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 (CLas), severely limits citrus production worldwide. We investigated how oxytetracycline (OTC) trunk injection affects the citrus holobiont, examining its ability to suppress CLas and improve tree performance while assessing compartment-specific responses of the microbiome and resistome. A field experiment was conducted in CLas-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 CLas 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.