Growth and physiological response of in vitro seedlings to exogenous melatonin under iron deficiency stress in grape

Abstract:【Objective】Exogenous melatonin (MT) can effectively alleviate stress symptoms in many crops, enabling the crop to resist adversity and be recovered from the salt-damaged or deficient environment for a short period of time. The experiment was conducted to study the effects of exogenous MT treatment on the growth and physiology of grapevines under mild iron deficiency stress conditions, so as to analyze the effect of exogenous MT on the alleviation of iron deficiency symptoms in grapevines and its alleviation efficacy.【Methods】The experimental materials were grape seedlings in the test tube grown in laboratory succession, and the cultivar was Crimson Seedless . Four treatments were set up as below: CK (Fe concentration of 50 μmol·L-1 without addition of exogenous MT), CK+MT (Fe concentration of 50 μmol·L^-1 with MT concentration of 140 μmol·L^-1 ), -Fe (Fe concentration of 30 μmol·L^-1 without addition of exogenous MT), and -Fe+MT (Fe concentration of 30 μmol·L^-1 with MT concentration of 140 μmol·L^-1 ). The iron and MT concentrations added in the experiment were based on the previous experiments. The test was conducted by selecting healthy and uniformly growing seedlings, cutting the canes into 1.0-1.5 cm in length with a single bud, and transferring them to the B5 solid medium before starting different treatments as described above. Each treatment was transferred to 20 vials of seedlings, with 2 plantlets per vial, and replicated 3 times. The transferred seedlings were incubated at (27±1) ℃ with a light intensity of 3000 lx and 2∶1 ratio of light to dark period, and the grapevine growth was observed regularly. At day 50 after inoculation, grape seedlings in the test tube were removed from triangular vials and the plant height, stem thickness, plant dry and fresh weight, leaf area and root system indicators were measured. Leaf area and root system indicators were scanned with an Epson root system scanner (STD 4800) and analyzed with a software (Win RHIZO 5.0, Canada). Moreover, those indicators were also determined, including leaf chlorophyll, malondialdehyde, proline, soluble protein and soluble sugar contents, SOD, POD and NR activities as well as root vigor. The data were comprehensively analyzed to study the effects of exogenous MT on the growth of grapevine in vitro seedlings under mild Fe-deficiency stress, and to investigate the alleviating effect of melatonin on Fe-deficiency stress at the physiological level.【Results】In terms of growth indexes, grapevines showed poor growth with yellowing of leaves, dwarf vines and stunted root development under Fe deficiency stress. After the addition of MT, plant growth recovered better and all indexes were significantly better improved than those with Fe deficiency treatment. It was suggested that exogenous melatonin may improve the growth environment of grapevine roots, promote plant growth and improve the resistance of grapevines to iron deficiency adversity by participating in the synthesis pathway of essential substances and regulating the amount of phytohormone synthesis. In terms of physiological indicators, the chlorophyll a, chlorophyll b, carotenoids and total chlorophyll contents in leaves significantly increased by the addition of exogenous MT compared with CK and -Fe treatments, and the most significant effect was relieved by -Fe treatment, indicating that exogenous MT had a good protective effect on chlorophyll structure of grapevine leaves under Fe deficiency stress. Under Fe deficiency stress, the SOD activity as well as Pro and MDA contents of grapevines increased significantly compared with CK treatment, but POD activity decreased; after adding exogenous MT, POD activity increased, and MDA content and SOD activity decreased rapidly. The content of osmoregulatory substances was significantly higher in leaves with -Fe treatment compared with CK, while the addition of MT to CK and -Fe effectively reduced the Pro, soluble protein and soluble sugar contents with the stress treatment (-Fe). Compared with the Fe deficiency treatment, root vigor and NR activity were significantly enhanced by adding exogenous MT.【Conclusion】Under mild Fe deficiency stress, the root volume and leaf area of grape seedlings increased and the vines grew well after the addition of MT. Leaf photosynthetic pigment content, POD and NR activity as well as root vigor increased significantly, while MDA, Pro, soluble protein and soluble sugar contents as well as SOD activity decreased significantly. The 140 μmol·L^-1 MT treatment was effective in alleviating the symptoms of iron deficiency with in vitro seedlings, protecting the plants from physiological damage and ensuring normal growth and development.