Compartmental and functional responses of the citrus microbiome and resistome to the systemic delivery of oxytetracycline by trunk injection

Huanglongbing (HLB), caused by Candidatus Liberibacter asiaticus ( C Las), severely limits citrus production worldwide. We investigated how oxytetracycline (OTC) trunk injection affects the citrus holobiont, examining its ability to suppress C Las and improve tree performance while assessing compartment-specific responses of the microbiome and resistome. A field experiment was conducted in C Las-infected sweet orange trees, integrating qPCR pathogen quantification, fruit yield and juice quality measurements, functional pathway analysis, and genome-resolved profiling across leaves, bark, fibrous roots, and the rhizosphere at three time points after injection. OTC reduced C Las abundance in leaves and improved fruit yield and juice quality without altering microbial diversity. No clear OTC-associated shifts in microbial functional pathways were observed in aboveground compartments, and resistome profiles were strongly compartment-dependent but showed no detectable response to OTC treatment. However, pronounced functional shifts were detected in belowground compartments, with consistent reductions in carbon-, nitrogen-, and phosphorus-related pathways and declines in several taxa and metagenome-assembled genomes associated with nutrient turnover. In contrast, stress-tolerance and xenobiotic-responsive microorganisms were enriched. In addition, these belowground responses were associated with low-abundance, rare taxa rather than by changes in alpha diversity or the dominant community, revealing a hidden functional reconfiguration that was concentrated in the root and rhizosphere compartments most relevant to nutrient cycling and long-term soil health. These findings demonstrate that systemically delivered OTC induces targeted, compartment-specific reorganization of microbiome functions rather than broad disruption. By linking physiological improvement with functional and genome-resolved microbial responses, this study highlights the broader ecological consequences of antibiotic interventions in perennial crops.