﻿A new species of Pseudopoda (Araneae, Sparassidae) from China, with the description of different and distinctive internal ducts of the female vulva

﻿Abstract One new species of the genus Pseudopoda Jäger, 2000, Pseudopodadeformis Gong & Zhong, sp. nov. (♂, ♀), is described and documented with digital images from Shennongjia Forestry District, Hubei Province, China, based on morphology and DNA barcodes. This new species is separated from other Pseudopoda species by the unique type of internal ducts of the female vulva that are curved longitudinally, forming a narrow triangle or trapezoidal shape. In addition, DNA barcodes for this species are provided.


Introduction
Pseudopoda Jäger, 2000 is currently the largest genus in the family Sparassidae Bertkau, 1872. It comprises 251 species, of which 152 are recorded from China, representing 60.6% of the global species (WSC 2023). The genus has been recorded in areas from South Asia (49 species in Nepal, India, Bhutan, and Pakistan), East Asia (154 species in China and Japan) and Southeast Asia (50 species in Myanmar, Thailand, Laos, and Vietnam) (WSC 2023).
While examining specimens recently collected from Shennongjia Forestry District of Hubei Province, central China, we found some huntsman spiders. The spiders described in this paper were identified as a new species based on comparison with other Pseudopoda species. The male palp of this new species has a slender embolus, and the female vulva has unique internal ducts. We used DNA barcodes of the species to match the sexes and for future use in identification.

Material and methods
Specimens were examined and measured with an Olympus SZX7 stereomicroscope. Positions of tegular appendages are given according to clock positions, based on the left palp in ventral view. Male and female copulatory organs were examined and illustrated after dissection from the spider bodies; vulvae were cleared in a warm 10% potassium hydroxide (KOH) solution. All photographs were captured with a KUY NICE industrial digital camera (20.0 megapixels) mounted on an Olympus CX43 dissecting microscope and assembled using Helicon Focus 3.10.3 image stacking software. Photographic images were then edited using Adobe Photoshop CC 2018. All measurements were obtained using an Olympus SZX7 stereomicroscope and are given in millimetres (mm).
Leg measurements are shown as: total length (femur, patella, tibia, metatarsus, tarsus). Number of macrosetae is listed for each segment in the following order: prolateral, dorsal, retrolateral, ventral; in femora and patellae ventral spines are absent and thus the fourth article is omitted in the setation formula (Gong and Zhong 2020).
To obtain DNA barcodes, one mitochondrial gene (mitochondrial cytochrome oxidase subunit I [COI]) and one nuclear gene (Internal Transcribed Spacer 2 [ITS2]) were amplified and sequenced for four specimens. Primers (Folmer et al. 1994), PCR conditions and other information (e.g., extraction, amplification and sequencing procedures) are the same as in Zhang et al. (2021). The accession numbers are provided in Table 1. For phylogenetic inference, we used the dataset (COI + ITS2) from Cao et al. (2016) and added the new sequences of Pseudopoda deformis Gong & Zhong, sp. nov. Phylogenetic analyses are the same as in Cao et al. (2016) and Zhang et al. (2017). Bayesian inference strongly supported the monophyly of the P. deformis Gong & Zhong,sp. nov. (Fig. 5 (2000) according to the following combination of characters: male palp ( Fig. 1A-C) with membranous conductor or absent, embolus arising on the left side of the tegulum and generally curved, RTA arising from tibia, basally or mesially and furcate or not; epigyne ( Fig Etymology. The specific name is derived from the Latin word deformis, -a, -um, meaning distorted, referring to the shape of the internal ducts of the female vulva.  Gong & Zhong, sp. nov. are similar to those of P. jiangi Zhang, Jäger & Liu, 2023(Zhang et al. 2023, P. lushanensis (Wang, 1990) (Quan et al. 2014: figs 4A-C, 5A-C) and P. shuqiangi Jäger &Vedel, 2007 (Jäger andVedel 2007: figs 73-75) in having a long, filiform embolus. They can be distinguished from the two congeners by the following combination of characters: (1) Embolus arising from tegulum at 1:00-o'clock position, then curving (2) The basal part of embolus is oval (circular in P. jiangi); (3) The tip of the conductor is straight and extends to approximately the tip of the cymbium in ventral view (not in P. lushanensis and P. shuqiangi); and (4) RTA arising medially from tibia (subdistally in P. lushanensis; basally in P. shuqiangi) (Fig. 1A-C). The females of this species can be separated from other Pseudopoda species by their unique internal ducts of the vulva, which are curved longitudinally, forming a narrow triangle or trapezoidal shape (Fig. 2D-I).
Epigynal field almost as wide as long, the anterior margins of lateral lobes forming a V-shape, median margin of lateral lobes united, internal duct systems not visible through cuticle, fertilisation ducts arising postero-laterally. In the dorsal view, internal duct systems differ extremely, and there is no regularity in the direction and structure of internal pipeline (Fig. 2).
Habitat. The specimens were collected on leaves at night with bare hands (Fig. 4A, B). Distribution. Known only from Hubei Province, China (Fig. 4C).
Remarks. The monophyly of Pseudopoda deformis Gong & Zhong, sp. nov. is highly supported by molecular phylogenetic results based mainly on Chinese Pseudopoda species (Fig. 5).

Discussion
We examined all specimens from Shennongjia (4 males, 7 females) and found no variation in the male palp. However, the females had different internal ducts in their vulva, which is not known to occur in other Pseudopoda spiders. In this paper, matching of the sexes of Pseudopoda deformis Gong & Zhong, sp. nov. was done using morphological and molecular data (Figs 3, 5). Females of this species resemble most Pseudopoda  Outgroups species having the median margin of the lateral lobes converged medially with the anterior part V-shaped, but they can be distinguished from these species by the longitudinally bent internal duct system of the vulva ( Fig. 2A-F). As shown in Fig. 2G-I, the female vulva is divided into three parts schematically to show the course of the internal duct system, leading to an interesting discovery. The first part is represented by a red line and is U-shaped in all three females examined. The third part is represented by a green line and is an inverted S-shaped. Variability occurs in the second part which is shown by the blue line, and there are irregularities to this variation.