The Effect of High Temperature on the Reproductive Success of Trianthema portulacastrum by Haley Anne Branch A thesis submitted in conformity with the requirements for the degree of Master of Science Ecology and Evolutionary Biology University of Toronto © Copyright by Haley Anne Branch 2016 The Effect of High Temperature on the Reproductive Success of Trianthema portulacastrum Haley Anne Branch Master of Science Ecology and Evolutionary Biology University of Toronto 2016 Abstract Plant reproduction is highly sensitive to rising temperatures, which can lead to pollen abortion, and lower yield in many crop species. It remains uncertain whether wild plant species adapted to hot climates are able to reproduce at high temperature. I studied heat sterility thresholds in Trianthema portulacastrum, a weedy species found throughout the tropics and subtropics, often on barren soils where temperature exceeds 40°C. Plants were grown at seven day/night temperatures: 30/24°C, 33/24°C, 36/24°C, 40/24°C, 44/24°C, 24/40°C, and 40/40°C. Pollen viability significantly declined with increasing temperature, but this did not significantly affect percent pollen germination or seed set. In contrast, seed set was significantly reduced under high night temperature. The results show high night temperatures have a greater impact on reproduction than day temperature, indicating T. portulacastrum is using a night escape strategy to maintain reproductive success in its natural habitat. ii Acknowledgments I would like to thank my supervisor, Prof. Rowan Sage, who inspired me to pursue an academic career in plant ecology and helped formulate the ideas for this thesis. He encouraged me to think outside the box, taught me to question everything, and has shown me the importance of passion in research throughout my MSc and during my undergraduate degree. Secondly, thank you to Prof. John Stinchcombe and Prof. Art Weis. I have greatly appreciated your advice and support throughout my Master’s. Thirdly, to Prof. Asher Cutter, thank you for allowing me access to your lab equipment, without you this thesis would not have been completed. A special thank you to Dr. Corlett Wood, who generously gave her time to assist me with my statistical methods. Thank you to my friends and colleagues at the University of Toronto. Stefanie Sultmanis, Colin Bonner, Matt Stata, Vanessa Lundsgaard-Nielsen, and Dr. Roxana Khoshravesh helped me with various aspects of my study and engaged with me in numerous scientific discussions. Michael Foisy, thank you for joining me at libraries and coffee shops while I took on the task of writing this thesis. To my husband, Graham Hassell, thank you for your unconditional support throughout my degree. You listened patiently as I practiced my talks over and over again, waded through unfamiliar jargon to edit my work, and dedicated much time to assist me in nighttime laboratory excursions. For all that you have done, I am grateful. Thank you to my parents, Donald and Cathy Branch, for providing me with the foundation to pursue science intellectually and creatively. To my big sister, Cara-Lynn, you have rooted for me since the very beginning, thank you. This research was funded by an NSERC CGS M scholarship. iii Table of Contents Acknowledgments .................................................................................................................... iii Table of Contents ..................................................................................................................... iv List of Figures ............................................................................................................................v List of Tables ........................................................................................................................... vi Abbreviations .......................................................................................................................... vii Introduction ...........................................................................................................................1 1.1 Heat Studies in Wild Species .........................................................................................6 Methods ...............................................................................................................................12 2.1 Growth Conditions .......................................................................................................12 2.2 Experiment 1: Variable day temperature, constant night temperature ........................13 2.3 Experiment 2: Elevated night temperature ..................................................................14 2.4 Analysis of Anther and Pollen Development ...............................................................14 2.5 Pollen Viability Characterization .................................................................................15 2.6 Characterizing Reproductive Success ..........................................................................16 2.7 Statistical Analysis .......................................................................................................16 Results .................................................................................................................................19 Discussion ...........................................................................................................................25 Conclusion and Future Directions .......................................................................................29 6 References .........................................................................................................................31 iv List of Figures Figure 1-1 Heat-induced abortion in eight diverse genera……………………………………4 Figure 1-2 Average temperatures in the Mojave Desert……………………………………..10 Figure 1-3 Trianthema portulacastrum growth and morphology…………………………....11 Figure 2-1 Flower and air temperature during control and 44/24°C treatments………….....14 Figure 3-1 Anther and pollen development in T. portulacastrum…………………………...20 Figure 3-2 Heat stress affects pollen and anther development…………………..…………..21 Figure 3-3 High temperature affects pollen viability but has no effect on yield.....................22 Figure 3-4 T. portulacastrum does not exhibit apomictic reproduction………..……………23 v List of Tables Table 2-1 Control treatment growing conditions…………………………………………...12 Table 2-2 T. portulacastrum growth stage and timing of sampling………………………...18 Table 3-1 Night growth temperature effects on reproduction………………………………24 vi Abbreviations AB – Aniline blue ANOVA – Analysis of variance ATS – Alexander’s Triple Stain DAPI – 4’,6-diamidino-2-phenylindole FDA – Fluorescein diacetate FITC – Fluorescein isothiocyanate HNT – High night temperature HSP – Heat shock protein ROS – Reactive oxygen species TT – Treatment temperature vii Introduction Climate change is one of the greatest threats of this century. Average global temperatures are expected to increase by up to 2°C by 2050, along with increased frequency and intensity of extreme heat events (Collins et al., 2013). This will have profound impacts on the Earth’s biota, as most biological processes have optimal thermal ranges. Plant reproduction is among the most sensitive of processes to increases in temperature, because a relatively small rise in temperature above 30°C can lead to heat-induced sterility (Hedhly, Hormaza, and Herrero, 2008; Schlenker and Roberts, 2009; Harsant et al., 2013; Sage et al., 2015). Because of this, heat stress is of particular concern with climate warming in regions of the world that currently experience maximum growing season temperatures above 30°C (Hedhly, Hormaza, and Herrero, 2008; Jagadish et al., 2010; De Storme and Geelen, 2014). For example, mean daily temperature during the growing season in Sub-Saharan Africa are frequently above 30°C, and maize production in this region can experience >10% yield decline for every additional 1°C increase in average temperature above 25°C (Lobell et al., 2011). Heat inhibition can increase to over 20% yield loss when compounded by the effects of drought (Lobell et al., 2011). Similarly, maize and soybean in the United States could experience a 17% yield decline for each degree increase in temperature (Lobell and Asner, 2003). At the International Rice Research Institute in the Philippines, Peng et al. (2004) compiled annual temperature and rice yield data over an eleven- year period. Temperatures were consistently near 30°C during the day, but night temperatures increased more rapidly over the time period, such that for each 1°C increase in night temperature rice yield declined 10% (Peng et al., 2004). This pattern is observed in many plant genera, raising concerns about yield in a wide range of crops as the Earth warms during a period when population expansion will substantially increase food demands. Warming temperatures can affect vegetative growth and development in many ways. Early in development, high temperatures disrupt cell elongation and differentiation (Bita and Gerats, 2013), reduce membrane stability and function (Maestri et al., 2002; Bita and Gerats, 2013), and lower the ability of seeds to germinate (Balyan and Bhan, 1986; Wahid et al., 2007). Plants that have germinated can experience lower shoot and root growth because of increased respiration and decreased resource assimilation (Wahid et al., 2007; Bita and Gerats, 2013). Throughout development, growth can be greatly impacted by the effects of heat stress