See discussions, stats, and author profiles for this publication at: https://www.researchgate.net/publication/303397768 Bruguiera hainesii, a critically endangered mangrove species, is a hybrid between B. cylindrica and B. gymnorhiza (Rhizophoraceae) Article in Conservation Genetics · May 2016 DOI: 10.1007/s10592-016-0849-y CITATIONS READS 10 1,037 15 authors, including: Jean W H Yong Koji Takayama Swedish University of Agricultural Sciences Museum of Natural and Environmental History, Shizuoka 138 PUBLICATIONS 4,291 CITATIONS 155 PUBLICATIONS 679 CITATIONS SEE PROFILE SEE PROFILE M. Nazre Alison K.S. Wee Universiti Putra Malaysia Guangxi University 47 PUBLICATIONS 439 CITATIONS 25 PUBLICATIONS 252 CITATIONS SEE PROFILE SEE PROFILE Some of the authors of this publication are also working on these related projects: Organellar genomics of tropical trees View project Plant Genetic Profiling View project All content following this page was uploaded by Jean W H Yong on 30 July 2016. The user has requested enhancement of the downloaded file. Conserv Genet DOI 10.1007/s10592-016-0849-y RESEARCH ARTICLE Bruguiera hainesii, a critically endangered mangrove species, is a hybrid between B. cylindrica and B. gymnorhiza (Rhizophoraceae) 1 2 3 4 Junya Ono • Jean W. H. Yong • Koji Takayama • Mohd Nazre Bin Saleh • 1,5 1 6 7 Alison K. S. Wee • Takeshi Asakawa • Orlex Baylen Yllano • Severino G. Salmo III • 8 9 10 Monica Suleiman • Nguyen Xuan Tung • Khin Khin Soe • 11 1 Sankararamasubramanian Halasya Meenakshisundaram • Yasuyuki Watano • 12 1,13 Edward L. Webb • Tadashi Kajita Received: 20 November 2015 / Accepted: 3 May 2016 Ó Springer Science+Business Media Dordrecht 2016 Abstract Bruguiera hainesii (Rhizophoraceae) is one of hainesii samples from five populations were heterozygous at the two Critically Endangered mangrove species listed in the both loci, with one allele was shared with B. cylindrica, and IUCN Red List of Threatened Species. Although the species the other with B. gymnorhiza. For chloroplast DNA markers, is vulnerable to extinction, its genetic diversity and the the two haplotypes found in B. hainesii were shared only by evolutionary relationships with other Bruguiera species are B. cylindrica. These results suggested that B. hainesii is a not well understood. Also, intermediate morphological hybrid between B. cylindrica as the maternal parent and B. characters imply that the species might be of hybrid origin. gymnorhiza as the paternal one. Furthermore, chloroplast To clarify the genetic relationship between B. hainesii and DNA haplotypes found in B. hainesii suggest that other Bruguiera species, we conducted molecular analyses hybridization has occurred independently in regions where including all six Bruguiera species using DNA sequences of the distribution ranges of the parental species meet. As the two nuclear genes (CesA and UNK) and three chloroplast IUCN Red List of Threatened Species currently excludes regions (intergenic spacer regions of trnL-trnF, trnS-trnG hybrids (except for apomictic plant hybrids), the conserva- and atpB-rbcL). For nuclear DNA markers, all nine B. tion status of B. hainesii should be reconsidered. Keywords Mangrove Á Hybridization Á Endangered Electronic supplementary material The online version of this species Genetics article (doi:10.1007/s10592-016-0849-y) contains supplementary Á material, which is available to authorized users. & Tadashi Kajita 8 Institute for Tropical Biology and Conservation, Universiti [email protected] Malaysia Sabah, Kota Kinabalu, Malaysia 9 1 Department of Biology, Graduate School of Science, Chiba Mangrove Ecosystem Research Centre, Hanoi National University, Chiba, Japan University of Education, Hanoi, Vietnam 10 2 Life Sciences, Singapore University of Technology and Department of Botany, University of Yangon, Union of Design, Singapore, Singapore Myanmar, Yangon, Myanmar 11 3 Museum of Natural and Environmental History, 5762 Oya, Biotechnology Programme, M. S. Swaminathan Research Suruga-ku, Shizuoka-shi, Shizuoka, Japan Foundation, Chennai, India 12 4 Faculty of Forestry, Putra Malaysia University, Department of Biological Science, National University of Seri Kembangan, Malaysia Singapore, Singapore, Singapore 13 5 College of Forestry, Guangxi University, Nanning, Guangxi, Iriomote Station, Tropical Biosphere Research Center, China University of the Ryukyus, 870 Uehara, Taketomi-cho, Yaeyama-gun, Okinawa 907-1541, Japan 6 Biology Department, College of Science and Technology, Adventist University of the Philippines, Silang, Philippines 7 Department of Environmental Science, School of Science and Engineering, Ateneo de Manila University, Quezon City, Philippines 123 Conserv Genet Introduction protection is needed for the remaining B. hainesii indi- viduals as well as carrying out further research to deter- Mangroves are an intertidal forest ecosystem distributed in mine minimum viable population size. subtropical to tropical regions of the world (Tomlinson Although B. hainesii is a highly prioritized mangrove 1986; Polidoro et al. 2010; Spalding et al. 2010). The major species for conservation, its genetic background and the constituents of mangroves are woody plants with unique evolutionary relationships with other Bruguiera species are characteristics, such as aerial roots, viviparous propagules not well understood. Schwarzbach and Ricklefs (2000) and high tolerance to salinity, which are adapted to inter- provided the most comprehensive molecular phylogenetic tidal environment. 73 species and hybrids from 20 families study for the genus Bruguiera to date, but B. hainesii was are known as ‘‘true mangroves’’ (Spalding et al. 2010), and not included. In studies that discussed phylogenetic rela- they form mangrove forests at coastal zones in 123 coun- tionships within the genus based on morphological features tries and territories of the world. Despite the important of the species, the genus Brugueira is generally divided ecological services provided by the mangroves (Fosberg into two groups (Tomlinson 1986; Sheue et al. 2005; Duke 1971; Robertson and Duke 1987; Ong 1993; Primavera and Ge 2011). One group has larger leaves and larger 1998; Dahdouh-Guebas et al. 2005), they are threatened by solitary-flowered inflorescences and includes B. gym- land use change (Field 1998; Valiela et al. 2001; Alongi norhiza (L.) Lamk., B. sexangula (Lour.) Poir. and B. 2002). Several reports warn that 20–35 % of world’s exaristata Ding Hou. The other group has smaller leaves, mangrove area has been lost in the last two decades smaller and multiple-flowered inflorescences of relatively (Valiela et al. 2001; FAO 2007; Polidoro et al. 2010). This small size and includes B. cylindrica (L.) Bl. and B. critical loss of mangrove area may increase the extinction parviflora Wight & Arnold ex Griffith. Although some risk of mangrove species. Although the majority of man- authors have placed B. hainesii in the multiple-flowered grove species are widespread and not considered to be group (Duke and Ge 2011), this species exhibits an inter- threatened with extinction, 16 % (11 species) of the 73 true mediate state for these traits having larger flowers in mangroves are categorized as threatened by extinction multiple-flowered inflorescences (Hou 1957, 1958) (Polidoro et al. 2010). (Fig. 1). The intermediate morphological features of B. Bruguiera hainesii C. G. Rogers (Fig. 1) is one of the hainesii may imply the species is of hybrid origin. two mangrove species classified as ‘‘Critically Endangered In this study, we used molecular markers to analyse (CR)’’ within the IUCN Red List of Threatened Species genetic relationships between B. hainesii and all other (Duke et al. 2010). This species has a wide geographic Bruguiera species, especially in terms of hybrid origin. distribution extending from Myanmar and Thailand To understand the genetic background of the critically through the Malay Archipelago to Papua New Guinea endangered species is crucially important because hybrid (Tomlinson 1986; Sheue et al. 2005). However, fewer than taxa are not to be included on the IUCN Red List of 250 mature individuals are currently known, and the spe- Threatened Species (IUCN 2015). We used chloroplast cies is considered to be the rarest mangrove species DNA (cpDNA) and two single-copy nuclear DNA (Kochummen 1989; Sheue et al. 2005; Polidoro et al. markers to elucidate clear genetic relationships of the 2010). Polidoro et al. (2010) suggested that urgent species. Fig. 1 Flowers of six species of genus Bruguiera. a B. parviflora, b B. cylindrica, c B. hainesii, d B. exaristata, e B. sexangula, f B. gymnorhiza. Scale bar = 2cm 123 Conserv Genet Materials and methods Jaruwattanaphan et al. (2013). After separating each DNA band, we re-amplified the obtained DNA and performed Plant materials direct sequencing following the method described above. DNA sequences were aligned in MEGA6 (Tamura et al. We collected leaf samples of B. hainesii, and other five 2013) using the Clustal W algorithm (Thompson et al. species of genus Bruguiera from the localities listed in 1994) included in the software, and alignments were sub- Table 1. We used Rhizophora stylosa Griff. as an outgroup. sequently manually corrected. For R. stylosa samples, Leaf samples were dried using silica gel powder and kept sequences of trnS-trnG IGS could not be completely in plastic bags for subsequent DNA extraction. determined due to poly-A site located at about 300 bp from the trnG gene. Thus, only 300-bp from the trnG end of the DNA extraction sequence