SEMI-HYDROPONIC CULTIVATION AS A SELECTION TOOL FOR ALUMINUM TOLERANCE IN FORAGE GRASSES

O aprimoramento constante das ferramentas de selecao se faz necessario, no sentido de otimizar a eficiencia de um programa de melhoramento para tolerância ao aluminio. Desta forma, o objetivo deste trabalho foi estabelecer um volume de vaso ideal ao cultivo semi-hidroponico de gramineas forrageiras submetidas ao estresse por aluminio em solucao nutritiva. O delineamento experimental utilizado foi o inteiramente casualizado, com quatro repeticoes, sendo cada repeticao constituida por uma planta por vaso. Os tratamentos foram dispostos em esquema fatorial 4x5, sendo o primeiro fator constituido por quatro genotipos de gramineas forrageiras (Brachiaria Humidicola; Brachiaria brizantha cv. Piata; Panicum maximum cv. Massai e Panicum maximum cv. Mombaca) e o segundo constituido de cinco volumes distintos de vasos plasticos onde as plântulas foram cultivadas (0,2; 0,25; 0,3; 0,35 e 0,4 dm3). As plântulas foram cultivadas em sistema semi-hidroponico com solucao nutritiva rica em aluminio (3 mg L-1). Foram mensuradas a altura de plantas, massa seca da parte aerea e comprimento de raiz. O cultivo semi-hidroponico foi eficaz na diferenciacao de genotipos de gramineas forrageiras em relacao a tolerância ao aluminio. Volumes de vasos proximos a 0,3 dm3 favoreceram o desenvolvimento dos atributos do sistema radicular e da parte aerea em gramineas forrageiras cultivadas em solucao nutritiva com a toxidez de aluminio.


INTRODUCTION
In Brazil, the beef cattle production is characterized by farming systems that use grasslands, providing 99% of the diet for cattle herds.The pastures managed in extensive system, with low use of fertilizers or, in most cases, without the addition of these inputs, so that the productivity levels of livestock systems are low, and over time production, declines are evident (DIAS-FILHO, 2011).
New areas opening for cattle pasture is getting more restrict, and gradually limited to marginal areas with low fertile soil and high acidity, rich in aluminum and other toxic elements (MACEDO, 2009).Aluminum is toxic to most species of cultivated plants by promoting the inhibition of root growth and, consequently, hindering the development of plants (CANÇADO et al., 2001).Thus, the operation of these pasture land need, increasingly, the selection of forage that is adapted to those conditions (EUCLIDES, 2000).
Character aluminum tolerance in forage species has received attention from breeding programs (OLIVEIRA et al., 2013;BITENCOURT et al., 2011;MATINS et al., 2011).According to Ryan et al. (1993), the Al 3+ is present predominantly in the root area of the plant, and the apexes of the roots its critical site of toxicity.Thus, methods that measure restricting root growth receive more attention as a parameter in the evaluation of Al 3+ toxicity (CRESTANI et al., 2011;CRESTANI et al., 2009;VOSS et al., 2006;MAZZOCATO et al., 2002).
Reviews in field conditions have traditionally been used in the plant tolerance study about Al 3+ toxic, by depicting more faithfully the natural growing conditions (MISTRO et al., 2001;CAMARGO et al., 1995;FERREIRA et al., 1986).However, most interference from external factors of hard control in this type of study priority the cultivation in nutrient solution to facilitate the work by environment control and ease visualization of Al 3+ effect on interested attribute (LANA et al., 2013;MACEDO et al., 2011;REIS et al., 2009).Crestani et al. (2009) signalize that growing in nutrient solution also allows evaluating large number of genotypes in short period of time in the early stages of plant development, providing significant improvements to the efficiency of selection.
Different methods based on cultivation in nutrient solutions have been developed in evaluation of tolerance and sensitivity to Al 3+ in annual and perennial species.Cardoso et al. (2004) proposed semi-hydroponics method in nutrient solution in which the species of interest is transplanted, still in seedling stage for a polyethylene vase containing an inert substrate, suspended over a container containing nutrient solution.In this way, contact with the substrate allows plant root system realize gas exchange, while the rest of the roots are immersed in the nutritive solution prepared for each study.For toxicity studies Al 3+ , Furlani and Hanna (1984) showed nutrient solution composed of a series of chemical elements essential for plant associated with the use of aluminum -potassium decahydrate as element source.
However, in all hydroponics system, priority should be given a homogeneous distribution of root system on the nutrient solution.Whereas the availability of nutrients to plants is also affected by morphological and physiological attributes of each plant species (SCHENK and BARBER, 1979), attributes such as length, volume and surface roots must be considered in experimental design.Moreover, plant roots have a high demand for oxygen by the high respiration rate (MARSCHNER, 1995), provide the necessary aeration system becomes essential to plant development in full growth medium.
The constant improvement of methodologies is necessary in order to optimize the evaluation process efficiency and selection of genotypes tolerant to aluminum.Thereby, the aim of this study was to establish the ideal vase volume semihydroponic cultivation of forage grasses subjected to stress by aluminum in nutrient solution.

MATERIALS AND METHODS
The experiment was conducted in greenhouse at the Experimental Station of the Federal University of Tocantins, Campus of Gurupi, at coordinates 11°43'45" latitude and 49°04'07" longitude.The experimental design completely randomized, with four replicates, each replicate consisting of one plant per vase.

350-356p.
The treatments were arranged in a factorial 4 x 5, the first factor consists of four genotypes of forage grasses (Brachiaria humidicola; Brachiaria brizantha cv.Piatã; Panicum maximum cv.Massai and Panicum maximum cv.Mombaça) and the second consisting of five different volumes of plastic pots where seedlings were grown (0.2; 0.25; 0.3; 0.35 and 0.4 dm 3 ).
The seeds of the genotypes germinated on filter paper, spaced from each other in one centimeter.The paper was rolled up and moistened with distilled water.These rolls were placed in germination chamber at 25°C, as described by Brasil (1992).After 60 hours, the rollers were removed and the germinating plants were selected for uniformity.Subsequently, these plantlets were placed individually in plastic vases.
In cultivation of seedlings of grasses used the double vase methodology adapted from Cardoso et al. (2004), where cylindrical plastic pots of 75 dm diameter containing washed sand, were superimposed with the aid of a support, a polyethylene tray measuring 45 cm long, 30 cm wide and 12.0 cm deep, containing nutrient solution (Figure 1).In shared double vases (20 plots per tray) of the roots grown in liquid medium and solid medium part as a way to avoid the necessity of artificial aeration of the solution.Seedlings received daily irrigation with nutrient solution similar to that defined by Furlani and Hanna (1984), however, with a 50% reduction in the concentration of calcium.The aluminum stress was simulated by adding to the nutritive solution at a single concentration of 3 mg L -1 , in the form of aluminum -potassium decahydrate .After 25 days elapsed germination, the plants were picked, separate the shoot roots, and evaluated the plant height (PH), measured in centimeters from the ground base to the apex of the larger sheet; and the root length (RL), measured in centimeters from the insertion base end to the main root.
Subsequently they were identified accommodated in paper bags and placed in a forced ventilation oven at a temperature of 60°C for 72 hours.After this period, the material was weighed to determine the dry weight of the aerial part (DWAP).
The data were submitted to regression analysis, assessing the significance of the betas and the coefficients of determination using the statistical program SigmaPlot 11.0 software (SIGMAPLOT, 2008).

RESULTS AND DISCUSSION
All genotypes showed quadratic response depending to the increase in the volume of growing vases.The determination coefficients were significant (p <0.05) in all genotypes.The same occurred in both regression coefficients (β -1 and β -2 ).The increase in the vases caused a significant effect on the root length (RL) of cultivated grasses (Figure 2).

350-356p.
By observing the maximum response point in RL of the four genotypes, it appears that there are differences in vase volume ideal for the full root development.Grasses B. humidicola, Piatã and Massai had similar maximum points (0.258; 0.262 and 0.266 dm 3 , respectively), while Mombaça grass was more demanding in volume vase, reaching maximum response point RL with vases of 0.292 dm 3 .
The four grass species decreased in the RL when grown in vases 0.2 dm 3 .This behavior can have occurred by the fact that this volume vase has not accommodated a sufficient root portion to find the respiration rate of the root system.Low availability of oxygen to roots can be because small volume of substrate available for the roots, it undertook the process of gas exchange by root metabolism.With lack of oxygen, oxidative phosphorylation is blocked and metabolism starts work anaerobically, severely affecting plant growth (SAIRAM et al., 2008).
Similar reduction in RL occurred in the four genotypes grown in pots of 0.35 dm 3 and 0.4 dm 3 .However, when subjected to higher volumes vase, the root system shows growth and space exploration laterally, less inducement to deepen roots, resulting in delay of contact with the nutrient solution.Since most of the absorbent root hairs do not exploit the nutrient solution, with an expected frame nutritional failure and consequent decrease in plant growth.According to Barber (1995), the plants tend to uncontrolled emission of roots, as a mechanism to increase efficiency in the interception of nutrients.Prado et al. (2011), evaluating seedlings Panicum maximum cv.Tanzânia in nutrient solution, observed reductions of up to 87% in root length, due to weak absorption of macronutrients.
As for plant height (PH), all genotypes showed quadratic response due to the increase in the volume of growing vases.The determination coefficients were significant (p<0.05) in all genotypes.The same observed in both regression coefficients (Figure 3).It showed shorter variation in the volume of vases that provided maximum increase in PH (4.48% difference between the four genotypes) compared to the volumes observed for RL.The genotypes B. humidicola, Piatã, Massai and Mombaça have peak in PH with potted 0.290; 0.299; 0.303 and 0.301 dm 3 , respectively.
The results observed in PH are reflections of root development of genotypes.In the 0.2 dm³ vases, the shoot elongation was compromised by the low metabolic rate, as a result of hypoxia that the root system was subjected.Studies show that lack of oxygen in the soil is able to significantly reduce the photosynthetic capacity in non-tolerant grasses (CAETANO and DIAS-FILHO, 2008;SOUSA and SODEK, 2002).Ramos et al. (2011), by evaluating physiological and metabolic changes B. Brizantha plants subjected to conditions of root hypoxia, observed a significant reduction in photo synthetic rate, due to stomatal imbalances when compared to plants under normal conditions.
About the dry weight of the aerial part (DWAP), all genotypes showed quadratic response due to the increase in the volume of growing vases.The determination coefficients were significant (p<0.05) in all genotypes.The same observed in both regression coefficients (Figure 4).The grasses B. humidicola, Piatã, Massai and Mombaça show maximum increment of points similar DWAP (0.289; 0.302; 0.296 and 0.299 dm 3 , respectively), with change of 4.5% between them.
Overall, the genotypes grown in pots of 0.2 dm 3 compromise the results presented at the three characteristics studied.This presents insufficient dimension to accommodate a substrate portion that allows the plant to perform the process of breathing and gas exchange medium by full root tissue, not supplying the breathing demand of the plant.Deficit in the biochemical phase of photosynthesis, providing energy for cell metabolism is precarious, and results in lower PH and DWAP, directly correlated to energy supply (TAIZ and ZEIGER, 2009).
When grown in vases 0.35 dm 3 and 0.4 dm 3 , genotypes found aggravating for its vegetative growth, a decrease in all attributes, indicating that these volumes are too big for semi-hydroponic cultivation.In that condition, the initial root portion is located exclusively in the substrate layer with reduced exposure of roots to nutrient solution, damaging essential nutrients absorption for metabolism.According to Tavares et al. (2012), in an experiment with genre grasses Cynodon and Digitaria, the layer of 0-10 cm responsible for about 70% of the mass and number of roots, responsible for 49% of absorption and accumulation of nutrients in shoot.
Despite the four species have presented different responses depending on the vase volume used in semi-hydroponic cultivation, it can say that volumes close to 0.3 dm³ promoted greater development for the studied attributes.However, it is important to alert that for each genotype, there is an ideal volume for better measurement of stressful effect on the plant, and it is up to the researcher to carry out prior determination of the ideal vase volume according to the genotype studied.In addition to satisfying reliability and experiment significance, the use of vases with smaller volumes can have real gains, with economy vases substrate and physical space, and temporal gains, with the agility to obtaining results.
For situations of stress for aluminum, where all treatments are equally exposed to stressful situation, the use of vases with ideal volume gives real answers of the effect of toxic aluminum in the metabolism of genotypes, excluding external factors that can distort the observations and macular selection tolerant individuals.

CONCLUSIONS
• The semi-hydroponic cultivation in aluminum increased nutrient solution is effective in differentiation of forage grasses genotypes in relation to aluminum tolerance.
• Vase volumes near 0.3 dm 3 promote greater development for root and aerial part attributes in forage grasses grown in nutrient solution with aluminum toxicity.

Figure 2 .
Figure 2. Root length of forage grasses genotypes depending on the volume of vases for semi-hydroponic cultivation in nutrient solution, 25 days after germination, Gurupi-TO 2015.

Figure 3 .
Figure 3. Height of plant genotypes of forage grasses depending on the volume of vases to semi-hydroponic cultivation in nutrient solution, 25 days after germination, Gurupi-TO 2015.

Figure 4 .
Figure 4. Dry weight of the aerial part of forage grasses genotypes depending on the volume of vases to semi-hydroponic cultivation in nutrient solution, 25 days after germination, Gurupi-TO 2015.