THE POPULATION GENETICS OF TWO TEMPERATE RAINFOREST TREES, LAGAROSTROBOS FRANKUNll (Hook f.) Quinn (HUON PINE), AND ATHEROSPERMA MOSCHATUM Labill. (SASSAFRAS). BY ALISON SHAPCOTT Submitted in fulfilment of the requirements for the degree of Doctor of Philosophy. University of Tasmania Hobart. July 1993 DECLARATION: This thesis contains no material which has been accepted for the award of any other degree or diploma in any University, and contains no copy or paraphrase of material previously written by another person, except where due reference is made in the text ~~ A. Shapcott. page ii ACKNOWLEDGEMENTS : I would like to thank the following people: - My University supervisors, Jim Reid and Jamie Kirkpatrick; - Members of my NRCP steering committees, Mick Brown, Steve Harris, John Hickey; and Jayne Balmer, for their advice and encouragement; - Georgina Davis for field assistance and artwork; - Gavin Moran for electrophoresis recipes; - Mike Battaglia, Brad Potts, Greg Jordan, Leon Barmutta, and Rob Wiltshire for statistical and computing assistance, and Myron Zalucki(Qld Uni), Jane Hughes, Doug Ward and Mark Kingston (Griffith Uni) for computer programs, and Carla Catteral for computer access; - Murray Baseler, Islay Robertson, Peter Bird, Peter Brown, Parks and Wildlife archaeologists, Johnathon Marsden-Smedley, Mike Peterson, Richard Shapcott and Isabel Shapcott Mark Neyland, Phil Barker, Jenny Read, Brian Thompson, John Hunter, W. Jones, and others, for collections, site locations, or field assistance; - Jean Jarman, Kristen Williams, Rene' Vaillancourt and many others. I would also like to thank the Department of Plant Science at the University of Tasmania for the use of facilities and the Department of Parks and Wildlife Hobart for financial support as well as the Forestry Commission Tasmania, Department of Conservation and Environment Victoria, and the N.S.W. national Parks and Wildlife Service for co-operation. I gratefully acknowledge the National Rainforest Conservation Program for funding of both my salary and project costs, without which this thesis would not have been possible. page iii TABLE OF CONTENTS: Acknowledgements iii Abstract 1 General Introduction 3 Chapter 1 : Seedfall in Huon pine its dispersal and establishment. 11 Introduction 11 Methods 12 Results 15 Discussion 23 Chapter 2 : Reproductive, sex and size structure of Huon pine stands. 28 Introduction 28 Field Methods 30 Data Analysis 32 Results 36 Discussion 45 Chapter 3 : Population genetic analysis of Huon pine sites. 54 Introduction 54 Field Methods 55 Laboratory Methods 57 Statistical Methods 59 Results 64 Discussion 82 Chapter4: Variation within and among Atherosperma moschatum populations.89 Introduction 89 Field Methods 91 Electrophoretic Methods 92 Statistical Methods 99 Results 102 Discussion 118 Chapter 5: The spatial genetic structure of Atherosperma moschatum stands. 126 Introduction 126 Methods 127 Results 130 Discussion 137 General Discussion 141 References 154 Appendix 1 : Study sites 172 Lagarostrobos franklinii sites 172 Atherosperma moschatum sites 180 Appendix 2: Huon pine height distributions at the study sites. 186 Appendix 3 : Huon pine genotype frequencies 188 Appendix 4: Sassafras genotype frequencies 191 page iv ABSTRACT: The population genetics of two temperate rainforest tree species endemic to south eastern Australia were studied. Both species are long-lived and members of ancient families. There are parallels between the two species even though one was a gymnosperm and the other an angiosperm. For example, both species reproduce both vegetatively and sexually. Lagarostrobos franklinii (Huon pine) (Podocarpaceae) is mostly dioecious and wind pollinated, while Atherosperma moschatum (sassafras) (Monimiaceae), is monoecious or dioecious and insect pollinated. Both have potential for long distance seed dispersal, L.franklinii by water and A. moschatum by wind. The population genetics of both species was studied from stands throughout their geographic range using isozyme analysis. Most genetic diversity was found within rather than among sites. Genetic diversity among sites was low but generally consistent with expectations for each species (Hamrick and Godt 1979). Atherosperma moschatum had much more diversity among sites than Huon pine, with its mainland sites differentiating significantly from its Tasmanian ones. In Huon pine, most differentiation was found in isolated sites. Diversity within sites was also low in Huon pine but was much greater in sassafras. The structure of genotypes within stands was examined using spatial autocorrelation. In both species trees of like genotypes were found to be clustered at short distances. This genetic substructuring was found regardless of population size, density, distance from other stands, level of inbreeding, history, etc. Most sites deviated from Hardy-Weinberg expectations with deficiencies of heterozygotes, and high levels of allelic fixation, and were effectively inbred. The size structure and floristics within stands were investigated and used to assist in the interpretation of the patterns of genetic variation, inbreeding and stand dynamics found in each species. There was much variation in size structures and regeneration modes between sites in both species and neither appeared to require large scale disturbances for regeneration. The two species varied in the relationships between site environmental/ecological similarity and genetic similarity. In both species there was as much diversity in genetic variability and size structure in small isolated stands as there was in stands within larger assemblages. page 1 The proportion of trees contributing to the reproductive population, as well as the proportion of each gender type within that population, were estimated for Huon pine stands. On average thirty percent of Huon pine trees greater than one metre tall were reproductively active in the mast year recorded, and overall there were equal proportions of male and female trees. The relationships between reproduction and gender expression, with size structure, density, floristics, inbreeding and genotypes were investigated. Stands were also compared to identify if there were geographical or climatic trends in the distribution of these characteristics. Reproduction was found to increase with increasing tree size and also with more open canopies. Sites with similar proportions of females were found to also have similar species compositions. The distribution of reproductive trees and gender types within stands was investigated using spatial autocorrelation. The results were compared with genotypic distributions within the same stands. Although there was no direct correlation between gender type and genotype, both genotype and gender type were clustered at the same spatial scale, suggesting that such clustering may have a strong vegetative component. Huon pine seed production was estimated at one site and seed dispersal was investigated. Very large quantities of seed were shed. Seed dispersal laterally was negligible, but potential for dispersal down water courses was great as it stayed afloat for extended periods. Huon pine seed germination was investigated both in the field and under experimental conditions. Germination generally was slow, and with a low success rate. However seed in the field germinated at particular daylengths (regardless of temperature) in two consecutive seasons. Both species showed evidence that vegetative reproduction and localised pollen and seed dispersal have led to the development of family clusters, leading to inbreeding, and local fixation of allelic proportions. However infrequent long distance gene flow has probably reduced population differentiation. The population viability of each species was discussed. page2 GENERAL INTRODUCTION : The conservation and maintenance of biodiversity implies not only the maintenance of species and ecosystem diversity, but the diversity within species (Hopper and Coates 1990, Lacy 1988). The diversity within species may give flexibility to adapt to changing conditions and to survive major perturbations such as disease, or may be the source for future new species (Ellstrand and Antonovics 1985). In this era of increased domestication and genetic manipulation of species, it is also important that the range of genetic variation in natural populations be conserved and understood (Brown 1978, Hopper and Coates 1990). As there is not time to study each species, conserving and understanding the dynamics of, for example, major canopy or habitat forming species may ensure survival of much of their dependent biota (Boyce 1992). In order to determine the viability of a species, it is important to understand how the species functions at many levels, from individuals, to stands and populations and how each level interacts (Boyce 1992). Viable populations can adapt to changing conditions and recover or re-establish after major perturbations (Shaffer 1987, Boyce 1992). Information that may assist in the determination of species/population viability includes its regeneration patterns and requirements, and age structures (Boyce 1992, Lande 1988). Regeneration is influenced by ability to reproduce, the relative importance of sexual and vegetative reproduction and by seed germination requirements (Lande 1988, Harman and Franklin 1989). The viability and survival of offspring may depend on their genetic makeup, especially if the species is under strong selection from a changing environment (Brown 1989, Coates 1992, Ennos 1989, Muona