TITLE:
Fine Tuning of Real Time PCR as a First Tool for the Detection of G143A Substitution in Venturia inaequalis Samples
AUTHORS:
Ceren Turan, Irene Maja Nanni, Lucia Landi, Alessandro Pirondi, Marina Collina
KEYWORDS:
Venturia inaequalis, qPCR, Strobilurin, Cytochrome b
JOURNAL NAME:
American Journal of Plant Sciences,
Vol.12 No.6,
June
30,
2021
ABSTRACT: Apple
scab caused by Venturia inaequalis (Cke.) Wint. is the most important disease of apple trees worldwide and
requires a high number of fungicide applications. The G143A substitution in the
inhibitor binding site of cytochrome b of V. inaequalis confers a high level
of resistance to strobilurins targeting the bc1 complex. The aim of this work was to substitute the labor intensive in vitro assays, with the faster
quantitative PCR. An allele-specific qPCR method with a newly designed primer
set was successfully developed to quantitatively determine the frequency of
QoI-resistant A143 allele in populations of V.
inaequalis. To be able to suggest that the molecular method could be
applied as unique and robust technique, we carried out in vitro sensitivity test to trifloxystrobin; first testing the relative germination and
subsequently confirmed with the quantification of mutated allele frequencies by
qPCR on forty-nine Italian V. inaequalis populations. qPCR gave a
similar pattern to that obtained using in
vitro conidial germination test in predominantly sensitive and resistant
populations, the variability between these two tests was observed in some
heterogeneous populations. The qPCR assay developed in this study efficiently
quantifies the A143 allele and we can conclude that this method could be useful
for the study of the fungicide resistance at population level in the fields,
giving a quick response also with a large amount of samples.