Morphological and Molecular Identification of Neopestalotiopsis Species Causing Leaf Blight and Shoot Dieback on Avocado in Tropical Orchards

Abstract

Leaf blight and shoot dieback symptoms causing severe decline in avocado orchards were observed in tropical production regions characterized by high humidity and frequent rainfall. Symptomatic leaves and shoots exhibiting necrotic lesions, chlorosis, and progressive twig dieback were collected for isolation and identification of the associated fungal pathogen. Fungal cultures consistently produced fast-growing colonies with white to gray aerial mycelia and dark sporulating structures on culture media. Microscopic examination revealed fusiform, multi-septate conidia with characteristic apical and basal appendages typical of the genus Neopestalotiopsis. Considerable variation in colony texture, pigmentation, and sporulation intensity was observed among isolates recovered from different orchards. Pathogenicity assays performed on healthy avocado seedlings and detached leaves reproduced typical blight and dieback symptoms, confirming the pathogenic nature of the isolates. Re-isolation of the fungus from inoculated tissues fulfilled Koch’s postulates. Molecular characterization based on sequencing of the internal transcribed spacer (ITS), translation elongation factor 1-α (TEF1-α), and β-tubulin gene regions confirmed the isolates as species belonging to Neopestalotiopsis. Phylogenetic analysis grouped the obtained isolates with reference strains of pathogenic Neopestalotiopsis species with strong bootstrap support, demonstrating close genetic relationships among isolates from geographically distinct orchards. Environmental conditions prevailing in tropical orchards, particularly prolonged leaf wetness and elevated humidity, favored disease development and pathogen dissemination. The occurrence of Neopestalotiopsis species on avocado represents an emerging threat to tropical avocado cultivation because of their capacity to cause extensive foliar damage and shoot decline.