Publications by authors named "Deepak Bhatnagar"

120 Publications

The COVID-19 pandemic: lifestyle and cardiovascular risk factors.

Authors:
Deepak Bhatnagar

Curr Opin Lipidol 2021 02;32(1):71-73

Division of Diabetes, Endocrinology and Gastroenterology, University of Manchester, Manchester, UK.

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http://dx.doi.org/10.1097/MOL.0000000000000725DOI Listing
February 2021

Characterization of morphological changes within stromata during sexual reproduction in .

Mycologia 2020 Sep-Oct;112(5):908-920. Epub 2020 Aug 21.

Center for Integrated Fungal Research, Department of Entomology and Plant Pathology, North Carolina State University , Raleigh, NC 27695.

contaminates agricultural products worldwide with carcinogenic aflatoxins that pose a serious health risk to humans and animals. The fungus survives adverse environmental conditions through production of sclerotia. When fertilized by a compatible conidium of an opposite mating type, a sclerotium transforms into a stroma within which ascocarps, asci, and ascospores are formed. However, the transition from a sclerotium to a stroma during sexual reproduction in is not well understood. Early events during the interaction between sexually compatible strains of were visualized using conidia of a green fluorescent protein (GFP)-labeled strain and sclerotia of an mCherry-labeled strain. Both conidia and sclerotia of transformed strains germinated to produce hyphae within 24 h of incubation. Hyphal growth of these two strains produced what appeared to be a network of interlocking hyphal strands that were observed at the base of the mCherry-labeled sclerotia (i.e., region in contact with agar surface) after 72 h of incubation. At 5 wk following incubation, intracellular green-fluorescent hyphal strands were observed within the stromatal matrix of the mCherry-labeled strain. Scanning electron microscopy of stromata from a high- and low-fertility cross and unmated sclerotia was used to visualize the formation and development of sexual structures within the stromatal and sclerotial matrices, starting at the time of crossing and thereafter every 2 wk until 8 wk of incubation. Morphological differences between sclerotia and stromata became apparent at 4 wk of incubation. Internal hyphae and croziers were detected inside multiple ascocarps that developed within the stromatal matrix of the high-fertility cross but were not detected in the matrix of the low-fertility cross or the unmated sclerotia. At 6 to 8 wk of incubation, hyphal tips produced numerous asci, each containing one to eight ascospores that emerged out of an ascus following the breakdown of the ascus wall. These observations broaden our knowledge of early events during sexual reproduction and suggest that hyphae from the conidium-producing strain may be involved in the early stages of sexual reproduction in . When combined with omics data, these findings could be useful in further exploration of the molecular and biochemical mechanisms underlying sexual reproduction in .
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http://dx.doi.org/10.1080/00275514.2020.1800361DOI Listing
August 2020

Two New Reference Genomes Reveal a Large Insertion Potentially Contributing to Isolate Stress Tolerance and Aflatoxin Production.

G3 (Bethesda) 2020 Oct 5;10(10):3515-3531. Epub 2020 Oct 5.

USDA-ARS, Crop Protection and Management Research Unit, Tifton, GA 31793

Efforts in genome sequencing in the genus have led to the development of quality reference genomes for several important species including , , and However, less progress has been made for As part of the effort of the USDA-ARS Annual Aflatoxin Workshop Fungal Genome Project, the isolate NRRL3357 was sequenced and resulted in a scaffold-level genome released in 2005. Our goal has been biologically driven, focusing on two areas: isolate variation in aflatoxin production and drought stress exacerbating aflatoxin production by Therefore, we developed two reference pseudomolecule genome assemblies derived from chromosome arms for two isolates: AF13, a MAT1-2, highly stress tolerant, and highly aflatoxigenic isolate; and NRRL3357, a MAT1-1, less stress tolerant, and moderate aflatoxin producer in comparison to AF13. Here, we report these two reference-grade assemblies for these isolates through a combination of PacBio long-read sequencing and optical mapping, and coupled them with comparative, functional, and phylogenetic analyses. This analysis resulted in the identification of 153 and 45 unique genes in AF13 and NRRL3357, respectively. We also confirmed the presence of a unique 310 Kb insertion in AF13 containing 60 genes. Analysis of this insertion revealed the presence of a bZIP transcription factor, named , which may contribute to isolate pathogenicity and stress tolerance. Phylogenomic analyses comparing these and other available assemblies also suggest that the species complex of is polyphyletic.
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http://dx.doi.org/10.1534/g3.120.401405DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7534430PMC
October 2020

Use of Dual RNA-seq for Systems Biology Analysis of and Interaction.

Front Microbiol 2020 3;11:853. Epub 2020 Jun 3.

Department of Plant Biology, Southern Illinois University, Carbondale, IL, United States.

The interaction between and is complex, and the identification of plant genes and pathways conferring resistance to the fungus has been challenging. Therefore, the authors undertook a systems biology approach involving dual RNA-seq to determine the simultaneous response from the host and the pathogen. What was dramatically highlighted in the analysis is the uniformity in the development patterns of gene expression of the host and the pathogen during infection. This led to the development of a "stage of infection index" that was subsequently used to categorize the samples before down-stream system biology analysis. Additionally, we were able to ascertain that key maize genes in pathways such as the jasmonate, ethylene and ROS pathways, were up-regulated in the study. The stage of infection index used for the transcriptomic analysis revealed that produces a relatively limited number of transcripts during the early stages (0 to 12 h) of infection. At later stages, in , transcripts and pathways involved in endosomal transport, aflatoxin production, and carbohydrate metabolism were up-regulated. Multiple WRKY genes targeting the activation of the resistance pathways (i.e., jasmonate, phenylpropanoid, and ethylene) were detected using causal inference analysis. This analysis also revealed, for the first time, the activation of resistance genes influencing the expression of specific genes. Our results show that seems to be reacting to a hostile environment resulting from the activation of resistance pathways in . This study revealed the dynamic nature of the interaction between the two organisms.
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http://dx.doi.org/10.3389/fmicb.2020.00853DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7285840PMC
June 2020

Spectral-Based Screening Approach Evaluating Two Specific Maize Lines With Divergent Resistance to Invasion by Aflatoxigenic Fungi.

Front Microbiol 2019 22;10:3152. Epub 2020 Jan 22.

Southern Regional Research Center, USDA-ARS, New Orleans, LA, United States.

In an effort to control aflatoxin contamination in food and/or feed grains, a segment of research has focused on host resistance to eliminate aflatoxin from susceptible crops, including maize. To this end, screening tools are key to identifying resistant maize genotypes. The traditional field screening techniques, the kernel screening laboratory assay (KSA), and analytical methods (e.g., ELISA) used for evaluating corn lines for resistance to fungal invasion, all ultimately require sample destruction. A technological advancement on the basic BGYF presumptive screening test, fluorescence hyperspectral imaging offers an option for non-destructive and rapid image-based screening. The present study aimed to differentiate fluorescence spectral signatures of representative resistant and susceptible corn hybrids infected by a toxigenic (SRRC-AF13) and an atoxigenic (SRRC-AF36) strain of , at several time points (5, 7, 10, and 14 days), in order to evaluate fluorescence hyperspectral imaging as a viable technique for early, non-invasive aflatoxin screening in resistant and susceptible corn lines. The study utilized the KSA to promote fungal growth and aflatoxin production in corn kernels inoculated under laboratory conditions and to provide actual aflatoxin values to relate with the imaging data. Each time point consisted of 78 kernels divided into four groups (30-susceptible, 30-resistant, 9-susceptible control, and 9-resistant control), per inoculum. On specified days, kernels were removed from the incubator and dried at 60°C to terminate fungal growth. Dry kernels were imaged with a VNIR hyperspectral sensor (image spectral range of 400-1000 nm), under UV excitation centered at 365 nm. Following imaging, kernels were submitted for the chemical AflaTest assay (VICAM). Fluorescence emissions were compared for all samples over 14 days. Analysis of strain differences separating the fluorescence emission peaks of resistant from the susceptible strain indicated that the emission peaks of the resistant strain and the susceptible strains differed significantly ( < 0.01) from each other, and there was a significant difference in fluorescence intensity between the treated and control kernels of both strains. These results indicate a viable role of fluorescence hyperspectral imaging for non-invasive screening of maize lines with divergent resistance to invasion by aflatoxigenic fungi.
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http://dx.doi.org/10.3389/fmicb.2019.03152DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6988685PMC
January 2020

A Rapid and Nondestructive Method for Simultaneous Determination of Aflatoxigenic Fungus and Aflatoxin Contamination on Corn Kernels.

J Agric Food Chem 2019 May 23;67(18):5230-5239. Epub 2019 Apr 23.

Southern Regional Research Center , USDA-ARS , New Orleans , Louisiana 70124 , United States.

Conventional methods for detecting aflatoxigenic fungus and aflatoxin contamination are generally time-consuming, sample-destructive, and require skilled personnel to perform, making them impossible for large-scale nondestructive screening detection, real-time, and on-site analysis. Therefore, the potential of visible-near-infrared (Vis-NIR) spectroscopy over the 400-2500 nm spectral range was examined for determination of aflatoxigenic fungus infection and the corresponding aflatoxin contamination on corn kernels in a rapid and nondestructive manner. The two A. flavus strains, AF13 and AF38, were used to represent the aflatoxigenic fungus and nonaflatoxigenic fungus, respectively, for artificial inoculation on corn kernels. The partial least-squares discriminant analysis (PLS-DA) models based on different combinations of spectral range (I: 410-1070 nm; II: 1120-2470 nm), corn side (endosperm or germ side), spectral variable number (full spectra or selected variables), modeling approach (two-step or one-step), and classification threshold (20 or 100 ppb) were developed and their performances were compared. The first study focusing on detection of aflatoxigenic fungus-infected corn kernels showed that, in classifying the "control+AF38-inoculated" and AF13-inoculated corn kernels, the full spectral PLS-DA models using the preprocessed spectra over range II and one-step approach yielded more accurate prediction results than using the spectra over range I and the two-step approach. The advantage of the full spectral PLS-DA models established using one corn side than the other side were not consistent in the explored combination cases. The best full spectral PLS-DA model obtained was obtained using the germ-side spectra over range II with the one-step approach, which achieved an overall accuracy of 91.11%. The established CARS-PLSDA models performed better than the corresponding full-spectral PLS-DA models, with the better model achieved an overall accuracy of 97.78% in separating the AF13-inoculated corn kernels and the uninfected control and AF38-inoculated corn kernels. The second study focusing on the detection of aflatoxin-contaminated corn kernels showed that, based on the aflatoxin threshold of 20 and 100 ppb, the best overall accuracy in classifying the aflatoxin-contaminated and healthy corn kernels attained 86.67% and 84.44%, respectively, using the CARS-PLSDA models. The quantitative modeling results using partial least-squares regression (PLSR) obtained the correlation coefficient of prediction set ( R) of 0.91, which indicated the possibility of using Vis-NIR spectroscopy to quantify aflatoxin concentration in aflatoxigenic fungus-infected corn kernels.
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http://dx.doi.org/10.1021/acs.jafc.9b01044DOI Listing
May 2019

The bZIP transcription factor Afap1 mediates the oxidative stress response and aflatoxin biosynthesis in Aspergillus flavus.

Rev Argent Microbiol 2019 Oct - Dec;51(4):292-301. Epub 2019 Mar 22.

Institute of Food Science and Technology, Chinese Academy of Agricultural Sciences/Key Laboratory of Agro-products Quality and Safety Control in Storage and Transport Process, Ministry of Agriculture, Beijing 100193, PR China. Electronic address:

Aflatoxin is a carcinogenic secondary metabolite produced mainly by Aspergillus flavus and Aspergillus parasiticus, which can seriously endanger the health of humans and animals. Oxidative stress is a common defense response, and it is known that reactive oxygen species (ROS) can induce the synthesis of a series of secondary metabolites, including aflatoxin. By using mutants lacking the afap 1 gene, the role of afap1 gene in oxidative stress and aflatoxin synthesis was assessed. The growth of the mutant strains was significantly inhibited by the increase in the concentration of HO, inhibition was complete at 40mmol/l. However, in the quantitative analysis by HPLC, the concentration of AFB1 increased with the increased HO until 10mmol/l. Following an analysis based on the information provided by the NCBI BLAST analysis, it was assumed that Afap1, a basic leucine zipper (bZIP) transcription factor, was associated with the oxidative stress in this fungus. Treatment with 5mmol/l HO completely inhibited the growth of the mutant strains in afap 1 but did not affect the growth of the CA14PTs strain (non-mutant strain). In addition, the concentration of AFB in the mutant strains was approximately ¼ of that observed in the CA14PTs strain. These results suggested that Afap1 plays a key role in the regulation of oxidative stress and aflatoxin production in A. flavus.
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http://dx.doi.org/10.1016/j.ram.2018.07.003DOI Listing
May 2020

Use of Visible-Near-Infrared (Vis-NIR) Spectroscopy to Detect Aflatoxin B on Peanut Kernels.

Appl Spectrosc 2019 Apr 20;73(4):415-423. Epub 2019 Feb 20.

2 USDA-ARS, Southern Regional Research Center, New Orleans, LA, USA.

Current methods for detecting aflatoxin contamination of agricultural and food commodities are generally based on wet chemical analyses, which are time-consuming, destructive to test samples, and require skilled personnel to perform, making them impossible for large-scale nondestructive screening and on-site detection. In this study, we utilized visible-near-infrared (Vis-NIR) spectroscopy over the spectral range of 400-2500 nm to detect contamination of commercial, shelled peanut kernels (runner type) with the predominant aflatoxin B (AFB). The artificially contaminated samples were prepared by dropping known amounts of aflatoxin standard dissolved in 50:50 (v/v) methanol/water onto peanut kernel surface to achieve different contamination levels. The partial least squares discriminant analysis (PLS-DA) models established using the full spectra over different ranges achieved good prediction results. The best overall accuracy of 88.57% and 92.86% were obtained using the full spectra when taking 20 and 100 parts per billion (ppb), respectively, as the classification threshold. The random frog (RF) algorithm was used to find the optimal characteristic wavelengths for identifying the surface AFB-contamination of peanut kernels. Using the optimal spectral variables determined by the RF algorithm, the simplified RF-PLS-DA classification models were established. The better RF-PLS-DA models attained the overall accuracies of 90.00% and 94.29% with the 20 ppb and 100 ppb thresholds, respectively, which were improved compared to using the full spectral variables. Compared to using the full spectral variables, the employed spectral variables of the simplified RF-PLS-DA models were decreased by at least 94.82%. The present study demonstrated that the Vis-NIR spectroscopic technique combined with appropriate chemometric methods could be useful in identifying AFB contamination of peanut kernels.
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http://dx.doi.org/10.1177/0003702819829725DOI Listing
April 2019

Breastfeeding and cardiovascular risk factors.

Curr Opin Lipidol 2019 02;30(1):45-47

Nuffield Department of Population Health, University of Oxford, Oxford, UK.

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http://dx.doi.org/10.1097/MOL.0000000000000564DOI Listing
February 2019

Monitoring Metabolite Production of Aflatoxin Biosynthesis by Orbitrap Fusion Mass Spectrometry and a D-Optimal Mixture Design Method.

Anal Chem 2018 12 3;90(24):14331-14338. Epub 2018 Dec 3.

Oil Crops Research Institute , Chinese Academy of Agricultural Sciences , Wuhan 430061 , People's Republic of China.

Aflatoxins, highly toxic and carcinogenic to humans, are synthesized via multiple intermediates by a complex pathway in several Aspergilli, including Aspergillus flavus. Few analytical methods are available for monitoring the changes in metabolite profiles of the aflatoxin biosynthesis pathway under different growth and environmental conditions. In the present study, we developed by a D-optimal mixture design a solvent system, methanol/dichloromethane/ethyl acetate/formic acid (0.36/0.31/0.32/0.01), that was suitable for extracting the pathway metabolites. The matrix effect from dilution of cell extracts was negligible. To facilitate the identification of these metabolites, we constructed a fragmentation ion library. We further employed liquid chromatography coupled with high-resolution mass spectroscopy (UHPLC-HRMS) for simultaneous quantification of the metabolites. The limit of detection (LOD) and limit of quantitation (LOQ) were 0.002-0.016 and 0.008-0.05 μg/kg, respectively. The spiked recovery rates ranged from 81.3 to 100.3% with intraday and interday precision less than 7.6%. Using the method developed to investigate the time-course aflatoxin biosynthesis, we found that precursors, including several possible toxins (with a carcinogenic group similar to aflatoxin B), occurred together with aflatoxin, and that production increased rapidly at the early growth stage, peaked on day four, and then decreased substantially. The maximum production of aflatoxin B and aflatoxin B occurred 1 day later. Moreover, the dominant branch pathway was the one for aflatoxin B formation. We revealed that the antiaflatoxigenicity mechanism of Leclercia adecarboxylata WT16 was associated with a factor upstream of the aflatoxin biosynthesis pathway. The design strategies can be applied to characterize or detect other secondary metabolites to provide a snapshot of the dynamic changes during their biosynthesis.
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http://dx.doi.org/10.1021/acs.analchem.8b03703DOI Listing
December 2018

Whole genome comparison of Aspergillus flavus L-morphotype strain NRRL 3357 (type) and S-morphotype strain AF70.

PLoS One 2018 2;13(7):e0199169. Epub 2018 Jul 2.

Food and Feed Safety Unit, USDA, New Orleans, Louisiana, United States of America.

Aspergillus flavus is a saprophytic fungus that infects corn, peanuts, tree nuts and other agriculturally important crops. Once the crop is infected the fungus has the potential to secrete one or more mycotoxins, the most carcinogenic of which is aflatoxin. Aflatoxin contaminated crops are deemed unfit for human or animal consumption, which results in both food and economic losses. Within A. flavus, two morphotypes exist: the S strains (small sclerotia) and L strains (large sclerotia). Significant morphological and physiological differences exist between the two morphotypes. For example, the S-morphotypes produces sclerotia that are smaller (< 400 μm), greater in quantity, and contain higher concentrations of aflatoxin than the L-morphotypes (>400 μm). The morphotypes also differ in pigmentation, pH homeostasis in culture and the number of spores produced. Here we report the first full genome sequence of an A. flavus S morphotype, strain AF70. We provide a comprehensive comparison of the A. flavus S-morphotype genome sequence with a previously sequenced genome of an L-morphotype strain (NRRL 3357), including an in-depth analysis of secondary metabolic clusters and the identification SNPs within their aflatoxin gene clusters.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0199169PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6028093PMC
December 2018

Modulation of cardiovascular risk factors.

Curr Opin Lipidol 2018 06;29(3):269-270

Diabetes Centre, Royal Oldham Hospital, Oldham.

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http://dx.doi.org/10.1097/MOL.0000000000000516DOI Listing
June 2018

Identification of candidate resistance genes of cotton against infection using a comparative transcriptomics approach.

Physiol Mol Biol Plants 2018 May 22;24(3):513-519. Epub 2018 Mar 22.

1School of Plant, Environmental, and Soil Sciences, Louisiana State University Agricultural Center, Baton Rouge, LA USA.

A comparative transcriptome analysis was performed using the genes significantly differentially expressed in cotton, corn and peanut in response to aflatoxin producing fungus with an objective of identifying candidate resistance genes in cotton. Two-way analyses identified 732 unique genes to be differentially regulated by the fungus with only 26 genes common across all three crops that were considered candidate resistance genes with an assumption that these genes have specific roles in conferring the resistance trait. Genes of membrane cellular component involved in DNA binding with involvement in defense responses were highly represented among the differentially expressed unique genes. Most (six) of these genes coded for 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase superfamily proteins. Genes encoding helix loop helix protein, alcohol dehydrogenase and UDP glycosylation transferase which were upregulated in response to both atoxigenic and toxigenic strains of , could be potential resistance candidate genes for downstream functional manipulation to confer resistance.
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http://dx.doi.org/10.1007/s12298-018-0522-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5911267PMC
May 2018

The Mycotox Charter: Increasing Awareness of, and Concerted Action for, Minimizing Mycotoxin Exposure Worldwide.

Toxins (Basel) 2018 04 4;10(4). Epub 2018 Apr 4.

Department of Plant Pathology, Throckmorton Plant Sciences Center, 1712 Claflin Avenue, Kansas State University, Manhattan, KS 66506, USA.

Mycotoxins are major food contaminants affecting global food security, especially in low and middle-income countries. The European Union (EU) funded project, MycoKey, focuses on “Integrated and innovative key actions for mycotoxin management in the food and feed chains” and the right to safe food through mycotoxin management strategies and regulation, which are fundamental to minimizing the unequal access to safe and sufficient food worldwide. As part of the MycoKey project, a Mycotoxin Charter (charter.mycokey.eu) was launched to share the need for global harmonization of mycotoxin legislation and policies and to minimize human and animal exposure worldwide, with particular attention to less developed countries that lack effective legislation. This document is in response to a demand that has built through previous European Framework Projects—MycoGlobe and MycoRed—in the previous decade to control and reduce mycotoxin contamination worldwide. All suppliers, participants and beneficiaries of the food supply chain, for example, farmers, consumers, stakeholders, researchers, members of civil society and government and so forth, are invited to sign this charter and to support this initiative.
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http://dx.doi.org/10.3390/toxins10040149DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5923315PMC
April 2018

Carbon Dioxide Mediates the Response to Temperature and Water Activity Levels in Aspergillus flavus during Infection of Maize Kernels.

Toxins (Basel) 2017 12 22;10(1). Epub 2017 Dec 22.

Department of Entomology and Plant Pathology, 223 Partners III, P.O. Box 7567, North Carolina State University, Raleigh, NC 27695, USA.

is a saprophytic fungus that may colonize several important crops, including cotton, maize, peanuts and tree nuts. Concomitant with colonization is its potential to secrete mycotoxins, of which the most prominent is aflatoxin. Temperature, water activity (a) and carbon dioxide (CO₂) are three environmental factors shown to influence the fungus-plant interaction, which are predicted to undergo significant changes in the next century. In this study, we used RNA sequencing to better understand the transcriptomic response of the fungus to a, temperature, and elevated CO₂ levels. We demonstrate that aflatoxin (AFB₁) production on maize grain was altered by water availability, temperature and CO₂. RNA-Sequencing data indicated that several genes, and in particular those involved in the biosynthesis of secondary metabolites, exhibit different responses to water availability or temperature stress depending on the atmospheric CO₂ content. Other gene categories affected by CO₂ levels alone (350 ppm vs. 1000 ppm at 30 °C/0.99 a), included amino acid metabolism and folate biosynthesis. Finally, we identified two gene networks significantly influenced by changes in CO₂ levels that contain several genes related to cellular replication and transcription. These results demonstrate that changes in atmospheric CO₂ under climate change scenarios greatly influences the response of to water and temperature when colonizing maize grain.
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http://dx.doi.org/10.3390/toxins10010005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5793092PMC
December 2017

The Pathogenesis-Related Maize Seed () Gene Plays a Role in Resistance to Infection and Aflatoxin Contamination.

Front Plant Sci 2017 17;8:1758. Epub 2017 Oct 17.

Food and Feed Safety Research Unit, United States Department of Agriculture - Agricultural Research Service, Southern Regional Research Center, New Orleans, LA, United States.

is an opportunistic plant pathogen that colonizes and produces the toxic and carcinogenic secondary metabolites, aflatoxins, in oil-rich crops such as maize ( L.). Pathogenesis-related (PR) proteins serve as an important defense mechanism against invading pathogens by conferring systemic acquired resistance in plants. Among these, production of the PR maize seed protein, (AC205274.3_FG001), has been speculated to be involved in resistance to infection by and other pathogens. To better understand the relative contribution of to resistance and aflatoxin production, a seed-specific RNA interference (RNAi)-based gene silencing approach was used to develop transgenic maize lines expressing hairpin RNAs to target . Downregulation of in transgenic kernels resulted in a ∼250-350% increase in infection accompanied by a ∼4.5-7.5-fold higher accumulation of aflatoxins than control plants. Gene co-expression network analysis of RNA-seq data during the -maize interaction identified as a network hub possibly responsible for regulating several downstream candidate genes associated with disease resistance and other biochemical functions. Expression analysis of these candidate genes in the -RNAi lines demonstrated downregulation (vs. control) of a majority of these -regulated genes during infection. These results are consistent with a key role of in resistance to infection and aflatoxin accumulation in maize kernels.
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http://dx.doi.org/10.3389/fpls.2017.01758DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5651032PMC
October 2017

Peanuts that keep aflatoxin at bay: a threshold that matters.

Plant Biotechnol J 2018 05 17;16(5):1024-1033. Epub 2017 Oct 17.

International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Hyderabad, Telangana, India.

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.
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http://dx.doi.org/10.1111/pbi.12846DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5902767PMC
May 2018

Temporal Effects on Internal Fluorescence Emissions Associated with Aflatoxin Contamination from Corn Kernel Cross-Sections Inoculated with Toxigenic and Atoxigenic .

Front Microbiol 2017 15;8:1718. Epub 2017 Sep 15.

Southern Regional Research Center, United States Department of Agriculture-Agricultural Research Service, New OrleansLA, United States.

Non-invasive, easy to use and cost-effective technology offers a valuable alternative for rapid detection of carcinogenic fungal metabolites, namely aflatoxins, in commodities. One relatively recent development in this area is the use of spectral technology. Fluorescence hyperspectral imaging, in particular, offers a potential rapid and non-invasive method for detecting the presence of aflatoxins in maize infected with the toxigenic fungus . Earlier studies have shown that whole maize kernels contaminated with aflatoxins exhibit different spectral signatures from uncontaminated kernels based on the external fluorescence emission of the whole kernels. Here, the effect of time on the internal fluorescence spectral emissions from cross-sections of kernels infected with toxigenic and atoxigenic , were examined in order to elucidate the interaction between the fluorescence signals emitted by some aflatoxin contaminated maize kernels and the fungal invasion resulting in the production of aflatoxins. First, the difference in internal fluorescence emissions between cross-sections of kernels incubated in toxigenic and atoxigenic inoculum was assessed. Kernels were inoculated with each strain for 5, 7, and 9 days before cross-sectioning and imaging. There were 270 kernels (540 halves) imaged, including controls. Second, in a different set of kernels (15 kernels/group; 135 total), the germ of each kernel was separated from the endosperm to determine the major areas of aflatoxin accumulation and progression over nine growth days. Kernels were inoculated with toxigenic and atoxigenic fungal strains for 5, 7, and 9 days before the endosperm and germ were separated, followed by fluorescence hyperspectral imaging and chemical aflatoxin determination. A marked difference in fluorescence intensity was shown between the toxigenic and atoxigenic strains on day nine post-inoculation, which may be a useful indicator of the location of aflatoxin contamination. This finding suggests that both, the fluorescence peak shift and intensity as well as timing, may be essential in distinguishing toxigenic and atoxigenic fungi based on spectral features. Results also reveal a possible preferential difference in the internal colonization of maize kernels between the toxigenic and atoxigenic strains of suggesting a potential window for differentiating the strains based on fluorescence spectra at specific time points.
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http://dx.doi.org/10.3389/fmicb.2017.01718DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5605554PMC
September 2017

Recent developments and applications of hyperspectral imaging for rapid detection of mycotoxins and mycotoxigenic fungi in food products.

Crit Rev Food Sci Nutr 2019 16;59(1):173-180. Epub 2017 Oct 16.

c Southern Regional Research Center, Agricultural Research Service-United States Department of Agriculture , New Orleans , LA , USA.

Mycotoxins are the foremost naturally occurring contaminants of food products such as corn, peanuts, tree nuts, and wheat. As the secondary metabolites, mycotoxins are mainly synthesized by many species of the genera Aspergillus, Fusarium and Penicillium, and are considered highly toxic and carcinogenic to humans and animals. Most mycotoxins are detected and quantified by analytical chemistry-based methods. While mycotoxigenic fungi are usually identified and quantified by biological methods. However, these methods are time-consuming, laborious, costly, and inconsistent because of the variability of the grain-sampling process. It is desirable to develop rapid, non-destructive and efficient methods that objectively measure and evaluate mycotoxins and mycotoxigenic fungi in food. In recent years, some spectroscopy-based technologies such as hyperspectral imaging (HSI), Raman spectroscopy, and Fourier transform infrared spectroscopy have been extensively investigated for their potential use as tools for the detection, classification, and sorting of mycotoxins and toxigenic fungal contaminants in food. HSI integrates both spatial and spectral information for every pixel in an image, making it suitable for rapid detection of large quantities of samples and more heterogeneous samples and for in-line sorting in the food industry. In order to track the latest research developments in HSI, this paper gives a brief overview of the theories and fundamentals behind the technology and discusses its applications in the field of rapid detection and sorting of mycotoxins and toxigenic fungi in food products. Additionally, advantages and disadvantages of HSI are compared, and its potential use in commercial applications is reported.
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http://dx.doi.org/10.1080/10408398.2017.1363709DOI Listing
June 2019

Interactions between water activity and temperature on the Aspergillus flavus transcriptome and aflatoxin B production.

Int J Food Microbiol 2017 Sep 26;256:36-44. Epub 2017 May 26.

Food Hygiene and Safety, University of Extremadura, Cáceres, Spain. Electronic address:

Effects of Aspergillus flavus colonization of maize kernels under different water activities (a; 0.99 and 0.91) and temperatures (30, 37°C) on (a) aflatoxin B (AFB) production and (b) the transcriptome using RNAseq were examined. There was no significant difference (p=0.05) in AFB production at 30 and 37°C and 0.99 a. However, there was a significant (p=0.05) increase in AFB at 0.91 a at 37°C when compared with 30°C/0.99 a. Environmental stress effects using gene ontology enrichment analysis of the RNA-seq results for increasing temperature at 0.99 and 0.91 a showed differential expression of 2224 and 481 genes, respectively. With decreasing water availability, 4307 were affected at 30°C and 702 genes at 37°C. Increasing temperature from 30 to 37°C at both a levels resulted in 12 biological processes being upregulated and 9 significantly downregulated. Decreasing a at both temperatures resulted in 22 biological processes significantly upregulated and 25 downregulated. The interacting environmental factors influenced functioning of the secondary metabolite gene clusters for aflatoxins and cyclopiazonic acid (CPA). An elevated number of genes were co-regulated by both a and temperature. An interaction effect for 4 of the 25 AFB genes, including regulatory and transcription activators occurred. For CPA, all 5 biosynthetic genes were affected by a stress, regardless of temperature. The molecular regulation of A. flavus in maize is discussed.
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http://dx.doi.org/10.1016/j.ijfoodmicro.2017.05.020DOI Listing
September 2017

Dietary adherence and cardiovascular risk.

Authors:
Deepak Bhatnagar

Curr Opin Lipidol 2017 04;28(2):214-215

aDiabetes Centre, The Royal Oldham Hospital, Oldham bUniversity of Manchester cCentre for Endocrinology and Diabetes, University of Salford, Salford, Manchester, UK.

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http://dx.doi.org/10.1097/MOL.0000000000000406DOI Listing
April 2017

The TLR4-NOS1-AP1 signaling axis regulates macrophage polarization.

Inflamm Res 2017 Apr 24;66(4):323-334. Epub 2016 Dec 24.

Center for Biosciences and Biomedical Engineering (BSBE), Indian Institute of Technology Indore (IITI), Indore, 453552, MP, India.

Objective: Macrophages polarize to proinflammatory M1 or anti-inflammatory M2 states with distinct physiological functions. This transition within the M1-M2 phenotypes decides the nature, duration and severity of an inflammatory response. Although there is a substantial understanding of the fate of these phenotypes, the underlying molecular mechanism of transition within the M1-M2 phenotypes is not well understood. We have investigated the role of neuronal nitric oxide synthase (NOS1)-mediated regulation of activator protein 1 (AP-1) transcription factor in macrophages as a critical effector of macrophage phenotypic change.

Materials And Methods: Raw 264.7 and THP1 macrophages were stimulated with LPS (250 ng/ml) to activate the inflammatory signaling pathway. We analyzed the effect of pharmacological NOS1 inhibitor: TRIM (1-(2- Trifluoromethylphenyl) imidazole) on LPS-induced inflammatory response in macrophages.

Results: We determined that NOS1-derived nitric oxide (NO) facilitate Fos and Jun interaction which induces IL-12 & IL-23 expression. Pharmacological inhibition of NOS1 inhibits ATF2 and Jun dimer. Switching of Fos and Jun dimer to ATF2 and Jun dimerization controls phenotype transition from IL-12 IL-23 IL-10 to IL-12 IL-23IL-10 phenotype, respectively.

Conclusion: These findings highlight a key role of the TLR4-NOS1-AP1 signaling axis in regulating macrophage polarization.
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http://dx.doi.org/10.1007/s00011-016-1017-zDOI Listing
April 2017

A Network Approach of Gene Co-expression in the / Pathosystem to Map Host/Pathogen Interaction Pathways.

Front Genet 2016 21;7:206. Epub 2016 Nov 21.

Department of Plant Biology, Southern Illinois University, Carbondale IL, USA.

A gene co-expression network (GEN) was generated using a dual RNA-seq study with the fungal pathogen and its plant host during the initial 3 days of infection. The analysis deciphered novel pathways and mapped genes of interest in both organisms during the infection. This network revealed a high degree of connectivity in many of the previously recognized pathways in such as jasmonic acid, ethylene, and reactive oxygen species (ROS). For the pathogen , a link between aflatoxin production and vesicular transport was identified within the network. There was significant interspecies correlation of expression between and for a subset of 104 , and 1942 genes. This resulted in an interspecies subnetwork enriched in multiple genes involved in the production of ROS. In addition to the ROS from , there was enrichment in the vesicular transport pathways and the aflatoxin pathway for . Included in these genes, a key aflatoxin cluster regulator, AflS, was found to be co-regulated with multiple ROS producing genes within the network, suggesting AflS may be monitoring host ROS levels. The entire GEN for both host and pathogen, and the subset of interspecies correlations, is presented as a tool for hypothesis generation and discovery for events in the early stages of fungal infection of by .
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http://dx.doi.org/10.3389/fgene.2016.00206DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5116468PMC
November 2016

Crystal structure and biochemical investigations reveal novel mode of substrate selectivity and illuminate substrate inhibition and allostericity in a subfamily of Xaa-Pro dipeptidases.

Biochim Biophys Acta Proteins Proteom 2017 Feb 2;1865(2):153-164. Epub 2016 Nov 2.

High Pressure and Synchrotron Radiation Physics Division, Bhabha Atomic Research Centre, Mumbai, India. Electronic address:

Xaa-Pro dipeptidase (XPD) catalyzes hydrolysis of iminopeptide bond in dipeptides containing trans-proline as a second residue. XPDs are found in all living organisms and are believed to play an essential role in proline metabolism. Here, we report crystal structures and extensive enzymatic studies of XPD from Xanthomonas campestris (XPDxc), the first such comprehensive study of a bacterial XPD. We also report enzymatic activities of its ortholog from Mycobacterium tuberculosis (XPDmt). These enzymes are strictly dipeptidases with broad substrate specificities. They exhibit substrate inhibition and allostericity, as described earlier for XPD from Lactococcus lactis (XPDll). The structural, mutational and comparative data have revealed a novel mechanism of dipeptide selectivity and substrate binding in these enzymes. Moreover, we have identified conserved sequence motifs that distinguish these enzymes from other prolidases, thus defining a new subfamily. This study provides a suitable structural template for explaining unique properties of this XPDxc subfamily. In addition, we report unique structural features of XPDxc protein like an extended N-terminal tail region and absence of a conserved Tyr residue near the active site.
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http://dx.doi.org/10.1016/j.bbapap.2016.10.016DOI Listing
February 2017

Amelioration of oxidative and inflammatory changes by Swertia chirayita leaves in experimental arthritis.

Inflammopharmacology 2016 Dec 14;24(6):363-375. Epub 2016 Oct 14.

School of Biochemistry, Devi Ahilya University, Khandwa Road, Indore, MP, 452001, India.

The present study was aimed to determine the therapeutic effects of Swertia chirayita leaves against oxidative and inflammatory injuries in Freund's complete adjuvant (FCA) induced arthritic rats. The extract was evaluated for its phytoconstituents and various invitro antioxidant properties followed by its in vivo effects. The hydroethanolic extract of S. chirayita leaves (SCE) was orally administered (200 mg/kg body weight, per day, p.o.) and the effect on the liver lipid peroxidation (LPO), antioxidant status, protein carbonyl formation along with the histopathology of liver were evaluated after induction of adjuvant arthritis. The markers of inflammation and arthritis, such as tumor necrosis factor-α (TNF-α), interleukin 1α (IL-1α), inhibition of paw edema, along with the histological and radiographic changes in the arthritic ankle joint were studied with and without SCE administration. The result showed the presence of major phytoconstituents, such as phenolic, flavonoid and terpenoid content in SCE. HPLC analysis revealed the presence of swertiamarin and amarogentin in high concentration. The extract also showed in vitro antioxidant potential which has positive correlation with the phytoconstituents. The result of in vivo study showed elevated malondialdehyde (MDA) and carbonyl content indicative of LPO and protein oxidation, respectively, with compromised intracellular antioxidant defense system in arthritic rats, which were significantly normalized after SCE treatment. The increase in serum proinflammatory cytokines (TNF- α and IL-1α) and paw edema of arthritic rats was significantly suppressed by SCE. Histology and radiographic analysis of arthritic ankle joints indicated abnormal changes. Marked reduction in inflammation and arthritic changes were observed after treatment with SCE. The present investigation suggests that hydroethanolic extract of S. chirayita leaves exhibit potential immunomodulatory effects, which may possibly be due to boosting the intracellular antioxidant defense.
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http://dx.doi.org/10.1007/s10787-016-0290-3DOI Listing
December 2016

RNA sequencing of an nsdC mutant reveals global regulation of secondary metabolic gene clusters in Aspergillus flavus.

Microbiol Res 2016 Jan 2;182:150-61. Epub 2015 Sep 2.

USDA, ARS, Southern Regional Research Center 1100 Robert E Lee Blvd., New Orleans, LA 70124, USA. Electronic address:

The filamentous fungus, Aspergillus flavus (A. flavus) is an opportunistic pathogen capable of invading a number of crops and contaminating them with toxic secondary metabolites such as aflatoxins. Characterizing the molecular mechanisms governing growth and development of this organism is vital for developing safe and effective strategies for reducing crop contamination. The transcription factor nsdC has been identified as being required for normal asexual development and aflatoxin production in A. flavus. Building on a previous study using a large (L)-sclerotial morphotype A. flavus nsdC mutant we observed alterations in conidiophore development and loss of sclerotial and aflatoxin production using a nsdC mutant of a small (S)-sclerotial morphotype, that normally produces aflatoxin and sclerotia in quantities much higher than the L-morphotype. RNA sequencing analysis of the nsdC knockout mutant and isogenic control strain identified a number of differentially expressed genes related to development and production of secondary metabolites, including aflatoxin, penicillin and aflatrem. Further, RNA-seq data indicating down regulation of aflatrem biosynthetic gene expression in the nsdC mutant correlated with HPLC analyses showing a decrease in aflatrem levels. The current study expands the role of nsdC as a globally acting transcription factor that is a critical regulator of both asexual reproduction and secondary metabolism in A. flavus.
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http://dx.doi.org/10.1016/j.micres.2015.08.007DOI Listing
January 2016

The Mediterranean diet.

Curr Opin Lipidol 2016 Feb;27(1):89-91

aDiabetes Centre, Royal Oldham Hospital, Oldham bCentre for Endocrinology and Diabetes, Univesrity of Manchester, Manchester cUniversity of Salford, Salford, UK.

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http://dx.doi.org/10.1097/MOL.0000000000000266DOI Listing
February 2016

A novel gastrointestinal microbiome modulator from soy pods reduces absorption of dietary fat in mice.

Obesity (Silver Spring) 2016 Jan 20;24(1):87-95. Epub 2015 Nov 20.

MicroBiome Therapeutics, New Orleans, Louisiana, USA.

Objective: Simplification of diets, low in variety but high in energy, contributes to the loss in diversity observed in the obese gastrointestinal (GI) microbiome. A novel GI microbiome modulator (GIMM) as a dietary intervention was developed.

Methods: Mice were fed either an obesogenic diet (ObD) or an ObD containing 15% activated soy pod fiber (ObD-ASPF) for 30 days. The diets were isocaloric and balanced for macronutrient content. ASPF is a novel fiber preparation from whole soy pods that is activated to produce glyceollins.

Results: Mice fed ObD-ASPF did not gain body fat. This was associated with decreased absorption of calories (P < 0.05) and increased fecal excretion of triglycerides, which may be attributed to decreased bile acid secretion (P < 0.05). A shift (P < 0.05) in abundances of microbiota in 10 genera was observed. Mice fed ObD-ASPF had elevated plasma concentrations of the anti-inflammatory IL-10 (P < 0.05) and decreased (P < 0.05) plasma concentrations of the neutrophil chemoattractant CXCL1.

Conclusions: A novel dietary intervention derived from soy pods that acts to hinder absorption of dietary fat and glucose in mice was developed. More studies with this GIMM in animal models of diet-induced nonalcoholic fatty liver diseases, type 2 diabetes, and autism are needed.
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http://dx.doi.org/10.1002/oby.21197DOI Listing
January 2016

The impact of gestational hypercholesterolaemia on origins of disease.

Atherosclerosis 2015 Dec 22;243(2):652-3. Epub 2015 Oct 22.

Centre for Endocrinology and Diabetes, Core Technology Facility, University of Manchester, 46 Grafton Street, Manchester, M13 9NT, UK; Diabetes Centre, The Royal Oldham Hospital, Rochdale Road, Oldham, OL1 2JH, UK.

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http://dx.doi.org/10.1016/j.atherosclerosis.2015.10.012DOI Listing
December 2015

Genome-Wide Transcriptome Analysis of Cotton (Gossypium hirsutum L.) Identifies Candidate Gene Signatures in Response to Aflatoxin Producing Fungus Aspergillus flavus.

PLoS One 2015 14;10(9):e0138025. Epub 2015 Sep 14.

School of Plant, Environmental and Soil Sciences, Louisiana State University Agricultural Center, Baton Rouge, LA, 70803, United States of America.

Aflatoxins are toxic and potent carcinogenic metabolites produced from the fungi Aspergillus flavus and A. parasiticus. Aflatoxins can contaminate cottonseed under conducive preharvest and postharvest conditions. United States federal regulations restrict the use of aflatoxin contaminated cottonseed at >20 ppb for animal feed. Several strategies have been proposed for controlling aflatoxin contamination, and much success has been achieved by the application of an atoxigenic strain of A. flavus in cotton, peanut and maize fields. Development of cultivars resistant to aflatoxin through overexpression of resistance associated genes and/or knocking down aflatoxin biosynthesis of A. flavus will be an effective strategy for controlling aflatoxin contamination in cotton. In this study, genome-wide transcriptome profiling was performed to identify differentially expressed genes in response to infection with both toxigenic and atoxigenic strains of A. flavus on cotton (Gossypium hirsutum L.) pericarp and seed. The genes involved in antifungal response, oxidative burst, transcription factors, defense signaling pathways and stress response were highly differentially expressed in pericarp and seed tissues in response to A. flavus infection. The cell-wall modifying genes and genes involved in the production of antimicrobial substances were more active in pericarp as compared to seed. The genes involved in auxin and cytokinin signaling were also induced. Most of the genes involved in defense response in cotton were highly induced in pericarp than in seed. The global gene expression analysis in response to fungal invasion in cotton will serve as a source for identifying biomarkers for breeding, potential candidate genes for transgenic manipulation, and will help in understanding complex plant-fungal interaction for future downstream research.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0138025PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4569580PMC
May 2016