scholarly journals Quantitative Proteomic Analysis for High- and Low-Aflatoxin-Yield Aspergillus flavus Strains Isolated From Natural Environments

2021 ◽  
Vol 12 ◽  
Author(s):  
Tao Li ◽  
Zhaowei Zhang ◽  
Yu Wang ◽  
Ying Li ◽  
Jiang Zhu ◽  
...  
Keyword(s):  
Aspergillus Flavus ◽  
Molecular Mechanisms ◽  
Multiple Reaction Monitoring ◽  
Aflatoxin Contamination ◽  
Field Conditions ◽  
Mass Spectrometry Analysis ◽  
High Yield ◽  
Effective Control ◽  
Natural Environments ◽  
Tandem Mass

The molecular mechanisms underlying aflatoxin production have been well-studied in strains of the fungus Aspergillus flavus (A. flavus) under artificial conditions. However, aflatoxin biosynthesis has rarely been studied in A. flavus strains isolated from field conditions with different aflatoxin-producing ability. In the present study, tandem mass tag (TMT) labeling and high-performance liquid chromatography (HPLC) coupled with tandem-mass spectrometry analysis were used for proteomic quantification in natural isolates of high- and low-aflatoxin-yield A. flavus strains. Additionally, findings obtained using the TMT-labeling method were validated using the high-resolution multiple reaction monitoring (MRM-HR) method. In total, 4,363 proteins were quantified, among which 1,045 proteins were differentially expressed between the high- and low-aflatoxin-yield A. flavus strains. Bioinformatics analysis showed that the up-regulated proteins were significantly enriched in carbon-related metabolism and the biosynthesis of secondary metabolites, whereas the down-regulated proteins were enriched in oxidative phosphorylation. Moreover, GST proteins were found to be significantly down-regulated in high-yield A. flavus strains; this result contradicted previous findings obtained from A. flavus strains grown under artificial conditions. In summary, our study provides novel insights into aflatoxin regulation in A. flavus under field conditions and could facilitate the development of various strategies for the effective control of aflatoxin contamination in food crops.

Letter: High-Performance Liquid Chromatography/Electrospray Mass Spectrometry Analysis of the Mycotoxin Aurofusarin

2008 ◽  
Vol 14 (5) ◽  
pp. 329-333 ◽  
Author(s):  
Torsten Neuhof ◽  
Robert Köppen ◽  
Matthias Koch ◽  
Irene Nehls
Keyword(s):  
Mass Spectrometry ◽  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
High Performance ◽  
Multiple Reaction Monitoring ◽  
Food Samples ◽  
Mass Spectrometry Analysis ◽  
Tandem Mass ◽  
Fragmentation Pattern ◽  
Spectrometry Analysis

High-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) can be used for simultaneous quantification of various mycotoxins in contaminated food samples. Therefore, multi-mycotoxin methods have been developed in the last couple of years. To enlarge these methods for further analytes, we have developed a LC-MS/MS method for the quantification of the mycotoxin aurofusarin. Additionally, further LC-MS n experiments were performed to demonstrate the fragmentation pattern of aurofusarin. Applicable multiple reaction monitoring (MRM) transitions of aurofusarin were found and optimized by parameter variation of the tandem mass spectrometer. The applicability of the developed method was tested by analysis of naturally contaminated wheat.

Comparative Susceptibility of Four Experimental Peanut Lines and the Cultivar Florunner to Preharvest Aflatoxin Contamination

1985 ◽  
Vol 12 (2) ◽  
pp. 70-72 ◽  
Author(s):  
P. D Blankenship ◽  
R. J Cole ◽  
T. H Sanders
Keyword(s):  
Aspergillus Flavus ◽  
Laboratory Method ◽  
Aflatoxin Contamination ◽  
Critical Assessment ◽  
Field Conditions ◽  
Temperature Conditions ◽  
Laboratory Screening ◽  
Comparative Susceptibility ◽  
Moisture Conditions

Abstract Four peanut genotypes, selected as resistant to invasion by Aspergillus flavus in laboratory screening with rehydrated, stored seed and Florunner cultivar were subjected to preharvest drought and temperature conditions conducive to A. flavus invasion and aflatoxin contamination. Preharvest aflatoxin contamination of peanuts has been previously correlated with geocarposphere temperature and moisture conditions during drought. All genotypes tested were highly contaminated with aflatoxin. This study indicates that a critical assessment should be made of the value of using the current laboratory method to select germplasm for resistance to A. flavus invasion and assuming resistance to aflatoxin contamination under field conditions.

scholarly journals Transcriptional Regulation of Aflatoxin Biosynthesis and Conidiation in Aspergillus flavus by Wickerhamomyces anomalus WRL-076 for Reduction of Aflatoxin Contamination

Toxins ◽  
2019 ◽  
Vol 11 (2) ◽  
pp. 81 ◽  
Author(s):  
Sui Hua ◽  
Siov Sarreal ◽  
Perng-Kuang Chang ◽  
Jiujiang Yu
Keyword(s):  
Aspergillus Flavus ◽  
Molecular Mechanisms ◽  
Field Experiments ◽  
Regulatory Gene ◽  
Aflatoxin Contamination ◽  
Economic Losses ◽  
Expression Levels ◽  
Wickerhamomyces Anomalus ◽  
Cultural Medium ◽  
Major Producer

Aspergillus flavus is a ubiquitous saprophytic fungus found in soils across the world. The fungus is the major producer of aflatoxin (AF) B1, which is toxic and a potent carcinogen to humans. Aflatoxin B1 (AFB1) is often detected in agricultural crops such as corn, peanut, almond, and pistachio. It is a serious and recurrent problem and causes substantial economic losses. Wickerhamomyces anomalus WRL-076 was identified as an effective biocontrol yeast against A. flavus. In this study, the associated molecular mechanisms of biocontrol were investigated. We found that the expression levels of eight genes, aflR, aflJ, norA, omtA, omtB, pksA, vbs, and ver-1 in the aflatoxin biosynthetic pathway cluster were suppressed. The decreases ranged from several to 10,000 fold in fungal samples co-cultured with W. anomalus. Expression levels of conidiation regulatory genes brlA, abaA, and wetA as well as sclerotial regulatory gene (sclR) were all down regulated. Consistent with the decreased gene expression levels, aflatoxin concentrations in cultural medium were reduced to barely detectable. Furthermore, fungal biomass and conidial number were significantly reduced by 60% and more than 95%, respectively. The results validate the biocontrol efficacy of W. anomalus WRL-076 observed in the field experiments.

scholarly journals Transcriptome Analysis Identified Coordinated Control of Key Pathways Regulating Cellular Physiology and Metabolism upon Aspergillus flavus Infection Resulting in Reduced Aflatoxin Production in Groundnut

2020 ◽  
Vol 6 (4) ◽  
pp. 370
Author(s):  
Pooja Soni ◽  
Spurthi N. Nayak ◽  
Rakesh Kumar ◽  
Manish K. Pandey ◽  
Namita Singh ◽  
...  
Keyword(s):  
Aspergillus Flavus ◽  
Molecular Mechanisms ◽  
Coordinated Control ◽  
Aflatoxin Production ◽  
Aflatoxin Contamination ◽  
Rna Seq ◽  
Resistant Genotype ◽  
Cellular Physiology ◽  
Global Health Issue ◽  
Key Pathways

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. A total 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.

scholarly journals Transcriptomic responses of Aspergillus flavus to temperature and oxidative stresses during aflatoxin production

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Fei Tian ◽  
Sang Yoo Lee ◽  
So Young Woo ◽  
Hwa Young Choi ◽  
Seongeun Heo ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Aspergillus Flavus ◽  
Antioxidant Properties ◽  
Culture Method ◽  
Aflatoxin Production ◽  
Aflatoxin Contamination ◽  
Stress Conditions ◽  
Effective Control ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Transcriptomic Response

AbstractAflatoxin is a group of polyketide-derived carcinogenic and mutagenic secondary metabolites produced by Aspergillus flavus that negatively impact global food security and threaten the health of both humans and livestock. Aflatoxin biosynthesis is strongly affected by the fungal developmental stage, cultivation conditions, and environmental stress. In this study, a novel float culture method was used to examine the direct responses of the A. flavus transcriptome to temperature stress, oxidative stress, and their dual effects during the aflatoxin production stage. The transcriptomic response of A. flavus illustrated that the co-regulation of different secondary metabolic pathways likely contributes to maintaining cellular homeostasis and promoting cell survival under stress conditions. In particular, aflatoxin biosynthetic gene expression was downregulated, while genes encoding secondary metabolites with antioxidant properties, such as kojic acid and imizoquins, were upregulated under stress conditions. Multiple mitochondrial function-related genes, including those encoding NADH:ubiquinone oxidoreductase, ubiquinol-cytochrome C reductase, and alternative oxidase, were differentially expressed. These data can provide insights into the important mechanisms through which secondary metabolism in A. flavus is co-regulated and facilitate the deployment of various approaches for the effective control and prevention of aflatoxin contamination in food crops.

scholarly journals The Phosphoproteomic Response of Okra (Abelmoschus esculentus L.) Seedlings to Salt Stress

2019 ◽  
Vol 20 (6) ◽  
pp. 1262 ◽  
Author(s):  
Chenliang Yu ◽  
Qinqfei Wu ◽  
Chendong Sun ◽  
Mengling Tang ◽  
Junwei Sun ◽  
...  
Keyword(s):  
Salt Stress ◽  
Stress Responses ◽  
Molecular Mechanisms ◽  
Crop Yields ◽  
Differentially Expressed ◽  
Abelmoschus Esculentus ◽  
Mass Spectrometry Analysis ◽  
Tandem Mass ◽  
Differentially Expressed Proteins ◽  
Affinity Enrichment

Soil salinization is a major environmental stresses that seriously threatens land use efficiency and crop yields worldwide. Although the overall response of plants to NaCl has been well studied, the contribution of protein phosphorylation to the detoxification and tolerance of NaCl in okra (Abelmoschus esculentus L.) seedlings is unclear. The molecular bases of okra seedlings’ responses to 300 mM NaCl stress are discussed in this study. Using a combination of affinity enrichment, tandem mass tag (TMT) labeling and high-performance liquid chromatography–tandem mass spectrometry analysis, a large-scale phosphoproteome analysis was performed in okra. A total of 4341 phosphorylation sites were identified on 2550 proteins, of which 3453 sites of 2268 proteins provided quantitative information. We found that 91 sites were upregulated and 307 sites were downregulated in the NaCl/control comparison group. Subsequently, we performed a systematic bioinformatics analysis including gene ontology annotation, domain annotation, subcellular localization, and Kyoto Encyclopedia of Genes and Genomes pathway annotation. The latter revealed that the differentially expressed proteins were most strongly associated with ‘photosynthesis antenna proteins’ and ‘RNA degradation’. These differentially expressed proteins probably play important roles in salt stress responses in okra. The results should help to increase our understanding of the molecular mechanisms of plant post-translational modifications in response to salt stress.

scholarly journals Flavonoids Modulate the Accumulation of Toxins From Aspergillus flavus in Maize Kernels

2021 ◽  
Vol 12 ◽  
Author(s):  
Lina Castano-Duque ◽  
Matthew K. Gilbert ◽  
Brian M. Mack ◽  
Matthew D. Lebar ◽  
Carol H. Carter-Wientjes ◽  
...  
Keyword(s):  
Aspergillus Flavus ◽  
Molecular Mechanisms ◽  
Fungal Infections ◽  
Defense Responses ◽  
Aflatoxin Contamination ◽  
Resistant Line ◽  
Fungal Treatment ◽  
Susceptible Line ◽  
A Genome ◽  
Maize Kernels

Aspergillus flavus is an opportunistic fungal pathogen capable of producing aflatoxins, potent carcinogenic toxins that accumulate in maize kernels after infection. To better understand the molecular mechanisms of maize resistance to A. flavus growth and aflatoxin accumulation, we performed a high-throughput transcriptomic study in situ using maize kernels infected with A. flavus strain 3357. Three maize lines were evaluated: aflatoxin-contamination resistant line TZAR102, semi-resistant MI82, and susceptible line Va35. A modified genotype-environment association method (GEA) used to detect loci under selection via redundancy analysis (RDA) was used with the transcriptomic data to detect genes significantly influenced by maize line, fungal treatment, and duration of infection. Gene ontology enrichment analysis of genes highly expressed in infected kernels identified molecular pathways associated with defense responses to fungi and other microbes such as production of pathogenesis-related (PR) proteins and lipid bilayer formation. To further identify novel genes of interest, we incorporated genomic and phenotypic field data from a genome wide association analysis with gene expression data, allowing us to detect significantly expressed quantitative trait loci (eQTL). These results identified significant association between flavonoid biosynthetic pathway genes and infection by A. flavus. In planta fungal infections showed that the resistant line, TZAR102, has a higher fold increase of the metabolites naringenin and luteolin than the susceptible line, Va35, when comparing untreated and fungal infected plants. These results suggest flavonoids contribute to plant resistance mechanisms against aflatoxin contamination through modulation of toxin accumulation in maize kernels.

A novel and sensitive LC-MS/MS method for the quantitation of ceftiofur in pharmaceutical preparations and milk samples

2020 ◽  
Vol 23 ◽  
Author(s):  
Serkan Levent ◽  
Saniye Özcan ◽  
Aysun Geven ◽  
Nafiz Öncü Can
Keyword(s):  
Quantitative Estimation ◽  
Signal To Noise Ratio ◽  
Multiple Reaction Monitoring ◽  
Pharmaceutical Preparations ◽  
Calibration Method ◽  
Negative Ion ◽  
Cow Milk ◽  
Tandem Mass ◽  
Ionization Source ◽  
Liquid Chromatography Tandem Mass

Introduction:: In the present study, a sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was described for the determination of ceftiofur (CEF) in cow milk and pharmaceutical preparations. CEF is an antibiotic compound, which is commonly used in the treatment of animal diseases such as respiratory system, soft tissue, and foot infections, as well as postpartum acute puerperal metritis. One of the critical features of CEF is its prescription while breastfeeding of cows; in accordance, its quantitative estimation is essential to assess its residual amounts. Methods:: In the method reported herein, after simple protein precipitation using acetonitrile, the pre-treated samples were introduced in to an LC-MS/MS instrument equipped with a Chromolith® High-Resolution RP-18 series HPLC column (100 mm × 4.6 mm from Merck KGaA, Germany). Electrospray ionization was employed as the ionization source in the triplequadrupole tandem mass spectrometer. Results:: For the calibration method using solvent-based standards; LOQ was 3.038 ng/mL, 12.15 ng/mL, and LOD was 1.215 ng/mL and 6.076 ng/mL for ESI+ and ESI- modes, respectively. On the other hand, for the method of matrix-matched standards; LOQ was 1.701 ng/mL, 10.13 ng/mL, and LOD was 0.486 ng/mL and 5.929 ng/mL for ESI+ and ESI- modes, respectively as obtained from signal to noise ratio. Conclusion:: Applicability of both positive and negative ion modes was tested, and the analyte was detected via multiple reaction monitoring. The distorting effects of the milk matrix on the MS ionization and quantitation of CEF were overcome by using matrix-matched calibration for the first time.

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