Chloroplast DNA Intraspecific Phylogeography of Prairie Cordgrass (Spartina pectinata Bosc ex Link)

Chloroplast DNA (cpDNA) sequences are appropriate for studying intraspecific phylogeography. Comparing cpDNA phylogeny with different cytotypes provides insights into the origin and establishment of polyploid species. We have previously defined the geographic distribution of cytotypes in prairie cordgrass throughout the different regions of the United States. In this study, chloroplast haplotype variation is explored in 54 populations of prairie cordgrass, using nine noncoding chloroplast regions. These chloroplast analyses were combined with the cytotype surveys of prairie cordgrass to infer the phylogeography and to elucidate the origin of the different cytotypes. In this study, we identified three haplotypes, Prairie Cordgrass Group 1 (PCG1), Prairie Cordgrass Group 2 (PCG2), and Prairie Cordgrass Group 3 (PCG3). In general, related haplotypes were strongly associated with geographic distribution. Useful insertions–deletions (indels) were also found among prairie cordgrass populations. The PCG1 haplotypes collected inthe East North Central to the New England regions of the US, were polyploid (tetraploid, hexaploid, and octoploids), while the PCG2 haplotypes, found in southern South Dakota, Iowa, and Missouri, were primarily octoploids, but also included a small number of tetraploids. The PCG3 haplotypes were octoploids and were collected in North Dakota, South Dakota, and Minnesota.     

Chloroplast DNA variation in the genusSecale (Poaceae)

The chloroplast genomes of 44 accessions ofSecale were surveyed for restriction site polymorphisms. The accessions were chosen to represent the geographic as well as taxonomic range of the genus. Using 12 restriction enzymes a total of 348 sites were detected. Twenty-nine mutation sites were phylogenetically informative and used in a cladistic analysis. Further, a 0.1 kb insertion separatedSecale from the outgroup species. Only the annual speciesS. sylvestre was distinct from the rest of the taxa. Cultivated rye together with both wild annual and wild perennial accessions were mixed among each other. Sequence divergence (p) among taxa ofSecale was low, varying from 0.000 to 0.005, suggesting a rather recent origin of the genus.     

Characterization of microsatellite loci in Spartina species (Poaceae)

The cordgrasses in the genus Spartina have become model organisms for studying biological invasions from both ecological and genetic perspectives. Here we characterize 11 disomic loci in Spartina alterniflora that show promise for population studies and for studying hybridization events between S. alterniflora and S. foliosa. Comparisons among invasive and native S. alterniflora populations showed that levels of allelic variation are lower in invasive populations. In addition, nearly all loci that amplified in S. foliosa populations and in a swarm of S. alterniflora×foliosa hybrids were polymorphic. We also found that several loci amplified successfully in other Spartina species.     

Hybridization between introduced smooth cordgrass (Spartina alterniflora; Poaceae) and native California cordgrass (S. foliosa) in San Francisco Bay, California, USA

Introduced Spartina alterniflora (smooth cordgrass) is rapidly invading intertidal mudflats in San Francisco Bay, California. At several sites, S. alterniflora co-occurs with native S. foliosa (California cordgrass), a species endemic to California salt marshes. In this study, random amplified polymorphic DNA markers (RAPDs) specific to each Spartina species were identified and used to test for hybridization between the native and introduced Spartina species in the greenhouse and in the field. Greenhouse crosses were made using S. alterniflora as the pollen donor and S. foliosa as the maternal plant, and these crosses produced viable seeds. The hybrid status of the crossed offspring was confirmed with the RAPD markers. Hybrids had low self-fertility but high fertility when back-crossed with S. foliosa pollen. Hybrids were also found established at two field sites in San Francisco Bay; these hybrids appeared vigorous and morphologically intermediate between the parental species. Field observations suggested that hybrids were recruiting more rapidly than the native S. foliosa. Previous work identified competition from introduced S. alterniflora as a threat to native S. foliosa. In this study, we identify introgression and the spread of hybrids as an additional, perhaps even more serious threat to conservation of S. foliosa in San Francisco Bay.     

Extent and degree of hybridization between exotic (Spartina alterniflora) and native (S. foliosa) cordgrass (Poaceae) in California, USA determined by random amplified polymorphic DNA (RAPDs)

Spartina alterniflora, smooth cordgrass, native to the eastern USA, was introduced into south San Francisco Bay ≈ 25 years ago. It has spread by purposeful introduction of rooted plants and dispersal of seeds on the tides. Previous work suggested that S. alterniflora was competitively superior to the native California cordgrass, S. foliosa, and that the two species hybridized. The present study determined the spread of S. alterniflora and S. foliosa × alterniflora hybrids in California and examined the degree of hybridization. We used nuclear DNA markers diagnostic for each species to detect the parental species and nine categories of hybrids. The California coast outside San Francisco Bay contained only the native species. All hybrid categories exist in the Bay, implying that several generations of crossing have occurred and that hybridization is bidirectional. Hybrids were found principally near sites of deliberate introduction of the exotic species. Where S. alterniflora was deliberately planted, we found approximately equal numbers of S. alterniflora and hybrid individuals; S. foliosa was virtually absent. Marshes colonized by water-dispersed seed contained the full gamut of phenotypes with intermediate-type hybrids predominating. The proliferation of hybrids could result in local extinction of S. foliosa. What is more, S. alterniflora has the ability to greatly modify the estuary ecosystem to the detriment of other native species and human uses of the Bay. To the extent that they share these engineering abilities, the proliferation of cordgrass hybrids could grossly alter the character of the San Francisco Bay.     

Chloroplast DNA diversity within and among populations of the allotetraploid Prunus spinosa L.

High chloroplast DNA (cpDNA) diversity was found within and among populations of Prunus spinosa sampled from seven European deciduous forests. A study of 12% of the total chloroplast genome detected 44 mutations, which were distributed over 24 haplotypes; four were common to two or more populations and the rest were unique haplotypes. The most-abundant and widely distributed haplotype was H2 (frequency = 41% approximately). Six of the seven populations were polymorphic. All of the six polymorphic populations had ”private” haplotypes (frequency < 5%) in addition to common haplotypes. The UPGMA dendrogram demonstrated a correlation between populations and their geographical locations. The total diversity was high (h_T= 0.824) and a major portion of it was within populations (h_s= 0.663). The level of population subdivision for unordered alleles was low (G_ST= 19.5%) and for ordered alleles was lower (N_ST= 13.6%). No phylogeographic structure could be demonstrated in the present geographical scale. High polymorphism in the cpDNA of P. spinosa has to be considered carefully when planning phylogenetic studies involving this species. Received: 20 September 1999 / Accepted: 10 November 1999     

Random amplified polymorphic DNA variation among remnant big bluestem (Andropogon gerardii vitman) populations from Arkansas' grand prairie

Random amplified polymorphic DNA (RAPD) analysis was used to characterize genetic diversity and genetic distinctiveness of Andropogon gerardii from remnant Arkansas prairies. Six oligonucleotide primers, which generated 37 RAPD bands, were used to analyse 30-32 plants from six Grand Prairie populations, Baker Prairie (Arkansas Ozarks), two Illinois prairies and two cultivars. Genetic diversity of the Arkansas remnants ranged from 82.7 to 99.3%, with 89% of the total genetic variation within and 11% among populations. The partitioning of genetic variation was consistent with that reported for other outcrossing perennial grasses, using the more conservative allozyme markers. Principal component analysis indicated a northern and southern association within Arkansas' Grand Prairie. Although there was no genetic structuring at the landscape level, the Illinois prairies and cultivars were different from all Arkansas prairies tested. There was significant within-population structuring in four of the seven Arkansas remnants, with a negative relationship between genetic similarity and geographical distance. The three nonstructured populations were from a linear railroad remnant, suggesting different population-level dynamics from nonlinear prairies. The results of this study indicated that small isolated remnant big bluestem populations were not genetically depauperate and that genetic relationships among populations could not be predicted solely on geographical proximity.     

One hundred years of Spartina altemiflora (Poaceae) in Willapa Bay, Washington: random amplified polymorphic DNA analysis of an invasive population

Spartina altemiflora (smooth cordgrass) has invaded Willapa Bay, Washington, covering bare mud flats and seagrass beds. At current expansion rates it threatens to occupy most of the intertidal habitat in the bay within 40 years. Although a major mitigation effort is underway, nothing is known about the genetic structure of the Willapa Bay S. altemiflora population or the underlying causes for the dramatic increase in seed set that has occurred in the last 15 years. Random amplified polymorphic DNA (RAPD) analysis was assessed as a method of DNA fingerprinting to analyse the genetic history and current structure of the Willapa population. The results suggest that all S. alterniflora clones in Willapa Bay are descended from a single genet. Given the limited genetic variability expected in a clonally founded population, we discuss the utility of RAPDs as genetic markers to explore the large morphological and reproductive differences reported among Willapa clones. Cluster analysis of RAPD fragments was used to show relatedness among S. alteniflora genets and may be helpful in locating the most prolific clones for mitigation efforts. Overall, RAPDs show great potential in genetic analysis of S. altemiflora, particularly in invasive populations where isozyme variability is low or nonexistent.     

Chloroplast DNA diversity in Calluna vulgaris (heather) populations in Europe

Variation in the chloroplast genome of Calluna vulgaris (heather), the dominant species of northwest European heath communities, was analysed using polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLPs) and microsatellites. No length polymorphisms were detected in the 100-200 base pair (bp) fragments amplified by the conserved microsatellite primers, and sequencing revealed that the repeat regions were interrupted relative to the corresponding sequence in Nicotiana tabacum. In contrast, PCR-RFLP analysis revealed high levels of haplotype diversity within populations (hS = 0.443, hT = 0.842), as well as substantial differentiation between populations (GST = 0.473). Diversity and differentiation were higher in southern Europe than in northern Europe. Interpreted in the light of data from allozyme studies and pollen core records, the results suggest that the main glacial refugia for C. vulgaris were located in southwest Europe, including northern Spain, the Pyrenees and the Massif Central region of France. There is also evidence for diffuse survival of the species at more northerly latitudes.     

Chloroplast DNA haplotype variation and population differentiation in Sorbus aucuparia L. (Rosaceae: Maloideae)

Intra-specific chloroplast DNA (cpDNA) variation was studied in Sorbus aucuparia L., an entomophilous, mid-or early successional tree producing fleshy fruits. Eight PCR-amplified fragments of the chloroplast genome were screened for restriction fragment length polymorphisms, using one or two 4 bp-cutter restriction endonucleases. cpDNA variation was investigated on two geographical scales: (1) among four regions in France and Belgium; and (2) within the Belgian region. A total of 150 individuals from six populations were analysed. Fourteen polymorphisms were detected in six of the cpDNA fragments. All polymorphisms probably resulted from insertions or deletions, and allowed the identification of 12 haplotypes. The level of genetic differentiation computed on the basis of haplotype frequencies was similar on the two geographical scales considered (G(STc) = 0.286 among regions, G(STc) = 0.259 among populations within the Belgian region). These values are much lower than those obtained in nine previously studied temperate tree species, which are all wind-pollinated, late-successional species producing dry fruits. These results might primarily be accounted for by the contrasting life history traits of S. aucuparia. In order to obtain insights into the relative contribution of pollen and seeds to gene flow, G(STc) was also compared with previously obtained G(ST) estimates based on allozyme data.     

Genetic evidence for hybridization between the native Spartina maritima and the introduced Spartina alterniflora (Poaceae) in South-West France: Spartina × neyrautii re-examined

 Spartina alterniflora, a perennial grass native to the North American Atlantic coast, was introduced during the 19th century in western Europe (Southern England and western France) where it hybridized with the native Spartina maritima. In England, the sterile hybrid S. × townsendii gave rise by chromosome doubling to the highly fertile allopolyploid Spartina anglica, which has now invaded many salt marshes and estuaries in western Europe, and has been introduced in several continents. In South-West France, another sterile hybrid was discovered in 1892 in the Bidassoa Estuary, and named Spartina × neyrautii. According to their morphology, some authors suggested that S. × neyrautii and S. × townsendii result from reciprocal crosses. During the 20th century, the hybridization site was severely disturbed, and surviving of S. × neyrautii was questioned. In this paper, various Spartina populations are investigated in the Basque region (France and Spain), and compared to the hybrid taxa formed in England (S. × townsendii and S. anglica). The samples were analyzed using molecular fingerprinting (RAPD and ISSR) and Chloroplast DNA sequence (trnL-trnT spacer, trnL intron and trnL-trnF spacer). In the Bidassoa estuary, a hybrid isolated clone has been found, that displays additive species-specific nuclear markers of S. maritima and S. alterniflora, and that is subsequently considered as a surviving clone of S. × neyrautii. The molecular analyses indicate that S. × neyrautii and S. × townsendii share the same maternal (S. alterniflora), and paternal (S. maritima) parental species, but also that the two independent hybridization events have involved different parental (nuclear) genotypes in England and in South-West France. Received July 12, 2002; accepted October 4, 2002 Published online: March 20, 2003     

Chloroplast DNA variation of Panax (Araliaceae) in Nepal and its taxonomic implications

The restriction site and size variation of five PCR amplified fragments of noncoding chloroplast DNA (cpDNA) was examined in material from 13 populations ofPanax from Nepal and China. Fourteen restriction endonucleases produced 81 restriction site and length variations from the large single-copy region of cpDNA, 27 of which are polymorphic. The cpDNA dataset suggests two distinct groups ofPanax from Nepal (clades I and II). Clade I consists of two populations ofP. pseudoginseng subsp.pseudoginseng, and clade II is composed of material referable toP. pseudogingeng subsp.himalaicus (vars.himalaicus, angustifolius, andbipinnatifidus). The three accessions ofP. pseudoginseng subsp.japonicus andP. ginseng studied from China had cpDNA characters that differed from the HimalayanPanax. The highly distinctive cpDNA profile and morphology ofP. pseudoginseng subsp.pseudoginseng sensu Hara (1970) from central Nepal support its status as a separate species, which has an extremely restricted distribution.     

Variation in self-fertility and the reproductive advantage of self-fertility for an invading plant (Spartina alterniflora)

The factors responsible for the reproductive success or failure of individuals in small, founding populations have received little attention. Previous work on a small population of smooth cordgrass (Spartina alterniflora) invading San Francisco Bay, California found that most clones flower prolifically but set little or no seed, while a few clones have high rates of viable seed set, producing most of the seeds in the population. This study first identifies recruitment from seeds as the main source of new smooth cordgrass plants during invasion and then tests the influence of growing conditions and pollination treatment on viable seed set among clones established in San Francisco Bay. Field transplants indicated that a clone's seed set rate was not strongly dependent on its site of establishment. Low and high nutrient greenhouse treatments also had little effect on viable seed set rates within most clones. In contrast, pollination treatment (self-pollination or outcrossing) had a major effect on viable seed set rates. Most clones had high seed set rates after outcross-pollination, but clones varied widely in their selfing capacity. Zero or low viable self-seed set rates were most common; however, a few clones had high viable self-seed set rates, comparable to outcross seed set rates. A clone's selfing capacity was significantly correlated across years (r=0.89, P<0.001), and capacity to set viable self-pollinated seeds in the greenhouse was significantly correlated with the clone's rate of viable seed set in the field . In this growing population where cross-pollination is limited, only the clones with high selfing ability had high viable seed set rates in the field. Among primarily outcrossing plant invaders, variation in self-fertility among individuals may be a common phenomenon, with important implications for genetic differentiation, effective population size and patterns of spatial spread during an invasion.     

Extensive intraspecific chloroplast DNA (cpDNA) variation in the alpine Draba aizoides L. (Brassicaceae): haplotype relationships and population structure

Chloroplast DNA (cpDNA) sequence variation is currently the most widely used tool for the inference of phylogenetic relationships among plants at all taxonomic levels. Generally, noncoding regions tend to evolve faster than coding sequences and have recently been applied to the study of phylogenetic relationships among closely related taxa. An implicit assumption of many of these studies is that intraspecific cpDNA variation is either absent or low and therefore will not interfere with the reconstruction of interspecific relationships. A survey of cpDNA sequence variation in the common alpine plant species Draba aizoides L. was undertaken to assess levels of intraspecific cpDNA sequence variation. These levels were compared to levels of interspecific sequence divergence between D. aizoides and related alpine Draba species. Intraspecific cpDNA sequence divergence was extensive in D. aizoides, and intraspecific differences were often larger than interspecific differences. cpDNA haplotype relationships were explored using a maximum parsimony approach and minimum-spanning networks. Results from both methods were largely congruent but comparisons provided interesting insights into the presumed evolutionary history of cpDNA haplotypes. A combined effect of cpDNA introgression and complex lineage sorting was inferred to explain the pattern of cpDNA variation found in D. aizoides. Our results suggest that intraspecific cpDNA variation can be extensive and that intraspecific variation needs to be taken into account when inferring phylogenetic relationships among closely related taxa.     

Ribosomal DNA variation within and among individuals ofLisianthius (Gentianaceae) populations

Restriction endonuclease fragment analysis of nuclear ribosomal DNA (rDNA) was completed on 25 individuals each from seven populations of theLisianthius skinneri (Gentianaceae) species complex in Panama. Seven restriction enzymes were used to determine the amount and type of rDNA variation within and among individuals of the populations. No restriction site variation was seen within populations or individuals although site differences were seen among populations. Spacer length variation within and among individuals of populations was mapped to the internal transcribed spacer (ITS) region between the 18S and 5.8S rRNA genes, a region inLisianthius rDNA that previously was shown to exhibit length differences among populations. This is the first reported case of such variation within and among individuals of populations for the ITS region. Presence or absence of ITS spacer length variation is not correlated with levels of isozymic heterozygosity within populations. No detectable length variation within individuals or populations was seen in the larger intergenic spacer (IGS). Although populations varied with respect to IGS length, all individuals of a given population had a single and equivalent IGS length.     

Chloroplast DNA variation and evolution in the genus Lens Mill

 Chloroplast DNA (cpDNA) restriction site diversity was assessed by 21 enzyme/probe combinations in 30 accessions of six Lens species, including the recently recognized L. lamottei and L. tomentosus. A total of 118 fragments were scored and 26 restriction site mutations were identified. The cpDNA restriction pattern supports circumscribing L. lamottei and L. tomentosus as independent species. The value of the data for reconstructing phylogeny in the genus is discussed. The cpDNA of all 13 accessions of the lentil’s wild progenitor, L. culinaris subsp. orientalis, differed from that of the single lentil cultivars used in this study. This diversity indicates that other populations of this subspecies from Turkey and Syria examined by Mayer and Soltis (1994) are potentially the founder members of lentil. Examination of L. lamottei×L. nigricans hybrids between accessions having different restriction patterns showed paternal plastid inheritance in L. nigricans. Received: 2 July 1996 / Accepted: 19 July 1996     

The evolution of Spartina anglica G. E. Hubbard (Gramineae): genetic variation and status of the parental species in Britain

The perennial salt marsh grass Spartina anglica is one of the classic examples of allopolyploid speciation. It originated on the south coast of England at the end of the nineteenth century following chromosome doubling in S. × townsmdii , a hybrid between the native British S. marilima and a species introduced from the United States, S. alterniflora. The nature of the origin of S. anglica is beyond doubt; however, it is not known whether it had a single or multiple origin. In order to address this problem we undertook a survey of the genetic variation in the parental species of S. anglica using isozyme electrophoresis. The results show that S. alterniflora has no detectable variation and that S. maritima has extremely low levels of variation. These results, unfortunately, prevent the question of a single or multiple origin from being answered. Possible reasons for the low levels of variation and its influence on the future of the species are discussed. Another problem concerning the parental species is the rapid decline of S. maritima in Britain. It is often assumed that the major factor in this regression is the invasion of its habitats by S. anglica. We have examined the status of S. marilima throughout its range in Britain and have found that S. anglica rarely co-occurs with S. maritima. We propose that the decline of S. maritima is largely due to the physical erosion of its habitats and that this erosion may produce suitable habitats for colonization by S. anglica.     

Inbreeding depression in smooth cordgrass (Spartina alterniflora, Poaceae) invading San Francisco Bay

The magnitude of inbreeding depression in invading plant populations is often presumed to be small and of little consequence. The purpose of this study was to assess the magnitude of inbreeding depression in a pollen-limited, partially self-incompatible, invading plant population. The magnitude and timing of inbreeding depression were compared among ten maternal plants sampled from a population of smooth cordgrass (Spartina alterniflora) invading San Francisco Bay. Selfed and outcrossed progeny were compared for embryo abortion, survival of seedlings, and growth/survival at the end of the first growing season in three greenhouse environments. Estimates of inbreeding depression varied among environments, with competitive environment > high-nutrient environment > low-nutrient environment. Population-level estimates of inbreeding depression ranged from 0.61 to 0.81; however, maternal plants varied significantly in their magnitude of inbreeding depression, ranging from 0.1 to 0.97. The 95% confidence interval for inbreeding depression for some maternal plants included zero. There was a significant negative correlation between the overall magnitude of inbreeding depression and self-fertility rate among maternal plants. The few maternal plants with high self-fertility carried relatively little genetic load, and their selfed progeny are likely to survive on open mudflats. The noncompetitive, pollen-limited growing conditions associated with invasion may allow self-fertility to spread in this population.     

Chloroplast DNA restriction fragment length polymorphisms (RFLPs) in Musa species

Summary Taxonomic and phylogenetic determinations within the genus Musa are established using a numerical, morphology-based scoring system. However, within this system, the classification and relationships of some types are disputed. The application of chloroplast DNA (cpDNA) restriction fragment length polymorphism (RFLP) analysis to Musa taxonomy provided valuable, supplemental information about the classification of, and relationships between, Musa species and subspecies. Whole-cell DNA was extracted from lyophilized Musa leaf-blade tissue and digested with various restriction enzymes, Southern blotted onto nylon membranes, and probed using radioactively labeled heterologous orchid cpDNA fragments. Phylogenies were inferred from cpDNA RFLP patterns using PAUP software. The relationships between most species examined were as expected; however, some species (M. beccarii and M. basjoo) did not conform to the conventional morphology-based phylogeny.