Research Article |
Corresponding author: Natalia Kirichenko ( nkirichenko@yahoo.com ) Academic editor: Erik J. van Nieukerken
© 2018 Natalia Kirichenko, Paolo Triberti, Shigeki Kobayashi, Toshiya Hirowatari, Camiel Doorenweerd, Issei Ohshima, Guo-Hua Huang, Min Wang, Emmanuelle Magnoux, Carlos Lopez-Vaamonde.
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.
Citation:
Kirichenko N, Triberti P, Kobayashi S, Hirowatari T, Doorenweerd C, Ohshima I, Huang G, Wang M, Magnoux E, Lopez-Vaamonde C (2018) Systematics of Phyllocnistis leaf-mining moths (Lepidoptera: Gracillariidae) developing on dogwood (Cornus spp.) in Northeast Asia, with the description of three new species. ZooKeys 736: 79-118. https://doi.org/10.3897/zookeys.736.20739
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During an ongoing DNA-barcoding campaign of the leaf-mining moths that feed on woody plants in Northeast Asia, four lineages of the genus Phyllocnistis (Gracillariidae, Phyllocnistinae) were discovered on dogwood (Cornus spp): P. cornella Ermolaev, 1987 on C. controversa Hemsl. (Japan: Hokkaido) and three new species – one feeding on C. controversa, C. florida L. and C. macrophylla Wall. in Japan (Honshu, Shikoku, Kyushu), a second species on C. macrophylla in China (Yunnan) and a third on Siberian dogwood Cornus alba L. in Russia (Siberia). All these species showed differences in morphology, in the barcode region of the cytochrome c oxidase I gene and in two nuclear genes (histone H3 and 28S ribosomal RNA). No correlation was found between the deep mitochondrial splits observed and the Wolbachia infection pattern. Based on both morphological and molecular evidence, the three recently discovered lineages are described here as new species: P. indistincta Kobayashi & Triberti, sp. n. (Japan), P. saepta Kirichenko, Ohshima & Huang, sp. n. (China) and P. verae Kirichenko, Triberti & Lopez-Vaamonde, sp. n. (Russia). In addition, the authors re-describe the adult morphology of P. cornella, provide the first record of this species from Japan and highlight the diagnostic characters that allow these Cornus-feeding Phyllocnistis species to be distinguished.
Phyllocnistis , new species, Cornus , Russia, Japan, China, DNA barcoding, nuclear genes, Wolbachia
Leaf-mining micromoths of the family Gracillariidae have been the focus of recent taxonomic studies in particular in the Palaearctic region (
To date, 109 species have been recognised within the genus Phyllocnistis (
Phyllocnistis species are found in six biogeographical realms: 36 species are known from the Oriental realm, 28 species from the Neotropics, 18 species from the Palaearctic, 17 species from Australasia, 16 species from the Nearctic and four species from the Afrotropics; five species occur in more than one realm, partly through introductions (
Until recently, only one species, P. cornella Ermolaev, 1987, has been recorded feeding on dogwood Cornus (Cornales: Cornaceae). It was described from the Russian Far East, from Kunashir, the southernmost island of the Kuril Islands, on C. controversa (
The Eastern part of Eurasia, with its vast forests and little explored mountain ranges, still holds many new species of Lepidoptera. During an ongoing DNA-barcoding campaign of Gracillariidae leaf-mining moths from Northeast Asia, four divergent Phyllocnistis lineages feeding on Cornus spp. were detected: two from Japan, one from China and one from Russia. DNA barcoding, nuclear data and morphology of adults confirmed the presence of three new species that are described here: P. indistincta Kobayashi & Triberti sp. n. (Japan: Honshu, Shikoku, Kyushu), P. saepta Kirichenko, Ohshima & Huang sp. n. (China: Yunnan) and P. verae Kirichenko, Triberti & Lopez-Vaamonde, sp. n. (Russia: Siberia). In addition, the authors also re-describe the adult morphology of the closely related East Asian P. cornella, highlighting diagnostic characters that help to distinguish Cornus-feeding Phyllocnistis and provide the first records of P. cornella from Japan (Hokkaido). Female genitalia and pupal morphology of P. cornella are described here for the first time.
In total, 212 specimens have been studied (Suppl. material
Japan. Leaf mines and individuals of P. cornella were sampled on Hokkaido (2 specimens) and P. indistincta on Honshu (137 specimens), Shikoku (1 specimen) and Kyushu (19 specimens) on different Cornus species (Suppl. material
China. Leaf mines of P. saepta with one living larva and two pupae were sampled on C. macrophylla in one location in Yunnan Province (Weixi) in July 2016 (Suppl. material
Russia. Leaf mines with individuals of P. verae were collected on Siberian dogwood C. alba in Siberia (Krasnoyarsk, near village Borovoe, along the river Yenisei) in a forested area, in June–July 2015. Overall, 11 larvae were preserved in 96 % ethanol and six adults were pinned (Suppl. material
Photographs of leaf mines were taken in the field and in the laboratory using an OLYMPUS μ1060 digital camera (in Japan) and a digital camera Sony Nex3 (in Russia). Additionally, leaf mines of P. indistincta were scanned using an EPSON GT7400. Pupae of P. indistincta were dried under room temperature and sputter-coated with a 60:40 mixture of gold-palladium for examination with a scanning electron microscope (SEM) HITACHI SU1510 (Hitachi Ltd., Tokyo, Japan), with a lanthanum hexaboride (LaB6) source, at an accelerating voltage of 15 kV. Mounted moths of Cornus-feeding species were photographed using an OLYMPUS E-500 digital camera (P. indistincta). Other species were photographed using Zeiss Axiocam MRc 5 digital camera mounted on a Zeiss V.20 stereo microscope. Phyllocnistis cornella, P. saepta and P. verae specimens were photographed up to 30 times and then focus stacking was applied using Helicon Focus (http://www.heliconsoft.com/) or Zeiss Axiovision software.
Twenty five genitalia slides of P. cornella (two slides), P. indistincta (16), P. saepta (one) and P. verae (six) were prepared and analysed (Suppl. material
The description of forewing pattern follows
ELKU Entomological Laboratory, Kyushu University, Fukuoka, Japan.
OPU Osaka Prefecture University, Osaka, Japan.
HAU Hunan Agricultural University, Hunan, China.
SIF SB RAS Sukachev Institute of Forest, Siberian Branch of the Russian Academy of Sciences, Krasnoyarsk, Russia.
RMNH Naturalis Biodiversity Centre, Leiden, The Netherlands.
The authors DNA barcoded 14 specimens of Phyllocnistis spp. feeding on Cornus: two specimens of P. cornella, five P. indistincta, two P. saepta and five P. verae. DNA barcodes of another 33 specimens belonging to closely related Phyllocnistis species (listed above, see Materials and methods) were involved to estimate interspecific distances amongst Phyllocnistis. Nine of them (i.e. seven specimens of P. labyrinthella, one P. unipunctella and one Phyllocnistis sp.1 from Salix) were obtained by the authors, whereas DNA barcodes of the other 24 Phyllocnistis specimens were taken from BOLD and/or GenBank (Suppl. material
The following primers were used for amplification and sequencing: LCO1490 (5’ GGT CAA CAA ATC ATA AAG ATA TTG G 3’) and HCO2198 (5’ TAA ACT TCA GGG TGA CCA AAA AAT CA 3’) for the COI gene (
DNA was extracted using NucleoSpin tissue XS kit (Macherey-Nagel, Germany) according to the manufacturer’s protocol. The COI barcoding fragment (658 bp) was amplified via PCR at the standard conditions for the reaction (
DNA sequences, along with the voucher data, images and trace files, were deposited in the Barcode of Life Data Systems (BOLD) (
Barcode Index Numbers (BINs) (
Fourteen barcoded specimens of Cornus-feeding Phyllocnistis spp. were screened for infection by Wolbachia spp. (Rickettsiaceae). The two genes, wsp and fbpA, were amplified with the three sets of primers: (1) wspecF (5’-CATACCTAT TCGAAGGGATAG-3’), wspecR (5’-AGCTTCGAGTGAAACCAATTC-3’) and (2) wsp81F (5’-TGGTCCAATAAGTGATGAAGAAAC-3’), wsp691R (5’-AAAATTAAACGCTACTCCA-3’), for the gene wsp (respectively 438 bp and ~600bp) and (3) fbpa-F1 (5’-GCTGCTCCRCTTGGYWTGAT-3’), fbpa-R1(5’-CCRCCAGARAAAAYYACTATTC-3’), for the gene fbpA (429 bp) to test for the presence of other Rickettsiaceae following the standard protocols (
PCR was run on a DNA-cycling machine 9800 Fast Thermal Cycler (Applied Biosystems – Foster City, CA). Gel electrophoresis was applied to visualise the amplified products in 1.5 % agarose gel using the gel electrophoresis apparatus (RunOne, EmbiTec, San Diego, CA). The gel was stained with ethidium bromide (EtBr) at a concentration of 0.5 μg/mL for 30 minutes (
In total, 47 DNA barcodes of 13 Phyllocnistis species feeding on Cornaceae, Salicaceae, Rutaceae, Vitaceae, Fabaceae, and Oleaceae were analysed (Fig.
A neighbour-joining tree based on the COI barcode fragment of the 47 Phyllocnistis spp. specimens collected in Eurasia and beyond on woody plants from six families (indicated in the figure). Host plant genus / species is given in the figure for those leaf miners which were obtained directly from their mines; for specimens collected in other ways the host plant species remained unknown.
The sequences of Cornus-feeding species formed distinct clusters, with a maximum intraspecific divergence varying from 0 to 1.2 % versus a nearest-neighbour distance, varying from 6.1 to 13.6 %, when comparing Cornus-feeding species in pairs (Table
Intra- and interspecific genetic divergences in DNA barcode sequences among the studied Phyllocnistis species*.
Species1 | Phylocnistis cornella | P. indistincta | P. verae | P. saepta | Phyllocnistis sp. 2 | Phyllocnistis sp. 3 | P. citrella | Phyllocnistis sp. 4 | P. labyrinthella | P. extrematrix | P. unipunctella | Phyllocnistis sp. 1 | P. gracilistylella |
Phylocnistis cornella | [0] | ||||||||||||
P. indistincta | 7.3 | [0.2] | |||||||||||
P. verae | 12.6 | 11.9 | [0.3] | ||||||||||
P. saepta | 13.6 | 11.8 | 6.1 | [1.2] | |||||||||
Phyllocnistis sp. 2 | 13.8 | 13.2 | 8.9 | 10.6 | [−] | ||||||||
Phyllocnistis sp. 3 | 14.8 | 15.7 | 14.1 | 13.4 | 13.6 | [−] | |||||||
P. citrella | 14.9 | 13.9 | 15.5 | 15.4 | 14.9 | 13.0 | [0.3] | ||||||
Phyllocnistis sp. 4 | 15.5 | 15.9 | 14.5 | 16.7 | 14.9 | 12.1 | 8.4 | [−] | |||||
P. labyrinthella | 16.7 | 13.1 | 14.9 | 14.5 | 13.2 | 10.7 | 10.2 | 12.9 | [1.4] | ||||
P. extrematrix | 17.1 | 15.3 | 16.5 | 15.6 | 15.2 | 13.5 | 10.6 | 13.7 | 5.6 | [1.2] | |||
P. unipunctella | 18.9 | 17.8 | 16.3 | 16.4 | 15.7 | 12.0 | 11.8 | 14.3 | 9.9 | 11.8 | [−] | ||
Phyllocnistis sp. 1 | 19.5 | 18.3 | 18.2 | 19.2 | 16.8 | 12.6 | 11.8 | 11.4 | 13.8 | 14.3 | 12.8 | [−] | |
P. gracilistylella | 20.8 | 18.8 | 19.4 | 17.7 | 17.2 | 14.7 | 11.9 | 12.7 | 14.5 | 15.1 | 15.7 | 6.8 | [0.3] |
Similar minimum interspecific distances were observed between other representatives of the genus Phyllocnistis developing on Salicaceae: 5.6 % in the pair P. labyrinthella – P. extrematrix, 6.8 % in P. gracilistylella – Phyllocnistis sp.1 and 9.9 % in P. unipunctella – P. labyrinthella (Fig.
The closest neighbour to the Cornus-feeding Phyllocnistis group turned out to be an undescribed species (i.e. Phyllocnistis sp. 2) collected from Vitaceae in Japan, with a minimal interspecific distance of 8.9 % between P. verae and sp. 2. The cluster of Salicaceae-feeding Phyllocnistis was another close neighbour to the cluster of Cornaceae-feeding Phyllocnistis, with the minimal interspecific divergence (13.1 %) observed between P. verae and P. labyrinthella (Table
Besides three new species on Cornus, DNA barcoding data revealed four putative new species in the east: three species in Japan feeding on Parthenocissus (Vitaceae), Ligustrum (Oleaceae) and Derris (Fabaceae), respectively and one in the Russian Far East feeding on Salix (Salicaceae) (Fig.
Sequences of the nuclear genes histone H3 and 28S were obtained for previously barcoded 14 specimens of Cornus-feeding Phyllocnistis spp. (Fig.
H3 explicitly delimited four clusters, corresponding to the four lineages defined by COI: P. cornella, P. indistincta, P. saepta and P. verae. The gene 28S supported the clusters of P. saepta and P. verae, but it did not show any divergence between P. cornella and P. indistincta (Fig.
In both H3 and 28S, the number of diagnostic mutations was the least in the pairs: P. cornella – P. indistincta, (two mutations in H3 and no mutation in 28S) and P. saepta – P. verae (four mutations in H3 and two in 28S). The highest number of mutations was detected in the pair P. cornella – P. saepta, i.e. 15 and 19 mutations in 28S and H3, respectively.
Out of the 14 specimens of the four Cornus-feeding Phyllocnistis species screened for the genes wsp and fbpA, only one specimen of P. indistincta (MICRU069-16) from Japan (Honshu, Nara, Tateriko, ex Cornus kousa) showed the presence of an infection (Suppl. material
The four species described below, all feeding on Cornus, are morphologically very similar. The forewing pattern is characterised by a longitudinal fascia (lf), more or less defined, running along the costal margin proximally and then bent inwardly distally. Three costal and four apical ciliary strigulae are present, the first costal forming a transverse fascia (tf) crossing the wing. The male genitalia have slender valvae, rounded apically and a membranous phallus, finely pleated in the distal half, without cornuti and with a long phallobase/ductus ejaculatorius. The female genitalia have a cup-shaped antrum, a thin ductus and a large bursa with two flattened signa usually bearing a short median projection. The set of these characters allows this group of species to be differentiated quite easily from other congeners in the Palaearctic (
1 | Forewing with inner margin of longitudinal fascia (lf) well-defined | 2 |
– | Forewing with inner margin of lf indistinct, well-defined only distally; larva on Cornus controversa, C. macrophylla, C. kousa and C. florida in Honshu, Shikoku and Kyushu (Japan) | indistincta |
2 | Forewing with transverse fascia (tf) interrupted in middle; larva on C. alba in Siberia (Russia) | verae |
– | Forewing with tf not interrupted | 3 |
3 | Forewing with strigula-shaped dark dorsal dot, distally inner margin of costal fascia connecting with short dark strigula, lf reaching tf; larva on C. macrophylla in Yunnan (China) | saepta |
– | Forewing with dark dot absent or, if present, rounded; lf not reaching tf; larva on C. controversa in Hokkaido (Japan) and Kunashir Island (Russia) | cornella |
This Latin adjective, declined in the feminine gender, means “indistinct”. It is related to the longitudinal fascia in the forewing pattern that is basally indistinct in this species.
Forewing lustrous-white with a longitudinal white-yellow fascia and with an indistinct inner margin, three costal and four apical ciliary strigulae; male genitalia with phallus about as long as phallobase; female corpus bursae with two signa, similar in shape, the distal signa larger than the central signa.
Unlike other Cornus-feeding Phyllocnistis, the forewing pattern of P. indistincta has a well-defined outer margin of lf and an indistinct inner margin. Male genitalia differ from P. saepta by the phallus (about as long as phallobase or slightly shorter in P. indistincta); female genitalia are only distinguishable from P. verae by the size and the shape of the two signa.
Holotype (♂): Japan: Honshu, Menashi, Imai, Soni, Uda, Nara Prefecture, 34.52N, 136.11E, 630 m, ex Cornus controversa, 7.VI.2008 (larva), 16.VI.2008 em., SK853, S. Kobayashi leg. (deposited in OPU).
149 (5♂, 19♀, 125 exs). All specimens were collected in Japan.
Host Cornus controversa: Honshu, 16 exs, Nasu-Kashidoro, Nasu Imperial Villa, Yumoto, Nasu, Tochigi Prefecture, 30.IX.2006, 1–10.X.2006 em., Rear. Nos 808-1–808-10, 809-1–809-5, K. Niimi leg.; 3 exs, same locality and data, 2.VIII.2007 col., 6–8.VIII.2007 em., 982-1–982-3; 1♂ 2♀ 2exs, Okuyamada, Takayama, Nagano Prefecture, 2.VIII.2010 (larva), 2–15.VIII.2010 em., MICRU068-16, MICRU067-16, SK323♂, TRB2020♂, S. Kobayashi leg.; 1♂ 3♀, Oshirakawa, Nagawa, Matumoto, Nagano Prefecture, 17.IX.2011 (larva), 29.IX.2011 em., S. Kobayashi leg. (deposited in OPU); 4 exs, Alps Park, Matsumoto, Nagano Prefecture, 9.X.2015 (larva), 24.X.2015 em., S. Yagi & T. Hirowatari leg. (deposited in ELKU); 1 ex, Fuji-Kawaguchiko, Yamanashi Prefecture, 3.VIII.2015 (larva), 11.VIII.2015 em., S. Kobayashi leg.; 1 ex, Kirara, Dando, Shidara, Aichi Prefecture, 30.IX.2008 (larva), 6.X.2008 em., S. Kobayashi & T. Hirowatari leg.; 2♀ 4 exs, Mt. Mikusa, Nose, Osaka Prefecture, 21.V.2008 (larva), 30.V.–8.VI.2008 em., SK154, 322; S. Kobayashi & T. Hirowatari leg.; [Soni, Uda, Nara Prefecture, S. Kobayashi leg.]: 1♀, Konagao, 24.V.2008 (larva), 27.V.2008 em., SK146; 1♂ 1♀ 4 exs, Konagao, 11–8.VI.2008 (larva); 17.VI.2008 em., 1 ex, Konagao, 12.X.2008 (larva), 23.X.2008 em.; 2 exs, Kumatawa, Konagao, 30.V.2015 (larva), 3.VI.2015 em.; 1♀, Imai, 12.VII.2008 (larva), 15.VII.2008 em., SK148; 1 ex, Imai, 12.X.2008 (larva), 19–20.X.2008 em.; Menashi: 1♂ 2 exs, 23.V.2008 (larva), 3–10.VI.2008 em., SK320; 1♀ 2 exs, 7.VI.2008 (larva), 16.VI.2008 em., SK321; 1 ex, 14.VI.2008 (larva), 20.VI.2008 em.; 1 ex, 17.VI.2008 (larva), 21–24.VI.2008 em.; 1 ex, 5.VII.2008 (larva); 17.VII.2008 em.; 4 exs, 3–4.VII.2009 em., 27.VI.2009 (larva); 1 ex, 17.VI.2017 (pupa), 27.VI.2017 em. Shikoku, 1 ex, Fagus-no-mori, Kamagatani, Sawadani, Naka, Tokushima Prefecture, 24.VIII.2010 col., 27.VIII.2010 em., S. Kobayashi leg. Kyushu, Hikosan, Fukuoka Prefecture, 6 exs, 6–10.VII.1954; 7 exs, 15–26.VI.1955, H. Kuroko leg. 2 exs, Kirishima Spa., Kagoshima, 6.X.1964, H. Kuroko leg. (deposited in OPU).
Host C. florida: Honshu, [Menashi, Imai, Soni, Uda, Nara Prefecture, S. Kobayashi leg.]: 1♀, 7 exs, 26.VII.2008, 28.VII.–3.VIII.2008 em., SK164♀; 2♀ 17 exs, 27.VI.2009 (larva), 2–8.VII.2009 em., SK580 (vein); 1 ex, 19.VII.2010 (larva), 24–30.VII.2010; em.: 2 exs, Okukōchi-Sansō, Imai, 17.VI.2017 (pupa), 27.VI.2017 em., mine on upper side of bracts. Kyushu, Fukuoka, 1.VII.2016 (larva), 1.VII.2016 (1 adult, em. 11.VII.2016, rearing No. IsO-777) (deposited in OPU).
Host C. kousa: Honshu, 8 exs, Nasu-Kashidoro, Nasu Imperial Villa, Yumoto, Nasu, Tochigi Prefecture, 28.VII.2006 (cocoon), 5–8.VIII.2006 em., M. Murase leg.; 3 exs, 1.X.2008(larva), 7.X.2008em., Ohyoriai, Nagawa, Matsumoto, Nagano Prefecture, S. Kobayashi & T. Hirowatari leg.; Nara Prefecture: 2♀ 2 exs, Kawakami, 5.VIII.2011 (larva), 11–12.VIII.2011em., S. Kobayashi leg., slides TRB4151, TRB4153; 2exs, Tateriko, Nosegawa, 29.VII.2008 (larva), 4–18.VIII.2008 em., S. Kobayashi & T. Hirowatari leg.; Tottori Prefecture: 3exs, Mt. Daisen, 4.vii.1965, 9.VII.1965 em. (H. Kuroko), host: [Yamaboshi no Sōhō = (bracts)] (deposited in OPU).
Host C. macrophylla: Honshu, Nagano Prefecture, Matsumoto, N. Hirano leg.: 2 exs, Okada, 4–18.X.2004; 1 ex, Tomikusa, Anan, 30.VI.2006. 1♀, Tawamine, Konagao, Soni, Uda, Nara Prefecture, 24.V.2008 (larva), 27.V.2008 em. slide TRB4148, S. Kobayashi leg.; 2 exs, Same locality, 17.VII.2011 (larva), 20–25.VII.2011 em., S. Kobayashi leg.; 1 ex, Mt. Daisen, Tottori Prefecture, 4.VII.1965, 12.VII.1965, H. Kuroko leg. (deposited in OPU). Kyushu, 1♂, 2♀, Ito campus (Kyushu Univ.), Nishi-ku, Fukuoka, 26.V.2017 (larva), 31.V–9.VI.2017 em., S. Yagi, T. Hirowatari, K. M. M. Kyaw & C. Tsuji leg. (deposited in ELKU).
Pupa (6): Honshu, 1 ex, Oshirakawa, Nagawa, Matsumoto, Nagano, Cornus controversa, 17.IX.2011 (larva), 29.IX.2011 (preserved), S. Kobayashi leg.; Soni, Uda, Nara Prefecture, S. Kobayashi leg., Cornus controversa: 1 ex, Konagao, 4.VI.2011 (larva), 17.VI.2011 (preserved); 4 exs, Menashi, Imai, 17.VI.2017 (larva), 19.VI.2017 (preserved), (deposited in OPU).
Larva (2): Honshu, 1 ex, Takayama, Gifu Prefecture, C. controversa, 6.X.2014, RMNH.INS.30395, E.J. van Nieukerken & S. Richter leg. (deposited in RMNH); Kyushu, 1 ex, Mt. Hikosan, Soeda, Fukuoka, Cornus kousa, 1.VII.2016, rearing No. IsO-764, I. Ohshima leg. (deposited in OPU).
(Fig.
Head. Frons and vertex smooth, lustrous white. Antennae and labial palpi white yellowish.
Thorax. Tegulae, thorax and legs white. Forewing lustrous white, subapical area orange with a small apical black spot; a yellow-orange lf along costa from base to middle, margined with dark brown on both sides, then bent inwards distally, inner margin often indistinct in basal 2/3 not touching transverse fascia. Cilia white with a tf from costal 2/3 to dorsal 1/2, sometimes interrupted in the middle and three dark brown costal strigulae before apex; a black apical spot, giving origin to four divergent dark brown apical strigulae, one extending to upper part of costal cilia, the second and third to apex, the fourth to upper part of terminal cilia; terminal cilia white with a fuscous fringe line near termen. Hindwing lustrous white. This species and other members of the genus Phyllocnistis share an R1 vein arising from the apical half of the discoidal cell in the forewing (Fig.
Abdomen. Mostly white yellowish dorsally, white ventrally. In the male, coremata present on segment 8, consisting of a pair of elongate, dilated extensions bearing a terminal cluster of long slender scales. In the female, dorsally on segment 8, a pair of tufts of scales longer than those covering the segment.
Male genitalia (Fig.
Female genitalia (Fig.
(Fig.
(Figs
Phyllocnistis indistincta, mines on leaves of Cornus spp. in Japan (Honshu, Nara Prefecture). A–D Cornus controversa E–I C. florida J, K C. macrophylla A, B Konagao, Kumatawa, 670 m C, D, H–J type locality: Soni, Imai, Menashi, 630 m E, F Konagao, Tawamine, 685 m G, K Imai, Oku-Kochi Sanso, 455 m I mine on bract. Arrows show mines (A, C, G, J) and pupation site (B, D, E, F, H, I, K). Scale bar: 50 mm.
Biology of Phyllocnistis indistincta in Japan (Honshu) and its hostplant, Cornus controversa. A, B, D, K, I Nara Prefecture, Soni, 400–630 m C, E Yamanashi Prefecture, Kawaguchi-ko, 880–950 m F–J, M type locality: Nara Prefecture, Soni, Imai, Menashi, 630 m A habitat B, C serpentine mines, lower side of leaves D old mine, same side E young mines, upper side of leaves F old mines, same side G, H later instar larva I cocoon fold J pupa, dorsal view K same, ventral view L same, lateral sview M resting posture of the adult, dorsal-lateral view. Arrows show mines. Scale bars: 2 mm (G, H), 1 mm (I–M).
Biology of Phyllocnistis indistincta in Japan (Honshu) and its host plants. A–H type locality: Nara Prefecture, Soni, Imai, Menashi, 630 m, Cornus florida I, J Nara Prefecture,Obako-dake, 1000 m, C. kousa K Yamanashi Prefecture, Yamanaka-ko, C. kousa, 1000 m L Nara Prefecture, Soni, Konagao, 685 m, C. macrophylla A habitat B host plant C flowers D–F, G later instar larva H cocoon fold I host plant J mines and branches of host plant K, L serpentine mines on upper side of leaves. Arrows show mines. Scale bars: 2 mm (G, H).
Biology of Phyllocnistis indistincta on Cornus florida in Japan (Honshu, Nara Prefecture, Soni, Imai, Oku-Kochi Sanso, 455 m). A habitat B serpentine mines on upper side of leaves C flowers and bracts, an arrow shows mine D serpentine mines on upper side of bracts E same, mines and cocoon folds F cocoon fold. Arrows show mines (C, D) and pupation site (D, E). Scale bar: 2 mm (F).
In the Nasu Imperial Villa of Tochigi Prefecture,
In Japan in 2008–2016, larvae were observed from June to October. The overwintering form of this species is unknown.
(Fig.
Japan: Honshu (Tochigi, Nagano, Yamanashi, Aichi, Nara, Osaka and Tottori Prefectures), Shikoku (Tokushima Prefecture), Kyushu (Fukuoka and Kagoshima Prefectures).
In the type series of P. indistincta, some specimens have a forewing pattern which differs in the following points: 1) a well-defined inner margin of lf (Figs
The species name, verae is a patronym in commemoration of Mrs. Vera Kirichenko, the mother of the first author.
Forewing lustrous-white with a complete lf, three costal and four apical ciliary strigulae, tf interrupted; male genitalia with phallus shorter than phallobase; female corpus bursae with two signa, similar in size and shape.
Forewing pattern of P. verae is distinguished from P. saepta and P. cornella by the interrupted tf. In male genitalia, length of the phallus and the phallobase is similar to P. indistincta, but with a higher number of ventral setae (42–50). In the female genitalia, the two signa are very similar in shape and size, while they are different in the other Cornus-feeding species.
Holotype (♂): Russia, Krasnoyarsk, near village Borovoe, along the river Yenisei (left bank), rock (skala) Berkut, 55.97N, 92.55E, 144 m, ex Cornus alba, 7.VII.2016 (larva), 14.VII.2016 em., No. 6-3, TRB4200, N. Kirichenko leg. (deposited in SIF SB RAS).
(5): 4♂, TRB4116, TRB4152, TRB4199, TRB4225; 1♀, Russia, Krasnoyarsk, near village Borovoe, along the river Yenisei, rock (skala) Berkut, ex Cornus alba, 28.VI.2015, TRB4116, N. Kirichenko leg. (deposited in
Larvae (11): Russia, Krasnoyarsk, near village Borovoe, along the river Yenisei, rock (skala) Berkut, Cornus alba, 3 larvae, 28.VI.2015, 4 larvae, 5.VII.2016, 4 larvae, 7.VII.2016, N. Kirichenko leg. (deposited in SIF SB RAS).
(Figs
Head, thorax, legs and hindwing do not differ from the other Cornus-feeding species. Forewing lustrous white, subapical area orange with a small dark spot; lf well-defined on both sides; cilia white with tf always interrupted in the middle, three dark brown costal and four apical strigulae.
Abdomen. Like in cornella.
Male genitalia (Fig.
Female genitalia (Fig.
Not studied.
(Fig.
Biology of Phyllocnistis verae on Cornus alba in Russia (type locality: Krasnoyarsk, village Borovoe, left bank of Yenisei River, 144 m). A habitat B branch with mined leaves on the lower side C mine with feeding larva D fragment of mine with young larva (transmitted light) E same, incident light F line of frass and feeding larva (transmitted light) G opened mine H sap-feeding larvae, dorsal view I, J pupation near leaf margin K, L pupa. Arrows show mines (B, C), larva (D, E, F, G), frass (G, F), pupation site (I–K). Scale bars: 3 mm (D, E), 5 mm (F, G), 1 mm (H, L), 2 mm (J, K).
In 2015, by the 5th of July, when insect mines were found in nature, most larvae were at their final stage and some already had pupated. It suggests that larval development of the first generation may have started in late May. Thus, adults of the first generation can be on the wing in mid July. There are no records of the second generation. The overwintering stage remains unknown.
(Fig.
Russia, Siberia. Occurs in the southern part of Krasnoyarsk Kray, in the suburb of Krasnoyarsk. In 2015–2017, no mines were found on Cornus spp. in other regions of Asian Russia (Tyumen, Omsk, Novosibirsk Oblasts, Khanty-Mansi Autonomous Okrug, Tomsk, Kemerovo, Irkutsk Oblasts, Altai Kray, the Republic of Buryatia and Transbaikalia), nor in the Russian Far East (Amur Oblast, Primorskiy Kray, the Island Sakhalin).
The name saepta is the past participle of the Latin verb saepio, that means “to block” and refers to the strigulae-shaped blotch present on the dorsum of the forewing.
Forewing lustrous white, lf well-defined, touching tf without interrupting it, a dorsal dark brown blotch, strigula-shaped, is present in the first third, while the inner margin of lf, apically, shows a hint of a dark line along the cell; male genitalia with a small number of ventral setae (14) and phallobase about twice the length of phallus.
P. saepta is distinguished from other Cornus-feeding species by the presence of a strigula-shaped blotch on the dorsum of the forewing. It also differs from P. cornella by a long lf, touching tf, from P. verae by tf not interrupted and from P. indistincta by well-defined lf.
Holotype (♂): China, Yunnan, Weixi County, Diqing city, 27.16N, 99.26E, 2800 m, ex Cornus macrophylla, 19.VII.2016 (larva), 22.VII.2016 em., TRB4256, No. IsO-793, G.H. Huang & M. Wang leg. (deposited in HAU).
Larva (1) and pupa (1): China, Yunnan, Weixi County, Diqing city, Cornus macrophylla, 18.VII.2016 col., Nos IsO-790 and IsO-790-bis (deposited in HAU).
(Fig.
Head. Like P. indistincta.
Thorax. Tegulae and thorax white, legs not present. Forewing lustrous white, subapical area orange with a small apical black spot; lf well-defined, touching tf without interrupting it. A dorsal dark brown blotch, strigula-shaped, is present in the first third, while the inner margin of lf, apically, shows a hint of a dark line along the cell (Fig.
Abdomen. Not studied.
Male genitalia (Fig.
Female genitalia. Unknown.
Not studied.
(Fig.
Biology of Phyllocnistis saepta on Cornus macrophylla in China (type locality: Yunnan Province, Weixi, 2800 m). A host plant B, C serpentine mine on the upper side of the leaf D pupation site under leaf fold on the upper side of the leaf E serpentine mine on the low side of the leaf F pupation site between two veins at a leaf margin. Arrows show mine (B, C, E), pupation site (D, F). Scale bars: 18 mm (C), 4 mm (D), 8 mm (F).
Adults of Phyllocnistis indistincta (Japan: Honshu, Kyushu) and P. saepta (China). A–E P. indistincta A holotype male, ex Cornus controversa, Honshu, Nara Prefecture [Suppl. material
In 2016, in China (Yunnan) late instar larvae were found at the end of July.
(Fig.
Only one location is known so far in China – Yunnan Province, Weixi.
Phyllocnistis cornella Ermolaev, 1987: 39–40; Seksyaeva, 1997: 429.
Forewing lustrous white, lf from base to 1/2, then bent inwards, not touching tf, inner margin indistinct basally; in female genitalia, bursa copulatrix with two signa usually similar in shape or smaller, with a more strongly curved median projection, the caudal signum up to twice as large as the central one.
P. cornella is very similar to P. saepta. It mainly differs in the forewing pattern, with lf not reaching tf. Hokkaido specimens show a similar forewing pattern. Unlike P. verae, tf is not interrupted. The male genitalia, drawn by Ermolaev, do not show any particular differential characters. Females are indistinguishable from P. indistincta, but differ from P. verae by size and shape of signa.
The type series of P. cornella Ermolaev comprised 21 specimens (
(20): 2♂, 12♀, Russia, Kunashir Island, Alekhino, ex C. controversa, 24–25.VIII.1984; 3♂, 3♀, Russia, Kunashir Island, Tretyakovo, ex C. controversa, 25–27.VIII.1984, V.P. Ermolaev leg. The paratypes are unreachable for investigation (see Remarks).
(Figs
Adults of Phyllocnistis verae (Russia) and P. cornella (Japan: Hokkaido). A–B P. verae, ex Cornus alba, Krasnoyarsk A holotype male [Suppl. material
Forewing colouration and patterns of Phyllocnistis indistincta from Cornus florida, C. controversa and C. kousa (Japan: Honshu). A, D, E ex Cornus florida B, C, F, G ex C. controversa H ex C. kousa I adult emerged from mined bract of C. florida A, D, E type locality: Nara Prefecture, Soni [Suppl. material
Forewing colouration and patterns of Phyllocnistis indistincta from Cornus controversa and C. macrophylla (Japan: Honshu, Shikoku, Kyushu). A–I ex C. controversa J–L ex C. macrophylla A–C type locality, Honshu, Nara Prefecture, Soni, Imai, Menashi [Suppl. material
(Fig.
Head. Like P. indistincta.
Thorax. Tegulae, thorax and legs white. Forewing lustrous white, subapical area copper-coloured with a small apical black spot; lf from base to 1/2, then bent inwards, not touching tf, inner margin indistinct basally. Cilia white, tf not interrupted, three dark brown costal and four apical strigulae (Fig.
Abdomen. As in P. indistincta.
Male genitalia (Fig.
Female genitalia (Fig.
Male genitalia of Phyllocnistis indistincta (Japan: Honshu). A–C paratype, ex Cornus controversa, Nara Prefecture, gen. slide SK163 [Suppl. material
Male genitalia of Phyllocnistis verae (Russia), P. septae (China), and P. cornella (Russia). A–C P. verae, holotype, ex Cornus alba, Krasnoyarsk, gen. slide TRB4200 [Suppl. material
Female genitalia of Phyllocnistis indistincta (Japan: Honshu, Nara Prefecture). A, C paratype, ex Cornus controversa, gen. slide SK146 [Suppl. material
Female genitalia of Phyllocnistis verae (Russia) and P. cornella (Japan: Hokkaido). A–C P. verae A paratype, ex Cornus alba, Krasnoyarsk, gen. slide TRB4116 [Suppl. material
Pupa of Phyllocnistis indistincta from Cornus controvera, Japan (Honshu, Nara Prefecture) [Suppl. material
Not studied. No description was provided in
Original description of mine: “Mine is green whitish without frass, more often on the lower side of the leaves of Cornus controversa” (
No data were provided in the original description of
The host plant is Cornus controversa Hemsl. (
Russia: Kunashir island, Alekhino and Tretyakovo (Ermolaev 1897); Japan: Hokkaido, Sorachi, Kamifurano (present study).
In his paper,
In Figure
Overall, 17 species of Phyllocnistis are presently known in the Asian part of Russia, China and Japan (Table
Phyllocnistis species occurring in the Asian part of Russia, China and Japan, and their host plants.
No. | Phyllocnistis species1 | Host plant family and species | Distribution | Reference |
---|---|---|---|---|
5 | P. chlorantica Seksyaeva, 1992 | Chloranthaceae: Chloranthus japonicus | Russia: Russian Far East (Primorskiy Kray); Japan (Hokkaido, Honshu, Kyushu) | Kobayashi et al. 2011, |
6 | P. shizukagozen Kobayashi & Hirowatari, 2011 | Chloranthaceae: Chloranthus serratus, Sarcandra glabra | Japan (Honshu, Kyushu) | Kobayashi et al. 2011 |
1 | P. cornella Ermolaev, 1987 | Cornaceae: Cornus controversa | Russia: Russian Far East (Kunashir); Japan (Hokkaido) |
|
2 | P. indistincta Kobayashi & Triberti sp. n. | Cornaceae: Cornus controversa, C. florida, C. macrophylla, C. kousa | Japan (Honshu, Shikoku, Kyushu) | present paper |
4 | P. saepta Kirichenko, Ohshima & Huang sp. n. | Cornaceae: Cornus macrophylla | China (Yunnan) | present paper |
3 | P. verae Kirichenko, Triberti & Lopez-Vaamonde sp. n. | Cornaceae: Cornus alba | Russia: Siberia (Krasnoyarsk Kray) | present paper |
7 | P. hyperbolacma (Meyrick, 1931) | Daphniphyllaceae: Daphniphyllum macropodum subsp. humile | Japan (Honshu) |
|
8 | P. selenopa Meyrick, 1915 | Meliaceae: Melia azedarach | Japan (Honshu, Shikoku, Kyushu) |
|
13 | P. embeliella Liu & Zeng, 1989 | Myrsinaceae: Embelia lacta | China (Guangdong) |
|
9 | P. citrella Stainton, 1856 | Rutaceae:Citrus spp., Poncirus trifoliata, Fortunella spp. | Japan (Honshu, Shikoku, Kyushu, Ogasawara), China (Jiangsu, Zhejiang, Fujian, Hubei, Hunan, Guangdong, Hainan, Chongqing, Guizhou, Guangxi, Sichuan, Yunnan) |
|
14 | P. wampella Liu & Zeng, 1985 | Rutaceae: Clausena lansium | China (Guangdong) |
|
10 | P. gracilistylella Kobayashi & Jinbo, Hirowatari, 2011 | Salicaceae: Salix gilgiana, S. gracilistyla, S. integra, S. serissaefolia | Japan (Honshu, Kyushu) | Kobayashi et al. 2011 |
11 | P. labyrinthella (Bjerkander, 1790) | Salicaceae: Populus alba, P. tremula (Siberia, Russian Far East), P. nigra (Siberia) | Russia: Siberia (Novosibirsk, Kemerovo, Irkutsk Oblasts, Krasnoyarsk Kray, Republic of Altai, Republic of Yakutia); Russian Far East (Khabarovsk Kray, Primorskiy Kray, Kuril Islands) |
|
12 | P. saligna (Zeller, 1839) | Salicaceae: Salix fragilis, S. kochiana (Siberia); Salix sp. (Russian Far East); S. babylonica, S. bakko, S. chaenomeloides, S. gilgiana, S. gracilistyla, S. integra, S. miyabeana, S. reinii, S. sachalinensis, S. serissaefolia (Japan) | Russia: Siberia (Novosibirsk, Kemerovo Oblasts); Russian Far East (Primorskiy Kray); Japan (Hokkaido, Honshu, Kyushu, Shikoku), China (Heilongjiang, Jilin, Liaolin, Hebei, Shanxi) |
|
15 | P. unipunctella (Stephens, 1834) | Salicaceae: Populus nigra, P. balsamifera (Siberia); P. nigra, P. suaveolens (Russian Far East); P. nigra var. italica (Japan) | Russia: Siberia (Novosibirsk, Irkutsk Oblasts, Republic of Yakutia), Russian Far East (Primorskiy Kray, Kuril Islands), Japan (Hokkaido, Honshu), |
|
16 | P. toparcha Meyrick, 1918 | Vitaceae: Vitis spp., V. ficifolia var. lobata, V. flexuosa, Ampelopsis glandulosa var. heterophylla, Parthenocissus tricuspidata, Cayratia japonica | Japan (Hokkaido, Honshu, Shikoku, Kyushu, Ryukyu) |
|
17 | P. vitella Ermolaev, 1987 | Vitaceae: Vitis amurensis | Russia: Russian Far East (Primorskiy Kray) |
|
Further candidate species | ||||
18 | Phyllocnistis sp. 1 | Salicaceae: Salix sp. | Russia: Russian Far East (Primorskiy Kray) | Col: N. Kirichenko |
19 | Phyllocnistis sp. 2 | Vitaceae: Parthenocissus tricuspidata | Japan (Honshu) | Col: E.J. van Nieukerken |
20 | Phyllocnistis sp. 3 | Fabaceae: Derris trifoliata | Japan (Okinawa) | Col: A. Kawakita |
21 | Phyllocnistis sp. 4 | Oleaceae: Ligustrum japonicum | Japan (Shikoku) | Col: A. Kawakita |
Endosymbiotic bacteria like Wolbachia may manipulate host reproduction and significantly affect mitochondrial divergence within insect species (
Phyllocnistis species develop on different Cornus species: P. indistincta on C. controversa, C. kousa, C. macrophylla (native to Japan) and on C. florida (introduced in Japan from North America), P. saepta on C. macrophylla (native to China) and P. verae on C. alba (native to Siberia). The related species P. cornella was described from the Russian Far East from Cornus controversa. The above-mentioned Cornus species correspond to four subgenera: Kraniopsis (C. alba and C. macrophylla), Mesomora (C. controversa), Syncarpea (C. kousa) and Cynoxylon (C. florida) (
Dogwoods are widely used for ornamental purposes in gardens and landscaping and their distribution is ubiquitous in temperate Eurasia. The North American C. florida is planted in Japan as an ornamental tree. Cornus alba, that is native in Russia, Mongolia, Northeast China, Manchuria and North Korea, has been introduced in Europe (
In addition to the three new species of Phyllocnistis discovered on Cornus in Northeast Asia, the analysis of DNA barcodes allowed the detection of a further four putative new species of Phyllocnistis in this region, though on plant families other than Cornaceae (Table
We thank E.J. van Nieukerken (The Netherlands), H. Kuroko, A. Kawakita, N. Hirano, K. Niimi, M. Murase, S. Yagi, C. Tsuji (Japan), G. Deschka (Austria), M. Jones (USA), A. Laštůvka, Z. Laštůvka (Czech Republic), A. Cama, J. Nel (France) and P. van Wielink (The Netherlands) for providing specimens and / or DNA barcodes of Phyllocnistis spp., J.C. Koster (The Netherlands) for preparing the genitalia slide of P. cornella, C. van den Berg (The Netherlands) for helping with collection of P. cornella in Japan, S.V. Baryshnikova and M.G. Ponomarenko (Russia) for checking the collections of their institutes for Cornus-feeding Phyllocnistis and for their useful remarks. Special thanks to R. Brito and G.R.P Moreira (Brazil) for their careful reading of the latest version of our manuscript, to D. Lees (UK) for checking the English, to the reviewers R. Rougerie (France) and D. Wagner (USA) and to the editor E.J. van Nieukerken for their insightful comments and suggestions. N. Kirichenko was supported by a fellowship of LE STUDIUM®, Institute for advanced studies – Loire Valley, France (grant No. INRA-URZF-007); French Embassy in Russia, Bourse Metchnikov (grant No. 908981L, Campus France) and by the Russian Foundation for Basic Research (grant No. 15-29-02645). T. Hirowatari. and I. Ohshima were supported by JSPS KAKENHI (grant No. JP16H05766).
Table S1. Phyllocnistis species involved in the study.
Figure S1. The DNA barcoded specimens of Cornus-feeding Phyllconistis tested for presence of Wolbachia and other Rickettsiaceae