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Dichrorampha Dinarica, New Species, a Century of Confusion in European Lepidopterology (Lepidoptera: Tortricidae) Resolved by Combining Morphology and DNA Barcoding

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Dichrorampha Dinarica, New Species, a Century of Confusion in European Lepidopterology (Lepidoptera: Tortricidae) Resolved by Combining Morphology and DNA Barcoding Zootaxa 3389: 41–50 (2012) ISSN 1175-5326 (print edition) www.mapress.com/zootaxa/ Article ZOOTAXA Copyright © 2012 · Magnolia Press ISSN 1175-5334 (online edition) Dichrorampha dinarica, new species, a century of confusion in European lepidopterology (Lepidoptera: Tortricidae) resolved by combining morphology and DNA barcoding PETER HUEMER1, 4, BOYAN ZLATKOV2 & JOAQUIN BAIXERAS3 1Tiroler Landesmuseen Betriebsgesellschaft m.b.H., Naturwissenschaftliche Sammlungen, Feldstr. 11 a, A-6020 Innsbruck, Austria. E-mail: [email protected] 2Department of Zoology and Anthropology, Faculty of Biology, Sofia University "St. Kliment Ohridski, 8 Dragan Tsankov Blvd., 1164 Sofia, Bulgaria. E-mail: [email protected] 3Cavanilles Institute of Biodiversity and Evolutionary Biology, University of Valencia, P.O. Box 22085, 46071-Valencia, Spain E-mail: [email protected] 4 Corresponding author Abstract Dichrorampha dinarica, new species, is described from the Dinaric mountains (Macedonia, Bosnia and Herzegovina). The new species was formerly confused with D. ligulana (Herrich-Schäffer, 1851) from the Alps and D. rilana Drenowsky, 1909 from the Rila mountains (Bulgaria). Therefore, a re-description of D. rilana is provided based on topotypical specimens. The diag- nostic morphological characters of the involved species, supported by significant divergence of the mtDNA barcode (COI frag- ment of 658 bp), are discussed in detail. Key words: Lepidoptera, Tortricidae, Dichrorampha, misidentifications, new species, Europe, high mountains, endemism, DNA barcode Introduction In the context of the current global biodiversity crisis, a comprehensive inventory of species is one of the greatest challenges facing taxonomists. Our knowledge of species diversity of mega-diverse insect orders such as Lepidoptera is still far from complete, with estimates of the described species richness varying from 157,000 (Nieukerken et al. 2011) to 175,000 species (Kristensen & Skalski 1999). The Lepidoptera fauna of Europe encompasses about 9500 described species (Lopez-Vaamonde et al. 2010) and is considered to be the world´s best- known fauna. Alpha-taxonomy is thought to be largely resolved in most groups. However, despite of the long history of exploration, encompassing more than 200 years, new species are still regularly discovered and described, particularly from easternmost Europe, the Mediterranean area, and the lesser explored mountain systems. Accessibility of remote areas as well as recently established molecular methods have increased considerably such discoveries (Huemer & Hausmann 2009, Huemer & Hebert 2011, Lopez-Vaamonde et al. 2011). The European Alps and other alpic mountain systems in particular seem to harbor examples of cryptic diversity and/or overlooked species, many of them locally endemic (Huemer 2011). The identities of other species have been confused over decades resulting in misidentifications (Mutanen et al. 2012, Segerer et al. 2011). In this paper we discuss an example of long standing confusion within the genus Dichrorampha and its taxonomic and faunistic consequences. Dichrorampha is a well known and diverse genus of Tortricidae radiated primarily on Asteraceae with a mainly Holarctic distribution. The world catalogue (Baixeras et al. 2010) lists 114 described species, 53 of which are known from Europe (Karsholt & Nieukerken 2011). Few supplementing taxa have been described recently, among them the western alpine D. tarmanni (Huemer 2009), which is not yet incorporated in the aforementioned catalogue. During an excursion to the high mountains of Macedonia (Mount Korab) we collected a series of a species which has been known for more than a century as either D. ligulana or D. rilana. Careful re-examination of Accepted by J.W. Brown: 22 May 2012; published: 16 Jul. 2012 41 literature and the still traceable material, as well as additional samples of D. rilana indicate a hitherto undescribed species from the Dinaric mountain chain. Material and methods We examined about 70 dried, pinned, and partially set specimens belonging to the three species which are involved into this study. Genitalia preparation followed standardized techniques (Robinson 1976). The vesica eversions were made using the method of Zlatkov (2011). Photographs of adults were taken through an Olympus SZX 10 binocular microscope and Olympus E 3 digital camera and processed with Helicon Focus 4.3 software. Images were edited using Adobe Photoshop Lightroom 2.3 software. Genitalia were photographed with an Olympus E1 digital camera through an Olympus BH2 microscope. Legs from four specimens of Dichrorampha dinarica sp. n., two of D. rilana and one of D. ligulana were processed at the Canadian Centre for DNA Barcoding (CCDB, Biodiversity Institute of Ontario, University of Guelph) to obtain DNA barcodes (COI fragment of 658 bp) using the standard high-throughput protocol described in Ivanova et al. (2006), and all resulted in a full barcode fragment. Furthermore, we had access to 237 sequences (> 550 bp) belonging to 35 species of Dichrorampha. Barcode sequences were submitted to GenBank and have the following accession numbers: JX150983-JX150989. Details on the sequences and voucher specimens of species covered in this paper can be obtained from the public project LADIL “Lepidoptera of the Alps - Dichrorampha ligulana-group” in the Barcode of Life Data Systems (BOLD; Ratnasingham and Hebert 2007). Sequence divergences were quantified using the Kimura 2-parameter model implemented within the analytical tools on BOLD (Kimura 1980). The following abbreviations are used for depository institutions: NHMW—Naturhistorisches Museum, Vienna, Austria; NMNHS—National Museum of Natural History, Sofia, Bulgaria; TLMF—Tiroler Landesmuseum Ferdinandeum, Innsbruck, Austria. Results Identifications and misidentifications—a confusing history Tortrix (Grapholitha) ligulana was described by Herrich-Schäffer (1848–1855) based on an unspecified num- ber of specimens of unknown origin. Razowski (1971) fixed the identity of this species by designation of a lecto- type and figured its unmistakable male genitalia. This phenotypically quite characteristic species (Fig. 1) was later described as Dichrorampha plusiana from an unspecified number of specimens from Engadin, Wallis and Piemont (Staudinger 1871). Though Staudinger based his description on specimens belonging to two different taxa, D. ligu- lana and D. thomanni, designation of a lectotype by Huemer (1991) fixed the identity of D. plusiana as a junior subjective synonym of D. ligulana. For about 50 years D. ligulana was considered an alpine endemic until Rebel (1904) recorded a male from Bosnia (Treskavica) and later (Rebel 1913) added additional records from Herzegov- ina (Maglic) and Montenegro (Durmitor). The species was subsequently recorded from Slovakia (Bengtsson et al. 1991), and according to Karsholt & Razowski (1996), it also occurs in Bulgaria, thus giving the impression that this species is widely distributed on the Balkan Peninsula. It was therefore not surprising when one of us (P.H.) col- lected a species of Dichrorampha in the high mountains of Macedonia externally matching D. ligulana. Examina- tion of genitalia revealed that they are quite distinct from those of D. ligulana. We therefore suspected that historical records of D. ligulana from this area could belong to this newly discovered species. However, efforts to obtain material referred to in historical works have been of limited success. Examination of the male genitalia of the single specimen of alleged D. ligulana from Bosnia and Herzegovina deposited in NHMW were identical to those of our Macedonian species, and it seems very likely that all other records from the Dinaric mountains also refer to the Macedonian species. The record of D. ligulana from Slovakia, which was questioned in the original paper owing to its unusual locality (steppe habitat at low altitudes of 200 m) (Bengtsson et al. 1991), was discov- ered to reflect a simple correspondence error overlooked in the galley proofs (Bengtsson in litt.). Finally, the occur- rence in Bulgaria remains doubtful as we have been unable to trace any material or reliable records from this country. It seems likely that such records all refer to D. rilana, a species matching D. ligulana externally, adding to the confusion. 42 · Zootaxa 3389 © 2012 Magnolia Press HUEMER ET AL. Dichrorampha rilana was described from three female specimens - according to Rebel who had studied the types they are in fact males (Obraztsov 1953) - collected on 10.vi.1907 in the Rila mountains (Bulgaria) at an ele- vation of 2000 m (Drenowsky 1909, 1910). Drenowsky (1909) provided a diagnosis and later (Drenowsky 1910) compared the species with D. ligulana and gave an extensive differential diagnosis, pointing out several distin- guishing phenotypical characters. However, the identity of the species remained obscure for several decades, and despite of all efforts we have been unable to trace any syntypes. Neither Obraztsov (1953) in his revision of Palae- arctic Dichrorampha nor other authors as Danilevskii & Kuznetsov (1968) or Razowski (2003) could locate this material. Most probably the syntypes were lost after Drenowsky deposited his collection in Skopje. However, Kuz- netsov (1989) figured the male and female genitalia of alleged D. rilana and added Albania to its distribution area. The full accordance of these figures with our species from Macedonia should have been clear evidence to the iden- tity, but such was not the
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