15urn:lsid:arphahub.com:pub:E0185C18-FE79-5ADE-9877-ED333312DD4Furn:lsid:zoobank.org:pub:EF082B8D-8FD0-41B8-BC24-3D1190FEC17FNota LepidopterologicaNL0342-75362367-5365Pensoft Publishers10.3897/nl.49.191476191476Research ArticleCrambidaePyraloideaTaxonomyIberian PeninsulaSouthern Europe and MediterraneanRedescription and DNA barcoding of Agriphilahispanodeliella Błeszyński, 1959, bona sp., stat. rev. from the Iberian Peninsula (Lepidoptera, Pyraloidea, Crambinae)GirdleyJohnhttps://orcid.org/0000-0001-7976-74391GarreManuelhttps://orcid.org/0000-0002-5846-86211RubioRosa Maríahttps://orcid.org/0000-0002-0109-78741OrtizAntonio S.aortiz@um.eshttps://orcid.org/0000-0002-3877-60961Department of Zoology and Physical Anthropology, University of Murcia, Campus de Espinardo, 30100 Murcia, Spain; john@birdtours.co.uk; mgarre@um.es; rmrubio@um.es; aortiz@um.esUniversity of MurciaMurciaSpainhttps://ror.org/03p3aeb86
Subject Editor: Théo Léger
20260806202649109116A1C38750-8C74-5628-A137-C4DBF1137C423428BCB5-5AB0-4DE9-A945-67C3656EE60D1203202607052026John Girdley, Manuel Garre, Rosa María Rubio, Antonio S. OrtizThis is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.https://zoobank.org/3428BCB5-5AB0-4DE9-A945-67C3656EE60D
The crambid moth Agriphilahispanodeliella Błeszyński, 1959, bona sp., stat. rev. is redescribed from the southern part of the Iberian Peninsula for the first time since its original description as a subspecies of Agriphiladeliella (Hübner, 1813). The range of this taxon is questioned depending on the specimens re-identified from other parts of the Iberian Peninsula and North Africa. A mitochondrial COI barcode sequence was generated for some specimens and compared with other Agriphila species.
Fauna Ibérica: Noctuoidea II (PID2023-149039NB-I00) of the Spanish Ministry of Research and Science, Fundación Séneca FSRM/10.13039/100007801 (23058/GERM/25) from the Comunidad Autónoma de la Región de Murcia and part of DNA sequencing was supported by Genome Canada (Ontario Genomics Institute) in the framework of the iBOL program, WP 1.9.Introduction
The genus Agriphila Hübner, 1825 is distributed around temperate Eurasia and North America and includes twenty-one species in Europe (Leraut 2012), and twelve in the Iberian Peninsula (Revilla 2025).
Agriphiladeliella (Hübner, 1813) was described from Germany and is distributed throughout Continental Europe, Asia Minor, North Africa, Transcaucasia, the West Siberian Plain and Afghanistan (Slamka 2008). In the Iberian Peninsula, it was first recorded by Zerny (1927) in Albarracín (Teruel). Subsequently, Monteiro (1962) found it in Serra da Estrela in Portugal and in 2000s several records from Spain were compiled: Zaragoza (Pérez De-Gregorio 2004) and Segovia, Burgos, Soria and Madrid (Pérez De-Gregorio and Requena 2008). The species was briefly described by Slamka (2008) and Leraut (2012) emphasizing the diagnostic value of the pectinate antennae in males and the prominent pointed frons in both sexes. The moths are on the wing from August to October in dry and open habitats covered by grasses, host plants for their larvae.
Błeszyński (1959) described Agriphilahispanodeliella Błezyński, 1959 based on five male specimens captured in Albarracín (Teruel, Spain) by Anthon Schmidt and deposited in the Magyar Nemzeti Museum collection in Budapest (Hungary). According to the author’s description, there are differences with A.deliella based in the ground color of the forewing, color of veins and antennae, size of the prominence of the frons, and male genitalia. However, Błeszyński (1965) later downgraded A.hispanodeliella to a subspecies of A.deliella without providing the justification for this decision. Slamka (2008) considers A.hispanodeliella to be merely a local variant of A.deliella.
In this article, we provide a re-description of Agriphilahispanodeliella Błeszyński, 1959, bona sp., stat. rev. collected from Sierra Nevada Natural Area (Almeria, Spain). All collected samples have some consistent morphological features and mtDNA sequences (COI), which differ from Agriphiladeliella.
Material and methodsSpecimens examined
Spain. Province of Almería: Bayárcal, Puerto de la Ragua, 2,100 m, 37°05’22.58”N, -3°00’50.60”E, 3 ♂, 29.ix.2024, leg. J. Girdley, gen. prep. JG632, gen. prep. JG634, Barcodes IBLAO3507-25, IBLAO3557-25, IBLAO3558-25, in RCBA-UMU; 2 ♂, 4.ix.2015, leg. M. Garre, gen. prep. 0654MG; 3 ♂, 20.ix.2025, leg. D. Grundy & J. Girdley, in RCBA-UMU; Paterna del Río, Collado de Gabiarra, 2,120 m, 37°04’31.45”N, -2°55’50.02”E, 1 ♂, 4.ix.2016, leg. M. Garre, Barcode IBLAO3586-25, in RCBA-UMU; Paterna del Río, Fuente París, 1,940 m, 37°03’16.24”N, -2°55’47.30”E, 1 ♂, 6.viii.2016, leg. M. Garre, in RCBA-UMU; Laujar de Andarax, Cruz del Pescadero, 2,210 m, 37°04’56.63”N, -2°55’50.35”E, 1 ♂, 23.viii.2022, leg. M. Garre, in RCBA-UMU.
Morphological study
All specimens were examined externally in order to evaluate possible differences in their ground color, wing markings and wingspan. Furthermore, dissections were performed using a standard procedure (Robinson 1976) with minor modifications and genitalia were mounted in permanent microscopic preparations in Dimethyl Hydantoin Formaldehyde (DMHF). Images of the mounted adults (Fig. 1A, B) were taken with a Nikon 950D camera, were z-stacked using the software Zerene (Zerene Systems, Richland, Washington, USA) and were edited with Thumbplus 10 and the morphology of male genital structures (Fig. 2A, B) was studied using a Zeiss Stemi 508 stereomicroscope with a Zeiss Axiocam 208 digital camera. Adult and genital images of Agriphiladeliella are published in Lepiforum.de and are published here under Friedmar Graf permission (Figs 1C, 2C, 2D). All specimens are deposited in the Research Collection of Animal Biology (RCBA-UMU) in the Department of Zoology and Physical Anthropology of the Universidad de Murcia (Spain).
Adult males: A. Typical habitus of Agriphilahispanodeliella Błeszyński, 1959; B. Agriphilahispanodeliella specimen variation. Photographs by R.M. Rubio & A.S. Ortiz; C. Agriphiladeliella (Hübner, 1813). Modified from pictures photographed by F. Graf.
Male genitalia: A, B. Agriphilahispanodeliella Błeszyński, 1959. Whole genitalia except phallus, and Phallus with everted vesica. Photographs by A.S. Ortiz; C, D. Agriphiladeliella (Hübner, 1813). Whole genitalia except phallus, and Phallus with everted vesica. Modified from pictures photographed by F. Graf.
For DNA extraction, two legs were removed from the specimen to sequence the 658 base-pair long barcode segment of the mitochondrial COI gene (cytochrome c oxidase 1, 5’ terminus). The tissue samples were submitted to the Canadian Centre for DNA Barcoding (CCDB, Biodiversity Institute of Ontario, University of Guelph) to obtain DNA barcodes using the high-throughput protocol described in deWaard et al. (2008), which can be accessed at www.dnabarcoding.ca/pa/ge/research/protocols. The DNA extracts are currently stored at the CCDB and the sequences are deposited in GenBank according to the iBOL data release policy. Voucher data, images, sequences and trace files for A.hispanodeliella are publicly available on the public dataset (https://doi.org/10.5883/DS-AGRHISPA) in the Barcode of Life Data System (BOLD) (Ratnasingham and Hebert 2007). Sequence divergences for the barcode region were calculated using the Kimura 2-parameter (K2P) model and the degrees of interspecific genetic variation were calculated using the analytical tools of BOLD. All the new and related species sequences were downloaded and aligned with the CLUSTAL algorithm of the MEGA6 software (Tamura et al. 2013), including all nucleotide pairs with the pairwise deletion option. Bootstrap values were calculated with 1000 replicates, and initial neighbor-joining (NJ) and maximum likelihood (ML) trees based on distance were constructed with the MEGA6 software. All trees produced identical topologies; therefore, only the ML tree showing the sequences from each BIN grouped together is presented (Fig. 3). In order to assess the COI divergences between Agriphilahispanodeliella and the other Agriphila species from Europe, we included all nucleotide pairs sites with the pairwise deletion option. The sequences of the similar Agriphila species according to Slamka (2008)A.deliella (Hübner, 1813) (BOLD:AAP8280; n=9), A.tristella (Denis & Schiffermüller, 1775) with three different BINs (BOLD:AAB9062; n=15; BOLD:ACK2123; n=13; BOLD:ADU0513; n=1), A.selasella (Hübner, 1813) (BOLD:AAE6817; n=16) and A.aeneociliella (Eversmann, 1844) (BOLD:ADD9929; n=1) were obtained from the public database in BOLD. We selected Catoptriabolivari (Agenjo, 1947) (BOLD:ACF1471; n=3), which also belongs to the subfamily Crambinae as outgroup to root the tree.
Maximum-likelihood tree of Agriphila species, obtained from 56 nucleotide COI sequences. The depth of each branch shows divergence within lineages. Bootstrap values are provided at major nodes. The scale bar represents 1% genetic difference.
https://binary.pensoft.net/fig/1673432ResultsAnimaliaLepidopteraCrambidaeC2DFFBFB-C7CB-56DE-A226-F9E8D61FF474AgriphilahispanodeliellaBłeszyński, 1959,bona sp. stat. rev.Agriphilahispanodeliella Błeszyński, 1959, Acta zool. cracov. 4: 148, Pl. 13, fig. 3, Pl. 14, fig. 7 (TL: Spain, Albarracín).Agriphiladeliellahispanodeliella Błeszyński, 1959, Microlepidoptera Palaearctica, Crambinae 1(1): 233, (2): Pl. 13, fig. 145, Pl. 55, fig. 145.Diagnosis.
Agriphilahispanodeliella is similar in appearance to A.deliella and was previously considered to be its subspecies. According to previous authors (Błeszyński 1959, 1965; Slamka 2008; Leraut 2012), A.hispanodeliella differs from A.deliella in several characters: a larger wingspan (A.deliella: 27 mm), forewings with a darker ground color and more distinct markings, unicolorous antennae, and more prominent frons. Regarding the male genitalia, A.hispanodeliella is characterised by a narrower valva apically, while in A.deliella the valva is clearly rounded. The costal arm of A.hispanodeliella is not clearly separated from the valva and projects slightly, while in A.deliella is well developed and clearly separated. In addition, the uncus and gnathos are slender in A.hispanodeliella, while in A.deliella they are broader. The aedeagus is also narrower in A.hispanodeliella and is relatively larger in A.deliella. In conclusion, our study shows that A.hispanodeliella can be distinguished from A.deliella based on differences in wingspan, forewing pattern, and the morphology of the genitalia.
The COI sequences of the sampled specimens had 658 nucleotide bp and indicate significant divergence with mean distances to A.deliella of 3.5% (Table 1). Mean divergence with the other congeneric sequences was 8.33% with maximum distance to the three different BINS of A.tristella (9.2%) and minimum distance to A.selasella (6.8%) (Table 1).
Interspecific mean K2P (Kimura 2-Parameter) divergences (mean pairwise distances) based on the analysis of COI fragments (>500 bp) among Agriphilahispanodeliella and other congeneric Agriphila species (TRIS: A.tristella; SEL: A.selasella; HIS: A.hispanodeliella; DEL: A.deliella).
TRIS ADU0513
TRIS ACK2123
TRIS AAB9062
SEL
HIS
DEL
A.aeneociliella
8.97
8.97
9.42
7.90
7.30
6.99
A.tristella_ADU0513
4.86
4.71
10.03
9.27
8.66
A.tristella_ACK2123
2.58
9.88
8.98
8.66
A.tristella_AAB9062
10.03
9.27
9.57
A.selasella
6.84
6.38
A.hispanodeliella
3.50
Description.
Male (Fig. 1A). Wingspan 28–33 mm. Frons porrect with distinct pointed apex. Head, frons, patagium, tegula, thorax and abdomen covered with pale yellow scales. Labial palpi porrect, about 4× the eye, pale yellow on outer side, brown on inner side. Maxillary palpi pencil-like, pale yellow. Antennae of male short pectinate, weakly ringed in brown and greyish white. Forewings beige to creamy, with a distinct longitudinal white stripe from base to termen, which is edged with a short and narrow brown line on the lower side and a broad and long dark brown stripe on the upper side; above this last stripe, two short parallel white lines, also edged with brown, the one closest to the costa more faintly marked; along de dorsum, a broad brown band crossed by a short whitish line and bordered on its upper side by another, barely perceptible whitish line. Marginal dark dots. Fringes pale beige. Hindwings uniform pale beige. Fringes white.
Variation.
Most specimens show a ground color and wing markings that enhance contrast in the forewing, but some specimens with almost uniformly pale beige forewing and few markings (Fig. 1B).
Male genitalia
(Fig. 2A). Uncus slender, narrowing to hooked apex. Gnathos slightly longer than uncus, distally round. Valva broad and long, with ventral margin somewhat angled medially, tapering to a relatively pointed apex. Costal arms are weakly developed, scarcely visible. Aedeagus straight, with parallel lateral margins and a distinct ventral thorn. Cornuti absent (Fig. 2B).
Female genitalia.
Unknown.
Bionomics.
The studied specimens inhabit open areas above 1,900 m within the supra- and oromediterranean belts of the Sierra Nevada Natural Area. These habitats are rich in grasses, included Corynephoruscanescens (L.) P. Beauv. (Lorite 2016), which is presumed to serve as larval host plant.
Distribution.
Endemic to Spain. Actually, only known in some localities in the centre and south-east of the Iberian Peninsula.
Genetic remarks.
BIN BOLD:AGX2565 (n=4; sequence length 658 bp). Based on COI divergence, A.hispanodeliella is separated from the related species Agriphiladeliella (BOLD:AAP8280; n=9) by mean pairwise divergence in COI-5P of 3.50% (Table 1, Fig. 3).
Discussion
The taxonomic status of A.hispanodeliella has been controversial since Błeszyński (1965) downgraded it to a subspecies of A.deliella because the available data did not support its status as a species. Subsequently, Slamka (2008) noted that only the examination of female genitalia and biological investigation would solve this problem definitively. Agriphilahispanodeliella differs in the DNA barcode from the other European Agriphila species that have been compared in this paper by an average of 7.76% (range 2.58–10.03%) (Table 1) and belongs to a phylogenetically isolated lineage that is well supported by morphology and genetic data. The sequence divergence between A.hispanodeliella and A.deliella is 3.5% suggesting the presence of an isolated lineage in the Iberian Peninsula, which is within the range reported for the COI mitochondrial gene in other congeneric insect species, supporting their genetic distinctiveness (Hebert et al. 2003, 2004). Despite species delimitation is well supported by both morphology and barcodes, the distribution and possible sympatric occurrence of A.hispanodeliella and A.deliella in the north of the Iberian Peninsula remains unresolved. New molecular data sets with data from these areas will be crucial to resolve this issue.
Acknowledgements
Thanks are due to Dave Grundy and Dr. Antonio Vives for their help and collaboration, and Friedmar Graf for sharing photographs of Agriphiladeliella adults and genitalia. This study has been partially funded by the project Fauna Ibérica: Noctuoidea II (PID2023-149039NB-I00) of the Spanish Ministry of Research and Science, Fundación Séneca FSRM/10.13039/100007801 (23058/GERM/25) from the Autonomous Community of the Region of Murcia (Spain) and part of DNA sequencing was supported by Genome Canada (Ontario Genomics Institute) in the framework of the iBOL program, WP 1.9. Collecting permits were issued by the Environmental Authority of the Junta of Andalusia.
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