Physiological, transcriptomic, and metabolomic analyses reveal that sp. YSD J2 inoculation improves the accumulation of flavonoids in L. var. sativus.

Plant growth-promoting microorganisms (PGPMs), such as sp. YSD J2, promote plant development and stress resistance, while their role in flavonoids accumulation still needs to be further understood. To investigate the complex flavonoid biosynthesis pathway of L. var. sativus (tigernut), we compared sp. YSD J2 inoculation (YSD J2) and water inoculation (CK) groups. YSD J2 significantly elevated the content of indole-3-acetic acid (IAA) and orientin. Furthermore, when analyzing flavonoid metabolome, YSD J2 caused increased levels of uralenol, petunidin-3--glucoside-5--arabinoside, luteolin-7--glucuronide-(2 → 1)-glucuronide, kaempferol-3--neohesperidoside, cyanidin-3--(2″--glucosyl)glucoside, kaempferol-3--glucuronide-7--glucoside, quercetin-3--glucoside, luteolin-7--glucuronide-(2 → 1)-(2″-sinapoyl)glucuronide, and quercetin-4'--glucoside, which further enhanced antioxidant activity. We then performed RNA-seq and LC-MS/MS, aiming to validate key genes and related flavonoid metabolites under YSD J2 inoculation, and rebuild the gene-metabolites regulatory subnetworks. Furthermore, the expression patterns of the trans cinnamate 4-monooxygenase (), flavonol-3--L-rhamnoside-7--glucosyltransferase (), shikimate -hydroxycinnamoyltransferase (), chalcone isomerase (), flavonol synthase (), and anthocyanidin synthase () genes were confirmed by qRT-PCR. Additionally, 4 transcription factors (TF) (especially bHLH34, Cluster-37505.3) under YSD J2 inoculation are also engaged in regulating flavonoid accumulation. Moreover, the current work sheds new light on studying the regulatory effect of sp. YSD J2 on tigernut development and flavonoid biosynthesis.