Transcriptome profiling reveals differential expression of virulence genes in Xylella fastidiosa under nutrient-rich and xylem-like conditions

Xylella fastidiosa is a xylem-limited, insect-vectored pathogen. To capture the metabolic changes that underlie its adaptation to both nutrient-rich and xylem-like environments, we performed transcriptome analyses on two reference strains, Temecula1 and 9a5c, sampled at early and late exponential phases in a complex laboratory medium (PWG) and a defined, xylem-mimicking medium (PIM6). Across eight conditions (24 transcriptomes), over 90% of annotated coding sequences were expressed, although 40-80% of transcripts exhibited low abundance (TPM < 100). In contrast, a core set of highly abundant transcripts was seen in both strains and media, including the non-coding RNAs 6S and RNase P, as well as multiple colicin- and microcin-like toxins, proteases, lipases, and stress-response factors. Comprehensive pathway reconstruction confirmed the activity of all essential metabolic pathways, while revealing conditional enrichment of stress-related and defense modules under nutrient limitation. We annotated 5' and 3' untranslated regions for thousands of transcripts and defined over 900 operons across both genomes. Differential expression analysis demonstrated that PIM6 medium confers a far greater transcriptional reprogramming between early and late phases than PWG. Strain-to-strain comparisons revealed that up to 12.6% of orthologous genes were differentially regulated, emphasizing divergent regulatory networks even under identical growth conditions. This multi-dimensional transcriptome atlas not only refines X. fastidiosa genome annotation but also pinpoints candidate regulators, non-coding RNAs, and operon variants for functional validation. By illuminating the dynamic gene expression landscapes that enable xylem colonization and environmental resilience, our study lays a foundation for targeted disruption of key pathways in this important plant pathogen.