Ceratocystis paradoxa, the causal agent of stem-bleeding disease of the coconut palm, causes great losses to the global coconut industry. As the mechanism of pathogenicity of C. paradoxa has not been determined, an exogenous gene marker was introduced into the fungus. In this study, pCT74-sGFP, which contains the green fluorescent protein (GFP) gene, and the hygromycin B resistance gene as a selective marker, was used as an expression vector. Several protoplast release buffers were compared to optimize protoplast preparation. The plasmid pCT74-sGFP was successfully transformed into the genome of C. paradoxa, which was verified using polymerase chain reaction and green fluorescence detection. The transformants did not exhibit any obvious differences from the wild-type isolates in terms of growth and morphological characteristics. Pathogenicity tests showed that the transformation process did not alter the virulence of the X-3314 C. paradoxa strain. This is the first report on the polyethylene glycol-mediated transformation of C. paradoxa carrying a ‘reporter’ gene GFP that was stably and efficiently expressed in the transformants. These findings provide a basis for future functional genomics studies of C. paradoxa and offer a novel opportunity to track the infection process of C. paradoxa.

Coconut (Cocos nucifera L.) cultivation is increasingly threatened by destructive fungal pathogens that reduce yield and compromise tree health, particularly in tropical regions in China. To address this challenge, the present study aimed to isolate and identify antagonistic bacteria with potential biocontrol activity against key pathogens of coconut, including Ceratocystis paradoxa (crown rot), Pestalotiopsis menezesiana (leaf blight), and Curvularia oryzae (leaf spot). A total of 65 bacterial strains were isolated from 58 soil samples collected from healthy coconut orchards. Among these, strain X1 exhibited the most vigorous antagonistic activity, with inhibition rates exceeding 70% against multiple pathogens in both plate confrontation and mycelial growth rate assays. Morphological, physiological, biochemical, and molecular (16S rDNA and gyrB gene sequencing) analyses confirmed the identity of strain X1 as Bacillus velezensis. Optimization of fermentation conditions for strain X1 revealed that maximum antimicrobial activity was achieved using a medium containing 2% glucose, 3% peptone, 0.3% NaH₂PO₄·2H₂O, and 0.4% Na₂HPO₄·2H₂O, at 28 °C and pH 7.0–7.5, 5% inoculum, 50–70 mL liquid volume in 250 mL flasks, 72 h fermentation, and agitation at 180–220 rpm. These findings highlight B. velezensis X1 as an up-and-coming biocontrol agent with dual functionalities: effective suppression of coconut fungal pathogens and potential plant growth promotion. Its application could significantly reduce the dependence on synthetic fungicides, offering an eco-friendly alternative for integrated disease management in coconut farming.

Areca palm yellow leaf phytoplasma (AYLP) and Areca palm velarivirus 1 (APV1) are two critical pathogens associated with yellow leaf in areca palm, resulting in devastating damage to areca production. However, evidence of their coinfection remains unclear. Areca palms showing yellow leaf symptoms (suspected to be caused by phytoplasma and velarivirus 1) were surveyed and sampled around Hainan Island of China, and the pathogens were identified and analyzed in the study. The results showed that infectious leaf yellowing symptoms of areca palms primarily occurred in eastern and southern cities and counties of Hainan Island. Molecular identification revealed coinfection of ‘Candidatus Phytoplasma asteris’ (16SrI-B subgroup) and APV1 in individual areca palm samples. Among 520 samples tested by qPCR, AYLP was detected in 23.46% (n = 122), APV1 in 53.46% (n = 278), and a coinfection with both AYLP and APV1 in 10.96% (n = 57). A statistically significant difference in APV1 detection rate was observed between asymptomatic and symptomatic plantations (P < 0.0001), suggesting that APV1 infection likely represents an important risk factor for yellowing development in cultivated areca palm populations. These results confirm the coinfection of AYLP and APV1 on Hainan Island, offering epidemiological evidence for targeted disease management.