Fungi constitute an important part of the soil ecosystem, playing key roles in decomposition, cycling processes, and biotic interactions. Molecular methods have been used to assess fungal communities giving a more realistic view of their diversity. For this purpose, total DNA was extracted from bulk soils cultivated with tomato (STC), vegetables (SHC), and native forest (SMS) from three sites of the Taquara Branca river basin in Sumaré County, S？o Paulo State, Brazil. This metagenomic DNA was used as a template to amplify fungal 18S rDNA sequences, and libraries were constructed in Escherichia coli by cloning PCR products. The plasmid inserts were sequenced and compared to known rDNA sequences in the GenBank database. Of the sequenced clones, 22 were obtained from the SMS sample, 18 from the SHC sample, and 6 from the STC sample. Although most of the clone sequences did not match the sequences present in the database, individual amplified sequences matched with Glomeromycota (SMS), Fungi incertae sedis (SMS), and Neocallimastigomycota (SHC). Most of the sequences from the amplified taxa represent uncultured fungi. The molecular analysis of variance (AMOVA) indicated that fluctuations observed of haplotypes in the composition may be related to herbicide application. 1. Introduction Despite the importance of soil microbial communities in regulating soil ecosystem-level processes, such as the nutrient cycle and organic matter decomposition, little is known about the structure of these microbial communities and the factors that influence it in soils. This lack of knowledge arises, in part, from the enormous complexity of soil microbial communities, which are estimated to contain more than 4,000 different genomic equivalents in a single gram of soil [1]. Because of their broad ecological range, ready adaptation abilities, and wide spectrum of nutrient sources, filamentous and yeast-like fungi are able to colonize many different niches or substrates [2]. As integral components in the soil ecosystem, fungi play an important role as major decomposers of plant residues, releasing nutrients that sustain and stimulate plant growth [3]. In spite of their importance, there are very few reports on the fungal communities in soil. Comparative studies have reported that microbial communities can change in response to soil disturbances, and differences have been observed between microbial communities in fields with different histories of soil amendment, irrigation, tillage, and plant community structure [4]. Knowledge of soil microorganisms is expanding with the advent
References
[1]
D. H. Buckley and T. M. Schmidt, “The structure of microbial communities in soil and the lasting impact of cultivation,” Microbial Ecology, vol. 42, no. 1, pp. 11–21, 2001.
[2]
J. I. Mitchell and A. Zuccaro, “Sequences, the environment and fungi,” Mycologist, vol. 20, no. 2, pp. 62–74, 2006.
[3]
K. Ritz and I. M. Young, “Interactions between soil structure and fungi,” Mycologist, vol. 18, no. 2, pp. 52–59, 2004.
[4]
M. A. Liebig, D. L. Tanaka, and B. J. Wienhold, “Tillage and cropping effects on soil quality indicators in the Northern Great Plains,” Soil and Tillage Research, vol. 78, no. 2, pp. 131–141, 2004.
[5]
V. Torsvik and L. ？vre？s, “Microbial diversity and function in soil: from genes to ecosystems,” Current Opinion in Microbiology, vol. 5, no. 3, pp. 240–245, 2002.
[6]
R. M. Pereira, E. L. da Silveira, D. C. Scaquitto et al., “Molecular characterization of bacterial populations of different soils,” Brazilian Journal of Microbiology, vol. 37, no. 4, pp. 439–447, 2006.
[7]
E. L. da Silveira, R. M. Pereira, D. C. Scaquitto et al., “Bacterial diversity of soil under eucalyptus assessed by 16S rDNA sequencing analysis,” Pesquisa Agropecuária Brasileira, vol. 41, no. 10, pp. 1507–1516, 2006.
[8]
S. P. Val-Moraes, M. J. Valarini, R. Ghini, E. G. M. Lemos, and L. M. Carareto-Alves, “Diversidade de bactérias de solo sob vegeta？？o natural e cultivo de hortali？as,” Revista Ciência Agron？mica, vol. 40, no. 1, pp. 7–16, 2009.
[9]
E. A. N. Pedrinho, E. G. M. Lemos, R. M. Pereira et al., “Avalia？？o do impacto do lodo de esgoto na microbiota do solo utilizando o gene 16s rRNA,” Arquivos do Instituto Biológico, S？o Paulo, vol. 76, no. 3, pp. 443–448, 2009.
[10]
R. Ghini and M. M. H. Zaroni, “Rela？？o entre coberturas vegetais e supressividade de solos a Rhizoctonia solani,” Fitopatologia Brasileira, vol. 26, pp. 10–15, 2001.
[11]
B. van Raij, J. C. de Andrade, H. Cantarella, and J. A. Quaggio, Análise Química Para Avalia？？o da fertilidade de Solos Tropicais, Instituto Agron？mico, Campinas, Brasil, 2001.
[12]
J. P. Martin, “Use of acid, rose Bengal and streptomycin in the plate method for estimating soil fungi,” Soil Science, vol. 69, pp. 215–232, 1950.
[13]
E. Smit, P. Leeflang, B. Glandorf, J. D. Van Elsas, and K. Wernars, “Analysis of fungal diversity in the wheat rhizosphere by sequencing of cloned PCR-amplified genes encoding 18S rRNA and temperature gradient gel electrophoresis,” Applied and Environmental Microbiology, vol. 65, no. 6, pp. 2614–2621, 1999.
[14]
J. Sambrook, E. F. Fritsch, and T. Maniatis, “Gel electrophoresis of DNA,” in Molecular Cloning Laboratory Manual, C. Nolan, Ed., vol. 1, pp. 6.1–6.62, Cold Spring Harbor Laboratory Press, New York, NY, USA, 1989.
[15]
B. Ewing, L. Hillier, M. C. Wendl, and P. Green, “Base-calling of automated sequencer traces using phred. I. accuracy assessment,” Genome Research, vol. 8, no. 3, pp. 175–185, 1998.
[16]
S. F. Altschul, T. L. Madden, A. A. Sch？ffer et al., “Gapped BLAST and PSI-BLAST: a new generation of protein database search programs,” Nucleic Acids Research, vol. 25, no. 17, pp. 3389–3402, 1997.
[17]
J. D. Thompson, D. G. Higgins, and T. J. Gibson, “Clustal W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position specific gap penalties and weight matrix choice,” Nucleic Acids Research, vol. 22, no. 22, pp. 4673–4680, 1994.
[18]
P. Hall, BioEdit—Version 5.0.6. Raleigh, North Carolina State University, Department of Microbiology, 2001.
[19]
K. Tamura, D. Peterson, N. Peterson, G. Stecher, M. Nei, and S. Kumar, “MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods,” Molecular Biology and Evolution, vol. 28, no. 10, pp. 2731–2739, 2011.
[20]
D. Swofford, G. Olsen, P. Waddel, and M. Hillis, “Phylogenetic inference,” in Molecular Systematics, D. M. Hills, C. Mortiz, and B. K. Mable, Eds., pp. 407–514, Sinauer Association, Sunderland, Mass, USA, 1996.
[21]
M. Nei, “Genetic distance between populations,” American Naturalist, vol. 106, no. 949, pp. 283–292, 1972.
[22]
M. Nei and S. Kumar, Molecular Evolution and Phylogenetics, Oxford University Press, Oxford, UK, 2000.
[23]
S. Schneider, D. Roeslli, and L. Excoffier, ARLEQUIN: Software for Population Genetics Data Analysis. Version 2.000, University of Geneva, Geneva, Switzerland, 2000.
[24]
A. P. Martin, “Phylogenetic approaches for describing and comparing the diversity of microbial communities,” Applied and Environmental Microbiology, vol. 68, no. 8, pp. 3673–3682, 2002.
[25]
M. C. Jahnel, E. J. B. N. Cardoso, and C. T. S. Dias, “Determina？？o do número mais provável de microrganismos do solo pelo método de plaqueamento por gotas,” Revista Brasileira de Ciência do Solo, vol. 23, no. 3, pp. 553–559, 1999.
[26]
T. H. Jukes and C. R. Cantor, “Evolution of protein molecules,” in Mammalian Protein Metabolism, H. N. Munro, Ed., pp. 21–132, Academic Press, New York, NY, USA, 1969.
[27]
J. Felsenstein, “Confidence limits on phylogenies: an approach using the bootstrap,” Evolution, vol. 39, pp. 783–791, 1985.
[28]
M. K？rschens, “Soil organic matter and environmental protection,” Archives of Agronomy and Soil Science, vol. 50, pp. 3–9, 2004.
[29]
A. D. Halvorson, B. J. Wienhold, and A. L. Black, “Tillage, nitrogen, and cropping system effects on soil carbon sequestration,” Soil Science Society of America Journal, vol. 66, no. 3, pp. 906–912, 2002.
[30]
D. W. Reeves, “The role of soil organic matter in maintaining soil quality in continuous cropping systems,” Soil and Tillage Research, vol. 43, no. 1-2, pp. 131–167, 1997.
[31]
K. R. Olson, S. A. Ebelhar, and J. M. Lang, “Effects of 24 years of conservation tillage systems on soil organic carbon and soil productivity,” Applied and Environmental Soil Science, vol. 2013, Article ID 617504, 10 pages, 2013.
[32]
F. A. A. M. de Ley and J. M. Lynch, “Functional diversity of the rhizosphere,” in Proceedings of the 4th International Workshop on Plant-Growth Promoting Rhizobacteria, A. Ogoshi, K. Kobayashi, Y. Homma, F. Kadoma, N. Kondo, and S. Akico, Eds., pp. 38–43, Organisation for Economic Co-operation and Development, Paris, France, 1997.
[33]
K. Hedlund, “Soil microbial community structure in relation to vegetation management on former agricultural land,” Soil Biology and Biochemistry, vol. 34, no. 9, pp. 1299–1307, 2002.
[34]
S. J. Grayston, G. S. Griffith, J. L. Mawdsley, C. D. Campbell, and R. D. Bardgett, “Accounting for variability in soil microbial communities of temperate upland grassland ecosystems,” Soil Biology and Biochemistry, vol. 33, no. 4-5, pp. 533–551, 2001.
[35]
J. S. States, “Useful criteria in the description of fungal communities,” in The Fungal Community, D. T. Wicklow and G. C. Carroll, Eds., pp. 185–200, Marcel Dekker, New York, NY, USA, 1981.
[36]
L. M. Zavatti and R. B. Abakerli, “Resíduos de agrotóxicos em frutos,” Pesquisa Agropecuária Brasileira, vol. 34, no. 3, pp. 473–480, 1999.
[37]
A. Nesci, G. Barros, C. Castillo, and M. Etcheverry, “Soil fungal population in preharvest maize ecosystem in different tillage practices in Argentina,” Soil and Tillage Research, vol. 91, no. 1-2, pp. 143–149, 2006.
[38]
H. Swer, M. S. Dkhar, and H. Kayang, “Fungal population and diversity in organically amended agricultural soils of Meghalaya, India,” Journal of Organic Systems, vol. 6, no. 2, 2011.
[39]
D. A. Wardle, “A comparative assessment of factors which influence microbial biomass carbon and nitrogen levels in soil,” Biological Reviews of the Cambridge Philosophical Society, vol. 67, no. 3, pp. 321–358, 1992.
[40]
G. D. Bending, M. K. Turner, and J. E. Jones, “Interactions between crop residue and soil organic matter quality and the functional diversity of soil microbial communities,” Soil Biology and Biochemistry, vol. 34, no. 8, pp. 1073–1082, 2002.
[41]
R. G. Burns, “Enzyme activity in soil: location and a possible role in microbial ecology,” Soil Biology and Biochemistry, vol. 14, no. 5, pp. 423–427, 1982.
[42]
J. Borneman, P. W. Skroch, K. M. O'Sullivan et al., “Molecular microbial diversity of an agricultural soil in Wisconsin,” Applied and Environmental Microbiology, vol. 62, no. 6, pp. 1935–1943, 1996.
