First Report of Brown Spot Caused by <i>Alternaria alternata</i> on <i>Prunus persica</i> in Brazil

Peach production plays a key role in the agricultural sector of Southeastern Brazil. In 2023, the country produced a total of 200,710 tons of peaches with a production value of approximately R$ 646,834,000.00 (IBGE, 2025). Alternaria alternata is a widely distributed plant pathogen known to cause fruit rot and leaf spot symptoms in peach trees (Prunus persica) (Thomidis et al., 2007; Abata et al., 2016; Alam et al., 2019). In September 2024, dark brown spots with black halos, measuring around 1.5-2 cm, were observed on the leaves of peach trees grown in an experimental orchard located in Piracicaba, São Paulo, Brazil (22°42'28”S, 47°37'42”W). A field survey indicated that 80% of the plants showed symptoms of the disease. Alternaria sp. was isolated from symptomatic tissues and cultured on V8 medium by surface-disinfecting the samples with 70 % ethanol and 10 % sodium hypochlorite. The isolates were incubated at 25 °C under a 12-hour photoperiod using black light (F30W T12 Sylvania) for five days. Colonies ranging from grey to olivaceous brown and dark blackish brown, producing conidia of varying color from light to dark, consistently grew from the samples. Conidia were obclavate to obpyriform in shape, measuring 16-39 × 7-17 μm, and containing 3-7 transverse septa. The morphological characteristics of the conidia were consistent with the descriptions available for A. alternata (Marin-Felix et al., 2019). Genomic DNA was extracted from a single-spore isolate using the Wizard® Genomic DNA Purification Kit. The internal transcribed spacer (ITS), partial glyceraldehyde-3-phosphate dehydrogenase (gapdh), partial DNA-directed RNA polymerase II second largest subunit (rpb2), and partial translation elongation factor 1-alpha (tef1-α) genes were amplified and sequenced. The sequences were deposited in GenBank (ITS - PX118603, gapdh - PX139704, rpb2 - PX139705 and tef1-α - PX139706), and revealed 99-100% identify with those of the ex-type isolate of A. alternata CBS 916.96 (AF347031, AY278808, KC584375 and KC584634). Phylogenetic analysis of the concatenated loci performed using Bayesian Inference, placed our isolate in a well-supported clade together with CBS 916.96. Pathogenicity tests were conducted on potted plants, 1.5 years old, with young shoots emerging after pruning, under controlled greenhouse conditions (25 ± 5 °C). A conidial suspension (5 × 10⁴ conidia.mL⁻¹) containing 0.05% Tween 20 was sprayed onto fully expanded leaves until runoff using an airbrush. Control plants were sprayed with distilled water under the same conditions. After inoculation, all plants were kept in a moist chamber in the dark at 23 ± 2 °C for 24 hours and then transferred to the greenhouse. The experiment followed a completely randomized design with five replicates, each consisting of one potted plant. The assay was performed twice. Fifteen days after inoculation, small chlorotic spots developed on the adaxial leaf surface of inoculated plants, evolving into characteristic brown spot symptoms by 40–45 days. Symptomatic leaf tissues were processed for pathogen reisolation and identified based on morphology and molecular sequencing. Control plants remained asymptomatic. To the best of our knowledge, this is the first report of A. alternata causing brown spot on peach trees in Brazil. While known as a postharvest pathogen, its impact on leaves had not been recorded. Foliar diseases affect photosynthesis and shift disease management to consider the entire crop cycle.