ylenchulus semipenetrans is a soil-borne pathogen that causes substantial damage and economic losses to citrus crops worldwide. Due to the high toxicity of chemical nematicides to humans and the environment, biocontrol bacteria have emerged as a promising alternative for managing citrus nematodes. This study aimed to screen bacterial strains for their efficacy to control T. semipenetrans and assess their impact on citrus plant growth. A total of 107 bacterial strains were isolated from the soil and roots of infested citrus trees. Among these, five strains exhibited significant nematicidal activity against T. semipenetrans. Four bacterial densities were tested for each strain: 3.6 × 10⁵, 2.5 × 10⁴, 3.6 × 10³, and 1.2 × 10³ cells/ml. These strains were tested both individually and in combination to evaluate their efficacy. The five strains were identified as Variovorax paradoxus, Bacillus pseudomycoides, Bacillus simplex, Bacillus cereus, and Paracoccus speluncae based on physiological, biochemical, and molecular (16S rRNA gene sequences) analyses. Juvenile mortality (J2s) and egg hatching inhibition were positively correlated with bacterial concentration and exposure duration. The highest juvenile mortality (100%) was observed with a combination of all five bacteria (3.6 × 10⁵ cells/ml) after 96 h, while B. cereus alone achieved 98.98% mortality. The maximum nematicidal activities of the bacterial filtrates were generally observed between the 4th and 6th days of incubation, coinciding with peak bacterial growth and biomass production. The selected isolates also demonstrated the ability to produce indole acetic acid and solubilize phosphorus. In greenhouse experiments, the five isolates reduced T. semipenetrans populations by up to 62.96% compared to the control. Additionally, all rhizosphere bacteria and their combination significantly enhanced plant growth parameters (p < 0.0001). Notably, P. speluncae BR21 has not previously been tested for nematicidal effects on any nematode, making this the first documented report of its nematicidal potential.
