J. Jpn. Bot. 91: 33–40 (2016)

Sargassum fusiforme (Fucales, Phaeophyceae) Has No Characteristic Stem in the Genus Sargassum

a, b a Hiromori Shimabukuro *, Toshinobu Terawaki and Goro Yoshida

aNational Research Institute of Fisheries and Environment of Inland Sea, Fisheries Research Agency, 2-17-5 Maruishi, Hatsukaichi, Hiroshima, 739-0452 JAPAN; bNational Research Institute of Fisheries Engineering, Fisheries Research Agency, 7620-7, Hasaki, Kamisu, Ibaraki, 314-0408 JAPAN *Corresponding author: [email protected]

(Accepted on July 1, 2015)

Sargassum fusiforme is a very useful product for fisheries and is well known in Japan; however there are few detailed reports about its morphological characteristics. This species was initially assigned to the genus Cystophyllum and was subsequently categorized in the genus Turbinaria because of a lack of distinction between its leaves and vesicles. Recently, this species was categorized in the genus Sargassum based on the results of molecular studies; however, a debate has arisen between researchers who propose that the species belongs to the genus Sargassum and those in Japan who maintain that it belongs to the genus Hizikia. This difference in opinion is caused by a large morphological variation in the leaves among the different latitudes in which the species grows. The leaves of S. fusiforme growing in temperate regions are narrow, while those growing in low-latitude regions are lanceolate with dentate margins. The branches of Sargassum spp. generally arise from the stems on the holdfast; however, the branches of S. fusiforme arise directly from the holdfast. The biennial S. fusiforme has no stem, which is a characteristic of the genus Sargassum. Having a short stem or not is not important as a characteristic of genus Sargassum.

Key words: Fucales, Hijiki, Hizikia, Phaeophyceae, Sargassum fusiforme, stem morphology.

Sargassum fusiforme (Harvey) Setchell cultivated in many areas of China and Korea is distributed in Taiwan, Japan, the Korean (Ohno 2004, Terasaki et al. 2009, Hong-Yu et al. Peninsula, and South China (Yoshida 1983, 2010). 1998, Tseng and Lu 2000, Huang 2003, Lee Sargassum fusiforme plays important roles et al. 2011). This species is a well-known and in shallow water marine ecosystems, including important brown alga in East Asia. After drying serving as nursery grounds (egg-laying sites) and and boiling, the leaves, vesicles, and lateral feeding grounds for various fish and shellfish branches of S. fusiforme are eaten, and they were (Murase et al. 2000, Shoji 2009). Because S. an important food source since ancient times fusiforme develops the highest-density seaweed in Japan. More recently, the health benefits of beds in the highest locations of the intertidal fucoxanthin and other extracts of the thalli have zone (Fig. 1), its canopies weaken the velocity of attracted attention, and this species has been ocean currents (Nishihara et al. 2011).

—33— 34 植物研究雑誌 第 91 巻 第 1 号 2016 年 2 月

morphological and anatomical observations of S. fusiforme.

Materials and Methods Specimens deposited at the National Research Institute of Fisheries and Environment of Inland Sea (FEIS #) were used in this study; Togawa, Choshi City, Chiba Prefecture (Mar. 27, 2010, FEIS 979); Jougashima, Miura City, Kanagawa Prefecture (Mar. 19, 2010, FEIS 730– 731); Furue, Kanoya City, Kagoshima Prefecture (Mar. 9, 2010, FEIS 1328–1330). Additional Fig. 1. Habitat of Sargassum fusiforme. Covering ground in the intertidal region, Okake, Ibusuki, samples were collected by us and were deposited Kagoshima Pref. (March 14, 2013) at the FEIS # after this study (Fig. 2: Izumo, Higashimuro County, Wakayama Prefecture Sargassum fusiforme was first recorded as (Mar. 5, 2010, FEIS 734–735); Minatomachi, Cystophyllum fusiforme by Harvey (1859) when Takehara, Hiroshima Prefecture (Nov. 10, specimens were collected in Shimoda City, 2009, FEIS 732–733); Maruishi, Hatsukaichi Shizuoka Prefecture, Japan. Subsequently, Yendo City, Hiroshima Prefecture (Sep. 26, 2011, (1907) categorized the species in the genus FEIS 736; May 29, 2012, FEIS 1613); Wada, Turbinaria on the basis of the characteristics Oshima County, Yamaguchi Prefecture (Jan. of its narrow, club-like leaves. Setchell (1931) 16, 2013, FEIS 1331–1333); Okake, Ibusuki reported this species growing in Hong Kong City, Kagoshima Prefecture (Mar. 14, 2013, as Sargassum fusiforme, belonging to the FEIS 1337–1340); Itarashiki, Shimajiri County, subgenus Bactrophycus. However, Okamura Okinawa Prefecture (Mar. 11, 2004, FEIS 1334– (1932) assigned the independent genus Hizikia 1336). All the specimens were collected between for this species, because the plant is difficult to March 2004 and March 2013, by snorkeling distinguish on the basis of the morphological or on foot, at nine sites (Fig. 2), and specimens characteristics of its leaves and vesicles. For a period, this species was designated under the genus Sargassum in China (Tseng 1984, Tseng and Lu 2000) and under the genus Hizikia in Japan (Yoshida 1983, 1998). Stiger et al. (2000) assigned this species to the genus Sargassum (subgenus Bactrophycus) on the basis of the analysis of the internal transcribed spacer-2 DNA sequences. In addition, Cho et al. (2012) reassessed Sargassum in Korea and included S. fusiforme on the basis of the analysis of four gene regions. However, the taxonomic issues are not entirely resolved because morphological characteristics are an important aspect of taxonomic study and few reports have presented the detailed morphology Fig. 2. Map showing collection sites of specimens used of this species. In this paper, we present detailed in this study. Scale bar: 500 km. February 2016 The Journal of Japanese Botany Vol. 91 No. 1 35

Fig. 3. Thallus of Sargassum fusiforme collected in Maruishi, Hatsukaichi, Hiroshima Pref., before becoming the dry specimen (FEIS 1613). Scale bar: 20 cm. collected by the authors additionally in Maruishi, Fisheries Technology and Development Center Hatsukaichi, Hiroshima Prefecture, to clarify (KFE) for comparison with other Sargassum the morphological characteristics of this species species. were also used in this study. The morphological features of all the Results samples were observed macroscopically and From the specimens we collected and from microscopically by using a stereomicroscope those deposited in the herbarium, Sargassum (Olympus SZX7). Hand-sectioned receptacle fusiforme was observed to have the following specimens stained in 0.5% aniline blue/water morphological characteristics. were observed under a biological microscope (Nikon Eclipse E600). All images were imported Sargassum fusiforme (Hervey) Setchell into Adobe® PhotoShop® Elements10 for plate (1931). assembly. Japanese name: Hijiki. We utilized specimens deposited in FEIS, The thallus is 20–80 cm in height and Kagoshima University, Faculty of Fisheries sometimes >100 cm in length (Fig. 3). The (KAGF), and the Kagoshima Prefectural thallus in the first year after sprouting from 36 植物研究雑誌 第 91 巻 第 1 号 2016 年 2 月

Figs. 4–6. Drawings showing holdfast and lower part Figs. 7–9. Leaves of Sargassum fusiforme. Fig. 7. of Sargassum fusiforme collected by authors from Cylindrical and narrow form of leaves collected from Maruishi, Hatsukaichi, Hiroshima Pref. Fig. 4. Lower Maruishi, Hatsukaichi, Hiroshima Pref. (FEIS 1613). part of thalli, germinating for the first time. Fig. 5. Fig. 8. Narrow oblanceolate form of leaves collected Lower part of perennial thalli, main branches arising from Furue, Kanoya, Kagoshima Pref. (FEIS 1328). directly from holdfast. Fig. 6. Lower part of thalli, main Fig. 9. Oblanceolate with dentate margin form of branch branching alternately. Scale bar: 5 mm leaves collected from Itarashiki, Shimajiri County, Okinawa Pref. (FEIS 1334). Scale bar: 2 cm. the embryo has a very small discoid holdfast with smooth surface, at the center of the holdfast (diameter, 2–3 mm) and has a small stem [Fig. 4]). This small discoid holdfast becomes a (length, 2–5 mm; diameter, about 2 mm; terete filamentous-shaped holdfast with growth (Fig. February 2016 The Journal of Japanese Botany Vol. 91 No. 1 37

and acute apex, up to 40 mm in length, about 1–2 mm in width. Few cryptostomata scattered on the surface of the leaf, midrib not present (Fig. 7). Leaves of specimens collected in Kagoshima Prefecture: narrow, oblanceolate with slightly dentate margin (Fig. 8). Leaves of specimens collected in Okinawa Prefecture: oblanceolate with dentate margins (Fig. 9). Leaves attached of specimens collected in all sites: upper part of the main and lateral branches, cylindrical to narrow with smooth surface and acute apex, simple, 10–15 mm in length, 1–2 mm in width. Vesicles are fusiform, 10–20 mm in length, 2–3 mm in width, with smooth margin and acute apex; cryptostomata scattered on the surface (Figs. 10, 11). Medullary strand (Takahashi et al. 2000) present in the vesicles. The plant is dioecious. Form of receptacles arising in clusters from the axils of leaves is cylindrical, bat-like with smooth surface, simple, 2–4 mm in length, 1 mm in width. Receptacles are longer and narrower in male plant than in Figs. 10–11. Vesicles of Sargassum fusiforme. Fig. 10. female plant (Figs. 12, 13). Male and female Fusiform form of vesicles collected from Maruishi, conceptacles scattered on receptacles, 120–200 Hatsukaichi, Hiroshima Pref. (FEIS 1613). Fig. 11. μm in diameter. Oogonia 80–100 μm in diameter Fusiform form of vesicles collected from Itarashiki, Shimajiri County, Okinawa Pref. (FEIS 1334). Scale (Figs. 14, 15). bar: 2 cm. Sargassum fusiforme grew on rocks, rock beds, and concrete blocks in the intertidal region. In some cases, thalli covered the ground in high 5). The transverse section (diameter, 1–2 mm) of densities. The Sargassum fusiforme life cycle is the filamentous holdfast is terete with a smooth as follows: initial leaves appear in September surface, branching and complex. The main to October (fall) from embryos or holdfasts that branches of first-year thalli arise from the top of over-summered on rocks; thalli grow best in the small stem, while the main branches of most winter to spring and mature in March to April thalli with a filamentous holdfast arise directly (spring). from the holdfast (Fig. 5). The primary branches The growth stages of filamentous holdfast in (diameter, 2–3 mm) are terete with a smooth this species could be observed at the Maruishi surface, mostly unbranched. Where branched, site. In its first year, S. fusiforme has a small the lower part of the main branch branches discoid holdfast and a stem (Fig. 16A, white alternately (Fig. 6). The lateral branches are arrowhead). However, a part of the holdfast very short (up to 10–20 mm in length), attached becomes extended in September to October distichously to the primary branch at intervals of (Fig. 16B, C, black arrowhead), after which, the 10–30 mm. filamentous organs arise from the lengthened Leaves collected in temperate region: simple, holdfast (Fig. 16D, arrows). When the holdfast cylindrical to linear form with smooth margin grows and enlarges, the stem fuses with the 38 植物研究雑誌 第 91 巻 第 1 号 2016 年 2 月

Figs. 12–15. Reproductive organs of Sargassum fusiforme collected by Authors from Maruishi, Hatsukaichi, Hiroshima Pref. Fig. 12. Close-up of female receptacles (arrows). Scale bar: 5 mm. Fig. 13. Close-up of male receptacles (arrows). Scale bar: 5 mm. Fig. 14. Transverse section of female receptacle. Scale bar: 200 µm. Fig. 15. Transverse section of male receptacle. Scale bar: 200 μm.

Fig. 16. Development process of filamentous holdfast from small discoid holdfast ofSargassum fusiforme, before becoming the dry specimen (FEIS 736). A. White arrowhead indicates small discoid holdfast. B, C. Black arrowheads indicate developed holdfast. D. Arrows indicate filamentous holdfast from developed holdfast. February 2016 The Journal of Japanese Botany Vol. 91 No. 1 39

have taken place since Harvey (1859), who named the species Cystophyllum fusiforme. Because of the difficulty in distinguishing leaves from vesicles, Yendo (1907) categorized this species under the genus Turbinaria. Although several morphological variations exist within this species, taxonomic assignment should not be based solely on the results of molecular experiments, and these results should be confirmed by morphological studies. The classification of S. fusiforme into Sargassum proved to be problematic because Fig. 17. Complex filamentous holdfast of Sargassum of the morphological variation within this fusiforme, before becoming the dry specimen (FEIS 979) collected from Togawa, Choshi, Chiba Pref. species and its lack of stem. However, leaves Scale bar: 2 cm. and vesicles of S. fusiforme, unlike those of the genus Turbinaria, are structurally clearly holdfast and disappears (Fig. 17). distinguishable. Additionally, the presence or absence of a stem is not an important Discussion characteristic of Sargassum; for example, the Sargassum fusiforme, the representative main axis of S. horneri arises directly from species of the temperate Sargassum, is widely the holdfast. The important characteristics of distributed across Japan from Hokkaido to the subgenus Bactrophycus are that leaves and Okinawa Island and adjacent waters. Sargassum branches can be clearly distinguished, and that fusiforme specimens grown at different simple (unbranched) reproductive organs arise latitudes show some phenotypic differences, from the axils of leaves. Therefore, S. fusiforme especially in their leaves; however, S. fusiforme can be included in the subgenus Bactrophycus of can be recognized by the following set of the genus Sargassum based on its morphological morphological characteristics: (1) filamentous characteristics, which is in agreement with the holdfast, (2) terete main branch with smooth molecular analysis result. surface, (3) fusiform-shaped leaves and vesicles, Sargassum fusiforme is a useful product for and (4) bat-like receptacles arising from the axils fisheries and is widely distributed in East Asia. of leaves. Here, we have provided detailed observations of Three Sargassum species in Japan have the morphological characteristics of this species. a filamentous holdfast: S. hemiphyllum, S. When large morphological variation exists nipponicum, and S. fusiforme. These three within a genus, as in the case of Sargassum, species are perennial brown macroalgae. taxonomic studies of species must incorporate Specimens of S. hemiphyllum and S. nipponicum observations of this variation as well as deposited in the FEIS, KAGF, and KFE herbaria molecular analyses. exhibited main branches arising from the top of the stems of the filamentous holdfast. However, We sincerely thank Dr. Ryuta Terada the main branches of S. fusiforme arise directly from the Kagoshima University (Faculty of from holdfasts rather than from stems. Sargassum Fisheries), and Mr. Shigemitsu Tokunaga from fusiforme has no stem and the main branches are the Kagoshima Prefectural Fisheries Technology alternately branched on the lower part (Fig. 6). and Development Center for allowing us Taxonomic discussions of S. fusiforme examine the specimens housed in the herbaria of 40 植物研究雑誌 第 91 巻 第 1 号 2016 年 2 月 their institutions. Pacific waters. 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a b a 島袋寛盛 ，寺脇利信 ，吉田吾郎 ：ヒジキ（ヒバマタ 目，褐藻綱）はホンダワラ属に特有な茎をもたない ヒジキは日本沿岸域に広く分布し，近年では ら螺旋状に主枝が生じる．しかし，越年したヒジキは付 Sargassum とする中国の研究者と Hizikia であると主張 着器から直接主枝が生じ，明確な茎は存在しなかった． する日本の研究者の間で議論が行われたが，最近では Sargassum にとって茎の有無は，属を分ける上での重 DNA 解析も行われ，Sargassum に含まれている．ヒジ 要な特徴ではなく，葉と気胞が独立し，葉腋から生じ キは日本でも広く知られている海藻類であるが，今まで る生殖器床が分枝しないことから形態的にもヒジキは 形態的な特徴に関する詳細な報告はほとんどなかった． Sargassum の Bactrophycus 亜属に含まれる． 温帯域に生育するヒジキは棒状の葉を有し，低緯度に生 （a国立研究開発法人 瀬戸内海区水産研究所， 育する藻体の葉は披針形で縁辺に鋸歯を有していた．従 b国立研究開発法人 水産工学研究所） 来 Sargassum は付着器から生じた円柱形の茎の頂部か