PROCEEDINGS OF THE FIFTEENTH SYMPOSIUM ON THE NATURAL HISTORY OF THE BAHAMAS Edited by Robert Erdman and Randall Morrison Conference Organizer Thomas Rothfus Gerace Research Centre San Salvador Bahamas 2016 Cover photograph - "Pederson Cleaning Shrimp" courtesy of Bob McNulty Press: A & A Printing © Copyright 2016 by Gerace Research Centre. All rights Reserved. No part of this publication may be reproduced or transmitted in any form or by any means, electric or mechanical, including photocopy, recording, or any information storage and retrieval system, without permission in written form. ISBN 978-0-935909-16-6 The 15th Symposium on the Natural History of the Bahamas IDENTIFICATION OF PALMS ON SAN SALVADOR ISLAND IN THE GENERA COCCOTHRINAX AND LEUCOTHRINAX [FAMILY ARECACEAE] USING MOLECULAR METHODS; A PRELIMINARY REPORT Randall E. Cross1, Patricia Fuentes1, Tyler Jacobson1, Lee B. Kass2, and Anna M. Goebel1 1Department of Biological Sciences Florida Gulf Coast University 10501 FGCU Blvd. South, Ft. Myers, FL 33965 2L.H. Bailey Hortorium Department of Plant Biology Cornell University, Ithaca, NY, 14853 ABSTRACT cothrinax, and Thrinax as sister to Hemithrinax, both of which were not previously reported. A Two species of Coccothrinax (C. ar- combined approach of molecular and morphologi- gentata) and (C. inaguensis) are reported to occur cal characters is needed to better identify and dis- on San Salvador Island, the Bahamas, but it is not cover the relationships among the many species of clear whether they are indeed two separate spe- Coccothrinax. cies, or whether they are actually the same species and merely varietal forms or ecotypes. The high INTRODUCTION degree of morphological variation in the Coc- cothrinax and potential hybridization suggests that Coccothrinax Sargent (Sargent, 1899) is a molecular approach is necessary to better under- the most diverse and widely distributed genus of stand these palms on San Salvador. Hoping to use palm in the Caribbean (Roncal et al. 2008). Hen- the gene PRK (phosphoribulokinase-like protein derson (1995) recognized 14 species, but 64 spe- 2, exons 4 and 5) as a bar code for species identi- cies and subspecies are recognized in the World ty, we generated sequence data from ten speci- Checklist of Arecaceae (Govaerts et al., 2006, ac- mens representing what we identified as both typ- cessed 2014). Evidence of the diversity of the ical and atypical morphological forms of both C. group, and/or difficulty in identifying individuals argentata and C. inaguensis. The sequences were to the species level lies in the fact that an addi- compared to previously published C. argentata tional 42 species, subspecies, or varieties were and C. inaguensis sequences as well as nine other suggested at one time, but are not currently recog- Coccothrinax species or subspecies and speci- nized (ibid.). mens from four closely related genera. Using San Salvador Island is home to several Bayesian analyses we identified the genus Coc- dense populations of Coccothrinax that are be- cothrinax as monophyletic, but relationships with- lieved to consist primarily of the silver palm, C. in the genus were not resolved and the species argentata (Jacquin) L.H. Bailey (Bailey, 1939). identities of the Coccothrinax specimens we col- A second species, C. inaguensis Read (Read, lected was similarly not resolved. Three of the 1966), was reported by Smith (1982, 1993) to atypical specimens were identified as Leu- grow among populations of C. argentata on the cothrinax which we did not recognize from mor- southern end of San Salvador Island. However, phology perhaps due to their immature state and Kass and Smith (Kass, 1986, p. 66; Kass, 2009, p. their location within a Coccothrinax-dominated 130) hypothesized that C. inaguensis may be a community. The Bayesian analyses of published “varietal form” of C. argentata. Given the lack of data strongly supported Zombia as sister to Coc- clarity of the species occurring on San Salvador, 53 The 15th Symposium on the Natural History of the Bahamas our goal was to document the occurrence, distri- south Florida through the Bahamas, the Greater bution, and abundance of these two species or va- and, Lesser Antilles and occurs on San Salvador. rieties across San Salvador. The genus Coccothrinax is broadly distributed Coccothrinax individuals dominate throughout the Bahama archipelago, Caribbean, Coastal Coppice, Coccothrinax-shrub subcommu- South Florida, and Cuba. Many of the species are nities, and define these coastal habitats (Kass more narrowly distributed, being endemic to one 2009, p6; Smith 1993 pp. 4, 11). Our initial ob- or a few islands (Table 2). servations indicated that there were not only sev- Our attempts at field identification to spe- eral distinct dense populations of C. argentata at cies or varietal forms were frustrating due to the Rocky Point, Grotto Beach, and Sandy Hook as variability we observed in morphological charac- documented by Kass (2009), but the palms also ters. Of course we realized that our frustration occurred in low density populations or as isolated may have stemmed from our lack of competence individuals across broad regions of the island. and experience with the systematics of the taxa Thus the broad distribution of palms suggested involved (Cross and Goebel, pers. comm.). How- that gene flow could be occurring across much of ever, it was also possible that the published sys- the island and that the populations at Rocky Point, tematics (Henderson et al., 1995) of Coccothrinax Grotto Beach and Sandy Hook might not be as was lacking clear definitive diagnostic sterile genetically distinct as we first thought. characters, or that the current taxonomic status of When we started measuring densities, we the group is limited for sterile characters and often noticed that it was very difficult to determine the overlapping for reproductive characters. Examin- species identity of many specimens as either C. ing reproductive structures such as flowers or argentata or C. inaguensis using reported mor- fruits may have assisted in accurate identification. phological distinguishing characters such as the Recognizing limitations in morphological silvery color of the bottom leaf surface in C. ar- characters used to identify the species, we began gentata and the inverted umbrella-shaped leaves to use a molecular genetic approach to identify of C. inaguensis (Henderson et al., 1995). The species of Coccothrinax and to determine their variation in silver color was significant not only abundance and distribution on the island. A mo- between individuals but also among leaves in the lecular analysis might also inform us of good same individual. Similarly, the leaf shape (invert- morphological diagnostic characters and we report ed umbrella) also varied among geographically our first attempt here. A combined morphological close individuals, as well as within individuals. and molecular approach will certainly benefit not Kass previously questioned the use of overlapping only our understanding of the group on San Sal- morphological characters to discriminate between vador, but of palms throughout the Caribbean. these purported species (Kass 1986, p. 66; Kass 2009, p. 130; Kass, unpublished). Molecular Approach The high degree of morphological varia- tion in the Coccothrinax may be due to phenotyp- DNA barcoding (Hebert et al., 2003a, ic plasticity due to local environmental interac- Hebert et al., Ratnasingham et al., 2003b) is a mo- tions producing ecotypes, or it may be due to dis- lecular approach to identifying species by com- tinct genetic lineages (species or varietal types) paring a genetic marker of an unknown individual occurring sympatrically, or possibly due to hy- to a genetic database composed of organisms bridization among varietal types or species. Po- whose species identities are known. The rbcL and tential intrageneric hybridization has been sug- the matK genes combined have been suggested as gested (Henderson et al. 1995) and intergeneric appropriate genes to use as barcodes in flowering hybridization with Leucothrinax has been reported plants (CBOL Plant Working Group, 2009; Au- by Nauman (1990) in South Florida. Leucothrinax subel, 2009) as have the nuclear internal tran- morrisii H. Wendl. (formerly Thrinax morrisii, see scribed spacer region and the plastid trnH-psbA Lewis and Zona 2008) is broadly distributed from intergenic spacer (Kress et al., 2005) or other 54 The 15th Symposium on the Natural History of the Bahamas combinations of the four genes (Kress and Erick- and placed in the annex herbarium of the Baha- son, 2007). However, only the two genes PRK mas National Herbarium (BNH) at the Gerace Re- (phosphoribulokinase-like protein 2, exons 4 and search Centre (Kass et al. 1998). We also exam- 5) and RPB2, (subunit 2 of RNA polymerase II) ined five atypical appearing specimens growing have sufficient DNA data within databases (Gen- among typical appearing C. argentata from Sandy Bank and EMBOL) to differentiate between mul- Hook and Rocky Point populations (Table 1). tiple species of Coccothrinax (Roncal et al., Specimens were noted as atypical if they had unu- 2008). We chose to look at the PRK DNA region, sually shaped or unusually large leaves compared as a first attempt at species identification because to individuals growing within a distance of one it was technically easier to obtain sequence data meter and/or the length of the inflorescence was from than RPB2. unusually long compared to the leaves. Roncal et al. (2008) provided a phylogeny of the Caribbean palms (Arecaceae) that included Table 1. New DNA sequences obtained from nine species of Coccothrinax, as well as individu- San Salvador Island. als of the genera Hemithrinax, Thrinax, Leu- cothrinax, Zombia, Schippia and Cryosophilia and Sample Locality on Sequence lat/long seven other genera in the tribe Cryosophileae. We name San Salvdor info used a subset of their data with the goals of de- Coccothrinax argentata: termining the species identity of the palms on San 1. RP1 Rocky Point 24.10691 uni- Salvador, as well as identifying whether the palms 74.518582 directional might be distinct lineages, ecotypes, or possibly 2.