RESUMEN
In Argentina, olive (Olea europaea L.) is cultivated in the mountainous, warm, arid northwest (Andes range), where Fusarium solani (blue sporodochia) is frequently found to be causing death of nursery and young field plants (1). Recently, olive orchards were established more than 1,600 km to the southeast (Pampas) in a plain with a temperate and humid climate and in the arid Patagonia, both influenced by the Atlantic Ocean. This area includes part of Buenos Aires and Rio Negro provinces. In March 2008, 10-year-old 'Barnea' olive trees with high incidence of root rot, dried leaves, dead branches, and dead plants were observed in the Coronel Dorrego District of Buenos Aires Province, where oat, barley or other cereals are planted between rows of olive trees. Planting material originated from olive nurseries located in Mendoza Province, 1,200 km from Coronel Dorrego. Diseased roots were disinfected in 2% NaOCl and 70% ethanol, cut into small pieces, plated onto rose bengal-glycerin-urea medium, and incubated at 20°C with a 12-h photopheriod. A fungus was purified through successive transfers of hyphal tips from the margin of a sparsely growing colony onto 2% water agar (2). Colonies grown on Spezieller Nährstoffarmer agar (3) and carnation leaf-piece agar were used for morphological identification, and those on grown on potato dextrose agar were used for evaluation of pigmentation and colony growth rate. Sporodochium color, cream, was typical of F. solani (Mart.) Sacc. This isolate was deposited in the IMYZA Microbial Collection as INTA-IMC 73. Mycelium was cultured in liquid Czapek-Dox medium supplemented with sucrose, peptone, yeast extract, sodium nitrate, and vitamins for 4 days and fungal DNA was obtained with a DNA extraction kit. Primers ITS1 and ITS4 were used to amplify the internal transcribed spacer (ITS) region of ribosomal genes. The purified PCR product was sequenced and the DNA sequence compared with GenBank records. The sequence shared 100% identity with 27 entries for F. solani and 97% identity with F. solani obtained from olive in Nepal (4), corresponding to EU912432 and EU912433. The nucleotide sequence was registered in GenBank as JF299258. Pathogenicity was confirmed on 'Manzanilla' plants at the eight-leaf stage. Pieces of water agar with mycelium were applied to small wounds at the stem base and on roots of 10 plants and were covered with cotton soaked in sterile distilled water. Plants were incubated at 20°C and a 14-h photoperiod. On control plants, water agar pieces without mycelium were applied to the wounds. After 33 days, inoculated plants showed dark brown lesions (average length 1.4 cm) and leaf chlorosis. Two plants showed wilting with leaves remaining attached to branches. F. solani was reisolated from roots and stem bases of inoculated plants. Controls remained asymptomatic. To our knowledge, this is the first report of F. solani occurring on olive in the temperate part of the Pampas of Argentina where cereals, which are susceptible to Fusarium species, are grown with olive trees. Sporodochium color (cream) of these isolates differed from the blue color of previously reported isolates of F. solani on olive in northwestern Argentina (1). References: (1) S. Babbitt et al. Plant Dis. 86:326, 2002. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Ames, IA, 2006. (3) H. I. Nirenberg. Releases Fed. Biol. Res. Center Agric. For. (Berlin-Dahlem) 169:1, 1976. (4) A. M. Vettraino et al. Plant Dis. 23:200, 2009.
RESUMEN
In October 2007, blighted shoots were observed on highbush blueberry (Vaccinium corymbosum L. cv. O'Neal) plants in La Plata, Buenos Aires Province, Argentina. Isolations from surface-disinfested shoots onto carrot agar and Spezieller Nahrstoffarmer Agar (SNA) consistently yielded white colonies that produced black sclerotia, mainly near the edge of the culture plates, after 7 days. Sclerotia were transferred to SNA tubes and kept at 5°C for several months. The germination of sclerotia produced numerous 6 mm long initials, stipitate pale brown cup-shaped apothecia (10 × 6 mm) with eight-spored asci (137 × 7 µm) at 18°C and continuous light conditions. Asci with uniseriate ascospores were cylindrical and narrow at the base. Ascospores (11 to 12 × 4 µm) were hyaline, unicellular, smooth, and ellipsoid. This isolated fungus was morphologically identified as Sclerotinia sclerotiorum (Lib.) de Bary (2,3). The isolate was deposited in the IMYZA Microbial Collection as INTA-IMC 87. Mycelium was cultured in 100 ml of Czapek's-Dox medium, supplemented with sucrose, peptone, yeast extract, sodium nitrate, and vitamins (1), for 3 days and fungal DNA was obtained using a DNA extraction kit. ITS1 and ITS2 of ribosomal genes were amplified by PCR using universal primers (4) and the PCR product was sequenced. A BLAST algorithm search revealed 100% identity of the sequence with 12 GenBank entries for S. sclerotiorum. The nucleotide sequence was deposited in the GenBank with Accession No. JF277567. Pathogenicity testing was achieved by placing detached leaves of cvs. Emerald, Misty, and Start on water agar (WA) plates, inoculating with 9-mm2 mycelial blocks, and incubating at 20°C with 12 h of light. Young shoots of highbush blueberry, Misty and O'Neal, were inoculated by the cut shoot method with micropipette tips filled with mycelium and kept under greenhouse conditions at 24°C and 14 h of light. On control plants, WA blocks or WA-filled micropipette tips were used. Leaf blight was observed after 5 to 6 days and sclerotia appeared after 7 days on inoculated tissues. Shoot blight was recorded on inoculated plants after 5 days. The fungus was reisolated from inoculated tissues, with no symptoms showing on controls. To our knowledge, this is the first report of Sclerotinia rot caused by S. sclerotiorum in blueberry in Argentina. References: (1) J.F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual. Blackwell Publishing, Hoboken, NJ, 2006. (2). J. E. M. Mourde and P. Holliday. No. 513 in: CMI Descriptions of Pathogenic Fungi and Bacteria. Kew, Surrey, UK, 1976. (3) S. Umemoto et al. Gen. Plant Pathol. 73:290, 2007. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.