MORPHOLOGY AND TAXONOMY OF PROROCENTRUM MEXICANUM AND REINSTATEMENT OF PROROCENTRUM RHATHYMUM (DINOPHYCEAE)1

Dinoflagellates collected during red tide events in Bahia Mazatlan, Mexico during the early spring of 1999 and 2000 appeared under LM to belong to Prorocentrum mexicanum Osorio‐Tafall. Observations with SEM of those populations showed marked differences in shape and microornamentation from the related species, Prorocentrum rhathymum Loeblich III, Sherley and Schmidt. In P. mexicanum, the presence and dimensions of poroids, the uneven distribution of trichocyst pores not located in depressions, and the general architecture of the periflagellar region are more closely related to Prorocentrum caribbaeum Faust. Also, P. mexicanum has a three‐horned (sometimes two‐horned) spine and is deeper in the anterior than the posterior region, whereas P. rhathymum has a simple small spine and its sagittal view is oval. Furthermore, the number and distribution of trichocyst pores in the periflagellar area is different between the two species, being located on both valves in P. mexicanum and only on the right valve in P. rhathymum. To date, true P. mexicanum has been described only from plankton sampling, whereas P. rhathymum was frequently mentioned associated with floating detritus (macroalgae) but also forming red tides. Altogether, the evidence presented demonstrates that P. mexicanum (planktonic) and P. rhathymum (epibenthic) are distinct species and are not synonyms, as is often accepted.     

Phylogenetic relationships and biogeography of Ptilophora (Gelidiales,Rhodophyta) with descriptions of P. aureolusa,P. malagasya,and P. spongiophila from Madagascar

The genus Ptilophora currently includes 16 species occurring mostly in subtidal habitats of the Indo‐Pacific Ocean, but its global diversity and biogeography are poorly understood. We analyzed mitochondrial cox1, plastid rbcL and plastid psbA sequences from specimens collected in southern Madagascar during the 2010 Atimo Vatae expedition and studied their morphologies. Both morphological and molecular data sets demonstrated the presence of five species in southern Madagascar: Ptilophora hildebrandtii, P. pterocladioides, and three new species described here, P. aureolusa, P. malagasya, and P. spongiophila. Ptilophora aureolusa is distinguished by its compound pinnae with uniformly spaced pinnules. Ptilophora malagasya has an indistinct midrib and irregularly spaced pinnules. Ptilophora spongiophila, heavily coated with sponges, has cylindrical to flattened main axes, lateral and surface proliferations, and spatulate tetrasporangial sori. The species of Ptilophora found in Madagascar are endemic, except P. hildebrandtii, which also occurs in eastern Africa. Ptilophora comprises four phylogenetic groups that map to eastern Australia, Japan, western Australia/Southeast Asia/Madagascar/eastern Africa, and Madagascar/eastern Africa/Aegean Sea. Biogeographical analysis revealed that the ancestor of Ptilophora originated in Australia, but most of the species radiated from Madagascar.     

A survey of epiphytic dinoflagellates from the coastal waters of the island of Hawai‘i

Three hundred and sixty-nine macroalgal and non-algal samples were collected from six coastal sites around the island of Hawai‘i on a biweekly basis over a 1-year period to ascertain (1) the presence of potentially toxigenic benthic dinoflagellates, and (2) substrate and environmental preferences of the dinoflagellates. Twenty-six genera/species of dinoflagellates were encountered including the (potentially) toxigenic species Amphidinium sp., Coolia monotis, Gambierdiscus sp., Ostreopsis ovata, Prorocentrum concavum, Prorocentrum hoffmannianum, Prorocentrum lima, and Prorocentrum mexicanum. Twenty of the species are being reported for the first time as present in Hawaiian coastal waters. There was some evidence of macroalgal host preference (e.g., C. monotis on Tolypiocladia glomerulata), although host morphology preferences was greater (e.g., Gambierdiscus sp., and P. lima on filamentous turfs; O. sp. 1 on sheet-like macroblades). While some dinoflagellate groups were significantly correlated with nutrient concentrations (e.g., total dinoflagellates with nitrate + nitrite and phosphate concentrations), others were not (e.g., Gambierdiscus sp., Prorocentrum emarginatum, P. lima, and Sinophysis microcephalus). The presence of several potentially toxigenic dinoflagellate species in Hawai‘i merits future study on possible impacts of these dinoflagellates on coastal food webs and human health.     

Comparative phylogeographies of six species of hinged terrapins (Pelusios spp.) reveal discordant patterns and unexpected differentiation in the P. castaneus/P. chapini complex and P. rhodesianus

Using up to 2117 bp of mitochondrial DNA and up to 2012 bp of nuclear DNA, we analysed phylogeographic differentiation of six widely distributed species of African hinged terrapins (Pelusios spp.) representing different habitat types. Two taxa each live in savannahs or in forests and mesic savannahs, respectively, and the remaining two species occur in intermediate habitats. The species living in forests and mesic savannahs do not enter dry savannahs, whereas the savannah species may occur in dry and wet savannahs and even in semi‐arid steppe regions. We found no obvious correlation between habitat type and phylogeographic pattern: one savannah species (P. rhodesianus) shows phylogeographic structure, i.e. pronounced genetic differences among geographically distinct populations, and the other (P. nanus) not. One species inhabiting forests and mesic savannahs (P. carinatus) has phylogeographic structure, the other (P. gabonensis) not. The same pattern is true for the two ecologically intermediate species, with phylogeographic structure present in P. castaneus and absent in P. chapini. Nuclear evidence suggests that the latter two taxa with abutting and partially overlapping ranges are distinct, while mtDNA is only weakly differentiated. Pelusios castaneus shows pronounced phylogeographic structure, which could reflect Pleistocene range interruptions correlated with the fluctuating forest cover in West and Central Africa. Our results do not support the recognition of an extinct subspecies of P. castaneus for the Seychelles. Pelusios carinatus contains two well supported clades, which are separated by the Congo River. This species is closely related to P. rhodesianus, a taxon consisting of two deeply divergent mitochondrial clades. One of these clades is paraphyletic with respect to P. carinatus, but the two clades of P. rhodesianus are not differentiated in the studied nuclear markers and, again, paraphyletic with respect to P. carinatus. Using mtDNA sequences from the type material of P. rhodesianus, we were able to allocate this name to one of the two clades. However, owing to the confusing relationships of P. rhodesianus and P. carinatus, we refrain from taxonomic decisions.     

Blue mould decay of stored onion bulbs caused by Penicillium polonicum,P. glabrum and P. expansum

Onion bulbs (Allium cepa L.) can be stored for up to 12 months to ensure their availability all year, but one of the limiting factors is decay caused by blue mould fungi. Postharvest development of blue mould is not only economically important, but also poses a health hazard due to mycotoxin contamination. Onion bulbs of different yellow and red cultivars with blue mould symptoms were collected from three storage facilities in Serbia. Pathogenic isolates were identified and characterized based on their molecular (partial β‐tubulin gene), morphological (micro‐ and macromorphology, and colony growth) and biochemical properties (analyses of indole metabolites via Ehrlich test). Three Penicillium species were identified: P. polonicum, P. glabrum and P. expansum. Virulence on inoculated onion bulbs varied significantly within isolates of P. glabrum, but the most virulent isolate of P. polonicum and P. glabrum did not differ, nor did they differ from an isolate of P. expansum. Variability in virulence of individual P. glabrum isolates corresponded to differences in their molecular and macromorphological characters. P. glabrum and P. expansum were identified for the first time as causal agents of onion bulb decay in Serbia. Data from this investigation provide insight into diversity of economically important and possibly toxigenic blue mould fungi which brings attention to their presence in storage facilities and therefore the necessity of the application of prevention measures.     

Chemotaxonomic Considerations of the n‐Alkane Composition in Pinus heldreichii,P. nigra,and P. peuce

The n‐alkane composition in the leaf cuticular waxes of natural populations of Bosnian pine (Pinus heldreichii), Austrian pine (P. nigra), and Macedonian pine (P. peuce) was compared for the first time. The range of n‐alkanes was wider in P. nigra (C₁₆ – C₃₃) than in P. heldreichii and P. peuce (C₁₈ – C₃₃). Species also diverged in abundance and range of dominant n‐alkanes (P. heldreichii: C₂₃, C₂₇, and C₂₅; P. nigra: C₂₅, C₂₇, C₂₉, and C₂₃; P. peuce: C₂₉, C₂₅, C₂₇, and C₂₃). Multivariate statistical analyses (PCA, DA, and CA) generally pointed out separation of populations of P. nigra from populations of P. heldreichii and P. peuce (which were, to a greater or lesser extent, separated too). However, position of these species on the basis of n‐alkane composition was in accordance neither with infrageneric classification nor with recent molecular and terpene investigations.     

Histopathology in Pacific oyster (Crassostrea gigas) spat caused by the dinoflagellate Prorocentrum rhathymum

The recognition of an apparent association between seasonal oyster spat mortalities (up to 40%) and high Prorocentrum rhathymum density in the Little Swanport Estuary, Tasmania, prompted further experimental investigation into the toxicity by this dinoflagellate. Standard brine shrimp, haemolysis assays and intraperitoneal mouse bioassays revealed fast acting toxins in methanol but not aqueous extracts of P. rhathymum, with mice dying in less than 20 min. Oyster bioassays involved feeding spat (4 mm shell width) for 21 consecutive days on a diet of cultured P. rhathymum at simulated bloom densities (10⁴ cells ml⁻¹). No oyster mortality was observed, however, histopathological signs of thin, dilated gut tubules and sloughing of gut cells resembled those seen in affected field samples. In contrast to field samples, gill pathology was also observed in experimental exposure oysters.     

A red tide off the Myanmar coast: Morphological and genetic identification of the dinoflagellate composition

A red tide composed mainly of two Prorocentrum species was found near Kadan Island, southern Myanmar coast, on March 14, 2012. This is the first record of a red tide in the Myanmar coastal area. The red tide included three different harmful dinoflagellates: Prorocentrum rhathymum, Prorocentrum shikokuense and Alexandrium affine. Strains of these species were successfully cultured and examined with detailed morphological observation and DNA (28S rRNA gene) analyses. The strains were subjected to growth experiments under different temperatures to understand their growth capabilities. The experiments were carried out at four different temperature regimes (15, 20, 25 and 30 °C). A. affine exhibited low tolerance for the low temperature regime (15 °C), despite records of its presence in northern temperate regions, indicating this strain is adapted to the tropical environment in Myanmar. P. rhathymum and P. shikokuense exhibited broad tolerance to all given temperature ranges and showed high division rates, providing the physiological basis to form red tides.     

Phylogeny and taxonomic reassessment of pikas Ochotona pallasii and O. argentata (Mammalia,Lagomorpha)

We examined a cranial morphometric data set consisting of 186 specimens from the entire distribution range of Ochotona pallasii sensu lato and O. argentata, as well as 67 complete sequences of the COI gene and 28 sequences of the MGF and PRKCI introns from these and closely allied species. Our results show that the two allopatric morphologically similar taxa composing O. pallasii sensu lato – from Mongolia and adjacent territories and Kazakhstan – are paraphyletic relative to O. argentata. Genetic distances between these three taxa are larger than the intraspecific variation known for the subgenus Pika, in which the species under consideration belong; these distances are even larger than the interspecific differences among closely related species such as O. hyperborea, O. mantchurica and O. hoffmanni. Thus, the three focal taxa are recognized here as distinct species. Inspection of the type specimen of O. pallasii indicated that this specimen was not collected in Kazakhstan, has previously been theorized. The most probable place of the holotype's origin is Russian south‐eastern Altai (Chuyskaya Steppe); whatever its exact origin, it definitively originates from the ‘Mongolian’ taxon. Based on this evidence, the senior synonym for the Kazakh pika is O. opaca Argyropulo, 1930. Thus, we propose to recognize three separate species in the O. pallasii species group: O. pallasii (Mongolia and adjacent territories), O. opaca (eastern Kazakhstan) and O. argentata (Helan Shan Range, China).     

Genetic diversity and haplotype distribution of Pachymeniopsis gargiuli sp. nov. and P. lanceolata (Halymeniales,Rhodophyta) in Korea,with notes on their non‐native distributions

The red alga Pachymeniopsis lanceolata, formerly known as Grateloupia lanceolata, is a component of the native algal flora of northeast Asia and has been introduced to European and North American waters. It has been confused with a cryptic species collected from Korea and Italy. Our analyses of rbcL, cox3 and ITS from P. lanceolata and this cryptic species has revealed two distinct entities, forming a clade, which were clearly separated from its congeners and positioned with other Asian species. Here, we describe the cryptic species as P. gargiuli sp. nov., a species that differs from others by molecular sequence and subtle anatomical characters. We hypothesize that P. gargiuli may have been recently dispersed by anthropogenic vectors, possibly at or near the same time as was P. lanceolata. Our cox3 data set revealed that one haplotype of P. gargiuli, shared between Korea and Italy, and two haplotypes of P. lanceolata, commonly occurring in Korea and USA, are invasive haplotypes. This is the first report of the utility of the mitochondrial coding cox3 sequences in red algae.     

Molecular phylogeny of the spiny-surfaced species of the dinoflagellate Prorocentrum with the description of P. thermophilum sp. nov. and P. criophilum sp. nov. (Prorocentrales,Dinophyceae)

Spiny-surfaced species of Prorocentrum form harmful algal blooms, and its taxonomic identity is obscure due to the size and shape variability. Molecular phylogenies reveal two major clades: one for P. cordatum with sequences mainly retrieved as P. minimum, and the other for P. shikokuense with sequences also retrieved as P. dentatum and P. donghaiense. Several closely related clades still need to be characterized. Here, we provide nuclear SSU and LSU rRNA genes, and nuclear ITS region (ITS1-5.8S gene-ITS2) sequences of the strain CCMP3122 isolated from Florida (initially named P. donghaiense) and strains Prorocentrum sp. RCC6871–2 from the Ross Sea, Antarctica. We describe Prorocentrum thermophilum sp. nov. based on the strain CCMP3122, a species also distributed in the open waters of the Gulf of Mexico, New Zealand, and the Arabian Gulf; and Prorocentrum criophilum sp. nov. based on the strain RCC6872, which is distributed in the Antarctic Ocean and Arctic Sea. Prorocentrum thermophilum is roundish (~14 μm long, ~12 μm wide), with an inconspicuous anterior spine-like prolongation under light microscopy, valves with tiny, short knobs (5–7 per μm²), and several (<7) large trichocyst pores (~0.3 μm) in the right valve, as well as smaller pores (~0.15 μm). Prorocentrum criophilum is round in valve view (~11 μm long, 10 μm wide) and asymmetrically roundish in lateral view, the periflagellar area was not discernible under light microscopy, valves with very tiny, short knobs (6–10 per μm²), and at least 12 large pores in the right valve. Other potentially undescribed species of spiny-surfaced Prorocentrum are discussed.     

The correct position of flagellar insertion in Prorocentrum and description of Prorocentrum rhathymum sp. nov. (Pyrrhophyta)

For three Prorocentrum species, P. minimum (Pavillard) Schiller,P. rhathymum sp. nov., and P. triestinum Schiller, we have foundthat both flagella emerge from a single large pore. This arrangementis contrary to previous observations which suggested they arosefrom separate pores. Prorocentrum triestinum has only one anteriorpore, whereas P. minimum and P. rhathymum sp. nov. have an additionalauxiliary pore. The pores in all three species are formed bya series of small plates. The left and right valves on eachspecies can be identified by ornamentation unique to each valve.Prorocentrum rhathymum sp. nov., isolated from the planktonof Cinnamon Bay, Virgin Islands, is described. ² The Biological Laboratories, Harvard University, Cambridge,Massachusetts 02138 ³ Department of Botany, Duke University, Durham, North Carolina27706     

Diversity of bladed Bangiales (Rhodophyta) in western Mediterranean: recognition of the genus Themis and descriptions of T. ballesterosii sp. nov., T. iberica sp. nov., and Pyropia parva sp. nov.

The diversity of the bladed species of the red algal order Bangiales from the Iberian Mediterranean shores has been reassessed after a detailed study of this region. Prior to this study, 11 bladed species of Bangiales had been reported from Mediterranean waters: Porphyra atropurpurea, P. cordata, P. coriacea, P. dioica, P. linearis, P. purpurea, P. umbilicalis, Pyropia leucosticta, Pyropia koreana (as P. olivii), Py. elongata (as P. rosengurttii) and Py. suborbiculata. A combined analysis of the nuclear nSSU and the plastid rbcL genes together with detailed morphological studies has confirmed the presence of species within the genera Porphyra and Pyropia and also revealed a third, undescribed genus, Themis gen. nov. Porphyra linearis, Pyropia elongata and the introduced Pyropia koreana had been previously listed for the Mediterranean and were recorded in this study. An additional four species, including the introduced Pyropia suborbiculata and three new species: Pyropia parva sp. nov., Themis ballesterosii sp. nov., and Themis iberica sp. nov. were also observed. Hence, most of the Porphyra species traditionally reported along these shores were not reported in this survey. This new floristic Bangiales composition confirms the importance of the Mediterranean basin as a hotspot for biodiversity, possible endemics of ancient origin and high proportion of introductions. Our data also continue to confirm the extent of Bangiales diversity at regional and worldwide levels.