Intraspecific Genetic Distances Research Articles

Tackling the Taxonomic Challenges in the Family Scoliidae (Insecta, Hymenoptera) Using an Integrative Approach: A Case Study from Southern China.

Simple SummaryThe scoliid wasps are larval parasitoids of scarabaeoid beetles and pollinators of various plants and therefore are important in biological control and pollination. However, some species of these wasps are extremely morphologically similar and difficult to identify. In this study, we used an integrative approach of species delimitation, combining morphology with molecular data, to investigate the species of Scoliidae from southern China. On the basis of both morphological and molecular evidence, we recognized 22 morphospecies of 9 genera in two tribes, among which one undescribed cryptic species of the polytypic species Solia (Discolia) superciliaris Saussure, 1864, five newly recorded species and one pending subspecies were discovered. Our study indicates that such an integrative approach is a potent tool in the effort to tackle the taxonomic challenges in Scoliidae, and even in other diverse groups of Aculeata, of which sexual dimorphism and cryptic species are common.Species of the family Scoliidae are larval parasitoids of scarabaeoid beetles and pollinators of various plants. Despite their great importance in pest biological control and plant pollination, the taxonomy and systematics of these parasitoids are far from clear. Some species of the family are extremely morphologically similar and difficult to identify, especially in males. In this study, an integrative taxonomic approach, combining morphology and molecular data, was used to discriminate the species of Scoliidae from southern China. In total, 52 COI sequences belonging to 22 morphospecies of 9 genera in two tribes were obtained. The COI sequences worked well for the identification of all the studied species, with intraspecific genetic distances generally less than 2%, while interspecific distances ranged between 5.3% and 20.8%. The delimitations of the problematic species and subspecies of Scolia and Megacampsomeris are well solved by COI sequences, suggesting that DNA barcoding could be a useful identification tool for Scoliidae. Based on both morphological and molecular evidence, we discovered one undescribed cryptic species of the polytypic species Solia (Discolia) superciliaris Saussure, 1864, five newly recorded species, i.e., Scolia (Discolia) sikkimensis Bingham, 1896, Sericocampsomeris flavomaculata Gupta and Jonathan, 1989, Megacampsomeris asiatica (Saussure, 1858), Megacampsomeris pulchrivestita (Cameron, 1902) and Megacampsomeris shillongensis (Betrem, 1928) and one pending subspecies of Scolia (Discolia) watanabei (Matsumura, 1912) from China. Our study indicates that such an integrative approach, combing both molecular and morphological evidence, is a potent tool to tackle the taxonomic challenges in the family Scoliidae, or even, in other diverse groups of Aculeata, of which sexual dimorphism and cryptic species are common.

DNA Barcoding of Penaeidae (Decapoda; Crustacea): Non-Distance-Based Species Delimitation of the Most Economically Important Shrimp Family

The Penaeidae family includes some of the most economic and ecological important marine shrimp, comprising hundreds of species. Despite this importance and diversity, the taxonomic classification for penaeid shrimp has constantly been revised, and issues related to the species identification are common. In this study, we implemented DNA barcoding analyses in addition to single-gene species delimitation analyses in order to identify molecular operational taxonomy units (MOTUs) and to generate robust molecular information for penaeid shrimp based on the cytochrome oxidase subunit I (COI) mitochondrial gene. Our final data set includes COI sequences from 112 taxa distributed in 23 genera of penaeids. We employed the general mixed Yule coalescent (GMYC) model, the Poisson tree processes (PTP), and the Bayesian PTP model (bPTP) for MOTUs delimitation. Intraspecific and interspecific genetic distances were also calculated. Our findings evidenced a high level of hidden diversity, showing 143 MOTUs, with 27 nominal species not agreeing with the genetic delimitation obtained here. These data represent potential new species or highly structured populations, showing the importance of including a non-distance-based species delimitation approach in biodiversity studies. The results raised by this study shed light on the Penaeidae biodiversity, addressing important issues about taxonomy and mislabeling in databases and contributing to a better comprehension of the group, which can certainly help management policies for shrimp fishery activity in addition to conservation programs.

Application of COI Primers 30F/885R in Rotifers to Regional Species Diversity in (Sub)Tropical China

Rotifers are the most diverse group in freshwater zooplankton and play an important role in food webs and ecosystems. DNA barcoding has become a useful approach to investigate species diversity at local and regional scales, but its application is still limited by efficient primers for the group. To test a pair of primers 30F/885R recently designed for rotifers, we applied them to investigating regional species diversity in the freshwater of South China. We sequenced the cytochrome c oxidase subunit I (COI) gene of rotifers collected from the investigated 23 reservoirs in a large river basin and obtained 145 COI sequences from 33 species in 14 genera. The mean PCR success rate for all tested species was 50%. The 145 sequenced mtCOI in this study covered 33 of 64 identified morphological taxa, including most of the common species in the basin. The intraspecific genetic distance was calculated with a K2P model for 24 rotifer species occurring in the quantitative samples, in which 15 rotifers, such as Keratella cochlearis and Brachionus calyciflorus, had a genetic distance higher than 5%. The high intraspecific genetic differentiation indicates that cryptic species are probably common in (sub)tropical China.

Species diversity of freshwater shrimp in Henan Province, China, based on morphological characters and COI mitochondrial gene.

Freshwater shrimp are a rich species group, with a long and problematic taxonomic history attributed to their wide distribution and similar morphological characteristics. Shrimp diversity and species identification are important cornerstones for fisheries management. However, identification based on morphological characteristics is a difficult task for a nonspecialist. Abundant freshwater shrimp species are distributed in the waters of Henan Province, but investigations of freshwater shrimp are limited in this region, especially concerning molecular features. Here, we combined morphology and DNA barcodes to reveal the species diversity of freshwater shrimp in Henan province. A total of 1,200 freshwater shrimp samples were collected from 46 sampling sites, and 222 samples were chosen for further microscopic examination and molecular delimitation. We used tree‐based methods (NJ, ML, and bPTP) and distance‐based methods (estimation of the paired genetic distances and ABGD) to delimit species. The results showed that there were nine morphospecies based on morphological characteristics; all could effectively be defined by molecular methods, among which bPTP and ABGD defined 13 and 8 MOTUs, respectively. The estimation of the paired genetic distances of K2P and the p‐distances had similar results. Mean K2P distances and p‐distances within species were both equal to 1.2%. The maximum intraspecific genetic distances of all species were less than 2%, with the exception of Palaemon modestus and M. maculatum. Various analyses have shown that P. modestus and M. maculatum have a large genetic differentiation, which may indicate the existence of cryptic species. By contrast, DNA barcoding could unambiguously discriminate 13 species and detect cryptic diversity. Our results demonstrate the high efficiency of DNA barcoding to delimit freshwater shrimp diversity and detect the presence of cryptic species.

Morpho-species of the genus Leuciscus Cuvier, 1816 (Teleostei: Leuciscinae) from Iran revisited using molecular approaches

Leuciscus is a genus of fish belonging to subfamily Leuciscinae. While, it is widely distributed throughout Eurasia, our perception of the phylogenetic relationships among its members is mainly based on morphological characters. In this study, two mitochondrial (CYTB and COX1) and one nuclear (RAG1) genomic markers were used in order to assess the taxonomic relationships between two Iranian Leuciscus morpho-species. Analysis of the combined dataset using Maximum Likelihood (ML) and Bayesian Inference (BI) resulted in well-resolved trees, where most clades were supported by high statistical values. The trees revealed two clades corresponding with the Caspian Asp, Leuciscus aspius, and the Mesopotamian Asp, L. vorax. The mean intraspecific and interspecific genetic distance between Leuciscus species was 0.5% and 10.6%, respectively. The genetic distance between L. aspius sequences of Iran and Europe was 1.1% and the genetic distance between L. vorax sequences from Iran and those of Turkey and Syria was 0.8%. This confirmed that the morpho-species Leuciscus from the Caspian basin belongs to L. aspius, while those from the Tigris basin belong to L. vorax. Due to very small genetic distances between some Leuciscus members, specific species delimitation analyses are required to clarify genus taxonomy throughout its distribution range.

Molecular identification of Aquilaria species with distribution records in China using DNA barcode technology

ABSTRACTS Aquilaria species is one of the main plant resources that produce agarwood, which containing black resin with important economic and medicinal values. There are about 15 species known to the genus around the world, but only two can be found in China, i.e. A. sinensis and A. yunnanensis. In this study, A. sinensis and A. yunnanensis that endemic respectively to Hainan and Yunnan were sampled, on the basis of the investigation and observation of their main morphological features in plantation. Five primers, i.e. ITS2, matK, trnL-trnF1, trnL-trnF2, and trnH-psbA, were eventually selected for DNA barcoding. The results showed that the seed surface of A. sinensis is smooth or sparsely pubescent, and the seed appendages were long. While the seed surface of A. yunnanensis is densely covered with yellow hairs and the seed appendages are short. The trnL-trnF1 sequence fragment has significant intraspecific and interspecific genetic distances. However, the species identification success rate of ITS2+matK combination was finally screened to be the highest, which was verified by the BBA method of TaxonDNA. The phylogenetic trees cluster analysis revealed that the classification of A. sinensis and A. yunnanensis is significant, and there is geographic isolation between the two species. Therefore, on the premise of accurate identification of plant morphological characters, ITS2+matK combination can be used to accurately identify the Aquilaria species in China.

Identification of Cannabis Sativa L. Based on rbcL Sequence.

Objective To assess the feasibility of the rbcL sequence of chloroplast DNA as a genetic marker to identify Cannabis sativa L. Methods The rbcL sequences in 62 Cannabis sativa L. samples, 10 Humulus lupulus samples and 10 Humulus scandens DNA samples were detected, and 96 rbcL sequences of the Cannabaceae family were downloaded from Genbank. Sequence alignment was performed by MEGA X software, the intraspecific and interspecific Kimura-2-Parameter （K2P） genetic distances were calculated, and the system clustering tree was constructed. Results The rbcL sequence length acquired by sequencing of Cannabis sativa L. and Humulus scandens were 617 bp and 649 bp, respectively, and two haplotypes of Cannabis sativa L. were observed in the samples. The BLAST similarity search results showed that the highest similarity between the sequences acquired by sequencing and Cannabis sativa L. rbcL sequences available from Genbank was 100%. The genetic distance analysis showed that the maximum intraspecific genetic distance （0.004 9） of Cannabis sativa L. was less than the minimum interspecific genetic distance （0.012 9）. The results of median-joining network and system clustering tree analysis showed that Cannabis sativa L. and other members of the Cannabaceae family were located in different branches. Conclusion The rbcL sequence could be used as a DNA barcode for identifying Cannabis sativa L., and combined with comparative analysis of the rbcL sequence and system cluster analysis could be a reliable and effective detection method for Cannabis sativa L. identification in forensic investigation.

DNA barcoding for identification of fishes in Xiangjiaba reservoir area in the downstream section of the Jinsha river

The downstream section of the Jinsha River is a global biodiversity hotspot for fishes. However, cascade hydropower development has altered the original habitat and has had a huge impact on the area’s fish diversity. To assess the power of DNA barcoding and construct a DNA barcode library for Xiangjiaba reservoir area, 333 fish mitochondrial DNA barcodes (672 bp) employing a fragment of the mitochondrial cytochrome c oxidase subunit I gene (COI) were obtained for 57 species from the Xiangjiaba reservoir area in this study. The intra-specific genetic distances were below 2%, ranging from 0 to 1.69% (mean 0.19%). The interspecific genetic distances within genera ranged from 0.45 to 11.14% (mean 2.98%). The Bayesian inference (BI) tree and neighbor-joining (NJ) tree inferred that all species were unambiguously separated, with strong support values at the terminal branches except the node of Hemibarbus maculatus shared with its adjacent clade. The DNA barcoding gaps could accomplish fish species discrimination. These results revealed that DNA barcoding was an effective tool for the identification of fish species, and that it was possible to construct a robust DNA barcode reference library for fish species in the Xiangjiaba reservoir area. The results will contribute to future efforts in monitoring, conservation, and management of fish resources.

Molecular Identification and Toxin Analysis of Alexandrium spp. in the Beibu Gulf: First Report of Toxic A. tamiyavanichii in Chinese Coastal Waters.

The frequency of harmful algal blooms (HABs) has increased in China in recent years. Information about harmful dinoflagellates and paralytic shellfish toxins (PSTs) is still limited in China, especially in the Beibu Gulf, where PSTs in shellfish have exceeded food safety guidelines on multiple occasions. To explore the nature of the threat from PSTs in the region, eight Alexandrium strains were isolated from waters of the Beibu Gulf and examined using phylogenetic analyses of large subunit (LSU) rDNA, small subunit (SSU) rDNA, and internal transcribed spacer (ITS) sequences. Their toxin composition profiles were also determined using liquid chromatography-tandem mass spectrometry (LC-MS/MS). All eight strains clustered in the phylogenetic tree with A. pseudogonyaulax, A. affine, and A. tamiyavanichii from other locations, forming three well-resolved groups. The intraspecific genetic distances of the three Alexandrium species were significantly smaller than interspecific genetic distances for Alexandrium species. Beibu Gulf isolates were therefore classified as A. pseudogonyaulax, A. affine, and A. tamiyavanichii. No PSTs were identified in A. pseudogonyaulax, but low levels of gonyautoxins (GTXs) 1 to 5, and saxitoxin (STX) were detected in A. tamiyavanichii (a total of 4.60 fmol/cell). The extremely low level of toxicity is inconsistent with PST detection above regulatory levels on multiple occasions within the Beibu Gulf, suggesting that higher toxicity strains may occur in those waters, but were unsampled. Other explanations including biotransformation of PSTs in shellfish and the presence of other PST-producing algae are also suggested. Understanding the toxicity and phylogeny of Alexandrium species provides foundational data for the protection of public health in the Beibu Gulf region and the mitigation of HAB events.

Hydraena (s.str.) dinarica, new species (Coleoptera: Hydraenidae) along with further records of Hydraena spp. from Durmitor National Park, Montenegro and comments on the DNA barcoding problem with the genus.

BackgroundLong-palped Water Beetles were collected during a taxon expedition in Montenegro which involved citizen scientists, students and taxonomists. The material was collected from springs, brooks, fens and the Tara River, at altitudes between 600 m and 1450 m above sea level, using fine-meshed hand-nets and by manual checking of submerged substrates. The morphological species delimitation was supplemented and congruent with mtDNA sequences mainly obtained in the field using the newly-developed MinION-based ONTrack pipeline.New informationThe new species Hydraena dinarica Freitag & de Vries, sp. n. from Durmitor Mt. is described, illustrated and compared in detail to closely-related congeners of the H. saga d'Orchymont, 1930/H. emarginata Rey, 1885 species complex. Five additional species and female specimens of two unidentified morphospecies of the genus were also recorded in the vicinity of Durmitor National Park. New records and the first DNA barcodes for Hydraena biltoni Jäch & Díaz, 2012 (endemic to Montenegro) and H. morio Kiesenwetter, 1849 are provided. Further records of H. nigrita Germar, 1824, H. minutissima Stephens, 1829, H. subintegra Ganglbauer, 1901 and females of two unidentified morphospecies are commented upon. The resulting inter- and intraspecific genetic distances and some observations of low or zero sequence divergence between recently-diverged species of Hydraena Kugelann, 1794 are briefly discussed.

A novel dataset to identify the endemic herpetofauna of the New Caledonia biodiversity hotspot with DNA barcodes

New Caledonia is the smallest global biodiversity hotspot, yet has one of the highest levels of endemism for an insular region of its size. Lizards are the dominant vertebrate fauna, and, while ecologically important, can be difficult to identify and many are in decline due to anthropogenic threats. As an aid to facilitate identification, we generated a near-complete DNA barcode dataset for New Caledonian lizards, consisting of 601 mitochondrial CO1 sequences of 100 of the 107 described lizards, and a number of yet undescribed species. We use this dataset to assess the performance of CO1 in delimiting species recognised by other, more extensive data and in recovering phylogenetic signal. Most species had intraspecific genetic distances ≤3.7%. Most comparisons between described species were at least ~5% divergent, with the exception of three pairwise species comparisons showing interspecific distances > 2.5%. Maximum likelihood CO1 trees of the six most speciose genera recovered each as monophyletic and, although discordant with previously published ND2 trees using quantitative topology tests, showed similar patterns of intraspecific and interspecific divergence, supporting the utility of CO1 in taxonomic identification and species delimitation. Some species showed overlap between intra- and interspecific pairwise distances, suggesting cryptic taxa, a finding also supported by species delimitation analyses using GMYC and mPTP. This dataset not only provides the basis for economical and reliable identification of New Caledonian lizards encountered during biodiversity assessments, but also provides a potential tool for investigating the identity of native lizards and their ecosystem interactions, even from partial remains.

Molecular characterization of Lipoptena cervi from environmental samples collected in Poland.

The activity of Lipoptena cervi has intensified in Poland in recent years. The population genetics of this ectoparasite in Poland has never been described in the literature. The objectives of this study were to investigate the population genetics of L. cervi in selected regions of Poland, to evaluate molecular differences between L. cervi populations, and to determine phylogenetic relationships with other L. cervi sequences obtained in previous studies. In 2019, louse flies were sampled in natural mixed forests in five Polish voivodeships. Seven samples of L. cervi were collected from each voivodeship, and a total of 35 insects were analyzed molecularly. In the first step, Lipoptena spp. were identified to species level under a stereoscopic microscope. A fragment of the rRNA 16S gene was used as a marker to identify L. cervi by the PCR assay. The sequences were assigned accession numbers MT337409 to MT337416. A total of eight haplotypes were identified, two of which were dominant. In the obtained sequences, intraspecific pairwise genetic distances varied between 0.000 and 0.0496 (m = 0.0135; SD = 0.0149; SE = 0.0006; V = 110.11). Mean interpopulation diversity was d = 0.0135 (SE = 0.0027). The acquired nucleotide sequences were highly similar to the sequences from the Czech Republic (MF495940, AF322437), Lithuania (MN889542-MN889544) and Poland (MF541726–MF541729). The similarity with GenBank sequences ranged from 97.24% to 100%. This study revealed two dominant haplotypes of L. cervi in Poland, MT337410 and MT337413. Fragments of the analyzed sequences were detected in only one voivodeship. These findings suggest that the two dominant sequences are the oldest sequences that gave rise to the locally identified haplotypes. The lack of significant correlations with the sequences obtained in regions situated west of the research sites suggests the presence of other genetic populations in Europe.

Distinct barcodes for the Cereal leaf beetles Oulema melanopus and Oulema duftschmidi (Coleoptera: Chrysomelidae), two syntopical sibling species

Oulema melanopus (Linnaeus, 1758) and Oulema duftschmidi (Redtenbacher, 1874) (Coleoptera: Chrysomelidae) are two native West Palaearctic species developing on various cultivated and wild grasses. Along with O. obscura they are considered to be secondary pests of cereal crops. However, local outbreaks have been recorded recently and their status as secondary pests may evolve, especially as the use of broad-spectrum insecticides is now greatly reduced. Oulema melanopus and O. duftschmidi are considered to be sibling species. They are morphologically very close and difficult to distinguish from each other, which makes it difficult to study them. We tested the reliability of the standard barcode fragment (COI) for distinguishing between these species. A total of 92 samples of the two species, covering the majority of their natural range, was sequenced for the barcode fragment and inter- and intraspecific genetic distances were estimated. Our results confirm those of Bezděk & Baselga (2015, Acta Entomol. Mus. Nat. Prag. 55: 273-304) in that this marker cannot differentiate between all the species of the Oulema melanopus complex, which in the Mediterranean basin contains several described and possibly some undescribed cryptic species. However, this marker may be useful in an agricultural context in areas where only O. melanopus and O. duftschmidi occur (such as in cereal crops in France) where it can be used to reliably and rapidly separate all stages of these two taxa and can therefore help in studying their ecology and dynamics.

Description and identification of tuna larvae based on genetic and morphological analysis

Biological information of tuna larvae is essential for understanding spawning ground and time. However, morphological identification of fish larvae is difficult and questionable. Meanwhile, genetic identification is useful and helpful to confirm the morphological identification result. This study purpose is to confirm a morphological identification result with genetic identification and determine the tuna species exactly. Specimens were obtained from a survey conducted in the Banda Sea in October 2018. Samples were collected by a bongo net towed obliquely. The result of morphological identification is showed that the larvae having 37 to 42 myomeres, preopercular spine developed, moderate in-depth, moderate to large head and eyes identified as Thunnini larvae. The morphological result is supported by the genetic result, even though the larvae are not from the genus of Thunnus. The samples were identified as Katsuwonus pelamis, Euthynnus affinis, and Petenia splendida. The intraspecific pairwise genetic distance for each species of K. pelamis, E. affinis, and P. splendida is 4.4 %; 0.5 %; and 22.3%, respectively. Meanwhile, the average pairwise genetic distance between the three species ranges from 16.2% to 33.3%.

COⅠ and 16S rDNA Sequence Identification of Common Necrophagous Flies in Fujian Province

Objective To identify the species of common necrophagous flies in Fujian Province by gene fragment sequences of mitochondrial cytochrome c oxidase subunit Ⅰ （COⅠ） and 16S ribosomal deoxyribonucleic acid （16S rDNA）, and to explore the identification efficacy of these two molecular markers. Methods In total 22 common necrophagous flies were collected from the death scenes in 9 different regions in Fujian Province and DNA was extracted from the flies after morphological identification. The gene fragments of COⅠ and 16S rDNA were amplified and sequenced. All the sequences were uploaded to GeneBank and BLAST and MEGA 10.0 software were used to perform sequence alignment, homology analysis and intraspecific and interspecific genetic distance analysis. The phylogenetic trees of DNA fragment sequences of COⅠ and 16S rDNA of common necrophagous flies in Fujian Province were established by unweighted pair-group method with arithmetic means （UPGMA）, respectively. Results The flies were classified into 6 species, 5 genera and 3 families by morphological identification. The results of gene sequence analysis showed that the average number of interspecific and intraspecific genetic distance of 16S rDNA ranged from 1.8% to 8.9% and 0.0% to 2.4%, respectively. The average number of interspecific and intraspecific genetic distance of COⅠ ranged from 7.2% to 13.6% and 0.0% to 6.3%, respectively. Conclusion The gene sequences of COⅠ and 16S rDNA can accurately identify the species of different necrophagous flies, and 16S rDNA showed higher value in species identification of common calliphoridae necrophagous flies in Fujian Province.

Eight new species of the genus Anaplecta Burmeister, 1838 (Blattodea: Blattoidea: Anaplectidae) from China based on molecular and morphological data

In this study, we examine 500 specimens of Anaplecta collected from China, of which 26 samples were used for COI sequencing. We confirm eight new species, i.e., Anaplecta corneola Deng & Che sp. nov., Anaplecta staminiformis Deng & Che sp. nov., Anaplecta arcuata Deng & Che sp. nov., Anaplecta strigata Deng & Che sp. nov., Anaplecta furcata Deng & Che sp. nov., Anaplecta cruciata Deng & Che sp. nov., Anaplecta nigra Deng & Che sp. nov. and Anaplecta bicolor Deng & Che sp. nov. based on morphological and molecular data using ABGD and GMYC analyses. The results of ABGD and GMYC were basically consistent with the morphospecies of Anaplecta. The intraspecific and interspecific genetic distances of Anaplecta ranged from 0 to 6.6% and 16.8% to 31.8%, respectively. We found the male genitalia of Anaplecta to exhibit intraspecific variation, especially in the phallomeres.

New records of cascade frogs of the genus <i>Odorrana </i> (Amphibia: Anura: Ranidae) from Vietnam

Odorrana fengkaiensis was recorded for the first time from Vietnam based on a new amphibian collection from Cao Bang, Bac Kan, Bac Giang, Quang Ninh provinces. Sequences of the specimens from Vietnam were identical to those of the paratypes of O. fengkaiensis from China. In addition, O. trankieni was newly recorded from Hoa Binh and Bac Giang provinces, Northern Vietnam, including the first description of female individuals. Genetic divergence between O. trankieni from Vietnam and its sister species, O. nasuta from China, ranged from 1.42 to 2.66%. The intra-specific genetic distance within O. trankieni ranged from 0.68 to 1.88% between the northeastern and northwestern populations.

Morphological plasticity of the Aden Nerite, Nerita adenensis Mienis, 1978 (Gastropoda: Cycloneritida: Neritidae)

ABSTRACT Fourteen morphotypes of the Aden nerite (Nerita adenensis) from Chabahar Bay, the northern Gulf of Oman, were sequenced for mitochondrial cytochrome c oxidase I and nuclear ATPSα genes. COI sequences across all morphotypes yielded uncorrected intraspecific genetic distances (p) of between 0.013 and 0.017, whereas interspecific p was 0.11–0.16 among congeneric species. For ATPSα, across all morphotypes, p was 0.00–0.03, whereas between N. adenensis and congeneric species, p was 0.07–0.19. Morphological analysis revealed a high degree of plasticity in shell colour, pattern and structure, while characters of the operculum and crenulations on the outer lip were more stable. Our descriptions provided here will be helpful in identifying this problematic taxon, which our genetic analyses show is best regarded as a single variable biological species.

An update on the indigenous vascular flora of sub-Antarctic Marion Island: taxonomic changes, sequences for DNA barcode loci, and genome size data

The flora of sub-Antarctic Marion Island forms part of the unique South Indian Ocean Biogeographic Province, and is under threat from climate change and invasive species. Current information on the flora is necessary to rapidly identify and manage future changes. We conducted a literature search on the taxonomy of indigenous vascular plant species on Marion Island and found nomenclatural changes following taxonomic revisions for Austroblechnum penna-marina (Poir.) Gasper & V.A.O.Dittrich, Carex dikei (Nelmes) K.L.Wilson, Leptinella plumosa Hook.f., Notogrammitis crassior (Kirk) Parris, Phlegmariurus saururus (Lam.) B.Ollg., and Polypogon magellanicus (Lam.) Finot. Additionally, Ranunculus moseleyi Hook.f. was removed from our species checklist due to its long absence in floristic surveys, leaving 21 species in the indigenous vascular plant flora present on Marion Island. We also amplified and sequenced the universal plant barcoding loci rbcL and matK for 19 and 13 species, respectively, and found that ample interspecific genetic distance and minimal intraspecific genetic distance allowed for easy discrimination between species. Lastly, we obtained genome size estimates using flow cytometry for 12 species. Mean 2C genome size for species on Marion Island ranged from 0.44 to 21.44 pg, which is on the lower end of the known range for vascular plant species. We detected two distinct cytotypes in Poa cookii (Hook.f.) Hook.f. and one cytotype in all other species measured.

DNA Barcoding and geographical scale effect: The problems of undersampling genetic diversity hotspots.

DNA barcoding identification needs a good characterization of intraspecific genetic divergence to establish the limits between species. Yet, the number of barcodes per species is many times low and geographically restricted. A poor coverage of the species distribution range may hamper identification, especially when undersampled areas host genetically distinct lineages. If so, the genetic distance between some query sequences and reference barcodes may exceed the maximum intraspecific threshold for unequivocal species assignation. Taking a group of Quercus herbivores (moths) in Europe as model system, we found that the number of DNA barcodes from southern Europe is proportionally very low in the Barcoding of Life Data Systems. This geographical bias complicates the identification of southern query sequences, due to their high intraspecific genetic distance with respect to barcodes from higher latitudes. Pairwise intraspecific genetic divergence increased along with spatial distance, but was higher when at least one of the sampling sites was in southern Europe. Accordingly, GMYC (General Mixed Yule Coalescent) single‐threshold model retrieved clusters constituted exclusively by Iberian haplotypes, some of which could correspond to cryptic species. The number of putative species retrieved was more reliable than that of multiple‐threshold GMYC but very similar to results from ABGD and jMOTU. Our results support GMYC as a key resource for species delimitation within poorly inventoried biogeographic regions in Europe, where historical factors (e.g., glaciations) have promoted genetic diversity and singularity. Future European DNA barcoding initiatives should be preferentially performed along latitudinal gradients, with special focus on southern peninsulas.