
南太平洋海域調査研究報告 No.54（2014年12月）
OCCASIONAL PAPERS No.54（December 2014）

Citrus Genetic Resources
Grown on the Ryukyu Islands, JapanYAMAMOTO Masashi
Faculty of Agriculture, Kagoshima University

Abstract
The Ryukyu Islands are located southwest part of Japan. Various local citrus are grown in thissubtropical region. Since there are large geographical and climatic differences between the Ryukyu Islands
and the main islands (Honshu, Kyushu, and Shikoku) of Japan, there are unique local citrus genetic
resources on the Ryukyu Islands. Shiikuwasha (Citrus depressa) is an indigenous mandarin species in thisregion. This species is clearly distinguished from the mandarin grown in China and India based on theresults of isozyme and DNA analyses. DNA analysis also revealed that wide diversity exists in Shiikuwasha.
Although Kunenbo (C. nobilis) and Daidai (C. aurantium) have been grown in most the islands, they wereintroduced from China and/or Southeast Asia. Indigeneous Shiikuwasha and introduced species probably
played a part in the origin of many local citrus on the Ryukyu Islands. Various unique species in this region
such as Kabuchii (C. keraji), Oto (C. oto), and Rokugatsu-mikan (C. rokugatsu) have been widely
cultivated. Fruit of some local citrus contains high levels of polymethoxyflavonoids, one of the mostimportant health-promoting components of citrus.
Keywords: Amami, citrus, Okinawa, Ryukyu, Shiikuwasha

Introduction
The Ryukyu Islands are located southwest part of Japan. Various local citrus are grownin this subtropical region. Since there are marked geographical and climatic differences
between the Ryukyu Islands and the main islands (Honshu, Kyushu, and Shikoku) of Japan,
there are unique local genetic resources of citrus on the Ryukyu Islands. Thus, many
accessions were recorded and classified in several studies (INAFUKU-TERAMOTO et al. 2010,ISHIHATA et al. 1997, KINJO 2007, KITA et al. 2013, NAKANO et al. 2001, YAMAMOTO et al.
2006, 2008a) since these local citrus accessions are very important for studies on genetic
resources in citrus. The genetic relationships among them were partly clarified (YAMAMOTO etal. 1998, 2010a, 2010b, 2011). Recently, fruits produced in this region have attracted attention
because they contain high levels of health-promoting components (YAMAMOTO et al. 2008b).

10YAMAMOTO Masashi

Here, local genetic resources of citrus grown on the Ryukyu Islands, with regard to: 1)
the distribution of local citrus accessions grown on the Ryukyu Islands, 2) phylogenetic
relationships of local genetic resources of citrus grown on the Ryukyu Islands, and 3)
phytonutrient components of local citrus grown on the Ryukyu Islands, are discussed.

Distribution of Local Citrus Accessions Grown on the Ryukyu Islands
Among the local citrus accessions grown on the Ryukyu Islands, Shiikuwasha (Citrusdepressa Hayata) is the indigenous species (TANAKA 1926). Daidai (C. aurantium L.) andKunenbo (C. nobilis Lour.) were introduced from China or Southeast Asia. Since then, several
species, such as Kabuchii (C. keraji hort. ex Tanaka), Oto (C. oto hort. ex Yu. Tanaka), andRokugatsu-mikan (C. rokugatsu hort. ex Yu. Tanaka), have grown as new seedlings that havearisen by chance from indigenous or introduced species (TANAKA 1948) (Fig. 1 and Table 1).

ABCDEFGHIJKLFig. 1. Fruits of local citrus on the Ryukyu Islands. A: Shiikuwasha (Citrus depressa), B: Kunenbo (C.
nobilis), C: Daidai (C. aurantium), D: Buntan (C. maxima), E: Akamikan (C. tangerina), F:Shimamikan (C. sp.), G: Kabuchii (C. keraji), H: Keraji (C. keraji), I: Oto (C. oto), J: Tarogayo (C.
tarogayo), K: Rokugatsu-mikan (C. rokugatsu), L: Shiikuu, Kusa, or Tunugekunin (C. sp.).

11Citrus Genetic Resources Grown on the Ryukyu Islands, Japan

Table 1. Major local citrus grown on the Ryukyu Islands.

Common name

Latin name

OriginShiikuwasha

Citrus depressa Hayata

Authigene

KunenboC. nobilis Lour.
C. aurantium L.

Introduction
Introduction

Pummelo (Buntan)
Dancy (Obeni-mikan)

C. maxima (Burm.) Merr.

Introduction

C. tangerina hort. ex Tanaka

Introduction

Komikan (Shimamikan)

C . sp.

Probably introduction

Kabuchii
KerajiC. keraji hort. ex Tanaka

Chance seedlingC. keraji hort. ex Tanaka

Chance seedlingOto

C. oto hort. ex Yu. Tanaka

Chance seedling

Tarogayo

C. tarogayo hort. ex Tanaka

Chance seedling

Rokugatsu-mikan

C. rokugatsu hort. ex Yu. Tanaka

Chance seedling

Fusu, Kusa or Tunugekunin

C . sp.

Chance seedlingSour orange (Daidai)

Phylogenetic Relationships of Local Genetic Resources of Citrus
Grown on the Ryukyu Islands
The results of isozyme and cleaved amplified polymorphic sequence (CAPS) analyses ofchloroplast DNA (cpDNA) are shown in Table 2 and Table 3, respectively (YAMAMOTO et al.
2011, 2013).
The indigeneous Shiikuwasha and introduced Kunenbo probably played a role in theorigin of many local citrus on the Ryukyu Islands. Keraji, Kabuchii, and Oto are closely
related to Kunenbo based on the results of isozyme and DNA analyses. The type of cpDNA ofKeraji, Kabuchii, Tarogayo, and Oto is the same as that of Kunenbo. This suggests that theyarose from Kunenbo as a female ancestor because cpDNA is inherited maternally. On theother hand, the genetic influence of Japanese mandarins such as Shiikuwasha on Kabuchii andOto was observed. Their GOT-2 genotype was MA, and A is a characteristic allele of theJapanese mandarins, Shiikuwasha and Tachibana (HIRAI et al. 1986). The involvement ofShiikuwasha in the development of Rokugatsu-mikan and Fusu was identified; all of thempossessed A in GOT-2, a characteristic allele of Japanese mandarin.
In cpDNA analysis, Shiikuwasha belonged to both types 4 and 5 in Table 3. The cpDNA
divergence of this species grown in Okinawa was also reported by URASAKI et al. (2005).
These results suggest a polyphyletic origin of Shiikuwasha.

12YAMAMOTO Masashi

Table 2. Isozyme genotype of major local citrus grown on the Ryukyu Islands.

Common name

Latin nameShiikuwasha
KunenboSour orange

Citrus depressa Hayata
C. nobilis Lour.

Genotype ofGOT-2
MAMM

C. aurantium L.

MMPummelo
Dancy
Komikan (Shimamikan)
KabuchiiKeraji
OtoRokugatsu-mikan

C. maxima (Burm.) Merr.
C. tangerina hort. ex Tanaka
C . sp.
C. keraji hort. ex Tanaka
C. keraji hort. ex Tanaka
C. oto hort. ex Yu. Tanaka
C. rokugatsu hort. ex Yu. Tanaka

MMMMMMMAMMMA
MAShiiku, Kusa,or Tunugekunin

C . sp.

MAC. tachibana (Makino) Tanaka
C. kinokuni hort. ex Tanaka
C. unshiu Marcow.

AAMMMM

ControlTachibana
Kinokuni
Satsuma mandarin

PonkanC. reticulata BlancoLemon
C. limon (L.) Burm. f.
Modified from YAMAMOTO et al . (2011).

MMSM

Table 3. Types of chloroplast DNA (cpDNA) based on CAPS analysis of major local citrus grown on theRyukyu Islands.

Type Common name1
Kunenobo, Sour orange, Pummelo, Kabuchii, Keraji, Oto, Tarogayo, Sweet orange, lemon
2
Khasi Papeda
3
Ichang Papeda, Yuzu
4
Shiikuwasha, Tachibana
5
Shiikuwahsa, Sunki, Cleopatra, Tankan
6Dancy, Ponkan, Satsuma mandarin, Kinokuni, Clementine
Modified from YAMAMOTO et al . (2013).

13Citrus Genetic Resources Grown on the Ryukyu Islands, Japan

Phytonutrient Components of Local Citrus Grown on the Ryukyu Islands
Citrus fruits are essential sources of some phytonutrient components. Among them,polymethoxyflavonoids (PMFs), unique components of citrus, show efficacy against
lifestyle-related diseases such as cancer and diabetes (KAWAI et al. 1999, LEE et al. 2010,MIYATA et al. 2008). The PMF content of Shiikuwasha and Kabuchii was much higher thanthat of Satsuma mandarin (C. unshiu), which is a leading citrus species in Japan (Table 4)(YAMAMOTO et al. 2008).

Table 4. Polymethoxyflavonoid content of juice and peelin Shiikuwasha and Kabuchii.

AccessionDate(y/m/d)

Polymetoxyflavanoid
Juice(pg/mL)

Peel(pg/g)

Shiikuwasha(C. depressa )2004/9/17
2004/10/27
2004/12/8

17.6
9.9
0.9

4,699.8
3,523.4
2,848.3

Kuriha(Citrus keraji )2004/9/17
2004/10/27
2004/12/8

20.6
12.1
1.9

3,584.7
2,833.4
2,628.0

Control
Miyagawa-wase 2004/10/27
2.0(C. unshiu )Modified from YAMAMOTO et al . (2008).

369.7

References
HIRAI, M., KOZAKI, I. and KAJIURA, I. 1986. Isozyme Analysis and Phylogenic Relationshipof Citrus. Japanese Society of Breeding, 36: 377-389.
INAFUKU-TERAMOTO, S., YAMAMOTO, M., KINJO, H., KITAJIMA, A., WADA, K. and
KAWAMITSU, Y. 2010. Local Citrus Genetic Resources and Their Polymethoxyflavones
Content in Northern Part of Okinawa Island. Horticultural Research (Japan), 9: 263-271(in Japanese with English Summary).
ISHIHATA, K., SHIOTA, H. and ONJO, M. 1997. Some Kinds of Tropical and Subtropical Fruit
Trees and Characteristics of Some Native Citrus Fruits in Amami Islands. Bulletin of theExperimental Farm, Faculty of Agriculture, Kagoshima University, 22: 1-13 (in Japanesewith English Summary).

14
YAMAMOTO Masashi

KAWAI, S., TOMONO, Y., KATASE, E., OGAWA, K. and YANO, M. 1999. Antiprofiferative
Activity of Flavonoids on Several Cancer Cell Lines. Bioscience, Biotechnology, andBiochemistry, 63: 896-899.
KINJO, H. 2007. Acid Citrus “Shiikuwasha” in Ryukyu Island. Studia Citrologica, 17: 137-148(in Japanese).
KITA, M., YAMAMOTO, M., INAMORI, H., SAKAUE, H., SEKITA, T., TAHARA, A. and MINORU, H.
2013. Exploration of Citrus Genetic Resources in Amami Archipelago in KagoshimaPrefecture, Japan. Annual Report on Exploration and Introduction of Plant Genetic
Resources, 29: 107-117 (in Japanese with English Summary).
LEE, Y. S., CHA, B. Y., SAITO, K., YAMAKAWA, H., CHOI, S. S., YAMAGUCHI, K., YONEZAWA,T., TERUYA, T., NAGAI, K. and WOO, J. T. 2010. Nobiletin Improves Hyperglycemia andInsulin Resistance in Obese Diabetic ob/ob Mice. Biochemical Pharmacology, 79:
1674-1683.
MIYATA, Y., SATO, T., IMADA, K., DOBASHI, A., YANO, M. and ITO, A. 2008. A Citrus
Polymethoxyflavonoid, Nobiletin, Is a Novel MEK Inhibitor That Exhibits Antitumor
Metastasis in Human Fibrosarcoma HT-1080 Cells. Biochemical and BiophysicalResearch Communications, 366: 168-173.
NAKANO, M., NESUMI, H. and YOSHIDA, T. 2001. Exploration for Citrus Genetic Resourceson the Nansei Archipelago, Japan. Annual Report on Exploration and Introduction ofPlant Genetic Resources, 17: 39-48 (in Japanese with English Summary).
TANAKA, T., 1926. Wild Citri of the Japanese Territories. Bulteno Scienca de la Fakultato
Terkultura, Kjusu Imperia Universitato, 2: 51-58 (in Japanese with English Summary).
TANAKA, Y. 1948. An Iconograph of Japanese Citrus Fruits. Vol. 2, 525 pp., Yokendo, Tokyo(in Japanese).
URASAKI, N., YOSHIDA, K., UEHARA, T., INOUE, H., ONDA, S., KINJO, H. and KAWANO, S.
2005. Single Nucleotide Polymorphism in Shiikuwasha (Citrus depressa Hayata)
Chloroplast DNA, trnL-trnF. Japanese Journal of Tropical Agriculture, 49: 246-251.
YAMAMOTO, M., FUKUDA, M., KOGA, T., KUBO, T. and TOMINAGA, S. 2010a. Examination ofthe Origin of Keraji (Citrus keraji), Local Citrus of Kikaijima Island, Kagoshima
Prefecture. Horticultural Research (Japan), 9: 7-12 (in Japanese with English Summary).
YAMAMOTO, M., KOUNO, R., NAKAGAWA, T., USUI, T., KUBO, T. and TOMINAGA, S. 2011.
Isozyme and DNA Analyses of Local Citrus Germplasm on Amami Islands, Japan.
Journal of the Japanese Society for Horticultural Science, 80: 268-275.
YAMAMOTO, M., KUBO, T. and TOMINAGA, S. 2008a. Exploration of Citrus depressa Hayata
Germplasm in Okinoerabu Island. Bulletin of the Experimental Farm, Faculty ofAgriculture, Kagoshima University, 32: 7-12 (in Japanese).
YAMAMOTO, M., MATSUMOTO, R., UECHI, Y., IJICHI, T., KUBO, T. and TOMINAGA, S. 2008b.
The Polymethoxy Flavones Content of Local Citrus Accessions on the Island of

15Citrus Genetic Resources Grown on the Ryukyu Islands, Japan

Kikaijima in Kagoshima Prefecture, Japan. Bulletin of the Faculty of Agriculture,
Kagoshima University, 58: 1-7 (in Japanese with English Summary).
YAMAMOTO, M., MATSUO, Y., KUNIGA, T., MATSUMOTO, R. and YAMADA, Y. 1998. Isozymeand RAPD Analyses of Shiikuwashas (Citrus depressa Hayata). Bulletin of the National
Institute of Fruit Tree Science, 30/31: 39-51 (in Japanese with English Summary).
YAMAMOTO, M., MATSUSHIMA, K., IJICHI, T., UECHI, Y., KAWAGUCHI, S., NAKANO, H.,NOMURA, T., TANIMURA, O., KUBO, T. and TOMINAGA, S. 2006. Exploration andPreservation of Local Citrus Germplasms in the Amami Archipelago. Bulletin of theExperimental Farm, Faculty of Agriculture, Kagoshima University, 29: 5-11 (in Japanesewith English Summary).
YAMAMOTO, M., TSUCHIMOCHI, Y., NINOMIYA, T., KOGA, T., KITAJIMA, A., YAMASAKI, A.,INAFUKU-TERAMOTO, S., YANG, X., YANG, X., ZHONG, G., NASIR, N., KUBO, T. andTOMINAGA, S. 2013. Diversity of Chloroplast DNA in Various Mandarins (Citrus spp.)
and Other Citrus Demonstrated by CAPS Analysis. Journal of the Japanese Society for
Horticultural Science, 82: 106-113.
YAMAMOTO, M., YAMASAKI, A., SHIMIZU, T., KITAJIMA, A., KUBO, T. and TOMINAGA, S.
2010b. SSR Analysis of Local Citrus Accessions Grown in Amami Islands. Horticultural
Research (Japan), 9 (suppl. 2): 86 (in Japanese).