Fusarium head blight (FHB) of wheat, caused principally by Fusarium graminearum in Canada, results in accumulation of mycotoxins in the grain. The fungus produces deoxynivalenol (DON) and its acetylated forms 3-ADON or 15-ADON. Increasing numbers of F. graminearum isolates collected in Manitoba between 1998 and 2004 were of the 3-ADON chemotype, whereas prior to 1998, the 15-ADON chemotype was considered the only significant cause of FHB in North America. Between 2008 and 2012, a series of studies were conducted at Carman (2008, 2009) and Glenlea, Manitoba (2008-2012) to monitor the ratio of 3-ADON to 15-ADON chemotypes recovered after wheat plots were artificially inoculated, usually in a 1:1 ratio of 3-ADON:15-ADON isolates of F. graminearum. Additional studies were conducted under controlled conditions. In 2008, the majority of isolates recovered from both Carman and Glenlea were of the 3-ADON chemotype. In subsequent years, however, the ratio was closer to 1:1 or sometimes with a slight predominance of 15-ADON isolates. Under controlled conditions, recovery of 15-ADON isolates predominated at lower incubation temperatures (18-22 degrees C) whereas at 28 degrees C, 3-ADON isolates were more prevalent. At moderate temperature (24 degrees C), results were mixed - seven samples had a 1:1 ratio, three had higher recovery of 3-ADON isolates, and two had higher recovery of 15-ADON. However, an examination of weather variables over the 5-year period of the field studies showed no correlation between recovery of chemotype and temperature or precipitation. ResumeLa brulure de l'epi (FHB) du ble, causee principalement au Canada par Fusarium graminearum, engendre des accumulations de mycotoxines dans les grains. Le champignon produit du deoxynivalenol (DON) et ses formes acetylees 3-ADON ou 15-ADON. De plus en plus d'isolats de F. graminearum collectes au Manitoba de 1998 a 2004 appartenaient au chimiotype du 3-ADON alors que, avant 1998, le chimiotype du 15-ADON etait considere comme la seule cause importante de FHB en Amerique du Nord. De 2008 a 2012, une serie d'etudes ont ete menees a Carman (2008, 2009) et Glenlea (2008-2012), au Manitoba, pour surveiller la proportion recuperee des chimiotypes du 3-ADON et du 15-ADON apres que les parcelles de ble avaient ete inoculees artificiellement, le rapport des isolats de F. graminearum produisant du 3-ADON et du 15-ADON etant habituellement de 1:1. Des etudes supplementaires ont ete menees dans des conditions controlees. En 2008, la majorite des isolats recuperes a Carman et a Glenlea appartenaient au chimiotype du 3-ADON. Toutefois, au cours des annees suivantes, le rapport etait plus pres de 1:1 ou parfois affichait une legere predominance des isolats produisant du 15-ADON. Dans des conditions controlees, les isolats recuperes appartenant au chimiotype du 15-ADON predominaient a des temperatures d'incubation plus basses (18 a 22 degrees C) tandis que, a 28 degrees C, les isolats appartenant au chimiotype du 3-ADON etaient plus nombreux. 24 degrees C, temperature consideree comme moderee, les resultats variaientsept echantillons affichaient un rapport de1:1, trois affichaient un rapport plus eleve de recuperation des isolats produisant du 3-ADON et deux, un rapport plus eleve d'isolats produisant du 15-ADON. Toutefois, un examen des variables meteorologiques des cinq annees qu'ont dure les etudes en champ a montre qu'il n'y avait aucune correlation entre la recuperation des chimiotypes et la temperature ou les precipitations.

Corn kernels colonized by Fusarium graminearum were spread between lines of barley (Hordeum vulgare) in an irrigated Fusarium head blight (FHB) nursery at a 2:1 ratio of isolates producing 15-acetyldeoxynivalenol (15 ADON) to isolates producing 3-acetyldeoxynivalenol (3 ADON) (in 2008) or 1:1 ratio (in 2009, 2010). In 2008, it was the first use in this field of a 3 ADON isolate after 9 years of artificial inoculation with exclusively 15 ADON isolates. Sets of three 2-row and three 6-row cultivars were used as checks throughout the nursery, representing a range of susceptibility to FHB. Composites of these cultivars were prepared and analysed for per cent F. graminearum infection, chemotype frequency and levels of deoxynivalenol (DON), 3 ADON and 15 ADON. Seed infection by F. graminearum ranged from 83.5% (2009) to 98.5% (2010), with little difference across the nursery. In 2008, the chemotype distribution of F. graminearum and DON levels on infected kernels showed a several-fold difference from the eastern side (67% 3 ADON isolates and 46 ppm DON) to the western end of the field (16% 3 ADON isolates and 13 ppm DON), but not in the subsequent 2 years. In both 2009 and 2010, the 3 ADON chemotype was recovered from only 18% and 13% of the seeds, and overall average DON levels were 18 ppm and 24 ppm, respectively. In 2008, the highest DON and 3 ADON levels were associated with the highest frequency of the 3 ADON chemotype. In all years, the recovery of isolates did not reflect the ratio of the initial inoculum, suggesting that other factors influenced the ultimate infection of the seed. However, the relative relationship among the barley cultivars was consistent throughout all 3 years: DON levels were much lower in the most resistant cultivars compared with the most susceptible cultivars in all years and composites. Genotypic response to DON accumulation did not show strong interaction with 3 ADON frequency, suggesting that the underlying genetic resistance of the barley cultivars is cross-applicable between chemotypes.

Fourteen samples consisting of three cultivars of hulless barley (Hordeum vulgare L.), grown commercially in southern Manitoba in 1994, were examined for levels of Fusarium species and seven fusarium trichothecene mycotoxins: deoxynivalenol (DON), diacetoxyscirpenol, 15-acetyldeoxynivalenol (15ADON), fusarenone-X, HT-2 toxin, neosolaniol and T-2 toxin. Four fractions from each sample were analysed. These consisted of kernels with the hulls still attached, kernels whose hulls had been removed at harvest, kernels that had been dehulled in the laboratory using a deawner, and kernels that had been pearled in the laboratory to 60% of their original weight. A composite sample of the hulls, obtained from the laboratory dehulling, was also tested. Mycotoxins were determined by gas chromatography-mass spectrometry. DON and 15ADON were the only trichothecenes detected in this study. The greatest numbers of propagules of Fusarium spp, and the highest concentrations of DON and 15ADON were found in the hulls. The average concentrations of F. graminearum Schwabe and DON in barley were reduced: 90 and 49%, respectively, as a consequence of laboratory dehulling; 95 and 59%, respectively, as a result of dehulling during harvest; and 99 and 86%, respectively, as a result of pearling. Results suggest that growing hulless cultivars shows promise for management of mycotoxin problems associated with FHB.

One hundred and twenty-seven isolates of Fusarium pseudograminearum were obtained from seeds and vegetative parts of cereals and other gramineae grown in western Canada. Culture of the isolates on agar selective for Fusarium graminearum allowed a successful differentiation from F. graminearum. Sterilized rice (40% moisture content) was inoculated with a single germinated spore of isolates identified as F. pseudograminearum and was incubated at 23 °C. After 14 days of incubation, mycelium was taken from each culture, and the DNA was extracted to detect the tri5 gene by polymerase chain reaction analysis and confirm the species identity. After 21 days of incubation, mycotoxins were quantified in dried rice. All isolates contained the tri5 gene. Deoxynivalenol (DON) was produced in 125 of the 127 isolates, 3-acetyldeoxynivalenol (3-ADON) in 122 isolates, 15-acetyldeoxynivalenol (15-ADON) in 2 isolates, diacetoxyscirpenol (DAS) in 17 isolates, and zearalenone in 100 isolates; both nivalenol (NIV) and fusarenon X (FX) were detected in 1 isolate. There appears to be three chemotypes: DON–3-ADON, DON–15-ADON, and NIV. Neither of the two 15-ADON producers or the NIV producer formed DAS. This is the first time that the production of DAS and FX is associated with F. pseudograminearum. None of the isolates produced HT-2 toxin or T-2 toxin at detectable levels.

Producer samples of Canada western red spring (CWRS) wheat (Triticum aestivum) and Canada western amber durum (CWAD) wheat (Triticum turgidum var. durum) grown in Manitoba, Saskatchewan, and Alberta between 2000 and 2002 were composited by year and crop district, then analyzed for fungal infection and contamination by trichothecenes and inoniliformin. One hundred and fourteen CWRS and 79 CWAD samples were composited from 1136 CWRS and 643 CWAD producer samples. Alternaria altentata was the most frequently isolated species from each province. Levels of Alternaria alternata, Bipolaris sorokiniana, and Fusarium graminearum were highest in Manitoba and eastern Saskatchewan, whereas Clodosporium spp., Pyrenophora tritici-repentis, and Stagonospora nodorum were highest in samples from further west. The frequency of the toxigenic species F graminearum greatly exceeded that reported 20 or more years earlier. Samples from all 3 years were analyzed for the presence of eight trichothecenes, and from 2002, for moniliformin. Deoxynivalenol (DON) was found almost exclusively in wheat from the eastern Prairies, at levels up to 7.9 ppm, where it sometimes cooccurred with 15-acetyl deoxynivalenol. HT-2 was detected at low levels in 7 of 79 CWAD composites but not in the CWRS composites. Low levels of moniliformin were found in one CWRS and nine CWAD composite samples, the first time that this compound has been reported from naturally infected wheat in North America. Not detected were nivalenol, fusarenon X, trichothecin, 3-acetyl deoxynivalenol, neosolaniol, diacetoxyscirpenol, and T-2 toxin.

Species-specific PCR was used for the identification of nine Fusarium species in pure mycelial culture. A PCR-based method was compared with the whole seed agar plate method and trichothecene analysis for three toxin-producing Fusarium species using 85 grain samples of wheat, barley, oat, corn and rye. A simple SDS-based DNA extraction system followed by potassium acetate precipitation resulted in consistent PCR amplification of DNA fragments from cultures and grain samples. The species-specific PCR assays correctly identified pure cultures of Fusarium avenaceum ssp. avenaceum (9 isolates), Fusarium acuminatum ssp. acuminatum (12 isolates), Fusarium crookwellense (7 isolates), Fusarium culmorum (12 isolates), Fusarium equiseti (11 isolates), Fusarium graminearum (77 isolates), Fusarium poae (10 isolates), Fusarium pseudograminearum (23 isolates), and Fusarium sporotrichioides (10 isolates). Multiplex PCR was developed for the simultaneous detection of F. culmorum, F. graminearum and F. sporotrichioides, the three most important trichothecene producing species in Canada. In grain samples, results of PCR assays for these same three species related well with whole seed agar plate method results and determination of Fusarium trichothecenes. The PCR assay described in this study can be used for routine detection and identification of Fusarium spp. in Canada.

From 1995 to 1997, spikes were collected from a total of 160 barley and 188 wheat fields from the Peace River region to the Three Hills area of Alberta, Canada. After threshing, 100 seeds from each field were surface sterilized, incubated on potato dextrose agar, and examined for the presence of the following pathogens: Fusarium graminearum, other Fusarium spp., Cochliobolus sativus, Pyrenophora tritici-repentis, Pyrenophora graminea, Pyrenophora teres, and Stagonospora nodorum. Fusarium graminearum was not detected in any seed samples. The most common Fusarium species isolated was Fusarium avenaceum. Maximum seed infection levels with E avenaceum in a single field were 51 and 37% for barley and wheat, respectively. Substantial levels of seed-borne P teres were found in barley with maximum infection levels of 82, 81, and 89% in 1995, 1996, and 1997, respectively. Stagonospora nodorum was also commonly found in both barley and wheat seed with maximum infection levels of 61 and 54%, respectively. Pyrenophora tritici-repentis, P graminea, and C. sativus were generally present at low levels.

Pathogen levels and seed viability of two samples each of barley (Hordeum vulgare) (B 1, 132), Canada western red spring wheat (Triticum aestivum) (RS1, RS2), and Canada western amber durum wheat (AD I, AD2) were assessed after heating seed at 50 or 70degreesC for up to 14 days. RS2 and B2, with an initial incidence of 23 and 84% of Fusarium graminearum, respectively, were also heated at 60degreesC for 24 days and 80degreesC for 10 days. Pathogen levels and seed viability were assessed by plating seed onto potato dextrose agar and wet filter paper, respectively. Fusarium graminearum was eliminated from RS2 after 15 days at 60degreesC, 5 days at 70degreesC, or 2 days at 80degreesC. In B2, F graminearum was eliminated after 21 days at 60degreesC, 9 days at 70degreesC, or 5 days at 80degreesC. After heating at 50 or 70degreesC, the observed frequency of Cochliobolus sativus in B1 declined slightly but significantly over time, whereas it substantially increased in B2. A significant decline in the incidence of C. sativits was observed in B2 after heating at 80degreesC. Pyrenophora teres was observed significantly more often in B I after heating at 50 or 70degreesC, whereas Pyrenophora tritici-repentis in AD1 and AD2 was unaffected by heating at 50degreesC. However, the detection of P tritici-repentis did significantly increase over time in AD2 when heated at 70degreesC. In AD1, heating at 70degreesC initially increased, then decreased the observed incidence of this pathogen. Cochliobolus sativus, P. teres and P. tritici-repentis were still viable in the samples after 14 days of heating at 70degreesC, but C. sativus was not detected after 10 days at 80degreesC. Germination of wheat tempered to 12, 14, and 16% moisture content was unaffected by heating at 70degreesC for 7 days, whereas barley tempered to the same moisture content had a slight decline in germination. Germination rates in most samples were unaffected by the treatment times and temperatures sufficient to eradicate E graminearum, but a significant decline in viability was recorded for AD2 and B1 heated at 70degreesC. The germination of B2 increased when heated at 70degreesC, but declined when heated at 80degreesC. It is recommended that thermotherapy be applied to control national and international movements of E graminearum and other heat-sensitive pathogens in germplasm used for research and breeding purposes.

Fusarium graminearum, Fusarium culmorum, and Fusarium avenaceum, isolated from Fusarium-damaged wheat harvested in western Canada, were cultured and evaluated for mycotoxin production. Extracts of the culture media were assayed for trichothecenes by gas chromatography-mass spectrometry and for moniliformin by liquid chromatography. Deoxynivalenol (DON) was found in 28 of 42 isolates of F. graminearum and 42 of 42 isolates of F. culmorum at levels ranging from 0.5 to 25.0 microg/g. 15-AcetylDON was found in 28 of 42 isolates of F. graminearum at levels ranging from 1.0 to 7.1 microg/g. 3-AcetylDON was found in 41 of 42 isolates of F. culmorum at levels ranging from 0.8 to 13.0 microg/g. Several other trichothecenes were assayed but not detected in the culture medium. Moniliformin was present in 40 of 42 isolates of F. avenaceum at levels ranging from 1.3 to 138.1 microg/g, but was not present in any of the isolates of F. graminearum or F. culmorum.

Survey results from 1993 to 1998 for fungi isolated from fusarium-damaged kernels of wheat are presented. There is compelling evidence that Fusarium gramineanrm has recently been spreading westward from southeastern Manitoba, replacing less pathogenic Fusarium species as the principal fusarium head blight (FHB) pathogen. This movement has been accompanied by increasing economic losses from the effects of FHB. Environmental factors such as a lower average daily temperature in June and July may be influential in limiting the damage from F. graminearum in the western prairies, where F. graminearum currently is rare. However, precipitation levels at anthesis equal to those in the areas presently affected by economic levels of FHB occur in many western crop districts, suggesting that precipitation levels during the period of anthesis will likely promote further westward spread of this pathogen. The potential role of infected seed as a mechanism for long-distance dispersal of F. graminearum is considered.

The mycoflora and levels of deoxynivalenol (DON) on oat seed (Avena sativa) grown in western Canada was determined by analyzing a total of 511 grain samples collected from 39 crop districts in Alberta, Saskatchewan, and Manitoba during 1995-1997. Fungi representing a minimum of 85 species were recovered. Alternaria alternata was the most frequently isolated species from each province. Levels of A. alternata, Bipolaris sorokiniana, and Fusarium graminearum were highest in Manitoba and eastern Saskatchewan, whereas Cladosporium species and Drechslera avenacea were highest in samples from Alberta and western Saskatchewan. DON levels ≥ 0.10 ppm were found in seeds from two Manitoba crop districts in 1996 and three in 1997, with a maximum level of 0.34 ppm. In 1997, DON was also detected in composite samples of seed from three Saskatchewan and two Alberta crop district composites.

A total of 136 seed samples of barley and 30 samples of oat grown in Manitoba in 1993 and 1994 were analyzed for the level and species of Fusarium infecting the kernels. Fusarium graminearum was the most commonly isolated of 11 Fusarium spp. In barley it ranged in mean frequency from 2% seed infection in samples from the northwestern (NW) region of Manitoba to 71% in samples from the central region of Manitoba. Oat had lower average levels of F. graminearum, ranging from nondetectable in the NW to 38% in central Manitoba. Definitive, reliable symptoms of fusarium head blight infection in harvested barley and oat were often absent. Due to the high frequency of F. graminearum in many samples, and this pathogen's known ability to produce deoxynivalenol, testing for this mycotoxin in these commodities appears warranted. This is especially true for barley grown in the central, eastern, or interlake regions of Manitoba in years when fusarium head blight is severe and the grain is destined for malting or pig feed.

From 1989 to 1993, composite samples of canola from crop districts in western Canada and, from 1991 to 1993, individual producer samples of canola from Ontario were tested for the presence of seedborne fungi. Each year, 600 seeds from each western crop district and between 150 and 300 seeds from each Ontario sample were surface disinfected before plating onto 20% V-8 agar. Seventy species representing 36 genera were recovered. Alternaria alternata was the most common species recovered, followed by Alternaria brassicae and Alternaria raphani. The frequency with which Alternaria alternata was recovered from seed was higher in samples from the more easterly provinces, whereas that of Alternaria brassicae and Alternaria raphani were highest in samples from the more westerly ones.

To determine the mycoflora of grain samples of white winter wheat (Triticum aestivum), 435 samples collected over three years were examined for the presence of fungi by plating surface disinfected seeds onto potato dextrose agar. At least 59 species representing 35 fungal genera were recovered from seed. Alternaria alternata, Epicoccum nigrum, and species of Arthrinium, Aspergillus, Cladosporium, Drechslera and Nigrospora infected more than 1% of the seeds every year. Bipolaris sorokiniana, Drechslera tritici-repentis, Fusarium graminearum, F. poae, and Septoria nodorum infected more than 1% of the seeds in one or two years. Yearly differences in the quantity and time of precipitation and the frequency of a number of fungi such as the pathogens B. sorokiniana, D. tritici-repentis, and S. nodorum, including a 100 fold increase in the frequency of F. graminearum between 1988 and 1989, were recorded. Whereas forty years ago B. sorokiniana was the most common pathogen recovered from Ontario wheat seed, F. graminearum was the most frequently detected pathogen in this study.

Growth of Fusarium moniliforme , F. proliferatum , and F. subglutinans on Czapek solution agar containing 20% saccharose resulted in both cultural differences and enhanced micromorphological features. F. moniliforme could be reliably distinguished from the other two species based on differences in colony color and texture. These differences were intensified by lowering the pH of the media from 7.7 to 4.4 without adversely affecting micromorphology.

The Fusarium spp. associated with samples of wheat seed containing tombstone kernels were identified for 454 samples collected in 1986 and 1987, representing six wheat classes from three provinces. The results indicate that the frequencies and distribution of the 13 species isolated were influenced by geographic location, growing conditions and wheat class. In 1986 and 1987, F. graminearum infected the most seeds and samples of Ontario-grown wheats. It was also the most frequently recovered species from Manitoba wheats in 1986, but was absent from the Saskatchewan wheats. In 1987, F. avenaceum was isolated from the greatest percentage of seeds from Manitoba and Saskatchewan. The greatest percentage of Manitoba samples in 1986 and 1987 were infected by F. sporotrichioides whereas F. acuminatum infected the most Saskatchewan samples. Only F. graminearum infected more than 7% of the seeds of any sample of Canada Western Red Spring, Canada Prairie Spring, Canada Eastern White Winter, Canada Eastern Red Winter, or Canada Eastern Red Spring class of wheat (to a maximum of 52%), whereas five species (F. acuminatum, F. avenaceum, F. equiseti, F. poae, and F. sporotrichioides) exceeded that level in one or more samples of the Canada Western Amber Durum class. The other seven Fusarium spp. (F. compactum, F. crookwellense, F. culmorum, F. proliferatum, F. oxysporum, F. pallidoroseum, and F. subglutinans) combined accounted for <5% of isolates. Overall, the lowest frequency of Fusarium spp. was found in the Canada Western Red Spring class of wheats, and the highest in the Canada Western Amber Durum class.Key words: Fusarium, wheat, disease, graminearum, avenaceum, scab