First Report of Stalk Rot Disease of Sugarcane Caused by Phaeocytostroma sacchari in Mexico.

In April 2009 and 2010, severe symptoms of stalk rot of sugarcane (Saccharum officinarum L.) plants cvs. MEX-79-431, MEX-69-749, and RB-73-9735 were observed in commercials fields located in southeast Michoacan state, Mexico. The diseased plants exhibited complete discoloration of foliage, ascendant necrosis and rot in the internal stalk tissue, and disintegration of vascular tissue. Symptoms were most evident in the nodes with intense purple coloration. Dead plants were observed. Two diseased plants of each cultivar were collected. Pieces of symptomatic stem tissue were surface sterilized with 2% sodium hypochlorite solution for 1 min, washed with sterile distilled water, dried on sterilized paper, and plated onto potato dextrose agar (PDA). Petri dishes were incubated at 22°C under continuous white light for 72 h. A fungus was consistently isolated. On PDA, colonies had sparse aerial mycelium in the center and dense in the margins with black masses of conidia. The fungus isolated was grown on dishes containing 2% water agar (WA) overlaid with pine needles and incubated at 22°C under continuous white light for 2 weeks to induce the formation of fruiting bodies. Pycnidia produced in WA were black, up to 500 μm in diameter, usually globose, blister shaped without peaks, scattered, and multilocular. Conidiophores were cylindrical, hyaline, 5 to 20 × 1.5 to 2 μm, and formed in the pycnidial cavity. Conidia were ellipsoidal to oblong, unicellular, pale brown to dark brown, 8.5 to 12.5 × 3 to 4.5 μm, biguttulate, and non-septate. Paraphyses were hyaline, aseptate, occasionally branched, and flexuous. On the basis of cultural and morphological characteristics, the fungus was identified as Phaeocytostroma sp. DNA from an isolate was extracted and the internal transcribed spacer region (ITS1-5.8S-ITS2) of rDNA was amplified using primers ITS1 and ITS4 (2). PCR products were purified and sequenced. The resulting sequence of 536 bp was deposited in GenBank (Accession No. KC893550). BLAST analysis showed a 99% similarity with the sequence of Phaeocytostroma sacchari (FR748047). Pathogenicity tests of an isolate of P. sacchari were performed on 6-month-old sugarcane plants (cvs. MEX-79-431, MEX-69-749, and RB-73-9735). A 1-cm-deep wound near the base of the stem was created with a sterilized needle. Mycelial plugs (9 mm diameter) of 6-day-old PDA cultures were deposited on wounds and wrapped with Parafilm. Four plants of each cultivar were inoculated and 12 control plants were treated similarly with PDA plugs instead of fungal inoculum. Plants were placed at 28°C and 95% relative humidity for 72 h. All the inoculated plants exhibited typical wilt symptoms 4 weeks after inoculation, whereas control plants remained healthy. P. sacchari was consistently re-isolated from artificially inoculated plants. To our knowledge, this is the first report of P. sacchari on sugarcane in Mexico. The occurrence of stalk rot disease of sugarcane caused by P. sacchari has been described causing severe losses in sugarcane-producing countries such as South Africa and India (1). References: (1) R. Viswanathan et al. Sugar Tech. 5: 61, 2003. (2) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.