Assessing fungicide resistance in populations of Alternaria in Idaho potato fields

Early blight, caused by the fungus Alternaria solani and brown leaf spot, caused by Alternaria alternata, are important diseases of potato crops in Idaho. In recent years growers have reported a reduction in efficacy of fungicides traditionally used in the past decade to control early blight. In 2009, a collection of A. solani 39 isolates were screened for resistance to azoxystrobin, pyraclostrobin, boscalid and famoxadone. Fungicide sensitivity testing was done using spiral plate dilution gradients. Results showed that of 39 isolates screened, all were resistant to azoxystrobin and three were resistant to boscalid. None were resistant to pyraclostrobin or famoxadone. In summer 2010, more isolates were collected (9 A. alternata and 26 A. solani) and the survey was expanded to include more fungicides with four different modes of action that targeted succinate dehydrogenase (SDH), methionine biosynthesis, mitochondrial respiration and multi-site contact activity. New isolates of A. solani and A. alternata were also collected from two additional sites. The results showed that 57% of the isolates were resistant to boscalid as well as an average of 63% of the isolates being resistant to the strobilurin fungicides. Seven and 15% of isolates were resistant to penthiopyrad (an SDH inhibitor), and pyrimethanil (a methionine biosynthesis inhibitor), respectively. However, none of the isolates were resistant to fluopyram (an SDH inhibitor) or a mixture of fluopyram and pyrimethanil. Although there appears to be cross resistance developing in Alternaria spp. to some of the new SDH inhibitors like penthiopyrad, others such as fluopyram are still showing limited to no resistance development in Alternaria spp. in Idaho.     

Effect of non-aflatoxigenic strains of Aspergillus flavus on aflatoxin contamination of pre-harvest peanuts in fields in China

Aflatoxin contamination of peanuts is one of the most concerns in peanut production in China. Applying non-aflatoxigenic Aspergillus flavus strains, based on competitive exclusion, has been proved to be a promising strategy to reduce aflatoxin contamination in pre-harvest peanuts. Two non-aflatoxigenic A. flavus strains collected in China, which have been proved effectively reducing aflatoxin in the laboratory, were mixed with high aflatoxin producer to the soil in peanut growing season. The two non-aflatoxigenic strains significantly (P ​< ​0.05) reduced aflatoxin contamination in peanut kernels under both normal and drought stresses in two fields. Compared to control, the total aflatoxin (sum of aflatoxin B₁ and B₂) was reduced 26.7–99.12% in field 1, and 84.96–99.33% in field 2. The aflatoxin was reduced 84.96–99.33% under drought stress in two fields. The present study indicated the non-aflatoxigenic A. flavus strains could be potential biocontrol agents for reducing aflatoxin contamination under field condition.     

Heritability of,and genotypic correlations between,aflatoxin traits and physiological traits for drought tolerance under end of season drought in peanut (Arachis hypogaea L.)

More rapid progress in breeding peanut for reduced aflatoxin contamination should be achievable with a better understanding of the inheritance of, aflatoxin trait and physiological traits that are associated with reduced contamination. The objectives of this study were to estimate the heritability of aflatoxin traits and genotypic (r_G) and phenotypic (r_P) correlations between drought resistance traits and aflatoxin traits in peanut. One hundred-forty peanut lines in the F_4:6 and F_4:7 generations were generated from four crosses, and tested under well-watered and terminal drought conditions. Field experiments were conducted under the dry seasons 2006/2007 and 2007/2008. Data were recorded for biomass (BIO), pod yield (PY), drought tolerance traits [harvest index (HI), drought tolerance index (DTI) of BIO and PY, specific leaf area (SLA), and SPAD chlorophyll meter reading (SCMR)], and aflatoxin traits [seed infection and aflatoxin contamination]. Heritabilities of A. flavus infection and aflatoxin contamination in this study were low to moderate. The heritabilities for seed infection and aflatoxin contamination ranged from 0.48 to 0.58 and 0.24 to 0.68, respectively. Significant correlations between aflatoxin traits and DTI (PY), DTI (BIO), HI, biomass and pod yield under terminal drought conditions were found (r_P = −0.25** to 0.32**, r_G = −0.57** to 0.53**). Strong correlations between SLA and SCMR with A. flavus infection and aflatoxin contamination were also found. Positive correlations between SLA at 80, 90, and 100 DAP and aflatoxin traits were significant (r_P = 0.13** to 0.46**, r_G = 0.26** to 0.81**). SCMR was negatively correlated with aflatoxin traits (r_P = −0.10** to −0.40**, r_G = −0.11** to −0.66**). These results indicated that physiological-based selection approaches using SLA and SCMR might be effective for improving aflatoxin resistance in peanut.     

Fungal species isolated from peanuts in major Kenyan markets: Emphasis on Aspergillus section Flavi

A survey was conducted in Nairobi, Nyanza and Western provinces in Kenya between March and July 2009 with 1263 peanut products sampled out of which 705 samples underwent microbial analysis. The study aimed at determining the incidence of fungal species – emphasis on Aspergillus section Flavi – associated with peanut products. A 0.5 kg representative sample was obtained from each surveyed vendor and the colony forming units (CFU) of fungal species determined. The samples were also analyzed for total aflatoxin level while isolates of Aspergillus flavus and Aspergillus parasiticus were screened for production of aflatoxin B1, B2, G1 and G2. Eight fungal species were detected in the samples and were in decreasing order of CFU/g of sample: A. flavus S-strain (467), A. flavus L-strain (341), Penicillium spp. (326), Aspergillus niger (156), Aspergillus tamari (27), Aspergillus alliaceus (21), A. parasiticus (10), and Aspergillus caelatus (5). The overall incidence of A. flavus S-strain in samples from Nairobi was 92 and 1425% higher than samples from Nyanza and Western regions, respectively. The combined incidence of A. flavus and A. parasiticus was varied significantly (p ≤ 0.05) with peanut product: peanut flour (69%), shelled raw peanuts (53%), spoilt peanuts (49%), boiled podded peanuts (45%), podded peanuts (39%), peanut butter (31%), fried peanuts (22%) and roasted peanuts (20%). Seventy three percent of A. flavus and A. parasiticus isolates produced at least one of the aflatoxin types, with 66% producing aflatoxin B1. The total aflatoxin level among peanut products ranged from 0 to 1629 μg/g; and there was a positive correlation (r = 0.2711) between the incidence of A. flavus and A. parasiticus, and total aflatoxin level. The high incidence of aflatoxin producing fungi in peanuts traded in Kenyan markets implies a risk of aflatoxin contamination, highlighting the need for stakeholders to promote sound practices at all stages of the peanut value chain in order to minimize market access by non-complying products.     

Aflatoxin contamination of corn under different agro-environmental conditions and biocontrol applications

Biological control of the fungus Aspergillus flavus has been shown to be effective in reducing aflatoxin contamination in corn. This study compared field application of a bioplastic-based formulation for delivering atoxigenic A. flavus isolates in Northern Italy and the Mississippi Delta.Due to an extremely hot and dry summer at the Italy site in 2012, aflatoxin contamination was approximately seven times higher than in 2011. In 2011, and 2012, application of bioplastic granules inoculated with the atoxigenic isolate A. flavus NRRL 30797 at 15 and 30 kg ha⁻¹ resulted in a reduction of aflatoxin contamination by 67.2 ± 4.1% and 94.8 ± 5.3%, respectively. The higher application rate was also effective when soil abundance of A. flavus was artificially increased by applying contaminated corn residues. At the Mississippi site, summer 2012 was also hot and dry, with high levels of aflatoxin contamination. In fields planted with non-Bt or Bt hybrids, application of biocontrol granules inoculated with A. flavus NRRL 30797 or NRRL 21882 at 30 kg ha⁻¹ reduced aflatoxin contamination to up to 89.6%. Field experiments on two continents showed that bioplastic-based A. flavus formulations markedly reduced aflatoxin contamination under different agro-environmental conditions and infestation intensities.     

Ear rot,aflatoxin accumulation,and fungal biomass in maize after inoculation with Aspergillus flavus

Aflatoxin, a toxin produced by the fungus Aspergillus flavus Link: Fries, occurs naturally in maize (Zea mays L.). Aflatoxin is a potent human carcinogen and is also toxic to livestock, pets, and wildlife. When contaminated with aflatoxin, the value of maize grain is markedly reduced. This investigation was conducted to compare ear rot, aflatoxin accumulation, and fungal biomass in maize single crosses with varying degrees of resistance to aflatoxin accumulation and to determine the relative importance of general combining ability (GCA) and specific combining ability (SCA) in the inheritance of resistance to ear rot, aflatoxin accumulation, and fungal biomass. Eight germplasm lines with different levels of resistance to aflatoxin accumulation were used as parents of a diallel cross. The cross was evaluated for visible ear rot, aflatoxin accumulation, and A. flavus infection in the grain. A. flavus infection was determined by quantitative real-time polymerase chain reaction (qRT-PCR) assays. Both GCA and SCA were significant sources of variation in the inheritance of the three traits although GCA accounted for a greater portion of the variation among single crosses. The interactions of GCA and SCA with years were highly significant for aflatoxin accumulation, but not significant for A. flavus infection. Estimates of GCA effects were highly significant for both reduced A. flavus infection and reduced aflatoxin accumulation for Mp313E, Mp715, and Mp717. Conversely, GCA effects associated with GA209 were significant for reduced levels of A. flavus infection and ear rot, but high levels of aflatoxin accumulation. Mp313E, Mp715, and Mp717 should be useful in breeding programs targeting both reduced levels of fungal infection and aflatoxin accumulation.     

Production of monoclonal antibodies against AFLM (Ver-1), a middle key protein involved in aflatoxin biosynthesis

Aflatoxins are potent carcinogens, mutagens and teratogens, and are harmful to both humans and animals. As many as 30 genes are involved in aflatoxin biosynthesis. Among them, aflM (ver-1) gene was predicted to encode a 28-kDa NADPH-dependent ketoreductase (AFLM), which catalyzed middle enzymatic steps in aflatoxin biosynthetic pathway. AFLM (Ver-1) was proved to be necessary for conversion of versicolorin A (VERA) to demethylsterigmatocystin (DMST) in aflatoxin B₁ (AFB₁) biosynthesis. For these reasons, aflM gene was cloned and specific monoclonal antibodies for AFLM was developed to better define potential pathways of AFLM involved in AFB₁ biosynthesis. Monoclonal antibodies 11B2-1D7 and 3G5-4E7 were successfully screened out by immunizing mouse. Immunoblot analysis revealed that both had high sensitivity and specificity to identify native AFLM protein in A. flavus with detection limit of 11 ​ng/mL and 8 ​ng/mL respectively. These results showed that it was suitable for quantitative detection of AFLM in A. flavus isolate. Further investigation revealed that aflatoxin accumulations of various A. flavus were not dependent on AFLM biosynthesis. Overall, this is the first report for development for AFLM monoclonal antibody development and application in A. flavus quantitative detection.     

Geographical distribution of Aspergillus flavus in peanut harvest period in China

In order to grasp the distribution of Aspergillus fiavus in the soil of peanut production areas in China, A. fiavus biomarkers were tested on 555 soil samples from 37 sampling points in 17 provinces, peanut fields in four agroecological zones(Southern area, Yangtze River Basin, Northern area, Northeast area). The results showed that(1) the cultivation amount of A. fiavus per gram of soil in the Yangtze River Basin is 1.30 times that of the southern area, 1.56 times that of the northern area, and...     

Southwestern corn borer damage and aflatoxin accumulation in conventional and transgenic corn hybrids

Southwestern corn borer (Diatraea grandiosella Dyar) is a major pest of corn (Zea mays L.) in the southern United States. In addition to the direct yield losses caused by southwestern corn borer, larval feeding on developing ears provides a site for fungi to enter the ear. Aspergillus flavus Link: Fries infection and the subsequent accumulation of aflatoxin in corn grain are major limitations to profitable corn production in the southern United States. This investigation was conducted to determine the effectiveness of transgenic corn hybrids expressing the δ-endotoxin insecticidal (CryIAb) proteins isolated from Bacillus thuringiensis (Bt) in reducing southwestern corn borer damage and aflatoxin accumulation. Ear damage and aflatoxin accumulation were compared among 10 pairs of conventional nonBt and transgenic Bt corn hybrids following infestation with southwestern corn borer and inoculation with A. flavus using kernel-wounding and nonwounding techniques. Both nonBt and Bt hybrids exhibited high levels of aflatoxin accumulation when inoculated with a kernel-wounding technique. When inoculated with a non-wounding technique and infested with southwestern corn borer, aflatoxin accumulation was significantly higher in nonBt than Bt hybrids. Aflatoxin accumulation was also significantly higher for nonBt hybrids inoculated with A. flavus and infested with southwestern corn borer than for hybrids that were only inoculated with A. flavus. Southwestern corn borer larval establishment was significantly higher on nonBt hybrids than on Bt hybrids. Larval survival was extremely low on the Bt hybrids. The results of this investigation indicate that these Bt hybrids should be effective in reducing aflatoxin contamination in areas where high southwestern corn borer infestations occur. The reduced levels of aflatoxin accumulation associated with Bt hybrids are likely a consequence of reduced insect damage rather than resistance to A. flavus infection or aflatoxin accumulation per se.     

Drought,pod yield,pre-harvest Aspergillus infection and aflatoxin contamination on peanut in Niger

Soil moisture and soil temperature affect pre-harvest infection with Aspergillus flavus and production of aflatoxin. The objectives of our field research in Niger, West Africa, were to: (i) examine the effects of sowing date and irrigation treatments on pod yield, infection with A. flavus and aflatoxin concentration; and (ii) to quantify relations between infection, aflatoxin concentration and soil moisture stress. Seed of an aflatoxin susceptible peanut cv. JL24 was sown at two to four different sowing dates under four irrigation treatments (rainfed and irrigation at 7, 14 and 21 days intervals) between 1991 and 1994, giving 40 different ‘environments’. Average air and soil temperatures of 28–34 °C were favourable for aflatoxin contamination. CROPGRO-peanut model was used to simulate the occurrence of moisture stress. The model was able to simulate yields of peanut well over the 40 environments (r² = 0.67). In general, early sowing produced greater pod yields, as well as less infection and lower aflatoxin concentration. There were negative linear relations between infection (r² = 0.62) and the average simulated fraction of extractable soil water (FESW) between flowering and harvest, and between aflatoxin concentration (r² = 0.54) and FESW in the last 25 days of pod-filling. This field study confirms that infection and aflatoxin concentration in peanut can be related to the occurrence of soil moisture stress during pod-filling when soil temperatures are near optimal for A. flavus. These relations could form the basis of a decision-support system to predict the risk of aflatoxin contamination in peanuts in similar environments.     

Association between aflatoxin contamination and drought tolerance traits in peanut

The current study investigates the association between drought tolerance traits and aflatoxin contamination in peanut grown under long-term drought. Two field experiments were conducted at Khon Kaen University, Thailand using a split–split plot design with three drought stress levels as main plots, 11 genotypes as sub-plots, and two soil inoculations of Aspergillus flavus treatments as sub-sub-plots. The effects of temperature, soil moisture and A. flavus population on kernel colonization and aflatoxin contamination, and drought tolerance traits viz. specific leaf area (SLA) and root length density (RLD) were measured. The results demonstrated that elevated soil temperatures and reduced soil moisture, favored aflatoxin production. Drought in combination with higher levels of A. flavus inoculum load in the soil resulted in an increase in the fungal populations in the soil which in turn resulted in increased kernel colonization and subsequent aflatoxin contamination. A combination of SLA and RLD, and kernel colonization had a significant influence on aflatoxin contamination under drought conditions in both seasons (r = 0.73** and 0.76**). The results revealed that drought tolerance traits (SLA and RLD) could be contributing to resistance to aflatoxin contamination suggesting that a combination of SLA, RLD and kernel colonization could be used as selection criteria in selecting parents for aflatoxin resistance.     

Screening,identification and antagonistic effect of antagonistic bacteria JTFM1001 against aflatoxin contamination in corn

Aflatoxin is a strong carcinogenic and toxic fungal toxin produced by Aspergillus flavus and other Aspergillus species,and can seriously threaten the health of consumers,thus becoming a global concern.Corn,as an important oil and economic crop,is highly susceptible contaminated by aflatoxin.In this study,antagonistic bacteria with strong inhibitory effect on aflatoxin were screened to provide support for the treatment aflatoxin contamination control in corn.Ten strains which have strong antagonistic effects against A.flavus were isolated from healthy corn from different corn producing areas in China.Among them,the antagonistic bacteria JTFM1001 through corn kernels in vivo and field experiment,the inhibition effect of aflatoxin contamination reached above 70%and 55%,respectively.And the strain was identified as Bacillus subtilis based on its morphological,physiological and biochemical characteristics and phylogenetic analysis of 16S rDNA.In addition,our data showed that it can colonize in the rhizosphere and survive for a long time,forming the dominant flora,with broad application prospect.Finally,we were surprised to find that the antibacterial metabolites secreted by the antagonistic bacteria was one of the mechanisms of its inhibition of A.flavus and aflatoxin.This will provide us with new ideas and perspectives on the effective prevention and control of aflatoxin contamination in corn and corn oil.     

Diallel analysis of aflatoxin accumulation in maize

Since its discovery in numerous feedstuffs, aflatoxin, a carcinogenic compound produced by the fungus Aspergillus flavus Link ex Fries, has caused much concern among consumers and producers alike. This toxin poses a serious economic threat to maize (Zea mays L.) producers of the southeastern and midwestern regions of the United States. Efforts to identify maize germplasm that is resistant to aflatoxin accumulation and to investigate the genetic basis of this resistance have been undertaken at numerous research institutions. The objectives of this study were to (1) evaluate aflatoxin accumulation in grain harvested from maize inbred lines and a diallel cross among these lines, (2) determine the importance of general and specific combining abilities in inheritance of resistance to aflatoxin accumulation, and (3) estimate general and specific combining ability effects associated with resistance to aflatoxin accumulation in the inbred lines and crosses among them. Eight inbred lines and a diallel cross of the maize lines were inoculated with an A. flavus spore suspension 12–14 d after silk emergence. Following harvest, aflatoxin content was determined from samples of grain. Statistical analyses performed using SAS general linear models (GLM) and DIALLEL-SAS indicated that general and specific combining ability were significant sources of variation in the inheritance of resistance to aflatoxin accumulation. The inbred line Mp313E, which was developed and released as a source of resistance to aflatoxin contamination, had significantly lower aflatoxin accumulation than other lines. Mo18W exhibited excellent general combining ability for reduced aflatoxin accumulation when crossed with the other lines. Both Mp313E and Mo18W could be useful in breeding programs to develop aflatoxin-resistant maize hybrids. Mp339, SC212M, and Ab24E demonstrated aflatoxin susceptibility as both inbreds and in single crosses.     

Improvement in biocontrol activity of Bacillus subtilis UTB1 against Aspergillus flavus using gamma-irradiation

Bacillus subtilis UTB1, a biocontrol bacterium isolated from Iranian pistachio nuts, has revealed to have antagonistic activity against aflatoxin-producing Aspergillus flavus R5. The strain UTB1 produces lipopeptide compounds and is able to degrade aflatoxin B1. In this study, a random mutagenesis generated using different doses of gamma irradiation (0.1–3 KGy) was applied on B. subtilis UTB1 to improve its antagonistic activity against A. flavus R5. Five hundred bacterial colonies were selected randomly after irradiation, and their effects against A. flavus R5 were assessed in a plate assay. Forty-five colonies (9%) exhibited higher inhibition activity as compared to the non-irradiated wild type. Eight colonies out of the 45 were selected based on different polymorphism patterns obtained by repetitive element sequence polymorphism-PCR (ERIC and BOX) analyses; six of which could significantly inhibit the fungal growth utilizing washed cells and cell-free supernatants as compared to the parental strain. According to thin-layer chromatograms, the production of lipopeptides including surfactin, fengycin and iturin families increased in these six mutants. A considerable inhibition of the fungal growth was observed using bioautography analysis, which associated with iturins production. A. flavus sporulation and aflatoxin content decreased significantly in pistachio nuts treated with mutants M419 and M464 as compared to the strain UTB1. These results suggest that both mutants M419 and M464 could be promising biocontrol candidates against A. flavus in pistachio nuts.     

Puroindoline genes introduced into durum wheat reduce milling energy and change milling behavior similar to soft common wheats

Grain physical characteristics and milling behavior of a durum wheat line in which both wild-type puroindoline genes were translocated and stabilized after backcrossing (Svevo-Pin) were compared with the parent line (Svevo). The only observed differences between grain characteristics were the mechanical resistance and starchy endosperm porosity revealed through vitreosity measurement. A significant increase of flour and a decrease of semolina yield and break milling energy were observed from Svevo-Pin in comparison with the non-recombinant parent line in accordance to the lower grain mechanical resistance and higher porosity measurements. Moreover, the particle size distribution shown for Svevo-Pin flour appeared consistent with a lower adhesion between starch granules and the protein matrix attributed to the presence of wild-type puroindolines. Coarse bran yield was conversely increased. This appeared to be due to a lower starchy endosperm recovery as a higher proportion of grain starch was found in this bran fraction. Flour from the durum parent line was inversely enriched in phytic acid, a cellular marker of the aleurone layer. Starch damage was also lower in Svevo-Pin flours in comparison with Svevo. All of the observed differences between translocation and parent lines were confirmed independent of the culture growth conditions (n = 12).     

Diallel analysis of fumonisin accumulation in maize

Contamination of maize (Zea mays L.) grain with aflatoxin, produced by the fungus Aspergillus flavus Link: Fries, or fumonisin, produced by the fungus Fusarium verticillioides (Saccardo) Nirenberg (Syn F. moniliforme Sheldon), greatly reduces its value and marketability. Host resistance to fungal infection and mycotoxin accumulation is considered a highly desirable way to reduce losses of both aflatoxin and fumonisin. Maize germplasm lines with resistance to aflatoxin accumulation have been developed in Mississippi. Three of the aflatoxin-resistant lines and five additional lines were used as parents to produce a diallel cross. The diallel cross was evaluated for fumonisin accumulation in field tests conducted in Mississippi in 2007 and 2008. General combining ability (GCA) was a highly significant source of variation in both 2007 and 2008, but specific combining ability (SCA) was not significant. GCA effects for reduced fumonisin levels were highly significant for Mp715, Mp717, and GA209. Mp715 and Mp717 were developed and released as sources of resistance to aflatoxin contamination and exhibit resistance as inbred lines and in hybrid combinations. These lines should be useful in developing maize lines and hybrids with resistance to both fumonisin and aflatoxin accumulation in grain.     

Development of new bioformulations using Trichoderma and Talaromyces fungal antagonists for biological control of sugar beet damping-off disease

The aims of this study were to develop new bioformulations using Trichoderma harzianum, Trichoderma asperellum, and Talaromyces flavus and some organic and inorganic carriers and evaluate their effects against Rhizoctonia solani, the fungal causal agent of sugar beet seedling damping-off disease. Selected fungal isolates were first re-cultured and maintained on potato dextrose agar (PDA) culture medium. Antagonistic effects of eight isolates of the above-mentioned antagonistic fungi were then evaluated against R. solani, through volatile metabolites and non-volatile metabolites production mechanisms under laboratory conditions. In volatile and non-volatile metabolite experiments, five and seven isolates caused significant reduction in R. solani growth respectively. Based on the results of laboratory experiments, the most effective antagonistic isolates (one isolate from each species) were selected for development of nine bioformulations using peat, rice bran and talc as carriers. The effectiveness of developed bioformulations was then evaluated in controlling sugar beet damping-off disease in a greenhouse experiment where sugar beet seeds were coated with bioformulations and were sown in pasteurized field soil pre-inoculated with R. solani. Results of the greenhouse experiment 60 days after sowing showed that all bioformulations increased the number of healthy seedlings significantly (compared to the untreated control) with different rates. According to the results, the most effective bioformulation was Talc-T. harzianum followed by Peat-T. flavus, Talc-T. flavus and Rice bran-T. harzianum. In general, in both laboratory and greenhouse experiments, T. flavus was the most effective fungal antagonist followed by T. harzianum and T. asperellum. Based on the results of this study it is concluded that Trichoderma and Talaromyces employing different mechanisms might be potential biocontrol agents for controlling R. solani-induced sugar beet damping-off disease.     

Prevalence of isolates of Botrytis cinerea resistant to multiple fungicides in Chilean vineyards

Gray mold (Botrytis cinerea) is an important disease of grapevines (Vitis vinifera) and requires several fungicide treatments to achieve a satisfactory control in Chile. Furthermore, a high resistance risk has developed because of the extensive use of specific fungicides with a single-site mode of action. The aim of this study was to determine the presence of resistance to the multiple fungicides currently used against B. cinerea in Chile. During 2007–2009 and 2011, 214 isolates of B. cinerea from 36 commercial vineyards were examined for sensitivity to anilinopyrimidines (cyprodinil or pyrimethanil), demethylation inhibitors (DMIs) (tebuconazole), hydroxyanilides (fenhexamid) and phenylpyrroles (fludioxonil). Of all of the isolates, 62.7% were resistant to anilinopyrimidines, 7.2% to DMIs, 27.1% to hydoxyanilides and 44.8% to phenylpyrrole derivates. Overall, 16 (12.3%) of the isolates were sensitive; although none was resistant to all four of the fungicide classes, and 50 (38.5%), 51 (39.2%), and 13 (10.0%) isolates were resistant to one, two and three fungicides, respectively. In vitro, resistant isolates were capable of causing fruit rot on wounded apples pretreated with any one of the five fungicides at label concentrations. To the best of our knowledge, this is the first report of isolates having simultaneous resistance to anilinopyrimidines, DMIs, phenylpyrroles and hydroxyanilides in B. cinerea populations from grapevines in Chile. Therefore, fungicide resistance is a serious problem that questions the sustainability of the current gray mold control strategy, which relies almost exclusively on fungicides with single-site modes of action.     

Aflatoxin in maize,a multifaceted answer of Aspergillus flavus governed by weather,host-plant and competitor fungi

Aspergillus section Flavi is able to produce aflatoxins (AFs) in vitro down to 0.85 a_w with a potential maximum occurring between 0.95 and 0.99 a_w, while in the field AFs increased significantly with kernel humidity below 0.95 a_w. In order to clarify this apparent discrepancy, a 3-year field trial with artificial inoculation of maize ears with Aspergillus flavus strains was organised. The co-occurrence of Aspergillus section Flavi and Gibberella fujikuroi species complex (Gfsc) was observed. The incidence of A. flavus was significantly influenced by the year and negatively related to Gfsc incidence. In 2012, when the highest temperature and the lowest rain were registered, aflatoxin B₁ (AFB₁) content was the highest and a_w < 0.95 in kernels was measured early throughout the growing season. In 2013 and 2014, the temperature was lower and rain more abundant, and a_w decreased below 0.95 only close to harvest and AFB₁ contamination was limited. The possibility of describing/predicting reasonably well a_w dynamic based on temperature, (degree day) was confirmed. With a_w > 0.95, a positive correlation between AFB₁ production rate and a_w was found, but a negative correlation resulted with a_w < 0.95. Other factors than a_w play a role, but a_w = 0.95 should be considered as an indicator of suitable conditions for rapid aflatoxin accumulation.