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Introduction: Peanut is susceptible to infection of Aspergillus fungi and conducive to aflatoxin contamination, hence developing aflatoxin-resistant variety is highly meaningful. Identifying functional genes or loci conferring aflatoxin resistance and molecular diagnostic marker are crucial for peanut breeding., Objectives: This work aims to (1) identify candidate gene for aflatoxin production resistance, (2) reveal the related resistance mechanism, and (3) develop diagnostic marker for resistance breeding program., Methods: Resistance to aflatoxin production in a recombined inbred line (RIL) population derived from a high-yielding variety Xuhua13 crossed with an aflatoxin-resistant genotype Zhonghua 6 was evaluated under artificial inoculation for three consecutive years. Both genetic linkage analysis and QTL-seq were conducted for QTL mapping. The candidate gene was further fine-mapped using a secondary segregation mapping population and validated by transgenic experiments. RNA-Seq analysis among resistant and susceptible RILs was used to reveal the resistance pathway for the candidate genes., Results: The major effect QTL qAFTRA07.1 for aflatoxin production resistance was mapped to a 1.98 Mbp interval. A gene, AhAftr1 (Arachis hypogaea Aflatoxin resistance 1), was detected structure variation (SV) in leucine rich repeat (LRR) domain of its production, and involved in disease resistance response through the effector-triggered immunity (ETI) pathway. Transgenic plants with overexpression of AhAftr1 (ZH6) exhibited 57.3% aflatoxin reduction compared to that of AhAftr1 (XH13) . A molecular diagnostic marker AFTR.Del.A07 was developed based on the SV. Thirty-six lines, with aflatoxin content decrease by over 77.67% compared to the susceptible control Zhonghua12 (ZH12), were identified from a panel of peanut germplasm accessions and breeding lines through using AFTR.Del.A07., Conclusion: Our findings would provide insights of aflatoxin production resistance mechanisms and laid meaningful foundation for further breeding programs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Production and hosting by Elsevier B.V.)

Background: Aflatoxin contamination caused by Aspergillus fungi has been a serious factor affecting food safety of peanut (Arachis hypogaea L.) because aflatoxins are highly harmful for human and animal health. As three mechanisms of resistance to aflatoxin in peanut including shell infection resistance, seed infection resistance and aflatoxin production resistance exist among naturally evolved germplasm stocks, it is highly crucial to pyramid these three resistances for promoting peanut industry development and protecting consumers' health. However, less research effort has been made yet to investigate the differentiation and genetic relationship among the three resistances in diversified peanut germplasm collections., Results: In this study, the Chinese peanut mini-mini core collection selected from a large basic collection was systematically evaluated for the three resistances against A. flavus for the first time. The research revealed a wide variation among the diversified peanut accessions for all the three resistances. Totally, 14 resistant accessions were identified, including three with shell infection resistance, seven with seed infection resistance and five with aflatoxin production resistance. A special accession, Zh.h1312, was identified with both seed infection and aflatoxin production resistance. Among the five botanic types of A. hypogaea, the var. vulgaris (Spanish type) belonging to subspecies fastigiata is the only one which possessed all the three resistances. There was no close correlation between shell infection resistance and other two resistances, while there was a significant positive correlation between seed infection and toxin production resistance. All the three resistances had a significant negative correlation with pod or seed size. A total of 16 SNPs/InDels associated with the three resistances were identified through genome-wide association study (GWAS). Through comparative analysis, Zh.h1312 with seed infection resistance and aflatoxin production resistance was also revealed to possess all the resistance alleles of associated loci for seed infection index and aflatoxin content., Conclusions: This study provided the first comprehensive understanding of differentiation of aflatoxin resistance in diversified peanut germplasm collection, and would further contribute to the genetic enhancement for resistance to aflatoxin contamination., (© 2022. The Author(s).)

Aflatoxin B 1 (AFB 1 ) and aflatoxin B 2 (AFB 2 ) are the most common aflatoxins produced by Aspergillus flavus in peanuts, with high carcinogenicity and teratogenicity. Identification of DNA markers associated with resistance to aflatoxin production is likely to offer breeders efficient tools to develop resistant cultivars through molecular breeding. In this study, seeds of 99 accessions of a Chinese peanut mini-mini core collection were investigated for their reaction to aflatoxin production by a laboratory kernel inoculation assay. Two resistant accessions (Zh.h0551 and Zh.h2150) were identified, with their aflatoxin content being 8.11%-18.90% of the susceptible control. The 99 peanut accessions were also genotyped by restriction site-associated DNA sequencing (RAD-Seq) for a genome-wide association study (GWAS). A total of 60 SNP (single nucleotide polymorphism) markers associated with aflatoxin production were detected, and they explained 16.87%-31.70% of phenotypic variation (PVE), with SNP02686 and SNP19994 possessing 31.70% and 28.91% PVE, respectively. Aflatoxin contents of accessions with "AG" (existed in Zh.h0551 and Zh.h2150) and "GG" genotypes of either SNP19994 or SNP02686 were significantly lower than that of "AA" genotypes in the mean value of a three-year assay. The resistant accessions and molecular markers identified in this study are likely to be helpful for deployment in aflatoxin resistance breeding in peanuts.

Aflatoxin is considered a "hidden poison" due to its slow and adverse effect on various biological pathways in humans, particularly among children, in whom it leads to delayed development, stunted growth, liver damage, and liver cancer. Unfortunately, the unpredictable behavior of the fungus as well as climatic conditions pose serious challenges in precise phenotyping, genetic prediction and genetic improvement, leaving the complete onus of preventing aflatoxin contamination in crops on post-harvest management. Equipping popular crop varieties with genetic resistance to aflatoxin is key to effective lowering of infection in farmer's fields. A combination of genetic resistance for in vitro seed colonization (IVSC), pre-harvest aflatoxin contamination (PAC) and aflatoxin production together with pre- and post-harvest management may provide a sustainable solution to aflatoxin contamination. In this context, modern "omics" approaches, including next-generation genomics technologies, can provide improved and decisive information and genetic solutions. Preventing contamination will not only drastically boost the consumption and trade of the crops and products across nations/regions, but more importantly, stave off deleterious health problems among consumers across the globe.

Background: Aflatoxin contamination caused by Aspergillus flavus is a major constraint to peanut industry worldwide due to its toxicological effects to human and animals. Developing peanut varieties with resistance to seed infection and/or aflatoxin accumulation is the most effective and economic strategy for reducing aflatoxin risk in food chain. Breeding for resistance to aflatoxin in peanut is a challenging task for breeders because the genetic basis is still poorly understood. To identify the quantitative trait loci (QTLs) for resistance to aflatoxin contamination in peanut, a recombinant inbred line (RIL) population was developed from crossing Zhonghua 10 (susceptible) with ICG 12625 (resistant). The percent seed infection index (PSII), the contents of aflatoxin B 1 (AFB 1 ) and aflatoxin B 2 (AFB 2 ) of RILs were evaluated by a laboratory kernel inoculation assay., Results: Two QTLs were identified for PSII including one major QTL with 11.32-13.00% phenotypic variance explained (PVE). A total of 12 QTLs for aflatoxin accumulation were detected by unconditional analysis, and four of them (qAFB1A07 and qAFB1B06.1 for AFB 1 , qAFB2A07 and qAFB2B06 for AFB 2 ) exhibited major and stable effects across multiple environments with 9.32-21.02% PVE. Furthermore, not only qAFB1A07 and qAFB2A07 were co-localized in the same genetic interval on LG A07, but qAFB1B06.1 was also co-localized with qAFB2B06 on LG B06. Conditional QTL mapping also confirmed that there was a strong interaction between resistance to AFB 1 and AFB 2 accumulation. Genotyping of RILs revealed that qAFB1A07 and qAFB1B06.1 interacted additively to improve the resistance to both AFB 1 and AFB 2 accumulation. Additionally, validation of the two markers was performed in diversified germplasm collection and four accessions with resistance to aflatoxin accumulation were identified., Conclusions: Single major QTL for resistance to PSII and two important co-localized intervals associated with major QTLs for resistance to AFB 1 and AFB 2 . Combination of these intervals could improve the resistance to aflatoxin accumulation in peanut. SSR markers linked to these intervals were identified and validated. The identified QTLs and associated markers exhibit potential to be applied in improvement of resistance to aflatoxin contamination.

Aflatoxin contamination in peanuts poses major challenges for vulnerable populations of sub-Saharan Africa and South Asia. Developing peanut varieties to combat preharvest Aspergillus flavus infection and resulting aflatoxin contamination has thus far remained a major challenge, confounded by highly complex peanut-Aspergilli pathosystem. Our study reports achieving a high level of resistance in peanut by overexpressing (OE) antifungal plant defensins MsDef1 and MtDef4.2, and through host-induced gene silencing (HIGS) of aflM and aflP genes from the aflatoxin biosynthetic pathway. While the former improves genetic resistance to A. flavus infection, the latter inhibits aflatoxin production in the event of infection providing durable resistance against different Aspergillus flavus morphotypes and negligible aflatoxin content in several peanut events/lines well. A strong positive correlation was observed between aflatoxin accumulation and decline in transcription of the aflatoxin biosynthetic pathway genes in both OE-Def and HIGS lines. Transcriptomic signatures in the resistant lines revealed key mechanisms such as regulation of aflatoxin synthesis, its packaging and export control, besides the role of reactive oxygen species-scavenging enzymes that render enhanced protection in the OE and HIGS lines. This is the first study to demonstrate highly effective biotechnological strategies for successfully generating peanuts that are near-immune to aflatoxin contamination, offering a panacea for serious food safety, health and trade issues in the semi-arid regions., (© 2017 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Aflatoxin contamination caused by Aspergillus flavus significantly threatens food safety and human health. Resistance to aflatoxin is a highly complex and quantitative trait, but the underlying molecular and biochemical mechanisms are poorly understood. The present study aims to identify the resistance‐related metabolites in groundnut that influence the defense mechanism against aflatoxin. Here, metabolite profiling of resistant (55–437) and susceptible (TMV‐2) groundnut genotypes, which exhibited contrasting levels of resistance to A. flavus growth and aflatoxin accumulation under pathogen‐ or mock‐inoculated treatments, was undertaken using liquid chromatography and high‐resolution mass spectrometry (LC‐HRMS). Non‐targeted metabolomic analysis revealed key resistance‐related metabolites belonging to phenylpropanoids, flavonoids, fatty acids, alkaloids, and terpenoid biosynthetic pathways. The phenylpropanoids ‐ hydroxycinnamic acid amides (HCAAs) and lignins were among the most abundantly accumulated metabolites in the resistant genotype compared to the susceptible genotype. HCAAs and lignins are deposited as polymers and conjugated metabolites to strengthen the secondary cell wall, which acts as a barrier to pathogen entry. Further, histochemical staining confirmed the secondary cell wall thickening due to HCAAs and lignin depositions. Quantitative real‐time PCR studies revealed higher expressions of phenylalanine ammonia‐lyase (PAL), 4‐coumarate: CoA ligase (4CL), cinnamoyl CoA reductase (CCR2), cinnamoyl alcohol dehydrogenase (CAD1), agmatine hydroxycinnamoyl transferase (ACT), chalcone synthase (CHS), dihydroflavonol 4‐reductase (DFR) and flavonol synthase (FLS) in the pathogen‐inoculated resistant genotype than in the susceptible genotype. This study reveals that the resistance to aflatoxin contamination in groundnut genotypes is associated with secondary cell wall thickening due to the deposition of HCAAs and lignins. [ABSTRACT FROM AUTHOR]

Aflatoxin-affected groundnut or peanut presents a major global health issue to both commercial and subsistence farming. Therefore, understanding the genetic and molecular mechanisms associated with resistance to aflatoxin production during host--pathogen interactions is crucial for breeding groundnut cultivars with minimal level of aflatoxin contamination. Here, we performed gene expression profiling to better understand the mechanisms involved in reduction and prevention of aflatoxin contamination resulting from Aspergillus flavus infection in groundnut seeds. RNA sequencing (RNA-Seq) of 16 samples from different time points during infection (24 h, 48 h, 72 h and the 7th day after inoculation) in U 4-7-5 (resistant) and JL 24 (susceptible) genotypes yielded 840.5 million raw reads with an average of 52.5 million reads per sample. Atotal of 1779 unique differentially expressed genes (DEGs) were identified. Furthermore, comprehensive analysis revealed several pathways, such as disease resistance, hormone biosynthetic signaling, flavonoid biosynthesis, reactive oxygen species (ROS) detoxifying, cell wall metabolism and catabolizing and seed germination. We also detected several highly upregulated transcription factors, such as ARF, DBB, MYB, NAC and C2H2 in the resistant genotype in comparison to the susceptible genotype after inoculation. Moreover, RNA-Seq analysis suggested the occurrence of coordinated control of key pathways controlling cellular physiology and metabolism upon A. flavus infection, resulting in reduced aflatoxin production. [ABSTRACT FROM AUTHOR]

Aflatoxin contamination in groundnut is an important qualitative issue posing a threat to food safety. In our present study, we have demonstrated the efficacy of certain good agricultural practices (GAPs) in groundnut, such as farmyard manure (5 t/ha), gypsum (500 kg/ha), a protective irrigation at 90 days after sowing (DAS), drying of pods on tarpaulins after harvest in farmers' fields. During 2013--2015, 89 on-farm demonstrations were conducted advocating GAPs, and compared with farmers' practices (FP) plots. Farmers' awareness of GAPs, and knowledge on important aspects of groundnut cultivation, were also assessed during our experimentation in the selected villages under study. Pre-harvest kernel infection by Aspergillus flavus, aflatoxin contamination, and pod yields were compared in GAPs plots, vis-à-vis FP plots. The cost of cultivation in both the plots was calculated and compared, based on farmer's opinion surveys. Results indicate kernel infections and aflatoxins were significantly lower, with 13-58% and 62-94% reduction, respectively, in GAPs plots over FP. Further, a net gain of around $23 per acre was realized through adoption of GAPs by farmers besides quality improvement of groundnuts. Based on our results, it can be concluded that on-farm demonstrations were the best educative tool to convince the farmers about the cost-effectiveness, and adoptability of aflatoxin management technologies. [ABSTRACT FROM AUTHOR]

Aflatoxin contamination is a global menace that adversely affects food crops and human health. Peanut seed coat is the outer layer protecting the cotyledon both at pre- and post-harvest stages from biotic and abiotic stresses. The aim of the present study is to investigate the role of seed coat against A. flavus infection. In-vitro seed colonization (IVSC) with and without seed coat showed that the seed coat acts as a physical barrier, and the developmental series of peanut seed coat showed the formation of a robust multilayered protective seed coat. Radial growth bioassay revealed that both insoluble and soluble seed coat extracts from 55-437 line (resistant) showed higher A. flavus inhibition compared to TMV-2 line (susceptible). Further analysis of seed coat biochemicals showed that hydroxycinnamic and hydroxybenzoic acid derivatives are the predominant phenolic compounds, and addition of these compounds to the media inhibited A. flavus growth. Gene expression analysis showed that genes involved in lignin monomer, proanthocyanidin, and flavonoid biosynthesis are highly abundant in 55-437 compared to TMV-2 seed coats. Overall, the present study showed that the seed coat acts as a physical and biochemical barrier against A. flavus infection and its potential use in mitigating the aflatoxin contamination. [ABSTRACT FROM AUTHOR]

The spatiotemporal trends in aflatoxin B1 (AFB1), fumonisin B1 (FB1), and deoxynivalenol (DON) accumulation were analyzed in a range of food commodities (maize, groundnut, pearl millet, rice, and wheat) in village settings in Unnao, Uttar Pradesh, India. Samples (n = 1549) were collected across six communities and six time points spanning a calendar year and were analyzed for mycotoxins using enzyme-linked immunosorbent assays. AFB1 and FB1 were common across surveyed villages, with moderate to high detection rates (45–75%) observed across commodities. AFB1 levels in maize and groundnuts and FB1 levels in maize and pearl millet frequently exceeded regulatory threshold levels of 15 μg/kg (AFB1) and 2 μg/g (FB1). DON was analyzed in wheat, with 3% of samples yielding detectable levels and none exceeding 1 μg/g. In rice, AFB1 levels were highest in the bran and husk and lower in the kernel. Commodity type significantly influenced AFB1 detection status, while commodity type, season, and visual quality influenced samples' legal status. Storage characteristics and household socioeconomic status indicators did not have significant effects on contamination. No significant effects of any variables on FB1 detection or legal status were observed. Data on mycotoxin contamination, combined with data on local dietary intake, were used to estimate spatiotemporal mycotoxin exposure profiles. Estimated seasonal per capita exposure levels for AFB1 (5.4–39.3 ng/kg body weight/day) and FB1 (~0–2.4 μg/kg body weight/day) exceeded provisional maximum tolerable daily intake levels (1 ng/kg body weight/day for AFB1 and 2 μg/kg body weight/day for FB1) in some seasons and locations. This study demonstrates substantial dietary mycotoxin exposure risk in Unnao food systems and serves as an evidentiary foundation for participatory food safety intervention in the region. • Aflatoxin B1 and fumonisin B1 are prevalent in the food systems, while deoxynivalenol is largely absent. • Mycotoxin contamination in Indian villages has distinct spatial and seasonal trends. • Food system composition and cropping calendars influence the distribution of mycotoxins. • Trends in contamination are associated with dietary intakes at levels that warrant public health intervention. [ABSTRACT FROM AUTHOR]