First Report of White Root Rot Caused by Rosellinia necatrix on Persea americana in South Africa

Avocados are an important fruit crop in tropical and subtropical areas of the world. In September 2016, rapidly declining trees (grafted on Phytophthora cinnamomi–tolerant rootstocks) were observed in orchards in Tzaneen, Limpopo, South Africa. Aboveground symptoms included leaf yellowing and wilting. In severe cases, the entire tree became brown and necrotic, resulting in death within a few weeks. Dark brown lesions were present at the base of the trunk, with white mycelial strands extending under the bark. Infected roots were necrotic and covered with a fine layer of mycelia. White shoestring mycelia were present in the surrounding soil. Symptomatic bark was surface disinfected and placed on half-strength potato dextrose agar (PDA) for 5 to 7 days at 25°C. Single aerial hyphae from the edges were transferred onto PDA and incubated at 25°C in the dark. Mycelia from isolates with characteristic pyriform swellings at the septa were used for DNA extractions with PrepMan Ultra Sample Preparation Reagent (Applied Biosystems). For all isolates, the internal transcribed spacer region of rDNA was amplified with primers ITS1/ITS4, beta-tubulin with T10/Bt2B, and actin with ACT-512F/ACT-783R; they were sequenced, subjected to BLAST analysis, and deposited in GenBank (MF611983, MG202164, and MG273315, respectively). All sequences shared 100% identity to Rosellinia necatrix Berl. Ex Prill (Petrini 1993) (EF592565, KY624310.1, and EF025603.1). To confirm pathogenicity, three isolates were used to inoculate 21 1-year-old, clonal Dusa rootstock plants. Sterilized bamboo sticks (2 × 0.5 cm) were placed on PDA containing R. necatrix and left for 7 days. Four colonized sticks (and noncolonized for controls) were placed in the soil around each plant, 3 cm from the stem. Plants were maintained in the greenhouse at 25°C for 4 to 6 weeks. Inoculated plants exhibited chlorosis, leaf wilting, and root rot. By 6 weeks leaves were brown and brittle, and plants had died. White mycelia grew in the soil and on roots. Control plants remained healthy during the trial. R. necatrix was consistently isolated from diseased roots and was morphologically identical to field isolates, fulfilling Koch’s postulates. In addition, 10 plants were used for stem inoculations with the same three isolates. The outer bark was removed with a cork borer (0.5 cm), and mycelial plugs were placed in the hole and wrapped with Parafilm. Five weeks postinoculation dark-brown lesions developed on stems and reisolations confirmed R. necatrix as the pathogen. Control plants had small lesions, but the pathogen was not isolated. One culture was deposited in the CMW collection at the Forestry and Agricultural Biotechnology Institute, University of Pretoria, South Africa (CMW 50481). R. necatrix threatens fruit tree crops in tropical and temperate climates and is important on avocado in Spain and Israel (CABI 1987; López et al. 2008; Schena et al. 2008). White root rot has been present on apples and pears in the Western Cape, South Africa, since the 1970s (Van der Merwe and Matthee 1974) and in the neighboring country of Mozambique (CABI 1987) but has not yet been reported on avocado in South Africa. The recent discovery poses an emerging threat to the industry that currently relies on P. cinnamomi–tolerant rootstocks for sustainable production. It also highlights the importance of restricting movement of plants and soil between areas. We suggest that this is most likely how the pathogen spread throughout southern Africa. To our knowledge, this is the first report of white root rot on avocado in South Africa.