Phylogeny and Megasystematics of Phagotrophic Heterokonts (Kingdom Chromista)

Heterokonts are evolutionarily important as the most nutritionally diverse eukaryote supergroup and the most species-rich branch of the eukaryotic kingdom Chromista. Ancestrally photosynthetic/phagotrophic algae (mixotrophs), they include several ecologically important purely heterotrophic lineages, all grossly understudied phylogenetically and of uncertain relationships. We sequenced 18S rRNA genes from 14 phagotrophic non-photosynthetic heterokonts and a probable Ochromonas, performed phylogenetic analysis of 210–430 Heterokonta, and revised higher classification of Heterokonta and its three phyla: the predominantly photosynthetic Ochrophyta; the non-photosynthetic Pseudofungi; and Bigyra (now comprising subphyla Opalozoa, Bicoecia, Sagenista). The deepest heterokont divergence is apparently between Bigyra, as revised here, and Ochrophyta/Pseudofungi. We found a third universal heterokont signature sequence, and deduce three independent losses of ciliary hairs, several of 1-2 cilia, 10 of photosynthesis, but perhaps only two plastid losses. In Ochrophyta, heterotrophic Oikomonas is sister to the photosynthetic Chrysamoeba, whilst the abundant freshwater predator Spumella is biphyletic; neither clade is specifically related to Paraphysomonas, indicating four losses of photosynthesis by chrysomonads. Sister to Chrysomonadea (Chrysophyceae) is Picophagea cl. nov. (Picophagus, Chlamydomyxa). The diatom-parasite Pirsonia belongs in Pseudofungi. Heliozoan-like actinophryids (e.g. Actinosphaerium) are Opalozoa, not related to pedinellids within Hypogyristea cl. nov. of Ochrophyta as once thought. The zooflagellate class Bicoecea (perhaps the ancestral phenotype of Bigyra) is unexpectedly diverse and a major focus of our study. We describe four new biciliate bicoecean genera and five new species: Nerada mexicana, Labromonas fenchelii (=Pseudobodo tremulans sensu Fenchel), Boroka karpovii (=P. tremulans sensu Karpov), Anoeca atlantica and Cafeteria mylnikovii; several cultures were previously misidentified as Pseudobodo tremulans. Nerada and the uniciliate Paramonas are related to Siluania and Adriamonas; this clade (Pseudodendromonadales emend.) is probably sister to Bicosoeca. Genetically diverse Caecitellus is probably related to Anoeca, Symbiomonas and Cafeteria (collectively Anoecales emend.). Boroka is sister to Pseudodendromonadales/Bicoecales/Anoecales. Placidiales are probably divergent bicoeceans (the GenBank Placidia sequence is a basidiomycete/heterokont chimaera). Two GenBank ‘opalinid’ sequences are fungal; Pseudopirsonia is cercozoan; two previous GenBank ‘Caecitellus’ sequences are Adriamonas. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editior: Patnck J. Keeling]     

Synchroma grande spec. nov. (Synchromophyceae class. nov., Heterokontophyta): an amoeboid marine alga with unique plastid complexes

Chromist algae including the Heterokontophyta are supposed to have evolved monophyletically by secondary endosymbiosis from a eukaryotic host cell that engulfed a eukaryotic red alga. The red algal endosymbiont was then reduced to a secondary plastid surrounded by four enveloping membranes. On the basis of the amoeboid marine alga Synchroma grande gen. et spec. nov., the Synchromophyceae are described here as a new class of Heterokontophyta. Their taxonomic position is characterized by 18S rRNA and rbcL gene phylogenies, morphology, and pigment composition. The so far unique feature of the Synchromophyceae is the occurrence of conspicuous chloroplast complexes representing multiplastidic red secondary endosymbionts. In these remarkable secondary endosymbionts, several primary chloroplasts are aggregated in a common periplastidial compartment and are collectively enveloped by an additional outer membrane pair. The discovery of this novel plastid morphology is highly relevant for research on algal evolution and is discussed in terms of the postulated monophyletic origin of Chromista.     

Description of Babesia duncani n.sp. (Apicomplexa: Babesiidae) from humans and its differentiation from other piroplasms

The morphologic, ultrastructural and genotypic characteristics of Babesia duncani n.sp. are described based on the characterization of two isolates (WA1, CA5) obtained from infected human patients in Washington and California. The intraerythrocytic stages of the parasite are morphologically indistinguishable from Babesia microti, which is the most commonly identified cause of human babesiosis in the USA. Intraerythrocytic trophozoites of B. duncani n.sp. are round to oval, with some piriform, ring and ameboid forms. Division occurs by intraerythrocytic schizogony, which results in the formation of merozoites in tetrads (syn. Maltese cross or quadruplet forms). The ultrastructural features of trophozoites and merozoites are similar to those described for B. microti and Theileria spp. However, intralymphocytic schizont stages characteristic of Theileria spp. have not been observed in infected humans. In phylogenetic analyses based on sequence data for the complete18S ribosomal RNA gene, B. duncani n.sp. lies in a distinct clade that includes isolates from humans, dogs and wildlife in the western United States but separate from Babesia sensu stricto, Theileria spp. and B. microti. ITS2 sequence analysis of the B. duncani n.sp. isolates (WA1, CA5) show that they are phylogenetically indistinguishable from each other and from two other human B. duncani-type parasites (CA6, WA2 clone1) but distinct from other Babesia and Theileria species sequenced. This analysis provides robust molecular support that the B. duncani n.sp. isolates are monophyletic and the same species. The morphologic characteristics together with the phylogenetic analysis of two genetic loci support the assertion that B. duncani n.sp. is a distinct species from other known Babesia spp. for which morphologic and sequence information are available.     

Phylogenetic and morphological diversity of novel soil cercomonad species with a description of two new genera (Nucleocercomonas and Metabolomonas)

Cercomonads are important components of microbial food webs in soils and aquatic sediments. Here, we investigated the general morphology, behaviour, life cycle and 18S rDNA phylogeny of cercomonad cultures from a German grassland soil habitat. We describe ten new species including two new genera from 23 strains. Three Cercomonas, two Eocercomonas and three Paracercomonas species are described. Based on large phylogenetic distance and distinct morphology, we erect two novel clade B genera near the root of the cercomonad tree. Nucleocercomonas nov. gen. bears a number of characters unusual for cercomonads: Its anterior flagellum is extremely long, it mostly does not glide, and in its most frequent life stage the cell body does not attach to the substratum, but produces unattached pseudopodia. Furthermore, it has a unique nucleus with a peripheral nucleolus that attaches to the nuclear envelope opposite the basal body connection. Metabolomonas nov. gen. is extremely metabolic. It is characterized by a very high beating frequency of the anterior flagellum, fast gliding, rapid changes in shape and strong cytoplasmic streams. A new genus Brevimastigomonas is erected for the previously described species Paracercomonas anaerobica. The general morphology of cercomonad species often does not correspond with their phylogenetic position: closely related species may have a very different morphology.     

The symbiont Capsaspora owczarzaki,nov. gen. nov. sp., isolated from three strains of the pulmonate snail Biomphalaria glabrata is related to members of the Mesomycetozoea

While investigating the resistance of some strains of Biomphalaria glabrata to infection with Schistosoma mansoni, a unicellular eukaryotic symbiont was noted in the snail haemolymph. It was similar in appearance to Nuclearia sp. reported from B. glabrata. Sequences comprising the 18S, ITS1, 5.8S, ITS2 and the beginning of the 28S rDNA gene regions were obtained from symbionts isolated from three strains of B. glabrata, and compared with the same sequences obtained from a culture of Nuclearia sp. 18S rDNA sequences were identical for all four isolates. 18S rDNA sequences were used in a phylogenetic analysis to produce minimum evolution, maximum parsimony, maximum likelihood and Bayesian trees. All four analyses indicated that the B. glabrata symbiont is not closely related to Nuclearia but instead to the Mesomycetozoea, a recently recognised clade of symbiotic eukaryotes. Based on phylogenetic analysis, life history and morphological differences, the symbiont is described as a new genus and species, Capsaspora owczarzaki. Distinguishing characters are the presence of life cycle stage(s) that occur within snail haemolymph; ability to kill and ingest digenetic trematode larvae; ability to undergo asexual fission to produce daughter cells; absence of flagella, a mucous sheath and membranes containing chitin, elastin, or collagen; and presence of long unbranching pseudopodia and a penetration process. Using both polymerase chain reaction (PCR) and culturing techniques, the S. mansoni-resistant Salvador and 13-16-R1 strains were found to be significantly more likely to harbour the symbiont than the susceptible M line strain. Small but consistent sequence differences were noted among symbiont isolates from different snail strains, raising the possibility that the symbiont has diverged in different snail lineages. This suggests further that the symbiont is not restricted to albino lab-reared snails. A role, if any, of the symbiont in resistance awaits further study.     

Phylogenetic relationships of three new microsporidian isolates from the silkworm, Bombyx mori

The pathogenicity, mode of transmission, tissue specificity of infection and the small subunit rRNA (SSU-rRNA) gene sequences of the three new microsporidian isolates from the silkworm Bombyx mori were studied. Out of the three, NIK-2r revealed life cycle features and SSU-rRNA gene sequence similar to Nosema bombycis, suggesting that it is N. bombycis. The other two, NIK-4m and NIK-3h, differed from each other as well as from N. bombycis. NIK-4m was highly pathogenic and did not show any vertical transmission, in accordance with the apparent lack of gonadal infection, whereas NIK-3h was less pathogenic and vertical transmission was not detected but could not be excluded. Phylogenetic analysis based on SSU-rRNA gene sequence placed NIK-3h and NIK-4m in a distinct clade that included almost all the Vairimorpha species and Nosema species that infect lepidopteran and non-lepidopteran hosts, while NIK-2r was included in a clade containing almost all the Nosema isolates that infect only lepidopteran hosts. Thus, we have presented molecular evidence that one of the three isolates is in fact the type species N. bombycis, while the other two isolates are Vairimorpha spp. There was distinct separation of microsporidian isolates infecting only lepidopteran hosts and those infecting lepidopteran and non-lepidopteran hosts, reflecting possible co-evolution of hosts and microsporidian isolates.     

Retama species growing in different ecological–climatic areas of northeastern Algeria have a narrow range of rhizobia that form a novel phylogenetic clade within the Bradyrhizobium genus

Sixty-seven isolates were isolated from nodules collected on roots of Mediterranean shrubby legumes Retama raetam and Retama sphaerocarpa growing in seven ecological–climatic areas of northeastern Algeria. Genetic diversity of the Retama isolates was analyzed based on genotyping by restriction fragment length polymorphism of PCR-amplified fragments of the 16S rRNA gene, the intergenic spacer (IGS) region between the 16S and 23S rRNA genes (IGS), and the symbiotic genes nifH and nodC. Eleven haplotypes assigned to the Bradyrhizobium genus were identified. Significant biogeographical differentiation of the rhizobial populations was found, but one haplotype was predominant and conserved across the sites. All isolates were able to cross-nodulate the two Retama species. Accordingly, no significant genetic differentiation of the rhizobial populations was found in relation to the host species of origin. Sequence analysis of the 16S rRNA gene grouped the isolates with Bradyrhizobium elkanii, but sequence analyses of IGS, the housekeeping genes (dnaK, glnII, recA), nifH, and nodC yielded convergent results showing that the Retama nodule isolates from the northeast of Algeria formed a single evolutionary lineage, which was well differentiated from the currently named species or well-delineated unnamed genospecies of bradyrhizobia. Therefore, this study showed that the Retama species native to northeastern Algeria were associated with a specific clade of bradyrhizobia. The Retama isolates formed three sub-groups based on IGS and housekeeping gene phylogenies, which might form three sister species within a novel bradyrhizobial clade.     

Arcobacter molluscorum sp. nov., a new species isolated from shellfish

Nineteen bacteria isolates recovered from shellfish samples (mussels and oysters) showed a new and specific 16S rDNA-RFLP pattern with an Arcobacter identification method designed to recognize all species described up to 2008. These results suggested that they could belong to a new species. ERIC-PCR revealed that the 19 isolates belonged to 3 different strains. The sequence of the 16S rRNA gene of a representative strain (F98-3^T) showed 97.6% similarity with the closest species Arcobacter marinus followed by Arcobacter halophilus (95.6%) and Arcobacter mytili (94.7%). The phylogenetic analysis with the16S rRNA, rpoB, gyrB and hsp60 genes placed the shellfish strains within the same cluster as the three species mentioned (also isolated from saline habitats) but they formed an independent phylogenetic line. The DDH results between strain F98-3^T and A. marinus (54.8% ± 1.05), confirmed that it represents a new species. Several biochemical tests differentiated the shellfish isolates from all other Arcobacter species. Although the new species was different from A. mytili, they shared not only the same habitat (mussels) but also the characteristic of being so far the only Arcobacter species that are simultaneously negative for urea and indoxyl acetate hydrolysis. All results supported the classification of the shellfish strains as a new species, for which the name Arcobacter molluscorum sp. nov. with the type strain F98-3^T is proposed (=CECT 7696^T = LMG 25693^T).     

Two novel species representing a new clade and cluster of Phytophthora

Phytophthora stricta sp. nov. and Phytophthora macilentosa sp. nov. are described based on morphological, physiological and molecular characters in this study. Phytophthora stricta represents a previously unknown clade in the rRNA internal transcribed spacer (ITS)-based phylogeny. Phytophthora macilentosa, along with nine other species, consistently forms a high temperature-tolerant cluster within ITS clade 9. These observations are supported by the sequence analysis of the mitochondrial cytochrome c oxidase 1 gene. Both species are heterothallic and all examined isolates are A1 mating type. Phytophthora stricta produces nonpapillate and slightly caducous sporangia. This species is named after its characteristic constrictions on sporangiophores. Phytophthora macilentosa produces nonpapillate and noncaducous sporangia, which are mostly elongated obpyriform with a high length to breadth ratio. Both species were recovered from irrigation water of an ornamental plant nursery in Mississippi, USA and P. stricta was also recovered from stream water in Virginia, USA.     

Morphological and molecular phylogenetic analysis of two Saprolegnia sp. (Oomycetes) isolated from silver crucian carp and zebra fish

Two Saprolegnia isolates, JY isolated from silver crucian carp (Carassius auratus gibelio Bloch) and BMY isolated from zebra fish (Brachydanio rerio Hamilton) came from infections occurring concurrently in different locations in China. To confirm whether the two isolates were from the same Saprolegnia clone, comparative studies have been carried out based on their morphological, physiological and molecular characteristics. Observations showed that morphologically (both asexual and sexual organs) the two isolates were broadly similar and both isolates underwent repeated zoospore emergence. Comparing 704 base pairs of internal transcribed spacer (ITS) region and the 5.8S rDNA, we found isolates JY and BMY shared an identical ITS sequence with a minor variation (99.6 % similarity). Forty available sequences for representatives Saprolegnia spp. belonged to four phylogenetically separate clades. The two studied isolates fell within clade I that comprised a group of isolates which showed almost an identical ITS sequence but had been identified as a number of different morphological species. Our findings suggest that isolates JY and BMY appear to belong to the S. ferax clade and this clade (I) contains a number of closely related phylogenetic species. This is distinct from the more common fish pathogenic isolates, which belong to the S. parasitica clade (III) and are characterized by having cysts decorated by bundles of long hooked hairs and two further clades (II and IV) containing largely saprotrophic or soil born species.     

A newly-identified lineage of Schistosoma

Because of their role in causing schistosomiasis, flukes of the genus Schistosoma are the best known of all digeneans. The genus has traditionally been divided into four familiar species groups. Here we report on three poorly known species of Schistosoma, one of which, Schistosoma hippopotami, is known from the hippopotamus, one of which is provisionally identified as Schistosoma edwardiense, another hippo parasite, and a third that has not previously been described. All were collected from freshwater snails obtained from Lake Edward, western Uganda, the type locality for both known hippo schistosomes. The three different kinds of schistosome cercariae differ from one another in size, and all are readily differentiated by their long tail stems from the cercariae of human-infecting species. Furthermore, each was recovered from a different genus of snail host, Biomphalaria sudanica, Bulinus truncatus or Ceratophallus natalensis. Molecular analysis, based on 8350 bases of combined nuclear and mitochondrial DNA, groups these three long tail-stem cercariae into a well supported clade that does not associate with any of the recognised species groups. The placement of this clade, basal to all African species plus several Asian species, suggests that there has been an ancient association between Schistosoma and hippos. This new African Schistosoma clade advocates the need for further modification of the traditional species group-based classification. Two of the four species groups are paraphyletic. It also suggests that Schistosoma has been remarkably plastic with respect to adapting to snail hosts-three distantly related genera of planorbid snails have been exploited by worms within a single clade. Finally, it adds a new layer of complexity to deciphering the origins of Schistosoma, often considered to be African but recently challenged as being Asian. In the late Cenozoic the distribution of hippo species straddled both Africa and Asia and they may have provided a means for the introduction of blood flukes to Africa.     

Supermatrix data highlight the phylogenetic relationships of photosynthetic stramenopiles

Molecular data had consistently recovered monophyletic classes for the heterokont algae, however, the relationships among the classes had remained only partially resolved. Furthermore, earlier studies did not include representatives from all taxonomic classes. We used a five-gene (nuclear encoded SSU rRNA; plastid encoded rbcL, psaA, psbA, psbC) analysis with a subset of 89 taxa representing all 16 heterokont classes to infer a phylogenetic tree. There were three major clades. The Aurearenophyceae, Chrysomerophyceae, Phaeophyceae, Phaeothamniophyceae, Raphidophyceae, Schizocladiophyceae and Xanthophyceae formed the SI clade. The Chrysophyceae, Eustigmatophyceae, Pinguiophyceae, Synchromophyceae and Synurophyceae formed the SII clade. The Bacillariophyceae, Bolidophyceae, Dictyochophyceae and Pelagophyceae formed the SIII clade. These three clades were also found in a ten-gene analysis. The approximately unbiased test rejected alternative hypotheses that forced each class into either of the other two clades. Morphological and biochemical data were not available for all 89 taxa, however, existing data were consistent with the molecular phylogenetic tree, especially for the SIII clade.     

Molecular phylogenetic analyses based on the nuclear rRNA genes and the intron–exon structures of the nuSSU rRNA gene in Dictyocatenulata alba (anamorphic Ascomycota)

Molecular phylogenies inferred from the nuclear small subunit rRNA gene (nuSSU), nuclear large subunit rRNA gene D1/D2 region (nuLSU), and ITS-5.8S rRNA gene (ITS) indicated that five cultures of the lichenized hyphomycete Dictyocatenulata alba isolated from Japan form a monophyletic clade with high bootstrap support, and a close relationship to the Ostropomycetidae (Lecanoromycetes, Pezizomycotina, Ascomycota). Insertion sequences were found in the nuSSU of all isolates [e.g., nine insertions in the strain JCM 5358 (Japan Collection of Microorganisms)], some of which were group I introns. Five new insertion positions were found among the D. alba isolates. Using BLAST, none of the insertion sequences of D. alba were closely related to those of fungi or other organisms deposited in public DNA databases. Insertion positions were similar in some isolates, and two positions were common to all isolates. Although all phylogenetic analyses based on nuSSU, nuLSU, and ITS revealed the monophyly of D. alba, the isolates were divided into two (in the nuSSU tree) or three (in the nuLSU and ITS trees) groups. Based on the phylogenetic analyses and the intron–exon structures, the five isolates identified as D. alba belong to three cryptic species and therefore D. alba should be considered a species complex. The very slow-growing, tough agar colonies of the isolates, the occurrence of the species on both slightly lichenized and nonlichenized surfaces of trees, or pebbles (rarely on soil), suggest that the members of the D. alba complex may be lichenized. The photobiont was not clearly identified in this study.     

Revisiting the Phylogenetic Position of Synchroma grande

ABSTRACT. Two new classes Synchromophyceae and Picophagea, belonging to the heterokonts, have been proposed recently in separate studies of 18S rRNA phylogenies. Here we revise the 18S phylogeny of these classes by including all available sequenced species and applying Bayesian and maximum likelihood methods; Synchroma grande groups with the photophagotrophic Chlamydomyxa labyrinthuloides with high statistical support. This clade is sister to Chrysophyceae, together they share a common ancestry. Our results show that the creation of class Synchromophyceae by Horn et al. was premature, because they did not include data from the closely related C. labyrinthuloides and Picophagus flagellatus species. A revision of these classes should include additional species and most likely multigene phylogenies.     

Molecular phylogeny and phenotypic variability of clinical and environmental strains of Aspergillus flavus

Aspergillus flavus is the second most common cause of aspergillosis infection in immunocompromised patients and is responsible for the production of aflatoxins. Little is known about the population structure of A. flavus, although recent molecular and phenotypic data seem to demonstrate that different genetic lineages exist within this species. The aim of this study was to carry out a morphological, physiological, and molecular analysis of a set of clinical and environmental isolates to determine whether this variability is due to species divergence or intraspecific diversity, and to assess whether the clinical isolates form a separate group. The amdS and omtA genes were more phylogenetically informative than the other tested genes and their combined analysis inferred three main clades, with no clear distinction between clinical and environmental isolates. No important morphological and physiological differences were found between the members of the different clades, with the exception of the assimilation of d-glucosamine, which differentiates the members of the clade II from the others.     

Sinorhizobium americanum symbiovar mediterranense is a predominant symbiont that nodulates and fixes nitrogen with common bean (Phaseolus vulgaris L.) in a Northern Tunisian field

A total of 40 symbiotic bacterial strains isolated from root nodules of common bean grown in a soil located in the north of Tunisia were characterized by PCR-RFLP of the 16S rRNA genes. Six different ribotypes were revealed. Nine representative isolates were submitted to phylogenetic analyses of rrs, recA, atpD, dnaK, nifH and nodA genes. The strains 23C40 and 23C95 representing the most abundant ribotype were closely related to Sinorhizobium americanum CFNEI 156(T). S. americanum was isolated from Acacia spp. in Mexico, but this is the first time that this species is reported among natural populations of rhizobia nodulating common bean. These isolates nodulated and fixed nitrogen with this crop and harbored the symbiotic genes of the symbiovar mediterranense. The strains 23C2 and 23C55 were close to Rhizobium gallicum R602sp(T) but formed a well separated clade and may probably constitute a new species. The sequence similarities with R. gallicum type strain were 98.7% (rrs), 96.6% (recA), 95.8% (atpD) and 93.4% (dnaK). The remaining isolates were, respectively, affiliated to R. gallicum, E. meliloti, Rhizobium giardinii and Rhizobium radiobacter. However, some of them failed to re-nodulate their original host but promoted root growth.     

Phylogeny of Phaeomollisia piceae gen. sp. nov.: a dark, septate, conifer-needle endophyte and its relationships to Phialocephala and Acephala

Dark, septate endophytes (DSE) were isolated from roots and needles of dwarf Picea abies and from roots of Vaccinium spp. growing on a permafrost site in the Jura Mountains in Switzerland. Two of the isolates sporulated after incubation for more than one year at 4 °C. One of them was a hitherto undescribed helotialean ascomycete Phaeomollisia piceae gen. sp. nov., the other was a new species of Phialocephala, P. glacialis sp. nov. Both species are closely related to DSE of the Phialocephala fortinii s. lat.-Acephala applanata species complex (PAC) as revealed by phylogenetic analyses of the ITS and 18S rDNA regions. Morphologically dissimilar fungi, such as Vibrissea and Loramyces species, are phylogenetically also closely linked to the new species and the PAC. Cadophora lagerbergii and C. (Phialophora) botulispora are moved to Phialocephala because Phialocephala dimorphospora and P. repens are the closest relatives. Several Mollisia species were closely related to the new species and the PAC according to ITS sequence comparisons. One DSE from needles of Abies alba and one from shoots of Castanea sativa formed Cystodendron anamorphs in culture. Their identical 18S sequences and almost identical ITS sequences indicated Mollisia species as closest relatives, suggesting that Mollisia species are highly euryoecious.     

Aquastella gen. nov.: A new genus of saprolegniaceous oomycete rotifer parasites related to Aphanomyces, with unique sporangial outgrowths

The oomycete genus Aquastella is described to accommodate two new species of parasites of rotifers observed in Brooktrout Lake, New York State, USA. Three rotifer species – Keratella taurocephala, Polyarthra vulgaris, and Ploesoma truncatum – were infected, and this is the first report of oomycete infection in these species. Aquastella attenuata was specific to K. taurocephala and Aquastella acicularis was specific to P. vulgaris and P. truncatum. The occurrence of infections correlated with peak host population densities and rotifers were infected in the upper layers of the water column. Sequencing of 18S rRNA and phylogenetic analysis of both species placed them within the order Saprolegniales, in a clade closely related to Aphanomyces. The Aquastella species were morphologically distinct from other rotifer parasites as the developing sporangia penetrated out through the host body following its death to produce unique tapered outgrowths. Aquastella attenuata produced long, narrow, tapering, finger-like outgrowths, whilst A. acicularis produced shorter, spike-like outgrowths. We hypothesize that the outgrowths serve to deter predation and slow descent in the water column. Spore cleavage was intrasporangial with spore release through exit tubes. Aquastella attenuata produced primary zoospores, whereas A. acicularis released spherical primary aplanospores, more typical of other genera in the Aphanomyces clade.