First report of charcoal rot caused by macrophomina phaseolina in sunflower in Turkey

Official URL: http://dx.doi.org/10.1094/PDIS-09-10-0631

Charcoal rot symptoms were observed in 2-month-old oilseed sunflower plants (Helianthus annuus L.) in Eskişehir Province of Turkey in June 2009. The disease was observed in 70% of the fields surveyed and incidence ranged from 10-50%. Symptoms were first observed in plants approaching physiological maturity and consisted of silver-gray lesions girdling the stem at the soil line, reduced head diameter compared to non-infected plants, and premature plant death. Pith in the lower stem was completely absent or compressed into horizontal layers. Black, spherical microsclerotia were observed in the pith area of the lower stem, underneath the epidermis, and on the exterior of the taproot. The internal stem had a shredded appearance. Later, the vascular bundles became covered with small, black flecks or microsclerotia of the fungus. Forty plant samples were collected from ten fields. After surface sterilization with 2% NaClO, outer tissues sampled from diseased tissues (2-3mm in length) of root and stems were removed and transferred to potato dextrose agar containing 250 mg l-1 of chloramphenicol. Petri plates were incubated for 7 days at 26±2°C in the dark. Ninety-eight percent of the fungal colonies were identified as Macrophomina phaseolina (Tassi) Goidanich based on gray colony color, colony morphology, and the size of the microsclerotia, which ranged from 80-90 μm in diameter, from both infected sunflowers and compared to pure cultures (1). All resulting cultures produced abundant microsclerotia. The only other sunflower pathogen known to form microsclerotia is Verticillium dahliae Kleb., whose microsclerotia are irregular in shape and 15-50 μm in diameter. Sequence-related amplified polymorphisms (SRAP) technique was used for diversity of M. phaseolina since it has proven to be more informative than AFLP, RAPD and SSR (2). Results showed a high level of genetic diversity (%60) among the 26 isolates of M. phaseolina. Sequencing of ITS region (3) showed high homology (>96%) to M. phaseolina (HQ380051). Pathogenicity tests for 20 isolates of M. phaseolina were carried out on three commercially used cultivars, SANAY, TUNCA and TR-3080. Groups of ten seedlings were grown separately in an autoclaved peat/soil mixture in 30-cm-diameter plastic pots in a greenhouse at 30 ± 2°C. Soil infestation was performed one day before sowing. Two-week-old cultures on Barley medium (4) were blended in distilled sterile water and adjusted to 105 sclerotia ml-1. Each pot received 250 ml of inoculant. Each treatment had three replications. Three pots for each cultivar were left un-inoculated. Within 3 weeks, 5-7 inoculated plants in each pot died. Identical disease symptoms were observed 30 days after inoculation; on the control plants no symptoms were observed. Microsclerotia were produced after 7 weeks at the stem base on 85% of the surviving plants. To our knowledge, this is the first report of M. phaseolina in sunflower in Turkey.