(2006). The barley cultivar Rihane, which covers >70% of the barley area in Tunisia, was used as a control. Disease severity was assessed 17 days after inoculation according to the rating scale described by Ceoloni (1980). The differential cultivars were scored for resistance (R) and susceptibility (S), and a matrix showing reaction patterns was constructed for the 79 pathotype responses vs. the 19 differential cultivars. Cluster analysis was performed on the pathotype matrix using the unweighted pair group method with arithmetic

averaging of darwin software (http://darwin.cirad.fr/darwin) to determine patterns of pathogenicity of Tunisian R. secalis AZD2281 research buy isolated from local barley landraces and the cultivar Rihane. The susceptibility percentage of 19 differential barley cultivars with known resistance genes to 79 R. secalis isolates sampled from different hosts (Rihane cv. and local

barley landraces) was calculated by host and by differential cultivar to determine the possible resistance genes. To detect new sources of resistance, the reaction spectrum of the 79 R. secalis isolates was compared with pairs of differentials with the same resistance genes Jet and Steudel and Kitchin and Abyssinian for differences in pathogenic reaction. Fungal mycelial DNA was extracted according to the Von Korff et al. (2004) method. Seven microsatellite loci

developed for R. secalis (Linde et al., 2005) were used to fingerprint the 79 isolates. Loci were amplified by multiplex PCR with group I (GA-SSR7, GA-SSR3, GA-R2 and CA-SSR1) Cyclopamine and II (TAC-SSR6, GA-SSR4 and TAC-SSR1) primers on either a Biometra T-gradient or an AB-GeneAmp PCR System 9700 thermocycler, subjected to capillary electrophoresis, and per-locus allele assignments were carried out using an ABI PRISM 310 Genetic Analyzer as described by Linde et al. (2005). SSR data were used to assess the level of genetic polymorphism and clustering. For each locus, we determined the total number of alleles and unique alleles by host and by virulence group. Patterns of genetic variation were determined by host through cluster analysis using the unweighted pair RG7420 in vivo group method as above. The relationship between variation in pathogenicity and the haplotype of microsatellite markers was compared for isolates having the same haplotype, by examining their reaction spectra to 19 differential cultivars. The ratio of the number of differential cultivars showing a coincident reaction to isolates with the same haplotype relative to the differential cultivars was used to calculate the degree of coincidence as described by Takeuchi & Fukuyama (2009). A total of 79 pathotypes were sampled from either Rihane cultivar (43) or local barley landraces (36) from 17 localities.