Pentaneurellakatterjokki Fittkau & Murray (Chironomidae, Tanypodinae): redescription and phylogenetic position

Abstract The monotypic genus Pentaneurella Fittkau & Murray was originally described based on larvae, pupal exuviae and pharate males. The latter prevented the observation of key features, such as wing dimensions, abdominal coloration pattern, and hypopygial apodemes (sternapodeme and phallapodeme), and the description of the adult male was considered incomplete by the authors. Herein, the adult female of Pentaneurellakatterjokki is described for the first time, and the adult male, pupa and larva are redescribed and figured based on specimens recently collected in Germany and Norway. We also discuss the phylogenetic position of Pentaneurella.

Non-biting midges of the genus Pentaneurella are medium-sized dipterans with a Palearctic distribution. Larvae are only known from springs and spring-fed streams in Swedish Lapland and from a mountain stream in northern Norway (Cranston and Epler 2013), although the genus has been consistently recorded through Europe (Saether and Spies 2013). Fittkau and Murray (1983) described Pentaneurella based on larvae, pupal exuviae and pharate males. The latter prevented the observation of key features, such as wing dimensions, wing venation, abdominal coloration pattern, and hypopygial apodemes (sternapodeme and phallapodeme), which are considered essential for male distinction in the subfamily Tanypodinae. Therefore, Pentaneurella katterjokki is redescribed and figured below as adult male, pupa and larva based on specimens recently collected in Germany and Norway. In addition, the adult female is described for the first time. Finally, the phylogenetic position of Pentaneurella is discussed.

Material and methods
Fourth instar larvae and pupae were sampled with hand nets, while adults were collected using emergence-and Malaise traps. Associations between different life stages were established using DNA barcoding. Alcohol-preserved specimens were dissected, the bodies cleared in 8% KOH, and slide-mounted in Euparal®. Measurement methods are according to Epler (1988). Morphological terminology and abbreviations follow Roback (1971) and Saether (1980), supplemented by Kowalyk (1985) for larval cephalic setation and Silva et al. (2011Silva et al. ( , 2014 for larval terminology. The color is described based on the specimens preserved in alcohol. The examined specimens are deposited in the NTNU University Museum insect collection (NTNU-VM) and Zoologische Staatssammlung München (ZSM), Germany. One leg was dissected off each specimen and submitted to the Canadian Centre for DNA Barcoding. Metadata, photos, sequences and trace-files are available in the Barcode of Life Data Systems (BOLD, www.boldsystems.org) through the dataset DS-PKATTER with doi: 10.5883/DS-PKATTER. GenBank accessions are HM421431, HM421434, HM421436, HM421438, HM421441 and MK402317 to MK402322. DNA extracts and partial COI gene sequences were generated using standard primers and bi-directional Sanger sequencing with BigDye 3.1 termination at the Canadian Centre for DNA Barcoding in Guelph. Protocols and original trace-files are available through the dataset DS-PKATTER in BOLD. Alignments were done on amino acid sequences and were trivial as indels were absent; only sequences > 300bp were used in the final alignment.
General coloration. Head pale brown with darker occipital margin; pedicel and antenna brown; maxillary palp pale brown. Thorax pale brown. Wing membrane transparent without marks. Legs brown to pale brown. Abdominal tergite I-VI white, T VII with continuous pale brown transverse band near proximal margin, VIII pale brown; hypopygium pale brown.
Systematics. In their comprehensive analyses of the Chironomidae subfamily Tanypodinae, Silva and Ekrem (2016) considered morphological characters across all life stages for all nine tribes within the subfamily, involving 54 genera and 115 species. In their study, Silva and   Pentaneurella was recovered as sister to Trissopelopia Kieffer in both analyses of equally weighted characters and by using implied weights. However, in an ongoing phylogenetic study of the subfamily Tanypodinae, which includes morphological evidence from modern and fossil chironomids (Silva and Baranov unpub. data), Pentaneurella turned out to be more closely related to the subgenera Reomyia, Schineriella and Zavrelimyia, within Zavrelimyia sensu Silva and Ekrem (2016), than to Trissopelopia, although with low support.
The male of Pentaneurella is morphologically similar to Larsia Fittkau, Pentaneura Philippi, Trissopelopia, and Zavrelimyia Fittkau. The bases of the lyrate tibial spurs are similar to the ones of Larsia and Pentaneura, and the absence of setae on tergite IX resembles Trissopelopia (Murray and Fittkau 1989). Nonetheless, Pentaneurella differs from Larsia in the presence of a distinctively notched, membranous anal point, while the presence of a distinct scutal tubercle separates adults of Pentaneurella from both Trissopelopia and Pentaneura. Moreover, Pentaneurella appears to be similar to Zavrelimyia sensu lato, only differing from the latter by having a scutal tubercle.
Regarding the immature stages, the pupa of Pentaneurella shows certain similarities to Krenopelopia Fittkau and Monopelopia Fittkau (Fittkau and Murray 1986). Monopelopia was recovered by Silva and Ekrem (2016) as sister to Nilotanypus Kieffer, and these two taxa as sister to Monopelopia (Cantopelopia) Roback. The presence of a basal lobe and thoracic comb, and anal macrosetae with adhesive sheaths, however, may be used to distinguish Pentaneurella from Krenopelopia and Monopelopia (Fittkau and Murray 1986). Larvae of Pentaneurella and Krenopelopia differ from Pentaneura and Trissopelopia by possessing a large peg sensilla which is firmly fused with the margin of antennal segment 2, forming a tuning-fork-like process. In both Pentaneurella and Krenopelopia the ligula has a lower middle tooth and inner teeth are curved outward. The absence of a dorsal pore, however, separates Pentaneurella from this genus (Cranston and Epler 2013). In addition, larvae of Pentaneurella appear to have cephalic setation and fork-like Lauterborn organs similar to those of Zavrelimyia sensu lato.
Remarks on distribution and ecology. In the Palaearctic, the subfamily Tanypodinae is represented by 29 genera, of which Anatopynia, Johannsen, Telmatopelopia Fittkau and Pentaneurella currently are unique to the region. The latter is a relatively common genus of non-biting midges initially recorded from northern Scandinavia. Currently, the genus has been recorded in Finland (Paasivirta 2014), France (Brown et al. 2007, Moubayed-Breil et al. 2012, Germany (Stur and Wiedenbrug 2006), Norway (Fittkau and Murray 1983), Russia (Ashe and O'Connor 2009), Slovakia (Šporka 2003), Spain (Hjorth-Andersen 2002), Sweden (Fittkau andMurray 1983, Bylén andRonny-Larsson 1994), Switzerland (Lods-Crozet 1998) and Turkey (Özkan 2006(Özkan , Kazanci et al. 2008. Herein, we record Pentaneurella from Central Norway. Several specimens were collected in the Rondane National Park, located in typical high mountain area, with large plateaus and several lentic and lotic systems. Little is known about the ecology of Pentaneurella. Immature stages seem to be cold stenothermic rheophiles and krenophiles. Larvae of Pentaneurella have been recorded inhabiting springs and spring-fed streams in Sweden and the Bavarian Alps as well as mountain streams in northern and Central Norway (Fittkau andMurray 1983, Stur andWiedenbrug 2006, own data). Moubayed-Breil et al. (2012) found Pentaneurella in low and middle mountain streams located in the eastern Pyrenees and Corsica, while Bylén and Ronny-Larsson (1994) recorded larvae of Pentaneurella being parasitized by the microsporidium Pernicivesicula gracilis Bylén & Ronny-Larsson, in a sample of midge larvae collected from a small river in southern Sweden. Furthermore, larvae of Pentaneurella were also recorded from a sand bed stream from insular Turkey (Özkan 2006).