Physiological and biochemical constraints on photosynthesis of leguminous plants induced by elevated ozone in open–top chambers

Thesis (MSc (Environmental Sciences))--North-West University, Potchefstroom Campus, 2012.

Air pollution is one of the most critical and urgent problems globally and is also a growing concern in southern-Africa. Rapidly growing cities, increased traffic on roads, use of non-renewable fuels, reliance on outdated industrial processes and a lack of implementation of environmental regulations, are all major factors that contribute to the poor air quality in most developing countries such as South Africa (Agrawal, 2005). As a lot of air pollution is due to vehicles, no evident solution appears to be in sight. As a result of anthropogenic emissions of nitrogen oxides (NOx) and volatile organic compounds (VOC), tropospheric ozone (O3) has increased drastically during the last centuries. Although there are many oxidising pollutants in the atmosphere, O3 is currently regarded as one of the most important air pollutants, since it causes more damage to vegetation world-wide than all the other air pollutants combined (Ashmore & Bell, 1991). In the Unites States of America, losses in the region of US$ 3 billion result each year from the impacts of O3 pollution on crops (Holmes, 1994). Holland et al. (2002) estimated that the agricultural damage in Europe as early as 1990 due O3 damage was in the order of ₤ 4.3 billion. The phytotoxicity of O 3 is due to its high oxidative capacity through the induction of reactive oxygen species (ROS) in exposed plant tissue, such as superoxide (O2 –), hydrogen peroxide (H2O2), hydroxyl radical (•OH) and singlet oxygen (1O2) (Malhorta and Khan, 1984). Specifically in southern Africa, there is a growing concern that the concentrations of O3 commonly found in the southern African troposphere may adversely affect natural vegetation, forests and crops (van Tienhoven and Scholes, 2003). While much research has been done in Asia, North America and Europe, little attention has been directed on Africa. Since agriculture plays a critical role in food security and economic growth in developing countries, it is of the utmost importance to understand and study the effect of air pollution on plants. The aim of this study was to identify and quantify the physiological and biochemical constraints imposed by O3 on two leguminous crops by analysing various parameters deduced from photosynthetic gas exchange and chlorophyll a fluorescence induction measured in parallel. In our first experiment, Phaseolus vulgaris genotypes (S156 and R123) with known differences in sensitivity to O3, were exposed to an elevated level of this pollutant at 80 nmol mol-1 in open-top chambers. The specific aim of this experiment was to identify the physiological and biochemical mechanism involved in the difference in resistant properties to O3 of the two genotypes. In the second experiment Pisum sativum plants were subjected to a concentration of 80 nmol mol-1 O3 and drought stress, singly or combined. The specific aims of this experiment were to evaluate whether a moderate drought stress in combination with O3 would have any additional effects on the physiological and biochemical mechanisms of the test plants. With respect to the first experiment: The sensitive genotype (S156) of Phaseolus vulgaris developed visual symptoms after 12 days of fumigation, ultimately developing into bronze-coloured lesions, which gradually joined together after 35 days of O3 exposure. A highly significant reduction of 58 % in the final pod weight occurred in the S156 genotype exposed to 80 nmol mol-1 O3. The latter decrease was mainly due to the pronounced decreases in CO2 assimilation as a result of a 61 % and 75 % decrease in the CO2 saturated rate of photosynthesis (Jmax) and carboxylation efficiency (CE), respectively. From the parameters obtained from the fluorescence data it could be concluded that the major effects responsible for the decrease in photosynthesis occurred in the reduction of end electron acceptors [δRo / (1-δRo)] and the efficiency of the conversion of trapped excitation energy to electron transport [ψ0 / (1-ψ0)]. The effect was also reflected by a decrease in the phenomological electron transport flux (ET/CS0). This was also the main reason for the reduced Jmax and CE in the S156 genotype. With respect to the second experiment: It was illustrated that elevated O3 levels of 80 nmol mol-1 reduced photosynthetic capacity of Pisum sativum without any accompanying visual injury throughout the experiment. CO2 gas exchange analysis indicated that inhibition of the mesophyll reactions as well as stomatal limitation were responsible for inhibition of photosynthesis in Pisum sativum. Analysis of the data revealed severe inhibition of the carboxylation efficiency (CE; Rubisco activity) and maximum rate of CO2 assimilation (Jmax; regeneration capacity of RuBP), ultimately leading to a marked reduction in CO2 assimilation (A370). The in vitro analysis revealed a highly significant O3 induced decrease in Rubisco activity in Pisum sativum test plants of up to 39 % corroborated the gas exchange data. As stomata regulate O3 uptake, our hypothesis was that the drought stress decreased O3 flux into the leaf due to stomatal closure. The stomatal conductance of the drought stressed treatments (DSCF and DSO3) was on average 56 % of that of the control plants (WWCF). This large decrease in stomatal conductance also illustrated by the scanning electron micrographs, showing closure of the stomatal aperature in the drought treatments. Analysis of the chlorophyll a fluorescence transients revealed inhibition of electron transport on the acceptor side of PSII, resulting from the inability of the inactive donor side to donate electrons. That means that the donor side, especially the oxygen evolving complex (OEC), was damaged. The chlorophyll a fluorescence data further supported the gas exchange data by confirming that the inhibition of CO2 assimilation was mainly due to impairment of the formation of end electron acceptors such as ATP and NADPH. The chlorophyll content decreased significantly in Pisum sativum plants exposed to O3. This was also reflected by the moderate decrease of 5 % and 4 % in the density of reaction centers per cross-section (RC/CS) calculated from the fluorescence transients, in the well-watered and drought stressed treatments, respectively. It could be assumed that the decreased chlorophyll content contributed to the decreases in biomass and yield production. It was also shown that O3 induced increases in the activity of the antioxidant enzyme, peroxidase (POD) after 20 days of fumigation in the O3- treated test plants, which, after 30 days of fumigation, increased by a highly significant 40 % and 41 % in the WWO3 and DSO3 plants, respectively. The additional drought stress induced on the DSO3 test plants showed no additional inhibitory effect on the test plants, indicating an ameliorating effect caused by the partial closing of the stomata. The latter finding proved the hypothesis set on the interaction between drought and O3 on P.sativum to be true. In conclusion: Using the resistant, R123 and sensitive, S156 bean genotypes as tool, valuable insight was gained into the inhibitory effect of O3 on plants. Although the R123 genotype of Phaseolus vulgaris exhibited no stress symptoms with respect to fluorescence and gas exchange data, the seed yield was affected. Photosynthesis was largely inhibited in the S156 genotype, mainly due to inhibition of the photosynthetic electron transport, resulting in decreased reduction of end electron acceptors, ultimately causing a decrease in CO2 assimilation. The above limitations ultimately lead to a large reduction in seed yield in S156. Our data show that the O3 sensitivity of S156 is mainly due to a weakness of the photosynthetic apparatus and electron transport chain. Especially PSII function, including the OEC, proved to be very vulnerable. Exposing P. sativum to O3 and drought stress simultaneously or singly lead to a drastic inhibitory effect on photosynthesis. Although it was shown that the decrease in stomatal conductance lead to amelioration of the O3-effect, the interaction was difficult to interpret as drought stress on its own has a constraints on photosynthesis.

Masters