GROWTH AND NUTRITION OF Paratecoma peroba SEEDLINGS IN DIFFERENT TUBE VOLUMES AND CONTROLLED-RELEASE FERTILIZERS

Paratecoma peroba is a native forest species of great economic and ecological interest, currently at risk of extinction from selective and illegal exploitation; however, there are no established parameters for the production of seedlings of this species. The objective of this study was to analyze the growth and nutritional balance of P. peroba seedlings grown in polypropylene tubes of different sizes with variable doses of controlled-release fertilizer (CRF). Three volumes of tubes (120, 180, and 280 cm3), four CRF doses (0, 2.5, 5, and 7.5 kg m-3), and a conventional mineral fertilizer (CF) were tested as indicated in the literature for native forest species: arranged in a completely randomized design in a 3 × 5 factorial scheme, with four replicates composed of 16 seedlings. After 180 days, the percentage of survival, growth variables, indices of the diagnosis and recommendation integrated system, and the average nutrient balance index of the seedlings were analyzed. Based on the total dry matter and the mean nutritional balance index, the combination of a 280 cm3 tube with a 7.5 kg m-3 CRF dose was the most effective in the production of P. peroba seedlings.


INTRODUCTION
Paratecoma peroba (Record and Mell) Kuhlm, popularly known as peroba-amarela, peroba-docampo e ipê-peroba is a tree species belonging to the family Bignoniaceae. These large trees can reach up to 40 m in height with a diameter of 40 to 80 centimeters (LORENZI, 2009). P. peroba is endemic to the Atlantic Forest and naturally occurs within the limited area south of Bahia, east of Minas Gerais, Espírito Santo, and north of Rio de Janeiro. Currently endangered due to intense exploitation, the species has great ecological and economic importance because of its natural beauty and wood quality. There are an estimated 8,500 adult individuals of P. peroba in nature (CNCFLORA, 2012).
In addition to the difficulty in finding matrices, the species has irregular seed production, with years of intense production followed by years of scarcity (LINS; NASCIMENTO, 2010). The seeds are recalcitrant, and the plants grow slowly in the nursery phase, taking around 6 to 8 months for seedling production (LORENZI, 2009). Therefore, there is a need to develop alternative and/or improved P. peroba propagation techniques to increase the availability of seedlings for species conservation projects.
Polypropylene tubes have advantages over plastic bags, such as the possibility of reuse, better formation of the seedling root system, lower substrate consumption, higher number of seedlings per area, and better handling, transport, and planting of seedlings (GONÇALVES et al., 2005). Despite these advantages, using the incorrect tube volume can result in deformation of the root system due to space limitations, with possible consequences on its performance in the field (JUNIOR et al., 2013).
Fertilizer application is required when using commercial substrates, which can result in increased production costs. Controlled-release fertilizers (CRFs) are polymer-coated fertilizers that release active nutrients in a controlled manner, extend the duration of nutrient release, and meets he metabolic needs of the plant (IRFAN et al., 2018). The use of CRFs in nurseries provides several benefits, such as reduced production costs, culture cycles, and labor.
Some techniques can illuminate the relationship between nutrients applied through fertilization and their assimilation by plants. Chemical analysis of the leaves, when well-interpreted, can be a useful tool for correcting nutritional deficiencies and imbalances in plants. In this sense, the Diagnosis and Recommendation Integrated System (DRIS) is used to interpret foliar chemical analysis. In short, the DRIS uses multiple relationships between nutrients and integrates them in a series of indices, thus taking into account the relationships and interactions between nutrients instead of the absolute and isolated concentration of each. The lower the negative DRIS index, the more nutrient-deficient the plant will be, and the higher the positive DRIS index, the more nutrient-rich the plant will be (WALWORTH et al.,1986).
Thus, the aim of this study was to analyze the growth and nutrition of P. peroba seedlings produced in different tube volumes and CRF doses. Therefore, we tested the following hypotheses: (i) the volume of the tube influences the growth of P. peroba seedlings; (ii) higher CRF doses enhance the growth and nutritional quality of seedlings of this species.

MATERIAL AND METHODS
The experiment was conducted in a university forest nursery located in the experimental area of the Department of Forest and Wood Sciences belonging to the Federal University of Espírito Santo (DCFM-CCA-UFES) in Jerônimo Monteiro, south of the state of Espírito Santo, from July 2017 to November 2017. The experimental area has a latitude of 20°47′S, a longitude of 41°23′W, and an altitude of 120 m. The region's climate is classified as Cwa (dry winter and rainy summer) by Köppen (1948).
The seeds were obtained from fruits selected from 20 matrix P. peroba trees, harvested in Reserva Natural da Vale S.A., in the municipality of Linhares, in ES, at 19°23′S and 40°04′W with an altitude of 43 m. The project was registered in SisGen for access to the genetic heritage (A796E13). After beneficiation, the largest and most vigorous seeds were selected. No pre-germination treatments were used on the seeds.
Conventional fertilization, proposed by Gonçalves et al. (2005), comprises soluble and nonencapsulated mineral nutrients to produce seedlings of native forest species. Initially, basic fertilization was carried out by adding nitrogen fertilizer (150 g m -3 N using ammonium sulfate), phosphorus fertilizer (300 g m -3 P2O5 using triple superphosphate), potassium fertilizer (100 g m -3 K2O using potassium chloride), and micronutrients in the form of 150 g of FTE (Fritted Trace Elements). After 50 days of sowing, cover fertilization was started with a nutrient solution applied to the substrate closest to the stem. For this solution, 200 g N (ammonium sulfate) and 200 g K20 (potassium chloride) were diluted in 100 L of water. Fertilizations were performed weekly. Therefore, the recommended monthly amount was divided into four parts. Table 1 shows that the chemical-physical characterization of the CRF Basacote® Mini 6 M formulation is 13-6-16, with a release time of 5 to 6 months.   Different doses of fertilizers were added to the commercial substrate (composed of Pinus bark, vermiculite, agro-industrial organic waste (Class A), wood sawdust, horse bed manure, and NPK, at 55% humidity) before filling the tubes, ensuring that each treatment received equal amounts of substrate. Samples of the substrate were collected for chemical characterization before implementing the experiment (Table 2), following the methodology of Silva (2009).
Sowing was carried out directly in the tubes filled with substrate and their respective doses of fertilizer. Each tube received two seeds, and was covered with a thin layer of the substrate. The tubes were placed in a polypropylene tray on raised beds under a screen with a 50% reduction in luminous intensity. Irrigation was carried out four times a day, for 7 min, through an automatic irrigation system with microsprinklers with flow rates from 31 to 152 lph. After 100 days of sowing, all seedlings were transferred to another area under full sun, receiving daily irrigation four times a day for 10 min.
After 180 days of sowing, the following variables were determined: survival (S, %); the height of the aerial part (H, cm), measured from the seedling neck to the insertion of the last leaf with the aid of a graduated ruler; diameter at root collar (DRC, mm), measured at the level of the tube edge using a digital caliper; number of leaves (NL), determined by counting the number of visible leaves; leaf chlorophyll content (LCC, µg cm -2 ), carried out on samples of intermediate seedling leaves through a portable chlorophyll meter SPAD-502; and the root volume (RV, cm³), obtained using a 500 mL beaker.
To determine the dry matter of the aerial part (DMAP, g plant -1 ), dry matter of the leaves (DML, g plant -1 ), and root dry matter (RDM, g plant -1 ), four seedlings per repetition were collected randomly, and each was divided into two parts: the aerial part and the root. The root system was separated from the soil by washing it with running water. Next, the material was packed in Kraft paper bags and placed in an oven with forced air circulation at 65 °C until constant weight, at which point it was weighed on an analytical balance (with a precision of 0.0001 g). The total dry matter (TDM, g plant -1 ) was the sum of the DMAP and the RDM. After obtaining these values, the Dickson Quality Index (DQI) was determined.
The DML was sent to the laboratory to determine the leaf content of the nutrients N, P, K, Ca, Mg, S, B, Zn, Mn, Fe, and Cu, according to Silva (2009). The Diagnosis and Recommendation Integrated System (DRIS) indices and the average nutritional balance indices (NBIm) were also calculated with the aid of the INAF program (Interpretation of Leaf Analysis) (GARCIA, 2013). To calculate the functions of the nutrient ratios, the method proposed by Beaufils (1973) with a k factor of 10 was used. Through the DRIS indices, it was possible to calculate the average nutritional balance index (NBIm), which is the sum of the DRIS indices in the module divided by the number of nutrients involved. The closer the value is to zero, the lower the nutritional imbalance of the plants and, potentially, the higher the production (MALAVOLTA et al., 1997;ARAÚJO et al., 2016).
For the variable S (%), the data were subjected to variance analysis, and when checking for significant differences, using the F-test at 5%, the doses were subjected to regression analysis to verify the optimal dose of the CRF through the first derivative estimators β0 and β1. The generalized mortality of seedlings grown at a CRF dose of 0 kg m -3 (control) made it impossible to continue the growth and nutrition assessments of this treatment. For the growth variables, principal component analysis (PCA) was applied to avoid the use of redundant and identical information contained in two or more correlated variables. The most important variable from the data set extracted through multivariate analysis was subjected to univariate analysis of variance. When checking for significant differences, using the 5% F-test, the treatments were compared by Tukey's test at the 5% and 1% probability. The analyses were performed using R Core Team (2018) version 3.5.0. Regression curves between seedling growth and NBIm were also fitted.
Tabela 4. Teor de nutrientes nas folhas (TN), índice DRIS (ID) e índice de balanço nutricional médio (IBNm) geradas a partir das normas DRIS para mudas de Paratecoma peroba em função do volume do tubete e da adubação com fertilizante de liberação controlada e adubação convencional. The results of the regression analysis between NBIm and TDM. which was the variable that best described the growth of P. peroba seedlings. are shown in Figure 4.  production of Paratecoma peroba seedlings.

DISCUSSION
Fertilization with CRF. among the factors tested. was crucial for the survival of seedlings of P. peroba. which were nutritionally demanding in the nursery phase. The high mortality rate of seedlings in the control treatment may be related to the ecophysiological characteristics of the species. Late secondary species show slower growth and a longer nursery time. requiring continuous fertilization during this period. Alternatively. the 80-day period with full sun exposure may have contributed to the increase in mortality in the poorly developed and poorly nourished seedlings from the control treatment.
Through analysis of the main components ( Figure 1; Table 3). it was possible to identify that TDM acts directly with the variables H. DRC. RV. RDM. DMAP. and DQI. This is because they act with the same signal as CP1; that is. when the value of one increase. the value of the other increases (multicollinearity). TDM was the variable that presented the greatest discriminatory power in CP1. and therefore. was the most important variable to describe and select P. peroba seedlings with superior growth and/or morphological quality.
The growth and quality of seedlings can be better described using the dry mass production variable of the root. the aerial part. and the combined total (FERNANDES et al. 2019). According to Larcher (2006). the difference between carbon gains via photosynthesis and losses via respiration (CO2 balance) results in vegetative growth; thus. the carbon not consumed in the respiratory process increases the dry mass of the plant. Therefore. it is believed that the higher the value of MST. the better the quality of the seedlings. and the better the survival and growth of plants after transplantation in the field.
In general. the cultivation of P. peroba seedlings were possible regardless of the size of tube used. However. among the treatments tested. the CRF combination at the dose of 7.5 kg m -3 with the 280 cm 3 tube proved to be the most effective for production of seedlings with maximum morphological quality (Figure 2; 3). The higher production of TDM in these treatments may be attributed to the synergistic action of improving the nutritional status of the plant and using larger tubes. which may include a greater absolute amount of substrate and fertilizer. as well as a greater available space for growth and nutrient exploration of the roots.
Previous literature suggests that forest species. independent of the ecological group. are responsive to the addition of CRF to the substrate. Navroski et al. (2016) demonstrated that Cedrela fissilis seedlings (late secondary) showed good growth when 5 kg m -3 of CRF was incorporated into the substrate. In seedlings of Peltophorum dubium (pioneer). Dutra et al. (2016) found growth improvement when they used CRF at doses between 5.4 and 8.2 kg m -3 . In addition. Rossa et al. (2015) found that CRF led to growth gains in seedlings of Anadenanthera peregrina (initial secondary) at doses of 5.54 to 6.68 kgm -3 and Schinus terebinthifolius (pioneer) at doses of 8.38 to 14.42 kgm -3 . Moraes Neto et al. (2003) experimented with 5 native species. such as Myroxylon peruiferum (climax). and found that doses between 3.2 and 4.8 kg m -3 of CRF led to the formation of quality seedlings in all 5 species. Research on the timing of seedling production indicates that the pioneer species stabilized in growth earlier (90 days) than the non- FLORESTA, Curitiba, PR, v. 50, n. 3, p. 1613-1622 pioneer species (150 to 180 days). Therefore. studies are needed to establish optimal CRF doses and formulations with adequate release times for each cultivated species.
In Calophyllum brasiliense and Toona ciliata. Lisboa et al. (2012) analyzed three tube volumes (115. 180. and 280 cm 3 ) and concluded that the most suitable tubes for C. brasiliense and T. ciliata were 180 cm 3 and 280 cm 3 . respectively. Gasparin et al. (2014) found that the use of the 280 cm 3 tube provided superior growth of Cabralea canjerana seedlings in the field phase. Melo et al. (2018) analyzed the influence of tube volume (30. 55. 110. 180. and 280 cm³) on the seedling growth of Mimosa caesalpiniifolia (pioneer) in the nursery and in the field. They found that. although the seedlings produced in the three larger volume containers showed greater growth in the nursery phase. this difference tended to disappear after implantation in the field. making it possible to produce seedlings of this species in any of the tested containers.
Thus. while larger tubes may result in larger seedlings. they require more space. substrate. and investment. By contrast. small containers are easy to handle and require less space and substrate; however. they can limit the growth of seedlings with a longer production cycle. nullifying and/or limiting the effect of fertilizers and. consequently. the nutritional balance of the seedlings. Brachtvogel and Malavasi (2010) found that the NL. DMAP. and TDM of Peltophorum dubium seedlings were similar under fertilization with NPK. regardless of the volume of the container. which did not occur in those under CRF. in which the growth was lower with the decrease in container volume. Gasparin et al. (2015) analyzed the combined effect of CRF doses (0. 3. 6. 9. and 12 kg m -3 ) on the growth and nutrition of Parapiptadenia rigida seedlings in the nursery with different tube volumes (50. 110. and 180 cm 3 ). They found that as the seedlings were exposed to higher doses of CRF. a higher N content was observed in the DMAP. However. no positive effect on seedling growth was observed on doses above 9.0 kgm -3 . Concerning the container volume. the seedlings grown in 180 cm 3 containers showed P. K. Mg. and S.
Conventional fertilization by Gonçalves et al. (2005) of native nursery seedlings was not effective for P. peroba seedlings when compared to CRF fertilization. possibly due to the absence of Mg. This can be seen in Table 4. where the Mg values and their respective DRIS indices were excessively low. It is also important to note that the Zn contents presented very high DRIS index values. The DRIS Ca and B indexes were excessively low in the treatments with 120 cm 3 tubes and CRF treatments at a dose of 2.5 kg m -3 . The levels of N in the DML of seedlings treated with conventional fertilization were slightly higher than those of seedlings cultivated with CRF. which was not reflected in gains in MST production.
Analysis of the nutritional status of P. peroba seedlings using NBIm revealed that the production of TDM was significantly associated with the nutritional status of the plants (R 2 =0.66; p<0.01). These results are compatible with the theoretical model suggested by Beaufils (1973). in which low values of NBIm. indicative of plants with balanced nutrition. mostly corresponded to samples with high production. confirming the importance of nutritional balance for the growth of P. peroba seedlings.

CONCLUSION
 The gains in growth and nutritional balance of Paratecoma peroba seedlings grown in higher CRF doses are limited when using tubes with volumes of 120 and 180 cm 3 .  Paratecoma peroba seedlings produced in a 280 cm 3 tube with a 7.5 kg m -3 dose of CRF can be considered suitable for planting in the field.