| Summary: | In 2017–2018, extensive symptoms of sudden decline and fruit rot were observed on date palms in southern Tunisia. Samples of diseased plants were randomly collected in six localities. Based on morphological identification, <i>Fusarium</i> was the most frequent fungal genus detected. A sequencing of translation elongation factor, calmodulin, and second largest subunit of RNA polymerase II genes was used to identify 63 representative <i>Fusarium</i> strains at species level and investigate their phylogenetic relationships. The main species detected was <i>Fusarium proliferatum</i>, and at a much lesser extent, <i>Fusarium brachygibbosum</i>, <i>Fusarium caatingaense</i>, <i>Fusarium clavum</i>, <i>Fusarium incarnatum,</i> and <i>Fusarium solani</i>. Pathogenicity on the <i>Deglet Nour</i> variety plantlets and the capability to produce mycotoxins were also assessed. All <i>Fusarium</i> species were pathogenic complying Koch’s postulates. <i>Fusarium proliferatum</i> strains produced mainly fumonisins (FBs), beauvericin (BEA), and, to a lesser extent, enniatins (ENNs) and moniliformin (MON). All <i>F. brachygibbosum</i> strains produced low levels of BEA, diacetoxyscirpenol, and neosolaniol; two strains produced also T-2 toxin, and a single strain produced HT-2 toxin. <i>Fusarium caatingaense</i>, <i>F. clavum</i>, <i>F. incarnatum</i> produced only BEA. <i>Fusarium solani</i> strains produced MON, BEA, and ENNs. This work reports for the first time a comprehensive multidisciplinary study of <i>Fusarium</i> species on date palms, concerning both phytopathological and food safety issues.
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