Mediterranean Lithophyllum stictiforme (Corallinales,Rhodophyta) is a genetically diverse species complex: implications for species circumscription,biogeography and conservation of coralligenous habitats

Lithophyllum species in the Mediterranean Sea function as algal bioconstructors, contributing to the formation of biogenic habitats such as coralligenous concretions. In such habitats, thalli of Lithophyllum, consisting of crusts or lamellae with entire or lobed margins, have been variously referred to as either one species, L. stictiforme, or two species, L. stictiforme and L. cabiochiae, in the recent literature. We investigated species diversity and phylogenetic relationships in these algae by sequencing three markers (psbA and rbcL genes, cox2,3 spacer), in conjunction with methods for algorithmic delimitation of species (ABGD and GMYC). Mediterranean subtidal Lithophyllum belong to a well‐supported lineage, hereby called the L. stictiforme complex, which also includes two species described from the Atlantic, L. lobatum and L. searlesii. Our results indicate that the L. stictiforme complex consists of at least 13 species. Among the Mediterranean species, some are widely distributed and span most of the western and central Mediterranean, whereas others appear to be restricted to specific localities. These patterns are interpreted as possibly resulting from allopatric speciation events that took place during the Messinian Salinity Crisis and subsequent glacial periods. A partial rbcL sequence from the lectotype of L. stictiforme unambiguously indicates that this name applies to the most common subtidal Lithophyllum in the central Mediterranean. We agree with recent treatments that considered L. cabiochiae and L. stictiforme conspecific. The diversity of Lithophyllum in Mediterranean coralligenous habitats has been substantially underestimated, and future work on these and other Mediterranean corallines should use identifications based on DNA sequences.     

Misleading morphologies and the importance of sequencing type specimens for resolving coralline taxonomy (Corallinales,Rhodophyta): Pachyarthron cretaceum is Corallina officinalis

Coralline red algae play a key role in the ecology of near shore marine ecosystems and are increasingly being used to study the effects of climate change in the marine environment. Corallines are very difficult to identify to species, and even to genus, using morpho‐anatomy, likely complicating studies of their ecology, physiology, and biodiversity. We sequenced a 296 base pair fragment of chloroplast DNA from a 187‐year‐old isolectotype specimen of Pachyarthron cretaceum, a morphologically distinct geniculate species, to demonstrate that coralline morphology is often misleading and that species names can only be applied unequivocally by comparing DNA sequences from type material with sequences from field‐collected specimens. Our results indicate that Pachyarthron cretaceum is synonymous with Corallina officinalis.     

Crusticorallina gen. nov., a nongeniculate genus in the subfamily Corallinoideae (Corallinales,Rhodophyta)

Molecular phylogenetic analyses of 18S rDNA (SSU) gene sequences confirm the placement of Crusticorallina gen. nov. in Corallinoideae, the first nongeniculate genus in an otherwise geniculate subfamily. Crusticorallina is distinguished from all other coralline genera by the following suite of morpho‐anatomical characters: (i) sunken, uniporate gametangial and bi/tetrasporangial conceptacles, (ii) cells linked by cell fusions, not secondary pit connections, (iii) an epithallus of 1 or 2 cell layers, (iv) a hypothallus that occupies 50% or more of the total thallus thickness, (v) elongate meristematic cells, and (vi) trichocytes absent. Four species are recognized based on rbcL, psbA and COI‐5P sequences, C. painei sp. nov., the generitype, C. adhaerens sp. nov., C. nootkana sp. nov. and C. muricata comb. nov., previously known as Pseudolithophyllum muricatum. Type material of Lithophyllum muricatum, basionym of C. muricata, in TRH comprises at least two taxa, and therefore we accept the previously designated lectotype specimen in UC that we sequenced to confirm its identity. Crusticorallina species are very difficult to distinguish using morpho‐anatomical and/or habitat characters, although at specific sites, some species may be distinguished by a combination of morpho‐anatomy, habitat and biogeography. The Northeast Pacific now boasts six coralline endemic genera, far more than any other region of the world.     

DNA Sequencing of Type Material Reveals Pneophyllum marlothii comb. nov. from South Africa and P. discoideum comb. nov. (Chamberlainoideae,Corallinales, Rhodophyta) from Argentina

A partial rbcL sequence from the type material of Spongites discoideus from southern Argentina showed that it was distinct from rbcL sequences of South African specimens to which that name had been applied based on morpho-anatomy. A partial rbcL sequence from an original syntype specimen, herein designated the lectotype, of Lithophyllum marlothii, type locality Camps Bay, Western Cape Province, South Africa, was identical to rbcL sequences of South African field-collected specimens assigned to S. discoideus. Based on phylogenetic analyses of rbcL and/or psbA sequences, both of these species belong in Pneophyllum and are transferred there as P. discoideum comb. nov. and P. marlothii comb. nov. The two species exhibit a distinct type of development where thick, secondary, monomerous disks are produced from thin, primary, dimerous crusts. Whether this type of development represents an example of convergent evolution or is characteristic of a clade of species within Pneophyllum remains to be resolved.     

Taxonomic circumscription of heterogeneous species Neogoniolithon brassica‐florida (Corallinales,Rhodophyta) in Japan

The nongeniculate species Neogoniolithon brassica‐florida (Harvey) Setchell et Mason is circumscribed as a polymorphic species with various gross morphologies due to it being synonymized with several previous species. However, small subunit rDNA and cox1 analyses showed that N. brassica‐florida was polyphyletic, and strongly imply that crustose species lacking any protuberances such as Neogoniolithon fosliei (Heydrich) Setchell et Mason and species with protuberances or branches such as N. brassica‐florida and N. frutescens (Foslie) Setchell et Mason should be treated as genetically different groups (species). Therefore, we propose the resurrection of N. frutescens. We also confirmed that N. trichotomum was distinguished from N. frutescens by slender uniform diameter branches, a conceptacle with a prominent ostiole, and large cox1 interspecific sequence differences. Male and female reproductive structures of N. trichotomum were illustrated for the first time. Neogoniolithon fosliei, was divided into three clades, each of which was recognized as a species complex based on interspecific level sequence differences within clade and morphological differences. Therefore, we treated the clade most similar to N. fosliei as N. fosliei complex (Clade B), and the other clades as respective complexes of N. cf. fosliei with yellow conceptacles (Clade A) or N. cf. fosliei with large conceptacles (Clade C). Of two species complexes (Clade A and B) were morphologically consistent with two entities of N. fosliei previously reported in the Ryukyu Islands, Japan, which is supported by their niche partitioning. DNA barcoding research of nongeniculate corallines can promote the finding of more reliable taxonomic characters and the understanding of their biological aspects.     

Discovery of the mineral brucite (magnesium hydroxide) in the tropical calcifying alga Polystrata dura (Peyssonneliales,Rhodophyta)

Red algae of the family Peyssonneliaceae typically form thin crusts impregnated with aragonite. Here, we report the first discovery of brucite in a thick red algal crust (~1 cm) formed by the peyssonnelioid species Polystrata dura from Papua New Guinea. Cells of P. dura were found to be infilled by the magnesium‐rich mineral brucite [Mg(OH)₂]; minor amounts of magnesite and calcite were also detected. We propose that cell infill may be associated with the development of thick (> ~5 mm) calcified red algal crusts, integral components of tropical biotic reefs. If brucite infill within the P. dura crust enhances resistance to dissolution similarly to crustose coralline algae that infill with dolomite, then these crusts would be more resilient to future ocean acidification than crusts without infill.     

Diversity of Chroothece (Rhodophyta,Stylonematales) including two new species

Chroothece has been reported from a range of freshwater environments, including streams, shallow ponds, trickling water on cliffs and moist soils, mostly in Europe and North America. The identification of genera and species by morphology is difficult because of overlaps in critical characters. To help clarify diversity within the genus, samples from Spain and from other regions (UK and Guam, western Pacific) were compared. Ecological and morphological data from field and cultured material were correlated with molecular data (rbcL gene sequences) that differentiate two new species: Chroothece thermalis I. Chapuis, P. Sánchez, M. Aboal & O. Necchi Jr., sp. nov. in a thermal spring and Chroothece lobata M. Aboal, B. A. Whitton, I. Chapuis, P. Sánchez & O. Necchi Jr., sp. nov. in a semi-arid stream. The results suggest recognition of four species, C. thermalis, C. lobata, C. richteriana Hansgirg and C. rupestris Hansgirg, from Spain. Morphology and ecology are useful to help distinguish these species, but the genus needs further study for possible cryptic diversity.     

Phylogenetic relationships of corallinaceae (Corallinales,Rhodophyta): taxonomic implications for reef‐building corallines

A new, more complete, five‐marker (SSU, LSU, psbA, COI, 23S) molecular phylogeny of the family Corallinaceae, order Corallinales, shows a paraphyletic grouping of seven well‐supported monophyletic clades. The taxonomic implications included the amendment of two subfamilies, Neogoniolithoideae and Metagoniolithoideae, and the rejection of Porolithoideae as an independent subfamily. Metagoniolithoideae contained Harveylithon gen. nov., with H. rupestre comb. nov. as the generitype, and H. canariense stat. nov., H. munitum comb. nov., and H. samoënse comb. nov. Spongites and Pneophyllum belonged to separate clades. The subfamily Neogoniolithoideae included the generitype of Spongites, S. fruticulosus, for which an epitype was designated. Pneophyllum requires reassesment. The generitype of Hydrolithon, H. reinboldii, was a younger heterotypic synonym of H. boergesenii. The evolutionary novelty of the subfamilies Hydrolithoideae, Metagoniolithoideae, and Lithophylloideae was the development of tetra/bisporangial conceptacle roofs by filaments surrounding and interspersed among the sporangial initials.     

An integrative systematic approach to species diversity and distribution in the genus Mesophyllum (Corallinales,Rhodophyta) in Atlantic and Mediterranean Europe

For the first time, a comprehensive assessment of Mesophyllum species diversity and their distribution in Atlantic Europe and the Mediterranean Sea is presented based on molecular (COI-5P, psbA) and morphological data. The distribution ranges were redefined for the four species collected in this study: M. alternans, M. expansum, M. macroblastum and M. sphaericum. Mesophyllum sphaericum, which was previously known only from a single maerl bed in Galicia (NW Spain), is reported from the Mediterranean Sea. The known range of M. expansum (Mediterranean and Macaronesia) was extended to the Atlantic Iberian Peninsula. The occurrence of M. alternans was confirmed along the Atlantic French coast south to Algarve (southern Portugal). Mesophyllum lichenoides was only recorded from the Atlantic, whereas M. macroblastum appears to be restricted to the Mediterranean Sea. A positive correlation was observed between maximum Sea Surface Temperature (SSTmax) and the depth at which M. expansum was collected, suggesting that this species may compensate for higher SST by growing in deeper habitats where the temperature is lower. The latter indicates that geographic shifts in the distribution of coastal species as a result of global warming can possibly be mitigated by changes in the depth profile at which these species occur. Mesophyllum expansum, an important builder of Mediterranean coralligenous habitats, may be a good target species to assess its response to climate change.     

Adeylithon bosencei gen. et sp. nov. (Corallinales,Rhodophyta): a new reef‐building genus with anatomical affinities with the fossil Aethesolithon

Adeylithon gen. nov. with one species, A. bosencei sp. nov., belonging to the subfamily Hydrolithoideae is described from Pacific coral reefs based on psbA sequences and morpho‐anatomy. In contrast with Hydrolithon, A. bosencei showed layers of large polygonal “cells,” which resulted from extensive lateral fusions of perithallial cells, interspersed among layers of vegetative cells. This anatomical feature is shared with the fossil Aethesolithon, but lacking DNA sequences from the fossils and the fragmentary nature of Aethesolithon type material, we cannot ascertain if Adeylithon and Aethesolithon are congeneric. Morpho‐anatomical features of A. bosencei were generally congruent with diagnostic features of the subfamily Hydrolithoideae: (i) outline of cell filaments entirely lost in large portions due to pervasive and extensive cell fusions, (ii) trichocytes not arranged in tightly packed horizontal fields, (iii) basal layer without palisade cells, and (iv) cells lining the canal pore oriented more or less perpendicular to roof surface and not protruding into the canal. However, it showed a predominant monomerous thallus organization and trichocytes were disposed in large pustulate, horizontal fields, although they were not tightly packed and did not become distinctly buried in the thallus. Only mature tetrasporangial conceptacles were observed, therefore the type of conceptacle roof formation remained undetermined. Adeylithon bosencei occurs on shallow coral reefs, in Australia, Papua New Guinea, and South Pacific islands (Fiji, Vanuatu). Fossil Aethesolithon is considered an important component of shallow coral reefs since the Miocene; fossil records showed a broad Indo‐Pacific distribution, but a long‐term process of range contraction in the last 2.6 million years, resulting in an overlap with the distribution of the extant Adeylithon. While the congeneric nature of extant and fossil taxa remained uncertain, similarities in morpho‐anatomy, habitat, and distribution may indicate that both taxa likely shared a common ancestor.     

Rocky shore turfs dominated by Corallina (Corallinales,Rhodophyta) in northern Japan†

Three intertidal sites dominated by Corallina turfs were investigated in Hokkaido, Japan. The sites (A, B and C) differed in slope, wave exposure and length of time exposed to air during tidal cycles. Monthly samples were analyzed for frond morphology and other features. Site A, the most wave-exposed site, was dominated by Corallina sp. X, an unknown species, and sites B and C by Corallina pilulifera Postels et Ruprecht. At the different sites the populations differed in conceptacle abundance, coverage by epiphytic Titanoderma corallinae (P. Crouan et H. Crouan) Woelkerling, Chamberlain et Silva, amount of contained sediment, numbers of axes per quadrat, numbers of branch fusions, branch entanglement, frond dryweight, frond length, amount of adventitious branching, numbers of epiphytes (exclusive of T. corallinae), and numbers of animal species. Ninety-one animal species were recorded from the turfs. Corallina is affected morphologically by conditions inherent in its microhabitat, including desiccation, epiphyte loading and the abundance of herbivores.     

New records of Peyssonnelia armorica and Peyssonnelia harveyana (Rhodophyta, Gigartinales) from Japan

Two species of the crustose red algal genus Peyssonnelia (Gigartinales, Peyssonneliaceae) are reported from Japanese waters for the first time. These species share the following combination of vegetative and reproductive features: thalli with appressed margins, perithallial filaments arising from the whole upper surface of each hypothallial cell (the Peyssonnelia rubra‐type anatomy), unicellular rhizoids, hypobasal calcification and spermatangia that are produced in double chains (the Peyssonnelia harveyana‐type spermatangial filament). However, they differ obviously from each other in the hypothallus orientation as seen from below, the perithallus structure in relation to the consistency of the crust, the origin of gonimoblasts and the elevation of the nemathecia. Peyssonnelia armorica is characterized by: (i) hypothallial filaments comprising a polyflabellate layer; (ii) easily separable perithallial filaments in a gelatinous matrix; (iii) gonimoblasts originating exclusively from the auxiliary cell; and (iv) semi‐immersed (slightly elevated) nemathecia. Peyssonnelia harveyana is characterized by: (i) hypothallial filaments arranged in parallel rows; (ii) closely packed perithallial filaments in a firm matrix; (iii) gonimoblsts originating from both the auxiliary cell and the connecting filament; and (iv) conspicuously elevated nemathecia.     

Taxonomic and molecular studies of the family Sebdeniaceae (Sebdeniales,Rhodophyta): new species of Lesleigha gen. nov. and Crassitegula from Hawaii,east Asia and Lord Howe Island

The genus Lesleigha gen. nov., based on L. hawaiiensis sp. nov. from Oahu in the Hawaiian Islands, is characterized morphologically and anatomically and strongly supported as distinct from the other genera of the family Sebdeniaceae (Sebdenia and Crassitegula) primarily on the basis of mitochondrial COI-5P and nuclear SSU and LSU ribosomal DNA sequences. Characters distinguishing Lesleigha from Sebdenia include the repent, dorsiventral habit, nemathecial tetrasporangia, and irregularly cruciate tetrasporangial division pattern. Although these features are equally displayed by the recently described Crassitegula from Bermuda, that genus appears to differ from Lesleigha anatomically by its lack of a fusion cell subtending the gonimoblast, although in the absence of carposporophytes it is virtually necessary that DNA-sequence analyses be conducted before a distinction between Lesleigha and Crassitegula can be certain. Besides Lesleigha hawaiiensis, L. howensis sp. nov. is newly described from Lord Howe Island in the southern Pacific, and the Japanese and Korean Sebdenia yamadae is transferred to Lesleigha as L. yamadae, based on habit and anatomical conformity as well as DNA sequences. In addition, a new species of Crassitegula, C. imitans, is described from Lord Howe Island. The existence of further sebdenialean taxa is documented although not formally treated, indicating that substantial additional diversity most likely still awaits discovery in this reputedly species-poor order.     

Taxonomic contributions to Hapalidiales (Corallinophycidae,Rhodophyta): Boreolithothamnion gen. nov., Lithothamnion redefined and with three new species and Roseolithon with new combinations

Phylogenetic analyses of rbcL gene sequences and of concatenated rbcL, psbA, and nuclear SSU rRNA gene sequences resolved the generitype of Lithothamnion, L. muelleri, in a clade with three other southern Australian species, L. kraftii sp. nov., L. saundersii sp. nov., and L. woelkerlingii sp. nov. Cold water boreal species currently classified in Lithothamnion and whose type specimens have been sequenced are transferred to Boreolithothamnion gen. nov., with B. glaciale comb. nov. as the generitype. The other species are B. giganteum comb. nov., B. phymatodeum comb. nov., and B. sonderi comb. nov., whose type specimens are newly sequenced, and B. lemoineae comb. nov., B. soriferum comb. nov., and B. tophiforme comb. nov., whose type specimens were already sequenced. Based on rbcL sequences from the type specimens of Lithothamnion crispatum, L. indicum, and L. superpositum, each is recognized as a distinct species and transferred to the recently described Roseolithon as R. crispatum comb. nov., R. indicum comb. nov., and R. superpositum com. nov., respectively. To correctly assign species to these three genera based only on morpho-anatomy, specimens must have multiporate conceptacles and some epithallial cells with flared walls. The discussion provides examples demonstrating that only with phylogenetic analyses of DNA sequences can the evolution of morpho-anatomical characters of non-geniculate corallines be understood and applied at the correct taxonomic rank. Finally, phylogenetic analyses of DNA sequences support recognition of the Hapalidiales as a distinct order characterized by having multiporate tetra/bisporangial conceptacles, and not as a suborder of Corallinales whose tetra/bisporangial conceptacles are uniporate.     

Sequencing type material resolves the identity and distribution of the generitype Lithophyllum incrustans,and related European species L. hibernicum and L. bathyporum (Corallinales,Rhodophyta)

DNA sequences from type material in the nongeniculate coralline genus Lithophyllum were used to unambiguously link some European species names to field‐collected specimens, thus providing a great advance over morpho‐anatomical identifi‐cation. In particular, sequence comparisons of rbcL, COI and psbA genes from field‐collected specimens allowed the following conclusion: the generitype species, L. incrustans, occurs mostly as subtidal rhodoliths and crusts on both Atlantic and Mediterranean coasts, and not as the common, NE Atlantic, epilithic, intertidal crust reported in the literature. The heterotypic type material of L. hibernicum was narrowed to one rhodolith belonging in Lithophyllum. As well as occurring as a subtidal rhodolith, L. hibernicum is a common, epilithic and epizoic crust in the intertidal zone from Ireland south to Mediterranean France. A set of four features distinguished L. incrustans from L. hibernicum, including epithallial cell diameter, pore canal shape of sporangial conceptacles and sporangium height and diameter. An rbcL sequence of the lectotype of Lithophyllum bathyporum, which was recently proposed to accommodate Atlantic intertidal collections of L. incrustans, corresponded to a distinct taxon hitherto known only from Brittany as the subtidal, bisporangial, lectotype, but also occurs intertidally in Atlantic Spain. Specimens from Ireland and France morpho‐anatomically identified as L. fasciculatum and a specimen from Cornwall likewise identified as L. duckerae were resolved as L. incrustans and L. hibernicum, respectively.     

High species richness and genetic diversity of the genus Caloglossa (Delesseriaceae,Rhodophyta) in New Ireland,Papua New Guinea

Distribution patterns and genetic diversity of Caloglossa species have been studied in various regions but are still poorly understood around the tropical western Pacific where many marine organisms show high species richness. We explored the diversity of Caloglossa species in New Ireland, Papua New Guinea using morphological and genetic analyses. Seven species of Caloglossa (C. adhaerens, C. leprieurii, C. ogasawaraensis, C. bengalensis, C. postiae, C. saigonensis and C. vieillardii) were collected from eight sites; the latter four species are new records in Papua New Guinea. All specimens collected were cultured to compare their morphology under the same culture condition. Reproductive structures of gametophytes and/or sporophytes were described for the first time in some species from these collections. In C. leprieurii, gall‐like structure appeared in most gametophyte strains but not in tetrasporophytes. Such galls were produced from wing cells, and spermatangia and/or procarps were formed on many galls. The LSU rDNA and rbcL analyses revealed high intraspecific diversity in many species although the number of samples was limited and the sampling sites were within 200 km. We revealed high species richness and intraspecific genetic variation in Papua New Guinea, suggesting a hot spot for the diversity of Caloglossa.