First report of Lasiodiplodia theobromae causing dieback of Common Fig ( Ficus carica ) in Morelos, Mexico

The common fig (Ficus carica L.) is widely cultivated in semi-arid and subtropical regions. In June 2025, five-year-old fig trees cv. Black Mission exhibiting dieback symptoms were observed in Morelos, Mexico (18°52'29.59"N, 98°47'01.65" W). Among 650 trees assessed, disease incidence was 18%. Affected trees exhibited foliar chlorosis, progressive branch dieback, canopy decline, and retention of dried leaves on affected shoots. Fungal isolates were obtained from twenty symptomatic fig trees. Tissues were surface-disinfected using 3% NaOCl for 5 min, rinsed twice with autoclaved distilled water, and then placed on potato dextrose agar (PDA; Bioxon®) medium. Ten single-spore isolates with morphological characteristics consistent with Lasiodiplodia spp. were obtained and maintained on PDA. The plates were incubated at 25°C ± 2 °C for 14 days, during which colonies produced abundant white aerial mycelium that turned grey after 14 days and developed black pycnidia. Microscopic examination revealed conidia that were initially hyaline, aseptate, thick-walled, subovoid to ellipsoid with rounded ends and granular content. Mature conidia became dark brown, one-septate, and longitudinally striate. Conidia measured 18.33–23.63 (mean = 20.66) µm in length and 11–15.3 (mean = 12.48) µm in width (n = 100 per isolate). Two representative isolates were selected for molecular and pathogenicity analyses, and deposited in the Culture Collection of Phytopathogenic Fungi of the Department of Agricultural Parasitology at Chapingo Autonomous University under accession numbers UACh-LH1 and UACh-LH2. For molecular identification, genomic DNA was extracted from the mycelium following the cetyltrimethylammonium bromide method (Doyle and Doyle 1990). The internal transcribed spacer (ITS) region, parts of the beta-tubulin (β-tubulin) and partial sequences of the translation elongation factor 1-alpha (TEF) genes were amplified by PCR, and sequenced using the primer pairs ITS5/ITS4 (White et al. 1990), Bt2a/Bt2b (Glass and Donaldson 1995) and EF1-728F/986R (Carbone and Kohn 1999), respectively. BLASTn analyses showed that UACh-LH1 had 99.79, 100, and 99.32% identity, whereas UACh-LH2 had 100, 99.77, and 96.67% identity for ITS, β-tubulin, and TEF, respectively, compared with the ex-neotype culture CBS 164.96 of Lasiodiplodia theobromae. Sequences were deposited in GenBank under accession numbers ITS: PZ051210, PZ091917; β-tubulin: PZ056668, PZ279902; TEF: PZ279900, PZ279901. A phylogenetic analysis using concatenated sequences with the Neighbor-Joining method placed isolates UACh-LH1 and UACh-LH2 in the same clade as L. theobromae. Pathogenicity was evaluated in two pathogenicity assays, each independently repeated twice. In the first assay, one-year-old healthy potted fig plants cv. Black Mission were sprayed with a conidial suspension (1.75 × 10⁵ spores mL⁻¹), with five plants inoculated with UACh-LH1 and five with UACh-LH2. Five control plants received sterile distilled water. All plants were incubated at 25 ± 2 °C and 80% relative humidity. Inoculated plants developed dieback symptoms and pycnidia after 35 days, whereas controls remained symptomless. A second assay was performed under field conditions using fig trees cv. Black Mission. For each isolate, 10 branches, 10 leaves, and 10 fruits were wound-inoculated with PDA plugs containing actively growing mycelium. Inoculated tissues were enclosed in poly-bags to maintain 100% relative humidity. Corresponding control tissues (10 branches, 10 leaves, and 10 fruits) were inoculated with sterile PDA plugs. Necrotic lesions developed after 7 days, whereas controls remained symptomless. Lesion length was measured using a digital caliper. The pathogen was successfully reisolated from all inoculated plants and tissues, whereas no fungi were recovered from controls, thus fulfilling Koch's postulates. The reisolated cultures exhibited the characteristic colony and conidial morphology of Lasiodiplodia, consistent with the original isolates. Both pathogenicity assays were independently repeated twice, with similar results. To our knowledge, this is the first report of Lasiodiplodia theobromae causing dieback of Ficus carica in Mexico. This report expands the documented host–disease distribution of L. theobromae in Mexico and provides a baseline for disease diagnosis and surveillance in fig-producing areas.