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Molnar, T. J., Walsh, E., Capik, J. M., Sathuvalli, V., Mehlenbacher, S. A., Rossman, A. Y., and Zhang, N. 2013. A real-time PCR assay for early detection of eastern filbert blight. Plant Dis. 97:813-818. Eastern filbert blight (EFB) is a devastating disease of European hazelnut, Corylus avellana, which causes economic losses in Oregon, where 99% of the U.S. crop is produced. The causal fungus, Anisogramma anomala, is native to eastern North America, where it is found associated with the American hazelnut (C. americana). Although C. americana is tolerant, EFB causes cankers, branch dieback, and death of C. avellana. Detection and identification of A. anomala is time consuming using conventional methods because the fungus can only be cultured from sporulating perithecia and the disease symptoms and signs only show 12 to 16 months after infection. In this study, a TaqMan real-time polymerase chain reaction (PCR) assay based on a ribosomal DNA internal transcribed spacer was developed for A. anomala. The assay was validated with multiple isolates of A. anomala, closely related species, common environmental microorganisms, and over 100 C. avellana samples. The real-time PCR assay detected as low as 0.12 pg of A. anomala genomic DNA, and positively diagnosed EFB on 82% of asymptomatic plants as early as 15 weeks from infection. The real-time PCR assay is more sensitive and faster than traditional diagnostic methods. It can facilitate hazelnut breeding and disease management by early and accurate diagnosis of EFB.

Eastern filbert blight (EFB) is a serious disease of European hazelnut, Corylus avellana L., which causes economic losses in Oregon (OR) where 99% of the U.S. crop is produced. The causal organism, Anisogramma anomala (Peck) E. Müller, is native east of the Rocky Mountains where it is found in association with C. americana Marshall. While C. americana is tolerant, EFB causes cankering, branch dieback, and death of C. avellana (3). EFB was first discovered in Washington State in the late 1960s (1). Since then, it has spread throughout the Willamette Valley of OR. In OR, ‘Gasaway’, an obsolete pollinizer, shows complete resistance to EFB, conferred by a dominant allele at a single locus (4). ‘Gasaway’ has been widely used in breeding at Oregon State University (OSU) to develop resistant cultivars that are used in most new orchards. In January 2008, cankers containing rows of dark brown elliptical stroma, characteristic of EFB, were first observed on more than 25 trees of ‘Gasaway’ growing at the Rutgers University research farms in Adelphia and North Brunswick, NJ. At that time, cankers were also found on 18 trees of ‘VR20-11’ growing on the research farms. ‘VR20-11,’ an offspring of ‘Gasaway’ that carries the same resistance gene, was released by OSU for use as a pollinizer for ‘Barcelona’, an EFB-susceptible but widely grown cultivar in OR. Additional cankers were observed on the New Jersey trees in January 2009 and 2010. To our knowledge, this is the first report of EFB on either cultivar under field conditions. The cankers are smaller than those on susceptible cultivars. Of 61 cankers on 10 trees of ‘Gasaway’, the average length was 11 cm with a range of 4 to 42 cm. Canker lengths on susceptible trees are typically 20 to 100 cm. The cankers appear otherwise alike with stromata, 2 to 4 × 2 mm, up to 2 mm high; perithecia upright, in the lower part of stroma; asci ellipsoid, 35 to 45 × 9 to 12 μm; and ascospores 8 to 11 × 4 to 5.5 μm, hyaline, smooth, ellipsoid, 2-celled, with the lower cell very short (1 to 1.5 μm long and wide). Genomic DNA was isolated from ascospores excised from cankers of ‘Gasaway’ and ‘VR20-11’. ITS1F and ITS2 primers were used to amplify and sequence the internal transcribed spacer 1 region (ITS1) of the rRNA genes (GenBank Accession Nos. HM565133 and HM565132). BLAST analysis of the 238-bp segments showed 99% homology with a sequence of A. anomala (EU683064). Phylogenetic analysis also confirmed that the two isolates are A. anomala. To test viability, ‘Gasaway’ cankers were excised and ascospore suspensions (1 × 106 spores ml–1) were applied to 15 trees of susceptible ‘Barcelona’ in March 2008 following the protocol of Johnson et al. (2). In December 2009, 12 of 15 inoculated trees expressed EFB. ‘Gasaway’ has shown no signs or symptoms of infection by A. anomala over several decades of exposure in OR, which is believed to have a limited diversity of the fungus due to a single-point introduction. Our findings suggest quarantine efforts must be bolstered to prevent further introductions of A. anomala into the Pacific Northwest to protect the viability of the U.S hazelnut industry. References: (1) A. D. Davison and R. M. Davidson, Jr. Plant Dis. Rep. 57:522, 1973. (2) K. B. Johnson et al. Phytopathology 84:1465, 1994. (3) K. B. Johnson and J. N. Pinkerton. Eastern filbert blight. Page 44 in: Compendium of Nut Crop Diseases in Temperate Zones. B. L. Teviotdale et al., eds. The American Phytopathological Society. St. Paul, MN. 2002. (4) S. A. Mehlenbacher et al. HortScience 26:410, 1991.