PROPOSAL OF THE GRACILARIALES ORD. NOV. (RHODOPHYTA) BASED ON AN ANALYSIS OF THE REPRODUCTIVE DEVELOPMENT OF GRACILARIA VERRUCOSA1

The mode of division of vegetative cells, formation of spermatangial parent cells, initiation of the carpogonial branch apparatus, and formation of tetrasporangial initials are homologous developmental processes that are documented for the first time in the type species of the economically important family Gracilariaceae, Gracilaria verrucosa (Hudson) Papenfuss from the British Isles. G. verrucosa is characterized by a supporting cell of intercalary origin that bears a 2-celled carpogonial branch flanked by two sterile branches, direct fusion of cells of sterile branches onto the carpogonium, formation of an extensive carpogonial fusion cell through the incorporation of additional gametophytic cells prior to gonimoblast initiation, gonimoblast initials produced from fusion cell lobes, schizogenous development of the cytocarp cavity, inner gonimoblast cells producing tubular nutritive cells that fuse with cells of the pericarp or floor of the cystocarp, absence of cytologically modified tissue in the floor of the cystocarp, and carposporangial initials produced in clusters or irregular chains. Spermatangial parent cells are generated in flaments from intercalary cortical cells that line an intercellular space forming a ‘pit’ or ‘conceptacle’. Tetrasporangial initials are transformed from terminal cells derived through division of an outer cortical cell. Tetrasporangia are cruciately divided. The Gracilariaceae is removed from Gigartinales and transferred to the new order Gracilariales. Their closest living relatives appear to be agarophytes belonging to the Gelidiales and Ahnfeltiales.     

Structure of reproductive apparatus of <Emphasis Type="Italic">Gracilaria/Gracilariopsis lemaneiformis</Emphasis> (Gracilariaceae,Rhodophyta)

Reproductive apparatus of Gracilaria/Gracilariopsis lemaneiformis collected from Qingdao city were studied with a light and a transmission electron microscope. The special superficial arrangement of spermatangium for this species was clearly observed, and the ultrastructure of spermatangial development revealed the similar cytodynamic pattern followed by all the Gracilariaceae members developed from spermatangial mother cells to spermatangium. The female reproductive apparatus before fertilization was also observed and trichogyne was found protruding above the cortex, contrary to the earlier reports. Tetrasporangium was formed by an outer cortical cell and the tetraspores became spherical and expended after being released.     

MOLECULAR DELINEATION OF SPECIES AND SPECIES RELATIONSHIPS IN THE RED ALGAL AGAROPHYTES GRACILARIOPSIS AND GRACILARIA (GRACILARIALES)1

Delineation of species in the economically important agarophyte genera Gracilaria and Gracilariopsis has proven extremely difficult using available morphological characteristics. In this study, we examine the usefulness of two transcribed spacers for molecular systematic studies of these genera. The polymerase chain reaction was used to amplify the internal transcribed spacers (ITSs) and the intervening 5.8S ribosomal DNA of the nuclear ribosomal repeat region. In addition, a plastid spacer region and flanking regions of coding genes were amplified from the RUBISCO operon. Both regions were sequenced for individuals and populations of Gracilariopsis lemaneiformis (Bory) Dawson, Acleto, et Foldvik to determine the usefulness of these spacers in delimiting populations. These studies reveal that there is as much variation among individuals of a population as there is between individuals of geographically separate populations. In addition, the ITS spacer regions were compared between different species of Gracilariopsis and Gracilaria. The nuclear ITS spacer region is conserved at a species level in both genera and provides phylogenetically informative characters that can be used to examine species interrelationships among relatively closely related taxa. However, because of the difficulties of aligning this entire region among species from the two genera, the ITS region is not useful for examining intergenera relationships. ITS interspecies sequence comparisons indicate that Gracilariopsis lemaneiformis from California is significantly different from G. lemaneiformis from China and that a species of Gracilariopsis from Peru is more closely related to G. lemaneiformis from North Carolina than it is to the other Gracilariopsis species examined. In addition, these studies indicate that Gracilaria chilensis Bird, McLachlan, et Oliveira from New Zealand and Gracilaria tenuistipitata Chang et Xia from southeast Asia are as closely related as are Gracilaria verrucosa (Hudson) Papenfuss, G. pacifica Abbott, and Gracilaria robusta Kylin. Phylogenetic analysis of aligned plastid spacer sequences from Gracilaria and Gracilariopsis taxa provide similar conclusions about species relationships.     

CHARACTERIZATION OF SCHIZOSERIS CONDENSATA,SCHIZOSERIDEAE TRIB. NOV. (DELESSERIACEAE,RHODOPHYTA)1

The Myriogramme group of Kylin contains two distinct clusters of genera that merit recognition at the tribal level. We previously established the tribe Myriogrammeae, and in this paper we erect the Schizoserideae based on a study of the type species of Schizoseris, S. laciniata (=S. condensata), from the southern hemisphere. The Schizoserideae is characterized by 1) marginal and diffuse intercalary meristems; 2) nuclei initially arranged in a plate in the median plane in meristematic and mature cells; 3) chloroplasts one to few, lobed or dissected; 4) microscopic veins absent; 5) procarps scattered, formed singly on either side of the blade with cover cells absent and consisting of a one- to two-celled lateral sterile group, a one- to two-celled basal sterile group, and a four-celled carpogonial branch in which the trichogyne passes beneath the lateral sterile group and emerges anterior to it; 6) auxiliary cell diploidized by a connecting cell cut off posteriolaterally from the fertilized carpogonium; 7) gonimoblast initial cut off laterally from one side of the auxiliary cell and giving rise to unilaterally branched gonimoblast filaments bearing carposporangia in branched chains; 8) gonimoblast fusion cell highly branched, candelabra-like, incorporating all but the basalmost cells of the carposporangial chains and radiating through the central cells in the floor of the cystocarp; 9) spermatangial and tetrasporangial sori formed from surface cells in both monostromatic and polystromatic portions on both sides of the blade; and 10) tetrasporangia formed primarily from cortical rather than from central cells. The Schizoserideae presently includes Schizoseris Kylin, Neuroglossum Kützing, Abroteia J. Agardh, and Polycoryne Skottsberg in Kylin and Skottsberg.     

Structure of reproductive apparatus of Gracilaria/ Gracilariopsis lemaneiformis (Gracilariaceae, Rhodophyta)

Reproductive apparatus of Gracilaria/Gracilariopsis lemaneiformis collected from Qingdao city were studied with a light and a transmission electron microscope. The special superficial arrangement of spermatangium for this species was clearly observed, and the ultrastructure of spermatangial development revealed the similar cytodynamic pattern followed by all the Gracilariaceae members developed from spermatangial mother cells to spermatangium. The female reproductive apparatus before fertilization was also observed and trichogyne was found protruding above the cortex, contrary to the earlier reports. Tetrasporangium was formed by an outer cortical cell and the tetraspores became spherical and expended after being released. Supported by the National Natural Sciences Foundation of China (Grant No. 40606034) and the National High-Tech Research and Development Program of China (Grant No. 2006AA10A413)     

Structure of reproductive apparatus of Gracilaria/Gracilariopsis lemaneiformis (Gracilariaceae,Rhodophyta)

Reproductive apparatus of Gracilaria/Gracilariopsis lemaneiformis collected from Qingdao city were studied with a light and a transmission electron microscope. The special superficial arrangement of spermatangium for this species was clearly observed, and the ultrastructure of spermatangial devel-opment revealed the similar cytodynamic pattern followed by all the Gracilariaceae members devel-oped from spermatangial mother cells to spermatangium. The female reproductive apparatus before fertilization was also observed and trichogyne was found protruding above the cortex, contrary to the earlier reports. Tetrasporangium was formed by an outer cortical cell and the tetraspores became spherical and expended after being released.     

Gracilaria vermiculophylla: A western Pacific species of Gracilariaceae (Rhodophyta) first recorded from the eastern Pacific

We report Gracilaria vermiculophylla (Ohmi) Papenfuss from the Pacific coast of North America based on a morphoanatomical revision and comparison of sequences of the nuclear gene coding for the small subunit of ribosomal RNA and the internal transcribed spacers of populations from Baja California, Mexico and Hiroshima, Japan. It is the first convincing report of this species out of its center of distribution in western north Pacific, where it has been considered as a synonym of former ‘G. verrucosa’ records. G. vermiculophylla also occurs in central California as indicated by internal transcribed spacer sequences of a previously unknown Gracilaria Greville material. In the northeastern Pacific G. vermiculophylla is characterized by a robust somewhat vermiform, well‐branched cylindrical thallus, with gradual cell size transition from cortex to medulla, deep spermatangial conceptacles, regular chains of carposporangia and carposporangial initials, downwardly oriented tubular nutritive cells, but rarely with upwardly oriented tubular nutritive cells. In some features of its cystocarp anatomy G. vermiculophylla is related to Gracilariopsis E. Y. Dawson or Hydropuntia Montagne and it is a relevant species for discussions about Gracilariaceae genera.     

LEACHIELLA PACIFICA,GEN. ET SP. NOV., A NEW PARASITIC RED ALGA FROM WASHINGTON AND CALIFORNIA

Leachiella pacifica, gen. et sp. nov., a marine alloparasitic red alga is described from Washington and California. Several species of Polysiphonia and Pterosiphonia are hosts for this parasite. The thallus is a white, multiaxial, unbranched pustule with rhizoidal filaments that ramify between host cells, forming numerous secondary pit connections with host cells. All reproductive structures develop from outer cortical cells. Tetrasporocytes, situated on stalk cells, undergo simultaneous, tetrahedral cleavage to form tetraspores. Spermatia are formed continuously by oblique cleavages of the elongate spermatial generating cells. This results in spermatial clusters consisting of 4–8 spermatia in an alternate arrangement. Carposporophyte development is procarpial. The carpogonium is part of a six-celled branch including a sterile cell that is formed by the basal cell. The carpogonial branch is attached laterally to an obovate supporting cell that also forms an auxiliary cell, presumably formed prior to fertilization. After fertilization the carpogonium temporarily fuses with the auxiliary cell apparently to transfer the diploid nucleus and initiate further fusion with the subtending supporting cell to form an incipient fusion cell. The auxiliary cell portion of this fusion cell divides to form gonimoblast initials that continue to divide, forming gonimoblast filaments whose terminal cells differentiate into carpospores. The remainder of the fusion cell enlarges by continual fusion with adjacent vegetative cells. The resultant carposporophyte consists of a basal, multinucleate fusion cell supporting a hemispherical cluster of gonimoblast filaments with terminally borne carpospores. Vegetatively, Leachiella resembles several other parasitic red algae but it is clearly separated by the procarp, carposporophyte development and structure, and tetrasporocyte cleavage.     

ONTOGENY OF SEXUAL CONCEPTACLES IN A SPECIES OF BOSSIELLA (CORALLINACEAE)1

The first evidence of conceptacle formation in male and female plants of Bossiella californica ssp. Schmittii is the secretion of a cap below the epithallium. Subsequent development involves the disintegration of the upper enucleate parts of cortical cells below the center of the cap concomitant with the growth of tissue surrounding these cells. Protoplasts in the lower parts of these cortical cells recover and become initials of reproductive cells. Differentiation of each initial in female plants results in a supporting cells bearing 1 or 2, 2-celled carpogonial filaments, and in male plants results in a basal cell bearing 2 or 3 spermatangial mother cells which bud off spermatangia. Following fertilization, supporting cells fuse and form a broad, thin fusion cell from the upper surface of which gonimoblast (carposporangial) filaments arise.     

PHYLOGENY OF ATLANTIC SPECIES OF GRACILARIACEAE (RHODOPHYTA) BASED ON SMALL SUBUNIT RIBOSOMAL GENE SEQUENCING

The complete small subunit ribosomal nuclear gene (ssu rDNA) sequence was determined for nine species of Gracilaria and one species of Gracilariopsis which are common on the American Atlantic waters. The sequences were aligned using the secondary structure as reference, including the published sequences of nine other species of Gracilariaceae. A matrix of 1736 sites was constructed with a proportion of 91% invariable sites and very few assumed indels events. All the inferred trees show three main lineages: 1) the strongly divergent lineages of Gracilariopsis; 2) the austral genera Curdiea / Melanthalia; and 3) the lineage of Gracilaria sensu stricto. The later encompasses the following groups: 1) Gracilaria chilensis from the Pacific ocean; 2) a group of cylindrical tropical species with "henriquesiana" spermatangial type; 3) a group of warm temperate cylindrical species with "verrucosa" spermatangial type; and 4) a group of flattened tropical species with mainly "textorii" spermatangial type. The relationship of a species described as Gracilaria pauciramosa from Venezuela was not inequivocally solved. The inferred phylogenetic groups are congruent with morphology and quality of agar.     

The morphology and life history of Mazoyerella gen. nov. (M. arachnoidea (Harvey) comb. nov.)—Rhodophyta,Ceramiaceae—from Southern Australia

Mazoyerella gen. nov., based on Corynospora arachnoidea Harvey from Tasmania, is placed provisionally in the Compsothamnieae tribe of the Ceramiaceae. All plants can produce elongate-ovoid monosporangia which reproduce the same phase. Sporophytes also bear polysporangia and sexual plants are dioecious, bearing either spermatangial heads or subterminal procarps followed by carposporophytes with nearly all gonimoblast cells becoming carposporangia and an involucrum of sterile filaments developing from sterile cells associated with the procarp. The life-history has been followed through successive generations in culture.     

CHARACTERIZATION OF MYHOGRAMME LNIDA, MYRIOGRAMMEAE TRIB. NOV (DELESSEIUACEAE, FWODOPHYTA)

The Myriogramme group of Kylin was found to contain two distinct clusters of genera that merit recognition at the tribal level. In this paper, we establish the tribe Myriogrammae based on a study of the type species of Myriogramme, M. livida, from the Southern Hemisphere. The Myriogrammae is characterized by 1) marginal and diffuse intercalary meristems; 2) nuclei arranged in a ring bordering the side walls of vegetative cells; 3) microscopic veins absent; 4) procarps scattered, formed opposite one another on both sides of the blade posterior to one or more vegetative pericentral cells (cover cells) and consisting of a carpogonial branch, a one-/to two-celled lateral sterile group and a one-celled basal sterile group; 5) auxiliary cell diploidized by a connecting cell cut off posteriolaterally from the fertilized carpogonium; 6) gonimoblast initial cut off distally from the auxiliary cell, generating one distal and one to two lateral gonimoblast filaments that branch in the plane of the expanding cystocarp cavity and later fuse to from an extensive, branched fusion cell; 7) spermatangial and tatrasporangial sori formed inside the margin on both sides of the blade by resumption of meristematic activity; and 8) tetrasporangia produced primarily from the central cells. The Myriogrammae currently includes Myriogramme Kylin , Gonimocolax Kylin , Haraldiophyllum A. Zinova , Hideophyllum A. Zinova, and a possible undescribed genus from Pacific North and South America. Genera are separated based primarily on features of gonimoblast and carposporangial development .     

A Light Microscope Study of Carposporophyte Development in Gracilaria verrucosa (Hudson) Papenfuss

Gracilaria verrucosa is a very common marine red alga of Greekcoasts. The diploid carposporophyte which develops attachedto the female gametophyte of Gracilaria is described. The immaturecystocarps are very small while the mature ones are globose,ostiolate and are borne profusely all over the surface of thethallus. The earliest observed fusion cell is small and fusesprogressively with adjacent vegetative cells to form a largemultinucleate cell. From this fusion cell gonimoblast initialsoriginate, dividing further and giving rise to a large numberof gonimoblast cells. The resultant carposporophyte consistsof a basal-central, multinucleate cell surrounded by a conicalor hemispherical mass of gonimoblast cells. Chains or clustersof successively maturing carpospores are borne from the terminalgonimoblast cells. The liberation of mature carpospores takesplace through the ostiole of the cystocarp. The liberated carposporeslack cell walls and are naked in a mucilage mass. Gracilaria verrucosa (Hudson) Papenfuss, Gigartinales, Gracilariaceae, Rhodophyta, carposporophyte, development     

GRACILARIOPSIS MCLACHLANII SP. NOV. AND GRACILARIOPSIS PERSICA SP. NOV. OF THE GRACILARIACEAE (GRACILARIALES,RHODOPHYCEAE) FROM THE INDIAN OCEAN1

Two new species of Gracilariopsis from the Indian Ocean are proposed—Gracilariopsis (Gp.) mclachlanii Buriyo, Bellorin et M. C. Oliveira sp. nov. from Tanzania and Gracilariopsis persica Bellorin, Sohrabipour et E. C. Oliveira sp. nov. from Iran—based on morphology and DNA sequence data (rbcL gene and SSU rDNA). Both species fit the typical features of Gracilariopsis: axes cylindrical throughout, freely and loosely ramified up to four orders, with an abrupt transition in cell size from medulla to cortex, cystocarps lacking tubular nutritive cells and superficial spermatangia. Nucleotide sequence comparisons of rbcL and SSU rDNA placed both species into the Gracilariopsis clade as distinct species from all the accepted species for this genus, forming a deeply divergent lineage together with some species from the Pacific. The new species are very difficult to distinguish on morphological grounds from other species of Gracilariopsis, stressing the importance of homologous molecular marker comparisons for the species recognition in this character‐poor genus.     

PHYLOGENY AND SYSTEMATICS OF THE MARINE ALGAL FAMILY GRACILARIACEAE (GRACILARIALES,RHODOPHYTA) BASED ON SMALL SUBUNIT rDNA AND ITS SEQUENCES OF ATLANTIC AND PACIFIC SPECIES1

We sequenced the small subunit rDNA and internal transcribed spacer region of Gracilariaceae from the tropical Atlantic and Pacific, with emphasis on flattened or compressed species. Sequence comparisons confirmed three main lineages of Gracilariaceae: Curdiea/Melanthalia, Gracilariopsis/Gracilariophila, and Gracilaria. The Curdiea/Melanthalia diverged early in the family. Gracilariopsis was paraphyletic, because at least one Gracilariophila species evolved from it. The Atlantic Gracilariopsis were monophyletic and separated from the Pacific lineages. The Gracilaria included all species referable to its own species and to Hydropuntia, which was paraphyletic, formed by distantly related lineages. The new combination Gracilaria pauciramosa (N. Rodríguez Ríos) Bellorin, M. C. Oliveira et E. C. Oliveira is proposed for Polycavernosa pauciramosa N. Rodríguez Ríos. Recognition of subgenera within Gracilaria, based on spermatangial arrangement, was not supported. Instead, infrageneric groups were delineated by geographic origins and combinations of reproductive characters. Most Pacific species with either “textorii” or “verrucosa” type spermatangia were deeply separated from Atlantic species. Within the Atlantic Gracilaria, a lineage encompassing mostly tropical cylindrical species with “henriquesiana” type spermatangia and distinctive cystocarp anatomy was recognized. A lineage was also retrieved for cold water stringy species with verrucosa type spermatangia. Several species from the western Atlantic are closely related to Gracilaria tikvahiae McLachlan with nearly identical morphology. On the other hand, most flattened species from the tropical Atlantic were closely related despite their diverse morphologies. The interpretation of our data in addition to the literature indicates that more populations from the Indo‐Pacific must be studied before a general picture of Gracilariaceae evolution can be framed.     

Reproductive morphology of Gastroclonium pacificum (Champiaceae,Rhodymeniales) from Japan

Morphological studies were undertaken on Gastroclonium pacificum (E.Y. Dawson) C.F. Chang et B.M. Xia (Champiaceae, Rhodymeniales) from Japan. We describe the details of male reproductive structures, the procarp and early post‐fertilization stages. This species has a solid axis, with both tetrasporangia and polysporangia, spermatangia are cut off from spermatangial parent cells, and a procarp is composed of a 4‐celled carpogonial branch and two 2‐celled auxiliary cell branches. The mature cystocarp lacks a conspicuous ostiole, a characteristic of the genus Gastroclonium. The most distinctive characteristic of the species is the tela arachnoidea, which is lacking in other species of Gastroclonium.     

STRUCTURE AND DEVELOPMENT OF THE CARPOSPOROPHYTE OF NEMALION HELMINTHOIDES (NEMALIONALES,RHODOPHYTA)1

Following fertilization, the carposporophyte of Nemalion helminthoides (Velley in With.) Batters differentiates into four distinct regions: the fusion cell, the sterile gonimoblast cells, the carposporangial mother cells and the carposporangia. The gonimoblast is formed by apically dividing, monopodial filaments of limited growth which may later become pseudodichotomous. Upon differentiation of a terminal carposporangium, a gonimoblast filament may continue to grow sympodially. A single carposporangial mother cell may produce carposporangia in several different directions as well as proliferate successive carposporangia within the sporangial walls that remain after carpospore liberation. As the carposporophyte matures, the gonimoblast initial, the stalk cell, the hypogynous and subhypogynous cells fuse. Except for the fusion cell, all cells of the carposporophyte show organelle polarity and contain a distally located, lobed chloroplast and proximal nucleus.