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This paper explores the diversity and taxonomy of species within Fragilaria sensu stricto, an abundant and ecologically important diatom genus, taking advantage of cultured and DNA ‐barcoded material. The goal is to facilitate the identification of European taxa within this complex, providing a unified view on morphological and molecular diversity. There is a general agreement that the separation of species within the group of Fragilaria is difficult because morphological descriptions of species are not consistent between authorities, ongoing taxonomic revisions have resulted in species described with standards of the late 20th and 21st centuries alongside descriptions based on 19th century (light microscopical) criteria, and because not all diagnostic characters can be seen in all specimens encountered in routine analyses. Consequent confusion could blur potentially important ecological distinctions between species. Our study demonstrated that some species defined on morphological criteria could be confirmed using the rbc L chloroplast gene as a genetic marker, for example, Fragilaria gracilis , Fragilaria tenera , Fragilaria perminuta , and Fragilaria subconstricta . However, even for those species, preliminary identifications based on morphology often differed from identifications based on phylogenetic clustering combined with detailed morphological study. Clades were well‐defined by rbc L, but based on morphology, the terminal taxa of these clades did not match the currently described Fragilaria species. To clarify recognition of these taxa, we describe three new species: Fragilaria agnesiae , Fragilaria heatherae , and Fragilaria joachimii . info:eu-repo/semantics/acceptedVersion

A previously unknown member of the Bacillariaceae was discovered almost simultaneously in four different brackish coastal wetlands on the Atlantic and Mediterranean coasts of the Iberian Peninsula. It appears to tolerate a wide range of salinities but was never common in samples where it occurred. The frustules were consistently hantzschioid (i.e. with the raphe systems always on the same side of the frustule) and the valve outline was asymmetrical about the apical plane, two features that have until recently been considered characteristic of Hantzschia. Molecular phylogenies based on rbcL and LSU rDNA indicated, however, that the new species does not belong in Hantzschia but among the several disparate lineages that comprise the paraphyletic genus Nitzschia. This finding, coupled with the recent discovery of other diatoms with constant hantzschioid symmetry but with a morphology very similar to the type species of Nitzschia, is discussed in relation to the status and characterization of Hantzschi...

Diatoms (Bacillariophyta) are ubiquitous microalgae which produce a siliceous exoskeleton and which make a major contribution to the productivity of oceans and freshwaters. They display a huge diversity, which makes them excellent ecological indicators of aquatic ecosystems. Usually, diatoms are identified using characteristics of their exoskeleton morphology. DNA-barcoding is an alternative to this and the use of High-Throughput-Sequencing enables the rapid analysis of many environmental samples at a lower cost than analyses under microscope. However, to identify environmental sequences correctly, an expertly curated reference library is needed. Several curated libraries for protists exists; none, however are dedicated to diatoms. Diat.barcode is an open-access library dedicated to diatoms which has been maintained since 2012. Data come from two sources (1) the NCBI nucleotide database and (2) unpublished sequencing data of culture collections. Since 2017, several experts have collaborated to curate this library for rbcL, a chloroplast marker suitable for species-level identification of diatoms. For the latest version of the database (version 7), 605 of the 3482 taxonomical names originally assigned by the authors of the rbcL sequences were modified after curation. The database is accessible at https://www6.inra.fr/carrtel-collection_eng/Barcoding-database.

Spermatogenesis and auxospore development were studied in the freshwater centric diatom Hydrosera triquetra. Spermatogenesis was unusual, lacking depauperating cell divisions within the spermatogonangium. Instead, a series of mitoses occurred within an undivided cell to produce a multinucleate plasmodium with peripheral nuclei, which then underwent meiosis. 32 or 64 sperm budded off from the plasmodium leaving a large residual cell containing all the chloroplasts. Similar development apparently occurs in Pleurosira, Aulacodiscus, and Guinardia, these being so distantly related that independent evolution of plasmodial spermatogenesis seems likely. After presumed fertilization, the Hydrosera egg cell expanded distally to form a triangular end part. However, unlike in other triangular diatoms (Lithodesmium, Triceratium), the development of triradiate symmetry was not controlled by the "canonical" method of a perizonium that constrains expansion to small terminal areas of the auxospore wall. Instead, the auxospore wall lacked a perizonium and possessed only scales and a dense mat of thin, apparently entangled strips of imperforate silica. No such structures have been reported from any other centric diatoms, the closest analogs being instead the incunabular strips of some raphid diatoms (Nitzschia and Pinnularia). Whether these silica structures are formed by the normal method (intracellular deposition within a silica deposition vesicle) is unknown. As well as being more rounded than vegetative cells, the initial cell is aberrant in its structure, since it has a less polarized distribution of the "triptych" pores characteristic of the species.

The large diatom genus Nitzschia and the morphologically similar Hantzschia are currently distinguished on the basis of cell symmetry and division. In Hantzschia, the eccentrically placed raphe systems of the two valves are always on the same side of the frustule (‘hantzschioid’ symmetry), never diagonally opposite (‘nitzschioid’ symmetry). Although some Nitzschia species produce hantzschioid cells, it has previously been thought that these cells never breed true but divide to give one nitzschioid daughter and one hantzschioid daughter. Sublittoral marine epipelon from Loch Goil, W Scotland, contained three undescribed species of Bacillariaceae whose cells were always hantzschioid, but which possessed a valve structure (silica flaps on either side of the raphe, no central raphe endings, a slit-like entrance to the raphe canal that narrows towards the poles, and discrete bar-like fibulae) linking them to N. spathulata and the type species of Nitzschia, N. sigmoidea. They also agreed with these species in c...

Many diatom and other microbial eukaryote morphospecies consist of a variable number of (pseudo)cryptic species, with obvious consequences for such fields as biogeography and community ecology. Here, we investigated the species limits of morphologically similar small–celled strains of the model diatom Sellaphora from the United Kingdom and Australia, using cox1 mitochondrial and rbcL chloroplast gene sequences. Based on cox1 sequence data, the sequenced strains belonged to six closely related lineages, presumably species, of which one corresponds to the previously described S. auldreekie D.G. Mann & S.M. McDonald. Although rbcL displayed less sequence variation, the same six lineages were also recovered in an rbcL phylogeny of the genus. Molecular species discovery was compared to mating trials involving three of the lineages, showing that they were reproductively isolated. Incomplete evidence from a fourth lineage suggested that it too was reproductively isolated. A posteriori examination of light microscope morphology revealed no simple metrics or presence/absence characters that could consistently separate all species of the auldreekie complex, even though some do differ in pole width or stria density. While it is premature to make conclusions about their biogeography, it is obvious that a number of cryptic Sellaphora species thus far undetected in the UK are easily found at several localities in warm–temperate Australia.

Amphoroid diatoms are recognized in the light microscope by their strongly dorsiventral frustules; their raphe system lies close to the ventral margin of the valve, and their girdle is much wider on the dorsal side. Historically, these diatoms were classified into the genus Amphora, although heterogeneity has long been pointed out based mainly on frustule fine structure, chloroplast arrangement, and molecular phylogeny. In this study detailed observations were undertaken on the frustule of an amphoroid species Amphora commutata. Distinct features of this species were the hyaline ventral side of the valve and hitherto unknown apical pores. The ontogeny of the valve and girdle bands was also reported. A molecular phylogenetic analysis of Amphora species, including representatives of the subgenera Amphora, Oxyamphora and Diplamphora, was performed, based on nuclear encoded small subunit (SSU) rDNA. The best tree showed paraphyly of amphoroid diatoms and also suggested that the subgenus Diplamphora, which inc...

We give a detailed account of sexual reproduction and auxospore development in the diatom genus Diploneis, principally from clonal cultures of the marine benthic Diploneis papula. Sexual reproduction of D. papula was apparently homothallic. After pairing side to side, cells entered meiosis, and each gametangium produced two gametes. Fertilization was physiologically anisogamous, and both gametes of one gametangium were active and those of the other gametangium passive so that the two zygotes were formed within or close to the ‘passive’ gametangium. Each gamete contained a single chloroplast. The zygote became surrounded by delicate incunabula that contained circular or elliptical scales; this confirmed recent observations that scales (or apparently homologous strips or plates) are quite commonly formed by the zygotes of raphid diatoms. Subsequently, a robust transverse perizonium was built up as the auxospore expanded, and the perizonium was comprised of a closed primary band at the centre and open secondary bands towards each pole. The ends of the secondary bands, which met along one side of the auxospore to form a suture as in other pennate diatoms, were curved inwards towards the centre of the auxospore. Novel features were the extension of the transverse perizonium to cover the tips of the auxospores (in other diatoms it stopped short of the poles, leaving an apical dome covered only by the incunabula) and a differentiation of the ends of the transverse bands into truncated and prolonged variants, which alternated along the suture. The longitudinal perizonium possessed the same highly conserved configuration as in other pennate diatoms, with a wide bifacial central band flanked by two other bands, which here differed in shape and structure. All the perizonial bands were fimbriate. The possible homology of the longitudinal perizonium to the thecae of vegetative cells was discussed. During formation of the initial cell, the protoplast contracted away from the perizonium, but the latter nevertheless moulded the outline shape of the initial valves, which were more strongly constricted than preauxospore cells and gametangia. Other Diploneis species produced either one or two gametes per gametangium.

Paedogamy (fusion of gametes produced within the same gametangium following meiosis) has rarely been reported in diatoms, with fewer than 10 confirmed examples. One of these, reported by L. Geitler, was in a diatom from Illmitz, Lake Neusiedl (Austria), identified as ‘Nitzschia frustulum var. perpusilla'. We observed uniparental auxosporulation in two Nitzschia clones isolated from the lower Ebro River (Catalonia, Spain), morphologically similar to Geitler's material and belonging to the N. inconspicua species complex. We established that the auxospores were formed paedogamously by Feulgen staining of the nuclei and time-lapse microscopy of living cells. However, reinvestigation of Geitler's original cytological preparations revealed differences between the Illmitz and Ebro material with respect to the length of the initial cells, the structure of the perizonium, and the timing of degeneration of superfluous haploid nuclei during gametogenesis, indicating a genetic and possibly a taxonomic separation. Sca...

The taxonomy of several small-celled, ecologically significant Nitzschia species, which are frequently confused with each other or whose names are misapplied, is clarified. Following an examination of type material and modern samples by light and electron microscopy, it was concluded that N. frustulum (Kutzing) Grunow, N. inconspicua Grunow, N. soratensis E. Morales & Vis and N. invisitata Hustedt are independent species. No morphological basis was found for separating N. frustulum var. subsalina Hustedt or N. boliviana E. Morales & Vis from N. inconspicua and they are therefore placed in synonymy with N. inconspicua. Nitzschia soratensis, described recently from Bolivia, has previously been misidentified in Europe, either as N. inconspicua (from which it differs most obviously in having more bluntly rounded poles, striae within the raphe canal that are composed of triplets, and fibulae that can be seen in light microscopy to widen at their bases) or as N. abbreviata Hustedt ex Simonsen (from which it dif...

Uniparental auxosporulation was observed in a monoclonal culture of a Sellaphora clone isolated from the epipelon of a fishpond in the Czech Republic. The cox1 sequence for the clone confirmed that it belonged to the Sellaphora pupula-bacillum species complex but showed significant differences from all previously characterized Sellaphora species, and it is therefore described as S. marvanii sp. nov. Protoplast, valve, and girdle structure resembled those of other Sellaphora species, but a novel finding for all diatoms was a change in girdle structure during the life cycle: the most advalvar girdle band (valvocopula) bore a single line of pores in enlarged postauxospore cells but was entirely plain in small cells and gametangia. The young auxospores were covered by incunabula containing large, delicate, ± circular scales, resembling those of centric diatom auxospores; similar scales have been reported in a few other raphid diatoms (Pseudo-nitzschia multiseries, Diploneis sp.) but contrast with the strip incunabula of some Nitzschia and Pinnularia and the helmet-like caps of Neidium. The scales persisted during auxospore expansion, mostly as two caps over the auxospore poles. The transverse perizonium comprised a very wide, closed primary band, flanked by numerous secondary bands whose open ends were strongly incurved toward the center. Initial valves were differentiated from their immediate descendants by the very strong external demarcation of the raphe sternum, irregular shape, and curved transapical profile.

SUMMARY The effects of salinity on the growth and valve morphology of five benthic estuarine diatoms (Nitzschia pusilla, N. frustulum, N. palea, N. filiformis var. conferta and Eolimna subminuscula), isolated from both freshwater and brackish/marine habitats, were investigated. The four Nitzschia strains grew well over a broad salinity range, though some (N. pusilla, N. frustulum) showed a broader salinity range tolerance (from fully saline down to at least 9.5 ppt) than others (N. palea, N. filiformis var. conferta had reduced growth at salinities of 16 ppt and above). Salinity significantly affected the valve morphology of the five strains studied. However, there was no consistent pattern in either the morphological characters affected or the direction of the effects. Although significant, the effects of salinity on valve morphology were very small and therefore it seems that the taxonomic usefulness of some of the classical taxonomical characters is not undermined.

Cox1 has been suggested as a barcode marker for diatoms but it has not been tested intensively in any group of closely related species outside Sellaphora. We evaluated the use of cox1, LSU and rbcL for phylogeny and identification in the taxonomically problematic but ecologically important freshwater diatom Nitzschia palea, for which LSU rDNA sequences, mating and morphological data had already been published, and for which DNA and/or clonal isolates were still available; some new isolates were added. The previous and new information concur in suggesting that N. palea is a complex of several or many species. Where cox1 sequences could be obtained, they were more variable than LSU and rbcL and discriminated between lineages that differed in their morphologies, mating compatibilities, LSU sequences or rbcL sequences. Repeated failures for some strains to recover cox1 sequences from DNA that yielded LSU and rbcL suggest that cox1 will be impractical as a universal barcode marker in diatoms until better prime...

Mann D.G. and PoulIckova A. 2010. Mating system, auxosporulation, species taxonomy and evidence for homoploid evolution in Amphora (Bacillariophyta). Phycologia 49: 183–201. DOI: 10.2216/09-08.1 Cytological characteristics of the mitotic cycle, sexual reproduction and auxospore formation are described for three members of the type group of Amphora: A. ovalis, A. copulata and A. minutissima. All have a single lobed ventral chloroplast, nuclei with granular heterochromatin and a single nucleolus (unusually large in A. ovalis). Amphora copulata is heterothallic, with two mating types that do not differ in gamete behaviour. All three species reproduce allogamously via fusion of two rearranged gametes per gametangium to produce elongate zygotes enclosed within the volume delimited by the gametangial thecae. Pairing is strictly via the ventral sides of the cells, with tight apposition of the cells, and expansion of the auxospores is strictly perpendicular to the gametangium long axes. Comparisons are m...

Several new diatom taxa have been described from nearly pristine habitats in New Zealand, but the new species Tabularia variostriata, described here, was abundant in a Wellington City suburban stream. It has typical features of Tabularia species: internally its pores are sunk down well below prominent short ribs, and externally its pores are occluded by cribra. It differs from other Tabularia species in having uniseriate striae with variable numbers of pores, the striae forming irregular edges to a wide sternum. The innermost pores on the longer striae appear enlarged as spacing of pores in striae increases near the edge of the sternum. The Australasian freshwater diatom Eunophora berggrenii, formerly known as Amphora berggrenii Cleve or ‘Eunophora sp. 1’, was studied and a lectotype was designated. Tasmanian specimens differed subtly from the type material in valve dimensions and may represent a different but closely related species.

Previous molecular genetic and life cycle studies have revealed the existence of a vigorously homothallic deme within the Sellaphora pupula species complex, which was given the provisional name of Sellaphora [pupula K-LB] Φ 'urban elliptical'. This is described here as S. bisexualis sp. nov., with a cox1 DNA-barcoded type from Scotland. A detailed description of frustule ultrastructure is given, the most complete for a Sellaphora species. The morphology of the terminal raphe fissure is variable, becoming vestigial in some small pre-auxospore valves, and the boundary between the raphe-sternum and striae is sometimes strongly, sometimes weakly marked by larger areolae and an external groove. The cingulum contains at least three plain bands. The pars interior of band 1 (the valvocopula), as well as being undulate with a pitch corresponding to the transapical ribs, bears a very fine fringe of short fimbriae. Sexual reproduction involves 'male' and 'female' gametangia, as in other allogamous species of Sellaphora, but in S. bisexualis cells of the same clone can differentiate into either sex, allowing homothallic reproduction and successive inbred generations. The proven distribution of the species includes sites in the UK and Australia.

Auxosporulation of the freshwater epipelic diatom Pinnularia nodosa (Ehrenb.) W. Sm. was studied in a clonal culture. Interphase cells possessed two chloroplasts with invaginated pyrenoids. The nucleus contained a single small body of heterochromatin at one end, also visible during most of meiotic prophase. During auxosporulation, induced by transfer of stationary-phase cells to fresh medium and suppressed by high nitrogen (N), an unpaired mother cell produced a single auxospore. Although meiosis II and nuclear fusion were not observed, indirect evidence indicated that auxosporulation was autogamous (rarely reported in pennate diatoms), rather than apomictic; paedogamy was excluded. The protoplast produced after meiosis either (1) matured into a "pseudozygote," via an asymmetrical contraction after meiosis I to form a single spherical cell at one end of the mother cell (pathway 1); or (2) constricted into two spherical cells (pathway 2). In pathway 2, the "pseudogametes" never fused and only one or none developed into a pseudozygote and then into an auxospore. Pathway 2 could be suppressed by continuous light. During metamorphosis of the spherical pseudozygote into an elongate young auxospore, a complete covering of thin siliceous incunabular strips was formed, separate from the organic wall formed around the pseudozygote when first formed and from the perizonium. Mature auxospores produced via pathway 2 had 60% of the volume as those produced via pathway 1 and had smaller chloroplasts (through loss of fragments during protoplast cleavage), but they achieved exactly the same lengths, suggesting that absolute length is monitored during expansion.

Examination of Grammatophora marina from rough and clonal cultures showed that cell size changes were more flexible than is generally reported for diatoms. Allogamous sexual auxosporulation took place through copulation between small male cells and larger female cells, but only in mixed rough culture and never in clonal cultures. Auxospores were also formed without copulation in clonal cultures (‘uniparental auxosporulation’) and these, like sexual auxospores, developed through formation of a perizonium, which consisted of a series of transverse bands. All of these bands, including the primary band, were open. Circular scales were present in the auxospore wall before initiation of perizonium formation and irregular, elongate structures lined the suture of the transverse perizonium. Perizonium and scales resembled those of another araphid pennate diatom, Gephyria media. Initial cells were formed within the perizonium and consisted of an initial epivalve with a simplified structure, an initial hypo...

Ditylum brightwellii (T. West) Grunow in Van Heurck is now a model organism for population genetic studies of marine phytoplankton, but observations of sexual reproduction in this species are sparse and there are controversial aspects to the identification of its gametes. Culture studies and field observations of D. brightwellii from Wadsworth Cove (Castine), Maine (USA) showed that it was homothallic, producing and releasing two naked, spherical eggs from each oogonium and 64 uniflagellate sperm per spermatogonangium. Eggs and sperm were produced by larger and smaller cells, respectively, and were formed in nutrient-replete medium, without special procedures for induction. Size regeneration was achieved from a putatively fertilized egg via a true auxospore. Vegetative cell enlargement occurred in senescent cultures and resulted in intermediate-sized cells. The size structure of natural populations of D. brightwellii was monitored weekly in autumn 2004 and was unimodal throughout, with slight shifts in si...

The cytology and life cycle of Pinnularia cf. gibba was examined in nine clones from three Scottish localities. This freshwater epipelic diatom is heterothallic and produces two isogametes per gametangium in type IC auxosporulation (Geitler's classification [1973]). The zygote undergoes a highly unusual metamorphosis before beginning expansion, becoming shortly linear-lanceolate; this is accompanied by formation of a complete covering of thin, oxidation-resistant strips and scale-like structures (at the poles), which are quite separate from the perizonium formed during auxospore expansion. Observations of similar incunabular structures in P. acidojaponica show that these elements are siliceous. The P. cf. gibba perizonium also has unusual features, including a remarkably wide primary band. Trikaryotic and haploid auxospores are sometimes formed and haploid 'zygotes' mature and expand like diploids, but do not develop into mature initial cells. Several phases of mucilage secretion take place, from the gametangia, zygotes and auxospores. Triplets of gametangia and polyspermy occurred with high frequency; this and the systematic significance of variation in auxospore, incunabula and perizonium structure, are discussed. Aspects of the taxonomy of the P. gibba group are treated in supplementary material provided on the European Journal of Phycology website.

Clones identified as Eunotia bilunaris were isolated from two freshwater habitats in New Zealand and one in Tasmania. Experimental studies of sexual reproduction showed that the mating system was strictly heterothallic. All clones were morphologically similar and, in mixtures of compatible clones, were able to produce a viable F1, regardless of geographical origin. There was nevertheless some geographically related variation in ITS rDNA sequences, suggesting that dispersal and concerted evolution are unable to prevent allopatric divergence. Isolates from two of the three localities also showed significant ITS polymorphisms. Auxosporulation followed a similar pattern to that in other Eunotia species, being isogamous with a single functional gamete per gametangium; plasmogamy was effected via two fusing papillae. Details of auxospore structure, initial cell size, and vegetative valve metrics (width, striation densities) show that conspecificity with a recently studied European E. bilunaris is doubtful. The ...

The identity and nomenclatural history of several smallcelled naviculoid taxa are revisited. The species discussed here are important from the ecological point of view since they are often dominant in benthic freshwater communities. The original concepts of several species that have suffered major taxonomic drift due to their entangled nomenclatural history are discussed, and forgotten epithets are resurrected. We examined the original material of Navicula aggerica E. Reichardt, Navicula atomoides Grunow, N. crassulexigua E. Reichardt, N. minima Grunow, N. minima var. typica R. Ross, N. minutissima (Kütz.) Grunow, N. saugerresii Desm., N. seminulum Grunow, N. seminulum var. intermedia Hust., N. seminulum var. radiosa Hust., N. stroemii Hust., N. subbacillum Hust., N. subseminulum Hust., N. tantula Hust., N. vasta Hust., N. ventraloides Hust., Stauroneis fonticola Hust., and Synedra minutissima Kütz. Several of these names were regarded as synonyms in many floristic works and, as such, remained forgotten or ignored. Analyses using light and scanning electron microscopy indicate conspecificity of Navicula minima (= Sellaphora seminulum sensu auct. nonnull.) with Sellaphora saugerresii (Desm.) C.E. Wetzel et D.G. Mann comb. nov., which has priority against N. minima. Synedra minutissima is lectotypified and transferred to Halamphora minutissima (Kütz.) C.E. Wetzel et Compère comb. nov. Navicula minutissima (Kütz.) Grunow 1860, nom. illeg. and Navicula minima Grunow pro parte, typo excl. designate one and the same species (valid and legitimate), currently known as Sellaphora aggerica (E. Reichardt) Falasco et Ector. We consider Sellaphora atomoides (Grunow) C.E. Wetzel et Van de Vijver comb. nov. (= Eolimna tantula sensu auct. nonnull.) and Sellaphora nigri (De Not.) C.E. Wetzel et Ector comb. nov. (= Eolimna minima sensu auct. nonnull.) to be separate species, although morphologically very similar. Sellaphora crassulexigua (E. Reichardt) C.E. Wetzel et Ector comb. nov. and Sellaphora subseminulum (Hust.) C.E. Wetzel comb. nov. are rarely encountered, but usually found in calcareous springs and aerial habitats, respectively. All species are transferred to the genus Sellaphora on the basis of their valve morphology, pending molecular studies confirming the monophyly of the group once living material of each can be located and brought into clonal culture. Additionally, 64 established taxa from Navicula s.l., Eolimna or Naviculadicta are formally transferred to Sellaphora. Navicula subminuscula Manguin is formally transferred to the genus Craticula Grunow.

Nitzschia fonticola (Grunow) Grunow is a member of Nitzschia sect. Lanceolatae, a group of taxonomically intractable but ecologically important and widespread diatoms. We investigated the morphology and life cycle in three clones of N. fonticola and all exhibited reduced sexuality, with pedogamous production of auxospores in unpaired gametangia. The auxospores of all clones contained tangles of striplike elements that lay outside the perizonium and were distinct from it in structure and ontogeny. We introduce a new term, incunabula, to refer to such components of the auxospore wall. Semicryptic variation was detected: one clone differed from the other two in valve size and shape, stria density, and fibula density, as well as its nuclear large subunit ribosomal DNA (LSU rDNA) sequence. The implications of reduced sexuality for the taxonomy of sect. Lanceolatae are discussed. A lectotype is designated for N. fonticola from among original material of Grunow, and the application of the name is clarified further by designating illustrations and the LSU sequence AM182191 from one of our clones as epitypes.

Homothallic sexual reproduction and auxosporulation were studied in monoclonal cultures and seminatural populations of the freshwater epipelic diatom Navicula cryptocephala Kutz. Gametangia paired via the girdle, one gamete was formed per gametangium (and hence one zygote per pair of gametangia), and gamete fusion took place without the formation of any copulation envelope or copulation canal. Superfluous nuclei from meiosis survived unusually long, so that gametes and young zygotes were probably functionally polyploid; later, all but two haploid nuclei degenerated. Expanded auxospores had a swollen center, but during formation of the initial valves, the auxospore contracted away from the perizonium to produce linear-lanceolate valves. The pattern of reproductive behavior found in N. cryptocephala can be classified as type IIA2a auxosporulation in Geitler's system. The same type of zygote and auxospore formation seen in clonal cultures was observed in seminatural material from four lakes in Scotland and the Czech Republic. Variation in nuclear structure and auxosporulation in the N. cryptocephala species complex is discussed, as is the evolution of type II auxosporulation (one zygote per pair of gametangia) from type I auxosporulation (two zygotes per pair). The penalty of smaller numbers of zygote produced in type II may be outweighed by formation of larger auxospores (prolonging the vegetative phase) or more vigorous auxospores. The variation present among members of the N. cryptocephala complex indicates that biogeographical analyses based on use of the name N. cryptocephala, as performed recently to support the ubiquity hypothesis of protist distributions, are almost meaningless.

Clonal cultures of Pseudo-nitzschia pungens were isolated at various times from seven sites in the North Sea. During the mitotic cell cycle, the two plate-shaped chloroplasts were girdle-appressed during interphase and mitosis. After cytokinesis, the chloroplasts moved onto the parental valve and remained there during the formation of the new hypovalve and until separation and re-arrangement of the sibling cells within the cell chain had been completed. Clones were almost always heterothallic and cultures of opposite mating type isolated from different localities were compatible. Meiosis I was cytokinetic and accompanied by chloroplast division. Meiosis II involved karyokinesis but not cytokinesis and preceded the rearrangement and contraction of the two gametes. Sexual reproduction involved physiological anisogamy. With one exception, gamete behaviour was clone-specific, gametes being active in clones of one mating type but passive in clones of the other mating type. Auxospore development was accompanied...

Frustule structure and symmetry, protoplast morphology, and auxosporulation are described for the raphid diatom genus Petroneis, separated from Navicula in 1990. Several features, such as the constant cis symmetry of frustules, flap-like occlusions of the areolae, the opening of the central external raphe endings into a ‘tear-drop’–like groove, valve-appressed plastids, secretion of mucilage capsules during vegetative cell division, and aspects of sexual reproduction, suggest a close relationship between Petroneis and Lyrella; this is supported by a parsimony analysis. Molecular phylogenetic analysis of selected raphid diatoms, based on rbcL sequences, also indicates that Petroneis and Lyrella are sister genera and supports their separation into the family Lyrellaceae, order Lyrellales. This group is ancient, since Lyrella is known from the Palaeocene.

The availability of extensive experimental data and remarkable intra- and interspecific variation in breeding behaviour make Achnanthes Bory sensu stricto an especially good model for studying the reproductive and population biology of pennate diatoms. In most Achnanthes species studied, auxospore formation is accompanied by biparental sexual reproduction, but we found uniparental auxosporulation in Achnanthes cf. subsessilis. Auxosporulation appears to be apomictic and follows contraction of the contents of unpaired cells and then a mitotic division, which is normally acytokinetic: one nucleus aborts before the cell develops into an auxospore. Rarely, both daughter nuclei survive and cytokinesis produces two auxospores (two auxospores per mother cell is highly unusual in pennate diatoms); one may abort. Expansion of auxospores is not accompanied by deposition of a transverse perizonium, but a longitudinal perizonium is produced and consists of a wide central strip (structurally similar to the araphid val...

SUMMARY The auxosporulation of Licmophora communis is allogamous and dioecious. Pairing between sessile, shortstalked cells of compatible clones is followed by meiosis and gametogenesis, to form two gametes in each gametangium. The behavior of the gametes differs between the gametangia. In the male gametangium, the gametes detach from the frustule, round up, and migrate out of the gametangium after its dehiscence at the broader, unattached pole. In the female gametangium, both gametes remain attached to the adjacent theca over almost their whole length and do not move. Plasmogamy therefore occurs within the female gametangium and this is where the zygotes are formed and remain. After fertilization, the zygotes detach from the thecae of the female gametangia, contract, and become ellipsoidal, before expanding parallel to the apical axis of the gametangium. We review the types of auxosporulation in other pennate diatoms and the systems used for classifying these. Dioecy and cis-type anisogamy (in which one gametangium produces active gametes and the other produces passive gametes), as in L. communis, are probably primitive within the pennate group (although there is no information on the AsterionellopsisRhaphoneis clade). However, size can also be restored in various araphid pennates by allogamous sexual reproduction involving the formation of only one gamete per gametangium, or in rare cases by automixis or (apparently) vegetative enlargement.

The diatom genus Eunotia is unusual among raphid diatoms in having a raphe system consisting of two short slits that are not integrated into the primary pattern center. This and other characteristics, particularly the presence of rimoportulae, are consistent with the hypothesis that Eunotia is a basal lineage within the raphid group. We studied auxosporulation in E. bilunaris (Ehrenberg) Mills and E. tropica Hustedt for comparison with other raphid pennate diatoms and with araphid pennates; E. bilunaris was studied in parental and F1 generations. Like araphid pennates, E. bilunaris and E. tropica are heterothallic. Clones of the same mating type did not interact sexually, and intraclonal sexual reproduction was absent or very rare. Clones retained the same sex throughout the life cycle, as shown by experiments using abrupt size reduction to produce clones of similar age but different size and using subclones derived from a single initial cell within six mitotic generations. Unlike in araphid pennate diatoms, in the Eunotia species the gametes are not visibly or behaviorally differentiated. Gametogenesis is merogenous, because the gametangium formed a supernumerary cell as well as a single gametic cell, both undergoing meiosis II to form a surviving functional nucleus and a nucleus that quickly degenerated. Plasmogamy is via papillae that grew out toward each other from the ends of the gametangia to create a copulation canal. After plasmogamy, the gametes moves bodily into the copulation canal, producing an elongate zygote, which expands to form a curved sausage-like auxospore.

The diatom genus Toxarium Bailey has been treated as a pennate because of its elongate shape and benthic lifestyle (it grows attached to solid substrata in the marine sublittoral). Yet its valve face lacks all structures that would ally it with the pennates, such as apical labiate processes, a midrib (sternum) subtending secondary ribs and rows of pores extending perpendicularly out from the midrib, or a raphe system. Instead, pores are scattered irregularly over the valve face and only form two distinct rows along the perimeter of the valve face. In our nuclear small subunit rDNA phylogenies, Toxarium groups with bi- and multipolar centrics, as sister to LampriscusA. Schmidt. Thus, the genus acquired a pennate-like shape and lifestyle independently from that of the true pennates. The two species known, T. hennedyanum Grunow and T. undulatum Bailey, differ only in a single feature: the valve perimeter of the former shows only a central expansion, whereas that of the latter possesses in addition a regular undulation. Yet both forms were observed in our monoclonal cultures, indicating that the two taxa represent extremes in a plasticity range. Toxarium resembles another elongate and supposedly araphid diatom, Ardissonea De Notaris, in being motile. Cells can move at speeds of up to 4 μm.s - 1 through secretion of mucilage from the cell poles or they remain stationary for longer periods, when they form short polysaccharide stalks. Division during longer periods of quiescence leads to the formation of small colonies of linked or radiating cells.

Cell division, the mating system, and auxosporulation were studied in the marine epipelic diatom Seminavis cf. robusta Danielidis & D. G. Mann. The interphase protoplast contains two girdle-appressed chloroplasts, each with an elongate bar-like pyrenoid, and also a central nucleus, located in a bridge between two vacuoles. Before cell division, the chloroplasts divide transversely and translocate onto the valves. The nucleus relocates to the ventral side for mitosis. After cytokinesis and valve formation, the chloroplasts move back to the girdle, showing a constant clockwise movement relative to the epitheca of the daughter cell. Seminavis cf. robusta is dioecious, and sexual reproduction is possible once cells are less than 50 μm. In crosses of compatible clones, gametangia pair laterally, without the formation of a copulation envelope, and produce two gametes apiece. The intensity of sexualization increases as cells reduce further in size below the 50-μm threshold. At plasmogamy, the gametangia dehisce fully and the gametes, which were morphologically and behaviorally isogamous, fuse in the space between the gametangial thecae. The auxospore forms a transverse and longitudinal perizonium. After expansion is complete, there is an unequal contraction of the protoplast within the perizonium, creating the asymmetrical shape of the vegetative cell. Apart from this last feature, almost all characteristics exhibited by the live cell and auxospores of Seminavis agree with what is found in Navicula sensu stricto, supporting the classification of both in the Naviculaceae. Haploid parthenogenesis and polyploid auxospores were found, lending support to the view that change in ploidy may be a significant mechanism in diatom evolution.

Amphora angusta Gregory, Amphora ventricosa Gregory and Amphora macilenta Gregory have been misunderstood for the last 140 years. Gregory described these species from sublittoral sediments off the west coast of Scotland. Although the illustrations were excellent for their time (1857), it is impossible to be confident of identification using Gregory's paper, and subsequent authors have created such confusion that records of these taxa must be regarded as untrustworthy unless backed by photographic evidence. In this paper, the first to describe in detail the recently established genus Seminavis, we show that A. angusta is in fact a species of Amphora Ehrenberg, whereas A. ventricosa sensu Gregory represents two independent species of the genus Seminavis D. G. Mann, namely Seminavis ventricosa (Gregory) Garcia-Baptista (non S. ventricosa sensu Garcia-Baptista 1993) and Seminavis arranensis Danielidis & D. G. Mann, sp. nov. The form usually known as A. ventricosa is neither of these and requires a new name, S...

The allogamous raphid diatom Achnanthes longipes C. A. Agardh possesses a complex breeding system involving interactions between three types of clone: monoecious, unisexual and bisexual. Previous studies showed that these three types can be crossed with each other, with a tendency for sexual characteristics to be inherited: inbred monoecious lineages gave rise to monoecious or, very rarely, to bisexual clones, while inbred unisexual lineages yielded unisexual and bisexual clones. The current paper reports on the progeny of crosses between monoecious and unisexual clones and their inbred offspring. All three types of clone appeared in the F1 and F2, although unisexual clones of opposite sex to the parental clone were not found. Inbreeding depression was observed and also a tendency for ‘normal’ auxosporulation (producing two auxospores per pair of gametangia) to be replaced by ‘reduced’ or ‘intermediate’ auxosporulation (producing one auxospore per pair). In addition, patterns of incompatibility were obser...

Ten polymorphic microsatellite markers were developed for the benthic freshwater diatom Sellaphora capitata and tested on 40 isolates from a Belgian pond. Genotyping was very successful (95%). The number of alleles per locus ranged from three to 12 (mean 6.6) and expected heterozygosities from 0.2 to 0.86 (mean 0.67). This is the first time that microsatellite markers have been developed for a freshwater or benthic diatom.

This paper continues a series of studies of the allogamous raphid diatom Achnanthes longipes, which has a complex reproductive system combining unisexual, bisexual and monoecious behaviour. Following earlier work on the effects of inbreeding in the progeny of crosses between two unisexual clones, we studied the progeny of clones that are capable of a high degree of selfing in monoclonal culture (‘monoecious clones’). Three generations of selfed progeny were examined. In addition, we investigated the F1 generation obtained after crossing two different monoecious clones. Monoecious clones produced monoecious or, more rarely, bisexual progeny, but did not give rise to unisexual progeny. As in inbred lineages made by crossing closely related unisexual clones, inbreeding in monoecious lineages leads to a reduction in the number of gametes formed by the gametangia, from two to one. Inbred clones exhibit marked inbreeding depression and only three inbred generations were possible in monoecious lineages. In the t...

Lyrella cassiteridum sp. nov. is described from the Isles of Scilly, SW Britain. It is an epipelic species and was found in samples taken from marine sandflats in the intertidal and uppermost subtidal zones. In LM, L. cassiteridum is most similar to L. abrupta, from which it can be distinguished by its larger size and coarser structure. The ultrastructure of the frustule is documented and discussed with respect to that of other Lyrella species and the related genus Petroneis. A brief review of the nomenclatural history of Lyrella is given.

A new raphid pennate diatom genus, Eunophora, and three new species are described from highland lakes and streams in Tasmania and New Zealand. Eunophora tasmanica and E. indistincta are only found in Tasmania; E. oberonica and a fourth species (Eunophora sp. 1) also occur in New Zealand. The presence of polar rimoportulae in E. tasmanica and E. oberonica, the relatively simple structure and arrangement of the raphe system and the stria pattern indicate that Eunophora belongs to the subclass Eunotiophycidae. However, it differs from the other genera in this subclass in the amphoroid symmetry of the cells, the length of the raphe slits and the non-coaxial internal central raphe endings; also unusual are the position of the raphe on the valve face instead of on the ventral mantle and the presence of many small discoid or band-like chloroplasts. Eunophora may represent a link between the Eunotiophycidae and the amphoroid genera of the Bacillariophycidae. Eunophora is characteristic of dystrophic to (ultra-)ol...

Monoecious, bisexual and unisexual clones of Achnanthes longipes were isolated from the Black Sea and studied in laboratory culture. Clones differed in their growth characteristics : in monoecious clones the cells formed tufted aggregations while in other clones they were more dispersed. Bisexual and unisexual clones exhibited intraclonal (monoecious) reproduction, but only at a very low frequency and usually within a more restricted size range than in monoecious clones. Interclonal crosses were made in all possible pairwise combinations. Abundant auxosporulation took place in all crosses, except where unisexual clones of the same sex were incubated together. Auxosporulation was more vigorous and occurred over a wider size range in interclonal crosses than during monoecious reproduction. Sexual reproduction is isogamous. In the commonest pattern of auxosporulation, two paired gametangia each produce two gametes, which fuse to give two auxospores. More rarely (9% of pairs), the gametangia produce only one ...

Stauroneis phoenicenteron, the type species of Stauroneis, Stauroneis anceps (two moiphotypes) and S. legumen have Geitler's type IC auxosporulation, which has been demonstrated previously also in S. anceps var. siberica. Type IC auxosporulation is possessed by raphid diatoms with quite different valve, girdle and protoplast structure. This, coupled with the variation that occurs between type IC diatoms in, for example, the structure of the copulation envelope, auxospore shape, and the structure of the caps present at the ends of the auxospores, suggests that type IC auxosporulation is not in itself a reliable indicator of close systematic relationship. However, the particular kind of type IC auxosporulation found in Stauroneis also occurs in Craticula and “Navicula” protracta (but not in Navicula sensu stricto). All available evidence, from cell wall and protoplast characters, is consistent with the idea of a close relationship between these taxa and with the idea that the freshwater Stauroneis species b...

Sexual reproduction and protoplast structure are used to check the classification of the naviculoid genus Placoneis Mereschkowsky (Bacillariophyta). The gametangia are closely associated within a robust mucilage capsule throughout auxosporulation, and pairing is ± random with respect to asymmetries of the frustule and protoplast. Two gametes are produced per gametangium, which are morphologically identical but differentiated into active and passive (physiological anisogamy). The supernumerary nuclei from meiosis II begin to degenerate soon after telophase II, but can still be detected in the initial cells. The gametes become rearranged within the gametangia after meiosis II and plasmogamy takes place via a single, simple aperture. The auxospores expand parallel to the apical axes of the gametangia. The classification of Placoneis in the Cymbellales, as the sister group to the cymbelloid and gomphonemoid diatoms, is supported by valve and protoplast structure, the method ofchloroplast division, an...

cGametogenesis and auxospore development have been studied in detail in surprisingly few centric diatoms. We studied the development of sperm, eggs and auxospores in Actinocyclus sp., a radially symmetrical freshwater diatom collected from Japan, using LM and electron microscopy of living cultures and thin sections. Actinocyclus represents a deep branch of the 'radial centric' diatoms and should therefore contribute useful insights into the evolution of sexual reproduction in diatoms. Spermatogenesis was examined by LM and SEM and involved the formation of two spermatogonia (sperm mother-cells) in each spermatogonangium through an equal mitotic division. The spermatogonia produced a reduced 'lid' valve, resembling a large flat scale with irregular radial thickenings. Sperm formation was merogenous, producing four sperm per spermatogonium, which were released by dehiscence of the 'lid' valve. The sperm were spindle-shaped with numerous surface globules and, as usual for diatoms, the single anterior flagellum bore mastigonemes. One egg cell was produced per oogonium. Immature eggs produced a thin layer of circular silica scales before fertilization, while the eggs were still contained within the oogonium. Sperm were attracted in large numbers to each egg and were apparently able to contact the egg surface via a gap formed between the long hypotheca and shorter epitheca of the oogonium and a small underlying hole in the scale-case. Auxospores expanded isodiametrically and many new scales were added to its envelope during expansion. Finally, new slightly-domed initial valves were produced at right angles to the oogonium axis, after a strong contraction of the cell away from the auxospore wall. At different stages, Golgi bodies were associated with chloroplasts or mitochondria, contrasting with the constancy of Golgi-ER-mitochondrion (G-ER-M) units in some other centric diatoms, which has been suggested to have phylogenetic significance. Electron-dense bodies in the vacuole of Actinocyclus are probably acidocalcisomes containing polyphosphate.

The genus Dickieia is re-established for Navicula ulvacea, N. subinflata and N. expecta. Dickieia cells contain one complex chloroplast per cell; this consists of two ‘K’-shaped halves, linked by an isthmus containing the single discoid pyrenoid. The pyrenoid lies near one side of the girdle at the centre, while the nucleus lies opposite. Neither the nucleus nor the chloroplast move significantly within the cell during the cell cycle. The valves contain round poroids with hymenate occlusions. The raphe-sternum lacks accessory ribs and the central internal raphe endings are simple and straight. The epitheca contains several narrow, porous girdle bands. Auxosporulation is unusually variable in D. ulvacea and can be allogamous (with isogamy) or automictic, though the gametangia are always paired. Plasmogamy is brought about principally through expansion of the gametes until they meet and fuse. The orientation of the auxospores bears no fixed relation to the gametangia. Haploid parthenogenesis and the formati...

Sexual reproduction and auxospore development are described for a marine species of Bacillariaceae related to Nitzschia reversa. Oogamy, recently claimed to occur in N. pungens f. multiseries, was not found. No copulation envelope is produced and reproduction is isogamous. The orientations of the auxospores and gametangia bear no fixed relation to each other. A similar mode of auxosporulation occurs in some other species of Bacillariaceae, such as N.fonticola, N. angustata and apparently also N. subcurvata. Certain features are also shared by N. longissima, which shares a similar cell shape, with long rostrate apices. Two of the four plastids present in the zygotes and auxospores are often contracted and apparently degenerate. Oogamy is unlikely to occur in any members of the Bacillariaceae, though some slight doubt lingers; an old report (by Pascher), of the formation of four protoplasts (gametes?) within frustules of Nitzschia cf. hantzschiana, requires reinvestigation. The nature of the caps that cover...

Diatoms are a diverse lineage with species that can be difficult to identify or cryptic, but DNA barcoding, a molecular technique, can assist identification and facilitate studies of speciation and biogeography. The most common region used for DNA barcoding, COI-5P, can distinguish diatom species, but has not displayed universality (i.e., successful PCR amplification from diverse taxa). Therefore, we have assessed the following alternative markers: ∼1400 bp of rbc L; 748 bp at the 3′ end of rbc L ( rbc L-3P); LSU D2/D3 and UPA. Sellaphora isolates were used to determine each marker's ability to discriminate among closely related species and culture collection material was utilized to explore further marker universality. All of the alternative markers investigated have greater universality than COI-5P. Both full and partial (3P) rbc L regions had the power to discriminate between all species, but rbc L-3P can be sequenced more easily. LSU D2/D3 could distinguish between all but the most closely related species (96%), whereas UPA only distinguished 20% of species. Our observations suggest that rbc L-3P should be used as the primary marker for diatom barcoding, while LSU D2/D3 should be sequenced as a secondary marker to facilitate environmental surveys.

The history and characteristics of the genus Craticula are reviewed, to justify its separation from Navicula and from the Naviculaceae. Members of the genus have previously been put in Navicula sect. Orthostichae; they include such species as those formerly known as Navicula cuspidata, N. ambigua, N. perrotettii and N. halophila. The closest relatives of Craticula seem to be Stauroneis sensu stricto and the diatoms allied to “Navicula” protracta.

Pseudostriatella oceanica gen et. sp. nov. is a marine benthic diatom that resembles Striatella unipunctata in gross morphology, attachment to the substratum by a mucilaginous stalk and possession of septate girdle bands. In light microscopy, P. oceanica can be distinguished from S. unipunctata by plastid shape, absence of truncation of the corners of the frustule, indiscernible striation and absence of polar rimoportulae. With scanning electron microscopy, P. oceanica can be distinguished by a prominent but unthickened longitudinal hyaline area, pegged areolae, multiple marginal rimoportulae and perforated septum. The hyaline area differs from the sterna of most pennate diatoms in being porous toward its expanded ends; in this respect, it resembles the elongate annuli of some centric diatoms, such as Attheya and Odontella. 18S rDNA phylogeny places P. oceanica among the pennate diatoms and supports a close relationship between P. oceanica and S. unipunctata, but the genetic distance between them...