Materials and MethodBacterial Strains and DNA ExtractionA collection of standard bacterial strains containing E. amylovora strains and several species of bacteria confirmed by biochemical, carbohydrate and virulence tests for identification of E. amylovora isolates (data not shown) were exploited to estimate the specificity test (Table 1). Furthermore, approximately 208 symptomatic plant samples were used to evaluate the performance of two PCR methods and the LAMP assay. This collection was obtained from various plant tissues (e.g. flowers, shoots, leaves, fruits and branches) belonging to apple, pear and quince cultivars from different regions of Iran, during the spring and summer of 2009 and 2010. To prepare the samples, the same method as (Gorris et al. 1996) was used: 100 μl of each dilution and other standard bacteria routinely cultured on Luria-Bertani (LB) agar or LB agar medium and incubated at 28°C for 48 hours. Below, total genomic DNA from each strain was isolated by lysing bacterial pellets from 1 ml broth culture, incubated overnight in DNA extraction buffer, purified with phenol-chloroform-isoamyl alcohol (25:24: 1) and precipitated with isopropyl alcohol (Llop et al. 1999; Schaad et al. 2001). Finally, each strain DNA was eluted in 100 µl of elution buffer and stored at -20 ◦C before further evaluation. whenever pure bacterial cultures were used, the optical density of the bacterial culture was measured using a spectrophotometer at 600 nm (2 × 107 CFU/ml), and one μl of each dilution of bacterial suspension was added directly to the LAMP and PCR reaction mixture . For samples of infected plants, however, Plant Mini Kit (Qiagen) was used. DNA isolation was performed according to the manufacturer's protocol for...... half of the paper ...... clearly specific-specific for E. amylovora. Comparison of LAMP with Conventional and Nested PCR Assays To evaluate the ability of LAMP and other PCR methods to detect E. amylovora in naturally infected plant material (described above), first the LAMP method was tested with infected plant material. Among the three methods, the LAMP assay showed the highest power for pathogen detection in all symptomatic samples (Table 3). Unlike single PCR and nested PCR, electrophoresis is not required in the LAMP method and the detection time is reduced to 45 minutes. The existence of the pathogen in the positive samples was confirmed by its isolation in the culture medium. These remarkable results demonstrated that, on the one hand, the method has higher specificity and sensitivity than single PCR and, on the other hand, it is slightly better than PCR nested in a single closed tube.