Physiological Effects Induced by Copper Excess in Peach Rootstock MrS 2/5 and GF 677 Growth In Vitro

In this work the physiological effect of copper excess on the peach rootstock MrS 2/5 and GF 677 was studied by means of an in vitro system. Two different copper treatments (10 and 100 μM CuSO4) for each rootstock were carried out and compared to a control containing 0.1 μM of CuSO4. Relative growth rate and copper concentration inside rootstock tissues were measured after 21 days from the beginning of the treatments and used to compare the genotypes’ reaction to copper excess. The GF 677 in vitro plantlets did not show any significant decrease in relative growth rate, while it increased in MrS 2/5 plantlets exposed to 100 μM CuSO4. In both rootstocks, the copper concentration in plantlet tissues increased when CuSO4 changed from 0.1 to 10 and 100 μM. INTRODUCTION Copper (Cu) is an essential micronutrient for plant growth and development (Fernandes and Heriques, 1991). The Cu role in plants is quite complex, being involved in protein and carbohydrate metabolism, chlorophyll biosynthesis, respiration and photosynthesis. Most metalloenzymes containing Cu are involved in the catalysis of reactive oxygen species, as in the copper-zinc superoxide dismutase isoform, that is mostly associated with chloroplasts in higher land plants (Asada et al., 1977). Via the polyphenol metabolism, copper affects cell wall lignification, thus influencing the plant’s water balance (Fernandes and Heriques, 1991). Optimal copper levels for plant nutrition are quite narrow (8-20 ppm) (Howeler, 1983; Stevenson 1986), and deleterious effects usually appear when copper concentration slightly exceeds these optimal levels. Copper excess induces a wide range of biochemical effects and metabolic disturbances in plants that are responsible for a strong inhibition of growth, which is sometimes accompanied by anomalous development. Copper concentrations in non-contaminated soils and sediments are usually low (20-30 ppm) (Salomons and Forstner, 1984). Moreover, due to the strong binding of Cu with organic matter and other soil colloids, only a limited fraction of the total amount of soil Cu is available for plant uptake. Sewage sludge amendments and cupric fungicide treatments can gradually increase the Cu levels in cultivated soils. In some vineyard cultivation areas, the copper content in soil has reached concentrations of 500 ppm (Brun et al., 1998), which could be potentially toxic for plants. Fruit tree rootstocks provide the means to regulate growth, precocity, fertility and yield of the scion cultivar (Beckman et al., 1992) and to solve problems associated with soils, climate or phyto-sanitation (Loreti, 1988), modified water balance, nutrient uptake and other characteristics of the grafted cultivar (Alvino et al., 1991). In peach cultivation, the rootstocks GF 677 (P. persica × P. amygdalus) and P. cerasifera L. “MrS 2/5” are diffuse because of their ability to growth in several adverse soil conditions. The rootstock MrS 2/5 is widely used for its reduced pathogen and nematode sensitivity, while the hybrid GF677 is most known for its tolerance to drought conditions and for resistance to iron-induced chlorosis (Carrera, 1992) The aim of this work was to study by means of an in vitro system the physiological effects of copper excess on the peach rootstocks MrS 2/5 and GF677. In Proc I IS Rootstocks – Decid. Fruit Eds. M.Á. Moreno Sánchez and A.D. Webster Acta Hort 658, ISHS 2004 52 vitro treatments can be, in fact, a fast and useful method to evoke copper toxicity symptoms and to study genotype reactions to copper excess. MATERIALS AND METHODS The in vitro shoots utilised for copper toxicity experiments were obtained from stock cultures of GF 677 (P. persica × P. amygdalus) and P.s cerasifera sel. MrS 2/5. Stock cultures were kept in MAGENTA vessels filled with 50 ml of Murashige and Skoog (1962) medium (MS) supplemented with 30 g l sucrose, 7 g l Difco Bacto-agar, 6-benzylaminopurine (BAP) 2.7x10μM, gibberellic acid (GA3) 5.8 x10 μM and indole3-butyric acid (IBA) 3x10μM (MS-Control medium). The pH was adjusted to 5.2 before autoclaving the medium at 120°C for 20 min. Plantlet growth conditions were a temperature of 24°C and 16/8h day/night photoperiod at a light intensity of 55 μmol PAR m s. After 3 weeks, the terminal (1 cm) part of the shoots (obtained from the stock cultures) was excised and weighed under sterile conditions. Explants were transferred (6 explants vessel) into two MAGENTA vessels containing: a) MS-Control medium (0.1 μM CuSO4); b) MS-Control medium + 10 μM CuSO4; c) MS-Control medium + 100 μM CuSO4. Growth conditions were set as previously described. Plantlets were harvested for analysis of dry weight after 21 days ( 2 DW ) from the beginning of the treatments. Dry weight was recorded on oven dried (60°C) plantlets until constant weight was reached. Relative growth rate (RGR) was calculated as: