Exploring the impact of titanium dioxide nanoparticles (nTiO₂) at varied concentrations in combination with Azospirillum brasilense on wheat growth and physiology

1 Introduction

Wheat (Triticum aestivum) is the main food crop for the one-third population of the world (FAO, 2023). The fertile land of Iraq, Syria, Jordan, Palestine, and Lebanon is the origin of the wheat crop. The wheat crop can be cultivated in a wide range of environmental conditions from arid, semi-arid, and dry regions of different countries, event it can also be a well-grown crop in the cool and wet regions of North America and Europe (Mujeeb-Kazi et al., 1996). Wheat crop is playing significant role to control the global food security and also provide the sustainability in agriculture production. It is reported by the FAO that only wheat crop is providing the 60 % of daily calories and protein needs for the people of word (FAO, 2017). But with the increasing global population, is a need for time to enhance wheat production events under the current challenges of climatic change, poor soil fertility, drought stress, and event under disease outbreaks (Hussai et al., 2021; Luo et al., 2022).

Nanoparticles are particles with a size of 1–100 nm having unique characteristics. Uses of nanoparticles in agriculture have been explored to enhance crop yield and overcome the different problems of farming. Foliar application of Nano fertilizers can improve plant physiology, can reduce the use of agrochemicals, and can save the plant from different kinds of adverse environmental conditions (Yu et al., 2024). The application of nanoparticles in agriculture is a promising development for contemporary farming methods, with numerous potential advantages (Muhammad et al., 2019). By utilizing their special physicochemical qualities, nanoparticles have the potential to greatly boost agriculture outputs. nTiO2 can act as nanocarriers for compounds that are necessary for growth and development of the plants (Khodakovskaya et al., 2012). Increased agricultural productivity is the outcome of precision delivery system, which makes sure that plants get the resources they require at the correct moment and location. Through their ability to catalyze biological processes, nanoparticles can have a significant impact on photosynthesis, respiration, and the metabolism of nutrients in plants (Ibrahim et al., 2020).

Nanoparticles such as zinc oxide, silver, and copper can improve the plant growth and physiological properties of wheat crops, but the use of the proper dose of titanium dioxide nanoparticles (nTiO₂) does not study enough. Different researchers reported the positive effect of nTiO₂ on the wheat crop (Khodakovskaya et al., 2012) and many are against it (Ibrahim et al., 2020; Muhammad et al., 2019). Some mentioned the suitable dose and some reported using its little quantity to improve particular growth traits (Ibrahim et al., 2020; Muhammad et al., 2019; Khodakovskaya et al., 2012). A titanium dioxide nanoparticle (nTiO₂) is becoming the most effective nanoparticle to improve and boost the different chemical reactions in plants. nTiO₂ is highly reactive and has the most effective catalytic properties. nTiO₂ can improve photosynthesis, chlorophyll content, nutrient uptake, and efficiency in plants. nTiO₂ has a positive effect on different traits, but it also harms human health and the environment (Khodakovskaya et al., 2012).

Different organic soil ammendments have the postive impact on plant growth and development (Jiang et al., 2023; Zhang et al., 2020). Some microbes have the positive effect on crop growth and some have the negative effect (Wang et al., 2023). Azospirillum brasilense is a nitrogen-fixing, gram-negative bacterium that were isolated from the maize plant roots in 1979 brazil (Baldani et al., 1983) since it is reported to be the growth-promoting bacteria in different crops such as wheat, rice, maize, etc. A. brasilense can produce different kinds of the growth hormones such as cytokinin and auxins that can enhance the cell division and different physiological processes of the wheat plant that result to enhance crop yield. A. brasilense can also improve the soil structure and fertility in the soil and can also break the chelate molecules from the soil to make available for plant uptake. Several studies reported that A. brasilense can enhance wheat crop growth under adverse environmental conditions and can also control the negative effect of different chemicals in the soil. A. brasilense has the great potential to improve soil fertility and plant growth, so it can use as a bio-fertilizer for sustainable crop production (Baldani et al., 1983).

Azospirillum brasilense can also control the negative effect of different nanoparticles in the plant by enhancing cell division with the production of cytokinins in the plant. Some nanoparticles harm wheat crops when used in higher amounts, so the use of A. brasilense in crop production can alter the plant morphology and can alleviate the negative effect (Singh et al., 2018). A different study was reported to control the toxicity of silver nanoparticles with A. brasilense. A. brasilense with the wheat seed inoculation is recommended by many researchers to enhance growth and yield under adverse environmental conditions (Baldani et al., 1983) but little study available with the nTiO₂. So, the present study was planned to understand the role of different doses of titanium dioxide nanoparticles (nTiO₂) with the seed inoculation of Azospirillum brasilense, on the growth and physiology of wheat crops.

We hypothesized that the Azospirillum brasilense can control the negative effect of nTiO₂ on wheat growth and yield. Our current study will also provide a suitable dose of nTiO₂ that can enhance yield and can also provide directions to understand the combined effect of Azospirillum brasilense and nTiO₂ on the growth and physiology of wheat crops.

2 Materials and methods

3 Results

Data regarding the growth and yield of wheat (Table 3) shows that the titanium dioxide nanoparticles (nTiO₂) and Azospirillum brasilense significantly effected on wheat plant. Highest plant height (99.35 cm), spike length (13.27 cm) and number of spikelets per spike (26.23) were noticed in T₄ when nTiO₂ applied @30 mg/L with the seed inoculation of wheat with A. brasilense followed (Plant height: 98.32 cm, spike length: 12.97 cm, number of spikelets per spike: 25.12) by T₃ when applied nTiO₂ @30 mg/L only and lowest results were seen in T₉ (Plant height: 89.23 cm, spike length: 07.12 cm, number of spikelets per spike: 16.42) when nTiO₂ applied @60 mg/L without A. brasilense. Number of grains per spike, 1000-grain weight and grain yield were also significantly affected by the nTiO₂ and Azospirillum brasilense. nTiO₂ application improve the plant growth but when applied on lower concentration till 30 mg/L after that it adversely effected on growth and yield related parameters. Highest grain per spike (49), 1000-grain weight (38.23 g) and grain yield (5191 kg/ha) were noticed in T₄ followed by T₃ (Grains per spike: 47, 1000-grain weight: 36.34 g, grain yield: 5045 kg/ha) and lowest were noticed in T₉ (Grains per spike: 29, 1000-grain weight: 18.35 g, grain yield: 4162 kg/ha).

Photosynthetic rate, leaf area index (LAI), and stomatal conductance were positively affected with the application of nTiO₂ till 30 mg/L and with the higher concentration of nTiO₂ all mentioned parameters negatively affected but A. brasilense seed inoculation overcome the negative effect and improved the Photosynthetic rate, leaf area index and stomatal conductance (Table 4). Highest photosynthetic rate (9.32 μmol m⁻² s⁻¹), LAI (3.91), and stomatal conductance (262 mmolm⁻² s⁻¹) was noticed in T₄ followed by T₃ (photosynthetic rate: 9.19 μmol m⁻² s⁻¹, LAI: 3.78, stomatal conductance: 256 mmolm⁻² s⁻¹) and lowest were observed in T₉ (photosynthetic rate: 7.33 μmol m⁻² s⁻¹, LAI: 2.03, stomatal conductance: 168 mmolm⁻² s⁻¹). Electrolyte leakage linked with the physical damage of cell membrane. Highest electrolyte leakage was noticed in T₉ (68 %) Followed by T₁₀ (65 %) and lowest were noticed in T₄ (21 %).

Chlorophyll content, crop growth rate (CGR), relative growth rate (RGR) and proline contents of wheat significantly affected by all treatments (Table 5). nTiO₂ application with lower concentration is helpful to improve mentioned parameters and with A. brasilense seed inoculation, it is more beneficial. Highest chlorophyll content (84.92 %), CGR (10.33 gm^-2day⁻¹), RGR (89 %) and proline contents (48.32 μg/g) were noticed in T₄ followed by T₃ (chlorophyll content: 83.33 %, CGR: 10.01 gm^-2day⁻¹, RGR: 87 %, proline contents: 47.34 μg/g) and lowest results were observed in T₉ (chlorophyll content: 74.34 %, CGR: 07.34 gm^-2day⁻¹, RGR: 65 %, proline contents: 38.53 μg/g). Data regarding N-P-K uptake by wheat plant shows that nutrients uptake also significantly affected with all studied treatments. Highest N-P-K uptake (0.064–1.92–6.19 mg/g) was noticed in T₄ followed by T₃ (0.050–1.83–6.00 mg/g) and lowest (0.018–0.93–5.72 mg/g) was noticed in T₉. nTiO₂ application above of 30 mg/L negatively affected on nutrients uptake but A. brasilense seed inoculation also helpful to overcome this damage (Table 6).

Antioxidant enzymes protect the plants from oxidative stress and detoxify reactive oxygen species (ROS). Ascorbate peroxidase (APX), catalase (CAT), and guaiacol peroxidase (POD), Superoxide dismutases (SOD) activities increase with the application of A. brasilense in all treatments but with the application of nTiO₂ their activities increased till the application of 30 mg/L but after that their activities decreased (Figs. 1–4). Highest APX (3.29 Umg⁻¹ Protein), CAT (7.32 Umg⁻¹ Protein), POD (7.43 Umg⁻¹ Protein) and SOD activities (289 Umg⁻¹ Protein) were noticed in T₄ when nTiO₂ applied @30 mg/L with A. brasilense, followed by T₃ (APX: 3.16 Umg-1Protein, CAT: 7.03 Umg⁻¹ Protein, POD: 7.05 Umg-1Protein, SOD: 281 Umg⁻¹ Protein) when only nTiO₂ applied @30 mg/L without A. brasilense application. Lowest enzymatic activities (APX: 2.26 Umg⁻¹ Protein, CAT: 3.76 Umg⁻¹ Protein, POD: 3.81 Umg⁻¹ Protein, SOD: 227 Umg⁻¹ Protein) were seen in T₉ when nTiO₂ applied @60 mg/L without A. brasilense.

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Fig. 1

Effect of nTiO₂ with A. brasilense on APX (ascorbate peroxidase) of wheat plant. B (A. brasilense bacterial strain), nTiO₂ (titanium dioxide nanoparticles).

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Fig. 2

Effect of nTiO₂ with A. brasilense on CAT (catalase) of wheat plant. B (A. brasilense bacterial strain), nTiO₂ (titanium dioxide nanoparticles).

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Fig. 3

Effect of nTiO₂ with A. brasilense on POD (guaiacol peroxidase) of wheat plant. B (A. brasilense bacterial strain), nTiO₂ (titanium dioxide nanoparticles).

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Fig. 4

Effect of nTiO₂ with A. brasilense on SOD (Superoxide dismutases) of wheat plant. B (A. brasilense bacterial strain), nTiO₂ (titanium dioxide nanoparticles).

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4 Discussion

Titanium dioxide nanoparticles (nTiO₂) foliar application significantly affected the growth, yield, and physiology of wheat crops, but up to a certain concentration. Our results follow the results of many researchers such as Zhang et al. (2017), Saber and Abdeen (2021) reported the positive effect of nTiO₂ on plant growth but with the lower concentration. Different researchers reported different concentrations, but our study conducted in the hot region of the desert shows that the nTiO₂ application up to 30 mg/L positively affected wheat growth and yield, but after that with the increase of the concentration it negatively affected. A 10.14 % increase in wheat grain yield was noticed with the application of nTiO₂ @30 mg/L but when we increase the nTiO₂ concentration @40 mg/L grain yield decreased by 3.93 % and this percentage increased with increasing the nTiO₂. Zhang et al. (2017) also reported that the higher concentration of nTiO₂ negatively affected plant growth. Several researchers (Wang et al., 2017) also reported that the nTiO₂ application on plants in high-temperature areas negatively affects growth and yield, so there might be the cause of our results that the nTiO₂ application up to 30 mg/L is beneficial and after that, it adversely affects. nTiO₂ application with lower concentration can improve the chlorophyll contents in the leaf, so it is very beneficial to improve the growth and yield-related parameters such as plant height, spikelets per spike, and grain weight. Nour et al. (2019) also reported that the foliar application of nTiO₂ increases the root-shoot ratio in the plant, which can enhance the plant growth rate and yield. Seed inoculation of agricultural crops with soil bacteria showed the positive effect on growth and yield (Qin et al., 2022). Zaheer et al. (2019) reported that the A. brasilense inoculation enhances the nutrient uptake from the soil and enhances the growth hormones produced in the plant, helpful to increase wheat growth and yield. Combining the application of A. brasilense with nTiO₂ @30 mg/L is more beneficial as compared to the other treatments. It was also observed that the A. brasilense controls the negative effect of the higher concentration of nTiO₂.

A. brasilense inoculation with the wheat seed also enhances the leaf area index and chlorophyll contents in the leaf by the production of more phytohormones such as cytokinin, gibberellins, and Indole-3-acetic acid (Lu et al., 2022). Combining the application of nTiO₂ @30 mg/L with seed inoculation is more beneficial to increase the chlorophyll contents and leaf area index. This increase in the LAI and chlorophyll content also enhances the photosynthetic rate and stomatal conductance. Increasing the photosynthetic rate by up to 23 % was noticed with A. brasilense seed inoculation. Wang et al. (2017) reported that the nanoparticle foliar application enhances and facilitates the plant for more nutrient's uptake from the soil and on the other hand availability of the A. brasilense in the root zone enhances nutrients available for plant uptake (Zaheer et al., 2021) so their combine application enhance the growth hormones and physiological processes in the wheat crop that enhance the LAI and Chlorophyll contents. More LAI and chlorophyll contents lead to a higher photosynthetic rate and stomatal conductance. A. brasilense and nTiO₂ application enhance the cytokinin production in the plant that enhances the cell division, leaves expansion, and chlorophyll accumulation, all these things directly enhance the photosynthetic rate, LAI, stomatal conductance, and chlorophyll contents (Zaheer et al., 2019; Nour et al., 2019; Saber and Abdeen 2021).

Electrolyte leakage (EL) is used to determine the cell membrane damage due to any stress condition. Higher EL decreases crop yield and growth-related parameters (Zhang et al., 2019). It was observed that nTiO₂ application higher than @30 mg/L leads towards higher EL in wheat plant leaf, which means higher EL level converting the higher nTiO₂ application towards the stress condition. 68.29 % increase in EL was noticed with the application of nTiO₂ @60 mg/L but the application of A. brasilense with the same nTiO₂ dose decrease this level up to 57.32 %. A. brasilense application with nTiO₂ @30 mg/L showed the best results. Zaheer et al. (2022) reported that A. brasilense decreases the EL by enhancing the antioxidant enzyme production in wheat plants with higher growth hormone production. Pii et al. (2019) also reported that the A. brasilense decrees EL in the wheat plant by increasing cell growth with the production of cytokinin which is essential for cell division. A. brasilense control the negative effect of the higher concentration of nTiO₂ with the production of cytokinin, Zaheer et al. (2022) and Zaheer et al. (2019) reported that the A. brasilense stimulate the production of enzymes and allow more cell division and cell expansion.

Many researchers reported the positive effect of nTiO₂ and A. brasilense on the nutrient's uptake by the plant (Wang et al., 2017; Singh et al., 2015) but no one explains the proper concentration of nTiO₂. Our results show the positive effect of nTiO₂ on the macronutrient (N-P-K) uptake by the wheat plant but till the 30 mg/L concentration, with the higher concentration adverse effect was noticed. A. brasilense application increases the nutrient's uptake by the wheat by improving the soil fertility status and availability of nutrients and improves the root development that is also very beneficial for the NPK Uptake with the nTiO₂ (Lin et al., 2021; Singh et al., 2015). A. brasilense seed inoculation with 30 mg/L nTiO₂ is very effective for nutrient uptake and improving wheat crop growth.

Leaf relative water contents and Proline content is significantly affected by the application of nTiO₂ and A. brasilense. Combining the application of nTiO₂ @30 mg/L with A. brasilense is more effective as compared to the other treatments. Different researchers show the different effects of nTiO₂ on RWC, Wang et al. (2017) reported the negative effect of nTiO₂ on leaf RWC due to the damage to the cell membrane and disturb the plant water mechanism but on the other hand, nTiO₂ have the positive effect on RWC but when to use in lower concentration, with increasing the higher concentration RWC becomes decreased. Lower concentrations of nTiO₂ improve the water use efficiency in the wheat plant that helpful in main the leaf turgor pressure and RWC (Ma et al., 2019). A. brasilense enhance the water and nutrients uptake from the soil which is helpful to enhance root growth, which enhances the water availability for wheat growth and maintains higher leaf RWC under stress condition (Zaheer et al., 2019). A. brasilense improves the RWC to improve the stomatal conductance and reduce the transpiration loss. Pereyra et al. (2012) reported the production of osmo protectants with the seed inoculation of A. brasilense that can enhance leaf turgor pressure and RWC.

nTiO₂ enhance the proline content in wheat crop due to an increase in the root shoot ratio, physiological process, and plant growth. Wang et al. (2017) reported higher proline accumulation with the application of nTiO₂ which saves the plant from oxidative damage. Our results also supported the same thing but till the application of nTiO₂ @30 mg/L on higher concentrations it effects negatively on proline content. Seed inoculation with A. brasilense increases the proline contents and antioxidant enzyme activity (APX, CAT, POD, and SOD) in wheat crops (Zaheer et al., 2021; Zaheer et al., 2022). Ghosh et al. (2016) observed higher SOD and CAT activities with nTiO₂ application, but POD activity was unaffected. Wang et al. (2017) also reported higher SOD, CAT, POD, and APX activities with nTiO₂ application in maize. Cytokinin production with the A. brasilense maintaining physiological functions under stress conditions and reduce the production of reactive oxygen species (ROS), Lower the ROS and availability of nutrients and favorable growth conditions enhance the antioxidant enzyme activity (Zaheer et al., 2021). Synthesis of the P5CS gene can increase the proline formation with increasing the cytokinin production in the plant cell. More microbial activities and organic matter in the soil leads towards positive impact on the plant growth (Zhang et al., 2021). Zaheer et al., (2021,2022) reported the higher cytokinin formation with the seed inoculation of A. brasilense that can leads towards the higher proline production.

5 Conclusions

Titanium dioxide nanoparticles (nTiO₂) are effective for wheat growth, yield, and physiological parameters, but their concentration more than of 30 mg/L adversely affected wheat growth and yield-related parameters. Combining application of nTiO₂ @30 mg/L with A. brasilense is very effective for the wheat crop. It is recommended to not use the higher concentration of nTiO₂ for crop production and to use A. brasilense for sustainable agriculture.

Funding

Researchers Support Project Number (RSPD2024R708), King Saud University, Riyadh, Saudi Arabia.