Te. Bunting et al., IDENTIFICATION OF MAGNAPORTHE-POAE BY PCR AND EXAMINATION OF ITS RELATIONSHIP TO OTHER FUNGI BY ANALYSIS OF THEIR NUCLEAR RDNA ITS-1 REGIONS, Phytopathology, 86(4), 1996, pp. 398-404
Magnaporthe poae is the causal agent of summer patch disease in turfgr
asses. Identification of this heterothallic, root-infecting fungus has
been difficult because of the lack of diagnostic structures in nature
and the time required for the fungus to produce perithecia in culture
. Potential probes for the identification of M. poae were obtained by
producing a partial genomic library of M. poae isolate 73-15 in the pG
EM3Zf(+) plasmid vector and subsequently screening cloned DNA for stro
ng hybridization to 73-15 isolate genomic DNA. Using Southern blot ana
lysis, a 2.7-kb DNA clone (pMp2-7) that hybridized to six confirmed is
olates of M. poae from NJ, PA, NY, and RI was identified. Except for o
ne isolate of Colletotrichum graminicola, the probe did not hybridize
to 42 other fungal isolates that commonly inhabit the turfgrass enviro
nment. Oligonucleotides that would prime amplification of a 453-nucleo
tide (nt) fragment from M. poae DNA, but not from C. graminicola DNA,
were developed based on sequence from one end of pMp2-7. These oligonu
cleotides also primed amplification of a similar-sized fragment of one
isolate of M. rhizophila (PREM 45952) DNA. DNA from another isolate o
f M. rhizophila (Mr-2 from PA) did not amplify using these primers. To
examine the relationship of M. poae to other fungal species, the inte
rnal transcribed spacer region (ITS-1) of nuclear ribosomal DNA for me
mbers of the Magnaporthe genus and several other fungi was sequenced.
Sequences of three isolates of M. poae, identified through mating, and
isolate PREM 45952 were very similar, having only five variable sites
of 236. Isolate Mr-2 had seven sites variable to the M. poae isolates
, two that were identical to variable sites in PREM 45952. Using maxim
um parsimony analysis, the M. rhizophila and M. poae isolates grouped
in 75% of bootstrap replications. M. rhizophila isolates formed a mono
phyletic group away from the M. poae isolates in four of five equally
parsimonious trees, but this grouping was not supported by more than 5
0% of bootstrap replications.