern Atlantic Ocean widely from France to Norway and south into the Mediterranean Sea. Thus far, all western Atlantic collections of this species are confined to the outer coast of Rhode Island, and at present are not found in Narragansett Bay or in Long Island Sound along the Connecticut coast. Molecular and morphological studies confirm the identity of this newly introduced invasive species.     

ASSESSING PHYLOGENETIC AFFINITIES AND SPECIES DELIMITATIONS IN KLEBSORMIDIALES (STREPTOPHYTA): NUCLEAR‐ENCODED rDNA PHYLOGENIES AND ITS SECONDARY STRUCTURE MODELS IN KLEBSORMIDIUM,HORMIDIELLA, AND ENTRANSIA1

Nuclear‐encoded SSU, group I intron, and internal transcribed spacer (ITS) rDNA sequences were obtained for 16 strains of green algae representing species of Klebsormidium, Hormidiella attenuata, and Entransia fimbriata (for taxonomic authorities, see Table S1 in the supplementary material). The SSU phylogeny resolved a well‐supported clade Klebsormidiales in the Streptophyta that comprised authentic Klebsormidium isolates described recently in a monograph by G. M. Lokhorst and various strains from culture collections. The H. attenuata and En. fimbriata pair was the sister group of Klebsormidium. Certain isolates from culture collections previously identified as “Klebsormidium” emerged as Trebouxiophyceae. Strains assigned to Koliella, Gloeotila, and Stichococcus previously allied with Klebsormidium because of shared morphological and ultrastructural characteristics also belonged to Trebouxiophyceae. Group I introns inserted at Escherichia coli position 516 were found in K. nitens and SAG strain 384‐1, and at position 1506 in H. attenuata and En. fimbriata. Introns were not observed in other Klebsormidiales. Unambiguous alignment of ITS regions of Klebsormidiales was only possible after thermodynamic folding had predicted eight conserved helical domains. The ITS phylogeny provided support for five of the morphospecies recognized by Lokhorst (K. flaccidum, K. elegans, K. bilatum, K. crenulatum, K. mucosum), but the sequences of K. dissectum, K. fluitans, and K. nitens formed an unresolved clade. The species with the earliest origin in the Klebsormidium phylogeny was K. flaccidum. The incongruence between Lokhorst’s morphology‐based cladograms and the ITS phylogenies demonstrated the need for a critical reappraisal of the taxonomy and the morphological and molecular species concept in Klebsormidium on the basis of a more extensive taxonomic and geographic sampling strategy.     

COMPARATIVE MOLECULAR EVOLUTION OF NEWLY DISCOVERED PICOCYANOBACTERIAL STRAINS REVEALS A PHYLOGENETICALLY INFORMATIVE VARIABLE REGION OF β‐PHYCOERYTHRIN1

The genetic diversity and phylogenetic position of 10 strains of picocyanobacteria from the Arabian Sea were examined using partial sequences from three loci: 16S rDNA, RNA polymerase rpoC1, and two elements of the phycoerythrin (PE) locus, cpeA and cpeB which encode for the α and β subunit of PE. Nine of the strains showed nearly identical spectral phenotypes based on the in vivo excitation spectrum for PE fluorescence emission and appear to be strains synthesizing a phycourobilin (PUB)–lacking PE. These strains include one, Synechococcus sp. G2.1, already known to be closely related to filamentous cyanobacteria and not to the commonly studied 5.1 subcluster of marine Synechococcus. The 10th strain was a PE‐lacking strain that was of interest because it was isolated from open‐ocean conditions where picocyanobacteria with this phenotype are relatively uncommon. Phylogenetic analysis of the concatenated 16S rDNA and rpoC1 data sets showed that none of the previously described strains were members of the 5.1 subcluster of marine Synechococcus, nor were they closely related to strain G2.1. Instead, they form a well‐supported and previously undescribed clade of cyanobacteria that is sister to Cyanobium. Thus, these strains represent the first PE‐containing Cyanobium from oceanic waters, and the lineage they define includes a strain with a PE‐lacking phenotype from the same environment. Analysis of the PE sequence data showed the PE apoprotein has evolved independently in the G2.1 lineage and the Cyanobium‐like lineage represented by the study strains. It also revealed a hypervariable region of the β‐subunit not described previously; variation in this region shows a pattern among a wide range of PE‐containing organisms congruent with the phylogenetic relationships inferred from other genes. This suggests that the PUB‐lacking spectral phenotype is more likely to have evolved in distantly related phylogenetic lineages by either divergent or convergent evolution than by lateral gene transfer. Both the conserved PE gene sequences and the inferred amino acid sequences for the hypervariable region show considerable divergence among Prochlorococcus PEs, red algal PEs, PUB‐containing PEs from the marine Synechococcus 5.1 subcluster, PEs from the Cyanobium‐like strains, and PEs from other cyanobacteria (including strain G2.1). Thus, it appears that the hypervariable region of the PE gene can be used as a taxon‐specific marker.     

Rapid detection of “Candidatus Phytoplasma mali” by recombinase polymerase amplification assays

Isothermal recombinase polymerase amplification (RPA) assays for the specific detection of “Candidatus Phytoplasma mali (Ca. P. mali),” the causal agent of apple proliferation, were developed. The assays amplify a fragment of the imp gene and amplimers were detected either by fluorescence in real‐time mode (TwistAmp^®exo assay) using a fluorophore‐labelled probe or by direct visualization employing a lateral flow device (TwistAmp^®nfo assay/Milenia^®HybriDetect). The RPA assays specifically amplified DNA from “Ca. P. mali” strains, and cross‐reactivity with other phytoplasmas or plant DNA was not observed. The limit of detection was determined with a cloned imp standard, and positive results were obtained down to 10 copies with both RPA assay formats. In comparison with a TaqMan real‐time PCR assay based on the same target gene, the RPA assays were equally sensitive, but results were obtained faster. Simplified nucleic acid extraction procedures from plant tissue with Tris‐ and CTAB‐based buffers revealed that crude Tris–DNA extracts were a suitable source for RPA tests while larger concentrations of CTAB were inhibitory. This is the first report of RPA‐based assays for the detection of “Ca. P. mali”. The assays are suitable for high‐throughput screening of plant material and point‐of‐care diagnostic and can be potentially combined with a simplified DNA extraction procedure.     

THE CONTRIBUTION OF SUB‐SAHARAN AFRICAN STRAINS TO THE PHYLOGENY OF CYANOBACTERIA: FOCUSING ON THE NOSTOCACEAE (NOSTOCALES,CYANOBACTERIA)1

To date, phylogenies have been based on known gene sequences accessible at GenBank, and the absence of many cyanobacterial lineages from collections and sequence databases has hampered their classification. Investigating new biotopes to isolate more genera and species is one way to enrich strain collections and subsequently enhance gene sequence databases. A polyphasic approach is another way of improving our understanding of the details of cyanobacterial classification. In this work, we have studied phylogenetic relationships in strains isolated from freshwater bodies in Senegal and Burkina Faso to complement existing morphological and genetic databases. By comparing 16S rDNA sequences of African strains to those of other cyanobacteria lineages, we placed them in the cyanobacterial phylogeny and confirmed their genus membership. We then focused on the Nostocaceae family by concatenated analysis of four genes (16S rDNA, hetR, nifH, and rpoC1 genes) to characterize relationships among Anabaena morphospecies, in particular, Anabaena sphaerica var. tenuis G. S. West. Using a polyphasic approach to the Nostocaceae family, we demonstrate that A. sphaerica var. tenuis is more closely related to Cylindrospermospsis/Raphidiopsis than to other planktonic Anabaena/Aphanizomenon. On the basis of phylogeny and morphological data, we propose that these three significantly different clusters should be assigned to three genera.     

Phylogenetic relationships among <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> (Bacillaceae: Bacillales) strains based on a comparison of SSU rRNA sequences and genome profiling

Bacillus thuringiensis Berliner has previously been classified via the serological identification of flagellar antigens. However, the phylogenetic relationships among strains of B. thuringiensis cannot be investigated by serotyping. Furthermore, high levels of homology have been found in gene sequences among various strains, complicating the determination of their evolutionary relationships. In order to elucidate the phylogenetic relationships within B. thuringiensis, we analyzed 40 strains belonging to typical serotypes using two approaches: an analysis of small subunit (SSU) rRNA sequences and genome profiling (GP) based on temperature gradient gel electrophoresis of random PCR products. The SSU rRNA analysis resulted in all 40 strains forming a single cluster with Bacillus cereus Frankland & Frankland. The distances among the subclusters were too small to further classify the strains. On the other hand, the phylogenetic analysis based on GP resulted in three clusters of B. thuringiensis strains. These results suggest that GP is a better method for the determination of phylogenetic relationships within B. thuringiensis.     

Chemotaxonomy of Trichoderma spp. Using mass spectrometry-based metabolite profiling

In this study, seven Trichoderma species (33 strains) were classified using secondary metabolite profile-based chemotaxonomy. Secondary metabolites were analyzed by liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS-MS) and multivariate statistical methods. T. longibrachiatum and T. virens were independently clustered based on both internal transcribed spacer (ITS) sequence and secondary metabolite analyses. T. harzianum formed three subclusters in the ITS-based phylogenetic tree and two subclusters in the metabolitebased dendrogram. In contrast, T. koningii and T. atroviride strains were mixed in one cluster in the phylogenetic tree, whereas T. koningii was grouped in a different subcluster from T. atroviride and T. hamatum in the chemotaxonomic tree. Partial least-squares discriminant analysis (PLS-DA) was applied to determine which metabolites were responsible for the clustering patterns observed for the different Trichoderma strains. The metabolites were hetelidic acid, sorbicillinol, trichodermanone C, giocladic acid, bisorbicillinol, and three unidentified compounds in the comparison of T. virens and T. longibrachiatum; harzianic acid, demethylharzianic acid, homoharzianic acid, and three unidentified compounds in T. harzianum I and II; and koninginin B, E, and D, and six unidentified compounds in T. koningii and T. atroviride. The results of this study demonstrate that secondary metabolite profiling-based chemotaxonomy has distinct advantages relative to ITSbased classification, since it identified new Trichoderma clusters that were not found using the latter approach.     

GENETIC VARIABILITY AND MOLECULAR PHYLOGENY OF DINOPHYSIS SPECIES (DINOPHYCEAE) FROM NORWEGIAN WATERS INFERRED FROM SINGLE CELL ANALYSES OF rDNA1

The objectives of this study were to determine rDNA sequences of the most common Dinophysis species in Scandinavian waters and to resolve their phylogenetic relationships within the genus and to other dinoflagellates. A third aim was to examine the intraspecific variation in D. acuminata and D. norvegica, because these two species are highly variable in both morphology and toxicity. We obtained nucleotide sequences of coding (small subunit [SSU], partial large subunit [LSU], 5.8S) and noncoding (internal transcribed spacer [ITS]1, ITS2) parts of the rRNA operon by PCR amplification of one or two Dinophysis cells isolated from natural water samples. The three photosynthetic species D. acuminata, D. acuta, and D. norvegica differed in only 5 to 8 of 1802 base pairs (bp) within the SSU rRNA gene. The nonphotosynthetic D. rotundata (synonym Phalacroma rotundatum[Claparède et Lachmann] Kofoid et Michener), however, differed in approximately 55 bp compared with the three photosynthetic species. In the D1 and D2 domains of LSU rDNA, the phototrophic species differed among themselves by 3 to 12 of 733 bp, whereas they differed from D. rotundata by more than 100 bp. This supports the distinction between Dinophysis and Phalacroma. In the phylogenetic analyses based on SSU rDNA, all Dinophysis species were grouped into a common clade in which D. rotundata diverged first. The results indicate an early divergence of Dinophysis within the Dinophyta. The LSU phylogenetic analyses, including 4 new and 11 Dinophysis sequences from EMBL, identified two major clades within the phototrophic species. Little or no intraspecific genetic variation was found in the ITS1–ITS2 region of single cells of D. norvegica and D. acuminata from Norway, but the delineation between these two species was not always clear.     

East Tibetan Lakes Harbour Novel Clusters of Picocyanobacteria as Inferred from the 16S–23S rRNA Internal Transcribed Spacer Sequences

Planktonic picocyanobacteria abundance and diversity were investigated in nine lakes on the East Tibetan Plateau spanning a salinity gradient of 0.4–22.6 g l⁻¹. The investigation was conducted using epifluorescence microscopy (EFM) and terminal restriction fragment polymorphism analysis of 16S–23S rRNA internal transcribed spacer (ITS) PCR amplicons followed by sequence analyses of large ITS clone libraries of seven selected samples. EFM showed that picocyanobacteria comprised 7–19% of the total prokaryotic cells found in surface water. Most of the clones were classified into six clusters and grouped within the “picocyanobacterial clade”, which consists exclusively of freshwater Synechococcus. Four new phylogenetic clusters and one new subcluster of Synechococcus spp. were found, none of which are members of any known picocyanobacterial clusters. The new clusters and subcluster were the most abundant picocyanobacteria (about 96% of the sequences) in the samples collected. Sequence analyses indicated that members of the four new Synechococcus groups were only found in freshwater lakes (<1.0 g l⁻¹ of total dissolved solid), while members of the new subcluster were found in all the investigated Tibetan lakes, over a large salinity gradient of 0.4–22.6 g l ⁻¹. This suggests that there is ecologically significant microdiversity within the observed Synechococcus group as defined by ITS sequences. Collectively our study demonstrated abundant and potentially novel Synechococcus in East Tibetan lakes that are likely the result of evolutionary adaptations to regional conditions.     

IDENTIFICATION AND ASSESSMENT OF DOMOIC ACID PRODUCTION IN OCEANIC PSEUDO‐NITZSCHIA (BACILLARIOPHYCEAE) FROM IRON‐LIMITED WATERS IN THE NORTHEAST SUBARCTIC PACIFIC1

We identified and investigated the potential toxicity of oceanic Pseudo‐nitzschia species from Ocean Station Papa (OSP), located in a high‐nitrate, low‐chlorophyll (HNLC) region of the northeast (NE) subarctic Pacific Ocean. Despite their relatively low abundances in the indigenous phytoplankton assemblage, Pseudo‐nitzschia species richness is high. The morphometric characteristics of five oceanic Pseudo‐nitzschia isolates from at least four species are described using SEM and TEM. The species identified are Pseudo‐nitzschia dolorosa Lundholm et Moestrup, P. granii Hasle, P. heimii Manguin, and P. cf. turgidula (Hust.) Hasle. Additional support for the taxonomic classifications based on frustule morphology is provided through the sequencing of the internal transcribed spacer 1 (ITS1) rDNA. Pseudo‐nitzschia species identification was also assessed by the construction of ITS1 clone libraries and using automated ribosomal intergenic spacer analysis (ARISA) for environmental samples collected during the Subarctic Ecosystem Response to Iron Enrichment Study (SERIES), conducted in close proximity to OSP in July of 2002. Based on ITS1 sequences, the presence of P. granii, P. heimii, P. cf. turgidula, and at least five other putative, unidentified Pseudo‐nitzschia ITS1 variants was confirmed within iron‐enriched phytoplankton assemblages at OSP. None of the oceanic isolates produced detectable levels of particulate domoic acid (DA) when in prolonged stationary phase due to silicic acid starvation. The lack of detectable concentrations of DA suggests that either these strains produce very little or no toxin, or that the physiological conditions required to promote particulate DA production were not met and thus differ from their coastal, toxigenic congeners.