First record of Paegniodes Eaton, 1881 (Ephemeroptera, Heptageniidae) from Thailand with description of a new species Author Boonsoong, Boonsatien https://orcid.org/0000-0002-8166-0021 Animal Systematics and Ecology Speciality Research Unit (ASESRU), Department of Zoology, Faculty of Science, Kasetsart University, Bangkok 10900, Thailand fscibtb@ku.ac.th Author Auychinda, Chonlakran https://orcid.org/0000-0003-3282-3506 Department of Biology and Health Science, Mahidol Wittayanusorn School, Salaya, Nakhon Pathom, Thailand Author Sartori, Michel https://orcid.org/0000-0003-3397-3397 Museum of Zoology, Palais de Rumine, Place Riponne 6, CH- 1005 Lausanne, Switze rland & University of Lausanne (UNIL), Department of Ecology and Evolution, CH- 1015 Lausanne, Switzerland Author Khanyom, Nuttakun Animal Systematics and Ecology Speciality Research Unit (ASESRU), Department of Zoology, Faculty of Science, Kasetsart University, Bangkok 10900, Thailand text ZooKeys 2021 2021-05-10 1036 153 170 http://dx.doi.org/10.3897/zookeys.1036.64880 journal article http://dx.doi.org/10.3897/zookeys.1036.64880 1313-2970-1036-153 9A2C63ECD583418F82773DAEC567EED6 DF7518270C4755979E945A1534237C20 Paegniodes sapanensis Boonsoong, Sartori & Auychinda sp. nov. Figures 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 Materials examined. Holotype . 1 female mature larva in alcohol, deposited in ZMKU , Thailand , Nan province , Bo Kluea district , Sapan waterfall , 19°11'25.8"N , 101°11'56.3"E , 800 m , 21.III.2020 , B. Boonsoong leg. Paratypes . 3 larvae in ethanol, deposited in ZMKU , same data as holotype ; 2 larvae in ethanol, GBIFCH00834844 , deposited in MZL same locality as holotype, 26.XI.2019 , B. Boonsoong leg. ; 1 male subimago (reared from larva), 2 female subimagos (reared from larvae), 2 larvae , all in ethanol, deposited in ZMKU , same locality as holotype, 29.XI.2020 , B. Boonsoong leg. Description of larva. Body length 16.2 mm (holotype) 10.0-13.5 mm (exuvia) 7.2-11.5 mm (immature), caudal filaments ca 1.5 x of body length (immature). General colouration dark brown with pale markings on tibiae and abdominal tergites. Colouration (Figs 1 , 10A-C ). Head, thorax, legs, and abdomen dorsally dark brown. Head, thorax, legs, and abdomen ventrally whitish. Caudal filaments brown. Figure 1. Paegniodes sapanensis sp. nov., habitus of larva (Holotype). Head. Head capsule . Ovoid in shape and flattened, 1.8-2.5 mm in length, 2.5-4.1 mm in width, brown, without distinct markings (Fig. 2A ); head capsule margins smooth. Compound eyes and base of ocelli black. Figure 2. Paegniodes sapanensis sp. nov., larval morphology A head and pronotum B labrum C left mandible D enlargement of left mandible (ventral view) E right mandible F enlargement of right mandible (ventral view). Antenna (Figs 2A , 10A ). Antennae length slightly longer than head width, scape and pedicel dark brown, flagellum light brown. Labrum (Fig. 2B ). Triangular, width of labrum 1/3 of head capsule, with median notch on anterior margin, anterior margin with row of long, hair-like setae; dorsal surface with hair-like setae except at notch and nearby area. Left mandible (Fig. 2C ). Outer and inner incisors acute with serrated margins, row of bristles located near inner incisor, row of fine serrated spines locate on base of outer and inner incisors, margin between inner incisor and mola slightly concave and smooth, one distinct denticle ventrally near mola area, setae present at apex of mola (Fig. 2D ). Lateral margin without row of setae. Right mandible (Fig. 2E ). Outer and inner incisors acute with serrated margins, row of bristles located near inner incisor, row of fine serrated spines locate on base of outer and inner incisors, margin between inner incisor and mola slightly concave, tuft of setae locate on inner margin near mola, setae present at apex of mola (Fig. 2F ). Lateral margin without row of setae. Hypopharynx (Fig. 3A ). Lingua subequal to superlingua, longer than broad, with medial tuft of long, stout setae. Superlingua distally almost straight, nearly square; each superlingua with notch on anterior margin, lateral margin rounded, with fine, long, simple setae along laterodistal margin. Figure 3. Paegniodes sapanensis sp. nov., larval morphology A hypopharynx B maxilla C labium (ventral view and dorsal view) D enlargement of labium (dorsal view). Maxilla (Fig. 3B ). Crown of galea-lacinia with a row of eight comb-shaped setae, apex of maxilla with three canines; two dentisetae, and one row of long setae on inner margin, with a row of submarginal setae on ventral surface; maxillary palpi two-segmented, apical one 1.6 x length of basal segment; ventral surface of segment II with row of dense setae forming brush-like structure, apex of last segment apically lanceolate. Labium (Fig. 3C ). Labium with U-shaped separation between glossae; shape of glossae conical, inner margin covered with dense setae, outer margin with row of setae; paraglossae moderately expanded laterally, with dense apical setae; basal segment of palp slightly longer than length of apical segment; apical segment slightly pentagonal, apex with broad projection (Fig. 3D ); with dorsal transverse row of setae apically and setae brush ventrally. Thorax. Foreleg (Fig. 4A ) Coxa with well-developed, round dorsal plate; tibia subequal to femur in length, tarsus about 1/4 length of tibia; femur with regular row of bristles on outer margin and many scattered, mostly spatulate setae on dorsal surface, ventrally with whitish elongated oval area (Fig. 4B ); tibia and apex of tarsus pigmented in light yellow; tarsus brown to dark; claw with a submedian denticle and three apical denticles (Fig. 4C ). Middle and hind legs as foreleg but with patellar-tibial (fusion) sutures, apex of tibia with cluster of setae on inner surface, and claw with two or three apical denticles. Mesosternum with a distinct transverse yellow macula (Fig. 4D ). Figure 4. Paegniodes sapanensis sp. nov., larval morphology A foreleg (dorsal view) B foreleg (ventral view) C tarsal claw of foreleg D sternum. Abdomen. Terga (Fig. 6A ). Terga pale brown, each tergum with two submedian pairs of pale dots; mature larva with distinct brown median stripes on terga II-VII, with brown oblique stripes on lateral margin of terga III-VII, posterior margin of each tergum with row of strong and acute denticles, posterolateral projections extended into acute projections. Gills (Fig. 5 ). Gills on abdominal segments I-VII; gill I (Fig. 5A ) smaller than others, dorsal lamellae 1/4 in length of well-developed fibrilliform portion; gill II-VII similar in shape (Fig. 5B-G ), lamellae much longer than fibrilliform portion, tracheation clearly visible, proximal half of lamellae margin thickened and sclerotised, gill IV (Fig. 5D ) relatively larger than others. Figure 5. Paegniodes sapanensis sp. nov., larval morphology A gill I B gill II C gill III D gill IV E gill V F gill VI G gill VII. Caudal filaments (Fig. 6B ). Cerci subequal to paracercus in length, paracercus laterally with long setae on both margins of each segment, similar setae located on inner margins of cerci only. Figure 6. Paegniodes sapanensis sp. nov., larval morphology A terga II-X (dorsal view) B cerci and paracercus. Diagnostic characters of larval stage. The main diagnostic characters are: i) lamellae of gill I ca 1/4 of fibrilliform portion, ii) mandibles without dense hair-like setae on lateral margin, iii) basal segment of maxillary palp without hairlike setae on margins, and iv) apical segment of labial palp slightly pentagonal with broad projection at apex. Description of adult stages. Male subimago (in ethanol Fig. 7 , living Fig. 10E ) Body length 8.5 mm, cerci 17.5 mm, forewing 10.9 mm, hindwing 1.5 mm. Colouration (Fig. 7 ). Head, thorax and abdomen dorsally yellowish brown. Head, thorax, and abdomen ventrally light yellow. Legs yellow. Caudal filaments brownish. Figure 7. Paegniodes sapanensis sp. nov., male subimago A head and thorax (dorsal view) B forewing C hindwing D head and thorax (ventral view) E foreleg F middle leg G hind leg H abdomen (dorsal view) I abdomen (lateral view) J abdomen (ventral view) K genitalia (dorsal view) L genitalia (ventral view). Head (Fig. 7A ). Compound eyes separated by 3.0 x width of median ocellus. Thorax (Fig. 7A, D ). Pronotum and mesonotum each with pair of dark dots; forewings semitransparent, veins yellowish to brown (Fig. 7B ); hindwings 0.14 size of forewings (Fig. 7C ). Legs yellowish to yellowish brown; femora brown with proximal and distal light maculae; tibiae uniformly brown; tibiae slightly shorter than femora. Forelegs (Fig. 7E ): length of leg segments: femur 2.5 mm; tibia 2.3 mm; tarsus 2.0 mm (tarsal segments in order of decreasing length: 2>3>4>1>5). Midlegs: (Fig. 7F ) length of leg segments: femur 2.4 mm; tibia 2.1 mm; tarsus 1.2 mm (tarsal segments in order of decreasing length 2>1>5>3>4). Hindlegs (Fig. 7G ): length of leg segments: femur 2.8 mm; tibia 2.2 mm; tarsus 1.2 mm (tarsal segments in order of decreasing length: 1≥5>2≥3>4). Each leg with two claws; one blunt, and one sharp and hooked. Abdomen . Dorsally with ornamentation as in Fig. 7H , tergum I with transverse dark band, terga II-VII with distinct, reddish median band; laterally with pattern as in Fig. 7I , with clearly oblique stripes on terga III-VII but those on terga II and VIII less visible; all sterna predominantly light yellow (Fig. 7J ). Combined length of two terminal segments of gonopods half the length of basal one (Fig. 7K, L ). Penis lobes jointed at base, apices separated, each penis lobe apex slightly expanded laterally. Styliger plate with concave posterior margin, but median part convex (Fig. 7L ). Cerci reddish to brown. Female subimago (in ethanol Fig. 8 , living Fig. 10D, F ) Body length 12.8 mm, cerci 20.0 mm, forewing 14.4 mm, hindwing 2.6 mm. Colouration (Fig. 8 ). Head, thorax and abdomen dorsally brown. Head, thorax and abdomen ventrally light brown. Legs yellow. Caudal filaments brown. Figure 8. Paegniodes sapanensis sp. nov., female subimago A head and thorax (dorsal view) B forewing C hindwing D head and thorax (ventral view) E foreleg F middle leg G hind leg H abdomen (dorsal view) I abdomen (lateral view) J abdomen (ventral view) K genitalia (ventral view). Head (Fig. 8A ). Compound eyes separated by 3.0 x width of median ocellus. Thorax (Fig. 8A, D ). Pronotum and mesonotum each with pair of dark dots, sternum light brown; forewings semitransparent, veins yellowish to brown (Fig. 8B ); hindwings 0.18 the size of the forewings (Fig. 8C ). Legs yellowish to yellowish brown; femora brown with proximal and distal light maculae; tibiae uniformly brown; foretibiae equal to femora; tibiae of midleg and hindleg slightly shorter than femora. Forelegs (Fig. 8E ): length of leg segments: femur 3.1 mm; tibia 3.1 mm; tarsus 1.7 mm (tarsal segments in order of decreasing length: 2>3>5>1>4). Midlegs (Fig. 8F ): length of leg segments: femur 2.9 mm; tibia 2.7 mm; tarsus 1.4 mm (tarsal segments in order of decreasing length 2>1≥3≥5>4). Hind leg (Fig. 8G ): length of leg segments: femur 3.5 mm; tibia 3.0 mm; tarsus 1.2 mm (tarsal segments in order of decreasing length: 5>1≥2>3>4). Each leg with two claws; one blunt, and one sharp and hooked. Abdomen . Dorsally with ornamentation as in Fig. 8H , tergum I with dark band, with distinct reddish median band on terga II-VII; laterally with pattern as in Fig. 8I , with clearly oblique stripes on terga III-VII but those on terga II and VIII less visible; all sterna predominantly light yellow (Fig. 8J ); subgenital plate (sternum VII) distally rounded and subanal plate (sternum IX) extended, with shallow median notch (Fig. 8K ); cerci reddish brown, with tiny setae on surface. Description of egg. (dissected from female subimago). Length ca 155-175 µm , width ca 80-95 µm ; elongate and oval in shape (Fig. 9A, B ); chorionic surface covered with hexagonal and pentagonal mesh ridges, with 1-4 knob-terminated coiled threads (KCTs) in between (Fig. 9C ); tagenoform micropyle in equatorial area (2 or 3 micropyles clearly visible on the same side) (Fig. 9D ). Figure 9. Paegniodes sapanensis sp. nov., scanning electron micrographs of egg (dissected from subimago) A eggs B general outline C chorionic surface D micropyle. M = micropyle, KCTs = knob-terminated coiled threads. Diagnostic characters of imaginal stage. The diagnostic characters to distinguish our new species from P. cupulatus are: i) the median stripes on abdominal terga and ii) lateral margins of genital plates slightly concave near apex. Figure 10. Paegniodes sapanensis sp. nov., habitus (live) A immature larva B male larva C female larva D closer view of female subimago E male subimago F female subimago. Etymology. The specific epithet is named for the Sapan waterfall (Bo Kluea district; tourist attraction of Nan province, Thailand), where the holotype is known. Distribution. Nan province. Biological aspects. The specimens were collected from tropical mountain streams (Fig. 11A ) which are slightly disturbed by tourist activities. The larvae of the new species were found in flowing areas and the littoral zone of the streams, underneath a mostly cobble substrate (Fig. 11B, C ). Figure 11. Habitats of larvae of Paegniodes sapanensis sp. nov. A tributary of a Sapan stream B cobble substrate with bottom sand and gravel C stream bank with cobble. Molecular analysis. Two COI sequences of Paegniodes were retrieved from BOLD system and GenBank (Table 1 ). One sequence from BOLD (THMAY162-12) was based on a specimen from the same locality (Namtok Sa Pan) as this study and this specimen was identified as P. dao . However, our Kimura 2-parameter (K2P) analysis revealed that intraspecific genetic divergence of the three sequences is very low (0.03%), and we considered all sequences as belonging to the same species. In addition, the interspecific distances (COI) between the new species and Paegniodes cupulatus ranged from 11.43-11.73%. Figure 12. Distribution of the genus Paegniodes Eaton, 1881.