NLR genes are crucial components of the effector-triggered immunity (ETI) system, responsible for recognizing pathogens and initiating immune responses. Although NLR genes in many plant species have been extensively studied, the diversity of NLR genes in citrus remains largely unknown. Our analysis revealed significant variations in the copy numbers of NLR genes among these species. Gene duplication and recombination were identified as the major driving forces behind this diversity. Additionally, horizontal gene transfer (HGT) emerged as the principal mechanism responsible for the increase in NLR gene copy number in A. buxifolia. The citrus NLR genes were classified into four categories: TIR-NBS-LRR (TNL), CC-NBS-LRR (CNL), RPW8-NBS-LRR (RNL), and NL. Our findings indicate that TNL, RNL, and CNL genes originated from NL genes through the acquisition of TIR and RPW8 domains, along with CC motifs, followed by the random loss of corresponding domains. Phylogenetic analysis suggested that citrus NLR genes originated alongside the species and underwent adaptive evolution, potentially playing crucial roles in the global colonization of citrus. This study provides important insights into the diversity of citrus NLR genes and serves as a foundational dataset for future research aimed at breeding disease-resistant citrus varieties.

HLB is a severe and devastating disease affecting citrus plants, for which no cure is currently available. The pathogen CLas is an unculturable, phloem-limited α-proteobacterium associated with HLB. Phages, known for their ability to directly lyse pathogenic bacteria, have been widely recognized as effective control agents in both medical and agricultural fields. In this study, we identified 191 prophages across 48 Liberibacter genomes using comparative genomics and phylogenetic methods. These prophages were classified into 17 distinct types, with 13 novel types, named NF1 to NF13. Diversity analysis revealed that the number of prophages per genome ranged from one to eight, with an average of four. Annotation showed that the genome sizes of NF1–NF13 ranged from 10,160 bp to 72,736 bp, with an average size of 27,616 bp, containing between 7 and 66 genes (average: 27 genes). Further functional enrichment of the sequences within 5 kb upstream and downstream of the integration site identified six main aspects, involving four core biological processes: genome structure modification, gene expression regulation, stress response activation, and metabolic pathway modulation. It is speculated that the insertion of the prophage significantly impacts the host. Six virulence factors, including carB, clpC, flhA, algW, rffG, and IlpA, were identified in the prophages. Interestingly, prophages containing carB or clpC are predominantly found in CLas, which may contribute to the global colonization of CLas. Notably, among the 35 identified Type 4 prophages, 19 were found to be active. Type 4 is the only active prophage type, making it a promising candidate for developing phage therapy against CLas. This study is significant for a deeper understanding of the diversity of Liberibacter and the interactions between prophages and their hosts.

NLR genes are crucial components of the effector-triggered immunity (ETI) system, which recognizes pathogens and initiates immune responses. While NLR genes in many plants have been extensively studied, the diversity of NLR genes in citrus remains largely unknown. In this study, we systematically identified 1,585 NLR genes across the genomes of 10 citrus species. Significant variations in NLR gene copy numbers were observed among these citrus species, with gene duplication and recombination identified as major driving forces. Citrus NLR genes were classified into TIR-NBS-LRR (TNL), CC-NBS-LRR (CNL), RPW8-NBS-LRR (RNL), and NL categories. It is believed that TNL, RNL, and CNL genes originated from NL genes through the acquisition of TIR, RPW8 domains and CC motifs, respectively, followed by the random loss of the corresponding domains. Phylogenetic analysis revealed that citrus NLR genes originated with the species and underwent adaptive evolution, potentially playing crucial roles in the global colonization of citrus. This study is importance for comprehensively understanding citrus NLR genes and provides a foundational dataset for research aimed at breeding disease-resistant citrus varieties.