ITS Markers Research Articles

DNA Barcoding of Two Medicinal Plants Using Molecular Markers

DNA barcoding is a novel method of species identification based on nucleotide diversity of conserved sequences. The establishment and refining of plant DNA barcoding systems are more challenging due to the high genetic diversity among different species. We, therefore, tested the potentiality of the ITS and rbcL markers for the identification of two medicinal species plants Solanum nigrum and Solanum villosum, which were collected from the Botanical garden of faculty of Science, Ain Shams Univ., Egypt. In this study, biological sequence homology and divergence of amplified sequences were studied using the Basic Local Alignment Tool (BLAST). Both DNA barcoding regions (ITS and rbcL) showed good universality amplification in the two species. The sequenced regions revealed conserved genome information for future identification of different medicinal plants belonging to these species. The amplified conserved barcodes revealed different levels of biological homology after sequence analysis. The results clearly showed that the use of these conserved DNA sequences as barcode primers would be an accurate way for species identification and discrimination. In this work, the ITS and rbcl markers were used to discriminate and confirm the identification of two medicinal plants, it was found that the viability and potentiality of ITS region in the identification process for the two plants used is more efficiency than rbcl, where rbcl confirm the identification of two plants at the generic level, while ITS at the species level. The findings of the study would be applicable in the medicinal industry to establish DNA based identification of different medicinal plant species to monitor adulteration.

The transferability of microsatellite loci from a homoploid to a polyploid hybrid complex: an example from fine-leaved Festuca species (Poaceae).

BackgroundMicrosatellite loci, or single sequence repeats (SSR), are widely used as powerful markers in population genetics. They represent an attractive tool for studying plants such as grasses, whose evolution is driven by hybridisation and polyploidization. However, the development of microsatellite markers has been challenging and time-consuming, especially for non-model organisms lacking available genome-wide sequence data. One straightforward and low-cost approach is to transfer the SSR loci developed for one species, or complex, to another closely-related one. This work evaluates the transferability of microsatellite loci from homoploid to allopolyploid complexes of fine-leaved Festuca species and to assess their use in two new species. The studied complex (F. amethystina—F. tatrae) is a useful model for research on the local adaptability of grasses with different ploidy levels. Since both species can be considered as rare or threatened (F. tatrae—as a mountain and narrow endemic species and F. amethystina—a mountain species with relict lowland populations), any tool enabling studies on genetic diversity and population genetics, such as SSR markers, could also be very useful in a conservation context.MethodsThe ploidy level within populations was estimated using flow cytometry. One diploid and one tetraploid population of F. amethystina and a diploid population of F. tatrae were chosen to test the transferability of SSR loci. Because our work describes the transfer of SSR nuclear markers designed originally for F. gautieri, a phylogenetic tree was prepared based on the ITS marker to assess the genetic distance between the studied complexes. The PCR products were separated on a high-resolution agarose gel, intended for SSR marker analysis. Appropriate solutions for the allotetraploid population and whole mixed-ploidy complex were implemented.ResultsFlow cytometry confirmed earlier data regarding DNA content in the investigated species and cytotypes. The phylogenetic ITS tree indicated a small genetic distance between F. gautieri complexes and the studied species. Ten microsatellite markers were successfully transferred. All markers were polymorphic. In total, 163 different alleles were scored from the 10 SSR loci. PCoA of accessions revealed well-separated groups corresponding to studied populations. Over 60% of the total variance is explained by differentiation within populations and one third among them.ConclusionsThe transferred markers are valid tools for the study of population genetics and inheritance relationships within cytotypes and species and between them. The presented markers can be used to study inbreeding depression in the Festuca species, and variations in the degrees of genetic diversity between different cytotypes in mountain and lowland areas. Our findings can also be applied to study conservation strategies for ensuring biodiversity at the genetic level in polyploid complexes.

Morphological variation, species delimits, and phylogenetic relationships of four important Ipomoea species in Sri Lanka

Genus Ipomoea is the largest of the family Convolvulaceae. It is a highly diverse genus distributed all around the world. In Sri Lanka, 13 members have been identified in Ipomoea; however, most of them have not been morpho-genetically characterized. In the present study, we selected I. batatas, I. aquatica, I. nil, and I. pes-caprae for the assessment based on certain criteria viz., food security, leafy vegetable, emergent weed, and coastal conservation capabilities. The morphological variation, species delimits, and phylogenetic relationships among these four species were assessed. The DNA was extracted from young leaf samples using CTAB method and subjected to PCR using ITS and rbcL as markers. The PCR products were subjected to sequencing. The DNA sequences were analyzed, and phylogenetic trees were generated by using PAUP to define the species delimits, identify the speciation and evolutionarily relationships in comparison to the homologous Ipomoea sequences available in GenBank. The tuber bearing ability, flower colour, and leaf shape are the key morphological descriptors that can be used to delimit the selected species. Both rbcL and ITS markers could be used to identify the four species studied distinctively. However, among the two markers, ITS provides a greater variation to identify the four species and their evolutionary relationships. Thus, the present study provides a key platform with usable morphological traits and DNA barcoding markers to characterize all Ipomoea species in Sri Lanka.

Effects of Past and Present-Day Landscape Structure on Forest Soil Microorganisms

Theory surrounding landscape ecology have been built on the species distribution of birds and plants, but increasing evidence now exists for below-ground organisms, whose dispersal may also be affected by above-ground landscape structures. Uncertainties remain for how communities of microorganisms respond to landscape structure over time, and whether some groups of microorganisms react more than others. Here, we investigated if fungal or bacterial diversity is driven by the amount of forest cover in the current or the past landscape. We tested the Habitat Amount Hypothesis (HAH) on ancient forests of the Cevennes national park, that experienced increased fragmentation 150 years ago, and are today surrounded by recent forests. As ancient forests are often more diverse in plant species, we hypothesized that the higher quantity of ancient forests in the landscape, the richer local fungal and bacterial communities would be. More precisely, we expected that ectomycorrhizal fungi, and pathotrophic fungi, often indicators of mature forests, would be also more sensitive to forest history and therefore to the quantity of ancient forests than bacteria and saprotrophic fungi. We sampled 40 soil cores per 0.5 ha, pooled in 8 composite samples per plot in 27 landscapes and sequenced ITS and 16S markers by Illumina-Mi seq. To identify functional groups of fungi, we relied on their taxonomy and the use of public databases. Our results partly follow the HAH, as fungal richness was positively related with the quantity of ancient forests in the landscape and not by the focal patch size. Ectomycorrhizal and pathotrophic fungi were positively affected by the ancient forest cover, and so were saprotrophic ones, but not bacteria. Local factors also shaped the communities such as soil composition and elevation, confirming classical patterns in soil microbial ecology. Interestingly, past landscape structure was better at explaining fungal community richness than contemporary landscape, suggesting a time lag in the response of communities to landscape modification and a potential extinction debt. Our results reveal the importance of below-ground communities in studies of landscape and historical ecology, with their structure and functions likely to be intimately linked with soil and landscape history.

<p><strong>Phylogenetic relationships of <em>Silene</em> sections <em>Lasiostemones</em> and <em>Sclerocalycine</em> (Caryophyllaceae) in Iran</strong></p>

A phylogenetic study of Silene sect. Lasiostemones and sect. Sclerocalycine in Iran is presented. A total of 72 accessions (43 of which obtained from GenBank) were analyzed using nuclear ITS and chloroplast rps16 markers. It was shown that sect. Lasiostemones is not a monophyletic group, including members which would belong to sect. Sclerocalycine (i.e. S. tenella, S. longipetala, S. lasiantha, S. marschallii, S. propinqua) and sect. Siphonomorpha (i.e. S. ghahremaninejadii, S. shahrudensis, S. saxatilis, S. ruprechtii, S. parrowiana).

Fungal isolates from marine sponge Chelonaplysilla sp.: Diversity, antimicrobial and cytotoxic activities

Abstract. Handayani D, Artasasta MA, Safira N, Ayuni DF, Tallei TE, Hertiani T. 2020. Fungal isolates from marine sponge Chelonaplysilla sp.: Diversity, antimicrobial and cytotoxic activities. Biodiversitas 21: 1954-1960. The purpose of this research was to study the diversity of fungi associated with marine sponges Chelonaplysilla sp. and their bioactivities. Fungal isolation was carried out by the multilevel dilution method in Saboraud Dextrose Agar (SDA). Twelve fungal isolates were successfully purified, then cultivated using rice for 4-6 weeks at room temperature and subsequently extracted using ethyl acetate. Antimicrobial activities of the fungal extracts were tested against Staphylococcus aureus, Escherichia coli, and Candida albicans by using the agar diffusion method. The extracts of isolates Ch05 and Ch12 showed a significant antagonistic effect against S. aureus and E. coli with the diameter that ranged from 15 to 17 mm. Using the brine shrimp lethality test (BSLT), six fungal extracts revealed cytotoxic activity with LC50 <100 µg/mL. Isolate Ch10 was the most potential fungus with the strong cytotoxic activity of LC50 of 0.90 µg/mL. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was conducted also for six potential fungal extracts against breast cancer cell (T47D). The isolate Ch05 showed moderate cytotoxic activity with IC50 of 83.69 µg/mL. The molecular identification was carried out for potential fungi using the ITS marker. The results showed that Ch02 was Aspergillus oryzae, Ch05 was Phomompsis sp., Ch06 was Penicillium simplicissimum, Ch10 was B. bassiana and Ch12 was Aspergillus mellinus. This study concluded that fungal isolates from marine sponge Chelonaplysilla sp. can be explored further for new sources of antimicrobial and anticancer compounds.

The new genus Jocatoa (Lecanoromycetes: Graphidaceae) and new insights into subfamily Redonographoideae

One new genus and two new species of Graphidaceae are described from tropical dry forests of Mexico, based on morphological and molecular data of the mtSSU, nuLSU and ITS markers. The new genus Jocatoa in subfamily Graphidoideae is described to accommodate the species Graphis agminalis (syn.: Medusulina texana), previously known from Colombia and North America. The new genus resembles Diorygma but differs by having simple paraphyses tips that do not form an epithecium. Jocatoa agminalis is similar to Diorygma monophorum but differs by having larger ascospores, hypostictic and stictic acids and by the type of paraphysis tips. In subfamily Redonographoideae, the two new species Gymnographopsis corticicola and Redonographa parvispora are described, representing the first corticolous species in the subfamily. Gymnographopsis corticicola is characterized by the smallest spores in the genus, the presence of norstictic acid, and a rectangular perispore that appears to be a newly recognized character state in fungi. Redonographa parvispora is characterized by warty periphysoids, small ascospores with 3 transverse septa, and norstictic acid. It also frequently develops a rectangular perispore. We present a phylogenetic analysis, based on the markers mtSSU, nuLSU and RPB2, that includes all the genera in the family Graphidaceae, with available sequences, to accommodate the new genus and to validate for the first time the position of Gymnographopsis. Diagnostic anatomical and ecological characters are discussed for Redonographoideae. Gymnographopsis is newly reported to the Northern Hemisphere.

Enzymatic hydrolysis of barley straw for biofuel industry using a novel strain of Trametes villosa from Paranaense rainforest

Agricultural practices generate lignocellulosic waste that can be bioconverted by fungi to generate value-added products such as biofuels. In this context, fungal enzymes are presented as an alternative for their use in the hydrolysis of cellulose to sugars that can be fermented to ethanol. The aim of this work was to characterize LBM 033 strain and to analyze its efficiency in the hydrolysis of cellulosic substrates, including barley straw. LBM 033 strain was identified as Trametes villosa by molecular techniques, through the use of the ITS and rbp2 markers and the construction of phylogenetic trees. The cell-free supernatant of T. villosa LBM 033 showed high titers of hydrolytic enzymatic activities, necessary for the hydrolysis of the holocellulosic substrates, hydrolyzing pure cellulose to cellobiose and glucose and also degraded the polysaccharides contained in barley straw to short soluble oligosaccharides. These results indicate that macro fungi from tropical soil environments, such as T. villosa LBM 033 can be a valuable resource for in-house, cost effective production of enzymes that can be applied in the hydrolysis stage, which could reduce the total cost of bioethanol production.

Calcareous sponges from the French Polynesia (Porifera: Calcarea).

Although the French Polynesian reefs are among the most well studied reefs of the world, sponges are still poorly known, with only 199 species or OTUs of sponges having been described from French Polynesia, 167 at an OTU level and 32 at a species level. From those 199 species, just five are calcareous sponges. As it is possible that this number is underestimated, the aim of the present work was to study the diversity of calcareous sponges from French Polynesia. Hence, different French Polynesian archipelagos were surveyed by SCUBA from 3 to 60 m of depth. Identifications were performed using morphological and molecular (ITS and C-LSU) tools. We found a total of nine species of Calcarea, comprising five different genera. Five species are new to science: Clathrina fakaravae sp. nov., Clathrina huahineae sp. nov., Ernstia variabilis sp. nov., Leucascus digitiformis sp. nov., and Leucandra tahuatae sp. nov. With the present work, the number of identified sponges from French Polynesia at a species level increased from 32 to 41. The only calcareous sponge previously known from French Polynesia that was recollected by our group was Leucetta chagosensis. Our results suggest that the Eastern Indo-Pacific Realm shows more affinity with the Central and the Western Indo-Pacific Realms. Four species supported these affinities: Ascandra cf. crewsi, previously known only from Papua New Guinea, Leucascus simplex from South Australia, and Leucetta chagosensis and L. microraphis, both widespread species in the Indo-Pacific. These two Leucetta species, however, most likely represent species complexes. Once again the molecular markers ITS and C-LSU helped in the identification of calcareous sponges, showing how important is an integrative taxonomy. Although our work has increased in 250% (6 spp to 15 spp) the diversity of calcareous sponges in French Polynesia, it is most possible that this number is still underestimated.

Green-Spinach, Red-Spinach, and Tree-Spinach (‘Three-Fold Spinach’ in Sri Lanka): An Insight into Phylogenetics and Consumer Preference

Three types of spinach, green spinach (GS), red spinach (RS), and tree spinach (TS) are consumed in Sri Lanka. GS, RS, and TS are referred to as Basella alba, Basella rubra, and Talinum fruticosum respectively. However, some taxonomists categorize GS and RS under B. alba causing an ambiguity. Due to the poor sanitation, consumers prefer to purchase greenhouse-grown spinach over field grown material. However, the taste parameters of field grown and greenhouse-grown spinach have not been assessed. The objectives of the present study were to resolve the taxonomic ambiguity between GS and RS, identify the evolutionary relationship of TS to other two species and to assess the organoleptic preference on the dishes prepared using greenhouse and field-grown shoot-tops of three spinach. The genomic DNA extracted from GS, RS, and TS, PCR amplified and sequenced for the barcoding markers rbcL, ITS, matK-trnT and atpB-rbcL. The sequences obtained along with other reported related sequences were subjected to phylogenetic analysis. A sensory test was carried out using the shoot-tops of three species grown under greenhouse and field conditions. The taste panelists were asked to rank the dishes for preferred levels of color, aroma, texture, bitterness, and overall taste and the data were subjected to the association analysis. The rbcL and ITS markers separate GS and RS into two well -supported clades, B. alba and B. rubra respectively. The polymorphisms of atpB-rbcL and matK-trnT markers support the definition of two species. The monophyly of B. alba and B. rubra with T. fruticosum must be the reason for the same palate in dishes and designation of all three species under “spinach” in Sri Lanka. The taste panel data demonstrated that there is no specific fondness for greenhouse or field grown materials enabling the popularization of greenhouse-grown spinach to answer the safety concerns.

PEMA: a flexible Pipeline for Environmental DNA Metabarcoding Analysis of the 16S/18S ribosomal RNA, ITS, and COI marker genes.

BackgroundEnvironmental DNA and metabarcoding allow the identification of a mixture of species and launch a new era in bio- and eco-assessment. Many steps are required to obtain taxonomically assigned matrices from raw data. For most of these, a plethora of tools are available; each tool's execution parameters need to be tailored to reflect each experiment's idiosyncrasy. Adding to this complexity, the computation capacity of high-performance computing systems is frequently required for such analyses. To address the difficulties, bioinformatic pipelines need to combine state-of-the art technologies and algorithms with an easy to get-set-use framework, allowing researchers to tune each study. Software containerization technologies ease the sharing and running of software packages across operating systems; thus, they strongly facilitate pipeline development and usage. Likewise programming languages specialized for big data pipelines incorporate features like roll-back checkpoints and on-demand partial pipeline execution.FindingsPEMA is a containerized assembly of key metabarcoding analysis tools that requires low effort in setting up, running, and customizing to researchers’ needs. Based on third-party tools, PEMA performs read pre-processing, (molecular) operational taxonomic unit clustering, amplicon sequence variant inference, and taxonomy assignment for 16S and 18S ribosomal RNA, as well as ITS and COI marker gene data. Owing to its simplified parameterization and checkpoint support, PEMA allows users to explore alternative algorithms for specific steps of the pipeline without the need of a complete re-execution. PEMA was evaluated against both mock communities and previously published datasets and achieved results of comparable quality.ConclusionsA high-performance computing–based approach was used to develop PEMA; however, it can be used in personal computers as well. PEMA's time-efficient performance and good results will allow it to be used for accurate environmental DNA metabarcoding analysis, thus enhancing the applicability of next-generation biodiversity assessment studies.

Revisiting Metarhizium and the description of new species from Thailand.

Over the last two decades the molecular phylogeny and classification of Metarhizium has been widely studied. Despite these efforts to understand this enigmatic genus, the basal lineages in Metarhizium are still poorly resolved. In this study, a phylogenetic framework is reconstructed for the Clavicipitaceae focusing on Metarhizium through increased taxon-sampling using five genomic loci (SSU, LSU, tef, rpb1, rpb2) and the barcode marker ITS rDNA. Multi-gene phylogenetic analyses and morphological characterisation of green-spored entomopathogenic Metarhizium isolates from Thailand and soil isolates of M. carneum and M. marquandii reveal their ecological, genetic and species diversity. Nineteen new species are recognised in the Metarhizium clade with narrow host ranges: two new species are found in the M. anisopliae complex – M. clavatum on Coleoptera larvae and M. sulphureum on Lepidoptera larvae; four new species are found in the M. flavoviride complex – M. biotecense and M. fusoideum on brown plant hoppers (Hemiptera), M. culicidarum on mosquitoes, M. nornnoi on Lepidoptera larvae; three new species M. megapomponiae, M. cicadae, M. niveum occur on cicadas; five new species M. candelabrum, M. cercopidarum, M. ellipsoideum, M. huainamdangense M. ovoidosporum occur on planthoppers, leafhoppers and froghoppers (Hemiptera); one new species M. eburneum on Lepidoptera pupae; and four new species M. phuwiangense, M. purpureum, M. purpureonigrum, M. flavum on Coleoptera. Of these 19 new species, seven produce a sexual morph (M. clavatum, M. eburneum, M. flavum, M. phuwiangense, M. purpureonigrum, M. purpureum, and M. sulphureum) and asexual morphs are found in the remaining new species and also in M. sulphureum, M. purpureonigrum and M. purpureum. Metarhizium blattodeae, M. koreanum and M. viridulum are new records for Thailand. An alternative neotype for Metarhizium anisopliae is proposed based on multi-gene and 5′tef analyses showing that CBS 130.71 from Ukraine is more suitable, being from a much closer geographical location to Metchnikoff’s Metarhizium anisopliae. This isolate is distinct from the neotype of Metarhizium anisopliae var. anisopliae proposed by M. Tulloch from Ethiopia (ARSEF 7487). Six new genera are established for monophyletic clades subtending the core Metarhizium clade, including Keithomyces, Marquandomyces, Papiliomyces, Purpureomyces, Sungia, and Yosiokobayasia. Metarhizium carneum, M. aciculare, and M. neogunnii are combined in Keithomyces and one new combination for M. marquandii in Marquandomyces is proposed. Purpureomyces is introduced for species producing purple stromata including a new combination for M. khaoyaiense and two new species P. maesotensis and P. pyriformis. Papiliomyces contains two new combinations for M. liangshanense and Metacordyceps shibinensis. The genus Sungia is proposed for the Korean species M. yongmunense on Lepidoptera pupa and Yosiokobayasia for the Japanese species M. kusanagiense also on Lepidoptera pupa. A synoptic and dichotomous key to the accepted taxa is provided together with tables listing distinguishing morphological characters between species, host preferences, and geography.

Ecological time series and integrative taxonomy unveil seasonality and diversity of the toxic diatom Pseudo-nitzschia H. Peragallo in the northern Adriatic Sea

Pseudo-nitzschia H. Peragallo (1900) is a globally distributed genus of pennate diatoms that are important components of phytoplankton communities worldwide. Some members of the genus produce the neurotoxin domoic acid, so regular monitoring is in place. However, the identification of toxic members in routine samplings remains problematic. In this study, the diversity and seasonal occurrence of Pseudo-nitzschia species were investigated in the Gulf of Trieste, a shallow gulf in the northern Adriatic Sea. We used time series data from 2005 to 2018 to describe the seasonal and inter-annual occurrence of the genus in the area and its contribution to the phytoplankton community. On average, the genus accounted for about 15 % of total diatom abundance and peaked in spring and autumn, with occasional outbreaks during summer and large inter-annual fluctuations. Increased water temperature and decreased salinity positively affected the presence of some members of the genus, while strong effects could be masked by an unsuitable definition of the species complexes used for monitoring purposes. Therefore, combining morphological (TEM) and molecular analyses by sequencing the ITS, 28S and rbcL markers, eight species were identified from 83 isolated monoclonal strains: P. calliantha, P. fraudulenta, P. delicatissima, P. galaxiae, P. mannii, P. multistriata, P. pungens and P. subfraudulenta. A genetic comparison between the isolated strains and other strains in the Mediterranean was carried out and rbcL was inspected as a potential barcode marker in respect to our results. This is the first study in the Gulf of Trieste on Pseudo-nitzschia time series from a long-term ecological research (LTER) site coupled with molecular data. We show that meaningful ecological conclusions can be drawn by applying integrative methodology, as opposed to the approach that only considers species complexes. The results of this work will provide guidance for further monitoring efforts as well as research activities, including population genetics and genomics, associated with seasonal distribution and toxicity profiles.

Fungia fungites (Linnaeus, 1758) (Scleractinia, Fungiidae) is a species complex that conceals large phenotypic variation and a previously unrecognized genus

Recent molecular phylogenetic analyses of scleractinian corals have resulted in the discovery of cryptic lineages. To understand species diversity in corals, these lineages need to be taxonomically defined. In the present study, we report the discovery of a distinct lineage obscured by the traditional morphological variation of Fungia fungites. This taxon exists as two distinct morphs: attached and unattached. Molecular phylogenetic analyses using mitochondrial COI and nuclear ITS markers as well as morphological comparisons were performed to clarify their phylogenetic relationships and taxonomic positions. Molecular data revealed that F. fungites consists of two genetically distinct clades (A and B). Clade A is sister to a lineage including Danafungia scruposa and Halomitra pileus, while clade B formed an independent lineage genetically distant from these three species. The two morphs were also found to be included in both clades, although the attached morph was predominantly found in clade A. Morphologically, both clades were statistically different in density of septal dentation, septal number, and septal teeth shape. These results indicate that F. fungites as presently recognized is actually a species complex including at least two species. After checking type specimens, we conclude that specimens in clade A represent true F. fungites with two morphs (unattached and attached) and that all of those in clade B represent an unknown species and genus comprising an unattached morph with only one exception. These findings suggest that more unrecognized taxa with hitherto unnoticed morphological differences can be present among scleractinian corals.

<p><strong><em>Allium xinlongense</em></strong><strong> (Amaryllidaceae, Allioideae), a new species from western Sichuan</strong></p>

Allium xinlongense from western Sichuan Province, China, is described as a new species. Its diagnostic morphological characters were confirmed to be valid by comparing them to those of related species, such as A. maowenense, A. chrysanthum, A. rude, A. xichuanense, A. chrysocephalum, and A. herderianum. Molecular phylogenetic analysis (combining ITS and rpL32-trnL markers) indicated that this new species is a member of Allium sect. Daghestanica. A comprehensive description of this new species is provided, including habitat environment and detailed morphological traits.

<p><strong>Morphological and molecular evidence for a new species from China: <em>Piper semi-transparens</em> (Piperaceae)</strong></p>

We conducted morphological and molecular analyses to describe a new species of Piperaceae, Piper semi-transparens, which was observed in Hainan, Guangdong and Guangxi Provinces, China. Detailed morphological comparisons between the newly discovered species and other members of Piper indicate that Piper semi-transparens is similar to P. bonii and P. bonii var. macrophyllum, from which it differs in having glabrous leaf blades; stem densely short and unbranched pubescent when young, glabrous when mature; veins 5(–7); stamens 2; stigmas 3, rarely 4; spikes color white when mature; floral bracts glabrous; pericarp white and semi-transparent. Molecular analyses based on the ITS marker of nuclear ribosomal DNA strongly support that P. semi-transparens is a new species in the genus Piper, most closely related to P. bonii and P. bonii var. macrophyllum.

Allium species of section Rhizomatosa, early members of the Central Asian steppe vegetation

The genus Allium is one of the largest genera in the family of Amaryllidaceae. It comprises over 1000 species, and the number is still increasing. The new intergeneric classification system, based on molecular sequence data, subdivided the genus into 15 subgenera and more than seventy sections. In some sections, the phylogenetic position of species remained unresolved. This is also the case regarding the sections Caespitosoprason and Rhizomatosa. These two sections are confined to Central Asia, where they inhabit steppe habitats. For this study we conducted a phylogenetic analysis of most of the taxa of sections Caespitosoprason and Rhizomatosa with the nuclear marker ITS and three different chloroplast regions (trnQ-rps16, trnL-trnF and rpl32-trnL). Taxonomical remarks, identification key and distribution maps for all species of section Rhizomatosa (including all species of section Caespitosoprason) are presented. A newly discovered Allium bellulum from the Katun Valley (Altai, Russia) was cytologically compared (flow cytometry and karyology) with the other two known localities from Tuva, Russia, and the Buchtarma Valley, Altai, Kazakhstan, and the disjunctive distribution discussed. Divergence time estimates of the species of section Rhizomatosa and their distribution are in agreement with the origin and climate/landscape history of the Central Asian steppe since the Lower Miocene.

Đặc điểm hình thái và di truyền của ba loài thuộc chi trinh nữ (Mimosa)

Mimosa is a large genus of about 400 species of herbs and shrubs belonging to the Fabaceae family (Leguminosae). Many common species of this genus were reported with several important biological activities including antibacterial, antifungal, anti-inflammation and antioxidant. This study is aimed to investigate morphological characteristics and genetic relationships among 3 species of Mimosa genus: Mimosa pudica, Mimosa pigra and Mimosa diplotricha. In this context, morphological characteristics indicated the similarities and differences of stems, leaves, flowers and fruits. It is also showed that all of three species have some typical traits of Mimosa genus such as stem with thorns, touched- sensitive leaves, light pink flowers and flowers are grouped into fluffy globular clusters. Individual characteristics of each species recorded show differences such as leaves, stems, flowers and fruits in size, color and shape. This study combined the morphological report with DNA barcoding data to improve the data. Phylogeny indicated that Mimosa pigra and Mimosa diplotricha are close relationship rather than Mimosa pudica basied on ITS marker and matK sequences analysis.

Characterization of Vittatidera zeaphila (Nematoda: Heteroderidae) from Indiana with molecular phylogenetic analysis of the genus.

In the summer of 2016, a field of corn (Zea mays) in Spencer County, Indiana was observed with heavily stunted plants, and from the affected roots a large number of cysts were recovered. Soil samples were submitted to one of us (JF), who extracted the nematode cysts and sent them to the USDA-ARS, Mycology and Nematology Genetic Diversity and Biology Laboratory (MNGDBL), Beltsville, MD for morphological and molecular identification. Cysts and the recovered second-stage juveniles (J2) that were examined morphologically conformed to the measurements of Vittatidera zeaphila, the goose cyst nematode originally described from Tennessee, USA in 2010. The molecular analysis of J2 showed the sample from Spencer County matched exactly with V. zeaphila according to ribosomal DNA markers ITS, 28S, and 18S, and with mitochondrial cytochrome oxidase I (COI). The nuclear marker heat shock protein 90 (Hsp90) was also analyzed for the first time from the Indiana population of V. zeaphila. Similarities to existing cyst nematode sequences are reported herein. Geographically, although the county is across the Ohio River from Kentucky, the previously reported Hickman County, Kentucky location and Indiana detection are approximately 200 miles apart. To the best of our knowledge, this is the first report of V. zeaphila in Indiana.

Huriella flakusii (Teloschistaceae, lichenized Ascomycota), a new species from the Colca Canyon region in Peru

AbstractA new species, Huriella flakusii, is described from the arid region of the Colca Canyon in southern Peru, and is characterized by squamulose, orange thalli and abundant, crowded and aggregated, usually flexuous when compressed, reddish apothecia. The thalline cortex, proper margin and hypothecium are paraplectenchymatous. The ascospores are broadly ellipsoid, 10‒15 × 5.0‒9.5 µm, with rather thin septa, 2‒4 µm. Phylogenetic analyses of the DNA sequences of ITS, nuLSU and mrSSU rDNA markers revealed that the new species is nested within the Amundsenia-Squamulea subclade in the subfamily Xanthorioideae, and has a sister group relationship to Huriella loekoesiana from South Korea.