A specific primer PCR and RFLP assay for the rapid detection and differentiation in planta of some Mycosphaerella species associated with foliar diseases of Eucalyptus globulus

H.A.G.C. Kularatne; A.C. Lawrie; P.A. Barber; P.J. Keane

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A specific primer PCR and RFLP assay for the rapid detection and differentiation in planta of some Mycosphaerella species associated with foliar diseases of Eucalyptus globulus

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A specific primer PCR and RFLP assay for the rapid detection and differentiation in planta of some Mycosphaerella species associated with foliar diseases of Eucalyptus globulus

H.A.G.C. Kularatne, A.C. Lawrie, P.A. Barber and P.J. Keane

Mycological Research, Vol.108(12), pp.1476-1493

2004

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Abstract

It is difficult to accurately identify Mycosphaerella species associated with leaf diseases of Eucalyptus based on morphological characters, as there is considerable overlap between very similar species and subspecies, and isolation from the host is not easy. Thus, a PCR and RFLP assay based on the ITS region of nr DNA was developed for the rapid detection and differentiation of M. nubilosa, M. cryptica and two non-sporing unidentified Mycosphaerella species isolated from the foliage of trees in resistant and susceptible families of E. globulus in a seed orchard at Kinglake West, Victoria, Australia. The M. nubilosa primer pair MNF/MNR was highly specific. A PCR-RFLP system based on the primer pair MCF/MCR, coupled with two restriction enzymes (DdeI and Tru1I), differentiated M. cryptica, M. nubilosa, M. tasmaniensis and M. aff. vespa. One of the unidentified field-isolated Mycosphaerella species was identified as M. grandis on the basis of ITS sequence data while the other species remains unidentified. A PCR-RFLP system based on the primer pair U1F/U1R, coupled with the restriction enzyme Sty1, differentiated between the two unidentified species. Unexpectedly, unlike isolation and culture studies, these assays detected M. nubilosa, M. cryptica and M. grandis in all single lesions examined on both juvenile and adult leaves, and on both highly resistant and highly susceptible E. globulus trees at this site.

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Title: A specific primer PCR and RFLP assay for the rapid detection and differentiation in planta of some Mycosphaerella species associated with foliar diseases of Eucalyptus globulus
Authors/Creators: H.A.G.C. Kularatne (Author/Creator)
A.C. Lawrie (Author/Creator)
P.A. Barber (Author/Creator)
P.J. Keane (Author/Creator)
Publication Details: Mycological Research, Vol.108(12), pp.1476-1493
Publisher: Cambridge University Press
Identifiers: 991005543938007891
Murdoch Affiliation: School of Biological Sciences and Biotechnology
Language: English
Resource Type: Journal article

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