Septoria and Stagonospora Diseases of Cereals - CIMMYT ...

Genetic Variability in a Collection of Stagonospora
nodorum Isolates from Western Australia
N.E.A. Murphy, 1 R. Loughman, 2 E.S. Lagudah, 3 R. Appels, 3 and M.G.K. Jones1 1 WA State Agriculture Biotechnology Centre, Division of Science and Engineering, Murdoch University,
Murdoch, Australia
2 Agriculture Western Australia, Western Australia
3 Plant Industries, CSIRO, Canberra, Australia
Abstract
Stagonospora nodorum isolates were collected from the Western Australian cereal-belt during 1993. These isolates
and a subset of isolates taken from a single location were used to assay the level of variation within the pathogen
population. The isolates were compared using anonymous nuclear DNA markers. Three low copy number and a single
high copy number probe were used to generate restriction fragment length polymorphisms (RFLPs). The collection
exhibited a high genotypic diversity for the high copy number probe. This is consistent with the high level of sexual
reproduction previously found in the fungal population. Only minor differences between the total collection of isolates
and the subset of isolates taken from the single location were found.
Septoria nodorum blotch is one
of the most important leaf diseases
of wheat (Triticum aestivum L.) in the
Western Australian cereal-belt
(Murray and Brown, 1987). It is
caused by Stagonospora nodorum
(Berk.) Castellani & E. Germano
(teleomorph Phaeosphaeria nodorum
(E.Muller) Hedjaroude). Previous
work on the variability of S. nodorum
has been conducted principally on
phenotypic traits such as
aggressiveness (Allingham and
Jackson, 1981) and culture color
(Scharen and Krupinsky, 1970).
These studies have shown that there
is considerable variation between
isolates collected within relatively
small distances (Allingham et al.,
1981). More recently studies have
assayed the genotypic variability
using molecular techniques.
RFLPs were used to look at two
populations of S. nodorum in the USA
using a hierarchical system of
sampling (McDonald et al., 1994).
Considerable genetic variation was
found to occur on a relatively small
scale. The same set of eight probes
were used to study a population of
S. nodorum isolates gathered in
Switzerland and two populations
from the USA using a similar
hierarchical sampling system. The
Swiss population was genetically
similar to the populations from the
USA, providing evidence for gene
flow between the populations (Keller
et al., 1997a) and the gene diversity
for four of the seven RFLP loci tested
was not significantly different
between the populations (Keller et
al., 1997a).
The aim of this investigation was
to examine the level of genetic
diversity among a collection of
Western Australian S. nodorum
isolates using RFLPs and to compare
the results obtained with similar
work done in other regions of the
world.
Materials and Methods
A total of 118 isolates from 38
locations were used in the study.
Thirty-nine of the isolates were
obtained from an existing collection.
Of these 39 historical isolates, 26
87
were from a single site at
Badgingarra Research Station,
recognized as an area where high
levels of disease severity could be
expected. These isolates were
captured as single ascospores in
spore traps during 1991. The
remaining 79 isolates were collected
from 26 locations throughout the
cereal belt in 1993. They were
isolated as ascospores from wheat
stubble remaining from the 1992
crop.
The cultures of S. nodorum were
revived on wheat meal agar plates to
confirm their identity through
pycnidiospore production. Mycelia
was cultured in liquid broth and the
DNA extracted (Lagudah et al., 1991).
The genomic DNA was digested
with HindIII and run on an agarose
gel. A capillary alkali method was
used to transfer the DNA onto the
membrane. Three low copy number
probes (pASN15, pASN125 and
pJSN73) were used to produce a
multilocus haplotype, and a single
high copy number probe (pSNS4)
was used to generate a fingerprint for