Internal Transcribed Spacer Sequence Variation Research Articles

Molecular characterization of Trichoderma spp. isolates in mushroom‐growing farms in Turkey and their effects on Agaricus bisporus production

AbstractMushroom production is highly affected by green mould disease, which causes severe losses worldwide. This study aimed to characterize Trichoderma spp. isolates collected from Agaricus bisporus‐ and Pleurotus ostreatus‐growing facilities based on their molecular features and evaluate their aggressiveness in mushroom production in vitro and in vivo. Fifty‐six Trichoderma isolates from mushroom farms in Aegean and Marmara regions were analysed for molecular characteristics. In a phylogenetic tree constructed based on internal transcribed spacer (ITS) and tef1 sequences, they were classified into five groups, which included Trichoderma spp. such as T. aggressivum, T. europaeum, T. pleuroticola, T. virens, T. harzianum and T. afroharzianum. Phylogenetic analyses revealed that tef1 sequences could distinguish between T. aggressivum f. aggressivum and T. aggressivum f. europaeum more specifically. Consequently, tef1 sequence variation identified 15 T. aggressivum f. europaeum isolates compared with ITS sequence variation. In vitro conditions based on the dual‐culture tests of the mycelial growth of A. bisporus showed growth inhibition between 50% and 75% depending on the isolates. Two Trichoderma isolates with different ITS and tef1 sequences (3/9 and 7/5 isolates, respectively) and more lytic and mycoparasitic activities were analysed further to determine their effects on yield and mushroom quality parameters. Therefore, T. aggressivum f. aggressivum is not the only species that affects mushroom production, and T. aggressivum f. aggressivum and T. aggressivum f. europaeum isolates significantly (p < 0.05) inhibited A. bisporus growth in vitro and in vivo.

Genetic Differentiation and Widespread Mitochondrial Heteroplasmy among Geographic Populations of the Gourmet Mushroom Thelephora ganbajun from Yunnan, China.

The mitochondrial genomes are generally considered non-recombining and homoplasmic in nature. However, our previous study provided the first evidence of extensive and stable mitochondrial heteroplasmy in natural populations of the basidiomycete fungus Thelephora ganbajun from Yunnan province, China. The heteroplasmy was characterized by the presence of two types of introns residing at adjacent but different sites in the cytochrome oxidase subunits I (cox1) gene within an individual strain. However, the frequencies of these two introns among isolates from different geographical populations and the implications for the genetic structure in natural populations have not been investigated. In this study, we analyzed DNA sequence variation at the internal transcribed spacer (ITS) regions of the nuclear ribosomal RNA gene cluster among 489 specimens from 30 geographic locations from Yunnan and compared that variation with distribution patterns of the two signature introns in the cox1 gene that are indicative of heteroplasmy in this species. In our samples, evidence for gene flow, abundant genetic diversity, and genotypic uniqueness among geographic samples in Yunnan were revealed by ITS sequence variation. While there was insignificant positive correlation between geographic distance and genetic differentiation among the geographic samples based on ITS sequences, a moderate significant correlation was found between ITS sequence variation, geographical distance of sampling sites, and distribution patterns of the two heteroplasmic introns in the cox1 gene. Interestingly, there was a significantly negative correlation between the copy numbers of the two co-existing introns. We discussed the implications of our results for a better understanding of the spread of stable mitochondrial heteroplasmy, mito-nuclear interactions, and conservation of this important gourmet mushroom.

Differentiation Among Rodentibacter Species Based on 16S-23S rRNA Internal Transcribed Spacer Analysis.

The internal transcribed spacer (ITS) regions of Rodentibacter pneumotropicus, R. heylii, R. rarus, R. ratti, and R. heidelbergensis and of a Rodentibacter- related β-hemolytic Pasteurellaceae taxon isolated from laboratory rodents were studied for their feasibility to discriminate among these species. The 6 species analyzed showed species-specific ITS patterns that were shared by the type strains and clinical isolates and that allowed their identification. Nevertheless, differentiating between the ITS band patterns of R. pneumotropicus and R. ratti is visually challenging. In all species tested, sequence analysis of the ITS fragments revealed a larger ITSile+ala, which contained the genes for tRNAIle(GAU) and tRNA Ala(UGC), and a smaller ITSglu with the tRNAGlu(UUC) gene. The ITS sequences varied among the 6 species evaluated, displaying identity levels ranging from 62% to 86% for ITSile+ala and 68% to 90% for ITSglu. Overall, ITS amplification proved to be a reliable method to differentiate among these important Pasteurellaceae species of laboratory rodents. Moreover, the ITS sequence variations recorded here might facilitate the design of probes for specific identification of these species. The ability to diagnose these organisms to the species level could increase our understanding of their clinical significance.

Plastome phylogeny and lineage diversification of Salicaceae with focus on poplars and willows.

Phylogenetic relationships and lineage diversification of the family Salicaceae sensu lato (s.l.) remain poorly understood. In this study, we examined phylogenetic relationships between 42 species from six genera based on the complete plastomes. Phylogenetic analyses of 77 protein coding genes of the plastomes produced good resolution of the interrelationships among most sampled species and the recovered clades. Of the sampled genera from the family, Flacourtia was identified as the most basal and the successive clades comprised both Itoa and Poliothyrsis, Idesia, two genera of the Salicaceae sensu stricto (s.s.) (Populus and Salix). Five major subclades were recovered within the Populus clade. These subclades and their interrelationships are largely inconsistent with morphological classifications and molecular phylogeny based on nuclear internal transcribed spacer sequence variations. Two major subclades were identified for the Salix clade. Molecular dating suggested that species diversification of the major subclades in the Populus and Salix clades occurred mainly within the recent Pliocene. In addition, we found that the rpl32 gene was lost and the rps7 gene evolved into a pseudogene multiple times in the sampled genera of the Salicaceae s.l. Compared with previous studies, our results provide a well‐resolved phylogeny from the perspective of the plastomes.

Species delimitation of Chinese hop-hornbeams based on molecular and morphological evidence.

Species delimitation through which infers species boundaries is emerging as a major work in modern systematics. Hop‐hornbeam species in Ostrya (Betulaceae) are well known for their hard and heavy woods. Five species were described in China and their interspecific delimitations remain unclear. In this study, we firstly explored their distributions in all recorded field sites distributed in China. We then selected 110 samples from 22 natural populations of five species from this genus and one type specimen of O. yunnanensis, for molecular barcoding analyses. We sequenced four chloroplast (cp) DNA fragments (trnH–psbA, trnL–trnF, rps16, and trnG) and the nuclear internal transcribed spacer (ITS) region for all samples. Sequence variations of Ostrya from four cpDNA fragments identified three groups that showed no correspondence to any morphological delimitation because of the incomplete lineage sorting and/or possible interspecific introgression in the history. However, phylogenetic analyses of ITS sequence variations discerned four species, O. japonica, O. rehderiana, O. trichocarpa, and O. multinervis while O. yunnanensis nested within O. multinervis. Morphological clustering also discerned four species and showed the complete consistency with molecular evidence. Moreover, our phylogenetic analyses‐based ITS sequence variations suggested that O. trichocarpa comprised an isolated lineage different from the other Eurasian ones. Based on these results, hop‐hornbeams in China should be treated as four separate species. Our results further highlight the importance of ITS sequence variations in delimitating and discerning the closely related species in plants.

Phylotranscriptomic analyses in plants using Betulaceae as an example

AbstractWith the advance of next‐generation sequencing techniques, phylogenetic analyses based on genomic data are becoming frequent in plants. However, the efficacy of different methods used to retrieve orthologs for phylogenetic studies based on transcriptome sequences is rarely compared and examined for closely related species with low genetic differentiation. In this study, we used both low‐copy gene‐based approaches and mapping‐based approaches to construct a phylogeny of Betulaceae. We found that both approaches showed no distinct differences in phylogeny construction and associated bootstrap support, although mapping‐based methods seemed to be superior to the former in all analyses particularly when the summed length of orthologs was not long enough. Phylogenetic relationships within the family, largely consistent with those previously based on chloroplast DNA and nuclear internal transcribed spacer sequence variations, received high support for all clades and subclades. However, we found that support values for the sister relationship between two polyploid genera were lower than those between diploid groups if only a few orthologs were sampled. This may result from ortholog misidentification due to genome duplications.

High Levels of Genetic Diversity within One Population of Rheum tanguticum on the Qinghai-Tibet Plateau have Implications for Germplasm Conservation

Rheum tanguticum, a source plant for the traditional Chinese medicine Dahuang (rhubarb), is an endangered species. In this study, we first assessed genetic diversity using 102 internal transcribed spacer (ITS) sequences cloned from one population of R. tanguticum on the Qinghai-Tibet Plateau. In total, 1773 polymorphic sites were identified, including 707 sites in the ITS1 region, 773 sites in the ITS2 region, and 293 sites on the 5.8S gene. Overall, transitions were the most common type of mutation and represented 77% of all base substitutions found within the ITS regions of R. tanguticum. We found that the average intra-specific genetic distance was 0.1260±0.0755 and ranged from 0 to 0.3618. Furthermore, a neighbor-joining phylogenetic tree constructed using these sequences strongly supported the conclusion that abundant intra-specific variation exists within the ITS regions of R. tanguticum. Interestingly, we found that eightyseven clones of R. tanguticum had high similarity to the ITS sequence of R. rhabarbarum and ten clones had high similarity to the ITS sequence of R. officinale; however, five clones of R. tanguticum had similarity to Fagopyrum esculentum. Therefore, we observed a significant amount of ITS sequence variation at both the intra-genomic and intra-specific levels in R. tanguticum. Based on these results, we suggest that in situ conservation is preferable for R. tanguticum protection.

ITS sequence variation and concerted evolution in the natural hybrid species Malus toringoides

The internal transcribed spacer (ITS) region of nuclear ribosomal DNA (nrDNA) is one of the most used molecular characters in plant systematics. Our previous studies based on morphological analysis and ITS sequence variation suggested that Malus toringoides (Rehd.) Hughes is derived from hybridization between M. transitoria (Batal.) Schneid. and M. kansuensis (Batal.) Schneid. To further understand the variation pattern of ITS sequences in M. toringoides, and to elucidate the evolutionary processes that affect ITS sequence variation after hybridization, we sampled 99 accessions from multiple populations of the hybrid and parental species, and then obtained totally 254 ITS sequences by cloning and sequencing. Our ITS variation data demonstrates three outcomes of ITS repeats after hybrid speciation. ∼ 27–41% of M. toringoides have only M. transitoria type ITS sequence, ∼ 40–70% have M. transitoria type ITS sequence plus one or two chimeric ITS sequences generated by recombination between parental ITS sequences, and six accessions retain both parental type ITS sequences. The plausible evolutionary processes that created the observed ITS variations were inferred to be the joint actions of recombination, concerted evolution, pseudogenization and backcrossing. Our study provides further understandings of the variation model of ITS repeats after hybridization as well as the evolution of M. toringoides after its hybrid speciation.

Genetic diversity assessment of Diplocyclos palmatus (L.) C. Jeffrey from India using internal transcribed spacer (ITS) sequences of nuclear ribosomal DNA

Internal transcribed spacer (ITS) region of nuclear ribosomal DNA from 20 populations of Diplocyclos palmatus (L.) C. Jeffrey belonging to five different geographical locations (Bihar, Jharkhand, Maharashtra, Madhya Pradesh, and Tamil Nadu) of India were sequenced. Analysis of nucleotide sequences reveals polymorphism among the populations [π = 0.01482, θw = 0.01180 (0.00236) and total variance 3.260]. AMOVA was used to partition the genetic diversity and tested whether there is any hierarchy of ITS sequence variation among individuals. The genetic differentiation between the populations is high (ΦST = 0.591). The maximum likelihood tree reveals the evolution of D. palmatus under reproductive isolation and under different environmental conditions which may be most probably due to long distance distribution, and possibility of genetic exchange among the populations of D.palmatus distributed in Bihar and Jharkhnad. Key words : Diplocyclos palmatus (L.) C. Jeffrey, genetic diversity, internal transcribed spacer (ITS), nrDNA.

Cortinarius species diversity in British Columbia and molecular phylogenetic comparison with European specimen sequences

Throughout the world, the diversity of fungi remains poorly characterized and Cortinarius is a classical example of a difficult, species-rich, and under-researched mushroom genus. Here, we sequenced and analyzed ribosomal internal transcribed spacer (ITS) sequence barcodes from herbarium specimens to improve understanding of Cortinarius species diversity in British Columbia (B.C.), Canada. Starting with 962 specimen sequences, 617 from B.C. herbaria, we present a maximum likelihood tree showing 179 putative Cortinarius species in British Columbia. As a working definition, we considered a “species” to be a monophyletic clade that included a reliably identified reference sequence, with a maximum of 3% ITS sequence variation. If no reference sequence was available, “species” were groups sharing 97% or more sequence identity. By these criteria, 110 putative B.C. species matched European species and 12 B.C. species matched species exclusively found in the Americas. Of the 56 B.C. species that did not match an identified reference sequence, some may be new to science, while others likely represent described species without available sequences. By depositing sequences from B.C. specimens into GenBank and BOLD, and by providing our alignment to TreeBASE, we have supplied the resources necessary to improve accuracy in identifications of Cortinarius in future systematic and ecological studies.

Evolutionary history of an alpine shrub Hippophae tibetana (Elaeagnaceae): allopatric divergence and regional expansion

Increasing evidence suggests that geological or climatic events in the past promoted allopatric speciation of alpine plants in the Qinghai-Tibetan Plateau and adjacent region. However, few studies have been undertaken to examine whether such allopatric divergences also occurred within a morphologically uniform species. In the present study, we report the evolutionary history of an alpine shrub species, Hippophae tibetana, based on examining chloroplast DNA (cpDNA) and nuclear ribosomal internal transcribed spacer (ITS) DNA variations. We sequenced two cpDNA fragments (trnL-F and trnS-G) and the nuclear ITS region in 183 individuals collected from 21 natural populations. Ten chlorotypes and 17 ITS types were identified. Phylogenetic analyses of both chlorotypes and ITS sequence variations suggested two distinct lineages distributed in the eastern and western region, respectively. On the basis of the fast and low plant substitution rates, these two lineages were estimated to have diverged from each other between 1 and 4 million years ago, during the period of the major glaciations and orogenic processes. In addition, ITS has undergone the accelerated evolution in two populations in the southern Himalaya isolated by the high mountains with a surprising accumulation of the private variations. The east-west split was also supported by an analysis of molecular variance, which partitioned around 91% of the total cpDNA variance between these two groups of populations. A single chlorotype was found for most populations in eastern or western region, suggesting a recent postglacial expansion within each region. Star-phylogeny and mismatch analyses of all chlorotypes within the eastern group of populations suggested an earlier regional expansion before the Last Glacial Maximum (LGM). The local fixture of the different chlorotypes in multiple populations suggested more than one refugia remained for eastern or western region. Coalescent tests rejected the hypothesis that all current populations originated from a single refugium during the LGM. Instead, they supported hypothesis that two lineages diverged before the late Pleistocene. These findings, when taken together, suggested that this species had experienced long allopatric divergence and recent regional range expansions in response to orogenic processes and the climate changes. The evolutionary history of this shrub species highlights importance of geographical isolations to the intraspecific divergence of alpine plants occurring in the world's ruff. (c) 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2011, 102, 37-50.

Evolutionary lineages of nickel hyperaccumulation and systematics in European Alysseae (Brassicaceae): evidence from nrDNA sequence data.

Nickel (Ni) hyperaccumulation is a rare form of physiological specialization shared by a small number of angiosperms growing on ultramafic soils. The evolutionary patterns of this feature among European members of tribe Alysseae (Brassicaceae) are investigated using a phylogenetic approach to assess relationships among Ni hyperaccumulators at the genus, species and below-species level. Internal transcribed spacer (ITS) sequences were generated for multiple accessions of Alysseae. Phylogenetic trees were obtained for the genera of the tribe and Alyssum sect. Odontarrhena. All accessions and additional herbarium material were tested for Ni hyperaccumulation with the dimethylglyoxime colorimetric method. Molecular data strongly support the poorly known hyperaccumulator endemic Leptoplax (Peltaria) emarginata as sister to hyperaccumulator species of Bornmuellera within Alysseae. This is contrary to current assumptions of affinity between L. emarginata and the non-hyperaccumulator Peltaria in Thlaspideae. The lineage Bornmuellera-Leptoplax is, in turn, sister to the two non-hyperaccumulator Mediterranean endemics Ptilotrichum rupestre and P. cyclocarpum. Low ITS sequence variation was found within the monophyletic Alyssum sect. Odontarrhena and especially in A. murale sensu lato. Nickel hyperaccumulation was not monophyletic in any of three main clades retrieved, each consisting of hyperaccumulators and non-hyperaccumulators of different geographical origin. Nickel hyperaccumulation in Alysseae has a double origin, but it did not evolve in Thlaspideae. In Bornmuellera-Leptoplax it represents an early synapomorphy inherited from an ancestor shared with the calcicolous, sister clade of Mediterranean Ptilotrichum. In Alyssum sect. Odontarrhena it has multiple origins even within the three European clades recognized. Lack of geographical cohesion suggests that accumulation ability has been lost or gained over the different serpentine areas of south Europe through independent events of microevolutionary adaptation and selection. Genetic continuity and strong phenotypic plasticity in the A. murale complex call for a reduction of the number of Ni hyperaccumulator taxa formally recognized.

Detection of multiple infections by Monocystis strains in a single earthworm host using ribosomal internal transcribed spacer sequence variation

Monocystis sp. are sporocyst-forming apicomplexan parasites common in seminal vesicles of the earthworm Lumbricus terrestris where they may account for temporary castration. This study describes the internal transcribed spacer (ITS) region of the ribosomal cistron of Monocystis sp. This region, including ITS-1, the 5.8S ribosomal RNA gene, and ITS-2, was PCR amplified, cloned, and sequenced for Monocystis sp. isolated from the seminal vesicles of several wild-caught L. terrestris. Our analysis revealed substantial polymorphisms, also within single host organisms, indicating intra-host diversity of parasites. These genetic markers are the first that allow distinction of Monocystis sp. genotypes, opening new avenues for the study of parasite diversity within and between hosts.

History and evolution of alpine plants endemic to the Qinghai‐Tibetan Plateau: Aconitum gymnandrum (Ranunculaceae)

How Quaternary climatic oscillations affected range distributions and intraspecific divergence of alpine plants on the Qinghai-Tibetan Plateau (QTP) remains largely unknown. Here, we report a survey of chloroplast DNA (cpDNA) and nuclear ribosomal internal transcribed spacer (ITS) DNA variation aimed at exploring the phylogeographical history of the QTP alpine endemic Aconitum gymnandrum. We sequenced three cpDNA fragments (rpl20-rps12 intergenic spacer, the trnV intron and psbA-trnH spacer) and also the nuclear (ITS) region in 245 individuals from 23 populations sampled throughout the species' range. Two distinct lineages, with eastern and western geographical distributions respectively, were identified from a phylogenetic analysis of ITS sequence variation. Based on a fast substitution rate, these were estimated to have diverged from each other in the early Pleistocene approximately 1.45 Ma. The analysis of cpDNA variation identified nine chlorotypes that clustered into two major clades that were broadly congruent in geographical distribution with the two ITS lineages. The east-west split of cpDNA divergence was supported by an amova which partitioned approximately half of the total variance between these two groups of populations. Analysis of the spatial distribution of chlorotypes showed that each clade was subdivided into two groups of populations such that a total of four population groups existed in the species. It is suggested that these different groups derive from four independent glacial refugia that existed during the Last Glacial Maximum (LGM), and that three of these refugia were located at high altitude on the QTP platform itself at that time. Coalescent simulation of chlorotype genealogies supported both an early Pleistocene origin of the two main cpDNA clades and also the 'four-refugia' hypothesis during the LGM. Two previous phylogeographical studies of QTP alpine plants indicated that such plants retreated to refugia at the eastern/south-eastern plateau edge during the LGM and/or previous glacial maxima. However, the results for A. gymnandrum suggest that at least some of these cold-tolerant species may have also survived centrally on the QTP platform throughout the Quaternary.

Intraspecific and within-isolate sequence variation in the ITS rRNA gene region ofPythium mercurialesp. nov. (Pythiaceae)

Sixteen Pythium isolates from diverse hosts and locations, which showed similarities in their morphology and sequences of the internal transcribed spacer (ITS) region of their rRNA gene, were investigated. As opposed to the generally accepted view, within single isolates ITS sequence variations were consistently found mostly as part of a tract of identical bases (A-T) within ITS1, and of GT or GTTT repeats within the ITS2 sequence. Thirty-one different ITS sequences obtained from 39 cloned ITS products from the 16 isolates showed high sequence and length polymorphisms within and between isolates. However, in a phylogenetic analysis, they formed a cluster distinct from those of other Pythium species. Additional sequencing of two nuclear genes (elongation factor 1 alpha and beta-tubulin) and one mitochondrial gene (nadh1) revealed high levels of heterozygosity as well as polymorphism within and between isolates, with some isolates possessing two or more alleles for each of the nuclear genes. In contrast to the observed variation in the ITS and other gene areas, all isolates were phenotypically similar. Pythium mercuriale sp. nov. (Pythiaceae) is characterized by forming thin-walled chlamydospores, subglobose to obovoid, papillate sporangia proliferating internally and smooth-walled oogonia surrounded by multiple antheridia. Maximum likelihood phylogenetic analyses based on both ITS and beta-tubulin sequence data place P. mercuriale in a clade between Pythium and Phytophthora.

ITS sequence variation supports the hybrid origin ofMalus toringoidesHughes

Malus toringoides (Rehd.) Hughes was suggested to have originated from hybridization between Malus transitoria Schneid. and Malus kansuensis Rehd., followed by repeated backcrossing to one of the putative parents. In the present study, the sequence information of the internal transcribed spacer (ITS) of nuclear ribosomal DNA (nrDNA) was used to re-examine the origin of this species. A total of 69 accessions from three natural populations (Maerkang, Xiaba and Kehe, Aba Autonomous Region, Sichuan, China) of M. toringoides and 10 accessions of its putative parents were analyzed. Using Malus angustifolia (Ait.) Michx., Malus ioensis (Wood) Britt. and Malus doumeri Chev. as outgroups, our phylogenetic analysis of the ITS sequences of M. toringoides and its putative parents showed that M. toringoides was not monophyletic, and two different types of ITS sequences which were obtained from each of the six accessions of M. toringoides were found to have clustered separately with those of the two putative parent species on the gene tree. A comparison of the sequence variation between M. toringoides and its putative parents revealed an additive variation pattern of ITS sequences in the putative hybrid species. These results are consistent with the previous morphological and amplified fragment length polymorphism (AFLP) data which suggested that M. toringoides was of hybrid origin. Our ITS data provide new molecular evidence for the hybrid origin hypothesis of M. toringoides and these results are of great importance for future study on hybridization, polyploid speciation and evolution of the genus Malus Miller.

Phylogenetic relationships, evolutionary origin, taxonomic status, and genetic structure of the endangered local Lower Elbe river (Germany) endemic Oenanthe conioides (Nolte ex Rchb.f.) Lange (Apiaceae): ITS and AFLP evidence

Oenanthe conioides is an endangered local endemic of the Lower Elbe river region in Germany where it is found in areas with freshwater tides. Internal transcribed spacer (ITS) and amplified fragment length polymorphism (AFLP) markers were used to investigate its phylogenetic relationships, evolutionary origin, taxonomic status, and genetic structure. Oe. conioides is most closely related to Oe. aquatica and Oe. fluviatilis, and cannot be distinguished from Oe. aquatica by ITS sequence variation. Oe. aquatica is found mainly in standing or slow-flowing freshwater habitats. The AFLP analysis indicated that Oe. conioides may constitute a monophyletic lineage nested within Oe. aquatica. Considering this result on the background of the sympatric distribution of the two species and their ecological differentiation, it is hypothesized that Oe. conioides arose sympatrically from Oe. aquatica through ecological divergence. It is concluded that Oe. conioides is best considered a subspecies of Oe. aquatica. AFLP analysis of Oe. conioides revealed low levels of genetic differentiation among populations reflecting the small geographical distance, the recent decline in population number and size, the connection of populations through the river, and their presumably high population dynamics. Considering this pattern of genetic variation, it is concluded that no individual population can be singled out as being particularly important from a genetic point of view. Also, all populations could be used as source material should the establishment of new populations or the enlargement of small populations be considered as a measure to consolidate the persistence of the species.

The liverwort Marchantia foliacea forms a specialized symbiosis with arbuscular mycorrhizal fungi in the genus Glomus

Microscopic evidence suggests that fungi forming endosymbioses with liverworts in the Marchantiales are arbuscular mycorrhizal (AM) fungi from the Glomeromycota. Polymerase chain reaction amplification of ribosomal sequences confirmed that endophytes of the New Zealand liverwort, Marchantia foliacea, were members of the genus Glomus. Endophytes from two Glomus rDNA phylotypes were repeatedly isolated from geographically separated liverwort samples. Multiple phylotypes were present in the same liverwort patch. The colonizing Glomus species exhibited substantial internal transcribed spacer sequence variation within phylotypes. This work suggests that certain liverwort species may serve as a model for studying DNA sequence variation in colonizing AM phylotypes and specificity in AM-host relationships.

Unexpected high divergence in nrDNA ITS and extensive parallelism in floral morphology of Pedicularis (Orobanchaceae)

Internal transcribed spacer (ITS) region sequences of nuclear ribosomal DNA (nrDNA) were used in the phylogenetic reconstruction of Pedicularis, a genus with strong adaptive radiation. Forty-two species representing 12 greges of the genus were all, except P. resupinata, sampled from the Hengduan Mountain region, China. A high level of ITS sequence variation was found among the species distributed in such a small geographical area, which had been rarely reported in other groups. The great ITS divergence in Pedicularis could be explained by a relatively ancient origin and diversification of the genus followed by migration of different floristic components into the Hengduan Mountains, or accelerated rates of molecular evolution in parasitic lineages of Orobanchaceae. In the present ITS phylogeny, almost all the main clades are not consistent with the high hierarchical taxa within Pedicularis, which implies that significant parallel evolution occurred in floral morphology of the genus, and that undue attention has been paid to corolla characters in the intragenus classifications.

Phylogenetic relationships in Anthemis L. (Compositae, Anthemideae) based on nrDNA ITS sequence variation

SummaryOberprieler, C.: Phylogenetic relationships in Anthemis L. (Compositae, Anthemideae) based on nrDNA ITS sequence variation. Taxon 50: 745–762. 2001. ISSN 0040‐0262.Sequences of the nrDNA internal transcribed spacer (ITS) region were analysed for 29 representatives of Anthemis L. and six outgroup taxa of Anthemideae (Compositae) to study the delimitation and the infrageneric taxonomy of this large and widespread Eurasian and African genus. Species of Anthemis included all subgenera and sections traditionally used in supraspecific classifications of the genus. Maximum parsimony (MP), maximum likelihood (ML), and distance (neighbor joining, NJ) analyses were carried out for phylogenetic reconstructions. The results suggest that the genus, in its traditional circumscription, is not monophyletic due to the consistent placement of a member of Tripleurospermum amidst the Anthemis representatives. Subdivision of Anthemis into two subgenera (A. subg. Anthemis, A. subg. Cota) is supported, with the exception of the placement of A. sect. Chiae and sect. Odontostephanae which fall outside A. subg. Anthemis. Within A. subg. Anthemis, life form dependent delimitation of A. sect. Anthemis (annuals) against A. sect. Hiorthia (perennials) is not supported, whereas the monophyly of other sections (A. sect. Maruta, A. sect. Rascheyanae) is confirmed including the placement of Ammanthus Boiss. & Heldr. into Anthemis subg. Anthemis.