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Tipula (Pterelachisus) recondita Pilipenko & Salmela, sp. n. is described. The new species is collected from two localities: Finland, Kittilä (North boreal ecoregion) and Russia, Primorski kray (Zone of temperate broadleaf and mixed forests). Although variation in the structure of male hypopygium between the Finnish and Russian populations is observed, DNA barcode sequences differ only by three nucleotides (0.2 % K2P distance), supporting presence of one widespread species. K2P minimum distances between the new species and 17 other species of the subgenus range from 5.3 to 15.8 % (mean 8.8 %). The new species is forest-dwelling, known from an old-growth herb-rich forest (Finland) and Quercus mongolica forest (Russia). The new species is perhaps closest to Tipula (Pterelachisus) imitator Alexander and in lesser extent to Tipula (Pterelachisus) pauli Mannheims; the inner gonostylus of both species are illustrated.
Crane flies, Tipulinae, taxonomy, Finland, Russia, COI
Tipulidae (Diptera, Nematocera), or long-palped crane flies, are medium to large sized true flies. Globally, 4269 tipulid species and subspecies are known, of these 1322 occupying the Palaearctic region (
Tipula (Pterelachisus) Rondani is a northern hemisphere subgenus, totaling over 200 species and subspecies (
DNA barcoding is a molecular-based method used in the identification and delimitation of species, having usually considerable congruence with morphology-based identifications (
In the present article we provide a description of Tipula (Pterelachisus) recondita Pilipenko & Salmelasp. n. collected from Europe (Finland) and Asia (Russian Far East). Both sexes of the new species are richly illustrated. In addition, mtDNA sequences (COI) were used to assess (i) the conspecific status of disjunct Finnish and Russian populations and (ii) genetic divergence between the new species and 17 consubgeneric species.
Material and methodsTotal DNA of Tipula (Pterelachisus) recondita Pilipenko & Salmela sp. n. specimens was extracted using a modified non-destructive salt extraction method (
The DNA barcode region (cythocrome oxidase subunit I) was amplified and sequenced from all specimens using universal primers LCO1490: 5’-GGGTCAACAAATCATAAAGATATTGG-3’ and HCO2198: 5’-TAAACTTCAGGGTGACCAAAAAATCA-3’ (
For other species (totaling 17 species and 26 specimens, Table 1) DNA barcodes were obtained at the Canadian Centre for DNA Barcoding. Legs or 2–3 abdominal segments of the specimens were placed in 96% ethanol in a 96-well lysis microplate and dispatched to the Biodiversity Institute of Ontario where DNA was extracted and sequenced using standard protocols and primers (
Tipula (Pterelachisus) specimens used in DNA barcoding (COI). Species and associated BOLD Sample ID are according to HOLPT project, available in http://www.boldsystems.org/. Co-ordinates are given in WGS84 decimal format.
Sample ID, species | Year | Country | Locality | N | E |
---|---|---|---|---|---|
JES-20110456|Tipula_cinereocincta | 2005 | Finland | Heinävesi | 62.419 | 28.596 |
JES-20120024|Tipula_cinereocincta | 2007 | Finland | Savonranta | 62.251 | 28.877 |
JES-20120065|Tipula_angulata | 2006 | Canada | Ontario | 45.483 | -76.081 |
JES-20120064|Tipula_entomophthorae | 2003 | Canada | Manitoba | 54.9 | -101.43 |
JES-20120004|Tipula_jutlandica | 2008 | Finland | Parikkala | 61.565 | 29.559 |
JES-20110501|Tipula_laetibasis | 2002 | Finland | Tuupovaara | 62.442 | 30.606 |
JES-20110497|Tipula_luridorostris | 2006 | Finland | Taivalkoski | 65.785 | 28.321 |
JES-20120011|Tipula_mats._pseudohortensis | 2007 | Finland | Inkoo | 60.018 | 23.822 |
JES-20110475|Tipula_mats._pseudohortensis | 2007 | Finland | Siuntio | 60.213 | 24.135 |
JES-20110092|Tipula_mutila | 2009 | Finland | Enontekiö | 68.639 | 22.552 |
JES-20110204|Tipula_mutila | 2008 | Finland | Kiuruvesi | 63.52 | 26.69 |
JES-20120095|Tipula_mutila | 2007 | Finland | Kittilä | 68.33 | 24.64 |
JES-20120014|Tipula_octomaculata | 2008 | Finland | Lieksa | 63.217 | 30.218 |
JES-20120031|Tipula_octomaculata | 2006 | Finland | Taivalkoski | 65.693 | 28.32 |
JES-20110494|Tipula_pauli | 2007 | Russia | Primorski kray | 47.94 | 137.72 |
JES-20110495|Tipula_pauli | 1995 | Russia | Moscow region | 56.02 | 37.11 |
JES-20110502|Tipula_pseudovariipennis | 2006 | Latvia | Tukums | 56.998 | 23.003 |
JES-20110035|Tipula_recondita_sp._n | 2009 | Finland | Kittilä | 67.634 | 25.416 |
JES-20110034|Tipula_recondita_sp._n | 2009 | Finland | Kittilä | 67.634 | 25.416 |
JES-20110036|Tipula_recondita_sp._n | 2006 | Russia | Primorski kray | 43.125 | 131.4 |
JES-20120038|Tipula_stenostyla | 2009 | Finland | Kittilä | 67.634 | 25.416 |
JES-20110408|Tipula_submarmorata | 2009 | Finland | Jyväskylä | 62.236 | 25.679 |
JES-20120041|Tipula_truncorum | 2010 | Finland | Enontekiö | 69.183 | 21.521 |
JES-20110345|Tipula_varipennis | 2007 | Finland | Ranua | 66.017 | 26.852 |
JES-20120032|Tipula_varipennis | 2005 | Finland | Jyväskylä | 62.213 | 25.793 |
JES-20110401|Tipula_varipennis | 2009 | Finland | Jyväskylä | 62.236 | 25.679 |
JES-20110222|Tipula_wahlgreni | 2008 | Finland | Kiuruvesi | 63.52 | 26.69 |
JES-20110450|Tipula_winthemi | 2009 | Finland | Lammi | 61.091 | 25.002 |
JES-20120026|Tipula_winthemi | 2008 | Finland | Virolahti | 60.465 | 27.426 |
In order to assess the COI divergence between the new species and 17 Holarctic Tipula (Pterelachisus) species, we calculated Kimura two-parameter (K2P) (
Maximum Likelihood tree based on COI sequences (mtDNA) of 17 Tipula (Pterelachisus) species. Numerical values denote to Bootstrap values after 1000 replications. In the tree Bootstrap value 26 refers to the clade including Tipula recondita sp. n. and Tipula jutlandica and value 11 refers to the clade including Tipula recondita sp. n., Tipula jutlandica, Tipula stenostyla and Tipula matsumuriana psedohortensis. Scale bar: nucleotide substitutions per site.
The morphological terminology used here mainly follows
Specimens were studied with a Zoom Stereo Microscope. Photographs were taken with a Canon PowerShot A640 camera and processed using Combine ZP software. All drawings were prepared from photographs.
[Comparative morphological material examined. Tipula (Pterelachisus) imitator Alexander: Russia, Shikotan Island, Kray Sveta cape, 25.VII.1965, V. Ermolenko, 1 male (ZMKU); Russia, Iturup Island (Kuril Is), Kurilsk env., 5.VII.1963; Krilov & Krivolutskaya, 1 male (ZISP). Tipula (Pterelachisus) pauli Mannheims: Russia, Moscow region, Chashnikovo, 29.V.1995, V. Pilipenko, 1 male (VPM).]
Taxonomyurn:lsid:zoobank.org:act:CFBAD0A0-AC21-4067-88E4-C15BCA35CC56
Holotype: Male, in alcohol (NCBN). “Finland, Lkoc: Kittilä, Iso Mustavaara, old-growth herb-rich forest, 67.6340°N, 25.4160°E, 30.V.–1.VII. 2009, J. Salmela leg.” (white label, printed) “Tipula (Pterelachisus) recondita sp. n./ Pilipenko &
ATGCTTTTATTATAATTTTTTTTATAGTTATACCTATTATAATTGGAGGATTTGGAAATTGATTAGTAC
CTTTAATATTAGGTGCCCCTGATATAGCCTTTCCTCGAATAAATAATATAAGTTTTTGAATATTACCTC
CTTCACTTACTCTTTTATTAGCTAGTAGTATAGTCGAAAACGGTGCGGGGACTGGATGAACCGTTTATC
CCCCACTCTCATCTAGAATTGCCCATACAGGAGCTTCAGTTGATTTAGCCATTTTTTCTCTTCATTTAG
CTGGAATTTCTTCAATTTTAGGAGCAGTAAATTTTATTACTACAGTAATTAATATACGATCAAGAGGAA
TTACTTTAGACCGAATACCTTTATTTGTTTGATCGGTAGTAATTACTGCAGTATTATTACTACTCTCTT
TACCTGTATTAGCGGGAGCTATTACTATACTTTTAACTGATCGAAATTTAAATACATCATTTTTTGATC
CTGCAGGAGGTGGAGATCCAATTCTTTACCAACATTTATTT
Finland, Lkoc: Kittilä, Iso Mustavaara Nature reserve, herb-rich old-growth forest, 67.6340°N, 25.4160°E, 30.V.–1.VII. 2009, Malaise trap, J. Salmela leg., 2 males (ZMUT, in alcohol [BOLD sample ID JES-20110035] and a pinned specimen). DNA barcode (524 bp) of paratype (JES-20110035|FINTI035-11) is identical to the holotype sequence. Russia, Far East, Primorski kray, Kedrovaya Pad’, oak forest (Quercus mongolica), 43.1301°N, 131.5041°E, 7.VII. 2006 V. Pilipenko leg., 3 males and 3 females, deposited in ZSIP (BOLD sample ID JES-20110036), ZMUM, VPM. DNA barcode (524 bp) of paratype male (JES-20110036|FINTI036-11) differs from holotype at three positions (212=C, 473=T, and 515=G). In other words, intraspecific K2P distance between Finnish and Russian specimens was 0.2 %.
Rather small yellowish brown Tipula species (body length: 11 mm male, 12.3 mm female; wing length 11–12.6 mm male, 12.5–13.5 mm female). Scape, pedicel and base of 1st flagellomere yellowish, other flagellomeres brown. Caudal margin of male 9th tergite with a median notch, bearing no tooth or other elevated structures. Outer gonostylus narrow, about as long as inner gonostylus, slightly bent sub-basally. Lower beak of inner gonostylus apically rounded, black. Outer basal lobe of inner gonostylus with 3–4 stout black spines.
Male. Head gray pruinose, sparsely covered with dark hairs. Base of rostrum gray pruinose, otherwise dark brown, shining. Nasus distinct, tip with light bristles (Fig. 2a). Palpi brownish. Lengths of palpal segments (n=2): p1 128-147, p2 307-309, p3 317-365, p4 309-333 and p5 1207-1359. Scape, pedicel and base of 1st flagellomere yellowish, other flagellomeres brown. Scape cylindrical (length 442–466, width 119–120, n=2). Pedicel globular (length 132–134, width 134–135, n=2). Flagellar segments cylindrical, covered with silvery, erect and thick pubescence. Verticils black, shorter than respective segments (Fig. 2a). Lengths of flagellomeres (n=2): f1 371–398, f2 312–314, f3 298–316, f4 289–326, f5 297–324, f6 296–325, f7 291, f8 270–289, f9 257–261, f10 227–230 and f11 100. Thorax. General coloration dark brown, with gray pruinosity (Fig. 2b). Pronotum with light hairs. Prescutum with four longitudinal brown bands; lateral bands short, median bands distinctly separated. Anepisternum, katepisternum and anepimeron with dense, gray pruinosity. Scutum, scutellum, laterotergite and mediotergite unicolorous, dark brown. Coxae brown, with light hairs. Trochanters yellowish, with light hairs. Proximal part (ca. two thirds) of femora yellowish, turning dark brown toward tips. Tibiae and tarsi dark brown, spur formula 1:2:2. Tarsal claws smooth. Legs covered with dark brown – black bristles. Stem of halter yellowish, knobs infuscated. Wings with marmorate pattern, length (n= 5) 11.9 mm (11–12.6 mm), venation as in Fig. 2c. R1+2 is variable, reach or not reach Costa. Wing cells c and sc yellowish, other cells brown tinged (see Figs. 2b, 2c). Pterostigma distinct. Abdomen yellowish brown, with a narrow dorsal stripe (Fig. 2b). Hypopygium (Fig. 3a) dark brown. Caudal margin of 9th tergite with a median notch, bearing no tooth or other elevated structures (Figs. 3g–h). Caudal margin of 9th tergite oblique (Finnish specimens) or almost horizontal, truncated (Russian specimens) (Figs. 3g–h). Outer gonostylus narrow, about as long as inner gonostylus, slightly bent sub-basally (Figs. 3b, d). Lower beak of inner gonostylus apically rounded, black. Beak of inner gonostylus rather narrow and elongated in lateral view (Figs. 3b, d), tip roundish and proximal margin oblique, notched in posterior view (Fig. 3c). Outer basal lobe of inner gonostylus with 3–4 stout black spines. Aedeagal guide as in Figs. 3e–f. Sperm pump hairy between posterior immovable apodemes, apex of aedeagus pointed (Figs. 3i–j).
Female. Wing length (n=3) 12.8 mm (12.5–13.5 mm), body length (n=3) 12.3 mm (12–13 mm). Generally similar to male (Fig. 2e). Antenna short (2.4 mm), not extending to wing base (Fig. 2d). The wing’s marmorate pattern more intensive than in male (Fig. 2f). Ovipositor (Figs. 4a, b) elongate, similar to that of most other tipulines; 8th tergite dark brown, 9th tergite narrow dull dark brown, 10th tergite shining chestnut brown. 8th sternite dull dark brown anteriorly, grading to shining yellow posteriorly. Cerci narrow, yellow, slightly longer than 10th tergite. Hypogynial valves yellow, reaching mid-length of cerci, relatively wide, gradually narrowing (Fig. 4c).
Tipula (Pterelachisus) recondita Pilipenko & Salmela, sp. n. a Holotype male, head, lateral view (Finland) b paratype male, habitus, lateral view (Russia) c paratype male, wing (Russia) d paratype female, head, dorso-lateral view (Russia) e paratype female, habitus, lateral view (Russia) f paratype female, wing (Russia). Scale bars: a, d 1 mm; c, f 2.5 mm; b, e 5 mm.
Tipula (Pterelachisus) recondita Pilipenko & Salmela, sp. n., paratype males a hypopygium, lateral view (Russia) b outer and inner gonostylus, lateral view (Finland) c outer and inner gonostylus, posterior view (Finland) d outer and inner gonostylus, lateral view (Russia) e aedeagal guide, lateral view (Finland) f aedeagal guide, lateral view (Russia) g 9th tergite, dorsal view (Finland) h 9th tergite, dorsal view (Russia) i sperm pump and aedeagus, ventro-lateral view (Finland) j sperm pump and aedeagus, ventro-lateral view (Russia). Scale bars: 0.5 mm.
Tipula (Pterelachisus) recondita Pilipenko & Salmela sp. n., paratype female (Russia) a–c Tipula (Pterelachisus) pauli Mannheims d and Tipula (Pterelachisus) imitator Alexander (e, f, g, h, i). a female terminal abdominal segments and cerci, dorsal view b female terminal abdominal segments, cerci and hypovalva, lateral view c female hypovalva and 8th sternite, dorsal view d–e male inner gonostylus, lateral view f outer and inner gonostylus, posterior view g 9th tergite, dorsal view h aedeagal guide, lateral view i sperm pump and aedeagus, ventro-lateral view. Scale bars: 0.5 mm.
The species epithet is from reconditus (Latin, adjective) meaning hidden, concealed. This word refers to the rarity and apparent low detectability of the new species, so far known only from two sites in the Palaearctic region.
Tipula (Pterelachisus) recondita Pilipenko & Salmela, sp. n. is known from North Europe (Finland) and Asia, Russian Far East. The Finnish collecting site in Kittilä, Iso Mustavaara, is a state-owned Nature Reserve (Lehtojensuojelualue), included in the Natura2000 network of conservation areas. It is part of the biogeographical province of Lkoc (Lapponia kemensis pars occidentalis) and lies in the North boreal vegetation zone. The collecting site is an old-growth mixed forest, dominated by birch (Betula pubescens), goat willow (Salix caprea) and Norway spruce (Picea abies), with scattered aspen (Populus tremula) trees. Lower vegetation is characterized by herbs and shrubs such as Calypso bulbosa, Daphne mezereum, Actaea erythrocarpa, Ribes spicatum, Filipendula ulmaria and Geranium sylvaticum. Decaying trees, especially goat willow and birch, are abundant in the site. The Russian collecting site is located in the Kedrovaya Pad’ Nature Reserve, within the temperate broadleaf and mixed forest zone, in an oak forest (Quercus mongolica) growing on limestone outcrops on the southern slope of a mountain range. Lower vegetation is characterized by Lespedeza bicolor, Spodiopogon sibirieus, Astra ageratoides, Carex siderosticta, Artemisia keiskeana, Lathyrus davidii and Calamagrostis brachytricha.
Tipula (Pterelachisus) recondita Pilipenko & Salmela, sp. n. is rather easily distinguished from other Holarctic Tipula (Pterelachisus) species. The new species is distinctive in characters of the male hypopygium, especially that of the 9th tergite. There are several Tipula (Pterelachsus) species with a U-shaped median notch or an emargination in the caudal margin of the tergite, but usually having a tooth or other elevated structures at the mid-point (e.g. Tipula (Pterelachisus) angulata Loew [
Morphologically the new species is perhaps the closest to two Palaearctic species, namely Tipula (Pterelachisus) imitator and Tipula (Pterelachisus) pauli. The former species has a median notch in 9th tergite, but also a distinct tooth at the midpoint (Fig. 4g); the outer basal lobe of inner gonostylus bears one conspicuous black spine, not 3–4 smaller ones (Fig. 4e). For other differences, see Figures 4f, h, i. Tipula pauli also has a median notch in 9th tergite and a small but discernible tooth in the midpoint; the lower beak of inner gonostylus is roundish and black, but the outer basal lobe bears no stout, black spines (Fig. 4d). Tipula (Pterelachisus) imitator is known from Japan and Kuril Islands and Tipula (Pterelachisus) pauli from Europe, Altay and Russian Far East (Oosterbroek 2012, V. Pilipenko pers. obs.).
Based on COI divergence, the new species is apparently rather isolated from the members of the subgenus Pterelachisus (Fig. 1). Among the other species vs. the new species, interspecific distances varied from 5.3 % (Tipula winthemi Lackschewitz) to 16.1 % (Tipula laetibasis Alexander). Mean of the minimum interspecific distances was 8.8 %. According to K2P divergence, the new species is closest to Tipula winthemi (5.3 %), Tipula jutlandica Nielsen (5.5 %), Tipula stenostyla Savchenko (6.6 %) and Tipula pauli (6.8 %); distances between the other species range from 7.4 to 16.1 %. In other words, no very close relatives were present in the pair-wise comparisons of COI sequences. For example, much shorter interspecific K2P distances were found between Tipula varipennis/Tipula pseudovariipennis (1.5 %), Tipula mutila/Tipula wahlgreni (2.2 %), Tipula stenostyla/Tipula winthemi (3.7 %). However, it must be emphasized that Tipula imitator was not included in COI analysis, due to the lack of fresh material. Given to the morphological similarity of the new species and Tipula imitator, it is likely that their barcoding distances would be similar to those three comparisons given above.
There are some morphological differences (9th tergite, inner gonostylus) between Finnish and Russian specimens, perhaps due to the long distance and lack of gene flow between the populations. These differences, however, are here considered to be intraspecific variation. Very small K2P divergence of COI gene (0.2 %) between Finnish and Russian specimens also substantiates the presence of one widespread, but disjunct, species. In rare cases (see
The new species is most probably a very rare tipulid. Despite the rather long tradition of crane fly taxonomy and faunistics in North Europe, this species has hitherto remained unnoticed. One of the authors (JS) has within 12 years identified some 70 000 crane flies from a Finnish Malaise trapping material consisting of 476 sampling sites and ca. 1670 Malaise trapping months. Thus, despite this relatively large sampling effort, only three specimens from a single locality have been caught. The true range of the species is Palaearctic, whether disjunct or not remains to be seen. In Northwestern Europe the species is likely to occur in the north boreal zone (for further information on boreal ecoregions or vegetation zones, see e.g.
To our regret we were not able to examine the holotype male of Tipula imitator (D. Furth, pers. comm.). Description of that species was based on a single male specimen (
Special thanks to Pjotr Oosterbroek (Amsterdam) for his help with the vast Tipula (Pterelachisus) literature. It would have been much more laborious for us to resolve the status of the new species without Pjotr’s help. Metsähallitus, Lapin luontopalvelut (Finnish Natural Heritage Services) provided a permission to collect insects in Iso Mustavaara Nature Reserve. Comments by John Kramer (Leicester), Fenja Brodo (Ottawa), anonymous referee and Vladimir Blagoderov (London) improved manuscript. Aleksanteri Hihnavaara (Ponku) is thanked for such a cheerful field company. Barcode sequences were obtained at the Canadian Centre for DNA Barcoding based in the Biodiversity Institute of Ontario at the University of Guelph. Their work was supported by funding from the Government of Canada through Genome Canada and the Ontario Genomics Institute in support of the International Barcode of Life Project. Finnish Barcode of Life (FinBOL) is also thanked for a help in the barcoding process. JS was financially supported by Finnish Cultural Foundation, Oskar Öflunds Stiftelse and Societas pro Fauna et Flora Fennica and EJV by Betty Väänäsen Rahasto.