* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download Population Dynamics of the Zuurberg Cycad and the Predicted
Plant evolutionary developmental biology wikipedia , lookup
Gartons Agricultural Plant Breeders wikipedia , lookup
History of botany wikipedia , lookup
Plant breeding wikipedia , lookup
Plant defense against herbivory wikipedia , lookup
Plant stress measurement wikipedia , lookup
Ecology of Banksia wikipedia , lookup
Plant nutrition wikipedia , lookup
Plant physiology wikipedia , lookup
Plant morphology wikipedia , lookup
Plant use of endophytic fungi in defense wikipedia , lookup
Flowering plant wikipedia , lookup
Biosequestration wikipedia , lookup
Ornamental bulbous plant wikipedia , lookup
Plant reproduction wikipedia , lookup
Photosynthesis wikipedia , lookup
Glossary of plant morphology wikipedia , lookup
Plant ecology wikipedia , lookup
Population Dynamics of the Zuurberg Cycad and the Predicted Impact of Climate Change By Karishma Singh Submitted in fulfillment of the requirements for the degree of Magister Scientiae in the Faculty of Science at the Nelson Mandela Metropolitan University January 2012 Supervisor: Prof E.E. Campbell Co-Supervisor: DR D.R. du Preez DECLARATION I, Karishma Singh (204043743), hereby declare that the dissertation for Masters in Science is my own work and that it has not previously been submitted for assessment or completion of any postgraduate qualification to another University or for another qualification. Karishma Singh Table of Contents........................................................................................i List of Figures.............................................................................................iii List of Tables...............................................................................................iv List of Plates.................................................................................................v Acknowledgements.................................................................................vi Abstract..........................................................................................................vii Chapter 1: Introduction.................................................................................1 Chapter 2: Literature Review......................................................................4 2.1 Cycad biogeography, morphology, anatomy and physiology, conservation and demography......................................................................4 2.1.1 Cones...............................................................................................5 2.1.2 Stem.................................................................................................6 2.1.3 Roots...............................................................................................6 2.1.4 Leaves.............................................................................................7 2.1.5 Toxicity............................................................................................8 2.1.6 Photosynthesis.............................................................................8 2.2 South African genera.................................................................................9 2.2.1 Encephalartos Lehm....................................................................9 2.2.2 Stangeria T.Moore......................................................................13 2.3 Pollination and Seed-dispersal..............................................................14 2.4 Propagation and Germination................................................................16 2.5 Conservation Status.................................................................................17 2.6 Demography...............................................................................................19 2.7 Climate Change.........................................................................................20 2.2.1 The causes and effects of Climate Change..........................20 2.7.2 What does climate change mean for C3 plants?..................22 Chapter 3: Materials and Method............................................................23 3.1 Demography...............................................................................................23 3.1.1 Study Sites...................................................................................23 3.1.2 Field Measurements...................................................................28 3.1.3 Data Analysis...............................................................................39 3.2 Germination................................................................................................30 3.3 Photosynthetic physiology.....................................................................30 Chapter 4: Results........................................................................................31 4.1 Demography...............................................................................................31 4.2 Growth Rates.............................................................................................36 4.3 Germination................................................................................................37 4.4 Photosynthetic Physiology.....................................................................39 Chapter 5: Discussion.................................................................................41 Chapter 6: Conclusion and Recommendations.................................50 References......................................................................................................52 Appendix A......................................................................................................62 Appendix B......................................................................................................65 ii List of Figures Figure 1: Distribution of Encephalartos longifolius in the Eastern Cape.........................13 Figure 2: The locality of the three study sites chosen for demographic study the cycad Encephalartos longifolius (Jacq.) Lehm.....................................................................24 Figure 3: Vegetation map showing the dominant vegetation types in all three study sites according to STEP (Vlok and Brown 2006) and Mucina and Rutherford (2006)....................................................................................................................26 Figure 4: Frequency of plant height of Encephalartos longifolius at (a) Kaboega, (b) Zuurberg, (c) Woodridge, measured in June 2010.......................................................32 Figure 5: The relationship between height and age at (a) Kaboega, (b) Zuurberg, (c) Woodridge measured in June 2010............................................................................34 Figure 6: : Relationship between plant diameter and height at (a) Kaboega, (b) Zuurberg, (c) Woodridge.........................................................................................................35 Figure 7: Frequency of change in plant height of Encephalartos longifolius at Kaboega, measured in July 2011.............................................................................................36 Figure 8: Change in plant height in juvenile and adult plants of Encephalartos longifolius at Kaboega, measured berween June 2010 and July 2011......................................................................................................................37 Figure 9: Overall germination of Encephalartos longifolius under controlled conditions over a 1 year period.................................................................................................38 Figure 10: Germinated Encephalartos longifolius seeds under controlled conditions..............................................................................................................38 iii Figure 11: The number of rotten Encephalartos longifolius seeds at three different moisture conditions................................................................................................39 Figure 12: The photosynthesis-irradiance relationship of Encephalartos longifolius seedlings (Vertical bars = ± S.E., n = 3) (Harrison et al. 1985).....................................................................................................................40 Figure 13: The photosynthesis–temperature relationship of Encephalartos longifolius seedlings (Vertical bars = ± S.E., n=3)........................................................................40 Figure 14: Fossil records of cycad abundance with changing temperature and carbon dioxide levels through geological time (Plass 1956; Buckley 1999).....................................................................................................................45 Figure 15: Projections of past, present and future temperature levels and carbon dioxide concentrations according to the IPCC, 2007 Assessment Report and Midgley et al. 2007......................................................................................................................46 Figure 16: The relationship between plant height and age at all three study sites combined...............................................................................................................65 List of Tables Table 1: Encephalartos species endemic to the Eastern Cape......................................11 Table 2: A Shapiro-Wilks test for normality for the demographic data collected at all three sites......................................................................................................................31 iv List of Plates Plate 1: Lignotuber on Encephalartos longifolius stem in response to fire........................................................................................................................62 Plate 2: Leaning-over effect of Encephalartos longifolius.............................................62 Plate 3: Encephalartos longifolius trampled and knocked over by large mammals...............................................................................................................63 Plate 4: Encephalartos longifolius cone damaged by baboons.....................................63 Plate 5: The base of Encephalartos longifolius stem impacted on by porcupines.............................................................................................................64 v Acknowledgements I would like to thank the following people and organizations: • Prof EE Campbell (Supervisor) for giving me an opportunity to do this project, as well as for her constant support, guidance and understanding throughout the project. • DR D.R Du Preez (Co-Supervisor) for his assistance with GIS, physiology, experiments and other additional information as well his dedication and sacrifices towards my physiology experiments • Nelson Mandela Metropolitan University and Prof Andrew Leitch for providing me with funding for my project. • Prof RM Cowling for allowing me to come along on his honours excursions. • The honours class of 2010 and 2011 for their assistance with sampling. • The Botany Department for making it possible for me to complete my studies. • The staff of the Ria Olivier Herbarium for their assistance in helping me obtain the relevant information. • The South African National Biodiversity Institute (SANBI); the South African National Parks (SANParks) and the Department of Economic Affairs Environment and Tourism (DEAT) for providing me with permits to sample in the respective study sites. • Ian Ritchie at Kaboega for allowing me to tag and measure the cycads as well as driving me to the cycad locality. • MR Neave Campbell at Woodridge College for allowing me to tag and measure the cycads on his property as well as access to the property at any given time. • My colleagues Adriaan, Anusha, Chan, Clayton, Merika, Nuette, Sibulele and Wynand for helping me with sampling and providing me with assistance throughout my project. . vi Abstract Cycads first appeared about 300 million years ago and historical data indicates that they survived fluctuations of global temperature and carbon dioxide concentrations and reached peak abundance during periods where temperature and carbon dioxide concentrations were much higher than the present conditions as well as the predicted increased levels. With Africa being one of the most vulnerable regions to climate change and in the absence of an evaluation of predicted impacts of climate change on cycads, this study aims to contribute to our understanding of responses of Encephalartos longifolius to increased temperature. Encephalartos longifolius (Jacq.) Lehm is an Eastern Cape endemic and like most cycads has been around for millions of years with very little change to its basic structure. Photosynthetic rates showed E.longifolius seedlings have C3 photosynthesis and even under stress they do not switch over to CAM photosynthesis. The photosynthetic rates of seedlings showed a typical C3 -plant type response under elevated carbon dioxide levels. Increased temperatures could be detrimental to the species but coupled with elevated carbon dioxide levels the growth of Encephalartos longifolius are postulated to outweigh the negative impacts of increased temperatures. Whilst climatic conditions in the Eastern Cape might not impact the abundance of Encephalartos longifolius, the species however is declining rapidly to the present near-threatened status. Demographic studies revealed a high percentage of juvenile numbers in the wild; however juvenile plants are impacted on by animals thereby jeopardizing their survival. Adult plants are heavily impacted on by animals; this reduces the probability of new offspring being produced. Cycads are also very slow growing which is the primary cause of their ruin in the wild. Encephalartos longifolius grows at just over approximately 1 cm per year with growth being more rapid in the juvenile plants. Once juvenile plants reach approximately 60 cm they stop growing in diameter but only get taller. Seed germination is also a very slow process with an optimum temperature of 28°C and a growth medium of at least 50% moisture. Regardless of the Eastern Cape predicted to become the hottest province in South Africa as a result of climate change, cycads will be able to adapt to the changing environment and conservation plans should focus on animal and human impacts that are the major force causing decreasing abundance of Encephalartos longifolius in the wild. vii Key words: cycads, photosynthetic rates, climate change, demographic studies, slow-growing, predicted impacts, C3 photosynthesis. viii Chapter 1 Introduction Cycads are known as the most primitive living seed bearing plants belonging to an ancient order the Cycadales (Chamberlain 1919; Giddy 1974). Cycads belong to a group of plants known as gymnosperms (plants which bear naked seeds) and their reproductive organs are produced in cones (Chamberlain 1919) Cycads are divided into three families namely Cycadaceae, Stangeriaceae and Zamiaceae which are distributed in the temperate and tropical regions of both the Southern and Northern hemisphere on the continents of: Asia, Africa, Central America and Australia (Anderson et al. 2007; Giddy 1974). In South Africa the two common genera of cycads are Encephalartos Lehm. and Stangeria T.Moore (Chamberlain 1919), with Encephalartos being the largest genus having 37 South African species of which 15 are found in the Eastern Cape and 8 are endemic to the province (Giddy 1974; Germishuizen et al. 2006). The key to distinguishing between the Encepharlartos species is the morphology of the median leaflets i.e leaves occuring at the middle of the leaf stalk (Giddy 1974). Encephalartos longifolius (Jacq.) Lehm is also known as the Zuurbeg cycad chosen as the study species (Giddy 1974; Donaldson 2003). This most attractive foliage species has long leaves, and stems are mostly erect with height ranging between 4 m and 6 m and diameter of between 30 cm and 55 cm. Grows well in full sun and adequate moisture conditions where the soil is not too damp, and is semi-hardy to frost (Chamberlain 1919; Giddy 1974; Donaldson 1997). Encephalartos longifolius like most cycads are slow-growing (Chamberlain 1919). The seeds of are nutrient rich and are heavily predated by monkeys, baboons, birds and rodents (Donaldson 1993). Like most cycads Encephalartos longifolius is an endangered species and according to the 1997; 2006 and 2010 IUCN Red List of threatened plants 82% of the world’s cycads were listed as threatened and is suffering a major extinction crisis (Anderson et al. 2007). Some species are dying out naturally (Donaldson 2003) but the major extinction factor is human activities, therefore permits are needed for possession of cycads (Giddy 1974; Donaldson et al. 2003). 1 Cycads have survived for over 300 million years with little change in their basic structure and neither fire nor drought presents any real threat to their survival (Giddy 1974). Some cycads have CAM photosynthesis but the majority of the South African species have C3 photosynthetic pathway (D.R. du Preez, pers.comm). Cycads first appeared during the late Carboniferous period and early Permian periods where temperatures and carbon dioxide levels are similar to that of the present and began to flourish in abundance (Microsoft Encarta 2005). The carboniferous period was then followed by a glaciations period which caused a slight decline in cycad abundance, however the populations recovered and it was during the Jurassic period that cycads reached their peak abundance (Chamberlain 1919; Plass 1956). Historically the earth has been much hotter and drier and carbon dioxide concentrations were much higher compared to the present (Plass 1956), however after the last cold period which ended about 15 000 years ago the earth continued to get warmer and warmer with much larger risks compared to changes that have occurred during the past decades (Scholze et al. 2006; IPCC, 2007). In the next 200 years global temperatures are estimated to increase by 1.5 -5.5ºC as a result of increased greenhouse gas emissions and have become one of the most important issues concerning scientists with a number of predictable environmental impacts (Marshak 2005; IPCC, 2007). Atmospheric carbon dioxide in particular is rising at an alarming rate and such a rate of increase will have far reaching effects on the earth’s resources especially primary production as primary production depends on the balance between respiration and photosynthesis (Turpie et al. 2002). According to the South African Country Study on Climate Change temperatures in South Africa are predicted to increase by 2.5ºC by the year 2050 which could result in major changes in the rainfall patterns, a strain on water resources, lower yields in food production and far reaching impacts on the biodiversity of plants in particular (Midgley et al. 2007; DEDEA, 2011). It is predicted that the impact of elevated carbon dioxide levels on C3 plants will result in an increase in the efficiency of photosynthesis, thus accelerating plant growth, however whilst C3 plants will show an increase in photosyntheesis in correlation with initial carbon dioxide increase over time photosynthesis will level out even though carbon dioxide continues to increase (Cheng et al. 2000; Ehleringer and Cerling 2002). 2 The aim of the study was to predict the impacts of climate change on Encephalartos longifolius (Jacq) Lehm. The objectives of the study were to determine population vulnerability, plant growth rates, and using germination rates predict what impact climate change in the Eastern Cape will have on Encephalartos longifolius. The hypothesis of the study is that the change in climate predicted for the Eastern Cape will have no effect on the survival of Encephalartos longifolius. 3 Chapter 2 Literature Review 2.1 Cycad biogeography, morphology, anatomy, physiology, conservation and demography Cycads are an ancient group of gymnosperms that can be divided into three families namely Cycadaceae, Zamiaceae and Stangeriaceae- the extant Cycadales (Chamberlain 1925; Donaldson et al. 2003). They are restricted to tropical and subtropical regions of Asia, South Africa, North and South America and Australia where they are well adapted to survive adverse environmental conditions (Chamberlain 1925; Whiting 1963). In Africa there are currently 69 recognized cycad species (Donaldson et al. 2003). The largest genus Encephalartos as well as Stangeria which is the smallest genus are both endemic to Africa (Giddy 1974; Donaldson et al. 2003). On the African continent cycads are distributed throughout 16 countries as well as on the Indian Ocean islands of Madagascar, Seychelles, Comores and Zanzibar (Donaldson et al. 2003; Anderson et al. 2007). South Africa stands as the most important center for diversity with more than 50% of the known African cycads found in the country (Donaldson et al. 2003). Cycads are an interesting group because they are morphologically intermediate between fern and flowering plants in evolution (Hertrich 1951). Cycads have survived for over the past 300 million years with little modification to their basic structure and it’s for this reason they are referred to as living fossils (Chamberlin1919; Giddy 1974). Fossil records for cycads extend at least the late Carboniferous early Permian period (about 300-315 million years ago) (Chamberlain 1919; Donaldson et al. 2003). There are many misconceptions of the age of individual plants (Hertrich 1951; Giddy 1974) but the age of cycads are generally estimated by counting the number of whorls of persistent leaf bases on the stem (Giddy 1974). All cycads are slow-growing plants and most of the species are arborescent meaning that they can either produce a single trunk which is marked with rings of growth after attaining a certain height (Chamberlain 1919; 4 Hertrich 1951). The slow growth rate and reproduction of cycads is the primary cause of their ruin in the wild. Cycads were once found worldwide during the early1800’s to the mid 1900’s (Brenner et al. 2003). A probable explanation for the long term survival of cycads is that they can survive drought and fire and are remarkably resistant to predators and pathogens; and are well equipped against environmental threats due to their ability to biosynthesize a variety of protective and secondary compounds such as glycosides and non-protein amino acids (Brenner et al. 2003). They thrive in habitats of moderate moisture, neutral or slightly acidic substrates and abundant sunlight (varies from species to species), with many species being adapted to low rainfall areas as excessive moisture causes root damage (Donaldson 1997). 2.1.1 Cones Cycads are dioecious and it is not possible to determine the sex without the presence of cones (Chamberlain 1919; Giddy 1974). Male and female cones occur on separate plants and are succulent consisting of a fleshy axis with numerous spirally attached sporophylls (Chamberlain 1919; Giddy 1974; Tang 1987). The seed cone sporophylls bear two ovules (Giddy 1974). The pollen cones are more slender compared to the seed cones. Male cones appear compact at first and later elongate with maturity to reveal the microsporangia or pollen sacs on the under surface of the sporophylls after which the pollen is shed once the pollen sacs are mature (Chamberlain 1919; Giddy 1974). In majority of the species the female cones are usually shorter, broader and sometimes nearly globular while the male cones are erect and triangularly shaped (Chamberlain 1919; Hertrich 1951). Cone appearance varies according to species and climatic conditions, male and female cones first start appearing towards the end of summer (late February) and grows steadily for about three months before attaining maximum size (Hertrich 1951). Plants are generally considered to be poikilothermic but they possess a number of physiological and morphological features that enable them to maintain tissue temperatures that differ from atmospheric temperatures (Tang 1987). In cycads endogenous heat production (thermogenesis) is widespread and common among male 5 cones whilst female cones exhibit very weak thermogenesis (Tang et al. 1987). Thermogenesis in male cones is known to occur in a circadian pattern during pollen shedding and this is the general pattern in all male cones because the basic morphology of male cycad cones is similar across all genera (Chamberlain 1925). 2.1.2 Stem Cycads stems are usually unbranched however when branching occurs in the columnar types this is a result of an injury to the stem (Giddy 1974). Stems are thick and slowgrowing and branching rarely occurs but in the event of branching up to 10 stems may originate from a single base (Chamberlain 1919; Giddy 1974). The age of cycads can be estimated by counting the armor of leaf bases on the trunk however some species for example Encephalartos longifolius have heavy stems that become compressed as a result of the weight and may reduce the visibility of leaf bases over time (Chamberlain 1919; Giddy 1974). Stem injury also allows for the plant to be more susceptible to pathogens and rotting (Whiting 1963; Giddy 1974; Vorster et al. 2004). Leaning over of the stem is often visible in some of the taller cycads, this is because the root system is not very extensive and the plants are heavy so as they get tall enough to be affected by the wind they tend to lean over (Chamberlain 1919). Externally the stems of cycads consist of a thick layer of spirally arranged persistent leafbases and internally there is a large pith in the center surrounded by a corky layer and a thin layer of wood even though they are known to be hardy plants (Chamberlain 1919; Whiting 1963; Giddy 1974). Mucilage canals are found in all parts of the plant particularly in structures such as the cortex, the pith, leaves and sporophylls (Giddy 1974). The stems of cycad seedlings in particular possess lignotubers (a swelling at the base of the stem just above the soil level) that helps the seedling survive fires by sending out new growth if the top growth is damaged (Chamberlain 1925; Donaldson 2003). 2.1.3 Roots Cycad seedlings have taproots which soon form several lateral branches which end in smaller fibrous roots that extends down to great depts. providing the plant with small amount s of water when the surface soil is very dry (Chamberlain 1919). Subterranean 6 (below the surface) cycad species have very large tuberous roots which are probably climatic adaptation and columnar species do have stems for water storage (Giddy 1974). An apogeotropic root (roots that grow upwards and form coralloid masses just above the surface of the soil) is an unusual feature in cycads (Chamberlain 1919). This feature allows for roots to contain a symbiotic blue-green alga Anabaena (Chamberlain 1919; Giddy 1974), that assist the plant by nitrogen fixation and assimilation therefore allowing the plant to live on nutrient poor soils (Giddy 1974; Whiting 1963). Studies have shown that the symbiotic algae in the roots fix up to 38 Kg of atmospheric nitrogen per hectare per year (Whiting 1963). The depth and size of roots provide the plant with anchorage and support during heavy winds however as plants get taller they are more prone to leaning over due to heavy winds (Chamberlain 1919; Whiting 1963). The roots system also contributes to the plants survival during fires, (Whiting 1963). 2.1.4 Leaves Cycad leaves are an attractive feature which gives the plant its popularity as a decorative plant (Chamberlain 1919). Leaves consist of a strong central stalk or rachis from which numerous leaflets are produced (Chamberlain 1919). Leaves are neither strictly alternate nor opposite and when they overlap the overlapping maybe either upwards or downwards and the leaves of mature plants appear in whorls (Chamberlain 1919; Giddy 1974). The shape, margins and colour of the leaflets varies from species to species providing distinguishing characters between species (Giddy 1974). The new leaves are very delicate and susceptible to injury but once they reach maturity and attain their full size the tissues harden and leaves become hard and leathery and extremely strong and resistant to moisture and dryness extremes (Chamberlain 1919; Giddy 1974). The length of the leaves vary from a few centimeters in the smallest plants to about 1-2 m in the larger plants (Whiting 1963). Leaves may persist for up to 5 years before a new set arises and once the new set arise the old leaflets turn yellow and fall off but the leafstalks may remain for several years before falling out (Chamberlain 1919). These persistent leafbases constitute the outer layer of the stem known as the characteristic armor that serve to protect the stem from the hazards of fire as well as provides an estimate of plant age (Chamberlain 1919; Giddy 1974). The structure of the epidermis of the leaflet is also adapted to the hot dry climate in which many species grow. It consists of a highly cutinized upper surface that is almost impervious to water while the stomata 7 or pores on the under surface of the leaflet are closed except when the air is moist, this feature is adapted to minimize water loss and aids the plant surviving low moisture supply (Chamberlain 1919; Whiting 1963; Giddy 1974). 2.1.5 Toxicity Cycads for many years have been a food source for people and animals, however the toxicity of the plant is very high (Chamberlain 1919; Whiting 1963, Giddy 1974). The plant is known to contain numerous toxic ingredients such as cycasin and macrozamin that is lethal to the consumer and when ingested continuously could result in irreversible paralysis of the extremities (Whiting 1963). The poisonous properties of the South African cycads Encephalartos is mainly found in the cones and seeds of the genus (Chamberlain 1925; Giddy 1974). In the seeds the kernels of all species have been tested and proven to be extremely toxic, the toxic effect resides primarily in the liver of the consumer; other organs such as the stomach, lungs and heart tissue also showed signs of damage when a post mortem conducted on a rabbit that consumed kernels as part of an experiment was examined (Giddy 1974). Similar research has shown that sheep and cows also have negative effects from consuming young cycad cones and in some cases it resulted in the animal’s death (Schneider et al. 2002). There are no known cases of poisoning following ingesting cycad seeds and cones in humans; however children are warned to take precaution in consuming the plant. 2.1.6 Photosynthesis There are three photosynthetic pathways that exist amongst terrestrial plants namely: C3, C4 and CAM photosynthesis (Ehleringer and Cerling 2002). The photosynthetic pathway of cycads is either CAM or C3 photosynthesis, however most South African species are C3 plants (D.R. du Preez, pers comm). C3 photosynthesis is a multi step process because two molecules of the phosphoglyceric acid (PGA) are formed when carbon dioxide combines with ribulose bisphosphate (RuBP), a 5-carbon molecule, in a reaction catalyzed by an enzyme known as RuBP carboxylase (Ehleringer 1979). The initial product formed in this pathway is PGA, a 3-carbon molecule and the energy sources used to drive this pathway are ATP and NADPH that come from the light reactions of photosynthesis (Ehleringer 1979). This pathway is also known as the most primitive pathway and one of its major disadvantages is a process known as 8 photorespiration (a process that takes place in the light in which carbon dioxide is evolved from photosynthetic tissues), however further research has shown that photorespiration arises because the enzyme RuBP carboxylase has an oxygenase activity as well as a carboxylase activity (Ehleringer 1979; Ehleringer and Cerling 2002). Therefore this oxygenase activity of RuBISCO is dependent on [CO2/O2] ratio and temperature and as result of this the efficiency of the C3 pathway increases as atmospheric carbon dioxide increases (Ehleringer and Cerling 2002). Crassulacean Acid Metabolism (CAM) is also another photosynthetic pathway employed by some cycads (REF). CAM photosynthesis is a modified form of C4 photosynthesis where C4 photosynthesis can be described as morphological and biochemical elaboration of the C3 pathway where CO2 is first combined with phosphoenolpyruvate (PEP) catalysed by the enzyme PEP carboxylase to form a 4-carbon molecule called oxaloacetic acid (OAA) (Ehleringer 1979). In CAM photosynthesis both the catalytic enzymes from the C3 and C4 pathways are present in the same cells, but the RuBP carboxylase is active during the day when the stomata is closed and CO2 taken up during the night is fixed and PEP carboxylase is active during the night when the stomata is open allowing for the enzyme to fix atmospheric CO2 (Ehleringer 1979). A disadvantage of CAM photosynthesis is that as a result of only nocturnal stomatal opening, the maximum rate of photosynthesis is much lower compared to that of C3 and C4 plants, however CAM plants are better adapted to survive extreme periods of drought than C3 and C4 plants (Ehleringer 1979). 2.2 South African genera 2.2.1 Encephalartos Lehm The genus Encephalartos belongs to the family Zamaiceae Horan. and are Cycadalean plants with radiospermic seeds borne in compact cones with a central axis (Anderson et al. 2007). Previously there were sixteen species of Encephalartos that were confined to South Africa and at present there are 37 species found in South Africa (Chamberlain 1925; Germishuizen et al. 2006). All species in this genus are classified as long-lived plants with the average adult span varying from 150 years to > 1000 years depending on 9 the species (Donaldson 2008). Like all cycads Encephalartos are dioecious plants and in the female cones the megasporophylls woody and peltate bearing a single pair of lateral ovules whereas in the male cones pollen is produced in simple cones with sporangiophores peltate and cones having two lateral lobes each bearing several abaxial, di-to tetrasporangiate and a shortly stalked synangia (Chamberlain 1919; Anderson et al. 2007). The cones of the species Encephalartos longifolius are the heaviest of all the South African species with the female cones weighing up to about 36 kg and produce about 600 seeds whereas the male cone is much smaller and weighs about 30 kg (Whiting 1963; Donaldson 1993). The leaflets of Encephalartos are pinnate with a central leafstalk with numerous leaflets that have parallel veins (Chamberlain 1919; Giddy 1974). Young leaves are covered in fine hairs that disappear as the plant matures, and stems are mostly erect and grow up to 10 m or more in height and are characterized by densely packet persistent leaf bases (Chamberlain 1919; Whiting 1963; Giddy 1974). The distribution of this genus ranges from tropical to warm-temperate and is widespread through central and northern South America, Australia and central and southern Africa. In South Africa most occur in the Eastern and Western Cape, Kwa-Zulu Natal, and in the Free State regions (Germishuizen et al. 2006; Giddy 1974). The Eastern Cape Province is one of several centers of diversity for the genus and 8 species are endemic to the province (Table 1) (Giddy 1974; Donaldson 2003; Vorster et al. 2004; Germishuizen et al. 2006): 10 11 tree altensteinii cycad arenarius R.A. cycad horridus Broadest leaves in the genus, Albany cycad Zuurberg cycad Encephalartos latifrons Lehm. Encephalartos longifolius cycad princeps R.A. Dyer cycad s River trispinosus (Hook) R.A. Bushman’ Encephalartos Dye4r Small size plant, single bushy Kei River Encephalartos diameter in height and 35-40 cm in Branched reclining stems, 1-3 m Medium to large size plant. cm in diameter stem, 1-4 m in height and 30-40 and 30-55 cm in diameter (Jacq.) Lehm. multi-stemmed, 1-6 m in height Most attractive foliage, erect 30-45 cm in diameter multi-stemmed, 1-3 m in height, in height and 30-40 cm in (Jacq.) Lehm. single to multi stemmed, 1-3 m Medium, low growing plant, Blue Encephalartos stemmed, 1-3 m in height and 2035 cm in diameter cycad cycadifolius Vulnerable Vulnerable Near Threatened Critically endangered Endangered moisture conditions sunlight and low Grows well in full moisture sunlight and low Grows well in full moderate moisture sunlight and Grows well in full climatic conditions Adapted to hot dry frost- hardy sunlight and is Grows well in full completely frost - sunlight and is Grows well in full not frost-hardy Medium size plant, single to multi rare blue in colour shade and high Grows well in half semi-hardy to frost sunlight or shade; Grows well in full Comments moisture conditions. Least concerned Endangered Vulnerable List 2010 (SANBI 2011) according to the IUCN Red Data National conservation status 20-30 cm in diameter, green or stemmed, 1-2 m in height, and Medium size plant, multi cm in diameter time, 4-5 m in height and 30-40 initial erect trunk reclines over Medium to large evergreen plant , Description (Jacq.) Lehm. Bedford Encephalartos Dyer KwaQaba Encephalartos Lehm. Bread Name Common Encephalartos Species Table 1: Encephalartos species endemic to the Eastern Cape Habitat requirements for Encephalartos species include forest, savanna and grassland, and thicket with savanna having the greatest species diversity and species within this habitat have a high tolerance limit to a range of conditions (Donaldson 2008). Species found occurring in the thicket, grassland and forest habitats are specific to these habitats. In South Africa most Encephalartos species have a decreasing population trend and are classified as vulnerable to critically endangered (SANBI, 2011). Encephalartos longifolius(Jacq.) Lehm which was the species of focus for this study is also an Eastern Cape endemic commonly known as Suurberg cycad is widely distributed in the southwestern parts of the province (Figure 1). This species was one of the first cycads to be scientifically recorded in South Africa in 1772 by Carl Peter Thunberg (Donaldson et al. 2003). According to the IUCN Red Data List in 1978 there were approximately 4500 wild plants of E.longifolius in the Eastern Cape, however as a result of human impacts the wild populations has been declining ever since and currently there are <700 plants left in the wild (IUCN, 2010). Encephalartos longifolius currently has a near threatened conservation status and if the population continues to decline the species will soon become endangered to critically endangered (SANBI, 2011). 12 Eastern Cape Figure 1: Distribution of Encephalartos longifolius in the Eastern Cape. 2.2.2 Stangeria T.Moore This genus was classed with ferns until the cones were discovered and only one species Stangeria eriopus (Kunze) Nash is endemic to South Africa (Chamberlain 1919). Belonging to the family Stangeriaceae which is a small family that has originated from Gondwana, Stangeria are perennial, fern-like herbs with subterranean, tuberous, branched or unbranched stems (Donaldson et al. 2003; Anderson et al. 2007). The stem may sometimes branch into 10 to12 heads and each head may produce a cone at the same time (Giddy 1974). Leaves are pinnate with taenopteroid venation or with many parallel veins and margins maybe serrated, entire of deeply fringed according to the locality (Giddy 1974; Anderson et al 2007). Male and female cones are silvery pubescent at first becoming brownish upon maturity. Female cones are much bigger than the male cones but both have overlapping cone scales (Giddy 1974). 13 Stangeria eriopus is found in eastern coastal grassland and forest habitats and is distributed in the Eastern Cape and Kwa-Zulu Natal (Giddy 1974; Germishuizen et al. 2006). 2.3 Pollination and Seed-dispersal Pollination usually occurs in March or April when the microstrobulus elongates to its maximum length and pollen is shed from the microsporangia (Donaldson 1997). Cones first emerge during January to February (Donaldson 1997). Female cones start disintegrating between September and November when seeds are released (Donaldson 1997). Seed germination occurs immediately after the cone bursts and seeds are released (Giddy 1974). Cycads do not exist in isolation and many of their known interactions with other species are specific to one or few cycad species (Donaldson et al 2003). Over the years research has shown that cycads have evolved symbiotic interactions with arbuscular mycorrhizae and nitrogen fixing blue-green algae (Whiting 1963; Giddy 1974). Seed dispersal by vertebrates and invertebrates as well as environmental factors heavy rains and in particular wind is central to individual species biology and ecosystem function (Tiffney 2004). Dispersal has many advantages to the species such that it allows for seeds to escape from potential predators, seed colonization at favourable growth sites, and competition reduction between parent and the offspring and between siblings (Tiffney 2004). Seed dispersal also influences distribution of individual taxa and plant community structure (Tiffney 2004). Cycads are postulated to be only wind-pollinated (Chamberlain 1925) and as a result of male and female cones occurring on separate plants, a male and female plants must cone at the same time and grow close enough to one another for wind pollination to be effective in order to produce a fertile seed in the wild (Giddy 1974). Wind pollination forces the pollen grains between the open apical scales from where they gradually shift spirally downwards through the center axis of the cone (Giddy 1974). As a result of wind dispersal of pollen immense an amount of pollen is wasted (Giddy 1974). Nevertheless wind dispersal and pollination is highly effective when it does occur. A study conducted by Donaldson (1997) showed that there was a significant decrease in seed set when wind was excluded from the experiment in comparison to wind inclusive experiments. 14 In contrast, some have suggested a role for Insects in cycad pollination (Giddy 1974; Schneider et al. 2002), however this is highly doubtful because for example bees collecting pollen on male cones show no evidence of visiting female cones growing nearby because female cones do not produce pollen and in the event that they are found on female cones it is probably incidental (Giddy 1974). The role of pollination by certain weevils for example a curculionid weevil Antliahinus zamiae is also doubtful because they are found mainly on female cones where they seek out the young immature seeds in which they lay their eggs (Giddy 1974; Schneider et al. 2002). However subsequent experimental studies have showed that insects do play an important role in cycad pollination for example Suinyuy et al (2009) found that three beetle species namely Metacucujus encephalarti, Erotylidae sp. and Porthetes hispidus are potential pollinators of a South African cycad Encephalartos friderici-guilielmi Lehm. Insects are known to play a significant role in dispersal, as pollen may sometimes get stuck on the legs or body of insects and carried over variable distances where the fate of the pollen is unpredictable (Schneider et al. 2002). Other fauna such as birds, dassies, baboons, vervet monkeys and fruit-eating bats all feed on cycads and indirectly aid in pollination or seed dispersal (Giddy 1974). Baboons and monkeys often remove the entire cycad cone and carried some distance away from the mother plant where the fleshy pulp is consumed and poisonous kernels are spat out, this often results in extensive damage to the meristamatic tissue of the cones (Giddy 1974). Several bird species particularly the Crowned Hornbill, the Brown-necked (Cape) Parrot and the Trumpeter Hornbill also aid in pollination/dispersal by breaking down the cones with their sharp beaks and exposing the drupaceous red seeds or by carrying away seeds over long distances and eating them during the halts on the way to the roosting site (Giddy 1974). Hand pollination of cycads has become popular over the years especially amongst cycad growers who wish to increase their seed stocks (Giddy 1974). Hand pollination aims to achieve what wind pollination does however this is dependent on correct timing and acquiring of pollen (Giddy 1974). Hand pollination can be achieved by collecting 15 large quantities of pollen and refrigerating it, after which it can be lightly sprinkled on the female cone, however this methodology does not always guarantee viable seed production (Dehgan and Yeun 1983). As a result of seed predation mast-seeding in cycads evolved (Janzen 1971). Mastseeding is a phenomenon in which synchronous seed production of seeds occurs within a plant population in one year followed by a long interval in which few or no seeds are set (Janzen 1971). This phenomenon is said to have evolved as a behavioral defense against predators because large amounts of seed production at a single time allows for predation satiation therefore during the period between the mast years predator numbers are said to decline as there is a shortage in their food resource (Ballardie and Whelan 1986). Seed dispersal however is very important to the species because it provides survival for the species such as in response to environmental changes it provides a mechanism for geographic migration in plant populations; it allows the plant, seeds and seedlings in particular to escape from predators; it provides seeds with the ability to spread over large areas therefore allowing for cross-pollination to occur as well as distance also inhibits the spread of pests in new generations; competition between parent and offspring and amongst siblings is also reduced (Tiffney 2004). 2.4 Propagation and Germination Propagation of cycads can be achieved by growing cycads from seeds or offsets and branches (Giddy 1974). Growing cycads from seeds usually occurs when the seed is viable (fully developed and fertile). Fertility of seeds can be determined in the following ways (Giddy 1974): 1) immersing seeds in deep water after removing the fleshy covering, infertile seeds will float whilst heavier fertile seeds will sink to the bottom; 2) cut seeds open longitudinally and see whether a fertile embryo is present, a fertile embryo is attached to a thin tightly coiled suspensor and extends to about half the seed length whilst an infertile embryo has an empty hollow at the micropylar end of the seed. Cycads are known as prolific seed producers with individual female cones producing approximately 500 seeds per season (The Cycad Newsletter, 2007). Seeds have an 16 inner stony portion called the sclerotesta and an outer fleshy layer called the sarcotesta, with both portions being separated by a paper-thin membrane (Whiting 1963). Seeds are usually grayish-brown in colour and the size of a pecan nut, this feature appears to be constant for a given species (Whiting 1963; Giddy 1974). Seed germination is often a very slow process and three separate dormancies are involved in germination such as: 1) The inhibitory effect of the fleshy coat; 2) the embryo that needs to reach maturity before the seed can germinate; and 3) the thick stony (sclerotesta) layer which is thick and prevents entry of water into the seed (Dehgan 1983). Seed germination in the wild is dependent on environmental variables and symbiotic relationships whereas laboratory germination is dependant on temperature and moisture and these two parameters vary from species to species (Dehgan 1983). Propagating cycads from suckers that form at the base of the adult plants involves opening the hole alongside the stem until the entire stem of the sucker and its roots are exposed and then carefully sliced off with the use of a sharp-bladed spade (Giddy 1974). Cuttings are then treated with fungicide and left for a few days to allow damaged portions to dry out before being planted (Giddy 1974). 2.5 Conservation status Cycads are group of plants that have global conservation significance, with 40% of the world’s cycads falling into globally recognized biodiversity hotspots, and a staggering 82% of the world’s cycads species being listed as threatened (Donaldson 2003). In South Africa cycads are classified as protected or specially protected plants. The sustainable utilization of wild populations has been adopted at regional, national and provincial level (Donaldson et al. 2003). In the past Gauteng declared all cycads Specially Protected Plants and required those whom had cycads in their possession to apply for a permit, however Gauteng was not the only province to protect cycads, other provinces and strict protective laws and heavy penalties were laid down by the Nature Conservation Ordinances of each province (Giddy 1974). Currently due to an increase in declining cycad numbers, cycads are listed in the CITES (Convention on International Trade in Endangered Species) Appendix 1 which states that a permit is required for 17 import and export of all cycad species and permission is required from national and regional authorities such as DEAT, SANBI and SANParks involving any activities on wild populations (CITES, 2003). Cycads have been and are still under threat of extinction despite the many attempts made to conserve the plants and their habitats. The threatened status of African cycads was recently revised as part of a global revision and as part of the 2010 IUCN Red List of Threatened Plants and according to the most recent data three South African cycad species are extinct in the wild, 12 taxa are critically endangered, 4 are classified as endangered and 10 as vulnerable, with a further 4 taxa classified as near threatened and only 4 as being of least concern (SANBI, 2011). Some cycad species are dying out naturally, but the major threat to the survival of cycads is humans. Human activities have long been responsible for the decline in cycad numbers (Giddy 1974; Donaldson 2003). Some of these activities include (Osborne 1995; Donaldson 2003): • Habitat destruction • Alien vegetation • Traditional use of species for medicinal or magical purposes • Trade in wild plant collections • Urban and rural land demands • Removal of plants by collectors or traders • Road and dam construction • Agricultural development and afforestation. 18 2.6 Demography Demographic studies are an important tool in assessing the population viability of cycad populations and are beneficial in cycad conservation because the main goal of the study is to estimate the growth rate, (increase or decrease in population numbers), of the population and the extinction probabilities (Gallego 2007). Demographic studies of cycads was poorly understood in the past because they are time consuming, labor intensive and data can be inaccurate but regardless of this it has now become a powerful conservation tool because it is simple to implement and allows for long-term monitoring of individuals within a population (Miguel et al. 2006; Gallego 2007). Survival and fecundity are the most important parameters in estimating growth rate where survival can be estimated by the probability of seedlings becoming adults over a set period of time, this is achieved by measuring the height and diameter of a set number of individuals as well as counting the leaf bases for an indication of the age of an individual (Clarke and Clarke 1987; Giddy 1974; Gallego 2007). Fecundity can be estimated by the probability of adults producing offspring that survive well into the seedling stages this is achieved by counting the number of seeds that germinated from seeds produced in the previous reproductive season, counting the proportion of seed-producing females in the population and, estimating the average number of seeds produced per female (Clarke and Clarke 1987; Miguel et al. 2006; Gallego 2007). For effective implementation of management plans and long-term conservation strategies demographic studies must be conducted for a minimum period of three years on a single population with regular monitoring intervals (Miguel et al. 2006; Gallego 2007). Apart from providing useful information for conservation demographic studies can also contribute towards many aspects of cycad biology such as herbivory rates, the role of disturbance, and the growth of new leaves (Clarke and Clarke 1987; Gallego 2007). 19 2.7 Climate Change 2.7.1 The causes and effects of climate change Global temperatures and carbon dioxide levels have fluctuated throughout history. The carboniferous period was the only geological period in which global temperatures and carbon dioxide levels were similar to that of the present (average temperatures of 12°C past compared to15.5°C of the present); (CO 2 levels of 350 ppm in the past and 390ppm in the present) (Plass 1956; Buckley 1999; IPCC, 2007). At present global warming is one of the most important issues concerning scientists. The issue focuses on the increasing accumulation of greenhouse gases especially carbon dioxide in the atmosphere caused by industrial activities, and deforestation (IPCC, 2007; Sheikh and Gorte 2008). Since the 1800’s industrial activity has steadily increased and so has the atmospheric carbon dioxide levels and global temperature (IPCC, 2000). The impact of human activity on atmospheric carbon dioxide levels and other greenhouse gases has become one of the most debated topics of all time. The burning of fossil fuels (oil, natural gas and coal) and clearing of vegetation are the most common impacts that result in the increase of atmospheric carbon dioxide (IPCC, 2000). Global temperature is predicted to increase by an additional 1.5°C and 5.5ºC over the next century (IPCC, 2007). The projected temperature increase is a global average with some areas being cooler and some being hotter. Higher temperatures will occur at high latitudes (the poles), inlands, and in the interiors of continents (IPCC, 2007). According to the Third Assessment Report of the Intergovernmental Panel on Climate Change (IPCC, 2001), Africa is one of the most vulnerable continents in the world to climate change. During the 20th century average surface temperatures have already increased by approximately 0.7° C and are predicted to increa se further by the end of the century (IPCC, 2007). In South Africa the Eastern Cape Province is expected to experience the highest temperature increases towards the north-west interior and along the coast the lowest increases are likely to occur (Midgley et al. 2007; Johnston et al. 2011). Average surface temperatures for the Eastern Cape are expected to increase by approximately 2° C by 2050 with major impacts on the environmental, social and economic aspects of the province (DEDEA, 2011; Johnston et al. 2011). Temperature increases could result in increased intensity in droughts and increases in evaporation rates, as well as changes in the regions rainfall patterns (Johnston et al. 2011). 20 Analysis of bubbles trapped in ice indicates that carbon dioxide concentration in the atmosphere has varied widely during the last 1700,000 years and closely parallels variation in global temperatures (Molles 2008). Atmospheric carbon dioxide concentrations are expected to increase by double its current levels by the 21st century and these increases will not only affect the global climate but it will also affect the structure and processes of ecological systems from populations to landscapes (IPCC, 2007; Midgley et al. 2007). It has also been predicted that larger amounts of atmospheric carbon dioxide could increase the rate of photosynthesis in areas with adequate water and soil nutrients. This could counter global warming because it would remove carbon dioxide from the troposphere therefore helping to slow global warming (Miller 2007). However recent studies have indicated that this counter current effect would be temporary due to the following reasons (Miller 2007): 1) increased photosynthetic rates would slow down as plants reach maturity and take up less carbon dioxide from the atmosphere, 2) carbon dioxide stored by plants would be returned to the atmosphere in the same composition when plants die, decompose or burn, 3) increased photosynthetic rates decrease the amount of carbon stored in the soil because more carbon dioxide is utilized for photosunthesis , and 4) tree growth might temporarily slow carbon dioxide emissions in the southern hemisphere but will probably increase carbon dioxide emissions in the northern hemisphere because the northern hemisphere has a higher deforestation rate. While the exact nature of increases in temperature and carbon dioxide concentrations is not known there are predicted outcomes such as: • Melting ice and snow (IPCC, 2007) • Sea level rise, which will result in further global impacts (IPCC, 2007) • Changing ocean currents (IPCC, 2007) • Warmer and more acidic seas (IPCC, 2007) • Inundation of coastal areas (IPCC, 2007) • Increased mortality of coral building reefs (IPCC, 2007) • Food production may increase in some areas and decrease in others (IPCC, 2007 21 • Release of carbon dioxide and methane from arctic soils, accelerating global warming (IPCC, 2007) With focus on cycads the following outcomes of climate change could impact the plants in a positive or negative way: • Increased frequency of intense storms (IPCC, 2007). • More intense and more frequent heat waves in temperate regions (IPCC, 2007). • Increased summer drought in semiarid regions (IPCC, 2007). • Dieback of forests due to increased incidents of disease and insect attack (IPCC, 2007). • Increased wildfires in dry regions (IPCC, 2007). • Widespread extinction of plant and animal species (IPCC, 2007). • Changes in precipitation and weather extremes (IPCC, 2007). 2.7.2 What does climate change mean for C3 plants? C3 plants will show an exponential rise in photosynthesis in response to increasing temperatures, but once the plant reaches its optimum temperature photosynthesis will start decreasing (Ward et al. 1999; Kim et al. 2007). This decrease in photosynthesis (CO2 uptake) is a result of decreased enzymatic activity of RUBISCO therefore reducing the ability to fix carbon dioxide thus stimulating photorespiration (O2 uptake) (Ehleringer and Cerling 2002). Temperature increases are unfavorable to C3 plants, however climate change involves and increase in temperature and carbon dioxide concentrations which occur concomitantly (Ward et al. 1999; Kim et al. 2007), therefore C3 plants will be favoured as a result of increased CO2 concentrations because the increase in C3 photosynthetic efficiency that takes place under elevated atmospheric carbon dioxide levels will outweigh the reduction of photosynthesis that is attributable to higher temperatures (Sage 1996; Ward et al. 1999; Ehleringer and Cerling 2002). Higher levels of carbon dioxide will increase the efficiency of photosynthesis, thus accelerating plant growth; however while C3 plants will show increased productivity in correlation with initial carbon dioxide increase, productivity will eventually level off over time despite a further carbon dioxide increase (Tissue et al. 1995). A possible explanation for this is that increased photosynthesis would slow as plants reach maturity and take up less carbon dioxide from the atmosphere (Cheng et al. 2000; Norikane et al. 2010). 22 Chapter 3 Materials and Methods A demographic study of Encephalartos longifolius was conducted to determine the health of the population in the Eastern Cape. Germination experiments were set up to determine germination requirements of the species and the photosynthesis pathway of cycads were determined by measuring the gaseous exchange of Encephalartos longifolius seedlings. Photosynthetic rates were also used to determine the influence of elevated temperatures and carbon dioxide levels on cycad seedlings and projection models of elevated temperatures and carbon dioxide concentrations through geological time was used to predict the impact future climate changes will have on cycads. 3.1 Demography Selection of study sites were dependent on the presence of wild populations of Encephalartos longifolius, as well as the permission granted by the landowners and conservation authorities to work on the property and with the plants. 3.1.1 Study Sites The three study sites chosen were Kaboega, Zuurberg Mountain Range and Woodridge College (Figure 2) which were all within the Eastern Cape region and apart from having the presence of Encephalartos longifolius in common the study sites differed from each other. 23 Woodridge Figure 2: The locality of the three study sites chosen for demographic study of the cycad Encephalartos longifolius (Jacq.) Lehm. • Kaboega The Kaboega area (Figure 2) is surrounded by the Greater Addo National Elephant Park. It lies on the Kaboega River against the Northern slopes of the Zuurberg Mountains, surrounded by rugged mountains and densely aromatic vegetation. It is a hydrographic area situated at 33° 21' S and 25º 22’ S (R.M. Cowling, pers.comm). Kaboega is an arid to semi-arid area receiving a mean annual rainfall of about 455 mm per year. Rainfall patterns tend to be evenly distributed throughout the year with two peaks occurring in February to March and October to November (I. Ritchie, pers.comm). The temperature is highly variable with a summer maximum of 32ºC and minimum of 15ºC and a winter maximum of 15ºC and minimum of 5ºC with frost occurring frequently in winter (I. Ritchie, pers.comm) 24 This Kaboega area is home to many wildlife species such as baboons, kudu, bushbuck, vervet monkeys, hartebeest and many other small mammals as well as many bird species (I. Ritchie, pers.comm). The vegetation of the area according to STEP (Vlok and Euston-Brown 2002) and Mucina and Rutherford (2006) as illustrated in Figure 3 includes: • Suurberg Quartzite Fynbos This vegetation type occurs on the mountain slopes and plateaus on sandy soils and the typical structural type are grassy fynbos with localized patches of dense ericaceous and proteoid fynbos. On south-facing slopes this vegetation type can be replaced by thicket vegetation as a result of overprotection from fire or replaced by grassland due to over-burning. This vegetation type has a least threatened conservation status and endemic taxa found in it are Oldenburgia grandis (small tree), and Euryops hypnoides, E. polytrichoides (low shrubs) (Mucina and Rutherford 2006). • Albany Thicket A brief description of this vegetation type includes: it is very dense, semisucculent, thorny and forest-like with an average height of 2-3m. Dominant species found in Albany Thicket are Olea africana subsp. africana (Wild olive), Allophylus decipiens (false current), Gymnosporia buxifolia (spike thorn), Schotia latifolia (bosboerboon), and Euphorbia triangularis (tree euphorbia) (Mucina and Rutherford 2006). • Southern Mistbelt Forest This vegetation type occurs in the ravines and on mountain slope This vegetation type is described as multi-layered i.e. it has two layers of tall trees, followed by a dense shrubby understorey and a well-developed herb layer. Trees are usually 15-20m tall and grow on soils that are loamy deep soils with a high nutrient status. This vegetation type has a least threatened conservation status and endemic species occurring are Herbs: Plectranthus rehmannii, P. elegantululus, Streptocarpus candidus, S. bolusii, S. silvaticus, S. fanniniae, Pyrrosia africana and tall shrub: Eugenia zuluensis (Mucina and Rutherford 2006). 25 Albany Thicket Kowie Thicket Southern Mistbelt Forest Sundays Thicket Suurberg Quartzite Fynbos Van Stadens Forest Thicket Woodridge Figure 3: Vegetation map showing the dominant vegetation types in all three study sites using STEP (Vlok and Euston-Brown 2002) and Mucina and Rutherford (2006). • Zuurberg Mountain Range The Zuurberg Mountain (Figure 2) is a major landscape feature that forms part of the Greater Addo National Elephant Park and is situated at 33° 21’ 4” S and 25º 44’ 38” E in the Eastern Cape (Kerley and Boshoff 1997). Height above sea level varies between 250 m to 970 m. Mean annual rainfall of the area is approximately 722 mm per year with peaks in rainfall pattern during spring and autumn (Vlok et al. 2003). Winter months are driest and during the summer months thunderstorms occur which can be the cause for lightening fires. Summer temperatures reach a maximum of about 29ºC and a minimum of about 19ºC (Kerley and Boshoff 1997). Zuurberg is home to many wildlife species such as baboons, impala, bushback, vervet monkeys, and many small mammals and in past times megaherbivores such as elephants and black rhino were present in the area (Kerley and Boshhoff 1997; Kamineth 2004). The dominant vegetation type found at Zuurberg is 26 • Sunday’s Thicket (Figure 3) Species occurring in this vegetation type include: tree euphorbias (Euphorbia triangularis) and Kiepersoi (Cussonia spicata) that emerge above the tree canopy; abundant species sneezewood (Pteroxylon obliquum), wild olive (Olea europaea subsp. europaea), bosboerboon (Schotia latifolia); characteristic species Hippobromus pauciflorus and spekboom (Portulacaria afra) only occurs on the driest sites and is never dominant. (Pierce and Mader 2006). • Kowie Thicket (Figure 3) This vegetation type has a least threatened conservation target and occurs mainly on steep, north-facing dry slopes with soil types ranging from clayey over the Darlington Dam; sandstone and shale (Wittenberg group); and Algoa group calcareous sandstone in the south. Vegetation found on moister southfacing slopes support thorny thickets that are dominated by shrubs (Carissa, Putterlickia, Azima, Gymnosporia), low evergreen trees (Euclea, Pappea, Cussonia, Hippobromus, Schotia, Ptaeroxylon) and fewer succulent trees abd shrubs. Endemic taxa found in the vegetation include Faucaria nemorosa (succulent herb), Wahlenbergia kowiensis (herb), and Albuca crudenii (Geophytic herb). Kowie Thicket is bioclimatically the core of the Albany Thicket Biome and the major node of the Albany Centre of Endemism. The location of this vegetation type is adjacent to a wide variety of other vegetation types namely: Albany Coastal Belt, Suurberg Shale Fynbos, Albany Broken Veld, Suurberg Quartzite Fynbos, Great Fish Noorsveld, Southern Coastal Forest and Eastern Cape Thronveld. Co-occurring of species from different vegetation types may result from these overlapping areas which are indicative of varying floristic influences on this vegetation. The description of Kowie Thicket was according to Mucina and Rutherford (2006). 27 • Woodridge College The Woodridge College (Figure 2) is a private co-educated school in the Eastern Cape situated along the N2 highway on the edge of the Van Stadens River gorge near a small town called Thornhill which is roughly about 40 km between Port Elizabeth and Jeffrey’s Bay (Woodridge official website 2010). The College is situated on a beautiful country estate, where its environment is actively used for outdoor activities and environmental awareness programmes (Woodridge official website 2010). Woodridge also runs various projects such as establishment of an organic vegetable garden, recycling, and the setting up of cameras for leopard monitoring in the Baviaan’s Kloof (W. Berrington, pers.comm). The climate at Woodridge is the similar to Port Elizabeth in that it varies considerably. The Port Elizabeth area has both summer and winter rainfall however the maximum rainfall is experienced during winter due to its westerly position, where it catches the tail end of the cold fronts that bring winter rainfall to the Western Cape (Schovell 2007). The Woodridge College is situated on a plateau which drops steeply into the Van Stadens River gorge (W. Berrington, pers.comm). The vegetation of the plateau was formally grassland which has now been transformed to alien invasive vegetation due to high human impacts (W. Berrington, pers.comm). The dominant vegetation type of this study area is: • Van Stadens Forest Thicket Mosaic (Figure 3) This vegetation type is a mosaic thicket that consists of a complex mixture of Gamtoos thicket and subtropical forest species. The dominant species occuring in this vegetation type include: dominant trees Cape chestnut (Calodendrum capense) and red current (Rhus chirindensis) and the rare tree Steculia alexandri is characteristic. Tree euphorbias (Euphorbia triangularis) are present but are never dominant (Mucina and Rutherford 2006). 3.1.2 Field measurments At each of the study sites approximately 50 cycads were tagged and measured. Plants were randomly selected and the number of plants measured was dependant on the 28 number of individuals present at each of the study sites. Plants were individually tagged by nailing numbered metal plates to the middle of the trunk using 25 mm long nails thereby making it minimally conspicuous to animals in the area. Once tagged the height, and diameter were measured, and the age of the plant was recorded by counting the number of leaf base whorls. In the Kaboega area plants were tagged and measured on the south-facing slopes that are moister than the north-facing slopes. Initial measurements were taken in June 2010 during periods of high rainfall (396 mm per year) and temperatures ranged between 7°C and 13°C (SA Weather Service, 2010). After a year m easurements of the same individuals were taken in July 2011 during periods of medium rainfall and temperatures ranged between 16°C and 22°C. In the Zuurberg area species were also measured on the south-facing slopes. Because tagging was not permitted a diagram of plant positions was drawn at the time of the initial measurements taken in June 2011 during periods of high rainfall and heavy winds, with temperatures ranging from 17°C and 20°C. Follo w-up measurements were taken at the end of November 2011 with temperatures ranging between 16°C and19°C. In the Woodridge area species were tagged and measured in April 2011 during periods of minimal rainfall and surrounding temperatures ranging between 22°C and 25°C. Follow up measurements were taken at the end of November 2011, temperatures ranging between 19°C and 22°C. 3.1.3 Data Analysis The frequency of height classes for the plants at each of the study sites was analysed using 20 cm bin sizes. Plants ranged from less than 5 cm to 219 cm in height. The relationship of height and diameter and age was determined using the number of leaf base whorls as a measure of age (Giddy 1974). Prior to statistical analysis data was tested for normality using Shapiro-Wilks test in Statistica ® 10.0. 29 A Kruskal-Wallis test fro multiple independent samples was used to determine if there was a significant difference in the population size structure of the three study sites. 3.2 Germination Encephalartos longifolius seeds were soaked in distilled water for 2 days to remove the fleshy outer layer and then soaked in fungicide for 3 days after which they were soaked in distilled water for a further 4 days to allow for imbibition. Seeds were then potted into 20 seed trays containing vermiculite. Each tray contained 10 seeds, and was placed in a dark growth cabinet with temperatures ranging from 28°C to 33°C. The germination process started in September 2010 and seeds were watered twice a week using a spray bottle to prevent any disturbance to the vermiculite. Each of the seed trays were numbered and trays 1-15 were placed away from the heat source, and trays 16-20 were placed close to the heat source in order to determine the effect of heat on germination. In February 2011 each of the seed trays was wrapped with commercial cling wrap in order to keep the humidity constant. 3.3 Photosynthetic physiology Five seedlings were used to determine whether Encephalartos longifolius has C3 or CAM metabolism. Photosynthesis-Irradiance relationships were determined at midday and light was measured from 0-1500 µmol m-2 s-1. The effect of temperature on photosynthesis was determined by exposing five seedlings to a range of different temperatures in a growth chamber starting with an initial temperature of 20°C. Temperature was adjusted every 3 days by 3°C. An AD C LCpro+ portable photosynthesis system Infrared Gas Analyser (IRGA) was used to determine the photosynthetic rates and irradiance. The cuvette used was a broad chamber that had an area of 6.25 cm2 and all measurement were conducted at a light intensity above the Ik value i.e. 200 µmol m-2 s-1. The carbon dioxide concentration inside the chamber was approximately 421 ppm which was close to the average atmospheric concentrations. The photosynthesis-irradiance relationship was presented using the Harrison et al. (1985) PI Curve. 30 Chapter 4 Results 4.1 Demography Demographic data were non-parametric at all three sites (Table 2). Table 2: A Shapiro-Wilks test for normality for the demographic data collected at all three sites Sites Kaboega Zuurberg Woodridge Statistical W p W p W p Height 0.892 <0.05 0.875 <0.05 0.931 <0.05 Diameter 0.890 <0.05 0.958 <0.05 0.921 <0.05 Age 0.900 <0.05 0.874 <0.05 0.911 <0.05 parameters The Zuurberg and Woodridge populations had more juveniles (<60 cm tall) in comparison to Kaboega (Figure 4). Only the Zuurberg population had a J-shaped demography (Figure 4b) indicative of a growing population. At Kaboega juvenile numbers comprised only a quarter of the population (12 individuals < 60 cm tall) (Figure 4a). The population is growing at Kaboega, however the number of offspring produced per reproductive season is low. The Zuurberg population, by contrast, had almost 60% of the population as juveniles (Figure 4b, 28 individuals). Juvenile were also abundant in the Woodridge population (almost 40% of the population, Figure 4c). Gaps in the height classes could be a result of individuals being destroyed by animal impacts, human activities or fires. A Kruskal- Wallis test showed that there was a significant difference between the height of the populations at Kaboega and Zuurberg (H= 7.59; N= 100; p< 0.05). 31 >220 200-219 180-199 160-179 140-159 120-139 100-119 12 >220 200-219 180-199 160-179 140-159 120-139 >220 200-219 180-199 160-179 140-159 120-139 100-119 80-99 60-79 40-59 a 100-119 12 80-99 60-79 40-59 20-39 12 80-99 60-79 14 40-59 14 20-39 0-19 Frequency (number of individuals per height class) 14 20-39 0-19 Frequency (number of individuals per height class) 16 0-19 Frequency (number of individuals per height class) 16 N = 50 10 8 6 4 2 0 Height classes (cm) b 10 N = 50 8 6 4 2 0 Height (cm) 16 c 10 N = 43 8 6 4 2 0 Height (cm) Figure 4: Frequency of plant height of Encephalartos longifolius at (a) Kaboega, (b) Zuurberg, (c) Woodridge, measured in June 2010. 32 There was a significant correlation between plant height and age at all three study sites (Figure 5, Kaboega: R= 0.92, N = 50, p < 0.05; Zuurberg: R= 0.97, N = 50, p < 0.05; and Woodridge: R= 0.95, N = 43, p < 0.05). As a result height can be used as a predictor of age without having to count the number of rings of leaf bases to obtain the age. There was a significant correlation between plant height and age at all three study sites combined (Figure 16, Appendix B: R= 0.93, N= 143, p < 0.05). As a result of this the relationship between plant height and age can be applied to all Encephalartos longifolius plants and other Encephalartos species with similar life histories. The relationship of diameter and height (Figure 6) also showed a significance correlation at Zuurberg: (R= 0.459; N= 50; p < 0.05); and Woodridge: (R= 0.671, N= 50; p < 0.05). In these populations plants grew thicker as they got taller but only up until approximately 60 cm after which the diameter did not increase further even though the stem continued to grow taller. 33 a 450 y = 0.9155x + 17.193 400 Height (cm) 350 300 250 200 150 100 50 0 0 50 100 150 200 250 300 350 400 450 Age (Years) 250 b y = 1.0542x + 11.639 Height (cm) 200 150 100 50 0 0 50 100 150 200 250 Age (Years) 250 c y = 1.185x + 17.937 Height (cm) 200 150 100 50 0 0 50 100 150 200 250 Age (Years) Figure 5: The relationship between height and age at (a) Kaboega, (b) Zuurberg, (c) Woodridge measured in June 2010. 34 Diameter at breast height (cm) 160 a 140 y = -0.0181x + 110.13 120 R = 0.008 2 100 juveniles 80 adults 60 y = 1.4365x + 11.519 40 R = 0.6336 Linear (juveniles) 2 Linear (adults) 20 0 0 50 100 150 200 250 300 350 400 450 Height (cm) Diameter at breast height (cm) 200 b 180 160 y = 0.5931x + 73.337 140 R = 0.1658 2 120 juveniles 100 adults 80 Linear (juveniles) 60 40 y = 0.4279x + 65.303 20 R = 0.0773 Linear (adults) 2 0 0 Diameter at breast height (cm) 250 50 100 150 Height (cm) 200 250 c 200 y = -0.1006x + 158.33 2 R = 0.0277 150 Juveniles Adults 100 Linear (Juveniles) y = 0.2672x + 86.562 50 Linear (Adults) 2 R = 0.0218 0 0 50 100 150 200 250 Height (cm) Figure 6: Relationship between plant diameter and height at (a) Kaboega, (b) Zuurberg, (c) Woodridge. 35 4.2 Growth Rates Using the regression of height and age growth rates Encephalartos longifoilus can be calculated as 1.1 cm + 0.1 cm (S.E.) per year (Figure 5). This low growth rate is confirmed by using the follow-up measurements of plants at Kaboega that showed that only 18% of the population increased in height, 76% of the population had no height increase and 6% of the individuals in the population were destroyed as a result of animal impacts (Figure 7). The 18% of individuals with a height increase were mainly juvenile plants (Figure 8). At Zuurberg and Woodridge there was no significant increase in plant height over the short period Zuurberg: H= 0, N= 100, p < 0.05; Woodridge: H= 0.014, N= 86, p < 0.05). 80 Height Frequency (%) 70 60 50 40 30 20 10 0 0 1 2 Height (cm) Figure 7: Frequency of change in plant height of Encephalartos longifolius at Kaboega, measured in July 2011. 36 2.5 Height (cm) 2 1.5 1 0.5 0 0 50 100 150 200 250 300 350 400 450 Height (cm) Figure 8: Change in plant height in juvenile and adult plants of Encephalartos longifolius at Kaboega, measured between June 2010 and July 2011 (Vertical Bars = +S.E.). 4.3 Germination For the first five months of the germinating period no seeds germinated (Figure 9) because moisture conditions were too low (a soil moisture of 35% of dry mass). Once moisture conditions were increased (raised to 76%) seeds began to germinate. There was a steady increase in germination for the next three months (Figure 9), after which no further germination occurred. Germination was higher at 28°C (Figure 10), compare d to 33°C, because higher temperatures resulted in lower moisture conditions (28%). The germination success decreased as temperature increased and soil moisture decreased (Figure 10). High moisture conditions resulted in more seeds germinating however due to a longterm persistence of high soil moisture conditions rotting of seeds occurred (Figure 11). At a low percentage moisture content no rotting occurred (Figure 11), indicating that high temperatures rapidly lower soil moisture content therefore making germination conditions unfavorable. Whilst high moisture levels maybe beneficial to seed germination too much moisture can also be detrimental to the seed in the long-term. 37 The overall germination rate of Encephalartos longifolius was only 19.5% (Figure 9). Germination Rate (%) 25 20 15 10 5 t Se pt em be r Au gu s Ju ly Ju ne ay M Ap ril ar ch M ct ob er No ve m be r Ja nu ar y Fe br ua ry O Se pt em be r 0 Months (September 2010-2011) Figure 9: Overall germination of Encephalartos longifolius under controlled conditions over a 1 year period. 35 Frequency (%) 30 25 20 15 10 5 0 28 29 31 33 Temperature (°C) Figure 10: Frequency of germinated Encephalartos longifolius seeds under controlled conditions. 38 Number of seeds rotted 12 10 8 6 4 2 0 28 35 76 Soil moisture content (%) Figure 11: The number of rotten Encephalartos longifolius seeds at three different moisture conditions. 4.4 Photosynthetic physiology Seedlings of Encephalartos longifolius have C3 photosynthesis (Figure 12) – they fix carbon in the day. The saturation irradiance of Encephalartos longifolius seedlings was 600 µmol m-2 s-1 (Figure 12). Photosynthesis increased exponentially with an increase in temperature (Figure 13, solid line). The optimum temperature for Encephalartos longifolius was 33°C after which photosynthesis declined (Figure 13, dotted line). 39 Figure 12: The photosynthesis-irradiance relationship of Encephalartos longifolius seedlings (Vertical bars = ± S.E., n = 3) (Harrison et al. 1985). Figure 13: The photosynthesis–temperature relationship of Encephalartos longifolius seedlings (Vertical bars = ± S.E., n=3). 40 Chapter 5 Discussion The photosynthetic response of Encephalartos longifolius to increased temperatures was as expected for C3 ornamental or woody plants (Sage 1996; Ehleringer and Cerling 2002). Whilst temperature increases photosynthesis this is only as short-term response of the plant and as the temperature approaches the plants optimum range, photosynthesis (CO2 uptake) begins to level out (Marler and Willis1997). This is a typical response of a C3 plant (Sage 1996; Ehleringer and Cerling 2002) but differs from the response of CAM plants that are more adapted to high temperatures by fixing carbon dioxide at night when temperatures are much lower, therefore CAM plants have a much higher water use efficiency because stomata are closed during the day (Ehleringer 1979; Nobel and Hartsoc. 1986; Drennan and Nobel 2000). The temperature rise predicted for the Eastern Cape together with predicted decreases in rainfall will increase the severity of droughts (Midgley et al. 2007). In response to drought cycads will have increased stomatal closure therefore reducing photosynthesis and transpiration and lowering the cost of growth (Ward et al. 1999). Other responses of C3 woody plants to drought include senescing and dropping of many older leaves thereby resulting in large reductions in leaf area in order to minimize water loss (Marler and Willis 1997; Ward et al. 1999). As a result of drought the high temperatures and low moisture conditions are expected to be detrimental to seed germination of Encephalartos longifolius. The germination trials of this study found that Encephalartos longifolius germinates best at temperatures between 28°C-30°C a nd moisture conditions between 50% and 75%. The predicted increased transpiration rates as a result of increased temperature could result in reduced seed moisture content that could contribute to seed dormancy (Giddy 1974). If germination takes long to be initiated seed dormancy will persist and eventually result in infertility (Giddy 1974; Dehgan 1983) this coincides with the initial slow germination rates of this study. Increase in wildfires is also associated with predicted temperature rise (IPCC, 2007) and studies done by Donaldson (1995) showed that Encephalartos species have a high 41 incidence of mast-seeding in response to fire (Janzen 1971). This was comparable to the demographic data collected at the Kaboega study site in which low juvenile numbers resulted from fires that swept the Southern slopes approximately 8 years ago triggering mast-seeding (I. Ritchie, pers comm). Other studies have found fire to have a deleterious effect on Australian cycad species seeds (Hill 2003). While no such data exist for Encephalartos longifolius it is assumed that intensity in wildfires could be a critical factor in the long-term viability of the population especially if seeds and seedlings are killed by fire thereby disrupting regeneration. Seedlings are sometimes more susceptible to fire due to the possession of lignotubers (a swelling at the base of the stem just above the soil level) that helps them survive fires by sending out new growth if the top growth is damaged (Plate 1, Appendix A) (Chamberlain 1925). Adults are also vulnerable to fires which probably account for the gaps in the older classes as recorded at Kaboega (Chamberlain 1925; Donaldson 2003). Whilst C3 plants may be disadvantaged in hot dry climates due to photorespiration Zuo et al. (2004) found little effect of increased temperature on the function of the chloroplasts of Cycas multipinnata and Cycas panzhihuaensis which was indicative of cycads adapting to the prevailing climatic conditions. The exponential photosynthesis response of Encephalartos longifolius to temperature increases is too simplified to derive conclusions of climate change impacts on cycads especially since climate change does not only involve temperature rise but it also involves a rise in carbon dioxide concentrations. It is predicted that as a result of global carbon dioxide concentrations rising at an alarming rate global temperature will also rise (IPCC, 2007), therefore the rise is carbon dioxide and temperature are going to occur concomitantly (IPCC, 2001; Kim et al. 2007). While temperature increases in the Eastern Cape seem to be unfavorable to C3 plants, a rise in carbon dioxide concentrations also need to be taken into consideration since it is the basic raw material that plants require for photosynthesis (Ehleringer 1979). Based on the typical response of C3 woody plants to elevated temperature assumptions were made that Encephalartos longifolius will follow the similar C3 response to elevated carbon dioxide concentrations. An increase in carbon dioxide concentrations is postulated to favor plant growth and development especially in C3 plants (Ehleringer and Cerling 2002). Even though Encephalartos longifolius showed an exponential rise with increasing temperature, the temperature optimum for plant growth 42 will be much higher in response to predicted elevated CO2 concentrations and photorespiration will be inhibited therefore making photosynthesis much more efficient (Ward et al. 1999; Kim et al. 2007). Elevated carbon dioxide levels will also mitigate the impacts of drought by producing a more convoluted root system which will ease the accessibility of water therefore increasing water use efficiency (Ward et al. 1999). The roots of Encephalartos longifolius are well known for their symbiotic relationship with blue-green alga Anabaena. As a result of elevated carbon dioxide concentrations this symbiotic relationship will be enhanced and will help the plant to adapt to hot dry climatic conditions (Saxe et al. 1998). Newton et al. (1994) also found that the enhancement of this symbiotic relationship by elevated CO2 concentrations will be of great benefit to slow-growing plants and cycads are well known for their slow-growth rates (Chamberlain 1919). This was comparable to the data of this study that showed that Encephalartos longifolius grew just over 1.1 cm per year and then only in the juvenile plants. Growth and biomass of Encephalartos longifolius is also assumed to increase as a result of increased carbon dioxide levels because according to Norby et al. (1999) carbon dioxide is essential for wood formation in woody plants therefore increased concentrations of carbon dioxide will speed up wood formation and thickening resulting in greater plant height and diameter. Increased carbon levels will also contribute to more active and extensive root systems, as well as further enhance cone and seed production and leaf stomatal density (Atwell et al. 2003; Yazaki et al. 2005). Increased carbon dioxide levels are presumed to be beneficial to cycad growth, however these are only short-term responses as for all C3 woody plants and in the long-term photosynthesis will eventually reach equilibrium as plants reach maturity and take up less carbon dioxide regardless of further CO2 increases (Tissue et al. 1995; Cheng et al. 2000; Norikane et al. 2010). Whilst increased carbon dioxide concentrations might prove to be more beneficial to C3 plants in comparison to temperature by outweighing the effects of increased temperatures the interaction of these two climate variables need to be considered for possible climate change impacts on plants. The outcomes of the interaction of these two variables on the Eastern Cape environment according to the predictions of the IPCC (2007) and Midgley et al. (2007) may prove to be a disadvantage to Encephalartos 43 longifolius over time. Whilst there is no data to support this theory a possible impact of climate change on the Eastern Cape environment that could impact Encephalartos longifolius is the intense frequency of storms which would result in an extreme leaningover effect of the taller plants in the population (Plate 2, Appendix A) due to its nonextensive root system (Chamberlain 1919). Based on theoretical understanding and historical data the hypothesis of this study was accepted. Cycads existed in a time when temperature and carbon dioxide levels were much higher than either the present (Figure 14) and the levels predicted for the future (Figure 15). When cycads first appeared in the late Carboniferous period the average global temperatures were 12°C and carbon dioxide co ncentrations were 350 ppm which is similar to the present temperature of 15.5°C and carbon dioxide concentration of 390 ppm (Figure 14 and Figure 15) (Plass 1956; Buckley 1999; IPCC, 2007). During the first glaciations period when global temperatures and carbon dioxide concentrations rapidly decreased this resulted in a decrease in cycad abundance (Figure 14); however when temperature and carbon dioxide concentrations began to raise again in the late Permian period, cycad abundance started to increase in a recovery phase (Figure 14) (Buckley 1999). It was during the Jurassic period that cycads reached their peak abundance were average temperatures were about 25°C and carbon dio xide concentrations about 1200 ppm, after their numbers began to decline and this continues to the present (Figure 14) (Plass 1956; Buckley 1999). 44 Figure 14: Fossil records of cycad abundance with changing temperature and carbon dioxide levels through geological time (Plass 1956; Buckley 1999). A comparison of past and present climatic conditions reveal that even though the average global temperature and carbon dioxide concentrations of the Jurassic period were much higher than that of the present (Figure 15), cycads still reached their peak abundance at that time thereby indicating that they were well adapted to such conditions. 45 Figure 15: Projections of past, present and future temperature levels and carbon dioxide concentrations according to the IPCC, 2007 Assessment Report and Midgley et al. 2007. Even though global temperatures and carbon dioxide levels are expected to increase in the future with average temperatures of about 21°C and carbon dioxide concentrations of about 1000 ppm (Figure15) , historically the earth has been much hotter and drier and had a higher concentration of carbon dioxide compared to present times, therefore assumptions can be made that future increases in temperature and carbon dioxide levels as outlined in (Figure 15) will not impact on the abundance of existing cycad populations. Based on the studies of McElwain (1998) on stomatal data, it is believed that cycads were well adapted to fluctuations in temperature and carbon dioxide levels and were able to acclimate themselves to the changes in these two climatic variables thereby allowing them to persist in the environment for many years. Regardless of predicted climate changes in the Eastern Cape Encephalartos longifolius will not be directly impacted by this change because increase in temperature and carbon dioxide concentrations will occur gradually over time, therefore allowing the species to adapt and acclimate to the changes, as predicted for cycad Dioon edule to fluctuations in CO2 levels and stress induced conditions (Vovides et al. 2002). 46 Encephalartos longifolius is a near threatened species and whilst climate change may not directly impact the species there are many other factors that are contributing to its declining population numbers especially in the wild. The demographic data of this study attempted to determine the vulnerability of the species in the wild by using the presence of juvenile numbers as an indicator of the health of the population with more juveniles indicating a healthy and growing population and fewer juveniles indicating an unhealthy population (Gallego 2007). Juvenile numbers at Kaboega were low. A preliminary study of the health of cycad species Encephalartos longifolius in 2008 showed the population to be healthy with a large number of juveniles present (Wakeford et al. 2008). Two years later the initial results of this study showed a rapid decline in juvenile numbers, and a further decline the year after. Juvenile numbers at Zuurberg and Woodridge were fairly high which indicates these populations are healthy. Ruling out the impacts of climate change the major drivers to this decreasing population trend of Encephalartos longifolius are human and animal impacts. Whilst this study did not focus on human and animal impacts, data from past studies and visual observance was used to explain the population trends found at the three study sites. The animal impacts at Kaboega on cycad species include: trampling and knocking over by large mammals (Plate 3, Appendix A); baboons having the highest impact as they feed on cones and seeds therefore preventing seeds from germinating (Plate 4, Appendix A); warthogs dig out seedlings to consume roots and bulbs; and porcupines eat mainly at the base of the trunk therefore making it more vulnerable to be knocked over by large mammals (Plate 3 and Plate 5, Appendix A) (R.M. Cowling, pers.comm; I. Ritchie, pers.comm). As a result of such animal impacts the declining juvenile population could also be a result of the mast-seeding phenomenon in which a large amount of seeds are produced synchronously within a plant population in one year followed by an interval in which few or no seeds are set in order to minimize predator satiation (Janzen 1971). This theory was tested by Donaldson (1993) on the genus Encephalartos in which he found that low levels of seed predation resulted from species that produced seeds sporadically. Encephalartos longifolius reproduces sporadically which is contributing to this declining juvenile number because even though mast-seeding is effective in limiting predator impacts on the plant, the survival of the seed is very low and sometimes germination does not occurs (Ruth et al. 1986; Li et al. 2007). 47 Animal impacts at Zuurberg were of a lesser extent than Kaboega, the results of this study for Zuurberg showed a bimodal distribution pattern in height classes which coincides with the results found by Kamineth (2004) on the impacts of megahebivores on tree euphorbias. The presence of megaherbivores in an area results in larger individuals being felled rather than juveniles, juveniles would eventually mature into reproductive adults and produce a cohort of juveniles and the ages of these cohorts would depend on the absence of the megaherbivores from the area. Megaherbivores were present in the area before the 1750’s (Kamineth 2004), which is almost about 261 years ago which explains the gaps in the older height classes (200-219 cm), and no cycad older than 220 years was encountered in this study (personal observations). Apart from the impacts of megaherbivores other animals such as baboons and porcupines also impact the species but to a much lesser extant in comparison to Kaboega (Donaldson 2008). Animal impacts are not always a disadvantage to the population. Some impacts prove to be quite beneficial to the population especially in terms of seed dispersal (Giddy 1974). Birds in particular are well-known for the role in seed dispersal as they carry seeds away from the parent plant over long distances (Giddy 1974). The results of this study at the Woodridge population support this theory because the population of cycads at Woodridge had a very scattered distribution (Giddy 1974; Tiffney 2004; personal observations). Dispersal increases the seeds chances of survival as it allows seeds to escape from potential predators and pests; it reduces parent-offspring and sibling competition and seeds are spread over large areas (Tiffney 2004), which would explain the high juvenile numbers at Woodridge. Apart from animal impacts, human impacts are one of the major threats responsible for declining wild populations and removal of plants for trade is the most common human impact (Osborne 1995; Donaldson 2003). As expected, wild populations are vulnerable to human impacts. In 1978 the IUCN records showed that approximately 4500 Encephalartos longifolius plants were present in the wild, currently this number has declined between 50% and 70% (Anderson et al. 2007; Donaldson 2003; Donaldson 2008). In the past plants at Zuurberg were removed for the following purposes: trade to collectors, used in private collections as ornamental plants, medicinal uses and for illegal 48 seed harvesting and selling of seedlings. Currently the remaining individuals have tracking devices inserted into their trunks which has been effective in minimizing removal rates (Wilson, pers.comm). Low adult numbers and gaps in the adult height classes at Woodridge are a result of habitat destruction during previous years; however restricted access to the school and continued monitoring has contributed to the currently high juvenile numbers (W. Berrington, pers.comm). The threats of climate change on Encephalartos longifolius are poorly understood, however the response of C3 plants to elevated temperatures and carbon dioxide concentrations coupled with historical data and responses of other cycad species to environmental stress indicate that Encephalartos longifolius will be able to adapt to the predicted climate change for the Eastern Cape and persist in the environment for the next 200 years, but particular concern needs to be given to the threats of human and animal impacts on the abundance of wild populations. 49 Chapter 6 Conclusion and Recommendations The main focus of this study was to gain insights on the potential impacts of predicted climate change in the Eastern Cape on Encephalartos longifolius and the results of this study can be used as a baseline study for all South African cycads since very little is known about the photosynthetic physiology of these extraordinary plants. Like all cycads Encephalartos longifolius is very-slow growing which is often the primary cause of their decline in the wild. Seeds that take long to germinate often lose their viability and seedlings that take long to mature are more susceptible to predator attacks, however the concomitant relationship between predicted increase in temperature and carbon dioxide could prove to be beneficial to the plants growth rate. Low juvenile numbers in the wild are also a matter of concern for the long-term persistence of the plants in the environment. The data of this study has shown that Encephalartos longifolius has a typical C3 response to increasing temperatures and elevated carbon dioxide concentrations. Whilst increased temperatures may be detrimental to the plant coupled with elevated carbon dioxide levels this could prove to be beneficial to the plant; however these responses will only be short-term. The impacts of climate change on cycads is poorly understood but based on historical data cycads tend to be well adapted to handle fluctuating environmental conditions and stress. Over the years cycads have acclimated to climatic changes and will continue to do so in the future. The primary feature of cycads are their extensive root system that allows them to adapt to periods of drought and low moisture and its symbiotic relationship with soil microbes provides survival for the plant in nutrient poor soils. Predicted climate change for the Eastern Cape is concluded to have no impact on the survival of Encephalartos longifolius, however the population is declining in the wild and based on present knowledge animal and human impacts are a major cause of concern for the plants ruin in the wild. 50 Although this study serves as a preliminary investigation into the population dynamics of Encephalartos longifolius and its response to predicted climate change in the Eastern Cape it has been very valuable in identifying the physiology of cycads and the major contributors to declining plant numbers in the wild therefore this study can be used as a baseline study for other South African Encephalartos species with similar life histories. I recommend the following for future studies and effective conservation measures: • Photosynthetic physiology of cycads needs to be investigated in greater depth with particular focus on South African cycads. • Experiments need to be set up in order to directly assess animal impacts on wild populations. • Monitoring should be designed to enable an assessment of population size and age-structure dynamics; coning and mast-seeding events; and the response of seeds, seedlings and adult plants to fires. • Habitat condition should be assessed especially in relation to alien invasive abundance. • Experiments need to be set up to monitor pollinator abundance and movements which could provide useful information for increasing seed production in ex situ conservation. Cycads are an extraordinary group of plants that can persist in our environment for many years to come provided that threats to their survival are minimized as best possible. 51 References Anderson JM, Anderson HM, Cleal CJ (2007). Brief history of the gymnosperms: classification, biodiversity, phytogeography and ecology. Strelitzia 20. South African National Biodiversity Institute, Pretoria. Atwell BJ, Henry ML, Whitehead D (2003). Sapwood development in Pinus radiata trees grown for three years at ambient and elevated carbon dioxide partial pressures. Tree Physiology 23: 13-21. Ballardie RT, Whelan RJ (1986). Masting, seed dispersal and seed predation in the cycad Macrozamia communis. Oecologia 70: 100-105. Brenner ED, Stevenson DW, Twigg RW (2003). Cycads: evolutionary innovations and the role of plant derived neurotoxins. Trends in Plant Science 8: 446-452. Buckley R (1999). The fossil cycads: a brief review. Available from site http://www.plantapalm.com. Accessed on 23/03/2010. Chamberlain CJ (1919). The living cycads. University of Chicago Press, Illinois. Chamberlain CJ (1925). The origin of cycads. Science New Series 61: 73-77. Cheng W, Sims DA, Luo Y, Coleman JS, Johnson DW (2000). Photosynthesis, respiration, and net primary production of sunflower stands in ambient and elevated atmospheric CO2 concentrations: an invariant NPP:GPP ratio? Global Change Biology 6: 931-941. Clarke DA, Clarke DB (1987). Temporal and environmental patterns of reproduction in Zamia skinneri, a tropical rain forest cycad. Journal of Ecology 75: 135-149. 52 CITES (2003). Review of significant trade cycads. Available from site http://www.cites.org/eng/app/index.php. Accessed on 27/12/2011. Dehgan B (1983). Propagation and growth of cycads. A conservation strategy. Pro, Fla, State Hort, Soc, 96: 137-137. Dehgan B, Yuen CKKH (1983). Seed morphology in relation to dispersal, evolution, and propagation of Cycas L. Botanical Gazette 144: 412-418. Department of Economic Development and Environmental Affairs (DEDEA) (2011). Eastern Cape climate change response strategy. Accessed on Available from site www.deaet.ecprov.gov.za. Accessed on 27/12/2011. Donaldson JS (1993). Mast-seeding in the cycad genus Encephalartos: a test of the predator satiation hypothesis. Oecologia 94: 262-271. Donaldson JS (1995). Understanding cycad life histories: an essential basis for successful conservation. In: Donaldson Js (ed) Cycad conservation in South Africa: issues, priorities, and actions. Cycad Society of South Africa, Stellenbosch, South Africa, pp 8-13. Donaldson JS (1997). Is there a floral parasite mutualism in cycad pollination? The pollination biology of Encephalartos villosus (Zamiaceae). American Journal of Botany 84: 1398-1406. Donaldson JS (2003). Cycads Introduction. In: Donaldson JS (ed) Cycads. Status survey and conservation action plan. IUCN/SSC. Cycad Specialist Group. IUCN, Gland, Switzerland and Cambridge, UK. Donaldson JS, Hill KD, Stevenson DW (2003). Cycads of the world: an overview. In: Donaldson JS (ed) Cycads. Status survey and conservation action plan. IUCN/SSC. Cycad Specialist Group. IUCN, Gland, Switzerland and Cambridge, UK, pp 3-8. 53 Donaldson JS (2008). South African Encephalartos species. Succulents and cycads. NDF Workshop Case Study 4: 1-20. Drennan PM, Nobel PS (2000). Responses of CAM species to increasing atmospheric CO2 concentrations. Plant, Cell and Environmental 23: 767-781. Ehleringer JR (1979). Photosynthesis and photorespiration: biochemistry, physiology and ecological implications. HortScience 14: 217-222. Ehleringer JR, Cerling TE (2002). C3 and C4 photosynthesis. Encyclopedia of Global Environmental Change volume 2. John Wiley & Sons, LTD, Chichester. Gallego CL (2007). Cycad conservation. Guidelines for establishing your own population monitoring program. The Cycad Newsletter 30: 36-38. Germishuizen G, Meyer NL, Steenkamp Y, Keith M (2006). (eds) In: A checklist of South African plants. South African Botanical Diversity Network Report No 41. SABONET, Pretoria, South Africa. Giddy C (1974). Cycads of South Africa. Purnell and Sons (S.A.) (PTY) LTD, Cape Town, South Africa. Harrison WG, Platt T, Lewis MR (1985). The utility of light saturation models for estimating marine primary productivity in the field: a comparison with conventional “stimulated” in situ methods. Canadian Journal of Fish and Aquatic Science 42: 864-872. Hertrich W (1951). Palms and Cycads. CF Braun and Co, Albambra, California. Hill KD (2003). Regional overview: Australia. An overview. In: Donaldson JS (ed) Cycads. Status survey and conservation action plan. IUCN/SSC. Cycad Specialist Group. IUCN, Gland, Switzerland and Cambridge, UK, pp 20-24. 54 IPCC (2000). Special Report on emissions scenarios. [Nakicenovic N and Swart R (eds.)] Cambridge University Press, UK. IPCC (2001). Climate Change 2001: Impacts, adaptation and vulnerability. Contribution of Working Group II to the Third Assessment Report of the Intergovernmental Panel on Climate Change. [McCarty JJ, Canziani NA, Dokken DJ, White KS (eds.)]. Cambridge University press, UK. IPCC (2007). Climate Change 2007: Synthesis Report. Contribution of working groups I, II and III to the Fourth Assessment Report of the Intergovernmental Panel on Climate Change [Pachauri RK and Reisinger A. (eds).] IPCC, Geneva, Switzerland. IUCN (1997). IUCN Red List of Threatened Species. Available from site http://www.redlist.org. Accessed on 20/05/2010. IUCN (2006). IUCN Red List of Threatened species. Available from site http://www.redlist.org. Accessed on 20/05/2010. IUCN (2010). IUCN Red List of Threatened Species. Available from site http://iucnredlist.org/apps/redlist/details/41935/0. Accessed on 27/12/2011. Janzen DH (1971). Seed predation by animals. Ann Rev Ecol Syst 2: 465-492. Johnston P, Coop L, Lennard C (2011). Climate change projections and impacts for the Eastern Cape region of South Africa. Climate Systems Analysis Group. Cape Town, South Africa. Kamineth AI (2004). The population dynamics and distribution of tree euphorbias in thicket. BSc Masters Project, Department of Botany, Nelson Mandela Metropolitan University, Port Elizabeth, South Africa. (Unpublished). 55 Kerley G, Boshoff A (1997). A Regional and National conservation and development opportunity. Terrestrial Ecology Research Unit. Department of Zoology, University of Port Elizabeth, Port Elizabeth, South Africa. Report No. 17: 4-61. Kim SH, Gitz DC, Sicher RC, Baker JT, Timlin DJ, Reddy VR (2007). Temperature dependence of growth development and photosynthesis in maize under elevated CO2. Environmental and Experimental Botany 61: 224-236. Li H, Zhang Z (2007). Effects of mast seeding and rodent abundance on seed predation and dispersal by rodents in Prunus armeniaca (Rosaceae). Forest Ecology and Management 242: 511-51. Marler TE, Willis LE (1997). Leaf gas-exchange characteristics of sixteen cycad species. Journal of American Society of Horticulture Science 122: 38-42. Marshak J (2003). Earth: portrait of a planet. 2nd edition. WW Norton and Company Inc, New York. Microsoft Encarta (2005). World conservation monitoring center. Microsoft Coropration [CD]. Miller GT (2007). Living in the environment: principles, connections, and solutions 15th edition. Thomson Learning, Canada. Molles MC (2008). Ecology: concepts and applications 4th Edition. McGraw-Hill Companies Inc, New York. McElwain JC (1998). Do fossil plants signal palaeoatmospheric CO2 concentration in the geological past? Phil Trans R Soc Land B. 353: 83-96. Midgley G, Chapman R, Mukheibir P, Tadross M, Hewitson B, Wand S, Schulze R, Lumsden T, Horan M, Warburton M, Kgope B, Mantlana B, Knowles A, Abayomi A, Ziervogel G, Cullis R, Theron A (2007). Impacts, vulnerability and adaptation in key 56 South African sectors: an input into the long-term mitigation scenarios process. University of Cape Town, South Africa. Miguel A, Farrera P, Vovides AP, Octavio-Aguilar P, Gonzalez-Astorga J, de la CruzRodriguez J, Jonapa RH, Villalobos-Mendez SM, Perez-Farrerra MA (2006). Demography of the cycad Ceratozamia mirandae (Zamiaceae) under disturbed and undisturbed conditions in a Biosphere Reserve of Mexico. Plant Ecology 187: 97-108. Mucina L and Rutherford MC (2006). The vegetation of South Africa, Lesotho and Swaziland, Strelitzia 19. South African National Biodiversity Institute, Pretoria. Newton PCD, Clark H, Bell CC, Glasgow EM, Campbell BD (1994). Effects of elevated CO2 and stimulated seasonal changes in temperature on the species composition and growth rates of pasture turves. Annuals of Botany 73: 53-59. Nobel PS, Hartsock TL (1986). Short-term and long-term responses of CAM plants to elevated CO2. Plant Physiology 82: 604-606. Norby RJ, Wullschleger SD, Gunderson CA, Johnson DW, Ceulemans R (1999). Tree responses to rising CO2 in field experiments: implications for the future forest. Plant, Cell and Environment 22: 683-714. Norikane A, Takamura T, Morokuma M, Tanaka M (2010). In vitro growth and single-leaf photosynthetic response of Cymbidium plantlets to super-elevated CO2 under cold cathode fluorescent lamps. Plant Cell Reports 29: 273-282. Osborne R (1995). An overview of cycad conservation in South Africa. In: Donaldson JS (ed) Cycad conservation in South Africa: issues, priorities and actions. Cycad Society of South Africa, pp 1-7. Plass GN (1956). Carbon dioxide and climate. American Scientist 44: 302-316. 57 Pierce SM, and Mader AD (2006). Subtropical Thicket Ecosystem Programme Handbook. Integrating the natural environment into land use decisions at the municipal level: towards sustainable development. Centre for African Conservation Ecology (ACE). Report Number 47 (Second Edition). Nelson Mandela Metropolitan University, South Africa. Ruth T, Whelan B, Whelan RJ (1986). Masting, seed dispersal and seed predation in the cycad Macrozamia communis. Oecologia 70: 100-105. Sage R (1996). Atmospheric modification and vegetation responses to environmental stress. Global Change Biology 2: 79-23. SA Weather Service (2010). Eastern Cape Climate. Available from site http://www.weathersa.co.za. Accessed on 27/07/2010. Saxe H, Ellsworth DS, Heath J (1998). Tree and forest functioning in an enriched CO2 atmosphere. New Phytologist 139: 395-436. Schneider D, Wink M, Sporer F, Ad Lounibos P (2002). Cycads: their evolution, toxins, herbivores and insect pollinators. Naturwissenchaften 89: 281-294. Scholze M, Knorr W, Arnell NW, Prentice IC. (2006). A Climate-change risk analysis for world ecosystems. Proceedings of the National Academy of Science of the United States of America 103: 13116-13120. Schovell I (2007). Van Stadens River Conservancy programme: priority land parcels audit. Wildlife and Environment Society of South Africa. 58 Sheikh PA, Gorte RW (2008). Climate change and international deforestation: legislative analysis. CRS Report for Congress. Code number RL34634. South African National Biodiversity Institute (SANBI) (2011). Red Data List of South African Plants version 2011.1. Available from Site http://redlist.sanbi.org/search.php?sppsearch=Encephalartos+ Accessed on 20/09/2011. Suinyuy TN, Donaldson JS, Johnson SD (2009). Insect pollination in the African cycad Encephalartos friderici-guilielmi Lehm. South African Journal of Botany 75: 682-688. Tang W (1987). Heat production in cycad cones. Botanical Gazette 142: 165-174. Tang W, Sternberg L, Price D (1987). Metabolic aspects of thermogenesis in male cones of five cycad species. American Journal of Botany 74: 1555-1559. The Cycad Newsletter (2007). Conservation of cycads. Education and Scientific Research volume 1: 10-16. Tiffney BH (2004). Vertebrate dispersal of seed plants through time. Annual Reviews 35: 1-29. Tissue DT, Griffin KL, Thomas RB, Strain BR (1995). Effects of low and elevated CO2 on C3 and C4 annuals. II. Photosynthesis and leaf biochemistry. Oecologia 101: 21-28. Turpie JK, Lechmere-Oertel R, Sigwela AM, Antrobus G, Donladson J, Robertson H, Skowno A, Knight A, Koelle B, Mills A, Kerley G, Leiman A, Van Zyl H (2002). The ecological and economic implications of conversion to game farming in the xeric succulent thicket of the Eastern Cape, South Africa. In: Turpie, J.K. (ed) An Ecologicaleconomic appraisal of conservation on commercial farmland in four areas of South Africa. National Botanical Institute, Cape Town. 59 Vlok JHJ, Euston-Brown D (2002). The patterns within and the ecological processes that sustain the subtropical thicket vegetation in the planning domain for the Subtropical Thicket Ecosystem Planning (STEP) Project. Terrestrial Ecology Research Unit Report 40. University of Port Elizabeth, Port Elizabeth. Vlok JHJ, Euston-Brown DIW, Cowling RM (2003). Acocks’ Valley Bushveld 50 years on: new perspectives on the delimitation, characterization and origin of Subtropical Thicket vegetation. South African Journal of Botany 69: 27-51. Vorster P, Van der Bank FH, Van der Bank M, Wink M (2004). Phylogeny of Encephalartos: some Eastern Cape species. Botanical Review 70: 250-259. Vovides AP, Etherington JR, Dresser PQ, Groenhop A, Iglesias C, Ramirez JF (2002). CAM-cycling in the cycad Dioon edule Lindl. in its natural tropical deciduous forest habitat in central Veracruz, Mexico. Botanical Journal of the Linnean Society 138: 155162. Wakeford SA, Schmidt J, Baker L (2008). Botany Honours Excursion at Kaboega. Nelson Mandela Metropolitan University, Port Elizabeth. Unpublished. Ward JK, Tissue DT, Thomas RB, Strain BR (1999). Comparative responses of model C3 and C4 plants to drought in low and elevated CO2. Global Change Biology 5: 857-867. Whiting MG (1963). Toxicity of cycads. Economic Botany 17: 270-302. Woodridge official website Available from site www.woodridgecollege.co.za. Accessed on 23/03/2011. Yazaki K, Maruyama Y, Mori S, Koike T, Funada R (2005). Effects of elevated carbon dioxide on wood structure and formation in trees. Plant responses to air pollution and global change. Springer-Verlag, Tokyo. 60 Zuo BY, Zhang Q, Jiang GZ, Chen CJ (2004). The response of ultrastructure and function of chloroplast from cycads to doubled CO2 concentrations. Botanical Review 70: 72-78. Personal communication Dr DR DuPreez. Head of School, Botany Department, Faculty of Science, Nelson Mandela Metropolitan University, Port Elizabeth, South Africa Email: [email protected] Wesley Berrington. Van Stadens Wildflower Reserve, Port Elizabeth South Africa. Email: [email protected] Cowling RM. Researcher in Botany Department,, Faculty of Science, Nelson Mandela Metropolitan University, Port Elizabeth, South Africa. Email: [email protected] Ritchie I. Land manager at Kaboega, Eastern Cape, South Africa Email: [email protected] Wilson, Game Ranger at Zuurberg Pass Park, Eastern Cape, South Africa. Cell: 0733171939. 61 Appendix A List of Plates Plate 1: Lignotuber on Encephalartos longifolius stem in response to fire. Plate 2: Leaning-over effect of Encephalartos longifolius 62 Plate 3: Encephalartos longifolius trampled and knocked over by large mammals Plate 4: Encephalartos longifolius cone damaged by baboons 63 Plate 5: The base of Encephalartos longifolius stem impacted on by porcupines. 64 Appendix B Additional Graph 500 450 400 Age (Years) 350 y = 0.9202x - 9.1828 300 250 200 150 100 50 500 450 400 350 300 250 200 150 100 50 0 0 Height (cm) Figure 16: The relationship between plant height and age at all three study sites combined. 65