﻿Two new Ptychoptera Meigen, 1803 (Diptera, Ptychopteridae) from the Western Palaearctic

﻿Abstract Ptychopteraxanthopleura Dvořák, Oboňa & Manko, sp. nov. from Azerbaijan and Georgia, and Ptychopterastaryi Dvořák, Oboňa & Manko, sp. nov. from Bulgaria are described. P.xanthopleurasp. nov. differs from the other member of the lacustris group mainly by having almost completely yellow pleurae, and by the shape of the epandrium and gonocoxites. The diagnostics of P.staryisp. nov. and P.incognita Török, Kolcsár & Keresztes, 2015 based on male genitalia are provided.


Introduction
Ptychopteridae comprises two extant subfamilies: Bittacomorphinae Shiner and Ptychopterinae Alexander. Bittacomorphinae is found in the Nearctic, East Palearctic, and Oriental regions (e.g., Nakamura and Saigusa 2009). Ptychopterinae is widely distributed in the world except for the Australasian Region (Hancock et al. 2006;Fasbender 2017).
The adults live in marshy and moist habitats, near suitable substrates for the larvae as well as quite some distance from the larval habitat (Oosterbroek 2006). The larvae are aquatic to semi-aquatic, living in muddy water, shallow pools in marshes, Sphagnum pools, or the margins of streams (e.g., Andersson 1997;Wiberg-Larsen et al. 2021).
Our present study reports the description of two new Ptychoptera species, one from the Balkans and one from the Caucasus.
All voucher specimens are deposited and accessible in the private collection of the 1 st author. Genitalia of Ptychoptera staryi sp. nov. and P. xanthopleura sp. nov. were macerated in 10% KOH and dehydrated using a series of dehydrating alcoholic solutions (70%, 80%, 96%). After that, parts of genitalia were mounted on permanent slides using Canada balsam as mounting medium.
Photographs of specimens were taken using a Pentax K-50 camera and a reverse-mounted Vivitar 28 mm 1: 2.0 MC lens, Motic SMZ-1 68 stereomicroscope equipped with Canon EOS 1200D camera and EOS utility software, and with Leica M205C stereomicroscope equipped with Leica DFC295 digital camera. Focus stacking was performed using Adobe Photoshop.
Phylogenetic analysis in the case of P. xanthopleura sp. nov.
We undertook phylogenetic analyses to understand the relationships between the species in the subgenus Paraptychoptera. Cladistic analyses of 53 morphological characters on antennae, wing, and male terminalia (Table 1) were selected based on Keresztes et al. (2021) andFasbender (2014). The list of morphological characters is described in detail by Keresztes et al. (2021).
The morphological data matrix was created and managed with Mesquite 3.5 (Maddison and Maddison 2019) and analysed using Maximum parsimony and Bayesian analysis. For maximum parsimony analysis, we used TNT "Tree Analysis using New Technologies" v. 1.5 (Goloboff and Catalano 2016). A "traditional" search based on 1000 replicates of Wagner trees, through 'tree bisection reconnection' (TBR) branch swapping holding 100 trees by the collapsing rule: 'min. length=0'. Subsequently, we selected the best tree, in terms of species topology and population phylogeographical clades, and resampled with 100 000 replicates using a standard bootstrap procedure. Values at nodes represented absolute frequencies and frequency differences (GC, Group present/Contradicted).
For visualisation of the phylogenetic tree, we used FigTree v. 1.4.4 (http:// tree.bio.ed.ac.uk/software/figtree/), and the character state and statistical values visualisation were plotted onto the trees using Adobe Photoshop.

Taxonomic account
Genus Ptychoptera Meigen, 1803 Diagnosis. Medium sized to large (7-15 mm), slender, black lustrous Nematocera, often with lighter markings on thorax and/or abdomen. Antennae, wings, abdomen, and legs long and slender. Ocelli absent; antenna with 15 to 16 segments. Thorax with deep, posteriorly directed transverse suture. Wing with markings, in particular along the crossveins and where veins bifurcate; spurious vein present on either side of crossvein R-M and wing membrane with a distinct fold between veins A 1 and C u A 2 (Oosterbroek 2006;Fasbender 2014). Table 1. Matrix of the 53 morphological data (based on the Keresztes et al. 2021) used in the phylogenetic analyses. For description of characters and character states, see Keresztes et al. (2021). '?' -missing data. border of Imereti and Samtskhe-Javakheti regions, brook and spring, south slope of Zekari pass, 2 050 m a. s. l., 41°49'23"N, 42°51'09"E, 17.VII.2019, leg. G. Vinçon. Description. Male. Head: Frons, vertex, and occiput black with metallic blue shine, mouthparts including palpi pale yellow, scape and pedicel yellowish orange, antennal flagellomeres a somewhat darker, tending to pale brown.
Thorax: Scutum, paratergite, and mediotergite blackish with metallic blue shine; scutellum, pleurotergite, katepisternum, and katepimeron brownish black with lighter metallic blue shine; other parts yellow. Halteres yellow with light brown knob. Legs yellow except brown extreme apex of femora and tibiae, tarsi somewhat darkened.
Wing length 10 mm (holotype, Fig. 1c). Wing almost hyaline, veins yellowish brown, distinct spots brownish black, forming more or less three stripes, at base of wing at the level of crossvein h, from C to Cu. Middle stripe touching vein C, running through cross-veins up to middle part of vein Cu, isolated spot before end of vein Cu. Isolated spot on around middle of R 1 . Third stripe consist of three large, almost touching spots: at the tip of R 1 and fork of R 2 + 3 , one on fork vein R 4 + 5 and one on fork vein M 1 + 2 . Small spot at the end of vein R 3 . Abdomen: Tergum 1 dark shiny brown with yellow apex, sternum 1 yellow. Tergum 2 brown basally and apically, yellow in middle, sternum 2 yellow. Tergum 3 yellow basally, brown apically, sternum 3 yellow. Remaining terga and sterna brown, sternum 4 yellow basally. Auxiliar copulatory organ yellow.
Male genitalia (Fig. 2): Hypopygium almost 2× as wide as long, widest in basal quarter, medially with very deep emargination. Epandrial claspers simple and long (length/width ratio ca. 5.8) and covered by long pale hairs, the longest hair up to 1.75 longer than width of epandrium. Apical stylus of gonostylus robust and rounded apically, secondary lobe long, reaching almost 0.75 of apical stylus length. Media lobe of basal lobe of gonostylus long and sharply pointed (saber-like); anterior lobe of basal lobe of gonostylus bulbous apically with several setae at extreme apex.
Female. The authors have an immature female which was sampled in Lesser Caucasus (Georgia, Kakheti region, Ilto river, above (N of) the Chart'ala village, 790 m a. s. l., 42°8'18"N, 45°7'32"E, 8.VII.2019, leg. P. Manko & G. Vinçon). The characters correspond to the above-described new species. However, its identity cannot be confirmed in this stage of ontogenesis/development and could be solved after collecting more specimens of the genus Ptychoptera from the Transcaucasia.
Etymology. The name reflects predominantly yellow pleurae (Fig. 1b), which are unique for the Western Palaearctic species.
Differential diagnosis. According to the presence of auxiliary sexual organ and shining pleurae, P. xanthopleura sp. nov. belongs to the subgenus Paraptychoptera and according to male genitalia and the maximum parsimonious tree based on 53 morphological characters (see Fig. 3 and Table 1), the nearest species is P. lacustris and belongs to the highly divergent monophyletic unit, the lacustris group, including five species, P. xanthopleura sp. nov., P. lacustris, P. castor, P. hel- ena, and P. pollux (Fig. 3). Ptychoptera xanthopleura sp. nov. is close to but differs from the most similar species P. lacustris mainly by having an almost completely yellow pleurae, the shape of the hypopygium (epandrial claspers, secondary lobe of gonostylus, and medial lobe of basal lobe of gonostylus).  Thorax: Predominantly black with silvery pubescent pleurae. Pronotum, epimeron 3 and metanotum 3 yellow. Fore and mid coxae and trochanters yellow, hind coxae brownish black basally, yellow apically, coxae also yellow. Almost all legs are missing. Halteres whitish yellow with a darker knob. Figure 5. Epandrium, gonocoxite and gonostylus, and hypandrium of P. staryi sp. nov., P. incognita, and P. albimana. Abbreviations: ECP = epandrial clasper, EL = epandrial lobe, EPI = epiproct, GBM = medial lobe of basal lobe of gonostylus, GCT = gonocoxite, GAS = apical stylus of gonostylus, GAT = tertiary lobe of apical stylus of gonostylus, PPA = apical process of paramere, HBD = basal division of hypandrium, HMW = membranous window of terminal division of hypandrium, HTD = terminal division of hypandrium.

Ptychoptera staryi
Wing length 12 mm (Fig. 4b). Wing with yellowish veins and infuscated spots on fork vein Rs+R, all cross-veins, end of R 1 vein up to its fork with R 2 vein, on fork vein R 4 +R 5 and on fork vein M 1 +M 2 . Legs: femur pale, darker in apical ¹⁄5, almost black on extreme apex; tibia pale brown in basal ½, darker apically and almost black on extreme apex; tarsomere 1 almost black, tarsomeres 2 and 3 dark brown, tarsomeres 4 and 5 pale brown.
Male genitalia: similar to P. incognita. Epandrial clasper slightly curved outwards with simple obtuse apex; anterior projection of ventromesal lobe sharp, posterior projection bow-shaped backwards; space between both projections is rounded, almost semi-circular. Gonocoxite and gonostylus: apical process of paramere with a U-shaped dark structure with thick edges; paramere base rounded, convex; width to height ratio of dorsal gonocoxal lobe 0.5; dorsal gonocoxal lobe with dense tiny dark hairs. Aedeagus: sides of lateral ejaculatory process distinctly convex, basal projections markedly convergent; transition to lateral ejaculatory process smooth, undulate. Hypandrium: width to length ratio 1.2; apex of hypogynial valves start under basal division of hypandrium. See also differential diagnosis.
Female. Unknown. Etymology. The name is dedicated to our colleague Jaroslav Starý and his life jubilee. (Jaroslav discovered the holotype of new species in his own material and provided it for the description).
Differential diagnosis. The new species is very similar to P. incognita Török, Kolcsár & Keresztes, 2015 (see also Table 2). After comparing the holotype of P. staryi sp. nov. with individuals of P. incognita (material used in Török et al. 2015: 2 individuals from Bulgaria, 17 individuals from Romania), we found that a diagnosis was possible on the basis of differences in male genitalia (Fig. 5, marked with arrows), namely: (i) shape of the plate on ventral parts of epandrium is of Projections of the plate on ventral site in obtuse angle, space between projections rounded, more than semicircle.

Gonocoxite and gonostylus
Width to height ratio of dorsal gonocoxal lobe 0.5; dorsal gonocoxal lobe with dense tiny dark hairs. The hairs of margins of gonocoxite and gonostylus much less dense and finer. Chitinisation of the proximo-lateral processes of the gonocoxite not developed, processes light coloured.
Width to height ratio of dorsal gonocoxal lobe 0.6; dorsal gonocoxal lobe with sparse small hairs. The hairs of margins of gonocoxite and gonostylus dense and stronger. Chitinisation of the proximo-lateral processes of the gonocoxite strongly chitinised, dark.
Width to height ratio of dorsal gonocoxal lobe 0.4; dorsal gonocoxal lobe with tiny dark hairs at the top.
The hairs of margins of gonocoxite and gonostylus dense and stronger. Chitinisation of the proximo-lateral processes of the gonocoxite not developed, processes pale coloured.

Aedeagus
Sides of lateral ejaculatory process distinctly convex, basal projections markedly convergent; transition to lateral ejaculatory process smooth, undulate.
Sides of lateral ejaculatory process slightly convex, basal projections divergent; transition to lateral ejaculatory process steep, in a right or acute angle to the axis of aedeagus.
Sides of lateral ejaculatory process slightly convex, basal projections almost parallel; transition to lateral ejaculatory process steep, in an obtuse angle to the axis of aedeagus.
Hypandrium Width and length ratio 1.2; apex of hypogynial valves start under basal division of hypandrium.
Width and length ratio 0.9; apex of hypogynial valves start on bases of basal division of hypandrium.
Width and length ratio 0.9; apex of hypogynial valves start over basal division of hypandrium. a different shape and orientated at a different angle in P. staryi; (ii) chitinisation (sclerotisation) of the proximo-lateral processes of the gonocoxite is not developed and these processes are light coloured in P. staryi in contrast to P. incognita with strong chitinisation and dark colouration; (iii) the hairs of margins of gonocoxite, gonostylus, and epandrium are much less dense and finer in contrast to P. incognita (not visible in Fig. 5).