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Origanum vulgare L, recognized throughout the world as a popular medicinal and flavoring herb, contains a wide array of medicinally active components, including phenolic glucosides, flavonoids, tannins, sterols and high amounts of terpenoids. Especially the latter are often extracted by hydrodistillation resulting in the so-called essential oil that is rich in monoterpenes (e.g. carvacrol, thymol, linalyl acetate) and/or sesquiterpenes (e.g. (E)-beta-caryophyllene, germacrene D, bicyclogermacrene, 13-caryophyllene oxide). Water stresses in the arid and semiarid regions of the world severely affect growth and productivity of oregano. To determine the variation in essential oil and gene expression pathway of Iranian oregano under prolonged water stress, two native subspecies of 0. vulgare (subsp. virens and subsp. gracile) were studied. The plants, grown in pots, were subjected to three water stress conditions, i.e. no stress, mild stress (60 +/- 5% FMC) and moderate stress (40 +/- 5% FMC). The studied subspecies exhibited significant differences in essential oil content, compositions, and patterns of gene expression under water stress conditions. The essential oil of O. vulgare subsp. gracile was rich in the phenolic monoterpene carvacrol (46.86-52.07%), whereas the sesquiterpene hydrocarbon (Z)-alpha-bisabolene (39.17-42.64%) was the major constituent in the oil of O. vulgare subsp. virens. Both the mild and moderate water stresses significantly increased the essential oil content of O. vulgare subsp. gracile, but did not significantly change the essential oil content of O. vulgare subsp. virens nor the level of carvacrol and (Z)alpha-bisabolene in the investigated subspecies. Interestingly, the amount of (E(beta-caryophyllene in O. vulgare subsp. virens was significantly increased under water stress conditions. Gene expression studies supported the above findings and demonstrated that there are two different pathways affecting the biosynthesis of the terpenoid precursors geranyl pyrophosphate (GPP) and farnesyl pyrophosphate (FPP). In O. vulgare subsp. gracile, HMGR, Ovtps2 and CYP71D180 transcript were up -regulated under mild and moderate water stress conditions. Transcription of FPPS was apparently down -regulated in water stressed O. vulgare subsp. gracile. Investigation of terpene synthases expression levels in oregano subspecies demonstrated that Ovtps2 and Ovtps6 controlled the concentration of carvacrol and (E)-beta-caryophyllene in oregano essential oils, respectively. (C) 2016 Elsevier Masson SAS. All rights reserved.