Biol Fertil Soils (2010) 46:851859 DOI 10.

1007/s00374-010-0490-8

ORIGINAL PAPER

Adequacy of contrasting sampling methods for root mass quantification in a slash-and-burn agroecosystem in the eastern periphery of Amazonia
Joo Thiago Rodrigues de Sousa & Christoph Gehring

Received: 3 June 2009 / Revised: 21 July 2010 / Accepted: 23 July 2010 / Published online: 12 August 2010 # Springer-Verlag 2010

Abstract Root research needs to optimize sampling schemes to address different scales and dimensions of variability within a framework of feasible effort and, in some cases, acceptable soil perturbation. Both large and small sample volumes are utilized in root research, and this study is designed to help field researchers in optimizing root sampling. We evaluate the performance of three contrasting sampling strategieslarge monoliths (LM, 25 dm3), small monoliths (SM, 31 dm3), and root auger (RA, 5196.3 cm3). We compare root biomass estimates obtained in a shifting cultivation agroecosystem in eastern Amazonia. We sampled a slash-and-burn field at 1/2 and 1 1/2 years of cultivation, and a paired 3-year-old spontaneous secondary forest regrowth down to 1 m soil depth, and we distinguish roots by diameter class and taxonomic origin (palm vs. non-palm roots). LMs are the only reliable quantification method for coarse roots. Both SM and RA methods are inadequate due to high variability and the frequent failure to detect the presence of coarse roots in the sampled soil. The SM method is acceptable for mid-sized (25 mm) roots, and both methods are adequate for fine root (<2 mm) quantification. Labor costs of LM, SM and RA methods are similar since reduced sampling effort in small sample volumes is compensated by increased costs in sample handling. The small sample volumes of SM or RA may be preferable in situations focusing on fine root dynamics, with the necessity to avoid major perturbations, or in repeated sampling schemes. Both SM and RA samplings gave a strong and systematic overestimation of
J. T. Rodrigues de Sousa : C. Gehring (*) Maranho State University, Cidade Universitria Paulo VI s/n Tirirical, 65000-000 So Luis, Maranho, Brazil e-mail: christophgehring@yahoo.com.br

root biomass in all fractions compared to the LM values, with differences being less pronounced for SM. We establish comparability between the three sampling schemes with transformation equations which are remarkably similar between shifting cultivation and secondary forest, between roots of the babassu palm and those of other origin and down the 1-m soil profile. Thus, future field studies in similar environments can utilize SM or RA fine root sampling and subsequently transform data to LM estimates. We recommend the SM rather than the RA method because of the lower data variability and the lower degree of root mass overestimation. Keywords Fine roots . Coarse roots . Root auger . Root biomass . Monolith . Sampling scheme . Variability

Introduction Knowledge of root stocks and of fine root dynamics is crucial for quantifying belowground carbon stocks (Sommer et al. 2000), nutrient cycling and buildup (Attiwill and Adams 1993; Szott et al. 1991; Sommer et al. 2004; Rasmussen et al. 2010), competition for water and nutrients (Noordwijk and Willingen 1986; Schroth et al. 1996), plant adaptive strategies and niche partitioning (Fitter 1987), vegetative regeneration (Miyanishi and Kellman 1986; Kauffman 1991), etc. However, contrary to aboveground vegetation, our knowledge on root systems remains very limited, especially in heterogeneous spontaneous woody vegetation. This discrepancy is due to the considerable labor costs and serious methodological problems involved in root quantification. Whereas isotopic methods have proven to be successful in studying root distribution, nutrient uptake (Wahid 2001;

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Peek and Forseth 2005; Gttlicher et al. 2008) and partially fine root turnover (Hendricks et al. 1997; Majdi et al. 2005; Rasmussen et al. 2010), destructive excavation remains the only means for root biomass (C and nutrient stocks) quantification (Bhm 1979). However, the high labor demand limits feasibility of large-scale root excavations, whereas, the high spatial variability of differing root components on scales ranging from centimeters to meters frequently compromises the reliability of root data (Casper and Jackson 1997). For woody vegetation, root biomass and C stocks are usually dominated by the coarse (i.e., >5 mm diameter) root components, which account for about 70% of total root biomass (Cairns et al. 1997). Though fine roots are quantitatively less important than coarse roots, they are decisive for nutrient uptake, belowground competition, net primary productivity, belowground carbon turnover and buildup (Eissenstat et al. 2000; Gill and Jackson 2000), nitrogen buildup (Rasmussen et al. 2010), and soil organic matter formation (Six et al. 2002). This paper evaluates the performance of contrasting sampling strategies for quantifying fine and coarse roots and establishes comparability between root estimates generated by these sampling schemes. In a slash-and-burn agroecosystem of the eastern periphery of Amazonia, we compare three of the most common sampling schemes (Bhm 1979; Smit et al. 2000): large monoliths, small monoliths and root augers. These schemes represent a sequence of decreasing volume of sampled soil and increasing quantity of internal replications. Research is based on the hypothesis that large monoliths due to the large volume of sampled soilresult in root biomass estimates which are closest to reality and therefore can serve as basis (true values) for comparisons with the other two sampling schemes. We explore data variability, betweenscheme relationships, and other relevant parameters in a cultivation and a secondary forest fallow site for fine and coarse roots, palm and non-palm roots, and throughout 1-m soil profile. We furthermore give estimates of labor costs associated with the three sampling schemes. This way, we provide researchers with a decision basis for developing root sampling schemes which optimally fit to their respective necessities and specific field settings, and we establish comparability between studies under similar environmental settings which utilized different sampling schemes.

Materials and methods Study region and research sites The research was conducted in a slash-and-burn shifting cultivation agroecosystem in the eastern periphery of

Amazonia, on So Luis island, Maranho State, Brazil (241S, 4416W). Annual rainfall amounts to about 2,000 mm, mainly concentrated in a 7-month rainy season. Dominance of slash-and-burn shifting cultivation in smallholder agriculture remains unbroken. Root data were generated in a slash-and-burn shifting cultivation field and in a nearby spontaneous 3-year-old secondary forest site, 1 m down the soil profile. Sites represent the cultivation and fallow phases of slash-andburn agriculture. We quantified roots at the end of rainy season both in the secondary forest and in the cultivation site at 1/2 and 1 1/2 years cultivation. Cropping sequence of the cultivation site was maize and upland rice intercropped with beans (caupi) and cassava, typical for smallholder agriculture throughout large parts of Amazonia. Weeding intensity diminished along the cropping sequence and resulted in a gradual transition from cultivation to fallow phase. Initial secondary regrowth was well established at cassava harvesting after 1 1/2 years. Both cultivation and fallow sites have suffered an unknown but high number of similar preceding slash-andburn land use cycles. The intention of this study is to cover a wide range of environmental conditions and thereby guarantee the general validity of our results for other field studies with similar agro-environment. We therefore base our investigations both on spontaneous 3-year-old secondary forest and on 1/2 and 1 1/2 years cropping, and cover 1 m soil depth. Aboveground biomass of the 3-year-old secondary regrowth site was estimated at 30.5 t ha1 and of the cultivation site after 1 1/2 years cultivation at 6.8 t ha1. The share of the babassu palm in aboveground biomass was estimated at 14% in the fallow site and 26% in the cultivation site after 1 1/2 years (unpublished data). The two study sites also differed substantially in their root biomass estimates (Table 1). Topsoil (010 cm) total root mass was three times as high in the secondary forest site (15.7 t ha1) than in the cultivation site (4.7 t ha1). Soils of the two study sites are classified as Typic Paleustult. They are deeply weathered. Texture is dominated by fine sand and is quite homogenous throughout the soil profile of both sites, with the absence of impermeable rock or clay horizons which could affect vertical root distribution (Table 2). This thorough weathering and vertical homogeneity is typical for the Oxisols and Ultisols which dominate throughout the humid tropics (Sanchez 1976). Soils are acidic and infertile. In contrast to soil physical parameters, soil chemistry differed between the two study sites as a consequence of vegetation burning, with significantly higher pH, extractable P and cation levels in the cultivation than in the fallow site (Table 3).

Biol Fertil Soils (2010) 46:851859 Table 1 Key root biomass estimates for 01-m depth of the cultivation and the secondary forest sites (values are site medians, n =15 sampling points estimated by the LM method) Area Total root mass (t ha1) 1.98 2.39 Fine roots (<2 mm) (t ha1) 0.78 1.27

853 % Babassu palm root biomass share 16.5 45.5

Cultivation site Secondary forest site

Sampling scheme We compare the accuracy and costs involved in three contrasting sampling methods, (1) RAroot auger: five cores per sampling point totalling 981.7 cm3 volume per 10-cm soil layer, (2) SMmicro-monolith: three cores per sampling point totalling 3 dm3 volume per 10-cm soil layer, and (3) LMlarge monoliths: 25 dm3 volume per 10-cm soil layer. Given the large soil volume, LM values are assumed to be the closest ones to the in situ situation and thus represent the true values of root mass and composition. Sampling is paired on a total of 1013 sampling points according to Fig. 1. Root classification We distinguish roots according to two criteria, (1) root diameter and (2) taxonomic origin. Definitions of fine and coarse roots differ between species and root positions (Majdi et al. 2005; Rasmussen et al. 2010), with the most commonly used diameter thresholds being 2 and 5 mm (Castellanos et al. 2001; Sierra et al. 2003). We therefore distinguish between fine (<2 mm), intermediate (25 mm), and coarse (>5 mm) roots. We combined two diameter classes (5 10 mm and >10 mm) in a single coarse root class in order to reduce the very high variability of these two diameter classes. We furthermore distinguish roots by their taxonomic origin into roots of the dominant babassu palm (Attalea speciosa C.Martius) and all other roots. This distinction was facilitated by the characteristic brown-red color of the lignified roots of the babassu palm.

Table 4 gives a general characterization of the dataset of this study, covering a total of 1013 sampling points within the two study sites. The wide spectrum of ecological conditions in this study is reflected in the wide range of values for all key variables. Root processing Roots were washed over a 0.73-mm mesh sieve 23 times (SM and RA samples) or 35 times (LM samples) for the complete removal of soil particles, and then they were oven-dried for 5 days at 70C. The secondand most laboriousstage of root processing involved (1) separation of roots into four diameter classes and two taxonomic groups, and (2) manual separation of roots from charcoal and other debris with a pince. No attempt was made to distinguish between live and dead roots due to the lack of objective and unambiguous selection criteria (Sierra et al. 2003). Roots were finally weighed on a fine balance for subsequent calculation of root stock estimates. Precision and cost of sampling methods We evaluate the precision of SM and RA methods by considering the LM as reference method. For this purpose, we apply the following criteria: Degree and significance of differences in biomass estimates between the methods and down the soil profile;

Table 2 Soil physics of the cultivation and the secondary forest site (total n =96 samples), mean (standard deviation) soil bulk density, and median % texture, methods according to Embrapa (1979) Soil density (g.cm3 standard deviation) 1 1/2-year-old cultivation site 010 cm 1.24 1040 cm 1.38 40100 cm 1.50 3-year-old secondary forest site 010 cm 1.29 1040 cm 1.31 40100 cm 1.34 % coarse sand (20.2 mm) % fine sand (0.020.05) % silt (0.050.002) % clay (<0.002)

(0.03) (0.04) (0.04) (0.04) (0.05) (0.08)

33 35 33 31 37 35

58 55 55 58 53 54

2 3 3 3 3 5

7 7 8 8 7 8

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Table 3 Soil chemistry in the 1 1/2 year-old cultivation site and the secondary forest site (medians, total n =96 samples taken between 01-m soil depth), analyses conducted according to standard methods used by the Agronomic Institute of Campinas (IAC 2001) Soil org. matter (g kg1)a 1 1/2 year-old cultivation site 010 cm 34.8 1040 cm 18.6 40100 cm 9.0 3 year-old secondary forest site 010 cm 25.2 1040 cm 14.9 40100 cm 9.5
a b c

pH (CaCl2)

P (mg1)b

K (mg kg1)b

Ca (mg kg1)c

Mg (mg kg1)c

5.3 5.4 4.9 5.1 4.8 4.4

18.7 10.4 4.6 3.8 1.9 1.3

6.3 2.8 2.6 17.7 3.0 2.9

723.2 311.2 130.2 307.0 135.7 63.3

125.4 95.4 64.1 99.4 58.5 28.5

Walkley&Black digestion Extraction with synthetic ion exchange resins Extraction with 1 M KCl

Similarity of values assessed as the relative size of intercept and deviation of slopes from 1:1 inclination; Predictability of values: R2 and p values of regression models; Percentage of false zero estimates (i.e., the failure to detect the presence of the respective root diameter class with SM or RA even though present, as stated by LM values); and Variability of estimates (%CV).

dominant babassu palm vs. all other roots). Costs were integrated over soil depth, with excavation costs being higher and processing costs lower in the deeper soil horizons. Costs were computed in man-days, assuming well-trained personnel and 8 h labor per man-day. Statistical analyses We tested all data for normality of distribution (KolmogorovSmirnov and Lilliefors tests). With the exception of fine root percentages quantified by the RA method, all data were normally or ln-normally distributed, as checked visually (histograms) and by KolmogorovSmirnov and Lilliefors tests against normality; ln-normally distributed data were ln-transformed to achieve normality and allow for parametric procedures. Possible redundancy of relationships between variables was visually evaluated by plotting residuals against predicted values. Residual values were always normally distributed, no correlation between residuals and predicted values was apparent, and no redundancies were detected with the Duncans test. Significance of ANOVAs and of regression analyses are given as ***p < 0.001, **p <0.01, *p <0.05, and n.s. p >0.05. Statistical analyses were conducted with Statistica 7.0 (StatSoft 2004).

We quantify labor costs of the three contrasting sampling schemes by compiling costs of sampling (excavation), drysieving and/or washing, manual cleansing (removal of charcoal and other debris), and separation into four size classes and two taxonomic groups (i.e., roots of the

Results Figure 2a and b compares total root biomass (left) and fine root biomass (right) estimates obtained by the three sampling methods throughout the 1-m soil profile. Large and systematic differences between the three sampling methods are evident for all diameter size classes and down the 1-m soil profile. RA estimates are consistently higher than LM values, with differences being significant at almost all soil depths. The SM-based estimates are intermediate,

Fig. 1 Paired root sampling with a large monolith, b small monolith, and c root auger sampling positions

Biol Fertil Soils (2010) 46:851859 Table 4 Characterization of the dataset of the three contrasting sampling schemes (over both study sites, medians of 1013 sampling points)
Total roots at 010 cm (t ha1) Large monolith Small monolith Root auger 6.16 5.87 15.29 Fine roots at 010 cm (t ha1) 3.61 5.31 14.85 % fine roots at 0100 cm 31.1 69.1 88.4 % babassu roots at 0100 cm 27.8 41.3 33.5

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with differences with LM values being non-significant in most cases, especially in the topsoil. Table 5 gives the median percentages of deviation of SM and RA estimates of total and of fine root estimates, as well as the respective estimates of babassu palm shares in total and fine root biomass as compared to LM values. Both SM and RA samplings overestimate the share of babassu palm roots for unknown reasons. Overestimation of total root biomass was consistently smaller than that of fine root biomass. Relative root biomass overestimation increases with soil depth probably due to the decreasing absolute root biomass values with soil depth. Root mass overestimation is considerably more serious with RA sampling, with overestimation up to half a magnitude higher than with SM sampling. Table 6 shows the results of regression analyses between SM and RA estimates and LM values. Relationships are lnlinear in all cases, and are more significant for fine roots than for mid- and coarse-sized roots. The inclination of regression lines is steeper for RA- than for SM-based
Fig. 2 Total root biomass (a) and fine root biomass (b) estimates (SE bars) generated by the three root sampling methods large monolith (full circles), small monolith (open circles), and root auger (full triangles). Significant differences between methods within the same soil layer are indicated with stars

estimates, resulting in the strong and significant root biomass overestimation of RA-based studies already shown in Fig. 2 and Table 5. Figure 3 exemplifies the relationships for fine root estimates obtained by RA sampling relative to LM values. Relationships are remarkably similar between the two study sites inspite of the large differences in absolute values. The capability of the differing root sampling methods to detect the presence of all root fractions in the soil (as stated by LM results) is a further important criterion on the quality and reliability of root sampling methods. Both SM and RA methods always detected fine roots (both babassu and of other origin) when present. False zero estimates generated by SM were 0% (010 cm layer) and 30% (1040 cm layer) for roots thicker than 5 mm in diameter, and 0% at both depths for roots with 25 mm diameter. These values were higher for RA data, with 63% and 60% false zero estimates for roots thicker than 5 mm diameter at 010 cm and 10 40 cm soil layers, and 50% and 20% for roots with 25 mm diameter at 010 cm and 1040 cm soil layers, respectively. Both SM and RA sampling schemes must therefore be considered unreliable for coarse diameter roots. The frequent failure of SM and RA samplings to detect coarse roots did not impede the overestimation of coarse or total root biomass, but may have been a reason for the lower degree of overestimation of coarse roots than of fine roots, as stated in Fig. 2 and Table 5. Table 7 shows the variability associated with the three contrasting sampling schemes down the soil profile. Variability increases with decreasing root density down the soil profile, and is higher for the coarse than for the fine diameter classes in all three sampling methods, reflecting

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Table 5 Pairwise comparison of root estimates generated with small monolith and with root auger: % deviation from large monolith values Small monolith Soil depth 010 cm 1020 cm 2030 cm 4050 cm 7080 cm 90100 cm average Fine roots 101.1 421.5 296.3 286.9 All roots 59.1 197.9 145.3 138.2 % Babassu in fine roots 141.0 121.8 139.3 128.5 % Babassu in all roots 243.6 189.7 129.8 187.9 Root auger Fine roots 432.3 981.2 659.7 1,024.6 1,032.8 1,906.6 659.6 All roots 123.1 166.4 284.5 513.0 397.7 473.1 275.6 % Babassu in fine roots 95.4 83.3 108.7 100.9 120.9 169.0 105.1 % Babassu in all roots 243.1 187.9 59.6 157.2 85.4 157.5 137.9

natural variability of root occurrence. Variability is, however, also affected by root sampling method, and is substantially higher with RA than with LM and SM. Table 8 gives an overview of the labor demand per root sample (i.e., composite of 15 subsamples per sampling point and soil depth) for the three sampling methods. Whereas labor requirements for on-field sampling involving small sample volumes are only half (SM) or one third (RA) of those associated with the LM method, the labor requirements for washing and especially for manual cleansing and separation of samples into diameter classes are higher. Overall costs for the three sampling methods are very similar (0.230.25 man-days per sample).

Discussion The choice of root sampling schemes depends on the research objectives, which may focus on root stocks (mainly dominated by mid-sized and coarse roots) as opposed to a focus on more dynamic processes driven by fine roots (nutrient uptake, root turnover, or productivity). Root sampling schemes also necessarily reflect the feasi-

bility of associated costs and the acceptability of associated perturbations. Root biomass variability differs with root diameter, species, soil profile, and time (Espleta and Clark 2007; Rasmussen et al. 2010). Coarse and mid-sized roots dominate in multi-cycle regrowth of multi-cycle slashand-burn agroecosystems due to the predominance in vegetative resprouting and clonal connections between woody plant individuals (Baar 1997). Excessive variability of coarse root estimates both with SM and RA samplings and the frequent failure of SM and RA samples to detect the presence of coarse roots put into question the meaningfulness of coarse root estimates generated by these methods. Sample volumes of both methods obviously are insufficient to adequately cover coarse-grained spatial variability of large-diameter roots. Thus, large-volume excavations are the only meaningful manners to adequately cover this root component. Findings in this respect have been reported by Bhm (1979) and Wiesenmller (1999). Spatial variability of fine roots presumably is lower than that of coarse roots, and fine root quantification therefore requires smaller soil sample volumes. Fine roots can be estimated with fair precision with SM and RA sampling

Table 6 Regression analyses between large monolith (SM) and root auger (RA) samplings and values generated with large monoliths (LM) which are assumed to represent the true values of root biomass

Equation LM/SM Fine roots (<2 mm) Mid-sized roots (52 mm) Coarse roots ( >5 mm) All roots LM/RA Fine roots (<2 mm) Mid-sized roots (52 mm) Coarse roots ( >5 mm) All roots

R2

ln ln ln ln

(LM)= 0.65+0.970 * ln (SM) (LM)=6.51+0.076 * ln (SM) (LM)=5.0+0.442 * ln (SM) (LM)=3.94+0.544 * ln (SM)

0.43 0.34

** n.s. * n.s. *** n.s. 0.06 ***

*P <0.05; **P <0.01; ***P < 0.001

ln (LM)= 6.31+1.510 * ln (RA) ln (LM)=4.97+0.205 * ln (RA) ln(LM)= 1.85+1.110 * ln(RA) ln (LM)= 4.33+1.368 * ln (RA)

0.78 0.41 0.56

Biol Fertil Soils (2010) 46:851859 Fig. 3 ln-linear relationship between fine root estimates generated by root auger (RA) and by large monolith (LM) methods. Different symbols represent the cultivation site after 1 1/2 years cultivation and the 3-year-old fallow site

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schemes, given the existence of sufficient internal replications. The small soil sample volumes of SM and RA schemes make significant internal replications per sampling point both possible and a necessity. Bhm (1979) discuss the required number of internal replications, Noordwijk et al. (1985) recommend sample replications of >22 for fine root quantification in Dutch grasslands and row crops fields. Variability of RA data was consistently higher than that of LM or SM sampling for all root components and throughout the soil profile (Table 7). We see such high variability associated with RA data as a serious drawback of this method. We do not have information whether increased internal replication (i.e., more than the five internal replicates applied in this study) would have reduced variability (and at the same time increased labor expenses). In a study involving grass biomass estimation with RA, Bengough et al. (2000) did not detect a systematic reduction of %CV between four and 100 replications, suggesting that variability of RA data is rather a consequence of insufficient sample volumes than of lack of internal replications. Spatially systematic root sampling schemes have been proposed for fine root biomass and length estimation in

planted forest regrowth (Berish and Ewel 1988), but are rather meaningless in heterogeneous slash-and-burn agroecosystems with unpredictable spatial relationships between aboveground vegetation and fine roots. Schroth and Kolbe (1994) propose sample homogenization followed by 10% subsampling as a manner to economize on labor effort. Vertical fine root distribution can be modeled, and root sampling can economize on some soil horizons by efficient vertical interpolation (Wiesenmller et al. 1998; Cook and Kelliher 2006). Whereas local vertical and horizontal variation in soil rootability can strongly affect root distribution and pose challenging problems in many field settings such as rocky outcrops or aggregated soil layers (Noordwijk et al. 1985), this source of variability may be considered unproblematic under the deeply weathered soils with homogenous soil texture (Table 2) which predominate in large areas of Amazonia (Sombroek 1984; Chauvel et al 1987) and in the humid tropics worldwide (Sanchez 1976). Both SM and RA methods overestimate root biomass (Fig. 2 and Table 5). Differences were large for all root components and soil horizons, though in the case of SM, frequently non-significant. We do not know the reason for

Table 7 Variability associated with the three root sampling methods: %CV of investigated root classes down the soil profile Large monolith (LM) Root class >5 mm 52 mm <2 mm All roots 010cm 127.3 81.1 63.0 73.0 1040cm 107.9 56.8 60.6 67.3 40100cm 169.5 96.5 83.1 80.3 Small monolith (SM) 010cm 104.0 52.8 50.0 50.1 1040cm 196.2 136.0 49.3 63.8 40100cm Root auger (RA) 010cm 216.4 132.7 42.3 33.5 1040cm 232.3 123.5 31.2 39.2 40100cm 238.6 133.4 36.8 60.5

858 Table 8 Estimates of labor demand involved in the three root sampling schemes First stage: field sampling Method N internal replicates 1 3 5 min/ replicate 60 10 4 min/ sample 60 30 20 Second stage: washing min/ replicate 20 10 8 min/ sample 20 30 40

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Third stage: separation and weighing min/ replicate 30 20 10 min/ sample 30 60 50

Total costs per root sample total min 110 120 110 man-day/ sample 0.23 0.25 0.23

LM SM RA

LM large monolith, SM small monolith, RA root auger

this overestimation. Soil compaction may have played a certain role, but cannot account for the high degree of overestimation, since our soils are sandy (Table 2) and therefore, presumably, not subject to serious compaction problems during sampling. Overestimation of total root biomass by SM and RA sampling methods was consistently smaller than fine root overestimation associated with these methods. The failure of small volume SM and RA samplings to hit coarse roots and the corresponding high portion of false zero estimates may have been one of the reasons for this finding. The lack of standardized root sampling procedures makes comparisons of published data problematic both on a regional and global scale (Robinson 2004) and even within studies (Berish and Ewel 1988). An important objective of this paper is to establish comparability between different root sampling strategies. The two study sites (cultivation site at 1/2 and 1 1/2 years and 3-year-old fallow site) differ strongly in biomass and composition of aboveground and of root biomass, and even in soil chemistry (Tables 1 and 3). These large differences are the outcome of site selection which strived to cover a wide range of environmental conditions in order to ensure the general applicability of our results. Even so, relationships between the sampling schemes remain remarkably constant between both sites and throughout the soil profile inspite of the large differences in absolute values, as shown in Fig. 3. Relationships are also similar for babassu roots and roots of other taxonomic origin. This robustness of the here described relationships makes us confident in respect to their general applicability, a prerequisite for their utility in other studies conducted within similar environmental settings (i.e., slash-and-burn agroecosystems on deeply weathered sandy soils). Root sampling strategies are always a compromise between the desirable precision and practical feasibility. The high labor costs constitute the main limiting factor in the pursuit of meaningful and efficient root sampling schemes (Schroth and Kolbe 1994). Fine root processing (i.e., washing, manual removal of debris by handpicking, and separation into different root classes) is the most laborious

part of root quantification according to our results and to those of other studies (Vogt et al. 1998; Wiesenmller 1999). However, correct procedures are crucial for the quality of generated data since root mass losses caused by washing and processing amount to 2040% of total root biomass (van Noordwijk and Norris 1979; Robinson 2004) and far larger portions of the fine or very fine roots. Overall costs for the three sampling methods are very similar (0.230.25 man-days per sample) since reduced labor demand for excavating smaller sample volumes is compensated by the high labor demand for sample processing. In field situations in which root sampling does not need to minimize associated perturbations, we therefore recommend LM sampling as single sampling method.

Conclusions This paper is designed to provide researchers with a decision basis for designing root sampling schemes with maximized efficiency, and to establish comparability between different root sampling strategies. As to be expected, coarse roots need to be quantified in large monoliths. Root sampling by small sample volumes does not reduce labor demand. Therefore, we recommend the LM method as the single sampling scheme in field situations in which the associated perturbation does not pose a problem. In field situations in which major soil perturbations associated with large sample volumes need to be avoided, fine roots can be quantified with reasonable precision with SM or RA sampling. However, the resulting values should be transformed to account for the root biomass overestimation associated with small sample volumes. We recommend the use of SM rather than RA sampling for this purpose due to the lower root biomass overestimation, lower portion of false zero estimates, and the lower data variability of this method. Our paper provides field researchers with transformation equations valid for fine and mid-sized (<5 mm) roots over a wide range of environmental conditions within slash-and-burn agroecosystems on highly weathered sandy soils of the humid tropics.

Biol Fertil Soils (2010) 46:851859 Acknowledgments We gratefully acknowledge the financial support by the Brazilian Research Council CNPq (CT-Universal 02/2006) as well as a research grant provided to the first author by the Research Fund of Maranho State (FAPEMA).

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