CONCLUSIONS: Biofortification of legumes using diverse techniques such as plant breeding, agronomic practices, genetic modification, and nano-technological approaches presents a sustainable strategy to address micronutrient deficiencies of underprivileged populations. The widespread issue of chronic malnutrition, commonly referred to as \"hidden hunger,\" arises from the consumption of poor-quality food, leading to various health and cognitive impairments. Biofortified food crops have been a sustainable solution to address micronutrient deficiencies. This review highlights multiple biofortification techniques, such as plant breeding, agronomic practices, genetic modification, and nano-technological approaches, aimed at enhancing the nutrient content of commonly consumed crops. Emphasizing the biofortification of legumes, this review employs bibliometric analysis to examine research trends from 2000 to 2023. It identifies key authors, influential journals, contributing countries, publication trends, and prevalent keywords in this field. The review highlights the progress in developing biofortified crops and their potential to improve global nutrition and help underprivileged populations.

UNASSIGNED: Zinc (Zn) content is of great importance in healthy human diet, crop productivity and stress tolerance in soils with zinc deficiency. The genes used to increase yield per unit area such as semi-dwarf 1 (sdw1) is commonly considered to reduce mineral content of grain.
UNASSIGNED: In the present study, influence of sdw1.d, a widely used allele for short plant height in barley breeding, on zinc accumulation and tolerance to zinc deficiency were investigated. A near isogenic line of sdw1.d allele, its recurrent parent Tokak 157/37 and donor parent Triumph were grown in zinc-deficient and-sufficient hydroponic cultures. Two experiments were conducted until heading stage and physiological maturity.
UNASSIGNED: In zinc-deficient conditions, sdw1.d allele increased shoot dry weight by 112.4 mg plant-1, shoot Zn concentration by 0.9 ppm, but decreased root Zn concentration by 6.6 ppm. It did not affect grain characteristics, but increased grain Zn content. In zinc-sufficient conditions, sdw1.d allele increased shoot Zn content, and decreased root Zn content. sdw1.d did not affect grain weight but increased grain Zn concentration by about 30% under zinc-sufficient conditions. The results showed that sdw1.d allele has no negative effect on tolerance to zinc deficiency, and even promotes tolerance to zinc deficiency by more Zn translocation. It was revealed that sdw1.d allele improves Zn accumulation under both zinc-deficient and zinc-sufficient condition. The sdw1.d allele could contribute to solving the problems in plant growth and development caused by zinc-deficiency via improving tolerance to zinc-deficiency. It could also provide a better Zn biofortification.

The integration of zinc nanoparticles (Zn NPs) with biochar offers a transformative approach to sustainable agriculture by enhancing plant productivity and human nutrition. This combination improves soil health, optimizes nutrient uptake, and increases resilience to environmental stressors, leading to superior crop performance. Our literature review shows that combining Zn NPs with biochar significantly boosts the crop nutrient composition, including proteins, vitamins, sugars, and secondary metabolites. This enhancement improves the plant tolerance to environmental challenges, crop quality, and shelf life. This technique addresses the global issue of Zn deficiency by biofortifying food crops with increased Zn levels, such as mung beans, lettuce, tomatoes, wheat, maize, rice, citrus, apples, and microgreens. Additionally, Zn NPs and biochar improve soil properties by enhancing water retention, cation exchange capacity (CEC), and microbial activity, making soils more fertile and productive. The porous structure of biochar facilitates the slow and sustained release of Zn, ensuring its bioavailability over extended periods and reducing the need for frequent fertilizer applications. This synergy promotes sustainable agricultural practices and reduces the environmental footprint of the traditional farming methods. However, potential ecological risks such as biomagnification, nanoparticle accumulation, and toxicity require careful consideration. Comprehensive risk assessments and management strategies are essential to ensure that agricultural benefits do not compromise the environmental or human health. Future research should focus on sustainable practices for deploying Zn NPs in agriculture, balancing food security and ecological integrity and positioning this approach as a viable solution for nutrient-efficient and sustainable agriculture.

A validated marker system is crucial to running an effective genomics-assisted breeding program. We used 36 Kompetitive Allele-Specific PCR (KASP) markers to genotype 376 clones from the biofortified cassava pipeline, and fingerprinted 93 of these clones with DArTseq markers to characterize breeding materials and evaluate their relationships. The discriminating ability of the 36-quality control (QC) KASP and 6602 DArTseq markers was assessed using 92 clones genotyped in both assays. In addition, trait-specific markers were used to determine the presence or absence of target genomic regions. Hierarchical clustering identified two major groups, and the clusters were consistent with the breeding program origins. There was moderate genetic differentiation and a low degree of variation between the identified groups. The general structure of the population was similar using both assays. Nevertheless, KASP markers had poor resolution when it came to differentiating the genotypes by seed sources and overestimated the prevalence of duplicates. The trait-linked markers did not achieve optimal performance as all markers displayed variable levels of false positive and/or false negative. These findings represent the initial step in the application of genomics-assisted breeding for the biofortified cassava pipeline, and will guide the use of genomic selection in the future.

Selenium (Se) is a nutrient whose daily intake is often below the recommended levels in people. Biofortification with Se is a method to increase this intake by raising the Se concentration in tomato fruits, an effect dependent on sources and modes of application. Additionally, Se application can promote the enhancement of other compounds in tomato fruits, altering their metabolism, which may increase the fruit\'s shelf life. This study aimed to determine how different strategies of applying a multi-nutrient fertilizer containing Se (SeMNF) can increase the Se content and other bioactive compounds and enhance the shelf life of tomato (Solanum lycopersicum L.) fruits. Different foliar fertilization strategies involving the use of SeMNF were evaluated in field trials conducted on commercial tomato crops. Indeterminate-growth tomatoes were used, and different Se doses and application strategies were tested. Harvesting was conducted in three phases according to fruit ripening. Each harvested fruit was assessed for the Se content, macro and micronutrients, total phenolic compounds, vitamin C, antioxidant activity, carotenoids, pH, total titratable acidity, and total soluble solids in tomato fruits. Doses of 15 g ha-1 of Se, split into three applications, increased the Se content in the fruits at 1 and 2 harvests. The application of SeMNF at Se doses above 10 g of Se ha-1 increased firmness, days of ripening, and the nutritional quality of the tomatoes (higher contents of carotenoids (+39%), lycopene (+33%), antioxidant activity (+16%), total phenolic compounds (+38%), and vitamin C (+14%) in a dose-dependent effect of the application strategy used. These results contributed to an increase in the shelf life of tomatoes, consequently reducing food waste.

Iodine and selenium deficiencies are widespread both in developed countries and developing countries. The soil is the fundamental source of iodine and selenium for plants, and iodine and/or selenium-depleted soil restrains the cultivation of crops to cover recommended daily intakes of iodine and selenium. Although food fortification strategies, including salt iodization, increase the dietary intake of these minerals, their global deficiencies have not been eliminated. Therefore, new strategies have been developed to prevent iodine and selenium deficiencies, and biofortification is one of them. The aim of this review is to assert the outcomes of the studies that investigate the optimum conditions for biofortification with iodine and selenium and to recognize the role of biofortification practices as a potential solution for preventing iodine and selenium deficiencies. The findings of studies show that biofortification with iodine and selenium can be a solution for iodine and selenium deficiencies. Agronomic biofortification is currently a more convenient method to increase selenium and iodine contents in plants. However, the most effective agronomic biofortification conditions are crucial to acquire biofortified food. Moreover, increasing the awareness of the producers and consumers on biofortification has a determinative role in the achievement of biofortification practices for human health. Although research about iodine and selenium biofortification has been increased, the effectiveness of biofortified foods to meet recommended daily intakes is still unknown. More research is needed to understand most effective biofortification conditions for plants and bioavailability of biofortified foods for humans.

Biofortification of green leafy vegetables with pro-vitamin A carotenoids, such as β-carotene, has remained challenging to date. Here, we combined two strategies to achieve this goal. One of them involves producing β-carotene in the cytosol of leaf cells to avoid the negative impacts on photosynthesis derived from changing the balance of carotenoids and chlorophylls in chloroplasts. The second approach involves the conversion of chloroplasts into non-photosynthetic, carotenoid-overaccumulating chromoplasts in leaves agroinfiltrated or infected with constructs encoding the bacterial phytoene synthase crtB, leaving other non-engineered leaves of the plant to sustain normal growth. A combination of these two strategies, referred to as strategy C (for cytosolic production) and strategy P (for plastid conversion mediated by crtB), resulted in a 5-fold increase in the amount of β-carotene in Nicotiana benthamiana leaves. Following several attempts to further improve β-carotene leaf contents by metabolic engineering, hormone treatments and genetic screenings, it was found that promoting the proliferation of plastoglobules with increased light-intensity treatments not only improved β-carotene accumulation but it also resulted in a much higher bioaccessibility. The combination of strategies C and P together with a more intense light treatment increased the levels of accessible β-carotene 30-fold compared to controls. We further demonstrated that stimulating plastoglobule proliferation with strategy P, but also with a higher-light treatment alone, also improved β-carotene contents and bioaccessibility in edible lettuce (Lactuca sativa) leaves.

Biofortification of provitamin A in maize is an attractive and sustainable remedy to the problem of vitamin A deficiency in developing countries. The utilization of molecular markers represents a promising avenue to facilitate the development of provitamin A (PVA)-enriched maize varieties. We screened 752 diverse tropical yellow/orange maize lines using kompetitive allele-specific PCR (KASP) makers to validate the use of KASP markers in PVA maize breeding. To this end, a total of 161 yellow/orange inbred lines, selected from among the 752 lines, were evaluated for their endosperm PVA and other carotenoid compounds levels in two separate trials composed of 63 and 98 inbred lines in 2020 and 2021, respectively. Significant differences (p < 0.001) were observed among the yellow maize inbred lines studied for all carotenoid profiles. An inbred line TZMI1017, introduced by the International Institute of Tropical Agriculture (IITA) showed the highest level of PVA (12.99 µg/g) and β-carotene (12.08 µg/g). The molecular screening showed 43 yellow maize inbred lines carrying at least three of the favorable alleles of the KASP markers. TZMI1017 inbred line also carried the favorable alleles of almost all markers. In addition, nine locally developed inbred lines had medium to high PVA concentrations varying from 5.11 µg/g to 10.76 µg/g and harbored the favorable alleles of all the KASP PVA markers. Association analysis between molecular markers and PVA content variation in the yellow/orange maize inbred lines did not reveal a significant, predictable correlation. Further investigation is warranted to elucidate the underlying genetic architecture of the PVA content in this germplasm. However, we recommend strategic utilization of the maize-inbred lines with higher PVA content to enhance the PVA profile of the breeding program\'s germplasm.

Folate, a vital water-soluble vitamin (B9), requires specific attention as its recommended daily intake frequently is not reached in countries without mandatory fortification. In this regard, biofortification with microorganisms like Bifidobacterium and Streptococcus offers a compelling approach for enhancing food with natural folates. A randomized, nonblinded, and monocentric human pilot study is conducted to assess the bioavailability of a folate-biofortified fermented whey beverage, comprising 3 intervention days and a controlled replenishment phase before and during the assay. Folate plasma concentration (5-CH3-H4folate) is determined using a stable isotope dilution assay and LC-MS/MS detection. Biokinetic parameters (cmax and tmax) are determined, and areas under the curve (AUC) normalized to the basal folate plasma concentration are calculated. An average bioavailability of 17.1% in relation to the 5-CH3-H4folate supplement, ranging from 0% to 39.8%, is obtained. These results reiterate the significance of additional research into folate bioavailability in general and dairy products. Further investigations are warranted into folate-binding proteins (FBP) and other potential limiting factors within the food and individual factors. In summary, biofortification via fermentation emerges as a promising avenue for enhancing the natural folate content in dairy and other food products.

Micronutrient deficiencies (MNDs) particularly zinc (Zn) and iron (Fe) remain widespread in sub-Saharan Africa (SSA) due to low dietary intake. Wheat is an important source of energy globally, although cultivated wheat is inherently low in grain micronutrient concentrations. Malawian wheat/Am. muticum and Malawian wheat/T. urartu BC1F3 introgression lines, developed by crossing three Malawian wheat varieties (Kenya nyati, Nduna and Kadzibonga) with DH-348 (wheat/Am. muticum) and DH-254 (wheat/T. urartu), were phenotyped for grain Zn and Fe, and associated agronomic traits in Zn-deficient soils, in Malawi. 98% (47) of the BC1F3 introgression lines showed higher Zn above the checks Paragon, Chinese Spring, Kadzibonga, Kenya Nyati and Nduna. 23% (11) of the introgression lines showed a combination of high yields and an increase in grain Zn by 16-30 mg kg -1 above Nduna and Kadzibonga, and 11-25 mg kg -1 above Kenya nyati, Paragon and Chinese Spring. Among the 23%, 64% (7) also showed 8-12 mg kg -1 improvement in grain Fe compared to Nduna and Kenya nyati. Grain Zn concentrations showed a significant positive correlation with grain Fe, whilst grain Zn and Fe negatively and significantly correlated with TKW and grain yield. This work will contribute to the efforts of increasing mineral nutrient density in wheat, specifically targeting countries in the SSA.