First Report of Xylella fastidiosa in Avocado in Costa Rica

Abstract

Since the late 1990s, chlorotic mottling, marginal scorch, deformation of leaves, defoliation, shortening of internodes, and branch dieback have been observed in avocado trees (Persea americana Mill.) in Costa Rica. The symptoms are not uniformly distributed in the tree, so some branches are symptomatic while others are not. These symptoms are similar to several leaf scorch diseases caused by the bacterium Xylella fastidiosa Wells (2,4). This bacterium has been detected in coffee and citrus plants in Costa Rica. Of 227 avocado trees tested by double-antibody sandwich (DAS)-ELISA with X. fastidiosa specific antiserum (Agdia Inc., Elkhart, IN) from 2000–2004, 188 were positive. Results of ELISA tests of individual trees varied with the season and branches tested. Fifteen greenhouse-grown, ELISA-negative avocado seedlings were grafted with budwood from an ELISA-positive tree. Eight of these developed scorch symptoms and one also showed chlorotic mottling and deformation, showing that the disease is graft transmitted. All of these features are characteristic of diseases caused by X. fastidiosa (2,4). Transmission electron microscopy of leaf petioles from three field trees positive by ELISA, revealed rod-shaped bacilli approximately 1.6 to 2.0 μm long and 0.3 μm in diameter with a rippled cell wall inside xylem vessels and embedded in a matrix; morphology and measurements that are consistent with those reported for X. fastidiosa (2). DNA extraction and PCR attempts have been limited by mucilaginous sap from avocado. Positive PCR results (approximately 472-bp band) were obtained from two of the grafted seedlings and seven field trees from two distinct geographical locations (Alajuela and San José provinces) with DNA extractions from the plant sap using DNeasy Plant Mini Kit (Qiagen GmbH, Hilden, Germany) following a modified protocol (1) and nested PCR (3). Four of the PCR products, including one from the grafted seedlings, were cloned and sequenced in duplicate. GenBank sequences EU021997 to EU022000 present 99 to 100% sequence identity to a Pierce's disease strain from California (Temecula1) and 94 to 95% to a citrus variegated chlorosis strain from Brazil (Found-5). Several attempts have been made to isolate the bacterium in ‘periwinkle wilt’ and buffered cysteine-yeast extract media with negative results, probably because of the rapid production of mucilaginous sap when the avocado tissues were sampled. To our knowledge, this is the first report of X. fastidiosa in avocado trees.