Apple replant disease resistance in different apple rootstocks evaluated using microbiomic and metabolomic analyses

Apple replant disease (ARD) is a critical obstacle to the sustainable development of apple (Malus domestica Borkh.) agriculture. The use of resistant rootstocks has been adopted as a method for the long-term, effective prevention and control of ARD. This study evaluated the responses of seven apple rootstocks to ARD stress, including five that have shown ARD resistance (14-30, 14-54, Jizhen 1 [J1], Jizhen 2 [J2], and Jizhen 3 [J3]) and two that are strongly and weakly ARD-resistant (G935 and M26, respectively). We measured plant growth and physiological indices, and determined ARD tolerance using membership function and principal component analyses (PCA). J3 exhibited the best performance among all rootstocks, maintaining higher biomass, photosynthetic rates, chlorophyll content, and superoxide dismutase (SOD) and peroxidase (POD) activity under ARD stress. The rootstocks displayed decreasing ARD resistance in the order: J3 > 14-54 >14-30 > G935 > M26 > J1 > J2. High-throughput sequencing, liquid chromatography–mass spectroscopy, and metabolomics analyses revealed that the high-resistance J3 and low-resistance J2 rootstocks had distinct rhizosphere microbial and metabolic profiles. J3 was associated with greater abundance of beneficial microbes, such as those in the genera Bacillus, Pseudomonas, Pontibacter, and Glomus, which were in turn correlated with improved soil nutrient availability and ARD tolerance. Metabolomic analysis identified 108 differential metabolites including flavonoids and carboxylic acids; compounds such as 4',5,7-trihydroxy-3-methoxyflavanone and trans-3-feruloylcorosolic acid were positively associated with both beneficial microbes and ARD resistance. Thus, the J3 rootstock gained enhanced ARD tolerance through recruiting beneficial rhizosphere microbes and modulating soil metabolites. These results provide a scientific basis for screening ARD-resistant apple rootstocks and improving apple tree breeding programs.