Research Article |
Corresponding author: Chun-Cai Yan ( skyycc@mail.tjnu.edu.cn ) Corresponding author: Xiao-Long Lin ( lin880224@gmail.com ) Academic editor: Fabio Laurindo da Silva
© 2023 Wen-Bin Liu, Ying Wang, Kang-Zhu Zhao, Cheng-Yan Wang, Jun-Yu Zhang, Chun-Cai Yan, Xiao-Long Lin.
This 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.
Citation:
Liu W-B, Wang Y, Zhao K-Z, Wang C-Y, Zhang J-Y, Yan C-C, Lin X-L (2023) New species, a new combination, and DNA barcodes of Parachironomus Lenz, 1921 (Diptera, Chironomidae). ZooKeys 1153: 121-140. https://doi.org/10.3897/zookeys.1153.98542
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The genus Parachironomus has a cosmopolitan distribution including 85 valid described species worldwide. Species records and studies of the genus in the Tibetan Plateau are scarce. In this study, the genus Parachironomus from China is revised and two new species, Parachironomus wangi Liu & Lin, sp. nov. and Parachironomus nankaiensis Liu & Lin, sp. nov., are described based on adult morphology and molecular data. Paracladopelma demissum Yan, Wang & Bu is placed in the genus Parachironomus as a new combination. A neighbor-joining tree was reconstructed based on all known Parachironomus COI DNA barcodes. A key to adult males of the genus Parachironomus from China is also provided.
COI, new combination, new species, taxonomy, Tibetan Plateau
The genus Parachironomus was erected by
The DNA barcodes corresponding to the 658-bp fragment of the mitochondrial gene cytochrome c oxidase I (COI) has been identified as the core of a global bio-identification system at the species level (
The Tibetan Plateau is located in southwest China, with a vast territory and diverse terrain. The Tibetan Plateau is one of the most important areas of biodiversity in the world because of its unique environmental and regional units, which breed unique biological communities and many unique and rare wild animals and plants. Some interesting species were discovered during the investigations of insect diversity in the Tibetan Plateau. In this paper, one new combination and two new species are proposed and described. The partial COI sequences of species in Parachironomus with DNA barcode analysis is conducted. A key to the known Chinese adult males of the genus is presented.
The examined specimens were caught using sweep net and light trap, stored in the dark at 4 °C, and preserved in 85% ethanol before molecular and morphological analyses. Genomic DNA was extracted from the thorax and leg using a Qiagen DNA Blood and Tissue Kit at Tianjin Normal University, Tianjin, China (TJNU), following the standard protocol except for the final elution volume of 100 µl. After DNA extraction, the exoskeleton of each specimen was mounted in Euparal on a microscope slide together with the corresponding antennae, legs, wing, and abdomen, following the procedures outlined by
The color pattern of all species is described based on the specimen preserved in ethanol. Digital photographs of slide-mounted specimens were taken with a 300-dpi resolution using Nikon Eclipse 80i with Nikon Digital Sight DS-Fil camera at TJNU.
The universal primers LCO1490 and HCO2198 (
Raw sequences were assembled and edited in Geneious Prime 2020 (Biomatters Ltd., Auckland, New Zealand). Alignment of the sequences was carried out using the MUSCLE (
In this study, the partial COI sequences of Parachironomus were submitted to online ABGD web interface (https://bioinfo.mnhn.fr/abi/public/abgd/abgdweb.html). We used the K2P nucleotide substitution model. The prior intraspecific divergence was set at between 0.001 and 0.1. The minimum relative gap width was 1.0 and other parameters were defaulted.
The holotype of two new species is deposited at the College of Fisheries and Life Science, Shanghai Ocean University (SHOU) and College of Life Sciences, Tianjin Normal University, Tianjin, China (TJNU).
In this study, five COI sequences were obtained, and 19 COI sequences of Parachironomus were downloaded from BOLD, totaling 24 COI sequences. All sequences could be translated successfully into amino acids without indels and stop codons. MEGA analysis showed that the average total length of the sequence was 658 bp, and that there were 427 conserved sites, 231 variable sites, 196 parsimony informative sites, and 35 singleton sites. The mean nucleotide base compositions were 27.1% A, 17.9% C, 16.2% G, and 38.8% T for COI genes (Table
The contents and nucleotide substitutions of COI gene sequences of Parachironomus.
T (%) | C (%) | A (%) | G (%) | ii | si | sv | R | |
---|---|---|---|---|---|---|---|---|
1st | 48.3 | 8.2 | 40.0 | 3.5 | 144 | 30 | 44 | 0.7 |
2nd | 25.3 | 18.5 | 27.6 | 28.6 | 205 | 11 | 1 | 13.3 |
3rd | 42.9 | 27.0 | 13.6 | 16.5 | 216 | 0 | 0 | 3 |
Avg | 38.8 | 17.9 | 27.1 | 16.2 | 565 | 41 | 44 | 0.9 |
TT | TC | TA | TG | CT | CC | CA | CG | |
1st | 76 | 11 | 18 | 1 | 10 | 5 | 2 | 0 |
2nd | 49 | 5 | 0 | 0 | 5 | 35 | 0 | 0 |
3rd | 93 | 0 | 0 | 0 | 0 | 58 | 0 | 0 |
Avg | 218 | 16 | 18 | 2 | 14 | 98 | 2 | 0 |
AT | AC | AA | AG | GT | GC | GA | GG | |
1st | 18 | 2 | 62 | 5 | 1 | 0 | 4 | 2 |
2nd | 0 | 0 | 59 | 1 | 0 | 0 | 1 | 61 |
3rd | 0 | 0 | 29 | 0 | 0 | 0 | 0 | 36 |
Avg | 18 | 2 | 151 | 6 | 2 | 0 | 4 | 98 |
The neighbor joining tree based on available COI DNA barcodes of the Parachironomus revealed two species new to science (Fig.
Kimura 2-parameter pairwise genetic distances based on COI barcodes of the Parachironomus.
Species | Pairwise genetic distances | ||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
P. tenuicaudatus|MG172661 | |||||||||||||||||||||||
P. varus|MZ606874 | 0.13 | ||||||||||||||||||||||
P. digitalis|MZ627020 | 0.14 | 0.13 | |||||||||||||||||||||
P. frequens|MZ629002 | 0.15 | 0.15 | 0.13 | ||||||||||||||||||||
P. parilis|MZ627840 | 0.13 | 0.09 | 0.14 | 0.14 | |||||||||||||||||||
P. gracilior|MZ624787 | 0.13 | 0.12 | 0.14 | 0.13 | 0.10 | ||||||||||||||||||
P. siljanensis|KC250820 | 0.14 | 0.10 | 0.13 | 0.15 | 0.11 | 0.11 | |||||||||||||||||
P. cf. vitiosus|HQ937673 | 0.14 | 0.13 | 0.15 | 0.13 | 0.15 | 0.15 | 0.14 | ||||||||||||||||
P. monochromus|MZ657902 | 0.15 | 0.14 | 0.14 | 0.14 | 0.14 | 0.15 | 0.15 | 0.13 | |||||||||||||||
P. vitiosus|MZ660486 | 0.12 | 0.14 | 0.14 | 0.13 | 0.13 | 0.15 | 0.14 | 0.07 | 0.13 | ||||||||||||||
P. subalpinus|JF870871 | 0.10 | 0.13 | 0.14 | 0.14 | 0.12 | 0.13 | 0.14 | 0.14 | 0.14 | 0.14 | |||||||||||||
P. elodeae|KM571020 | 0.12 | 0.10 | 0.14 | 0.14 | 0.12 | 0.13 | 0.13 | 0.11 | 0.13 | 0.11 | 0.11 | ||||||||||||
P. vitiosus group | 0.14 | 0.13 | 0.15 | 0.13 | 0.14 | 0.15 | 0.14 | 0.00 | 0.13 | 0.08 | 0.13 | 0.11 | |||||||||||
P. potamogeti | 0.13 | 0.12 | 0.14 | 0.14 | 0.14 | 0.15 | 0.14 | 0.15 | 0.12 | 0.13 | 0.12 | 0.12 | 0.15 | ||||||||||
P. swammerdami|LC329152 | 0.13 | 0.12 | 0.14 | 0.12 | 0.13 | 0.14 | 0.12 | 0.13 | 0.15 | 0.12 | 0.13 | 0.11 | 0.13 | 0.12 | |||||||||
P. delinificus|KC750457 | 0.14 | 0.13 | 0.13 | 0.11 | 0.14 | 0.13 | 0.14 | 0.13 | 0.14 | 0.13 | 0.14 | 0.13 | 0.13 | 0.14 | 0.14 | ||||||||
P. cayapo | 0.15 | 0.13 | 0.16 | 0.14 | 0.14 | 0.15 | 0.14 | 0.15 | 0.14 | 0.14 | 0.15 | 0.13 | 0.15 | 0.15 | 0.14 | 0.14 | |||||||
P. paradigitalis|MZ660327 | 0.12 | 0.07 | 0.13 | 0.14 | 0.10 | 0.13 | 0.11 | 0.11 | 0.13 | 0.13 | 0.13 | 0.09 | 0.11 | 0.13 | 0.10 | 0.12 | 0.12 | ||||||
P. biannulatus|MZ658910 | 0.13 | 0.11 | 0.15 | 0.15 | 0.12 | 0.15 | 0.13 | 0.13 | 0.13 | 0.13 | 0.13 | 0.09 | 0.13 | 0.12 | 0.11 | 0.15 | 0.14 | 0.10 | |||||
P. demissum comb. nov. |XL592 | 0.14 | 0.14 | 0.11 | 0.13 | 0.14 | 0.13 | 0.14 | 0.13 | 0.15 | 0.13 | 0.13 | 0.13 | 0.12 | 0.13 | 0.13 | 0.12 | 0.14 | 0.13 | 0.15 | ||||
P. nankaiensis sp. nov. |XL599 | 0.14 | 0.13 | 0.12 | 0.13 | 0.13 | 0.12 | 0.14 | 0.14 | 0.15 | 0.14 | 0.12 | 0.13 | 0.14 | 0.15 | 0.14 | 0.13 | 0.13 | 0.12 | 0.15 | 0.13 | |||
P. demissum comb. nov. |XL590 | 0.14 | 0.14 | 0.11 | 0.13 | 0.14 | 0.13 | 0.13 | 0.12 | 0.14 | 0.12 | 0.12 | 0.13 | 0.12 | 0.13 | 0.13 | 0.12 | 0.13 | 0.12 | 0.15 | 0.01 | 0.12 | ||
P. wangi sp. nov. |XL601 | 0.14 | 0.14 | 0.14 | 0.15 | 0.14 | 0.15 | 0.16 | 0.14 | 0.15 | 0.14 | 0.14 | 0.14 | 0.14 | 0.15 | 0.12 | 0.14 | 0.16 | 0.14 | 0.15 | 0.12 | 0.14 | 0.12 | |
P. wangi sp. nov. |XL602 | 0.14 | 0.14 | 0.14 | 0.15 | 0.14 | 0.15 | 0.16 | 0.14 | 0.15 | 0.14 | 0.14 | 0.14 | 0.14 | 0.15 | 0.12 | 0.14 | 0.16 | 0.14 | 0.15 | 0.12 | 0.14 | 0.12 | 0.00 |
When the interspecific genetic distance is greater than the intraspecific genetic distance, barcode gaps will appear through the frequency histogram of genetic data. There is an obvious barcode gap in the genetic distance of all Parachironomus COI sequences, which fully confirms the feasibility of COI as a DNA barcode (Fig.
Paracladopelma demissum Yan, Wang & Bu, 2012: 291.
Holotype. Male (TJNU: 11430), China, Sichuan Province, Yajiang County, Sandaoqiao Town, 30.01532°N, 101.05134°E, 2460 m a.s.l., 9.VI.1996, light trap, leg: X. H. Wang. Paratypes. One male (TJNU: 11730), China, Sichuan Province, Ya’an City, Baoxing County, Xinglong Elementary School, Xihe River, 30.25800°N, 102.50387°E, 1100 m a.s.l., 19.VI.1996, light trap, leg: X. H. Wang. One male (TJNU: 12886), China, Sichuan Province, Kangding County, Wasigou River, 30.04191°N, 102.09454°E, 2000 m a.s.l., 15.VI.1996, light trap, leg: X. H. Wang. One male (TJNU: 13079), China, Sichuan Province, Shimian County, Nanyahe River, 29.11156°N, 102.23265°E, 1040 m a.s.l., 16.VI.1996, light trap, leg: X. H. Wang.
Two males (SHOU: XL590, XL592), China, Xizang Autonomous Region, Rikaze City, Yadong County, Xiasima Town, 27.48986°N, 88.90572°E, 3032 m a.s.l., 20.VII.2014, light trap, leg: X. L. Lin.
The species can be distinguished from known species of Parachironomus by the following combination of characters: AR 0.58–0.67, 0.62; tergite IX with shoulder-like posterior margin; anal point triangular base, constricted in the middle, with a ridge mesally; anal tergite bands V-shaped and fused; superior volsella with a bare triangular projection apically; gonostylus slender, parallel-sided, curved medially, gradually tapered to the top.
Adult males (n = 6, unless otherwise stated). Total length 3.08–3.60, 3.35 mm. Wing length 1.60–1.89, 1.75 mm. Total length/wing length 1.81–2.10, 1.93. Wing length/length of profemur 2.16–2.63, 2.32.
Coloration. Thorax yellowish brown with pale brown spots. Femora and tibiae of front legs yellowish green with distal parts brown, anterior 1/2 of tarsi I yellowish green, remainder of front legs dark brown; femora, tibiae and basal 1/2 of tarsi I of mid and hind legs yellowish green, remaining dark brown. Abdomen yellowish green to dark brown, with tergites I–V yellowish green, tergites VII, VIII, and hypopygium dark brown.
Head
(Fig.
Head (A–C) and thorax (D–F) of Parachironomus species A P. demissum (Yan, Wang & Bu), comb. nov. B P. wangi Liu & Lin, sp. nov. C P. nankaiensis Liu & Lin, sp. nov. D P. demissum (Yan, Wang & Bu), comb. nov. E P. wangi Liu & Lin, sp. nov. F P. nankaiensis Liu & Lin, sp. nov. Scale bars: 200 μm.
Thorax
(Fig.
Wing
(Fig.
Legs. Front tibia with 3 subapical setae, 80–84, 82; 83–97, 91 and 85–112, 101 (5) μm long. Combs of mid tibia 37–41, 39 μm wide with 18–25, 22 μm long spur, and 41–55, 48 μm wide with 20–34, 27 μm long spur; combs of hind tibia 37–41, 39 μm wide with 23–27, 25 μm long spur, 60–66, 63 μm wide with 28–34, 32 μm long spur. Tarsomere 1 of mid and hind legs without sensilla chaetica. Lengths (in μm) and proportions of legs as in Table
Lengths (in μm) and proportions of legs of Parachironomus demissum (Yan, Wang & Bu), comb. nov., adult male (n = 6).
fe | ti | ta1 | ta2 | ta3 | |
---|---|---|---|---|---|
P1 | 632–815, 755 | 498–640, 586 | 901–1049, 996 | 341–434, 407 | 276–352, 329 |
P2 | 575–810, 731 | 524–660, 600 | 317–370, 349 | 134–180, 169 | 99–133, 122 |
P3 | 730–910, 841 | 698–890, 802 | 496–549, 521 | 245–290, 275 | 187–246, 221 |
ta4 | ta5 | LR | BV | SV | |
P1 | 273–326, 305 | 152–170, 162 | 1.63–1.81, 1.69 | 1.94–1.95, 1.94 | 1.25–1.37, 1.34 |
P2 | 64–90, 84 | 66–70, 69 | 0.55–0.64, 0.58 | 3.75–3.90, 3.79 | 3.47–3.93, 3.81 |
P3 | 112–140, 126 | 80–96, 87 | 0.61–0.71, 0.65 | 3.01–3.07, 3.05 | 2.88–3.22, 3.15 |
Hypopygium
(Figs
Hypopygium A P. demissum (Yan, Wang & Bu), comb. nov. B P. wangi Liu & Lin, sp. nov. C P. nankaiensis Liu & Lin, sp. nov. Anal point D P. demissum (Yan, Wang & Bu), comb. nov. E P. wangi Liu & Lin, sp. nov. F P. nankaiensis Liu & Lin, sp. nov. Superior volsella G P. demissum (Yan, Wang & Bu), comb. Nov. H P. wangi Liu & Lin, sp. nov. I P. nankaiensis Liu & Lin, sp. nov. Scale bars: 50 μm.
China (Sichuan and Xizang).
Holotype. Male (SHOU: XL601), China, Xizang Autonomous Region, Shannan City, Gongga County, Gangdui Town, 29.27888°N, 90.82323°E, 3586 m a.s.l., 18.VII.2014, sweep net, leg: X.L. Lin. Paratypes. One male (TJNU: XL602), collecting data as holotype.
The species can be distinguished from known species of Parachironomus by the following combination of characters: frontal tubercles absent; tergite IX with triangular posterior margin; anal point slightly wider at base, parallel-sided, swollen apically; superior volsella wide, parallel-sided, bent inward at 1/3 distance from apex, apically rounded, free microtrichia; inferior volsella not reaching beyond anal tergite margin; gonostylus gradually widened distally, curved and parallel-sided, apically rounded.
Adult males (n = 2, unless otherwise stated). Total length 3.63–3.91, 3.77 mm. Wing length 1.85–1.91, 1.88 mm. Total length/wing length 1.96–2.05, 2.01. Wing length/length of profemur 2.74–2.79, 2.77.
Coloration. Thorax yellowish brown with pale brown spots. Front legs dark brown; femora and basal 1/3 of tarsi I of mid and hind legs yellowish brown, remaining dark brown. Abdomen pale yellow to yellowish brown, with tergites I–VI pale yellow, tergites VII, VIII, and hypopygium yellowish brown.
Head
(Fig.
Thorax
(Fig.
Wing
(Fig.
Legs. Front tibia with three subapical setae, 82–91, 87; 84 (1) and 105 (1) μm long. Combs of mid tibia 26–33, 30 μm wide with 15–25, 20 μm long spur, and 21 μm wide with 28 μm long spur; combs of hind tibia 22–26, 24 μm wide with 22–34, 28 μm long spur, 38–42, 40 μm wide with 34–38, 36 μm long spur. Tarsomere 1 of mid and hind legs without sensilla chaetica. Lengths (in μm) and proportions of legs as in Table
Lengths (in μm) and proportions of legs of Parachironomus wangi Liu & Lin, sp. nov., adult male (n = 2).
fe | ti | ta1 | ta2 | ta3 | |
---|---|---|---|---|---|
P1 | 664–695, 680 | 469–503, 486 | 858–940, 899 | 455–489, 472 | 337–355, 346 |
P2 | 692–700, 696 | 604–638, 621 | 348–362, 355 | 171–189, 180 | 126–135, 131 |
P3 | 726–751, 739 | 783–801, 792 | 560–594, 577 | 291–302, 297 | 220–231, 226 |
ta4 | ta5 | LR | BV | SV | |
P1 | 282–290, 286 | 147–152, 150 | 1.83–1.87, 1.85 | 1.62–1.67, 1.65 | 1.27–1.32, 1.30 |
P2 | 83–84, 84 | 75–78, 77 | 0.57–0.58, 0.57 | 3.50–3.61, 3.56 | 3.70–3.72, 3.71 |
P3 | 127–130, 129 | 90–93, 92 | 0.72–0.74, 0.73 | 2.82–2.86, 2.84 | 2.61–2.69, 2.65 |
Hypopygium
(Figs
Name after Prof. Xin-Hua Wang, for his outstanding contribution towards increasing our knowledge of aquatic insect taxonomy; noun in nominative case.
China (Xizang).
Holotype. Male (SHOU: XL599), China, Xizang Autonomous Region, Shannan City, Gongga County, Gangdui Town, 29.27888°N, 90.82323°E, 3586 m a.s.l., 18.VII.2014, sweep net, leg: X.L. Lin.
The species can be distinguished from known species of Parachironomus by the following combination of characters: frontal tubercles small; squama with seven setae; anal tergite bands V-shaped, separated; superior volsella narrower at base, curved and expanded in the distal part, with a bare lamellar projection as wide as apex of volsella; inferior volsella reaching slightly beyond anal tergite margin; gonostylus slender, slightly curved in the middle, tapered to the apex.
Adult male (n = 1). Total length 3.90 mm. Wing length 2.51 mm. Total length/wing length 1.56. Wing length/length of profemur 2.64.
Coloration. Thorax dark brown with dark spots. Legs brown. Abdomen yellowish green to brown, tergites I–V yellowish green, tergites VI–VIII yellowish brown, hypopygium brown.
Head
(Fig.
Thorax
(Fig.
Wing
(Fig.
Legs. Front tibia with three subapical setae, 80, 87 and 93 μm long. Combs of mid tibia 33 μm wide with 19 μm long spur, and 29 μm wide with 27 μm long spur; combs of hind tibia 30 μm wide with 20 μm long spur, 65 μm wide with 36 μm long spur. Tarsomere 1 of mid and hind legs without sensilla chaetica. Lengths (in μm) and proportions of legs as in Table
Lengths (in μm) and proportions of legs of Parachironomus nankaiensis Liu & Lin, sp. nov., adult male (n = 1).
fe | ti | ta1 | ta2 | ta3 | |
---|---|---|---|---|---|
P1 | 950 | 744 | 1135 | 576 | 431 |
P2 | 889 | 825 | 448 | 258 | 187 |
P3 | 1056 | 1072 | 675 | 385 | 292 |
ta4 | ta5 | LR | BV | SV | |
P1 | 315 | 140 | 1.53 | 1.94 | 1.49 |
P2 | 120 | 91 | 0.54 | 3.30 | 3.83 |
P3 | 162 | 89 | 0.63 | 3.02 | 3.15 |
Hypopygium
(Figs
Name after Nankai University, the institution of study and work for Prof. Xin-Hua Wang; noun in nominative case.
China (Xizang).
1 | Tergite IX with shoulder-like caudal margin | 2 |
– | Tergite IX with triangle caudal margin | 4 |
2 | Superior volsella without projection; gonostylus with constriction in middle | P. frequens (Johannsen) |
– | Superior volsella with a projection; gonostylus gradually tapered to the top | 3 |
3 | AR 0.58–0.67; anal tergite bands fused; inferior volsella not reaching beyond anal tergite margin | P. demissum (Yan, Wang & Bu), comb. nov. |
– | AR 1.60; anal tergite bands separated; inferior volsella reaching slightly beyond anal tergite margin | P. nankaiensis Liu & Lin, sp. nov. |
4 | Superior volsella short, bearing two apical setae and with folds on inner margin; gonostylus widened basally | P. gracilior (Kieffer) |
– | Superior volsella elongate, bearing an apical seta and a subapical seta and without folds; gonostylus widened distally | 5 |
5 | Mid and hind tibiae each with 1 spur; superior volsella straightly, widened at base | P. poyangensis Yan |
– | Mid and hind tibiae each with 2 spurs; superior volsella curved, widened in the distal part | 6 |
6 | Superior volsella slightly curved; gonostylus slender, with distinct expansion in distal 1/3 | P. monochromus (van der Wulp) |
– | Superior volsella bent inward at1/3 distance from apex; gonostylus parallel-sided, gradually widened distally | P. wangi Liu & Lin, sp. nov. |
In this study, the holotype of Paracladopelma demissum were examined, and the original description has been modified. The distinguishing feature of Parachironomus are that the superior volsella usually has a distinct preapical tooth as well as setae arising from distinct pits (
Parachironomus wangi Liu & Lin, sp. nov. resembles Parachironomus biannulatus Staeger, 1839 in having similar shapes of the superior volsella and posterior margin of tergite IX, but can be separated by the following combination characters: AR 1.54–1.55, anal point parallel-sided and gonostylus expanded apically in P. wangi Liu & Lin, sp. nov, whereas AR 3.0–3.6, the anal point is constricted in the middle, and the gonostylus is tapered to the apex in P. biannulatus.
Parachironomus nankaiensis Liu & Lin, sp. nov. resembles Parachironomus cayapo Spies, Fittkau & Reiss, 1994 in having similar shapes of anal point, inferior volsella, and anal tergite bands, but can be separated from the latter by the following combination characters: squama seven setae, the superior volsella expanded in the distal part and with a bare lamellar projection, plus with the gonostylus tapered to the apex. In contrast, the squama of P. cayapo is bare, the superior volsella is not widened in the distal part and has no projection, and the gonostylus has a protruding hump.
The results of molecular identification and morphological taxonomy are consistent, indicating that DNA barcodes and traditional morphological taxonomy are complementary in this case; therefore, the DNA barcode can be used as a simple method to supplement traditional morphological taxonomy for Parachironomus.
Biogeographically, the three species examined in this study are all distributed in the Tibetan Plateau at an altitude of more than 3,000 meters (Fig.
In conclusion, this study not only enriches the database of Chironomidae in China, but also provides baseline data for the protection of the environment and biodiversity in the Tibetan Plateau.
We thank Dr. M. Spies (Zoologische Staatssammlung München, Germany) for providing input on various levels of this work. Financial support from National Natural Science Foundation of China is gratefully acknowledged (grant numbers: 32170473, 31801994, 31900344).