Antifungal activity of Euphorbia species against moulds responsible of cereal ear rots.

AIMS: This work aimed to identify secondary metabolites from aerial parts of Euphorbia species functional for control of toxigenic Fusarium species responsible of cereal grain rots. METHODS AND RESULTS: Aerial parts of Euphorbia serpens, Euphorbia schickendantzii and Euphorbia collina were sequentially extracted with hexane, ethyl acetate and methanol. The extracts were tested against strains of Fusarium verticillioides and Fusarium graminearum by microdilution tests. The hexane extract of E. collina provided the lowest IC50 s on both fungal species. Further fractionation showed that cycloartenol (CA) and 24-methylenecycloartanol are associated to the moderate inhibitory effect of the hexane extract on fungal growth.Sublethal concentrations of CA and 24MCA blocked deoxynivalenol (DON) and fumonisins production.CA and 24MCA co-applied with potassium sorbate, a food preservative used for Fusarium control, synergized the growth inhibition of fungi. The mixtures reduced mycotoxins accumulation when applied at sublethal concentrations. CONCLUSIONS: CA and 24MCA inhibited both fungal growth and mycotoxins production. This fact is an advantage respect to potassium sorbate which increased the mycotoxins accumulation at sublethal concentrations. SIGNIFICANCE AND IMPACT OF THE STUDY: CA and 24MCA synergized potassium sorbate and their mixtures offer a lower mycotoxigenic risk than potassium sorbate for control of the Fusarium species.

Volatiles mediate host-selection in the corn hoppers Dalbulus maidis (Hemiptera: Cicadellidae) and Peregrinus maidis (Hemiptera: Delphacidae).

Volatile organic compounds (VOCs) released by plants are generally involved in host recognition and host selection for many phytophagous insects. However, for leafhoppers and planthoppers, host recognition is mainly thought to involve a phototactic response, but it is not clear if a host plant could be selected based on the volatile cues it emits. In this study we evaluated olfactory responses in dual choice tests of two Hemiptera species, Dalbulus maidis (De Long) (Cicadellidae) and Peregrinus maidis (Ashmead) (Delphacidae), vectors of maize-stunting diseases, to three maize (Zea mays L.) germplasms, a temperate and a tropical hybrid and a landrace. VOCs emitted by the germplasms were collected and identified using gas chromatography-mass spectrometry. The temperate hybrid released significantly more VOCs than the tropical hybrid and the landrace, and its volatile profile was dominated by (±)-linalool. D. maidis preferred odours emitted from the temperate hybrid, whereas P. maidis preferred odours from the tropical hybrid and the landrace over the temperate one. In order to test if linalool plays a role in the behavioural responses, we assayed this compound in combination with the tropical hybrid, to provide other contextual olfactory cues. D. maidis was attracted to the tropical hybrid plus a 0.0001% linalool solution, indicating that this compound could be part of a blend of attractants. Whereas addition of linalool resulted in a slight, though not significant, reduction in host VOC attractiveness for P. maidis. Both hopper species responded to olfactory cues in the absence of supplementary visual cues.

Nutraceutical Properties of Herbal Infusions from Six Native Plants of Argentine Patagonia.

Six native plants of South America traditionally consumed in the Patagonian region (southern Argentina and Chile), namely: Adesmia boronioides Hook. f., Apium australe Thouars, Buddleja globosa Hope, Drimys andina (Reiche) R. Rodr. & Quezada, Dysphania multifida L. and Solidago chilensis Meyen were investigated to determine the nutraceutical properties of infusions of their aerial parts. The infusions were characterized in terms of their antioxidant activity, phenolic and flavonoid content, profile of phenolic compounds, general toxicity and cytotoxicity on two different human cell lines: T84 (derived from colon cancer) and HTR8/SVneo (not derived from cancer). Twenty-nine compounds, mainly phenolic acids and flavonoids, were identified. This is the first analysis of phenolic compounds in infusions from native plants of Patagonia. D. andina, B. globosa and S. chilensis showed high levels of antioxidants, even higher than those of Green Tea. The content of phenolic compounds correlated significantly with the antioxidant activity of the samples analyzed. The toxicity test indicated that the use of A. australe, B. globosa and D. multifida seems safe, but a moderate consumption is suggested for A. boronioides, D. andina and S. chilensis until more exhaustive and long-term results are available. Moreover, A. boronioides and S. chilensis showed anticancer potential due to their antiproliferative activity on human cancer cell lines.

Effect of guaianolides in the meiosis reinitiation of amphibian oocytes.

Sesquiterpene lactones (STLs) are a large and structurally diverse group of plant metabolites generally found in the Asteraceae family. STLs exhibit a wide spectrum of biological activities and it is generally accepted that their major mechanism of action is the alkylation of the thiol groups of biological molecules. The guaianolides is one of various groups of STLs. Anti-tumour and anti-migraine effects, an allergenic agent, an inhibitor of smooth muscle cells and of meristematic cell proliferation are only a few of the most commonly reported activities of STLs. In amphibians, fully grown ovarian oocytes are arrested at the beginning of meiosis I. Under stimulus with progesterone, this meiotic arrest is released and meiosis progresses to metaphase II, a process known as oocyte maturation. There are previous records of the inhibitory effect of dehydroleucodin (DhL), a guaianolide lactone, on the progression of meiosis. It has been also shown that DhL and its 11,13-dihydroderivative (2H-DhL; a mixture of epimers at C-11) act as blockers of the resumption of meiosis in fully grown ovarian oocytes from the amphibian Rhinella arenarum (formerly classified as Bufo arenarum). The aim of this study was to analyze the effect of four closely related guaianolides, i.e., DhL, achillin, desacetoxymatricarin and estafietin as possible inhibitors of meiosis in oocytes of amphibians in vitro and discuss some structure-activity relationships. It was found that the inhibitory effect on meiosis resumption is greater when the lactone has two potentially reactive centres, either a α,ß-α',ß'-diunsaturated cyclopentanone moiety or an epoxide group plus an exo-methylene-γ-lactone function.

Antifungal and antimycotoxigenic metabolites in Anacardiaceae species from northwest Argentina: isolation, identification and potential for control of Fusarium species.

AIMS: The purpose of this research was to identify antifungal compounds from leaves of Schinus and Schinopsis species useful for the control of toxigenic Fusarium species responsible of ear rot diseases. METHODS AND RESULTS: Leaves of Schinopsis (S. lorentzii and S. haenkeana) and Schinus (S. areira, S. gracilipes and S. fasciculatus) were sequentially extracted with dichloromethane, ethyl acetate and methanol. The antifungal activity of the fraction soluble in methanol of these extracts (fCH2Cl2, fAcEt and fMeOH, respectively) was determined by the broth microdilution method and the disc-diffusion method. The minimum inhibitory dose (MID), the diameter of growth inhibition (DGI) and the minimum concentration for 50% inhibition of fungal growth (MIC50) were calculated. The fCH2Cl2 and fAcEt of the Schinopsis species had the lowest MID and MIC50 values and the highest DGI. The antifungal compounds were identified as lupeol and a mix of phenolic lipids. The last one had the highest antifungal activity with MIC50 31-28 µg g(-1) and 165-150 µg g(-1) on Fusarium graminearum and Fusarium verticillioides, respectively. The identified metabolites completely inhibited fumonisin and deoxynivalenol production at lower concentrations than ferulic acid, a natural antimycotoxigenic compound. CONCLUSIONS: It was proven that lupeol and phenolic lipids were inhibitors of both fungal growth and mycotoxin production of toxigenic Fusarium species. This fact is specially interesting in the control of the toxigenic Fusarium species because several commercial antifungals showed to stimulate mycotoxin biosynthesis at sublethal concentrations. SIGNIFICANCE AND IMPACT OF THE STUDY: Control of toxigenic Fusarium species requires compounds able to inhibit both fungal growth and mycotoxin production. Our results suggest that the use of lupeol as food preservative and the phenolic lipids as fungal growth inhibitors of F. verticillioides and F. graminearum did not imply an increase in mycotoxin accumulation.

Toxigenic potential of Fusarium graminearum isolated from maize of northwest Argentina.

Twenty six isolates of Fusarium graminearum from grains of maize hybrids harvested in ±west Argentina were grown on autoclaved rice grain to assess their ability to produce type B trichothecenes. Chemical analysis indicated that 38% of isolates were nivalenol (NIV) producers only, 31% were major NIV producers with high DON(deoxynivalenol)/NIV ratios, 8% were major DON producers with minor NIV production, and 23% were DON producers only. Isolates showed a high variability in their toxigenic potential which was not related to fungal biomass. The distribution of the different chemotypes as well as the high and the low trichothecene-producing Fusarium isolates could not be associated to a geographical origin. Our results confirmed for the first time that isolates of Fusarium graminearum from maize of northwest Argentina are able to produce DON and NIV. A substancial contamination with both NIV and DON is likely in maize from northwest Argentina. Their contents should be quantified in regional surveillances for mycotoxin contamination.

Development and validation of a micromethod for fast quantification of 5-n-alkylresorcinols in grains and whole grain products.

A 96-well plate micromethod was developed to measure 5-n-alkylresorcinols (5nARs) in cereal grains and food derived products. The 5nARs reacted in alkaline alcoholic medium with Fast Blue RR ½ZnCl2 salt to yield coloured azo-derivatives. The highest sensitivity for 5nARs was obtained at 490 nm with 0.025% ethanolic Fast Blue RR and 5% K2CO3. This reaction showed good linearity for olivetol (0.05-0.20 µg). Contents of 5nARs determined in cereal grains and derived products by the new Fast Blue RR micromethod were highly correlated (R(2)=0.9944) with those obtained by a Fast Blue B method currently used. A Bland-Altman analysis indicated a small positive bias near to zero (R(2)=0.0401), suggesting that the methods can be interchangeably used. The new reaction is completed in 15 min and the coloured products are read within the 15 min after completion. The micromethod offers a fast analysis of 5nARs in cereal grains and derived products with low consumption of reagents and solvents.

Toxigenic potential of Fusarium graminearum isolated from maize of northwest Argentina

Twenty six isolates of Fusarium graminearum from grains of maize hybrids harvested in ±west Argentina were grown on autoclaved rice grain to assess their ability to produce type B trichothecenes. Chemical analysis indicated that 38% of isolates were nivalenol (NIV) producers only, 31% were major NIV producers with high DON(deoxynivalenol)/NIV ratios, 8% were major DON producers with minor NIV production, and 23% were DON producers only. Isolates showed a high variability in their toxigenic potential which was not related to fungal biomass. The distribution of the different chemotypes as well as the high and the low trichothecene-producing Fusarium isolates could not be associated to a geographical origin. Our results confirmed for the first time that isolates of Fusarium graminearum from maize of northwest Argentina are able to produce DON and NIV. A substancial contamination with both NIV and DON is likely in maize from northwest Argentina. Their contents should be quantified in regional surveillances for mycotoxin contamination.

Antimicrobial activities of sesquiterpene lactones and inositol derivatives from Hymenoxys robusta.

Six compounds from the aerial parts of the Argentinean plant Hymenoxys robusta (Rusby) Parker were isolated and their structures elucidated using extensive spectroscopic analyses. These compounds comprise two inositol derivatives and four 3,4-seco-pseudoguaianolides, including vermeerin. Bioactivity assays of these compounds against bacterial and fungal pathogens showed that only vermeerin possessed antimicrobial activity specific against Staphylococcus aureus, and showed no toxicity when exposed to human-derived macrophages.

Effect of biosynthetic intermediates and citrate on the phenyllactic and hydroxyphenyllactic acids production by Lactobacillus plantarum CRL 778.

AIM: To evaluate the influence of biosynthetic precursors, intermediates and electron acceptors on the production of antifungal compounds [phenyllactic acid (PLA) and hydroxyphenyllactic acid (OH-PLA)] by Lactobacillus plantarum CRL 778, a strain isolated from home-made sourdough. METHODS AND RESULTS: Growth of fermentative activity and antifungal compounds production by Lact. plantarum CRL 778 were evaluated in a chemically defined medium (CDM) supplemented with biosynthetic precursors [phenylalanine (Phe), tyrosine (Tyr)], intermediates [glutamate (Glu), alpha-ketoglutarate (α-KG)] and electron acceptors [citrate (Cit)]. Results showed that the highest PLA production (0.26 mmol l(-1)), the main antifungal compound produced by Lact. plantarum CRL 778, occurred when greater concentrations of Phe than Tyr were present. Both PLA and OH-PLA yields were increased 2-folds when Cit was combined with α-KG instead of Glu at similar Tyr/Phe molar ratio. Similarly, glutamate dehydrogenase (GDH) activity was significantly (P < 0.01) stimulated by α-KG and Cit in Glu-free medium. CONCLUSION: Phe was the major stimulant for PLA formation; however, Cit could increase both PLA and OH-PLA synthesis by Lact. plantarum CRL 778 probably due to an increase in oxidized NAD(+). This effect, as well as the GDH activity, was enhanced by α-KG and down regulated by Glu. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study where the role of Glu and GDH activity in the PLA and OH-PLA synthesis was evidenced in sourdough lactic acid bacteria (LAB) using a CDM. These results contribute to the knowledge on the antifungal compounds production by sourdough LAB with potential applications on the baked goods.

Species diversity and toxigenic potential of Fusarium graminearum complex isolates from maize fields in northwest Argentina.

Members of the Fusarium graminearum species complex (Fg complex) are the causal agents of ear rot in maize and Fusarium head blight of wheat and other small grain cereals. The potential of these pathogens to contaminate cereals with trichothecene mycotoxins is a health risk for both humans and animals. A survey of ear rot isolates from maize collected in northwest Argentina recovered 66 isolates belonging to the Fg complex. A multilocus genotyping (MLGT) assay for determination of Fg complex species and trichothecene chemotypes was used to identify 56 of these isolates as F. meridionale and 10 isolates as F. boothii. F. meridionale was fixed for the nivalenol (NIV) chemotype, and all of the F. boothii isolates had the 15-acetyldeoxynivalenol (15ADON) chemotype. The results of genetic diversity analysis based on nine variable number tandem repeat (VNTR) loci supported the hypothesis of genetic isolation between F. meridionale and F. boothii, and provided little evidence of geographic substructure among populations of the dominant pathogen species, F. meridionale. This is the first study to indicate that F. meridionale and F. boothii may play a substantial role in the infection and trichothecene contamination of maize in Argentina. In addition, dominance of the NIV chemotype among Fg complex isolates from Argentina is unprecedented, and of significant concern to food safety and animal production.

A molecular based strategy for rapid diagnosis of toxigenic Fusarium species associated to cereal grains from Argentina.

Fusarium species are worldwide causal agents of ear rot in cereals. Their toxigenic potential is a health risk for both humans and animals. In Argentina, most identification of these fungi has been based on morphological and cross-fertility criteria which are time consuming and require considerable expertise in Fusarium taxonomy and physiology. DNA based approaches have been reported as rapid, sensitive and specific alternatives to identify the main fumonisin and trichothecene-producing Fusarium species. In this work, we used PCR assays and the partial sequence of TEF1-alpha gene (Translation Elongation Factor-1 alpha) to identify the fumonisin and trichothecene-producing species in Fusarium isolates from diverse regions of Argentina. The relative efficiency and reliability of those methods to improve mycotoxin risk prediction in this country were also assessed. Species-specific PCR assays were targeted toward multicopy IGS (Intergenic Spacer of rDNA units) and on the toxin biosynthetic genes FUM1 (fumonisins) and TRI13 and TRI7 genes (trichothecenes). PCR assays based on FUM1 gene and IGS sequences allowed detection and discrimination of the fumonisin producers Fusarium proliferatum and Fusarium verticillioides. Molecular identification of nonfumonisin producers from Gibberella fujikuroi species complex was possible after determination of TEF1-alplha gene sequences, which indicated the presence of Fusarium subglutinans, Fusarium andiyazi and Fusarium thapsinum. TEF-1 alpha gene sequences also allowed discrimination of the different species of the Fusarium graminearum complex (F. graminearum sensu lato) as F. graminearum sensu stricto, Fusarium meridionale and Fusarium boothii. The last two species belonged to NIV chemotype and were detected for the first time in the subtropical region of Argentina while F. graminearum sensu stricto was DON producer only, which was also confirmed by specific PCR assays based on TRI137/TRI7 genes. Our results indicated that the PCR assays evaluated in this work are reliable diagnostic tools to detect the main toxigenic Fusarium species associated to cereal grains in Argentina. An extensive epidemiological survey based on the approach presented in this work is currently in progress to know the mycotoxigenic hazard of Fusarium species in cereal grains from the subtropical region of Argentina.

Sesquiterpene lactones from Centaurea tweediei.

Aerial parts of Centaurea tweediei from Argentina afforded as the main constituent the sesquiterpene lactone onopordopicrin and minor amounts of a new heliangolide, a new guaianolide, a new eudesmanolide, a new eudesmane acid and the lignans arctigenin and matairesinol.

Sesquiterpene lactones and other constituents of Disynaphia multicrenulata from Argentina.

Flowers and leaves of Disynaphia multicrenulata from Argentina afforded a large number of known germacradienolides and heliangolides, a new germacradienolide, a known melampolide, a new parthenolide derivative, one known and seven new cronquistiolides, a new 4Z-melampolide, three known and five new eudesmanolides, a new elemadienolide, a known grazielolide, an isoguaiagrazielolide, two diepoxygermacran-8,12-olides, three common kauranoic acids, pinoresinol, jaceosidin and the sesquiterpene oplopanone. Structures were elucidated by high-field 1H NMR spectrometry. The relationship to the known chemistry of other members of the Disynaphiinae is discussed.