Systematics and evolutionary dynamics of insect-fern interactions in the specialized fern-spore feeding Cuprininae (Lepidoptera, Stathmopodidae).

Fern-spore-feeding (FSF) is rare and found in only four families of Lepidoptera. Stathmopodidae is the most speciose family that contains FSF species, and its subfamily Cuprininae exclusively specializes on FSF. However, three species of Stathmopodinae also specialize on FSF. To better understand the evolutionary history of FSF and, more generally, the significance of specialization on a peculiar host, a phylogenetic and taxonomic revision for this group is necessary. We reconstructed the most comprehensive molecular phylogeny, including one mitochondrial and four nuclear genes, of Stathmopodidae to date, including 137 samples representing 62 species, with a particular focus on the FSF subfamily, Cuprininae, including 33 species (41% of named species) from 6 of the 7 Cuprininae genera. Species from two other subfamilies, Stathmopodinae and Atkinsoniinae, were also included. We found that FSF evolved only once in Stathmopodidae and that the previous hypothesis of multiple origins of FSF was misled by inadequate taxonomy. Moreover, we showed that (1) speciation/extinction rates do not differ significantly between FSF and non-FSF groups and that (2) oligophage is the ancestral character state in Cuprininae. We further revealed that a faster rate of accumulating specialists over time, and thus a higher number of specialists, was achieved by a higher transition rate from oligophagages to specialists compared to the transition rate in the opposite direction. We finish by describing three new genera, Trigonodagen. nov., Petalagen. nov., and Pediformisgen. nov., and revalidating five genera: Cuprina, Calicotis, Thylacosceles, Actinoscelis, Thylacosceloides in Cuprininae, and we provide an updated taxonomic key to genera and a revised global checklist of Cuprininae.

Gelechiaomelkoi sp. nov. - a new species from the Russian Altai Mountains related to the Nearctic Gelechiamandella Busck, 1904 (Lepidoptera, Gelechiidae), with a synopsis of Gelechia from the Altai Republic of Russia.

Gelechiaomelkoi sp. nov. is described from the Ukok plateau and South Chuisky ridge in the Altai Mountains of Russia. The adult of the new species, including its male genitalia, is illustrated and compared with species most similar in morphology and DNA barcodes-G.sororculella (Hübner, 1817) and G.jakovlevi Krulikovsky, 1905 from the Palaearctic region, as well as G.mandella Busck, 1904 from Canada. This last species is redescribed based on adult specimens, including the genitalia of both sexes, and a lectotype is designated. Gelechiasirotina Omelko, 1986 is recorded from the Altai Republic for the first time. An updated list of six species of Gelechia from the Altai Mountains of Russia is given. Dorsal habitus photographs of all species are provided. The male genitalia of the lectotype of G.jakovlevi is illustrated for the first time.

DNA barcode library for European Gelechiidae (Lepidoptera) suggests greatly underestimated species diversity.

For the first time, a nearly complete barcode library for European Gelechiidae is provided. DNA barcode sequences (COI gene - cytochrome c oxidase 1) from 751 out of 865 nominal species, belonging to 105 genera, were successfully recovered. A total of 741 species represented by specimens with sequences ≥ 500bp and an additional ten species represented by specimens with shorter sequences were used to produce 53 NJ trees. Intraspecific barcode divergence averaged only 0.54% whereas distance to the Nearest-Neighbour species averaged 5.58%. Of these, 710 species possessed unique DNA barcodes, but 31 species could not be reliably discriminated because of barcode sharing or partial barcode overlap. Species discrimination based on the Barcode Index System (BIN) was successful for 668 out of 723 species which clustered from minimum one to maximum 22 unique BINs. Fifty-five species shared a BIN with up to four species and identification from DNA barcode data is uncertain. Finally, 65 clusters with a unique BIN remained unidentified to species level. These putative taxa, as well as 114 nominal species with more than one BIN, suggest the presence of considerable cryptic diversity, cases which should be examined in future revisionary studies.

The myth of monophagy in Paralobesia (Lepidoptera: Tortricidae)? A new species feeding on Cypripedium reginae (Orchidaceae).

The genus Paralobesia Obraztsov, 1953 is found primarily in eastern North America and consists of 18 described and several undescribed species. Prior to 1900, all North American Paralobesia were assumed to be P. viteana (Clemens). However, rearing experiments by William Kearfott in the early 1900s suggested that species of Paralobesia were monophagous and could be separated by host. Recently, a species of Paralobesia was reared from showy lady's slipper, Cypripedium reginae Walter (Orchidaceae), during a study of two populations of this orchid in eastern Ontario and southwestern Québec. Although originally assumed to be P. cypripediana (Forbes), which was described from specimens reared from Cypripedium in Manitoba, DNA barcode data and genital morphology confirmed that this was a new species similar to P. cypripediana and P. monotropana (Heinrich). Herein, we describe P. marilynae, sp. n., and provide specifics of its discovery and life history. Rearing records indicate that Paralobesia can span the range from strictly monophagous to polyphagous, even for very similar species with similar feeding habits, and that host records should be combined with morphological and molecular data when circumscribing species in this genus. This work is part of a complete systematic revision of Paralobesia currently in progress.

A DNA Barcode Library for North American Pyraustinae (Lepidoptera: Pyraloidea: Crambidae).

Although members of the crambid subfamily Pyraustinae are frequently important crop pests, their identification is often difficult because many species lack conspicuous diagnostic morphological characters. DNA barcoding employs sequence diversity in a short standardized gene region to facilitate specimen identifications and species discovery. This study provides a DNA barcode reference library for North American pyraustines based upon the analysis of 1589 sequences recovered from 137 nominal species, 87% of the fauna. Data from 125 species were barcode compliant (>500bp, <1% n), and 99 of these taxa formed a distinct cluster that was assigned to a single BIN. The other 26 species were assigned to 56 BINs, reflecting frequent cases of deep intraspecific sequence divergence and a few instances of barcode sharing, creating a total of 155 BINs. Two systems for OTU designation, ABGD and BIN, were examined to check the correspondence between current taxonomy and sequence clusters. The BIN system performed better than ABGD in delimiting closely related species, while OTU counts with ABGD were influenced by the value employed for relative gap width. Different species with low or no interspecific divergence may represent cases of unrecognized synonymy, whereas those with high intraspecific divergence require further taxonomic scrutiny as they may involve cryptic diversity. The barcode library developed in this study will also help to advance understanding of relationships among species of Pyraustinae.

The Lepidoptera of White Sands National Monument, Otero County, New Mexico, USA 10. A remarkable new white species of Chionodes Hübner (Gelechiidae).

The U.S. National Park Service initiated a 10-year study, in late 2006, of the Lepidoptera at White Sands National Monument, Otero County, New Mexico. Chionodes bustosorum sp. n., described here, was discovered in 2010, during the third year of the study. The male imago and male genitalia are illustrated, and its DNA barcode is compared to that of seven other species of Chionodes from western North America.

Coleophora ericarnella Baldizzone, a new species of the C. pyrrhulipennella group (Lepidoptera: Coleophoridae) from the South-Eastern Alps.

Coleophora ericarnella Baldizzone, n. sp. is described from the East-Central Alps region (Italy and Slovenia). It belongs to the C. pyrrhulipennella group. Its larval host plant is Erica carnea (Ericaceae). The adult habitus and genitalia as well as its larval case are illustrated and compared with those of Coleophora pulchripennella Baldizzone, 2011 and C. pyrrhulipennella Zeller, 1849. DNA barcodes also distinguish all three species from each other in congruence with genitalia morphology.

Systematics and biology of some species of Micrurapteryx Spuler (Lepidoptera, Gracillariidae) from the Holarctic Region, with re-description of M. caraganella (Hering) from Siberia.

During a DNA barcoding campaign of leaf-mining insects from Siberia, a genetically divergent lineage of a gracillariid belonging to the genus Micrurapteryx was discovered, whose larvae developed on Caragana Fabr. and Medicago L. (Fabaceae). Specimens from Siberia showed similar external morphology to the Palearctic Micrurapteryx gradatella and the Nearctic Parectopa occulta but differed in male genitalia, DNA barcodes, and nuclear genes histone H3 and 28S. Members of this lineage are re-described here as Micrurapteryx caraganella (Hering, 1957), comb. n., an available name published with only a brief description of its larva and leaf mine. Micrurapteryx caraganella is widely distributed throughout Siberia, from Tyumen oblast in the West to Transbaikalia in the East. Occasionally it may severely affect its main host, Caragana arborescens Lam. This species has been confused in the past with Micrurapteryx gradatella in Siberia, but field observations confirm that Micrurapteryx gradatella exists in Siberia and is sympatric with Micrurapteryx caraganella, at least in the Krasnoyarsk region, where it feeds on different host plants (Vicia amoena Fisch. and Vicia sp.). In addition, based on both morphological and molecular evidence as well as examination of type specimens, the North American Parectopa occulta Braun, 1922 and Parectopa albicostella Braun, 1925 are transferred to Micrurapteryx as Micrurapteryx occulta (Braun, 1922), comb. n. with albicostella as its junior synonym (syn. n.). Characters used to distinguish Micrurapteryx from Parectopa are presented and illustrated. These findings provide another example of the potential of DNA barcoding to reveal overlooked species and illuminate nomenclatural problems.

Coleophoranepetellae Baldizzone & Nel, a new species of the C.lixella group (Lepidoptera, Coleophoridae) from France and Italy.

Coleophoranepetellae Baldizzone & Nel, sp. n. is described from the southern Alps (Italy and France). It belongs to the Coleophoralixella species group. Its host plants are Nepetanepetella L. (Lamiaceae) and an unidentified Poaceae. The fifth instar larva, its case, the adult habitus, and genitalia are illustrated. The species is compared to Coleophoranevadella Baldizzone, 1985, here newly confirmed from France and whose larvae feed on Nepetalatifolia DC. in the Eastern Pyrénées. DNA barcodes are shown to be distinct and congruent with morphological differences among species of the lixella group. Barcodes revealed that Coleophoratricolor Walsingham, 1889, formerly known only from Great Britain, is also present in France and Greece.

RésuméColeophoranepetellae Baldizzone & Nel, sp. n. du groupe de Coleophoralixella Zeller, 1849., est décrite des Alpes méridionales (Italie et France). La plante-hôte de ponte est la Lamiacée Nepetanepetella L. et celle de la larve à maturité est une espèce non-identifiée de Poaceae. La chenille L5 et son fourreau, l'habitus et les genitalia mâles et femelles sont figurés. L'espèce est comparée à Coleophoranevadella Baldizzone, 1985, espèce ici confirmée pour la France où elle est inféodée à Nepetalatifolia DC. dans les Pyrénées-Orientales. Enfin, la validité de cette nouvelle espèce est confirmée par l'étude des codes-barres ADN du groupe lixella. Les codes-barres ont aussi révélé la présence de Coleophoratricolor Walsingham, 1889 en France et en Grèce alors que l'espèce n'était auparavant connue que de la Grande-Bretagne.

Plutella australiana (Lepidoptera, Plutellidae), an overlooked diamondback moth revealed by DNA barcodes.

The genus Plutella was thought to be represented in Australia by a single introduced species, Plutella xylostella (Linnaeus), the diamondback moth. Its status as a major pest of cruciferous crops, and the difficulty in developing control strategies has motivated broad-ranging studies on its biology. Prior genetic work has generally supported the conclusion that populations of this migratory species are connected by substantial gene flow. However, the present study reveals the presence of two genetically divergent lineages of this taxonin Australia. One shows close genetic and morphological similarity with the nearly cosmopolitan Plutella xylostella. The second lineage possesses a similar external morphology, but marked sequence divergence in the barcode region of the cytochrome c oxidase I gene, coupled with clear differences in genitalia. As a consequence, members of this lineage are described as a new species, Plutella australiana Landry & Hebert, which is broadly distributed in the eastern half of Australia.

A molecular phylogeny for yponomeutoidea (insecta, Lepidoptera, ditrysia) and its implications for classification, biogeography and the evolution of host plant use.

BACKGROUND: Yponomeutoidea, one of the early-diverging lineages of ditrysian Lepidoptera, comprise about 1,800 species worldwide, including notable pests and insect-plant interaction models. Yponomeutoids were one of the earliest lepidopteran clades to evolve external feeding and to extensively colonize herbaceous angiosperms. Despite the group's economic importance, and its value for tracing early lepidopteran evolution, the biodiversity and phylogeny of Yponomeutoidea have been relatively little studied. METHODOLOGY/PRINCIPAL FINDINGS: Eight nuclear genes (8 kb) were initially sequenced for 86 putative yponomeutoid species, spanning all previously recognized suprageneric groups, and 53 outgroups representing 22 families and 12 superfamilies. Eleven to 19 additional genes, yielding a total of 14.8 to 18.9 kb, were then sampled for a subset of taxa, including 28 yponomeutoids and 43 outgroups. Maximum likelihood analyses were conducted on data sets differing in numbers of genes, matrix completeness, inclusion/weighting of synonymous substitutions, and inclusion/exclusion of "rogue" taxa. Monophyly for Yponomeutoidea was supported very strongly when the 18 "rogue" taxa were excluded, and moderately otherwise. Results from different analyses are highly congruent and relationships within Yponomeutoidea are well supported overall. There is strong support overall for monophyly of families previously recognized on morphological grounds, including Yponomeutidae, Ypsolophidae, Plutellidae, Glyphipterigidae, Argyresthiidae, Attevidae, Praydidae, Heliodinidae, and Bedelliidae. We also assign family rank to Scythropiinae (Scythropiidae stat. rev.), which in our trees are strongly grouped with Bedelliidae, in contrast to all previous proposals. We present a working hypothesis of among-family relationships, and an informal higher classification. Host plant family associations of yponomeutoid subfamilies and families are non-random, but show no trends suggesting parallel phylogenesis. Our analyses suggest that previous characterizations of yponomeutoids as predominantly Holarctic were based on insufficient sampling. CONCLUSIONS/SIGNIFICANCE: We provide the first robust molecular phylogeny for Yponomeutoidea, together with a revised classification and new insights into their life history evolution and biogeography.

Shared but overlooked: 30 species of Holarctic Microlepidoptera revealed by DNA barcodes and morphology.

This study reports 30 species of Lepidoptera previously known from either the Palearctic or the Nearctic that are newly recorded as Holarctic. For 28 of these species, their intercontinental distributions were initially detected through DNA barcode analysis and subsequently confirmed by morphological examination; two Palearctic species were first detected in North America through morphology and then barcoded. When possible, the origin and status of each species (introduced, overlooked Holarctic species, or unknowingly re-described) is discussed, and its morphology is diagnosed and illustrated. The species involved include Tineidae: Scardia amurensis Zagulajev, Triaxomera parasitella (Hübner), Nemapogon cloacella (Haworth), Elatobia montelliella (Schantz), Tinea svenssoni Opheim; Gracillariidae: Caloptilia suberinella (Tengström), Parornix betulae (Stainton); Phyllonorycter maestingella (Müller); Yponomeutidae: Paraswammerdamia albicapitella (Scharfenberg), P. conspersella (Tengström); Plutellidae: Plutella hyperboreella Strand; Lyonetiidae: Lyonetia pulverulentella Zeller; Autostichidae: Oegoconia deauratella (Herrich-Schäffer), O. novimundi (Busck); Blastobasidae: Blastobasis glandulella (Riley), B. maroccanella (Amsel), B. tarda Meyrick; Depressariidae: Agonopterix conterminella (Zeller), Depressaria depressana (F.); Coleophoridae: Coleophora atriplicis Meyrick, C. glitzella Hofmann, C. granulatella Zeller, C. texanella Chambers, C. vitisella Gregson; Scythrididae: Scythris sinensis (Felder & Rogenhofer); Gelechiidae: Altenia perspersella (Wocke), Gnorimoschema jalavai Povolný, Scrobipalpa acuminatella (Sircom), Sophronia gelidella Nordman; Choreutidae: Anthophila fabriciana (L.); and Tortricidae: Phiaris bipunctana (F.). These cases of previously unrecognized faunal overlap have led to their redescription in several instances. Five new synonyms are proposed: Blastobasis glandulella (Riley, 1871) = B. huemeri Sinev, 1993, syn. nov.; B. tarda Meyrick, 1902 = Neoblastobasis ligurica Nel & Varenne, 2004, syn. nov.; Coleophora atriplicis Meyrick, 1928 = C. cervinella McDunnough, 1946, syn. nov.; C. texanella Chambers, 1878 = C. coxi Baldizzone & van der Wolf, 2007, syn. nov., and = C. vagans Walsingham, 1907, syn. nov. Lectotypes are designated for Blastobasis tarda Meyrick and Coleophora texanella Chambers. Type specimens were examined where pertinent to establish new synonymies. We identify 12 previously overlooked cases of species introductions, highlighting the power of DNA barcoding as a tool for biosurveillance.

Allopatry as a gordian knot for taxonomists: patterns of DNA barcode divergence in arctic-alpine lepidoptera.

Many cold adapted species occur in both montane settings and in the subarctic. Their disjunct distributions create taxonomic complexity because there is no standardized method to establish whether their allopatric populations represent single or different species. This study employs DNA barcoding to gain new perspectives on the levels and patterns of sequence divergence among populations of 122 arctic-alpine species of Lepidoptera from the Alps, Fennoscandia and North America. It reveals intraspecific variability in the barcode region ranging from 0.00-10.08%. Eleven supposedly different species pairs or groups show close genetic similarity, suggesting possible synonymy in many cases. However, a total of 33 species show evidence of cryptic diversity as evidenced by the presence of lineages with over 2% maximum barcode divergence in Europe, in North America or between the two continents. Our study also reveals cases where taxonomic names have been used inconsistently between regions and exposes misidentifications. Overall, DNA barcodes have great potential to both increase taxonomic resolution and to make decisions concerning the taxonomic status of allopatric populations more objective.

A review of the North American genus Epimartyria (Lepidoptera, Micropterigidae) with a discussion of the larval plastron.

The indigenous North American micropterigid genus Epimartyria Walsingham,1898 is revised. Three species are recognized, including Epimartyria auricrinella Walsingham, 1898 which occurs widely over much of the northeastern United States and Canada, a new species, Epimartyria bimaculella Davis & Landry from northwestern United States and Canada, and Epimartyria pardella (Walsingham, 1880) from northern California to northern Oregon. The larva of Epimartyria auricrinella is described in detail, supplemented with illustrations of the external structure of the larval integument. The larval plastron is described and illustrated for Epimartyria, and this is compared with the plastrons of Neomicropteryx Issiki, 1931 and Micropterix Hübner, 1825. COI barcode sequences show that the three species are genetically distinct, congruent with morphological differences. Marked haplotype divergence within some Epimartyria auricrinella populations appears to be unrelated to morphology, geography or phenology.

Pyrosequencing for mini-barcoding of fresh and old museum specimens.

DNA barcoding is an effective approach for species identification and for discovery of new and/or cryptic species. Sanger sequencing technology is the method of choice for obtaining standard 650 bp cytochrome c oxidase subunit I (COI) barcodes. However, DNA degradation/fragmentation makes it difficult to obtain a full-length barcode from old specimens. Mini-barcodes of 130 bp from the standard barcode region have been shown to be effective for accurate identification in many animal groups and may be readily obtained from museum samples. Here we demonstrate the application of an alternative sequencing technology, the four-enzymes single-specimen pyrosequencing, in rapid, cost-effective mini-barcode analysis. We were able to generate sequences of up to 100 bp from mini-barcode fragments of COI in 135 fresh and 50 old Lepidoptera specimens (ranging from 53-97 year-old). The sequences obtained using pyrosequencing were of high quality and we were able to robustly match all the tested pyro-sequenced samples to their respective Sanger-sequenced standard barcode sequences, where available. Simplicity of the protocol and instrumentation coupled with higher speed and lower cost per sequence than Sanger sequencing makes this approach potentially useful in efforts to link standard barcode sequences from unidentified specimens to known museum specimens with only short DNA fragments.

DNA barcodes for 1/1000 of the animal kingdom.

This study reports DNA barcodes for more than 1300 Lepidoptera species from the eastern half of North America, establishing that 99.3 per cent of these species possess diagnostic barcode sequences. Intraspecific divergences averaged just 0.43 per cent among this assemblage, but most values were lower. The mean was elevated by deep barcode divergences (greater than 2%) in 5.1 per cent of the species, often involving the sympatric occurrence of two barcode clusters. A few of these cases have been analysed in detail, revealing species overlooked by the current taxonomic system. This study also provided a large-scale test of the extent of regional divergence in barcode sequences, indicating that geographical differentiation in the Lepidoptera of eastern North America is small, even when comparisons involve populations as much as 2800 km apart. The present results affirm that a highly effective system for the identification of Lepidoptera in this region can be built with few records per species because of the limited intra-specific variation. As most terrestrial and marine taxa are likely to possess a similar pattern of population structure, an effective DNA-based identification system can be developed with modest effort.

Integration of DNA barcoding into an ongoing inventory of complex tropical biodiversity.

Inventory of the caterpillars, their food plants and parasitoids began in 1978 for today's Area de Conservacion Guanacaste (ACG), in northwestern Costa Rica. This complex mosaic of 120 000 ha of conserved and regenerating dry, cloud and rain forest over 0-2000 m elevation contains at least 10 000 species of non-leaf-mining caterpillars used by more than 5000 species of parasitoids. Several hundred thousand specimens of ACG-reared adult Lepidoptera and parasitoids have been intensively and extensively studied morphologically by many taxonomists, including most of the co-authors. DNA barcoding - the use of a standardized short mitochondrial DNA sequence to identify specimens and flush out undisclosed species - was added to the taxonomic identification process in 2003. Barcoding has been found to be extremely accurate during the identification of about 100 000 specimens of about 3500 morphologically defined species of adult moths, butterflies, tachinid flies, and parasitoid wasps. Less than 1% of the species have such similar barcodes that a molecularly based taxonomic identification is impossible. No specimen with a full barcode was misidentified when its barcode was compared with the barcode library. Also as expected from early trials, barcoding a series from all morphologically defined species, and correlating the morphological, ecological and barcode traits, has revealed many hundreds of overlooked presumptive species. Many but not all of these cryptic species can now be distinguished by subtle morphological and/or ecological traits previously ascribed to 'variation' or thought to be insignificant for species-level recognition. Adding DNA barcoding to the inventory has substantially improved the quality and depth of the inventory, and greatly multiplied the number of situations requiring further taxonomic work for resolution.

Combining DNA barcoding and morphological analysis to identify specialist floral parasites (Lepidoptera: Coleophoridae: Momphinae: Mompha).

Close interactions between insects and plants have played a major role in the evolution of both these diverse groups of organisms. Studying these interactions, however, can be difficult because many insects, especially parasites, impinge most strongly on plants during larval stages when they are morphologically difficult to identify, and many belong to diverse groups for which most species remain undescribed. We used DNA barcoding to identify nondescript lepidopteran larvae that regularly parasitize flower buds of the coastal dune endemic Camissoniopsis cheiranthifolia (Onagraceae). We obtained cytochrome oxidase 1 mitochondrial DNA sequences from 201 parasite specimens from across the host geographical range. The Barcode of Life Database Identification System combined with Bayesian analysis grouped all 15 parasite haplotypes in a distinct, monophyletic clade within the genus Mompha (Lepidoptera: Coleophoridae: Momphinae), a group known to be host specialists on plants of the Onagraceae. Species identity and phylogenetic affinities within Mompha could not be confirmed because few barcode sequences exist from this diverse and poorly known group of moths. However, morphological analysis, including detailed dissection of genitalia for a subsample of 23 reared adults and comparison with known species of Mompha, also indicated that the larvae parasitizing C. cheiranthifolia constitute a distinct and undescribed species within this genus. Knowing that floral parasitism of C. cheiranthifolia involves a single, putatively host-specific microlepidopteran greatly facilitates formulating and testing hypotheses concerning how floral parasitism has promoted the evolution of striking floral diversity within this species. More generally, DNA barcoding combined with morphological analysis can greatly hasten identification of problematic specimens and enhance our understanding of the diversity, ecology and evolution of plant-insect interactions.

A universal DNA mini-barcode for biodiversity analysis.

BACKGROUND: The goal of DNA barcoding is to develop a species-specific sequence library for all eukaryotes. A 650 bp fragment of the cytochrome c oxidase 1 (CO1) gene has been used successfully for species-level identification in several animal groups. It may be difficult in practice, however, to retrieve a 650 bp fragment from archival specimens, (because of DNA degradation) or from environmental samples (where universal primers are needed). RESULTS: We used a bioinformatics analysis using all CO1 barcode sequences from GenBank and calculated the probability of having species-specific barcodes for varied size fragments. This analysis established the potential of much smaller fragments, mini-barcodes, for identifying unknown specimens. We then developed a universal primer set for the amplification of mini-barcodes. We further successfully tested the utility of this primer set on a comprehensive set of taxa from all major eukaryotic groups as well as archival specimens. CONCLUSION: In this study we address the important issue of minimum amount of sequence information required for identifying species in DNA barcoding. We establish a novel approach based on a much shorter barcode sequence and demonstrate its effectiveness in archival specimens. This approach will significantly broaden the application of DNA barcoding in biodiversity studies.