Heavy metal contamination, particularly cadmium (Cd), poses a significant threat to agricultural productivity and environmental health. Cd, a non-essential and undegradable element for plants, accumulates in soil due to excessive fertilizer and pesticide use and industrial wastewater irrigation. Cd accumulation in plants leads to various negative physiological effects, including chlorophyll reduction, decreased photosynthesis, metabolic disorders, and oxidative stress. Oxidative stress arises from the production of reactive oxygen species (ROS), causing cellular damage. Previous research has shown that various amendments, including biochar and microorganisms, can mitigate Cd toxicity. Biochar, an environmentally friendly soil amendment, possesses several advantageous properties, such as high carbon content, large surface area, and abundant functional groups, all contributing to its potential to reduce Cd uptake and enhance plant growth. Biofertilizers, containing beneficial microorganisms like *Bacillus* sp., can also chelate heavy metals and improve plant health. Cotton, a major cash crop in China, is particularly vulnerable to Cd contamination. This study aimed to evaluate the combined effects of biochar and biofertilizer on Cd-contaminated cotton growth, focusing on Cd accumulation, oxidative stress response, and photosynthetic performance.

Extensive literature documents the detrimental effects of Cd on plant growth and physiology. Studies have shown Cd's negative impact on chlorophyll content and photosynthetic rates, ultimately reducing crop yields. The toxicity of Cd is linked to its stimulation of ROS production, leading to oxidative stress and damage to cellular components. Plants respond to this stress by producing antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) to scavenge ROS. Numerous studies have investigated the use of various soil amendments to remediate Cd contamination. Biochar, with its high carbon content and large surface area, has shown promise in reducing Cd bioavailability and improving plant growth. Similarly, biofertilizers, containing beneficial microorganisms, have demonstrated the ability to reduce Cd uptake and enhance plant health. However, research on the combined effects of biochar and biofertilizer on Cd-contaminated cotton remains limited. The current study addresses this gap.

Soil samples were collected from a local cotton field and their physicochemical properties were determined. Biochar, derived from cotton straw, and biofertilizer (*Bacillus* composite) were prepared and analyzed. Cd-contaminated soil was created by adding CdCl2·2.5H2O to the soil samples to achieve concentrations of 0, 1, 2, and 4 mg Cd kg⁻¹ soil. A randomized complete block design was used with twelve treatments (five replicates per treatment): control (no Cd, biochar, or biofertilizer), biochar treatment, biofertilizer treatment, and combinations of Cd and amendments. Cotton seeds (*Gossypium hirsutum* L. ‘Xinluzao 53’) were sown in pots containing the treated soil. After 3 true leaves appeared, 5 seedlings per pot were selected for further cultivation. At the boll stage, cotton plants were harvested, and dry weights of roots, stems, leaves, and bolls were measured. Cd concentrations were determined using atomic absorption spectrophotometry. The transfer coefficients of Cd in different plant organs were calculated. Chlorophyll content was determined spectrophotometrically, and photosynthetic parameters (net photosynthetic rate, stomatal conductance, transpiration rate, and intercellular CO2 concentration) were measured using a Li-6400 portable photosynthesis system. The activities of SOD, CAT, and POD were measured spectrophotometrically, along with MDA content and electrolyte leakage. Redundancy analysis (RDA) was employed to investigate the relationships between Cd absorption, transport, cotton growth indices, and physiological indicators. Statistical analyses (SPSS 23.0) were performed using Duncan’s new multiple range test (α = 0.05).

The study found that Cd significantly reduced cotton dry weight, chlorophyll content, and photosynthetic parameters. The highest Cd concentration (4 mg kg⁻¹) resulted in the lowest dry weight (60.63 g). Biochar and biofertilizer significantly increased dry weight across all Cd treatments. The maximum dry weight (91.17 g) was achieved with the biofertilizer treatment at 0 mg Cd kg⁻¹. Cd accumulated primarily in the roots, followed by leaves and stems, with the lowest accumulation in bolls. Biochar and biofertilizer significantly reduced Cd accumulation in all plant organs, particularly in bolls (P < 0.05). The amendments also decreased the transfer coefficient of Cd from roots to stems. Biochar and biofertilizer significantly increased chlorophyll a and b contents, improving photosynthetic parameters. The application of biochar and biofertilizer enhanced the activities of SOD and CAT enzymes and reduced MDA content and electrolyte leakage rate. RDA analysis showed that Cd content in cotton organs was negatively correlated with growth indices and antioxidant enzyme activities, while being positively correlated with MDA and electrolyte leakage. Biochar and biofertilizer treatment significantly decreased Cd accumulation in stems by an average of 27.50% (biochar) and 25.14% (biofertilizer) across different Cd levels.

The findings demonstrate that both biochar and biofertilizer effectively mitigate the negative effects of Cd on cotton growth and physiology. The increased dry weight and improved photosynthetic parameters in the treatment groups strongly support their role in enhancing cotton productivity under Cd stress. The reduction of Cd accumulation in cotton organs, particularly in the bolls, is crucial for ensuring food safety. The enhanced activities of antioxidant enzymes indicate the protective role of these amendments in reducing Cd-induced oxidative stress. The RDA analysis further supports the observed relationships between Cd content, growth parameters, and antioxidant enzyme activities. The positive effects of the amendments are likely due to a combination of mechanisms including reduced Cd bioavailability, increased nutrient availability, and enhanced antioxidant defense systems. These results suggest that biochar and biofertilizer could be valuable tools for sustainable cotton production in Cd-contaminated soils.

Biochar and biofertilizer proved effective in reducing Cd accumulation, improving photosynthetic efficiency, and enhancing antioxidant defense mechanisms in cotton plants subjected to Cd stress. This study highlights the potential of these amendments as sustainable remediation strategies for Cd-contaminated agricultural lands. Further research should explore the optimal application rates of biochar and biofertilizer for different Cd concentrations and soil types. Investigating the long-term effects of these amendments on soil health and cotton productivity is also warranted.

The study was conducted under controlled pot conditions, which may not entirely reflect the complexity of field environments. The specific bacterial composition of the biofertilizer was not detailed, which could limit the generalizability of the findings. The study focused on a single cotton variety, and the results may vary across different cultivars. The duration of the experiment was limited to a single growing season. Long-term studies are needed to assess the sustainability of biochar and biofertilizer applications.