The nymph of Gilliesia Peters & Edmunds, 1970 (Ephemeroptera: Leptophlebiidae), with description of a new species from Thailand Author Boonsoong, Boonsatien Author Sartori, Michel text Zootaxa 2015 3981 2 253 263 journal article 10.11646/zootaxa.3981.2.6 4ecba3c3-55a4-46cf-b276-0b0fa6c4f24c 1175-5326 237013 7C5C4334-8C6C-473C-9C6C-0B92BCF90883 Gilliesia ratchaburiensis sp. nov. ( Figs 1–40 ) Material examined. HOLOTYPE : male imago (reared from nymph), THAILAND , Ratchaburi Province, tributary of Phachi River, 13°28.6’ N , 99°14.5’ E , 230 m , 30.iii.2013 , B. Boonsoong leg.. ALLOTYPE: female imago (reared from nymph), same data as holotype . PARATYPES : 4 mature nymphs, same data as holotype ( ZMKU ); 4 nymphs mature nymphs, same data as holotype ( MZL ); 9 nymph, 4.ii.2013 , same locale as holotype ; 1 male imago (reared from nymph), 15.xi.2014 , same locale as holotype (all in ZMKU ); 1 male imago (reared from nymph), 15.xi.2014 , same locale as holotype ( MZL ). Additional material. 7 nymphs, 2.ii.2013 , F. David leg.; 65 nymphs, 16.ii.2013 , F. David leg.; 6 nymphs, 30.ii.2013 , F. David leg.; same locale as holotype ( ZMKU ). FIGURE 1. Gilliesia ratchaburiensis sp. n. (1) male mature nymph, dorsal view. Description. Mature nymph ( Fig. 1 ) (in alcohol). Body length between 3.0–4.0 mm for male (n=10) and 3.5– 5.0 mm for female (n=10). Head prognathous, antennae more than 2 times width of head ( Fig. 1 ). Mouthparts ( Figs 2–16 ): Labrum with pronounced anteromedian emargination and row of setae on anterior margin directed ventrally ( Figs.2, 3 , 14 ). Dorsal surface of labrum with scattered setae and no transverse row of setae. Ventral surface of labrum with distal row of bristles ( Figs. 4 , 14 ). Left and right mandibles as in Figs 15–16 , with well-developed prostheca, without seta on outer margin. Superlinguae of hypopharynx rounded, not extended laterally, with rows of setae along anterior margin; apex of lingua cleft ( Fig. 5 ); with two dense tufts of setae. Segment 1 of maxillary palpi subequal in length to segment 3. Length of segment 3 more than ca 1.6 times of length segment 2, apex of segment 3 with numerous long and thin setae, progressively longer from base to apex ( Figs 6–7 ). Maxillae with crown of setae covering inner 3/4 of apex of galea ( Figs 6–8 ); with 3 pectinate submarginal setae (ps, Figs 8–9 ), apex of maxilla with three reduced canines (c1–c3, Fig. 9 ), proximal dentiseta (pd) and distal dentiseta (dd) entire ( Figs 8–9 ). Labium as in Figs 10–13 ; segment 2 of palpi slightly longer than 1/2 length of segment 1; segment 3 subequal in length to segment 2, triangular, with distinct stout bristles and thin setae ( Fig. 12 ); glossae distinctly ventral to paraglossae; narrow at top and wider at base, densely covered with long setae on ventral surface ( Fig. 13 ). Legs ( Figs 17–19 ): all legs with stout bristles on outer margin of femur and inner margin of tibia; tibio-patellar suture present on mid- and hindlegs; foreleg ( Fig. 17 ) with dark pigmentation on dorsal surface of femur, apex of tibia, and in middle of tarsi; midleg ( Fig. 18 ) with stout bristles on dorsal surface of femur; hindleg ( Fig. 19 ) with few short bristles on outer margin; tarsal claw ( Fig. 20 ) rather stout and not elongated, apex moderately hooked and narrow, with row of ca 10 teeth increasing in size towards apex. Abdominal terga with distinctive color pattern; terga 2–7 with the same pattern, with two median spots and two lateral spots; terga 2–9 with median longitudinal pale line; tergum 10 with two median spots ( Fig. 1 ). Posterolateral expansions present on abdominal segment 9 only ( Fig. 21 ). Gills ( Fig. 22 ): gills on segments 1–7 alike; long, slender and slightly forked at 2/5 distance from base; tracheae unbranched; no setae on gill margins. Caudal filaments with whorls of fine hair-like setae; terminal filament ( 3.4–3.8 mm ) slightly longer than cerci ( 2.9–3.3 mm ). FIGURES 2–5. Nymphal details of Gilliesia ratchaburiensis sp. n. (2) labrum in dorsal view; (3) labrum in ventral view; (4) median cleft of labrum, ventral view; (5) hypopharynx, ventral view. Male imago ( holotype in alcohol). Body length 4.24 mm , fore wing 4.18 mm , hind wing 0.40 mm . Compound eyes black; antennae pale, pedicel brown, ocelli white with dark base ( Fig. 23 ). Vertex of head and notum brown. Forefemora ( Fig. 26 ) brown, with two white spots basally and medially; tibiae pale, brown at base and apex; tarsi pale, each segment distally brownish; ratios of segments in fore legs, 0.50: 1.00 ( 1.72 mm ): 0.03: 0.37: 0.18: 0.15: 0.08. Mid- and hindlegs paler, with same markings as in foreleg. Midlegs ( Fig. 27 ): ratios of segments, 0.77: 1.00 ( 0.89 mm ): 0.13: 0.05: 0.05: 0.10. Hind legs ( Fig. 28 ): ratios of segments, 0.78: 1.00 ( 0.93 mm ): 0.11: 0.05: 0.05: 0.09. Forewing transparent, cross veins yellowish-brown; those in C-Sc, Sc-R1 and R1- Rs surrounded by dark brownish clouds forming markings as shown in Fig. 24 ; MP2 independent of vein MP1; wing base brown; two long intercalaries between CuA and CuP ( Fig. 24 ). Hind wing very small with distinct acute costal projection near apex ( Fig. 25 ). Abdomen dark brown, terga 2–7 with one pair of median longitudinal white stripes arising from anterior margin; all terga with brown posterior margins; terga 8–9 entirely brown, tergum 10 with two pale spots ( Fig. 29 ). Caudal filaments missing. Genitalia ( Fig. 33 ): forceps length 0.4 mm (ratios of segments 1.00 ( 0.32 mm ): 0.15: 0.09), slightly curved, first segment long and curved, second broader than third segment. Styliger plate with deep, V-shaped, median incision. Penis anchor-like, apical portion of each lobe bent laterally but not ventrally, sharply pointed, no spine arising from tip of penis lobe. FIGURES 6–9 . Nymphal maxillae details of Gilliesia ratchaburiensis sp. n. (6) right maxilla, ventral view; (7) left maxilla, dorsal view; (8) ventral view of maxilla; (9) SEM view of canines and dentisetae; abbreviation: c: canine, dd: distal dentiseta, pd: proximal dentiseta, ps: pectinate submarginal setae. Female imago (allotype in alcohol). Body length 4.70 mm , fore wing 4.61 mm , hind wing 0.41 mm . Body color darker than in male. Vertex of head brown; compound eyes black; ocelli white with dark base ( Fig. 30 ). Pronotum pale, with dark brown pattern as in Fig. 30 . Meso- and metanotum yellowish-brown, lateral and ventral parts of abdomen pale. All legs missing. Abdominal tergal patterns similar to that of male. Color pattern of wing similar to that of male but crossveins darker ( Fig. 31 ). Apex of sternum 9 with U-shaped, deep, median incision ( Fig. 34 ). Egg . General shape ovoid; chorionic surface with small uniform tubercles subequal in size, each turbercle with indistinct ridges and shallow groove between turbercles ( Figs. 35–38 ). Diagnosis and discussion. Male imagos of Gilliesia ratchaburiensis n. sp. can be distinguished from those of G. hindustanica and G. pulchra by the following characters: 1) proportions of forelegs, 2) fore femur coloration, 3) abdominal terga patterns, 4) shape of apex of penis lobes, and 5) shape of hind wing ( Table 1 ). TABLE 1. Comparison of imaginal characteristics of three species of Gilliesia (Gillies 1951, Zhou 2004).

G. hindustanica G. pulchra	G. ratchaburiensis
forelegs	tibiae equal in length to tarsi tibiae shorter than tarsi	tibiae longer than tarsi
fore femur	dark brown grey to brown	dark brown with median pale

abdominal terga dark brown with pitch brown translucent white on terga 1–8, dark brown on terga 1–10, with on terga 1–8 with distinct markings distinct pale markings Nymphs of Gilliesia can be distinguished from those of the two Leptophlebiinae genera distributed in Southeast Asia ( Dipterophlebiodes and Habrophlebiodes ) by the posterolateral spines of the abdomen, gill shape, characteristics of the glossae, and maxillary palpi segment 3 ( Table 2 ). Gilliesia seems to be more similar to Dipterophlebiodes than to Habrophlebiodes by the following characters: posterolateral spines well-developed on abdominal segment 9 only, and gills slender and slightly forked at>1/3 distance from base ( Table 2 ).

penis	apex of the penis lobes broad, each lobe bent laterally and then ventrally	apex of the penis taped, each lobe bent laterally and then ventrally	apex of the penis lobes sharpened, each lobe bent laterally
apex of female sternum 9	with V-shaped deep median cleft	with V-shaped deep median cleft	with U-shaped deep median cleft
shape of the hind wing	costal projection well developed and rounded, apex located about 1/2 distance from base	costal projection well developed and rounded, apex located about 2/3 distance from base	costal projection well developed and acute, apex located about 3/4 distance from base

FIGURES 10–13 . Nymphal details (SEM) of Gilliesia ratchaburiensis sp. n. (10) labium in ventral view; (11) labium in dorsal view; (12) apical segment of labium; (13) paraglossae, ventral view. FIGURES 14–22 . Nymph of Gilliesia ratchaburiensis sp. n. (14) labrum, dorsal view (left) and ventral view (right); (15) right mandible, ventral view; (16) left mandible, dorsal view; (17) foreleg, dorsal view; (18) foreleg, dorsal view; (19) foreleg, dorsal view; (20) forleg claw;(21) abdominal segment 8–10, dorsal view; (22) gill 4. FIGURES 23–29 . Male imago of Gilliesia ratchaburiensis sp. n. (23) head, dorsal view; (24) forewing; (25) hindwing; (26) foreleg; (27) middle leg; (28) hind leg; (29) abdomen in dorsal view FIGURES 30–34 . Imagos of Gilliesia ratchaburiensis sp. n. (30) head of female imago, dorsal view; (31) forewing of female imago; (32) hindwing of female imago; (33) male genitalia, ventral view (inset in lateral view); (34) female genitalia, ventral view. FIGURES 35–38 . SEM pictures of eggs extracted from a female imago of Gilliesia ratchaburiensis sp. n. (35) general outline; (36–37) detail of pole; (38) detail of the chorionic structure. FIGURES 39–40 . Inner-apical angle of maxilla of Gilliesia ratchaburiensis sp. n. (39) and Habrophlebiodes prominens (40), view in optical microscope. Gilliesia nymphs have all attributes of the subfamily Leptophlebiinae as defined by Kluge (1994 , 2009 ): 1) apex of maxilla with three reduced canines; proximal and distal dentisetae present and entire; 2) no transverse row of setae on the dorsal face of labrum; 3) tibio-patellar suture present on mid- and hindlegs; and 4) superlinguae of hypopharynx rounded, not extended laterally. All these characters are considered by Kluge (1994) as plesiomorphic, hence the non-ranking plesiomorphon Leptophlebia /fg2 proposed by him ( Kluge 2009 ). Besides Gilliesia , Leptophlebiinae encompasses the following genera: Leptophlebia Westwood, 1940 (with three subgenera Leptophlebia s.s. , Paraleptophlebia Lestage, 1917 and Neoleptophlebia Kluge, 1997 ), Habrophlebiodes Ulmer, 1920 and Dipterophlebiodes Demoulin, 1954 . Many authors treat Paraleptophlebia at the genus level, however. Gilliesia gills resemble somewhat those of Leptophlebia s.s. , being large in the proximal part before the fork, but contrary to Leptophlebia s.s. the fork is not suddenly constricted to form a single filament but regularly decreasing, much more similar to what is found in Habroleptoides Schönemund, 1929 (Habrophlebiinae) . Gilliesia tarsal claws are rather stout and not as elongated as in Leptophlebia s.l. They are more similar to those of Dipterophlebiodes or Habrophlebiodes , the row of teeth extending over ¾ of the claw length, and teeth increasing slightly in size distally. TABLE 2 . Comparison of nymphal characteristics of three related genera ( Habrophlebiodes , Dipterophlebiodes and Gilliesia ). Habrophlebiodes Dipterophlebiodes Gilliesia posterolateral expansions of well developed on well developed on well developed on abdominal abdomen abdominal segments 8 and 9 abdominal segments 9 only segments 9 only

gills 4	gills long, more slender and deeply forked at 1/6 basally	gills long, slender and deeply forked at 1/3 basally	gills long, slender and slightly forked at 2/5 basally
hypopharynx	lingua rectangular or cordate with two dense tufts of setae, superlingua slightly extended laterally	lingua rectangular with two dense tufts of setae, superlingua not extended laterally	lingua rectangular with two dense tufts of setae,, superlingua not extended laterally
glossae	taped, with scattered long hairs on ventral surface	conical, with marginal long hairs on ventral surface	narrow taped, with dense thickened-long hairs on ventral surface
maxilla palpi segment 3	length shorter than 1.6 times the length of segment 2; apical-pointed, with scattered setae	length shorter than 1.6 times the length of segment 2; apical-blunted, with few setae	length more than 1.6 times the length of segment 2; apicalblunted, with numerous setae

In Gilliesia , the posterolateral spines are only developed on abdominal segment 9, as in Dipterophlebiodes , whereas they are also present on segment 8 in Leptophlebia s.l. (strongly developed in Leptophlebia s.s. but more reduced in some Paraleptophlebia ) and Habrophlebiodes . In addition, maxillary canines of Gilliesia are more reduced than in Habrophlebiodes , ( Figs 39–40 ), whereas Peters (1972) did not mention the details of maxilla in Dipterophlebiodes . The shape of Gilliesia superlinguae resembles those of Dipterophlebiodes , which are not extended laterally, whereas in Habrophlebiodes they are slightly extended ( Peters & Edmunds 1970, Fig. 200 ). Gilliesia possess two dense tufts of setae on ventral side of the lingua, as in Habrophlebiodes ; this character is not mentioned for Dipterophlebiodes ( Peters 1972, Fig. 5 , p. 54 ). At the imaginal stages, too, Gilliesia is easily distinguished from Leptophlebia s.l. by the presence of a hindwing costal process, and by the male penis lobes which do not possess “a ventral appendage arising from apex of each penis lobe” sensu Peters & Edmunds (1970, p. 177) . The eggs of Gilliesia , here also described for the first time, resemble those of Habrophlebiodes by the arrangement of chorionic tubercules (see Kang & Yang 1994 , Fig. 17 ). Eggs of Leptophlebia s.l. possess a very different chorionic structure (see for instance Koss 1968 , Figs 30–36 ), without tubercles. The genus Gilliesia was previously known from northern India ( Gillies 1951 , Peters & Edmunds 1970 ) and mainland China ( Zhou 2004 ). The discovery of a new species of this genus in Thailand extends its distribution to part of the Indo-Malayan mountain system in Southeast Asia, the locality of the new species being located in the Tenasserim Mountain Range. We may expect that some Oriental species described under the generic name Paraleptophlebia at the nymphal stage only may actually be members of Gilliesia . Among them, Paraleptophlebia spina Kang & Yang, 1994 from Taiwan could be a good candidate, because of the reduced maxillary canines ( Kang & Yang 1994, Fig. 8 D ) and the shape of the eggs (ibid, Fig. 19 ). Etymology. The specific epithet is named for the province, where the type material was collected: Ratchaburi province, Thailand . Biological notes. The type-locality is located in a bamboo forest. The stream is characterized by a pronounced slope (25 % on the 200 m ) and an alternation of riffles, pools and waterfalls up to 10 m high. The stream bed is mostly composed of boulders, cobbles, sand and leaf litter. The nymphs of Gilliesia ratchaburiensis n. sp. were found in leaf bags of Bambusa bambos and Lagerstroemia floribunda in both pools and riffles habitats ( David & Boonsoong 2014 ). Nymphs were found mostly together with those of Habrophlebiodes prominens Ulmer, 1939 and Isca janiceae Peters & Edmunds, 1970 .