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Proc. Arthropod. Embryol. Soc. Jpn. 50, 11–13 (2016)Ⓒ 2016 Arthropodan Embryological Society of Japan
ISSN 1341–1527[SHORT COMMUNICATION]

Is Abdominal Tergal Chaetotaxy Reliable for Species Diagnosis ofJapanese Soil-Dwelling Mundochthonius Pseudoscorpions(Pseudoscorpiones: Chthoniidae)?
Hajime OHIRA1), Shingo KANEKO2) and Tadaaki TSUTSUMI2)
Graduate School of Symbiotic Systems Science and Technology, Fukushima University, 1 Kanayagawa, Fukushima, Fukushima
960–1296, Japan2)
Department of Environmental System Management, Faculty of Symbiotic Systems Science, Cluster of Science and Technology,
Fukushima University, 1 Kanayagawa, Fukushima, Fukushima 960–1296, Japan
E-mail: ohira.hajime@gmail.com (HO)1)

Pseudoscorpions are small arachnid animals living invarious habitats, i.e., in leaf litter, under stones or barks, increvices of rocks along seashore, in caves, and even in desert.
Studies on the embryonic and postembryonic developments
and reproductive biology, including reproductive behavior, ofpseudoscorpions have used specimens collected directly fromthe field, because it is difficult to establish cultures (e.g.,Morikawa, 1960, 1962; Sakayori, 1989, 2002b, 2003, 2014b;
Kato and Tsutsumi, 2004). Japanese soil-dwelling
pseudoscorpions are mainly classified based on the chaetotaxyof carapace and abdominal tergites, and pedipalpal femur
morphology (Morikawa, 1960; Sato and Sakayori, 2015).
However, these features are variable within species(Tsutusmi, 2012; Sakayori, 2014a) and chaetotaxy is known tochange with growth (Sakayori, 2014b).
In the previous studies dealing with the developmental
andreproductive
biologiesonpseudoscorpions,
Muncochthonius species have also been used as materials inJapan (Kato and Tsutsumi, 2004). In the identification of
Mundochthonius species and subspecies (Chamberlin, 1929;
Morikawa, 1954; Sakayori, 2002a, 2009), including M.
japonicus japonicus Chamberlin, M. j. scolytidis Morikawa, M.
kiyoshii Sakayori, and M. itohi Sakayori, abdominal tergal
chaetotaxy (ATC, Fig. 1) in the first, second, and third
abdominal segments are used as the most importantcharacters for classification (Sakayori, 2010), and three types
of ATC are currently distinguished: “4-4-4” in M. j. japonicus,“4-4-6” in M. j. scolytidis, and “4-6-6” in M. kiyoshii and M.
itohi (Sakayori, 2010). The three numerals that are used todescribe ATC represent the number of tergal setae that occur
in the first to third abdominal segments and are basically evennumbers, because tergal setae are symmetrical and not on the
median line. However, ATCs with an odd number are
frequently observed and recognized as an irregularity and/or
variation within a species. For example, in M. itohi, several

irregular types of ATC such as “4-5-6” and “4-6-7” are foundin addition to their regular type of ATC, “4-6-6” (Sakayori,
2009; Tsutsumi, 2012). Moreover, Tsutsumi (2012) reported
that one of the total 374 samples examined in M. itohi had anirregular “4-4-6” ATC, which is the ATC typical to M. j.
scolytidis. It may be possible that the ATC of the regular typein one species might be found as an irregular type in otherspecies. Therefore, it should be examined whether the ATC isreliable for the identification of Muncochthonius species. In
the present study, we test the reliability of ATC in theidentification of Mundochthonius species, using molecularphylogenetic analysis that was used for critical taxonomical

Fig. 1 Mundochthonius itohi Sakayori, dorsal view.
Chaetotaxy is shown only on abdominal tergal plates.
Right pedipalp and legs are omitted.

12

H. OHIRA, S. KANEKO AND T. TSUTSUMI

revisions of pseudoscorpions (e.g., Murienne et al. 2008;
Harrison et al. 2014).
Mundochthonius specimens were collected in Japan from
Kanayama (37 43 49.7 N, 140 3 13.6 E) and Sobara (37 41
22.1 N, 140 4 0.5 E) in Kitashiobara, Fukushima Prefectureand from Chinone, which is the type locality of M. itohi (36 35
27.5 N, 140 34 27.6 E), in Hitachiota, Ibaraki Prefecture.
Specimens were extracted from leaf litter and directly placed
into absolute ethanol using Tullgren funnels. Extracted
specimens were transferred to and preserved in absoluteethanol. One pedipalp was removed using a thin tungsten
needle under a stereomicroscope and transferred to a PCR
tube for DNA extraction; the body was then stored in a 0.2-ml
tube with absolute ethanol and then processed into a
microscopic slide preparation using a conventional procedure(cf. Okajima, 2006). Specimen sex was determined, and setae
in the first, second, and third abdominal tergal plates werecounted. All specimens examined were deposited in ourlaboratory at Fukushima University.
Total genomic DNA was extracted from one pedipalp
using an improved method of DNA extraction from a single
pollen grain (Suyama, 2011). Partial sequences of 18S
ribosomal RNA gene were amplified using the following
primer sets: 1F-5R, 3F-18Sbi, and 18Sa2.0–9R (Giribet et al.,
1996; Whiting et al., 1997). All nucleotide sequences wereautomatically aligned using MAFFT version 7 (Katoh andStandley, 2013). The sequence gaps were treated as missing
data using trimAI (Capella-Gutiérrez et al., 2009). Two speciesof Chthonius, which belongs to the same tribe as
Mundochthonius, Chthoniini, were used as outgroups for our
phylogenetic analysis; their sequences were obtained from
GenBank (accession numbers JN018288 and JN018289).
Phylogenetic reconstruction was performed using theneighbor-joining method (Saitou and Nei, 1987), and nodal
support of the phylogenetic tree was measured with 1000
bootstrap replicates (Felsenstein, 1985) and implemented in
MEGA version 6 (Tamura et al., 2013).
The specimens examined, 18 Mundochthonius
individuals, showed several variations in ATC, including odd
numbers of setae (“4-6-7”, “4-5-6”, and “5-4-6”), and thevariations are not related to sex (Table 1). Two of these types
of ATC, “4-6-7” and “4-5-6”, had already been reported(Sakayori, 2009; Tsutsumi, 2012), but “5-4-6” was a new
irregular type. The neighbor-joining dendrogram based on thepartial sequences of nuclear DNA 18S ribosomal RNA gene(1244 bp) revealed three distinct clades with high bootstrap
values (Clades 1–3, Fig. 2). The present analysis revealed thatthese three clades cannot be clearly distinguished by the ATC
types. The ATC types “4-6-6” and “4-4-6” predominated in
the clades 2 and 3 respectively, but different ATC types “4-56” and “5-4-6” were included as minorities in the clades 2 and3. The clade 1 included three different types “4-4-6”, “4-6-6”,

Table 1 Specimens examined in this study
Individual
Locality
Sex*
ATC**
DDBJ accession no.
M1
Kanayama
F
4-6-7
LC082336
M2
Kanayama
F
4-6-6
LC082337
M3
Kanayama
M
4-6-6
LC082339
M4
Kanayama
M
4-6-6
LC082338
M5
Kanayama
M
4-4-6
LC082343
M6
SobaraF
4-6-6
LC082341
M7
SobaraM
4-6-6
LC082342
M8
SobataM
4-4-6
LC082340
M9
ChinoneM
4-6-6
LC082344
M10
ChinoneM
4-5-6
LC082345
M11
ChinoneF
4-5-6
LC082346
M12
ChinoneF
4-4-6
LC082350
M13
ChinoneF
5-4-6
LC082351
M14
ChinoneF
4-6-6
LC082349
M15
ChinoneM
4-4-6
LC082352
M16
ChinoneM
4-6-6
LC082347
M17
ChinoneF
4-4-6
LC082353
M18
ChinoneF
4-6-6
LC082348*
F: female, M: male.
**
Abdominal tergal chaetotaxy (ATC) represents the number ofsetae in the first, second, and third abdominal tergal plates.

Fig. 2 Neighbor-joining dendrogram of Japanese soildwelling Mundochthonius inferred from the partial
sequences (1244 bp) of 18S ribosomal RNA gene.
Operational taxonomic units (OTUs) were indicatedby the individual number of Mundochthonius shownin Table 1 (M1-M18) with the ATC in parenthesis.
Chthonius dacnodes [GenBank accession number
JN018288] and C. ischnocheles [JN018289]) were usedas outgroups. Bootstrap support values are shown
above the branches.

13

ABDOMINAL TERGAL CHAETOTAXY OF MUNDOCHTHONIUS

and “4-6-7” as well. The present study revealed that the ATCcannot clearly define the monophyletic clades, and this
implies that the ATC should not be a reliable taxonomic
diagnosis for Mundochthonius.
Sakayori (2014a) pointed out that the pedipalpal femur
morphology (length:width ratio of the pedipalp femur) is not a
useful character in Allochthonius species identification. The
pedipalpal femur morphology has been employed also in theclassification of Mundochthonius as another useful diagnosis(Sato and Sakayori, 2015). Its reliability has to be also tested
for this genus, and simultaneously more reliable and practical
diagnoses in Mundochthonius identification should besurveyed.

Touka Syobo, Fukuoka.
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Acknowledgments: The present study was supported by
the Research Project for Regeneration of Harmonies between
Human Activity and Nature in Bandai-Asahi National Park in
Fukushima University.

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