Wastewater and substrates on the growth of Anadenanthera colubrina L. seedlings

ABSTRACT The production of seedlings of forest species for reforestation of biomes is a practice that restores the conditions of the environment, but water and substrates that favor the growth of seedlings are required. However, in the semi-arid region, water is a limiting factor, requiring the use of alternative sources such as treated domestic sewage. In this context, the objective was to evaluate the growth of Anadenanthera colubrina L. seedlings produced in substrates (soil + bovine manure and soil + coconut fiber) fertilized with treated domestic sewage effluent diluted in public-supply water in proportions (100, 75, 50 and 25% of domestic effluent) and using water from the supply network as a control. The treatments were arranged in a completely randomized design, in a split-plot with three replicates. The treated domestic effluent (TDE) used in the experiment came from the domestic sewage treatment plant of the decanter-digester type of the Projeto de Assentamento Milagre Apodi/RN. The study was conducted in a seedling production nursery at UFERSA, with growth and vigor evaluations at 30, 60, 90 and 120 days after transplanting. The production of seedlings fertigated with domestic effluent caused differences in all growth variables, with improvement in phytomass production. Seedlings fertigated with a dilution of 50% TDE and 50% water supply showed better results when produced with substrate composed of soil and coconut fiber. The seedlings showed good quality, and those cultivated with 50% and 75% of wastewater showed better quality after 120 days of cultivation.


Conflict of interest:
The authors declare no conflict of interest related to the publication of this manuscript.

INTRODUCTION
Water scarcity imposes abiotic stresses, which are the most important factors in the limitation of the plant's production capacity, so the artificial supply of water, via irrigation, is an important instrument to reduce or mitigate the impacts of climate fluctuations on production (TORRES et al., 2019).
The main advantage of using treated domestic effluents in agriculture is the recovery of a resource of great importance for agriculture -water; the constituents of these effluents are products that can increase soil fertility for containing nutrients that are essential to plants, due to the organic matter added to it, with the consequent formation of humus (OLIVEIRA, 2012).
Gaspar, Bezerra and Mota (2020) observed that the use of treated effluents in domestic sewage treatment plants promoted better growth of citronella (Cymbopogon winterianu) seedlings when compared to the use of public-supply water and that the behavior of plants followed the same trend, making use of the nutrients available in the treated effluents to increase their growth rate.
The use of substrate composed of bovine manure and coconut fiber is of paramount importance for seedling development, as it provides conditions for the root system to develop until the moment the seedling is transplanted to the field.Anadenanthera colubrina is an arboreal legume that belongs to the Fabaceae family and has an economic importance, besides being widely used in conservation plantations (LORENZI, 1998).For belonging to the Fabaceae family, it has potential for biological nitrogen fixation and is a significant option to be introduced in forest restoration areas or agroforestry systems (SOUZA et al., 2012).Thus, the objective of this study was to evaluate the effect of treated domestic sewage effluent and substrates on the growth of A. colubrina seedlings.

MATERIAL AND METHODS
The experiment was carried out in a greenhouse of the Department of Agronomic and Forestry Sciences of the Federal Rural University of the Semi-Arid Region (UFERSA), in Mossoró, RN,Brazil (5º 11' S,37º 20' W and 18 m altitude), from January to June 2019.
Anadenanthera colubrina seeds were collected from vigorous trees of spontaneous occurrence within a radius of 50 m, in the municipalities of Apodi-RN and Governador Dix -Sept Rosado, beside the Apodi/Mossoró river.
The materials used to make up the two substrates were: soil, aged bovine manure and coconut fiber.The soil used was collected from the subsurface layer (20 -40 cm) of a Latossolo Vermelho-Amarelo (Oxisol) under Caatinga, located in the rural area of the municipality of Mossoró/RN.The bovine manure used was acquired in the rural area of the municipality of Governador Dix-Sept Rosado/RN.The coconut fiber used was a commercial type, which contains as basic raw material the residue from Cocos nucifera industrialization, in which the shell is milled.
After preparing the growing substrates used in the experiment, samples were collected for physical-chemical characterization at the Soil Fertility Laboratory (LASAP) of UFERSA (Table 1).
The public-supply water used in the experiment to irrigate the seedlings came from the hydraulic network of the UFERSA campus.On the other hand, the domestic sewage effluent came from the Domestic Sewage Treatment Plant of the Projeto de Assentamento Milagre Apodi (RN), composed of 20 residences and approximately 90 people, producing a daily volume of 20 m 3 of domestic wastewater.To estimate the amount of nutrients supplied to the plants through the depths of domestic sewage (DE) and public-supply water (PSW), samples were collected for physical-chemical characterization (Table 2).
The seedlings were manually irrigated, twice a day, using a graduated cup to measure the amount of water applied, and the volume used was sufficient to maintain the maximum water capacity of the substrate in the pot, with a volume of 3 L.The maximum water capacity in the substrate inside the growing container was determined twice a week by means of tensiometry and water retention curve for each treatment.Irrigation was performed by gradually adding water to the substrate and collecting the drained water.
The evaluations of growth and development of A. colubrina seedlings were carried out at 30, 60, 90 and 120 days after transplantation (DAT), determining the following variables: a) Plant height (PH, cm) -obtained by the distance from the stem to the insertion of the last leaf in development, using a graduated ruler; b) Stem diameter (SD, mm)measured at 2.0 cm from the collar, with a digital caliper; c) Root length (RL, mm) -obtained by measuring from the plant collar to the tip of the root, with a graduated ruler; d) Number of leaves -obtained by simply counting the fully expanded leaves; e) Plant fresh matter: determined by individual weighing of roots, stem and leaves of the plants (g plant -1 ); f) Plant dry matter -determined by individual weighing of roots, stem and leaves of the plants after drying in an oven with forced air circulation at 70 °C until reaching constant mass (g plant -1 ); g) Dickson Quality Index (DQI) -determined as a function of plant height (PH), stem diameter (SD), shoot dry matter (SDM) and root dry matter (RDM), according to Dickson, Leaf and Hosner (1960).
The data were subjected to analysis of variance by the F test at 5% significance level, and the means were compared by Tukey test at 5% significance level, using the statistical program Sisvar® (FERREIRA, 2011).

RESULTS AND DISCUSSION
According to the results presented in Table 3, for plant height, A. colubrina seedlings showed significant difference from each other when produced in different substrates (soil + manure; soil + coconut fiber) when irrigated with the five different concentrations of wastewater.However, when comparing the five concentrations considering the Soil + Manure substrate they showed no significant difference from each other, and the means ranged from 12.34 cm for the concentration of 25% PSW + 75% TDE to 7.0 cm for the concentration of 75% PSW + 25% TDE.In the production of seedlings using the Soil + Coconut Fiber substrate, plant height showed significant differences when comparing the concentrations of irrigation water; the combinations 75% PSW + 25% TDE and 50% PSW + 50% TDE differ from the others statistically with values of 20.94 cm and 26.83 cm, respectively.
In a study evaluating growth parameters and dry matter accumulation of Calophyllum brasiliense seedlings, Artur et al. (2007) recorded a decrease in height when bovine manure was added to the substrate.Different results were obtained by Silva et al. ( 2013), who recorded growth of Ormosia arborea after the addition of bovine manure to the substrate.A. colubrina seedlings probably prefer more porous substrates that allow better aeration to the roots, which favors their development.
For stem diameter, the means differed for the two substrates and the different water concentrations, with minimum of 1.32 cm (25% of wastewater) obtained with manure and maximum of 3.10 cm (50% of wastewater) obtained with coconut fiber.According to Pereira et al. ( 2017), shoot height combined with stem diameter, as well as dry biomass, are indicators of quality of seedlings of forest species.
Brito et al. ( 2017), in a study conducted with domestic effluent application and substrates on Tabebuia aurea seedlings, concluded that the application of domestic sewage in seedlings promoted a positive response in stem diameter.In the study, the lowest means were observed for stem diameter with irrigation using 25% PSW and 75% TDE, while the lowest means were observed when the seedlings were irrigated with 100% PSW.
These results corroborate those found in the present study, where the highest means were obtained using 25% PSW + 75% TDE and 50% PSW + 50% TDE for the Soil + Manure and Soil + Coconut Fiber substrates, respectively.The waters with the lowest concentration of domestic effluent showed intermediate behavior for this characteristic, which can be attributed to high levels of N, P and K in the composition of the water with higher concentration of domestic effluent (Table 2).
In a study applying seven doses of treated domestic effluent in 'sabiá' (Mimosa caesalpiniaefolia Benth) seedlings and different substrates, Rebouças et al. (2018) concluded that, regardless of the fertilization management employed, the effluent proved to be an important source of nutrients, since it promoted increments in stem diameter and plant height, and these parameters are directly related to the seedling quality index.These findings corroborate those of the present study.
Table 4 shows the means of Dickson quality index and shoot dry matter of A. colubrina seedlings, as a function of different wastewaters and substrates.
Regarding the DQI, seedlings irrigated with water at the concentrations 50% PSW + 50% TDE and 25% PSW+75% TDE in the Soil + Coconut Fiber substrate were the ones with the highest means.This index is an indicator of seedling quality, as it considers the sturdiness and balance of biomass distribution through a balanced formula, including the relationships of morphological parameters, such as total dry biomass, shoot dry biomass, root dry biomass, shoot height and stem diameter.According to the criterion of Hunt (1990), DQI lower than 0.20 indicates that the seedlings do not have good final quality to be transplanted to the field, and the higher the DQI, the higher the quality of the seedling.Thus, the seedlings produced with Soil + Bovine Manure substrate and irrigated with 25% PSW + 75% TDE are on the threshold of DQI for good quality seedlings.Seedlings produced with coconut fiber had higher DQI, differing significantly from those produced with manure, except when irrigation was performed with public-supply water.
In relation to shoot dry matter, seedlings produced with Soil + Coconut Fiber substrate and irrigated with 50% PSW+50% TDE differed from the others, with value of 2.56 g.This behavior indicates the preference of A. colubrina for the Soil + Coconut Fiber substrate when irrigated with publicsupply water and with the same proportions of treated domestic sewage; this combination probably allowed better balance of nutrients for the seedlings.
Seedlings produced in the Soil + Bovine Manure substrate and irrigated with 25% PSW + 75% TDE had higher shoot dry matter production, differing significantly from the others.Biomass production is one of the best characteristics to evaluate the quality of seedlings, despite being destructive, because it reflects the net photosynthesis of the plant, that is, it is possible to analyze the growth in height and diameter obtained by the forest species during its permanence in the nursery and whether or not it is able to go to the field (ALVES; FREIRE, 2017).
Chemical factors such as pH, EC, C/N ratio and high salt concentration in irrigation water may be determinant for plant development, as well as the preference of the species for a given growing environment.
The means of plant height showed no difference as a function of the application of the different wastewaters in the initial growth stage (30 and 60 DAS), while in the growth stage at 90 days after planting the combination 50% PSW + 50% TDE was superior, with 21.73 cm, and also at 120 days after planting stood out among the others with the highest height, 24.35 cm, under 25% PSW + 75% TDE (Table 5).The height considered as ideal for forest species to leave the nursery stage for field conditions is still a controversial and highly debated subject.Schorn and Formento (2003) recommend height between 15 and 20 cm as appropriate for the seedlings to leave the nursery and be ready for the definitive field.In this study, reduced height was observed in seedlings irrigated with 100% PSW at 120 days of cultivation (Table 5).However, some Cerrado species have shown difficulty to reach this height standard, even after one year in the nursery, as reported by Pilon and Durigan (2013).
Regarding stem diameter, when A. colubrina seedlings were evaluated at 30 DAP, there was a difference between the applied waters, and the highest mean was equal to 2.08 mm, under 50% PSW + 50% TDE.Stem diameter is an important characteristic, since the higher its value, the greater the vigor, sturdiness and resistance of the plant (GUIMARÃES et al., 2009).Similar results were observed by Bezerra et al. (2005), with the increase in sewage sludge doses, for the stem diameter of colored cotton (Gossypium hirsutum L. r. latifolium Hutch).
Regarding root length, there were no differences during all 90 DAP, and these results agree with Souza et al. (2012), who found no significant difference in root length for A. colubrina, 'sabiá', tamarind, 'jucá' and 'mulungú' when evaluating the production of seedlings of forest essences irrigated with public-supply water, reject brine and fish farming effluent.At 120 DAP, the seedlings irrigated with 25% PSW + 75% TDE showed higher mean and stood out from the others.
For the Dickson Quality Index, until 60 days, the waters led to the same behavior.At 90 days, seedlings irrigated with 50% PSW + 50% TDE had the highest means and the others had lower means.After 120 days, the waters 50% PSW + 50% TDE and 25% PSW + 75% TDE promoted the highest means, not differing from each other.
Significant correlations were found between DQI, plant height, stem diameter and days after emergence, indicating a satisfactory result because the evaluation of these variables is non-destructive, hence facilitating and enabling experimentation in forest nurseries.When studying the quality of Trema micrantha (L.) Blume seedlings, Fonseca et al. (2002) reported that DQI is highly correlated with all morphological parameters, corroborating the results in this study.DQI is considered one of the most complete indices for assessing the quality of forest seedlings, as it includes in its calculation the relationships between the morphological parameters height, diameter, shoot dry matter weight and root dry matter weight, in addition to total biomass (ABREU et al., 2015).
According to José, Davide and Oliveira ( 2005), the higher the DQI, the higher the quality of the seedling, within that lot.DQI represents a good piece of information regarding the quality of seedlings, since it considers in its calculation the sturdiness and also the balance in biomass distribution in the seedlings (VIEIRA; WEBER, 2015).
For plant height, seedlings produced with the two types of substrate showed no differences when comparing the means at 30 days (Table 6).However, after this period the seedlings produced with Soil + Coconut Fiber obtained higher means than those produced with Soil + Bovine Manure.These results obtained in Table 6 are not in agreement with those reported by Kratka and Correia (2015), who evaluated the growth of Myracrodruonur undeuva Alemão in substrates prepared with sewage sludge, organic compost and bovine manure, and found that substrates with bovine manure showed excellent results.This is due to the supply of nutrients provided by this component for the seedlings, as well as the improvements of soil fertility and aeration, in addition to the water supply (CARVALHO et al., 2003).
In a study with production of A. colubrina seedlings using substrate of Latossolo Distrófico (Oxisol) at different base saturations, Bernardino et al. (2005) observed height ranging from 19.80 to 47.55 cm.The means obtained with Soil + Coconut Fiber were within this range.
The results of plant height in this experiment, regardless of the type of substrate, are consistent with those reported by Chaves, Carneiro and Barroso (2006), who evaluated the growth of Anadenanthera macrocarpa seedlings produced in fertilized substrate, consisting of agro-industrial residues, and found means ranging from 4.27 to 29.19 cm.
For stem diameter, there was no difference between the substrates only until 60 days.However, in the other evaluation periods, the substrate Soil + Coconut Fiber promoted stem diameter higher than that obtained with Soil + Bovine Manure.The diameter of native seedlings is of great importance because it dictates the survival of plants in the definitive field.According to Carneiro (1995), seedlings should have a larger stem diameter aiming at a better balance of shoot growth.
For root length, the substrate composed of Soil + Coconut Fiber led to results that were higher and different from those obtained with Soil + Bovine Manure during all evaluation phases.Substrate porosity is very important for root development, and the substrate with coconut fiber was the one that most favored root length.
In addition, these materials increase organic matter content and water storage capacity, resulting in greater availability to plants (SOUTO et al., 2005), as well as substrate aeration, promoting the dynamic balance of the water-soil-plant-atmosphere system (CUNHA et al., 2012).
For DQI, the two substrates did not show significant differences at 30 and 60 days.After this period, the seedlings produced with the Soil + Coconut Fiber substrate showed higher means at 90 and 120 days.
Considering the criterion of Hunt (1990), DQI lower than 0.20 indicates that the seedlings are not with good final quality to be transplanted to the field, and the higher the DQI, the higher the quality of the seedling.In this case, seedlings produced with Soil + Coconut Fiber substrate can be considered of good quality.

CONCLUSIONS
A. colubrina seedlings fertigated with nutrient solution containing 50% sewage effluent and 50% public-supply water showed better results when produced in substrate composed of soil and coconut fiber.The substrate composed of soil and coconut fiber promotes greater growth of A. colubrina seedlings.The seedlings showed good quality; however, plants that were cultivated with 50% and 75% of wastewater showed higher quality at 120 days of cultivation.

Table 1 .
Physical-chemical attributes of the growing substrates.

Table 3 .
Means of plant height and stem diameter of A. colubrina, as a function of different wastewaters and types of substrate at 120 days after transplanting.
Means followed by the same lowercase letter in the rows and uppercase letter in the columns do not differ from each other, at 5% significance level by Tukey test.

Table 4 .
Means of Dickson quality index and shoot dry matter of A. colubrina seedlings, as a function of different wastewaters and types of substrate, at 120 days after transplanting.

Table 5 .
Means of plant height (PH), stem diameter (SD), root length (RL) and Dickson Quality Index (DQI) of A. colubrina plants, irrigated with wastewater levels according to the evaluation times.

Table 6 .
Means of plant height (PH), stem diameter (SD), root length (RL) and Dickson Quality Index (DQI) of A. colubrina plants, evaluated in different types of substrate in four evaluation times.Means followed by the same uppercase letter in the columns do not differ from each other, at 5% significance level, by Tukey test.