Effects of Glyphosate-Based Herbicide on Primary Production and Physiological Fitness of the Macroalgae Ulva lactuca

Abstract

The use of glyphosate-based herbicides (GBHs) worldwide has increased exponentially over the last two decades increasing the environmental risk to marine and coastal habitats. The present study investigated the effects of GBHs at environmentally relevant concentrations (0, 10, 50, 100, 250, and 500 μg·L^-1) on the physiology and biochemistry (photosynthesis, pigment, and lipid composition, antioxidative systems and energy balance) of Ulva lactuca, a cosmopolitan marine macroalgae species. Although GBHs cause deleterious effects such as the inhibition of photosynthetic activity, particularly at 250 μg·L^-1, due to the impairment of the electron transport in the chloroplasts, these changes are almost completely reverted at the highest concentration (500 μg·L^-1). This could be related to the induction of tolerance mechanisms at a certain threshold or tipping point. While no changes occurred in the energy balance, an increase in the pigment antheraxanthin is observed jointly with an increase in ascorbate peroxidase activity. These mechanisms might have contributed to protecting thylakoids against excess radiation and the increase in reactive oxygen species, associated with stress conditions, as no increase in lipid peroxidation products was observed. Furthermore, changes in the fatty acids profile, usually attributed to the induction of plant stress response mechanisms, demonstrated the high resilience of this macroalgae. Notably, the application of bio-optical tools in ecotoxicology, such as pulse amplitude modulated (PAM) fluorometry and laser-induced fluorescence (LIF), allowed separation of the control samples and those treated by GBHs in different concentrations with a high degree of accuracy, with PAM more accurate in identifying the different treatments.

Keywords: energetic metabolism; glyphosate; oxidative stress; pesticide; photobiology.

Figure 1

Chlorophyll transient kinetics (OJIP curves or Kautsky plots) in Ulva lactuca following a 48-h exposure to a glyphosate-based herbicide formulation in different concentrations (mean, n = 18).

Figure 2

The energy fluxes (absorbed (ABS/CS, trapped (TR/CS), transported (ET/CS) and dissipated (DI/CS)) and the number of available reaction centers per cross-section (RC/CS), in Ulva lactuca following a 48-h exposure to a glyphosate-based herbicide formulation in different concentrations (mean ± s.d., n = 18, different letters indicate significant differences between treatments at p < 0.05).

Figure 3

Photosystem II and ETC-related photochemical traits: (A) oxidized quinone pool; (B) reaction center turnover rate (N); (C) the energy needed to close all reaction centers (S_M); (D) the net rate of PS II RC closure (M₀); (E) smallest possible normalized total area when each QA is reduced only once (S_S); (F) the probability that a PSII chlorophyll molecule function as an RC (γ_RC), in Ulva lactuca following a 48-h exposure to a glyphosate-based herbicide formulation in different concentrations (mean ± s.d., n = 18, different letters indicate significant differences between treatments at p < 0.05).

Figure 4

Photosystem II and PS I photochemical traits: (A) active oxygen-evolving complexes (OECs); (B) grouping probability between the two PSII units (P_G); (C) contribution or partial performance due to the light reactions for primary photochemistry (TR₀/DI₀); (D) the contribution of the dark reactions from quinone A to plastoquinone (ψ₀/(1 − ψ₀)); (E) reaction center II density within the antenna chlorophyll bed of PS II (RC/ABS); (F) the contribution of PSI reducing its end acceptors (δ_R0/(1 − δ_R0); (G) the equilibrium constant for the redox reactions between PS II and PS I (ψ_E0/(1 − ψ_E0); (H) electron transport from PQH₂ to the reduction of PS I end electron acceptors (RE₀/RC)), in Ulva lactuca following a 48-h exposure to a glyphosate-based herbicide formulation in different concentrations (mean ± s.d., n = 18, different letters indicate significant differences between treatments at p < 0.05).

Figure 5

Red region laser-induced fluorescence in Ulva lactuca following a 48-h exposure to a glyphosate-based herbicide formulation in different concentrations (mean, n = 30, different letters indicate significant differences between treatments at p < 0.05).

Figure 6

Ascorbate peroxidase (APX, (A)), superoxide dismutase (SOD, (B)), catalase (CAT, (C)), and glutathione reductase (GR, (D)) enzymatic activities in Ulva lactuca following a 48-h exposure to a glyphosate-based herbicide formulation in different concentrations (mean ± s.d., n = 3, different letters indicate significant differences at p < 0.05).

Figure 7

Total fatty acid content: (A) fatty acid ratios (saturated to unsaturated fatty acids ratio [SFA/UFA], polyunsaturated to saturated fatty acids ratio [PUFA/SFA] and double-bound index [DBI]); (B) and fatty acid relative abundance profile (%); (C) in Ulva lactuca following a 48-h exposure to a glyphosate-based herbicide formulation in different concentrations (mean ± s.d., n = 3, different letters indicate significant differences at p < 0.05).

Figure 8

Energy balance: (A) energy available [Ea]; (B) energy consumption rate [ETS]; (C) cellular energy allocation [CEA]) in Ulva lactuca following a 48-h exposure to glyphosate-based herbicide formulation in different concentrations (mean ± s.d., n = 5, different letters indicate significant differences at p < 0.05).

Figure 9

Energy availability: (A) carbohydrates; (B) proteins; (C) lipids in Ulva lactuca following a 48-h exposure to glyphosate-based herbicide formulation in different concentrations (mean ± s.d., n = 5, different letters indicate significant differences at p < 0.05).

Figure 10

Linear discriminant analysis (LDA) using the (A) OJIP dataset and (B) the laser-induced fluorescence (LIF) dataset of Ulva lactuca following a 48-h glyphosate-based herbicide formulation in different concentrations. Ellipses group samples with lower statistical distance based on Euclidean resemblances.