This study aimed to characterize the genetic diversity of the Passiflora genus in Colombia using molecular markers. The study: 1) Analyzed 213 Passiflora individuals representing 151 species and 15 subgenera using chloroplast and mitochondrial DNA markers. This identified 280 chloroplast and 372 mitochondrial haplotypes. 2) Chloroplast DNA provided better resolution of relationships than mitochondrial DNA, separating the Decaloba subgenus and revealing geographic structuring of some groups. 3) Results only partially agreed with morphology-based taxonomy, suggesting divergence between genomes and mixed modes of organelle transmission in the genus. The study helped elucidate the evolution and systematics of Passiflora but also highlighted remaining questions

1. Study of the genetic diversity of the genus Passiflora L. and its distribution in Colombia John Ocampo École Supérieure d’Agronomie de Montpellier Université de Montpellier II Ph.D. ocampo.john@gmail.com

3. Geo COPPENS d’EECKENBRUGGE

5. Xavier SCHELDEMANGinés-Mera Fellowship

7. Passion fruit priorities: - Collection and characterization - Taxonomy problems - Conservation problems

8. General objective To study the genetic diversity of the genus Passiflora L. and its biogeographic distribution in Colombia.

10. To characterize the morphologic and molecular diversity at inter- and intra-specific levels.

12. 1. Passifloraceae family Taxonomy 18 genera ca. 650 species America Africa Asia Killip (1938) 22 subgenera GenusPassifloraL. ca. 573 species America Asia Feuillet & McDougal (2003) 4 subgenera Astrophea DecalobaDeidamioides Passiflora

15. Psilanthus

16. Murucuja

17. Pseudomurucuja

18. Apodogyne- Climbers - America South Asia and Australia - 2n = 12, 24, 36, 72 Killip (1938)

20. Tacsonia

21. Distephana

22. Manicata

23. Dysosmia- Climbers Killip (1938) - American - 2n = 18, 20 (Dysosmia)

25. Tryphostemmatoides- Climbers - American - 2n = ?? Killip

28. Pollinationsyndromes

29. 3. Distribution of thegenusPassifloraL. 22 species Subg. Decaloba

30. 4. Uses Other… Ornamental Food Pharmacopeia Liquors Medicinal

31. 5. Cultivatedspecies in Colombia

32. Economic importance – International markets P. edulis f. flavicarpa Yellow maracuja - Maracuyá P. ligularis Sweet granadilla P. edulis f. edulis Purple maracuja - Gulupa

33. Economic importance – International markets P. tripartita var. mollissima Curuba de Castilla P. tarminiana Curuba India P. maliformis Cholupa

34. Economic importance – International markets P. quadrangularis Giant granadilla - Badea P. popenovii Granadilla de Quijos P. alata Maracuja doce

35. Study Components I- Diversity distribution in Colombia (GIS) 1.a. Biogeography and conservation 1.b. Potential distribution and adaptation II- Morphological diversity (descriptors) III- Genetic diversity (CAPS – DNAcp/mt)

36. I – Diversity Distribution in Colombia (SIG) 1a. Biogeography and conservation of Colombian Passifloraceae .

37. Reported species HERBARIA National PSO - HAU - COL - FAUC - CUVC - FMB VALLE - MEDEL - CHOCO – TOLI - CAUP AFP - UIS – CDMB - COAH – JAUM - HUQ SURCO Foreign P : Museum d’Histoire Naturelle – Paris – France MO : Missouri Botanical Garden – USA MA : Real Jardin Botánico – Madrid – Espagne K :Kew Garden – London - Grande-Bretagne NY : New York Botanical Garden– USA LITERATURE Killip (1938, 1960), Escobar (1988, 1989) etc. 3.375 georeferenced data

38. Maximum Diversity in the American Andean Region, especially in Colombia and Ecuador Diversity Centre Diversity distribution of the American Passion Fruits 3.375 georeferenced data 16 4 23.5º N 167 reported species (26 new species for the country) Tropic of Cancer 20 14 20 70 15 17 10 10 18 55 25 13 78 31 48 53 30 32 36 167 0º 90 127 73 48 Tropic of Capricorn 2 16 -23.5º S 19 3

39. Species spatial distribution High density in the central mountain region. Low density North-East and South of the Andes. Very marked low density in lowlands, especially in the Orinoquian and Amanzonian regions. DIVA-GIS software

41. Access permits (MMA - Ministerio del Medio Ambiente)

42. Public order555 collections from 17 of 32 departments TOTAL : 3.930 OBSERVATIONS

43. Elevation Species distribution by Biogoegraphic region Caribbean (38) Diversity concentrated in the Andean Region Pacific (36) Andes (123) Orinoquia (19) Amazonia (44)

44. Pacific Andes Caribbean Amazonia Orinoquia Colombian Biogeography & Passifloracea Species

45. Passifloraceae distribution in Colombia (Maximum Distance vs. Circular Area) Species of continental distribution Species of regional distribution and common endemic species R2= 0.779 P. vitifolia P. foetida P. auriculata P. quadrangularis P. nitida P. maliformis P. mollis P. mixta Endemic species P. cumbalensis P. menispermifolia P. ligularis P. suberosa P.arborea P. oerstedii P. capsularis P. seemannii P. coriaceae P.misera P. biflora P. spinosa P..coccinea P. rubra P. antioquiensis P. micropetala P. serratodigitata P. involucrata D. parviflora 1000 400

47. Maximum Distance MaxD

48. Number of records

49. Date of recordsUICN Categories EX:Extinct EW:Extinct in the wild CR:Critically endangered EN:Endangered VU:Vulnerable NT:Near threatened LC:Least concern DD:Deficient Data

50. 1. Diversity and potentiel distribution 1b. Diversity distribution and in situ conservation of the Colombians Passifloraceae.

51. Software DIVA-GIS Speciesrichness and diversity Herbarium data: 3.330 Literature data: 45 Collection data: 555 3.930 georeferenced data

52. 1 2 Nine ultra- diverse zones in the Andes, four sub-explored (1, 6, 8, 9) 3 4 Species richness 5 6 7 8 9 Species potential distribution “Hotspot” Software DIVA-GIS

53. Protectedareas In situ conservation “Hotspot”

55. Crop systemsSoftware DIVA-GIS

57. Nine ultra-diverse (hotspot) and non-protected zones in the Andes.

58. There is no geographic relation between richness and endemism.

59. There is a need for urgent conservation strategies.

62. Tenerife (El Cerrito – Valle del Cauca) – 2800 m

63. El Moral (El Cerrito – Valle del Cauca) – 2400 m

64. San Joaquín (El Tambo – Cauca) – 1800 m

65. Paraguacito (Buenavista – Quindío) – 1250 m

66. Arauca (Risaralda – Caldas) – 1000 m

67. In situ (collections)

68. Plant material 124 accessions 60 species 9 subgenera (Killip, 1938)

69. List of 127 descriptors

70. Relative variance components – Quantitative characters 50

71. d. Factor loadings from a Principal Component Analysis - PCA COMPONENTS Descriptors 1 2 3 4 5 ACP - Factor loadings 83% total variance -0.088 0.215 -0.023 0.041 -0.892 STDI 0.759 LEMS 0.353 0.365 0.152 0.041 LELC 0.019 0.571 -0.686 0.009 -0.195 -0.017 0.040 0.011 0.014 LENN 0.962 PENN 0.648 0.407 0.013 -0.055 0.158 0.135 -0.287 0.010 0.145 PDDI 0.847 PDLF -0.943 -0.054 -0.136 -0.007 -0.044 -0.054 -0.113 -0.006 -0.046 PDBS -0.973 BRLR 0.723 0.296 0.186 -0.091 0.240 FLPL 0.774 0.501 -0.021 0.235 0.042 Cp.1: Flower length 0.595 -0.024 0.095 0.028 FLPW 0.713 FLSL 0.716 0.577 0.036 0.243 -0.028 0.520 0.034 0.074 -0.108 FLSW 0.754 Cp.2: Flower width and bract shape FLLE 0.960 0.179 0.045 0.017 0.090 0.063 0.086 -0.100 0.119 0.904 FLHL FLHL 0.824 0.208 0.026 -0.080 -0.137 FLCN 0.723 -0.521 0.034 -0.007 -0.329 Cp.3: Stem with, peduncle branching and leaf length 0.529 0.651 0.082 0.097 0.171 FLFL FLSF 0.677 0.658 0.076 0.061 -0.049 0.454 0.693 0.067 0.073 0.263 FLOL 0.064 0.047 0.033 0.099 0.964 FLSL 0.236 0.099 0.363 0.214 0.711 FLOP Cp.4:Number of nectaries on leaf margin -0.160 -0.221 -0.131 0.241 BRWI/BRLE 0.718 0.699 FLNC/FLHD 0.056 0.042 0.043 0.368 Expl.Var 7.610 6.496 3.336 1.256 1.238 Prp.Totl 0.317 0.271 0.139 0.052 0.052 Cp.5:Leaf serration % total varianza 46.028 16.999 11.138 4.913 3.986

72. Manicata Distephana PCA – Three-dimensional plot of the scores for Passiflora accessions Tacsonia Passiflora Psilanthus Dysosmia Decaloba Tryphostemmatoides Astrophea

73. 32 Discriminant qualitative characters between subgenera

74. e. Qualitative data dendrogram (nNeighbour Joining, 32 characters) n = 12 n = 6 n = 10 n = 9 n = 6 n = ?? Distances Sokal & Michener

75. Conclusions The new descriptors list has permitted to rank species together with taxonomy in a coherent way. However, there are some exceptions. The analysis is coherent with Killip's classification (1938) at the subgenus level. Descriptors related to the flower permit a higher discrimination between subgenera and species.

76. III. Genetic diversity (CAPS – DNAcp/mt)

78. 151 species

80. Passion Fruit National Collections – Blois (France) – 22 %

81. Royal Botanical Garden – Meise (Belgium) – 2 %

83. Outgroup Previousstudies PASSIFLORA Yockteng & Nadot (2004) Nuclear Sequences ncpGS - 91 species - 17 subgenera (Killip, 1938) DECALOBA ASTROPHEA

84. Previousstudies PASSIFLORA Hansen et al. (2006) Chloroplas Sequences TrnL / TrnT - 57 species - 16 subgenera (Killip, 1938) DECALOBA outgroup ASTROPHEA

85. Cytoplasmatic genome inter-specific variation Chloroplast 280 haplotypes Mitochondria 372 haplotypes Chlorotypes are easier to interpret than mitotypes

86. Observed variation per primer-enzyme combination (haplotypes) Mitochondrial Chloroplastic

87. AFTD of Chloroplastic data OUTGROUP

89. Passiflora - little structure

90. Astrophea

91. TryphostemmatoidesNJ, distance de Sokal & Michener

94. The data set must be interpreted taking into account the geological context of tropical flora separation and the formation of the Andean mountain range.

96. Thanks!