Taxonomic revision of Telemidae (Arachnida, Araneae) from East and Southeast Asia

Abstract Species of the spider family Telemidae Fage, 1913 from East and Southeast Asia are revised. Four new genera are erected: Mekonglema Zhao & Li, gen. nov. with the type species Mekonglema bailang Zhao & Li, sp. nov. (♂♀, Yunnan, China), Siamlema Zhao & Li, gen. nov. with the type species Siamlema changhai Zhao & Li, sp. nov. (♂♀, southern Thailand), Sundalema Zhao & Li, gen. nov. with the type species Sundalema bonjol Zhao & Li, sp. nov. (♂♀, Sumatra), and Zhuanlema Zhao & Li, gen. nov. with the type species Zhuanlema peteri Zhao & Li, sp. nov. (♂♀, northern Laos). Eight additional new species are described: Mekonglema kaorao Zhao & Li, sp. nov. (♂♀, northern Laos), M. walayaku Zhao & Li, sp. nov. (♂♀, Yunnan, China), M. yan Zhao & Li, sp. nov. (♂♀, Yunnan, China), Pinelema daguaiwan Zhao & Li, sp. nov. (♂♀, Guangxi, China), P. shiba Zhao & Li, sp. nov. (♂♀, Guangxi, China), P. tham Zhao & Li, sp. nov. (♂♀, northern Laos), Siamlema suea Zhao & Li, sp. nov. (♂♀, southern Thailand), and Sundalema khaorakkiat Zhao & Li, sp. nov. (♂♀, southern Thailand). Thirty species are transferred from the genus Telema Simon, 1882 to the genera Pinelema Wang & Li, 2012, Sundalemagen. nov., and Telemofila Wunderlich, 1995. Seychellia xinpingi Lin & Li, 2008 is transferred to Mekonglemagen. nov. as M. xinpingicomb. nov. Furthermore, the genus Pinelema is divided into seven species groups based on male morphological characters.


Introduction
The spider family Telemidae Fage, 1913 currently includes 85 species in ten genera (Li 2020). Telemids are tiny spiders, whose body length ranges from 0.9 to 2.2 mm, with much longer legs relative to their body. The dispersal ability of telemids is poor, resulting in high endemism (Lin and Li 2010;Wang and Li 2010b;Zhao et al. 2018a, b).
Telemids occur in tropical rainforests and karst caves in the southern Holarctic, Ethiopian, Oriental, Neotropical, and Australasian Realms (except Australia and New Zealand) (WSC 2020). Telema tenella Simon, 1882 occurs in Spain and France and is the type species of the genus Telema Simon, 1882 and the only known telemid species from Europe. Thirty-four Asian species were classified in the poorly defined genus Telema prior to the current study, and a revision of Asian Telema is necessary. Furthermore, the placement of Seychellia xinpingi Lin & Li, 2008 from southwestern China requires examination because the shape of the bulbal apophysis of the male differs from the generotype Seychellia wiljoi Saaristo, 1978, from Seychelles.
The goal of this paper is to revise all Asian telemid species using combined morphological and molecular approaches, adding new materials collected from Southeast Asia and southwestern China.

Materials and methods
Specimens used in this paper were collected by sifting leaf litter in rainforests or collected by hand from caves. All samples were examined and measured using a LEICA M205C stereomicroscope. The habitus, left male palp, and endogyne were photographed using an Olympus C7070 wide zoom digital camera. Images were montaged using Helicon Focus Lite 7.5.6 software. Female genitalia were removed and treated in lactic acid before being photographed. All measurements are given in millimetres. Leg measurements are shown as: total length (femur, patella, tibia, metatarsus, tarsus). For SEM images, the tibial glands on leg III were photographed using a Hitachi SU8010 Environmental Scanning Electron Microscope.
For molecular phylogenetic analyses, we used all available materials of Asian telemids as well as the type species Telema tenella and Seychellia wiljoi. Our analyses contain 57 of 62 known Asian species from the type localities and 12 potentially new species. Five species for which we did not obtain molecular data are Sundalema acicularis (Wang & Li, 2010) comb. nov., Pinelema claviformis (Tong & Li, 2008) comb. nov., Telemofila malaysiaensis (Wang & Li, 2010) comb. nov., Telema nipponica (Yaginuma, 1972), and Pinelema spina (Tong & Li, 2008) comb. nov. We used two segestriid species as outgroups, as Segestriidae is considered the sister lineage of Telemidae (Shao and Li 2018). In total, 73 taxa were included in our molecular dataset.
Genomic DNA was extracted from the legs or prosoma using TIANamp Genomic DNA Kit DP304 (TIANGEN Co., Beijing, China). Two nuclear loci, Histone 3 and Wingless (H3 and Wnt), were amplified for subsequent molecular phylogenetic analyses. Primer information and PCR protocols are shown in Suppl. material 1: Table S1.
All amplicons were sequenced using an ABI 3730 automated sequencer, and raw sequences were corrected manually in BioEdit (Hall 1999). Sequence alignments were produced using Clustal W in MEGA 5 (Tamura et al. 2011), and the sequences were checked for stop codons after translation to amino acid sequences. 1 Tibial glands plate-shaped ( Diagnosis. Pinelema can be distinguished from Telema by the following: beltshaped tibial glands (arrows on Fig. 1B) (vs. plate-shaped), the presence of a cymbial apophysis (vs. absent), and the triangular, trapezoidal, or tube-like embolus (vs. duckbill shaped). The endogyne is extended distally (vs. no extension).
Comments. All 26 new combinations are supported by morphological characters, such as the presence of a cymbial apophysis and the extended tip of the receptacle. Including the new species described in this paper increases the total number of Pinelema species to 54, making it the most speciose genus in Telemidae.
Composition. According to the morphological characters of the male palp, 54 Pinelema species have been divided into seven species groups as well as six species not attached to a species group. The composition of species groups is discussed below.
Remarks. For the diagnoses and descriptions of this group (except P. bailongensis and P. curcici), see Zhao et al. (2018b). Other material examined. 1♂ (molecular voucher, IZCAS), same data as holotype. Diagnosis. For differences between P. bailongensis and P. curcici, see Wang and Li (2016); for differences between P. bailongensis and the other eleven species in this group, see Zhao et al. (2018b).
Distribution. China (Guangxi, site 6 in Fig. 30), known only from the type locality. Diagnosis. Pinelema curcici resembles P. liangxi but can be distinguished by the following: the presence of eyes (vs. absent); the embolic tip is much narrower than the embolic base ( Description. See Wang and Li (2016). Distribution. China (Yunnan, site 9 in Fig. 30), known only from the type locality.

The cunfengensis-group Figures 2C, 30
Diagnosis. This group resembles the feilong-group by having a triangular and short embolus relative to the bulb length but can be distinguished by the following: the bulb is bent at a right angle dorso-distally ( Fig. 2C) (vs. not bent), and the junction of the bulb and cymbium is located dorso-mesially on the bulb (Fig. 2C) Zhao & Li, 2017) or absent (P. spirae (Lin & Li, 2010) comb. nov.). Bulb bent dorso-distally at a right angle (Fig. 2C)   Diagnosis. Pinelema spirae comb. nov. resembles P. cunfengensis but can be distinguished by the following: the absence of eyes (vs. presence); the different shape of the embolus (Fig. 2C), the ratio of bulbal length/width is 1.39 (vs. 1.22), and the length ratio of the embolus/bulb is 0.38 ( Description. See Lin and Li (2010). Distribution. China (Guangxi, site 20 in Fig. 30), known only from the type locality. Figures 3A, 31 Diagnosis. This group resembles the xiezi-group by the short embolus relative to the bulb but can be distinguished by the triangular shape of the embolus (Fig. 3A) (vs. trapezoidal).
Distribution. China (Guizhou) and Vietnam (Bac Kan) (sites 10-11 in Fig. 31 Etymology. The species name refers to the type locality; noun in apposition. Diagnosis. Pinelema daguaiwan sp. nov. resembles P. pacchanensis by the bulb being concave ventrally but can be distinguished by the following: the bulb is more deeply concave (arrow in Fig Palp. Tibia two times longer than patella, cymbium 2.47 times longer than tibia, 1.88 times longer than femur, cymbial apophysis finger shaped (Fig. 4C); bulb concave ventrally, and shaped as in Fig. 4C, D, length ratio of bulb/cymbium about 0.9; embolus triangular and membranous, 1/5 as long as bulb (   Neck of receptacle membranous, as long as diameter of receptacle (Fig. 5A); receptacle tip globular with several membranous tubes, much wider than neck (Fig. 5A).
Distribution. China (Guizhou, site 10 in Fig. 31), known only from the type locality.  Diagnosis. This group resembles the feilong-group by the embolus which is short relative to the bulb but can be distinguished by the following: the length ratio of the embolus/bulb ranges from 0.57 to 0.79 ( Fig. 3C) (vs. 0.33 to 0.49), and the shape of the embolus is a long isosceles triangle ( Fig. 3C) (vs. equilateral triangle).
Distribution. China (Guangxi, site 15 in Fig. 31), known only from the type locality.    Description. See Wang and Li (2010b). Distribution. China (Guangxi, site 16 in Fig. 31), known only from the type locality.

The xiezi-group Figures 8, 32
Diagnosis. This group resembles the feilong-group by the short embolus relative to the bulb but can be distinguished by the trapezoidal shape of the embolus (Fig. 8) (vs. triangular).
Description. See Tong and Li (2008b). Distribution. China (Guizhou, site 7 in Fig. 32), known only from the type locality. Diagnosis. Pinelema laensis resembles P. breviseta comb. nov. but can be distinguished by the following: the bulb does not protrude ventro-distally (Fig. 8, and   Diagnosis. Pinelema oculata comb. nov. resembles P. grandidens comb. nov. but can be distinguished by the following: the eyes are present (vs. absent); the dorsal bend between the embolus and bulb is ca. 180° (Fig. 8)   Description. See Lin and Li (2010). Distribution. China (Guangxi, site 10 in Fig. 32), known only from the type locality. Diagnosis. Pinelema spinafemora comb. nov. resembles P. cucurbitina comb. nov. but can be distinguished by the following: the bulb protrudes ventro-distally (Fig. 8) (vs. not protrude), the ventral bend between the embolus and bulb is right-angled (Fig. 8) (vs. acute-angled).

Species group uncertain
Description. See Wang and Li (2010b). Distribution. China (Guangxi, site 14 in Fig. 32), known only from the type locality. Diagnosis. Pinelema nuocnutensis resembles P. mikrosphaira comb. nov. but can be distinguished by the following: the eyes are vestigial (vs. present), the width ratio of the bulb/palpal tibia is 2.0 ( Fig. 9) (vs. 4.0); the beak-shaped embolus (  Other material examined. 1♂ (molecular voucher, IZCAS), same data as holotype. Diagnosis. Pinelema spirulata resembles P. dengi comb. nov. but can be distinguished by the following: the embolus is twisted (Fig. 9) (vs. cylindrical); the distal part of the receptacle is five times wider than the receptacle neck (cf. Zhao  Etymology. The species name refers to the type locality; noun in apposition. Diagnosis. Pinelema tham sp. nov. resembles P. zhenzhuang but can be easily distinguished by the following: the embolus is hawk-beak-shaped (Figs 9, 10B) (vs. needle-shaped), the tip of the embolus is directed ventro-prolaterally (Figs 9, 10B) (vs. ventrally). The terminal part of the receptacle is 3.50 times wider than the neck (Fig.  11A) (vs. six times).

Comments. The placement of this species in
Apneumonella is doubtful because the males of both A. oculata Fage, 1921 (the type species of Apneumonella) and A. jacobsoni are unknown, and females of the above two species provide little information regarding their generic belonging. To test the relationship of A. jacobsoni to A. oculata, molecular data of A. oculata is necessary. Etymology. The generic name is a combination of "Mekong" referring to the Mekong-Lancang River which encompasses the distributional range of the genus, and "-lema", a convention used because it is part of the genus Telema, which was the first genus described in Telemidae. The gender is feminine.
Distribution. China (Yunnan) and Laos (Luang Prabang) (sites 2-6 in Fig. 33). Diagnosis. This species resembles M. yan sp. nov. but can be distinguished by the following characters: abdominal scutae present in the male (vs. absent); the embolus is sclerotized (Fig. 12B-D) (vs. unsclerotized), the bulb length/width ratio is ca. 1.2 (Fig.  12C, D) (vs. ca. 1.7); the receptacle is short and almost straight (Fig. 13A) (vs. long and U-shaped). This species also resembles M. xinpingi comb. nov. but can be differentiated by the following: the absence of eyes (vs. presence), the nearly ellipsoid shape of the bulb (Fig. 12C, D) (vs. droplet shaped), the fin-like embolus (Fig. 12C) (vs. cone shaped), and the receptacle is short and not swollen distally (vs. receptacle long and swollen distally).

Etymology. The species name refers to the type locality; noun in apposition.
Diagnosis. This species resembles M. walayaku sp. nov. but can be distinguished by the following characters: the bulb is nearly globular (Fig. 14C, D) (vs. ellipsoidal), the tip of the embolus is not sclerotized (Fig. 14B) (vs. well sclerotized). The tip of receptacle is 1.50 times wider than the neck (Fig. 15A) (vs. three times).
Distribution. Laos (Luang Prabang, site 3 in Fig. 33), known only from the type locality. Etymology. The species name refers to the type locality; noun in apposition. Diagnosis. This species resembles M. kaorao sp. nov. but can be distinguished by the following characters: the bulb is ellipsoidal (Fig. 16C, D) (vs. nearly globular), the tip of the embolus is well sclerotized (arrow on Fig. 16B) (vs. unsclerotized); the neck of the receptacle is ca. three times thinner (vs. 1.50 times thinner) than the distal part of the receptacle (Fig. 17A).
Comments. This species is transferred to Mekonglema gen. nov. because it shares a similar shape of the copulatory organs with M. bailang sp. nov., the type species of the genus. This placement is also supported by molecular phylogenetic analyses (Fig. 34). Etymology. The species refers to the type locality; noun in apposition.  Diagnosis. Mekonglema yan sp. nov. resembles M. bailang sp. nov. but can be distinguished by the following characters: an abdominal scuta is absent in the male (vs. present); the tip of the embolus is unsclerotized (Fig. 18B-D) (vs. sclerotized), the bulbal length/width ratio is approximately 1.7 (Fig. 18C, D) (vs. 1.2); the receptacle is long and U-shaped (Fig. 19A) (vs. short and nearly straight).
Etymology. The generic name is derived from "Siam", referring to the old name of Thailand, and "-lema" is a convention from Telema, the type genus of the family. Feminine in gender.
Description. Total length: 0.92-1.11 (male), 1.04-1.20 (female). Carapace 0.41-0.48 long. Sternum with sparse setae. Eyes normally developed, vestigial, or absent. Leg formula: 1-2-4-3, tibia I 0.97-1.05 long, leg glands belt shaped (arrows on Fig.  1D). Male palp: cymbium, tibia, patella, and femur robust relative to bulb, femur longer than tibia and cymbium, dorsal spine present on distal part of palpal tibia, distinct cymbial apophysis located prolaterally on mesial part; bulb droplet-shaped    Diagnosis. Females belonging to Sundalema gen. nov. can be distinguished from those of other genera by the following: a sclerotized and spiral receptacle (Fig. 24A) (vs. unsclerotized or not spiral). Males belonging to Sundalema gen. nov. resemble species in the bailongensis-group by having a long embolus but can be distinguished by the embolus lacking a spiral ridge and being nearly L-shaped (Fig. 25B-D) (vs. a distinct spiral ridge present on the nearly straight embolus).
Comments. The females of this genus are easily distinguished from all other genera, as their receptacles are long, spiral, and sclerotized; thus, a female has been chosen to represent the holotype.  fig. 4A, B).
Distribution. Thailand (Prachuap Khiri Khan, site 9 in Fig. 33), known only from the type locality. Description. See Wang and Li (2010a). Distribution. Thailand (Krabi, site 10 in Fig. 33), known only from the type locality.
Composition. Telema auricoma, T. guihua, T. nipponica, T. tenella and T. wunderlichi. The composition is supported not only by morphological characters but also by molecular phylogenetic analyses ( Fig. 34; T. nipponica is not included).
Distribution. Eurasian disjunctive range, known from Southern Europe (France and Spain, one species) and East Asia (Japan and southwestern China, four species, sites 13-16 in Fig. 33).

Telema auricoma Lin & Li, 2010
Telema auricoma Lin and Li 2010: 3, fig. 1  Comments. All known specimens of this species are female, and molecular barcoding data from all eight populations examined shows no differences (unpublished data). This indicates that the species may be parthenogenetic, a character that may allow it to easily disperse more broadly than gametogenetic species. Other material examined. 1♀ (molecular voucher, IZCAS), same data as holotype.
Diagnosis. This species resembles T. wunderlichi but can be distinguished by the following characters: the small body size (vs. larger size); the embolus is membranous (vs. sclerotized), and the tip of the embolus is blunt (cf. Lin and Li 2010: fig. 6E) (vs. sharply pointed).
Description. See Lin and Li (2010). Distribution. China (Guizhou, site 14 in Fig. 33), known only from the type locality. Diagnosis. This species resembles T. tenella but can be distinguished by the thumb-like shape of the embolus (cf. Yaginuma 1974: fig. 4), whereas in T. tenella, the embolus is short and triangular (cf.   fig. 2C, D).
Comments. The placement of T. pecki in this genus is dubious because its leg formula is 1-2-4-3 (Brignoli 1980) and the embolus is triangular (cf. Brignoli 1980: figs 1, 2). These characters are inconsistent with the genus characters of Telemofila. However, we have been unable to examine the types of T. pecki, and molecular data from this species is lacking.  Diagnosis. This species resembles T. malaysiaensis comb. nov. but can be distinguished by the following: bigger body size, bean-shaped receptacle (cf. Wang and Li 2010a: fig. 8C, D) (vs. globular); claw-like embolus (cf. Wang and Li 2010a: fig. 8A, B) (vs. sickle-shaped). This species can be distinguished from T. samosirensis by the embolus being three times shorter than the bulb (cf. Wang and Li 2010a: fig. 9A, B) (vs. two times shorter).
Description. See Wang and Li (2010a). Distribution. Singapore (Site 17 in Fig. 33). Diagnosis. This species can be distinguished from T. fabata comb. nov. and T. malaysiaensis comb. nov. by the length of the embolus which is equal to the radius of the bulb (cf. Wunderlich 1995: fig. 17) (vs. length of embolus 1/3 as long as the diameter of the bulb in two similar species).
Genus Zhuanlema Zhao & Li, gen. nov. http://zoobank.org/D5FC4E70-33EA-4741-88FF-85005FF3C759 Type species. Zhuanlema peteri sp. nov. from Luang Prabang Province, Laos. Etymology. The generic name is derived from "Zhuan", referring to the Chinese pinyin "zhuan", indicating that the apex of the embolus is twisted, and "-lema" which is a convention from the type genus of the family. Feminine in gender.
Diagnosis. The new genus resembles species in the bailongensis-group but can be distinguished by the following characters: the apex of the embolus is twisted (Fig.  28C) (vs. tube-like embolus), the cymbial apophysis is located basally and four times shorter than the width of the cymbial base (Fig. 28C, D) (vs. cymbial apophysis located mesially or sub-basally, and longer or 1-3 times shorter than the width of the cymbial base). Females can be distinguished by the sclerotized receptacle (Fig. 29A) (vs. membranous).

Sequence data and model selection
For 71 telemid taxa, a total of 71 and 67 sequences were successfully generated for H3 (331 base pairs, bp) and Wnt (330 bp), respectively. All sequences were submitted to GenBank (Suppl. material 1: Table S2). The best-fit Akaike information criterion (AIC) model of the concatenated BI dataset was SYM+I+G.

Molecular phylogenetic results
The topology of both the ML and BI trees are consistent at the genus level, branches to all tips are long, indicating distinct genetic divergence of each lineage (Fig. 34). Telemidae consists of two major clades: one of Telema and one of the other eight genera. Telema includes four species (molecular data of T. nipponica was not acquired), and the remaining Telema species are clustered in Pinelema, Sundalema gen. nov., and Telemofila (new combinations in Fig. 34), supporting the hypothesis of previously incorrect generic placement of those species.
Pinelema is monophyletic with high support values (BS = 98 and PP = 100), although most interior nodes are not well supported (Fig. 34). Telemofila is robustly monophyletic (BS = 100 and PP = 100), and molecular data support the transfer of Telemofila fabata comb. nov. to this genus (Fig. 34). Mekonglema xinpingi comb. nov. did not cluster with the type species of Seychellia, so the hypothesis of the incorrect generic placement of M. xinpingi comb. nov. is supported (Fig. 34). The generic placement of Apneumonella jacobsoni is ambiguous because material of the type species A. oculata is lacking. The four new genera (Mekonglema gen. nov., Siamlema gen. nov., Sundalema gen. nov., and Zhuanlema gen. nov.) are monophyletic and (excluding Siamlema gen. nov.) strongly supported by both ML and BI (Fig. 34). This result is consistent with morphological delimitation (Fig. 34).

Discussion
In this paper we constructed a molecular phylogeny of Telemidae from East and Southeast Asia for the first time, and our results have changed the taxonomic framework of Asian telemids. We revised the taxonomic status of these Asian telemids, introducing four new genera and 12 new species, and 31 species were transferred to other genera.
When researchers are delimiting genera of Telemidae, qualitative morphological characters should be considered. These characters include: for both sexes, the leg formula and the shape of tibial glands; for males, the presence or absence of a cymbial apophysis, the length ratio of the palpal femur/cymbium, and the accessory structures on the palpal tibia or bulb; for females, the shape of the receptacle and the presence/ absence of membranous/sclerotized tubes within the receptacle. Quantitative morphological characters of reproductive organs should be considered when researchers are delimiting different species in the same genus. These characters include the ratio of the embolus/bulb length, the angle between the embolus and bulb, and the width ratio of the receptacle tip/neck, etc.
Despite the large genetic difference between Telema species in East Asia and T. tenella (e.g. the long branch between T. tenella and Asian Telema spp. in Fig. 34) this genus occupies a small morphospace. This is also supported by paleontological evidence, as the morphological characters of T. moritzi Wunderlich, 2004 (extinct species in Baltic amber, ca. 45 million years ago) are difficult to distinguish from extant Telema spp. (Wunderlich 2004). The occurrence of Telema spp. in the entire Palearctic is intriguing given their extremely weak dispersal ability and requires further investigation. It is possible that they dispersed from Europe to Asia (or vice versa) in ancient warm stage, then many species became extinct when the climate cooled. Only a few species survived in refuges like southern Europe and the tropical and subtropical areas of Asia, and this has resulted in the current fragmented distribution.