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<i>Lasiodiplodia</i> (family <i>Botryosphaeriaceae</i>) is a widely distributed fungal genus that causes a variety of diseases in tropical and subtropical regions. During 2020–2021, a routine survey of fruit tree plants was conducted in five Egyptian Governorates, and fresh samples exhibiting dieback, decline, leaf spot and root rot symptoms were collected. Collection from eight different symptomatic leaves, twigs, branches and roots of fruit trees yielded 18 <i>Lasiodiplodia</i>-like isolates. The sequencing data from the internal transcribed spacer region (ITS), partial translation elongation factor 1-alpha (<i>tef1-a</i>) and <i>β-tubulin</i> (<i>tub2</i>) were used to infer phylogenetic relationships with known <i>Lasiodiplodia</i> species. Two isolates obtained from black necrotic lesions on <i>Phoenix dactylifera</i> leaves were identified as a putative novel species, <i>L. newvalleyensis</i> sp. nov., and were thus subjected to further morphological characterization. The results of isolation and molecular characterization revealed that <i>L. theobromae</i> (<i>n</i> = 9) was the most common species on <i>Mangifera indica</i>, <i>Citrus reticulata</i>, <i>C. sinensis</i>, <i>Ficus carica</i>, <i>Prunus persica</i>, <i>Prunus armeniaca</i> and <i>Pyrus communis</i> trees. <i>Lasiodiplodia pseudotheobromae</i> (<i>n</i> = 5) was isolated from <i>M. indica</i>, <i>Prunus persica</i> and <i>C. sinensis</i>. <i>Lasiodiplodia laeliocattleyae</i> (<i>n</i> = 2) was isolated from <i>C. reticulata</i>. Pathogenicity test results suggested that all <i>Lasiodiplodia</i> species were pathogenic to their hosts. The present study is considered the first to characterize and decipher the diversity of <i>Lasiodiplodia</i> species associated with fruit trees in Egypt, using the multi-locus ITS, <i>tef1-a</i> and <i>tub2</i> sequence data, along with morphological and pathogenic trials. To our knowledge, this is the first report of <i>L. newvalleyensis</i> on <i>Phoenix dactylifera</i> and <i>L. laeliocattleya</i> on <i>C. reticulata</i> in Egypt and worldwide.