Contents lists available at SciVerse ScienceDirect
Geoderma
j ourna l homepage: www.e lsev ie r .com/ locate /geoderma
Soil microbial biomass and activity under natural and regenerated forests andconventional sugarcane plantations in Brazil
Danielle Karla Alves da Silva a, Nicácio de Oliveira Freitas a, Renata Gomes de Souza a,Fábio Sérgio Barbosa da Silva b, Ademir Sérgio Ferreira de Araujo c,⁎, Leonor Costa Maia a
a Programa de Pós-Graduação em Biologia de Fungos, Departamento de Micologia, Universidade Federal de Pernambuco, Av. Prof. Nelson Chaves s/n, 50670‐420 Recife, PE, Brazilb Universidade de Pernambuco—Campus Petrolina (UPE), BR 203 Km 2, 56300‐000 Petrolina, PE, Brazilc Universidade Federal do Piauí, Centro de Ciências Agrárias, Programa de Pós-Graduação em Agronomia, Campus da Socopo, CEP 64000‐000, Teresina, PI, Brazil
⁎ Corresponding author. Tel.: +55 86 3215 5740; faxE-mail address: [email protected] (A.S.F. de Ar
Article history:Received 4 December 2011Received in revised form 13 June 2012Accepted 20 June 2012Available online 18 August 2012
Keywords:Enzyme activityAtlantic forestMicrobial respirationMicrobial biomass C
The objective of this study was to evaluate the changes in soil microbial biomass and activity after the regener-ation of deforested and cultivated soil in Brazil. Soil sampling was carried out in June and December 2007 (wetand dry seasons, respectively), at four areas, including: a native forest (NF), a 10 years old regenerated forest(RF10), a 20 years old regenerated forest (RF20) and a conventional sugarcane plantation (CL). Microbial bio-mass C, substrate-induced respiration and cellulase, saccharase, hydrolysis of fluorescein diacetate and dehydro-genase activities were analyzed. Themicrobial variables varied significantly among different areas depending onthe season. Usually, larger differences were observed in the RF10 than in others, in dry season, for soil microbialbiomass C and enzyme activities. Themultidimensional scaling analysis showed that the RF10 has distinctmicro-bial and biochemical characteristics comparing with the other areas. In general form, microbial activity, as mea-sured by enzyme activities, is higher in native forest and regenerated lands than in cropland.
The Atlantic Forest is the second largest tropical forest in theAmerican continent (Tabarelli et al., 2005). Nowadays, this forest isfragmented and only 11 to 16% of its original area still exists becauseof substantial deforestation for conversion to croplands (Ribeiro et al.,2009).
Forest conversion to agriculture can decrease soil C storage, as natu-ral vegetation is cut and replaced by crops that support lower soil C con-tents and plant biomass (Houghton, 1990). However, there is evidencethat, after extensive use, agricultural lands that are abandoned and thatundergo subsequent regeneration of forest may recover C storage topre-agricultural levels. However, the rate of recovery depends on thetime frame one considers and whether the land was previously usedas cropland or pasture (Post and Kwon, 2000). Therefore, this regener-ation process may promote the recovery of soil properties, mainly soilbiological processes, which are important for soil fertility and plantgrowth. Microorganisms mineralize, oxidize, reduce and immobilizemineral and organic materials in soil (Kennedy and Doran, 2002). Anychanges in the soil may alter the number or the activity of soil microor-ganisms, which can, in turn, affect soil biochemical processes and, ulti-mately, influence soil fertility and plant growth (Munier-Lamy andBorder, 2000).
: +55 86 3215 5743.aujo).
rights reserved.
Microbial biomass is the living component of soil organic matter(SOM) (Jenkinson and Ladd, 1981), and its activity has been shownto be an early and sensitive indicator of soil changes (Araújo et al.,2010; Chander and Brookes, 1993). Soil respiration is one of themost widely used parameters for quantifying microbial activity insoil (Anderson and Domsch, 1990). Moreover, soil microbial biomassproduces enzymes that either are involved in the cellular metabolismor may be active as abiotic enzymes. Soil enzyme activity is one of thefirst soil properties that are altered when the system is disturbed.Thus, it has long been considered to be an indicator of soil quality be-cause it controls both the supply of nutrients to plants and microbialgrowth (Burns, 1978).
The hypothesis of this study was that changes in soil microbialbiomass and activity should be expected when an abandoned croplandis naturally regenerated over a long time period. Thus, the main objec-tives of this studywere (a) to evaluate the changes in soil microbial bio-mass and activity after regeneration of deforested and cultivated soiland (b) to identify the effect of time on these changes in a Braziliantropical forest.
2. Materials and methods
The study was conducted in the “Usina São José”, which has247 km2, and is located at Igarassu city, Pernambuco State, NortheastBrazil (7°50′20″S, 35°00′10″W). The climate is As’ (Köppen classifica-tion), hot and humid, with a mean precipitation of 2000 mm yr−1
and an annual mean temperature of 30 °C with minimum and
Table 1Soil chemical and physical properties (0–20 cm) in both sampling periods (June andDecember, 2007) in sites under a native forest (NF), a 10 years old regenerated forest(RF10), a 20 years old regenerated forest (RF20) and a conventional sugarcane planta-tion (CL).
Available P was extracted by Mehlich 1. CEC = cation exchange capacity; OM = organicmatter.
258 D.K.A. da Silva et al. / Geoderma 189–190 (2012) 257–261
maximum temperatures of 22 °C and 35 °C, respectively. The driest pe-riod extends from September to December and the rainy season fromJanuary to August (Lima et al., 2008). The soil type is a Haplic Acrisol(FAO-UNESCO) dystrophicwith predominant geomorphological forma-tion from clay–sand sedimentary rocks (Barreiras Group) and depositsfrom the Plio-Pleistocenic age (Trindade et al., 2008).
Soil sampling was carried out in June and December of 2007 (wetand dry seasons, respectively) at four areas: native forest (NF),10 years old regenerated forest (RF10), 20 years old regenerated forest(RF20) and conventional sugarcane plantation (CL). Native vegetationis dominated by Tapirira guianensis Aubl. (Anacardiaceae), Protiumheptaphyllum Marchand (Burseraceae), Clusia nemorosa G. Mey.(Clusiaceae), Margaritaria nobilis L.f. (Euphorbiaceae) and Pouteriapeduncularis (Mart. & Eichler) Baehni (Sapotaceae). The regeneratedforests are populated by Albizia saman (Jacq.) F.Muell. (Fabaceae),Gustavia augusta L. (Lecythidaceae), Xylopia frutescens Aubl.(Annonaceae), Casearia sylvestris Sw. (Flacourtiaceae), Erythroxylumsquamatum Sw. (Erythroxylaceae) and Apeiba tibourbou Aubl.(Tiliaceae). Previously, these regenerated lands were used extensivelyfor sugarcane crop production and after that, these areas were aban-doned and the vegetation grew spontaneously, occurring natural regen-eration. In this region, each sugarcane plantation is changed every fiveyears and it is fertilized with vinasse (300 m3 ha−1y−1). Immediatelybefore harvest, the field is manually burned to ease the harvesting pro-cess. More detailed information about the sites and soil usage historyare available in Trindade et al. (2008) and Kimmel et al. (2008),respectively.
Each area (NF, CL, RF10 and RF20) was composed of four indepen-dent sites. In each independent site, ten subsamples were taken ran-domly, at 0–20 cm depth, and formed a composed sample per site,totaling four composed soil samples per area. The soil samples weretransported to laboratory on ice in a cooler. The field-moist sampleswere stored in sealed plastic bags at 4 °C for microbial analysis.
The remaining soil samples were air-dried. Soil samples wereground to evaluate chemical and physical properties. Soil chemicaland physical analyses (Table 1) were done in the Soil Laboratory ofthe “Instituto Agronômico de Pernambuco” (IPA). Soil pH was deter-mined in a 1:2.5 soil/water extract. Exchangeable Al, Ca and Mg weredetermined by extracting with 1 M KCl. Available P and exchangeableK were extracted by Mehlich-1 and determined by colorimetry andphotometry, respectively (Tedesco et al., 1995).
Microbial biomass C (MBC) was determined according to Vance etal. (1987) with extraction of C from fumigated and unfumigated soilswith 0.5 mol L−1 K2SO4. MBC was measured using dichromate diges-tion and an extraction efficiency coefficient of 0.38 was used to convertthe difference in soluble C between fumigated and unfumigated soil inMBC.
Substrate-induced respiration (SIR), a measure of active microbialbiomass, was measured according to Anderson and Domsch (1978)using a glucose solution. Fluorescein diacetate (FDA) hydrolysis was de-termined according to the method of Swisher and Carroll (1980), anddehydrogenase (DHA) activity was determined using the method de-scribed in Casida et al. (1964), which is based on the spectrophotomet-ric determination of triphenyltetrazolium formazan (TTF) released by5 g of soil during 24 h at 37 °C. Saccharase and cellulase activitieswere estimated according to Schinner and von Mersi (1990).
The data were subjected to analysis of variance (ANOVA) to detectsignificant differences among the areas studied; when a significant pvalue was detected, the means were compared by the least significantdifference (LSD) test (pb0.05). Non-metric multidimensional scaling(MDS) analysis was performed by using the Primer 6.0 software.
3. Results
The results showed that each microbial variable varied significant-ly among different areas depending on the season; typically, larger
differences were observed in the RF10 than in others. The soil MBCwas largely unchanged among the evaluated areas in the wet season,but in the dry season was observed a high amount of soil MBC in RF10than RF20 and CL (Fig. 1a). Likewise, SIR showed no significant differ-ences among sites during the wet season (Fig. 1b). On the other hand,in the dry season, NF showed lower values of SIR than did CL andRF10. The soil analysis revealed high organic matter (OM) contentin RF10 in both sampling periods. In addition, the soil of RF10 pres-ented soil physical properties distinct from the other areas, showingthree times more silt and more clay than the other areas (Table 1).
Enzyme activities showed different behaviors among the areas. Cel-lulase and saccharase activities showed similar patterns to those ob-served for SIR during the wet season (Fig. 2a, b). However, in the dryseason, the highest values for saccharase and cellulase activities wereobserved in the RF10 and NF, respectively. FDA hydrolysis showedhigher values in the RF10 site than in the others for both evaluationsfollowed byRF20 andNF (Fig. 3a),while that the lowest valueswere ob-served in CL. However, dehydrogenase activity (Fig. 3b) presentedhigher values in RF10 and RF20 than in the other sites during the wetseason, whereas in the dry season the values remained similar amongsites.
The MDS analysis, which shows relationships between several soilattributes, indicated that the RF10 has distinct microbial and bio-chemical characteristics comparing with the other sites, which aresimilar among them (Fig. 4).
4. Discussion
As an active component of soil organic C (SOC), microbial biomassis an important attribute of soil quality and also serves as a sensitiveindicator of change and future trends in organic C (Jenkinson et al.,1976; Zhang et al., 2008). Typically, soil microbial biomass content in-creases in soil under systems with high plant diversity such as nativeforest or regenerated lands (Hackl et al., 2004). Soil microbial biomassdepends on organic matter as substrate; therefore, the vegetationcover promotes a high availability of fresh organic matter that stimu-lates the soil microorganisms (Chen et al., 2005).
Fig. 1. Soil microbial biomass (a) and substrate-induced respiration (b) in sites under anative forest (NF), a 10 years old regenerated forest (RF10), a 20 years old regeneratedforest (RF20) and a conventional sugarcane plantation (CL). Bars sharing the samesmall letter are not significantly different between areas and the same capital letterare not significantly different between season within each area (Pb0.05).
Fig. 2. Soil cellulase (a) and saccharase (b) activity in sites under a native forest (NF), a10 years old regenerated forest (RF10), a 20 years old regenerated forest (RF20) and aconventional sugarcane plantation (CL). Bars sharing the same small letter are not sig-nificantly different between areas and the same capital letter are not significantly dif-ferent between season within each area (Pb0.05).
259D.K.A. da Silva et al. / Geoderma 189–190 (2012) 257–261
In the regenerated lands, the higher amount of soilMBC inRF10 thanin other areas, in dry period, may be related with higher clay and OMcontent found in this area (Table 1). It suggests that soil clay can protectmicrobial biomass, in dry period, as reported by Muller and Hoper(2004). The main mechanism is the physical protection of soil organicmatter and microbial biomass by clays (van Veen et al., 1984). In addi-tion, high OMcontent promotes an increase in soil microbial biomass asa response to more available C sources (Santos et al., 2012).
In soil under cropland, the soil microbial biomass content variesaccording to soil management. For sugarcane production, the applica-tion of vinasse promotes the addition of a high C content organic source.Although agriculture can significantly decrease soil microbial biomass(Islam and Weil, 2000), our data suggest that the agricultural practicesutilized in sugarcane plantation, such as use of vinasse, maintained themicrobial biomass content.
For regenerated lands, our results are similar to those found forother tropical sites where accumulation of plant litter and soil nutrientspromoted an increase in soil microbial status (Groffman et al., 2001;Templer et al., 2005). In the wet season, SIR method showed similarvalues for soil microbial biomass in all areas. Although SIR may accu-rately detect changes in land use (Anderson and Domsch, 1990) ourdata showed that the microbial community did not respond to glucoseadditions. The SIR technique is sufficiently sensitive to discriminatechanges in catabolic diversity (Nsabimana et al., 2004) in a short timefollowing addition of organic substrates (Degens, 1998). Probably, inwet season, the soil moisture stimulated the soil microbial biomassand the additional source of C, added in the SIR method, did not pro-mote an increase in soil microbial activity. However, SIR was sensitiveto show differences between the areas with high SIR in RF10 and CL.
Soil enzymes are responsible for controlling different reactions andmetabolic processes in the biogeochemical cycling of nutrients, andthese enzymatic activities provide insight into the natural and anthropo-genic disturbances occurring in the soil ecosystem (Trevors, 1984). The
analysis of cellulase and saccharase activities is highly recommendedfor evaluation of forest soils (Schinner and von Mersi, 1990). Ourdata on activity of these enzymes demonstrated differences betweenareas, as well as between seasons. In fact, in some cases the activity ofenzymes in the natural area differed from the other areas. In the dryseason, the increase in cellulase activity under natural forest and sac-charase activity under regenerated forest probably occurred due tothe high deposition of fresh organic matter.
The hydrolysis of FDA may broadly represent soil microbial activity(Schnurer and Rosswall, 1982). The trends observed for this enzyme ac-tivity in response to deforestation and changes in land use are in agree-ment with other studies showing lower values in cultivated soils whencompared to the corresponding undisturbed or regenerated soils(Acosta-Martínez et al., 2007). In both seasons, the regenerating forest(RF10) presented high microbial activity of this enzyme, which isrelated to the decomposition of organic substrates, indicating that inthis area there is a trend to increase soil fertility in the long term(Caravaca et al., 1964). The high values of organic matter in RF10 con-firm that observation. Furthermore, areas in the regeneration processare characterized by intense growth of vegetation that provides organicmatter readily available to the microbiota (Groffman et al., 2001). Thesoil dehydrogenase activity reflects soil oxidative power and providesa way to quantify the total number of viable microbial cells (Gianfredaet al., 2005). In thewet season,we observed high activity of this enzymein RF20 than in NF and CL, indicating that regenerated forests presentmore metabolically active microorganisms. Our result is in agreementwith the findings of Nogueira et al. (2006) who observed higher dehy-drogenase activity in regeneration forest than in native forest andcropland.
Analyses of different variables, taken together, can provide a betterpicture of the status of soil processes and functions (Acosta-Martínezet al., 2007). The difference found in the MDS analyses are related to
Fig. 3. Hydrolysis of fluorescein diacetate (a) and dehydrogenase (b) activity in sitesunder a native forest (NF), a 10 years old regenerated forest (RF10), a 20 years oldregenerated forest (RF20) and a conventional sugarcane plantation (CL). Bars sharingthe same small letter are not significantly different between areas and the same capitalletter are not significantly different between season within each area (Pb0.05).
260 D.K.A. da Silva et al. / Geoderma 189–190 (2012) 257–261
the regeneration time, as shown by the area in the early stage of regen-eration (RF10) where the dynamic balance of soil differed from theother sites. On the other hand, RF 20 presented characteristics similarto NF, indicating that, after 20 years of regeneration, the soil is recover-ing its stability. The similarity betweenNF and CL is probably “artificial”,considering that both aremore stable. Edaphic similarities between cul-tivated areas and preserved Atlantic Forest were also referred by Freitaset al. (2008). These results revealed that natural and regenerated foreststended to have an increase in soil microbial biomass and activity. Thesefindings show the importance of soil conservation, including the sus-tainable use of forest to improve soil microbial status, as also reportedby Martens et al. (2004).
Fig. 4. Non-metric multidimensional scaling (MDS), regardless of the sampling period,based on data from dehydrogenase activity, hydrolysis of fluorescein diacetate, saccha-rase activity, cellulase activity, substrate induced respiration and microbial biomass insites under a native forest (NF), a 10 years old regenerated forest (RF10), a 20 years oldregenerated forest (RF20) and a conventional sugarcane plantation (CL).
5. Conclusions
These results indicate that soil microbial variables varied in differentseasons. Usually, in wet season there is no variation in soil microbial bio-mass among intact forest, regenerated forest and active agricultural sitesin tropical soil. While that, in dry season, these microbial variables variedbetween sites. In general form,microbial activity, asmeasured by enzymeactivities, is higher in native forest and regenerated lands than incropland. We observed that the areas in the regeneration processare recovering the soil quality, with tendency to stability. Due to sim-ilarities observed between the sites with more time of regenerationand native forest, we can assume that new conditions of balanceare being achieved in this regeneration area.
Acknowledgments
The authors acknowledge Professor Ana Carolina Lins e Silva, Thom-as Kimmel and Ladivânia Nascimento who, respectively provided infor-mation regarding the plant species and the areas and helped with thesoil collections. This work was supported by the “Coordenação deAperfeiçoamento de Pessoal de Nível Superior (CAPES)” and “ConselhoNacional de Desenvolvimento Científico e Tecnológico (CNPq)”, whichprovided aMaster scholarship to Danielle K. A. Silva, a Ph.D. scholarshipto Nicacio Freitas and research grants to Ademir S. F. Araújo and LeonorC.Maia. This researchwas a contribution to the project “Sustainability ofremnants of the Atlantic rainforest in Pernambuco and its implicationsfor conservation local development”, a Brazilian–German Scientific Col-laborationwithin the program “Science and Technology for the AtlanticRainforest” that is funded by CNPq (590039/2006-7) and BMBF (01 LB0203 A1) and permitted and logistically supported by Usina São JoséS.A/Grupo Cavalcanti Petribú.
References
Acosta-Martínez, V., Cruz, L., Sotomayor-Ramírez, D., Pérez-Alegría, L., 2007. Enzymeactivities as affected by soil properties and land use in a tropical watershed. Ap-plied Soil Ecology 35, 35–45.
Anderson, J.P.E., Domsch, K.H., 1978. A physiological method for the quantitative mea-surement of microbial biomass in soils. Soil Biology and Biochemistry 10, 215–221(Applied Soil Ecology 33, 30–38.).
Anderson, J.M., Domsch, K.H., 1990. Application of eco-physiological quotients (qCOand qD) on microbial biomass from soils of different cropping histories. Soil Biolo-gy and Biochemistry 22, 251–255.
Araújo, A.S.F., Silva, E.F.L., Nunes, L.A.P.L., Carneiro, R.F.V., 2010. The effect of convertingtropical native savanna to Eucalyptus grandis forest on soil microbial biomass. LandDegradation and Development 21, 540–545.
Burns, R.G., 1978. Soil Enzymes. Academic Press, New York.Caravaca, F., Alguail, A., Azcón, R., Roldán, A., 1964. Formation of stable aggregates in
rizosphere soil of Juniperus oxycedrus: effect of AM fungi and organic amendments.In: Casida, L.E., Klein, D.A., Santoro, T. (Eds.), Soil dehydrogenase. Soil Science, 98,pp. 371–376.
Chander, K., Brookes, P.C., 1993. Residual effects of zinc, copper and nickel in sewagesludge on microbial biomass in a sandy loam. Soil Biology and Biochemistry 25,1231–1239.
Degens, B.P., 1998. Microbial functional diversity can be influenced by the addition ofsimple organic substrates to soil. Soil Biology and Biochemistry 30, 1981–1988.
Freitas, N.O., Silva, F.S.B., Maia, L.C., 2008. Edge effect on soil biochemical and microbi-ological activities in an Atlantic forest fragment in the state of Pernambuco, Brazil.Bioremediation, Biodiversity and Bioavailability 2, 62–67.
Gianfreda, L., Rao, M.A., Piotrowska, A., Palumbo, G., Colombo, C., 2005. Soil enzyme ac-tivities as affected by anthropogenic alterations: intensive agricultural practicesand organic pollution. Science of the Total Environment 341, 265–279.
Hackl, E., Bachmann, G., Zechmeister-Boltenstern, S., 2004. Microbial nitrogen turnoverin soils under different types of natural forest. Forest Ecology and Management188, 101–112.
Houghton, R., 1990. The global effects of tropical deforestation. Environmental Scienceand Technology 24, 414–422.
Islam, K.R., Weil, R.R., 2000. Land use effects on soil quality in a tropical forest ecosys-tem of Bangladesh. Agriculture Ecosystem and Environment 79, 9–16.
261D.K.A. da Silva et al. / Geoderma 189–190 (2012) 257–261
Jenkinson, D.S., Ladd, J.N., 1981. Microbial biomass in soil: measurement and turn-over. In: Paul, E.A., Ladd, J.N. (Eds.), Soil biochemistry. Marcel Dekker, NewYork, pp. 415–471 (Org.).
Jenkinson, D.S., Powlson, D.S., Wedderburn, R.W.M., 1976. The effects of biocidal treat-ments on metabolism in soil—III. The relationship between soil biovolume, mea-sured by optical microscopy, and the flush of decomposition caused byfumigation. Soil Biology and Biochemistry 8, 189–202.
Kennedy, A., Doran, J., 2002. Sustainable agriculture: role of microorganisms. In: Bitton,G. (Ed.), Encyclopedia of Environmental Microbiology. John Wiley & Sons, NewYork, pp. 3116–3126 (Org.).
Kimmel, T., Piechowski, D., Gottsberger, G., 2008. The history of fragmentation of thelowland Atlantic Forest of Pernambuco, Brazil. Bioremediation, Biodiversity Bio-availability 2, 1–4.
Lima, A.L.A., Rodal, M.J.N., Lins-e-Silva, A.C.B., 2008. Phenology of tree species in a frag-ment of Atlantic Forest in Pernambuco—Brazil. Bioremediation Biodiversity Bio-availability 2, 68–75.
Martens, D.A., Reedy, E.T., Lewis, D.T., 2004. Soil organic carbon content and composi-tion of 130-year crop, pasture, and forest land-use managements. Global ChangeBiology 10, 65–78.
Muller, T., Hoper, H., 2004. Soil organic matter turnover as a function of the soil claycontent: consequences for model applications. Soil Biology and Biochemistry 36,877–888.
Munier-Lamy, C., Border, O., 2000. Effect of triazolle fungicide on the cellulose decom-position by the soil microflora. Chemosphere 41, 1029–1035.
Nogueira, M.A., Albino, U.B., Brandão-Júnior, O., Braun, G., Cruz, M.F., Dias, B.A., Duarte,R.T.D., Gioppo, N.M.R., Menna, O., Orlandi, J.M., Raimam, M.P., Rampazo, L.G.L.,Santos, M.A., Silva, M.E.Z., Vieira, F.P., Torezan, J.M.D., Hungria, M., Andrade, G.,2006. Promising indicators for assessment of agroecosystems alteration amongnatural reforested and agricultural land use in southern Brazil. Agriculture Ecosys-tem and Environment 115, 237–247.
Nsabimana, D., Haynes, R.J., Wallis, F.M., 2004. Size, activity and catabolic diversity ofthe soil microbial biomass as affected by land use. Applied Soil Ecology 26, 81–92.
Post, W.M., Kwon, K.C., 2000. Soil carbon sequestration and land-use change: processesand potential. Global Change Biology 6, 317–328.
Ribeiro, M.C., Metzger, J.P., Martensen, A.C., Ponzoni, F.J., Hirota, M.M., 2009. The BrazilianAtlantic Forest: howmuch is left, and how is the remaining forest distributed? Impli-cations for conservation. Biological Conservation 142, 1141–1153.
Santos, V.B., Araújo, A.S.F., Leite, L.F.C., Nunes, L.A.P.L., Melo, W.J., 2012. Soil microbialbiomass and organic matter fractions during transition from conventional to or-ganic farming systems. Geoderma 170, 227–231.
Schinner, F., von Mersi, W., 1990. Xylanase, CM-cellulase and invertase activity in soil:an improved method. Soil Biology and Biochemistry 22, 511–515.
Schnurer, J., Rosswall, T., 1982. Fluorescein diacetate hydrolysis as a measure of totalmicrobial activity in soil and litter. Applied and Environmental Microbiology 43,1256–1261.
Swisher, R., Carroll, G.C., 1980. Fluorescein diacetate hydrolysis as an estimator of mi-crobial biomass on coniferous needle surfaces. Microbial Ecology 6, 217–226.
Tabarelli, M., Pinto, L.P., Silva, J.M., Hirota, M., Bedê, A.L., 2005. Challenges and opportu-nities for biodiversity conservation in the Brazilian Atlantic Forest. ConservationBiology 19, 695–700.
Tedesco, M.J., Gianello, C., Bissani, C.A., 1995. Análises de solos, plantas e outrosmateriais. UFRGS, Porto Alegre. 1995.
Templer, P.H., Groffman, P.M., Flecker, A.S., Power, A.G., 2005. Land use change and soilnutrient transformations n the Los Haities region of the República Dominicana. SoilBiology and Biochemistry 37, 217–225.
Trevors, J.T., 1984. Effect of substrate concentration, inorganic nitrogen, O2 concentration,temperature and pH on dehydrogenase activity in soil. Plant and Soil 77, 285–293.
Trindade, M.B., Lins-e-Silva, A.C.B., Silva, H.P., Figueira, S.B., Schessl, M., 2008. Fragmen-tation of the Atlantic Rainforest in the northern coastal region of Pernambuco,Brazil: recent changes and implications for conservation. Bioremediation, Biodiversityand Bioavailability 2, 5–13.
van Veen, J.A., Ladd, J.N., Frissel, M.J., 1984. Modelling C and N turnover through the mi-crobial biomass in soil. Plant and Soil 76, 257–274.
Vance, E.D., Brookes, P.C., Jenkinson, D.S., 1987. An extraction method for measuringsoil microbial biomass C. Soil Biology and Biochemistry 19, 703–707.
Zhang, N., Wan, S., Li, L., Bi, J., Zhao, M., Ma, K., 2008. Impacts of urea N addition on soilmicrobial community in a semi-arid temperate steppe in northern China. Plant andSoil 311, 19–28.
