Edaphologia, No. 104: 11–18, March 29, 2019 11

Juvenile gracilliseta (Collembola, ) with four anal spines

Naoyuki Matsumoto1, Yasuhiko Suma2, and Taizo Nakamori3

16-22-2 Bunkyo, Chitose 066-0052, Japan

26-7-32 Harutori, Kushiro 085-0813, Japan

3Graduate School of Environmental and Information Sciences, Yokohama National University, 79-7 Tokiwadai, Hodogaya,

Yokohama 240-8501, Japan

Corresponding author: Naoyuki Matsumoto ([email protected])

Received 19 June 2018; Accepted 1 November 2018

Abstract Desoria gracilliseta colonizes a fruticose lichen, Cladonia sp., in a snowy habitat in Hokkaido, Japan. Ju- veniles appear in October, continue to develop under snow, and mature in May. Recent surveys from mid-September to mid-October revealed the presence of a Tetracanthura-like collembolan, in addition to D. gracilliseta juveniles. Body lengths of the Tetracanthura-like collembolan remained consistent throughout the duration of the surveys averaging 0.49 mm, whereas D. gracilliseta juveniles grew steadily from 0.54 mm to 0.8 mm. Both forms were almost indis- tinguishable except that the Tetracanthura-like collembolan had four anal spines on abdominal segment VI and short spines on the head and thorax segment II. Large individuals distinctive of D. gracilliseta collected in March were also included for comparisons in morphology and DNA sequences. Antennal segment IV and the furca developed with in- creasing body length. DNA barcode analysis, using mitochondrial cytochrome c oxidase subunit 1 and 16S ribosomal RNA genes confirmed the genetic identities of the Tetracanthura-like collembolan, and juveniles and subadults of D. gracilliseta to be D. gracilliseta. The significance of morphological changes in D. gracilliseta was discussed in terms of survival strategy against predation.

Key words: Anal spine, Desoria gracilliseta, DNA barcoding, morphological change, Tetracanthura sp.

The taxonomic entity of Tetracanthura is regarded as Introduction dubious. Juveniles possessing anal spines have been found The collembolan Desoria gracilliseta (Börner, 1909) is in (Najt, 1983), and members of Tetracanthura are active even under snow and univoltine on a fruticose lichen considered as juveniles of Isotoma or related taxa (Potapov, (Cladonia sp.) in a cold temperate region (Matsumoto et 2001). Isotoma and Desoria are closely related, both belong- al., 2018). Desoria gracilliseta was absent during summer ing to the subfamily Isotominae (e.g., Greenslade and Pota- months. Juveniles of ca. 0.5 mm long were first observed in pov, 2015). Consequently, the coexistence of ud-Tetra and D. October and grew steadily during the winter to attain body gracilliseta, as well as their morphological similarities except lengths up to 2 mm by May. Subsequent surveys from mid- anal spines made us to hypothesize that ud-Tetra metamor- September to mid-October in the same site revealed the pres- phosed into juvenile D. gracilliseta. ence of a similar collembolan that possessed four anal spines. Metamorphosis is commonly known in Isotomidae and The collembola was tentatively assigned to Tetracanthura sp. Hypogastruridae. Fjellberg (1976) referred to the phenomenon (referred to as ud-Tetra), but differed from other known spe- of morphological change associated with seasonal change as cies belonging to the genus (Tanaka, 1982). Small numbers cyclomorphosis. Cyclomorphosis usually accompanies the of juvenile D. gracilliseta were present together with a large change in the furca and is considered an adaptation to facili- number of ud-Tetra in September. They grew larger and even- tate dispersal on the snow surface (Leinaas, 1981a, 1981b; tually outnumbered ud-Tetra with time. Hågvar, 1995; Sawahata, 2005). In the case of Hypogastrura 12 Naoyuki Matsumoto, Yasuhiko Suma, and Taizo Nakamori lapponica (Axelson, 1902), cyclomorphic change was not Large specimens, distinctive of D. gracilliseta collected associated with specific instars (Leinaas, 1981b). Epitoky on March 20, 2016, were used as reference since adult or includes morphological alterations in size and shape of anal large subadult specimens are required for taxonomic determi- spines, claws, mucro, and macrochaetae, and occurs during nations (Potapov, 2001). Specimens of ud-Tetra and juvenile sexual maturity (Fjellberg, 1977). Ecomorphosis occurs in and subadult stages of D. gracilliseta, also collected from the response to environmental factors such as rising temperatures same site, were used for DNA sequence analysis. DNA was and lowered soil moisture and includes the appearance of anal extracted from individual specimens according to Aoyama et spines (Hart, 1978). Our observations showed that ud-Tetra al. (2015). DNA sequences of partial regions of mitochondrial populations were replaced by D. gracilliseta populations but cytochorome c oxidase subunit 1 (CO1) and 16S ribosomal lacked the direct evidence to indicate metamorphosis such as RNA (16S rRNA) genes were determined as described by specimens undergoing ecdysis. Then, we conducted a detailed Potapov et al. (2010). Sequences were aligned using Muscle study of morphological development and population dynam- (Edgar, 2004; https://www.ebi.ac.uk/Tools/msa/muscle/) for ics throughout the growth cycle of the two types and applied both genes, and then cleaned using Gblocks 0.91b (Castresana DNA barcoding technique to confirm the identities of ud-Tetra 2000; http://molevol.cmima.csic.es/castresana/GBLOCKS_ and D. gracilliseta. server.html) for the 16S rRNA gene. To assess the associa- tion among ud-Tetra, juvenile and subadult individuals of D. Materials and methods gracilliseta, p-distances among specimens were estimated us- Sampling was made in two spots which were distant ing MEGA7 (Kumar et al., 2016) for both genes. Specimens ca. 100 m from each other and separated by a paved road in of Desoria trispinata (MacGillivray, 1896), congeneric with D. site B (Matsumoto et al., 2018). Site B had been cleared in gracilliseta, Tetracanthella sylvatica Yosii, 1939, a species of the late 1990’s and sparsely planted with Picea glehnii (Fr. spined genus, and Sahacanthella saoriae Nakamori & Pota- Schm.) Masters in Chitose, Hokkaido, northern Japan (42˚46’ pov, 2017, the species of spined genus used by Potapov et al. N, 141˚36’ E). A lichen, Cladonia sp., colonized the exposed (2017), were also included in the analysis (Table 1). The se- soil consisting predominately of pumice beneath the P. glehnii quences obtained were submitted to GenBank under accession canopy. Eight pieces of thallus of the lichen (7 × 7 cm) were numbers LC384348–LC384353 and LC384828–LC384833. sampled along with ca. 1 cm thick of pumice from each spot Specimens were deposited to the Tottori Prefectural Museum from September 16 to October 15 in 2017 to study population under accession numbers TRPM-AAr-0000756–TRPM- dynamics of ud-Tetra and juvenile D. gracilliseta. Collembo- AAr-0000761. lans were collected in 100% isopropanol with Tullgren fun- nels, sorted under a dissecting microscope, and preserved in Results 70% ethanol. The 2017 observations showed that numbers of ud-Tetra, Specimens of ud-Tetra and juvenile D. gracilliseta were first observed on September 16, peaked in late September, and arbitrarily sampled and mounted in Hoyer’s medium on glass dropped to zero by mid October (Table 2). Body lengths of slides for morphological comparison. Mounted specimens col- ud-Tetra remained consistent throughout the observation pe- lected from spots 1 and 2 in 2017 were photographed along riod, ranging from 0.47 to 0.5 mm (Table 2). Desoria gracil- with a ruler with a digital camera (Stylus SH-1, Olympus), liseta numbers were initially near zero but soon outnumbered and enlarged photos were used to determine body lengths as ud-Tetra in both spots (Table 2). Individuals of D. gracilliseta a distance between both ends of head and abdominal segment were initially larger than those of ud-Tetra and grew with time VI. Up to 20 individuals were determined. Antennal segment from 0.54 to 0.8 mm whereas the size of ud-Tetra remained IV and the furca of ud-Tetra and juvenile D. gracilliseta col- static throughout the sampling interval (Table 2). lected from spot 1 on October 4, 2016, were photographed us- Ud-Tetra possessed four anal spines on abdominal seg- ing an Olympus DP22 digital camera mounted on an Olympus ment VI, which was distinct from segment V (Fig. 1). When BX53 biological microscope. Lengths of antennal segment IV comparing individuals collected on the same date, ud-Tetra and the furca were precisely determined, using the DP2-SAL (Fig. 2A) closely resembled juvenile individuals of D. gracil- Olympus firmware. Number of individuals determined varied liseta (Fig. 2B) except that juveniles were slightly larger and from 11 to 20. lacked anal spines. Short spines were present on the head and

Juvenile Desoria gracilliseta with anal spines 13

Table 1. Voucher sample information and GenBank accession number of sequence data used in molecular analysis.

Voucher sample information GenBank accession number Reference Taxa Individual Museum ID COI gene 16S rRNA ID gene Desoria gracilliseta subadult ZI03 TRPM-AAr-0000760 LC384350 LC384830 Present study Desoria gracilliseta subadult ZI04 TRPM-AAr-0000761 LC384351 LC384831 Present study D. gracilliseta juvenile ZI05 TRPM-AAr-0000758 LC384352 LC384832 Present study D. gracilliseta juvenile ZI06 TRPM-AAr-0000758 LC384353 LC384833 Present study Tetracanthella sp. (ud-Tetra) ZI24 TRPM-AAr-0000756 LC384348 LC384828 Present study Tetracanthella sp. (ud-Tetra) ZI02 TRPM-AAr-0000757 LC384349 LC384829 Present study Desoria trispinata KI09 TRPM-AAr-0000691 LC213064 LC213078 Potapov et al. (2017) Desoria trispinata KI10 TRPM-AAr-0000692 LC213065 LC213079 Potapov et al. (2017) Tetracanthella sylvatica RI73 TRPM-AAr-0000695 LC213068 LC213082 Potapov et al. (2017) Tetracanthella sylvatica RI74 TRPM-AAr-0000696 LC213069 LC213083 Potapov et al. (2017) Sahacanthella saoriae RI63 TRPM-AAr-0000684 LC209816 LC209818 Potapov et al. (2017) Sahacanthella saoriae RI64 TRPM-AAr-0000685 LC209817 LC209819 Potapov et al. (2017)

Table 2. Change in number of individuals and body lengths of Tetracanthura sp. (ud-Tetra) and Desoria gracilliseta during the observation period.

Date of sampling Spot 1 Sept. 16 Sept. 24 Sept. 30 Oct. 5 Oct. 9 No. individualsa) Tetracanthura sp. 1 52 47 6 0 D. gracilliseta 0 18 60 91 102 % Tetracanthura sp. 100 74.3 43.9 6.2 0 body length in mmb) 0.49±0.03 (0.44–0.53) 0.48±0.04 (0.4–0.52) 0.47±0.04 (0.42–0.5) Tetracanthura sp. n.d.c) –- n=20 n=20 n=6 0.54±0.05 (0.47–0.66) 0.60±0.09 (0.48–0.76) 0.64±0.1 (0.52–0.94) 0.70±0.1 (0.53–0.93) D. gracilliseta n=18 n=20 n=20 n=20

Date of sampling Spot 2 Sept. 26 Oct. 1 Oct. 6 Oct. 11 Oct. 15 No. individualsa) Tetracanthura sp. 45 15 9 1 0 D. gracilliseta 2 45 140 32 48 % Tetracanthura sp. 95.7 25 6 3 0 body length in mmb) 0.49±0.02 (0.45–0.53) 0.50±0.02 (0.44–0.51) 0.48±0.03 (0.45–0.5) 0.48 Tetracanthura sp. –- n=20 n=15 n=9 n=1 0.58±0.03 (0.56–0.6) 0.60±0.05 (0.54–0.69) 0.67±0.11 (0.54–0.89) 0.71±0.12 (0.56–0.9) 0.80±0.19 (0.53–1.28) D. gracilliseta n=2 n=20 n=20 n=20 n=20 a) Figures indicate total numbers of individuals collected from eight lichen samples (7 × 7 × 1 cm) in 2017. b) Mean ± standard deviation (min.–max.). c) Not determined. 14 Naoyuki Matsumoto, Yasuhiko Suma, and Taizo Nakamori

100 μm

Fig. 1. Four anal spines on abdominal segment VI of Tetracanthura sp. (ud-Tetra) collected on Oc- tober 4, 2016.

thorax segment II of ud-Tetra (Fig. 3) but were absent from nal segment IV was short and ellipsoidal with length/width ra- D. gracilliseta, regardless of body size. Macrochaetae were tios of 1.93 and 2.02 for ud-Tetra and juvenile D. gracilliseta, restricted mostly to abdominalFig. segments1 III to VI in ud-Tetra respectively, and the furca, especially the dens, was relatively (Fig. 2A). In contrast, juvenile D. graciliseta had long, erect short with manubrium/dens ratios 1.5 for the former and 1.64 macrochaetae from thorax segment II to abdominal segment for the latter (Table 3). VI, which were more prominent in the posterior part of the DNA sequences were almost identical among un-Tetra, body (Fig. 2B) as was the case with adults. Macrochaetae and juveniles and subadults of D. gracilliseta for both CO1 were often serrate even in ud-Tetra. A black spot was present and 16S rRNA genes and distinct from those of D. trispinata, on the head of most individuals of juvenile D. gracilliseta but T. sylvatica and S. saoriae (Table 4). Un-Tetra, and juveniles absent in ud-Tetra. Ud-Tetra possessed bidentate mucro (Fig. and subadults of D. gracilliseta constituted a single cluster for 4A) whereas mucro of D. gracilliseta were tridentate and both genes with p-distance values (data not shown). elongated (Fig. 4B). Morphological development between D. gracilliseta and Discussion ud-Tetra was compared among the different stages, includ- Tanaka (1982) noted that, in the genus Tetracanthura, ing subadult individuals of D. gracilliseta collected in March mature had never been found and that individuals of T. (mean body length 1.77 mm). The large subadult D. gracilli- litoralis were all of the same instar. Our findings agreed with seta possessed a long, ellipsoidal antennal segment IV (length/ his results and revealed morphological changes in a species width ratio 3.36) (Fig. 5A) and developed furca with long of Isotomidae. Although we were unable to directly observe dens (manubrium/dens ratio 2.14) (Table 3, Fig. 5B). Among the process of metamorphosis, by employing DNA barcod- ud-Tetra and juvenile D. gracilliseta, antennal segment IV and ing using CO1 and 16S rRNA genes, we demonstrated that the furca were undeveloped (Fig. 2A, B). Additionally, anten- ud-Tetra was a form of juvenile D. gracilliseta. The ud-Tetra Juvenile Desoria gracilliseta with anal spines 15

A

B

500 μm Fig. 2. Tetracanthura sp. (ud-Tetra) (A) and juvenile Desoria gracilliseta (B), collected simulta- neously on October 4, 2016.

Fig. 2

20 μm

Fig. 3. Small spines on the head and thorax segment II of Tetracanthura sp. (ud-Tetra) collected on October 4, 2016.

. Fig. 3 16 Naoyuki Matsumoto, Yasuhiko Suma, and Taizo Nakamori

A A

B

B

10 μm Fig. 4. Bidentate mucro of Teracanthura sp. (ud-Tetra) (A) and tridentate mucro of juvenile Desoria gracilliseta (B) collected simultaneously on October 4, 2016.

Fig. 4 200 μm

Fig. 5. Antennal segments (A) and the furca (B) of a subadult individu- al of Desoria gracilliseta collected on March 20, 2016.

Table 3. Comparisons in the size of antennal segment IV and furcaFig. in Tetracanthura 5 sp. (ud-Tetra) and Desoria gracilliseta at two different growth stages. Antennal segment IV Furca Length (μm) Width (μm) Length/width Manubrium Dens Manubrium/dens Tetracanthura sp. (n=20) 51.9±1.7 26.2±0.7 1.93±0.09 57.2±3.2 88.7±3.5 1.50±0.08 D. gracilliseta juvenile (n=11) 62.2±3.0 30.2±2.6 2.02±0.11 72.2±2.7 121.3±3.9 1.64±0.07 D. gracilliseta subadult (n=20) 190.9±19.3 57.1±4.9 3.36±0.28 204.8±40 436.6±73.8 2.14±0.18

Table 4. Mean p-distance among Tetracanthura sp. (ud-Tetra), Desoria gracilliseta, and related taxaa).

COI gene 16S r RNA gene D. gCOIraciliseta gene Tetracanthura sp.16S r RNAD. gene gracilliseta Tetracanthura sp. Desoria gracilliseta D. graciliseta 0 Tetracanthura sp. 0D. gracilliseta Tetracanthura0.001 sp. 0.001 DesoriaDesoria gracilliseta trispinata 0 0.164 0 0.164 0.001 0.2110.001 0.211 Desoria trispinata 0.164 0.164 0.211 0.211 TetracanthellaTetracanthella sylvatica sylvatica 0.179 0.179 0.179 0.179 0.197 0.1970.197 0.197 SahacanthellaSahacanthella saoriae saoriae 0.160 0.160 0.160 0.160 0.178 0.1780.178 0.178 aa)) Two Two sam samplesp eachles each were comwerepared. compared. Juvenile Desoria gracilliseta with anal spines 17 form exhibited four anal spines and numerous short spines on to active escape with developed antennae, furca, and mac- the head and thorax segment II which were not present in ad- rochaetae from predation in advanced instars. Species with vanced instars. Other alterations from spined juveniles includ- a large furca tend to make single, but long jumps, to escape ed the elongation of antennal segments and the development from predators, and the antennae are endowed with sensory of a third tooth on the mucro. Macrochaetae were fewer on the structures, particularly at the tip (Hopkin, 1997). Morphologi- juvenile than on advanced instars as described by Chiritiansen cal changes in D. gracilliseta from the juvenile to the adult (1992) and restricted to the posterior part of the body in the involved the development of furca and sensory organs such juvenile. Since body lengths of the ud-Tetra form were short as antennae and macrochaetae, which should facilitate the and remained the same, the metamorphosis occurred synchro- detection of predators and long jumps. Tridentate mucro is nously at a specific growth stage, possibly the first instar. considered to catch the ground more firmly to facilitate long Morphological changes commonly occur especially in jumps. Juveniles of D. gracilliseta with undeveloped furca Isotomidae and Hypogastruridae, but the type of metamorpho- may use anal spines and short spines on the head and thorax sis we observed on D. gracilliseta seems incongruous with segment II for defence. Although we were unable to reveal the any of the phenomena previously reported. While cyclomor- role of the spines experimentally, it seems more cost-effective phosis accompanies the change in the furca (Leinaas, 1981a) for the juvenile with a limited amount of energy reserve to and is not associated with specific instar (Leinaas, 1981b), D. defend passively with spines than to escape actively from pre- gracilliseta had anal spines in the juvenile (possibly the first dation, using the furca. Further studies are necessary to reveal instar) and failed to show any changes in the furca except the the roles of anal spine and morphological changes in Desoria dens. While epitoky occurs during sexual maturity (Fjellberg, gracilliseta. 1977), the loss of anal spines occurred in the juvenile in the case of D. gracilliseta. Ecomorphosis includes the appearance Acknowledgements of anal spines in response to environmental changes (Hart, Thanks are due to Chika Sumida (Yokohama National 1978); however in D. gracilliseta, annal spines were lost dur- University) and Takato Nakayama (Hokkaido National Ag- ing growth. ricultural Research Center) for technical assistance in DNA Many collembolans develop anal spines at different barcoding experiments and in microphotoscopy, respectively. growth stages (e.g., Suma et al. 2003, Suma and Watanabe, We also wish to thank Denis A. Gaudet (formerly Agriculture 2005), but their roles may vary among species. The ud-Tetra Canada) for the correction of English and anonymous review- form of juvenile D. gracilliseta possessed anal spines, which ers for their invaluable comments. The study was partly sup- was consistent with the observations by Suma and Yamauchi ported for Taizo Nakamori by JSPS under the Japan–Russia (1999) who described a pair of anal spines present exclusively Research Cooperative Program. on small individuals of Lophognathella choreutes Böner, 1908. On the contrary, in the genus Dimorphacanthella, small 摘 要 juveniles with two anal spines become“ Tetracanthella” when 松本直幸 1・須摩靖彦 2・中森泰三 3（1 〒 066-0052 北海道千 they develop the second pair of anal spines (Potapov et al., 歳市文京 6-22-2・2 〒 085-0813 北海道釧路市春採 6-7-32・3 横 2010). In Xenylla uniseta Da Gama, 1963, anal spines are 浜国立大学環境情報研究院 〒 240-8501 神奈川県横浜市保土ヶ undeveloped and inconspicuous in the first instar whereas 谷区常盤台 79-7）： 4 本の尾角をもつホソゲツチトビムシの幼 the adults possess anal spines (Winkler et al., 2011). These 個体． authors did not refer to the function of anal spines; however, 北海道において，地衣類タカネゴケに生息するホソゲツチ since they were found at different growth stages, anal spines トビムシは年 1 化性で，10 月に幼個体が出現し，積雪下にお are considered to have different roles. The most powerful idea いても成長を続け，5 月に成熟した後は，消失する．今回 9 月 is that anal spines are used for defence from predation as in 中旬から 10 月中旬において，さらに詳しく調査したところ， the case of neck teeth or toothed crest in the water flea, Daph- 小型のホソゲツチトビムシに加え，4 本の尾角を有する Tetra- nia pulex Leydig, 1860 (Havel and Dodson, 1984), canthura 様のトビムシが得られた．Tetracanthura 様トビムシ The morphological changes in D. gracilliseta observed in の体長は約 0.49 mm と調査期間を通じて一定であったのに対 the present study suggest a shift in its survival strategy from し，ホソゲツチトビムシは成長を続け，最初 0.54 mm であっ passive defense with spines against predation in the juvenile たのが調査終了時には 0.8 mm となっていた．Tetracanthura 18 Naoyuki Matsumoto, Yasuhiko Suma, and Taizo Nakamori

様トビムシには尾角と頭部および胸部第 2 節に小さなトゲが evolutionary genetics analysis version 7.0 for bigger datasets. あることを除けば，両者の形態は酷似していた．さらに 3 月 Molecular Biology and Evolution, 33: 1870–1874. に採取したより大型のホソゲツチトビムシを加え，形態的特 Leinaas, H. P., 1981a. Cyclomorphosis in the furca of the winter 徴と DNA バーコーディングによる比較を行った．三者の遺伝 active Collembola Hypogastrura socialis (Uzel). Entomo- 的同一性が確認され，触角第４節と跳躍器は成長と共に発達 logica Scandinavica, 12: 35–38. した．ホソゲツチトビムシにおける形態変化を捕食に対する Leinaas, H. P., 1981b. Cyclomorphosis in Hypogastrura lap- 適応戦略の面から考察した． ponica (Axelson, 1902) (=H. frigida [Axelson, 1905] syn. キーワード： 尾角，Desoria gracilliseta，DNA バーコーディ nov.) (Collembola, Poduridae). Morphological adaptations ング，形態変化，Tetracanthura sp. and selection for winter dispersal. Zeitschrift für Zoologische Systematik und Evolutionsforschung, 19: 278–285. References Matsumoto, N., Suma, Y. and Koike, K., 2018. Reproduction and Aoyama, H., Saitoh, S., Fujii, S., Nagahama, H., Shinzato, N., growth of Collembola under snow in a cold temperate region. Kaneko, N. and Nakamori, T., 2015. A rapid method of non- Edaphologia, 102: 11–21. desctructive DNA extraction from individual . Ap- Najt, J., 1983. Modifications morphologiques liées à l’écomor- plied Entomology and Zoology, 50: 419–425. phose chez les Collemboles Isotomidae. Pedobiologia, 25: Castresana, J., 2000. Selection of conserved blocks from multiple 337–348. alignments for their use in phylogenetic analysis. Molecular Potapov, M., 2001. Synopses on Palaearctic Collembola. Volume Biology and Evolution, 17: 540–552. 3. Isotomidae. Abhandlungen und Berichte des Naturkunde- Christiansen, K., 1992. Springtails. The Kansas School Naturalist, museums, Görlitz, 73: 1–603. 39: 1–16. Potapov, M. B., Bu, Y., Huang, C. W., Gao, Y. and Luan, X. Y., Edgar, R. C., 2004. MUSCLE: Multiple sequence alignment with 2010. Genetic switch-over during ontogenesis in Dimorpha- high accuracy and high throughput. Nucleic Acids Research, canthella gen. n. (Collembola, Isotomidae) with barcoding 32: 1792–1797. evidence. ZooKeys, 73: 13–23. Fjellberg, A., 1976. Cyclomorphosis in Isotoma hiemalis Schött, Potapov, M., Nakamori, T., Saitoh, S., Kuznetsova, N., and Ba- 1983 (mucronata Axelson, 1900) syn. nov. (Collembola, benko, A., 2017. New or little-known taxa of Anurophorinae Isotomidae). Revue d’Ecologie et de Biologie du Sol, 13: (Collembola) with anal spines from East Asia with notes on 381–384. DNA barcode. Zootaxa, 4318: 312–324. Fjellberg, A., 1977. Epitoky in species (Collembola, Sawahata, T., 2005. Morphological change of winter-active Col- Isotomidae). Revue d’Ecologie et de Biologie du Sol, 14: lembola Hypogastrura bokusi Yosii. Edaphologia, 78: 11–13. 493–495. Suma, Y. and Yamauchi, S., 1999. On the larval morphology of Greenslade, P., and Popapov, M., 2015. Biology, affinity and de- Lophognathella horeutes Böner, especially anal spine of ab- scription of an unusual aquatic new genus and species of Iso- dominal segment VI. Sylvicola, 17: 49–56. (In Japanese) tomidae (Collembola) from high altitude lakes in Tasmania. Suma, Y., Yamauchi, S., and Nodasaka, Y., 2003. Some records on European Journal of Entomology 112: 334–343. the Collembola from the Osorezan mountain range, Aomori Hågvar, S., 1995. Long distance, directional migration on snow Pref., Japan, especially ecomorphosis of Granisotoma reinieri in a forest collembolan, Hypogastrura socialis (Uzel). Acta (Folsom). Journal of the Natural History of Aomori, 8: 19–24. Zoologica Finnica, 196: 200–205. (In Japanese) Hart, J. W., 1978. Ecomorphisms in Proisotoma vesiculata Fol- Suma, Y. and Watanabe, T., 2005. Collembolan fauna of Sapporo I. som. Proceedings of the Indiana Academy of Science, 88: jezoensis 31: 81–88. (In Japanese) 191–193. Tanaka, S., 1982. Two new species of the genus Tetracanthura Havel, J. E. and Dodson, S. I., 1984. Chaoborus predation on typi- Martynova (Collembola: Isotomidae) from Japan. Edapholo- cal and spined morphs of Daphnia pulex: Behavioral obser- gia, 25/26: 21–32. vations. Limnology and Oceanography, 29: 487–494. Winkler, D., Korda, M. and Traser, Gy., 2011. Two new species of Hopkin, S. P., 1997. Biology of the (Insecta: Collem- Collembola new for the fauna of Hungary. Opuscula Zoologica bola). Oxford University Press, Oxford. Budapest, 42: 199–206. Kumar, S., Stecher, G. and Tamura, K., 2016. MEGA7: Molecular