Abstract Background Plants can recruit beneficial microbes to enhance their ability to defend against pathogens. However, in contrast to the intensively studied roles of the rhizosphere microbiome in suppressing plant pathogens, the collective community-level change and effect of the phyllosphere microbiome in response to pathogen invasion remains largely elusive. Results Here, we integrated 16S metabarcoding, shotgun metagenomics and culture-dependent methods to systematically investigate the changes in phyllosphere microbiome between infected and uninfected citrus leaves by Diaporthe citri, a fungal pathogen causing melanose disease worldwide. Multiple microbiome features suggested a shift in phyllosphere microbiome upon D. citri infection, highlighted by the marked reduction of community evenness, the emergence of large numbers of new microbes, and the intense microbial network. We also identified the microbiome features from functional perspectives in infected leaves, such as enriched microbial functions for iron competition and potential antifungal traits, and enriched microbes with beneficial genomic characteristics. Glasshouse experiments demonstrated that several bacteria associated with the microbiome shift could positively affect plant performance under D. citri challenge, with reductions in disease index ranging from 65.7 to 88.4%. Among them, Pantoea asv90 and Methylobacterium asv41 identified as “recruited new microbes” in the infected leaves, exhibited antagonistic activities to D. citri both in vitro and in vivo, including inhibition of spore germination and/or mycelium growth. Sphingomonas spp. presented beneficial genomic characteristics and were found to be the main contributor for the functional enrichment of iron complex outer membrane receptor protein in the infected leaves. Moreover, Sphingomonas asv20 showed a stronger suppression ability against D. citri in iron-deficient conditions than iron-sufficient conditions, suggesting a role of iron competition during their antagonistic action. Conclusions Overall, our study revealed how phyllosphere microbiomes differed between infected and uninfected citrus leaves by melanose pathogen, and identified potential mechanisms for how the observed microbiome shift might have helped plants cope with pathogen pressure. Our findings provide novel insights into understanding the roles of phyllosphere microbiome responses during pathogen challenge.