Natural Control of Corn Postharvest Fungi Aspergillus flavus and Penicillium sp. Using Essential Oils from Plants Grown in Argentina

The objective in this study was to evaluate the antifungal activity of essential oils from native and commercial aromatic plants grown in Argentina against corn postharvest fungi and to link the essential oil bioactivity with lipid oxidation and morphological changes in fungus cell membrane. Essential oil (EO) of oregano variety Mendocino (OMen), Cordobes (OCor), and Compacto (OCom), mint variety Inglesa (Mi), and Pehaujo (Mp), Suico (Sui); rosemary (Ro), and Aguaribay (Ag) were tested in vitro against 4 corn fungi: A. flavus (CCC116–83 and BXC01), P. oxalicum (083296), and P. minioluteum (BXC03). The minimum fungicidal concentration (MFC) and the minimum inhibitory concentration (MIC) were determined. The chemical profiles of the EOs were analyzed by GC‐MS. Lipid oxidation in cell membrane of fungi was determined by hydroperoxides and related with essential oil antifungal activity. The major compounds were Thymol in OCor (18.66%), Omen (12.18%), and OCom (9.44%); menthol in Mi and Mp; verbenone in Sui; dehydroxy‐isocalamendiol in Ag; and eucaliptol in Ro. OCor, Omen, and OCom showed the best antifungal activity. No antifungal activity was observed in Ag and Ro EO. The hydroperoxide value depended on the fungi (P < 0.001) and the antimicrobial agent (P < 0.001).Membrane lipids were oxidized by Sui EO in A. flavus BXC01 and A. flavus CCC116–83 (0.021 and 0.027 meqO₂/kg, respectively). The results suggest that the EOs of OCor, OMen, OCom, Mi, Mp, and Sui grown in Argentina can be used as natural alternatives to control fungi that produce mycotoxin in maize.     

Evaluation of chemically characterised essential oils of Coleus aromaticus,Hyptis suaveolens and Ageratum conyzoides against storage fungi and aflatoxin contamination of food commodities

The result of the present investigation explores the efficacy of chemically characterised essential oils (EOs) of Coleus aromaticus, Hyptis suaveolens and Ageratum conyzoides as antifungal and antiaflatoxigenic agent against some storage fungi and the toxigenic strain of Aspergillus flavus (Saktiman 3NSt). Minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) of these EOs were also determined against the toxigenic strain of A. flavus (Saktiman 3NSt). The EO from C. aromaticus was found to be most effective exhibiting MIC and MFC at 0.1μL mL^?1. The EOs also completely checked aflatoxin B₁ synthesis in concentration‐dependent manner. In addition, fumigation of stored wheat samples with EOs exhibited remarkable protection (>80%) from fungal infestation showing their efficacy during in vivo storage conditions. Based on the results of the present investigation, the EOs of C. aromaticus, H. suaveolens and A. conyzoides may be recommended as novel plant‐based antifungal and aflatoxin B₁ suppressor over the synthetic preservatives.     

Effect of UV-C irradiation on inactivation of Aspergillus flavus and Aspergillus parasiticus and quality parameters of roasted coffee bean (Coffea arabica L.)

ABSTRACT

This study investigated the antifungal effect of ultraviolet-C (UV-C) against Aspergillus flavus and Aspergillus parasiticus on roasted coffee beans. Also, any changes in the quality of the roasted coffee beans were measured after UV-C irradiation. As UV-C irradiation time increased (0–2 h), the number of surviving A. flavus and A. parasiticus spores significantly (P < .05) decreased. The reduction values of A. flavus in round part, crack part, and whole roasted coffee beans were 2.16, 0.71, and 1.58 log₁₀ CFU g^?1, respectively, and the reduction values of A. parasiticus in round part, crack part, and whole roasted coffee beans were 1.03, 0.37, and 0.72 log₁₀ CFU g^?1, respectively, after 2 h of UV-C irradiation. Field emission scanning electron microscopy showed that the morphology of A. flavus and A. parasiticus spores included expanded wrinkles that were deformed by UV-C irradiation. The Hunter colours were significantly reduced (P < .05). There was no significant change (P > .05) in moisture content, but the pH was significantly decreased (P < .05). Most of the sensory parameters did not change, but there was a significant difference (P < .05) in flavour. Based on this study, 2 h of UV-C irradiation was effective in reducing 90% of A. flavus, but it was not effective against A. parasiticus present on roasted coffee beans. Also, Hunter colour, pH, and sensory parameters (flavour) were changed by UV-C irradiation.     

Effects of Tropical Citrus Essential Oils on Growth, Aflatoxin Production, and Ultrastructure Alterations of Aspergillus flavus and Aspergillus parasiticus

Ethyl acetate extracts and hydrodistillated essential oils from five cultivars of tropical citrus epicarps were evaluated for their inhibitory activities against Aspergillus fumigatus, Aspergillus niger, Aspergillus flavus, Aspergillus parasiticus, and Penicillium sp. using disk diffusion and broth microdilution assays. Essential oils prepared from kaffir lime (Citrus hystrix DC) and acid lime (Citrus aurantifolia Swingle) epicarps exhibited stronger antifungal activity to all fungi than their ethyl acetate extracts with minimum inhibitory concentration and minimum fungicidal concentration values of 0.56 and 1.13 mg/ml (dry matter), respectively, against aflatoxin-producing A. flavus and A. parasiticus. The dominant components of the essential oil from kaffir lime were limonene, citronellol, linalool, o-cymene, and camphene, whereas limonene and p-cymene were major components of acid lime essential oil. Pure limonene, citronellal, and citronellol were five to six times less fungicidal than the natural essential oils, indicating the synergistic activity of many active compounds present in the oils. Kaffir and acid lime essential oils significantly reduced aflatoxin production of A. flavus and A. parasiticus, particularly lime essential oil, which completely inhibited growth and aflatoxin production of A. flavus at the concentration of 2.25 mg/ml. Target cell damage caused by acid lime essential oil was investigated under transmission electron microscopy. Destructive alterations of plasma and nucleus membrane, loss of cytoplasm, vacuole fusion, and detachment of fibrillar layer were clearly exhibited in essential-oil-treated cells.     

Original article: Antifungal activities of cinnamon oil against Rhizopus nigricans,Aspergillus flavus and Penicillium expansum in vitro and in vivo fruit test

The postharvest pathogens such as R. nigricans, A. flavas and P. expansum are the causal agents of jujube or orange fruit, therefore, in vitro and in vivo antifungal activities of cinnamon oil to inactivate these fungi were investigated. Cinnamaldehyde is the main constituent of cinnamon oil. The minimum inhibitory concentrations of cinnamon oil against Rhizopus nigricans, Aspergillus flavus and Penicillium expansum were 0.64% (v/v), 0.16% (v/v) and 0.16% (v/v), respectively. The antifungal activity of cinnamon oil against A. flavus and P. expansum was stronger than that against R. nigricans and the activity was improved with increasing its concentration. In an in vivo study, cinnamon oil with concentrations of 2.0% (v/v) and 3.0% (v/v) showed complete control the growth of fungi in wound‐inoculated Lingwu Long Jujube and Sand Sugar Orange fruits. These results revealed that cinnamon oil has a good potential to be as a natural antifungal agent for fruit applications.     

二氢杨梅素抑制黄曲霉菌生长的机制

本研究探讨了二氢杨梅素对黄曲霉的抗真菌活性和潜在的抗真菌机制。首先,通过抑菌实验证明,二氢杨梅素对黄曲霉孢子和菌丝的最小抑菌质量浓度（minimum inhibitory concentration,MIC）均为4 mg/mL。通过荧光增白剂（calcofluor white,CW）和碘化丙锭（propidium iodide,PI）染色实验证明,二氢杨梅素处理后黄曲霉细胞壁和细胞膜受损。与对照组相比,1/2 MIC和MIC组黄曲霉细胞内容物释放量（OD260 nm）分别增加了3.14 倍和5.93 倍,细胞外相对电导率和pH值均升高,MIC的二氢杨梅素对黄曲霉的呼吸抑制率达25.82%。这些结果表明,二氢杨梅素通过破坏细胞壁和细胞膜的完整性以及干扰呼吸代谢发挥抗菌活性。此外,二氢杨梅素在MIC时能够完全抑制黄曲霉在花生和玉米籽粒上的萌发,因此,二氢杨梅素可作为一种有效的抗真菌天然化合物应用于粮食及农产品储藏中。     

Synergistic antifungal effects of thyme essential oil and Lactobacillus plantarum cell‐free supernatant against Penicillium spp. and in situ effects

This study evaluates the synergistic antifungal effects between thyme essential oils and Lactobacillus plantarum cell‐free supernatant (LCFS) against Penicillium spp. and in situ antifungal activity in rice grains. Thyme essential oil and LCFS showed remarkable antifungal activities against Penicillium spp. with the minimum inhibitory concentration (MIC) of 40 and 80 µL/mL, respectively. The analysis of fractional inhibitory concentration (FIC) index showed the antifungal synergism between thyme essential oil and LCFS against Penicillium spp. with FIC index of 0.5. This synergism also resulted in fourfold reduction in their MICs when applied in combination. The antifungal modes of action were characterized by observing the changes in cell membrane permeability and degradation of fungal cell wall. The combination of thyme essential oil and LCFS (2 × MIC of each) showed remarkable in situ antifungal effect and completely inhibit the growth of Penicillium in rice seeds. The results suggested the possible applications of the observed synergism on actual crops.

Practical applications

Essential oils are used as preservative in food industry and high concentration of essential oil is associated with negative organoleptic characteristics. This study presented a novel approach for synergistic antifungal effects by using the combination of thyme essential oil and Lactobacillus plantarum cell‐free supernatant (LCFS) against Penicillium spp. and systematic evaluation of the antifungal effect by using fractional inhibitory concentration (FIC) index method. This approach will be a role model for future research on synergism and overcome the major drawbacks of using live bacteria and the negative effects arising from antimicrobial activities of essential oils.     

Chemical composition of Ocimum basilicum L. essential oil and its efficacy as a preservative against fungal and aflatoxin contamination of dry fruits

The study presents fungal and aflatoxin contamination of some dry fruits and Ocimum basilicum essential oil (EO) as a plant‐based preservative. During mycoflora analysis, 2045 fungal isolates were recorded from dry fruits and 40% isolates of Aspergillus flavus were toxigenic in nature. The EO of O. basilicum exhibited strong fungitoxicity against toxigenic strain of A. flavus. Its minimum inhibitory concentration (MIC) was recorded at 1.0 μL ml^?1, and it completely inhibited aflatoxin B₁ production at 0.5 μL ml^?1. The oil exhibited broad fungitoxic spectrum and considerably reduced A. flavus isolates from dry fruits when used as fumigant in closed storage containers at 1.0 μL ml^?1. The chemical profile of the EO was standardised through GC–MS analysis. Based on antifungal potency, antiaflatoxigenicity and efficacy as fumigant during storage conditions, O. basilicum EO may be recommended as a botanical preservative for enhancing the shelf life of dry fruits and edible products during storage.     

Antibacterial mode of action of seed essential oil of Eleutherococcus senticosus against foodborne pathogens

This study investigated the antibacterial mechanism of action of the seed essential oil of Eleutherococcus senticosus (ESEO) against foodborne pathogenic bacteria. Preliminarily, the ESEO (1000 μg disc^?1) showed potential antibacterial effect as diameter of inhibition zones (12.0 ± 0.2–37.0 ± 2.0 mm) against the tested foodborne pathogens. The MIC and MBC values of ESEO against the tested bacteria were found in the range of 125–500 and 500–1000 μg mL^?1, respectively. At MIC concentration, the ESEO had potential inhibitory effect on the cell viability of the tested pathogens. In addition, SEM analysis showed the inhibitory effect of ESEO as confirmed by considerable morphological alterations on the cell wall of B. cereus ATCC 13061 and E. coli O157:H7 ATCC 43889. Moreover, the ESEO revealed its mode of action against foodborne pathogens on membrane integrity as confirmed by release of extracellular ATP, 260‐nm absorbing materials and leakage of potassium ions. These findings confirm that the ESEO can be used as a potential antibacterial agent in food industry to inhibit the growth of various foodborne pathogens.     

Inactivation mechanisms of electron beam irradiation on Listeria innocua through the integrity of cell membrane,genomic DNA and protein structures

As a non-thermal sterilisation technology, electron beam irradiation (EBI) has attracted great interest for microbial inactivation in food preservation. This study aims to investigate the effects of EBI on membrane permeability, physiological status, morphological structure, genome integrity and protein structures of Listeria innocua irradiated at doses of 0.75, 1.50, 2.25, 3.00, 3.75 and 5.00 kGy. The results showed that EBI noticeably reduced the total microbial counts of L. innocua by more than 7 log CFU mL⁻¹ with 5.00 kGy treatment. The cell membrane permeability increased, resulting in the leakage of intracellular substances and changes in cell physiological status, which was proven by the cell staining and electron microscopy (EM) observations. Moreover, the integrity of genomic DNA and protein secondary structure, but not the protein primary structure were also disrupted. These findings provide the intrinsic mechanisms for the inactivation of L. innocua affected by EBI, which could serve as a theoretical basis for a better application of EBI in food sterilisation.     

Evaluation of Cinnamomum tamala Oil and Its Phenylpropanoid Eugenol for Their Antifungal and Antiaflatoxigenic Activity

During screening of 20 plant extracts against toxigenic strain of Aspergillus flavus (SK 3NSt), the extract of Cinnamomum tamala was found to exhibit absolute fungitoxic activity (100% growth inhibition). Hence, essential oil of C. tamala was extracted and selected for further investigations. The selected oil was subsequently standardised through physico-chemical and fungitoxic properties. Minimum inhibitory concentration (MIC) of the oil for absolute inhibition of growth of the toxigenic strain of A. flavus (SK 3NSt) was found to be 150 μl/l and oil showed fungicidal nature at its respective MIC. The oil had a broad fungitoxic spectrum. It was found to be absolutely inhibitory to almost all the 11 fungi tested when its fungitoxic spectrum was assayed. The antifungal potency of oil was found to be greater in comparison to some prevalent synthetic fungicides. The oils had the potency to withstand high inoculum density. The oil remained active up to 2 years and was thermostable. In addition, the oil showed significant efficacy in arresting aflatoxin B₁ secretion by the toxigenic strain (SK 3NSt) of A. flavus at 750 μl/l. GC-MS analysis of the oil led to the identification of main components of oil viz. eugenol (45.58%), β-pinene (10.03%), β-myrcene (9.73%), β-ocimene (4.51%), β-costol (3.88%) and thujyl alcohol isomer (2.51%). The efficacy of C. tamala oil as aflatoxin B₁ suppressor is being reported for the first time. Eugenol, the major component of oil showed absolute antiaflatoxigenic efficacy even at 250 μl/l. The high LD₅₀ value (16.94 ml/kg body weight) of oil recorded on mice indicates its non-mammalian toxicity and suggests the recommendation of the oil as a novel and safe post-harvest biological preservative of food commodities for their preservation from contaminating fungi.     

苯乳酸对单增李斯特菌的细胞膜完整性和通透性的影响

苯乳酸(D-(+)-3-Phenyllactic acid,PLA)是近年来在多种发酵食品中发现的天然高效抑菌小分子。本文采用流式细胞术、荧光显微镜、扫描电子显微镜等方法,研究了苯乳酸对食源性致病菌单增李斯特菌(L. monocytogenes 10403s)细胞膜通透性和完整性的影响。结果表明,苯乳酸最小抑菌浓度(Minimum inhibitory concentration,MIC)作用于该菌1 h后,细胞的通透率达到最高。通过流式细胞仪观察发现,通透细胞数占细胞总数的90.6%,且细胞膜的完整性随着作用时间的延长破坏程度越明显,且在作用1 h后,达到91.9%的破坏率。荧光显微镜直观地展现了细胞的损伤比例。在苯乳酸与L.monocytogenes 10403s作用1、3和6 h后,扫描电镜观察细胞形态有不同程度的皱缩、变形、细胞表面出现一定的孔洞,作用6 h后的细菌出现断裂、抱团和粘连现象。苯乳酸能够影响L.monocytogenes 10403s的细胞膜的通透性和完整性,为其在食品保鲜领域良好的应用提供一定的理论基础。     

金属抗菌肽SIF4对金黄色葡萄球菌细胞通透性的影响机制

为探明金属抗菌肽SIF4对金黄色葡萄球菌细胞通透性的影响机制,从胞外碱性磷酸酶活性、细胞表面电位、细胞表面疏水性、细胞内膜通透性和胞内生物大分子泄漏等角度考察了SIF4对细胞通透性的影响。研究发现,SIF4可破坏细胞壁结构完整性,随着抗菌肽质量浓度和温育时间的延长,胞外碱性磷酸酶活性也同步增长,2MIC组与TritonX-100组无显著差异（P>0.05）;细胞表面电位与SIF4质量浓度呈负相关关系;细胞表面疏水性与SIF4质量浓度呈良好的量-效正相关关系;细胞内膜通透性与抗菌肽质量浓度和温育时间呈正相关;胞内生物大分子泄漏与抗菌肽质量浓度和温育时间呈正相关,温育1 h时,胞内蛋白质泄漏呈差异性显著（P<0.05）,温育3 h后,2MIC与TritonX-100细胞内生物大分子泄漏差异基本不显著（P>0.05）。结果表明,SIF4可增强细胞通透性并使胞内物质泄漏,还能增强细胞表面疏水性和降低细胞表面电位,使细胞聚沉并诱导细胞坏死。     

Assessment of chemically characterised Rosmarinus officinalis L. essential oil and its major compounds as plant‐based preservative in food system based on their efficacy against food‐borne moulds and aflatoxin secretion and as antioxidant

The study explores antifungal, anti‐aflatoxigenic and antioxidant efficacy of Rosmarinus officinalis essential oil (ROEO) and its major compounds. In addition, the mode of action of ROEO and its practical efficacy as preservative have been assessed. GC‐MS analysis of ROEO identified 16 compounds; α‐pinene, 1,8‐cineole and camphor being the major compounds. The minimum concentration for inhibition of growth and aflatoxin B₁ secretion against A. flavus (LHP‐6) was found to be 1.5, >5.0, 4.0 and 3.0 μL mL^?1 and 1.25, >5.0, 3.5 and 3.0 μL mL^?1 for ROEO, α‐pinene, 1,8‐cineole and camphor, respectively. The IC₅₀ value through DPPH analysis and percentage inhibition of linoleic acid peroxidation of ROEO were 0.042 μL mL^?1 and 71.05%, respectively. The targeted site of antifungal action of ROEO was confirmed as plasma membrane through ergosterol measurement and TEM analysis. Moreover, ROEO significantly protected Piper nigrum fruits against mould infestation upto 6 months in in vivo trial.     

Effect of lucerne preservation method on the feed value of forage

BACKGROUND: Natural climatic wilt (NCW) and induced industrial wilt (IIW) are widely used as preservation methods for lucerne. Both of these methods reduce the quality of green forage due to respiration under NCW and heat damage under IIW. We compared the influence of these two preservation methods on nutritive value across a wide range of harvest conditions. RESULTS: Cell wall content and cell wall‐linked nitrogen values were higher (P < 0.05) in IIW than NCW. The preservation methods differed significantly (P < 0.05) in terms of soluble fraction, insoluble potentially degradable fraction of dry matter and effective degradability of dry matter. Nitrogen disappearance kinetics showed that the interactions of preservation by cut and preservation by phenological state were significant for the effective degradability of nitrogen. Organic matter digestibility was higher in lucerne preserved by NCW than IIW whereas cell wall digestibility was higher in lucerne preserved by IIW than NCW. Digestible organic matter intake did not differ between preservation methods (P > 0.05). CONCLUSION: Natural climatic wilt forage hay presents similar feed value to the induced industrial wilt alfalfa. Copyright © 2009 Society of Chemical Industry     

Enhanced antimicrobial activity of nisin in combination with allyl isothiocyanate against Listeria monocytogenes,Staphylococcus aureus,Salmonella Typhimurium and Shigella boydii

This study was designed to evaluate the synergistic antimicrobial effect of nisin and allyl isothiocyanate (AITC) against Listeria monocytogenes, Staphylococcus aureus, Salmonella Typhimurium and Shigella boydii. The synergistic interactions between nisin and AITC were observed against all foodborne pathogens, showing the fractional inhibitory concentrations <1. The populations of L. monocytogenes and S. aureus at the combined treatment of nisin and AITC were decreased to below 1 log CFU mL^?1 after 10‐h incubation at 37 °C. The changes in fatty acid profiles of all strains were substantially influenced by nisin alone and the combined treatment of nisin and AITC. A good agreement was observed among cell viability, membrane permeability and depolarisation activity in response to nisin and AITC. The results suggest that nisin and AITC as synergistic inhibitors could be an effective approach to achieve satisfactory antimicrobial activity against a wide range of foodborne pathogens.     

Fermentation of table cream by Lactobacillus plantarum strains: effect on fungal growth,aflatoxin M1 and ochratoxin A

Aflatoxin M1 (AFM₁) and ochratoxin A (OTA) are two main mycotoxins in milk and dairy products. In the present work, the ability of four Lactobacillus strains (L. plantarum PTCC 1058, L. plantarum LP3, L. plantarum AF1 and L. plantarum LU5) to remove AFM1 and OTA in fermented cream was studied during 24 h fermentation. The antifungal activity of the mentioned lactobacilli against the defined fungi (Aspergillus flavus PTCC 5004, Aspergillus parasiticus PTCC 5018, Aspergillus nidulans PTCC 5014, Aspergillus ochraceus PTCC 5060) was also evaluated. The results showed that the cell counts of all strains were increased by 64–70% during fermentation. All Lactobacillus strains decreased the amount of AFM₁ significantly (P ≤ 0.05) in the range of 26–52%, which the highest AFM₁-reducing effect was related to L. plantarum LU5 (from 0.5 to 0.24 μg kg⁻¹). The mean OTA removal by Lactobacillus strains in fermented cream also ranged from 32 to 58%. Amongst Lactobacillus strains, the cell-free culture supernatants of L. plantarum LU5 showed the highest (inhibition zone of 26.7 ± 1.2 mm) and L. plantarum LP3 and L. plantarum PTCC 1058 the lowest antifungal activities. The fermented creams contained Lactobacillus strains exhibited the highest and lowest antifungal activities against A. ochraceus and A. parasiticus, respectively. L. plantarum LU5, with the inhibition zone of 27.6 ± 0.9 mm, was the most effective fungal inhibitor, while L. plantarum PTCC 1058 had the lowest antifungal activity.     

Study of texture properties of ‘laba’ garlic in different color states and their change mechanisms

In addition to color, texture of ‘laba’ garlic can also change. To study texture properties of ‘laba’ garlic and their change mechanisms during pickling, texture, ultrastructures, membrane permeability, water status, wall polysaccharides contents, and enzyme activities of garlics in three color states (white, green, and yellow) were studied. The results showed that the firmness, fracturability, membrane permeability, malonaldehyde, protopectin, and chelator-soluble pectin contents, and cellulase and polygalacturonase activities of garlic decreased significantly (P < 0.05), while the water, cellulose, and water-soluble pectin contents, and pectin methylesterase (PME) activity increased after pickling. In addition, the more content of free water and less content of immobilised water were contained in ‘laba’ garlic than in fresh garlic owing to wall degradation and water translocation, especially in yellow garlic. In summary, texture changes in ‘laba’ garlic are mainly dependent on pectin composition, water status, and PME activity.     

Inhibition of biofilm formation,adhesion and invasion in Caco-2 cells by foodborne pathogens using phyto-mediated synthesised silver nanoparticles from industrial wastes

Certain foodborne diseases are associated with antibiotic resistance, a significant problem throughout the world. Silver nanoparticles (AgNPs) using industrial waste from Eucalyptus camaldulensis and sericin, a protein derived from Bombyx mori, were synthesised by a one-step approach. Spherical-shaped nanoparticles with the average size of 17.19 nm exhibited strong antioxidant activity. The minimum bactericidal concentrations against foodborne pathogens including Bacillus cereus, Listeria monocytogenes, Staphylococcus aureus, Escherichia coli O157:H7, Klebsiella pneumoniae, Salmonella Typhimurium, Shigella sonnei, Vibrio cholerae and Vibrio parahaemolyticus were between 2.96 and 11.83 µg/mL. Killing against L. monocytogenes and E. coli O157:H7 was observed within 4 h. Treatment with AgNPs at 0.25 – 0.5 × MIC significantly reduced biofilm production in all isolates (P < 0.05). AgNPs significantly impeded adhesion to and invasion of human epithelial Caco-2 cells by L. monocytogenes and E. coli O157:H7 (P < 0.05). Biocompatibility assessment of AgNPs with Caco-2 and human red blood cells demonstrated no toxic effects.