Spiroplasma kunkelii and Spiroplasma citri, both helical‐shaped cell wall‐less bacteria, are the causative agents of corn stunt disease and citrus stubborn disease, respectively. Plants exhibiting natural resistance to these phytopathogenic spiroplasmas are currently lacking. Engineering artificial plant resistance using antimicrobial peptides (AMPs) has been conceived as a new approach to control the agronomically important spiroplasmal diseases. In preparation for such task, the present study focused on screening of AMPs that have potentials to curb the growth of S. kunkelii and S. citri. Four AMPs, including Novispirin T7, Caerin 1.1, Tricholongin and Dhvar4, were selected for in vitro growth inhibition test. A liquid assay method was developed for quick qualitative and quantitative evaluations of the AMPs. Our results demonstrated that Novispirin T7 and Caerin 1.1 were able to inhibit the growth of both phytopathogenic spiroplasmas with the efficacy comparable to that of tetracycline. Cell deformations were observed in spiroplasma cultures treated with these two peptides, indicating interactions of the AMPs with the spiroplasma cell membranes. The minimum inhibitory concentrations (MICs) of the AMPs against S. kunkelii and S. citri were determined. [ABSTRACT FROM AUTHOR]