Genetic structure of Japanese Chattonella marina (Raphidophyceae) populations revealed using microsatellite markers

Chattonella marina var. antiqua and C. marina var. marina (Raphidophyceae) are red tide‐forming, harmful phytoplankton species. We investigated the genetic diversity and genetic relationship among the populations using microsatellite markers to identify putative sources of C. marina var. antiqua and C. marina var. marina in Japanese coastal populations. A positive correlation between genetic divergence and geographical distance (isolation by distance) was recognized for C. marina var. antiqua. The C. marina var. antiqua populations were established throughout a geological time scale, and genetic divergence had progressed in each population with gene flow depending on geographic distances. In contrast, isolation by distance was not observed for C. marina var. marina populations, and the genetic divergence among populations was extremely high. The Tokyo Bay population of C. marina var. marina, which was first recognized in 2008, had many private alleles but was related to the Kagoshima Bay population. The Tokyo Bay population may have been established by several invasions from the Kagoshima Bay population and other regions.     

Isolation and characterization of chloroplast microsatellite markers in four mangrove species,Aegiceras corniculatum,Avicennia marina,Acanthus ilicifolius and Lumnitzera racemosa

One, three, seven, and six of polymorphic chloroplast microsatellite (cpSSR) markers were developed from four mangrove species, Acanthus ilicifolius, Aegiceras corniculatum, Avicennia marina, and Lumnitzera racemosa, respectively. Characterization of 229, 509, 369, and 216 individuals of A. ilicifolius, A. corniculatum, A. marina, and L. racemosa, collected from different natural mangrove populations (A. ilicifolius, 6; A. corniculatum, 14; A. marina, 10; L. racemosa, 6) in the southern coastline of China showed that these loci provide cpSSR markers with polymorphisms ranging from two to four alleles per locus and gene diversity between 0.005 and 0.675. Combining the polymorphic cpSSR loci of each species, 3, 5, 11, and 4 of cpSSR haplotypes were separately detected in populations of A. ilicifolius, A. corniculatum, A. marina, and L. racemosa in the southern coastline of China. These cpSSR markers will be useful for analyzing the maternal lineage distributions and population genetic structures of these four species.     

MORPHOLOGICAL AND GENETIC VARIATION IN THE POPULATIONS OF SARGASSUM HEMIPHYLLUM (PHAEOPHYCEAE) IN THE NORTHWESTERN PACIFIC1

Difficulty in species identification of Sargassum (Sargassaceae, Fucales) is partly attributed to the high polymorphism among its individuals and populations. This study aimed at assessing morphological and genetic variations in two varieties, var. hemiphyllum J. Agardh and var. chinense J. Agardh, of Sargassum hemiphyllum (Turner) C. Agardh, a widely distributed species in the northwestern Pacific. We investigated 26 measurable, five numerical, and 33 categorical morphological parameters associated with different branching levels of specimens from each of six localities within its distribution range using cluster analysis (CA) and principal coordinate analysis (PCoA). Leaf size of the primary and secondary branching levels and the vesicle size of the secondary branches of the specimens examined were determined to be the most important morphological parameters that were significantly different among populations. Change in leaf and vesicle length of individuals among the six populations followed a latitudinal gradient, with smaller leaves and vesicles associated with northern populations and larger ones in the southern populations. The possible influence of the gradual change in sea surface temperatures (SSTs) along this gradient in the northwestern Pacific on leaf and vesicle morphologies of this species was suggested. PCR‐RFLP analysis of the RUBISCO spacer in the chloroplast genome revealed two distinct and highly homogenous clades, a China clade and a Japan‐Korea clade, which corresponded to var. chinense and var. hemiphyllum, respectively. The formation of refugia along the “Paleo‐coast” in the East China Sea during glacial periods is suggested to have led to the vicariance of ancestral populations of S. hemiphyllum and thus to have promoted genetic differentiation. The massive freshwater outflow of the Yellow and Yangtze rivers may continue to act as a barrier, prolonging the allopatric distribution of the two varieties.     

Redefinition of Dodecatheon dentatum (Primulaceae) and rationale for use of varietal rank

The new variety,Dodecatheon dentatum var.utahense, represents a geographic stepping stone betweenD. dentatum var.dentatum of the Pacific Northwest andD. dentatum var.ellisiae of the Southwest. Rationale for using the infraspecific rank of variety includes historical precedence and preponderant usage, convenience of grouping multiple varieties into subspecies, and elimination of subjectivity in grading the magnitude of genetic differences.     

GENETIC DIVERGENCE AND ISOZYME NUMBER VARIATION AMONG FOUR VARIETIES OF ALLIUM DOUGLASII (ALLIACEAE)

Allium douglasii comprises four, allopatric varieties confined to the northwestern United States: var. columbianum, var. constrictum, var. douglasii, and var. nevii. Although no apparent karyotypic differences distinguish the four taxa, evidence from starch gel electrophoresis indicates that they are very distinct at genes coding for enzymes. The mean genetic identity for pairwise comparisons of populations of all four varieties is 0.84. This value is lower than identity values generally observed for subspecific taxa, and may be indicative of a long period of geographic separation and subsequent lack of gene flow among the varieties. However, a higher genetic identity of 0.93 was observed between var. constrictum and var. columbianum relative to other varietal comparisons. In addition, var. constrictum exhibited two biochemical autapomorphies: 1) loss of expression for the gene coding for Tpi-3, and 2) duplication or reactivation of an ADH gene. These data indicate that var. constrictum may be a recent derivative of var. columbianum. The distribution of genetic variation within and among populations of the putative progenitor/derivative pair also supports this hypothesis.     

Population genetic structure of annual and perennial populations of Zostera marina L. along the Pacific coast of Baja California and the Gulf of California

The Baja California peninsula represents a biogeographical boundary contributing to regional differentiation among populations of marine animals. We investigated the genetic characteristics of perennial and annual populations of the marine angiosperm, Zostera marina, along the Pacific coast of Baja California and in the Gulf of California, respectively. Populations of Z. marina from five coastal lagoons along the Pacific coast and four sites in the Gulf of California were studied using nine microsatellite loci. Analyses of variance revealed significant interregional differentiation, but no subregional differentiation. Significant spatial differentiation, assessed using θ_ST values, was observed among all populations within the two regions. Z. marina populations along the Pacific coast are separated by more than 220 km and had the greatest θ_ST (0.13–0.28) values, suggesting restricted gene flow. In contrast, lower but still significant genetic differentiation was observed among populations within the Gulf of California (θ_ST = 0.04–0.18), even though populations are separated by more than 250 km. This suggests higher levels of gene flow among Gulf of California populations relative to Pacific coast populations. Direction of gene flow was predominantly southward among Pacific coast populations, whereas no dominant polarity in the Gulf of California populations was observed. The test for isolation by distance (IBD) showed a significant correlation between genetic and geographical distances in Gulf of California populations, but not in Pacific coast populations, perhaps because of shifts in currents during El Niño Southern Oscillation (ENSO) events along the Pacific coast.     

PHYLOGEOGRAPHY OF THE PANTROPICAL SEA URCHIN EUCIDARIS IN RELATION TO LAND BARRIERS AND OCEAN CURRENTS

The pantropical sea urchin genus Eucidaris contains four currently recognized species, all of them allopatric: E. metularia in the Indo-West Pacific, E. thouarsi in the eastern Pacific, E. tribuloides in both the western and eastern Atlantic, and E. clavata at the central Atlantic islands of Ascension and St. Helena. We sequenced a 640-bp region of the cytochrome oxidase I (COI) gene of mitochondrial DNA to determine whether this division of the genus into species was confirmed by molecular markers, to ascertain their phylogenetic relations, and to reconstruct the history of possible dispersal and vicariance events that led to present-day patterns of species distribution. We found that E. metularia split first from the rest of the extant species of the genus. If COI divergence is calibrated by the emergence of the Isthmus of Panama, the estimated date of the separation of the Indo-West Pacific species is 4.7–6.4 million years ago. This date suggests that the last available route of genetic contact between the Indo-Pacific and the rest of the tropics was from west to east through the Eastern Pacific Barrier, rather than through the Tethyan Sea or around the southern tip of Africa. The second cladogenic event was the separation of eastern Pacific and Atlantic populations by the Isthmus of Panama. Eucidaris at the outer eastern Pacific islands (Galapagos, Isla del Coco, Clipperton Atoll) belong to a separate clade, so distinct from mainland E. thouarsi as to suggest that this is a different species, for which the name E. galapagensis is revived from the older taxonomic literature. Complete lack of shared alleles in three allozyme loci between island and mainland populations support their separate specific status. Eucidaris galapagensis and E. thouarsi are estimated from their COI divergence to have split at about the same time that E. thouarsi and E. tribuloides were being separated by the Isthmus of Panama. Even though currents could easily convey larvae between the eastern Pacific islands and the American mainland, the two species do not appear to have invaded each other's ranges. Conversely, the central Atlantic E. clavata at St. Helena and Ascension is genetically similar to E. tribuloides from the American and African coasts. Populations on these islands are either genetically connected to the coasts of the Atlantic or have been colonized by extant mitochondrial DNA lineages of Eucidaris within the last 200,000 years. Although it is hard to explain how larvae can cross the entire width of the Atlantic within their competent lifetimes, COI sequences of Eucidaris from the west coast of Africa are very similar to those of E. tribuloides from the Caribbean. F_ST statistics indicate that gene flow between E. metularia from the Indian Ocean and from the western and central Pacific is restricted. Low gene flow is also evident between populations of E. clavata from Ascension and St. Helena. Rates of intraspecific exchange of genes in E. thouarsi, E. galapagensis, and E. tribuloides, on the other hand, are high. The phylogeny of Eucidaris confirms Ernst Mayr's conclusions that major barriers to the dispersal of tropical echinoids have been the wide stretch of deep water between central and eastern Pacific, the cold water off the southwest coast of Africa, and the Isthmus of Panama. It also suggests that a colonization event in the eastern Pacific has led to speciation between mainland and island populations.     

Population variation and phylogeny in the endangered <Emphasis Type="BoldItalic">Chamaesyce skottsbergii</Emphasis> (Euphorbiaceae) based on RAPD and ITS analyses

Chamaesyce skottsbergii var. skottsbergii is federally listed as an endangered taxon, and is found in small and isolated populations restricted to calcareous soils in dry shrubland habitats on the Hawaiian islands of Oahu and Molokai. Concern over the genetic relationship among these disjunct populations arose as a result of threats to the habitat of the Oahu population. The populations were examined using random amplified polymorphic DNA (RAPD) markers and sequence analysis of the internal transcribed spacer (ITS) region of the rDNA cistron. Chamaesyce skottsbergii var. vaccinioides, a closely related variety found in several small populations on Molokai, was used for baseline comparison of the genetic divergence among populations. RAPD analysis demonstrated that variation within and among populations is the highest for any Hawaiian species examined. Polymorphism was greater than 95% within populations and was 99.4% at the species level. Similarly, measures of genetic similarity indicate that differentiation among these populations is higher than is known for some species. Both RAPD and ITS sequence analysis indicate that populations of C. skottsbergii var. skottsbergii on Oahu and Molokai are genetically distinct, and the extent of this genetic differentiation supports the recognition of these populations as distinct varieties. The Molokai population is in fact much more closely related to var. vaccinioides than to var. skottsbergii on Oahu, and thus should be recognized by the previously used variety name, C. skottsbergii var. audens. Further conservation measures for each of the varieties are addressed.     

TAXONOMY OF ZEA (GRAMINEAE). II. SUBSPECIFIC CATEGORIES IN THE ZEA MAYS COMPLEX AND A GENERIC SYNOPSIS

A compromise classification of the genus Zea, reflecting both phylogeny and practical needs, recognizes six taxa, as follows: Section Luxuriantes : Zea perennis. Zea diploperennis, Zea luxurians. Section Zea : Zea mays ssp. mexicana (Neo-volcanic Plateau), Zea mays ssp. parviglumis Iltis & Doebley ssp. n. var. parviglumis (Rio Balsas drainage, Pacific slope from Guerrero to Jalisco), Zea mexicana ssp. parviglumis var. huehuetenangensis Iltis & Doebley var. n. (Pacific slope, western Guatemala, Prov. Huehuetenango), Zea mays ssp. mays. The new subspecies is distinguished by smaller spikelets and rachis joints, the varieties by different habitats, blooming dates and their genetic behavior in relation to cultivated Zea mays. Zea mays ssp. mexicana is the ancestor of corn.     

INTERPOPULATIONAL VARIATION IN THE GAMETOPHYTES OF PELLAEA ANDROMEDAEFOLIA

The development and mature morphology of the gametophytes from both sexual and apogamous populations of the fern Pellaea andromedaefolia were investigated. While most sexual examples were indistinguishable, some differences were noted. An insular collection was distinctive in its variability and irregularity of form. Although the latter was a representative of var. pubescens, other collections of the variety could not be distinguished from var. andromedaefolia on the basis of gametophytic characteristics. The apogamous gametophytes were decidedly more variable in development and often very different from sexual thalli. The mature asexual thalli tended to be more irregular in form and usually sharply divergent from the typical cordate type characteristic of the sexual populations. Each of the five apogamous samples was unique with respect to gametophyte development. The differences among the gametophytes of the various populations do not correlate with the sporophytic characteristics which differentiate the two varieties of the species.     

Highly restricted gene flow and deep evolutionary lineages in the giant clam <Emphasis Type="Italic">Tridacna maxima</Emphasis>

The tropical Indo-West Pacific is the biogeographic region with the highest diversity of marine shallow water species, with its centre in the Indo-Malay Archipelago. However, due to its high endemism, the Red Sea is also considered as an important centre of evolution. Currently, not much is known about exchange among the Red Sea, Indian Ocean and West Pacific, as well as connectivity within the Indo-Malay Archipelago, even though such information is important to illuminate ecological and evolutionary processes that shape marine biodiversity in these regions. In addition, the inference of connectivity among populations is important for conservation. This study aims to test the hypothesis that the Indo-Malay Archipelago and the Red Sea are important centres of evolution by studying the genetic population structure of the giant clam Tridacna maxima. This study is based on a 484-bp fragment of the cytochrome c oxidase I gene from 211 individuals collected at 14 localities in the Indo-West Pacific to infer lineage diversification and gene flow as a measure for connectivity. The analysis showed a significant genetic differentiation among sample sites in the Indo-West Pacific (Φ_st = 0.74, P < 0.001) and across the Indo-Malay Archipelago (Φ_st = 0.72, P < 0.001), indicating restricted gene flow. Hierarchical AMOVA revealed the highest fixation index (Φ_ct = 0.8, P < 0.001) when sample sites were assigned to the following regions: (1) Red Sea, (2) Indian Ocean and Java Sea, (3) Indonesian throughflow and seas in the East of Sulawesi, and (4) Western Pacific. Geological history as well as oceanography are important factors that shape the genetic structure of T. maxima in the Indo-Malay Archipelago and Red Sea. The observed deep evolutionary lineages might include cryptic species and this result supports the notion that the Indo-Malay Archipelago and the Red Sea are important centres of evolution. Communicated by Biology Editor Dr. Ruth Gates     

Towards a legal framework for systematic conservation: identification and development of allele-specific PCR markers for conspecific varieties of an endangered perennial herb <Emphasis Type="Italic">Primula kisoana</Emphasis> Miquel based on sequence variation of chloroplast DNA

Primula kisoana var. kisoana (Primulaceae) is a narrow endemic found only in extremely restricted areas of central Honshu Island, Japan. Although the species is included as ‘Critically Endangered (CR)’ in the latest Japanese Red List, it is not covered by legislation such as the ‘Law for the Conservation of Endangered Species of Wild Fauna and Flora, Japan’. This poor conservation status is due, largely, to the presence of another conspecific variety, var. shikokiana, which is regarded as less threatened than var. kisoana. In this study, we investigated the genetic dissimilarity between the two varieties by examining sequence variation in three noncoding regions of chloroplast DNA. Ten distinct haplotypes were detected, none of which were distributed across populations and varieties. The genetic differentiation between the two varieties revealed by an AMOVA and a minimum spanning network suggests that var. kisoana and var. shikokiana should be conserved and managed as separate units. Using the allele-specific PCR method, four primer pairs were developed at the point mutations and insertion/deletion loci that exhibited only inter-variety variation. These markers will facilitate reliable identification of var. kisoana, assisting in the implementation of restoration programs including the introduction of seedlings and construction of a legal framework for conservation.     

Inter-simple sequence repeat (ISSR) diversity within Monarda fistulosa var. brevis (Lamiaceae) and divergence between var. brevis and var. fistulosa in West Virginia

Inter-simple sequence repeat (ISSR) banding patterns were used to examine genetic diversity within and among populations ofMonarda fistulosa var.brevis, a rare taxon restricted to several populations in limestone glades and barrens in eastern West Virginia and Virginia. More than 34% of the total ISSR diversity in var.brevis occurred among populations, which is high when compared to the few other rare species that have been examined for ISSR variation. Prior studies demonstrated that var.brevis is morphologically distinct from the more widespread var.fistulosa, and that the differences are maintained when the two varieties are grown together in a uniform environment. The present study utilizing ISSR markers indicated that the two varieties are distinct, though quite similar genetically, and this is concordant with prior investigations documenting their morphological and habitat differences. However, the ISSR results suggest that the two varieties have diverged relatively recently and/or there is a low level of gene flow between them.     

The presence of a cryptic barrier in the West Pacific Ocean suggests the effect of glacial climate changes on a widespread sea‐dispersed plant,Vigna marina (Fabaceae)

Ocean currents are an important driver of evolution for sea‐dispersed plants, enabling them to maintain reciprocal gene flow via sea‐dispersed diaspores and obtain wide distribution ranges. Although geographic barriers are known to be the primary factors shaping present genetic structure of sea‐dispersed plants, cryptic barriers which form clear genetic structure within oceanic regions are poorly understood. To test the presence of a cryptic barrier, we conducted a phylogeographic study together with past demographic inference for a widespread sea‐dispersed plant, Vigna marina, using 308 individuals collected from the entire Indo‐West Pacific (IWP) region. Chloroplast DNA variation showed strong genetic structure that separated populations into three groups: North Pacific (NP), South Pacific (SP) and Indian Ocean (IN) (F′_CT among groups = 0.954–1.000). According to the Approximate Bayesian computation inference, splitting time between NP and SP was approximately 20,200 years (95%HPD, 4,530–95,400) before present. Moreover, a signal of recent population expansion was detected in the NP group. This study clearly showed the presence of a cryptic barrier in the West Pacific region of the distributional range of V. marina. The locations of the cryptic barrier observed in V. marina corresponded to the genetic breaks found in other plants, suggesting the presence of a common cryptic barrier for sea‐dispersed plants. Demographic inference suggested that genetic structure related to this cryptic barrier has been present since the last glacial maximum and may reflect patterns of past population expansion from refugia.     

GENETIC STRUCTURE OF GIANT CLAM (TRIDACNA MAXIMA) POPULATIONS IN THE WEST PACIFIC IS NOT CONSISTENT WITH DISPERSAL BY PRESENT-DAY OCEAN CURRENTS

The Pacific marine biota, particularly species with long planktonic larval stages, are thought to disperse widely throughout the Pacific via ocean currents. The little genetic data available to date has supported this view in that little or no significant regional differentiation of populations has been found over large geographical distances. However, recent data from giant clams has demonstrated not only significant regional differentiation of populations, but routes of gene flow that run perpendicular to the main present-day ocean currents. Extensive surveys of genetic variation at eight polymorphic loci in 19 populations of the giant clam Tridacna maxima, sampled throughout the West and Central Pacific, confirmed that the patterns of variation seen so far in T. gigas were not unique to that species, and may reflect a fundamental genetic structuring of shallow-water marine taxa. Populations of T. maxima within highly connected reef systems like the Great Barrier Reef were panmictic (average F_ST < 0.003), but highly significant genetic differences between reef groups on different archipelagos (average F_ST = 0.084) and between West and Central Pacific regions (average F_ST = 0.156) were found. Inferred gene flow was high (N_em usually > 5) between the Philippines and the Great Barrier Reef, between the Philippines and Melanesia (the Solomon Islands and Fiji), and between the Philippines and the Central Pacific island groups (Marshall Islands, Kiribati, Tuvalu and Cook Islands). Gene flow was low between these three sets of island chains (N_em < 2). These routes of gene flow are perpendicular to present-day ocean currents. It is suggested that the spatial patterns of gene frequencies reflect past episodes of dispersal at times of lower sea levels which have not been erased by subsequent dispersal by present-day circulation. The patterns are consistent with extensive dispersal of marine species in the Pacific, and with traditional views of dispersal from the Indo-Malay region. However, they demonstrate that dispersal along present-day ocean surface currents cannot be assumed, that other mechanisms may operate today or that major dispersal events are intermittent (perhaps separated by several thousands of years), and that the nature and timing of dispersal of Pacific marine species is more complex than has been thought.     

Genetic structure of the red mangrove (<Emphasis Type="Italic">Rhizophora mangle</Emphasis> L.) on the Colombian Pacific detected by microsatellite molecular markers

Rhizophora mangle, one of the five species of the genus Rhizophora, is found widely distributed along the American and West African coasts. This species is one of the principal constituents of the mangrove ecosystem in Colombia and is also found within the most important economic activities for the communities that inhabit the littoral. In order to assess the degree of genetic diversity of R. mangle in five populations of the Colombian Pacific, nuclear microsatellite molecular markers were used. In 92 individuals sampled, it was found that 100% of the loci were polymorphic , and no private alleles were detected. The population structure of R. mangle in the Colombian Pacific, was highly significant (P < 0.001); however, the greatest differentiation was detected at the within-population level (94.62%). For the populations of La Plata, Virudó and Charambirá, the tendency toward panmixia could be the cause of the low differentiation among these three locations. Within populations, the genetic diversity revealed a deviation from Hardy–Weinberg equilibrium with high significance in Virudó and Tumaco, where it appears the intense anthropogenic activity has exercised strong pressure on the red mangrove, resulting in the possible fragmentation of the local landscape and therefore an increase in the rate of endogamy within these populations. Despite this situation, our study―one of the first developed in genetics of the red mangrove in Colombia―did not show evidence of recent bottleneck effects or deterioration in its genetic composition, which could be exploited to propose management and restoration programs for the zones where the forests of this species are degraded. Handling editor: K. Martens     

Seasonal occurrence and diel locomotor activity in littoral Cladocera in a mesohumic lake in Norway

The community structure and standing crop biomass of a mangrove forest in Futian Nature Reserve, Shenzhen, the People's Republic of China was studied. This mangrove, located towards the northern latitudinal limit of the mangrove development (22°N), had relatively simple structure and low diversity (the Shannon-Wiener index was 0.78). The three dominant species, namely Aegiceras corniculatum, Kandelia candel and Avicennia marina, possess importance values of 72, 19 and 9%, respectively. The average height of the mangrove community was 4.5 m with no vertical stratification. The values of tree density of A. corniculatum and A. marina were found to be 5290 and 260 ha^–1, respectively. The biomass of both A. corniculatum and K. candel was best estimated from regression equations using a combination of height and diameter at breast height as the independent variables. For Avicennia marina, there was no simple correlation between biomass and height or diameter. The regression models suggested by previous workers did not give satisfactory estimation of biomass of A. marina in this mangrove forest. The total biomass of this mangrove forest was 12.1 kg m^–2, with 73% of such production contributed by A. corniculatum and 8% by A. marina. Because of its small percentage, the inaccuracy in estimating biomass of A. marina did not affect the overall determination of biomass of the whole community. Average above-ground biomass was 8.7 kg m^–2 (72% of the total biomass) and the major components were aerial woody tissues, stems and branches. The root:shoot ratio of this plant community was 0.4:1.     

PHYLOGEOGRAPHY OF THE SPOTTED SAND BASS, PARALABRAX MACULATOFASCIATUS: DIVERGENCE OF GULF OF CALIFORNIA AND PACIFIC COAST POPULATIONS

Abstract Have the warm tropical waters and currents of the southern Gulf of California, Mexico (also known as the Sea of Cortez), formed a barrier to gene flow, resulting in disjunct populations in the upper gulf that are isolated from the outer Pacific Coast? Phylogeographic and genetic divergences of the spotted sand bass, Paralabrax maculatofasciatus, from three Gulf of California and two outer Pacific coastal locations were tested using mitochondrial DNA (mtDNA) control region sequences. Sequence data from two congeners that are sympatrically distributed along the outer Pacific Coast, the barred sand bass, P. nebulifer, and the kelp bass, P. clathratus, were used to gauge the levels of genetic divergences. Differences among the three species and between the northern gulf and outer Pacific coastal populations of P. maculatofasciatus also were analyzed using 40 allozymic presumptive gene loci. Allozyme and mtDNA analyses each revealed many fixed differences among the species. Three significant allozymic frequency differences and two fixed mtDNA substitutions differentiated the gulf and outer Pacific coastal populations of P. maculatofasciatus. Three unique mtDNA haplotypes and three unique allozyme alleles were identified from the outer Pacific coastal population. The gulf sites contained four unique mtDNA haplotypes and six unique allozyme alleles. Partitioning of the mtDNA variation revealed that 72% of the variance occurred between the gulf and outer Pacific Coast, 20% between sampling sites in the two regions, and 8% within the sites. There appears to be little gene flow across the waters of the southern Baja Penninsula, producing divergence estimated as 120,000 to 600,000 years between the outer Pacific coastal and the Gulf of California populations. This separation level may date to a hypothesized seaway closure near La Paz, Mexico, during the mid‐Pleistocene, and characterizes other fish populations. A second pattern of deeper allopatric species‐level divergences in some other fishes may date to a Pliocene closure of a mid‐Baja Penninsular seaway. Significant differences also were discerned in P. maculatofasciatus between the San Diego and central Baja California coastal sites and between the upper/central and the lower gulf locations. Variation between locations in the two regions may be indicative of larval retention and low adult migration, which needs to be tested further.     

Life cycle of Chattonella marina (Raphidophyceae) inferred from analysis of microsatellite marker genotypes

Red tides of Chattonella spp. have caused continuous damage to Japanese aquaculture, however, the life cycle of this organism remains incompletely understood. To further investigate this matter, we assessed genotypes at 14 microsatellite markers in three varieties of Chattonella marina, viz., C. marina var. antiqua, C. marina var. marina, and C. marina var. ovata, to establish whether Chattonella undergoes asexual diploidization or sexual reproduction. After genotyping 287 strains of C. marina, all but one of these strains was shown to be heterozygous for at least some loci, and thus, in the diploid state, suggesting that Chattonella strains undergo sexual reproduction. In addition, we performed single‐cell amplification on ‘small cells’ that are derived from vegetative cells under dark and low‐nutrient conditions. The results indicated the existence of two types of small cells. The ‘Small cell Type 1’ was found to be heterozygous, genotypically equivalent to the vegetative cells, and is therefore diploid. These small cells may change to resting cells (cysts) directly. The ‘Small cell Type 2’ was homozygous at all analyzed loci, suggesting that these small cells are haploid and may be derived by meiosis. As fusion between small cells has previously been observed, the ‘Small cell Type 2’ may be the gamete of Chattonella. We present a construct of the full life cycle of Chattonella marina based on our own and previous results.     

BIOSYSTEMATIC STUDIES IN THE BOUTELOUA CURTIPENDULA COMPLEX. III. POLLEN SIZE AS RELATED TO CHROMOSOME NUMBERS

Pollen size statistics are presented for 10 closely related species of Bouteloua and relationships between pollen size and chromosome numbers are presented for 13 populations of 5 species and 3 varieties. With 1 exception, all populations of all taxa conformed to a general pattern of pollen size dependent upon chromosome number. Chromosome numbers varied from 2n = 20 to 2n = ca. 103, with several independent aneuploid series. Statistical analyses were made of pollen size as related to chromosome number in the 3 varieties of B. curtipendula. These data showed that tetraploids (2n = 40) of var. tenuis had significantly greater pollen size and coefficient of variation than diploids (2n = 20) of the same variety. Similarly, aneuploids of var. curtipendula with 2n = 45 to 2n = 64 chromosomes had significantly larger and more variable pollen than tetraploids (2n = 40) of the same variety. Highly significant positive regression coefficients were obtained from analyses of chromosome numbers and mean pollen size, and chromosome numbers and coefficient of variation, for var. curtipendula. Regression coefficients for var. caespitosa populations with chromosome numbers over the hexaploid (2n = 60) level were not significant.