From Traditional to Omics-Driven: Emerging Strategies for Isolation, Cultivation, and Identification of Plant Endophytes.

Plant endophytes can regulate host plant growth, improve stress resistance, and facilitate the biosynthesis of secondary metabolites, with great research value and application potential. However, traditional approaches for the isolation, cultivation and identification of plant endophytes are constrained by low culturability, limited species diversity, and loss of their original ecological functions inside host tissues. In recent years, integrated multi-omics strategies combining metagenomics, metatranscriptomics, and metaproteomics have exhibited the greatest potential to mitigate culturability limitations by enabling genome-guided targeted strain isolation and in situ functional activity profiling, among which the cultivation and targeted isolation of endophytes benefit most from omics integration. These approaches drive a paradigm shift from conventional blind screening to precise targeted isolation, and from generic medium culture to omics-guided rational cultivation, greatly improving the accuracy of strain identification and functional characterization. Nevertheless, current omics-based strategies still face inherent limitations including high experimental costs, complex operational procedures, and challenging data interpretation. The most critical future direction lies in establishing standardized experimental protocols and shared resource databases, combined with microfluidic platforms and artificial intelligence-assisted bioinformatics analysis, to address the core bottlenecks restricting endophyte isolation, cultivation and identification. This review is the first to systematically summarize research progress on traditional approaches, omics technologies and emerging strategies for plant endophyte isolation, cultivation and identification, highlights prevailing challenges and developmental trends in this field, and provides methodological references for the efficient exploitation and sustainable utilization of plant endophyte resources.