Quantitative detection of pear-pathogenic Stemphylium vesicarium in orchards

ABSTRACT Isolates of Stemphylium vesicarium causing brown spot of pear can be distinguished from nonpathogenic isolates of S. vesicarium from pear or from other hosts on the basis of distinctive amplified fragment length polymorphism fingerprinting profiles. DNA fragments specific for isolates pathogenic to pear were identified and a quantitative polymerase chain reaction (PCR) was developed on the sequence from one of these specific DNA loci. This TaqMan PCR has a high sensitivity with a dynamic range for reliable quantification between 1 ng and 100 fg of DNA. The method detected pear-pathogenic isolates of S. vesicarium originating from four different European countries and various regions within those countries. No cross-reaction was found with either the nonpathogenic isolates of S. vesicarium tested or isolates belonging to other Stemphylium spp. or related fungi. The pathogen was detected on leaves with brown-spot symptoms originating from six different locations in The Netherlands, Italy, and Spain. Pear-pathogenic S. vesicarium populations were monitored on crop residues in two Dutch orchards between October 2007 and October 2008. Brown spot had been observed at both orchards at the end of the growing season of 2007. In one location, pear-pathogenic S. vesicarium was detected only sporadically on crop residues and no brown-spot symptoms were observed on fruit in 2008. At the other location, a pathogenic population was found on fallen pear leaves and on other crop residues but this population decreased during winter. From the beginning of the growing season in 2008 onward, the pathogen population could not be detected and the disease incidence was only 0.6%. The TaqMan PCR will allow more detailed studies on epidemiology of brown spot and on the effect of disease control measures.