Four new species of troglomorphic Coecobrya Yosii, 1956 (Collembola, Entomobryidae) from Thailand based on morphological and molecular evidence, with an updated key of Thai troglomorphic species Author Nilsai 1, Areeruk Division of Biological Science, Faculty of Science, Prince of Songkla University, Hat Yai, Songkhla, 90110, Thailand Author Detcharoen 1, Matsapume Division of Biological Science, Faculty of Science, Prince of Songkla University, Hat Yai, Songkhla, 90110, Thailand Author Godeiro 2, Nerivania Nunes Division of Biological Science, Faculty of Science, Prince of Songkla University, Hat Yai, Songkhla, 90110, Thailand Author Jantarit 3, Sopark Division of Biological Science, Faculty of Science, Prince of Songkla University, Hat Yai, Songkhla, 90110, Thailand text Subterranean Biology 2021 2021-12-10 41 1 42 http://dx.doi.org/10.3897/subtbiol.41.76926 journal article http://dx.doi.org/10.3897/subtbiol.41.76926 1314-2615-41-1 73326F5CC4BE431DBE6D601A69BD7FFA 482986A3D1D15408B0FE4F5EF7587312 Coecobrya whitteni Nilsai & Jantarit sp. nov. Figures 1A , 2 , 3 , 4 Type material. Holotype : female on slide. Thailand, Khon Kaen province, Chum Phae district, Tham Nayn Noi (note: "tham" = "cave" in Thai), altitude 359 m a.m.s.l., 16.8292°N , 101.9848°E . 3.XII.2020; S. Jantarit, A. Nilsai, K. Sarakhamhaeng and K. Jantapaso leg. (sample # THA_SJ_KKN04), dark zone of a cave, by entomological aspirator. Paratypes : same data as holotype, 11 specimens (three female and eight subadults on slides). Additional material : same data as holotype, 3 specimens (in ethanol) Holotype and 11 paratypes on slides deposited in NHM-PSU. Description. Habitus (Fig. 1A ). Medium size Entomobryidae . Body length 1.8-2.3 mm (holotype 1.8 mm). No scales. Eyes absent. Color: whitish in alcohol, without pigmentation. Four antennal segments. Body slender not bent nor humped at the level of Th. II. Abd. IV 3.80-5.63 times as long as Abd. III along the dorsal midline. Figure 1. The distribution map of the troglomorphic Coecobrya in Thailand and the habitus of the four new species. ( 1 ) A Coecobrya whitteni sp. nov. ( 2 ) B C. troglobia sp. nov. ( 3 ) C C. ellisi sp. nov. ( 4 ) D C. phitsanulokensis sp. nov. ( 5 ) C. chumphonensis Zhang & Nilsai, 2017 ( 6 ) C. sirindhornae Jantarit, Satasook & Deharveng, 2019 ( 7 ) C. polychaeta Zhang & Nilsai, 2017 and ( 8 ) C. cavicta Nilsai & Zhang, 2017. Scale bar: 1 mm. Pseudopores (Figs 3F , 4A-C ). Pseudopores present as round flat disks, smaller than mac sockets, except for the coxae and manubrium where psp are as large as mac sockets, present on various parts of the body: antennae, head, tergites, coxae and manubrium. On antennae, psp located ventro-apically between the tip of antennal segments and the chaetae of the apical row, or just below the apical row of chaetae (2 psp on Ant. I, 2-3 psp on Ant. II, and 3 psp on Ant. III). On the head, 1-2 psp located externally on each peri-antennal area. On tergites, 1+1 psp close to the axis from Th. II to Abd. IV (Figs 4A-C ). On coxae, 1-2 psp on coxae I, 2-3 psp on coxae II and 1-2 psp on coxae III, located close to longitudinal rows of chaetae. On manubrium, 2+2 dorso-apical ones (Fig. 3F ). Clypeus and mouthparts (Figs 2A-C , 2H, I , 3G ). Clypeal area with three long, smooth prefrontal; six middle chaetae (2+2 small ciliated chaetae of unequal size posteriorly and 1+1 rather long, thin, smooth chaetae anteriorly); and 1+1 long smooth lateral chaetae (Fig. 3G ). Prelabral and labral chaetae 4/5, 5, 4, all thin and smooth; the three median chaetae of the second row longer than two lateral ones (35-40 vs. 25 µm ) (Fig. 2H ). Distal border of the apical non-granulated area of the labrum with a relatively narrow median U- or V-form intrusion into the granulated area dorsally; apical edge without spines (Fig. 2H ). Ventro-distal complex of labrum well differentiated, asymmetrical, with 1+1 distal combs of 14-19 minute teeth on the right side and 14-16 strong and larger teeth on the left side (Fig. 2A ), and an axial pair of long sinuous tubules. Maxillary outer lobe with one basal chaeta, one apical chaeta (basal chaeta thicker than apical one) and three smooth sublobal hairs (55-60 vs. 35-38 µm ) (Fig. 2B ). Labial palp strongly modified for the genus, with 0, 5, 0, 4, 4 guards for papillae A-E, as described by Fjellberg (1999) for Entomobryidae or by Xu and Zhang (2015) and Jantarit et al. (2019) for Coecobrya . Lateral process of labial palp subcylindrical, as thick as normal chaetae, with tip beyond the apex of the labial papilla (Fig. 2C ). Mandible apex blunt and strong, asymmetrical (left with four teeth, right with five teeth) (Fig. 2I ); molar plate with three strong pointed basal teeth, and 3-(5) smaller inner distal teeth, identical in both mandibles. Maxilla capitulum with a three-toothed claw and several stout ciliated lamellae; lamella 2 large and broad, lamella 3 well developed; several other lamellae present as figured in Jantarit et al. (2019) . Figure 2. Coecobrya whitteni sp. nov. A Ventro-distal complex of labrum B Outer maxillary lobe C Labial palp D Ant. III organ E Chaetae of labial basis and ventral chaetotaxy of head F Mucro G Distal part of tita III and claw complex H Prelabral and labral chaetae I Mandibles. Figure 3. Coecobrya whitteni sp. nov., continued A Distal part of manubrium ventrally B Trochanteral organ C Posterior side of ventral tube and Lateral flap D Dorsal cephalic chaetotaxy E Anterior side of ventral tube F Manubrium plaque G Clypeal chaetae. Figure 4. Coecobrya whitteni sp. nov., continued A Chaetotaxy of dorsal Th. II- III B Chaetotaxy of dorsal Abd. I- III C Chaetotaxy of dorsal Abd. IV D Chaetotaxy of dorsal Abd. V. Antennae (Fig. 2D ). Antennae very long, approximately 3.78-5.14 times as long as cephalic diagonal. Antennal segments ratio as I: II: III: IV. 1: 2.08-2.20: 2.17-2.21: 3.75-4.40 (N = 3). Antennal segments not subdivided nor annulated. Antennal chaetal types not analysed in detail. Ant. I ventrally with many smooth spiny mic of various sizes in its basal part, many subcylindrical, hyaline sens in its middle to apical part, and many long smooth straight chaetae. The paddle-like chaetae (sensu Nilsai et al. 2017 ) on Ant II dorsally absent. Ant. III organ with typical five sens, sens one and four subequal, hyaline; sens five acuminate, and shorter; sens two and three swollen in paddle-like chaetae shape (Fig. 2D ). Ant. IV without apical bulb. Subapical organite not distinctly knobbed, swollen, slightly enlarged apically, inserted dorsally. Dorsal head chaetotaxy (Fig. 3D ). Dorsal cephalic chaetotaxy with four antennal chaetae (An3a2 and An3 as mac, An1-2 as mes), four anterior mac (A0, A2-3 and A5), three median (M1, M2 and M4) and eight sutural (S) mac (S0, S1-7); Gr. II with four mac; A0 as mac; 5-7+5-7 scale-like structures (usually 6+6) (sensu Jantarit et al. 2019 ) present below sutural mac, probably inside the integument; a pair of short cephalic trichobothria, external and close to the middle of the head (Fig. 3D ). Ventral head chaetotaxy (Fig. 2E ). Chaetae of labial basis all smooth (mrel1l2), chaetae m e and l1 subequal, r thin and shortest, and l2 longest. The r/m length ratio: 0.13-0.14 (Fig. 2E ). Postlabial chaetae X2, X and X4 smooth, acuminate and minute chaetae, of similar size, X3 normally absent but present as mic in one individual. On each side of the cephalic groove with 9 chaetae, of which, the proximal six always long and smooth, 7th and 9th as mics, 8th always long and ciliated, one specimen with three mics distally (Fig. 2E ). Tergite chaetotaxy (Figs 4A-D ). Th. II with two (m1, m2) medio-medial, one (m4) medio-sublateral and 18-20 posterior mac; 1+1 ms and 2+2 sens antero-laterally (Fig. 4A ). Th. III with 23-26 mac. 2+2 sens laterally (Fig. 4A ). Abd. I with six central mac (m4i, m2-4, m2i, m4p), 1+1 ms and 1+1 sens laterally (Fig. 4B ). Abd. II with two (m3, m3e) central and one (m5) lateral mac, 2+2 tric without modified chaetae, 1+1 sens laterally and 1+1 mic near internal tric (Fig. 4B ). Abd. III with one (m3) central and three (am6, pm6, p6) lateral mac. 3+3 tric without modified chaetae, 1+1 sens laterally, 1+1 mic near m3, ms not seen (Fig. 4B ). Abd. IV with four central mac (M, A5-6, B5) and six (D3, E2-4, E2p, F3) lateral mac, 2+2 tric and about 19 long S-like chaetae and one mic in the middle below psp, without modified chaetae (Fig. 4C ). Abd. V with at least 6 obvious mac and several mes to small mac, and 3+3 sens (Fig. 4D ). Abd. VI not analysed. S-chaetae formula from Th. II to Abd. V: 2+ms, 2/1+ms, 2, 2, ≈ 20, 3; ps not seen, as sens on Abd. IV 1/3 as long as S-like chaetae (Figs 4A-D ). Legs (Figs 2G , 3B ). Leg long; tita of leg III slightly longer than tita of legs I and II. Legs devoid of scales, covered with ordinary ciliated chaetae of various lengths, mic not seen. Trochanteral organ with 18-21 smooth, straight, unequal spine-like chaetae (Fig. 3B ). The distal whorl of tita III with 10 subequal ciliated mes, irregularly arranged, and usually claw I-II with dorso-apical clavate tenent hair (pointed in two specimens). A smooth, thin and long chaeta close to tenent hair (sensu Jantarit et al. 2019 ) absent. Claw III generally with dorso-apical clavate tenent hair (8 individuals with clavate tenant hair, 4 individuals with pointed tenant hair; holotype with pointed tenent hair Fig. 2G ). Ventro-distal smooth chaeta of tita III thick, erected, pointed, rather short. Pretarsal mic minute (2.5-3.0 µm ). Claw slender and elongated. Unguis of all claw with one small inner teeth at 53 % and a pair of unequal basal teeth at about 44-49 % of inner edge from basis. Unguiculus approximately 2/3 as long as inner edge of claw, rather swollen baso-externally, pointed apically, devoid of inner tooth, with at least five minute outer teeth, often inconspicuous, at 3/4 of its length (Fig. 2G ). Ventral tube (Figs 3C, E ). Ventral tube two to three times longer than wide. Lateral flaps with 6-7+6-7 smooth chaetae except for one specimen with one ciliated chaeta present in both sides (Fig. 3C ). Anterior face with 6-8+6-8 chaetae, 2(3) of them larger than others, all ciliated, arranged roughly asymmetrically (Fig. 3E ); posterior face with at least 12 chaetae, four apical chaetae longer and larger than others and about 8-13 proximal chaetae; two straight smooth chaetae and 6-11 either small finely ciliated or mics arranged asymmetrically (Fig. 3C ). Furcal complex (Figs 2F , 3A, F ). Tenaculum with four large teeth of decreasing size from the basal to the distal one of each ramus, on a prominent, irregular body, with a postero-basal strong serrated chaeta bent distally. Mucrodens 1.6 times longer than manubrium. Furcula without smooth chaetae. Manubrium with a dense cover of ciliated chaetae both dorsally and ventrally. Manubrial plaque with 2+2 psp and three to six ciliated chaetae (usually 5+5) (Fig. 3F ). Distal part of manubrium ventrally with 13+13 ciliate chaetae (Fig. 3A ). Dens without spines, annulated and covered with ciliated chaetae on both sides. Distal smooth part of dens slightly longer than mucro. Mucro strong and falcate, basal spine long, reaching the tip of the mucronal tooth (Fig. 2F ). Genital plate. Female genital plate with 2+2 genital mic, male genital plate not clearly seen. Ecology. Coecobrya whitteni sp. nov. was found in the dark zone of a cave on the surface of the muddy ground, near the bank of a stream. It was also found in the upper levels on stalagmites, clay, gravel and rock surfaces. This species was found mainly in eutrophic habitats where piles of guano were well-presented. There was a big colony of the intermediate roundleaf bat ( Hipposideros larvatus ) inside the cave. The air temperature of the habitat where this species was collected was 22.9-24.0 °C and the relative humidity was 85%. In the same cave, and in the same chamber, at least six different stygobitic species were also observed in the small puddles: Aequigidiella aquilifera , Dugesia deharvengi , Heterochaetella glandularis , Siamoporus deharvengi , Stenasellus rigali and Theosbaena cambodjiana . The co-occurrence of these six species was first observed in 1987 ( Association Pyreneenne de Speleologie 1988 ; Deharveng and Bedos 2000 , 2012 ) and from our observations they are all still present in rather large numbers of individuals. We also found a millipede ( Plusioglyphiulus saksit ) and a spider ( Speocera deharvengi ) in the same habitat where this new species was found. Most of these observed species only known as endemics to this cave. The discovery of this new species in Tham Nayn Noi emphasizes the importance of this cave in harboring the endemic subterranean fauna of the area. Etymology. Coecobrya whitteni sp. nov. is named in honor of the late Tony Whitten in appreciation of his enormous contributions to the discovery and conservation of karst and cave invertebrates across Asia. Remarks. Among the described troglomorphic Coecobrya species in Thailand, Coecobrya whitteni sp. nov. is near to C. cavicta Nilsai & Zhang, 2017 from Satun province, southern Thailand in the body length, dorsal head chaetotaxy, labial chaetae, number of sublobal hairs on maxillary outer lobe, number of mac on Abd.III and number of chaetae on ventrodistal part of manubrium. However, it differs from C. cavicta by the combination of various characters i.e. longer length of antennae, number of mac of dorsal tergits on Th. II (21-23 vs. 36-37), Th. III (23-26 vs. 35), Abd. II (2 vs. 3) and Abd. IV (11 vs. 18-20), number of inner teeth of claw (3 vs. 2) and number of smooth chaetae of trochanteral organe (18-21 vs. 15-16) (see Table 1 ). In fact, Coecobrya whitteni sp. nov. is most similar to C. troglobia sp. nov. which is described in this work since the caves where these two species are found are only 3.4 km apart in a straight line, separated by limestone mountains. The altitudes of the two caves differs by about 200 m. The two species are similar in body size, antennal length, number of sublobal hair on the maxillary outer lobe, number of chaetae on Gr. II, labial chaetae, number of mac on Abd. I-IV, number of inner teeth of the claw, tenent hair, similar number in ventral tube chaetae both anterior and posterior side as well as a lateral flap, number of chaetae on trochanteral organ (Table 1 ). However, Coecobrya whitteni sp. nov. differs from C. troglobia sp. nov. by the number of An dorsal mac on the head (4 vs. 5) and number of chaetae on Th. II (medio-medial mac = 2 vs. 3, medio-sublateral mac = 1 vs. 4) (Table 1 ). Coecobrya whitteni sp. nov. has in fact already been mentioned by Deharveng and Bedos (2000) as an undescribed cf. Coecobrya species (Fig. 31.8A page 625) from oligotrophic habitats. (Note that in this reference the cave is misnamed as Tham Kubio). Table 1. Comparison of troglomorphic Coecobrya in Thailand: C. chumphonensis Zhang & Nilsai, 2017, C. cavicta Nilsai & Zhang, 2017, C. polychaeta Zhang & Nilsai, 2017, C. sirindhornae Jantarit, Satasook & Deharveng, 2019, C. nupa Christiansen & Bellinger, 1992. C. whitteni Nilsai and Jantarit sp. nov., C. troglobia Jantarit and Nilsai sp. nov., C. ellisi Jantarit and Nilsai sp. nov., C. phitsanulokensis Jantarit and Nilsai sp. nov. Key: c = ciliated chaetae, s = smooth chaetae,? = not given in literature description. Characters indicated in parentheses are rarely observed.

Characters/species		C. chumphonensis	C. cavicta	C. polychaeta	C. sirindhornae	C. nupa	C. whitteni sp. nov.	C. troglobia sp. nov.	C. ellisi sp. nov.	C. phitsanulokensis sp. nov.
Body length		up to 2.82 mm	1.72 mm	up to 2.58 mm	up to 2.6 mm	2.0 mm	1.8-2.3 mm	2.0-2.4 mm	1.6-2.1 mm	1.8-2.3 mm
Ant./head ratio		3.70-4.48	2.67	5.91-7.12	8.0-12.3	6.5	3.78-5.14	4.0-6.8	3.5-6.4	3.1-4.2
Long smooth straight chaetae on Ant. I		absent	present	absent	present	?	present	present	present	present
No. of paddle-like chaetae on Ant. II		2-4	?	1	10-12	?	not seen	not seen	not seen	not seen
Number of chaetae on clypeus
-	Prefrontal area	3s	?	3s	3s	?	3s	3s	3s	3s
-	Facial area	8s	?	2s; 7-10c	2s; 7-10c	?	2s; 6c	2s; 4c	2s; 5c	2s; 4-7c
Number of chaetae on dorsal head
-	An area	2	4	4	1	?	4	5	3	5
-	M series	4	3	3	0	?	3	3	3	3
-	Group II (Gr. II)	6(7)	4	3	1	?	4	4	4(5)	4(5)
A0		mac	mac	mic	mes	?	mac	mac	mac	mac
Number of sublobal hairs on maxillary outer lobe		3	3	4	4	3	3	3	4	4
Lateral process of labial palp		short	short	long	long	short	long	long	long	long
Labial chaetae shape		mRel1l2	mrel1l2	M1m2rel1l2	m1m2rel1l2	M1m2rel1l2	mrel1l2	mrel1l2	m(M)1m2rel1l2	m1m2rel1l2
Postlabial chaetae X		minute	minute	normal	normal	?	minute	minute	normal	minute
Number of chaetae along cephalic groove		4s; 5-7c	3-4s; 7-8c	6-7s; 5-10c	7-9s; 3-7c	?	5-6s; 1-2c1-2mics	6s; 1c; 2mics	2-5s; 1-5c; 2mics	4-7s; 1-2c; 1-2(3-4)mics
Chaetotaxy of Th. II
-	Number of medio-medial mac	4-6	3	7(6)	3	2	2	3	3	3
-	Number of medio-sublateral mac	3	3	4	4	3	1	2	3	3
-	Number of posterior mac	25-32	30-31	29-40	15-18	15?	18-20	15-18	28-31	31-33
Number of mac on Th. III		32-35	35	35-43	32-35	15?	23-26	24-27	29-36	32-33
Number of mac on Abd. I		6-7	6-7	8-9	6	4	6	6	7	7
Number of central mac on Abd. II		3	3	4(3)	2	2	2	2	2	3(4)
Chaetotaxy of Abd. III
-	Number of central mac	2	1	1	1	1	1	1	2	2
-	Number of lateral mac	3	3	3	3	2?	3	3	3	3
-	ms	not seen	not seen	present	not seen	?	not seen	not seen	present	present
Chaetotaxy of Abd. IV
-	Number of central mac	7	7-9	6	6	4	5	5	8	9
-	Number of lateral mac	10-12	11	9	8	?	6	6	9	8
Ventral tube chaetae
-	anterior face	9-12c	?	12c	10c	?	6-8c	7-8c	6-7c	6-7c
-	posterior face	13	?	20-31	20-30	7	8-13	14-18	12-13	12-15
-	lateral flap	7(10)	?	9-12	7-8s (5c)	6	6-7s (1c)	7-9s (1-2c)	6-7s; 1-3c	7-8s
Tenent hair shape		usually pointed	pointed	pointed	pointed	pointed	clavate/pointed	clavate/pointed	pointed	pointed
Number of ungual inner teeth		3	2	3	3	3	3	3	3	2
Unguiculus outer edge shape		serrate	serrate	serrate	2-3 teeth	?	serrate	serrate	?	serrate
Smooth chaetae on trochanteral organ		12-22	15-16	15-25	12-18	16	18-21	19-23	17-23	19
Chaetae on manubrial plaque		4-7	4	4-10	3	5	3-6c	5c	5-6c	5-6c
Chaetae on ventrodistal part of manubrium		11-15c	13c	15-25c	8-10c	?	13c	?	10-12c	13+13c
Mucronal spine		nearly reaching mucronal apex	nearly reaching mucronal apex	nearly reaching mucronal apex	nearly reaching mucronal apex	beyond mucronal apex	nearly reaching mucronal apex	nearly reaching mucronal apex	nearly reaching mucronal apex	nearly reaching mucronal apex
Locality		Chumphon, Thailand	Satun, Thailand	Satun, Thailand	Satun, Thailand	Maui, Hawaii	Khon Kaen, Thailand	Khon Kaen, Thailand	Phetchabun, Thailand	Phitsanulok, Thailand