Applications of Molecular Markers and DNA Sequences in Identifying Fungal Pathogens of Cool Season Grain Legumes

Abstract

Molecular techniques have now been widely applied in many disciplines of biological sciences including fungal identification in microbial ecology and in plant pathology. In plant pathology, it is now common to use molecular techniques to identify and study plant pathogens of many agronomical and horticultural crops including cool season grain legume crops. In this chapter, we present two examples in which molecular techniques have been applied in order to identify and investigate multiple fungal pathogens causing two important diseases of chickpea and lentil. In each case, molecular techniques improved over traditional morphological identification and allowed timely and unambiguous identification of fungal pathogens. The first example involves identification of two Sclerotinia species (S. sclerotiorum and S. trifoliorum) causing stem rot of chickpea. Traditional method requires induction of carpogenic germination and observation of dimorphic ascospores in S. trifoliorum, which takes up to eight weeks. Taking advantage of the group I introns present in the nuclear small subunit rDNA of S. trifoliorum but absent in the same DNA region of S. sclerotiorum, a simple PCR amplification of the targeted DNA region allowed timely and reliable differentiation and identification of the species. The second example is of powdery mildew of lentil. Identification of powdery mildew fungi requires observing the teleomorphic (sexual) state of the pathogens, but this is not always available. In studying lentil powdery mildew in the US Pacific Northwest, we found that the powdery mildew on lentil does not fit previously reported species (Erysiphe pisi and E. diffusa). Further investigation confirmed that the lentil powdery mildew in the US is E. trifolii, a new pathogen of lentil. This discovery was mainly based on the rDNA ITS sequences and further confirmed by morphological and pathogenicity studies. These two examples demonstrate the important role of modern molecular techniques in solving practical agricultural problems. The ITS and adjacent rDNA could be ideal target regions for developing DNA barcodes for identifying these and related fungal species.