Method evaluation of Fusarium DNA extraction from mycelia and wheat for down-stream real-time PCR quantification and correlation to mycotoxin levels

Fredlund, Elisabeth; Gidlund, Ann; Olsen, M.; Börjesson, Thomas; Spliid, Niels Henrik; Simonsson, Magnus

doi:10.1016/j.mimet.2008.01.007

Cited by 84 publications

(41 citation statements)

References 17 publications

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“…These different values were obtained because of the presence of PCR inhibitors. The same problem was found by Fredlund et al (2008) who tested different types of Fusarium DNA extraction, but inhibitors were always present. Another factor which can affect the detection limit of the PCR is the different effectivity of the binding of degenerate primers in various Fusarium template DNAs.…”

Section: Resultssupporting

confidence: 63%

“…On the other hand, the methods based on DNA analysis are fast, sensitive and accurate. Many types of molecular methods were developed for the detection and identification of Fusarium species, based mainly on PCR and multiplex PCR techniques (Edwards et al 2002;Bluhm et al 2004;Jurado et al 2006;Kulik 2008a,b;Sampietro et al 2010) and on real-time PCR for quantification (Reischer et al 2004;Waalwijk et al 2004;Dyer et al 2006;Leišová et al 2006;Fredlund et al 2008). The Amplified fragment length polymorphisms (AFLP) and Random amplified polymorphic DNA (RAPD) were used for identification of different pathotypes of Gibberella zeae (Cumagun et al 2007).…”

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confidence: 99%

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

Pavlátová¹,

Novotný²,

Hodek³

et al. 2011

Czech J. Food Sci.

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Pavlátová L., Novotný D., Hodek J., Chrpová J., Ovesná J. (2011): Utilization of DNA microarrays for detection and identification of selected Fusarium species from the Czech Republic. Czech J. Food Sci., 29 (Special Issue): S93-S101.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.

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Section: Resultssupporting

confidence: 63%

mentioning

confidence: 99%

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

Pavlátová¹,

Novotný²,

Hodek³

et al. 2011

Czech J. Food Sci.

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“…In addition, long distance dispersal of viable F. graminearum spores is possible (MaldonadoRamirez et al 2005;Schmale et al 2012). We speculate whether the increased prevalence of FGSC observed in several European countries (Chandelier et al 2011;Jennings et al 2004;Stępień et al 2008;Waalwijk et al 2003) including the Nordic countries (Bernhoft et al 2010;Fredlund et al 2008;Nielsen et al 2011;YliMattila 2010) may have facilitated the introduction of the 15-DON genotypes into new areas like Norway and Denmark. As such, the prevalence of 15-ADON genotypes and associated mycotoxin levels in Norwegian cereals should be monitored.…”

Section: Discussionmentioning

confidence: 84%

“…infection levels in cereal seeds, as well as the increased levels of trichothecene contamination in cereal grains, observed in Norway in recent years (Bernhoft et al 2013). During the same period, members of the Fusarium graminearum species complex (FGSC) have become more prevalent in several European countries (Chandelier et al 2011;Jennings et al 2004;Stępień et al 2008;Waalwijk et al 2003) including the Nordic countries (Bernhoft et al 2010;Fredlund et al 2008;Nielsen et al 2011;Yli-Mattila 2010), partly replacing other Fusarium species.…”

Section: Introductionmentioning

confidence: 99%

Genetic and phenotypic diversity within the Fusarium graminearum species complex in Norway

et al. 2015

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As has been observed in several European countries, the frequency of Fusarium head blight (FHB) caused by members of the Fusarium graminearum species complex (FGSC) has increased in Norwegian cereals in recent years, resulting in elevated levels of deoxynivalenol in cereal grains. The objective of this study was to determine if this increase was associated with changes in FGSC composition within Norway. FGSC isolates collected from wheat, oats and barley in Norway during two periods, mainly 1993-1998 and 2004-2007, were characterized to determine species and trichothecene genotype composition and to assess levels of genetic variation and population structure. In vitro growth rates at different temperatures and aggressiveness in spring wheat were further characterized for a sub-selection of isolates. All Norwegian isolates were identified as F. graminearum. The 3-acetyl-deoxynivalenol (3-ADON) trichothecene type was dominant. However, isolates with the 15-ADON chemotype were detected in Norway for the first time and may represent a recent introduction of this trichothecene type. Bayesian-model based clustering and analyses of genetic differentiation indicated the persistence over the last 20 years of two sympatric and partially admixed populations of F. graminearum in Norway. Significant differences in average in vitro growth rates and aggressiveness were observed between these two populations. Our results demonstrate that the recent increase in prevalence of the FGSC in Norwegian cereals do not correspond to any dramatic changes in FGSC species or trichothecene chemotype composition. However, significant changes in population frequencies were observed among Norwegian F. graminearum.

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“…Likewise, reports from several other European countries indicate an increased prevalence of F. graminearum during the most of the last twenty years (Fredlund et al, 2008;Nielsen et al, 2011;Waalwijk et al, 2004;Xu et al, 2005). This is in contrast to studies performed during the 1990's where F. culmorum generally was regarded as the main DON producer in Norwegian cereals, and F. graminearum was registered at low frequencies (Henriksen and Elen, 2005;Kosiak et al, 2003).…”

Section: Fusarium Graminearum Fusarium Culmorum and Associated Mycotmentioning

confidence: 84%

Associations betweenFusarium species and mycotoxins in oats and spring wheat from farmers’ fields in Norway over a six-year period

Hofgaard

Aamot

Torp

et al. 2016

WMJ

110

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During the last ten years, Norwegian cereal grain industry has experienced large challenges due to Fusarium spp. and Fusarium mycotoxin contamination of small-grained cereals. To prevent severely contaminated grain lots from entering the grain supply chain, it is important to establish surveys for the most prevalent Fusarium spp. and mycotoxins. The objective of our study was to quantify and calculate the associations between Fusarium spp. and mycotoxins prevalent in oats and spring wheat. In a 6-year period from 2004-2009, 178 grain samples of spring wheat and 289 samples of oats were collected from farmers' fields in South East Norway. The grains were analysed for 18 different Fusarium-mycotoxins by liquid chromatography -mass spectrometry. Generally, the median mycotoxin levels were higher than reported in Norwegian studies covering previous years. The DNA content of Fusarium graminearum, Fusarium culmorum, Fusarium langsethiae, Fusarium poae and Fusarium avenaceum were determined by quantitative PCR. We identified F. graminearum as the main deoxynivalenol (DON) producer in oats and spring wheat, and F. langsethiae as the main HT-2 and T-2-toxins producer in oats. No association was observed between quantity of F. graminearum DNA and quantity of F. langsethiae DNA nor for their respective mycotoxins, in oats. F. avenaceum was one of the most prevalent Fusarium species in both oats and spring wheat. The following ranking of Fusarium species was made based on the DNA concentrations of the Fusarium spp. analysed in this survey (from high to low): F. graminearum = F. langsethiae = F. avenaceum > F. poae > F. culmorum (oats); F. graminearum = F. avenaceum > F. culmorum > F. poae = F. langsethiae (spring wheat). Our results are in agreement with recently published data indicating a shift in the relative prevalence of Fusarium species towards more F. graminearum versus F. culmorum in Norwegian oats and spring wheat.

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Method evaluation of Fusarium DNA extraction from mycelia and wheat for down-stream real-time PCR quantification and correlation to mycotoxin levels

Cited by 84 publications

References 17 publications

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

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

Genetic and phenotypic diversity within the Fusarium graminearum species complex in Norway

Associations betweenFusarium species and mycotoxins in oats and spring wheat from farmers’ fields in Norway over a six-year period

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