Spatial distribution , association patterns and richness of tree species in a seasonal forest from the Serra Geral formation , southern Brazil

Spatial distribution and association patterns of tree species in a seasonal forest from the Serra Geral formation, southern Brazil). Th is study aimed to determine the spatial distribution of the most abundant tree species, to look for relationships that may be responsible for their distribution and to compare the expected richness along the Serra Geral formation in the state of Rio Grande do Sul , Brazil. An one-hectare plot was divided in 100 contiguous 10 x 10 m sampling units and all individuals with at least 4.8 cm diameter at breast high were measured and identifi ed. Each sampling unit received a description of the spatial position through metric coordinates (x,y) in order to quantify the spatial pattern. Later, an association index among species was applied. Th e observed richness was rarefi ed and compared to other surveys located on the slopes of Serra Geral in Rio Grande do Sul. We surveyed 1,063 individuals belonging to 69 species and 33 families, and the Shannon’s diversity index showed a diversity of 3.35 nats (Pielou’s species evenness J’ = 0.79). Most species showed an aggregated distribution. Both association and dissociation patterns occurred among species leading to distinct forest patches. Th e rarefaction results showed a decrease of Atlantic species, which were replaced by typical seasonal forest species, when analyzing fl oristic contingents towards the west.


Introduction
In nature, organisms frequently show an aggregated distribution, forming patches, gradients or other types of spatial patterns (Legendre & Fortin 1989;Dale et al. 2002).Th e analysis of such patterns is a relevant statistical tool in the area of forest dynamics (Pélissier 1998;Franklin & Rey 2007), which helps in understanding species ecology and providing information for its management and conservation (Perry et al. 2002;Bell et al. 2010).In Brazil, studies about spatial distribution of plants were scarce in the last decade (Nascimento et al. 2002) but have started to increase in recent years (e.g.Anjos et al. 2004;Lindenmaier & Budke 2006;Giehl et al. 2007;Paludo et al. 2009;Higuchi et al. 2010;Rode et al. 2010).
Due to advances in computational techniques in the past few decades (Perry et al. 2002;Hooten et al. 2003), several models have been created for the analysis of spatial patterns.In this sense, Dale et al. (2002) and Perry et al. (2002) described concepts and provided a broad framework for making decisions on analytical approaches and interpretations of spatial patterns.Among these methods, the Spatial Analysis by Distance Indices or SADIE, proposed by Perry (1995), has been successfully used in recent studies (Ferguson et al. 2000;Winder et al. 2001;Xu & Madden 2004;Lindenmaier & Budke 2006;Giehl et al. 2007;Rechenmacher et al. 2007), and has been useful in describing and quantifying spatial patterns (Xu 2003).
Based on the spatial pattern provided by SADIE, Perry & Dixon (2002) created a new method of association analysis.By identifying the spatial heterogeneity as a patch or a gap, this method is more effi cient in detecting associations than a simple correlation co-effi cient (Donovan et al. 2007;Bell et al. 2010).Th rough the analysis of spatial pattern and association, Donovan et al. (2007) emphasized the importance of termites in a tropical forest in Malaysia.Bell et al. (2010) built co-occurrence networks based on statistical analysis of the SADIE method, providing valuable information on the area of biological control in the United Kingdom.Gabriel et al. (2007), using SADIE's method, verifi ed that the young individuals of Cupania vernalis Cambess.do not follow the distribution of adults in a forest remnant in southern Brazil.
Th e state of Rio Grande do Sul (southern Brazil) has, beyond areas of sandbanks and pioneer formations, humid and seasonal forests (Teixeira et al. 1986).Nowadays the best preserved areas are located on the slopes, along the Serra Geral and Serra do Sudeste formations, mainly in inaccessible areas (Jarenkow & Waechter 2001;Jurinitz & Jarenkow 2003).
Th e main entrance corridors of tropical species in this state occur in the east, through the Atlantic Corridor, and in the west, across the basins of the Paraná and Uruguay rivers (Rambo 1956;1961).Species interpenetration occurs on the hillside areas of Serra Geral (Jarenkow & Waechter 2001).Rambo (1956;1960) mentioned that as species move, from east to west, across the Serra Geral, there is a reduction trend in species richness, which is higher in Atlantic Forest stricto sensu and gradually becomes lower towards the seasonal forest.However, little has been discussed about this in recent years.Regardless of several structural studies about plant communities of the Serra Geral formation (Jarenkow 1994;Longhi et al. 1999;Longhi et al. 2000;Jarenkow & Waechter 2001;Lindenmaier & Budke 2006;Giehl et al. 2007;Sühs et al. 2010), the lack of standardized data, which occurs in most descriptive studies of terrestrial plant communities conducted in Brazil, makes it harder to compare their results (Sonego et al. 2007;Giehl & Budke 2011).
Th is study aimed to determine the spatial distribution of the most abundant tree species from a seasonal forest remnant in the central region of Rio Grande do Sul, Brazil, and to look for relationships that may be responsible for their distribution.Likewise, the horizontal structure, di-versity and changes in species richness along the slopes of the Serra Geral region were also evaluated.In this sense, we tried to answer the following questions: i) how is the spatial pattern of the most abundant species sampled?ii) is there an association between the spatial distribution of these species?iii) is there reduction in the species richness, from east to west, along the Serra Geral formation?

Study area
Th e study was conducted at Sinimbu, in the central region of Rio Grande do Sul State, Brazil, in a Private Reserve of Natural Heritage which belongs to the University of Santa Cruz do Sul.Th is reserve was offi cially created in 2009 (Brasil 2009) and has an area of 221.39 ha.Sühs et al. (2010) found 160 tree species in this area and observed that 12 are at risk of extinction.On the other hand, the same authors also found 11 exotic species and emphasized that these species must be controlled, especially the invasive ones.
According to Maluf (2000), the regional climate belongs to type STSBv, (subtropical, sub humid, with dry periods in summer).Th e average annual temperature is 19.3 ºC and average annual rainfall is 1,547 mm, recorded in Santa Cruz do Sul, the nearest station (data supplied by the Laboratory of Meteorology, University of Santa Cruz do Sul).Th e local soils belong to an association of Ciríaco and Charrua mapping units (Streck et al. 2002).

Data collection
A plot of 1 hectare (100 x 100 m) was set in the reserve domain and divided in 100 contiguous 10 x 10 m sampling units (central point near coordinates 29º23'22''S -52º32'00''W at 500 m.s.l.-map available in Sühs et al. (2010).Th e plot was set on a slope, because it had better forest, where we observed the presence of two non-rectilinear gullies along the sampling area.All individuals with diameter at breast height > 4.8 cm were measured and identifi ed.Voucher specimens were deposited in the HCB Herbarium at the University of Santa Cruz do Sul.Specimens were identifi ed through literature and by comparison with vouchers from the HCB Herbarium.Th e classifi cation of families of angiosperms was defi ned according to the Angiosperm Phylogeny Group (APGIII 2009) while Smith et al. (2006) was used to classify tree-like ferns.Th e nomenclature of species was based on Forzza et al. (2010).
Th e species were classifi ed according to their fl oristic contingents: western seasonal (basins of Paraná and Uruguay rivers), eastern wet (Atlantic contingent), wide geographical distribution, Araucaria forest and exotics (Jarenkow & Budke 2009, Jarenkow & Waechter 2001, Budke et al. 2004, Lindenmaier & Budke 2006, Sühs et al. 2010).We also grouped species by vertical distribution and in ecological groups of regeneration: "pioneer" for species that show an entirely heliophilous life cycle, "light-demanding" and "shade tolerant" for those who tolerate some degree of shade (Budke et al. 2008).Th e species were divided into three height groups: small (average height < 8 m); medium (average height ≥ 8 m < 12 m) and tall (average height ≥ 12 m).Estimated height values were given in Sühs et al. (2010).
Twelve samples of topsoil (0-20 cm depth) were randomly collected in the area and formed a single composite sample that was taken for chemical and texture analysis.Th e results were interpreted according to Comissão de Química e Fertilidade do Solo -RS/SC (2004).

Data analysis
For each species, the phytosociological parameters of density, frequency and dominance (derived from tree basal area) (Mueller-Dombois & Ellenberg 1974) were calculated to describe the tree community structure.A histogram was constructed (class interval determined statistically at 4.8 cm) to express the diameter classes of individuals.
In order to answer the fi rst question of our study, we applied the SADIE methodology to evaluate the spatial patterns of species (Perry 1995).Each sampling unit received a description of the spatial position through metric coordinates (x,y), mapped from an initial established point.Species showing more than 20 ind•ha -1 had their spatial distribution analyzed through the index of aggregation -I a (Perry 1995).Values of I a > 1 indicates aggregation (patches), I a < 1 indicates regularity and I a = 1 suggests randomness (Perry et al. 1996;Perry 1998).Negative values of aggregation indicates that a species-i does not occur in that position (gaps), providing further details on pattern analysis (Perry et al. 1999).
According to Perry (1998), the spatial pattern in a sampled population is quantifi ed by SADIE by measuring the total eff ort (in terms of distance moved), which the individuals in the observed sample must expend to move to extreme arrangements, in order to keep individuals either at uniformly (regularly) or aggregated (patchy) intervals.By comparing counts of the observed spatial pattern with random needed rearrangements, the degree of non-randomness within a set of data can be quantifi ed (Perry 1998).Once the sampling-unit counts are the same for each randomization, the original heterogeneity of the sampled data is preserved (Xu 2003).Finally, distribution maps were produced using the kriging technique from the index values generated for each cluster plot, in which the aggregation index of each species was interpolated along the sampling area.Th ese analyses were performed with the SADIEshell program (Perry et al. 1996;Conrad et al. 2006).
We used SADIES's association index (Perry & Dixon 2002) to assess the spatial association of all sampled individuals, species richness and among most abundant species in order to answer the second question of this study.Th is spatial association refers to the similarity between the spatial patterns of two data sets, and it was equivalent to a spatially explicit correlation coeffi cient that tests the level of association between two populations (Bell et al. 2010).According to Perry & Dixon (2002), a positive association is expected when the spatial distributions of two species join both in patches and gaps.In addition, if the spatial distribution of patches and gaps do not coincide, a spatial dissociation is expected.Th e spatial associations were undertaken with the SADIEshell program (Perry et al. 1996;Conrad et al. 2006).
Rarefaction curves from the number of sampled individuals (Koellner et al. 2004) were created to compare species richness to other studies developed on the slopes of the Serra Geral region in Rio Grande do Sul, Brazil.By this methodology, the expected richness can be estimated in a random previously determined individual sample (Waite 2000).Following this, the results of other studies carried out in the slopes of the Serra Geral formation, in Rio Grande do Sul, were adapted for the construction of the rarefaction curves.In this way, a matrix of species abundance was built and, based on Forzza et al. (2010), the species names were updated and their synonymy was verifi ed.Th e survey showing the lowest number of individuals was set as the basis for the comparison.Some studies were not included due to a low sampling eff ort or by omitting the names of species or grouping, which could weaken the comparison.Neophytes were excluded from this analysis but non-identifi ed taxa or taxa identifi ed at the level of family or genus were kept.Alpha (α) diversity was calculated through Shannon's diversity index with Pielou's evenness index (neophytes were excluded).Rarefaction and diversity analysis were performed within the PAST package (Hammer et al. 2001).
Spearman correlation coeffi cient (r s ) was used to verify whether there was a reduction of species richness from east to west along Serra Geral formation.In order to answer the third question of this study we further partitioned the amount of species from each fl oristic contingent (western seasonal and eastern wet).Correlation analyses were performed with JMP® soft ware (Version 9. SAS Institute Inc., Cary, NC, 1989-2007).

Results
Soil Th e soil samples showed high proportion of silt and low proportions of clay and sand, leading to a soil class "four", which group soils with medium content of organic matter, base saturation and pH.Th e cation exchange capacity (CEC) was considered high, in the same way as Calcium (Ca), Magnesium (Mg) and Sulfur (S).Zinc (Zn), Boron (B) and Manganese (Mn) levels were considered high, while copper (Cu) was considered medium.Th e Aluminum (Al) saturation was null and base saturation had a high rate (Tab.1).

Structure
Th is study surveyed 1,024 living trees belonging to 69 species and 33 families.Th e species with highest absolute densities were Casearia sylvestris, with 127 individuals (12.4%),Nectandra megapotamica, with 106 (10.3%),Sorocea bonplandii, with 94 (9.2%), Alchornea triplinervia, with 72 (7.0%), Cupania vernalis, with 67 (6.5%) and Allophylus edulis, with 51 (5.0%).Together these species totaled 50.4% of surveyed individuals (Tab.2).With respect to absolute frequency, a small number (13%) of the sampled species occupied higher values of frequency classes (between 20% and 60%), while 87% of sampled species had frequency classes between 0% and 20%.Nectandra megapotamica was the most frequent (AF = 59%).Regarding dominance, the overall basal area was 24.10 m²•ha -1 , where the highest values were recorded for N. megapotamica, which had 9.04 m²•ha -1 , A. triplinervia with 2.82 m²•ha -1 and Tetrorchidium rubrivenium with 2.12 m²•ha -1 .Together these species account for 58% of the total value of dominance.Nectandra megapotamica held 37.5% of the total basal area, with several large sized individuals (height and diameter values above average).Casearia sylvestris and Sorocea bonplandii had high values of density and frequency and low values of dominance because they showed several well distributed small trees.On the other hand, Tetrorchidium rubrivenium and Ocotea puberula had higher values of dominance with lower values of frequency and density.Concerning the diameter, 50% of sampled individuals had diameters between 4.8 and 9.7 cm (Fig. 1).
Th e spatial distributions of Alchornea triplinervia and Diospyros inconstans were signifi cantly associated to the spatial distribution of species richness per sampling unit or, as the case of Cupania vernalis, showed a dissociated pattern between overall species richness and plant species abundance (Tab.4).Th e spatial distributions of Casearia sylvestris, Alchornea triplinervia, Allophylus edulis, Myrsine umbellata and Tetrorchidium rubrivenium were signifi cantly associated with the overall number of individuals (Tab.4).Furthermore, we verifi ed that some species were signifi cantly associated (p < 0.025) with each other.On this way, regardless of their abundance, the spatial patterns (distribution) are similar both in areas of patches (high abundance) and in areas of gaps (low abundance).At the same time, some species showed signifi cant dissociation (p > 0.975) with each other, indicating that the spatial patterns in both areas (patches and gaps) did not match.According to this, Tab. 5 shows the signifi cant dissociation between Alchornea triplinervia and Cupania vernalis and, as expected, their associations with other species, which were frequently dissimilar; Diospyros inconstans showed signifi cant association with A. triplinervia and signifi cant dissociation with C. vernalis, while Myrsine guianensis showed signifi cant association with A. triplinervia and a dissociation trend with C. vernalis.Th e same occurred between Sorocea bonplandii and Allophylus edulis.Both of these species showed a tendency to be dissociated with each other but S. bonplandii showed a trend to be associated to Cabralea canjerana, which also showed a dissociation trend with A. edulis.Finally, we observed that Casearia sylvestris was signifi cantly associated to A. edulis and showed a trend to be dissociated to C. canjerana.Only C. vernalis did not show associations with any other species.

Alpha Diversity and Species Richness
Th e Shannon's diversity index (H') showed a diversity of 3.34 nats.ind with Pielou's evenness of J' = 0.79.According to the rarefaction procedure, the survey located in the farthest eastern region (municipality of Morrinhos do Sul) showed the highest species richness (92 species), followed by Montenegro, also located in the east (66) and this study, in central region (66 species), then Santa Maria in the west (55) and Vale do Sol in the central region (48) (Tab.3).Th e latter, although located a bit westward, had more characteristics from the eastern contingent than the present one, probably due to its elevation, justifying its ordinance before our survey in Tab. 3. We did not fi nd a reduction of species richness from east to west (r s = -0.61;p = 0.27).Aft er partitioning the amount of species of eastern and western fl oristic contingents, we observed a trend of increasing   species for the western seasonal species (r s = 0.80; p = 0.10) from east to west and a trend of decreasing for the eastern wet species (r s = -0.90;p = 0.03) in the same direction.Th ere was no correlation between eastern wet and western seasonal (r s = -0.60;p = 0.28).

Discussion
50% of the sampled individuals had a diameter between 4.8 and 9.7 cm (Fig. 1).Th is result highlights the importance of the understory and regenerating individuals in the community, while, if we had adopted a higher inclusion criterion (e.g. 10 cm DBH, as used in some studies in tropical and subtropical forests), we would have found only middle and upper layers and half of the surveyed individuals.
Spatial distribution and association patterns -Th ree species that showed aggregation in the present study also had aggregation in the study of Lindenmaier & Budke (2006), located in Cachoeira do Sul, Rio Grande do Sul, in a riverine forest.Although they used a higher inclusion criterion, Casearia sylvestris, Sorocea bonplandii and Cabralea canjerana showed signifi cant values of aggregation in their survey, the latter had a signifi cant value of aggregation in our study.Th is could indicate that these species present aggregation not only due to local factors but also as a natural behavior.
Most species showed significant or relatively high values of association or dissociation with other ones.In other words, all of them had some degree of association or dissociation, not occurring regularly in the horizontal space.Th e causes of such patterns are not well understood or studied, although credit has mainly focused on biotic and abiotic interactions.Nevertheless, some considerations can be made to understand part of these processes; if there is association between species, it indicates that they occupied the same areas (patches and gaps) in the survey.So we can infer that I) one of them is being (or has been) favored by the other (Levine, 1999;Brooker et al. 2008), which may indicate habitat changing in the way one or both species had been successfully established (facilitation); II) both are (or could have) benefi ting from a third biotic or abiotic factor(s); III)   (-) = signifi cant dissociation ; N = density (ind.ha - ); I a = index of aggregation; Mvj = average value of I a for gaps; Mvi = average value of I a for patches.Spi x Spp = P values obtained with association index between spatial distribution of species "i" (Spi) and spatial distribution of number of species for each sampling unit (Spp); Spi x Nind = P values obtained with association index between spatial distribution of species "i" and spatial distribution of number of individuals for each sampling unit (Nind).there is stand-specifi c competition between them (Goldberg & Barton 1992;Devictor et al. 2010) or it had existed at some moment; and IV) both species developed mechanisms to coexist among each other (e.g., creating microsites favorable for coexistence) or among other unknown factors (Christie & Armesto 2003;Silvertown 2004).
Alchornea triplinervia was associated with seven species (two signifi cant associations and 4 tendencies) and had only one dissociation (signifi cant).Th us, this species also had an association tendency with the spatial distribution of species richness per sampling unit, suggesting that this species occurs in areas with high heterogeneity.Th e question is why these interactions occur and whether this species itself is the cause of such heterogeneity.Th e association trends of Alchornea triplinervia with Myrsine umbellata and with Sorocea bonplandii can be emphasized.We can infer that aft er the last disturbance (e.g.canopy opening by the fall of a large tree), M. umbellata was one of the fi rst tree species which became established, due to its pioneer characteristic (Oliveira et al. 2005).Alchornea triplinervia, which is lightdemanding (Rôças et al. 1997) and occupied the highest layer in the present survey, was probably benefi ted (I) and could establish itself.Finally, S. bonplandii, which is shade tolerant (Budke et al. 2008), could establish itself (I) in the shade provided by A. triplinervia and occupy the lowest layer.At the same time and corroborating this hypothesis, there was a dissociation trend between Myrsine umbellata and Sorocea bonplandii, since their preferences (e.g.regarding luminosity requirements) are diff erent.
Nectandra megapotamica tends to be associated with Alchornea triplinervia and Tetrorchidium rubrivenium.This three species are light-demanding (Vaccaro et al. 1999;Budke et al. 2008) and were large trees in the survey (so they occupied the same stratum).It is still not known if there is some type of facilitation between them (I) or if there is another factor that could benefi t them (II), but we can infer that maybe there was (or still there is) competition between them (III) or these species have developed co-existing mechanisms (IV).Th e same can be inferred to the signifi cant associations that occurred between Casearia sylvestris with Allophylus edulis and Myrsine guianensis with Cabralea canjerana, for example.All of them are lightdemanding species (Budke et al. 2008) and had an average size in the survey, occupying the same stratum.
Cupania vernalis had signifi cant dissociation with the spatial distribution of species richness per sampling unit and tends to be dissociated with the spatial distribution of the number of individuals per sampling unit.Nevertheless, C. vernalis was the only species that did not have an association with other species.In a recent study, Gabriel et al. (2007) showed that the young individuals of C. vernalis did not follow the distribution of adults in a forest in southern Brazil.Th is may be an important strategy of this species and reinforces the hypothesis that it forms dense and dominant groups.
Alpha Diversity and Species Richness -Th e value of species diversity obtained through the Shannon Index (H') of 3.35 nats.indcan be considered high when compared to other surveys made on the slopes of the Serra Geral formation.We attribute this elevated value to natural disturbances in the location: I) due to the terrain's inclination and soil characteristics, the two gullies in the studied area probably interfered in the dominance of one species, and II) these gullies provide a permanent canopy gap, which favored the establishment of widely distributed pioneer or lightdemanding species with low density values.Th e infl uence and functioning of these disturbances in the richness of tropical and subtropical forests, has been demonstrated in current studies (Schintzer & Carson 2001;Sheil & Burslem 2003;Smith et al. 2005).Th ese disturbances play an important role in the structure of plant communities, leading to environmental heterogeneity in time and space at diff erent scales (Budke et al. 2010).
A reduction of species richness can be seen from east to west when comparing the farthest east survey, located in the tropical rain forest domain, with our survey and with the farthest west survey, both located in the seasonal forest domain.Th e latter had 40% less species than the farthest east one.However, the correlation analysis done with the total amount of species of each survey showed that there is not a clear reduction of species richness from east to west because some of the compared surveys showed an oscillation in their richness values, so that the reduction line from east to west was not rectilinear.Th is fact can be related to local factors and not regional, because canopy gaps, big tree falls, or other local disturbances may create microsites which infl uence species richness and forest structure (Christie & Armesto 2003;Smith et al. 2005;Budke et al. 2010).It is also worth noting that two surveys, despite being in relatively close longitudes (e.g., our survey and the survey done in Vale do Sol), may have diff erences in their richness due to the elevation factor (Budke et al. 2004).From this premise, the present survey (done at 500 m.s.l.) had less species from the east contingent and also more species from the west contingent than the survey located in Vale do Sol (done at 100-140 m.s.l.), which is located a few kilometers to the west.So we considered that the survey of Vale do Sol had more characteristics from the east contingent than the present one.
Aft er partitioning the amount of species of each fl oristic contingent (western seasonal, eastern wet and wide geographical distribution) for each survey, a pattern became clear with the correlation analysis: at the same time that a reduction of the eastern wet species from east to west occurs, there is an increase of western seasonal species in the same direction.From this, we can infer that beyond a species reduction process there is a replacing process of the Atlantic species to seasonal tree species towards the west.

Figure 1 .
Figure 1.Distribution of tree individuals by diameter class (class interval of 4.8 cm), obtained in a survey in a seasonal forest in the central region of Rio Grande do Sul State, Brazil.

Table 1 .
Physical and chemical parameters obtained with soil samples collected in a seasonal forest in central region of Rio Grande do Sul state, Brazil.

Table 2 .
Structural parameters of a seasonal forest in central region of Rio Grande do Sul state, Brazil, collected in 100 contiguous subplots of 10 x 10 m² each.

Table 3 .
Data of surveys realized in the slope of Serra Geral formation, in Rio Grande do Sul state, Brazil, aft er the adaptation procedure (revision of synonyms, exclusion of neophytes), ordered from east to west.

Table 5 .
Probability values (p) obtained with association analysis between trees species with at least 20 individual surveyed in a seasonal forest in central region of Rio Grande do Sul, Brazil.Signifi cant association (+) occurs when p < 0.025 and signifi cant dissociation (-) when p > 0.975.

Table 4 .
Values obtained with index of aggregation and association for species with at least 20 individuals surveyed in a seasonal forest in central region of Rio Grande do Sul state, Brazil.