Occurrence of Phacidiopycnis washingtonensis Causing Speck Rot on Stored Pink Lady Apple Fruit in Chile

Chilean apple (Malus × domestica Borkh.) industry has a production area of 37,207 ha (ODEPA 2015). Recently, speck rot, a new postharvest rot of stored apple caused by Phacidiopycnis washingtonensis Xiao & J.D. Rogers, has been reported in Washington State, U.S.A., and Germany (Kim and Xiao 2006; Weber 2011). Speck rot was observed during storage (150 to 180 days; 0°C) of Pink Lady apples harvested from orchards in Linares (35°52′S; 71°37′W) in the south-central region of Chile during 2014. Speck rot was characterized by an initial light brown skin discoloration that progressed to dark brown to black, with a firm rubbery texture that affected part of the fruit or the entire fruit. Superficial or partially immersed black pycnidia were observed on the surface of the decayed areas. Internally decayed tissue was spongy to firm with a brown-black to black appearance and could not be separated from apparently healthy tissue (Weber 2011). Fifteen diseased fruits were superficially disinfested using 75% ethanol for 30 s and cut. Small fragments (2 to 3 mm in length) were collected from the margins of internal diseased tissues and aseptically placed on acidified potato dextrose agar (APDA; 0.5 ml/liter of 92% lactic acid) for 7 days incubation at 20°C. Fifteen light gray to gray colonies with aerial mycelia and showing alternating growth rings were subcultured in APDA. Abundant black pycnidia with cream-colored cirrhi were often present after 7 to 10 days at 20°C in APDA cultures. The conidia on the APDA were smooth, unicellular, hyaline, ellipsoid to lacriform with flattened ends, and (5.2) 6.6 (7.9) µm × (2.9) 3.6 (4.7) µm. Based on morphology, this fungus was identified asPhacidiopycnis washingtonensis Xiao & J.J Rogers (Xiao et al. 2005). Conidia of P. washingtonensis differ in size and shape with P. piris (Xiao et al. 2005). To confirm its identity, the internal transcribed spacer (ITS) region of rDNA of four isolates (Mz-1, Mz-4, Mz-5, and Mz-9, GenBank Accession Nos. KP759279 to KP759282, respectively) was amplified and sequenced using primer ITS4/ITS5. BLASTn analyses showed 99% identity with P. washingtonensis (ex-type, AY608644). To prove pathogenicity, 12 mature Pink Lady apple fruits were superficially disinfected using 75% ethanol for 30 s and injured with a sterile 5-mm diameter cork borer. Then, apple fruits were inoculated with 40 µl of conidial suspension (106 conidia/ml) of four isolates (Mz-1, Mz-4, Mz-5, and Mz-9). An equal number of fruits treated with 40 µl of sterile water were used as negative controls. The fruits were placed into commercial apple box and incubated at 20°C for 15 days. An additional 12 fruits were disinfested, wounded, and inoculated as described above, including negative controls; however, these fruits were placed in a cold chamber at 0°C for 35 days. All inoculated apple fruits developed necrotic lesions measuring 26.1 to 31.3 mm and 60.1 to 70.4 mm in diameter for 0 and 20°C storage conditions, respectively. The negative controls remained healthy. Reisolations were performed on APDA and the presence of P. washingtonensis was morphologically confirmed in 100% of the symptomatic fruits. To our knowledge, this study is the first report of P. washingtonensis causing speck rot on cold stored apple fruit in Chile and South America. The impact of speck rot on the Chilean apple industry is unknown, but we determined a 0.2 to 13.9% prevalence (n = 9,489 apple fruits) of speck rot on stored Pink Lady apples. © 2016, American Phytopathological Society. All rights reserved.