The efficiency of nutrient uptake is partly modulated by root characteristics and by the amount of the specific nutrient applied . It was found that energy cane plants under Mn deficiency presented low Mn accumulation and uptake efficiency, which consequently led to biological damage to these plants.
This nutritional disorder of Mn promotes an increase in ROS, e.g., H2O2, which was observed in sorghum plants , maize , and also energy cane. Therefore, plants under Mn deficiency are susceptible to oxidative stress caused by ROS overproduction due to the poor ability of plants to induce antioxidant defense mechanisms. In the present study, an important impairment was found in the enzyme and non-enzyme antioxidant system of Mn-deficient plants.
The decrease in SOD activity occurred in plants with nutritional deficiency as Mn is a constituent of this enzyme [1, 13], which was also reported by  in sorghum plants and by  in sugarcane. Moreover, there was a decrease in glutathione peroxidase (GPOX) activity, which is important for H2O2 dismutation in water and oxygen . The decrease in the activity of another peroxidase (APX) was the main response of the plant to Mn deficiency , thus consisting of an important characteristic of response to this nutritional disorder. GPOX belongs to the class of enzymes that catalyze H2O2 reduction by removing electrons from donor molecules such as phenolic compounds , whose synthesis requires Mn  and which may play a relevant role in the reduction of enzyme activity.
Our data showed that Mn deficiency in energy cane decreased the total phenolic content. This may have occurred as Mn activates specific enzymes involved in phenol biosynthesis [9, 50]. Mn stabilizes the active conformation of the enzyme phenylalanine ammonia lyse (PAL), which catalyzes the non-oxidative deamination of phenylalanine in cinnamic acid , promoting the formation of phenols and other compounds. This loss of Mn deficiency in this non-enzymatic antioxidant compound also contributed to increase the H2O2 content, as phenols can reduce lipid peroxidation by eliminating active molecular oxygen and H2O2 in the plant metabolism .
The lower activity of GPOX associated with a lower phenol content, as found in energy cane, may have led to H2O2 accumulation. In addition, a decrease in SOD activity due to the lack of Mn can induce the accumulation of another harmful ROS, namely: O2•− [13, 48, 52]. The accumulation of H2O2 and O2•− in Mn-deficient plants was recently demonstrated in wheat . The accumulation of reactive species results in lipid peroxidation, as evidenced by the increase in MDA content in energy cane plants. This is the result of the lipid degradation of cell membranes and an evidence of oxidative damage  that has been reported as an effect of Mn deficiency in other grasses [8, 11].
Oxidative stress decreased leaf pigments of Mn-deficient energy cane plants due to membrane degradation of thylakoids and chloroplasts  and a decrease in the number of chloroplasts . In addition, Mn is required for the synthesis of lipids and proteins, which are structural components of this organelle [9, 13], as well as of carotenoids . In Mn deficient-plants, plastid and chloroplast gene transcription are reduced, which reduce the content of integral proteins in the organelle and their synthesis, affecting the integrity of the chloroplast and disorganizing the thylakoid membrane system .
The effects of Mn deficiency on pigment concentration are also a consequence of the lower quantum efficiency of PSII , a fact also found in energy cane. This results from the function of Mn as a component of the catalytic Mn4Ca cluster of the photochemical stage of photosynthesis responsible for water photooxidation . When Mn is deficient, it destabilizes and disintegrates the Mn complex in PSII . In addition, it increases the formation of singlet oxygen (1O2), which can degrade chlorophylls and the D1 protein of PSII  if not eliminated by carotenoids and tocopherols in the chloroplasts .
This damage caused by Mn deficiency, impairing the energy cane plant metabolism, explains the decrease of growth variables. It is noteworthy that Mn deficiency has a strong impact by reducing the growth of plant roots and shoots, which was observed in barley plants . Similarly, there were important reductions in the leaf area and dry mass of roots and shoots of energy cane, a fact that has also been described for other species [8, 14, 62].
Our study still evidences that energy cane with high fiber content is much more sensitive to Mn deficiency. This was evidenced as the deficiency of Mn in relation to plants under sufficient nutrition of Mn induced a decrease in plant mass production (total) of energy cane with high fiber content equal to 62%, which was much higher than that observed for energy cane with low fiber content, which reached only 22%, according to . This occurs as Mn is an important nutrient for photosynthesis  and for the synthesis pathway of vegetable fiber, especially lignin [50, 64]. Therefore, it is a nutrient that is very limiting to the production of this species when deficient, showing the importance of its adequate management. In this case, there is a need for strategies to mitigate this nutritional stress, among which the most promising is the use of Si.
We found that energy cane presents high Si absorption capacity regardless of the nutritional status in terms of Mn in the plant. Thus, the Si absorption capacity in energy cane indicates that it bears resemblance to sugarcane (S. officinarum L.), which is a Si-accumulating plant [20, 65]. This occurs as Poales present specific transporters in the cell membranes of the roots that are efficient for the absorption of Si . More recently, it was found that that the presence of transporters such as intrinsic proteins similar to nodulin 26 (NIP-III) in some species of Poales is a result of species evolution and indicates a high capacity to accumulate Si .
The high ability of energy cane to accumulate Si in the shoot increases the possibility of the biological benefits of the element in this species. The first benefit is the contribution of Si in increasing the accumulation and uptake efficiency of Mn regardless of the plant nutritional status. The increase in Mn absorption induced by Si was reported only in sugarcane under adequate Mn conditions and in energy cane with lower fiber content (type I) under Mn sufficiency and deficiency . Thus, even this species with higher fiber content, such as the one analyzed in our study, presents benefits from Si in nutrition with Mn in both conditions of Mn supply. Previously, similar results were reported in sorghum plants grown under Mn deficiency  and in corn and wheat plants grown without deficiency of this micronutrient . As the Mn uptake efficiency in this study takes into consideration the potential to accumulate Mn in the plant in relation to the amount of roots , it is clear that the benefit of Si in providing a higher Mn uptake efficiency was a reflection of the increment in the accumulation of Mn, especially in plants in the condition of deficiency (97%).
The increase in Mn absorption promoted by Si may have occurred due to the effect of the beneficial element on the modulation of H + -ATPase activity by the expression of genes that encode these proteins, which are involved in generating an electrochemical gradient for the active absorption of nutrients [27, 67, 68]. In addition, Si increases the activity of Mn-specific active transporters [2, 30, 69]. This effect of Si draws attention, as NRAMP proteins (Nramp 6 and Nramp 1) were recently identified as responsible for playing critical roles in the absorption and utilization of Mn in plants subjected to conditions of low availability of the micronutrient . Therefore, the fact that Si had a positive effect on this same family of transporters in other studies  reflects an important benefit of Si against the potential nutritional disorder of Mn deficiency.
In the face of limited availability, plants need to be efficient in acquiring Mn through the roots to increase stress tolerance . Thus, the effect of Si in enhancing this mechanism reflects an additional benefit to energy cane plants subjected Mn deficiency as a form of adaptation, also improving their performance in soils with adequate Mn fertilization.
It is also possible that the Mn that was absorbed in the first stage of the experiment for fifty days was remobilized to the plant shoot—a fact that was reported for iron in cucumber, barley, and sorghum plants [29, 70, 71]. These authors indicated that there was an increase in the expression of Si-induced genes encoding the biosynthesis of nicotianamine and in the YSL transporters responsible for the discharge of Fe into the phloematic vessels that reach the plant shoots. This can occur when Mn is precipitated in the apoplast or connected to the cell wall, being subsequently remobilized when there is a deficiency imposed to the plants . In barley plants, the use of Si increased the Fe content in young leaves of deficient plants, which was associated with the increased expression of genes involved in Fe acquisition . In addition, an increase in the translocation efficiency and accumulation of Fe in young leaves by Si was also observed in Fe-deficient sorghum plants .
We showed that the contribution of Si to energy cane plants is provided by the increase of Mn content in the shoot of plants deficient in this micronutrient, which was associated with the activation of the antioxidant enzymes of the defense system due an increase in the activity of SOD, GPOX and non-enzymatic antioxidants (phenols and carotenoids). This occurs as Mn is one of the cofactors of the SOD enzyme, composing the Mn-SOD isoform of this enzyme  and participating in the synthesis of phenols  and carotenoids by the activation of the phytoene synthetase enzyme in the synthesis of isoprenoids, which are precursors of these pigments .
The activity of SOD potentiated by Si possibly reduced the accumulation of the superoxide radical O2• − and promoted the formation of H2O2 through the catalysis of this enzyme. Furthermore, it was observed that GPOX activity potentially increased (76%) with the supply of Si, which is possibly associated with increased SOD activity to reduce the H2O2 content, maintaining homeostasis between production and promoting the elimination of ROS. In an extensive review by  on the effects of Si on antioxidant systems under various abiotic stresses, the authors concluded that the main actions of Si were associated with the increased activity of enzymes involved in the transformation of H2O2 into water (APX and CAT). In our study, the effect of Si on the elimination of this ROS was made clear by the increase in GPOX activity and phenol content (74%), both occurring with the function of eliminating H2O2  due to another abiotic stress – the nutritional deficiency.
The effect of Si on the increase of enzyme antioxidant activity was found in sorghum plants under Mn deficiency . Si did not effectively affect the activities of SOD and GPOX enzymes in energy cane with low fiber content, but it benefited the non-enzymatic response by increasing the phenol content , which was also observed in plants under nutritional stress of other species [74, 75]. This may be associated, in addition to the longer exposure time to the deficiency, to the sensitivity of the species. Thus, in the face of damage, the response of all antioxidant components evaluated was potentiated by Si, including enzymes, which suggests that Si can benefit the metabolism of plants under Mn deficiency even in longer periods and in plants under severe deficiency. A similar fact was observed in barley plants sensitive to Al-induced stress and treated with Si, resulting in higher production of antioxidant components than in more tolerant plants .
These antioxidant mechanisms potentiated by Si in Mn-deficient energy cane plants decreased the overproduction and accumulation of H2O2 contents and possibly of other ROS, reducing the MDA content. In this scenario, Si decreased membrane degradation in photosynthesizing pigments [24, 29, 71, 76], which occurred in energy cane as there was an increase in the content of Chl a, Chl b, and carotenoids in Mn-deficient plants.
In Mn-sufficient plants, the application of Si increased leaf pigments, as the micronutrient is involved in the synthesis of chlorophyll and proteins, as well as in chloroplast integrity [9, 13]. However, the use of Si in these plants did not affect lipid peroxidation and quantum efficiency of PSII, indicating that these plants did not present oxidative stress. On the other hand, in Mn-deficient plants, the benefit of Si was evidenced by increasing the quantum efficiency of PSII of energy cane plants, which was also reported in sorghum and corn plants by  and recently in soybeans plants . The fact that there was an improvement in electron transport in Mn-deficient plants through Si may have reduced ROS accumulation, since both the preservation and improvement in electron transport in PSII maintained the formation of singlet oxygen, superoxide ions, H2O2, and hydroxyl radicals .
The sum of these results resulted in a greater growth of plants under Mn deficiency, indicating an attenuation of the effects of Mn-deficiency by Si. The root, as the main element of nutrient absorption, was benefited in the presence of Si in Mn-deficient plants. This was a result of increased Mn absorption, as the nutrient influences root growth by participating in lipid metabolism, gibberellic acid production, and carbohydrate flux . Similarly, the leaf area and the total dry mass of the plant were benefited by the nutritional, biochemical, and physiological improvements, increasing the conversion to dry mass.
The first hypothesis of this study, which reported that Mn deficiency, regardless of Si, causes important damage in physiological and nutritional aspects, indicating that energy cane is sensitive to Mn deficiency, can be accepted. Thus, in a Mn-deficient plant, Si is not capable of fully reversing the nutritional and physiological damage, promoting growth that is similar to that of a Mn-sufficient plant. This is easy to understand as Si does not replace the nutritional functions of Mn, but attenuates the effects of this nutritional disorder in the plant. However, the biological effects of Si are clearly evidenced in Mn-deficient plants, as it is known that this element benefits plants under stress conditions , proving the second hypothesis of this study.
The present study proves the benefits of Si in Mn-deficient plants, as it increases the absorption of the micronutrient associated with the modulation of antioxidant defense systems, decreases the production of H2O2 and lipid peroxidation, and increases the contents of photosynthesizing pigments, avoiding oxidative damage and increases the leaf area and dry weight of energy cane plants with high fiber content.
The results of Si in attenuating Mn deficiency in Poaceae, which are Mn-demanding plants, are promising. In addition, plants without Mn deficiency can also benefit from Si, which opens perspectives for further research to check if these benefits can reach non-Poaceae plants with or without nutritional deficiency.