Infected Wheat Grains Research Articles

Fusarium langsethiae – a HT‐2 and T‐2 Toxins Producer that Needs More Attention

AbstractFusarium langsethiae is a toxigenic fungus that was formally described as a new species in 2004. This fungus was first detailed in the 1990s but was initially referred to as ‘powdery Fusarium poae’ having a spore morphology similar to F. poae but a mycotoxin profile like that of Fusarium sporotrichioides. The species has been isolated from infected oat, wheat and barley grains but has been reported as more problematic in the former crop rather than the latter two. Whilst the epidemiology of F. langsethiae remains unclear, the fungus has been shown to produce high levels of type‐A trichothecenes HT‐2 and T‐2 toxins in small‐grain cereals. HT‐2 and T‐2 toxins are two of the most potent trichothecenes capable of inhibiting protein synthesis in eukaryotes. In this regard, mycotoxin contamination caused by F. langsethiae is clearly a food and feed safety hazard. With the European Commission considering legislation of HT‐2 and T‐2 toxins, more information is required not only on the producer and conditions favouring mycotoxin production, but also on reliable methods of pathogen detection and reduction of cereal contamination. This review describes recent research concerning the known epidemiology of F. langsethiae and suggestions of what needs to be known about the fungus in order to be able to understand and employ measures for preventing its infection and contamination of cereals with HT‐2 and T‐2 toxins.

Utilization of DNA microarrays for detection and identification of selected Fusarium species from the Czech Republic

Fusarium is a serious phytopathogenic fungal genus with producting of many kinds of highly toxic secondary metabolites – mycotoxins. The consumption of Fusarium contaminated food and feed can cause dangerous mycotoxicoses both in humans and animals, therefore the detection of a wider range of Fusarium species in the samples of crops is very important. The aim of our work was to test the reliability of detection and identification of three Fusarium species in infected wheat grains by DNA microarrays versus classical mycological methods and by specific PCR. The in-house DNA microarrays for the detection and identification of the selected Fusarium species by using oligonucleotides probes were prepared. For hybridisation on DNA microarrays fluorescent labelled PCR products were used of part of the translation elongation factor 1 alpha. The conditions of hybridisation were optimised on fungal template DNA. The method of DNA microarrays was verified on artificially infected samples of wheat and tested on unknown infected wheat samples with simultaneous analysis by classical mycological methods and by specific PCR.

Differential detection of potentially hazardous Fusarium species in wheat grains by an electronic nose.

Fungal infestation on wheat is an increasingly grave nutritional problem in many countries worldwide. Fusarium species are especially harmful pathogens due to their toxic metabolites. In this work we studied volatile compounds released by F. cerealis, F. graminearum, F. culmorum and F. redolens using SPME-GC/MS. By using an electronic nose we were able to differentiate between infected and non-infected wheat grains in the post-harvest chain. Our electronic nose was capable of distinguishing between four wheat Fusaria species with an accuracy higher than 80%.

Acoustic sensing of deoxynivalenol in co-occurrence with zearalenone and T-2/HT-2 toxin in winter wheat cultivar Sirvinta from Lithuania

A fast acoustic screening method in combination with an ELISA method was applied for the detection of deoxynivalenol (DON) in co-occurrence with some trichothecenes such as zearalenone (ZEA) and T-2/HT-2 toxins in winter wheat grains (Triticum aestivum L.) of the most popular Lithuanian cultivar Sirvinta. Strong correlations between the amplitude of the acoustic signal that penetrated the wheat samples and DON as well as total amount of mentioned trichothecenes quantified by ELISA method were found (R2=0.787 and R2=0.794, respectively). It turned out that weak correlations between the acoustic signal and the concentrations of ZEA and T-2/HT-2 were found. These correlations suggest that DON is the marker of the Fusarium infected wheat grain. This statement is also strengthened by the fact that DON is the most significant trichothecene contaminant of cereal grains in the world and levels are generally higher in the wetter climates. The dependency determined between the acoustic signal and structural physical characteristics (1000-kernel-weight) of the grain matrix could therefore be the basis for the developed rapid acoustic method for cereal quality and safety control. Additional examination of microstructure of wholesome and Fusarium-affected kernels under the scanning electron microscope revealed that obvious damage to the starch granules and lack of protein matrix was caused by the increase in several enzyme activities in the infected grains. Moderate positive relationships between α-amylase, protease and xylanase activities, and the degree of Fusarium infection were found (for α-amylase R2=0.623, for protease R2=0.823, and for xylanase R2=0.682). An intriguing characterisation may contribute to explanations as to how and to what extent wheat starch granules of different sizes stay more or less intact by slightly infected grains while the wheat starch granules are damaged and/ or completely destroyed by seriously infected grains. The changes of the protease activity had a slightly detectable influence on the crude protein content in seriously infected grain samples which might have an effect on dough properties and the quality of bread.

The effects of fungicides on Fusarium spp. and their associated mycotoxins in naturally infected winter wheat grain

The effects of fungicides on Fusarium spp. and their associated mycotoxins in naturally infected winter wheat grain Field trials conducted at the Institute of Agriculture, Lithuanian Research Centre for Agriculture and Forestry (central part of Lithuania) in 2009 were aimed to evaluate the effect of fungicides on Fusarium Head Blight (FHB) in a naturally infected field. A single application of dimoxystrobin + epoxiconazole (Swing Gold), prothioconazole (Proline), metconazole (Juventus), tebuconazole (Folicur), prothioconazole + tebuconazole (Prosaro) was applied to winter wheat cv. ‘Zentos’ at the manufacturer's recommended doses at anthesis (BBCH 65). The FHB incidence and severity were assessed at milk and hard maturity stages. The percentage of Fusarium infected grain and deoxynivalenol (DON), zearalenone (ZEN) and T-2 toxin (T-2) concentrations in harvested grain were determined. In all fungicide treated plots a significant reduction of FHB incidence and severity was determined; however the fungicides did not exert any effect on the amount of Fusarium-infected grain as compared with the untreated control. A reduction of DON, ZEN and T-2 contents in grain was determined in tebuconazole treatments. Fusarium avenaceum (Fr.) Sacc, F. culmorum (W. G. Sm.) Sacc., F. poae (Peck) Wollenw, F. sporotrichioides Sherb. and F. tricinctum (Corda) Sacc were identified in wheat grain, F. poae was prevalent.

On-farm experiments over 5 years in a grain maize/winter wheat rotation: effect of maize residue treatments on Fusarium graminearum infection and deoxynivalenol contamination in wheat

Over the course of 5 years, different maize residue treatments were conducted on 14 zero tillage on-farm sites in Switzerland to evaluate their effect on the development of Fusarium head blight (FHB) and the contamination with the mycotoxin deoxynivalenol (DON) in winter wheat grains and wheat straw following grain maize. Two experimental series with three and five different treatments were carried out, respectively. Fusarium graminearum (Schwabe) was the predominant FHB-causing species with an overall incidence of 15% infected wheat grains. A significant correlation between symptoms in the field, F. graminearum incidence and DON content in wheat grains and wheat straw was observed. The average DON content in both wheat grains and wheat straw was approximately 5,000 μg/kg and thus several times higher than the European maximum limit of 1,250 μg/kg for unprocessed small-grain cereals for human consumption. Of all grain samples, 74% were above the maximum limit. Pooled over both experimental series, the average reduction of DON in grains through treatments of the maize residue compared with a control treatment ranged between 21 and 38%. The effect of various other factors, including the year, the wheat variety, the site, the maize hybrid and the production system was evaluated as well. The year and the wheat variety were the most important FHB influencing factors. Over all treatments, the variety Levis showed a fivefold higher average DON content compared with the variety Titlis. From different categories of maize residue particles, intact pieces of 5-15 cm length were strongly correlated with F. graminearum incidence and DON content in grains. During the time course of this study, the recommendation from a preliminary version of the internet-based DON forecasting system FusaProg to apply or to omit a fungicide treatment was correct in 32 out of 42 cases. The results are currently being used to optimise the FusaProg models. This study has shown that in a grain maize/winter wheat rotation, the DON content in wheat grains frequently exceeded the European maximum limit, even with a thorough treatment of maize residues and less susceptible wheat varieties. Hence, in order to reduce the contamination risk in a zero tillage system, the crop rotation needs to be modified.

Fusarium head blight and associated mycotoxin occurrence on winter wheat in Luxembourg in 2007/2008

Fusarium head blight (FHB) is among the major causes of reduced quality in winter wheat and its products. In addition, the causal fungi produce a variety of toxins. A relatively high FHB infection rate in winter wheat was observed in 2007 and 2008 in Luxembourg. A fusariotoxin survey was carried out in 17 different geographical locations. Three groups of Fusarium mycotoxins (trichothecenes A and B and zearalenone) were analysed by a multi-detection HPLC–MS/MS method. Fusarium strains were also investigated by morphological and molecular methods. In addition, questionnaires relating to cultural practices were sent to the farmers managing the 17 fields investigated. FHB prevalence ranged from 0.3 to 65.8% (mean: 8.5%) in 2007 and from 0 to 24.5% (mean: 8.3%) in 2008. Results of morphological and molecular identification showed that the most common species isolated from diseased wheat spikes was F. graminearum (33.1%), followed by F. avenaceum (20.3%) and F. poae (17.8%). The chemical analysis revealed that 75% of the investigated fields were contaminated by deoxynivalenol (DON, range 0–8111 µg/kg). The preceding crop was highly and significantly correlated to the number of grains infected and had a significant impact on disease prevalence (p = 0.025 and 0.017, respectively, Fisher's F-test). A trend was found for maize as the preceding crop (p = 0.084, Tukey's test) to predict the amount of DON in the fields. This is the first report on the occurrence of DON and ZON in naturally infected wheat grains sampled from Luxembourg.

Occurrence and Distribution of Microdochium and Fusarium Species Isolated from Durum Wheat in Northern Tunisia and Detection of Mycotoxins in Naturally Infested Grain

AbstractAn outbreak of Fusarium Head Blight of durum wheat occurred in 2004 being localized in sub‐humid and higher semi‐arid region of Northern Tunisia. A mycological survey carried out throughout these regions, revealed that 78% of the prospected fields were infested. Results of the morphological and molecular identification, showed that the most common species isolated from diseased wheat spikes was Microdochium nivale var. nivale (63.5%), followed by Fusarium culmorum (26%), F. pseudograminearum (9%) and F. avenaceum (1.5%). To evaluate mycotoxin content of naturally infected grain, the amounts of trichothecene mycotoxin deoxynivalenol (DON) in harvested grain from 45 fields were quantified by RIDASCREEN DON Enzyme Immunoassay Kit (ELISA). This study showed that the infection levels in freshly harvested grain were very low and the maximum deoxynivalenol (DON) level of the positive samples was 53 ppb. This is the first report on the natural occurrence of DON in naturally infected wheat grain sampled from Northern Tunisia.

First occurrence of resistance to strobilurin fungicides in Microdochium nivale and Microdochium majus from French naturally infected wheat grains

Microdochium nivale (Fr.) Samuels & Hallet and Microdochium majus (Wollenweber) belong to the Fusarium ear blight (FEB) fungal complex affecting cereals. In 2007 and 2008, major Microdochium sp. infestations were observed in France, and the efficacy of strobilurins was found to be altered in some field trials. The aim of this study was to determine the sensitivity to strobilurins of French isolates of Microdochium and to characterise the possible mechanisms of resistance. Half of the strains collected in 2007 were resistant to strobilurins, and most also displayed strong resistance to benzimidazoles. Strobilurin resistance was found mostly in M. majus isolates. Positive cross-resistance was observed between all strobilurins tested, but not with the phenylpyrrole derivative fludioxonil and the various classes of sterol biosynthesis inhibitors (SBIs). In most strains, resistance was correlated with the G143A substitution in cytochrome b, the molecular target of strobilurins. Two other mechanisms were also detected at lower frequencies. This is the first report of strobilurin resistance in Microdochium. Several resistance mechanisms have evolved independently in populations and may have different impacts on field efficacy. This makes the accurate detection and quantification of QoI resistance difficult. The management of field resistance and efficacy must be adapted to take these findings into account.

Survival of the Teliosporogenous Mycelia of Neovossia indica in Infected Wheat Grains

Survival of the Teliosporogenous Mycelia of Neovossia indica in Infected Wheat Grains

Fusarium DNA traceability along the bread production chain

SummaryWheat grain contamination with toxigenic Fusarium spp. is of great economic concern to cereal producers and to the grain processing industry and is of great relevance for the quality and safety of the final products. In particular, the bread production chain can potentially be a vehicle for mycotoxin ingestion above the tolerable total daily intake. A quantitative polymerase chain reaction‐based analytical approach has been developed as a possible tool to estimate and control the risk of mycotoxins, particularly deoxynivalenol (DON). This DNA‐based analytical method has been applied to detect and quantify contamination levels of Fusarium graminearum and Fusarium culmorum in naturally infected wheat grain samples. The persistence of Fusarium contamination was also detected along the bread production chain in wholemeal, flour and bread. A significant correlation was found between Fusarium DNA and DON concentration in all samples.

Deletion of a Chromosome Arm Altered Wheat Resistance to Fusarium Head Blight and Deoxynivalenol Accumulation in Chinese Spring.

Fusarium head blight (FHB), caused by Fusarium graminearum Schwabe, is an important disease of wheat worldwide. Production of deoxynivalenol (DON) in infected wheat grain by F. graminearum is a major safety concern when considering use of the grain as feed for livestock or for human consumption. Determining chromosome locations of FHB-related genes may facilitate enhancement of wheat resistance to FHB and DON accumulation. In this study, a set of 30 ditelosomic lines derived from Chinese Spring, a moderately FHB-resistant landrace from China, were evaluated for proportion of scabbed spikelets per inoculated spike in the greenhouse and for DON contamination in harvested grain over 2 years. Significant variation in the proportion of scabbed spikelets was observed among ditelosomic lines, ranging from 13 to 95%. Seven ditelosomic lines exhibited a greater proportion of scabbed spikelets and three of these also had greater DON content than Chinese Spring (P = 0.01), suggesting that those missing chromosome arms may carry genes that contribute to resistance to FHB. Six ditelosomic lines had a reduction in proportion of scabbed spikelets, suggesting that susceptibility factors or resistance suppressors may be on these missing chromosomal arms. Selection for low proportion of scabbed spikelets in general will select for low DON content.

Fusarium head blight reactions and accumulation of deoxynivalenol, moniliformin and zearalenone in wheat grains

Deoxynivalenol (DON), moniliformin (MON) and zearalenone (ZON) are the most important mycotoxins detected in Fusarium infected wheat grains in Austria. Our goal was to identify an evaluation method for Fusarium head blight (FHB) symptoms, which is closely related to the toxin content in grains of artificially inoculated wheat genotypes. In total 108 wheat lines with varying FHB resistance were treated with F. culmorum (DON producer), F. avenaceum (MON producer) or F. graminearum (ZON producer). Several FHB evaluation methods were investigated and the data were correlated with the mycotoxin content of the grains. Data on the visual scores of the FHB development in the field, percentage of Fusarium damaged kernels and specific grain weight showed acceptable correlations with the DON content (r=0.75-0.81). MON contamination was closely related to the percentage of Fusarium damaged kernels (r=0.70). No suitable method was identified to predict ZON concentrations. Only in grain samples of highly resistant genotypes consistently a low ZON content was measured.

Resistance to Fusarium head blight and deoxynivalenol accumulation in wheat

AbstractFusarium head blight (FHB), caused by Fusarium graminearum Schwabe (telomorph =Gibberella zeae (Schw.)), is an important wheat disease world‐wide. Production of deoxynivalenol (DON) by F. graminearum in infected wheat grain is detrimental to livestock and is also a safety concern in human foods. An international collection of 116 wheat lines was evaluated for FHB resistance and concentration of DON in grain. Plants were inoculated with mixed isolates of F. graminearum in the greenhouse by injecting conidia into a single spikelet of each spike and in the field by scattering F. graminearum‐infected wheat kernels on the soil surface. FHB symptoms were evaluated by visual inspection in both the greenhouse and field, and DON was analysed by HPLC. Significant differences in FHB ratings and DON levels were observed among cultivars. In the greenhouse test, visual symptoms varied from no spread of FHB from the inoculated spikelet to spread throughout the spike, and DON levels ranged from trace levels to 283 mg/kg. In the field test, DON ranged from 2.8 to 52 mg/kg. The greenhouse test identified 16 wheat lines from various origins that accumulated less than 2 mg/kg DON. These lines may be useful as sources for breeding wheat cultivars with lower DON levels. Correlation coefficients were significant between FHB symptom ratings, seed quality traits, and DON levels. Thus, the percentage of scabbed spikelets and kernels can be generally used to predict DON levels in harvested wheat grain. In breeding programmes, selection for plants having few scabbed spikelets and scabbed kernels is most likely to result in low DON levels.

Pathogenicity and Zearalenone Production by Different Fusarium graminearum Isolates in Artificially Infected Wheat Grain

Nine Fusarium graminearum isolates were investigated for their pathogenicity on wheat ears and zearalenone production. The investigated isolates originated from different geographical regions and included representatives of Group 1 and Group 2 of F. gramineraum species. Among these isolates significant differences in pathogenicity were found. Groups 1 and 2 included isolates with higher and lower pathogenicity. 44,4% of the isolates produced zearalenone in artificially infected wheat grains at concentrations ranging from 203 to 431 µg/kg. Isolates of both groups are capable of zearalenone synthesis.

Identification and determination of mellein in cultures of the fungus Septoria nodorum (Berk.) by thin-layer and high-performance liquid chromatography

In order to select in vitro wheat embryos with a high level of resistance to the fungal pathogen Septoria nodorum (Berk.), extracts from Septoria cultures grown on infected wheat grains were prepared. One of their toxins, mellein, was identified by thin-layer chromatography and its content determined by high-performance liquid chromatography using UV light and fluorescence detection. Synthetic (±)mellein was used as standard. We investigated two techniques of Septoria cultivation, different methods of mellein extraction and the stability of the extracted mellein. In these experiments, the content of mellein in fresh fungal cultures amounted to 46 mg/kg, but in the dry material only 21 mg per kg of dry matter were recoverable.

A study of the correlation between the amount of deoxynivalenol in grain of wheat and triticale and percentage ofFusarium damaged kernels

The correlation between the amount of deoxynivalenol (DON) and the percentage ofFusarium damaged kernels (FDK) in samples of wheat and triticale was studied.Samples of naturally infected wheat grain, collected in 1986, 1987 and 1988 and of triticale collected in 1986 were used.Additionally, artificial inoculated wheat samples (10 genotypes inoculated with 3F. Culmorum strains of weak, medium and severe pathogenicity and samples of 10 triticale genotypes inoculated withF. culmorum. andF. graminearun) were studied. Using statistical methods (the variance analysis, method of least significant difference (LSD), orthogonal contrast (OC) and minimum within groups sum of squares criterion (MSSC)), the samples were divided into two groups with respect to the attribute DON/FDK.To the first group belong samples of wheat and triticale, of which the heads were artificially inoculated with severely pathogenic strainsF. culmorum. In the samples of this group the amount of DON in kernels damaged withFusarium increased by 0,46 mg/kg per 1% of FDK.In the second group, consisting of naturally infected samples and samples from artificially inoculated heads the amount of DON increased 0,30 mg DON/kg per 1% of FDK.The equation for the calculation of approximated amount of DON in farm and commercial lots of wheat and triticale after examination of percentage of FDK is given.

Water relations of some Fusarium species from infected wheat ears and grain

The effects of water activity (aw) and temperature on growth of Fusarium avenaceum, F. culmorum, F. poae and F. tricinctum were studied in vitro on wheat extract agar. Optima and minima aw for growth of all species except F. poae were 0.995–0.99 and 0.89–0.90 at optimum temperatures. The optimum aw for growth of F. poae decreased from 0.995 to 0.98 as temperature increased from 25 to 30 °C. At high aw (>0.98) F. culmorum and F. poae grew much faster (6.8 mm day−1) than F. avenaceum and F. tricinctum (4–5 mm day−1), but at lower aw differences were less. Isolates of F. culmorum and F. tricinctum all grew at similar rates but isolates of F. avenaceum differed in growth rate. Growth of all four species was influenced by substrate (wheat, malt extract or potato dextrose agars) at both high and low aw.

THE SPREAD OF DISEASES CAUSED BY SPECIES OF VERTICILLIUM

An investigation has been made of the spread of Verticillium disease of tomato and antirrhinum by root contact between diseased and healthy plants, and by the growth of the fungi through the soil. V. alboatrum and V. Dahliae spread rapidly by root contact, while V. tricorpus, V. nigrescens and V. nubilum showed no appreciable spread. Where the tomato and antirrhinum hosts wilted and died the causal organism was invariably isolated from the soil near the roots of the dead plants, but where the inoculated host harboured species without showing disease symptoms, as in the case of V. tricorpus and V. nubilum in antirrhinum, the parasite was never obtained from the soil. Moreover, these species in antirrhinum, and V. nigrescens and V. nubilum in tomato would appear to be mild pathogens since when they were already well established V. alboatrum was later actually found to enter and parasitize the respective hosts and bring about its own typical disease symptoms.Spread through the soil in the absence of hosts was tested by the insertion into it of infected wheat grains followed by later attempts to isolate the fungi at varying distances from the point of infection. V. alboatrum, V. Dahliae and V. tricorpus showed little or no growth through the soil, whereas V. nigrescens and V. nubilum spread saprophytically to some extent, though never attaining sufficient concentration to cause appreciable infection of later planted tomatoes.The conclusion is reached that spread of disease caused by species of Verticillium takes place mainly by root contact, with a rapidity relatively proportional to that of the death of the host. This finding, together with the lack of spread from antirrhinum plants infected with the non‐lethal V. tricorpus and V. nubilum, suggests that the parasites remain in the vascular tracts until the death of the hosts.