Page 297 - PC2019 Program & Proceedings

PLANT CANADA 2019

P167. Development of an Immuno-PCR for the detection of pea root rot causal agent,
Aphanomyces euteiches
2
*1
Kaphle, S. ; C. Sheedy ; S. Chatterton
2
1 University of Lethbridge
2
Agriculture and Agri-Food Canada
Pea root rot, caused by the oomycete Aphanomyces euteiches, is one of the most destructive diseases of
pea. Effective measures to control this disease are lacking. An indirect ELISA and a real time immuno-
PCR (RT-iPCR) assay for the timely and sensitive detection of A. euteiches were developed using
antiserum specific to oospores of A. euteiches field isolates. The indirect ELISA exhibited a linear
working range of 62 to 500 oospores, while the RT-iPCR could detect as low as 1 oospore for all isolates
examined. To assess the performance of the RT-iPCR assay with soil samples, non-infested soils
representing a range of soil textures were collected from different locations in Alberta. An extraction
protocol was developed by spiking the treated soils with A. euteiches oospores and the RT-iPCR assay
was used to quantify oospores in the soil extracts. Detection levels in soil extracts were as low as 10
oospores per g of soil. Infested soils collected from different locations across Western Canada are being
tested by RT-iPCR for the presence of A. euteiches. To validate results obtained by the RT-iPCR assay, a
greenhouse bioassay was also performed and showed good agreement with RT-iPCR results. The current
RT-iPCR assay is more sensitive than quantitative PCR (qPCR) to detect low levels of oospores, and may
be an invaluable tool for field diagnostics.

Salina Kaphle (salina.kaphle@uleth.ca)

P168. Fusarium head blight of wheat in Alberta: species complex and related trichothecene
genotypes.
M. Hafiz; N. Schatz; M. Telfer; R. Gourlie; K. Turkington; Aboukhaddour, R.
AAFC

Fusarium head blight (FHB) is the most damaging fungal disease of wheat in Canada and Fusarium
graminearum is the primary causal organism of it. In this study infected wheat stem and grain samples
were collected from three field experiments in Alberta (Lethbridge, Lacombe and Beaverlodge) to
characterize the major Fusarium species associated with FHB. A Fusarium-selective medium was used to
isolate Fusarium species and the elongation factor 1 alpha (EF1α) gene was used for molecular
identification of the isolates. The results showed that, F. graminearum was recovered at very low rates
from Lethbridge and was not detected in Lacombe or Beaverlodge. F. avenaceum was the most dominant
species in Lethbridge (46%) as well as in Lacombe (59%), while F. culmorum was the most frequently
isolated species in Beaverlodge (51%). Trichothecene genotyping using two multiplex PCR assays based
on the Tri3 and Tri12 genes respectively, showed that the 3ADON was the most dominant genotype in all
type B trichothecene producing Fusarium species tested from these three experimental sites (F.
culmorum, F. pseudograminearum, F. acuminatum). In contrast, the 15ADON was the only genotype
recorded for all F. graminearum isolates. In conclusion, F. graminearum was not the main causal agent
associated with stem and grain samples from the three experiments, and more concern should be given to
the other Fusarium species involved in FHB complex.

Reem Aboukhaddour (reem.aboukhaddour@canada.ca)

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