An oligonucleotide microarray for the identification and differentiation of trichothecene producing and non-producing Fusarium species occurring on cereal grain.
ABSTRACT Cereal grain may be infected with a number of Fusarium species some of which are producers of highly toxic compounds such as the trichothecenes. Correct identification of these species is essential for risk assessment of cereal grain for human or animal consumption. Most of the available methods for identification are either time consuming or aimed at only one or a few target species. Microarray technology offers parallel analysis of a high number of DNA targets. In this study 57 capture oligonucleotides (CO) were designed based upon Fusarium ITS2 rDNA sequences, and used for microarray production. From this array COs could be selected that were able to hybridise specifically to labelled PCR products from the ITS region of Fusarium graminearum/Fusarium culmorum, Fusarium pseudograminearum, Fusarium poae, Fusarium sporotrichioides, Fusarium equiseti, Fusarium langsethiae and Fusarium tricinctum/Fusarium avenaceum. A few COs showed some cross hybridisation to non-target species. In a preliminary experiment it was shown that this cross hybridisation could be eliminated by increasing hybridisation stringency. The array could be used to detect individual Fusarium species in mixed samples and in environmental samples. This study demonstrates the feasibility of oligonucleotide microarrays for parallel detection of a number of Fusarium species.
- Phytopathology. 01/1996; 86(5).
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ABSTRACT: Existing PCR-based assays for the detection of the cereal pathogen Fusarium avenaceum were found to cross-react with F. tricinctum. An investigation into the phenetic relationship between these two species using two different marker systems revealed a close relationship between them despite their being from separate taxonomic sections. Whilst RFLP differences in the ITS regions surrounding the nuclear 5.8 S rDNA were insufficient to discriminate between isolates of the two species, they were clearly differentiated by RAPD profiling. RAPD fragments from F. avenaceum isolates were screened for hybridization to F. tricinctum DNA on Southern blots. One of 12 selected RAPD fragments showed no hybridization to genomic DNA from F. tricinctum. This fragment was cloned and sequenced, and the sequence obtained was used to design PCR primers. The primers were found to be specific for F. avenaceum, with no cross-reactions obtained with F. tricinctum or any other wheat pathogen assayed. The primers were able to differentiate between the two species in infected plant material, in contrast to the earlier assays.Plant Pathology 05/1998; 47(3):278 - 288. · 2.73 Impact Factor
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ABSTRACT: The Fusarium species predominantly found associated with Fusarium head blight (FHB) in wheat and other small-grain cereals all over Europe are F. graminearum, F. avenaceum and F. culmorum. Among the less frequently encountered species are several others which are less pathogenic or opportunistic, but also toxigenic. These include F. poae, F. cerealis F. equiseti F. sporotrichioides F. tricinctum and, to a lesser extent, F. acuminatum F. subglutinans F. solani F. oxysporum F. verticillioides F. semitectum and F. proliferatum. The species profile of FHB is due to several factors, primarily climatic conditions, particularly rain and the temperature at flowering stage, but also agronomic factors, such as soil cultivation, nitrogen fertilization, fungicides, crop rotation, and host genotype. The most frequently encountered Fusarium mycotoxins in FHB in Europe has proved to be deoxynivalenol and zearalenone produced by F. graminearum and F. culmorum with the former more common in southern (warmer) and the latter in northern (colder) European areas. Nivalenol was usually found associated with deoxynivalenol and its derivatives (mono-acetyldeoxynivalenols), together with fusarenone-X, formed by F. graminearum F. cerealis F. culmorum and, in northern areas, by F. poae. Moreover, from central to northern European countries, moniliformin has been consistently reported, as a consequence of the widespread distribution of F. avenaceum whereas the occurrence of T-2 toxin derivatives, such as T-2 toxin and HT-2 toxin, and diacetoxyscirpenol have been recorded in conjunction with sporadic epidemics of F. sporotrichioides and F. poae. Finally, beauvericin and various enniatins have recently been found in Finnish wheat colonized by F.avenaceum and F. poae.European Journal of Plant Pathology 08/2002; 108(7):611-624. · 1.61 Impact Factor