Phylogenetic implications of the internal transcribed spacers of nrDNA and chloroplast DNA fragments of <Emphasis Type="Italic">Musa</Emphasis> in deciphering the ambiguities related to the sectional classification of the genus

The genus Musa L. comprises of economically important bananas and has been divided into five sections based on their chromosome number and morphological characteristics viz., Australimusa, Callimusa, Eumusa, Rhodochlamys and Ingentimusa. However, this sectional classification has long been disputed. In this study, we present data on sequenced multiple DNA fragments from the maternally inherited chloroplast genome (atpB-rbcL spacer, trnK-matK intron and psbK-psbI spacer) and the biparentally inherited internal transcribed spacers of nuclear ribosomal DNA of three species of Musa along with data available in GenBank database to understand their implications on the sectional relationship and phylogeny of the genus Musa. In our findings, none of the five sections of Musa previously defined based on morphology was recovered in the molecular phylogeny analysis using cpDNA and nrDNA. Instead, the results corroborate with the suggestion that sectional classification in the genus Musa should be reviewed by combining together the section Eumusa and Rhodochlamys as one and section Australimusa and Callimusa as a single different section, along with that of Ingentimusa.     

Retrotransposon-based sequence-specific amplified polymorphism markers for the analysis of genetic diversity and phylogeny in <Emphasis Type="Italic">Malus</Emphasis> Mill. (Rosaceae)

Sequence-Specific Amplified Polymorphism (S-SAP)-analysis of 131 accessions of different species from five sections of the genus Malus was carried out to study genetic diversity, and to clarify phylogenetic and taxonomic issues. S-SAP-markers, based on long terminal repeat (LTR)-retrotransposons TRIM2 and dem1, were developed, which identified 679 polymorphic fragments in the studied Malus species. S-SAP technique proved to be effective for taxonomic studies in Malus. The obtained results generally support the existing sectional taxonomy in the genus Malus and allowed to determine the taxonomic status of some Russian landraces. The genetic diversity and taxonomic status of the Russian apple Antonovka landraces, widely used in breeding programs for their increased adaptation to abiotic stress and scab resistance, were clarified. All of them belong to M. domestica, section Malus. However, Antonovka Olginskaya might have a hybrid origin with some contribution from Gymnomeles species according to PCO analysis data. The taxonomic status was resolved for another Russian landrace, Yakutskaya, which exhibits increased winter hardiness and drought resistance; it belongs to section Gymnomeles, with a high resemblance to M. baccata.     

Actinomycetal complex of light sierozem on the Kopet-Dag piedmont plain

The population density of actinomycetes in the samples of light sierozem from the Kopet Dag piedmont plain (75 km from Ashkhabad, Turkmenistan) reaches hundreds of thousand CFU/g soil. The actinomycetal complex is represented by two genera: Streptomyces and Micromonospora. Representatives of the Streptomyces genus predominate and comprise 73 to 87% of the actinomycetal complex. In one sample, representatives of the Micromonospora genus predominated in the complex (75%). The Streptomyces genus in the studied soil samples is represented by the species from several sections and series: the species of section Helvolo-Flavus series Helvolus represent the dominant component of the streptomycetal complex; their portion is up to 77% of all isolated actinomycetes. The species of other sections and series are much less abundant. Thus, the percentage of the Cinereus Achromogenes section in the actinomycetal complex does not exceed 28%; representatives of the Albus section Albus series, Roseus section Lavendulae-Roseus series, and Imperfectus section belong to rare species; they have been isolated not from all the studied samples of light sierozem, and their portion does not exceed 10% of the actinomycetal complex.     

Psychrotolerant actinomycetes on plant substrates of tundra and taiga ecosystems

Studies of tundra flowering plants and mosses on the Тaymyr Peninsula have determined the presence of thousands and tens of thousands of colony formation units of psychrotolerant actinomycetes in 1 g of vegetation substrate, which is smaller than the amount of mesophilic forms by one to two orders of magnitude. Incubation of plants at 5°C has shown very small taxonomical variety of actinomycetes. The actinomycete complex is represented by species of the genus Streptomyces, belonging to the sections and series Albus Albus and Cinereus Achromogenes, and by the genus Micromonospora.     

Phylogenetic relationships within <Emphasis Type="Italic">Lactuca</Emphasis> L. (Asteraceae), including African species,based on chloroplast DNA sequence comparisons

Lettuce (Lactuca sativa L.) belongs to the genus Lactuca L. and is an important vegetable worldwide. Over the past decades, there have been many controversies about the phylogeny of Lactuca species due to their complex and diverse morphological characters and insufficient molecular sampling. In this study we provide the most extensive molecular phylogenetic reconstruction of Lactuca, including African wild species, using two chloroplast genes (ndhF and trnL-F). The sampling covers nearly 40 % of the total endemic African Lactuca species and 34 % of the total Lactuca species. DNA sequences from all the subfamilies of Asteraceae in Genebank and those generated from Lactuca herbarium samples were used to establish the affiliation of Lactuca within Asteracaeae. Based on the subfamily tree, we selected 33 ndhF sequences from 30 species and 79 trnL-F sequences from 48 species to infer relationships within the genus Lactuca using randomized axelerated maximum likelihood and Bayesian inference analyses. Biogeographical, chromosomal and morphological character states were reconstructed over the Bayesian tree topology. We conclude that Lactuca contains two distinct phylogenetic clades—the crop clade and the Pterocypsela clade. Other North American, Asian and widespread species either form smaller clades or mix with the Melanoseris species. The newly sampled African endemic species probably should be treated as a new genus.     

Phylogenetic position of the disjunct species <Emphasis Type="Italic">Musa ornata</Emphasis> (Musaceae): first approach to understand its distribution

Musa L. commonly known as the banana group is one of the most important and oldest food crops of humankind. Among the wild relatives with ornamental interest in the genus, Musa ornata Roxb. shows a disjunct distribution between Asia and North America (Mexico). The wild occurrence of this species in Mexico has led to speculation about the evolutionary relationships with its Asian relatives. This study examined the phylogenetic relationships between intercontinental specimens of this species and, based on registered evidence, explored the more likely hypothesis about the origins of its distribution. The phylogeny of intercontinental specimens, along with other representatives of the same genus, was carried out using three molecular markers (ITS, trnL-F, and atpB-rbcL) and applying three phylogenetic reconstruction methods: maximum parsimony, maximum likelihood, and Bayesian inference. The genetic analysis of the combined dataset grouped together all the Mexican and most Asian specimens, but the monophyly of the species was not supported. The relationships suggest that Mexican populations may have originated from an Asian invasion. However, several studies and historical documents suggest the presence of Musa in America long before the arrival of Europeans. Based on its current distribution, phylogenetic evidence, and fossil record, this species’ disjunct distribution could be explained in terms of an ancestral distribution range that encompassed America and Asia, followed by its subsequent restriction to the Old World and a secondary dispersal by humans. However, further studies are necessary to shed more light on the origins of this disjunct distribution.     

Discriminating power of microsatellites in cranberry organelles for taxonomic studies in <Emphasis Type="Italic">Vaccinium</Emphasis> and Ericaceae

Simple sequence repeats (SSRs) in chloroplast and mitochondrial DNA, which have not been previously developed in the Ericaceae or, more specifically, in the genus Vaccinium, can be powerful tools for determining evolutionary relationships among taxa. In this study, 30 chloroplast, 23 mitochondrial, and 1 mitochondrion-like SSRs were identified in cranberry (V. macrocarpon), and primer-pairs were developed and tested for each locus. Although no polymorphisms were detected for any of the 54 SSR loci in nine diverse cranberry genotypes, all primers were cross-transferable to some extent to a panel of 12 additional Vaccinium taxa and four non-Vaccinium Ericaceae species. A Neighbor-Joining tree of the estimated average squared distances resolved the species by genus and by section within Vaccinium. Similar topologies with increased branch support were observed in Bayesian inference trees constructed from the DNA sequences of six plastid and two mitochondrial SSR loci. Two multiplexing/poolplexing panels of M13 fluorescently labeled primers, which amplify 24 of the 54 markers, were developed and can serve as an efficient, cost-effective means for characterizing the basic molecular phylogeny of Vaccinium. Increased understanding of evolutionary relationships among Vaccinium species should facilitate interspecific hybridization and introgression efforts to improve economically important traits of commercial berry crops.     

Maiden hair ferns (<Emphasis Type="Italic">Adiantum</Emphasis> L., Pteridaceae–Vittarioideae) of North East India: diversity,phytochemistry and utilization

The genus Adiantum L. (Pteridaceae–Vittarioideae) are medicinal ferns with around 200 species and distributed in temperate and tropical regions. Several species of the genus are used in traditional medicine in different parts of the world including the Chinese Herbal Medicine, United States Pharmacopoeia and Ayurveda System of Medicine and Homeopathy. In the present paper, an updated and comprehensive data on species diversity of Adiantum found in the seven (7) states of North East India will be discussed. Altogether, nine (9) species are recorded and this includes A. capillus–veneris L. with two (2) races, A. caudatum L., A. edgeworthii Hook., A. flabellulatum L., A. hispidulum Sw., A. incisum Forssk., A. peruvianum Klotzsch, A. philippense L. with three (3) subspecies and A. raddianum C. Presl. Brief morphological parameters, utilization including medicinal, economical and traditional aspects, along with their bioactive components are described. The paper also aims to provide for the first time an updated compilation of species information including correct nomenclature, synonyms, distribution and propagation.     

Phylogenetic relationship of 40 species of genus Aloe L. and the origin of an allodiploid species revealed by nucleotide sequence variation in chloroplast intergenic space and cytogenetic insitu hybridization

Aloe species, which have been used as medicinal plants, belong to the Asphodelaceae family consisting of 530 species. In this study, genetic diversity and phylogenetic relationships among 40 Aloe species including a putative interspecies hybrid were analyzed using PCR band profiles from eight chloroplast intergenic space markers and nucleotide sequence diversity in the psbK–psbI intergenic region. A phylogenetic tree based on psbK–psbI sequences supported the revised classification of the genus Aloe as polyphyletic with several species be re-allocated into three genera Kumara, Aloidendron, and Aloiampelos. Further, the origin of the putative interspecies Aloe hybrid was characterized through molecular cytogenetics. Fluorescence and genomic insitu hybridization illustrated that the hybrid has a bimodal karyotype with a chromosome complement of 2n = 14, of which complementary halves were derived from two parental species, A. vera and A. arborescens. These findings revealed that the hybrid species was allodiploid. The phylogenetic analysis showed that A. arborescens was the maternal genome donor of the hybrid, as both have identical chloroplast genome sequences. We thus conclude that the allodiploid hybrid should be called A. arborescens × A.vera.     

Algological and Mycological Characterization of Soils under Pine and Birch Forests in the Pasvik Reserve

The structure of algological and mycological complexes in Al–Fe-humus podzols (Albic Podzols) under pine and birch forests of the Pasvik Reserve is characterized. The number of micromycetes is higher in more acid soils of the pine forest, while the species diversity is greater under the birch forest. The genus Penicillium includes the largest number of species. The greatest abundance and occurrence frequency are typical for Penicillium spinulosum, P. glabrum, and Trichoderma viride in pine forest and for Umbelopsis isabellina, Mucor sp., Mortierella alpinа, P. glabrum, Aspergillus ustus, Trichoderma viride, and T. koningii in birch forest. Cyanobacteria–algal cenoses of the investigated soils are predominated by green algae. Soils under birch forest are distinguished by a greater diversity of algal groups due to the presence of diatoms and xanthophytes. Species of frequent occurrence are represented by Pseudococcomyxa simplex and Parietochloris alveolaris in soils of the pine forest and by Tetracystis cf. aplanospora, Halochlorella rubescens, Pseudococcomyxa simplex, Fottea stichococcoides, Klebsormidium flaccidum, Hantzschia amphioxys, Microcoleus vaginatus, and Aphanocapsa sp. in soils under birch forest     

Remarks on genetic diversity and relationship of <Emphasis Type="Italic">Punica protopunica</Emphasis> and <Emphasis Type="Italic">P. granatum</Emphasis> assessed by molecular analyses

Here, two Punica species, viz., P. protopunica Balf. fil., reported as native to Socotra, and P. granatum L., were compared for the first time. Analysis of one P. protopunica and eleven P. granatum accessions was performed using three molecular markers, i.e., sequence related amplified polymorphism (SRAP), target region amplification polymorphism (TRAP), and intron targeted amplified polymorphism (ITAP), along with analysis of pgWD40 sequences, a gene involved in anthocyanin biosynthesis. All markers revealed the relationship between the two species and placed them at 33% similarity. SRAP, TRAP, and ITAP generated a total of 299, 260, and 160 bands, respectively. Of these, 78, 74, and 41 bands were specific for P. protopunica, and 92, 85, and 57 bands, respectively, were shared between both species. Sequence analysis of pgWD40~870 bp amplicons exhibited 100% identity among P. granatum accessions and 98% identity to that of P. protopunica. Phylogenetic analysis of WD40 sequences from monocot and dicot species, including both Punica species confirmed the relation between P. protopunica and P. granatum, supporting earlier reports that P. protopunica could be an ancestral species of P. granatum. Furthermore, the genetic diversity among and within P. granatum accessions from Egypt (3), Mexico (5), and Yemen (3) was assessed. Molecular marker-based relationships among region-bulked accessions was approximately the same (~90% similarity), whereas the degree of genetic variation was altered within each region. Specific bands (alleles) for accessions of each region along with those shared among them were identified. Thus, these bands could be used for pomegranate genotyping and breeding programs.     

Diversity of <Emphasis Type="Italic">Citrus</Emphasis><Emphasis Type="Italic">depressa</Emphasis> Hayata (Shiikuwasha) revealed by DNA analysis

Citrus depressa Hayata is an indigenous mandarin species on the Ryukyu Islands located in the subtropical region of Japan. We deduced its phylogenetic relationships by evaluating accessions grown on various Ryukyu Islands via cleaved amplified polymorphic sequence analysis of cpDNA and sequence-related amplified polymorphism (SRAP). The cpDNA results indicated that C. depressa could be classified into two types. SRAP revealed patterns of diversity within C. depressa consistent with our cpDNA results. These results indicate that maternal origin may influence or is correlated with the constitution of the nuclear genome of C. depressa. Another Japanese mandarin species, Citrus tachibana (Makino) Tanaka was distinguished from C. depressa by SRAP markers. Moreover, both C. depressa and C. tachibana could be distinguished from other Citrus species. Our results suggest that Japanese mandarin possesses a characteristic genome with the genus Citrus.     

Combining conservative and variable markers to infer the evolutionary history of Prunus subgen. Amygdalus s.l. under domestication

The genus Prunus L. is large and economically important. However, phylogenetic relationships within Prunus at low taxonomic level, particularly in the subgenus Amygdalus L. s.l., remain poorly investigated. This paper attempts to document the evolutionary history of Amygdalus s.l. and establishes a temporal framework, by assembling molecular data from conservative and variable molecular markers. The nuclear s6pdh gene in combination with the plastid trnSG spacer are analyzed with bayesian and maximum likelihood methods. Since previous phylogenetic analysis with these markers lacked resolution, we additionally analyzed 13 nuclear SSR loci with the δµ² distance, followed by an unweighted pair group method using arithmetic averages algorithm. Our phylogenetic analysis with both sequence and SSR loci confirms the split between sections Amygdalus and Persica, comprising almonds and peaches, respectively. This result is in agreement with biogeographic data showing that each of the two sections is naturally distributed on each side of the Central Asian Massif chain. Using coalescent based estimations, divergence times between the two sections strongly varied when considering sequence data only or combined with SSR. The sequence-only based estimate (5 million years ago) was congruent with the Central Asian Massif orogeny and subsequent climate change. Given the low level of differentiation within the two sections using both marker types, the utility of combining microsatellites and data sequences to address phylogenetic relationships at low taxonomic level within Amygdalus is discussed. The recent evolutionary histories of almond and peach are discussed in view of the domestication processes that arose in these two phenotypically-diverging gene pools: almonds and peaches were domesticated from the Amygdalus s.s. and Persica sections, respectively. Such economically important crops may serve as good model to study divergent domestication process in close genetic pool.     

Relationships among <Emphasis Type="Italic">Crataegus</Emphasis> accessions sampled from Hatay,Turkey, as assessed by fruit characteristics and RAPD

The genus Crataegus known as hawthorns, is the largest genus among the Maloideae, which comprises 265 species. Turkey is one of the genetic centers of Crataegus and there are more than 20 species found in Turkey. The fruits of Crataegus are used as food and have high flavonoid, vitamin C, glycoside, anthocyanidin, saponin, tannin, and antioxidant levels. In this study, we attempted to characterize 15 Crataegus accessions sampled from Hatay, located in Eastern Mediterranean region of Turkey. The accessions belonged to several species; C. aronia (L.) DC. var. aronia, C. aronia var. dentata Browicz, C. aronia var. minuta Browicz, C. monogyna Jacq. subsp. azarella (Griseb.) Franco, and C. orientalis Pall. ex M. Bieb. var. orientalis. Fruit characteristics of the accessions exhibited considerable variation. The multivariate, principle component and cluster analyses indicated that the accessions belonged to three groups: (1) C. aronia var. arona accessions; (2) C. aronia var. dentata accessions; and, (3) C. monogyna subsp. azarella and C. orientalis var. orientalis accessions. The principle component analysis results also revealed that the first three components explained 46, 21, and 14% of the variation, comprising a total of 81%. The fruit length and width, leaf area, and soluble solids contents were highly correlated characteristics for the first three components. The 19 RAPD primers generated a total of 107 bands, where 76 of these were polymorphic. The molecular data analyses by principle coordinate and clustering showed similar results to those of pomological characteristics. There were three groups, (1) C. aronia var. arona accessions; (2) C. aronia var. dentata accession; and, (3) C. monogyna subsp. azarella. C. orientalis var. orientalis accession grouped with C. aronia var. arona accessions. Therefore, it can be concluded that, overall, the diversity patterns of pomological and molecular data, generated by RAPD, for Crataegus are in good agreement and the accessions of C. aronia var. aronia, C. aronia var. minuta, C. monogyna subsp. azarella and C. orientalis var. orientalis accessions.     

Complete chloroplast DNA sequences of Georgian indigenous polyploid wheats (<Emphasis Type="Italic">Triticum</Emphasis> spp.) and B plasmon evolution

Three types of plasmon (A, B and G) are present for genus Triticum. Plasmon B is detected in polyploid species - Triticum turgidum L. and Triticum aestivum L. By now, 21 complete sequences of chloroplast DNA of the genus Triticum is published by different authors. Many inaccuracies can be detected in the sequenced chloroplast DNAs. Therefore, we found it necessary to study of plasmon B evolution to use only those sequences obtained by our method in our laboratory. Complete nucleotide sequences of chloroplast DNA of 11 representatives of Georgian wheat polyploid species were determined. Chloroplast DNA sequencing was performed on an Illumina MiSeq platform. Chloroplast DNA molecules were assembled using the SOAPdenovo computer program. Using T. aestivum L. subsp. macha var. palaeocolchicum as a reference, 5 SNPs were identified in chloroplast DNA of Georgian indigenous polyploid wheats. 38 and 56 bp inversions were observed in paleocolchicum subspecies. The phylogeny tree shows that subspecies macha, durum, carthlicum and palaeocolchicum occupy different positions. According the simplified scheme based on SNP and indel data the ancestral, female parent of all studied polyploid wheats is an unknown X predecesor, from which four lines were formed.     

Screening species of <Emphasis Type="Italic">Pilocarpus</Emphasis> (Rutaceae) as sources of pilocarpine and other imidazole alkaloids

Plants of the Pilocarpus genus (Rutaceae) are popularly known as jaborandi and are the only source of pilocarpine, an imidazole alkaloid used in eye-drops for the treatment of glaucoma as well as for the stimulation of sweat and lachrymal glands. Alkaloid extracts from leaf samples of seven species of Pilocarpus, from the states of São Paulo and Maranhão in Brazil, were analyzed using HPLC–ESI–MS/MS. The samples contained between 0.88 ± 0.04 and 1.00 ± 0.14% of alkaloids in relation to the dry weight of their leaves, with significant differences in results (P ≤ 0.05) found only between Pilocarpus microphyllus planted in the state of Maranhão and Pilocarpus spicatus, Pilocarpus trachyllophus, Pilocarpus pennatifolius and Pilocarpus jaborandi; as well as between Pilocarpus spicatus and Pilocarpus racemosus. Pilocarpine was not found in P. spicatus, whereas in the other species it ranged from 2.6 ± 0.1 to 70.8 ± 1.2% of total alkaloids. P. microphyllus planted in the state of Maranhão for pilocarpine extraction had the highest total alkaloid content, but it had only 35% of pilocarpine in relation to total alkaloids. Three other species contained more pilocarpine in relation to total alkaloids: P. jaborandi (70.8%), P. racemosus (45.6%) and P. trachyllophus (38.7%); and could be candidates for pilocarpine extraction. Differences in alkaloid content were significant for all these samples (P ≤ 0.05). Imidazole alkaloids were observed and partially characterized based on their retention times and high resolution mass. The seven species analyzed had different imidazole alkaloid profiles, but only one did not present quantifiable pilocarpine contents in its leaves. The Pilocarpus genus shows potential for the prospection of novel alkaloids.     

Electrophoretic variation in seed proteins and interrelationships of species in the genus <Emphasis Type="Italic">Oryza</Emphasis>

Comparative studies of two cultivated and sixteen wild species of the genus Oryza were carried out using one- and two-dimensional gel electrophoresis for variation in their seed proteins for interrelationships of these species. A number of polypeptides in the range of molecular weight 13–110 kDa were seen to vary. Under reducing conditions, polypeptides spread over the regions of mol. wt. 33–40.5, 25–27 and 19–21.5 kDa exhibited maximum variation in their patterns. Two-dimensional gel electrophoresis revealed the occurrence of disulphide-linked glutelin polypeptide pairs of mol. wt. 60, 58, 52 and 25 kDa breaking into a large and a small subunit each in the range of mol. wt. 18–40.5 and 16–25 kDa respectively in Oryza sativa. The number of such polypeptide pairs varied from 2 to 6 in different species and also in O. sativa showed variation in mol. wts. of their constituent subunits. The UPGMA dendrogram revealed that most of the Oryza species occurred in different clusters and subclusters and thus did not share very close relationships. The undisputed and closest relationship observed was that of cultivated rice O. sativa with the O. rufipogon followed by that with O. nivara. The African cultivated O. glaberrima occurring on the nearest branch of the same subcluster, thereby, supporting the phylogenetic of these species suggested in earlier studies. Eight diploid species and seven tetraploid species were included in one part of the dendrogram while the remaining two species with AA genome i.e. O. glumaepatula and O. meridionalis and one with FF i.e. O. brachyantha stood separately from these as scattered in the group of seven tetraploid species with BBCC, CCDD and HHJJ genomes. The tetraploids O. alta, O. latifolia and O. grandiglumis with CCDD genomes which occurred on the farthest part were distantly related with O. sativa. The cyanogen bromide peptide maps and two dimensional gel electrophoresis also supported the closest relationship between O. sativa and O. rufipogon.     

Genotyping-by-sequencing reveals the origin of the Tunisian relatives of cultivated carrot (<Emphasis Type="Italic">Daucus carota</Emphasis>)

Tunisia, other countries in northwestern Africa, and the Iberian Peninsula, represent a center of diversity of Daucus. The genus traditionally has included about 20–25 species worldwide, but a recent molecular study redefined and expanded Daucus to include representatives from nine other genera. By this classification, Daucus now contains about 40 species, with some of them having winged fruits in addition to its traditionally recognized spiny fruits. The taxonomy of Daucus in Tunisia has recently been studied with morphological data, concluding that D. carota subsp. capillifolius is a subspecies that co-occurs and crosses with subsp. carota. The present study extends these findings with additional data from Genotyping-by-Sequencing (GBS) from 33 Tunisian accessions of D. carota (93 individuals with two to three replicates per accession), including the first collections of subsp. gummifer from Tunisia. Placed in the context of additional GBS data, D. carota subsp. gummifer has separate origins from other collections of subsp. carota in Tunisia and/or immediately adjacent areas to the north in Italy or its surrounding islands. Our results add support to the utility of large SNP datasets for species-level phylogenetic studies in Daucus.     

SCAR markers that discriminate between <Emphasis Type="Italic">Dasypyrum</Emphasis> species and cytotypes

Dasypyrum is a genus with two species, an annual diploid Dasypyrum villosum and a perennial diploid or tetraploid D. breviaristatum. It is difficult to distinguish between the two, especially for a non-taxonomist. In this paper we demonstrate how to distinguish between the two species and between the two ploidy levels of D. breviaristatum. We achieve this using RAPD analysis to identify potential candidates for marker development and after carrying out principal component analyses and partitioning of genetic variance, developed SCARs and an identification key.