Fingerprinting methods to approach multitrophic interactions among microflora and microfauna communities in soil

The soil functioning and the response of the biota to external perturbations such as organic input are based on multitrophic interactions among a wide range of organisms. However, the various components of the soil microflora and microfauna are rarely addressed in ecological studies. We have developed a molecular approach based on terminal restriction fragment length polymorphism (T-RFLP) analysis to assess the community structures of protozoa and nematodes, together with bacteria and fungi that share the same soil environment. Two soils were characterized by a specific fingerprint for each of the four groups of organisms, showing the potential of all T-RFLP procedures to differentiate the community structures. The response of the soil biota to organic inputs was addressed using T-RFLP fingerprints together with physiological profiles of bacteria communities and global microbial activities and densities. Although the impact of compost or manure on the soil biota was only slightly noticeable from the global parameters measured, it was obvious from the community level analyses. However, the different components of the soil biota were altered to various extents, depending on the group of organisms and the soil–organic matter combination. The potential of the T-RFLP strategy to analyze simultaneously different biotic groups from the same soil DNA extract will facilitate the more systematic integration of eukaryotic organisms in ecological studies to investigate multitrophic interactions among the microflora and microfauna in relation with soil processes.     

Impact of fumigation with metam sodium upon soil microbial community structure in two Japanese soils

The effects of fumigation with sodium methyl dithiocarbamate (metam sodium) on the microbial community structure and function in 2 soils were investigated using a variety of techniques. In both soils ca. 50% and 90% of the populations of total and culturable bacteria, respectively, were killed by fumigation, with recovery to levels prevailing in control soils 26 d after cessation of fumigation. The size of the ammonium and nitrite oxidiser populations was reduced by up to 4 orders of magnitude by fumigation, with the latter showing a slight recovery 105 d later. There were substantial changes in the C-utilisation (Biolog GN) profiles in the fumigated soils even 105 d later. The number and pattern of amplified 16S ribosomal DNA restriction analysis (ARDRA) fragments was changed by fumigation, and there was a shift in the %G+C profile toward a greater proportion of lower %G+C classes in treated soils. It appeared that DNA released from killed cells remained for some time after fumigation, and masked the apparent community DNA profiles. This study demonstrates that the effects of fumigation on the soil microbial community structure and function were pronounced and for some parameters very persistent. However, the effects on broad-scale properties such as total or culturable bacterial numbers were less enduring.     

Composition of diazotrophic bacterial assemblages in bean-planted soil compared to unplanted soil

The effect of common bean (Phaseolus vulgaris L.) on the composition of nitrogen fixing bacterial assemblages in soil was studied by comparing planted and unplanted soil. The community composition was studied by terminal restriction fragment length polymorphism (T-RFLP) of the nitrogenase reductase gene (nifH). Principal component analysis (PCA) of T-RFLP profiles showed the separation of profiles from planted and unplanted soil. Terminal restriction fragments (T-RFs) corresponding to rhizobial bacteria were identified preferentially in planted soil; however most nifH T-RFs in soil could not be assigned to T-RFs simulated from a database of known diazotrophs. To specifically study rhizobial bacteria in the soil and nodules, PCR products from the alpha subunit of the nitrogenase enzyme (nifD) were analyzed by denaturing gradient gel electrophoresis (DGGE). DGGE results showed the specific stimulation of the rhizobial microsymbionts in planted soil.     

Impacts of methamidophos on the biochemical,catabolic, and genetic characteristics of soil microbial communities

Methamidophos is an organophosphate pesticide with high toxicity and may significantly affect soil microbes. However, the magnitude of this effect is unclear. We examined the effect of low and high inputs of methamidophos on the structure of the soil microbial community, and the catabolic activity and the genetic diversity of the bacterial community using the polyphasic approaches of microbial biomass, phospholipid fatty acids (PLFAs), community-level catabolic profiles (CLCPs), and amplified ribosomal DNA restriction analysis (ARDRA) patterns. Our results indicated that high methamidophos inputs significantly reduced total microbial biomass carbon (C_mic) and fungal biomass, but increased Gram-negative bacteria with no significant effects on the Gram-positive bacteria. Interestingly, CLCPs patterns showed that high methamidophos inputs also significantly improved the catabolic activity of Gram-negative bacteria. The ARDRA pattern showed that the genetic diversity of the bacterial community decreased under chemical stress. Furthermore, changes in the microbial parameters examined were less significant under low inputs than high inputs of methamidophos, suggesting a dosage effect of methamidophos on the microbial community. Our results provide the first evidence that methamidophos differentially affected components of the soil microbial community through inhibiting fungal growth but enhancing the biomass and catabolic activity of Gram-negative bacteria.     

Response of soil microbial communities to compost amendments

Soil organic matter is considered as a major component of soil quality because it contributes directly or indirectly to many physical, chemical and biological properties. Thus, soil amendment with composts is an agricultural practice commonly used to improve soil quality and also to manage organic wastes. We evaluated in laboratory scale experiments the response of the soilborne microflora to the newly created soil environments resulting from the addition of three different composts in two different agricultural soils under controlled conditions. At a global level, total microbial densities were determined by classical plate count methods and global microbial activities were assessed by measuring basal respiration and substrate induced respiration (SIR). Soil suppressiveness to Rhizoctonia solani diseases was measured through bioassays performed in greenhouses. At a community level, the modifications of the metabolic and molecular structures of bacterial and fungal communities were assessed. Bacterial community level physiological profiles (CLPP) were determined using Biolog™ GN microtiter plates. Bacterial and fungal community structures were investigated using terminal restriction fragment length polymorphism (T-RFLP) fingerprinting. Data sets were analyzed using analysis of variance and ordination methods of multivariate data. The impact of organic amendments on soil characteristics differed with the nature of the composts and the soil types. French and English spent mushroom composts altered all the biological parameters evaluated in the clayey soil and/or in the sandy silty clay soil, while green waste compost did not modify either bacterial and fungal densities, SIR values nor soil suppressiveness in any of the soils. The changes in bacterial T-RFLP fingerprints caused by compost amendments were not related to the changes in CLPP, suggesting the functional redundancy of soil microorganisms. Assessing the density, the activity and the structure of the soil microflora allowed us not only to detect the impact of compost amendment on soil microorganisms, but also to evaluate its effect at a functional level through the variation of soil disease suppressiveness. Differences in disease suppressiveness were related to differences in chemical composition, in availability of nutrients at short term and in microbial composition due to both incorporation and stimulation of microorganisms by the compost amendments.     

Evaluating the physical capture method of terminal restriction fragment length polymorphism for comparison of soil microbial communities

Terminal restriction fragment length polymorphism (T-RFLP) is a popular method of comparative microbial community analysis which is normally accomplished by tagging terminal restriction fragments (T-RFs) with a fluorescent primer. Here, we evaluate an alternative method of T-RFLP where T-RFs are physically captured using a biotinylated primer and streptavidin-coated beads. This eliminates one of the primary criticisms of T-RFLP, namely that T-RFs cannot be identified by sequence analysis, and also represents an alternative method for collecting T-RFLP profiles. Microbial communities from forest, agricultural, and turf soils were investigated using several sets of primers specific for different microbial groups. The physical capture method of T-RFLP resulted in similar profiles to those generated by fluorescent T-RFLP. The relationships among ecosystem types captured by both methods and revealed by ordination were virtually identical. The total variance in the profiles that was attributed to ecosystem type was approximately equal, or greater, when generated by the physical capture method, depending on the primers used. However, physical capture T-RFLP resolved fewer T-RFs than fluorescent T-RFLP, and this may reduce the sensitivity to changes in non-dominant populations within the community. Direct cloning and sequencing of physical capture T-RFs revealed that most bands were not comprised of sequences related to those in the database that would generate T-RFs of similar size. T-RFs should therefore be identified by sequencing, rather than by comparing the sizes of T-RFs to computer digests of database sequences. Physical capture T-RFLP should be a useful tool to identify T-RFs by sequencing, and for laboratories without economical access to equipment required to perform fluorescent T-RFLP.     

Eubacterial community structure and population size within the soil light fraction, rhizosphere, and heavy fraction of several agricultural systems

The hypothesis that soil light fraction and heavy fraction harbor distinct eubacterial communities and have differing numbers and sizes of bacterial cells was tested in three agronomic cropping systems. This hypothesis would imply that these soil fractions are distinct microbial habitats. Shoot residue and rhizosphere soil were also included in the analysis. Terminal restriction fragment length polymorphism (T-RFLP) of 16S ribosomal DNA was used to assay eubacterial community structure. T-RFLP profiles were affected by both soil fraction and cropping system, accounting for 35-50% of the variance in the profiles. T-RFLP profiles separated samples into two distinct eubacterial habitats: soil heavy fraction, which includes the mineral particles and associated humified organic matter, and soil light fraction/shoot residue and rhizosphere, which includes particulate soil organic matter. Differences were not based on organic C content of fractions alone; T-RFLP profiles were also differentiated by cropping system and by rhizosphere versus light fraction/shoot residue. Heavy fraction communities had the least amount of random variability in T-RFLP profiles, resulting in the clearest cropping system effects, while rhizosphere and shoot residue communities were the most variable. Profiles from organically managed corn soil were more variable than for either conventionally managed corn or alfalfa. The log number of bacterial cells per gram fraction was affected by soil fraction but not cropping system, being highest in the light fraction. The percentage of cells >0.18 μm³ was also greater in the light fraction than in other fractions. While bacterial cell density was generally correlated with C content of the soil fraction, heavy fraction did have a significantly greater number of cells per μg C than other soil fractions. The results show that habitat diversity in soil, related both to the amounts and types of organic matter, as well as other potential factors, are important in maintaining the high soil bacterial species diversity and evenness that is found in soil.     

RNA- and DNA-based profiling of soil fungal communities in a native Australian eucalypt forest and adjacent Pinus elliotti plantation

Total and active soil fungal communities in a native eucalypt forest and first rotation Pinus elliotti plantation were investigated by direct extraction of DNA and RNA from soil. Terminal restriction fragment length polymorphism (T-RFLP) analysis of internal transcribed spacer (ITS) and 18S rRNA profiles indicated that total and active fungal communities differed significantly in both forest types. This was supported by DGGE profile analysis on an individual plot basis for both forest types and when groups in the canonical analysis were redefined to allow comparison between forest types. Analyses of both ITS and 18S T-RFLP profiles indicated that conversion from native eucalypt forest to P. elliottii plantation may significantly alter total and active soil fungal communities. ITS DGGE (DNA) and 18S (RNA) profiles also suggested differences in fungal communities in the two forest types. No significant separation of the fungal communities in the two forest types was observed, however, when ITS DGGE (RNA) profiles were compared. Overall, the data suggest that conversion from native eucalypt forest to P. elliottii plantation at the Beerburrum State Forest in subtropical Australia has significantly altered soil fungal communities.     

Mathematical model for microbial degradation of pesticides in the soil

A theoretical model for the microbial degradation of pesticides in soil is presented. This model takes into account an important soil property, the adsorption capacity, and the probable modification of the soil microflora after the pesticide was introduced into the soil.The main assumptions of the model are that: (1) the degradation takes place simultaneously in two ways, by metabolism and by co-metabolism; (2) the pesticide exhibits a lethal effect on a sensitive fraction of the soil microflora; (3) a fraction of this microflora is insensitive and does not degrade the pesticide; (4) the various fractions of the soil microflora can grow at the expense of both the pesticide and soil carbon (metabolizing microorganisms) or of the soil carbon alone (co-metabolizing, sensitive and insensitive microbial populations).Two numerical examples are given to illustrate the soil adaptation phenomenon.The validity of different hypotheses included in the model when applied to the soil system are discussed by comparing the results of the simulations to some experimental data concerning the modification of the degrading soil microflora or by comparing some physico-chemical and biological parameter values used in the calculations to published values.     

Long-term exposure to Zn-spiked sewage sludge alters soil community structure

An 8 year study to investigate the effects of Zn-spiked sewage sludge additions on the microbial community structure and microbial processes was carried out in a field soil under pasture. The microbial community structure was evaluated using a combination of multiplex-terminal restriction fragment length polymorphism (M-TRFLP) and T-RFLP fingerprinting approaches. Soil respiration, microbial biomass and enzymatic activities were measured as indicators of soil microbial processes. Changes in the microbial community structure, with Zn additions were evident in all the microbial groups investigated (bacteria, fungi, archaea, actinobacteria and rhizobia/agrobacteria). The fungal community showed the greatest response to Zn additions compared to the other microbial communities measured. The relative abundance of several fungal terminal restriction fragments (TRFs) significantly increased in high Zn treated treatments, at the expense of others, some of which were lost from T-RFLP profiles completely. These results indicate that metal-spiked sludge application can have long-lasting impacts on the composition of the microbial community in pasture soils. Despite notable changes in community structure there was no significant long-term impact of Zn-spiked sludge applications on microbial respiration, biomass or enzyme activities.     

Non-parametric multivariate comparisons of soil fungal composition: Sensitivity to thresholds and indications of structural redundancy in T-RFLP data

Complex soil microbial data produced by molecular profiling techniques, such as terminal restriction fragment length polymorphism (T-RFLP), are often analysed using multivariate statistical methods. Despite this, there has been little evaluation of the sensitivity of multivariate methods to routine data manipulations such as the application of noise thresholds. We examine effects of three percentage area thresholds (0.1%, 0.5% and 1.0%; i.e. ranging from low to commonly applied) on non-metric multidimensional scaling (NMDS) ordinations of soil fungal T-RFLP profiles. We then interpret threshold effects by introducing the concept of structural redundancy in T-RFLP data.NMDS ordinations compared 20 soil samples (encompassing seven sites, two forest types, and three locations) for T-RFLP presence/absence data, which were produced using primers specific to the rDNA internal transcribed spacer (ITS) region, and three separate restriction enzymes (HinfI, AluI and TaqI). Increasing thresholds from 0.1% to 1.0% led to decreases in average number of detected reproducible fragments per site of 57% (HinfI), 75% (AluI), and 69% (TaqI). However, despite the removal of many fragments unique to site/forest type/location, the application of increasing thresholds did not significantly change NMDS patterns for any enzyme in both the complete data and in a more weakly structured sub-group (involving one forest type and two locations). Thus, our data indicate that application of area thresholds up to 1.0% would have minimal impact on NMDS-based comparisons of soil fungal composition.Robustness of NMDS patterns to considerable fragment loss with increasing threshold suggested a degree of ‘structural redundancy’ in our T-RFLP data – that is, multiple fragments per soil sample were interchangeable in the way they contributed to between-sample NMDS patterns. Indeed, for each restriction enzyme, we found 3–4 peels of data (i.e. mutually exclusive sub-sets) that were capable of reproducing overall patterns in presence/absence data at the 0.1% threshold. Despite potential in the T-RFLP method for production of more than one fragment per fungal species/phylotype, we present arguments to support at least some translation of structural redundancy in T-RFLP data to structural redundancy in fungal community comparisons (i.e. the between-soil similarity patterns can be explained by, or are imprinted in, multiple groups of fungal phylotypes). Our analyses highlight structural redundancy as an efficient method for identifying key phylotypes responsible for differences in community composition among soils, and as a promising starting point for improved understanding of many aspects of spatial soil ecology.     

Atrazine mineralization in bulk soil and maize rhizosphere

An experiment was carried out in the greenhouse in order to compare atrazine mineralization in bulk soil and maize rhizosphere at different development stages. After 4, 8 and 12 weeks, we have (1) measured the soil microbial biomass C, (2) characterized the C substrate utilization profiles of the culturable microflora, and (3) analyzed atrazine mineralization. Microbial growth was stimulated in planted soil and different C substrate utilization patterns were obtained in bulk and rhizosphere soils during the first 2 months. During this period, laboratory tests for atrazine biodegradation revealed a lower mineralization potential in bulk than in planted soil. Atrazine mineralization was stimulated to a greater extent after atrazine application in the greenhouse but again the presence of plants had a favorable effect. After 12 weeks of cropping, the atrazine mineralization potential decreased in planted soil with or without prior atrazine application.     

Characterization of cultivable heterotrophic bacterial communities in Cr-polluted and unpolluted soils using Biolog and ARDRA approaches

The bacterial communities of two soils with different chromium levels were characterized by Biolog carbon substrate utilization patterns and amplified 16S ribosomal DNA restriction analysis (ARDRA). For each bacterial community sample, cell suspensions containing 10,000 or 100 colony-forming units (CFU) were inoculated in each well of Biolog-GN microplates. The number of carbon compounds utilized by the bacterial community consisting of 100 CFU from unpolluted soil was significantly lower than that detected for the bacterial community consisting of 10,000 CFU. The size of inoculum did not substantially influence the percentage of carbon sources utilized by the Cr-polluted soil bacterial community. ARDRA approach was applied to about 100 bacterial isolates for each soil sample. A similar number of clusters for Gram-negative bacteria were found in both soils, but there were differences in percentages of isolates belonging to each group and specific genomic groups were found in each soil. Pseudomonas was the dominant taxon in both soils. Comparing the ARDRA clusters obtained from Gram-positive isolates it was evident that the culturable bacterial communities of Cr-polluted and unpolluted soils were dominated by the genus Arthrobacter and the genus Bacillus, respectively.     

长期施肥对农田土壤真菌的影响

不合理施肥所引发的土壤环境问题逐渐成为制约我国农业可持续发展的重要因素之一,而土壤真菌作为一类重要的土壤微生物,研究施肥措施对真菌群落的影响对促进农业生产具有重要意义。本研究以有20年历史的长期定位试验田为研究对象,利用末端限制性片段长度多态性分析技术,对长期定位施肥农田生态系统中不同施肥方式对土壤真菌群落的影响以及时间变化规律进行了系统研究。长期施肥定位试验包括EM堆肥(EM)、传统堆肥(OF)、化肥(CF)和不施肥(CK)处理。主要研究结果如下:在0~20 cm土层,施肥处理对土壤真菌多样性有显著影响,Shannon-Weiner多样性指数为2.64~3.53,Simpson集中性指数为0.03~0.08;EM和OF处理的Shannon-Weiner多样性指数均显著高于CF和CK;在3月、6月和10月,EM和OF处理与CF和CK处理相比,有较高的真菌多样性;Simpson集中性指数最高的是3月的CK处理,最低的是10月的EM和OF处理。冗余分析结果表明,土壤pH、有机质、总氮、有效磷和有效钾等对真菌影响显著。因此,长期施用有机肥与化肥相比可以提高土壤真菌多样性,改变其群落结构;与化肥处理相比,施用EM堆肥,不仅可以保持土壤可持续利用性,同时改善0~20 cm土层土壤真菌的生存环境;3种施肥处理对土壤真菌群落结构影响程度由强到弱:EMOFCF。     

Effects of the introduction of a biocontrol strain of Trichoderma atroviride on non target soil micro-organisms

The main objective of this study was to assess the impact of the application of an antagonistic strain of Trichoderma atroviride on the native microbial soil communities. The structures of the fungal and bacterial communities were assessed by T-RFLP (terminal restriction fragment length polymorphism) method, based on T-RFLP analysis of 18S and 16S rRNA genes, respectively. Results showed that the introduction of the strain I-1237 into two soils slightly modified the microbial diversity, only for a short period of time. Nine months post-inoculation resilience took place, resulting in similar structures of the fungal and bacterial communities in the inoculated and control soils.     

Molecular approaches to investigate herbicide-induced bacterial community changes in soil microcosms

Since biochemical and microbiological methods used to study microbial community changes induced by anthropogenic activities can be biased, the impact of two herbicides on soil microorganisms was investigated by culture-independent molecular techniques. The effect of three different amounts (the recommended field dose, tenfold, and 100-fold the dose) of propanil or prometryne on the bacterial community of a clay soil, two modalities of incubation (soil moisture at 70% of the field capacity and a soil-herbicide suspension, 1:10, w:v), and time of incubation were investigated by denaturing gradient gel electrophoresis (DGGE) and amplified rDNA restriction analysis (ARDRA). Two sets of primers for 16S rDNA were used to amplify total soil DNA. Sterile and non-sterile samples were used to determine, by HPLC, the amounts of herbicides adsorbed on soil and transformed by soil microorganisms. Prometryne persisted in soil longer than propanil. Propanil was removed significantly more by non-sterile than by sterile samples, while for prometryne, slight differences were observed. 3,4-Dichloroaniline, a product of propanil hydrolysis, was detected in non-sterile samples and increased with incubation time. Propanil did not affect soil bacteria significantly as indicated by DGGE and ARDRA, with the only exception being the soil-herbicide suspension. Despite a lower utilization of prometryne by soil microorganisms, DGGE analysis showed a more diverse banding than with propanil. Some bands were also detected in the DNA sample extracted from the soil-prometryne suspension, and could be representative of bacterial species utilizing the herbicide as a carbon source, in two very different soil microcosms.     

Endogeic earthworms differentially influence bacterial communities associated with different soil aggregate size fractions

Endogeic earthworm activities can strongly influence soil structure. Although soil microorganisms are thought to be central to earthworm-facilitated aggregate formation, how and where within the soil matrix earthworm-facilitated influences on soil microbial communities are manifested is poorly defined. In this study we used 16S rRNA gene-based terminal restriction fragment polymorphism (T-RFLP) analyses to examine bacterial communities associated with different aggregate size fractions (macroaggregates, microaggregates-within-macroaggregates and inner-microaggregates-within-macroaggregates) of soils incubated for 28 d with and without earthworms. We hypothesized that bacterial communities in different soil aggregate size fractions are differentially influenced by earthworm activities. Our results indicate significantly enhanced aggregate formation (both macroaggregates and microaggregates within macroaggregates) in earthworm-worked soils relative to soils receiving only plant litter. Although significant differences were found between bacterial communities of earthworm and litter-only treatments for all soil fractions, communities associated with earthworm-worked macroaggregate fractions exhibited the least similarity to all other soil fractions regardless of treatment. In addition to differences in terminal restriction fragment (T-RF) size distributions, T-RFLP profiles of earthworm-worked soil macroaggregates had significantly fewer T-RF sizes, further suggesting less species evenness and more extensive alteration of bacterial communities within this fraction. These findings suggest that, due to rapid occlusion of organic materials, microbial communities associated with microaggregates-within-macroaggregates formed during or shortly after passage through the earthworm gut are relatively inactive, and therefore change relatively little over time compared to macroaggregate populations as a whole.     

Eubacterial communities in different soil macroaggregate environments and cropping systems

Different positions within soil macroaggregates, and macroaggregates of different sizes, have different chemical and physical properties which could affect microbial growth and interactions among taxa. The hypothesis that these soil aggregate fractions contain different eubacterial communities was tested using terminal restriction fragment length polymorphism (T-RFLP) of the 16S ribosomal gene. Communities were characterized from two field experiments, located at the Kellogg Biological Station (KBS), MI, USA and the Ohio Agricultural Research and Development Center (OARDC), Wooster, OH, USA. Three soil management regimes at each site were sampled and management was found to significantly affect T-RFLP profiles. The soil aggregate erosion (SAE) method was used to isolate aggregate regions (external and internal regions). Differences between eubacterial T-RFLP profiles of aggregate exteriors and interiors were marginally significant at KBS (accounting for 12.5% of total profile variance), and not significant at OARDC. There were no significant differences among macroaggregate size classes at either site. These results are in general agreement with previous studies using molecular methods to examine microbial communities among different soil macroaggregate size fractions, although further study of communities within different aggregate regions is warranted. Analysis of individual macroaggregates revealed large inter-aggregate variability in community structure. Hence the tertiary components of soil structure, e.g. arrangement of aggregates in relation to shoot residue, roots, macropores, etc., may be more important than aggregate size or intra-aggregate regions in the determination of the types of microbial communities present in aggregates. Direct microscopic counts were also used to examine the bacterial population size in aggregate regions at KBS. The proportion of bacterial cells with biovolumes >0.18 μm³ was higher in aggregate interiors than in exteriors, indicating potentially higher activity in that environment. This proportion was significantly related to percent C of the samples, while total bacterial cell counts were not.     

Long-term suppression of Pythium ultimum in arid soil using fresh and composted municipal wastes

 The effect of addition of municipal solid waste (MSW) at different degrees of stabilisation on the biological properties of an arid soil was studied 24 months after application. This included effects on the indigenous soil microflora and soil enzyme activities in the presence and absence of Pythium ultimum. The addition of organic waste (fresh or composted) reduced populations of culturable bacteria and fungi and disease symptoms caused by P. ultimum, and resulted in heavier plants with longer and more extensively branched roots. Addition of organic waste increased the population size of culturable bacteria and fungi, while enzymatic activity of the soil was higher in soil amended with organic matter than in non-amended soil. Populations of biological control agents, such as Trichoderma and fluorescent pseudomonads, were larger in soil amended with organic matter. The addition of urban waste could therefore be a suitable technique with which to restore soil quality by stimulating biological control against plant pathogens such as P. ultimum. Received: 3 May 1999     

Polyphasic characterization of the bacterial community in an urban soil profile with in situ and culture-dependent methods

Bacterial communities of urban soils have not been thoroughly investigated up to now. Therefore, soil samples from the urban park Tiergarten in the centre of Berlin were taken from a profile in 15, 30 and 90 cm depth. The total number of bacteria (4′,6-diamidino-2-phenylindole (DAPI) counts) as well as biomass declined one order of magnitude from topsoil to subsoil. Soil texture changed comparably and water content and amount of organic matter dropped 3–10-fold. The number of culturable bacteria (colony forming units = CFU) also decreased with increasing soil depth. Amplified ribosomal DNA restriction analysis (ARDRA) revealed similar bacterial communities in the two upper soil layers in contrast to the deepest layer. The number of bacterial cells which were detected with probe EUB338 in relation to total cell counts differed between 43 and 35% in the three soil layers. With the probe active count method (PAC) this number could be increased up to 72% of total cell counts in topsoil whereas activation of cells declined with increasing depth. In relation to total cell counts (DAPI) α-Proteobacteria and β-Proteobacteria are equally distributed in all three depths, whereas γ-Proteobacteria declined within the soil profile. With the BIOLOG system we observed the general trend that the capability of utilizing diverse substrates decreased with soil depth whereas a few substrates, such as Tween 40 and Tween 80 could be utilised by the bacteria of all soil depths.