Ecklonia Maxima Extract Effect in Tissue Regeneration of Symbionts at in Vivo Heteroplasmic Grafting of Some Tomatoes

The research was conducted to determine the Ecklonia maxima extract effect in the symbiont accretion at the in vivo heteroplasmic grafting of some tomatoes. E. maxima or sea bamboo is a seaweed used for obtaining of organic extracts used as stimulators in horticulture because consists the natural plant hormones such as auxins and cytokinins which have optimal role in cell division, important activity for tissue regeneration. The experimental variants were grafted plants, combinations between different symbionts, cultivar fragments from Lycopersicon esculentum specie. The symbionts were two scions, F1 hybrids, 'Siriana' (Romanian cultivar), 'Abellus' (Dutch cultivar) and two rootstocks, 'Buzău' variety (Romanian cultivar), 'Emperador' F1 hybrid (Dutch cultivar). The algae extract used had auxins (11 mg/L) and cytokinins (0.3 mg/L). Two treatments were applied before grafting on scion and rootstock, 1 mL/500 mL water and a treatment at grafting on soil, 5 mL/L water. Control variant was without hormone extract. Determinations, observations and interpretations of the algae effect were made on symbionts. The best results on tissue regeneration were obtained in plants treated with sea bamboo extract compared to untreated control plants. The E. maxima extract influenced the tissue regeneration.


INTRODUCTION
Grafting has been used in agriculture for over 2000 years (Haroldesen et al., 2012) and in the horticultural industry (Rivard and Louws, 2006). The tomato grafting has seen important developments between 2010-2015 years at Horting Institute Bucharest, Romania. Blestos and Olympios, 2008 say that grafting of the Solanaceae is an environmentally friendly operation which reduces the incidence of crop infection by soilborne diseases and reduces the population of soil-borne pathogens in a similar way to the application of crop rotation.
Grafting can be defined as the natural or deliberate fusion of plant parts so that vascular continuity is established between them and the resulting genetically composite organism functions as a single plant (Yassin H. and Hussen S., 2015). Pina and Errea, 2005 say they existup different reasons that may have an influence on graft success: inherent system of celular incompatibility, formation of plasmodesmata, vascular tissue connections, and the presence of growth regulators and peroxidases. The in vitro micro-grafting technique could be solved some problems with incompatibility from scion and rootstock (Joley and Opitz, 1971; AI-Barazi and Schwabe, 1982; Sheibani and Villiers, 1995). For Wang and Kollmann, 1996, the formation of vascular connections is considered the basic requirement for a successful graft. Phytohormones coordinate cellular activities. E. maxima (sea bamboo) is a seaweed used for obtaining of organic extracts with hormones such as (auxins and cytokinins) which have optimal role in cell division, important activity during of the symbionts accretion at tomato grafting.

Biological material
The experiment was conducted in 2017, into a specialized greenhouse for production of grafted seedlings from the Department of Horticultural Cultures in Protected Spaces of the Horting Institute Bucharest. The research has followed of the E. maxima extract effect in the symbionts accretion at in vivo heteroplasmic grafting of some tomatoes. The experimental variants were grafted plants, combinations between different symbionts, cultivar fragments from L. esculentum specie. The symbionts were plant fragments from two scions, F1 hybrids, 'Siriana' (Romanian cultivar from the Research and Development Station for Vegetable Growing Buzău, Romania), 'Abellus' (Dutch cultivar from Rijk Zwaan company) and two rootstocks, 'Buzău' variety (Romanian cultivar from the Research and Development Station for Vegetable Growing Buzău, Romania), 'Emperador' F1 hybrid (Dutch cultivar from Rijk Zwaan company).
preparation for grafting with necessary materials (clips, blades), -in vivo heteroplasmic grafting in the phenophase of a two true leaves of the plants; manual cutting to 45 degree and splice grafting of the symbionts in silicone tube of the opposite poles, apical for scion and basal for rootstocks ( Fig. 1 and Fig. 2), interaction between symbiontsforming of callus in polyethylen tunnel with specific microclimate conditions (temperature of 23-25 0 C and relative air humidity of 98-100%), for to stimulate symbionts welding The tunnel was closed for 4 days and ventilated from 5 th day. So, it is realizing a forcing process for tissue regeneration.
transferring of grafted seedlings from tunnel in the growing greenhouse.

Biological treatment
The E. maxima algae extract used had auxins (11 mg/L) and cytokins (0.3 mg/L). Two treatments were applied before grafting, foliar on scion and rootstock, 1 mL/500 mL water and a treatment at grafting, on soil, 5 mL/L water. Control variant was without hormone extract. A trifactorial experience was organized; the experimental factors were: Control, c1with treatment. The variants of working were: a1b1c0, a1b1c1, a1b2c0, a1b2c1, a2b1c0, a2b1c1, a2b2c0, a2b2c1. The reception of the tomato seedlings was carried out after forcing and acclimatization, when the vascular tissues were regenerated.  The degree of callus forming of the grafted plants was determined using a scale ranging from 0 to 4 was used: 0=no callus, 1=25%, 2=50%, 3=75% and 4=100% callus formation on graft union surface (Celik, 2000), table 2. The results of the treatment with E. maxima extract on tissue regeneration at in compatible partners are shown in Fig. 3.

Fig. 3: Percentage of grafted tomatoes with tissue regeneration (%)
In Fig. 3, there is a direct linear correlation between untreated and treated variants with E. maxima extract and the percentage of grafted plants with tissue regeneration. The value of the determination coefficient shows that the significance of the correlation is very significant in the case of the percentage of grafted tomatoes with tissue regeneration (R 2 = 1). 95% of the grafted plants treated with E. maxima extract were have callus, then the callus cells were determined the tissue regeneration of the compatible partners compared to untreated grafted plants, with a grafting rate of 80%. Grafted plants that did not produce callus on the surface of the sections were dehydrated, the tissues oxidized, the medullary rays of the partners (scion and rootstock) were necrotic. Several researchers have studied the influence of the E. maxima extract on the plant processes.

V.
CONCLUSION Objective of this research was to analyze the effect of the E. maxima (sea bamboo) extract in tissue regeneration of the symbiots at the in vivo heteroplasmic grafting of some tomatoes. Results obtained allow concluding that the E. maxima extract has a biostimulant effect in the symbionts accretion at the in vivo heteroplasmic grafting of the investigated tomatoes. The best results on tissue regeneration were obtained on plants treated with sea bamboo extract compared to untreated plants, control; the significance of the correlation is very significant.