Genetic fingerprinting and aflatoxin production of Aspergillus section Flavi associated with groundnut in eastern Ethiopia

Abstract Background Aspergillus species cause aflatoxin contamination in groundnut kernels, being a health threat in agricultural products and leading to commodity rejection by domestic and international markets. Presence of Aspergillus flavus and A. parasiticus colonizing groundnut in eastern Ethiopia, as well as presence of aflatoxins have been reported, though in this region, no genetic studies have been done of these species in relation to their aflatoxin production. Results In this study, 145 Aspergillus isolates obtained from groundnut kernels in eastern Ethiopia were genetically fingerprinted using 23 Insertion/Deletion (InDel) markers within the aflatoxin-biosynthesis gene cluster (ABC), identifying 133 ABC genotypes. Eighty-four isolates were analyzed by Ultra-Performance Liquid Chromatography (UPLC) for in vitro aflatoxin production. Analysis of genetic distances based on the approximately 85 kb-ABC by Neighbor Joining (NJ), 3D-Principal Coordinate Analysis (3D-PCoA), and Structure software, clustered the isolates into three main groups as a gradient in their aflatoxin production. Group I, contained 98% A. flavus, including L- and non-producers of sclerotia (NPS), producers of B1 and B2 aflatoxins, and most of them collected from the lowland-dry Babile area. Group II was a genetic admixture population of A. flavus (NPS) and A. flavus S morphotype, both low producers of aflatoxins. Group III was primarily represented by A. parasiticus and A. flavus S morphotype isolates both producers of B1, B2 and G1, G2 aflatoxins, and originated from the regions of Darolabu and Gursum. The highest in vitro producer of aflatoxin B1 was A. flavus NPS N1436 (77.98 μg/mL), and the highest producer of aflatoxin G1 was A. parasiticus N1348 (50.33 μg/mL), these isolates were from Gursum and Darolabu, respectively. Conclusions To the best of our knowledge, this is the first study that combined the use of InDel fingerprinting of the ABC and corresponding aflatoxin production capability to describe the genetic diversity of Aspergillus isolates from groundnut in eastern Ethiopia. Three InDel markers, AFLC04, AFLC08 and AFLC19, accounted for the main assignment of individuals to the three Groups; their loci corresponded to aflC (pksA), hypC, and aflW (moxY) genes, respectively. Despite InDels within the ABC being often associated to loss of aflatoxin production, the vast InDel polymorphism observed in the Aspergillus isolates did not completely impaired their aflatoxin production in vitro.

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Evaluation of Post-harvest Aflatoxin Production in Peanut Germplasm with Resistance to Seed Colonization and Pre-harvest Aflatoxin Contamination

Peanut Science ◽

10.3146/pnut.31.2.0012 ◽

2004 ◽

Vol 31 (2) ◽

pp. 124-134 ◽

Cited By ~ 7

Author(s):

H. Q. Xue ◽

T. G. Isleib ◽

G. A. Payne ◽

G. OBrian

Keyword(s):

Genotypic Variation ◽

Fungal Growth ◽

Aflatoxin Production ◽

Aflatoxin Contamination ◽

Block Designs ◽

Plastic Bags ◽

Arachis Hypogaea L ◽

Seed Colonization ◽

Highly Correlated

Abstract Contamination of peanut (Arachis hypogaea L.) with aflatoxin produced by species of Aspergillus remains a problem for the U.S. peanut industry. Several peanut genotypes were reported to be resistant to in vitro seed colonization by Aspergillus flavus Link ex Fries (IVSCAF), to field seed colonization by A. flavus (FSCAF), or to preharvest aflatoxin contamination (PAC), but few to production of aflatoxin per se. Cotyledons of 39 peanut genotypes reportedly resistant to IVSCAF, FSCAF, or PAC, and eight susceptible to PAC were evaluated in four tests for their ability to support aflatoxin production after inoculation with A. flavus. Cultivars Perry and Gregory were used as checks in each test. Seed cotyledons were separated, manually blanched, inoculated with conidia of A. flavus, placed on moistened filter paper in petri dishes, and incubated for 8 d at 28 C. Dishes were arranged on plastic trays enclosed in plastic bags and stacked with PVC spacers between trays. Incomplete block designs were used for all tests. In each test, none of the genotypes examined was completely resistant to aflatoxin production, but significant genotypic variation was observed in the amount of total aflatoxin accumulated in seeds. Genotypes previously reported to be resistant to IVSCAF, FSCAF, or PAC exhibited differential abilities to support aflatoxin production. PI 590325, PI 590299, PI 290626, and PI 337409 supported reduced levels of aflatoxin, and their degree of resistance was consistent across tests. Fungal growth was highly correlated with aflatoxin production in three tests. The results from this study suggested that there were no absolute relationships of aflatoxin production resistance with IVSCAF, FSCAF, or PAC resistance, but that it should be possible to identify a genotype with high IVSCAF, FSCAF, or PAC resistance and reduced capacity for aflatoxin production by A. flavus.

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Assessment of the Effect of Satureja montana and Origanum virens Essential Oils on Aspergillus flavus Growth and Aflatoxin Production at Different Water Activities

Toxins ◽

10.3390/toxins12030142 ◽

2020 ◽

Vol 12 (3) ◽

pp. 142 ◽

Cited By ~ 3

Author(s):

Marta García-Díaz ◽

Jessica Gil-Serna ◽

Belén Patiño ◽

Esther García-Cela ◽

Naresh Magan ◽

...

Keyword(s):

Essential Oil ◽

Essential Oils ◽

Aspergillus Flavus ◽

Aflatoxin Production ◽

Aflatoxin Contamination ◽

Activity Levels ◽

Control Methods ◽

Mycotoxin Biosynthesis ◽

Satureja Montana

Aflatoxin contamination of foodstuffs poses a serious risk to food security, and it is essential to search for new control methods to prevent these toxins entering the food chain. Several essential oils are able to reduce the growth and mycotoxin biosynthesis of toxigenic species, although their efficiency is strongly influenced by the environmental conditions. In this work, the effectiveness of Satureja montana and Origanum virens essential oils to control Aspergillus flavus growth was evaluated under three water activity levels (0.94, 0.96 and 0.98 aw) using a Bioscreen C, a rapid in vitro spectrophotometric technique. The aflatoxin concentrations at all conditions tested were determined by HPLC-FLD. Aspergillus flavus growth was delayed by both essential oil treatments. However, only S. montana essential oil was able to significantly affect aflatoxin production, although the inhibition percentages widely differed among water activities. The most significant reduction was observed at 0.96 aw, which is coincident with the conditions in which A. flavus reached the highest levels of aflatoxin production. On the contrary, the treatment with S. montana essential oil was not effective in significantly reducing aflatoxin production at 0.94 aw. Therefore, it is important to study the interaction of the new control compounds with environmental factors before their application in food matrices, and in vitro ecophysiological studies are a good option since they provide accurate and rapid results.

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Aspergillus flavus NRRL 3251 Growth, Oxidative Status, and Aflatoxins Production Ability In Vitro under Different Illumination Regimes

Toxins ◽

10.3390/toxins10120528 ◽

2018 ◽

Vol 10 (12) ◽

pp. 528 ◽

Cited By ~ 5

Author(s):

Tihomir Kovač ◽

Bojan Šarkanj ◽

Biljana Crevar ◽

Marija Kovač ◽

Ante Lončarić ◽

...

Keyword(s):

Aspergillus Flavus ◽

Growth Period ◽

Aflatoxin Production ◽

Aflatoxin Contamination ◽

Oxidative Status ◽

Light Illumination ◽

Proper Understanding ◽

Experimental Strains ◽

Oxidative Species

Aspergillus flavus is the most important mycotoxin-producing fungus involved in the global episodes of aflatoxin B1 contamination of crops at both the pre-harvest and post-harvest stages. However, in order to effectively control aflatoxin contamination in crops using antiaflatoxigenic and/or antifungal compounds, some of which are photosensitive, a proper understanding of the photo-sensitive physiology of potential experimental strains need to be documented. The purpose of the study is therefore to evaluate the effect of visible (VIS) light illumination on growth and conidiation, aflatoxin production ability and modulation of A. flavus oxidative status during in vitro experiment. Aflatoxigenic A. flavus strain was inoculated in aflatoxin-inducing YES media and incubated under three different VIS illumination regimes during a 168 h growth period at 29 °C. VIS illumination reduced A. flavus mycelia biomass yield, both during growth on plates and in liquid media, promoted conidiation and increased the aflatoxin production. Furthermore, aflatoxin production increased with increased reactive oxidative species (ROS) levels at 96 h of growth, confirming illumination-driven oxidative stress modulation activity on A. flavus cells.

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Aflatoxin-inhibiting ability of Bacillus subtilis isolated from soil in southern Vietnam

Veterinariya, Zootekhniya i Biotekhnologiya ◽

10.36871/vet.zoo.bio.202004004 ◽

2020 ◽

Vol 1 (4) ◽

pp. 31-35

Author(s):

Le Thi Ngoc An’ ◽

◽

T. N. Gryazneva ◽

Nguyen Ngoc Hai ◽

◽

...

Keyword(s):

Bacillus Subtilis ◽

Animal Feed ◽

Aflatoxin Production ◽

Aflatoxin Contamination ◽

Biologically Active ◽

Southern Vietnam

Reducing the level of aflatoxin contamination of animal feed using soil-isolated cultures of B. subtilis, showed the prospects of using this type of bacteria for decontamination of feed. A total of 367 B. subtilis cultures were isolated from soil in southern Vietnam and screened for inhibition of aflatoxin production by Aspergillus in vitro. Of these, 34 isolates of biologically active B. subtilis were selected, of which 7 isolates were the most resistant to aflatoxin. These cultures of bacilli after 5 days of cultivation in a mixture with Aspergillus on crushed corn contributed to a 26,76-fold decrease in aflatoxin levels compared to the control. The data obtained indicate that B. subtilis isolates isolated from soil can inhibit aflatoxin in vitro.

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Activity and Anti-Aflatoxigenic Effect of Indigenously Characterized Probiotic Lactobacilli against Aspergillus flavus—A Common Poultry Feed Contaminant

Animals ◽

10.3390/ani9040166 ◽

2019 ◽

Vol 9 (4) ◽

pp. 166 ◽

Cited By ~ 2

Author(s):

Nimra Azeem ◽

Muhammad Nawaz ◽

Aftab Ahmad Anjum ◽

Shagufta Saeed ◽

Saba Sana ◽

...

Keyword(s):

Aspergillus Flavus ◽

High Performance ◽

Animal Feed ◽

Aflatoxin Production ◽

Aflatoxin Contamination ◽

Poultry Feed ◽

Poultry Production ◽

Ameliorative Effect ◽

Probiotic Lactobacilli

Aflatoxin contamination in human food and animal feed is a threat to public safety. Aflatoxin B1 (AFB1) can be especially damaging to poultry production and consequently economic development of Pakistan. The present study assessed the in vitro binding of AFB1 by indigenously characterized probiotic lactobacilli. Six isolates (Lactobacillus gallinarum PDP 10, Lactobacillus reuetri FYP 38, Lactobacillus fermentum PDP 24, Lactobacillus gallinarum PL 53, Lactobacillus paracasei PL 120, and Lactobacillus gallinarum PL 149) were tested for activity against toxigenic Aspergillus flavus W-7.1 (AFB1 producer) by well diffusion assay. Only three isolates (PL 53, PL 120, and PL 149) had activity against A. flavus W-7.1. The ameliorative effect of these probiotic isolates on AFB1 production was determined by co-culturing fungus with lactobacilli for 12 days, followed by aflatoxin quantification by high-performance liquid chromatography. In vitro AFB1 binding capacities of lactobacilli were determined by their incubation with a standard amount of AFB1 in phosphate buffer saline at 37 °C for 2 h. AFB1 binding capacities of isolates ranged from 28–65%. Four isolates (PDP 10, PDP 24, PL 120, and PL 149) also ceased aflatoxin production completely, whereas PL 53 showed 55% reduction in AFB1 production as compared to control. The present study demonstrated Lactobacillus gallinarum PL 149 to be an effective candidate AFB1 binding agent against Aspergillus flavus. These findings further support the binding ability of lactic acid bacteria for dietary contaminants.

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Aspergillus flavus and Aflatoxin Contamination of Leguminous Trees of the Sonoran Desert in Arizona

Phytopathology ◽

10.1094/phyto.2001.91.9.913 ◽

2001 ◽

Vol 91 (9) ◽

pp. 913-919 ◽

Cited By ~ 28

Author(s):

María L. Boyd ◽

Peter J. Cotty

Keyword(s):

Aspergillus Flavus ◽

Sonoran Desert ◽

Aflatoxin Production ◽

Common Species ◽

Aflatoxin Contamination ◽

Leguminous Trees ◽

Significant Growth ◽

Tree Legumes ◽

Aflatoxin B

Aspergillus spp. in section Flavi were frequently associated with desert tree legumes in uncultivated areas of the Sonoran Desert. Of 270 samples of debris and fruits of mesquite (Prosopis spp.), ironwood (Olneya tesota), acacia (Acacia spp.), and palo verde (Cercidium and Parkinsonia spp.), 87% were positive for A. flavus (S and L strains) and A. tamarii. A. flavus was the most common species (87%) among the 3,763 isolates examined. Mesquite pods were both the substrate from which A. flavus was recovered most frequently and the substrate from native habitats with the greatest aflatoxin content. In vitro, most desert legumes supported significant growth, reproduction, and aflatoxin production by A. flavus, with mesquite pods yielding 1 × 1010 propagules/g and 5,000 μg/kg of aflatoxin B1. Twenty percent of legume pods collected in the desert contained measurable quantities of aflatoxin, ranging from 1 to >2,500 μg/kg. Insect-damaged mesquite pods had significantly higher aflatoxin than intact pods. Legumes are apparently important reservoirs of aflatoxin-producing fungi and significant sources of aflatoxin contamination in the native Sonoran Desert habitats of Arizona.

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Mitigating Aflatoxin Contamination in Groundnut through A Combination of Genetic Resistance and Post-Harvest Management Practices

Toxins ◽

10.3390/toxins11060315 ◽

2019 ◽

Vol 11 (6) ◽

pp. 315 ◽

Cited By ~ 16

Author(s):

Manish K. Pandey ◽

Rakesh Kumar ◽

Arun K. Pandey ◽

Pooja Soni ◽

Sunil S. Gangurde ◽

...

Keyword(s):

Management Practices ◽

Genetic Resistance ◽

Aflatoxin Production ◽

Climatic Conditions ◽

Aflatoxin Contamination ◽

Harvest Management ◽

Stunted Growth ◽

Post Harvest ◽

Crop Varieties

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.

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Breeding Peanut for Resistance to Aflatoxin Contamination at ICRISAT

Peanut Science ◽

10.3146/at07-008.1 ◽

2009 ◽

Vol 36 (1) ◽

pp. 42-49 ◽

Cited By ~ 48

Author(s):

S. N. Nigam ◽

F. Waliyar ◽

R. Aruna ◽

S. V. Reddy ◽

P. Lava Kumar ◽

...

Keyword(s):

Rural Economy ◽

Management Practices ◽

Aflatoxin Production ◽

Aflatoxin Contamination ◽

Seed Infection ◽

Sources Of Resistance ◽

Breeding Lines ◽

Post Harvest ◽

Components Of Resistance

Abstract Peanut plays an important role in the livelihoods of poor farmers and in the rural economy of many developing countries. Aflatoxin contamination in peanut seeds, caused by Aspergillus flavus, hampers international trade and adversely affects health of consumers of peanut and its products. It can occur in the field when the crop is growing, during harvesting and curing, and in storage and transportation. Aflatoxin research on peanut at ICRISAT focuses on identification and utilization of genetic resistance to preharvest seed infection and aflatoxin production by A. flavus and pre and post harvest management practices to minimize contamination. Breeding for aflatoxin resistance has been a contentious issue in peanut for nearly four decades since the first report of host resistance to aflatoxin production by A. flavus. Despite global efforts, progress in aflatoxin resistance breeding has been limited due to the low level of resistance to different components of resistance (preharvest seed infection and aflatoxin production, and in vitro seed colonization by A. flavus), their variable performance due to high G × E interaction, lack of reliable screening protocols, and limited understanding of genetics of resistance. Efforts to combine the three independently inherited components of resistance did not produce expected results towards improving the host plant resistance to aflatoxin contamination. Although breeding lines have shown better performance for resistance to aflatoxin contamination at ICRISAT, they need wider evaluation under diverse growing conditions. The search for better sources of resistance in the cultivated and wild Arachis germplasm continues, and recent developments in the area of transgenic research through modification of aflatoxin biosynthesis pathway or use of genes with antifungal and anti-aflatoxin properties appear encouraging. Meanwhile, the available improved breeding lines coupled with pre and post harvest aflatoxin management practices can help to significantly reduce aflatoxin contamination in farmers' fields. It is expected that transgenic resistance against fungal infection and aflatoxin production in combination with conventional breeding efforts may lead to the development of agronomically superior peanuts that are free of aflatoxin contamination.

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Effects of Rest and Exercise on Cardiac Blood Volume Determinations

Nuklearmedizin ◽

10.1055/s-0038-1629844 ◽

1997 ◽

Vol 36 (08) ◽

pp. 259-264

Author(s):

N. Topuzović

Keyword(s):

Radionuclide Ventriculography ◽

Left Ventricular ◽

Peak Exercise ◽

Volume Determination ◽

Group I ◽

Lv Volumes ◽

Group Ii ◽

Lv Volume

Summary Aim: The purpose of this study was to investigate the changes in blood activity during rest, exercise and recovery, and to assess its influence on left ventricular (LV) volume determination using the count-based method requiring blood sampling. Methods: Forty-four patients underwent rest-stress radionuclide ventriculography; Tc-99m-human serum albumin was used in 13 patients (Group I), red blood cells was labeled using Tc-99m in 17 patients (Group II) in vivo, and in 14 patients (Group III) by modified in vivo/in vitro method. LV volumes were determined by a count-based method using corrected count rate in blood samples obtained during rest, peak exercise and after recovery. Results: In group I at stress, the blood activity decreased by 12.6 ± 5.4%, p <0.05, as compared to the rest level, and increased by 25.1 ± 6.4%, p <0.001, and 12.8 ± 4.5%, p <0.05, above the resting level in group II and III, respectively. This had profound effects on LV volume determinations if only one rest blood aliquot was used: during exercise, the LV volumes significantly decreased by 22.1 ± 9.6%, p <0.05, in group I, whereas in groups II and III it was significantly overestimated by 32.1 ± 10.3%, p <0.001, and 10.7 ± 6.4%, p <0.05, respectively. The changes in blood activity between stress and recovery were not significantly different for any of the groups. Conclusion: The use of only a single blood sample as volume aliquot at rest in rest-stress studies leads to erroneous estimation of cardiac volumes due to significant changes in blood radioactivity during exercise and recovery.

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Anamnestic factors that shape reproductive health of women with repeated unsuccessful popygamy in vitro fertilization

HEALTH OF WOMAN ◽

10.15574/hw.2016.114.140 ◽

2016 ◽

pp. 140-143

Author(s):

N.V. Cotsabin ◽

◽

O.M. Makarchuk ◽

Keyword(s):

High Risk ◽

Risk Group ◽

Polycystic Ovary ◽

High Risk Group ◽

Stress Factors ◽

Control Group ◽

Fertilization In Vitro ◽

Infertile Women ◽

Group I

The proportion of patients with multiple unsuccessful attempts of assisted reproductive technology (ART) is about 30% of all patients treated with the use of ART. Women with history of unsuccessful ART attempts - a special category of patients who require emergency attention and a thorough examination at the stage of preparation for superovulation stimulation,the selection of embryos and endometrium preparation for embryo transfer. The objective: to distinguish high-risk group of unsuccessful attempts based on a detailed analysis of anamnestic and clinical data of infertile women with repeated unsuccessful ART attempts that requires more in-depth study of hormonal features, ovarian reserve and condition of the endometrium. Materials and methods. For better understanding of the problem of repeated unsuccessful ART attempts and сreation of efficient infertility treatment algorithms for these couples we conducted a thorough analysis of anamnestic data of three groups of infertile women (105 patients), which were distributed by age: group I – younger than 35, the II group – from 35 to 40, the III group - over 40 years. These groups of patients were compared with each other and with the control group of healthy women (30 persons). Results. Leading stress factors in the percentage three times prevailed in the group of infertile women and had a direct connection with the fact of procedure «fertilization in vitro» and chronic stressors caused by prolonged infertility. Primary infertility was observed significantly more frequent in patients younger than 35 years (p <0.05), secondary infertility - mostly in the second and third experimental groups (p <0.05). Noteworthy significant percentage of wellknown causes of infertility and idiopathic factor in all groups, and the prevalence of tubal-peritoneal factor in the second and third experimental groups, and endocrine dysfunction in the I experimental group. The most common disorder among this category of woman was polycystic ovary syndrome. Frequency of usual miscarriage among patients of I ana II groups was two times higher than in the third group (p <0.05). Among the experimental groups the leading place belongs urinary tract infection, respiratory tract diseases, pathologies of the cardiovascular system. Data of the stratified analysis show an increase likelihood of repeated unsuccessful ART attempts under the influence of constant chronic stress (odds ratio OR=2.06; 95% CI: 0.95–3.17; p<0.05). Conclusions. Among infertile patients with repeated unsuccessful ART attempts must be separated a high risk group of failures. The identity depends on the duration of infertility, female age and leading combination of factors. Key words: repeated unsuccessful ART attempts, anamnesis, infertility, high risk.

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