Citrus- associated Alternaria species in the Mediterranean areas

Alternaria brown spot is one of the most important diseases of tangerines and their hybrids worldwide. Recently, disease outbreaks in Southern Italy, Spain, and Greece refocused the attention on the disease. Twenty representative cultures of Alternaria were selected from a collection of more than 150 isolates from leaves and fruits of cvs Fortune, Nova, Valencia, and Tangerine. They were characterized with specimen strains of A. tenuissima, A. alternata, A. arborescens, A. citri, A. toxicogenica, and A. limoniasperae (‘small-spored’ Alternaria species) to determine the etiology of the disease and evaluate the virulence of different isolates/species. Morphological characteristics and sporulation patterns separated most Alternaria isolates into three main groups corresponding to A. alternata, A. arborescens, and A. tenuissima, whose the first was the most abundant one. Phylogenetic analyses based on endopolygalacturonase (endoPG) and beta-tubulin genes, two anonymous genomics regions (OPA 1-3 and OPA 2-1), and the internal transcribed spacer (ITS) region produced a clustering of isolates largely confirming morphological results. The OPA 1-3 region was more suitable than the other tested regions for separating closely related ‘small-spored’ Alternaria species and revealed the existence of intra-species molecular variability. Investigated isolates showed different levels of virulence on leaves and fruits but it was not possible to identify a direct correlation between virulence and genetic/morphological groupings of isolates. The toxigenic potential of Alternaria strains was investigated. The twenty strains were assayed for the production of tenuazonic acid (TeA), alternariol (AOH), alternariol monomethyl ether (AME), altenuene (ALT) and tentoxin (TEN). The TeA was the most abundant toxin, produced in vitro in the range 0.2-20 mg/L. Most of the strains were able to synthesize AOH, AME and ALT, although at a lesser extent. The widespread occurrence of Alternaria in citrus and its ability to produce mycotoxins might represent a serious concern for producers and consumers.