﻿Three new species of the primitively segmented spider genus Songthela (Mesothelae, Liphistiidae, Heptathelinae) from Hunan Province, China

﻿Abstract Three new species of the primitively segmented spider genus Songthela Ono, 2000 are identified and described from Hunan Province, China, based on morphological characters of males and females: S.anhua Zhang & Xu, sp. nov. (♂♀), S.longhui Zhang & Xu, sp. nov. (♂♀), and S.zhongpo Zhang & Xu, sp. nov. (♂♀). All the new Songthela species belong to the multidentata-group according to male palp and female genital morphology.


Introduction
The primitively segmented spider family Liphistiidae Thorell, 1869 is the basal lineage among spiders, which contains species with a limited dispersal ability and high endemicity (Bristowe 1976;Haupt 2003;Xu et al. 2015a, b). Members of liphistiids retain some plesiomorphic arachnid traits, such as abdominal tergites (Fig. 1) and spinnerets situated in the middle of abdominal venter (Bristowe 1975;Haupt 2003;  Schwendinger and Ono 2011;Xu et al. 2015a). Although a few alternative taxonomic classifications for this group did exist (Kishida 1923;Haupt 1983Haupt , 2003Ono 2000;Schwendinger and Ono 2011), the classification system of a single family Liphistiidae consisting of eight genera in two subfamilies, Liphistiinae Thorell, 1869 and Heptathelinae Kishida, 1923, was established based on the evidence from morphology, monophyly, phylogeny, and fossils (Xu et al. 2015a, b), and has been widely accepted by most arachnologists since then (Schwendinger et al. 2019;Ono and Aung 2020;Yu et al. 2021). Recently, Li (2022) elevated the two subfamilies, Liphistiinae and Heptathelinae, to family ranks, Liphistiidae and Heptathelidae, but provided no additional justification for the taxonomic classification change. Breitling (2022) argued against Li's two-family classification system based on nomenclature, morphology, molecular data and fossils, and recommended to maintain a single family Liphistiidae. We agree with Breitling's argumentation, and follow the classification that we have previously proposed (Xu et al. 2015a(Xu et al. , b, 2021, which supports two subfamilies in the family Liphistiidae because all members of Liphistiidae share the same plesiomorphic traits mentioned above, although the members of Liphistiinae have signal lines around their burrow entrances, which are often used as one of the diagnostic characters to separate Liphistiinae from Heptathelinae. Currently, the genus Songthela Ono, 2000, belonging to the subfamily Heptathelinae, contains 35 described species, of which, 34 are distributed in southern China (Chongqing, Guizhou, Hubei, Hunan, Sichuan, Yunnan, Zhejiang); one species, S. sapana (Ono, 2010) is found in northern Vietnam (World Spider Catalog 2023). Until now, 20 Songthela species are known from Hunan Province, and they are divided into three species groups based on morphology and molecular data: bispina-group, multidentata-group, and unispina-group (details see Li et al. 2022).
In this study, we diagnose and describe three new Songthela species collected from Hunan Province, China based on male palp and female genital morphology.

Materials and methods
We collected the specimens alive from Hunan Province, China (Fig. 2), brought subadults back to the laboratory, and reared them until they reached maturation. We removed the right four legs of adults, preserved them in 100% ethanol, and stored them at -80 °C for molecular work. We preserved the remaining body of each specimen in 80% ethanol as vouchers for morphological examination. All type and voucher specimens are deposited at the College of Life Sciences, Hunan Normal University, Changsha, Hunan Province, China.
We examined and dissected the specimens using an Olympus SZ61 stereomicroscope. The soft tissues of female genitalia were removed and degraded using 10 mg/ml pancreatin (Bomei Biotech Company, Hefei, Anhui, China) for at least 3 h at the room temperature. Male palp and female genitalia were observed and photographed using the digital camera CCD mounted on an Olympus BX53 compound microscope, and then generated compound focused images using Helicon Focus v.6.7.1. All measurements are given in millimeters. Leg and palp measurements are given in the following order: leg total length (femur + patella + tibia + metatarsus + tarsus), palp total length (femur + patella + tibia + tarsus).
Abbreviations used are: ALE = anterior lateral eyes; AME = anterior median eyes; BC = bursa copulatrix; BL = body length; CL = carapace length; Co = conductor; CT = contrategulum; CW = carapace width; DT = dorsal extension of terminal apophysis of tegulum; E = embolus; GA = genital area; GS = genital stalk; MA = marginal apophysis of tegulum; OL = opisthosoma length; OW = opisthosoma width; PC = paracymbium; PLE = posterior lateral eyes; PME = posterior median eyes; RC = receptacular cluster; T = tegulum; TA = terminal apophysis of tegulum. Diagnosis. Males of Songthela differ from those of all other Heptathelinae genera by smooth conductor with one or two apical spines and by conductor middle portion having several teeth (Figs 3B, E, 4A-C, 6A, B); by embolus having a wide and flat opening (Figs 3D, 4D, 6D); and by contrategulum having densely serrated margin (Figs 3D, 4D, 6D); females of Songthela can be distinguished from those of all other liphistiid genera by two pairs of receptacular clusters separated from each other, median ones with obviously tubular genital stalks, four receptacular clusters situated at the anterior margin of the bursa copulatrix, or middle ones situated at the anterior margin of the bursa copulatrix and lateral ones located relatively dorsolaterally, or all four located dorsally (Figs 3J-M, 5A-F, 7A-H).

Songthela anhua
Palp. Prolateral portion of paracymbium unpigmented and unsclerotised, with several setae and spines on the tip (Fig. 3A-C). Contrategulum with an obviously triangular apophysis proximally and two irregular dentate edges distally (Fig. 3C, D). Tegulum with a helicoid marginal apophysis, a dentate dorsal extension of the terminal apophysis, and a thumb-like terminal apophysis retrolaterally (Fig. 3F, G). Conductor lamellar, fused with embolus ventroproximally, with a blade-shaped apical spine pointed to the one-third of opening of embolus proximally, and the middle portion inserted with several teeth (Fig. 3B-E, H, I). Embolus largely sclerotized with a wide opening, several ribbed ridges, and with a twisted top in ventral view ( Fig. 3B-E, G).
Female (XUX-2021-008; Fig. 1G). Carapace yellow brown and opisthosoma dark brown in alcohol, with 12 dark brown tergites attached a pair of thick bristles, the second to fifth larger than others and the fourth largest; sternum narrow, much longer than wide; a few pointed hairs running over ocular area; chelicerae robust with promargin of cheliceral groove with 12 denticles of variable size; legs with sturdy hairs and spines; 7 spinnerets. Female genitalia. Two pairs of receptacular clusters situated on the dorsal side of the bursa copulatrix; the middle pair of receptacular clusters with long genital stalks and larger than the lateral ones, the middle stalks separated from each other; the posterior margin of the bursa copulatrix sclerotized; the posterior margin of the genital area wide (Fig. 3J-M).
Variation. Males and females vary in body size, cheliceral teeth, and spinnerets. Range of measurements in males (N = 2): BL 11.94-13.28, CL 5.59-6.42, CW 5.07-5.70, OL 6.08-6.19, OW 4.80-4.85; the number of cheliceral teeth varies from 9-14 (N = 2); there are 7 or 8 spinnerets. Females (N = 3): BL 10. ; the number of cheliceral teeth varies from 12-13 (N = 3); there are 7 or 8 spinnerets. In addition, male palp and female genitalia also show intraspecific variations: in males, the left palp is slightly different from the right palp, e.g. the tegulum of left male palp with three teeth basally in ventral view (Fig. 3B), but they are missing in right male palp (Fig. 3E, F); the number of teeth located in middle portion of conductor also varies between left and right palp (Fig. 3C, E, H, I); in females, the relative position of the middle receptacular clusters situated on the dorsal side of the bursa copulatrix is slightly different (Fig. 3L, M).
Etymology. The species epithet, a noun in apposition, refers to the type locality.   (Fig. 4A, B, E, H-J), but can be distinguished from those of S. dapo and S. lingshang by tegulum with smaller dorsal extension of terminal apophysis (Fig. 4C, F), and conductor with slightly narrower base of apical spine (Fig. 4A, H-J); from S. multidentata by conductor with longer apical spine (Fig. 4A, B, E, H-J), and contrategulum with larger apophysis proximally (Fig. 4A, B, D); from S. pluma by tegulum with smaller terminal apophysis (Fig. 4F), and contrategulum with one irregular dentate margin (Fig. 4A, D); from S. xiujian by contrategulum with larger apophysis proximally (Fig. 4A, B, D); from S. anhua sp. nov. by apical spine of conductor needle-shaped (Fig. 4A, B, H-J), by tegulum with slightly smaller terminal apophysis and wider dorsal extension of terminal apophysis (Fig. 4C, F); from S. zhongpo sp. nov. by apical spine of conductor with slightly narrower base (Fig. 4A, H-J); from those of other species of multidentata-group by needle-shaped apical spine of conductor (Fig. 4A, E, H-J); from those of other Songthela species by middle part of the conductor with several small spines (Fig. 4A, B, E, H-J).

Distribution. Hunan (Anhua), China
Females of S. longhui sp. nov. can be distinguished from S. anhua sp. nov. by Yshaped median genital stalks, lateral receptacular clusters with distinct short genital stalks, and deeper depressions in dorsal view ( Fig. 5A-C); from S. pluma by lateral receptacular clusters with slightly longer genital stalks, and two larger and deeper depressions in dorsal view ( Fig. 5A-C); from S. zhongpo sp. nov. by median receptacular clusters with longer genital stalks ( Fig. 5A-C); from those of other species of multidentata-group by median receptacular clusters with longer genital stalks, and Y-shaped median genital stalks, lateral ones with distinct genital stalks ( Fig. 5A-C); from those of other Songthela species by four receptacular clusters located at dorsal side of bursa copulatrix and median genital stalks fused together basally ( Fig. 5A-C).
Palp. Paracymbium unpigmented and unsclerotised prolaterally, numerous setae and spines on the tip (Fig. 4A-C). Contrategulum with an arched apophysis proximally and irregular dentate edge (Fig. 4A, B, D). Tegulum with a semi-circular marginal apophysis and dentate dorsal extension of the terminal apophysis, and with a small terminal apophysis retrolaterally (Fig. 4C, F, G). Conductor having a long apical spine pointed to the one-third of opening of embolus proximally, the middle part covered with several small teeth, and the smooth base fused with embolus (Fig. 4A, B, E, H-J). Embolus largely sclerotized, with a wide and flat opening, several longitudinal ribs in middle and distal portion (Fig. 4A, B, D, E, G).
Female (XUX-2021-281; Fig. 1F). Carapace dark reddish brown and opisthosoma light brown, with 12 dark brown tergites attached a pair of thick bristles, the second to fifth larger than others and the fourth largest; sternum narrow, much longer than wide; a few pointed hairs running over ocular area; chelicerae robust with promargin of cheliceral groove with 12 denticles of variable size; legs with sturdy hairs and spines; 7 spinnerets. Measurements : BL 11.52,CL 5.20,CW 4.33,OL 5.73,OW 4.51 Female genitalia. Two pairs of receptacular clusters with distinctly genital stalks, situated on the dorsal wall of the bursa copulatrix; the median ones similar to or slightly larger than the lateral ones, the Y-shaped middle genital stalks; the posterior margin of the bursa copulatrix sclerotized, the posterior margin of the genital area wide, two deeper depressions in dorsal view (Fig. 5A-F).
Variation. Males and females vary in body size, cheliceral teeth and spinnerets. Range of measurements in males (N = 2): BL 10. . There are 6 or 7 spinnerets (N = 2). Females (N = 5): BL 5. . The number of cheliceral teeth varies from 12 to 13 (N = 5). In addition, male palp and female genitalia also show intraspecific variations: in males, the middle part of conductor with more teeth (Fig. 4H, I) or less teeth (Fig. 4J); tegulum with a relatively larger terminal apophysis (Fig. 4C) or slightly smaller (Fig. 4E). In females, the middle Y-shaped genital stalk fused totally with only two receptacular clusters separated from each other (Fig. 5A, D) or fused basally and separated in the middle (Fig. 5B, C, E, F).
Etymology. The species epithet, a noun in apposition, refers to the type locality.  (Fig. 6A, D, E, H-K), but can be distinguished from S. dapo by tegulum with smaller terminal apophysis (Fig. 6C, F, G); from S. lingshang by contrategulum with slightly larger teeth (Fig. 6D, F, G, L), conductor with fewer teeth in middle part (Fig. 6A, D, E, H-K), and tegulum with a small terminal apophysis (Fig. 6C, F, G); from S. xiujian by tegulum with arched marginal apophysis (Fig. 6G), and contrategulum with slightly smaller apophysis proximally (Fig. 6A, L); from S. anhua sp. nov. by apical spine of conductor narrower and longer (Fig. 6A, B, D, H-K); from S. longhui sp. nov. by apical spine of conductor with slightly narrower base (Fig. 6A, H-K), and surface of dorsal extension of the tegular terminal apophysis with several ridges (Fig. 6G); from those of other species of multidentata-group by contrategulum with two dentate margins in middle part (Fig. 6D, L), by apical spine of conductor with slightly wider base and gradually becomes elongated distally (Fig. 6A, D, E, H-K); from those of other Songthela species by middle part of the conductor with several teeth (Fig. 6A, B, D, E, H-K).

Distribution. Hunan (Longhui), China
Female of S. zhongpo sp. nov. can be distinguished from S. anhua sp. nov. by the lateral receptacular clusters with slightly longer genital stalks, and arched anterior margin of bursa copulatrix (Fig. 7A-D); from S. longhui sp. nov. by median receptacular clusters with shorter genital stalks ( Fig. 7A-D); from S. multidentata by median receptacular clusters with thicker genital stalks ( Fig. 7A-D), from S. tianzhu by lateral genital stalks slightly longer (Fig. 7A, D); from those of other species of multidentata-group by median receptacular clusters with slightly thicker genital stalks, and lateral ones with distinct genital stalks (Fig. 7A-H); from those of other Songthela species by two pairs of receptacular clusters situated on dorsal wall of bursa copulatrix (Fig. 7A-H).
Description. Male (holotype). Carapace black brown; opisthosoma dark brown, with 12 brown tergites attached a pair of thick bristles, the second to fifth larger than others and the fourth largest; sternum narrow, much longer than wide; a few pointed hairs running over ocular area; chelicerae robust with promargin of cheliceral groove with 11 denticles of variable size; legs with sturdy hairs and spines; 7 spinnerets. Measurements: BL 13.53,CL 5.96,CW 5.44,OL 7.19,OW 5.94  Palp. Paracymbium unpigmented and unsclerotised prolaterally, with several setae and spines on the tip (Fig. 6A-C). Contrategulum with a triangular apophysis proximally and with a dentate edge in distal and proximal portions, while with two dentate margins in the middle part (Fig. 6A, D, F, G, H-K). Tegulum with an arched helicoid marginal apophysis, a helicoid dorsal extension of the terminal apophysis, and a thumb-shaped terminal apophysis retrolaterally (Fig. 6C, E-G). Conductor fused with embolus basally and having several teeth in the middle part, the long apical spine with a spinule basally and pointed to the one-fourth of opening of embolus proximally (Fig. 6A, B, D-K). Embolus largely sclerotized, with a wide and flat opening, numerous longitudinal ribs in middle and distal portion (Fig. 6B, D-F).
Female genitalia. Four receptacular clusters situated on the dorsal side of the bursa copulatrix; the middle ones with thick genital stalks close to each other, fused together basally and separated from each other distally (Fig. 7A-C), or the middle genital stalks fused as one (Fig. 7D); the lateral receptacular clusters similar to or smaller than the middle ones; the posterior margin of the bursa copulatrix sclerotized, the posterior margin of the genital area wide and straight (Fig. 7A, D), or incurved in the middle (Fig. 7B, C).
Etymology. The species epithet, a noun in apposition, refers to the type locality.

Distribution. Hunan (Huaihua), China
Remarks. The three new species from Hunan Province, China can be assigned into the multidentata-group based on the following characters of both male palp and female genital morphology: 1) conductor of male palp with one apical spine and the middle part covered several teeth; 2) female genitalia with two pairs of receptacular clusters situated on the dorsal side of the bursa copulatrix; and 3) the posterior margin of the bursa copulatrix of female genitalia pigmented and sclerotised.