Research Article |
Corresponding author: Sho Tsukamoto ( esutukamoto153@gmail.com ) Academic editor: Pavel Stoev
© 2024 Sho Tsukamoto, Katsuyuki Eguchi.
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:
Tsukamoto S, Eguchi K (2024) Integrative taxonomy of Dicellophilus Cook, 1896 (Chilopoda, Geophilomorpha, Mecistocephalidae) in Japan, with a description of a new species. Zoosystematics and Evolution 100(3): 821-840. https://doi.org/10.3897/zse.100.121512
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The genus Dicellophilus Cook, 1896, is a peculiar genus from the point of view of distribution. Dicellophilus is distributed in three limited areas that are well separated from one another: central Europe (D. carniolensis), Honshu (D. pulcher), and the southwestern part of the USA (D. anomalus and D. limatus). In the present study, in a field survey conducted throughout Japan, specimens belonging to the genus Dicellophilus were collected from Tohoku to the Kansai region, Honshu. Morphological analysis, molecular phylogenetic analysis, and genetic distance among Dicellophilus in Japan and D. carniolensis revealed that specimens from Sendai-shi, Miyagi Pref., could be assigned to an undescribed species. This previously unrecognized species is herein described as D. praetermissus sp. nov. The new species can be distinguished from D. carniolensis and D. limatus by the number of pairs of legs (43 pairs in D. carniolensis and 45 in D. limatus, but 41 in D. praetermissus sp. nov.), from D. anomalus by the lack of a pair of setae on the posteromedian part of the clypeus and variable crenulation on the internal margin of the forcipular tarsungulum, and from D. pulcher based on the following combination of characteristics: both ends of the transverse suture not evidently convex forward; long rather than wide trochanteroprefemur; wide rather than long metasternite.
Cryptic species, DNA barcoding, geophilomorph centipede, molecular, phylogeny
The geophilomorph family Mecistocephalidae Bollman, 1893, is mainly distributed from temperate to tropical regions in both hemispheres, and species diversity is remarkably high in Japan (
Mecistocephalidae are morphologically characterized by a cephalic capsule and a forcipular segment that are evidently sclerotized and darker than the remaining trunk segments (
Among Mecistocephalidae, the genus Dicellophilus Cook, 1896, is a distinct genus from a morphological viewpoint, with the following diagnostic characteristics: a macropore near the center of the coxopleuron and a concave margin of the lateral side pieces of the labrum (
The distribution of Dicellophilus is limited to three areas that are well separated from one another: central Europe (D. carniolensis), Honshu of Japan (D. pulcher), and the southwestern part of the USA (D. anomalus and D. limatus) (
For a decade, the combination of morphological observation, molecular phylogenetic analyses, and DNA barcoding (“integrative taxonomy”) has helped detect undescribed species and reveal the genetic structure of the taxa concerned (
Inspired by these previous studies, the present study aims to reveal the genetic diversity and confirm whether the morphospecies D. pulcher involves unnoticed and undescribed cryptic species by using an integrative taxonomic approach.
Although our ongoing sampling efforts to taxonomically reveal the East Asian mecistocephalid faunas cover the whole of Japan and surrounding areas, the present study focused on Honshu (the largest island of mainland Japan), from which D. pulcher was described.
A total of 38 specimens morphologically identified as Dicellophilus pulcher, hitherto the only known Japanese species of the genus, were collected from Honshu from 2018 to 2021. The detailed collection sites of the examined specimens are shown in Fig.
The list of specimens that were used in the phylogenetic analyses. Geographic coordinates enclosed by parentheses are secondary due to the lack of information in the labels.
Specimen ID | Geographic coordinate of collection sites | Sequence data (Accession no.) | Remarks | ||
---|---|---|---|---|---|
COI | 16S | 28S | |||
Dicellophilus pulcher (Kishida, 1928) | |||||
TS20220518-01 | 35°16.45'N, 137°00.59'E (Aichi Pref.) | LC815129 | LC815167 | LC815205 | Tomoki Sumino leg. |
TS20210809-01 | 34°33.71'N, 136°27.33'E (Mie Pref.) | LC815130 | LC815168 | LC815206 | Tomoki Sumino & Fukube Sumino leg. |
TS20210809-02 | 34°33.71'N, 136°27.33'E (Mie Pref.) | LC815131 | LC815169 | LC815207 | Tomoki Sumino & Fukube Sumino leg. |
TS20220813-01 | 34°33.42'N, 136°24.13'E (Mie Pref.) | LC815132 | LC815170 | LC815208 | Tomoki Sumino leg. |
TS20230517-01 | 35°16.38'N, 137°00.96'E (Aichi Pref.) | LC815133 | LC815171 | LC815209 | Tomoki Sumino leg. |
TS20190413-01 | 35°25.15'N, 139°10.39'E (Kanagawa Pref.) | LC815134 | LC815172 | LC815210 | Sho Tsukamoto leg. |
TS20181214-01 | 35°37.02'N, 139°22.74'E (Tokyo Pref.) | LC815135 | LC815173 | – | Joe Kutsukake leg. |
TS20180714-01 | 35°27.46'N, 139°24.51'E (Kanagawa Pref.) | LC815136 | LC815174 | – | Joe Kutsukake leg. |
TS20191006-02 | 35°38.15'N, 139°15.71'E (Tokyo Pref.) | LC815137 | LC815175 | – | Sho Tsukamoto leg. |
TS20200924-01 | 35°52.42'N, 138°25.96'E (Yamanashi Pref.) | LC815138 | – | – | Joe Kutsukake leg. |
TS20200927-01 | 35°54.95'N, 138°20.55'E (Yamanashi Pref.) | LC815139 | LC815176 | – | Koshi Kawamoto leg. |
TS20201122-01 | (35°25.62'N, 135°23.69'E) (Kyoto Pref.) | LC815140 | LC815177 | LC815211 | Tatsumi Suguro leg. |
TS20210322-01 | 35°15.40'N, 139°44.43'E (Kanagawa Pref.) | LC815141 | LC815178 | – | Ryo Miyata leg. |
TS20210401-03 | 35°44.46'N, 139°31.01'E (Tokyo Pref.) | LC815142 | LC815179 | LC815212 | Mayu Susukida leg. |
TS20210411-08 | 35°26.84'N, 139°02.82'E (Kanagawa Pref.) | LC815143 | LC815180 | LC815213 | Namiki Kikuchi leg. |
TS20210424-07 | 35°25.88'N, 139°14.15'E (Kanagawa Pref.) | LC815144 | LC815181 | LC815214 | Katsuyuki Eguchi leg. |
TS20210504-01 | 35°21.91'N, 138°48.44'E (Shizuoka Pref.) | LC815145 | LC815182 | LC815215 | A topotype of D. pulcher; Sho Tsukamoto leg. |
TS20210418-01 | 34°34.94'N, 137°02.13'E (Aichi Pref.) | LC815146 | LC815183 | LC815216 | Katsuyuki Eguchi leg. |
TS20210522-01 | 35°44.78'N, 138°53.15'E (Yamanashi Pref.) | LC815147 | LC815184 | LC815217 | Takahiro Yoshida leg. |
TS20210530-02 | 35°41.19'N, 138°52.64'E (Yamanashi Pref.) | LC815148 | LC815185 | LC815218 | Namiki Kikuchi leg. |
TS20210722-01 | 35°57.71'N, 139°03.60'E (Saitama Pref.) | LC815149 | LC815186 | LC815219 | Mayu Susukida leg. |
TS20211004-01 | (36°08.54'N, 138°16.92'E) (Nagano Pref.) | LC815150 | LC815187 | LC815220 | Masaru Nonaka leg. |
TS20211030-04 | 36°51.75'N, 138°46.43'E (Niigata Pref.) | LC815151 | LC815188 | LC815221 | Katsuyuki Eguchi leg. |
TS20211018-02 | 35°26.99'N, 136°49.95'E (Gifu Pref.) | LC815152 | LC815189 | LC815222 | Katsuyuki Eguchi leg. |
TS20210919-01 | 34°46.18'N, 135°59.73'E (Kyoto Pref.) | LC815153 | LC815190 | LC815223 | Katsuyuki Eguchi leg. |
TS20210711-15 | 35°17.08'N, 136°32.74'E (Gifu Pref.) | LC815154 | LC815191 | LC815224 | Katsuyuki Eguchi leg. |
TS20210819-01 | 35°36.51'N, 139°23.48'E (Tokyo Pref.) | LC815155 | LC815192 | LC815225 | Katsuyuki Eguchi leg. |
TS20210523-01 | 35°27.82'N, 138°29.66'E (Yamanashi Pref.) | LC815156 | LC815193 | LC815226 | Katsuyuki Eguchi leg. |
TS20210531-01 | 37°05.59'N, 140°31.72'E (Fukushima Pref.) | LC815157 | LC815194 | LC815227 | Katsuyuki Eguchi leg. |
TS20210523-03 | 35°11.94'N, 138°31.29'E (Shizuoka Pref.) | - | LC815195 | LC815228 | Katsuyuki Eguchi leg. |
TS20210718-01 | 35°56.36'N, 139°15.30'E (Saitama Pref.) | LC815158 | LC815196 | LC815229 | Katsuyuki Eguchi leg. |
TS20210508-02 | 35°19.27'N, 139°18.72'E (Kanagawa Pref.) | LC815159 | LC815197 | LC815230 | Katsuyuki Eguchi leg. |
TS20210531-02 | 37°05.59'N, 140°31.72'E (Fukushima Pref.) | LC815160 | LC815198 | LC815231 | Katsuyuki Eguchi leg. |
TS20210728-01 | 35°14.75'N, 139°01.02'E (Kanagawa Pref.) | LC815161 | LC815199 | LC815232 | Sho Tsukamoto leg. |
TS20210728-02 | 35°13.32'N, 139°03.13'E (Kanagawa Pref.) | LC815162 | LC815200 | LC815233 | Sho Tsukamoto leg. |
Dicellophilus praetermissus sp. nov. | |||||
TS20201007-02 | 38°16.33'N, 140°32.69'E (Miyagi Pref.) | LC815125 | LC815163 | LC815201 | Sho Tsukamoto leg. |
TS20201007-03 | 38°16.33'N, 140°32.69'E (Miyagi Pref.) | LC815126 | LC815164 | LC815202 | Sho Tsukamoto leg. |
TS20201007-04 | 38°16.33'N, 140°32.69'E (Miyagi Pref.) | LC815127 | LC815165 | LC815203 | Sho Tsukamoto leg. |
TS20201007-05 | 38°16.33'N, 140°32.69'E (Miyagi Pref.) | LC815128 | LC815166 | LC815204 | Sho Tsukamoto leg. |
Dicellophilus carniolensis (Koch, 1847) (outgroup) | |||||
DNA102580/LBv792 | No data | KF569305 | HM453225 | HM453285 | Referred from |
Nannarrup innuptus Tsukamoto in |
|||||
TS20210503-09 | 34°51.39'N, 138°55.40'E (Shizuoka Pref.) | LC715530 | LC715605 | LC715680 | Referred from |
Each specimen was labeled with its unique specimen identification number in the form “TSYYYYMMDD-XX,” where TS is an abbreviation of the first author’s name, Tsukamoto Sho; YYYYMMDD designates the date on which the specimen was collected; and XX is the identification number assigned to each specimen collected on a particular date (e.g., TS20171010-01).
All of the type specimens of Dicellophilus designated in this paper were deposited at the Collection of Myriapoda, Department of Zoology, National Museum of Nature and Science, Tokyo (NSMT), and the Museum of Nature and Human Activities, Hyogo (MNHAH). The deposition site of each type specimen is shown in the “Taxonomic account” section. All non-type voucher specimens of Dicellophilus pulcher are managed by the first author.
For dissected specimens, the cephalic capsule, maxillae, mandibles, forcipular segment, and leg-bearing segments were made transparent using lactic acid to examine the anatomy and produce images. Multi-focused images of these body parts were produced using Affinity Photo 1.10.4 (https://affinity.serif.com/ja-jp/photo/) from a series of source images taken using a Canon EOS Kiss X9 digital camera attached to a Nikon AZ100 microscope and improved using Adobe Photoshop Elements 10 and Affinity Designer 1.10.5 (https://affinity.serif.com/ja-jp/designer/). Then, the body parts were measured directly using an ocular micrometer attached to the microscope.
The morphological terminology used in this study is in accordance with
Genomic DNA was extracted from one or two legs of each specimen in accordance with the Chelex-TE-ProK protocol described by
PCR amplification was performed in a MiniAmp Thermal Cycler (Thermo Fisher Scientific, Waltham, Massachusetts, USA) in a 10.5-μL reaction volume containing 5 μL of 2× PCR buffer for KOD FX Neo, 2 μL of 2 mM dNTPs, 0.3 μL of 10 pmol/μL forward and reverse primers, 0.2 μL of 1.0 U/μL DNA polymerase KOD FX Neo (TOYOBO KFX-201X5), and 1.0 μL of DNA template. The sequences of primers for mitochondrial cytochrome c oxidase subunit I (COI), 16S rRNA (16S), and nuclear 28S rRNA (28S) genes are shown in Table
Genes | Primer name | Sequence (5’ - 3’) | Source |
---|---|---|---|
COI | LCO-CH | TTT CAA CAA AYC AYA AAG ACA TYG G |
|
HCO-CH | TAA ACT TCT GGR TGR CCR AAR AAT CA | ||
16S rRNA | 16Sa | CGC CTG TTT ATC AAA AAC AT |
|
16Sbi | CTC CGG TTT GAA CTC AGA TCA | ||
28S rRNA | 28S D1F | GGG ACT ACC CCC TGA ATT TAA GCA T |
|
28S rD4b | CCT TGG TCC GTG TTT CAA GAC |
|
The amplification conditions for mitochondrial COI were as follows: 98 °C for 2 min; 5 cycles of 98 °C for 10 s; 45 °C for 30 s; and 68 °C for 45 s; 40 cycles of 98 °C for 10 s; 48.5 °C for 30 s (annealing step); 68 °C for 45 s; and 68 °C for 7 min. If the target fragment of COI was not appropriately amplified, then the annealing temperature was changed from 48.5 °C to 50 °C. PCR was performed again by omitting the first five cycles of annealing and the extension step.
The amplification conditions for mitochondrial 16S were as follows: 98 °C for 2 min; 35 cycles of 98 °C for 10 s; 45 °C for 30 s (annealing step); 68 °C for 45 s; and 68 °C for 7 min. If the target fragment of 16S was not appropriately amplified, then the annealing temperature was changed from 45 °C to 48 °C. The number of annealing cycles was changed from 35 to 45.
The amplification conditions for nuclear 28S were as follows: 98 °C for 2 min; 5 cycles of 98 °C for 10 s; 42 °C for 30 s; and 68 °C for 1 min; 30 cycles of 98 °C for 10 s; 50 °C for 30 s (annealing step); 68 °C for 1 min; and 68 °C for 7 min. If the target fragment of 28S was not appropriately amplified, then the annealing temperature was changed from 50 °C to 48 °C. The number of annealing cycles was changed from 30 to 40–45 cycles. In addition, PCR was performed again by omitting the first five cycles of annealing and the extension step.
The amplified products were incubated at 37 °C for 4 min and at 80 °C for 1 min using ExoSAP-ITTM Express (Thermo Fisher Scientific) to remove any excess primers and nucleotides. All nucleotide sequences were determined by direct sequencing using the ABI PRISM BigDyeTM Terminator Cycle Sequencing Kit ver. 3.1 (Thermo Fisher Scientific) or BrilliantDyeTM Terminator Cycle Sequencing Kit v. 3.1 (Nimagen, B.V., Nijmegen, Netherlands) equipped with an ABI 3130xl automated sequencer (Thermo Fisher Scientific). The sequences were assembled using ChromasPro 1.7.6 (Technelysium Pty Ltd., Australia) and deposited onto the DDBJ, EMBL, and GenBank databases under the accession numbers LC815125– LC815233 (Table
The sequences obtained using the abovementioned methods were used for phylogenetic analyses, together with the COI, 16S, and 28S sequences of Dicellophilus carniolensis (C.L. Koch, 1847) and Nannarrup innuptus Tsukamoto in
All sequences were aligned using MAFFT v. 7.475 (
Maximum-likelihood (ML) trees were created on the basis of the sequence dataset for each gene and concatenated using IQ-tree 1.6.12 (
Bayesian inference trees were created using ExaBayes 1.4.1 (
The aligned COI dataset used for phylogenetic analyses were also used to calculate genetic distances. Kimura two-parameter (K2P) distances were calculated using MEGA X (
The program “assemble species by automatic partitioning (ASAP)” was used to delimit “provisional” operational taxonomic units (POTUs). ASAP is a species delimitation program based on a hierarchical clustering algorithm that only uses pairwise genetic distances (
The present study preliminarily relied on the morphological information provided by
As mentioned above, the present study presupposes the monophyly of Dicellophilus, supported by the morphological evidence (
Each putative species recognized by following the abovementioned steps was also labeled in accordance with the study of
Depending on the availability of the morphological information necessary to formally describe and name species in the conventional manner of Linnaean Taxonomy (
COI was successfully sequenced for 38 specimens, 16S for 38 specimens, and 28S for 33 specimens (Table
Maximum-likelihood tree of Dicellophilus based on the concatenated dataset of COI, 16S, and 28S, with the results of species delimitation by ASAP. Note that specimens whose COI sequence was not determined were included in the conferred species if they belonged to the same concerning clade to easily understand the result of species delimitation. Nodal values are obtained from the ultrafast bootstrap (UFBoot), SH-like approximate likelihood ratio test (SH-aLRT), and posterior probability (PP). The asterisk (*) indicates 100% in UFBoot, SH-aLRT, and 1.0 in PP. Hyphen (-) indicates lower than 95% in UFBoot, 80% in SH-aLRT, or 0.95 in PP. Nodal values are not shown when UFBoot, SH-aLRT, and PP values are <95%, <80%, and <0.95, respectively. The unit of evolutionary distance is the number of base substitutions per site. A broken square shows that the clade consisted of specimens from eastern Honshu. Abbreviations: Ai = Aichi Pref.; Fs = Fukushima Pref.; Gi = Gifu Pref.; Kn = Kanagawa Pref.; Ky = Kyoto Pref.; Mi = Mie Pref.; My = Miyagi Pref.; Ni = Niigata Pref.; Nn = Nagano Pref.; Sh = Shizuoka Pref.; Si = Saitama Pref.; To = Tokyo Pref.; Yn = Yamanashi Pref.
The ML tree based on the COI dataset (Fig.
Maximum-likelihood tree of Dicellophilus based on the dataset of COI, with the results of species delimitation by ASAP. Nodal values are obtained from the ultrafast bootstrap (UFBoot), SH-like approximate likelihood ratio test (SH-aLRT), and posterior probability (PP). The asterisk (*) indicates 100% in UFBoot, SH-aLRT, and 1.0 in PP. Hyphen (-) indicates lower than 95% in UFBoot, 80% in SH-aLRT, or 0.95 in PP. Nodal values are not shown when UFBoot, SH-aLRT, and PP values are <95%, <80%, and <0.95, respectively. The unit of evolutionary distance is the number of base substitutions per site. A broken square shows that the clade consisted of specimens from eastern Honshu. Abbreviations: Ai = Aichi Pref.; Fs = Fukushima Pref.; Gi = Gifu Pref.; Kn = Kanagawa Pref.; Ky = Kyoto Pref.; Mi = Mie Pref.; My = Miyagi Pref.; Ni = Niigata Pref.; Nn = Nagano Pref.; Sh = Shizuoka Pref.; Si = Saitama Pref.; To = Tokyo Pref.; Yn = Yamanashi Pref.
The ML tree based on the 16S dataset (Fig.
Maximum-likelihood tree of Dicellophilus based on the dataset of 16S. Nodal values are obtained from the ultrafast bootstrap (UFBoot), SH-like approximate likelihood ratio test (SH-aLRT), and posterior probability (PP). The asterisk (*) indicates 100% in UFBoot, SH-aLRT, and 1.0 in PP. Hyphen (-) indicates lower than 95% in UFBoot, 80% in SH-aLRT, or 0.95 in PP. Nodal values are not shown when UFBoot, SH-aLRT, and PP values are <95%, <80%, and <0.95, respectively. The unit of evolutionary distance is the number of base substitutions per site. A broken square shows that the clade consisted of specimens from eastern Honshu. Abbreviations: Ai = Aichi Pref.; Fs = Fukushima Pref.; Gi = Gifu Pref.; Kn = Kanagawa Pref.; Ky = Kyoto Pref.; Mi = Mie Pref.; My = Miyagi Pref.; Ni = Niigata Pref.; Nn = Nagano Pref.; Sh = Shizuoka Pref.; Si = Saitama Pref.; To = Tokyo Pref.; Yn = Yamanashi Pref.
The ML tree based on the 28S dataset (Fig.
Maximum-likelihood tree of Dicellophilus based on the dataset of 28S. Nodal values are obtained from the ultrafast bootstrap (UFBoot), SH-like approximate likelihood ratio test (SH-aLRT), and posterior probability (PP). The asterisk (*) indicates 100% in UFBoot, SH-aLRT, and 1.0 in PP. Hyphen (-) indicates lower than 95% in UFBoot, 80% in SH-aLRT, or 0.95 in PP. Nodal values are not shown when UFBoot, SH-aLRT, and PP values are <95%, <80%, and <0.95, respectively. The unit of evolutionary distance is the number of base substitutions per site. A broken square shows that the clade consisted of specimens from eastern Honshu. Abbreviations: Ai = Aichi Pref.; Fs = Fukushima Pref.; Gi = Gifu Pref.; Kn = Kanagawa Pref.; Ky = Kyoto Pref.; Mi = Mie Pref.; My = Miyagi Pref.; Ni = Niigata Pref.; Nn = Nagano Pref.; Sh = Shizuoka Pref.; Si = Saitama Pref.; To = Tokyo Pref.; Yn = Yamanashi Pref.
Although there is no consistency of phylogenetic relationship among Clades A, B, and D. carniolensis in four datasets, each topology shows that Clade A is a distinct lineage from other Dicellophilus specimens.
The minimum K2P distance between congeneric morphospecies was 21% (D. carniolensis (accession no.: KF569305) vs. D. pulcher TS20230517-01 from Aichi Pref.). Thus, the intermorphospecific threshold induced by this dataset is 21%. However, the maximum K2P distance was 24% within D. pulcher (TS20191006-02 from Tokyo Pref. in Clade B vs. TS20201007-04 from Miyagi Pref. in Clade A).
The maximum K2P distance within Clade A was 0.6% (TS20201007-04 vs. TS20201007-02 and TS20201007-03), and that within Clade B was 15% (TS20210322-01 from Kanagawa Pref. vs. TS20210523-01 from Yamanashi Pref.).
The best five partitioning hypotheses inferred by the ASAP program are shown in Figs
All 38 specimens of D. pulcher (a combination of Clades A and B) examined in steps I–IV have 41 pairs of legs and can be distinguished from D. carniolensis and D. limatus by the number of pairs of legs (43 pairs in D. carniolensis and 45 in D. limatus). Examined 23 adult specimens can also be distinguished from D. anomalus, which has 41 pairs of legs, by the lack of a pair of setae on the posteromedian part of the clypeus and variable crenulation on the internal margin of the forcipular tarsungulum (
In addition, the 23 adult specimens, including a specimen from the type locality of D. pulcher (Subashiri, Oyama, Suntou-gun, Shizuoka Pref.), had the following diagnostic characteristics of D. pulcher (
On the other hand, four specimens of Clade A (TS20201007-02, TS20201007-03, TS20201007-04, and TS20201007-05) were morphologically different from the specimens of Clade B based on the following characteristics: both ends of transverse suture not evidently convex forward; long rather than wide trochanteroprefemur; wide rather than long metasternite (Table
Morphological comparison between D. pulcher and D. praetermissus sp. nov.
Putative species identification code | Species identified in the present study | Both ends of transverse suture | The width to length ratio of trochanteroprefemur |
The width to length ratio of sternite of ULBS |
---|---|---|---|---|
Dicellophilus sp. 0000-0003-3020-8454_0068 | Dicellophilus pulcher (Kishida, 1928) | evidently convex forward | 1: 0.9–1.1 | 1: 1.0–1.3 |
Dicellophilus sp. 0000-0003-3020-8454_0069 | Dicellophilus praetermissus sp. nov. | not evidently convex | 1: 1.3–1.4 | 1: 0.66–1.0 |
Family Mecistocephalidae Bollman, 1893
Genus Dicellophilus Cook, 1896
Dicellophilus sp. 0000-0003-3020-8454_0069
Holotype. 1 adult male, Baba, Akiu-machi, Taihaku-ku, Sendai-shi, Miyagi Pref., Japan (38°16.33'N, 140°32.69'E), 7 October 2020, coll. Sho Tsukamoto (labeled as TS20201007-02), deposited at the Collection of Myriapoda, Department of Zoology, NSMT.
Paratype. 1 adult male, Baba, Akiu-machi, Taihaku-ku, Sendai-shi, Miyagi Pref., Japan (38°16.33'N, 140°32.69'E), 7 October 2020, coll. Sho Tsukamoto (labeled as TS20201007-03), 1 adult female, Baba, Akiu-machi, Taihaku-ku, Sendai-shi, Miyagi Pref., Japan (38°16.33'N, 140°32.69'E), 7 October 2020, coll. Sho Tsukamoto (labeled as TS20201007-04), 1 adult male, Baba, Akiu-machi, Taihaku-ku, Sendai-shi, Miyagi Pref., Japan (38°16.33'N, 140°32.69'E), 7 October 2020, coll. Sho Tsukamoto (labeled as TS20201007-05), deposited at MNHAH.
The species name is a masculine adjective derived from “overlooked” in Latin. Since the description by
Trunk segments without dark patches; head 1.4 times as long as wide; lateral margin of cephalic plate abruptly converged posteriorly; clypeus with densely scattered setae; palaclypeal suture evidently converging posteriorly; both ends of transverse suture uniformly rounded; mandible with 6 lamellae; forcipular trochanteroprefemur longer than wide, with one small distal denticle; forcipular tarsungulum with evident and variably spaced notches; metatergite subtrapezoidal; metasternite trapezoidal, wide rather than long; forty-one pairs of legs.
General features
(Fig.
Cephalic capsule
(Fig.
Antenna
(Fig.
Dicellophilus praetermissus sp. nov. A–F. Holotype (TS20201007-02) G, H. Paratype (TS20201007-04) A. Antennal articles I–IV, dorsal; B. Antennal articles I–IV, ventral; C. Antennal articles V–VIII, dorsal; D. Antennal articles V–III, ventral; E. Antennal articles IX–XIV, dorsal; F. Antennal articles IX–XIV, ventral; G. Antennal article XIV, dorsal; H. Antennal article XIV, ventral. Arrows indicate apical sensillum; arrowheads indicate club-like sensillum. Scale bars: 0.5 mm (A–F); 0.1 mm (G, H).
Mandible
(Fig.
First maxillae
(Fig.
Second maxillae
(Fig.
Forcipular segment
(Fig.
Dicellophilus praetermissus sp. nov., holotype (TS20201007-02). A. Forcipular segment, dorsal; B. Forcipular segment, ventral; C. Right condylar process of forcipular coxosternite, dorsal; D. Right forcipular tarsungulum, dorsal; E. Poison calyx, dorsal. The arrow indicates the basal tubercle of the forcipular tarsungulum. Scale bars: 0.5 mm (A, B); 0.2 mm (C); 0.3 mm (D); 0.1 mm (E).
Leg-bearing segments
(Fig.
Dicellophilus praetermissus sp. nov., holotype (TS20201007-02). A. Tergite of leg-bearing segment 40, dorsal; B. Sternite of leg-bearing segment 40, ventral; C. Pretarsus of left leg 40, anterolateral. D. Pretarsus of left leg 2, posterolateral. The arrow indicates a subsidiary spine. Scale bars: 0.5 mm (A, B); 0.1 mm (C, D).
Ultimate leg-bearing segment
(Figs
Male postpedal segments
(Fig.
Female postpedal segments
(Fig.
Only known from the type locality.
Dicellophilus praetermissus sp. nov. most closely resembles D. pulcher but is distinguishable by the following combination of characteristics: both ends of transverse suture not evidently convex forward; the longer than wide trochanteroprefemur; the wide rather than long metasternite (Table
The record of D. latifrons Takakuwa, 1934 (= D. pulcher) from Sendai, Miyagi Pref. (
Mecistocephalus pulcher
Kishida, 1928:
Dicellophilus latifrons:
Dicellophilus japonicus:
Tygarrup monoporus:
Dicellophilus pulcher:
Dicellophilus sp. 0000-0003-3020-8454_0068
See Table
Mainly based on
The first section of the Subashiri trail of Mt. Fuji, Shizuoka Pref., Japan (
Honshu (Fukushima Pref. to Hyogo Pref.).
See remarks and the diagnosis of D. praetermissus sp. nov. for confirming how to distinguish D. pulcher from D. praetermissus sp. nov.
There are three junior synonyms under D. pulcher, which were synonymized by previous authors based on morphological examination (
No POTU delimitation hypotheses proposed by ASAP corresponded well to the principle in steps I–IV. This is because each of those hypotheses involves many POTUs, which were separated from each other with a K2P distance lower than the intermorphospecific threshold. Although Clade B could be divided into many more POTUs, such partitioning hypotheses can be rejected as oversplitting in accordance with the species delimitation criteria in step IV with the intermorphospecific threshold (21% in COI). Therefore, two putative species were recognized in the 38 D. pulcher specimens examined in steps I–IV, and they can be separately labeled as follows: Dicellophilus sp. 0000-0003-3020-8454_0068 (= Clade B; hereafter referred to as D. sp. 0068) and D. sp. 0000-0003-3020-8454_0069 (= Clade A; hereafter referred to as D. sp. 0069, Table
Only one validly named species of Dicellophilus from Japan, D. pulcher, was described on the basis of a specimen from the Subashiri trail of Mt. Fuji, Shizuoka Pref. (
As mentioned in the result section, many POTUs were divided from the COI dataset of Dicellophilus examined in the present study. When taking into account the overall genetic diversity of all examined specimens of Dicellophilus, i.e., D. pulcher, D. carniolensis and D. praetermissus sp. nov. (maximum genetic divergence in K2P: 24%), the genetic diversity within the morphospecies D. pulcher alone is quite high (15%).
Possibly, such an oversplit of POTU would have been caused by the algorithm ASAP and the quite high genetic divergence of D. pulcher in the dataset. According to
POTUs are divided by the possibility of panmixia, so it can be expected that each POTU will be a biological species. However, each POTU within the morphospecies D. pulcher detected in the present study is not regarded as a species until morphological evidence is discovered.
Dicellophilus specimens examined in Japan were collected from 34°33'N to 38°16'N on Honshu within a latitudinal band, which is congeners’ distribution. The authors and their collaborators have collected Dicellophilus exclusively from Miyagi Pref. to Kyoto Pref. but not from other areas, despite a comprehensive field trip in Japan (Fig.
According to the molecular phylogenetic analyses of Dicellophilus specimens in Japan, it is possible that there are two large populations among D. pulcher, viz., specimens from Eastern Honshu (Fukushima Pref. to Shizuika Pref. and one specimen from Gifu Pref.) and those from western Honshu (Gifu Pref. to Kyoto Pref.), because the monophyly was supported by the phylogenetic analysis based on the concatenated and COI datasets, respectively. However, the boundary between two populations is still not clear due to the lack of field surveys in the central part of Honshu.
In field surveys conducted by the authors in Japan, D. praetermissus sp. nov. was collected only in Sendai-shi, Miyagi Pref. (the northern part of Honshu). It is also noteworthy that D. pulcher has yet to be collected from the northern part of Honshu (from Aomori Pref. to Miyagi Pref). This result of field surveys shows that the distribution of D. praetermissus sp. nov. may segregate from D. pulcher, but further field surveys are needed around Miyagi Pref.
We are grateful to Dr Masaru Nonaka (visiting researcher of Tokyo Metropolitan University), Dr Namiki Kikuchi (Toyohashi Museum of Natural History), Dr Takahiro Yoshida (assistant professor of Tokyo Metropolitan University), Mr Joe Kutsukake (Tokyo Metropolitan University), Mr Koshi Kawamoto (Tokyo Metropolitan University), Ms Mayu Susukida, Mr Ryo Miyata, Mr Tatsumi Suguro (Keio Yochisha Elementary School), Mr Tomoki Sumino, and Mr Fukube Sumino for collecting and providing Dicellophilus specimens. We are further grateful to Dr Namiki Kikuchi and Mr Joe Kutsukake for assisting in collecting and taking photographs of Dicellophilus praetermissus sp. nov., respectively. We thank two reviewers for providing valuable comments and suggestions. We also would like to thank Enago (www.enago.jp) for the English language review. This study was supported by the following funds: the Fund for the Promotion of Joint International Research (Fostering Joint International Research (B), JSPS KAKENHI, no. 22KK0087, leader: Katsuyuki Eguchi, FY2022–2025), Grant-in-Aid for Scientific Research (C) (JSPS KAKENHI, no 23K05299, Leader: Emiko Oguri, FY2023–2026), the Tokyo Metropolitan University Fund for TMU Strategic Research (leader: Noriaki Murakami, FY2020–FY2022), and the Asahi Glass Foundation (leader: Katsuyuki Eguchi, FY2017–FY2023).