Cadmium (Cd) Tolerance and Phytoremediation Potential in Fiber Crops: Research Updates and Future Breeding Efforts

Abstract

Heavy metal pollution is one of the most devastating abiotic factors, significantly damaging crops and human health. One of the serious problems it causes is a rise in cadmium (Cd) toxicity. Cd is a highly toxic metal with a negative biological role, and it enters plants via the soil–plant system. Cd stress induces a series of disorders in plants’ morphological, physiological, and biochemical processes and initiates the inhibition of seed germination, ultimately resulting in reduced growth. Fiber crops such as kenaf, jute, hemp, cotton, and flax have high industrial importance and often face the issue of Cd toxicity. Various techniques have been introduced to counter the rising threats of Cd toxicity, including reducing Cd content in the soil, mitigating the effects of Cd stress, and genetic improvements in plant tolerance against this stress. For decades, plant breeders have been trying to develop Cd-tolerant fiber crops through the identification and transformation of novel genes. Still, the complex mechanism of Cd tolerance has hindered the progress of genetic breeding. These crops are ideal candidates for the phytoremediation of heavy metals in contaminated soils. Hence, increased Cd uptake, accumulation, and translocation in below-ground parts (roots) and above-ground parts (shoots, leaves, and stems) can help clean agricultural lands for safe use for food crops. Earlier studies indicated that reducing Cd uptake, detoxification, reducing the effects of Cd stress, and developing plant tolerance to these stresses through the identification of novel genes are fruitful approaches. This review aims to highlight the role of some conventional and molecular techniques in reducing the threats of Cd stress in some key fiber crops. Molecular techniques mainly involve QTL mapping and GWAS. However, more focus has been given to the use of transcriptome and TFs analysis to explore the potential genomic regions involved in Cd tolerance in these crops. This review will serve as a source of valuable genetic information on key fiber crops, allowing for further in-depth analyses of Cd tolerance to identify the critical genes for molecular breeding, like genetic engineering and CRISPR/Cas9.

1. Introduction

The rise in industrialization and urbanization has influenced the life of living organisms. Heavy metal mining leads to increased heavy metal (HM) pollution. Additionally, global warming increases the temperature and precipitation, which makes it easier for heavy metals to migrate and disturb the ecosystem, and this leads to a significant rise in cadmium pollution in soil [1,2,3]. The quantity of HMs is increased in the environment due to mining, chemical fertilizers, and pesticides [4]. The Ministry of Land and Resources of China stated that 10% of farmland in China has been contaminated with heavy metals [5]. Soil contamination caused by heavy metal toxicity is a crucial risk to our sustainable agriculture because of its deadly impact on crop yield and production. The growth and yield of crops have been severely affected by heavy metal toxicity in large areas. Heavy metals (HMs) can interfere with plant functioning, such as respiration, photosynthesis, and protein synthesis [6,7].

Cadmium is one of the most toxic metals, posing severe threats to the environment, such as plant damage, soil pollution, contamination of the food chain, and damage to aquatic life [8,9,10]. Long-term exposure to cadmium stress also affects human health and causes lung, urinary, kidney, pancreatic, and bladder cancer [11]. Cd stress and pollution have expanded rapidly in the last hundred years via human activities, such as waste emissions, mining, and fertilizer abuse (Figure 1) [12]. Cd stress seriously inhibits seed germination [13] and plant growth [14], and leads to the production of reactive oxygen species (ROS) and hydrogen peroxide (H₂O₂), which cause oxidation in cells and eventually lead to plant death [15,16]. ROS react with proteins, lipids, pigments, and nucleic acids, and therefore affect enzyme activity and induce membrane damage and the oxidation of lipids [17]. Since Cd can quickly enter the food chain and therefore can eventually enter the human body, it is necessary to prevent it from entering the food chain [18].

Plants use different techniques, including a reduction in Cd uptake, vacuolar sequestration, chelation, and detoxification, to mitigate Cd toxicity [19], and several genes are activated to tolerate heavy metals [20]. Therefore, achieving sustainable crop production in Cd-contaminated environments has become a primary research goal. The current techniques being used are traditional and modern genetic breeding tools to develop crop cultivars that are capable of curbing the uptake, accumulation, transportation, and detoxification of HMs [21].

Kenaf, jute, cotton, flax, and hemp are fast-growing crops with the ability to increase their biomass and resistance against HMs [22,23,24,25]. Hemp is well known for its role in the cleaning of contaminated soils owing to its large biomass, long root system, and ability to uptake and accumulate HMs [26,27]. Jute is tolerant to metal stress, and is the second most important source of fiber after cotton [28]. Kenaf is a highly significant fiber crop that accumulates large amounts of heavy metals. However, these heavy metals are unlikely to enter the food chain. As a result, kenaf has the potential to be used to remediate HM-polluted soils. This application is not limited to the remediation of soil pollution and can also have economic benefits [22]. Cotton is a valuable fiber crop with high tolerance to Cd stress, which makes it an excellent choice for the phytoremediation of HMs-contaminated soils [25]. Flax is a vital fiber crop that can tolerate heavy metal stress through various mechanisms. Most flax varieties can also tolerate Cd stress [29].

These are the most significant fiber crops grown in many parts of the world. They face the issue of Cd stress, which affects their growth and development. Among these crops, kenaf, hemp, and jute can potentially treat Cd-contaminated soils [22]. However, the molecular mechanisms through which these crops withstand abiotic stresses have not been fully determined [30,31]. Conventional and molecular breeding tools play a vital part in improving Cd tolerance in fiber crops. Molecular tools, like QTL mapping, transcriptome, TFs, GWAS, and genetic engineering, have been successfully employed to identify the important genes and their exploitation in the genetic improvement of fiber crops. Previous studies reported the success of these techniques regarding Cd tolerance in kenaf and hemp, but the complex genetic mechanism of Cd tolerance has still not been fully discovered [31,32,33,34,35]. A recent review paper summarized the phytoremediation potential of bast fiber crops, providing valuable insights about crop growth in contaminated soils [36]. The current review aims to summarize the recent advancements in the development of Cd tolerance in economically important crops. We hope that this comprehensive review will assist future researchers in tailoring the genetic makeup of crops to boost their tolerance to Cd stress.

2. Effects of Cd Toxicity on Fiber Crops

Kenaf is used to make rope, and textiles and is tolerant to drought and heavy metal stress [37].

2.1. Effects of Cd Stress on Kenaf

The toxic effects of Cd on growth, seed germination, seedling growth, and physiological traits in crops have been well-studied (Figure 2). Cd stress decreased the stem diameter, root length (RL), and biomass production of two kenaf cultivars when treated with 50 mg/kg Cd²⁺ [38]. One of the most visible effects of Cd toxicity on kenaf is a reduction in photosynthetic activity. Arbaoui et al. [39] observed that Cd stress reduced photosynthetic activity by reducing chlorophyll content [39]. Earlier experiments showed that Cd stress at 80 mg/kg reduced the emergence of the kenaf cultivar, Hongyou No. 2 [40]. Plant height, stem girth, and other traits in kenaf were reduced at different Cd concentrations of 0, 150, and 300 mg Cd kg⁻¹. Cd stress also decreased the activity of catalase (CAT) and ascorbate peroxidase (APX) enzymes [41]. In an earlier study, Li et al. [42] reported that Cd toxicity inhibited the growth and caused the oxidation of lipids in two cultivars (ZM412 and Fuhong 991) of kenaf. Cd stress significantly decreased the growth and activity of antioxidant enzymes, peroxidase (POD), catalase (CAT), and superoxide dismutase (SOD) in ZM412 as compared to Fuhong 991 [42]. Deng et al. [22] also showed that Cd stress affected kenaf growth by increasing the oxidation of lipids owing to enhanced MDA production [22]. The toxic effects of Cd on kenaf vary with changes in the concentration of Cd stress. The results showed an increase in the activity of the antioxidant enzymes, SOD, and POD at 10 mg L⁻¹ Cd stress, while a reduction in the activity of these enzymes was reported at 30 mg L⁻¹ Cd stress [33]. Cao et al. [43] observed that 200 µmol/L Cd toxicity increased ROS production and induced damaging effects in kenaf [43]. Cd stress has a detrimental effect on the genetic material of kenaf. A recent study showed that Cd stress of 300 µM reduced the DNA methylation level in the kenaf cultivar CP085 [30]. More studies are needed to analyze the impacts of Cd stress on water and nutrient uptake and crop yield.

2.2. Effects of Cd Stress on Hemp, Jute, Cotton, and Flax

The growth and yield of hemp crop are affected by Cd stress. Previous experiments have shown that Cd stress reduces hemp’s growth and physiological traits. Tang et al. [44] also found that chlorophyll and carotenoids in hemp decreased with Cd toxicity. As Cd stress increased, the values of these physiological traits also decreased [44]. The exposure of hemp to Cd stress decreased dry matter production, stomatal conductance, and transpiration rate [45]. Cadmium stress at 0, 25, 50, and 100 mg/kg concentrations reduced hemp plant photosynthesis, shoot, and root length [46]. Marabesi et al. [47] observed that hemp plants exposed to 25 mg·L⁻¹ Cd toxicity exhibited poor growth with reduced photosynthetic efficiency and premature senescence. However, lower Cd concentrations (2.5 and 10 mg·L⁻¹) did not influence the photosynthetic efficiency and plant height. The chlorophyll concentration was also lower at 10 mg·L⁻¹ Cd stress than at 2.5 mg·L⁻¹ [47]. Cd stress decreased lignin production and the substitution of xylan in hemp fibers [48]. Cd stress also affects the growth and yield of jute crops. Earlier studies showed that the concentration of CdCl₂ at 30 μM caused an increase in the oxidation of lipids and antioxidant activities in jute [49]. In another experiment, adding Cd²⁺ caused dose-dependent reductions in plant growth and biomass and increases in proline content in jute, with significant effects at 20 mg/L [50]. Cadmium stress affects the cotton crop’s morphology, physiology, and biochemistry.

Farooq et al. [51] conducted a study and tested the effects of three different Cd (0, 1.0, and 5.0 μM) stress levels on cotton seedlings. They documented that Cd stress decreased root growth and biomass production and resulted in the production of shorter plants. In addition to this, Cd (5 μM) also decreased antioxidant (SOD, POD, and CAT) activities, and increased MDA and H₂O₂ production [51]. Cd stress also affects the growth and properties of flax crop. Hancock et al. [52] found that Cd stress negatively affected plant fitness but did not affect plant growth [52]. A recent study showed that Cd stress (400 μM Cd) reduced the biomass of leaves, stem and roots in cotton seedlings. Cd stress also decreased the content of glycine (Gly), glutamic acid (Glu), and cysteine (Cys) [53]. Another study showed the ultrastructural aspects and pectin features of flax cellulosic fibers at the onset of secondary wall-thickening in 10-day-old seedlings in response to 0.5 mM cadmium (Cd). Based on PATAg staining, the cell wall texture of the treated fibers was disturbed, suggesting a modified adhesion between the cellulose microfibrils and matrix polysaccharides. The effect of Cd on the distribution of highly methylesterified homogalacturonans (recognized by the JIM7 antibody) was low in the primary and outer parts of the secondary wall while it was moderate in the cell junctions [54]. More studies are urgently needed to explore the effects of Cd stress on fiber crops’ biochemical and molecular properties. Existing studies are limited, and further experiments will shed light on the novel mechanisms in plants facing Cd toxicity. For instance, the toxic impacts of Cd on photosynthesis of flax and hemp have not been fully explored. How does cadmium stress affect osmolytes, growth hormones, and internal CO₂ concentration in leaves? What is the trend in fiber crop growth and fiber crop properties with an increase in the duration and dosage of Cd stress?

3. Cadmium Absorption and Accumulation in Kenaf (Hibiscus cannabinus L.)

An investigation of Cd absorption and accumulation in kenaf is critical to understand the phytoremediation potential. Arbaoui et al. [55] observed that kenaf accumulated 2.49 mg kg⁻¹ Cd after three months of exposure to Cd stress without showing any toxic effects. These findings showed the phytoremediation potential of kenaf [55]. Chen et al. [56] evaluated two kenaf cultivars, Guanghong01 (GH) (Cd-tolerant) and Yuanjiang (YJ) (Cd-sensitive), under Cd stress (10 mg L⁻¹ CdCl₂) conditions for 7 days. The plant height and biomass of cultivar Guanghong01 (GH) were higher than those of Yuanjiang (YJ). Chen et al. [56] found that Cd in the roots and shoots of GH increased to 1897.5 and 320.3 mg kg⁻¹, while in the YJ cultivar, the concentration of Cd in roots and shoots increased to 1706.1 and 238.1 mg kg⁻¹ [56]. In another study, Chen et al. [33] evaluated the kenaf cultivar Fuhong 991 under Cd stress conditions (10 mg L⁻¹ or 30 mg L⁻¹) for one week. Cd contents in the roots and shoots reached to 1986.5 and 321.1 mg kg⁻¹ when exposed to 10 and 30 mg L⁻¹ Cd, respectively [33].

A new study tested a Cd-sensitive cultivar ‘Z367’ (Z) and a Cd-tolerant cultivar ‘FY2’ (F). The plants were subjected to 0, 2.5, and 200 μmol/L Cd stress. The tolerant cultivar accumulated more Cd than the sensitive cultivar. The results indicated that cultivar F accumulated (1660.08 mg/kg DW) Cd in the roots and shoots (418.12 mg/kg DW) compared to cultivar Z (1347.25 mg/kg DW; 546.94 mg/kg DW). Tolerant cultivar ‘F’ exhibited higher ascorbic acid (AsA) and glutathione (GSH) activities under Cd stress [43]. Different kenaf cultivars have different tolerances to Cd stress, as proved by various studies. China Kenaf 13 is one of the most significant kenaf varieties. Five-day-old kenaf plants were grown under control and Cd stress (10, 50, 100, 150, 200 μmol L⁻¹) conditions and showed higher tolerance to Cd stress through the increased activities of antioxidant enzymes (SOD and POD) and non-enzymatic antioxidants like ascorbate [22]. Cd accumulation in kenaf varies among organs, including the shoot, root, and leaves. The tolerance level of different organs is different. Luo et al. [30] evaluated two kenaf cultivars and a hybrid to investigate Cd accumulation in different plant organs. Cd contents were higher in the root, followed by the leaves and stem, and the degree of tolerance was F₁ > CP089 > CP085. The Cd contents in the roots, stems, and leaves of hybrid F1 were 2390.78, 504.65, 336.95 mg/kg, which were higher than in the parents CP089 (2313.53, 453.04, 277.51 mg/kg), and CP085 (2123.40, 363.41, 247.57 mg/kg). A translocation factor analysis showed that the F₁ hybrid had a better Cd transportation ability than its parents [30]. Kenaf has a higher remediation potential for Cd. To investigate their phytoremediation ability, two kenaf varieties, HC-95 and HC-3, were evaluated under three different levels of Cd stress. The values of the bioconcentration factor showed that kenaf was a Cd accumulator. The results showed that Cd slowly moved from the roots to shoots in kenaf, and the rate of the translocation factor increased with the increase in Cd stress in a hydroponic solution [24].

A recent study conducted an in-depth investigation of the phytoremediation ability of kenaf cultivars grown in contaminated soils in Southern China. Cd uptake, translocation, and the accumulation of Cd in different tissues among kenaf cultivars were investigated. The authors observed that Cd uptake in phloem ranged from 47% to 61%, while in xylem, it ranged from 38% to 53%. Hence, Fuhong R1, Fuhong 952, and Fuhong 992 were selected for the phytoremediation of Cd-contaminated soils [57]. Pan et al. [58] tested the kenaf cultivar (Zhe-367) and revealed the mechanism of arbuscular mycorrhizal fungi (AMF) in the alleviation of Cd stress. These authors observed that AMF inoculation decreased Cd absorption, indicating that AMF significantly impacts Cd absorption. Another study was conducted to explore the phytoremediation potential of kenaf on Cd-contaminated soils [58]. Three kenaf cultivars, Hongyou No. 2, GGS, and HP, were tested under Cd stress conditions. A Cd stress of 0, 40, and 80 mg/kg was added to the soil, and the results showed that all kenaf cultivars had higher tolerance to Cd stress. The content of Cd in kenaf varieties was higher than the control, reaching 355 mg/kg in Hongyou No. 2. These kenaf cultivars had best potential for Cd uptake, were regarded as highly tolerant to Cd stress, and can be used for the phytoremediation of Cd-contaminated soils [40]. These studies showed that kenaf has a higher degree of Cd tolerance, as it accumulated high Cd contents in its underground and aerial parts. Most of the studies mentioned above showed kenaf’s response to low or moderate levels of Cd stress. Kenaf’s responses to higher levels of Cd stress require additional studies. The growth of kenaf under a lethal concentration of Cd would determine whether kenaf is an ideal candidate for the phytoremediation of Cd-contaminated soils. For example, what is kenaf’s response to 200 mg/kg Cd stress? The increase in Cd content in the roots and shoots following increased Cd stress should be further investigated.

Molecular Mechanism of Cd Tolerance in Kenaf

Several research studies have presented the molecular mechanism of Cd tolerance in kenaf crop. One of the most influential techniques is the identification of potential genes regulating Cd tolerance. Two contrasting Cd-sensitive kenaf cultivars (GH and YJ) were used in an experiment to determine Cd tolerance in kenaf. GH had a stronger Cd absorption and accumulation ability than YJ. Molecular analysis revealed that 2221 and 3321 genes were expressed in GH and YJ (

Table 1. Molecular analysis of Cd tolerance in kenaf.

Cultivars	Total Genes	Role	Reference
CP032		HcERF.C3 gene was proven to have a positive effect on Cd tolerance and Cd homeostasis via virus-induced gene-silencing analysis.	[59]
GH and YJ	2221 and 3321	Activating the antioxidant defense system, heavy metal transport and detoxification, substance transport, plant hormone, and calcium signals.	[56]
FH991	3926	Transport and catabolism, heavy metals’ transport, detoxification of heavy metals, antioxidant activities, carbohydrate and energy metabolism.	[33]
F and Z	3439	Phenylpropanoid biosynthesis, plant hormone signal transduction pathways, vacuolar compartmentalization of Cd, and Cd uptake and transport.	[43]
Zhe 367		AMF increases the expression of critical genes (Hc.GH3.1, Hc. AKR, and Hc. PHR1), which increases Cd tolerance in kenaf.	[58]
CP085, CP089 and their hybrid F1 seedlings		Expression of NPF2.7, NADP-ME, NAC71, TPP-D, LRR-RLKs, and DHX51 was altered due to cadmium stress; related to cytosine methylation regulation.	[30]

). In GH, 689 genes were upregulated and 1532 genes were downregulated, while 2451 and 870 genes were upregulated- and downregulated in YJ. In GH, more genes were upregulated, which showed that GH activated several genes to cope with Cd stress. This study identified vital genes, such as (SOD2, PODs, MT1, DTXs, NRT1, ABCs, CES, AP2/ERF, MYBs, NACs, and WRKYs. Further, KEGG and GO analyses indicated that these genes are involved in different pathways, including antioxidant activities, HM transport, calcium signaling, and hormone activity [56]. Overall, these results indicated that these genes can be further investigated for their role in Cd tolerance, and the validation of these results is essential. Most of the studies related to kenaf involve the use of one cultivar for transcriptome analysis. A comparative transcriptome analysis of kenaf cultivars is highly recommended.

Earlier, Chen et al. [33] evaluated one kenaf cultivar (FH991) to conduct physiological and transcriptome analyses under different doses of Cd stress. The transcriptome analysis identified 3926 genes (1206 up- and 2720 downregulated). The validation of 15 genes was achieved using real-time quantitative PCR. KEGG analysis showed that the DEGs identified under Cd stress were mainly involved in transport and catabolism, heavy metals’ transport, the detoxification of heavy metals, antioxidant activities, carbohydrates, and energy metabolism. HIPP22 and MT2 were engaged in heavy metal transport and detoxification, and POD19 was involved in antioxidant enzyme activity. Further investigation of these critical genes could help regulate Cd uptake and transportation in kenaf [33].

In a recent study, two kenaf cultivars were treated with Cd stress to conduct physiological and transcriptome analyses of Cd tolerance. Cd-tolerant cultivar F resisted Cd stress by producing more antioxidant enzymes and ascorbic acid (AsA). A total of 3439 DEGs were identified in both cultivars. Functional annotation analysis showed that phenylpropanoid biosynthesis and plant hormone signal transduction pathways were significantly enriched. The genes encoded cellulose, pectin, and hemicellulose, which are involved in the chelation of Cd in the cell wall. The results suggest that the genes Narmp3, ABCC3, and V-ATPases could contribute to the vacuolar compartmentalization of Cd. HcZIP2 was responsible for Cd uptake and transport [43]. Another study showed that several factors like AMF can regulate the expression of Cd-tolerant genes in kenaf. Under Cd stress, AMF improved the expression of several essential genes (Hc.GH3.1, Hc. AKR, and Hc.PHR1) that are critical to increasing Cd tolerance in kenaf [58].

DNA methylation analysis also provides novel insights into Cd tolerance in kenaf. In an earlier study, two kenaf cultivars, CP085 and CP089, and their hybrid F₁ seedlings were subjected to Cd stress. A methylation-sensitive amplification polymorphism (MSAP) analysis found that DNA methylation decreased by 16.9% under cadmium in F₁ and increased in parents CP085 and CP089 by 14.0% and 3.0%, respectively. Cd toxicity altered the expression of different genes (NPF2.7, NADP-ME, NAC71, TPP-D, LRR-RLKs, and DHX51) linked with the regulation of cytosine methylation. These authors found that a lower level of DNA methylation regulates gene expression and leads to a substantial increase in Cd tolerance [30].

Recently, Cao et al. [59] conducted a transcriptome analysis of Cd tolerance in kenaf seedlings. Kenaf seedlings were treated with a Cd stress of 200 μM CdCl₂·2.5 H₂O for 15 days. DEGs such as ZIP1, ZIP5, ABCG8, ABCC14, CAX18, VIT4, and HIPP20 were upregulated through the application of nitric oxide (NO) and involved in Cd absorption and transport. NO also enhanced the activities of SOD, POD, CAT, and glutathione reductase (GR). A transcriptome analysis revealed that DEGs were involved in different pathways, such as the ROS metabolism, nitrogen and sulfur metabolism, and cell wall biosynthesis [59].

Studies conducting a transcriptome analysis of Cd tolerance in kenaf are still limited. Comparative analyses of Cd tolerance in contrasting kenaf cultivars are rarely reported. More studies are needed to identify new genes linked with the transport and accumulation of heavy metals. Despite the studies that have been carried out, the mechanism of Cd tolerance in kenaf is not yet fully understood. Studies on DNA methylation could be further increased to investigate the role of low DNA methylation in activating new genes for Cd tolerance.

4. Uptake and Accumulation of Cd, and the Use of Agronomic Approaches to Increase Cd Tolerance in Cotton

HMs pollution causes significant environmental damage; therefore, urgent solutions are needed to counter this issue. Different agronomic approaches like the application of biochar and biofertilizers effectively improved the growth of cotton exposed to Cd stress. An earlier pot experiment was conducted to study the role of biochar and biofertilizers in increasing Cd tolerance in cotton. The results showed that biochar (3%) and biofertilizer (1.5%) reduced Cd uptake and increased the root dry weight of cotton. Cotton seedlings showed an increase in SOD (47.70% and 77.21%) and CAT (35.40% and 72.82%) activities in roots and leaves [60]. In a recent pot experiment, it was concluded that biochar application reduced Cd concentration in cotton roots by 24.9% and enhanced the activity of antioxidants like CAT, SOD, and POD [61]. Another way to reduce Cd stress in cotton is by using hormones and osmolytes. Earlier studies revealed that melatonin osmolyte improved Cd tolerance in cotton by improving its photosynthetic and biochemical traits. MT-treated cotton seedlings showed an improvement in the activities of SOD, POD, and CAT. More studies are needed to understand the role of biochar and other organic amendments in Cd tolerance in cotton, especially their role in improving root and shoot growth [62].

Cd stress (5 mg kg⁻¹) increased the Cd content in roots and reduced plant height, biomass, antioxidant enzymes activity, and photosynthetic traits, while modifiers (polyacrylate compound modifier, and organic polymer compound modifier) improved plant biomass, photosynthetic characteristics, and antioxidant enzyme activities (SOD, POD, and CAT). These liquid modifiers could provide a real solution to reducing Cd toxicity in the future, but they still need to be further investigated under modified doses of Cd stress [63].

Likewise, polymer amendments (PAs) were used for the phytoremediation of soils polluted with heavy metals because they efficiently absorb metal ions. The results showed that PA improved Cd tolerance in cotton by downregulating oxoglutaric acid, which increased the photosynthetic rate (7.71–46.20%) and antioxidant enzyme activity (15.49–45.50%) in leaves. PA also downregulated the expression of jasmonic acid, reducing the Cd concentration in soil and reducing the transfer of Cd from root to bolls by 54.39%. The effects of PA on root and shoot growth under Cd stress need further investigation [64]. A recent study also showed that polymer amendments altered the metabolites in soil and cotton leaves under Cd stress. Cd stress increased Cd content in soils and reduced enzyme activities. The application of PA increased the boll number, yield, and fiber strength to 14.17%, 21.04%, and 19.89%, respectively, compared with the Cd treatment. The PA also altered the nicotinate and nicotinamide metabolism pathway and proline metabolism pathway in cotton. Further studies will provide a comprehensive understanding of PA-mediated Cd tolerance in cotton. The effect of a PA on Cd accumulation and detoxification is critical to investigate in future studies. The role of PA in the regulation of growth hormones in cotton should be investigated. In this way, PA applications can be increased to study the changes in metabolites in Cd-treated cotton plants [65].

An earlier study investigated the role of nanoparticles in reducing Cd stress in cotton. Zinc oxide nanoparticles play a crucial role in developing Cd tolerance in cotton. ZnONPs improved the root and shoot growth in Cd-treated cotton seedlings compared to the control. ZnONPs also upregulated the activities of chlorophyll a, b, proteins, and antioxidant enzymes (SOD, POD, and CAT). Overall, these results indicate that the addition of ZnONPs can reduce Cd accumulation in cotton by activating the biochemicals (protein, SOD, CAT) and photosynthetic machinery [66]. Melatonin (MT) plays a key role in reducing Cd stress in cotton. The results of a recent study showed that the application of melatonin (100 μM MT) increased root volume, surface area, and root length, and regulated Cd transport in cotton plants affected by Cd stress [67]. Another study showed that MT application decreased Cd accumulation in cotton seedlings through the regulation of antioxidant enzyme activities and cell wall biosynthesis gene expression. Further studies are required to investigate the crosstalk of MT with other growth hormones in cotton grown under Cd stress [68]. The application of silicon (Si) (1 mM)) reduced Cd stress in cotton seedlings by improving photosynthesis, decreasing Cd uptake, and improving antioxidant enzymes activities. Further studies are required to expand on the use of Si to improve Cd tolerance in cotton [69].

There are few studies on the use of agronomic techniques to develop Cd tolerance in cotton. These research gaps should be covered in future research studies. What role do plant growth hormones like gibberellic acid (GA), abscisic acid (ABA), cytokinin (CK), and ethylene play in reducing Cd stress in cotton? More studies are required to address these issues. It would be better to screen a large number of cotton genotypes under Cd stress and apply growth hormones to select the best genotypes and treatments for further breeding programs. The application of Cd stress and improvements in cotton growth when using hormones can be studied at the seedling stage to screen for resistant seedlings. Hydroponic studies or the growth of plants in vermiculite would be ideal for these objectives. We are hopeful that agronomic approaches will be a more fruitful way to reduce Cd stress in cotton to ensure its growth in Cd-contaminated soils. Due to climate change and soil degradation, the threat of heavy metal pollution is increasing. Improving Cd tolerance in cotton may require more efforts and investment. With an increase in cadmium stress, the Cd tolerance of cotton will decrease; however, tolerance can be enhanced using the techniques mentioned above.

Molecular Mechanism of Cd Tolerance in Cotton

It is hard to improve polygenic traits using agronomic approaches. Hence, researchers are continuously working on the genetic improvement of cotton. Molecular techniques not only help us in detailed analyses of the genetic mechanisms of Cd tolerance but also help to accelerate the breeding of targeted traits that are hard or impossible to improve via conventional breeding. Molecular techniques allow scientists to precisely identify and manipulate targeted genes in crops and develop new varieties with desired traits [70,71]. A transcriptome analysis of cotton cultivars revealed the potential genes involved in Cd tolerance and their regulation mechanisms. In an earlier study, Chen et al. [25] conducted a transcriptome analysis of the root of cotton grown under three levels of Cd stress. A total of 1151 DEGs were identified in three Cd stress samples (

Table 2. Cadmium-responsive genes identified in cotton via transcriptome analysis.

Cultivars	Genes	Functions	References
C184 cotton species	1151 DEGs in roots	Binding action and catalytic activity, mainly metal iron binding, and some cellular and metabolic processes.	[25]
Han 242	4627 DEGs in the root, 3022 DEGs in the stem, and 3854 DEGs in the leaves	Heavy metal transporter-coding genes (CDF, ABC, and HMA), heat shock genes (HSP), and annexin genes	[72]
Cotton variety CCRI 45	5573, 7105, 7253, 25, 198, and 9 upregulated, and 6644, 7192, 7404, 9, and 59, downregulated, 195, 150, 150, 12, 24, 59 upregulated and 16, 11, 23, 38, 127, 66 downregulated differentially accumulated metabolites (DAMs)	These mechanisms include enhancing antioxidant capacity through regulating APX, flavonoids, and alkaloids, accumulating secondary metabolites associated with Cd chelation (like amino acids and derivatives), and controlling the transportation of cadmium ions through the activation of ABC transporters. In conclusion, this study offers new insights into how cotton responds to Cd stress through MT-mediated detoxification.	[73]
Cotton cultivar J-4B	LOC107894197, LOC107955631, LOC107899273	Improvement in leaf functions and plant growth.	[62]

). Most of the identified genes were involved in binding actions and catalytic activity, mainly metal iron binding, and some cellular and metabolic processes. A KEGG analysis showed that most of these genes were associated with the production of secondary metabolites like flavonoid biosynthesis, sucrose synthesis, and phenylalanine biosynthesis to enhance the activity of antioxidant enzymes, repair systems, and transport systems, and to reduce Cd toxicity. This study identified several genes involved in the biosynthesis of secondary metabolites, starch, and sucrose, as well as flavonoid biosynthesis, to improve the activity of antioxidant enzymes and transport and repair systems and reduce Cd stress. These pathways and genes can be further investigated to increase the Cd absorption ability of cotton cultivars [25].

This study also identified the 28 differentially expressed transcription factors in cotton roots. Twenty TFs were upregulated, including MYB, zinc finger (GATA-type, CCCH-type, and C2H2-type), leucine zipper, and NAC transcription factors. The main mechanisms of cotton tolerance to Cd stress were oxidation resistance, the thickening of physical barriers, and complete detoxification. Moreover, ethylene and brassinolide pathways were involved in the response of cotton to Cd stress. In addition, this study also identified the flavonoid biosynthesis, starch, and sucrose metabolisms under Cd stress [25].

In another experiment, Han et al. [72] identified the cotton genes and the potential mechanisms involved in Cd tolerance. An RNA-Seq analysis was conducted in cotton and 4627 DEGs were identified in the root, 3022 DEGs in the stem, and 3854 DEGs in the leaves. Some of these genes were related to heavy metal transporter-coding genes (CDF, ABC, and HMA), heat shock genes (HSP), and annexin genes. In the study, the researchers constructed an overexpression system of GhHMAD5 in Arabidopsis thaliana, which led to longer roots compared to roots of control plants. They studied the GhHMAD5-silenced cotton plants and noted their sensitivity to Cd toxicity. The authors stated that GhHMAD5 is critical in Cd tolerance [72]. A comparative transcriptome and metabolome analysis helped researchers to identify the genes and metabolites involved in Cd tolerance in cotton. This is perhaps the most valuable approach to deciphering the potential role of different factors involved in Cd tolerance. There are very few studies dealing with this aspect of cotton. To understand how Cd tolerance is regulated in cotton, a previous study analyzed the transcriptome and metabolites of the cotton variety CCRI 45 when subjected to different levels of Cd stress. The researchers investigated the role of melatonin in mediating Cd tolerance in cotton through a transcriptome and metabolome analysis. They found that at a concentration of 100 μmol L⁻¹, melatonin (MT) increased Cd accumulation in both roots and shoots. The exogenously applied MT also increased the synthesis of alkaloids and flavonoids and reduced the production of amino acids and lipids [73].

In another study, Cd-stressed cotton seedlings were rescued through the application of melatonin. The cotton seedlings were treated with 100 µM Cd with or without MT. MT affected several Cd transporters genes, such as LOC107894197, LOC107955631, and LOC107899273 in roots, which improved leaf functions and plant growth. The downregulation of Cd transporter genes by melatonin prevents Cd ion from being transported to leaf tissues. This indicates that the transporter genes that are differentially expressed (DEG) play a crucial role in the regulation of Cd transportation and sequestration in cotton, which is mediated by melatonin. According to the transcriptomic analysis, melatonin triggers the activation of mitogen-activated protein kinase (MAPK) signaling pathways to regulate stomatal adjustment and photosynthesis in Cd-stressed leaves. Additionally, melatonin protects intercellular organs, specifically ribosomes, from Cd-induced oxidative damage by promoting ribosomal biosynthesis and improving translational efficiency. These findings help explain the molecular basis for melatonin-mediated Cd stress tolerance in plants and offer valuable insight into the effective management of Cd accumulation in cotton [62].

Other molecular mechanisms involved the identification of an early sulfur-containing compound (cysteine), which plays a crucial role in sulfur metabolism. A genome-wide analysis of cysteine genes identified the GhCYS2 as an essential Cd-tolerance gene. Knocking down GhCYS2 in plants led to reduced levels of cysteine and glutathione. This, in turn, caused the accumulation of MDA and ROS within cells, disrupting the normal process of photosynthesis. As a result, the stomatal aperture of leaves decreased, and epidermal cells underwent distortion and deformation. Ultimately, all of these adverse effects led to plant wilting and a significant reduction in biomass. The association between Cd²⁺ and cysteine established in this study provides a valuable reference point for further investigation into cysteine synthesis genes’ functional and regulatory mechanisms [74]. There are few studies on the molecular mechanism of Cd tolerance in cotton. There are many TF families that have yet to be investigated and validated. The genes mentioned above should be used to develop transgenic cotton lines that can thrive under Cd stress conditions. Genetic engineering and CRISPR/Cas9 can exploit the genes of heavy metal transporters. There are few or no studies about the role of proteins in Cd tolerance in cotton. A comparative proteomic analysis will provide the protein profile of cotton cultivars exposed to Cd stress. Studies are urgently required to address the issues linked with Cd tolerance in cotton. The characterization of genes related to antioxidant enzyme activity would improve the antioxidant machinery of cotton under Cd stress. In addition to this, QTL mapping should be conducted to detect the genomic regions regulating Cd tolerance in cotton.

5. Cadmium Tolerance in Flax, and Absorption, Accumulation, and Growth Traits

Flax is used for multiple purposes, such as food and fiber. Cd accumulation in flax demonstrates its Cd tolerance ability. Flax could be a potential candidate for the phytoremediation of Cd-contaminated soils. Different flax varieties have different levels of resistance to Cd stress. Various studies have shown the Cd-accumulating nature of flax cultivars. A flax cultivar was grown on sand containing 0.1mM of Cd in the greenhouse. The results showed that the root and stem accumulated more Cd (750 and 360 mg/kg dry matter), exceeding the threshold value defined for hyperaccumulator plants. This ability increases the possibility of flax growth in Cd-accumulated soils. Hence, the basal stem and root play a key role in Cd accumulation in flax [75]. Flax’s ability to tolerate a lethal amount of Cd was tested by Smykalova et al. [76] using 25 varieties of flax plants grown under 19 mg/L Cd²⁺. Cd levels were established for the in vitro selection system to determine the flax lines with improved Cd accumulation and tolerance. Cd accumulation in the roots was affected by the regeneration of explants in each variety. Tolerant varieties were selected based on the observed traits. The results suggested that flax varieties can be used for the phytoremediation of Cd-contaminated soils [76].

Later, a four-year field experiment was conducted to investigate the Cd accumulation ability of six flax varieties grown in Cd concentrations of 10–1000 mg Cd kg⁻¹ soil. Cd content was higher in roots than shoots. An increase in Cd stress did not affect the flax cultivars, and it was found that flax was a Cd accumulator. The content of Cd was increased in roots following an increase in Cd stress. A more Cd-tolerant variety, cv. Jitka, was selected for further use and considered as a possible candidate for the phytoremediation of Cd-contaminated soils by Bjelková et al. [77].

Examining the difference in Cd accumulation and Cd tolerance makes it possible to select the best-performing flax variety for phytoremediation. Keeping this point in mind, House et al. [78] investigated the difference in Cd content in four flax varieties: AC McDuff, CDC Bethune, AC Emerson, and Flanders. Cd concentrations ranged from 0.39 to 2.42 mg/kg. Roots and leaves had higher Cd content compared to other plant organs. AC Emerson had a higher Cd content than AC McDuff across tissues and ages, including seeds [78]. A recent study by Brutch et al. [79] tested six types of fiber flax under Cd stress for 49 days in the Jacobsen apparatus to further investigate the Cd tolerance and accumulation of the flax cultivars. Two flax cultivars were selected due to their adult plant resistance, and it was concluded that the presence of Cd did not influence flax germination, and plants showed resistance to Cd stress. The flax variety Orshanskiy 2 had a higher tolerance to Cd stress. The stable growth of this variety indicates that it can grow well in Cd-affected soils. In a greenhouse experiment, susceptible flax varieties like Svetoch and Priziv 81 were tested under Cd stress [79]. The phytoremediation ability of flax varies from genotype to genotype. Recently, Zhao et al. [40] conducted a greenhouse experiment for a detailed investigation of the phytoremediation of trace metals using flax varieties (Y2I328, Y2I329, and Zhongyama No. 1). These varieties were tested against Cd at 0, 40, and 80 mg/kg. The results showed that a high Cd stress of Cd 80 mg/kg reduced the emergence rate of all cultivars, while a low dose of 40 mg/kg enhanced the emergence rate of all varieties. Hence, these varieties can remove Cd in contaminated soils [40]. In a field experiment, eight flax cultivars were evaluated for their Cd tolerance. The results showed that one cultivar (Zhongya 1) extracted a significant concentration of Cd and could be regarded as a Cd-accumulator for the phytoremediation of Cd-affected soils [80].

Most of the flax varieties are described as being tolerant to Cd stress. One of the most ideal ways of testing flax tolerance is to investigate the plant response under different Cd levels. For example, the growth of flax cultivar cv Hermes under 0.5 mM Cd for 18 days on culture medium resulted in Cd compartmentation in roots. The growth traits of Cd-treated flax were moderately changed, and protein carbonylation and antioxidant enzyme activities were higher in the roots. Cd stress also increased the activities of cell wall enzymes (peroxidase and pectin methylesterase). In this way Cd tolerance can be increased in plant [29].

5.1. Agronomic Approaches

Agronomic approaches, which include using growth hormones like salicylic acid (SA), are critical in reducing Cd stress in flax. Three earlier studies investigated the role of SA in increasing Cd tolerance in flax seedlings. The first study revealed that SA reversed the Cd-induced (0, 50, and 100 μM CdCl₂) changes in root traits in flax seedlings. SA-treated seedlings showed a reduction in Cd bioaccumulation factors (BAF) in roots. These results revealed that SA might be a critical factor affecting the Cd uptake, sequestration, and translocation processes, mainly in the roots of flax seedlings [81]. A recent study focused on the combined effect of three plant growth hormones, SA, 24-epibrassinolide (EBL), and sodium nitroprusside (SNP), on growth and the antioxidants system of flax subjected to cadmium stress. The combined application of these hormones enhanced antioxidant enzyme activities (SOD, POD, and CAT) and reduced the content of H₂O₂. This interactive effect of hormones is highly recommended to enhance Cd tolerance in flax [82].

Belkadhi et al. [83] treated flax seedlings with an exogenous application of SA to reduce Cd-induced oxidative stress. SA-treated seedlings showed higher levels of antioxidant enzyme activities and increased levels of H₂O₂-detoxifying enzymes to counter the toxic effects of Cd stress [83]. In another study, SA was applied to Cd-stressed flax seedlings to reduce the harmful effects of Cd stress (100 µM CdCl₂) and protect the phospholipids. The pretreatment of flax seedlings with SA for 8 h protected phospholipids from the toxic effects of Cd stress. SA increased the proportion of linolenic acid (18:3) in Cd-stressed flax seedlings [84]. These results showed that agronomic approaches are vital to protect flax seedlings from the increasing effects of Cd stress. The use of other plant growth hormones, such as JA, GA, CK, and ET, in flax is rarely reported. In future studies, it is critical to consider the role of nanoparticles and other fertilizers in reducing Cd stress. However, rapid changes in the climate, soil degradation, and industrialization threaten the growth and productivity of flax crops. Climate change, such as increasing temperatures, would enhance the uptake and toxicity of Cd in flax crops and ultimately reduce tolerance to cadmium stress. Secondly, soil degradation can decrease soil pH, making Cd more mobile and available for crop absorption. It is critical to employ the abovementioned techniques to combat the rising threats of climate change.

5.2. Role of Genetic Factors in Regulating Cd Tolerance in Flax

Flax is a very significant fiber crop [85,86] and a good accumulator of Cd, but molecular studies describing the role of genetic factors in Cd uptake, accumulation, and tolerance are limited. Transcriptome profiling is a powerful tool that allows us to comprehensively understand gene expression and regulation, and to identify critical genes involved in stress tolerance mechanisms. There are several ways to study transcriptomes, including expressed sequence tags (ESTs) and RNA sequencing. The choice of method depends on the availability of genomic resources and the plant type. RNA sequencing is the most effective and cost-effective method [87]. The flax genome and the availability of genetic maps help in the rapid identification of several genes related to abiotic stress tolerance [88,89]. The molecular mechanisms of flax tolerance to Cd stress are mostly unknown [90]. A proteomics analysis revealed the number and type of proteins involved in cadmium tolerance in flax. The insufficient number of molecular studies makes it hard to obtain a detailed understanding of Cd tolerance in flax.

A detailed proteomics analysis investigated the relative changes in proteins in flax cultivars exposed to Cd stress. Cd stress significantly changed the expression of 14 proteins, mainly related to defense, metabolism, storage, and cell structure. Cd-treated cell suspension cultures of contrasting cultivars were prepared, and two-dimensionally separated proteins were extracted to observe the changes using mass and spectrometric analysis. Two proteins, ferritin and glutamine synthetase, were upregulated in the cultivar cv. Jitka revealed that Cd tolerance in this variety is due to the maintenance of low Cd levels at sensitive sites using ferritin. However, the functional validation of these proteins requires further study [91]. A recent study also described the proteomics and metabolic changes in flax under Cd stress conditions. Four flax genotypes, including two fiber genotypes, an oilseed cultivar, and one transgenic line, were tested. This study reported the identification of 1400 different proteins representing diverse Cd tolerance mechanisms in flax. These proteins include metal-binding proteins, enzymes of flavonoids, transporters, and HSP70 proteins. Pipecolinic acid was involved in Cd accumulation, as determined through metabolome analysis. The accumulation of coumaric acid and cinnamic acid confirmed the role of flavonoids and polyphenols in Cd tolerance in flax [90].

Regarding genome-wide association studies, Khan et al. [92] provided a detailed overview of the gene families associated with heavy metals in flax. Based on a phylogenetic analysis, they identified twelve HMA genes and divided them into four HMA gene subfamilies. Two genes (LuHMA3, LuHMA4) were orthologs with Cd-associated genes in rice and maize. This HMA gene was highly conserved among subfamilies of flax and Arabidopsis. A gene ontology analysis suggested that most of these genes played a role in metal ion binding, catalytic activity, and transport. These genes have yet to be investigated for their role in Cd tolerance in flax [92]. Khan [93] utilized four major techniques to find genomic-based solutions to determine Cd content in flax. These techniques include GWAS, genomic selection (GS), genomic cross-prediction, and gene family identification. This study identified 198 ABC transporter and 12 heavy metals associated (HMA) genes in the flax genome, of which 9 were orthologous to Cd-associated genes. A transcriptomic analysis of eight tissues allowed for the functional annotation of these genes and confirmed the expression of the ABC transporter and HMA genes in flax seeds and other tissues. A panel of 168 flax accessions was sequenced using 43,000 SNPs. This study confirmed the effectiveness of the QTL-based technique in the identification of genomic regions associated with heavy metal tolerance. The identification of QTNs with minor and large effects has the potential to improve flax breeding [93]. A detailed understanding of the molecular mechanisms of Cd tolerance in flax has not yet been obtained. There are few or zero studies using GWAS, QTL, TFs, or transcriptome analyses. Conducting more studies to bridge the existing gaps and shed light on how to achieve genetic improvements in flax tolerance to Cd stress is essential.

6. Cd Uptake, Absorption, and Accumulation in Jute

Cd uptake in jute is determined by the amount of Cd is absorbed by the roots. Jute is one of the most significant fiber crops for the phytoremediation of heavy metals, particularly Cd [36,94,95]. To accurately determine the phytoremediation potential of jute, it is essential to screen a large number of germplasms under different levels of Cd stress. A review of earlier studies is provided as follows. Two jute varieties, BJC-7370 and CVE-3, were studied for their phytoremediation potential at the early growing stage. The jute varieties were tolerant and super-accumulators at the highest Cd level (10 mg L⁻¹). Cd was transported from roots to shoots, and the translocation rate was reduced with an increase in Cd in the nutrients solution. Cd contents in the roots were higher than in the shoots. The bioaccumulation factor was the ratio of heavy metals in the dry parts of the plant (roots or shoots), and values were calculated on a dry weight basis. At 5 mg L⁻¹ of Cd, the bioconcentration factor of the jute varieties was 497.08 and 311.52, respectively. However, this study tested the jute varieties using low doses of Cd. More studies should be conducted on adult plants’ response to higher doses of Cd stress. It can be concluded that jute varieties can tolerate low Cd levels due to their higher rate of seedling survivability and other morphological indicators, as well as the bioconcentration factor (BCF) [24]. Another study revealed the phytoremediation capacity of jute in Cd-contaminated paddy soils. Biomass and Cd concentration in three jute cultivars were studied, and then Cd uptake was observed. Each cultivar and organ had a different Cd content and different tolerance levels. The jute cultivar Lianhonghuangma had a higher Cd content than the other two cultivars. The plant capsule had the highest Cd content, at 4.75 mg/kg⁻¹, followed by the petiole, at 4.27 mg/kg⁻¹. The other four organs’ Cd content was in the order leaf > root > xylem > phloem. Lianhonghuangma had a higher Cd accumulation (53.3 g.hm⁻²) than ‘Huangma 179’ and ‘Minhouhongpi’. Xylem extracted 33.11–42.99% of the total Cd content taken up by the plant [96]. Hou et al. [97] used four jute cultivars to observe the phytoremediation potential of Cd-polluted soils. Bioaccumulation factors, transfer factors, and accumulation were studied. The cultivar Funong 4 had a greater biomass (21.53 and 26.18 t/hm²⁾ at the flowering and maturity stage than other cultivars. Guimacai1 had the highest accumulation of Cd (50.160 g/hm²). Guimacai1 can be recommended for the phytoremediation of Cd-contaminated soils [97]. A pot experiment was conducted to observe the rate of Cd absorption in jute roots grown in Cd-polluted soils. The results revealed that jute root absorbed a significant amount of Cd and could be regarded as a Cd-accumulator [98]. More studies are needed to screen more jute genotypes under Cd stress conditions to select the best Cd-accumulator genotype for the phytoremediation of contaminated soils.

6.1. Role of Agronomic Approaches in Cd Tolerance

Agronomic practices are crucial for mitigating the effects of Cd stress in jute. Although jute is primarily known for its ability to accumulate Cd, some studies have looked into food safety concerns related to jute. The Cd ratio found in jute was 0.7 mg/kg, which exceeds the threshold limit recommended for food safety. Therefore, soil amendments using recycling plant (ACARP) compost can help minimize the health risks posed by Cd in jute [99].

A separate study found that applying selenium (Se) can reduce the stress caused by Cd in jute cultivars. Jute was planted for 265 days, with or without the application of Se. The cultivar GB2762-2017 accumulated the highest amount of Cd (1.37 times) in the leaves when the Se dose was not applied. However, a Se dose reduced the Cd risk by 38.32%, and levels were reduced to less than 1. Therefore, it can be concluded that Se application can help minimize Cd enrichment in jute, possibly by preventing the roots from uptaking metal ions, reducing oxidative stress, and improving growth and photosynthesis [100]. Citric acid was found to positively affect Cd²⁺ stress (at a concentration of 20 mg/L) in jute. Applying citric acid at a concentration of (5mM) was shown to effectively reduce Cd²⁺ uptake and accumulation in both the roots and shoots of the plant. Furthermore, this application was found to increase the activities of antioxidant enzymes (SOD and POD) [50].

Graphene oxide (GO) is a versatile material with unique physicochemical properties that is used in various scientific fields, such as biochemistry, environmental, and plant science. However, there is still limited knowledge regarding how the presence of GO affects the growth and Cd tolerance of plants that are used for environmental remediation purposes. In this experiment, jute seedlings were grown in a Cd-contaminated aquatic system and exposed to varying GO concentrations (5, 10, and 20 mg/L). The study evaluated parameters such as biomass, reactive oxygen species, and antioxidant activities. Low concentrations of GO reduce oxidative stress by enhancing the activity of antioxidant enzymes and increase Cd uptake in jute, increasing Cd tolerance. Further studies could reveal the possible future use of GO to improve the phytoremediation ability of jute [101].

Despite all these research studies, the uptake of Cd and its regulation are still not fully understood, indicating limited progress in the development of a Cd-tolerant jute crop. There is a need to explore more agronomic approaches, such as using growth hormones and nanoparticles to reduce or increase Cd uptake in jute. The specific objectives of each study can vary. Using any of these approaches will significantly enhance jute’s ability to tolerate excessive Cd stress and its potential for the phytoremediation of Cd-contaminated soils. Combining several agronomic approaches could be a better way to enhance Cd tolerance in jute.

6.2. Molecular Factors Regulating Cd Tolerance in Jute

Studies on the molecular mechanisms of tolerance to heavy metal stress can help us improve jute’s genetic makeup [102]. Due to the rapid development in sequencing technology, the identification of genes expressed in specific tissues and organs is of tremendous importance. Gene sequencing technology has primarily promoted investigations into the molecular biology and genetics of jute. Several transcriptome studies have been reported in crops dealing with gene identification related to vegetative growth [103], abiotic stress tolerance [104], and fiber development [105]; however, transcriptome analysis of Cd tolerance is rarely reported in jute. Hauqe et al. [106] completed an expression profiling of microRNAs and their targets for the phytoremediation of heavy metals in jute, but they did not include the Cd in their study [106]. Rahman et al. [107] published a review article that explains the progress made in jute’s response to heavy metal stresses. However, the article did not report any molecular study related to Cd tolerance. This indicates that few studies have been conducted on jute’s tolerance to Cd stress. This highlights the importance of considering this matter to identify novel genomic regions that can increase the phytoremediation ability of jute [107]. Recently, Yang et al. [108] conducted GWAS and RNA-seq analyses to reveal the genetic basis of Cd accumulation and absorption in jute cultivars. The gene (COS02g_02406), linked with Cd-tolerance, and gene (COS06g_03984), associated with Cd-absorption, were identified in flavonoid biosynthesis pathways and ethylene response signaling pathways. These genes can be exploited using molecular breeding tools [108].

There are dozens of molecular studies dealing with Cd tolerance in other crops, which can serve as a reference point for jute. Studies that use QTL mapping, GWAS, TFs analyses, and CRISPR/Cas9 in jute are very rare or have not been reported. These techniques have tremendous importance in the development of heavy metal tolerance in crops [71,109,110,111], particularly in jute, regarding Cd tolerance, and could encourage further studies aiming to elucidate the genetic control of Cd uptake and transport, as well as the accumulation mechanisms, in jute. QTL mapping, GWAS, and TFs analyses should be explored in detail in future research to identify the potential genomic regions regulating Cd tolerance in jute and increase the genetic resources for future breeding programs. Additionally, the use of CRISPR/Cas9 could be beneficial for precisely editing targeted genes in jute.

7. Cadmium Uptake, Accumulation, and Tolerance in Hemp (Cannabis sativa)

Hemp is a crucial crop for the phytoremediation of heavy metals [112,113]. A detailed review of this aspect was published recently [114]. Hemp is best known for its phytoremediation potential and recent study demonstrated hemp’s ability to grow in Cd-contaminated soil. A pot experiment was conducted to evaluate hemp plants’ phytoremediation ability. Hemp plants showed better performance at 3 mg Cd Kg⁻¹ and exhibited their potential for phytoremediation [115]. Linger et al. [116] demonstrated that hemp plants had phytoremediation potential when 126 g of Cd was used (ha vegetation period)⁻¹ [116]. Linger et al. [117] showed that hemp roots had a higher tolerance to Cd stress of more than 800 mg (Cd) kg⁻¹(d.m.) [117]. Recently, two hemp varieties, Futura 75 and Kc Dora, were tested against three levels of Cd. Futura 75 accumulated more Cd in leaves under low Cd stress [118]. At higher levels of Cd stress (120 mg kg⁻¹), the translocation of Cd from the roots to shoots decreased, leading to an increase in Cd content in the roots. Kc Dora showed a greater translocation of Cd, leading to the same Cd levels being detected in the roots and shoots. The Cd content in seeds was lower than that in the three organs at all Cd levels (Table 3) [118].

Another study documented the potential of two hemp cultivars, Henola and Białobrzeskie, as Cd decontaminators. The cultivars were tested under 4 mg/kg and 8 mg/kg Cd levels. The Henola variety had the highest Cd content (0.51 mg/kg in seeds) and a higher bioconcentration factor (BCF) than Białobrzeskie. The bioconcentration factor (BCF) measures the amount of heavy metal a plant accumulates in its tissues compared to the content of heavy metals in the soil in which the plant is grown [112]. An earlier research study showed that Cd content in hemp shoots was (66 μg g⁻¹) [119]. Cd uptake and Cd accumulation depends on the cultivar and plant parts. Earlier studies showed that oil hemp had a Cd content of 0.020 mg kg⁻¹, which could make it helpful in the selection of more tolerant cultivars [120]. Likewise, the Cd content in hemp seeds was (1.3–4.0 mg kg⁻¹), as determined by Mihoc et al. [121].

A recent study investigated Cd accumulation and translocation in four hemp varieties. In a hydroponic experiment, plants were exposed to Cd stress (0 mg·L⁻¹ Cd and 2.5 mg·L⁻¹ Cd). Cd content in the roots ranged from 16.1 to 2274.2 mg·kg⁻¹ in all varieties, while translocation factors ranged from 6.5 to 193. Whole-plant bioconcentration factors ranged from 20 to 1051 mg·kg⁻¹ [122]. Similarly, the Cd concentration in the aboveground parts of the plant in highly Cd-contaminated soil was 1.9 and 21.4 times greater than in that in soils with low Cd contamination levels, as Thurston [123] described. There appears to be a lot of research on Cd accumulation and tolerance in hemp; Table 3 provides some crucial values related to Cd uptake. However, there is still a need to expand the hydroponic studies on Cd stress tolerance in hemp.

Interestingly, the early seedling stage can provide an indication of Cd tolerance in hemp [23]. A field experiment in Pakistan showed that hemp accumulated Cd with a shoots–roots ratio of 1–1.1/1.2–1.35. The concentration of Cd is represented as mg kg⁻¹ [124]. Recently, Nash et al. [125] exposed hemp plants to Cd stress (10 mg·L⁻¹) and revealed that the Cd content in leaves was 23.2 mg·kg⁻¹ dw [125]. The hemp cultivar Wanma No. 1 was shown to achieve the highest Cd uptake as shown by Guo et al. [126]. There are dozens of studies dealing with heavy metal accumulation and tolerance in hemp. However, a detailed review focusing on Cd is missing. Table 3 shows the critical cultivars of hemp and their ability to accumulate Cd.

Table 3. Cd contents in different key cultivars of hemp.

Hemp Cultivars	Cd Content/Accumulation	References
Purple Tiger	23.2 mg·kg−1 dw in leaves	[125]
KC Dora	3 µg g−1 in seeds	[118]
Henola	0.51 mg/kg in seeds	[112]
Apricot Auto, Alpha Explorer, Von, T1	1056.8, 2274.2, 512.4, 16.1 in roots. Concentration represented as mg·kg−1 dw	[122]
Futura 75 and Tisza	Both cultivars accumulated 0.28 and 0.21 mg/kg in roots	[127]
Hemp variety	Cd contents in stem and roots ranged from 0.1 to 0.4 mg/g of dry mass	[128]
Carmagnola	1.7 mg/kg in fiber	[129]
Felina 32	0.39 mg/kg dry weight	[130]
Uso 31	1.80 mg kg−1 in leaves	[131]
Yunma No. 1	279.0 mg/kg	[40]
Santhica 27	5.59 mg/kg of DW in roots	[132]

Table 3. Cd contents in different key cultivars of hemp.

Hemp Cultivars	Cd Content/Accumulation	References
Purple Tiger	23.2 mg·kg−1 dw in leaves	[125]
KC Dora	3 µg g−1 in seeds	[118]
Henola	0.51 mg/kg in seeds	[112]
Apricot Auto, Alpha Explorer, Von, T1	1056.8, 2274.2, 512.4, 16.1 in roots. Concentration represented as mg·kg−1 dw	[122]
Futura 75 and Tisza	Both cultivars accumulated 0.28 and 0.21 mg/kg in roots	[127]
Hemp variety	Cd contents in stem and roots ranged from 0.1 to 0.4 mg/g of dry mass	[128]
Carmagnola	1.7 mg/kg in fiber	[129]
Felina 32	0.39 mg/kg dry weight	[130]
Uso 31	1.80 mg kg−1 in leaves	[131]
Yunma No. 1	279.0 mg/kg	[40]
Santhica 27	5.59 mg/kg of DW in roots	[132]

7.1. Role of Agronomic Techniques in Increasing Cd Tolerance in Hemp

Many researchers have employed numerous agronomic tools to protect hemp from the increasing threat of Cd stress, which have proven fruitful. For example, hemp seedlings were treated with 0, 25, 50, and 100 mg Cd kg⁻¹ sand (DW), and 500 μM of salicylic acid was applied. SA enhanced photosynthetic activities and antioxidant enzyme (SOD, POD) activities. The roots had 25.5–29.5 times more Cd content than shoots, suggesting that hemp is a Cd-excluder rather than an accumulator [46]. Another study showed that secondary metabolites like proanthocyanidins can be used under Cd stress conditions to protect hemp from toxic effects, as shown in a study in which Yunnan hemp no. 1 (YM1) was subjected to 100 µmol/L CdCl₂ for four days to achieve the experimental objectives. Proanthocyanidins increased photosynthetic activity and altered metabolic pathways [27]. The use of Arbuscular mycorrhizal fungi (AMF) can help to enhance hemp’s tolerance to Cd stress by altering its photosynthetic system.

In one study, hemp cultivar Huoma No. 1 was treated with 80 mg/kg Cd and AMF. The results showed that AMF reduced the Cd content in the leaves, increasing the transpiration rate. Under AMF inoculation, the light saturation points of the plants reached 1448.4 μmol/m²/s, and the optical compensation point was 24.0 μmol/m²/s [45]. This study did not show AMF to have any effect on the molecular factors of hemp. Another study revealed the positive impact of AMF on Cd content in hemp. AMF inoculation increased Cd contents in the roots, but Cd enrichment was reduced in leaves. Malic acid and citric acid levels changed, and Cd’s bioavailability also changed [133]. Many unresolved issues must be taken into consideration in future studies, for example: how does AMF affect protein content in hemp? The Cd stress can be increased to explore the maximum potential of AMF in reducing Cd content in hemp. Silicon was found to enhance the absorption of Cd and increase root–shoot translocation in hemp plants without affecting their growth. When Santhica 27 plants were exposed to 20 μM Cd stress, their growth and photosynthesis system were reduced, but the plants showed a translocation factor higher than 1, indicating their potential for phytoremediation. The treatment of the plants with silicon decreased the Cd content and enhanced the synthesis of phytochelatin and glutathione, which helped the plants cope with oxidative stress. The Cd content in the roots was 5.59 mg/kg of DW [132].

A recent study showed that applying a molybdate foliar spray at concentrations of 0.5, 1.0, and 2.0 ppm to Cd-stressed hemp increased the dry biomass of roots (1.87 g), proline levels in roots (69.00 µg/g), polyphenols, and chlorophyll contents. These results suggest that molybdenum (Mo) can help alleviate Cd stress by promoting plant growth and improving biochemical traits. The effect of molybdenum (Mo) on secondary metabolites under Cd stress has not been reported [134]. Recently, biochar was used to enhance hemp growth and yield under Cd stress (Cd 12 mg kg⁻¹). Biochar application significantly enhanced the aboveground biomass of hemp by ~1.67-fold for shoots, and by ~2-fold for both seed number and seed weight [135]. Further studies are needed to determine the impact of Si on antioxidant enzyme activities (SOD, and POD), water use efficiency, and plant biomass. Additionally, more agronomic techniques should be explored to protect hemp growth under Cd stress. Growth hormones can also be investigated to determine their effects on plant growth and Cd uptake in contaminated soils.

7.2. Genetic Analysis of Cd Tolerance in Hemp

The most effective and reliable way to increase hemp tolerance to Cd stress is the identification of genes regulating different pathways related to Cd tolerance (Figure 3). A detailed review of the molecular mechanism of Cd tolerance in hemp has not been reported. The increased transcriptome abundance in hemp is associated with plant exposure to heavy metal stress. For this reason, extensive efforts have been made by hemp researchers to identify the potential genes linked with the uptake and accumulation of heavy metals. The hemp cultivar ‘purple tiger’ was recently treated with 0, 2.5, 10, and 25 mg kg⁻¹ of Cd to conduct a transcriptome analysis of Cd tolerance. The transcriptome analysis revealed that genes were more highly expressed in the roots than in the leaves. The genes CsHMA3, CsHMA1, CsHMA4, and CsHMA5 (

Table 4. List of key Cd-tolerant genes and proteins in hemp.

Variety	Cd Dose	Genes	Function	References
Purple Tiger	0, 2.5, 10 and 25 mg L−1	CsHMA3 CsHMA1, CsHMA4, and CsHMA5	Regulate Cd uptake via roots; transport to shoot, leaves, and lower tissues.	[47]
Hemp genome		113 MYB genes	CsMYB024-induced cannabidiol biosynthesis pathway.	[136]
Santhica 27	10 μM for one week	122 proteins in leaves affected by Cd stress	PCS1-1, which increases phytochelatin biosynthesis.	[137]
Yunnan Hemp No. 1		CsANR and CsLAR	Proanthocyanidins’ synthesis.	[27]
NA	NA	Glutathione-disulfide reductase (GSR) and phospholipase D-α (PLDα)	These genes are responsible for the accumulation and tolerance of Cd in hemp leaves.	[138]
Hemp		CBF/DREB	A positive correlation was found between Cd accumulation and increased proline content.	[134]
	Yunma No. 1 (Ym), Neimengguxiaolidama (Nx)	22 DEGs for heavy metal detoxification process in Nx and 118 in Ym.	Results showed that peroxidase-, glutathione-, and S-transferase-regulated genes were both significantly upregulated in Nx and Ym. Cd-tolerance protein/phytochelatin synthase I thioredoxin reductase-related genes were specifically upregulated in Ym.	[34]

) were only upregulated in root tissues under 10 mg·L⁻¹ Cd. These transporters are responsible for the uptake of Cd in the roots and its subsequent transportation to the shoots and leaves. Further functional validation of these genes is required to understand the regulatory pathways they control [47].

Notably, genes identified in hemp under Cd stress are linked with several functions, such as the regulation of metabolites, photosynthesis, or antioxidant enzyme activities. Cd stress affects the production of secondary metabolites. In an earlier study, 113 genes that belong to the MYB family, which affect plant growth and secondary metabolites, were identified. Seven genes were critical genes involved in hemp response to Cd stress. CsMYB024 expression was changed following Cd stress, which ultimately induced the cannabidiol biosynthesis pathway [136]. This study did not mention the functional characterization of other genes that might be involved in other Cd-tolerance mechanisms. However, these genes can serve as reference points for future studies. Another study revealed the role of metabolite genes in Cd tolerance in hemp. Two essential genes for proanthocyanidins synthesis in industrial hemp were identified, CsANR and CsLAR, via transcriptome analysis in hemp cultivar Yunnan hemp no. 1 under Cd stress. The overexpression of these genes was induced in Arabidopsis, and the results showed that transgenic plants were more tolerant to Cd stress and the secondary metabolites content was higher [27].

Cd stress also intensively affects proteins and biochemicals, which may affect hemp’s response to Cd stress. Earlier studies indicated that Cd stress (20 μM) inhibited growth in hemp crops. After one week of Cd stress, transcriptome and proteomics analyses were performed for leaves and roots. Cd stress stimulated the expression of genes related to the proline and phenylpropanoid pathways. Cd stress was typically involved in the stimulation of PCS1-1, which increased phytochelatin (PC) biosynthesis. Cd stress affected the accumulation of 122 proteins in leaves, which might be a meaningful finding for future studies. Cd stress also affected the content of glutathione in hemp plants. Glutathione is a key non-enzyme antioxidant component of the antioxidant defense system [137]. The genes contributing to Cd accumulation could be an ideal genetic material for the molecular breeding of hemp. Ahmad et al. [138] identified two essential heavy metal transporters genes, glutathione-disulfide reductase (GSR) and phospholipase D-α (PLDα), in hemp. These genes were responsible for the accumulation and tolerance of Cd in hemp. The Cd content in leaves was 151 mg kg⁻¹ [138]. TF families also regulate plant responses to Cd stress, as shown by Ali et al. [134]. The expression of CBF/DREB genes in Cd-stressed hemp plants was shown to have a positive correlation with the accumulation (R² = 0.751) of Cd in hemp [134]. Earlier papers have shown that the number of Cd-tolerant genes in hemp is limited. To develop Cd-tolerant hemp that could be an ideal candidate for the phytoremediation of heavy metals, it is necessary to validate, clone, and transform these genes functionally. However, the role of lignin and flavonoids in Cd tolerance has not been deeply studied to date. Potential molecular research tools like QTL mapping and GWAS could be used to manually control QTL and genetic Cd tolerance in hemp. Genetic engineering and CRISPR/Cas9 could exploit these genomic regions in molecular breeding.

8. Role of CRISPR/Cas9 in Cd Tolerance in Fiber Crops

CRISPR/Cas9 is one of the most powerful gene-editing tools, without any biological barrier. CRISPR/Cas9 is well known for its simplicity, cost-effectiveness, and efficiency. CRISPR/Cas9 comprises a Ca9 protein and single-guided RNA, known as sgRNA. sgRNA guides the Cas9 protein to the targeted gene, and then the Cas9 protein binds with the targeted gene to cut the gene, hence inducing mutation [139]. CRISPR/Cas9 has been effectively used to edit targeted genes in many crops (Figure 3). In cotton, CRISPR/Cas9 was used to knockout the gene GhRCD1 to develop Cd-tolerant mutants. The results show that the overexpression of GhRCD1 resulted in enhanced tolerance in cotton seedlings through regulating the GhbHLH12–GhMYB44–GhHMA1 transcriptional cascade [140]. For example, CRISPR/Cas9 was used to knock out the OsLCD gene to develop low-cadmium rice, demonstrating the effectiveness of the gene editing tool. Knockout of the OsLCD gene decreased cadmium content in mutant lines [141]. The above findings show that the use of CRISPR/Cas9 in fiber crops is limited and challenging, particularly in jute, hemp, kenaf, and flax; this should be addressed in future studies. The use of CRISPR/Cas9 in fiber crops can allow for targeted gene editing and minimize Cd stress. We strongly suggest using CRISPR/Cas9 to edit the genes mentioned in this review in future research studies.

9. Conclusions and Future Outlook

Heavy metal stress is one of the most devastating issues of the current century. It not only affects environments but also induces damage to crops, soil pollution, and contamination of the food chain, as well as posing a risk to human life by increasing the risk of, for example, kidney and lung cancers. With the increase in the human population, the demand for food and clothing is surging, which puts more pressure on crop breeders to adopt reliable and novel breeding methods for the genetic improvement of crops. Fiber crops, such as cotton, flax, hemp, jute, and kenaf, have a long history of use in textile products. These crops are extensively used for making clothes, ropes, threads, and other textile products. This review article briefly describes the effects of Cd stress on crops, Cd uptake and Cd regulation mechanisms, and the role of agronomic and genetic techniques in enhancing Cd tolerance. For a long time, plant breeders have been trying to improve Cd tolerance in fiber crops using conventional breeding methods. These methods mainly depend on diversity and phenotypic selection, which are very costly and laborious. Due to anthropogenic activities and industrialization, the Cd content in soil is increasing to an alarming level. The increase in Cd content in agricultural lands is damaging the growth and production of crops, severely impacting global food security.

Removing heavy metals from soils ensures food security and protects agricultural lands. One way to achieve this is through the use of crops that can perform phytoremediation. Hemp, jute, kenaf, and flax crops effectively remove heavy metals from contaminated soils. These crops have been extensively studied for their ability to uptake and accumulate Cd in their aboveground (leaves and shoots) and belowground (roots) parts. By accumulating Cd content, these crops can clean up agricultural lands to allow for their safe use in growing other food crops. We must develop Cd-tolerant crops to combat the growing threat of Cd stress. Different molecular tools, such as transcriptome analysis and proteomics analysis, have identified several genomic regions that regulate Cd uptake, translocation, and accumulation in fiber crops. The Cd tolerance of many of these genes and proteins remained unknown, as did their potential use to develop tolerant crops that show excellent phytoremediation abilities for contaminated soils. Additionally, the complete molecular mechanisms of Cd tolerance are not yet fully understood. The lack of a complete understanding of crop genomes, genes, and regulatory networks makes it challenging to effectively improve Cd tolerance in economically important crops. QTL, GWAS, and TFs analyses of Cd tolerance in jute, kenaf and flax are limited, which hinders our understanding of the nature of roots and shoots regarding their ability to accumulate Cd. Genetic engineering, CRISPR/Cas9, and GWAS have not been widely used to understand the mechanism of Cd tolerance in fiber crops. CRISPR/Cas9 can effectively edit Cd-tolerant genes to increase their ability to accumulate Cd. The critical genes mentioned in this paper require further functional validation and could possibly be used to breed Cd-tolerant crops. By using QTL mapping, GWAS and TFs, several new genomic regions that may be involved in Cd uptake and crop accumulation can be identified, increasing their phytoremediation ability.

Exposing fiber crops to different doses and durations of Cd is essential because this will help us better understand their ability to uptake and accumulate Cd. It is important to note that crops’ response to Cd stress varies from genotype to genotype and according to plant part. Previous studies did not screen a large number of germplasms under Cd stress, which is a key issue hindering the selection of tolerant genotypes. Screening a large number of germplasms may be a more reliable way of identifying Cd-tolerant genes. The regulation of secondary metabolites could help to increase plant tolerance to Cd stress. Growing fiber crops on contaminated soils for extended periods could enhance their ability to remove pollutants from the soil. Increasing the expression of genes linked to heavy metal transporters is essential and can be achieved by exposing the crops to higher doses of Cd stress. For example, many transcriptomes and TFs identified in fiber crops have not yet been exploited in molecular breeding programs. The combined use of physiological, transcriptome, proteomics, genomics, epigenomics, metabolome, and GWAS analyses can effectively enhance our understanding of Cd tolerance in fiber crops. This is the most powerful technique and can be used for the identification of novel traits and genomic regions regulating Cd tolerance in fiber crops. Varieties with higher levels of Cd in their roots and shoots should be grown over large areas to remediate Cd-contaminated soils effectively. Another approach is to develop a crop ideotype that combines morphological and physiological traits that could improve the yield of fiber crops under Cd stress. Critical steps in developing a crop ideotype for Cd tolerance are to enhance the chelation mechanism, subcellular compartmentation, and antioxidant system in crops to counter the effects of Cd stress. By developing crop ideotypes, we can accelerate future breeding programs. The mitigation of Cd toxicity in plants is crucial because it may become more hazardous in the future. It is better to grow crops with phytoremediation potential, use fertilizers, and use tolerant varieties. These efforts will lead to the mitigation of Cd toxicity in fiber crops. The use of agronomic approaches such as melatonin, biochar, SA, nanoparticles, and the foliar application of growth hormones has not been reported for all crops.