First Report of Stem Blight of Blueberry in California Caused by Neofusicoccum parvum.

In July 2013 in coastal (Santa Barbara County) California, commercial plantings of southern highbush blueberry (Vaccinium corymbosum) developed symptoms of a previously undiagnosed disease. Symptoms consisted of reddening and wilting of foliage, with leaves and small twigs later drying up. The bark of diseased branches was discolored and sunken; removal of this bark revealed a brown discoloration of the underlying wood. Approximately 5% of the planting was affected. When placed on acidified potato dextrose agar (A-PDA), surface disinfested pieces of symptomatic wood consistently yielded one type of fungus. On A-PDA, isolates produced extensive white aerial mycelium that turned dark gray after 4 to 5 days and formed pycnidia after 21 days. Three single-spore isolates were grown on PDA for 21 days for morphological and molecular characterization. Conidia were hyaline, smooth, and ellipsoid with round apices and truncated bases. Conidia measured 13 to 20 × 5 to 7.5 μm (n = 50; mean 16.7 × 6.1 μm), with a length/width ratio of 2.73. After 25 days, conidia became biseptate with a darker middle cell. rDNA sequences of the internal transcribed spacer (ITS) region of the isolates (GenBank KJ126847 to 49), amplified using primers ITS1 and ITS4 (5), were 99% identical to the holotype isolate of Neofusicoccum parvum Pennycook and Samuels (3) by a BLAST query (GU251125). Partial sequences of the translation elongation factor 1-alpha (EF1-α) gene (KJ126850 to 52), obtained using primers EF728Fa and EF986R (5), were 99% identical to N. parvum (GU251257). To demonstrate Koch's postulates, 14-day-old colonies of the three N. parvum isolates were grown on A-PDA. Using three blueberry cultivars (Abundance, Jewel, and Snowchaser), slits were cut beneath the epidermis of branches 1 cm diameter or less; one colonized agar plug (6 mm diameter) was placed into each cut and the epidermis was resealed with Parafilm. Ten inoculations (one inoculation per branch; two branches per plant) were made for each isolate and each cultivar; inoculated plants were maintained in a greenhouse. After 10 to 14 days, leaves on inoculated branches turned red and wilted, bark above and below the inoculation sites turned brown, and vascular tissue beneath the bark was also brown. After 21 days, diseased areas became sunken. N. parvum was recovered from all inoculated branches of all cultivars and matched the characteristics of the original isolates. Control branches, inoculated with sterile agar plugs, did not develop any symptoms and N. parvum was not isolated. This experiment was repeated with similar results. Many Botryosphaeriaceae species, including N. parvum, are associated with canker and dieback symptoms on blueberry worldwide (2). To our knowledge, this is the first documentation of stem blight caused by N. parvum on blueberry in CA. Blueberry is a rapidly expanding industry in the state, with 960 ha planted in 2005 increasing to 2,830 ha in 2012 (1). Drought stress predisposes plants to stem blight caused by Botryosphaeriacease species (4); therefore, expansion into arid areas of CA could increase the incidence and severity of N. parvum. References: (1) N. Amer. Blueberry Council. 2012 World Blueberry Acreage & Prod. Rept., 2013. (2) D. F. Farr and A. Y. Rossman. Fungal Databases. Syst. Mycol. Microbiol. Lab., online publication, ARS, USDA. Retrieved February 5, 2014. (3) S. R Pennycook and G. J. Samuels. Mycotaxon 24:445, 1985. (4) W. A. Sinclair and H. H. Lyon. Diseases of Trees and Shrubs, Second Edition. Comstock Publ. Assoc. 2005. (5) B. Slippers et al. Mycologia 96:83, 2004.