Talita Vieira-Pinto DIVERSIDADE DAS ALGAS CALCÁRIAS CROSTOSAS DO BRASIL BASEADA EM MARCADORES MOLECULARES E MORFOLOGIA DIVERSITY OF CRUSTOSE CORALLINE ALGAE FROM BRAZIL BASED ON MOLECULAR MARKERS AND MORPHOLOGY São Paulo, SP 2016 Talita Vieira-Pinto DIVERSIDADE DAS ALGAS CALCÁRIAS CROSTOSAS DO BRASIL BASEADA EM MARCADORES MOLECULARES E MORFOLOGIA DIVERSITY OF CRUSTOSE CORALLINE ALGAE FROM BRAZIL BASED ON MOLECULAR MARKERS AND MORPHOLOGY Tese apresentada ao Instituto de Biociências da Universidade de São Paulo, para a obtenção de Título de Doutor em Ciências, na Área de Botânica. Orientador(a): Mariana Cabral de Oliveira Colaborador(a): Suzanne Fredericq São Paulo, SP 2016 Vieira-Pinto, Talita Diversity of Crustose Coralline Algae (CCA) from Brazil based on molecular markers and morphology Número de páginas Tese (Doutorado) - Instituto de Biociências da Universidade de São Paulo. Departamento de Botânica, 2016. 1. Crostose coralline algae 2. DNA 3. Taxonomy I. Universidade de São Paulo. Instituto de Biociências. Departamento de Botânica. Comissão Julgadora: Prof(a). Dr(a). Prof(a). Dr(a). Prof(a). Dr(a). Prof(a). Dr(a). Prof(a). Dr(a). Orientador(a) Dedication To my family; To honor the memory of Dr. Rafael Riosmena-Rodriguez Acknowledgments I would like to thank Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP for providing me two scholarships (Proc. FAPESP 2012/0507-6 and 2014/13386-7) and for funding our research and courses and meetings I attended along these years. I would also like to thank my admirable advisor, Dr. Mariana Cabral de Oliveira for all she did for me and all she does to Brazilian science – you are a true inspiration and a whole model. I also thank Dr. Suzanne Fredericq, who welcomed me in her laboratory at the University of Louisiana at Lafayette and who made possible for me to live, learn and experience a lot more than I never thought I would in my whole life. I’ll be forever grateful for all these two great women did for me. Dr. Paulo Horta, for always being there for me, for guiding me in the first steps of this path and for being such a believer – making of us, his students, to believe too. Dr. Rafael Riosmena, a kind and generous soul that shared so much knowledge with all of us corallines enthusiastic and unfortunately left us too soon – Rest in Peace Prof Rafa. I acknowledge all my lab mates, Professors and technicians from LAM (Laboratório de Algas Marinhas “Édson José de Paula - University of São Paulo) from present and past, an extensive list of awesome people, who were very important and helped me in so many ways daily. Thanks to you all! I especially thank Carolina Azevedo, Carolina Franco, Mariana Mungioli, Natália Guimarães, my closest friends, for all the emotional support along these years. My lab mates from Dr. Fredericq’s Seaweed Lab (University of Louisiana at Lafayette - ULL), Joe, Dago, Thomas, Olga, Will and Sherry, awesome people that I have the pleasure to meet and learn a lot from. I especially thank Dr. Joseph Richards (or simply Joe) for his generosity and patience, and most of all, for all the time he spent helping me – I will never be able to thank you enough mate; you are a true professor and lucky will be your future students. I thank all the good friends I made along the way, people that I’ve had the pleasure to meet during conferences, workshops and courses I had the opportunity to attend, and with whom I learned a lot from; they are a lot of people and I would not be able to name each one, but especially want to thank some of them: Una, Aleisha, Mauricio, Katrina, Laurie, Lynn and Marycruz. Marina Sissini and Eduardo Bastos for sending me samples; I also thank Sisbiota Mar for the samples they sent me. Dr. Pablo Riul and Ronei Sarti for helping with the maps. Dr. Paul Gabrielson for giving me the topotype sample of Sporolithon episporum. Dr. Tom Pesacreta and Mike Purpera for letting me use the SEM and the facilities in the microscopy center at the ULL. All the security guards and staff and the Professors of the Biosciences Institute, especially from the Botany department from the University of São Paulo, who gave me support in many ways. The University of São Paulo itself. By all means, it was an honor walking through this campus, these buildings, these Institutes and classrooms for the last few years. My parents Maria and Marcio, my sister Gladys, my brother-in-law Cyro and my brother Cristiano. Without them, their support and endless love I would be NOTHING. I’ll be forever grateful for all the sacrifices you made to raise me. Not everybody knows, but my parents could never actually go to elementary school, but they struggled and battled for their children to have a better education and here I am, almost a Doctor, thanks to them (and to my sister who paid for my English course, who taught me to read and write at the age of 4 and who kept checking my lessons and grades over the years). And finally, I thank my husband Ivan Martins, for always being by my side, for being the best friend and supporter somebody could ask for and for the family we are building together – you and our precious baby mean everything to me. Thank you for your love and for sharing your life with me. Summary Resumo 01 Abstract 02 Introdução Geral * 03 Referências* 07 Chapter 1. Diversity and distribution of CCA along the Brazilian 10 coast with emphasis on Corallinales and Hapalidiales Introduction 11 Aims 34 Material and methods 34 Results 56 Discussion 142 Final remarks 165 References 167 Chapter 2. Diversity of the shallow waters Sporolithon 186 (Sporolithales, Rhodophyta) from South Atlantic coast Abstract 187 Introduction 188 Material and methods 190 Results 193 Discussion 198 Conclusions 200 References 201 Chapter 3. Molecular and Morphological Diversity of 225 Lithothamnion spp. Rhodoliths (Hapalidiaceae, Hapalidiales) from Deepwater Rhodolith Beds in the Northwestern Gulf of Mexico Abstract 226 Introduction 227 Material and methods 230 Results 235 Discussion 251 References 266 Conclusões gerais* 314 Appendix I - List of samples collected along the Brazilian coast. CD Appendix II - List of sequences downloaded from Genbak. CD Appendix III - Distance matrix - UPA CD Appendix IV - Distance matrix – rbcL-3P CD Appendix V - Distance matrix – COI-5P CD * Por exigência do programa de pós-graduação, como a tese foi redigida em Inglês, a introdução geral e conclusões gerais foram redigidas em português. Appendix VI - Distance matrix – psbA CD Appendix VII - UPA alignment matrix CD Appendix VIII - rbcL-3P alignment matrix CD Appendix IX - COI-5P alignment matrix CD Appendix X - psbA alignment matrix CD Appendix XI - Distance matrix UPA CD Appendix XII - Distance matrix psbA CD Appendix XIII - Distance matrix COI CD CD with pdf files Back cover Figure index Chapter 1. Diversity and distribution of CCA along the Brazilian coast with emphasis on Corallinales and Hapalidiales Fig. 1. Main morphological habits that coralline algae can 13 exhibit. Fig. 2. Spectrum of shapes and ramification exhibited by CCA. 14 Photos from D. Littler (Littler & Littler, 2013). Fig. 3. Simplified general sexual cycle/life history of the CCA 16 (adapted from Harvey 2005). The images were all newly generated in this study and are only representative, variations may occur in different genera. Fig. 4. Distribution of CCA species referred to Brazilian coast. 33 Fig. 5. Sample sites along the Brazilian coast and surrounding 35 Oceanic Islands. Fig. 6. Phylogram based on NJ analysis of UPA sequences. 59 Fig. 7. Phylogram based on NJ analysis of rbcL-3P sequences. 63 Fig. 8. Backbone of the phylogram based on ML analysis of 69 COI-5P sequences. Fig. 9. Detail of the Corallinales clade from the phylogram 71 based on ML analysis of COI-5P sequences. Fig. 10. Detail of the Hapalidiales clade I from the phylogram 73 based on ML analysis of COI-5P sequences. Fig. 11. Detail of the Hapalidiales clade II from the phylogram 74 based on ML analysis of COI-5P sequences. Fig. 12. Backbone of the phylogram based on ML analysis of 80 psbA sequences. Fig. 13. Detail of the Corallinales – Neogoniolithon clade from 82 the phylogram based on ML analysis of psbA sequences. Fig. 14. Detail of the Corallinales clade II from the phylogram 83 based on ML analysis of psbA sequences. Fig. 15. Detail of the Hapalidiales from the phylogram based 85 on ML analysis of psbA sequences. Fig. 16. Corallinaceae sp. 2; specimen IBC1228 – External 92 morphology and vegetative features. Fig. 17. Corallinaceae sp. 2; specimen IBC1228 – 94 Reproductive features. A-D. Fig. 18. Corallinaceae sp. 4; specimen IBC1222 – External 96 morphology and vegetative features. Fig. 19. Corallinaceae sp. 4; specimen IBC1222 – 98 Reproductive features. A-B. Fig. 20. Corallinaceae sp. 5; Specimen IBC1737 – External 101 morphology and vegetative features. Fig. 21. Corallinaceae sp. 5; specimen IBC1737 – 102 Reproductive features. A-B. Fig. 22. Lithophyllum sp. 3; specimens IBC1726 and IBC1878 104 – External morphology, vegetative and reproductive features. Fig. 23. Corallinaceae sp. 6; specimen IBC1917 - External 107 morphology and vegetative features. Fig. 24. Corallinaceae sp. 6; specimen IBC1917 – 109 Reproductive features. Fig. 25. Corallinaceae sp. 8; specimen IBC1919 - External 111 morphology and vegetative features. Fig. 26. Corallinaceae sp. 8; specimen IBC1919 – 112 Reproductive features. Fig. 27. Neogoniolithon sp. 1; specimen IBC1216 - External 114 morphology and vegetative features. Fig. 28. Neogoniolithon sp. 1; specimen IBC1216 – 115 Reproductive features. Fig. 29. Lithothamnion sp. 1; specimens IBC1704, IBC1564 118 and IBC1557 – External morphology. Fig. 30. Lithothamnion sp. 1; specimens IBC1704 and 119 IBC1907 - Vegetative features.