﻿Yunguirius gen. nov., a new genus of Coelotinae (Araneae, Agelenidae) spiders from southwest China

﻿Abstract A new genus of the subfamily Coelotinae F. O. Pickard-Cambridge, 1893, Yunguiriusgen. nov. is described, comprising two new species and three species previously described in Draconarius Ovtchinnikov, 1999, all from southwest China: Y.duogesp. nov. (♀), Y.xiangdingsp. nov. (♀), Y.ornatus (Wang, Yin, Peng & Xie, 1990) comb. nov. (♂♀) (the type species of Yunguiriusgen. nov.), Y.subterebratus (Zhang, Zhu & Wang, 2017) comb. nov. (♀), and Y.terebratus (Peng & Wang, 1997) comb. nov. (♂♀). Molecular analyses support Yunguiriusgen. nov. as a monophyletic group, with the Sinodraconarius clade as its sister group: Yunguiriusgen. nov. + (Hengconarius + (Nuconarius + Sinodraconarius)).


Introduction
The subfamily Coelotinae F. O. Pickard-Cambridge 1893 (Araneae: Agelenidae) is distributed worldwide (i.e., throughout Asia, Europe and North America) and is represented by 798 species in 37 genera (WSC 2023). Over the past decade, with the concerted efforts of arachnologists, this subfamily has achieved a basic and relatively stable framework, both in morphology and molecular phylogeny amongst the known genera (Chen et al. 2015(Chen et al. , 2016Li 2016, 2017;Okumura 2017Okumura , 2020Zhu et al. 2017;Li et al. 2018aLi et al. -c, 2019bOkumura and Zhao 2022). Herein, we focus on the taxonomy of the paraphyletic Draconarius-clades defined by Zhao and Li (2017).
Recently, when examining specimens collected from southwest China and comparing them with known species in the literature, we realized that they represent two undescribed species, and suspected that they may belong to a new genus. The two species are closely related to D. ornatus (Wang, Yin, Peng & Xie, 1990), D. terebratus (Peng & Wang, 1997), and D. subterebratus Zhang, Zhu & Wang, 2017. Therefore, morphological and phylogenetic studies were carried out on these closely related species to elucidate their taxonomy.

Sampling and morphological examination
All specimens studied in this paper were collected from southwest China. Fresh specimens were preserved in 95% ethanol with storage at -20 °C for DNA extraction and 75% ethanol for morphological study. Specimens were examined with a LEICA M205 C stereomicroscope. Photos were taken with an Olympus C7070 wide zoom digital camera (7.1 megapixels) mounted either on an Olympus SZX12 dissecting microscope or on an Olympus BX51 compound microscope. Images from multiple focal ranges were combined using Helicon Focus v. 6.80 photo stacking software program. The epigyne and male palp were dissected from the body for examination. The epigyne was treated in a warm 10% potassium hydroxide (KOH) solution. Images of the left male palp are presented. Measurements were obtained with a LEICA M205 C stereomicroscope and are given in millimetres. Eye diameters were measured as the maximum distance in either dorsal or frontal views. Leg measurements are given as follows: total length (coxa, trochanter, femur, patella, tibia, metatarsus, tarsus). Terminology follows Wang et al. (1990), Peng and Wang (1997) and Zhu et al. (2017). Abbreviations are as follows:

ALE
anterior lateral eye; ALE-PLE distance between ALE and PLE; AME anterior median eye; AME-ALE distance between AME and ALE; AME-AME distance between AME and AME; AME-PME distance between AME and PME; PLE posterior lateral eye; PME posterior median eye; PME-PLE distance between PME and PLE; PME-PME distance between PME and PME.

IZCAS
Institute of Zoology, Chinese Academy of Sciences;

MHBU
Museum of Hebei University.

Laboratory protocols and phylogenetic analyses
DNA barcodes were obtained for delimiting the species. A partial fragment of the mitochondrial cytochrome oxidase subunit I (CO1) gene was amplified and sequenced for the new and type species using primers LCO1490-oono (5'-CWACAAAYCATAR-RGATATTGG-3') and HCO2198-zz (5'-TAAACTTCCAGGTGACCAAAAAAT-CA-3'), following Zhao and Li (2017) and Zhao et al. (2020). GenBank accession numbers of CO1 are listed separately in Table 1.
To perform phylogenetic analyses, part of the molecular data of coelotine spiders from Zhao and Li (2017), Zhao et al. (2020), and Okumura and Zhao (2022) were collected. The new molecular dataset consists of eight genes: CO1, NADH dehydrogenase subunit I (ND1) gene, histone 3 (H3) gene, wingless gene and the ribosomal RNA genes 12S, 16S, 18S, and 28S. They were assembled from 72 species, 67 known species (with 26 type species from different genera) in 32 known genera of Coelotinae as the ingroup, and three species of Ageleninae and one species of Amaurobiidae as the outgroup, alongside three new sequences. GenBank accession numbers for all the above genes are shown in Suppl. material 1.
Phylogenetic relationships were inferred using both maximum likelihood (ML) and Bayesian inference (BI). First, the best-fit partitioning schemes and models were selected for the RAxML and MrBayes analyses using PartitionFinder v.2.1.1 (Lanfear et al. 2012). ML analysis was conducted in RAxML v.8.0.0 (Stamatakis 2006) using the GTRCAT substitution model for all partitions (partitioned by gene). A rapid bootstrap of 1, 000 replicate ML inferences was performed to determine the best-scoring ML tree and nodal support values. BI analyses were performed in MrBayes v.3.2.2 (Ronquist and Huelsenbeck 2003) with posterior distributions estimated by Markov chain Monte Carlo (MCMC) sampling. The appropriate model was selected for each partition (gene): the GTR + I + G model was favored for each partition, except that different models were selected for H3 (HKY + I + G), wingless (SYM + I + G) and 18S (K80 + I + G). Two simultaneous runs with four MCMC chains were performed for 10 million generations to ensure that the average standard deviation of the split frequency was below 0.01 and to obtain a well-supported consensus tree. Additional ML analysis was performed in IQ-TREE (Nguyen et al. 2015) using the ModelFinder function (-m MFP + MERGE) to select the best-fit model for each partition, and the option '-bb 1,000' to estimate nodal support values.

Results and discussion
By examining specimens collected from southwest China, we found that two species with particular external genital morphology could not be placed into existing genera. They are morphologically similar to three Draconarius species, D. ornatus, D. terebratus, and D. subterebratus (Wang 2003;Zhu et al. 2017). The epigynes of these five species all lack epigynal teeth but have a large central atrium. In the vulva, the copulatory ducts are broad, anteriorly extended and curved, and the spermathecal stalks are elongated. Males also show similar homologous characteristics, such as a thick embolus beginning at a 7 o'clock position, and a short cymbial furrow less than half the length of cymbium, although only two males out of five species have been described so far. All species are closely related to each other by the comprehensive characteristics mentioned above and differ from the type species D. venustus Ovtchinnikov, 1999 and the venustus group of Draconarius which share a pair of triangular epigynal teeth commonly (Wang 2003;Li et al. 2019a). Therefore, we establish a new genus, Yunguirius gen. nov., and herein transfer the three Draconarius species to it.
Our different phylogenetic analyses infer similar tree topologies ( Fig. 1) and strongly support Yunguirius gen. nov. as a monophyletic group (ML rapid bootstrap = 100 and 95; BI posterior probability = 1.00). Although the relationships within the        genus are unclear (two species lack molecular data), the other three species are indeed genetically closely related. The genus is sister to the Sinodraconarius clade (Hengconarius + (Nuconarius + Sinodraconarius)) and genetically distant from the genus Draconarius. The close relationship between Yunguirius gen. nov. and the Sinodraconarius clade can also be confirmed by having common morphological features such as bifurcated conductors and absent epigynal teeth, which obviously differ from Draconarius. Geographically, species of Yunguirius gen. nov. are restricted to southwest China (Fig. 5). Zoogeographic studies suggest that the genus-level distribution of coelotine spiders is regional, and the divergence and formation of these monophyletic genera are closely related to geological and climatic events in Eurasia (Zhao and Li 2017;Zhao et al. 2022 Etymology. The generic name is derived from the pinyin word "Yungui", referring to Yunnan-Guizhou Plateau where the genus is distributed, and "-rius" refers to the genus as part of its sister groups of genera: Nuconarius, Hengconarius, and Sinodraconarius. The gender is masculine.
Diagnosis. Morphological characteristics of Yunguirius gen. nov. resemble those of Nuconarius, Hengconarius, and Sinodraconarius, which are distributed in southeastern China, by cymbial furrow short, with a length less than half of cymbium ( Description. Small to very large spiders, body length 6.00 to 21.80. Carapace brown to black, tonneau-shaped, longer than or as long as abdomen, with longitudinal fovea and dark radial grooves; chelicerae black, with three promarginal and two retromarginal teeth; endites and labium brown to dark brown, anterior white with black hairs; sternum brownish to brown, longer than wide. Abdomen yellowishbrown, nearly oval, posterior widest, with four to six darker chevrons or speckles, or without any pattern. Leg formula 4 > 1 > 2 > 3 or 1 > 4 > 2 > 3. Male palp: patellar apophysis finger-shaped, retrolateral tibial apophysis large, lateral tibial apophysis small, median apophysis spoon-shaped; conductor large, with two branches; embolus thick, beginning at a 7 o'clock position, embolic base swollen; cymbial furrow short, with the length less than half of cymbium. Epigyne: posterior epigynal sclerite varying in shape; atrium very large, wide to narrow, with osteosclerotic lateral margin, inside white osteon, outside with brownish or brown markings and brown or darker folds; copulatory duct membranous, arising posteriorly, extending to anterior, opening anteriorly; spermatheca brown, spermathecal base swollen, spermathecal head long and line-shaped, extending anteriorly, opposite end swollen, lamellar or connected with a stalk. Distribution. Guizhou, Hunan, Sichuan and Yunnan, China (Fig. 5). Diagnosis. Yunguirius duoge sp. nov. resembles Y. terebratus by having rectangular posterior epigynal sclerite, subrounded atrium and dumbbell-shaped spermatheca at its first half. However, it can be distinguished from Y. terebratus as follows: 1) posterior margin of the epigyne narrow and pointed in the middle ( Fig. 2A) in Y. duoge sp. nov., but flat ( fig. 28 in Peng and Wang 1997) in Y. terebratus; 2) epigynal folds banded ( Fig. 2A) in Y. duoge sp. nov., but dentiform ( fig. 28 in Peng and Wang 1997) in Y. terebratus; 3) anterior copulatory duct close to each other (Fig. 2B) in Y. duoge sp. nov., but lapped ( fig. 29 in Peng and Wang 1997) in Y. terebratus; and 4) stalk of spermatheca extending laterally (Fig. 2B) in Y. duoge sp. nov., but extending anteriorly ( fig. 29 in Peng and Wang 1997) in Y. terebratus.