Evolution of Lycopodiaceae from results of sequencing spacer sequences of chloroplast rRNA genes

Nucleotide sequences of a chloroplast rDNA region including 8 bp from the 3' end of 23S rDNA-ITS2-4.5S rDNA-ITS3-5S rDNA-ITS4 (approximately 800 bp) were determined in 25 species of Lycopodiaceae and two species of the genus Isoetes. The rate of molecular evolution of spacers significantly varied in different Lycopsida taxa. A phylogenetic analysis by the neighbor-joining (NJ) method revealed that the family Lycopodiaceae is monophyletic. The topology of phylogenetic trees suggests the isolation of four or probably five genera in family Lycopodiaceae. For these genera, synapomorphic indels were detected. The obtained data were compared with the results of phylogenetic analysis of Lycopsida with regard to other sequences. The relationships of taxa within the family Lycopodiaceae is discussed.     

Reconsideration of systematic relationships within the order Euplotida (Protista, Ciliophora) using new sequences of the gene coding for small-subunit rRNA and testing the use of combined data sets to construct phylogenies of the Diophrys-complex

Comprehensive molecular analyses of phylogenetic relationships within euplotid ciliates are relatively rare, and the relationships among some families remain questionable. We performed phylogenetic analyses of the order Euplotida based on new sequences of the gene coding for small-subunit RNA (SSrRNA) from a variety of taxa across the entire order as well as sequences from some of these taxa of other genes (ITS1-5.8S-ITS2 region and histone H4) that have not been included in previous analyses. Phylogenetic trees based on SSrRNA gene sequences constructed with four different methods had a consistent branching pattern that included the following features: (1) the “typical” euplotids comprised a paraphyletic assemblage composed of two divergent clades (family Uronychiidae and families Euplotidae–Certesiidae–Aspidiscidae–Gastrocirrhidae), (2) in the family Uronychiidae, the genera Uronychia and Paradiophrys formed a clearly outlined, well-supported clade that seemed to be rather divergent from Diophrys and Diophryopsis, suggesting that the Diophrys-complex may have had a longer and more separate evolutionary history than previously supposed, (3) inclusion of 12 new SSrRNA sequences in analyses of Euplotidae revealed two new clades of species within the family and cast additional doubt on the present classification of genera within the family, and (4) the intraspecific divergence among five species of Aspidisca was far greater than those of closely related genera. The ITS1-5.8S-ITS2 coding regions and partial histone H4 genes of six morphospecies in the Diophrys-complex were sequenced along with their SSrRNA genes and used to compare phylogenies constructed from single data sets to those constructed from combined sets. Results indicated that combined analyses could be used to construct more reliable, less ambiguous phylogenies of complex groups like the order Euplotida, because they provide a greater amount and diversity of information.     

Sequence Variation of Ribosomal Internal Transcribed Spacers (ITS) in Commercially Important Phytoseiidae Mites

Preliminary work is needed to assess the usefulness of different markers at different taxonomic scales when a new group is analyzed, such as the commercially important Phytoseiidae mites. We investigate here the level of sequence variation of the nuclear ribosomal spacers ITS 1 and 2 and the 5.8S gene in six species of Phytoseiidae: Neoseiulus californicus, N. fallacis, Euseius concordis, Metaseiulus occidentalis, Typhlodromus pyri and Phytoseiulus persimilis. As expected, the 5.8S gene (148 base pairs) is markedly conserved and displays little variation in between genera comparisons. ITS1 and ITS2 show contrasting patterns: while the ITS2 is short (80–89 bp) and shows little variation, the ITS1 is longer (303–404 bp) and is very variable in sequence. This fact compromises reliable nucleotide homologies when comparing the genera. The comparison of ITS1 sequence similarity at the species level might be useful for species identification, however, the value of ITS in taxonomic studies does not extend to the level of the family. The intraspecific variations of ITS were investigated in three species: N. californicus, N. fallacis and E. concordis. The first species has identical ITS1 sequences and the last two display low polymorphism (2 nucleotide substitutions). The ITS2 and 5.8S sequences were identical in all three subspecies comparisons.     

Preliminary Analysis of Length and GC Content Variation in the Ribosomal First Internal Transcribed Spacer (ITS1) of Marine Animals

Length and guanine–cytosine (GC) content of the ribosomal first internal transcribed spacer (ITS1) were compared across a wide variety of marine animal species, and its phylogenetic utility was investigated. From a total of 773 individuals representing 599 species, we only failed to amplify the ITS1 sequence from 87 individuals by polymerase chain reaction with universal ITS1 primers. No species was found to have an ITS1 region shorter than 100 bp. In general, the ITS1 sequences of vertebrates were longer (318 to 2,318 bp) and richer in GC content (56.8% to 78%) than those of invertebrates (117 to 1,613 bp and 35.8% to 71.3%, respectively). Specifically, gelatinous animals (Cnidaria and Ctenophora) were observed to have short ITS1 sequences (118 to 422 bp) with lower GC content (35.8% to 61.7%) than the other animal taxa. Mollusca and Crustacea were diverse groups with respect to ITS1 length, ranging from 108 to 1,118 and 182 to 1,613 bp, respectively. No universal relationship between length and GC content was observed. Our data indicated that ITS1 has a limited utility for phylogenetic analysis as obtaining confident sequence alignment was often impossible between different genera of the same family and even between congeneric species.     

Taxonomy, phylogeny, and distribution of Puccinia graminis, the black stem rust: new insights based on rDNA sequence data

Puccinia graminis (Uredinales) is an economically important and common host-alternating rust species on Berberidaceae/Poaceae (subfamilies Pooideae and Panicoideae) that has been spread globally by human activities from an unknown center of origin. To evaluate the taxonomic implications, phylogenetic relationships, and distribution/spread of this complex species, we sequenced and cladistically analyzed the ITS1, 5.8S, and ITS2 regions from herbarium specimens on various host plants from Iran (17), Europe (1), and North America (4). The ITS region plus the 5.8S gene ranged from 686 to 701 bp, including the flanking partial sequences of the 18S and 28S rDNA. Our phylogenetic analysis included 54 bp of the 18S sequence, the entire ITS1 + 5.8S + ITS2, and 58 bp of the 28S sequence. A second analysis used only the last 42 bp of ITS1, and all the 5.8S and ITS2, to incorporate data from additional sequences downloaded from GenBank. In addition to variation in sequence length, there was variation in sequence content. The analysis does not support classical morphology-based taxonomic concepts of the P. graminis complex. Also, host range, host taxonomy, and geographic origin provide minor information on taxonomic relationships. Puccinia graminis is most probably monophyletic. Coevolutionary aspects can hardly be discussed because of lack of sequence data from alternate host specimens. The occurrence of unrelated fungal taxa on the same host species suggests that, besides coevolution with the host, host jumps and hybridization may have played an important role in the evolution of P. graminis. From rDNA data we conclude that the pathogen was introduced to North America at least twice independently. For a new taxonomic concept, we think the complex has to be split into at least two species. New morphological features and further features other than sequence data, however, must be checked for taxonomic value first and, if necessary, be considered.     

Phylogenetic Utility of the External Transcribed Spacer (ETS) of 18S–26S rDNA: Congruence of ETS and ITS Trees ofCalycadenia(Compositae)

The 3′ region of the external transcribed spacer (ETS) of 18S–26S nuclear ribosomal DNA was sequenced in 19 representatives ofCalycadenia/Osmadeniaand two outgroup species (Compositae) to assess its utility for phylogeny reconstruction compared to rDNA internal transcribed spacer (ITS) data. Universal primers based on plant, fungal, and animal sequences were designed to amplify the intergenic spacer (IGS) and an angiosperm primer was constructed to sequence the 3′ end of the ETS in members of tribe Heliantheae. Based on these sequences, an internal ETS primer useful across Heliantheaesensu latowas designed to amplify and sequence directly the 3′ ETS region in the study taxa, which were the subjects of an earlier phylogenetic investigation based on ITS sequences. Size variation in the amplified ETS region varied across taxa of Heliantheaesensu latofrom approximately 350 to 700 bp, in part attributable to an approximately 200-bp tandem duplication in a common ancestor ofCalycadenia/Osmadenia.Phylogenetic analysis of the 200-bp subrepeats and examination of apomorphic changes in the duplicated region demonstrate that the subrepeats inCalycadenia/Osmadeniahave evolved divergently. Phylogenetic analyses of the entire amplified ETS region yielded a highly resolved strict consensus tree that is nearly identical in topology to the ITS tree, with strong bootstrap and decay support on most branches. Parsimony analyses of combined ETS and ITS data yielded a strict consensus tree that is better resolved and generally better supported than trees based on either data set analyzed separately. We calculated an approximately 1.3- to 2.4-fold higher rate of sequence evolution by nucleotide substitution in the ETS region studied than in ITS-1 + ITS-2. A similar disparity in the proportion of variable (1.3 ETS:1 ITS) and potentially informative (1.5 ETS:1 ITS) sites was observed for the ingroup. Levels of homoplasy are similar in the ETS and ITS data. We conclude that the ETS holds great promise for augmenting ITS data for phylogenetic studies of young lineages.     

Lengths and nucleotide sequences of the internal spacers of nuclear ribosomal DNA in gymnosperms and pteridophytes

The nucleotide sequences of the internal transcribed spacers (ITS1 and ITS2) of the nuclear ribosomal DNA were analyzed in species belonging to gymnosperms and pteridophytes. The lengths of the ITSs of sixteen species of gymnosperms and seven species of pteridophytes were estimated. The gymnosperms have ITS1 regions larger than those observed in the pteridophytes and angiosperms (ca. 610–3100 bp versus 159–360 bp). On the other hand, the ITS2 regions appear to be of a conserved length (182–370 bp). We have determined the complete nucleotide sequences of ITS regions from four gymnosperm species and five pteridophyte species by cloning the PCR products. Sequence analysis showed the presence of three short tandem arranged subrepeats of about 70 bp in the 1112 bp ITS1 ofEphedra fragilis. Pyrimidine rich (about 90%) DNA segments of 40–50 bp were observed in the ITS1 ofGinkgo biloba. A highly conserved 16 bp long sequence known to be present in the ITS1 of the angiosperm species has been also found in the ITS1 ofCycas revoluta, Taxus baccata andEphedra fragilis. Dedicated to Prof.Emilio Battaglia.     

Molecular phylogenetics of the subtribeGentianinae (Gentianaceae) inferred from the sequences of internal transcribed spacers (ITS) of nuclear ribosomal DNA

The internal transcribed spacer (ITS) regions of 18S–25S nuclear ribosomal DNA from representatives of 23 species of the subtribeGentianinae and one outgroup species (Centaurium capitatum) were analyzed by polymerase chain reaction amplification and direct DNA sequencing. Within the taxa analyzed, the length of the ITS1 region varied from 221 to 233 bp, ITS2 from 226 to 234 bp. Of the aligned sequences of 497 positions, 151 sites involved gaps or nucleotide ambiguity, 133 were invariable and 213 showed divergence. In pairwise comparisons among the taxa of the subtribeGentianinae and the outgroup, sequence divergence ranged from 1.3% to 34.1% in ITS1, from 0 to 28.1% in ITS2 and from 0.6% to 27.5% in combined ITS1 and ITS2. Phylogenetic trees generated from ITS sequences were highly resolutive and principally concordant with morphological classifications for the major phylogenetic divisions in the subtribe. An ancient divergence leading to two evolutionary lines was suggested in the subtribe by both DNA sequence and morphological data. One line encompasses the generaGentiana, Crawfurdia andTripterospermum, morphologically characterized by their glands on the base of ovary and their plicate corolla, while the other line involves all other members of the subcribe surveyed, characterized by their epipetalous glands and simple corolla without plicae.Megacodon, with glands on the base of ovary but without plicae on its corolla, was revealed to be more related to the latter group than to the former.Comastoma, Gentianella andGentianopsis were shown to be well-defined monophyletic genera.Pterygocalyx showed much closer affinity toGentianopsis than to any other genus. Some conflictions were detected in the genusSwertia.     

Length Variation of the Nuclear Ribosomal DNA Internal Transcribed Spacer in the Genus Abies,with Reference to Its Systematic Utility in Pinaceae

The internal transcribed spacer (1TS) region (1TS1, ITS2 and 5.8S rDNA) of the nuclear ribosomal DNA (nrDNA) was amplified via PCR in 28 taxa of Abies Mill. The amplified fragments showed length polymorphism among species, with species from Central America and two species from North America having a length of approximately 2 500 base pairs (bp) and the remaining taxa having a length of approximately 1 700 bp based on 100 bp and 1 kb ladder standard markers. The complete sequencing of ITS of Abies bracteata showed that the shorter type is 1 697 bp (1TS1 is 1 296 bp, 5.8S + 1TS2 is 401 bp). For the longer one, the partial rrs1 and complete 5.8S + ITS2 sequencing revealed that thelength of 5.8S + ITS2 is the same as that of the shorter type. The length difference of ITS in Abies is mainly due to the length difference in the ITS1 region, a result similar to the previous findings in other genera of Pinaceae. Variation in ITS length seems well correlated with morphological and geographic characters in Abies, suggesting that the length variation may be a phylogenetically informative character within the genus, long ITS was also found in other genera of Pinaceae in the previous studies. The long length of ITS in the family makes the sequencing of the region and subsequent alignment of sequences among species or genera more difficult than in taxa with short ITS, such as angiosperms. Although the length variation of ITS in the genus Abies is significant, the homogenous of ITS sequence between the longer one and the shorter one is obvious if the insertion in the longer ITS is ignored.     

Application of ITS sequences of nuclear rDNA in phylogenetic and evolutionary studies of angiosperms

Nuclear rRNA genes (rDNA) in angiosperms are arranged in long tandem repeat ing units, much like those of other higher eukaryotes. Owing to rapid concerted evolution, the repeat units have homogenized or nearly so in most species. The internal transcribed spacer (ITS) of nuclear rDNA is composed of ITS1 and ITS2, which are seperated by 5.8S rDNA. The two spacers, ITS1 (187～298 bp) and ITS2 (187～252 bp), can be readily amplified by PCR and sequenced using universal primers. The sequences contain many vari able sites and potential informative sites among related species, and have been proven to be a useful molecular marker in phylogenetic and evolutionary studies of many angiosperm taxa. It can be used not only in classification and phylogenetic inferences at the levels of family, subfamily, tribe, genus and section, but also in reconstruction of reticulate evolution and de tection of the speciation via hybridization and polyploidization. But this region may not be useful for resolving phylogenetic relationships among families or taxa of higher hierarchy ow- ing to the rapid variation of the ITS sequences.     

MONOPHYLY OF ANEUPLOID ASTRAGALUS (FABACEAE): EVIDENCE FROM NUCLEAR RIBOSOMAL DNA INTERNAL TRANSCRIBED SPACER SEQUENCES

Evolutionary relationships within Astragalus L. (Fabaceae) were inferred from nucleotide sequence variation in nuclear ribosomal DNA of both New World and Old World species. The internal transcribed spacer regions (ITS) of 18S–26S nuclear ribosomal DNA from representatives of 26 species of Astragalus, three species of Oxytropis DC., and two outgroup taxa were analyzed by polymerase chain reaction amplification and direct DNA sequencing. The length of the ITS 1 region within these taxa varied from 221 to 231 bp, while ITS 2 varied in length from 207 to 217 bp. Of the aligned, unambiguous positions, approximately 34% were variable in each spacer region. In pairwise comparisons among Astragalus species and outgroup taxa, sequence divergence at these sites ranged from 0 to 18.8% in ITS 1 and from 0 to 21.7% in ITS 2. Parsimony analyses of these sequences resulted in a well-resolved phylogeny that is highly concordant with previous cytogenetic and chloroplast DNA evidence for a major phylogenetic division in the genus. These data suggest that the New World aneuploid species of Astragalus form a monophyletic but morphologically cryptic group derived from euploid species of Old World (Eurasian) origin, which are consequently paraphyletic.     

Comparative studies on the diversity of the edible mushroom Pleurotus eryngii: ITS sequence analysis, RAPD fingerprinting, and physiological characteristics

Verification of Pleurotus eryngii strains was assessed using ITS sequence analysis and RAPD fingerprinting. Sequence analysis of the ITS1–5.8S rDNA–ITS2 region of 24 strains of Pleurotus sp., which consisted of 22 strains of P. eryngii and the control strains P. ostreatus and P. ferulae, demonstrated that the DNA regions share mostly 99 % sequence identity, indicating that sequence-based analysis is not applicable for the verification of closely related mushroom strains. To verify the mushroom strains using RAPD, we amplified DNA fragments from the total cellular DNA of 24 mushroom strains with 18 different random primers, yielding 538 distinct DNA fragments ranging from 200–4000 bp. Analysis of the DNA fragment pattern showed that the 22 P. eryngii strains were clearly distinguished from the control strains P. ostreatus and P. ferulae, and could be categorized into five subgroups. Subsequent physiological studies on the development of fruiting bodies demonstrated the close correlation of the RAPD-based grouping with the phenotypical characteristics of mushroom fruiting bodies.     

Phylogenetic relationships in the Lejeuneaceae (Hepaticae) inferred using ITS sequences of nuclear ribosomal DNA

Sequences of the ITS1–5.8S–ITS2 region of nuclear ribosomal DNA were generated for 12 species from 9 genera of Lejeuneaceae and a single species of Jubulaceae (outgroup). The taxon sampling of Lejeuneaceae included representatives of the two widely recognized subfamilies, Lejeuneoideae and Ptychanthoideae. The molecular dataset was analysed independently and in combination with a morphological dataset. The nrITS dataset and the combined dataset resulted in identical topologies. The genus Bryopteris, sometimes treated as a separate family Bryopteridaceae, is nested within the Lejeuneaceae subfamily Ptychanthoideae. Lejeuneaceae subfamily Lejeuneoideae proved to be paraphyletic with the tribe Lejeuneeae sister to Ptychanthoideae, albeit without significant bootstrap support. The tribes Brachiolejeuneeae and Cheilolejeuneeae of Lejeuneoideae, established recently based on morphological evidence, are well supported in bootstrap analyses both of the ITS and the combined molecular–morphological datasets. The results support classifications of Lejeuneaceae based on morphological data and demonstrate the usefulness of the ITS region for phylogenetic studies within or among closely related genera of Lejeuneaceae.     

Chloroplast-Expressed Glutamine Synthetase (ncpGS): Potential Utility for Phylogenetic Studies with an Example from Oxalis (Oxalidaceae)

Chloroplast-expressed glutamine synthetase (ncpGS), a nuclear-encoded gene containing several introns, is introduced as a tool for phylogenetic studies at lower taxonomic levels. This gene is a member of a multigene family, but it diverged long ago from the cytosolic-expressed members of the family and appears to be single copy in the majority of taxa examined to date. The conservation of both coding sequence and position of introns has allowed the design of primers for use in a broad range of dicot taxa to amplify and sequence a region of ncpGS that contains four introns. The utility of this region in phylogenetic studies of congeneric species is illustrated by an example using eight Oxalis species. The four introns in these taxa are typical in size (76 to 136 bp), base composition (high T content), and structure (e.g., sequence of splice sites and putative branch points) for plant internal introns. Levels of variation among these ncpGS sequences compare favorably with those of the internal transcribed spacer of nuclear ribosomal DNA (ITS) from the same taxa, and results of phylogenetic analysis of ncpGS data are generally congruent with previous results using ITS.     

Phylogenetic relationships within the cyst-forming nematodes (Nematoda, Heteroderidae) based on analysis of sequences from the ITS regions of ribosomal DNA

The ITS1, ITS2, and 5.8S gene sequences of nuclear ribosomal DNA from 40 taxa of the family Heteroderidae (including the genera Afenestrata, Cactodera, Heterodera, Globodera, Punctodera, Meloidodera, Cryphodera, and Thecavermiculatus) were sequenced and analyzed. The ITS regions displayed high levels of sequence divergence within Heteroderinae and compared to outgroup taxa. Unlike recent findings in root knot nematodes, ITS sequence polymorphism does not appear to complicate phylogenetic analysis of cyst nematodes. Phylogenetic analyses with maximum-parsimony, minimum-evolution, and maximum-likelihood methods were performed with a range of computer alignments, including elision and culled alignments. All multiple alignments and phylogenetic methods yielded similar basic structure for phylogenetic relationships of Heteroderidae. The cyst-forming nematodes are represented by six main clades corresponding to morphological characters and host specialization, with certain clades assuming different positions depending on alignment procedure and/or method of phylogenetic inference. Hypotheses of monophyly of Punctoderinae and Heteroderinae are, respectively, strongly and moderately supported by the ITS data across most alignments. Close relationships were revealed between the Avenae and the Sacchari groups and between the Humuli group and the species H. salixophila within Heteroderinae. The Goettingiana group occupies a basal position within this subfamily. The validity of the genera Afenestrata and Bidera was tested and is discussed based on molecular data. We conclude that ITS sequence data are appropriate for studies of relationships within the different species groups and less so for recovery of more ancient speciations within Heteroderidae.     

Comparisons of phylogenetic hypotheses among different data sets in dwarf dandelions (Krigia,Asteraceae): Additional information from internal transcribed spacer sequences of nuclear ribosomal DNA

The internal transcribed spacer (ITS) region of the 18 S–25 S nuclear ribosomal DNA repeat was sequenced from 19 populations of the tribeLactuceae, including all species of dwarf dandelion (Krigia) and five outgroup genera. The incidence of length changes and base substitutions was at least two times higher for ITS 1 than ITS 2. Interspecific sequence divergence withinKrigia averaged 9.62% (1.61%–15.19%) and 4.26% (0%–6.64%) in ITS 1 and ITS 2, respectively. Intergeneric sequence divergence ranged from 15.6% to 44.5% in ITS 1 and from 8.0% to 28.6% in ITS 2. High sequence divergence and homoplasy among genera of tribeLactuceae suggest that the phylogenetic utility of ITS sequence data is limited to interspecific studies or comparisons among closely related genera. Trees generated from ITS sequences are essentially identical to those from restriction site comparisons of the entire nuclear ribosomal (nr) DNA region. The degree of tree resolution differed depending on how gaps were treated in phylogenetic analyses. The ITS trees were congruent with the chloroplast DNA and morphological phylogenies in three major ways: 1) the sister group relationship betweenKrigia andPyrrhopappus; 2) the recognition of two monophyletic sections,Krigia andCymbia, in genusKrigia; and 3) the monophyly of theK. occidentalis-K. cespitosa clade in sect.Cymbia. However, the two nrDNA-based trees are not congruent with morphology/chloroplast DNA-based trees for the interspecific relationships in sect.Krigia. An average of 22.5% incongruence was observed among fourKrigia data sets. The relatively high degree of incongruence among data sets is due primarily to conflict between trees based on nrDNA and morphological/cpDNA data. The incongruence is probably due to the concerted evolution of nrDNA repeating units. The results fromKrigia and theLactuceae suggest that nrDNA data may have limited utility in phylogenetic studies of plants, especially in groups which exhibit high levels of sequence divergence. Our combined phylogenetic analysis as a total evidence shows the least conflict to each of the individual data sets.     

Accelerated nrDNA evolution and profound AT bias in the medicinal fungus Cordyceps sinensis

Cordyceps sinensis is a reputed medicinal fungus growing parasitically on buried larvae of ghost moths in Asian high-altitude grassland ecosystems. We have analysed the intraspecific ITS nrDNA (ITS1, 5.8S gene, ITS2) variation among 71 sequences of C. sinensis available in EMBL/Genbank. The ITS sequences, submitted to Bayesian ML analyses, were distributed into five groups, referred to as A–E. Nine of the sequences (groups D and E) grouped with distantly related hypocrealean/clavicipitalean taxa and are interpreted as sequences erroneously accessioned under wrong taxon names. The remaining 62 sequences constituted three highly supported clades (groups A–C), that may represent cryptic (phylogenetic) species currently ascribed to C. sinensis. A remarkably high sequence divergence occurred in the 5.8S gene between the three groups. Sequences of groups B and C showed accelerated substitution rates and high AT nucleotide bias. We hypothesize that the accelerated evolution and AT bias have been caused by a shift in life historical attributes or ecology. We also suggest that the recorded differences in medicinal effects among C. sinensis populations may be attributed to the existence of genetically differentiated chemotypes in this morphotaxon.     

Internal transcribed spacer region evolution in Larix and Pseudotsuga (Pinaceae)

The nuclear ribosomal DNA (nrDNA) internal transcribed spacer (ITS) region has been characterized in the sister genera Larix and Pseudotsuga (Pinaceae). Complete sequences were obtained for seven species of Larix from North America and Eurasia and five species of Pseudotsuga from western North America and eastern Asia. ITS region lengths ranged from 1759 to 1770 bp in Larix and from 1564 to 1571 bp in Pseudotsuga. In both genera, ITS1 is three times as long as the 5.8S plus ITS2 and contains subrepeats as observed in other genera of Pinaceae. Secondary structure models predicted that the subrepeats fold into terminal stem and loop domains. ITS polymorphism detected within individuals of Larix and Pseudotsuga suggests a slow rate of concerted evolution among nrDNA loci. Except for the placement of L. sibirica, phylogenetic analyses of the ITS region agreed with previously reported restriction site analyses of Larix and Pseudotsuga. The data were not consistent with phylogenetic hypotheses for Larix based primarily upon ovulate cone characters, failing to support a derivation of the North American L. laricina from a short-bracted Eurasian lineage. The phylogenetic hypothesis did not conflict with a stepping stone model of evolution for Pseudotsuga, but a basal lineage could not be inferred for either genus.     

Revision of the kelp family Alariaceae and the taxonomic affinities of Lessoniopsis Reinke (Laminariales,Phaeophyta)

The morphologically diverse members of the Laminariales are separated into three families, the Alariaceae, Laminariaceae and Lessoniaceae, on the basis of developmental features exhibited at the stipeblade transition zone. We have investigated the relationships of lessoniacean taxa with those of alariacean and laminariacean affinity. This was done by completing phylogenetic analyses of 3 small-subunit (SSU) rRNA gene (113 bp), first internal transcribed spacer (ITS 1), 5.8S rRNA gene and second internal transcribed spacer (ITS2)(12 bp) sequence data from a variety of taxa. In summary, the Alariaceae and Lessoniaceae are polyphyletic. We present a restricted view of the Alariaceae, including only Alaria and Pterygophora of the genera usually placed in this family, and transfer the monotypic genus Lessoniopsis to this group. Current theories of kelp evolution and biogeography must be reconsidered in view of our data.