First Report of <i>Macrophomina pseudophaseolina</i> Causing Cladode Rot on <i>Opuntia stricta</i> in Brazil

Cactus cladodes (Opuntia stricta Haw.) are an important forage source for ruminant herds in northeastern Brazil, particularly during prolonged droughts. Under these conditions, severe water stress increases plant susceptibility to infection by soilborne fungi. In March and April 2024 (rainy season), cladodes of the ‘Orelha de Elefante’ forage palm were collected in the city of São Rafael, Rio Grande do Norte, Brazil (5°46’52.3” S, 36°52’57.1” W), showing symptoms of rot and necrosis on the cladodes, initially at the basal portion. Initial lesions were moist, light to dark brown, and progressed to whitish areas with longitudinal cracks, suggesting possible epidermal detachment. Disease incidence among symptomatic plants was estimated at 30% of the field. Symptomatic tissues from ten plants were cut into small pieces, surface-sterilized in 70% ethyl alcohol and 1.5% NaClO for 1 minute each, placed onto potato dextrose agar (PDA), and incubated at 25°C for 4 days. Dark to grayish fungal colonies developed, and pure cultures were obtained by hyphal tip isolation. Four isolates with similar morphology were used (CFC-1779, CFC-1780, CFC-1781, and CFC-1782). Based on colony morphology and the presence of microsclerotia, the isolates were preliminarily associated to Macrophomina spp. (Goidanich, 1947). For molecular detection, genomic DNA was extracted, and PCR was performed using species-specific primers: MpTefF/MpTefR for M. phaseolina, MsTefF/MsTefR for M. pseudophaseolina, and MeTefF/MeTefR for M. euphorbiicola (Santos et al., 2020). Amplification occurred only with MsTefF/MsTefR, identifying the isolates as M. pseudophaseolina (Sarr et al., 2014). The translation elongation factor (tef1-α) gene was amplified with primers EF1-728F/EF1-986R (Carbone and Kohn, 1999). A phylogenetic analysis using maximum likelihood and maximum parsimony confirmed the species identity. Sequences were deposited in GenBank with the accession numbers PQ679944 (CFC-1779), PQ679941 (CFC-1780), PQ679943 (CFC-1781), and PQ679942 (CFC-1782). Pathogenicity was tested on symptomless cladodes of ‘Orelha de Elefante’ forage palm, the test was repeated. After surface disinfestation of cladodes in 1.5% sodium hypochlorite for 3 minutes and subsequent drying, 7-day-old mycelial discs of each isolate were placed in 0.8 circular wounds made in the cladodes. PDA discs without mycelium served as controls. Cladodes were maintained at 25 ± 2 °C in a humid chamber for 7 days. Rot symptoms developed on cladodes inoculated with all isolates, whereas controls remained symptomless. Reisolation was performed to confirm Koch's postulates. The pathogens were reisolated and presented the same morphology as the inoculated isolates. Although several Botryosphaeriaceae species have been associated with cladode diseases, this is the first report of M. pseudophaseolina causing cladode rot in O. stricta in Brazil. M. pseudophaseolina has already been reported infecting crops such as Arachis hypogaea, Gossypium hirsutum, and Ricinus communis, in Brazil (Machado et al., 2019). Considering the strategic role of this crop as a forage source for ruminants in semiarid regions, the disease may threaten future production by causing direct losses in cladode yield, reducing forage availability during drought periods, and persisting in the soil through microsclerotia, which can favor its spread and long-term survival in cultivation areas.