Proposed Conservation Strategies for Diospyros celebica

1. Proposed Conservation Strategies for Diospyros celebica Group 1 • Prasit • Amy • Aileen • Zue • Rao • Salwana • Tedi • Tri • Zhuo

2. Presentation Outline • Background/Rationale • What are the critical biological information (BI) needed? • How can we generate the BI? • How can we translate these BI to environmental conservation strategies?

3. D. celebica

4. Taxonomy Kingdom PlantaeDivision MagnoliophytaClass MagnoliopsidaOrder EricalesFamily EbenaceaeGenus Diospyros Species Celebica

5. Population Status and Trends • Once a widespread species in Sulawesi, it is now comparatively rare, especially in the South • Exported since 18th century • When in forests, D. celebica tends to scatter irregularly Status of origin: Endemic to Sulawesi (Minahasa and Bolaang Mongondow/North Sulawesi; Parigi, Poso, Donggala, Toli-toli, Kolonodale and Luwuk/Central Sulawesi; Maros, Barru, Luwu and Mamuju/South Sulawesi) Floristic element : Eastern provinces ofMalesian element Major ecological region: Sulawesi

6. Rationale • Rare plant conservation programs must be guided by the species biological attributes. • Ecological and genetic processes often interact synergistically to influence the population viability and to determine the persistence of populations in the long run. • Conservation has a cost and the resources available for conservation programs are always limited. Thus, CS must not only be scientifically justified but also practical in terms of resource availability.

7. What Are The Critical BI? ECOLOGY GENETICS • Distribution & habitat • Demography • Germination • Phenology • Level of genetic diversity • Spatial genetic structure • Population differential • Mating system • Minimum population size

8. Ecological Distribution & Habitat Ecological interaction between plants and their environment can influence population growth rates via their effects on fecundity, growth, or survivorship of individuals (Blundell and Peart, 2001; Peters, 2003) The studies on population dynamics and demography patterns will lead to a better understanding of the natural processes that operate within the population

9. Distribution and Habitat

10. Legend : Natural distribution of D. celebica in Indonesia (Overall natural distribution)

11. Conservation status & measures • IUCN status available of the Vulnerable (VU) (International Union for Conservation Red Data Book, 1978) • In Sulawesi, D. celebica is protected and there is a quota system in place. The Indonesian government has already started a planting program. It has not, however, been planted on a large commercial scale

12. D. celebica is threatened by heavy exploitation since it is an important source of streaked ebony Threats & Utilization

13. D. celebica timber is used for piano keys, carvings, brush backs, inlaying, and parts of stringed instruments.

14. Germination Study Population Survey 1 5 Ecological Approach In Conserving Ebony 2 Phenology Spatial Distribution 4 3 Population Dynamics & Demographic Studies

15. 1. Population Survey Study Plot 200m • Tagging • Diameter measurement (DBH> 5cm) • Mapping & coordinate using GPS & GIS 200 m ? To know the population status and relative density of ebony in the area.

17. Involves population dynamics, species recruitment and mortality Demography Field survey Set-up plot Long-term & short-term survey

18. 2. Spatial Distribution Soil Information Climate Topography ? To get information on habitat preference of ebony (ridge, valley, slope) from established plot

20. Mt. Makiling Forest Reserve • 65 km SE Manila • shifting cultivation, burned and selectively logged • Mature secondary forest with natural mixed stand • Dominated by Celtis luzonica and Diplodiscus paniculatus • Tropical monsoon

22. Structural characteristics of main canopy species • 113 tree species and 6 palms, >5 cm dbh • Highest density - Celtis • Highest basal area -Diplodiscus • Max. dbh by Ficus (balete, strangling fig) • Max. mean height of 21m

23. 3. Pop’n Dynamics & Demographic Studies Regeneration Quadrat Plot • WHY??? • Monitor growth • Seedling recruitment and mortality of the seedling (3 years; short-term study 10m 3 . First year seedlings will be identified, marked & appearance will be recorded 10 m ? to know the changes taking place in the life cycle of ebony

24. 4. Phenology of Ebony Reproductive biology will determine gene flow (mating system, pollination, fruit dispersal, etc.) & recruitment rate of the species (Lee, 2006 personal comm’n) • STEPS: • Identify the ebony tree from the established plots/ • Check and measure every month • Do some ranking such as budding stage, peak bloom and mature seed of ebony. ?

25. Pollination Biology Observation Pollinators Study on Pollination Biology Conservation strategy Wind Conserve animals that are seed dispersal agents Dispersal strategy Observation Animals

26. Regeneration & species role • Flowering and fruiting occurs at the age of 5-7 years • Seeds remain viable for only a short time • Seeds vectors: • Bats • Birds • Monkeys • Found with Homalium celebicum

27. Germination study From established plot, set up seed trap for seedfall study of ebony Monthly seedfall collection will be done + seeds will be checked Correlation of seed weight to germination capacity of ebony For mature & sound seeds, weighing & germination test will be done to compare growth & survival from natural forest condition ? To know the germination trait of ebony for ex-situ conservation.

28. In summary…

29. Rationale for Genetic Info • Allelic richness could contribute to population growth thru its effect on evolutionary potential, or the ability of a species to respond to changes in its selective environment. • Reduced heterozygosity can result in decrease of population growth due to inbreeding depression. • Therefore, we need to know the genetic diversity partition within and among populations

30. Levels of Genetic Diversity Minimum Population Size 5 1 Genetics Approach Conserving Ebony 2 4 Spatial Genetic Structure Mating System 3 Population Differentiation

31. 1. Levels of Genetic Diversity ? To estimate level of genetic diversity that we need to generate information aThe scoring scheme follows closely those in Hillis et al.2 and Karp and Edwards49.

32. Why Choose SSR Marker ? • SSR- (microsatellite) marker was developed by (Weber & May 1989 in human and found to be abundant in plant by Morgante & Olivieri 1993) • Very high degree of polymorphism & codominance make them extremely informative • Practical number of loci is 10 • Few as 5 or 6 microsatellite loci can often answer many conservation genetic questions (e.g. paternity, pollen flow) that cannot be answered with 30 or more isozyme loci • High reliability (reproducibility) • If the resources is limited, we can use allozyme

33. D.celebica leaf samples will collected from 40 populations base on 40 area divide based on soil, climate and slope • From each population, about 30 adults will be sampled • DNA genomic will be extracted using Murray and Thompson (1980) and purified • using High Pure PCR Template Preparation • Kit • DNA genomic will be extracted using Murray and Thompson (1980) and purified • using High Pure PCR Template Preparation • Kit • Develop primer • PCR amplification • For genotyping, PCR product will be electrophoresed along with GeneScan ROX 400 internal size standard using DNA automated sequencer • Develop primer • PCR amplification • For genotyping, PCR product will be electrophoresed along with GeneScan ROX 400 internal size standard using DNA automated sequencer Genescan analysis software and GENOTYPER software v3.7 Genescan analysis software and GENOTYPER software v3.7 How to Get The Parameters ? • D.celebica leaf samples will collected from 40 populations base on 40 area divide based on soil, climate and slope • From each population, about 30 adults will be sampled Sample Collection DNA extraction Microsatellite analysis Analysis data

34. Result of Genetic Diversity • Low genetic diversity means  not enough sufficient gene pool for short term adaptation and long term evolutionary Need to enhance by the introduction of new alleles through introgression following hybridization for long term conservation

35. Significantly structured Need sampling strategy for ex-situ conservation 2. Spatial Genetic Structure ? To determine genetic structure within a population Moran I Coefficient analysis statistical analysis • random • Need capture all • Choose and select

36. 3. Population genetic structure Erikson & Ekberg 2001

37. Analysis Data from SSR Analysis To determine coefficient of population differentiation. Gst<0.050 low genetic differentiation 0.050<Gst<0.15 moderate genetic differentiation 0.151<Gst<0.250 large gene differentiation 0.250<Gst very large gene differentiation

38. if high population differentiation • Mean: --low gene flow --high inbreeding --low genetic diversity --high variation among population more populations need to be conservation

39. If low population differentiation • Mean: --high genetic flow --high genetic diversity --low variation among population --Low population genetic differentiation among population implies no preference in identification of population for in-situ conservation or germplasma collection for ex situ conservation need more bigger area, and few population

40. 4. Mating System and Gene Flow OUTCROSSING RATE General description (Gregorius 1989) “Random mating, the environmental influence on mating events, selfing and the consequences of selfing and other forms of inbreeding, and incompatibility systems” Data analysis by using multilocus mating system program (MLTR) Ritland (1996)

41. Mating System Example: • Predominantly outcrossing • Selfing Ex-situ conservation (field genebank) Outcrossing : collect many seeds from a few mother trees Selfing : collect seeds from many mother trees)

42. 5. Minimum Population Size How many individuals in a conserved population are needed to maintain evolutionary potential of population and to resist to inbreeding depression from generation to next generation ?

43. N=Number of individuals in population t =Number of generation Ht=Heterozygosity at time t Most breeders agree that : A small loss of heterozygosity by 1% from one generation to next generation will not be significant for breeding , and does not result in inbreeding depression. In other word, we accept Ht+1=99%*Ht and then calculate N=50. So 50 individuals is often referred to as the basic rule of conservation genetics under the conditions of absence of selection, randomness of mating, each individuals in reproductive phase. Theoretically H selection Ht Ht+1 H mutation

44. Minimum Population Size In fact, a population is in selection and non-randomness of mating, which are very difficult to be quantified, so a population size of 500 reproducible individualsis hoped to represent a safety device and to provide a better chance for the restoration of genetic variation by mutation against the loss (Hattemer 2005). If a population has less than 500 individuals, maybe all of them should be conserved.

45. CONCLUSION ECOLOGY GENETICS • Distribution & habitat • Demography • Germination • Phenology • Level of genetic diversity • Spatial genetic structure • Population differential • Mating system • Minimum population size

46. THANK YOU K L Dr.Kelvinand Dr.Lee

47. Demography