Zoosyst. Evol. 97 (1) 2021, 97-109 | DOI 10.3897/zse.97.52354 eee BERLIN A new representative of the genus Bryocyclops Kiefer, 1927 from a karst cave in north-eastern Thailand (Copepoda, Cyclopoida, Cyclopidae) and comments on the generic affinities Santi Watiroyram! 1 Division of Biology, Faculty of Science, Nakhon Phanom University, Nakhon Phanom, 48000, Thailand http://zoobank. org/C7&8&6D024-63 D4-474C-8F54-A9101E 246312 Corresponding author: Santi Watiroyram (santi.watiroyram@npu.ac.th) Academic editor: Kay Van Damme @# Received 23 March 2020 Accepted 29 August 2020 @ Published 3 February 2021 Abstract The seventh Thai species of Bryocyclops Kiefer, 1927 — Bryocyclops jayabhumi sp. nov. — was found in a karst cave in the Chai- yaphum Province of north-eastern Thailand. The new species differs from all previously-known species by the absence of an inner seta on the proximal endopod of the first four swimming legs. Bryocyclops jayabhumi sp. nov. is most similar to B. maholarnensis Watiroyram, Brancelj & Sanoamuang, 2015 — the monotypic species of Group VII, which was previously described from Thailand. However, the new species differs from B. maholarnensis by having the following characteristics: 1) posterior margin of urosomites serrated; 11) anal operculum triangular with acute-tip; 111) PI—P4 Enp-1 without an inner seta; tv) armature on the female P2—P3 Enp-2 and P4 Enp; v) a transformed spine on the male P3 Enp-2. In this study, the generic affinity of the genus Bryocyclops Kiefer, 1927 is discussed and redefined, based on the available literature concerning its principle morphology to fill the present knowledge gap about the characteristics of the genus. Key Words cave fauna, epikarst, groundwater, Southeast Asia, Thailand Introduction North-eastern Thailand (locally called ‘Isan’) is located on the Khorat Plateau and encompasses approximate- ly 200,000 km? or one-third of the country (Smith and Stokes 1997). Its major geographic features are the plains and mountain ranges along the region’s western edge (..e. Phetchabun Mountain), especially in the Loei, Nong Bua Lam Phu and Chaiyaphum Provinces. Two faunal groups (snails and geckos) have been intensively researched in the caves in this area (Ellis and Pauwells 2012; Pau- wels et al. 2014; Tumpeesuwan and Tumpeesuwan 2014; Tanmuangpak 2015), but few studies have focused on aquatic invertebrates, though the region boasts highly di- verse, cave-dwelling fauna — especially Copepoda (Wa- tiroyram et al. 2015, 2017). To date, four cave-dwelling copepods have been reported in north-eastern Thailand, including Bryocyclops maholarnensis Watiroyram, Brancelj & Sanoamuang, 2015; B. muscicola (Menzel, 1926); Elaphoidella bidens decorata (Daday, 1901); and E. namnaoensis Brancelj, Watiroyram & Sanoamuang, 2010 (Watiroyram et al. 2017; Watiroyram 2018). The genus has, so far, been divided into seven species groups (I-VII) and four subgenera: Bryocyclops s. str.; Haplocyclops Kiefer, 1952; Palaeocyclops Monchenko, 1972; and Rybocyclops Dussart, 1982. The latter three have been further split into the three different genera (Kief- er 1927; Lindberg 1953, 1956; Monchenko 1972; Dussart 1982; Rocha and Bjornberg 1987; Ranga Reddy and De- faye 2008; Watiroyram et al. 2015; Fiers and Van Damme 2017). Recently, a species of Group II — B. sogotraensis Mirabdullayev, Van Damme & Dumont, 2002 — was re-evaluated and given to a new genus: Thalamocyclops Fiers & Van Damme, 2017 (Rocha et al. 1998; Fiers 2002; Copyright Santi Watiroyram. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 98 Santi Watiroyram: Bryocyclops jayabhumi sp. nov. from a cave in north-eastern Thailand Fiers and Van Damme 2017). In addition, Fiers and Van Damme (2017) note that the species of Groups I, II and VII (Lindberg 1953; Watiroyram et al. 2015) share gener- ic affinities and a common lineage, but species of Group If (B. constrictus Lindberg, 1947; B. travancoricus Lind- berg, 1947) and Group IV (B. africanus Kiefer, 1932); Bryocyclops (Palaeocyclops) jankowskajae Monchenko, 1972 present different lineages that are clearly unrelated to Bryocyclops s. str. (Reid and Spooner 1998; Fiers 2012). Based on the available literature and the discovery of the new species described herein, this study discusses and up- dates the generic affinity of Bryocyclops Kiefer, 1927. Materials and methods Site description Chatyaphum Province sits in the westernmost edge of this plateau, mostly covered by Triassic-Tertiary sedimentary rocks and Permian limestone (Singtuen and Won-In 2018). The Prakai Phet cave of Chatyaphum Province, located in the Phetchabun Mountain Range, is made of dolomitic limestone karst formed in the Permian Period and it served as a refuge for several Pleistocene mammals (Filoux et al. 2014). This cave is about 1.5 km long, ending in a 45 m-deep abyss and only 15 m of the cave’s anterior 1s open to tourists. The present author visited the cave in the rainy season once a year — September and October 2017-2019 — and a single population of the new copepod on the floor and stalagmites, which are formed by dripping water, was collected at about 4-5 m from the cave’s entrance (Fig. 1). & Thailand 900 km Figure 1. The sampling site and habitat of B. javabhumi sp. nov.: A. Sampling site indicated with white circle (©); B. The entrance Sampling and specimen preparation Samples were collected using a hand net (60 um) from drip pools on the stalagmites and the cave floor and then fixed immediately in ca. 70% ethanol. Adult ani- mals were picked out and preserved with 70% ethanol in 1.5 ml microtube. Adult specimens were dissected under an Olympus SZ51 stereomicroscope in a mixture of glycerol and 70% ethanol (ratio ~ 1:10 v/v). Dis- sected specimens were mounted in pure glycerol and sealed with transparent nail polish. Permanent slides with dissected animals were examined with an Olympus compound microscope (CX31) at 1000= magnification. Pencil drawings were made with a drawing tube (an Olympus U-Da) mounted on a compound microscope, then the final drawings were scanned for correction in the CORELDRAW 12.0 graphic programme. Specimens for scanning electron microscopy (SEM) were dehydrat- ed in progressive ethanol concentrations (70%, 80%, 90%, 95%, 100% and 100% absolute ethanol) for 15 min each concentration. Specimens were dried in a critical point dryer using liquid carbon dioxide as the exchange medium. Dried specimens were mounted on stubs using adhesive tape under the stereomicroscope. Specimens were coated with gold in a sputter-coater. The SEM pho- tographs were made using a scanning electron micro- scope (LEO 1450 VP). The morphological terminology follows Huys and Boxshall (1991). Specimens were deposited at the Natural History Museum, London, United Kingdom (NHMUK) and the Nakhon Phanom University, Faculty of Science, Thailand (NPU). of the cave; C. Dripping water from the plant roots penetrating from the surface; D. pool on stalagmite. zse.pensoft.net Zoosyst. Evol. 97 (1) 2021, 97-109 Abbreviations The following abbreviations are used throughout the text and figures: A aesthetasc; Enp endopod; Exp exopod; Exp/Enp-n = exopodal segment n/endopodal segment n; P1—P6 swimming legs 1-6; Sp spine/spines. Taxonomic section Order Cyclopoida Rafinesque, 1815 Family Cyclopidae Rafinesque, 1815 Genus Bryocyclops Kiefer, 1927 Bryocyclops jayabhumi sp. nov. http://zoobank.org/8A037BC7-5D7F-4EC3-961B-EA 1 A6F9897F8 Figs 2-6 Type locality. A rimstone pool that is close to an entrance (see site description) in the Prakai Phet Cave (Fig. 1), Thung Luilai Subdistrict, Khon San District, Chatya- phum Province, north-eastern Thailand; coordinates of cave entrance: 16°29'03"N, 101°47'05"E, altitude: 573 m above sea level. Material examined. Holotype: one adult female dis- sected and mounted on one slide (NHMUK 2020.48); allotype: one adult male dissected and mounted on one Slide (NHMUK 2020.49); paratypes: one adult female dissected and mounted on one slide (NPU 2020-001), three adult females and three adult males preserved in 70% ethanol (NHMUK 2020.50—55), three adult females and three adult males preserved in 70% ethanol (NPU 2020-002). All specimens were collected from the type locality on 9 October 2017 by the author. Etymology. The species name is taken from the San- skrit words ‘jaya’ and ‘bhumi’, meaning ‘land of victory’ or ‘Chatyaphum’ in Thai, referring to the Chatyaphum Province, where the new species was collected. Description. Adult female. Habitus (Fig. 2A) cy- clopiform (n = 5). Body length, excluding caudal se- tae, 340 um, with prosome/urosome ratio of 1.6. Body surface ornamented with refractile points (not figured). Nauplius eye indiscernible. Prosome with length/width ratio of 1.9. Posterior margins of prosomites with smooth hyaline fringe. Cephalothorax completely fused, with several pairs of sensilla scattered dorsally on surface; pedigers 2—3 with pair of sensilla dorsally; greatest width at anterior part of cephalothorax (Fig. 2B, C). Urosome length 130-151 um; length/width ratio of 2.2 (n = 5). Genital double-somite (Fig. 2C, D) enlarged; anterior slightly wider than posterior; 1.3 times as wide as long (n = 5), with transverse sclerotised suture indicating ances- tral segmentation; with pair of dorsal sclerotised rounded 99 structures and P6 dorsolaterally; posterior margin with slightly irregular serrated hyaline fringe. Copulatory pore (Fig. 31) behind one-half length of segment; copulatory duct narrow, short, strongly sclerotised. Seminal recep- tacle with anterior expansion at about one-half length of anterior portion; lateral arm narrow, slightly posteriorly curved. Urosomites 3-4 (Fig. 2E, G) shorter than wide, 1.6 times as long as wide (n= 5); ornamented with trans- verse row of shallow pits dorsolaterally; narrow serrated hyaline fringe. Anal somite (Fig. 2E, G) short, 1.6 times as long as wide (n= 5); two dorsal sensilla at base of anal operculum; transverse row of spinules distally on ven- tral and dorsolateral side. Anal operculum (Fig. 2E) well developed, extended to tip of caudal ramus; triangular, acute tip; free margin smooth. Caudal ramus (Fig. 2E, G) asymmetrically conical, about 1.5 times as long as wide, with dorsal longitudinal keel. Lateral seta (II) bare, slight- ly shorter than caudal ramus, inserted at one-half of cau- dal ramus length. Outermost terminal seta (IIT) bipinnate, longer than caudal ramus, with spinules at insertion point on ventrolateral side. Outer terminal seta (IV) bipinnate, about 4.0 times as long as caudal ramus, with fracture plane. Inner terminal seta (V) bipinnate, about 6.0 times as long as caudal ramus, with fracture plane. Innermost terminal seta (VI) bare; short. Dorsal seta (VI) bipinnate, about 2.0 times as long as caudal ramus, inserted at distal end of longitudinal keel. Antennule (Fig. 3A). Eleven-segmented, not reach- ing posterior margin of cephalothorax, ornamented with refractile points. Armature formula as follows: 6.2.5.2.0.2.3.1+A.2.2.6+A; all setae smooth; aesthetascs slender, fused basally with seta as acrotheck. Antenna (Fig. 3B). Four-segmented, coxobasis with one distomedial seta. Enp-1 unarmed. Enp-2 with five distomedial setae. Enp-3 with seven apical setae, both ornamented with spinular row along lateral margin. All setae smooth. Mandible (Fig. 3C) with six strongly-chitinised teeth; dorsal seta on gnathobase. Palp reduced to one bare seta. Maxillule (Fig. 3D) with three strongly-chitinised teeth on precoxal arthrite; four bare setae and one pin- nate seta on inner margin. Coxobasis with three bare se- tae distally. Exp reduced to one bare seta. Enp with three bare setae. Maxilla (Fig. 3E) with precoxal endite with two pin- nate setae. Coxa with two endites: proximal endite with one bare seta; distal endite with two bare setae. Basis with two strong claw-like expansions, bare seta close to its base. Two-segmented Enp: Enp-1 with one bare seta; Enp-2 with three bare setae. Maxilliped (Fig. 3F). Four-segmented; syncoxa with transverse row of spinules and two pinnate setae. Basis and Enp-1—2 with one, one and two smooth setae, re- spectively. P1—P4 (Fig. 4A—D). P1I—P3 with two-segmented Exp and Enp; P4 with two-segmented Exp and one-segment- ed Enp. Armature formula (seta in Arabic numerals and spine in Roman numerals from outer-inner or outer-api- cal-inner margins) as follows: zse.pensoft.net Santi Watiroyram: Bryocyclops jayabhumi sp. nov. from a cave in north-eastern Thailand dl Figure 2. Bryocyclops jayabhumi sp. nov., SEM photographs of adult female: A. Habitus, dorsal view; B. Cephalothorax and pediger 2, dorsal view; C. Pediger 4—5 and genital double-somite, dorsal view; D. Urosome (without pediger 5), ventral view; E. Urosome 3—5 and caudal rami, dorsal view; F. P5 on pediger 5, dorsal view; G. Urosome 4—5 and caudal rami, ventral view; H-I. Left and right P6 on genital double-somite, dorsal view. zse.pensoft.net Zoosyst. Evol. 97 (1) 2021, 97-109 101 Figure 3. Bryocyclops jayabhumi sp. nov., adult female: A. Antennule; B. Antenna; C. Mandible; D. Maxillule; E. Maxilla; F. Maxilliped; G. Egg sacs on genital double-somite; H. Pair of spermatophore; I. Genital double-somite, ventral view. zse.pensoft.net 102 Santi Watiroyram: Bryocyclops jayabhumi sp. nov. from a cave in north-eastern Thailand =3 ss SeS> os SS > Lom SS Cf ee SS => SoCS oS = Oi = é Saxe S235 vary LES unl OS ra P2525 a2 2 Y} Figure 4. Bryocyclops jayabhumi sp. nov., adult female: A. P1; B. P2; C. P3; D. P4; E. PS. Coxa Basis Exp Enp Pl 0-0 1-I 1-0; III-2-3 0-0; 1-I+1-0 P2 0-0 1-0 1-0; III-2-3 0-0; 1-I+1-0 P3 0-0 1-0 1-0; III-2-3 0-0; 1-I+1-0 P4 0-0 1-0 1-0; III-2-2 0-I+1-0 P1 (Fig. 4A). Intercoxal sclerite with acute distal mar- gins. Coxa without inner seta. Basis with bare, slender outer seta and robust inner spine; setules on inner distal comer. Exp-1 with outer spine. Exp-2 with three spines on outer margin; two apical setae; blunt seta and two nor- mal setae on inner margin. Enp-1 without seta on inner zse.pensoft.net margin. Enp-2 with apical seta and spine; additional seta on outer margin. P2—P3 (Fig. 4B, C). Intercoxal sclerite, coxa, basis similar to P1, but basis without inner spine. Exp-1 with outer spine. Exp-2 with three outer spines, apical normal seta and blunt seta, inner blunt seta and two normal se- tae. Enp-1 without inner seta. Enp-2 with apical seta and spine; additional seta on the outer margin. P4 (Fig. 4D). Intercoxal sclerite with acute distal margins. Coxa without inner seta. Basis with slender outer seta. Two-segmented Exp. Exp-1 with outer spine. Zoosyst. Evol. 97 (1) 2021, 97-109 Exp-2 with three outer spines, apical normal seta and blunt seta, inner blunt seta and normal seta. One-seg- mented Enp, oval; with two apical pinnate setae, both shorter than segment. PS (Figs 2F, 4E). Completely fused to somite ventro- laterally, with one long and two short pinnate setae. Prox- imal (dorsal) seta arising from lateral prominence; slen- der, longer than 4.0 times of the remaining setae; distal (ventral) setae strong, subequal in length. P6 (Fig. 2H, I). Reduced to semi-circular plate, with three short elements: anterior seta articulated, two poste- he 103 rior setae fused to plate. Posterior seta as long as anterior one. Middle seta spiniform, shortest. Egg sac (Fig. 3G). Two large eggs attached ventrally to segment, with an average egg diameter of 60 um. Spermatophore (Fig. 3H). The spermatophores are generally bean-shaped. Adult male. Body length (Fig. 5A), excluding cau- dal rami, 320-335 um (mean 330 um, n = 5); smaller than female. General segmentation and ornamentation (Fig. SA—D) similar to female. Genital somite and uro- somite 3 not fused as in female, 1.8 times as long as an a WITH ep tTTTNy Lip \ ANN Higa mY Figure 5. Bryocyclops jayabhumi sp. nov., adult male: A. Habitus, dorsal view; B. Urosome, dorsal view; C. Urosome, ventral view; D. Urosome (without pediger 5), lateral view; E. Antennule. zse.pensoft.net 104 Santi Watiroyram: Bryocyclops jayabhumi sp. nov. from a cave in north-eastern Thailand wide (n= 5). Anal operculum (Fig. 5B, D) as in female, but shorter. Antennule (Fig. SE). Fifteen-segmented, genic- ulate. Armature formula as follows: 7+3A.4.2.2+A. 1.2.1.2.A.2.1+S8p.0.1.1.8+A. Antenna, mouthparts, Pl (Fig. 6A) and P5 similar to those in female. Sexual dimorphism 1s observed on P2— P4 and P6 as follows: P2 (Fig. 6B). Intercoxal sclerite with acute distal mar- gins. Coxa without inner seta. Basis with bare, slender outer seta; setules on inner distal corner. Two-segment- ed Exp, Exp-1 with outer spine. Exp-2 with three out- er spines, apical normal seta and blunt seta, inner blunt seta and two normal setae. Two-segmented Enp, Enp-1 ——— ZS ae ae Lr A rr = Le 25 ae Z2 oe = We We leo me LH f ese FL], P 6 y. C7