New species of Limnephilidae (Insecta: Trichoptera) from Europe: Alps and Pyrenees as harbours of unknown biodiversity Author Graf, Wolfram Author Vitecek, Simon Author Previšić, Ana Author Malicky, Hans text Zootaxa 2015 3911 3 381 395 journal article http://dx.doi.org/10.11646/zootaxa.3911.3.5 8ebf5688-2e69-4da4-b700-004354c3fe53 1175-5326 PMC4805176 25661619 244687 4E11C1AB-2614-4C4A-809D-EF7C5BE959D9 Anisogamus species delimitation and larval affiliation An analysis of the genetic distance of mtCOI between two Anisogamus males clearly supports differentiation of A. difformis and A. waringeri sp. nov. as separate species. Uncorrected p -distances recorded in both fragments of the mtCOI gene (i.e., 8.2 % and 9.6 %, in the “barcode region” and the “S 20 -Jerry region”, respectively; Table 1 ), are in line with interspecific distances commonly recorded in Limnephilidae (e.g., Graf et al. 2005 , Previšić et al. submitted) and other caddisfly families (e.g., Hydropsychidae ; Pauls et al. 2010 ). Haplotypes of the “barcode region” of the adult male of A. waringeri sp. nov. and the hitherto unknown larva collected at the same locality differed in a single base pair, whereas haplotypes of the “S 20 -Jerry region” were completely identical in these two specimens ( Table 1 ). Since these values are well within the intraspecific variability of mtCOI usually observed in caddisflies (e.g., Pauls et al. 2009 , 2010 ; Previšić et al. 2009 , 2014 ) data at hand enable confident affiliation of the larva and the male of A. waringeri sp. nov. FIGURE 6. Larval characters of A. waringeri sp. nov. and A. difformis . A, A. waringeri sp. nov. head, left lateral; B, A. waringeri sp. nov. head, frontal; C, A. difformis pronotum, left lateral; D, A . waringeri sp. nov. pronotum, left lateral; E, A. difformis pronotum, right lateral (arrows indicating row of pale yellow setae); F, A. waringeri sp. nov. pronotum, right lateral (arrows indicating row of all-dark setae). FIGURE 7. Larval characters of A. waringeri sp. nov. A, pronotum, dorsal; B, left lateral protuberance, left lateral (dotted circle indicating setal area sa 3, arrow indicating associated sclerite); C, abdominal sternum I, with the separated setal areas 1 ( sa 1) marked with a dotted circle; D, left side of abdominal segments IX and X and left anal proleg and claw, left lateral (upper arrow indicating posterolateral setae of abdominal dorsum IX, lower arrow indicating long stout seta on abdominal venter IX); E. larval case of A. waringeri sp. nov. TABLE 1. Inter- and intraspecific genetic distances of two mitochondrial cytochrome oxidase I (mtCOI) gene fragments recorded for Anisogamus species. Values below diagonal in second and third columns indicate number of nucleotide differences and above diagonal uncorrected pairwise distances ( p ) (shown as percents), respectively. Abbreviations are used to denote life stages; IM/M = adult male, L = larva. Species Stage Specimen codes mtCOI "barcode" Andiff01 Ansp01 fAns0101L Anisogamus difformis IM/M Andiff01 8.2 8.2 Anisogamus waringeri IM/M Ansp01 54 0.2 Anisogamus waringeri L fAns0101L 54 1 TABLE 1. (Continued) Species mtCOI "S 20 -Jerry" GenBank Access. No. Andiff01 Ansp01 fAns0101L "barcode" "S 20 -Jerry" Anisogamus difformis 9.6 9.6 KP 174661 KP 174658 Anisogamus waringeri 52 0 KP 174662 KP 174659 Anisogamus waringeri 52 0 KP 174663 KP 174660 Distribution & biogeography of Anisogamus species. The genus Anisogamus was established by McLachlan in 1874 based on the species A. difformis , and its type locality is situated in the Eastern Alps ( Austria , Carinthia, Saualpe, Stelzing (Kimmins 1949 )) . Collated distribution data for A. difformis suggest a panalpine presence of the species ( Fig. 3 D). Specimens of A. waringeri were collected at the Col de Jou, Mont Canigou, Pyrenées-Orientales, France . At a location close by, Décamps ( 1967 ) found putative A. difformis to be present (but very rare) in the valley of the Neste d’Aure at 1600 m a.s.l. and in the tributaries of the Têt river at 1100 m a.s.l. Specimens of A. waringeri sp. nov. were collected in the watershed of the Têt river, whereas the Neste d’Aure is some 125 km west of the recent collection points. Menéndez & González ( 2009 ) recorded A. difformis from the eastern Prepyrenees (Girona, Setcades), some 20 km south of the type locality of A. waringeri sp. nov. , and were re-identified by M. A. González as A. waringeri sp. nov. (pers. comm. M. A. González). From the same area, Stenophylax nurianus was described by Navás ( 1917 ) , illustrating a specimen similar to the genus Anisogamus , but the type specimen is lost (pers. comm. M. A. González), and the description and the figure itself do not allow certain identification. Further, this species was proposed by Schmid ( 1949 ) to be a synonym of A. difformis , based on his own collection and identification of 2 putative A. difformis specimens. Thus, we consider Stenophylax nurianus a nomen dubium in concordance with Malicky ( 2005 ) , justifying the description of A. waringeri sp. nov. We further conclude that A. waringeri sp. nov. is the single representative of the genus Anisogamus in the Pyrenees.