Nuclear Internal Transcribed Spacer Sequences Research Articles

Molecular faunistics of accidental infections of Gyrodactylus Nordmann, 1832 (Monogenea) parasitic on salmon Salmo salar L. and brown trout Salmo trutta L. in NW Russia

Salmon Salmo salar L. and brown trout S. trutta L. juveniles were examined for the presence of accidental monogenean ectoparasitic species of Gyrodactylus Nordmann, 1832 in the Baltic and White Sea basins of Russian Karelia in order to estimate the frequency of host-switching attempts on an ecological timescale. To collect phylogeographical information and for exact species identification, the parasites were characterised by nuclear internal transcribed spacer sequences of rDNA (ITS) and, for some species, also by their mitochondrial DNA (CO1 gene) sequences. Four accidental Gyrodactylus species were observed on salmon and brown trout. A few specimens of G. aphyae Malmberg, 1957, the normal host of which is the Eurasian minnow Phoxinus phoxinus (L.), were observed on lake salmon from the Rivers Kurzhma (Lake Kuito, White Sea basin) and Vidlitsa (Lake Ladoga, Baltic basin). G. lucii Kulakovskaya, 1952, a parasite of the northern pike Esox lucius L., was observed on salmon in the Kurzhma. In the River Vidlitsa, two specimens of G. papernai Ergens & Bychowsky, 1967, normally on stone loach Barbatula barbatula (L.), were found on salmon. On anadromous White Sea salmon in the River Pulonga in Chupa Bay, a few salmon parr carried small colonies of G. arcuatus Bychowsky, 1933, which were shown to have originated from the local three-spined stickleback Gasterosteus aculeatus L. consumed as prey. No specimens of Gyrodactylus salaris Malmberg, 1957 were observed, although the Pulonga is the nearest salmon spawning river to the River Keret', which is heavily infected with introduced G. salaris. In the River Satulinoja, Lake Ladoga, three specimens of G. lotae Gusev, 1953, from burbot Lota lota (L.), were collected from a single brown trout S. trutta. All nonspecific gyrodactylid infections on salmonids were judged to be temporary, because only a few specimens were observed on each of the small number of infected fishes. The prevalence of endemic G. salaris was also low, only 1% (Nfish = 296) in Lake Onega and 0.7% (Nfish = 255) in Lake Ladoga, while brown trout specific Gyrodactylus species were not observed on any of the 429 trout examined from the Ladoga basin. The host-specific and unspecific burden of Gyrodactylus spp. on these 'glacial relict' populations of salmon and brown trout was very low, suggesting a generalised resistance against the co-evolved freshwater parasite community, or some kind of 'vaccination' effect. These hypotheses deserve further testing.

Phylogenetic relationships of wild and cultivated species of Allium section Cepa inferred by nuclear rDNA ITS sequence analysis

Allium section Cepa consists of 12 species most of which are used by humans as condiment, vegetable or medicinal plants. Common onion (Allium cepa) and bunching onion (A. fistulosum) are cultivated species while all others are locally collected from the wild. Although common onion is the most important crop within Allium, its wild progenitor and origin are still not clear. We analyzed the phylogeny of Allium section Cepa using sequences of the nuclear ribosomal DNA internal transcribed spacer (ITS) region of 36 accessions representing eleven species of section Cepa, together with eight outgroup species with phenetic, cladistic, and model-based algorithms. These analyses confirmed section Cepa to be monophyletic and revealed three species groups within the section. These are (i) A. altaicum/A. fistulosum, (ii) A. farctum/A. roylei/A. asarense/A. cepa/A. vavilovii, and (iii) A. galanthum/A. oschaninii/A. praemixtum/A. pskemense. While the first two groups were statistically well supported for the last group support was low, although it resulted in all phylogenetic analyses conducted. Tree and network-based analyses grouped A. cepa within A. vavilovii, indicating the latter to be progenitor of the common onion. However, also an origin of A. cepa through hybridization of A. vavilovii with A. galanthum or A. fistulosum seems possible. We argue that a subdivision of section Cepa in subsections Cepa and Phyllodolon, although possible from our data, as well as the formal description of alliances do not seem reasonable in a small group of species.

An updated estimate of intergeneric phylogenetic relationships in the Australian VitaceaeThis article is one of a selection of papers presented at the symposium onVitisat the XVII International Botanical Congress held in Vienna, Austria, in 2005.

Previous molecular studies have suggested that the genus Cissus (Vitaceae) is not monophyletic. The majority of species are grouped, but four Australian taxa ( Cissus antarctica Vent., Cissus hypoglauca A. Gray, Cissus oblonga (Benth.) Planch., and Cissus sterculiifolia (F. Muell. ex Benth.) Planch.) form a distinct clade and Cissus opaca has now been recognized as belonging to the genus Clematicissus ( Clematicissus opaca (F. Muell.) Jackes & Rossetto). Here, we investigate relationships among the Australian Vitaceae by parsimony and Bayesian analysis of plastid trnL-trnF and nuclear internal transcribed spacer sequences and include a range of taxa that had not been previously investigated. We find no support for a close relationship between Nothocissus and the four distinct Australian Cissus, as suggested in previous morphology-based treatments. We find a robust sister relationship between Clematicissus and at least two southern American Cissus ( Cissus tweediana (Baker) Planch. and Cissus striata Ruiz & Pav.), suggesting a possible origin from an ancient southern progenitor. Finally, this study confirms the paraphyletic nature of Cayratia , with species occurring in two clades, both with Australian and non-Australian species. The necessity in future studies for data from additional and more tractable nuclear loci is also noted.

Phylogenetics of Cucumis (Cucurbitaceae)

Background: Melon, Cucumis melo, and cucumber, C. sativus, are among the most widely cultivated crops worldwide. Cucumis, as traditionally conceived, is geographically centered in Africa, with C. sativus and C. hystrix thought to be the only Cucumis species in Asia. This taxonomy forms the basis for all ongoing Cucumis breeding and genomics efforts. We tested relationships among Cucumis and related genera based on DNA sequences from chloroplast gene, intron, and spacer regions (rbcL, matK, rpl20-rps12, trnL, and trnL-F), adding nuclear internal transcribed spacer sequences to resolve relationships within Cucumis. Results: Analyses of combined chloroplast sequences (4,375 aligned nucleotides) for 123 of the 130 genera of Cucurbitaceae indicate that the genera Cucumella, Dicaelospermum, Mukia, Myrmecosicyos, and Oreosyce are embedded within Cucumis. Phylogenetic trees from nuclear sequences for these taxa are congruent, and the combined data yield a well-supported phylogeny. The nesting of the five genera in Cucumis greatly changes the natural geographic range of the genus, extending it throughout the Malesian region and into Australia. The closest relative of Cucumis is Muellerargia, with one species in Australia and Indonesia, the other in Madagascar. Cucumber and its sister species, C. hystrix, are nested among Australian, Malaysian, and Western Indian species placed in Mukia or Dicaelospermum and in one case not yet formally described. Cucumis melo is sister to this Australian/Asian clade, rather than being close to African species as previously thought. Molecular clocks indicate that the deepest divergences in Cucumis, including the split between C. melo and its Australian/Asian sister clade, go back to the mid-Eocene. Conclusion: Based on congruent nuclear and chloroplast phylogenies we conclude that Cucumis comprises an old Australian/Asian component that was heretofore unsuspected. Cucumis sativus evolved within this Australian/Asian clade and is phylogenetically far more distant from C. melo than implied by the current morphological classification.

Phylogenetics of Cucumis (Cucurbitaceae): Cucumber (C. sativus) belongs in an Asian/Australian clade far from melon (C. melo)

BackgroundMelon, Cucumis melo, and cucumber, C. sativus, are among the most widely cultivated crops worldwide. Cucumis, as traditionally conceived, is geographically centered in Africa, with C. sativus and C. hystrix thought to be the only Cucumis species in Asia. This taxonomy forms the basis for all ongoing Cucumis breeding and genomics efforts. We tested relationships among Cucumis and related genera based on DNA sequences from chloroplast gene, intron, and spacer regions (rbcL, matK, rpl20-rps12, trnL, and trnL-F), adding nuclear internal transcribed spacer sequences to resolve relationships within Cucumis.ResultsAnalyses of combined chloroplast sequences (4,375 aligned nucleotides) for 123 of the 130 genera of Cucurbitaceae indicate that the genera Cucumella, Dicaelospermum, Mukia, Myrmecosicyos, and Oreosyce are embedded within Cucumis. Phylogenetic trees from nuclear sequences for these taxa are congruent, and the combined data yield a well-supported phylogeny. The nesting of the five genera in Cucumis greatly changes the natural geographic range of the genus, extending it throughout the Malesian region and into Australia. The closest relative of Cucumis is Muellerargia, with one species in Australia and Indonesia, the other in Madagascar. Cucumber and its sister species, C. hystrix, are nested among Australian, Malaysian, and Western Indian species placed in Mukia or Dicaelospermum and in one case not yet formally described. Cucumis melo is sister to this Australian/Asian clade, rather than being close to African species as previously thought. Molecular clocks indicate that the deepest divergences in Cucumis, including the split between C. melo and its Australian/Asian sister clade, go back to the mid-Eocene.ConclusionBased on congruent nuclear and chloroplast phylogenies we conclude that Cucumis comprises an old Australian/Asian component that was heretofore unsuspected. Cucumis sativus evolved within this Australian/Asian clade and is phylogenetically far more distant from C. melo than implied by the current morphological classification.

Molecular Phylogeny of the East Asian–North American Disjunct Smilax Sect. Nemexia (Smilacaceae)

Phylogenetic relationships among the species of Smilax sect. Nemexia (Smilacaceae) were investigated by separate and combined cladistic analyses of plastid trnL‐F, rpl16, and nuclear internal transcribed spacer sequences. Results strongly support a clade of two well‐defined East Asian species, Smilax riparia and S. nipponica. Likewise, all North American species form a clade in which S. jamesii, the only species from western North America, is sister to the morphologically variable “S. herbacea complex” from eastern North America. Our data are inconclusive regarding whether this complex should be treated as one or more species. Smilax sect. Nemexia exhibits an East Asian–North American phytogeographic disjunction. The common ancestor of the group is hypothesized to have originated in Asia and to have spread northeastward to North America through the Bering Strait land bridge. Pleistocene glaciation likely fragmented the continuous distribution of the group into the three refugial areas where the species grow today: East Asia, northern California, and the eastern United States. On both continents, nonclimbing erect herbs evolved independently from climbing ancestors within the section.

A Beauveria phylogeny inferred from nuclear ITS and EF1-α sequences: evidence for cryptic diversification and links to Cordyceps teleomorphs

Beauveria is a globally distributed genus of soil-borne entomopathogenic hyphomycetes of interest as a model system for the study of entomo-pathogenesis and the biological control of pest insects. Species recognition in Beauveria is difficult due to a lack of taxonomically informative morphology. This has impeded assessment of species diversity in this genus and investigation of their natural history. A gene-genealogical approach was used to investigate molecular phylogenetic diversity of Beauveria and several presumptively related Cordyceps species. Analyses were based on nuclear ribosomal internal transcribed spacer (ITS) and elongation factor 1-alpha (EF1-α) sequences for 86 exemplar isolates from diverse geographic origins, habitats and insect hosts. Phylogenetic trees were inferred using maximum parsimony and Bayesian likelihood methods. Six well supported clades within Beauveria, provisionally designated A–F, were resolved in the EF1-α and combined gene phylogenies. Beauveria bassiana, a ubiquitous species that is characterized morphologically by globose to subglobose conidia, was determined to be non-monophyletic and consists of two unrelated lineages, clades A and C. Clade A is globally distributed and includes the Asian teleomorph Cordyceps staphylinidaecola and its probable synonym C. bassiana. All isolates contained in Clade C are anamorphic and originate from Europe and North America. Clade B includes isolates of B. brongniartii, a Eurasian species complex characterized by ellipsoidal conidia. Clade D includes B. caledonica and B. vermiconia, which produce cylindrical and comma-shaped conidia, respectively. Clade E, from Asia, includes Beauveria anamorphs and a Cordyceps teleomorph that both produce ellipsoidal conidia. Clade F, the basal branch in the Beauveria phylogeny includes the South American species B. amorpha, which produces cylindrical conidia. Lineage diversity detected within clades A, B and C suggests that prevailing morphological species concepts underestimate species diversity within these groups. Continental endemism of lineages in B. bassiana s.l. (clades A and C) indicates that isolation by distance has been an important factor in the evolutionary diversification of these clades. Permutation tests indicate that host association is essentially random in both B. bassiana s.l. clades A and C, supporting past assumptions that this species is not host specific. In contrast, isolates in clades B and D occurred primarily on coleopteran hosts, although sampling in these clades was insufficient to assess host affliation at lower taxonomic ranks. The phylogenetic placement of Cordyceps staphylinidaecola/bassiana, and C. scarabaeicola within Beauveria corroborates prior reports of these anamorph-teleomorph connections. These results establish a phylogenetic framework for further taxonomic, phylogenetic and comparative biological investigations of Beauveria and their corresponding Cordyceps teleomorphs.

A phylogeny of all species of Arceuthobium (Viscaceae) using nuclear and chloroplast DNA sequences

The genus Arceuthobium (dwarf mistletoes, Viscaceae) comprises 42 species that parasitize hosts in Pinaceae and Cupressaceae in the Old and New Worlds. Maximum parsimony analyses were conducted on two data partitions (separately and combined): nuclear ribosomal internal transcribed spacer (ITS) sequences for all 42 currently recognized species and chloroplast trnT-L-F sequences for 34 New World species. The Old and New World species were phylogenetically distinct using ITS, thus making subgenus Arceuthobium paraphyletic. Arceuthobium pendens and A. guatemalense comprise the basalmost clade of subgenus Vaginata, characterized by the presence of flabellate secondary branching. The trnT-L-F sequences, which vary widely in length depending upon taxon, contain three times less phylogenetic signal than ITS, although homoplasy for this partition is lower. Several of the clades obtained from analysis of nuclear ITS sequences are also recovered using trnT-L-F sequences such as A. guatemalense and A. pendens, the A. rubrum group, the A. vaginatum group, and the A. campylopodum group. The ITS + trnT-L-F tree is well resolved except for four internal nodes. A revised classification of the genus is discussed that recognizes only monophyletic species that are well differentiated by molecular data.

Phylogenetic Relationships among the Primarily North American Genera of Cichorieae (Compositae) based on Analysis of 18S–26S Nuclear rDNA ITS and ETS Sequences

Abstract Phylogenetic analyses of internal transcribed spacer (ITS), external transcribed spacer (ETS), and 5.8S gene sequences of 18S–26S nuclear rDNA from all 23 genera of Cichorieae with centers of diversity in North America (and Picrosia from South America) show that all but three of the genera (Glyptopleura, Krigia, and Phalacroseris) belong to a series of seven clades that are well supported by bootstrap values >90%. Phalacroseris, endemic to California, with a single species (P. bolanderi), is sister to a well-supported (>95% bootstrap) clade that includes all other principally North American genera (plus Picrosia). The seven clades with major support and their component genera are: 1) the Lygodesmia Clade: Chaetadelpha, Lygodesmia, and Shinnersoseris; 2) the Pinaropappus Clade: Marshalljohnstonia and Pinaropappus; 3) the Pyrrhopappus Clade: Picrosia and Pyrrhopappus; 4) the Microseris Clade: Agoseris, Microseris, Nothocalais, Stebbinsoseris, and Uropappus; 5) the Stephanomeria Clade: Munzothamnus,...

A phylogenetic analysis of Primulaceae s.l. based on internal transcribed spacer (ITS) DNA sequence data

Phylogenetic relationships in Primulaceae were investigated by analysis of nuclear rDNA ITS sequences. Thirty-four species of Primulaceae, two of Myrsinaceae and four outgroup taxa were analyzed. In accordance to the results of recently published papers on the phylogeny of Primulaceae we found the family to be paraphyletic and resolved the positions of some genera. Our results show (a) the rather basal position of Centunculus within Lysimachieae, the genus thus being rather distantly related to Anagallis, (b) the close relationship between Lysimachia sect. Lerouxia, Anagallis, Asterolinon, and Pelletiera, (c) the well-supported monophyly of a group consisting of the four genera Hottonia, Omphalogramma, Bryocarpum, and Soldanella, and (d) the affinity of Stimpsonia to the Myrsinaceae-Lysimachieae-Ardisiandra clade. The ITS sequence data do not provide sufficient information to resolve basal relationships within the Primulaceae s.l. There is evidence against the monophyly of the large genera Primula, Androsace, and Lysimachia. In contrast to the phylogenetic reconstructions based on plastid gene sequences, Cyclamen does not appear as a member of the Myrsinaceae-Lysimachieae clade, but its position remains unclear.

Phylogeny of Stephanomeria and related genera (compositae–lactuceae) based on analysis of 18S–26S nuclear rDNA ITS and ETS sequences

A phylogenetic analysis of DNA sequences from the internal transcribed spacer (ITS), the external transcribed spacer (ETS), and the 5.8S regions of 18S-26S nuclear rDNA from all diploid species of Stephanomeria and related genera shows that Stephanomeria does not include either Munzothamnus blairii (previously S. blairii) or Pleiacanthus spinosus (previously S. spinosa). Without these two taxa, Stephanomeria is a well-supported (100% bootstrap), monophyletic group of ten perennial and six annual species. Munzothamnus blairii and Pleiacanthus spinosus, both now considered members of monotypic genera, had been placed in Stephanomeria primarily because they have the same chromosome number as Stephanomeria and similar pollen surface features, but many disparities were ignored in previous classifications. Within Stephanomeria, an unsuspected sister relationship was detected between the montane S. lactucina and coastal S. cichoriacea. A second clade contained all the annual taxa and five of the perennial species. Among the annuals, strong bootstrap support was obtained for the previously recognized relationships between S. diegensis and S. exigua (98%) and between S. malheurensis and its progenitor, S. exigua subsp. coronaria (96%). Among the five perennial species that constitute a clade with the annuals, the recently described S. fluminea was shown to be sister to S. runcinata, and both of them were closely allied to S. tenuifolia and S. thurberi. The clade including the annuals (and five of the perennial species) was subtended by perennial lineages and pairwise divergence values among the annual taxa were much lower than among the perennial taxa as a group (though not too different than among the perennials in the same clade). The annuals probably originated recently within the genus.

Molecular phylogenetic relationships amongst Alternaria species and related fungi based upon analysis of nuclear ITS and mt SSU rDNA sequences

To elucidate relationships among Alternaria, Ulocladium, and Stemphylium species, nuclear internal transcribed spacer (ITS) and mitochondrial small subunit (SSU) ribosomal DNA (rDNA) sequences from 18 Alternaria, four Ulocladium and four Stemphylium spp. were determined and compared. Phylogenetic analysis of the ITS and SSU rDNA sequences, performed by the neighbour joining and maximum parsimony methods, revealed that the Stemphylium spp. were phylogenetically distinct from the Alternaria and Ulocladium spp. Most Alternaria spp. and the Ulocladium spp. were placed together in a large Alternaria/Ulocladium clade. Within this large clade, the Alternaria spp. clustered into several distinct species-clades, most of which correlated with species-groups previously established based upon morphological characteristics. The Ulocladium spp. were placed into two species-clades, each of which also included Alternaria spp. A. longissima was distantly related to the other Alternaria spp., as well as the Ulocladium and Stemphylium spp. Based upon ITS and nuclear 18S rDNA sequence identities, A. longissima was most closely related to Leptosphaeria .

Molecular evidence for allopolyploid speciation and a single origin of the narrow endemic Draba ladina (Brassicaceae)

Draba ladina (Brassicaceae) is a small alpine flower endemic to the Swiss Alps. It occurs exclusively at elevations between 2600 and 3000 m and is restricted to less than a dozen mountains in the Lower Engadin. Morphological characters and polyploidy suggest a hybrid origin. Potential diploid progenitor species are distributed widely and often occur sympatrically. To study the evolutionary history of D. ladina we assessed intra- and interspecific sequence variation at noncoding chloroplast DNA loci and nuclear rDNA ITS sequences in D. ladina and its presumed progenitor species D. aizoides, D. dubia, and D. tomentosa. A single ITS (Internal Transcribed Spacer) genotype was found in each of D. aizoides and D. dubia and two in D. tomentosa. Additivity of ITS sequences of D. aizoides and D. tomentosa was found in D. ladina, supporting the hypothesis of an allotetraploid origin. Intraspecific cpDNA variation was found in all diploid species, but not in D. ladina. The single chloroplast DNA haplotype found in the latter was closest to one cpDNA haplotype found in D. tomentosa, suggesting that D. tomentosa was the maternal parent. These results suggest that D. ladina is a relatively young, presumably postglacial, taxon with a single allopolyploid origin.

Inter- and intraspecific genetic variation in Caulerpa (Chlorophyta) based on nuclear rDNA ITS sequences

Caulerpa (Chlorophyta, Ulvophyceae) is a common marine tropical-subtropical genus of about 70 species, inhabiting the eulittoral zone on rocks and corals as well as mangroves. The genus is particularly diverse and abundant along southern Australian coasts, where it is thought to have originated. Here, we compare sequences from the nuclear ribosomal cistron among five species of Caulerpa, including nine populations of Caulerpa filiformis from two biogeographic regions : five from Australia and four from South Africa. Species relationships were well resolved by internal transcribed spacer (ITS) sequences and supported by high bootstrap values as follows: (C. geminata (C. simpliciuscula (C. trifaria (C. scalpelliformis (C. filiformis – Australia, C. filiformis – South Africa ))))). Nucleotide divergence within C. filiformis was low with four and five nucleotide differences present in ITS1 and ITS2 respectively. Unexpectedly, the 5.8S rRNA gene showed eight nucleotide differences between the Australian and South African populations and may indicate cryptic species. The Australian C. filiformis is probably not a South African introduction. These sequences provide an independent means for assessment of species relationships and appear, in so far as limited taxon sampling permits, to follow chloroplast ultrastructural groupings proposed 20 years ago.

Multiple trans-Arctic passages in the red alga Phycodrys rubens: evidence from nuclear rDNA ITS sequences

In order to investigate how episodes of geological and climatic change have influenced the distribution and evolutionary diversification of Arctic to cold temperate-North Atlantic seaweed species, intraspecific genetic variation was analyzed among isolates of the sublittoral, benthic red alga Phycodrys rubens (collected between June 1992 and January 1994). Rooted phylogenetic analyses of nuclear ribosomal DNA internal transcribed spacer (ITS) sequences and the plastid encoded Rubisco spacer sequences suggest that P. rubens invaded the North Atlantic from the Pacific shortly after the opening of the Bering Strait (3 to 3.5 million years ago), colonizing both the western and eastern Atlantic coasts. Based on these data we further hypothesize that P. rubens survived along the European coasts during the more recent Pleistocene glaciations, while becoming locally extinct along the North American Atlantic coasts. Following retraction of the last ice sheet, the western Atlantic coast was colonized a second time from the Pacific. The presence of two distinct genetic types (based on ITS and Rubisco sequences) along the European coasts is postulated to be a result of isolation and subsequent differentiation. This is likely because ice-free areas are known to have existed in northern Scotland and Norway during the last glaciation. The presence of an East Atlantic genetic type along the West Atlantic coast is believed to be a recent introduction (caused by human activity) of P. rubens to Newfoundland.