Low Intraspecific Variation Research Articles

Hymenochaete ametzii sp. nov. (Hymenochaetales, Basidiomycota), an endangered bark-dwelling species inhabiting old Quercus pyrenaica trees from the Iberian Peninsula

Bark-dwelling fungi represent a group of ecologically highly specialized organisms. This study deals with an undescribed species of Hymenochaete characterized by specifically inhabiting the bark of Pyrenean oak (Quercus pyrenaica), and producing effuse-reflexed basidiomata and mainly globose to subglobose basidiospores. Maximum Likelihood and Bayesian analyses of the nuclear ITS-LSU regions revealed that the sequences of H. ametzii form a monophyletic group with a low intraspecific variation and substantially different from closest taxa, further supporting its recognition as a species. The seven localities H. ametzii is known from are old forests with a long ecological continuity, containing large old Q. pyrenaica trees, and are located in the supramediterranean belt of the Mediterranean biogeographical region. Based on habitat availability and field counts of colonized trees, the global population of H. ametzii is estimated at 8,670 mature individuals. Considering that the traditional use of old Q. pyrenaica stands ceased several decades ago in the Iberian Peninsula, with a consequent decline in habitat quality and availability, it is concluded that H. ametzii is Endangered (EN) according to the IUCN criteria.

Variation in induced responses in volatile and non-volatile metabolites among six willow species: Do willow species share responses to herbivory?

Chemical variation is a critical aspect affecting performance among co-occurring plants. High chemical variation in metabolites with direct effects on insect herbivores supports chemical niche partitioning, and it can reduce the number of herbivores shared by co-occurring plant species. In contrast, low intraspecific variation in metabolites with indirect effects, such as induced volatile organic compounds (VOCs), may improve the attraction of specialist predators or parasitoids as they show high specificity to insect herbivores. We explored whether induced chemical variation following herbivory by various insect herbivores differs between VOCs vs. secondary non-volatile metabolites (non-VOCs) and salicinoids with direct effects on herbivores in six closely related willow species. Willow species identity explained most variation in VOCs (18.4%), secondary non-VOCs (41.1%) and salicinoids (60.7%). The variation explained by the independent effect of the herbivore treatment was higher in VOCs (2.8%) compared to secondary non-VOCs (0.5%) and salicinoids (0.5%). At the level of individual VOCs, willow species formed groups, as some responded similarly to the same herbivores. Most non-VOCs and salicinoids were upregulated by sap-suckers compared to other herbivore treatments and control across the willow species. In contrast, induced responses in non-VOCs and salicinoids to other herbivores largely differed between the willows. Our results suggest that induced responses broadly differ between various types of chemical defences, with VOCs and non-VOCs showing different levels of specificity and similarity across plant species. This may further contribute to flexible plant responses to herbivory and affect how closely related plants share or partition their chemical niches.

Potential of reproductive traits in functional ecology: A quantitative comparison of variability in floral, fruit, and leaf traits.

Despite their claimed low intraspecific variability, plant reproductive traits are less frequently used in functional ecology. Here we focused on underrepresented plant organs, i.e. flowers and fruits, by comparing their traits with well-established leaf traits. We evaluated 16 functional traits (six floral, six fruit, and four leaf traits) in a randomly selected group of woody species under comparable environmental conditions. We aimed to assess interspecific and intraspecimen variability and explore the potential of the proposed flower and fruit traits for ecological research. Traits related to the dry mass of flowers and fruits exhibited the highest interspecific variability, while carbon content traits in flowers and leaves had the lowest. At a specimen level, specific leaf area revealed the highest variation. Carbon content traits for all organs demonstrated the least intraspecimen variability, with flower carbon content being the least variable. Our study revealed connections between the newly proposed traits and widely recognized functional traits, uncovering intriguing links between the established traits and the floral and fruit traits upon which we focused. This complements the already well-recognized variability in plant form and function with additional insights into reproductive processes.

Characterization of Coniella granati Isolates Causing Pomegranate Decline in Italy.

Coniella granati, the causal agent of pomegranate crown rot, twig blight, and fruit decay, is an emerging worldwide pathogen with a heavy impact on pomegranate cultivation. In this study, we report the rapid spread of the fungus in Italian pomegranate orchards associated with crown rot symptoms and provide new results on fungal development, baseline sensitivity to different fungicides, and intraspecific variability by analyzing 11 isolates, representative of populations of the pathogen from comparable pomegranate orchards in different regions of Italy. In vitro assays showed that 25 to 30°C was the optimal range for both colony growth and conidial germination, corroborating the results previously obtained for Californian and Greek isolates. According to the baseline sensitivity assay on the response of colony growth and conidial germination to 10 fungicides, fludioxonil, thiophanate-methyl, tebuconazole, and cyprodinil were the most effective. Random amplified polymorphic DNA (RAPD) analysis, carried out using fourteen 10-mer primers, showed very low intraspecific variability (similarity coefficient >0.95), probably as a result of the recent spread of the pathogen in Italy and the uncommon occurrence of the sexual process as a source of genetic variability. In summary, this study provides new knowledge on C. granati that will be helpful for improving pomegranate crown rot management.

A chromosome-scale assembly reveals chromosomal aberrations and exchanges generating genetic diversity in Coffea arabica germplasm

In order to better understand the mechanisms generating genetic diversity in the recent allotetraploid species Coffea arabica, here we present a chromosome-level assembly obtained with long read technology. Two genomic compartments with different structural and functional properties are identified in the two homoeologous genomes. The resequencing data from a large set of accessions reveals low intraspecific diversity in the center of origin of the species. Across a limited number of genomic regions, diversity increases in some cultivated genotypes to levels similar to those observed within one of the progenitor species, Coffea canephora, presumably as a consequence of introgressions deriving from the so-called Timor hybrid. It also reveals that, in addition to few, early-occurring exchanges between homoeologous chromosomes, there are numerous recent chromosomal aberrations including aneuploidies, deletions, duplications and exchanges. These events are still polymorphic in the germplasm and could represent a fundamental source of genetic variation in such a lowly variable species.

Molecular Discrimination and Phylogenetic Relationships of Physalis Species Based on ITS2 and rbcL DNA Barcode Sequence

Plants of the genus Physalis are of economic interest because of their fleshy edible fruits with high nutritional value. Some species have high medicinal value with a long history of ethno-medicinal use to treat diverse diseases. There is therefore a need to correctly discriminate the different species of Physalis for proper utilization. Although most Physalis species have unique morphologies, their vegetative stages are identical, making it difficult to accurately identify them based on morphological characteristics. DNA barcoding has the potential to discriminate species accurately. In this study, ribulose bisphosphate carboxylase large (rbcL) and internal transcribed spacer 2 (ITS2) regions were used to discriminate Physalis species and to reveal their phylogenetic relationships and genetic diversity. Physalis plant samples were collected from seven counties in Kenya based on the availability of the germplasm. The voucher specimens were identified using the botanical taxonomy method and were deposited in the University of Nairobi herbarium. Genomic DNA was isolated from leaf samples of 64 Physalis accessions and used for PCR amplification and the sequencing of rbcL and ITS2 barcode regions. The discriminatory ability of the barcodes was based on BLASTn comparison, phylogenetic reconstruction and cluster analysis, and the determination of inter- and intra-specific distances. The nucleotide polymorphism, genetic diversity and distance of the identified Physalis species were determined using DnaSP and MEGA 11.0 software. Species discrimination was more robust using ITS2 sequences. The species identified and discriminated by ITS2 sequences were Physalis purpurea, Physalis peruviana and Physalis cordata. The rbcL sequences were only able to identify Physalis to the genus level. There was high interspecific and low intraspecific divergence within the identified Physalis species based on ITS2 sequences. The ITS2 barcode is an ideal DNA barcode for use in the discrimination of species, as well as in genetic diversity studies of Physalis accessions in Kenya.

Microbial Diversity of Marula Wine during Spontaneous Fermentation

Marula wine is produced from ripe fruits of the Sclerocarya birrea subspecies caffra tree through spontaneous fermentation. A few culture-based studies have shown that the fermentation is largely driven by yeasts, although, in the early stages, some lactic acid bacteria (LAB) and acetic acid bacteria may be detected. Some of the microbes may produce undesirable metabolites that lead to the spoilage and short shelf life of the wine. However, there is generally limited information on the microbial composition and its contribution to the chemical characteristics of the resultant marula wine. The aim of this study was to characterise the microbial population of marula wine from different localities in the Limpopo province, South Africa. MALDI-TOF and amplicon sequencing technique were used to identify microbial strains and to determine their diversity and changes in the different stages of fermentation. The phylogenetic relationships of LAB and S. cerevisiae were analysed using multilocus sequence typing. Bacterial species that were common in the different marula wines included Gluconobacter oxydans, Lactiplantibacillus plantarum, Levilactobacillus brevis, Lacitilactobacillus nagelii, Lentilactobacillus kefiri and Lentilactobacillus parabuchneri, and the yeasts were Hanseniaspora guiliermondii, Saccharomyces cerevisiae, Rhodotorula mucilaginosa and Pichia kudriavzevii. The MLST data indicated common microbiota from different marula wines with low intraspecific diversity, suggesting that the LAB and S. cerevisiae strains that are mainly responsible for the spontaneous fermentation of marula wine are similar irrespective of the geographical differences and production preferences.

Revisiting the bipolarity of Roaldia revoluta (Mitt.) P.E.A.S. Câmara & Carv.-Silva (Bryophyta, Pylaisiaceae)

AbstractThe occurrence of species in both polar regions (bipolarity) is a common phenomenon in the Antarctic flora. Considering the high morphological variation in polar regions due to extreme conditions, the use of molecular tools is indispensable for testing whether Arctic and Antarctic populations indeed belong to the same species. However, few phylogeographic studies of bipolar bryophytes have been conducted so far, especially when comparing molecular and morphological variation. Here, we assess the bipolarity and intraspecific variation of Roaldia revoluta, a strictly bipolar species of pleurocarpous mosses. Phylogenetic analyses based on ITS sequences clearly resolve R. revoluta as monophyletic and confirm its bipolar distribution pattern. Low intraspecific molecular variation in the markers ITS/26S and rpl16 was observed, and most specimens from both polar regions belong to a single haplotype, making it difficult to infer the origin and dispersal routes between both polar regions of R. revoluta. Morphometric analysis furthermore suggests that there are no significant morphological differences among populations from both polar regions and that morphological variation is mainly influenced by local environmental conditions. Our data do not unequivocally support the recent separation of the former intraspecific taxon Hypnum revolutum var. dolomiticum at the species level as Roaldia dolomitica.

Wood trait trade‐offs in desert plants: A triangular model to understand intra‐ and interspecific variations along an aridity gradient

AbstractThe hydraulic architecture of woody plants may show intra‐ and interspecific variations along aridity gradients as a result of wood trait trade‐offs, which depend on species‐specific abilities to withstand or evade drought. Plant hydraulic architecture can be defined as the set of different cell types involved in water transport through the soil–plant‐atmosphere continuum. At the present time, there are two models designed to understand trade‐offs among xylem wood functional traits, developed by Baas and Pratt, respectively. The first model presents a triangular trade‐off space where three main plant xylem functions are linked to each other to provide resistance to embolism in evolutionary time. The second model depicts xylem cell types associated with their main functions, namely vessel‐transport, parenchyma‐storage and fibre‐support. We propose here a new model considering the same relative xylem cell trait abundance for three desert species with different drought tolerance strategies along an aridity gradient of more than 300 mm. Hence, our model may account for both intra‐ and interspecific variations in wood traits for species from different localities along an aridity gradient. The tree Neltuma flexuosa showed little intraspecific variation in the medium drought resistance portion of the trade‐off space, suggesting this species is independent from rainfall. Larrea divaricata showed a similar low intraspecific variation, but the spatial location in the trade‐off space suggests it has a high tolerance to drought. The third species, Bulnesia retama, showed a higher intraspecific variation within the trade‐off space, changing its hydraulic architecture in more fibre area with increasing aridity. Our model seems to be a generalized tool to assess both intra‐ and interspecific variations in trade‐offs for hydraulic architecture among woody plants as a response to drought.

Evaluating the Utility of Five Gene Fragments for Genetic Diversity Analyses of Mytella strigata Populations

Mytella strigata (Hanley, 1843) is an invasive mussel species that has rapidly spread in China in recent years. Here, we tested the utility of three mitochondrial gene fragments, COI, 12S, and 16S, and two nuclear gene fragments, D1 28S and 18S-ITS1, for characterizing the levels of genetic diversity among and within populations using 191 M. strigata specimens collected in China to aid ongoing efforts to identify the origin of the invasion as well as molecular genetic studies. M. strigata exhibited two sex-associated haplogroups according to the COI and 12S sequences. The ratio of female-lineage to male-lineage COI and 12S sequences was 149:22 and 72:7, and the genetic distances between haplogroups were 6.56 and 9.17, respectively. Only one haplotype was detected among the 18S-ITS1 sequences (413 bp), and three haplotypes were detected among the D1 28S sequences (296 bp). The haplotype diversity of both the female-lineage COI and 12S sequences was greater than 0.5, and the nucleotide diversity of the 12S, 16S, D1 28S, and 18S-ITS1 sequences was less than 0.005 in all six populations in China. Our findings indicated that COI is the most useful gene fragment for genetic diversity studies of M. strigata populations; D1 28S and 18S-ITS1 sequences would be useful for species identification because of their low intraspecific diversity. Our genetic analysis of the COI sequences revealed Colombia as the most likely origin of M. strigata in China and showed that the invasive populations in China have recently experienced or are currently experiencing a population bottleneck.

Genetic variability and phylogenetic characterization of different populations of Meloidogyne izalcoensis and reaction of coffee genotypes to this new species detected in Brazil

AbstractA new root‐knot nematode (RKN), Meloidogyne izalcoensis, was detected on coffee in Brazil. This species was first described on coffee from El Salvador and later detected in Asia and Africa. There are no reports on the genetic diversity of this species and resistance to M. izalcoensis has not yet been studied in coffee cultivars. All populations were identified by esterase phenotype and sequence‐characterized amplified region (SCAR) species‐specific markers. Based on random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) markers, low intraspecific variability was detected among M. izalcoensis populations from Africa, Vietnam and Brazil, but the population from El Salvador showed fewer genetic differences from the other populations. Phylogenetically, all populations of M. izalcoensis from different locations (El Salvador, Kenya, Tanzania, Vietnam and Brazil) grouped with 90% and 69% bootstrap support for COII and HSP90 regions, respectively, indicating that these markers are highly conserved for the species. In addition, both markers enabled separation of M. izalcoensis populations from other important coffee Meloidogyne species, including M. exigua, M. paranaensis, M. incognita, M. arabicida and M. lopezi. In resistance studies, most genotypes tested that had resistance to other Meloidogyne spp. (Amphillo × Catuaí, Hybrid of Timor, IAPAR 59, IPR 99, IPR 100, IPR 102, IPR 103, IPR 105, IPR 106, IPR 107 and IPR 108) were found to be susceptible to M. izalcoensis, except for the rootstock cv. Apoatã IAC 2258, which proved to be moderately resistant, with a genetic segregation for this character of 43.8%.

Intraspecific Variation in Three Species of Semudobia (Diptera: Cecidomyiidae) Forming Fruit Galls on Betula (Betulaceae) in the Holarctic Region

We analyzed the Folmer fragment of the mitochondrial COI gene of three closely related species of Semudobia (Diptera: Cecidomyiidae): S. betulae (Winnertz), S. tarda Roskam, and S. skuhravae Roskam. These species form fruit galls on different birch species (Betula, Betulaceae) in the Holarctic Region. In Japan, larvae of S. betulae and S. tarda were collected from the native Japanese birch, B. platyphylla, whereas S. skuhravae was collected from the eastern Palearctic birch, B. ermanii. In the Netherlands, the three species were collected from intermediate forms between the Eurasian endemic birches, B. pendula and B. pubescens. Additional sequences of Canadian and European S. betulae and S. skuhravae were obtained from the Barcode of Life Data system (BOLD). In a maximum likelihood analysis, the individuals of each species formed strongly supported clades, but genetic divergence was high within each clade indicating species complexes. European and Canadian individuals of S. betulae and S. skuhravae each formed a single clade with low intraspecific variation, suggesting that these species were introduced together with their hosts to North America. In contrast, Japanese individuals of the three Semudobia species were remarkably distant from European and Canadian individuals, indicating that they have been isolated genetically for a long period on their native eastern Palearctic and Japanese hosts.

Strategies for Intraspecific Genotyping of Duckweed: Comparison of Five Orthogonal Methods Applied to the Giant Duckweed Spirodela polyrhiza.

The predominantly vegetative propagating duckweeds are of growing commercial interest. Since clonal accessions within a respective species can vary considerably with respect to their physiological as well as biochemical traits, it is critical to be able to track the clones of species of interest after their characterization. Here, we compared the efficacy of five different genotyping methods for Spirodela polyrhiza, a species with very low intraspecific sequence variations, including polymorphic NB-ARC-related loci, tubulin-gene-based polymorphism (TBP), simple sequence repeat variations (SSR), multiplexed ISSR genotyping by sequencing (MIG-seq), and low-coverage, reduced-representation genome sequencing (GBS). Four of the five approaches could distinguish 20 to 22 genotypes out of the 23 investigated clones, while TBP resolved just seven genotypes. The choice for a particular method for intraspecific genotyping can depend on the research question and the project budget, while the combination of orthogonal methods may increase the confidence and resolution for the results obtained.

Short communication: Zizyphus fruit fly (Carpomya incompleta (Becker), Diptera: Tephritidae) is expanding its range in Europe

Aim of study: To identify a new pest of jujube reported by the farmers in Spain. Area of study: The Iberian Peninsula (Spain). Material and methods: The insects were identified according to the mitochondrial cytochrome C oxidase subunit I (COI) gene and the main morphological features of this tephritid species, including the ocellar seta, the mesonotum and apical crossband in winds. Main results: Based on morphological characterization the insects were identified as Carpomya incompleta (Becker) (Diptera: Tephritidae), which was confirmed by the phylogenetic analysis with more than 94% of identity. Besides, the distance analysis showed very low intraspecific divergence in C. incompleta sequences. Research highlights: We report the presence of the zizyphus fruit fly in Spain for the first time. This presence has been reported to the Early Warning Systems on Alien Invasive Species of the Andalusian Government (Spain).

Morphological and molecular studies on the genus Gayralia (Ulotrichales, Chlorophyta) in northeastern Brazil with expansion of its species distribution

Abstract The genus Gayralia has a complex taxonomy, often with morphological and ontogenetic characteristics that overlap with those of other monostromatic green algae, mainly Monostroma. Molecular studies of monostromatic green algae are still rare, especially in Brazil, where G. brasiliensis was described originally. Considering the difficulties in delimiting genera and species based on morphological criteria, we investigated the genus Gayralia through ITS sequencing of field collections in the tropical northeastern region of Brazil. Our 14 newly generated sequences formed a fully supported clade with G. brasiliensis showing low intraspecific divergence (0–0.6%). The distribution of G. brasiliensis in Brazil was expanded to the northeastern Brazilian coast, with the first record for the states of Rio Grande do Norte, Paraíba, Alagoas, and Sergipe. In addition, two sequences from India identified as G. “oxysperma” were grouped in the G. brasiliensis clade, indicating that they correspond to the same genetic species. This finding extends the geographical distribution of G. brasiliensis to India. Our results place Monostroma kuroshiense within the Gayralia clade, which warrants a new combination – Gayralia kuroshiensis (F.Bast) comb. nov.

Molecular identification of Tetrahymena species.

Mitochondrial cox1 689 bp barcodes are routinely used for identification of Tetrahymena species. Here, we examine whether two shorter nuclear sequences, the 5.8S rRNA gene region and the intergenic region between H3 and H4 histone genes, might also be useful either singly or in combination with each other or cox1. We obtained sequences from ~300 wild isolates deposited at the Tetrahymena Stock Center and analyzed additional sequences obtained from GenBank. The 5.8S rRNA gene and portions of its transcribed flanks identify isolates as to their major clade and uniquely identify some, but not all, species. The ~330 bp H3/H4 intergenic region possesses low intraspecific variability and is unique for most species. However, it fails to distinguish between two pairs of common species and their rarer counterparts, and its use is complicated by the presence of duplicate genes in some species. The results show that while the cox1 sequence is the best single marker for Tetrahymena species identification, 5.8S rRNA, and the H3/H4 intergenic regions sequences are useful, singly or in combination, to confirm cox1 species assignments or as part of a preliminary survey of newly collected Tetrahymena. From our newly collected isolates, the results extend the biogeographical range of T.shanghaiensis and T.malaccensis and identify a new species, Tetrahymena arleneae n. sp. herein described.

Sporothrix brasiliensis and Feline Sporotrichosis in the Metropolitan Region of Rio de Janeiro, Brazil (1998-2018).

Feline sporotrichosis is enzootic in different regions of Brazil, especially in Rio de Janeiro. This study compared the genotype profiles of Sporothrix sp. isolated from cats in Rio de Janeiro between 1998 and 2018 and evaluated their association with clinical and epidemiological characteristics. One hundred nineteen Sporothrix sp. isolates from a cohort of cats with sporotrichosis seen at INI/Fiocruz were included. Clinical and epidemiological data were obtained from the medical records of the animals. T3B PCR fingerprinting was used for molecular identification of the Sporothrix species. All isolates were characterized as Sporothrix brasiliensis, with the observation of low intraspecific variation in 31 isolates (31.3%). The interval between lesion onset and first medical visit at INI/Fiocruz, as well as treatment duration until clinical cure, was longer in cats from the first decade of the epizootic. In addition, the frequency of the variables “good general status” and “presence of lymphadenomegaly” was higher among cats whose strains did not exhibit intraspecific variation. So far, S. brasiliensis has been the only species identified in feline cases of sporotrichosis since the beginning of the epizootic in Rio de Janeiro at INI/Fiocruz.

Molecular Characterization of Ctenotaenia marmotae (Frölich, 1802) Railliet, 1893 (Cyclophyllidea: Anoplocephalidae) Parasitizing Rodents of the Genus Marmota and Spermophilus from Eurasia

Cestodes Ctenotaenia marmotae are a widespread parasite of Eurasian ground squirrels. Being the only valid species of its genus, it has been recorded in ten host species from the genera Marmota and Spermophilus. Its definitive hosts live in similar ecological conditions and their area covers a wide geographical territory. Therefore, it remained unclear whether C. marmotae is a single species and how high the level of its genetic variability is. The present study analyzed the variability of two mitochondrial cestode genes from four host species from geographically distant localities. Phylogenetic analysis revealed that the newly obtained sequences form a species-level clade with already known sequences from the GenBank. Small genetic distances between the studied sequences indicate low intraspecific genetic variability within the C. marmotae, which may be a consequence of rapid expansion of the cestodes in ground squirrel species inhabiting the flat and mountain steppe landscapes in the past.

Genetic distance and phylogenetic relationships of some Echinostoma species (E. malayanum, E. revolutum, E. miyagawai) and Hypoderaeum conoideum (family Echinostomatidae) inferred from partial 28S rDNA sequence analysis

Echinostomiasis is a neglected disease caused by the intestinal flukes (family Echinostomatidae, suborder Echinostomata) and is common in communities in Asian countries, such as India, Indonesia, the Philippines, China, Malaysia, Singapore, Korea, Japan, Thailand, Laos, Cambodia and Vietnam. The genetic markers from the nuclear ribosomal transcription units are commonly used in genetic studies and phylogenetic analyses. A portion of the 28S rDNA sequence (domains D1–D3, of 1062–1067 bp for the final use) was obtained from the zoonotic Echinostoma malayanum (strain E/Amala-EMI3-TH), E. revolutum (strain Erevo-MSD15-TH), E. miyagawai (Emiya-RED11-TH), and Hypoderaeum conoideum (Hcono-RED42-TH) species; and used to perform an alignment for genetic distance estimation and phylogenetic analysis. The alignment was performed using 62 strains of 42 species from 19 genera of the family Echinostomatidae, including Echinoparyphium, Echinostoma, Artyfechinostomum, Patagifer, Neoacanthoparyphium, Hypoderaeum, Echinoparyphium, Drepanocephalus, Euparyphium, Chaunocephalus, Neopetasiger, Ribeiroia, Cathaemasia, Rhopalias, Isthmiophora, Petasiger, Moliniella, Pegosomum, and Schistosoma (family Schistosomatidae). The genetic distance estimation among 16 strains/10 species has shown a low intra-specific divergence level between strains within the same species, such as E. miyagawai (0–0.10%), E. revolutum (0.10–0.50%), and H. conoideum (0–0.10%), while between strains within the same genus it was higher (normally over 1.0%) and among strains/species between genera it was the highest (3.06–4.12%). The 28S rDNA sequence analysis and phylogenetic relationship well supported the Echinostoma/ Artyfechinostomum malayanum intergeneric taxonomy and the topology indicated clear, well-supported positions of member species in different genera in the family Echinostomatidae of the suborder Echinostomata. More sensu lato samples of the genera, are required for sequencing, particularly those of zoonotic species in the five genera: Artyfechinostomum, Echinostoma, Hypoderaeum, Echinoparyphium, and Isthmiophora. The resultant mitochondrial and nuclear data obtained from these species will be a good source to use to clearly assess the taxonomic and generic relationships.

DNA Barcoding for Dipterocarpus Species in Vietnam Based on Chloroplast Gene Region matK

Objective: Dipterocarp identification is challenging, based on traditional methods of morphological characteristics for species conservation and management. In the present study, the matK region was used as a DNA bar-code to identify the Dipterocarpaceae species in Vietnam. Methods: All the samples were collected at studied localities and identified by morphological methods by the botanist. DNA samples were extracted using the modified Cetyltrimethylammonium bromide method and amplified the matK gene by PCR. Sequencing was performed on an Avant 3100 Automated DNA sequencer system. Results: Thirty-three trees were analyzed, representing 11 species (eight in the Dipterocarpus genus, one in Hopea and one in Vativa) and showing low intraspecific divergence and interspecific divergence within and among Dipterocarpus species, with an average of 0.08% and 0.19%, respectively. However, there was zero distance between D. costatus, D. dyeri, D. intricatus and D. obtusifolius. Three Dipterocarpus species (D. alatus, D. condorensis and D. tuberculatus) were discriminated against, while others (D. costatus, D. dyeri and D. intricatus) showed too low divergent intraspecific lineages and suggested hybridization may be affected them. These results also indicated a low barcoding gap for Dipterocarpus species. The highest gene distance was found between Vatica and Dipterocarpus, with an average of 2.12%. Conclusion: In this study, we used the matK region as a tool to identify some Dipterocarpaceae species in Vietnam. The accurate identification of species may manage and conserve species effectively. Our results showed little sequence differentiation among most Dipterocarpus species. The lack of differentiation may be due to incomplete lineage sort and hybridization between sympatric species. In order to identify species, a combination of two or three chloroplast gene regions should be utilized in further studies. DNA barcoding gap should be analyzed within different genera in Dipterocarpaceae.