Effects of land use on soil organic carbon and microbial processes associated with soil health in southern Brazil

European Journal of Soil Biology 55 (2013) 117e123

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European Journal of Soil Biology journal homepage: http://www.elsevier.com/locate/ejsobi

Original article

Effects of land use on soil organic carbon and microbial processes associated with soil health in southern Brazil Daniel Bini a, Cristiane Alcantara dos Santos a, Kellen Banhos do Carmo a, Nagomi Kishino a, Galdino Andrade b, Waldemar Zangaro c, Marco Antonio Nogueira a, * a b c

Programa de Pós-Graduação em Microbiologia, Universidade Estadual de Londrina (UEL), Londrina, PR, Brazil UEL, Laboratório de Ecologia Microbiana, Depto. de Microbiologia, Londrina, PR, Brazil UEL, Laboratório de Micorrizas, Depto. de Biologia Animal e Vegetal, Londrina, PR, Brazil

a r t i c l e i n f o

a b s t r a c t

Article history: Received 2 August 2012 Received in revised form 19 December 2012 Accepted 31 December 2012 Available online 17 January 2013 Handling editor: Yakov Kuzyakov

Carbon plays a key role in determining soil health, which is defined as the soil’s capacity to maintain environmental functions and biological productivity. In this study, C cycling was evaluated in soils along a gradient of land use, from native forest (NF), reforested sites (secondary forest e SF, Araucaria angustifolia e AR, Pinus elliottii e PI), clear-cut P. elliottii stands (CT) to farmland (AG). NF, AR, and SF sites had lower litter C:N ratios than PI, CT, and AG sites. Soils under forests had more organic C than CT and AG soils, whereas soils with native species had more microbial biomass C than PI, CT and AG soils. Both metabolic quotient (qCO2) and dehydrogenase activity increased with land use. Multivariate analysis revealed that soils of AR and SF were similar to NF and differed from CT and AG, which had higher qCO2 and dehydrogenase activity, suggesting microbial stress. Litter C:N ratios and soil microbial biomass C, moisture, CO2 evolution, and cellulase activity discriminated most effectively between land uses. Reforestation with native species restored soil properties to levels similar to those in NF, being more sustainable, whereas reforestation with the exotic P. elliottii made soils more similar to AG soils. Ó 2013 Elsevier Masson SAS. All rights reserved.

Keywords: Araucaria angustifolia Pinus elliottii Reforestation Soil enzymes Soil quality

1. Introduction Most native vegetation in southern Brazil is classified as Mixed Ombrophilous Forest, which is often characterized by the conifer Brazilian Pine [Araucaria angustifolia (Bertoloni) Otto Kuntze]. This ecosystem has been reduced to up 0.8% of its original extent by deforestation and the establishment of farmland, pastures, or plantations of exotic timber species like Pinus sp. and Eucalyptus sp. Although fragments of native vegetation remain in protected areas, A. angustifolia is considered an endangered species [1]. These dramatic changes in vegetation cover are expected to modify soil C content and nutrient cycling [2e4], thus impacting environmental equilibrium and sustainability [5]. Soil health has been defined as the capacity of a soil to support ecosystem functions and sustain biological productivity and environmental quality, while promoting plant and animal health [6]. Poor soil management and the replacement of native forests by

* Corresponding author. Present address: Embrapa Soja, Laboratório de Biotecnologia do Solo. Cx. Postal 231, 86001-970 Londrina, PR, Brazil. Tel.: þ55 43 3371 6215; fax: þ55 43 3371 6100. E-mail addresses: [email protected], [email protected] (M.A. Nogueira). 1164-5563/$ e see front matter Ó 2013 Elsevier Masson SAS. All rights reserved. http://dx.doi.org/10.1016/j.ejsobi.2012.12.010

farmland or exotic timber species may compromise soil health. Because several chemical, physical, biological and biochemical properties are used to characterize soil health, identifying the most sensitive of these is useful for assessing the impacts of land use change. Levels of soil organic C, generally considered an indicator of soil health [7], depend on inputs of plant litter and rhizodeposition [2,8,9]. Changes in a site’s original vegetation or improper soil use may impair C cycling, decrease soil organic matter content, and increase CO2 emissions [8,10], thereby contributing to the greenhouse effect. Vegetation composition affects rhizodeposition and litter quality [3,4]. Plant litter with a high C:N ratio accumulates on the soil surface due to slow biodegradability [11], while litter with a C:N ratio lower than 25 is easily degraded, with net mineralization of nutrients [12]. Assessment of microbial processes related to C cycling can provide important information on C and nutrient contents in soils under different land-use systems. Soil respiration is an indicator of microbial activity that is usually correlated with organic matter content and microbial biomass [5,2]. The metabolic quotient (qCO2) [13] can be used as an ecological indicator, and expresses the metabolic effectiveness of soil microbial communities [14]. Soil microorganisms are also the source of several

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exoenzymes, including cellulase and amylase, which are involved in C mineralization. Dehydrogenases are active in living cells, and their relative activity levels are taken as an indicator of microbial activity [15]. Given that these microbial parameters and enzyme activities are commonly changed by land use [16], quantifying them can help evaluate the changes in soil microbial functions driven by changes in land use [5]. This study focused on two questions: (i) How does intensifying land use affect soil microbial soil attributes related to C cycling?, and (ii) Which microbial soil attributes best discriminate between different land-use systems? The aim of the study was to evaluate indicators of soil health related to C cycling in soils under different land use, and to identify sensitive indicators that have high potential as tools for monitoring the effects of land-use changes. 2. Material and methods 2.1. Site description and sampling

Each site differing in land use had at least 40 ha, within which eight transects (experimental plots of 15  5 m) were randomly established in order to provide eight independent soil samples, composed by 15 subsamples. Soil was randomly sampled at 0e 10 cm depth with a steel auger along each transect, pooled to form a composite sample, and sieved (2 mm). The litter layer was removed before sampling. Field-moist samples were stored at 5  C for microbial and biochemical analyses or air-dried for 72 h for chemical analysis. Gravimetric soil moisture was determined at 105  C for 24 h to express laboratory results on a dry basis and to standardize moisture levels whenever necessary. Litter samples were collected in eight randomly sited 0.25 m2 squares (0.5  0.5 m) at the same transects used for soil samplings. No distinction was made for the levels of litter decomposition. The samples were oven-dried at 60  C and ground in a mill (<0.5 mm) for chemical analysis. 2.2. Chemical and physical analyses

The study was carried out in November 2007 in Guarapuava, Paraná State, southern Brazil (25 230 S; 51270 W), in six sites subjected to different land use and long-term changes in soil use since native forests were clear cut 30e70 years ago (Table 1). The study region is 1120 m above sea level, with rolling hills and slopes ranging from 6 to 20%. Soils are clayey, acidic, naturally nutrientpoor (Table 2), and classified as Typic Acrudox [17]. Climate is classified as Cfb (humid subtropical mesothermic) under the Köppen system, with a poorly defined dry season. Average temperature is 22  C in austral springesummer (Sept.eMar.) and 18  C in austral autumnewinter (Apr.eAug.), with some occurrence of minimum temperatures below freezing. Mean annual rainfall ranges from 1400 to 1600 mm, with more intense rains between December and March. Apart from differing in land uses, environmental conditions were considered identical at all sites.

Total C in soil and litter was assessed by dichromate oxidation in acidic medium [18]. Total N was measured by semi-micro Kjeldahl [19] after sulfuric digestion to obtain the C:N ratios of soil organic matter and litter. Water-dispersed clay content was determined in 20 g of oven-dried (105  C) samples, after successive periods of shaking in distilled water and settling [20]. To determine soil bulk density (m/v), eight non-deformed samples were collected randomly at 0e10 cm depth with steel cylinders in each site, ovendried (105  C) for 24 h, and weighed. 2.3. Microbiological and biochemical analyses The length of filamentous fungal hyphae was estimated in 5 g of soil previously incubated with 2% pyrophosphate for 1 h to disperse

Table 1 Dominant species, age, soil use history and actual soil management in each sampling site representative of different land uses in Guarapuava, PR, southern Brazil. Intensification of land use (1 / 6)

Dominant species during the sampling

Plant age

History/current use

1. Native forest (NF), reference site.

Allophylus edulis, Araucaria angustifolia, Campomanesia xanthocarpa, Capsicodendron dinisii, Casearia decandra, Dicksonia sellowiana, Ilex paraguariensis, Ocotea pulchella, Pimenta pseudocaryophyllus, Vitex megapotamica, Zanthoxylum rhoifolium Araucaria angustifolia; understory with Capsicodendron dinisii, Fleurya aestuans, Geophila sp, Pteridium sp., Solanum paniculatum, Solanum pseudocapsicum Acacia plumosa, Mimosa scabrella, Nectandra megapotamica, Ocotea sp, Piper mikanianum, Pteridium sp, Solanum verbascifolium Pinus elliottii

Native

Preserved fragment of native forest (Atlantic forest) with 41 ha, open to public visitation and research in the municipal park, considered as climax vegetation. In addition to the arboreal species, the understory is also occupied by herbaceous species. Site previously cropped with Pinus for 40 yr before reforestation with A. angustifolia.

2. Reforestation with Araucaria angustifolia (AR), native species.

3. Secondary forest (SF), native species.

4. Reforestation with Pinus elliottii (PI), exotic species.

32 years

13 years

21 years

5. Clear cutting of P. elliottii (CT), exotic species.

Pinus elliottii seedlings, Solanum verbascifolium

1.5 year

6. Agricultural cropping (AG), annual crops.

Maize (Zea mays)

15 days

Site previously cropped with Pinus for 40 yr with natural regeneration of secondary forest after Pinus clear cutting, occurrence of accidental burning, and abandonment. Site cropped for more than 40 yr with Pinus. Ten years after planting the actual forest, 30% of the trees were cut; clear understory. The same as site 4, but harvested 1.5 year before. After clear cutting, pioneer gramineous and bushy vegetation started to re-establish among some spontaneous P. elliottii seedlings with 0.6 m high. Site under annual crops for more than 30 yr, initially conventional tillage, followed by no-tillage in the last 7 yr. Cropped with oat in the last winter for cattle grassing, but sometimes cropped with wheat or barley in years before. Cropped with soybean or maize in the summer. Use of mineral fertilizers, liming, and pesticides.

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were independent, we considered that the main exigencies for applying ANOVA were fulfilled, and thus the individual composite samples were treated as independent replicates [27]. Thus, the dataset was subjected to one-way ANOVA and means comparisons via Tukey’s test (p  0.05), according to an entirely randomized experimental design for comparisons between sites. Data were also subjected to multivariate analysis by principal component analysis (PCA) using Canoco software [28] to provide an integrated view of the variable patterns according to land use. Canonical discriminant analysis (CDA) was also performed to identify the soil properties most strongly associated with land use. Identification is based on low values in the Wilk’s Lambda multivariate statistical test due to low colinearity, and highly significant differences among the sites (P < 0.01). The individual contribution of each variable in the separation of sites along the diagram is expressed as the parallel discrimination coefficient (PDC), which is the product of the standardized canonical coefficient (SCC) and the correlation coefficient (r) between each variable and the canonical discriminant function.

Table 2 Chemical properties and granulometric fractions at 0e10 cm of soil depth in sites under different land uses in Guarapuava, PR, southern Brazil. Soil property

Land use NF

AR

SF

PI

CT

AG

Available P (Resin) (mg dm3) pH (H2O) Al (cmolc dm3) H þ Al (cmolc dm3) Ca (cmolc dm3) Mg (cmolc dm3) K (cmolc dm3) CEC (cmolc dm3) Base saturation (%) Sand (g kg1) Silt (g kg1) Clay (g kg1)

4 4.5 35 169 13 6 2.6 191 11 88 326 586

7 4.3 67 222 7 6 3.0 237 6 80 334 586

7 4.6 40 196 29 14 4.7 243 20 117 346 608

7 4.0 94 239 3 3 2.5 247 3 73 319 537

7 4.8 21 166 52 20 2.3 240 31 75 310 615

15 6.0 <1 50 114 61 4.8 231 78 91 346 563

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NF e native forest; AR e reforestation with Araucaria; SF e secondary forest; PI e reforestation with Pinus; CT e clear cutting of Pinus; AG e agricultural cropping.

soil aggregates and release hyphae. Counting procedures are fully described in Ref. [21]. To measure basal respiration, moisture was adjusted to 60% of water holding capacity (WHC) and samples incubated for 21 days. The NaOH trap was replaced every two days and titrated with standardized 0.5 mol L1 HCl [22]. Microbial biomass C (Cmic) was estimated via fumigatione -extraction [23]. Briefly, soil moisture was adjusted to 60% of WHC and fumigated with chloroform for 24 h at 25  C. Nonfumigated subsamples accounted for the soluble carbon background. After extraction with 0.5 mol L1 K2SO4, soluble organic carbon in the extracts was determined by oxidation with dichromate [24]. Cmic was calculated as the difference between fumigated and non-fumigated extracts, with a KC value of 0.33. Microbial biomass N (Nmic) [25] was determined using a KN value of 0.54, in the same extracts in which total N was determined after sulfuric digestion [19] and used to calculate the C:N ratio of the microbial biomass. The ratio between basal respiration and Cmic yielded the metabolic quotient (qCO2) [13]. Amylase (EC 3.2.1) and cellulase (EC 3.2.1.4) activities in soil were assessed in 10-g samples incubated for 24 h in phosphate buffer (pH 5.5), with specific substrates and temperatures (starch at 37  C for amylase and carboxymethylcellulose at 50  C for cellulase). The reducing sugars resulting from hydrolysis were quantified via spectrophotometry (690 nm) using the Prussian blue method [26]. To determine dehydrogenase activity, 5-g samples were incubated with 5 mL of 1% triphenyl tetrazolium chlorine (TTC) and incubated at 37  C for 24 h. Triphenyl tetrazolium formazan (TTF) was subsequently extracted with methanol, filtered, and read via spectrophotometry at 485 nm [15].

3. Results 3.1. Litter and soil organic carbon Litter amounts were highest in clear cut Pinus stands (CT) and declined as follows: CT > reforested Pinus elliottii stands (PI) > reforested A. angustifolia stands (AR) ¼ secondary forest (SF) ¼ native forest (NF) > farmland (AG) (Table 3). The highest C:N ratios for litter were found at the PI, CT, and AG sites, twice as high as those at the native species reforestation sites (AR and SF) and in NF. Soils at forest sites, whether native or reforested, generally contained more total organic C than the CT and AG sites (Table 3). The C:N ratio of soil organic matter was lowest at the SF site and highest at the AR and PI sites. 3.2. Soil physical traits Soil moisture at sampling was highest at the AR site, and declined as follows: AR > (NF ¼ SF ¼ PI) > CT > AG (Table 4). Soils under NF and reforested with native species (AR and SF) showed the lowest water-dispersed clay contents, and differed significantly from AG, which showed the highest contents (Table 4). The trend was similar for soil bulk density, with the lowest values at AR, NF, and SF, increasing at PI and CT, and the highest at the AG site. 3.3. Soil microbiological and biochemical traits SF soils contained the highest amounts of filamentous fungal hyphae, about three times more than NF and AG soils, which contained the lowest amounts (Table 5). Similarly, SF soils showed the highest levels of respiration, which decreased at the other reforested sites and was the lowest in the AG soils. Cmic was highest in AR and NF soils, and decreased in SF, PI, CT, and AG soils (Table 5). Metabolic quotient (qCO2) was lower at NF, AR, SF and PI soils,

2.4. Statistical analyses All data were examined for homogeneity of variances and normal distribution. Taking into account that the composite samples

Table 3 Litter and soil traits associated with C in sites under different land uses in Guarapuava, PR, southern Brazil. Trait

Land use

Litter (ton ha1) Litter C:N ratio Soil total C (g kg1) Soil organic matter C:N

10.5 23.1 40.4 11.4

NF

AR (1.7) (5.5) (4.5) (0.8)

c b ab ab

17.2 25.4 45.2 11.7

SF (4.9) (4.0) (6.7) (1.8)

c b a a

11.4 22.1 40.8 9.5

PI (3.0) (6.0) (5.8) (1.3)

c b ab b

22.4 43.3 40.4 12.7

CT (3.9) (8.0) (7.3) (1.3)

b a ab a

50.7 51.0 34.0 11.1

AG (25.7) a (7.6) a (6.4) bc (2.0) ab

3.8 44.1 30.1 11.6

(1.0) (8.1) (4.3) (0.8)

d a c ab

Means sharing the same letter are not statistically different (Tukey, p  0.05). Numbers in brackets represent the standard deviation (n ¼ 8). NF e native forest; AR e reforestation with Araucaria; SF e secondary forest; PI e reforestation with Pinus; CT e clear cutting of Pinus; AG e agricultural cropping.

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Table 4 Soil physical traits in sites under different land uses in Guarapuava, PR, southern Brazil. Trait

Moisture (%) Dispersed clay (g kg1) Bulk density (g cm3)

Land use NF

AR

SF

PI

CT

AG

52.7 (4.4) b 16.9 (7.0) b 0.62 (0.06) cd

65.3 (4.2) a 18.8 (14.6) b 0.54 (0.13) d

52.2 (2.8) b 20.6 (8.6) b 0.65 (0.06) cd

50.0 (2.0) b 30.0 (20.2) ab 0.73 (0.04) bc

44.1 (4.3) c 37.5 (20.0) ab 0.82 (0.12) b

35.7 (3.4) d 46.3 (12.5) a 1.04 (0.11) a

Means sharing the same letter are not statistically different (Tukey, p  0.05). Numbers in brackets represent the standard deviation (n ¼ 8). NF e native forest; AR e reforestation with Araucaria; SF e secondary forest; PI e reforestation with Pinus; CT e clear cutting of Pinus; AG e agricultural cropping.

increased at CT, and was the highest in AG soils. The C:N ratio of microbial biomass did not differ between sites (Table 5). Dehydrogenase activity was highest in AG soils, followed by NF and the other sites (Fig. 1A). Conversely, amylase activity was highest in AR soils, with similar activity in SF and PI, decreasing in NF, CT, and the lowest in AG soils (Fig. 1B). Cellulase activity showed more variation than amylase but a similar trend, with the highest levels in PI and AR soils, intermediate in NF, SF, and CT soils, and the lowest levels in AG soils (Fig. 1C). 3.4. Global data analysis Axis 1 of the PCA indicated that NF and sites reforested with native species (AR and SF) were more similar to each other than to the AG and CT sites (Fig. 2). The variables more associated with the native species cluster were soil respiration, Cmic, and microbial biomass C:N ratio. Conversely, the variables associated with AG and CT soils were dehydrogenase activity, qCO2, water-dispersed clay, and litter C:N ratio. Cellulase and amylase activities, and total organic C were negatively associated with the CT and AG sites. The PI site was the most dissimilar along axis 2 of the PCA, and was associated with cellulase activity and the C:N ratio of soil organic matter. The CDA resulted in four canonical discriminant functions (CDF), in which CDF1 explained 80.64%, CDF2 11.85%, CDF3 4.26%, and CDF4 3.2% of the discrimination among sites (Fig. 3). The functions that explained variation included variables with discriminating power (Cmic, soil respiration, soil moisture, litter C:N ratio, and cellulase activity), which were responsible for the distribution pattern of the sites in the dispersion diagram, while the other variables did not show significant discriminating power to separate sites. Thus, along the axis that represents the CDF1, the variables that more contributed to separate land use were soil moisture and Cmic, because they had a higher PDC in the most explanatory function (CDF1). On the CDF2 axis, soil respiration and litter C:N ratio were the most important variables to discriminate the sites (Table 6). 4. Discussion Organic matter has key influence on physical, chemical and biological soil attributes. Since changes in land use disturb the

equilibrium between the formation and mineralization of soil organic matter [4], they also change soil attributes. Quantification of attributes associated with C dynamics in soil is thus useful for forecasting the effects of land-use change on soil functions [10]. The lower C content of AG soils reflected the accelerated oxidation of organic matter and the lower inputs of organic material as compared to the native and reforested sites [10,29e32]. Less inputs of organic material lead to less protected soils, causing water loss, increases in water-dispersed clay and bulk density, and higher qCO2 and dehydrogenase activity [10,31,32]. Conversion of native forest to farmland strongly impacts physical and chemical soil attributes, as well as the soil microbial community associated with C cycling [10]. More organic C has been found in native and secondary forest soils than in agricultural soils [32,33]. Nevertheless, although secondary forest was more than 20 yr old in that study [32], organic C was still lower than in native forest soils, which highlights the difficulty to recover the original levels of soil organic C in some cases. In the present work, however, the secondary forest with 13 yr old had concentrations of organic C similar to the native forest soil. This difference can be attributed to the wetter and cooler climate conditions, making easier the recovery of soil organic C levels. Pinus plantations have been found to contribute weakly to organic C levels in mineral soil due to low levels of bioturbation [4]. Several other studies have reported decreases in soil organic C following the replacement of native vegetation by Pinus [3,4,8]. In this study, levels of soil organic C in CT were comparable to those in AG soils, despite the greatest amount of litter at CT site. This decrease is attributable to the litter quality of Pinus, which is less prone to decaying and slow in returning C to the soil [8]. Even 30 years after a native grassland field was afforested with Pinus taeda, the remnant soil organic C was found to derive from the original vegetation, and shows a negligent contribution from P. taeda litter [3]. Inputs of organic material with lower C:N ratios, despite less accumulated litter, resulted in more Cmic and organic C in sites with native vegetation than sites currently or recently with Pinus. Inputs of more easily degradable litter stimulate microbial activity and biomass due to the abundance of substrates for microbial use [11,35]. Reforestation with Araucaria or native species resulted in less alteration of soil microbial properties than reforestation with Pinus. Soils at long-term (>60 yr) reforestation with Pinus in southern

Table 5 Soil microbiological traits in sites under different land uses in Guarapuava, PR, southern Brazil. Trait

Land use NF

Hyphae (m g1) Soil respiration (mg CeCO2 g1 d1) Cmic (mg kg1) qCO2 (mg CeCO2 g1 Cmic h1) C:Nmic ratio

4.7 81.7 1582 2.2 12.4

AR (1.0) c (5.9) bc (379) ab (0.4) c (2.5) a

11.2 96.6 2033 2.1 15.6

SF (4.3) b (8.6) b (506) a (0.7) c (5.4) a

16.5 116.8 1342 3.7 8.9

PI (4.5) a (18.2) a (227) bc (0.9) bc (1.6) a

7.0 72.6 897 3.5 11.2

CT (3.1) bc (11.4) cd (198) cd (0.7) bc (5.5) a

7.0 87.1 778 5.1 14.5

AG (2.2) bc (19.2) bc (299) de (2.0) b (10.2) a

4.8 60.8 370 8.5 8.9

(1.8) c (8.7) d (205) e (3.9) a (6.6) a

Cmic ¼ carbon microbial biomass; qCO2 ¼ metabolic quotient; C:Nmic ¼ C to N ratio of the soil microbial biomass. Means sharing the same letter are not statistically different (Tukey, p  0.05). Numbers in brackets represent the standard deviation (n ¼ 8). NF e native forest; AR e reforestation with Araucaria; SF e secondary forest; PI e reforestation with Pinus; CT e clear cutting of Pinus; AG e agricultural cropping.

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Fig. 2. Principal component analysis (PCA) based on microbial, biochemical, physical and chemical soil attributes in soil under different land uses in Guarapuava, PR, southern Brazil. NF e native forest; AR e reforestation with Araucaria; SF e secondary forest; PI e reforestation with Pinus; CT e clear cutting of Pinus; AG e agricultural cropping. TOC e total organic carbon; C:N OM e C:N ratio of the soil organic matter; WDC e water-dispersed clay; qCO2 e metabolic quotient; C:N MB e C:N ratio of microbial biomass; Cmic e carbon microbial biomass.

Fig. 1. Dehydrogenase (A), amylase (B), and cellulase (C) activities in soil under different land uses in Guarapuava, PR, southern Brazil. NF e native forest; AR e reforestation with Araucaria; SF e secondary forest; PI e reforestation with Pinus; CT e clear cutting of Pinus; AG e agricultural cropping. Means sharing the same letter are not statistically different (Tukey, p  0.05). TTF ¼ triphenyl tetrazolium formazan. Vertical bars represent the standard deviation (n ¼ 8).

Brazil had less microbial biomass N and N-cycling than reforestation with Araucaria [34]. Similarly, a decrease in Cmic in soils reforested with Pinus has been observed in Korea [8]. In the present study, intensification of land use reduced microbial activity and Cmic, as has been observed in some Pinus plantations [3,4] and also in farmlands in Brazil [32,34]. Under stressful conditions, the soil microbial community is less effective at using environmental resources and especially organic carbon as substrate, resulting in higher qCO2 [13], which sometimes coincides with higher dehydrogenase activity [36,37]. High dehydrogenase activity in soils with low microbial biomass indicates a more intense electron flow due to respiration under adverse conditions [5] and this is in agreement with a higher qCO2 index. However, qCO2 must be interpreted carefully because higher qCO2 indices are also correlated with the availability of easily degradable organic substrates [29], and do not necessarily indicate stressful conditions. For this reason, qCO2 is best interpreted together with dehydrogenase activity and microbial biomass C rather than on its

own. Mature ecosystems have larger, more diverse, and more metabolically-effective microbial biomass [38], as observed in the soils reforested with native species, where the metabolic status of the microbial community represented by qCO2 was similar to that of the native ecosystem. Pinus plantation understories have a very low diversity of litter. The release of allelopathic substances like volatile organic compounds (e.g. terpenes) negatively affects the establishment of other plant species and microbial activity [8,12,36,37,39]. Conversely, sites reforested with native species had more diverse inputs of litter, in addition to a lower litter C:N ratio. The SF site had been

Fig. 3. Dispersion diagram for canonical discriminant analysis (CDA), relating the first and the second canonical discriminating functions (CDF1 and CDF2) on the standardized canonical coefficients for microbial biomass carbon, soil respiration, cellulase activity, soil moisture, litter C:N ratio. NF e native forest; AR e reforestation with Araucaria; SF e secondary forest; PI e reforestation with Pinus; CT e clear cutting of Pinus; AG e agricultural cropping.

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Table 6 Standardized canonical coefficient (SCC), coefficient of correlation (r) and parallel discrimination coefficient (PDC) for the canonical discriminating function 1 (CDF1) and 2 (CDF2), based on physical, chemical, and microbiological soil traits related to C cycling. Trait

Soil respiration Cmic Cellulase activity Litter C:N ratio Soil moisture

CDF1

CDF

2

SCC

r

PDC

SCC

r

PDC

0.236 0.689 0.001 0.476 0.799

0.233 0.413 0.060 0.393 0.592

0.055 0.285 0.0001 0.187 0.473

0.787 0.337 0.253 0.689 0.505

0.501 0.067 0.242 0.482 0.377

0.394 0.023 0.061 0.332 0.190

planted with Pinus, but after cutting and abandonment for approximately 13 yr, a secondary forest with native species had established spontaneously, providing enough time to restore most properties (e.g. litter C:N ratio, soil total C, Cmic, enzyme activity, and basal respiration) to near NF levels. A similar pattern was observed at the AR reforested site. These results illustrate how the use of native species helps to restore several soil properties associated with C cycling in sites with a long-term history of exotic timber species plantation. At sites with native vegetation, lower water-dispersed clay indicated more stable aggregates, as a result of higher microbial activity and biomass on soil aggregation [8]. Lower bulk densities were also observed in these soils owing to bioturbation. Not only do microorganisms affect soil physical properties, but soil physical properties also affect the microbial community. Soil management that leads to disruption of aggregates generally decreases the amounts of microsites, compromising the establishment and maintenance of the microbial community [40]. Less water-dispersed clay content coincided with more filamentous fungal hyphae in SF and AR, but not in NF. In a more stable ecosystem, several other factors, like soil organic matter content and quality, rhizodeposition, microbial biomass and the exopolysaccharides they produce, increase the stability of aggregates [8,9], reducing the relative importance of filamentous fungal hyphae in this process [21]. As with microbial properties, reforestation with native species also restored soil physical conditions, especially water-dispersed clay and bulk density, to levels similar to those in NF. Higher cellulase activity at the PI site is explained by the cellulose-rich litter [41,42]. However, despite the greater amount of litter at the CT site, cellulase activity was as low there as in AG soils. Less favorable conditions, such as lower levels of soil organic C and moisture, may have impaired soil microbial activity and enzyme release, resulting in lower activity [39,41]. The presence of living plants is an important stimulus for enzyme activity because microbial activity is increased in the rhizosphere [35]. However, amylase and cellulase are not only sensitive to the quantity and quality of the litter that reaches the soil, but also to physical and chemical properties of soil [35,39]. Even so, soil enzymes are useful tools for assessing soil quality following anthropogenic disturbances in ecosystems [16], in part because they reflect functional processes of soil microorganisms that cannot be assessed by culturing. In the multivariate PCA, axis 1 can be interpreted as a gradient of land use. Considering native forest and farmland as extremes of land use, the proximity of the reforested sites to either of these extremes reflects their level of change as a consequence of land use. Clear cut is an aggressive process caused by intensive traffic and soil compaction, like in farmland. In addition, the high C:N ratio and contents of lignin, resins, and cellulose result in accumulation of litter on soils cropped with Pinus and consequently slow down the cycling of C and nutrients [8,12,36,37]. The qualitative changes in litter following the replacement of native vegetation modify C

dynamics [4,37], and thus soil microbial and biochemical activities [34,43]. The CDA discriminated variables associated with key processes of C cycling in soil, such as Cmic, moisture, respiration, litter C:N ratio, and cellulase activity. Therefore, differences between sites are mainly attributed to differences in litter quality (C:N ratio) and soil moisture regimes as a consequence of land use, and effects on the soil microbial biomass and activity [44]. At sites reforested with native species, the lower litter C:N, in addition to the more diversified rhizodeposition in a more diverse plant community, favored microbial and biochemical processes. Thus, Cmic and basal respiration were sensitive indicators to discriminate between sites. Cellulase activity and litter C:N were also effective to separate sites, and were closely associated with Pinus, which showed the highest litter C:N ratios. 5. Conclusions The soil use for plantations of Pinus elliottii is less impacting than the agricultural use with farmlands, but after the forest clear cutting, the soil attributes tend to be similar to those of the farmland. On the other hand, reforestation with the native woody species A. angustifolia, or native species occurring spontaneously in the secondary forest, tends to retrieve the soil properties toward the preserved native forest fragment, improving sustainability. This is attributed to the diversified organic residues of easier degradation produced by native species, allowing better conditions for microbial community establishment and the processes they play on C cycling. Soil properties like dehydrogenase activity, qCO2, respiration, Cmic, litter C:N, soil moisture, and cellulase activity were better discriminators among different land uses and are promising tools for monitoring strategies of land use in similar conditions. Acknowledgments We thank Parque Municipal das Araucárias and the Cooperativa Agrária Agroindustrial (Guarapuava, Paraná) for allowing us to carry out field work. We also thank Mr. Jorge Elio Bini for providing information on the histories of the sites and support during field work. Brazil’s National Council for Scientific and Technological Development (CNPq) and Program for the Advancement of Upperlevel Personnel (CAPES) are acknowledged for fellowships. M.A. Nogueira and G. Andrade are CNPq scholars. This text was approved for publication by the Editorial Board of Embrapa Soja as manuscript 07/2012. References [1] FAO e Food and Agriculture Organization, Databook on Endangered Tree and Shrub Species and Provenances, in: FAO Forestry Paper (1986). Rome. [2] Y. Li, M. Xu, M. Zou, Y. Xia, Soil CO2 efflux and fungal and bacterial biomass in a plantation and a secondary forest in wet tropics in Puerto Rico, Plant Soil 268 (2010) 151e160. [3] M. Wiesmeier, D.P. Dick, C. Rumpel, R.S.D. Dalmolin, A. Hilscher, H. Knicker, Depletion of soil organic carbon and nitrogen under Pinus taeda plantations in southern Brazilian grasslands, Eur. J. Soil Sci. 60 (2009) 347e359. [4] W. Wilcke, J. Lilienfein, Soil carbon-13 natural abundance under native and managed vegetation in Brazil, Soil Sci. Soc. Am. J. 68 (2004) 827e832. [5] F. Bastida, J.L. Moreno, T. Hernández, C. García, Microbiological activity in a soil 15 years after its devegetation, Soil Biol. Biochem. 38 (2006) 2503e2507. [6] J.W. Doran, M. Safley, Defining and assessing soil health and sustainable productivity, in: C.E. Pankhurst, B.M. Doube, V.V.S.R. Gupta (Eds.), Biological Indicators of Soil Health, CAB International, New York, 1997, pp. 1e28. [7] O.H. Smith, G.W. Petersen, B.A. Needelman, Environmental indicators of agroecosystems, Adv. Agric. 69 (2000) 75e97. [8] C.W. Park, S. Ko, T.Y. Yoon, S. Han, K. Yi, W. Jo, L. Jin, S.J. Lee, N.J. Noh, H. Chung, Y. Son, Differences in soil aggregate, microbial biomass carbon concentration, and soil carbon between Pinus rigida and Larix kaempferi plantations in Yangpyeong, central Korea, For. Sci. Technol. 8 (2012) 38e46.

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