The Role of Stabilised Enzymes in Soil Microbial Ecology
According to Burns (1982) enzymes entrapped by humic colloids are essential for the successful competition of microbial cells in the hostile soil extracellular environment. Particularly the stabilised enzymes can catalyse the transformation of high molecular substrates into products that can be taken up by near microbial cells and can act as inducers or chemoattrac-tants. In the first case the products induce the microbial cell to synthesise and release the specific extracellular enzyme into...
Soil Proteomics
It is now recognised that also in soil microbiology there is a need to go beyond DNA analysis for a better understanding of soil functionality (Nan-nipieri et al. 2003). By characterising both DNA and RNA one might be able to determine the actual species effectively involved in the processes being measured. Different protocols involving the simultaneous extraction of RNA and DNA from soil have recentlybeen published (Duarte et al. 1998 Griffiths et al. 2000 Hurt et al. 2001 Weinbauer et al....
Degradation Kinetics of Introduced DNA in Soil
Two different approaches were used introduction of purified DNA into soil and introduction of cells or tissue material containing the DNA to be traced. Introduced recombinant plasmid DNA was detectable in different non-sterile soils for up to 60 days (end of experiment) by dot blot and Southern hybridisation and transformation of CaCl2-treated E. coli (Romanowski et al. 1992) and by quantitative PCR and electroporation of E. coli (Romanowski et al. 1993b). As an example, in loamy sand soil...
Enzymes as Decontaminating Agents
Transformation reactions of phenols and chlorophenols by laccases and peroxidases have widely been investigated under laboratory conditions. Their oxidative self-coupling (Dec and Bollag 2000 Duran and Esposito 2000 Bollag et al. 2003 Gianfreda et al. 2003) and cross-coupling reactions with humic precursors (Filazzola et al. 1999 Park et al. 1999 Itoh et al. 2000), humic substances themselves (Huang et al. 2002 Park et al. 2000), and soil-derived dissolved organic matter (Huang and Weber 2004)...
Effect of Stresses Salinity
The potential adverse effects of the accumulation in soil of soluble salts on the activities of soil enzymes were studied in laboratory experiments (Frankenberger and Bingham 1982) also involving gypsiferous soils (Garc a and Hern ndez 1997). Many enzyme activities were reduced with increasing electrical conductivity of saturated extracts and the degree of inhibition varied among the enzymes assayed and the nature and amounts of salts added. More recently, a different approach has been adopted,...
Relationship Between Enzyme Activities and Soil Physical Properties and Soil Depth
Soil enzyme activity is generally related to soil microbial population size and it is positively influenced by the amount of pores ranging from 30-200 m (Pagliai and De Nobili 1993). Marinari et al. (2000) showed that the addition of organic fertilisers improved soil physical and microbiological properties. They reported a significant linear correlation between acid phosphatase and dehydrogenase activities and soil total porosity, whereas benzoylargininamide hydrolysing activity was not...
Factors Affecting Soil Enzyme Activities
Due to the complexity of the soil environment it seems evident that several factors will affect enzyme activities in soil. As discussed by Gianfreda and Bollag (1996) and Dick and Tabatabai (1999), the activity of any enzyme in soil is the result of synthesis, persistence, stabilisation, regulation, and catalytic behaviour of the enzymatic proteins, present in the soil environment at the moment of the assay (Fig. 12.1). All these processes maybe dynamically interrelated and influenced by...
References
Aardema BW, Lorenz MG, Krumbein WE (1983) Protection of sediment-adsorbed transforming DNA against enzymatic inactivation. Appl Environ Microbiol 46 417-420 Alvarez AJ, KhannaM, Toranzos GA, Stotzky G (1998) Amplification of DNA bound on clay minerals. Mol Ecol 7 775-778 Agnelli A, Ascher J, Corti G, Ceccherini MT, Nannipieri P, Pietramellara G (2004) Microbial biomass, soil respiration and DGGE analysis of total and extracellular DNA in a forest soil profile. Soil Biol Biochem 36 859-868...
Cell Rupture Depends on Cell Type and Growth
Physical means to rupture cells are generally more efficient on Gramnegative than Gram-positive bacteria, due to the thicker peptidoglycan layer of the latter. The strength of the peptidoglycan layer is not only dependent on its thickness, but also on the degree of the peptide cross-links (Pisabaro et al. 1985). The number of cross-links can vary between species, but for each species it also depends on growth rate. Growth will necessarily imply that the wall must be weakened, to allow it to...
RRNA Approaches Focused on Specific Phylogenetic Groups
The presence of SSU rDNA domains with varying levels of conservation across taxa facilitates the design of primers and probes that target specific microbial groups at various taxonomic levels. Thus, conserved regions may be used for the design of high level taxon primers (see above Table 8.1), intermediately variable regions may be applied to analyses at the order through class level, and highly variable regions are suited for detection of specific genera and species (e.g. Heuer et al. 1999)....
Release of DNA from Organisms
The organisms living in soil encompass mainly prokaryotes, fungi, and various animals. Plants can grow their roots in soil and often also rhizomes and parts of the shoot. In order to obtain clues as to the presence of extracellular DNA in the environment including soil, the release of DNA from members of these groups has been examined. The experiments were mainly performed in the laboratory and only in few cases in natural field soils or in soil microcosms. The soil bacterium Bacillus subtilis...
Phylogenetic Oligonucleotide Arrays POAs
POAs primarily contain probes sequences derived from rRNA genes however, efforts have been made to incorporate other markers such as gyrB (Fukushima et al. 2003 Kakinuma et al. 2003) or rpoB (Dahll f et al. 2000). The probes incorporated into POAs may be designed on a species-specific basis, a hierarchical manner, or some combination thereof, such that several different taxonomic or phylogenetic levels of information are targeted. While species-specific probes can be easily generated on a...
Extraction of Enzymes from Soil
A cathepsin-like activity was measured in a soil extract in 1954 (Antoniani et al. 1954). However, according to Skujins (1967), Fermi and Sabrama-nian were the first to measure enzyme activities (protease and aminase activities, respectively) in a soil extract. Urease was isolated from a surface forest soil with phosphate buffer at pH 6 and purified in the United States by Briggs and Segal (1963). In Australia, Ladd (1972) determined protease and peptidase activities in several pasture and...
Soil Proteins as Metal Biosensors
It has been estimated that at least 30 of all proteins are metalloproteins, requiring one or more metal ions for proper functioning (Andreini et al. 2004 for examples, see Table 5.1). The specificity of protein-metal complexes and the affinity (and stoichiometric interaction) of proteins for metals in general provide an opportunity to detect or to determine biological availability of metal ions by assaying protein functions including enzymatic activity (Haraguchi 2004). Theoretically, two major...
Distribution of Enzyme Activities with Soil Particles
Enzyme activities have been measured in soil particles to better understand the relationships between biotic and abiotic constituents with enzyme activity of soil, by using a preservative fractionation method by Stemmer et al. (1998), which is quite soft to allow complete recovery of soil enzyme activities (Kandeler et al. 1999a,b,d Gerzabek et al. 2002 Poll et al. 2003). In the case ofxylanase the sum of the enzyme activities of all isolated fractions considerably surpassed the activity of the...
Framework for the Structure Function Grand Challenge
The great challenge in microbial ecology of the inaugural decade of the twenty-first century is to resolve and then understand the linkage between microbial community structure and its function. This goal is based on the fact that the primary value of microbes is their function and the belief that understanding the composition, relative abundance, biochemical diversity and dynamics of the microbial community will help us understand and eventually predict or manage function in microbial...
Biodegradation of Soil Polymers and C1 Compounds
The most ubiquitous polymers of natural origin found in soil are cellulose and chitin. Cellulolytic activity is critical for carbon recycling and chitin plays a unique role as the most abundant source of nitrogen in soil. Chitinolytic ability is ubiquitous among soil microorganisms (and also numerous higher organisms) and chitin is an important source of N in most environments. Many bacteria possess multiple chitinase genes and fungal chitinases may have dual roles in nutrition and hyphal cell...
Methodology for Soil Proteomics
Methods for extracting protein molecules from soils are entirely independent of the subsequent methods of protein analysis. If the analytical methods involve assessment of protein function, including enzyme activity, then the extraction methods must preserve function as much as possible. For example, the availability of colorimetric staining techniques for more than 300 enzymes (Manchenko 1994) coupled with nitrocellulose membrane-immobilisation of protein molecules extracted from environmental...
Molecular Detection of Functional Gene Signatures for Detecting Pathogens in Soil
Detection of pathogens and assessment of their activity in the environment is important for ecological studies as well as for public health. A variety of microbial pathogens that infect humans and animals are known to survive in soil, e.g. enteric pathogens such as Salmonella sp., Vibrio cholerae, Shigella sp., Campylobacter jejuni, Yersinia sp., Escherichia coli O157 h7 (Santamaria and Toranzos 2003) and other pathogens such as Mycobacterium bovis (Young et al. 2005). Pathogens enter the soil...
Tillage Cropping and Other Managements
Soil management influences microbial activity, microbial biomass and enzyme activity through changes in nutrient inputs and in the quantity and quality of plant residues entering the soil (Burns 1978a Ekenler and Tabatabai 2002a). Different tillage significantly alters soil properties and especially the spatial distribution within the soil profile of the various enzyme activities (Deng and Tabatabai 1996a Bergstrom et al. 1998a,b). Usually, enzyme activities of the surface layer of no-till...
Organic Amendments
In general, recent additions of organic residues can stimulate enzyme activities as the result of microbial proliferation or enzyme induction in response to the amendment, if inhibitory substances are not present in organic residues such as heavy metals in municipal solid waste (MSW) compost. Indeed the ratios between acid phosphatase, alkaline phosphatase, a-glucosidase, arylsulfatase, or urease activity and microbial biomass were significantly increased by manure regardless of crop type in...
Measurement of Soil Enzyme Activities
Soil enzyme activities have been often proposed as sensitive indicators of soil quality. As underlined by Schloter et al. (2003), the ideal soil microbio-logicalandbiochemicalindicator todeter minesoil quality wouldbesimple to measure, should work equally well in all environments and reliably reveal which problems existed where. The authors considered enzymatic assays very helpful methodologies because they are short-term laboratory procedures, usually performed under standardised environmental...
Bead Beating Efficiency and Bias
Bead beating has been a popular component of cell lysis protocols. A soil slurry (or a suspension of extracted cells) is mixed with small spheres (most commonly 0.1 or 0.2 mm spheres, sometimes mixed with larger beads) and shaken at high frequency. Bead beating exploits the collision between glass spheres to break the cells thus deformation is probably the main cause of cell disruption, although viscous shear may hypothetically be important at high speeds. Recognised critical physical...
The Potential Role of Rhizosphere Enzyme Activities
There are several concerns about the possibility of horizontal gene transfer between genetically modified plants and the soil microbial community. The hypothetical acquisition of these genes by the bacterial community raises great concern about the undesirable effects of altering the resistance level of indigenous bacteria and of disseminating such traits into the environment. For example, Acinetobacter spp., the phytopathogen Ralstonia solanacearum, and the bacterial pathogen Erwinia...
Detection of Antibiotic and Heavy Metal Resistance Genes
There have been relatively few studies focusing on detection of antibiotic or heavy metal resistance genes in situ, in environments such as soil. Most research has concentrated on detection of resistance genes in clinical and environmental isolates. Resistance genes have been detected in soil using conventional DNA extraction and PCR methods. Chee-Sanford et al. 2001 screened for eight tet genes in groundwater associated with swine production facilities. Tetracycline resistance genes were found...
The rRNA Approach
Ribosomal RNA genes, especially the small subunit SSU ribosomal RNA genes, have become the targets of choice for microbial ecologists involved in the in situ characterisation of soil-borne microbial communities. A number of properties of this marker have made it highly useful in such studies 1 Its ubiquitous presence in all known organisms, and its lack of horizontal gene transfer between organisms Sneath 1993 2 conserved secondary structure, facilitating sequence alignment Van de Peer et al....
Analytical Methods
The reliability of inferences about the presence or absence of specific polypeptides in a soil protein extract depends not only on the rigour of the extraction procedure, but also on the accuracy of the technique used to estimate the concentration of protein molecules in the extract, and on the sensitivity of the analytical technique used for protein detection. For most purposes, the determination of protein concentration in a given extract can be achieved through spectrophotometric analysis...
