Comparative genomics of a poinsettia-associated phytoplasma and functional validation of its SAP11-homologous effectors that induce plant branching

Phytoplasmas are insect-transmitted plant pathogens that manipulate host development through secreted effector proteins. While they are notorious for causing agricultural losses, in the ornamental plant poinsettia (Euphorbia pulcherrima), phytoplasma infection is uniquely harnessed to induce the commercially desirable free-branching trait. However, the effectors responsible for this phenotype have remained unknown. To address this question, we sequenced and analyzed the genome of 'Candidatus Phytoplasma pruni' PR2021, a strain associated with the high-branching cultivar Princettia Pink. Comparative genomics confirmed its species assignment and revealed an unusual effector repertoire. PR2021 lacks most previously described effectors but encodes two distinct SAP11 homologs, a family of effectors known to induce plant branching. Genomic context analysis showed that one homolog is located within a potential mobile unit (PMU) and is nearly identical to the SAP11 of the distantly related 'Ca. P. asteris', while the other is located outside PMU regions and is divergent in sequence and predicted structure. Functional assays using Agrobacterium-mediated transient expression in Nicotiana benthamiana demonstrated that each homolog independently induced significant branching, whereas co-expression did not enhance the phenotype, indicating overlapping functions. These findings establish a direct connection between poinsettia branching and SAP11-homologous effectors, providing the first experimental evidence linking phytoplasma effector activity to this horticulturally important trait. This work expands understanding of phytoplasma effector diversity and mobility, while offering a functional framework for developing pathogen-free strategies to modulate ornamental plant architecture.