PRODUCTION TECHNIQUES OF ARBOREAL SPECIES SEEDLINGS AND USE OF MULCHING IN AN ALTERED AREA

Among the methods used for the restoration of altered areas, the planting of native tree seedlings is one of the most traditional ones. The objective of this study was to evaluate the survival and initial growth of Casearia sylvestris Sw, Handroanthus heptaphyllus Vell. Mattos and Parapiptadenia rigida Benth. Brenan seedlings in altered areas, according to containers used in nursery seedlings production and use of mulching on planting, using morphological and physiological attributes. The experiment was conducted in a 2x2 factorial scheme. The type of containers used (plastic bag and tube) was considered, and the presence or absence of mulching around the seedlings. The three species presented similar survival averages at 24 months. The rates between the treatments were 72.5; 85.0 and 80.0%, respectively. The use of plastic bags had a positive impact on the height (IncH) and stem collar (IncDC), crown area (CA) and shoot dry mass (SDM) for C. sylvestris and H. heptaphyllus, while for P. rigida, the use of plastic bags favored the CA and the SDM. The presence of mulching favored the attributes IncH, IncDC, AC, and MSPA of H. heptaphyllus, as well as CA and SDM of C. sylvestris. Regarding the physiological parameters (relative levels of chlorophyll a and b and fl uorescence of chlorophyll a), no diff erence was observed between the treatments tested. It is recommended that for the planting of seedlings of C. sylvestris, H. heptaphyllus, and P. rigida, in altered areas by anthropization, they be produced in a 1.5 L plastic bag, aiming at greater fi eld growth. Also, in the planting of C. sylvestris and H. heptaphyllus the mulching should be used to favor their growth.


1.INTRODUCTION
The intense demand for natural resources, together with the expansion of agriculture, livestock and urbanization, led to the deforestation of large forest areas in Brazil, resulting in the reduction of biodiversity and natural resources (Ferraz and Engel, 2011). There has been an increasing awareness of environmental conservation and sustainable management on a global scale. Nevertheless, many sites are still altered or degraded, requiring intervention, for restoring their environmental functionality.
In the last decade, the reforestation of altered areas by anthropogenic action has attracted the attention of public policies turned to the restoration of ecosystem services, aiming to recover the physiognomy of vegetation and its biological function (Paula et al., 2016). There are many methods for recovering altered areas, and choosing the most appropriate depends on the level of alteration of the environment, natural regeneration capacity, available fi nancial resources, and other factors (Chazdon, 2008). Among the methods used for covering or recovering altered areas, the planting of native tree seedlings is one of the most traditional ones.
When seedlings with larger dimensions are chosen, in order to favor their survival and growth in the fi eld, they are often produced in plastic bags. Nevertheless, it should be noted that the use of this container may have disadvantages, such as the demand for a high amount of substrate, long production cycles and the possibility of folding of the root system (Davide et al., 2015).
On the other hand, polypropylene tubes can be used, since they have internal splines in order to lead the root system, being able to improve the quality of the seedlings, besides facilitating the productive process (Gonçalves et al., 2005). However, they can restrict the growth of the plants because of their reduced size, thus causing deformations in the roots.
It is noteworthy that, although using seedlings produced with adequate inputs are used, the success of planting is also related to the management techniques adopted after this stage (Lima Filho et al., 2019;Brancalion et al., 2019), considering that the establishment of seedlings in the fi eld is a critical phase for successful restoration (Campoe et al., 2014;Brancalion and Holl, 2020). Among the cultural treatments that may help the survival of the seedlings is the use of mulching after planting, aiming at reducing the surface temperature and the incidence of weeds, as well as maintaining moisture and reducing soil erosion (Silva et al.,2018).
In addition, it is still important to use the appropriate species, according to each site and purpose of planting. The species C. sylvestris, H. heptaphyllus and P. rigida, widely found in the south region, are among those recommended for use to cover altered areas. Although many studies have been carried out in order to evaluate factors that aff ect the development and establishment of tree species (Abreu et al., 2015;Zavistanovicz et al., 2020), these studies are usually not performed in altered environments.
Thus, it is important to know the morphological and physiological attributes of tree species, in order to predict their potential for use in restoration projects. This study aimed to evaluate the survival and initial growth of C. sylvestris, H. heptaphyllus and P. rigida seedlings in an altered area, according to the kind of container used in the production of seedlings and the use of mulching in the planting, through morphological and physiological attributes. It is believed that the highest survival rates, as well as greater growth of the seedlings and more adequate physiological attributes are verifi ed in those produced in plastic bags, along with the use of mulching.

2.MATERIAL AND METHODS
The study was developed in the Parque Estadual Quarta Colônia (PEQC) (29º27'57,39''S and 53º16'51,30''O), a conservation unit of integral protection located in the state of Rio Grande do Sul, in the Atlantic Forest Biome. According to the Köppen climate classifi cation, the climate of the region is classifi ed as "Cfa", with well-distributed rains over the months (> 40 mm per month), with the average temperature of the coldest month between -3 and 18 ºC and the hottest month with temperatures higher than 22 ºC (Alvares et al., 2013).
The PEQC area used for the experiment presents a history of anthropic intervention. This area is composed of small rural properties and a housing complex. After the evacuation of the area, the houses were demolished, but the debris was left behind and the soil was visibly compacted. Prior to the experiment, a vegetation composed basically of Brachiaria spp. (Brachiaria spp.), Andropogon bicornis L. (Burdock) and Eryngium horridum Malme (caraguata) was identifi ed, in addition to some individuals of Pinus sp. and Eucalyptus sp.

Production of seedlings
The choice of tree species to compose the experiment was based on phytosociological studies carried out in the region. Those with rapid to moderate growth that could cover the area in the short term and which are commonly found in nurseries in the region were also chosen. Thus, three native species of the South region were selected: C. sylvestris, H. heptaphyllus and P. rigida.
The production of seedlings was carried out at the Forest Nursery of the Federal University of Santa Maria. Polypropylene conical tubes with 180 cm³ of volume were used in the production. The substrate used in the fi lling consisted of the mixture of a commercial substrate Carolina Soil® composed of Sphagnum peat (CS) and carbonated rice husk (CRH), in the ratio 4:1 (v:v), with base fertilization being carried out with controlled release fertilizer (Osmocote® 18-05-09 Mini Prill) at the dosage of 6 g L -1 .
After 170 days of sowing, 50% of the seedlings were transplanted to polyethylene plastic bags of 1,500 cm³ (18 cm height x 12 cm wide) fi lled with substrate composed of subsoil, CS and CRH (1:1:1). A diff erent substrate composition for seedlings transplanted into plastic bags was used in view of the characteristics of that container (Abreu et al., 2015), where subsoil is generally used (Gonçalves et al., 2005). Subsequently, the seedlings remained in the nursery for eight months until planting in the fi eld. In this period, top-dressings were applied to all the plants, with ammonium sulphate ((NH 4 ) 2 SO 4 ) and potassium chloride (KCl), in accordance with the methodology of Gonçalves et al. (2005). At the time the seedlings were moved from the nursery to the fi eld, they presented height (H) and stem diameter (SD) varying according to the container used (Table 1).

Treatments and experimental design
The experiment consisted of four treatments in a 2x2 factorial scheme, with the factor "A" being composed of two types of containers: seedlings produced in tubes and transplanted into polyethylene plastic bags, where they remained until planting and seedlings produced and kept in tubes until planting; and factor "B" composed of the presence and absence of mulching around seedlings in the planting. Each species was analyzed individually, according to the treatments, and the species was not a treatment factor.
The experiment was conducted in split plots, with the plot being composed of the container treatments used in the production, and in the subsplit plots the presence or absence of mulching, in a randomized block design. Each plot of 10.5 m x 10 m consisted of a single species, with 35 individuals, according to the treatments, and space of 1.5 m x 2.0 m between plants and 2 m between blocks, with a total of fi ve blocks.

Area preparation and planting
Prior to planting, soil samplings were collected to characterize the area, with three samplings per block at depths of 0 to 20 cm and 20 to 40 cm, using a cutting blade. The chemical analysis of the soil was performed by the Soil Analysis Laboratory of the Federal University of Santa Maria (Table 2). Through the physical analysis of the soil, carried out at the Laboratory of Soil Physics of the UFSM, the textural class (pipette method) was determined, classifying it as sandy-loam.
Mechanized mowing was performed in the total area, followed by plot marking according to the treatments. Subsequently, pits with approximately 0.008 m³ (20 cm x 20 cm x 20 cm) were opened with the use of a digger. After the area was prepared, seedlings were manually planted, in October 2013. It is noteworthy that, at the time of planting, both the seedlings from plastic bags and those from the tubes had a well-structured lump, with a large amount of roots added to the substrate, as well as thin and light roots.
After planting, the seedlings were irrigated, with about 2 L of water per plant. Then, the mulching was allocated around the seedlings that received this treatment. The mulching was obtained from dry grass (Brachiaria spp.), collected in the area adjacent to the experiment and allocated in a radius of approximately 20 cm and height of 5 cm, around the stem of the plants. Replacements with the same material were done, every four months, during the fi rst year of the experiment. Being: OM -organic matter; V -base saturation; P-Mehlich -phosphorus extracted by the Mehlich method -1; K -potassium; CEC pH 7,0 -cation exchange capacity; Ca -calcium; Mg -magnesium; Al -exchangeable aluminum and H+Al -potential acidity. Sendo: MO -matéria orgânica; V -saturação por bases; P-Mehlich -fósforo extraído pelo método Mehlich-1; K -potássio; CTC pH 7,0 -capacidade de troca de cátions; Ca -cálcio; Mg -magnésio; Al -alumínio trocável e H+Al -acidez potencial.  The replanting of the seedlings that did not survive was performed 60 days after the planting. In addition, the experiment was periodically monitored, with control of leafcutter ants through granulated baits (based on Fipronil and Sulfl uramide) and weed control, performed by manual weeding around the seedlings (with subsequent replenishment of the mulching when it was dispersed), mowing in the crop rows and use of selective post-emergent herbicide (Sethoxydim 120 g/L) between the rows. 1.2 L/ha were applied with the aid of a backpack sprayer. The top-dressing in the experimental area was performed at 01, 12 and 24 months after planting, using Polyblen® controlled release fertilizer (N-P-K 18-08-18 + S + B).
The meteorological data of precipitation (mm) and average temperature (ºC), corresponding to the period of the experiment, were obtained from the National Meteorological Institute (INMET), located at the UFSM Campus, Santa Maria -RS ( Figure 1).

Evaluations of morphological and physiological attributes
The survival of the seedlings was evaluated at 24 months after planting, through visual analysis. The height (H), measured with a graduated ruler, and the stem diameter (SD), measured with a digital caliper, were measured at planting time and at 24 months of age, when the plants crown diameter was also measured (CrownD), with two horizontal perpendicular measurements (CrownD1 and CrownD2), with the aid of a graduated ruler, for later calculation of the crown area (CA), where CA= [(CrownD1+CrownD2)/4] 2* π. For the morphological attributes described, the six central plants of each subplot were measured. The evaluation of the shoot dry mass (SDM), at 24 months of age, was determined by sectioning one plant per replicate (representative of the plot) in the collection region, then the plant material (leaves, branches and stem) was allocated in brown paper envelopes identifi ed and submitted to oven drying with forced air circulation at 65 ºC, until constant weight. Subsequently, the material was weighed in a digital scale to obtain SDM.
For the physiological analyses of the seedlings, the relative levels of chlorophyll a (Cla) and b (Clb) were verifi ed; and chlorophyll a fl uorescence at 24 months after planting, in one representative plant per replicate. The evaluation of the relative chlorophyll content was performed using a chlorophyll meter (Clorofi LOG-Falker), in expanded leaves of the upper third of the plant, measuring one leaf per plant (two readings per leaf) (Barbieri Junior et al., 2012). The determination of chlorophyll a fl uorescence was performed using a portable light-modulated fl uorometer (Junior-Pam Chlorophyll Fluorometer), in expanded leaves attached to the plant, on sunny days, between 08:00 and 10:30 in the morning. Leaves of the upper third of the plant were selected and wrapped in foil for 30 minutes to adapt to the dark. Then, fl uorescence measurements were taken, obtaining the values of initial fl uorescence (F o ), maximum fl uorescence (F m ) and maximum quantum yield of photosystem II (F v / F m ). The physiological attributes were not determined in seedlings of P. rigida due to the small size of their leafl ets, making the contact between the devices and the leaf impossible.

Statistical analysis
The data were checked for the normality assumptions of the error distribution and homogeneity of variance, respectively, by the Shapiro-Wilk and Bartlett tests. Subsequently, the data were submitted to analysis of variance (ANOVA) and, when a diff erence between the treatments was found by the F test, the means were compared using the Student's t-test (p<0.05). The statistical software SISVAR (Ferreira, 2014) was used for these analyses.

3.RESULTS
The statistical analysis indicated that there was no interaction between the factors studied (container and mulching) for any variables investigated, therefore, they were analyzed and are presented in this section individually. The survival of the seedlings did not show any infl uence of the treatments used (p> 0.05), for the three species (Table 3). There was a general survival mean of 72.5% for C. sylvestris seedlings, 85.0% for H. heptaphyllus and 80.0% for P. rigida. In relation to the increment in height and stem diameter (IncH and IncSD), there was a signifi cant diff erence between the levels of the container factor in C. sylvestris seedlings, which presented higher values of IncH and IncSD when a plastic bag was used (53.15 cm and 13.58 mm, respectively) ( Table 3).
For H. heptaphyllus, the statistical analysis indicated a diff erence in both isolated factors (container and mulching), and for the recipient, the seedlings from the plastic bag obtained higher IncH and IncSD (131.02 cm and 22.66 mm respectively) than those produced in polypropylene tubes (107.98 cm and 16.49 mm, respectively). In relation to the mulching factor, the seedlings presented a higher increase in height and stem diameter when there was presence of mulch in their surroundings (127.69 cm and 21.22 mm, respectively) ( Table 3). It was also verifi ed that, for the P. rigida seedlings, unlike the species previously mentioned, the containers tested did not show infl uence on the shoot increase. However, there was a signifi cant diff erence between the mulching factor levels for the IncH variable, and a larger increase (138.65 cm) was observed in seedlings with no mulch coverage at planting.
Seedlings of C. sylvestris and H. heptaphyllus showed similar behavior regarding crown area (CA) and shoot dry mass (SDM), with signifi cant diff erence between both factors tested, but without interaction. For both species, in relation to the recipient, the highest values of CA and SDM were found in seedlings transplanted to plastic bags, with averages of 0.219 m² of CA and 90.97 g of SDM in C. sylvestris and 0.935 m² of CA and 308.20 g of SDM in H. heptaphyllus (Table 4). In addition, there was a positive eff ect for the presence of mulch, with values higher than 0.197 m² of CA and 80.10 g of SDM in C. sylvestris and 0.786 m² of CA and 252.00 g of SDM in H. heptaphyllus. For P. rigida, there were only diff erences between the types of containers. The seedlings from plastic bags presented higher averages, both in CA (1,971 m²) and in SDM (581,10 g), than those produced and kept in tubes (1.214 m² CA and 305.80 g SDM) (Table 4).
Regarding the physiological attributes, there was no signifi cant diff erence between the treatments Regarding the planting area, it should be noted that the weeds present in the area showed intense development in the fi rst months after the planting of the seedlings.

4.DISCUSSION
Among the species studied, there was a lower survival rate for seedlings of C. sylvestris (72.5%). Although it is considered a pioneer (Lorenzi, 2008), its high mortality shows less resistance of this species to tolerate the limiting factors (such as weed competition, compacted soil with a low content of organic matter, among others) that occurred in the planting area. This fact was corroborated by the small increase of the seedlings (Table 3), making them more susceptible to competition with weeds, with widespread occurrence in the area. This suggests the need for greater care in the management when planting C. sylvestris in altered areas, such as planting this species in areas where the fi rst stages of vegetation can be found.
On the other hand, the survival rates of H. heptaphyllus (85%) and P. rigida (80%) showed greater adaptability to altered site conditions. The success of planting native trees for forest restoration depends on the knowledge about the behavior of the tree species used, in relation to survival and growth in the face of adverse conditions (Martinez-Garza et al., 2013). Schievenin et al. (2012) report that the survival rate of the seedlings allows to infer what maintenance is needed, as well as whether the species used and the cultural treatments are adequate for the conditions of the area. In the present study, survival analysis demonstrated the specifi city of each species to tolerate adversities at the site of planting. In this case, we highlight that the area of the experiment was strongly impacted by anthropic action.
According to Silva et al. (2016), plantings with survival above 80%, in the Atlantic Forest Biome, can be considered satisfactory and they also point out that mortality is an inherent characteristic of the place where planting place and their ability to adapt to fi eld conditions. This condition was also verifi ed by Morais Junior et. al (2020), who, when conducting a study with fi ve native Atlantic Forest species, produced in diff erent containers and, afterwards, planted in a degraded area, verifi ed diff erent behavior for each species. While some showed a higher survival rate when produced in larger containers, for others this factor was irrelevant.
Martinez- Garza et al. (2013), when evaluating the behavior of 24 tree species planted in a restoration area on abandoned pastures in Mexico, found that survival varied greatly between species (from 9 to 96%), with the highest rates observed for pioneers.
The results found in this study, as well as in those cited above, reinforce the need to carefully choose the species to be used, through the analysis of their behavior in the fi eld, choosing species and methods that present high survival, avoiding unnecessary expenses with replanting. In addition, it should be considered that the establishment period of the seedlings is a critical moment for the recovery, to be successful, and care with planting, such as irrigation and weed mowing, in the fi rst months must be intensifi ed, in order to favor survival (Campoe et al., 2014). This fact was evidenced in the present study, in which there was a need for constant monitoring during the fi rst year, considering that the weeds present in the area showed intense development in the fi rst months after the planting of the seedlings, which corresponded to a period with high temperatures and constant precipitation. In this phase, the seedlings were still being established, therefore, in addition to the silvicultural practices aiming to diminish the weed competition, a greater frequency in the control of leafcutter ants was also necessary.
Increases in height and stem diameter, CA and SDM of seedlings of C. sylvestris and H. heptaphyllus, in addition to the CA and SDM of P. rigida, were favored by the use of the plastic bags, possibly due to the higher volume of the substrate provided by such container. Field growth for seedlings of forest species produced in diff erent recipient volumes has already been verifi ed by other studies (Gasparin et al., 2017;Zavistanovicz et al., 2020), and in most of them, a larger volume is associated to the greater development of the seedlings in the fi eld, as evidenced by Morais Junior et al. (2020).
This occurs because when the seedlings are produced in containers of small dimensions, they require a series of actions to favor their growth, due to the smaller volume of substrate in the container, less amount of available nutrients, as well as, less accumulation of water (Davide et al., 2015), while for larger containers, such as plastic bags, the opposite is evident. In addition, smaller container volumes, such as 180 cm³ tubes, may limit the growth of the root system of the seedlings (Abreu et al., 2015).
Thus, by controlling the amount of roots that the plant can produce, the volume of the recipient also determines the height of the shoot part (Ritchie et al., 2010), since the plants tend to present a balanced growth between the root and shoot part (Abreu et al., 2015).
It is suggested that the volume of the container used infl uences less the behavior of the seedlings when they are planted at suitable sites (Pinto et al., 2011). However, when the planting area presents limiting factors to survival and growth, as is the case in most of the altered areas, the authors emphasize that the volume of the container will be decisive for success in planting.
According to the species, it may occur that the type and volume of the container used do not signifi cantly interfere with the growth in height and stem diameter of seedlings after planting, as verifi ed in this study for P. rigida. Although the study by Ferraz and Engel (2011), shows that seedlings of this species presented higher growth when produced in containers of larger volume, in the nursery phase, this research did not consider their development in the post-planting, and sowing occurred directly in all containers, a fact that may have infl uenced the fi nal result.
The behavior observed in the present study for P. rigida is characteristic of fast-growing species, generally included in the pioneer group and that present a greater rusticity and capacity to tolerate less favorable environments and environmental conditions (Nunes et al., 2015). Although in the nursery phase the container with higher volume favors the growth of the seedlings, there is a tendency for this growth diff erence to decrease gradually after planting in the fi eld, as verifi ed for seedlings of E. contortisiliquum (Abreu et al., 2015), which presented high growth rates, as well as P. rigida.
It should be noted, however, that the seedlings of P. rigida produced in plastic bags showed a better performance in CA and SDM, a fact that was possibly favored by the more developed root system, providing the expansion of the CA and, consequently, of the SDM. According to Ritchie et al. (2010), the growth of the seedlings in the shoot area is generally correlated with the number of leaves of the plant, being a good estimate of their photosynthetic capacity. The authors point out that, in planting sites with high vegetative competition, as in the present study, the ability to capture and process sunlight is decisive for plant survival and growth. Therefore, seedlings with more developed shoot areas have advantages over smaller seedlings that tend to be suppressed by competing vegetation (Ritchie et al., 2010).
In cases where the species has an adequate development when produced in diff erent container volumes, as occurred for P. rigida, the choice of which to use should consider the fi nal objective of planting (restoration, production, others) and the costs involved during production, considering that the use of larger containers, according to Ferraz and Engel (2011), implies an increase in implantation costs, because of the higher consumption of substrates, the need for more space in the nursery and the higher cost of transportation.
Besides the container infl uence observed, it was verifi ed that the use of mulching favored the growth of plants of C. sylvestris and H. heptaphyllus. The benefi cial eff ects of mulching on the planting of tree species were also evidenced by Dostalek et al. (2007), who verifi ed that the growth of the seedlings was up to three times higher compared to those with no cover. The same authors point out that the species present diff erent behaviors regarding the presence of mulching and that diff erent materials must be tested to cover the soil in order to determine the one that best meets the needs of the area and the species used.
Diff erent from the other species of this study, in seedlings of P. rigida the use of mulching proved to be unfavorable for the morphological attributes, mainly for IncH. This result may have been infl uenced by the pioneering characteristic of the species, which, even in view of the growth potential of the seedlings produced in a plastic bag, did not need to invest in aerial growth to overcome the weed competition, which in turn had its growth inhibited by mulching. Therefore, as with the container, the use of mulching should be considered for each species individually, considering the diff erent responses obtained in each situation and their relevance. In plantings in altered areas, especially in those whose soil is exposed, the use of mulching should be analyzed regarding not only the benefi t it provides to the growth of the seedlings, but also the capacity of physical protection that it off ers, becoming a possible method for avoiding erosion.
Other soil covering techniques have been used in forest restoration plantations, aiming mainly at the control of weeds, such as the use of 'cardboards' mulch. Silva et al. (2018) found that the use of cardboard not only helped in the control of weeds, but also reduced the temperature at the soil surface, allowing for adequate development of seedlings in the fi eld, because of the improvement in soil conditions.
Besides that, it is necessary to evaluate the availability of the materials to be used, opting for materials found close to the planting area, in order to make the process feasible economically.
In the present study, the physiological attributes were not altered by the treatments used for both C. sylvestris and H. heptaphyllus. This indicates that even the seedlings that obtained lower growth, were in similar physiological condition in the diff erent treatments. Among the evaluated physiological attributes, the relationship between variable fl uorescence and maximum fl uorescence (F v /F m ), a measure of the intrinsic or maximum effi ciency of photosystem II (PSII), stands out (Araújo and Deminicis, 2009).
The values of F v /F m for both species were below the recommended (0.75-0.85) by Araújo and Deminicis (2009) suggesting that the plants were exposed to stress conditions, mainly C. sylvestris, due to the reduced value of F v /F m (0.485).
This fact was probably due to the high weed competition present at the planting site, because when plants are exposed to restrictive environmental factors (weed competition, nutritional defi ciency, water stress), it is common to observe falls in the value of this physiological attribute, being indicative of stress (Araújo and Deminicis, 2009).

5.CONCLUSIONS
The survival of C. sylvestris, H. heptaphyllus and P. rigida was not infl uenced by the treatments applied.
For the rapid growth of C. sylvestris, H. heptaphyllus and P. rigida in the altered area, seedlings produced in larger containers, such as 1.5 L polyethylene plastic bags, should be used.
For C. sylvestris and H. heptaphyllus, the use of mulching favors growth after planting, and it is appropriate to use it whenever it is available in the planting area.