Agarna malayi Tiwari 1952

© 2018 Academia Sinica, Taiwan Open Access The Mesocallis Matsumura (Hemiptera: Aphididae) of the Korean Peninsula with Descriptions of Two New Species Yerim Lee1, Mariusz Kanturski2, and Seunghwan Lee1,* 1Laboratory of Insect Biosystematics, Department of Agricultural Biotechnology, Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul 08826, Republic of Korea. E-mail: [email protected] (Lee) 2Department of Zoology, Faculty of Biology and Environmental Protection, University of Silesia in Katowice, Bankowa 9, 40-007 Katowice, Poland. E-mail: [email protected] (Received 9 November 2017; Accepted 18 March 2018; Published 23 April 2018; Communicated by Chiun-Cheng Ko) Citation: Lee Y, Kanturski M, Lee S. 2018. The Mesocallis Matsumura (Hemiptera: Aphididae) of the Korean peninsula with descriptions of two new species. Zool Stud 57:17. doi:10.6620/ZS.2018.57-17. Yerim Lee, Mariusz Kanturski, and Seunghwan Lee (2018) This study reviewed the aphid genus Mesocallis from the Korean peninsula. A total of five species are recognized using morphological and molecular evidence, including two new species, Mesocallis (Mesocallis) carpinicola sp. nov. and M. (Paratinocallis) occulta sp. nov., one newly recorded species, M. (M.) pteleae Matsumura, 1919, and the two species M. (M.) sawashibae (Matsumura, 1917) and M. (P.) corylicola (Higuchi, 1972). Species description, illustrations and distributional and biological data were provided for all Korean Mesocallis species along with a pictorial key. Pairwise distances, a neighbor-joining tree and a median-joining network are also given. Key words: Aphids, Biodiversity, Calaphidinae, COI, New record, Panaphidini. *Correspondence: E-mail: [email protected] BACKGROUND The genus Mesocallis Matsumura, 1919 (Aphididae: Calaphidinae: Panaphidini) is one of the least known aphid genera in the world. It is comprised of eight East Asian species in the two subgenera: Mesocallis Matsumura, 1919 and Paratinocallis Higuchi, 1972 (Favret 2017, Remaudière and Remaudière 1997). Most of the Mesocallis species occur on host plants belonging to the family Betulaceae, except for one species, M. (M.) fagicola Matsumura, 1919, from the family Fagaceae (Blackman and Eastop 2017, Holman 2009, Quednau 2003). This genus differs from the closely related genus Pterocallis Passerini, 1860 by its narrow elliptical-formed secondary rhinaria on the 3rd antennal segment and different spinal and marginal setae patterns on the abdomen tergites (Quednau 2003). Only two species have been recorded in the Korean peninsula thus far (Quednau and Lee 2001): Paik (1972) first recorded M. (M.) sawashibae (Matsumura, 1917) from South Korea and Quednau (1979) added M. (P.) corylicola (Higuchi, 1972), which was collected in North Korea. Since then, a DNA barcoding study on this group revealed that five distinct groups are distributed in South Korea (Lee et al. 2017). Between 2001 and 2016, a large number of Mesocallis samples were collected in South Korea and examined them together with the museum specimens of Mesocallis collected in North and South Korea between 1985 and 2001. For the freshly collected specimens, molecular analyses with the partial mitochondrial cytochrome oxidase subunit I (COI) were also conducted. The results of the morphological examination Zoological Studies 57: 17 (2018) doi:10.6620/ZS.2018.57-17 1 © 2018 Academia Sinica, Taiwan and molecular analyses identified a total of five valid species in the Korean peninsula. In the present study, we proposed two new species, M. (M.) carpinicola sp. nov. and M. (P. ) occulta sp. nov., and a new faunistic record of M. (M.) pteleae Matsumura, 1919. All five species were illustrated and keyed along with their morphological descriptions. A pictorial key to species of Mesocallis in the Korean Peninsula was provided. Pairwise distance analyses, neighbor-joining tree and median-joining network based on the partial COI were also given. MATERIALS AND METHODS Morphological identification: All specimens are deposited in the National Academy of Agricultural Science (NAAS), Jeonju-si, South Korea and the College of Agriculture and Life Sciences, Seoul National University, South Korea (CALS SNU). North Korean specimens were collected by Jan Havelka, Institute of Entomological Academy of Science of the Czech Republic, during scientific expeditions between 1985 and 2001. Aphid samples for this study were collected in South Korea between 2001 and 2015. All samples were preserved in 90% ethanol for over one month and then mounted in Canada balsam following Blackman and Eastop (2000) and Martin (1983). Illustrations for each species were taken by a digital camera attached to a microscope (Leica 400B, Leica Microsystems, Germany) at a resolution of 600 dpi. Measurements for each specimen were taken from the digital images using the analysis software Active measure ver. 3.0.3 (Mitani Co. Ltd, Japan). Aphid samples were identified using keys to Mesocallis species from Ghosh and Quednau (1990) and Quednau (2003). For further confirmation, DNA barcoding results were also applied. The abbreviations used in this study were listed as follows. CB: Chungcheongbukdo, South Korea; CN: Chungcheongnam-do, South Korea; GG: Gyeonggi-do, South Korea; GN: Gyeongsangnam-do, South Korea; GW: Gangwon-do, South Korea; JN: Jeollanam-do, South Korea; PB: Pyeonganbuk-do, North Korea; YG: Yanggang-do, North Korea; BL: body length, ANT: antennae, ANT I-VI: antennal segments I-VI, BASE: basal part of last antennal segment, PT: processus terminalis of last antennal segment, Ls ANT III: longest setae on ANT III, BD III: basal diameter of ANT III, URS: ultimate rostral segment, Cu: cubitus, Rs: radial sector, FEMUR III: hind femur, TIBIAE III: hind tibiae, HT I: first segment of hind tarsus, HT II: second segment of hind tarsus, SIPH: siphunculus, ABD I-VIII: abdominal tergite I-VIII. Main morphological characters like measurements (in mm), number of setae on antennal segments, number of rhinaria and body parts ratios of Korean Mesocallis are given in table 1. Holotypes and paratypes of the new species are deposited in CALS SNU. Paratypes will be also deposited in the Hemiptera collection of the Department of Zoology, the University of Silesia in Katowice (DZUS). DNA extraction and Molecular identification: Whole genomic DNA was extracted from each sample using the DNeasy Blood and Tissue Kit (Qiagen, Dusseldorf, Germany) according to the modified manufacturer’s protocols. We conducted the nondestructive method to confirm morphological features (Lee et al. 2017). A 658bp of the COI gene region was amplified using a universal primer set: LCO1490 5'-GGTCAACAAATCATAAAGATATTGG-3' and HCO2198 5'-TAAACTTCAGGGTGACCAAAAA ATCA-3'. Polymerase chain reaction (PCR) was performed with AccuPower PCR PreMix (Bioneer, Daejeon, Korea) in 20 ml reaction mixtures under the following conditions: initial denaturation at 94°C for 3 min; followed by 35 cycles at 94°C for 30s, an annealing temperature of 45.2°C for 30s, an extension at 72°C for 1 min; and the final extension at 72°C for 5 min. Successfully amplified samples were purified using a QIAquick PCR purification kit (Qiagen, Inc.), and then sequenced directly using an automatic sequencer (ABI PrismH 3730 XL DNA Analyzer) at Bionics Inc. (Seoul, Korea). A total of 45 COI sequences of five Mesocallis species were used for the analyses (Appendix 1). Sequences were aligned using MEGA 7 (Kumar et al. 2016). Intraand inter-specific distances were calculated using a pairwise distance method based on the Kimura-2-Parameter (K2P) model (Kimura 1980). Neighbor-joining analysis (NJ) based on K2P for the final data set of 658bp was also conducted. The COI haplotypes for the final dataset were analyzed using DnaSP ver.5.1. (Librado and Rozas 2009). Median-joining network (MJ) was constructed by Network ver. 5.0.0.1. (Polzin and Daneshmand 2003). page 2 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan RESULTS TAXONOMY Genus Mesocallis Matsumura, 1919 Mesocallis Matsumura, 1919: 103. Neocallis Matsumura, 1919: 104. Nippochaitophorus Takahashi, 1961: 247. Type species: Myzocallis sawashibae (Matsumura, 1917), by original designation. Generic Diagnosis: According to Quednau (2003), Mesocallis belongs to the Pterocallis group of genera. This genus is similar to Pterocallis Passerini but can be distinguished by having two setae on ABD VIII, black-bordered fore tibiae and Cu1b of forewing distinctly dark bordered. Representatives of Mesocallis can be also distinguished from Pterocallis by oval to narrow elliptical secondary rhinaria on ANT III, epicranial suture weakly developed but present on the head of apterous viviparous females. Host plant: Species belonging to this genus are known to feed on Alnus spp., Carpinus spp., Corylus spp., Ostrya spp. (Betulaceae) and Fagus sp. (Fagaceae). Distribution: This genus is known from SouthEast Asian countries (China, India, Japan, Korea, Nepal, and Siberia). Key to the subgenera and species of Mesocallis 1. ABD margin with a single pointed seta on ABD I-IV (subgenus Mesocallis) ...................................................... 2 - ABD margin with 2-3 setae on ABD I-IV (subgenus Paratinocallis) ................................................................... 7 2. Head vertex and ANT I-III pale ......................................... 3 - Head vertex including ANT I-III blackish ........................... 5 3. PT 0.80-1.25 × BASE, URS with 4-5 accessory setae ........ .............................................................. M. (M.) sawashibae - PT 0.60-0.70 × BASE, URS with 2-3 accessory setae ..... 4 4. ANT 0.59-0.61 × BL, ANT III with 9-11 secondary rhinaria, URS 0.80-0.90 × HT II (Gosh and Quednau 1990) ............. .................................................................... M. (M.) alnicola - ANT 0.61-0.75 × BL, ANT III with 4-7 secondary rhinaria, URS 0.50-0.55 × HT II (Gosh and Quednau 1990) ............. ............................................................. M. (M.) obtusirostris 5. ANT III with 6-9 secondary rhinaria, URS 0.06-0.07 mm with 2 accessory setae (Quednau 2003) .......... M. (M.) taoi Table 1. Biometric data of alate viviparous females of Mesocallis spp. in the Korean peninsula M. (M.) carpinicola sp. nov. M. (M.) pteleae M. (M.) sawashibae M. (P.) corylicola M. (P.) occulta sp. nov. Body parts (n = 20) (n = 20) (n = 20) (n = 20) (n = 20) Length of ANT I 0.05-0.06 0.05-0.07 0.05-0.07 0.05-0.06 0.04-0.06 ANT II 0.04-0.05 0.05-0.06 0.05-0.06 0.05-0.06 0.04-0.06 ANT III 0.29-0.36 0.33-0.43 0.30-0.39 0.28-0.40 0.21-0.33 ANT IV 0.21-0.23 0.23-0.27 0.19-0.26 0.23-0.32 0.12-0.22 ANT V 0.17-0.20 0.18-0.22 0.15-0.19 0.19-0.25 0.13-0.18 BASE 0.09-0.11 0.09-0.14 0.08-0.11 0.10-0.13 0.08-0.12 PT 0.06-0.08 0.08-0.10 0.08-0.11 0.11-0.13 0.08-0.11 FEMUR III 0.31-0.37 0.37-0.45 0.30-0.42 0.30-0.39 0.23-0.32 TIBIAE III 0.51-0.59 0.65-0.80 0.45-0.64 0.55-0.70 0.41-0.63 Ls ANT III 0.01 0.01 0.01 0.01 0.01 No. of setae on ANT I 3 3-4 3 3-4 3-5 ANT II 3-4 2-4 2-3 2-3 2-3 ANT III 4-5 5-9 6-11 4-6 3-4 BASE 1 1 1 1 1 SIPH 0 0 0 0 0 ABD VIII 2 2-3 2 2 2 Ratio of PT / BASE 0.60-0.80 0.71-0.91 0.80-1.25 0.92-1.10 0.83-1.10 PT / ANT III 0.19-0.24 0.21-0.29 0.21-0.32 0.30-0.39 0.33-0.38 SIPH / BL 0.03-0.04 0.02-0.03 0.03-0.04 0.03-0.06 0.03-0.07 SIPH / ANT III 0.14-0.18 0.09-0.14 0.12-0.15 0.13-0.22 0.15-0.33 SIPH / FEMUR III 0.14-0.18 0.08-0.13 0.12-0.16 0.13-0.20 0.16-0.30 SIPH / Cauda 0.56-0.75 0.33-0.45 0.44-0.60 0.50-0.78 0.56-0.86 Abbreviations used in this table are listed in list of abbreviations. page 3 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan - ANT III with 12-16 secondary rhinaria, URS 0.10-0.14 mm with 4 or more accessory setae ........................................ 6 6. PT 0.06-0.08 mm long, URS 0.10-0.11 mm long with 4-6 accessory setae, cauda with 10-15 setae ........................... ....................................... M. (M.) carpinicola Lee Y. sp. nov. - PT 0.08-0.10 mm long, URS 0.11-0.14 mm long with 8-9 accessory setae, cauda with 7-11 setae ..... M. (M.) pteleae 7. PT 0.6-0.7 × BASE, ABD dorsum with spinulous capitate spinal setae (Quednau 2003) ............. M. (P.) yunnanensis - PT 0.83-1.10 × BASE, ABD dorsum without spinulous capitate spinal setae ......................................................... 8 8. ANT 0.75-0.92 × BL and URS 0.80-1.00 × HT II ................. .................................................................. M. (P.) corylicola - ANT 0.60-0.77 × BL and URS 1.00-1.14 × HT II ................. .............................................. M. (P.) occulta Lee Y. sp. nov. Mesocallis (Mesocallis) carpinicola Lee Y., sp. nov. (Figs. 1, 2, 3a-b, 5, 10; Table 1) urn:lsid:zoobank.org:act:966B034F-6ABD-40AA-972B4FBDADB1ED2D Material examined: Holotype: 1 alate viviparous female, Jangsan-forest, Jangsan-ri, Maam-myeon, Goseong-gun, GN, South Korea, 35°02'48"N, 128°24'44"E, on Carpinus laxiflora, Yerim Lee and Hyoseok Lee leg., 3.v.2015, no. 150503YR-11. Paratypes: 19 alate viviparous females, same data as the holotype, including DNA voucher no. Mes1-2, Mes1-3, Mes1-4, Mes1-5. Etymology: The species name, carpinicola, is derived from the genus name of the host plant (Carpinus) and the Latin suffix ‘-cola’ (dweller, inhabitant). Diagnosis: This species is close to M. (M.) pteleae in having similar coloration and Cu1b of forewing dark bordered. However, the new species can be distinguished from M. (M.) pteleae by its shorter PT, 0.06-0.08 mm (0.08-0.10 mm in M. pteleae) and shorter URS length, 0.10-0.11 mm (0.11-0.14 mm in M. pteleae), ANT III with 4-5 setae (5-9 in M. pteleae), URS with 4-6 accessory setae (8-9 in M. pteleae), cauda with 10-15 setae (7-11 in M. pteleae). Description (Alate viviparous female): Color in life: Head pale yellow, head vertex and ANT I-III fuscous, distal 1/3 of ANT IV and ANT V darken, BASE and distal 1/2 of PT also darken. Thorax and abdomen pale yellow. Legs pale, fore and hind tibiae including 1/9 of femur and tarsus black, 1/9 of middle tibiae and tarsi dark. Wing vein Cu1b of forewing bordered with black pigment. SIPH pale. Cauda and anal plate pale. Morphology: Body oval, BL, 1.30-1.55 mm. Head smooth with a flat median protrusion on frons, head vertex with 0.01 mm pointed setae, epicranial suture weakly developed, head dorsum without tubercles. ANT 6-segmented, 0.92-1.05 mm long, ANT 0.66-0.78 × BL, ANT III longest with 1216 narrow transversely elliptical secondary rhinaria in a row on the whole segment, with 4-5 short and pointed setae, ANT III-VI imbricated with very short and inconspicuous setae, Ls ANT III × 0.50 BD III, ANT IV without secondary rhinaria, ANT V shorter than ANT IV, PT 0.60-0.80 × BASE. Rostrum not reaching to middle coxae, URS 0.10-0.11 mm long with 4-6 accessory setae, URS 1.00-1.22 × BASE, 1.22-1.38 × HT II. Thorax smooth, without tubercles. Fore coxae weakly enlarged, longest setae on TIBIAE III 0.67-1.00 × middle width of TIBIAE III, HT I with 7 chaetotaxy, HT II 0.080.09 mm long. Wing vein Rs of forewing weakly developed. Dorsal spinal setae sometimes on small elevations, conspicuous tubercles not developed, ABD VIII with a pair of spinal setae, ABD margin I-IV with a single seta on short cone-shaped marginal tubercles, 4th marginal tubercle 0.03 mm. SIPH cylindrical, truncated, 0.05-0.06 mm long. Cauda knobbed, 0.07-0.09 mm long, bearing 10-15 setae. Anal plate bilobed, each lobe with 6-8 setae. Distribution: So far, the species is known from Goseong-gun in the southern part of South Korea. Host plant: The species feed on the underside of leaves of Carpinus laxiflora (Betulaceae). Remarks: This species was first referred to as Mesocallis sp. in Lee et al. (2017). Mesocallis (Mesocallis) pteleae Matsumura, 1919 (Figs. 1, 2, 3c-d, 6; Table 1) Mesocallis pteleae Matsumura, 1919: 103. Agrioaphis corylicola Shinji, 1935: 1148. Agrioaphis hashibamii Shinji, 1935: 287. Material examined: 3 alate viviparous females, Mt. Beagunsan, Gwangyang-si, JN, South Korea, 35°06'26"N, 127°37'19"E, on Carpinus sp., Yerim Lee leg., 25.vii.2013, no. 130725YR-23, including DNA voucher no. Ca2; 33 alate viviparous females, Mt. Chilgapsan, Cheongyang-gun, CN, South Korea, 36°24'48"N, 126°53'03"E, on Corylus sp., Yerim Lee and Hyoseok Lee leg., 10.v.2014, no. 140510YR-1, including DNA voucher no. Co10-1, Co10-2, Co10-3, Co10-4, Co20; 2 alate viviparous females, Mt. Chilgapsan, Cheongyang-gun, CN, South Korea, 36°24'48"N, 126°53'03"E, on Corylus sp., Yerim Lee and Hyoseok Lee leg., 6.v.2016, no. 160506YR-2. Diagnosis: This species is similar to M. page 4 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan Fig. 1. Collection sites of Mesocallis spp. specimens collected in this study (●) and museum specimens (♦). N page 5 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan (M.) taoi Quednau by the color of ANT and black pigmented frons. However, it can be distinguished by its longer ANT III length, 0.33-0.43 mm (0.140.20 mm in M. taoi) and long and slender URS, 0.11-0.14 mm (0.06-0.07 mm in M. taoi). Redescription (Alate viviparous female): Color in life: Head pale, head vertex and ANT I-III fuscous, compound eye red, distal joint of ANT IVV, BASE and distal 1/2 of PT also dark. Thorax and abdomen pale. Legs pale, fore tibiae including 1/9 of femur black, distal margin of FEMUR III dark, tarsus dark. Wing vein Cu1b of forewing dark bordered. SIPH and cauda pale. Morphology: Body oval, BL 1.51-1.76 mm long. Head smooth with a flat median protrusion on frons, head vertex with 0.01 mm pointed setae, epicranial suture developed, head dorsum without tubercles. ANT 6-segmented, ANT 0.630.74 × BL, ANT III longest with 13-16 transversely elliptical secondary rhinaria in a row on the whole segment, ANT III with 8-10 short pointed setae, ANT III-VI imbricated, ANT IV without secondary rhinaria, ANT V shorter than ANT IV, BASE with a single inconspicuous seta, PT 0.71-0.91 × BASE. Rostrum not reaching to middle coxae, URS 0.110.14 mm long with 8-9 accessory setae, URS 0.861.27 × BASE, 1.22-1.50 × HT II. Thorax smooth, without tubercles. Fore coxae weakly enlarged, longest setae on TIBIAE III 1.00-1.05 × middle width of TIBIAE III, HT I with 7 chaetotaxy, HT II 0.08-0.10 mm long. Wing vein Rs of forewing weakly developed. Dorsal spinal setae sometimes on small elevations, conspicuous tubercles not developed, ABD VIII with a pair of spinal setae, Fig. 2. A pictorial key to the species of Mesocallis in the Korean peninsula. page 6 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan rarely with 3 setae, ABD margin I-IV with a single seta on short cone-shaped tubercles, 4th marginal tubercle 0.02 mm long. SIPH cylindrical, truncated, 0.03-0.06 mm long. Cauda knobbed, 0.09-0.11 mm long, bearing 8-11 setae. Anal plate bilobed, each lobe with 7-10 setae. Distribution: The species is known from Chungcheongnam-do in western Korea. (First record), China (Zhang and Zhong 1990), Japan (Higuchi 1972) Host plant: This species feeds on Alnus spp., Betula spp., Carpinus spp., Ostrya spp. (Holman 2009). In Korea, it was only found on Corylus spp. (Betulaceae). Matsumura (1919) gave Ptelea trifoliata (Rutaceae) as a host plant, however, it is unlikely to be a true host. Fig. 3. Representatives of Mesocallis (Mesocallis) of Korea in life: (a) alate viviparous female of M. (M.) carpinicola sp. nov.; (b) alatoid nymph of M. (M.) carpinicola sp. nov.; (c) alate viviparous female of M. (M.) pteleae; (d) alatoid nymph of M. (M.) pteleae; (e) alate viviparous female of M. (M.) sawashibae; (f) alatoid nymph of M. (M.) sawashibae. (a) (c) (e) (b) (d) (f) page 7 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan Mesocallis (Mesocallis) sawashibae (Matsumura, 1917) (Figs. 1, 2, 3e-f, 7; Table 1) Myzocallis sawashibae Matsumura, 1917: 374. Neocallis carpinicola Matsumura, 1919: 105. Nippochaitophorus moriokaensis Takahashi, 1961: 187. Material examined: 4 alate viviparous females, Seoul, South Korea, 37°33'59.53"N, 126°58'40.69"E, on Carpinus erosa, Woon-Ha Paik leg., 3.xi.1970, no. 6146; 1 alate viviparous female, Seoul, South Korea, 37°33'59.53"N, 126°58'40.69"E, on Carpinus koreana, Woon-Ha Paik leg., 6.v.1971, no. 6241; 1 alate viviparous female, Seoul, South Korea, 37°33'59.53"N, 126°58'40.69"E, on Carpinus erosa, Woon-Ha Paik leg., 6.v.1971, no. 6247; 1 alate viviparous female, Seoul, South Korea, 37°33'59.53"N, 126°58'40.69"E, on Carpinus cordata, Woon-Ha Paik leg., 21.x.1971, no. 6837; 2 alate viviparous females, Manmulsang-area, Mt. Geumgangsan, North Korea, 38°39'24.10"N, 128°6'17.85"E, on Carpinus cordata, Jan Havelka, 24.v.1988, no. 88HA2587; 1 alate viviparous female, Soholeub, Pocheon-si, GG, South Korea, 37°49'24"N, 127°08'18"E, on Carpinus laxiflora, Yerim Lee leg., 2.viii.2013, no. 130802YR-3, DNA voucher no. Ca4; 4 alate viviparous females, Girin-myeon, Inje-gun, GW, South Korea, 37°57'19"N, 128°19'09"E, on Carpinus sp., Duwal Keshari Ram leg., 26.ix.2013, no.130926Ram-6, including DNA voucher no. Ca52, Ca5-3, Ca5-4; 10 alate viviparous females, Mt. Manisan, Is. Ganghwado, Incheon-si, GG, South Korea, 37°36'41"N, 126°26'05"E, on Carpinus turczaninowii, Yerim Lee and Hyoseok Lee leg., 17.v.2014, no. 140517YR-1; 33 alate viviparous females, Yongdae-ri, Inje-gun, GW, South Korea, 38°11'02"N, 128°21'43"E, on Carpinus sp., Yerim Lee leg., 4.vi.2014, no. 140604YR-42, including DNA voucher no. Ca10; 13 alate viviparous females, Mt. Manisan, Is. Ganghwad, Incheon-si, GG, South Korea, 37°36'41"N, 126°26'05"E, on Carpinus turczaninowii, Yerim Lee and Hyoseok Lee leg., 21.vi.2014, no. 140621YR-3, including DNA voucher no. Ca9. Diagnosis: This species is morphologically close to M. (M.) obtusirostris Gosh by having pale head and body color, and partly pigmented ANT. However, it can be distinguished by having 6-12 secondary rhinaria in a row on basal 2/3 of ANT III (4-7 secondary rhinaria mostly restricted to basal half of ANT III in M. obtusirostris). Redescription (Alate viviparous female): Color in life: Head and antenna pale yellow, distal joint of Ant.III-VIb dark, compound eye red; Thorax and abdomen pale yellow; Legs pale; cu1b of forewing, slightly dark bordered; SIPH and cauda. Morphology: Body oval, BL 1.27-1.77 mm. Head smooth with a flat median protrusion on frons, head vertex with 0.01 mm pointed setae, epicranial suture weakly developed, head dorsum without tubercles. ANT 6-segmented, ANT 0.640.76 × BL, ANT III longest with 6-12 transversely elliptical secondary rhinaria in a row on 2/3 of segment, bearing 6-11 short pointed setae, ANT IV-VI imbricated with inconspicuous hair-like setae, ANT IV without secondary rhinaria, ANT V shorter than ANT IV, BASE with a single inconspicuous seta, PT 0.80-1.25 × BASE. Rostrum not reaching to middle coxae, URS 0.06-0.08 mm long with 4-5 accessory setae, URS 0.60-0.89 × BASE, 0.671.00 × HT II. Fore coxae weakly enlarged, longest setae on TIBIAE III 0.67-1.00 × middle width of TIBIAE III, HT I with 7 chaetotaxy, HT II 0.080.10 mm long. Wing vein Rs of forewing weakly developed. Thorax smooth, without tubercles. Dorsal abdominal setae sometimes on small elevations, conspicuous tubercles not developed, ABD VIII with a pair of spinal setae, ABD margin I-IV with a single seta on short cone-shaped tubercles, 4th marginal tubercle 0.02-0.04 mm long. SIPH cylindrical, truncated, 0.04-0.06 mm long. Cauda knobbed, 0.08-0.11 mm long, bearing 11-13 setae. Anal plate bilobed, each lobe with 5-10 setae. Distribution: This species is widely distributed in Korea (North and South). Also known from China (Zhang and Zhong 1990), Japan (Higuchi 1972), and East Siberia (Quednau and Shaposhnikov 1988). Host plant: This species was found on different Carpinus species (Betulaceae). In Korea also on Corylus sieboldiana (Paik 1972). Mesocallis (Paratinocallis) corylicola (Higuchi, 1972) (Figs. 1, 2, 4a-b, 8; Table 1) Paratinocallis corylicola Higuchi, 1972: 30. Material examined: 1 alate viviparous female, Mt. Gyeryongsan, Gyeryong-si, CN, South Korea, 36°20'7"N, 127°12'3"E, on Alnus japonica, WoonHa Paik leg., 17.ix.1963; 1 alate viviparous female, Daegwanryeong-myeon, Pyeongchanggun, GW, South Korea, 37°40'24"N, 128°42'22"E, host plant unknown, Woon-Ha Paik leg., 11-20. viii.1968; 3 alate viviparous females, Seoul, South page 8 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan Korea, 37°34'1"N, 126°57'5"E, on Physocarpus insularis, Woon-Ha Paik leg., 12.v.1971; 1 alate viviparous female, Mt. Myohyangsan, PB, North Korea, 40°1'3"N, 126°19'46"E, on Corylus heterophylla, Jan Havelka leg., no. 85HA0789; 3 alate viviparous females, Mt. Baekdusan, Naegokri, YG, North Korea, 41°7'52"N, 128°15'42"E, on Corylus heterophylla, Jan Havelka leg., 18.vi.1988, no. 88HA3031; 1 alate viviparous female, Danseong-myeon, Danyang-gun, CB, South Korea, 36°56'13"N, 128°19'24"E, on Corylus sp., Seunghwan Lee leg., 18.v.2001, no. 011517SH22; 4 alate viviparous females, Mt. Beagunsan, Gwangyang-si, JN, South Korea, 35°06'26"N, 127°37'19"E, on Corylus sieboldiana, Yerim Lee leg., 25.vii.2013, no. 130725YR-22, including DNA voucher no. Co2, Co3; 1 alate viviparous females, Mt. Manwolsan, Yangyang-gun, GW, South Korea, 37°57'11"N, 128°41'37"Eon Corylus sieboldiana, Yerim Lee leg., 14.viii.2013, no. 130814YR-4, DNA voucher no. Co4; 15 alate viviparous females, Mt. Hwangbyeongsan, Chahang-ri, Pyeongchang-gun, GW, South Korea, 37°42'27"N, 128°41'14"E, on Corylus sieboldiana, Yerim Lee leg., 15.viii.2013, no. 130815YR-11, including DNA voucher no. Co61, Co6-2, Co6-3, Co6-4, Co6-5; 1 alate viviparous females, Silnae-pass, Hwacheon-gun, GW, 38°09'05"N, 127°31'02"E, on Corylus sieboldiana, Yerim Lee leg., 1.v.2014, no. 140501YR-15, DNA voucher no. Co8-1; 2 alate viviparous females, Sangdong-ri, Yeongwol-gun, GW, South Korea, 37°47'09"N, 127°17'58"E, on Corylus sieboldiana, Yerim Lee leg., 12.vi.2014, no. 140612YR-23, DNA voucher no. Co15-1, Co15-2; 1 alate viviparous female, Is. Geojedo, GN, South Korea, 34°53'12"N, 128°37'25"E, on Corylus sp., Yerim Lee and Hyoseok Lee leg., 14.vii.2014, no. 140714YR-7, DNA voucher no. Co14. Diagnosis: This species is similar to M. (P.) yunanensis (Zhang) but it differs by having pointed setae on the dorsal abdomen, URS 0.80-1.00 × HT II (1.50-1.70 in M. yunanensis), and PT 0.90-1.10 × BASE (0.60-0.70 in M. yunanensis). Redescription (Alate viviparous female): Color in life: Head and antenna pale yellow, distal joint of ANT III-VIb dark. Thorax and abdomen Fig. 4. Representatives of Mesocallis (Paratinocallis) of in life: (a) alate viviparous female of M. (P.) corylicola; (b) alatoid nymph of M. (P.) corylicola; (c) alate viviparous female of M. (P.) occulta sp. nov.; (d) alatoid nymph of M. (P.) occulta sp. nov. (a) (c) (b) (d) page 9 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan 0.09 mm long with 4-6 accessory setae, URS 0.75-1.00 × BASE, 1.00-1.14 × HT II. Thorax smooth without tubercles. Fore coxae slightly enlarged, longest setae on TIBIAE III 1.50 × middle width of TIBIAE III, HT I with 8 chaetotaxy, HT II 0.07-0.09 mm long. Wing vein Rs weakly developed. Dorsal spinal setae sometimes on small elevations, conspicuous tubercles not developed, ABD VIII bearing a pair of short setae, ABD margin I-IV with 0.01-0.02 mm long of 2 setae on short cone-shaped tubercle, 4th marginal tubercle 0.01-0.02 mm. SIPH cylindrical, truncated, 0.04-0.06mm long. Cauda knobbed, 0.07-0.09 mm with 9-13 setae. Anal plate bilobed, each lobe with 7-10 setae. Distribution: So far, the new species is known from Gangwon-do, Gyeongsangnam-do, and Jeollanam-do in South Korea. Host plant: Representatives of the new species feed on the underside of leaves of Corylus sp. (Betulaceae). Remarks: This species was first recognized as a cryptic species of M. (P.) corylicola in the previous DNA barcoding study (Lee et al. 2017). Molecular identification The overall mean genetic distance was 7.33% for the 45 partial COI sequences of the 5 Mesocallis species. All species showed very low intraspecific distance level, ranging from 0% to 1.07% (Table 2). Among the five species, intraspecific distance was most varied in M. corylicola, with ranges of up to 1.07%; in contrast, 0% of intraspecific distance was observed among M. carpinicola sp. nov. In the remaining species, intraspecific distances were below 0.61%. Interspecific distance ranged from 6.21% to 12.72%. 11.81% to 12.72% of highest interspecific distances were observed between M. pteleae and M. corylicola (Table 2). M. corylicola and M. occulta sp. nov. showed 6.21% to 7.40% of lowest interspecific distances. The COI sequence NJ tree showed that the 45 sequences are clearly divided into five specific groups (Fig. 12). This result indicates that molecular identification based on COI barcoding region is effective for Mesocallis. Haplotype analysis results found 128 variable positions in the 658bp COI region. Overall, 19 different haplotypes were observed, consisting of 1 haplotype for M. carpinicola sp. nov., 3 for M. pteleae, 4 for M. sawashibae, 9 for M. corylicola and 2 for M. occulta sp. nov. (Fig. 13). DISCUSSION In this study, five Mesocallis species were recognized based on morphological and molecular evidence. Among them, morphological similarities were observed within two different species pairs, M. carpinicola sp. nov. and M. pteleae (e.g. having a similar coloration and Cu1b of forewing dark bordered), and M. corylicola and M. occulta sp. nov. (e.g. having a pale body color and length of URS 0.07-0.09 mm). In particular, the latter species pair shows only some length differences. M. occulta sp. nov. has recently been disclosed with the comprehensive DNA barcoding of Calaphidinae (Lee et al. 2017). This species could have been overlooked as a result of intraspecific variation in M. corylicola, considering its ambiguous morphology and overlapping host-plant range. In contrast to the similar morphological features, molecular analyses based on the COI region strongly supported the separation of five species with high interspecific distance values from 6.21% to 12.72%. Interspecific distances above 6.21% are much higher than the general species delimitation value of 2.5% in the subfamily Calaphidinae (Lee et al. 2017). Such a high genetic Table 2. Intraand inter-specific pairwise genetic divergence (%) based on the K2P model for five Mesocallis species M. carpinicola sp. nov. M. pteleae M. sawashibae M. corylicola M. occulta sp. nov. (n = 4) (n = 9) (n = 6) (n = 16) (n = 10) M. carpinicola sp. nov. 0 M. pteleae 7.71-8.01 0-0.61 M. sawashibae 9.92-10.10 9.88-10.59 0-0.46 M. corylicola 10.4-11.1 11.81-12.72 6.87-8.07 0.0-1.07 M. occulta sp. nov. 11.47-12.01 10.76-12.01 7.86-8.20 6.21-7.40 0-0.46 page 16 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan Fig. 12. Neighbor-joining tree of COI partial gene sequences of Mesocallis spp. (45 sequences of 5 species) page 17 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan distance was also supported in the NJ tree and MJ network (Figs. 12-13). Each species group was distinctly separated from other species groups. These results suggest that DNA barcoding can be applied for rapid and reliable species identification in this group. Haplotype diversity differed greatly depending on species. M. corylicola showed highest haplotype diversity with 9 haplotypes (Fig. 13). However, only 1 haplotype was found in M. carpinicola sp. nov. These differences may be due to differences in number of collection sites. M. corylicola is one of the most common species in the Korean peninsula and thus we could compare various populations from a number of regions of South Korea (Fig. 1). However, we could not make a comparison between various populations of M. carpinicola sp. nov. due to its distributional scarcity. To date, M. pteleae has been known to occur on various host plants belonging to the family Betulaceae. However, it is questionable whether this species really has a wide host plant spectrum because polyphagous species are relatively very rare in the subfamily Calaphidinae. In this study, we found that M. carpinicola sp. nov. has similar morphological features to M. pteleae and this species has only been collected on Carpinus laxiflora thus far. Carpinus spp. are some of the best known host plants of M. pteleae. Although we collected a single specimen of M. pteleae on Carpinus sp., a colony of M. pteleae (including females and nymphs) was only observed on Corylus spp. Among individuals of the above two species, high interspecific distances were detected, ranging from 7.71% to 8.01% (Table 2). Therefore, it will be necessary to reconfirm a true host plant range for M. pteleae by performing additional molecular works for different host plants associated groups, considering the Mesocallis group share some morphological similarities between its members. CONCLUSIONS We reviewed the genus Mesocallis in the Korean peninsula. Overall, five species - including two new species and one unrecorded species - are Fig. 13. Median-joining network for COI partial gene sequences of Mesocallis spp. (45 sequences of 5 species). page 18 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan now identified in the Korean peninsula. Our study demonstrated that five Mesocallis species are clearly separated with the strong morphological and molecular evidence. We provided a number of morphological diagnostic characters and demonstrated that DNA barcoding is effective enough for species identification in this group. This study only dealt with 50% of all known Mesocallis species and thus future studies should compare more species in the genus. Future studies should also conduct molecular phylogenetic studies and investigate M. pteleae’s true host plant ranges in order to contribute to our knowledge of the Mesocallis group’s evolutionary history. Acknowledgments: This work and the two new species names have been registered with ZooBank under urn:lsid:zoobank.org:pub:0AFFCE2F97DD-4E6E-8DD0-2ED5A6E586A2. We are grateful to Hyoseok Lee for assisting with the field collection together with Yerim Lee. Special thanks to Gwanseok Lee (NAAS) for specimen loan. This research was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) and funded by the Ministry of Education, Science (NRF2015R1D1A1A01059031); it was also supported by a grant from the National Institute of Biological Resources (NIBR), funded by the Ministry of Environment (MOE) of the Republic of Korea (NIBR201701203), and Research Institute for Agricultural and Life Sciences, SNU. We would like to thank the Editor and the Reviewers for all comments and suggestions; they greatly improved the first version of the manuscript. Mariusz Kanturski gratefully acknowledges the Scholarship for Outstanding Young Scientists from the Ministry of Science and Higher Education of Poland (1165/ E-340/STYP/12/17). Authors’ contributions: Conceptualization: YL. Data curation: YL. Formal analysis: YL. Funding acquisition: YL, SL. Investigation: YL. Project administration: YL. Specimen collection: YL. Writing - original draft: YL. Writing - review and editing: YL, MK. Competing interests: YL, MK and SL declare that we have no competing interest. Availability of data and materials: Our datasets from the manuscript are deposited in Genbank. Consent for publication: Not applicable. Ethics approval consent to participate: Not applicable. REFERENCES Blackman RL, Eastop VF. 2000. Aphids on the world’s crops. An Identification and Information guide. CAB International, Wallingford, 987 pp. Blackman RL, Eastop VF. 2017. Aphids on the world’s plants. http://www.aphidsonworldsplants.info/index.htm Accessed Aug. 2017. Ghosh AK, Quednau FW. 1990. The fauna of India and the adjacent countries. Homoptera, Aphidoidea, Part V. Drepanosiphinae. Technical and General Press, Calcutta 332 pp. Favret C. 2017. Aphid species file ver. 5.0. http://Aphid. SpeciesFile.org/ Accessed Aug. 2017. Higuchi H. 1972. A taxonomic study of the subfamily Callipterinae in Japan (Homoptera: Aphididae). Insecta Matsumurana 35(2):19-126. Holman J. 2009. Palaearctic Region. Springer, 1216 pp. Kimura M. 1980. A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16(2):111-120. Kumar S, Stecher G, Tamura K. 2016. MEGA7: Molecular Evolutionary Genetics Analysis version 7.0 for bigger datasets. Mol Biol Evol 33(7):1870-1874. doi:10.1093/ molbev/msw054. Lee Y, Lee W, Kanturski M, Foottit RG, Akimoto SI, Lee S. 2017. Cryptic diversity of the subfamily Calaphidinae (Hemiptera: Aphididae) revealed by comprehensive DNA barcoding. PloS One 12(4):e0176582. doi:10.1371/ journal.pone.0176582. Librado P, Rozas J. 2009. DnaSP v5: a software for comprehensive analysis of DNA polymorphism data. Bioinformatics 25(11):1451-2, doi:10.1093/bioinformatics/ btp187. Matsumura S. 1917. A list of the Aphididae of Japan, with description of new species and genera. J Coll Agric Hokkaido Imp Univ 7(6):351-414. Matsumura S. 1919. New species and genera of Callipterinae (Aphididae) of Japan. Trans Sapporo Nat Hist Soc 7:99114. Martin J. 1983. The identification of common aphid pests of tropical agriculture. Trop Pest Manage 29:197-212. doi:10.1080/09670878309370834. Paik WH. 1972. Illustrated encyclopedia of fauna and flora of Korea. Vol. 13. Insecta Ministry of Education, Seoul, 751 pp. Polzin T, Daneshmand SV. 2003. On Steiner trees and minimum spanning trees in hypergraphs. Oper Res Lett 31(1):12-20. doi:10.1016/S0167-6377(02)00185-2. Quednau FW. 1979. A list of drepanosiphine aphids from the Democratic People’s Republic of Korea with taxonomic notes and descriptions of new species (Homoptera). In Annales Zoologici 34(19):502-525. Quednau FW. 2003. Atlas of the drepanosiphine aphids of the world part II: Panaphidini Oestrund, 1923-Panaphidina Oestrund, 1923 (Hemiptera: Aphidinae: Calaphidinae). The American Entomological Institute, Florida, 301 pp. Quednau FW, Lee S. 2001. An annotated list of drepanosiphine aphids (Hemiptera: Aphididae) from Korea. Part I: page 19 of 20Zoological Studies 57: 17 (2018) © 2018 Academia Sinica, Taiwan Saltusaphidinae to Calaphidinae from Korea with the description of a new species. Fragm Faun 44:213-227. Quednau FW, Shaposhnikov GC. 1988. A list of drepanosiphine aphids from the Soviet Far East, with descriptions of new species (Homoptera: Aphidoidea). Can Entomol 120(11):1017-1032. doi:10.4039/Ent1201017-11. Remaudière G, Remaudière M. 1997. Catalogue of the world’s Aphididae: Homoptera Aphidoidea. Institut National de la Recherche Agronomique INRA, Paris, 473 pp. Shinji O. 1935. A key for distinguishing the Japanese Agrioaphis with descriptions of three new species. Oyo-DobutsugakuZasshi 7(6):281-287. (in Japanese) Takahashi R. 1961. Four new species of Callipterinae from Japan (Aphididae, Homoptera). Mushi 35(2):3-9. Zhang G, Liu L, He F, Zhong T. 1986. Drepanosiphinae new to China. Acta Entomologica Sinica 29:404. Zhang G, Zhong T. 1990. New species and new records of Callaphididae and Aphididae in Northeast China (Homoptera: Aphididae). Acta Entomologica Sinica 33(1):84-88. Appendix 1. Detailed collection information and Genbank accession numbers. (download) page 20 of 20Zoological Studies 57: 17 (2018)