Genetic Distinctness of Sorex caecutiens hallamontanus (Soricomorpha: Mammalia) from Jeju Island in Korea: Cytochrome Oxidase I and Cytochrome b Sequence Analyses

2012·0.2 MB·

by  Hung-Sun Koh

Anim. Syst. Evol. Divers. Vol. 28, No. 3: 215-219, July 2012
http://dx.doi.org/10.5635/ASED.2012.28.3.215
Short communication
Genetic Distinctness of Sorex caecutiens hallamontanus
(Soricomorpha: Mammalia) from Jeju Island in Korea:
Cytochrome Oxidase I and Cytochrome b
Sequence Analyses
Hung Sun Koh*, Kyung Hee Jang, Seong Teak In, Eui Dong Han, Jae Eun Jo,
Eui Jeong Ham, Seon Ki Jeong, Jong Hyek Lee, Kwang Seon Kim, Gu Hee Kweon
Department of Biology, Chungbuk National University, Cheongju 361-763, Korea
ABSTRACT
To examine genetic divergences of two endemic Sorex caecutienssubspecies from Korea(S. c. hallamontanus
in Korean Jeju Island and S. c. annexus in the mainland Korean Peninsula), we obtained partial cytochrome
oxidase I(COI) sequences(429bp) and complete cytochrome b sequences(1,140bp) from the two Korean
subspecies, and we compared these sequences to the corresponding sequences of S. caecutiens, obtained from
GenBank. We found that Jeju S. c. hallamontanusis one of three clades within S. caecutiens, with an average
Jukes-Cantor distance of 1.57% in the COI sequences and the distance of 2.07% and 11 fixed site differences
in the cytochrome bsequences, indicating that Jeju S. c. hallamontanusis one endemic subspecies with con-
cordant genetic distinctness, although further analyses with nuclear DNA sequences are necessary to confirm
these findings. However, S. c. annexusfrom the mainland Korean Peninsula was not divergent from S. c. mac-
ropygmaeus from northeastern China and adjacent Russia, indicating that S. c. annexus from the mainland
Korean Peninsula is another endemic subspecies with only morphological differences, although it is necessary
to reexamine the subspecies status of S. c. annexus.
Keywords:genetic divergence, cytochrome bgene, COI, Sorex caecutiens hallamontanus, Jeju Island
INTRODUCTION the Eurasian continent, the mainland Korean Peninsula, Sak-
halin, and Jeju, and the shrews from Jeju were identified as
The Laxmann’s shrew(Sorex caecutiens Laxmann 1788), S. caucutiens. Additionally, Ohdachi et al.(2005) classified
composed of 21 nominal subspecies, is found across the from morphometric analyses S. caecutiensfrom Jeju as a new
northern part of the Eurasian continent, including the main- subspecies of S. c. hallamontanusAbe and Oh.
land Korean Peninsula, and the peripheral islands of the far- To examine genetic divergences of the two subspecies of
eastern Russian Sakhalin and Japanese Hokkaido(Hutterer, Sorex caecutiensfrom Korea(S. c. hallamontanusin Korean
2005). Sorex c. macropygmaeusMiller is distributed in north- Jeju Island and S. c. annexus from the mainland Korean
eastern China and adjacent far-eastern Russia, while S. c. Peninsula) from cytochrome oxidase I(COI) and cytochrome
annexusThomas is found in the central and southern parts of b sequence analyses, we collected three specimens of S. c.
the mainland Korean Peninsula(Jones and Johnson, 1960). hallamontanusfrom Jeju Island(one specimen from Kwane-
From cytochrome b sequence analyses, Ohdachi et al. umsa and two specimens from Youngsil) and four specimens
(2001) found two distinct subgroups within Sorex caecutiens of S. c. annexus from the mainland Korean Peninsula(one
(one cluster from Hokkaido Island and the other cluster from specimen from Mt. Sobaek and three specimens from Mt.
Eurasian continent and Sakhalin Island), and Ohdachi et al. Odae). Small pieces of muscle were collected and preserved
(2003) reported that the Jeju S. caucutiensformed one of six in a deep freezer.
subclusters within the Continent-Sakhalin-Jeju cluster from Total cellular DNA was extracted using a genomic DNA
cc This is an Open Access article distributed under the terms of the Creative *To whom correspondence should be addressed
Commons Attribution Non-Commercial License(http://creativecommons.org/ Tel: [phone], Fax: [phone]
licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, E-mail: [email protected]
and reproduction in any medium, provided the original work is properly cited.
pISSN 2234-6953 eISSN 2234-8190 Copyright The Korean Society of Systematic Zoology
Hung Sun Koh, Kyung Hee Jang, Seong Teak In, Eui Dong Han, Jae Eun Jo, Eui Jeong Ham, Seon Ki Jeong, Jong Hyek Lee, Kwang Seon Kim, Gu Hee Kweon
extraction kit(Intron Co., Seoul, Korea). The COI gene was Korean Peninsula(six cytochrome b haplotypes of two S.
PCR-amplified using the RonM and NancyM primers, desi- caecutienssubspecies are deposited in GenBank under acce-
gned by Pfunder et al.(2004), and the PCR thermal cycle ssion nos. JX192946-JX192951). Within the 19 cytochrome
employed was as follows: 94�C for 5min; 94�C for 1min, bhaplotypes of S. caecutiens(six haplotypes from this study
53�C for 1min, 72�C for 1min(35 cycles); and 72�C for 5 and 13 haplotypes from GenBank), 104 sites(9.12%) were
min. The cytochrome b gene was PCR-amplified using the variable, and 82 sites(7.19%) were parsimony informative.
L14724 and H15149 primers, designed by Irwin et al.(1991), Maximum likelihood trees with the 19 haplotypes of S. cae-
and the PCR thermal cycle employed was as follows: 94�C cutiensare shown in Fig. 1. All 19 haplotypes were subdivid-
for 5min; 94�C for 1min, 55�C for 1min, 72�C for 1min ed into three distinct subgroups: ten haplotypes from Primor-
(25 cycles); and 72�C for 5min. The amplified products were ye, the mainland Korean Peninsula, and Sakhalin(Gp 1, Con-
purified using a DNA PrepMate kit with a silica-based ma- tinent-Sakhalin clade), seven haplotypes from Jeju Island(Gp
trix(Intron Co.) The purified PCR products were analyzed 2, Jeju clade), and two haplotypes from Hokkaido(Gp 3, Hok-
with an automated DNA Sequencer(Perkin Elmer 377; Perkin kaido clade). The average JC distances between Gps 1 and 2
Elmer, Norwalk, CT, USA) at Bioneer Co.(Seoul, Korea). was 2.07%, between Gps 1 and 3 was 6.27%, and between
We first obtained partial COI sequences(429bp) of the Gps 2 and 3 was 7.03%. In addition, Gp 2 was distinct from
seven specimens from the two subspecies of S. caecutiensin Gp 1, with 11 fixed site differences(site nos., 47, 178, 216,
Korea, and these sequences were compared to two haplotypes 219, 270, 609, 618, 690, 717, 726, and 898) among 29 parsi-
of S. caecutiens from Maoershan in northeastern China(JF mony informative sites, and the average JC distance between
444466 from Heilongjiang Sheng and JQ599823 from Liao- seven haplotypes from the mainland Korean Peninsula(Gp 1,
yeling Sheng), obtained from GenBank. We also obtained in part) and other three haplotypes from Primorye and Sak-
complete cytochrome b sequences(1,140bp) of the seven halin(Gp 1, the rest) was 0.69%.
specimens from the two subspecies of S. caecutiensin Korea, From the seven COI sequences(429bp), one haplotype
and these sequences were compared to the corresponding 13 (COIJeju01) was identified from S. c. hallamontanusin Jeju
sequences of S. caecutiens, obtained from GenBank: two ha- Island, and two haplotypes(COIKorea01 and COIKorea02)
plotypes(AB028561 and AB028560) from Hokkaido Island, were, from S. c. annexusin the mainland Korean Peninsula
four haplotypes(AB062725 and AB062727 to AB062729) (three COI haplotypes of two S. caecutiens subspecies are
from the mainland Korean Peninsula, two haplotypes(AB0 deposited in GenBank under accession nos. JX192952-JX1
62731 and AB028554) from Ussuriiskii, Primorye in far- 92954). Maximum likelihood trees with the five haplotypes
eastern Russia, one haplotype(AB028556) from Sakhalin of S. caecutiens(three haplotypes from this study and two
Island, and four haplotypes(AB062721 to AB062724) from haplotypes from GenBank) are shown in Fig. 2. One mono-
Jeju Island. genic haplotype of S. c. hallamontanusfrom Jeju Island(Gp
Sequence alignment, detection of parsimony informative 2) was different from four haplotypes of other S. caecutiens
sites, model selection, calculation of nucleotide distances, from the mainland Korean Peninsula and northeastern China
and tree constructions with 1,000 bootstrapped replications (Gp 1): the average JC distance between two haplotypes from
were carried out using MEGA5(Tamura et al., 2011): the the mainland Korean Peninsula(Gp 1, in part) and two hap-
Jukes Cantor(JC) model , which showed the lowest Bayesian lotypes from northeastern China(Gp 1, the rest) was 0.47%,
information criterion scores, was chosen, and maximum like- but the distance between two subgroups(Gps 1 and 2) was
lihood trees were constructed. In addition, another maximum 1.57%.
likelihood trees were constructed by PAUP version 4.0b
(Swofford, 2002): HKY85 substitution model was selected
by the PAUP, and heuristic searches were performed with DISCUSSION
step-wise addition options. Sorex isodon(AB062736 for cyto-
chrome band JF444467 for COI) was used as outgroup. In this study with the cytochrome bgene sequences(Fig. 1),
we found that S. caecutiensis composed of three distinct sub-
groups(Hokkaido subgroup from Hokkaido Island, Gp 3;
RESULTS Continent-Sakhalin subgroup from Primorye, the mainland
Korean Peninsula, and Sakhalin Island, Gp 1; and Jeju sub-
From the seven cytochrome b sequences(1,140bp), three group from Jeju Island, Gp 2). The Jeju subgroup(S. c. hal-
haplotypes(CBJeju01 to CBJeju03) were identified from S. lamontanus) was distinct from the other two subgroups(Hok-
c. hallamontanus in Jeju Island, and three haplotypes(CB kaido and Continent-Sakhalin), with the average nucleotide
Korea01 to CBKorea03), from S. c. annexusin the mainland distance of 7.03% and 2.07%, respectively. However, Ohda-
216 Anim. Syst. Evol. Divers. 28(3), 215-219
Genetic Distinctness of Jeju S. c. hallamontanus
A
64 CBKorea03
80 AB062727Korea
AB062728Korea
52
AB062725Korea
CBKorea01
Gp1
AB062731Primorye
AB028554Primorye
78
AB028556Sakhalin
CBKorea02
95 AB062729Korea
100 CBJeju01
94
AB062721Jeju
AB062723Jeju
CBJeju03 Gp2
100
S. caecutiens
hallamontanus AB062722Jeju
CBJeju02
AB062724Jeju
AB028561Hokkaido
Gp3
100 AB028560Hokkaido
AB062736
0.01
B AB062727Korea
88 AB062728Korea
CBKorea03
AB062725Korea
CBKorea01
Gp1
AB062731Primorye
AB028554Primorye
83
AB028556Sakhalin
92 CBKorea02
AB062729Korea
CBJeju01
100
78 AB062723Jeju
AB062721Jeju
AB062724Jeju Gp2
S. caecutiens CBJeju02
100 100
AB062722Jeju
hallamontanus
CBJeju03
100 AB028561Hokkaido
Gp3
AB028560Hokkaido
AB062736
0.01
Fig. 1. Maximum likelihood trees with 19 cytochrome b haplotypes(1,140bp) of Sorex caecutiens. The two trees(A and B) were
constructed by MEGA5 and PAUP 4.10b, respectively, and the bootstrap values ¤50% are reported at the internodes. Six haplotypes
(CBKorea01-CBKorea03 and CBJeju01-CBJeju03) were obtained from this study, and for other 13 haplotypes obtained from GenBank,
location name follows the accession number in each haplotype. Sorex isodon(AB062736) was used as outgroup.
Anim. Syst. Evol. Divers. 28(3), 215-219 217
Hung Sun Koh, Kyung Hee Jang, Seong Teak In, Eui Dong Han, Jae Eun Jo, Eui Jeong Ham, Seon Ki Jeong, Jong Hyek Lee, Kwang Seon Kim, Gu Hee Kweon
A
74 JF444466China
95 JQ599823China
Gp1
COIKorea01
S. caecutiens
COIKorea02
COIJeju01 Gp2
hallamontanus
JF444467
0.01
B JF444466China
JQ599823China
Gp1
81 COIKorea01
S. caecutiens 96 COIKorea02
hallamontanus
COIJeju01 Gp2
JF444467
0.01
Fig. 2. Maximum likelihood trees with five cytochrome oxidase I(COI) haplotypes(429bp) of Sorex caecutiens. The two trees(A
and B) were constructed by MEGA5 and PAUP 4.10b, respectively, and the bootstrap values ¤50% are reported at the internodes.
Two haplotypes(COIKorea01 and COIKorea02) from the mainland Korean Peninsula and one haplotype(COIJeju01) from Jeju Island
were obtained from this study, and for other two haplotypes(JF444466 and JQ599823), obtained from GenBank, location name
follows the accession number in each haplotype. Sorex isodon(JF444467) was used as outgroup.
chi et al.(2003) reported from their cytochrome bsequence A subspecies is an aggregate of phenetically similar popu-
analysis that S. caecutiensis composed of two distinct sub- lations of a species differing taxonomically from other pop-
groups(one cluster from Hokkaido Island and the other clus- ulations of that species(Mayr and Ashlock, 1991), and it was
ter from Eurasian continent, Sakhalin Island, and Jeju Island), advocated that a classification should reflect all available
and that four shrews of Sorex caecutiens from Jeju Island characters distributed as widely and evenly as possible over
formed one of six subclusters within the Continent-Sakhalin- the organisms studied(Huelsenbeck et al., 1996). Ohdachi et
Jeju cluster from Eurasian continent, the mainland Korean al.(2005) classified S. caecutiensfrom Jeju Island as a new
Peninsula, Sakhalin, and Jeju. subspecies of S. c. hallamontanusfrom morphometric analy-
Pfunder et al.(2004) noted that COI gene can be used for ses, whereas Tatsuo et al.(2005) reported that chromosomal
species identification of shrews and voles. From this COI characteristics of S. caecutiensfrom Jeju Island were essen-
sequence analysis with S. caecutiens(Fig. 2) S. c. hallamon- tially identical to those of S. caecutiens from the Eurasian
tanusfrom Jeju Island(Gp 2) was distinct from other S. cae- continent. In this paper with cytochrome band COI sequence
cutiens in the mainland Korean Peninsula and northeastern analyses(Figs. 1, 2), we found that S. c. hallamontanusis a
China(Gp 1), with the average JC distance of 1.57%. Thus, distinct clade, which is genetically divergent from other pop-
we concluded that endemic Jeju S. c. hallamontanus is one ulations of S. caecutiens. The importance of examining both
of three clades within S. caecutiens, with concordant genetic nuclear and mitochondrial loci when attempting to elucidate
distinctness in mtDNA cytochrome band COI sequences. patterns of genetic structure has been previously mentioned
Island populations should diverge over time(genetically (Rubinoff and Holland, 2005), and we propose further sys-
and morphologically) from the respective mainland species tematic analyses with nuclear DNA sequences to confirm our
populations(Johnson et al., 2000), although at the end of the findings.
last glacial, large areas of continental shelf were dry land, Bradley and Baker(2001) noted that a genetic distance of
facilitating exchange of plant and animal species by land ⁄2% based on the cytochrome bgene was typical of a pop-
bridge connections to what are now isolated islands(Lomo- ulation and infraspecific variation. Ohdachi et al.(2003) re-
lino et al., 2010). Thus, we considered that genetically dis- ported from their cytochrome bsequence analysis that phy-
tinct S. c. hallamontanusfrom Jeju Island has been geogra- logenetical relationships among the other five subclusters
phically isolated even during the last glacial. within the Continent-Sakhalin-Jeju cluster do not always re-
218 Anim. Syst. Evol. Divers. 28(3), 215-219
Genetic Distinctness of Jeju S. c. hallamontanus
flect the geographical proximity of their capture locations. geography. 4th ed. Sinauer Associates Inc., Sunderland, MA,
In addition, in this study(Figs. 1, 2) S. c. annexusfrom cen- pp. 1-878.
tral and southern part of the mainland Korean Peninsula(Gp Mayr E, Ashlock PD, 1991. Principles of systematic zoology.
1, in part) was not divergent from S. c. macropygmaeusfrom McGraw-Hill Inc., New York, pp. 1-475.
Ohdachi S, Dokuchaev NE, Hasegawa M, Masuda R, 2001. In-
northeastern China and nearby Russia(Gp 1, the rest), with
traspecific phylogeny and geographical variation of six spec-
the average JC distance of 0.69% in the cytochrome bsequ-
ies of northeastern Asiatic Sorexshrews based on the mito-
ences and the JC distance of 0.47% in the COI sequences.
chondrial cytochrome bsequences. Molecular Ecology, 10:
On the other hand, Jones and Johnson(1960) noted that S. c.
2199-2213.
annexus from the mainland Korean Peninsula is larger in
Ohdachi SD, Abe H, Han SH, 2003. Phylogenetical positions of
morphological characters than S. c. macropygmaeus from Sorexsp.(Insectivora, Mammalia) from Cheju Island and S.
northeastern China and adjacent Russia. Though the mainland caecutiensfrom the Korean Peninsula, inferred from mito-
Korean S. c. annexus has been considered as an endemic chondrial cytochrome bgene sequences. Zoological Science,
Korean subspecies, our results based on the two mitochond- 20:91-95.
rial gene sequences revealed that the mainland Korean sub- Ohdachi SD, Abe H, Oh HS, Han SH, 2005. Morphological re-
species is not divergent from its neighboring northeastern lationships among populations in the Sorex caecutiens/shinto
group(Eulipotyphla, Soricidae) in East Asia, with a descrip-
Chinese subspecies of S. c. macropygmaeus, requiring reex-
tion of a new subspecies from Cheju Island, Korea. Mam-
amination for the subspecies status of S. c. annexus.
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