1. Local Scale Patterns of Microbial Diversity.
We have concentrated on hypotheses related to the question: "What are the spatial patterns of diversity for microbial communities on local scales?" In particular, we are examining the relationship between plant and microbial diversity at two scales - (1) among species within communities and (2) across plant community types. Our focus for the within community analysis has been done at the scale of individual plants and has focused on the plant rhizosphere. For the between community analysis we are examining the hypotheses that microbial diversity is positively correlated with productivity (Hypothesis 1) and the diversity (Hypothesis 2) of the overlying plant community. We have also examined the impact of disturbance (annual tillage) on the composition and diversity of both the plant and microbial community. Our focus has been on soil communities and processes that they are likely to effect. To address these hypotheses, we are using a combination of field surveys and greenhouse experiments. The field work has utilized experimental plots that are part of the KBS LTER in which the management creates plant communities of different plant composition and productivity. We are also working in late successional old-fields at KBS, in which there is a gradient of plant productivity and diversity within each site.
A. Linking diversity in plant communities to soil microbial communities and processes. Our most recent investigations of this question have focused on comparisons across three experimental treatments on the KBS LTER that differ in plant community composition and diversity: successional old-fields (dominated by perennial herbs), 'weedy poplars' (poplars and perennial herbs) and monoculture poplars. Plant productivity is similar across these treatments, but the diversity and composition of the plant community is quite different. Biolog and PLFA assays show that the two treatments in which poplars are dominant species have similar microbial profiles - but inclusion of the successional fields in the analysis reveals that the perennial forbs have a different signature. These results suggest that characteristics of the dominant plant species in a community -more so then plant diversity- may strongly influence the composition and function of the soil microbial community.
B. Species vs. diversity effects on microbial communities and processes. We have recently initiated a collaboration with John Lawton and Andy Hector at the Centre for Population Biology at Silwood Park, United Kingdom, to further explore the effects of plant species and functional diversity on soil communities and processes. They have established an experiment (BioDepth) in which both the number of plant species and functional groups are varied independently. They find that plant productivity increases linearly with species number, but our results of soil communities indicate that soil processes and the composition of the microbial community are relatively independent of species number or functional groups in the overlying community. There are however, strong 'species composition' effects on several soil processes. We are conducting a more detailed sampling of the plots this fall (September 1999) to follow up on these results.
C. Bacterial Communities of Physical Soil Fractions. The extraordinarily high diversity of soil microbial communities suggests that there are unique structural features in soil that promote coexistence. We hypothesized that the light fraction (LF) of soil may provide a unique microbial habitat analogous to the rhizosphere and may provide additional structural heterogeneity in soil communities. Soil samples were taken from long-term alfalfa plots and continuous corn plots at KBS to compare they composition of these communities. Samples were sieved to isolate rhizosphere, shoot residue (>2mm), and bulk soil. Bulk soil was then separated into LF and heavy fraction (HF, soil aggregates and other minerals) by water centrifugation. Bacterial T-RFLP fingerprints were obtained by DNA extraction from each fraction, followed by PCR amplification with fluorescently-labeled eubacterial primers, and restriction with HhaI. Total bacterial population sizes and populations within cell size classes were determined by digital image analysis of soil smears stained with DTAF. Numbers of CTC-active cells were also determined. Soil bacterial populations were highest in soil fractions where freshly deposited substrate is concentrated (carbon hotspots). Hotspot communities were very different from bulk soil communities. In addition, LF and shoot residue communities were nearly identical but distinct from the rhizosphere. Importantly, cropping history also caused divergence of communities within each habitat type. A wide diversity of habitats is likely to be found in soils, particularly those with more heterogeneous plant assemblages.
D. Effect of disturbance (agronomic treatment) and plant community composition on microbial community structure as measured by quantifying 16S rRNA. To assess the influence of various agricultural treatments on the structure of soil microbial communities, we assessed community structures in soils maintained under distinct agricultural management practices and types of vegetation for seven years prior to sampling and in nearby undisturbed reference plots. Total RNA was extracted from soil samples and 16S rRNA-targeted oligonucleotide probes were used to quantify the abundance of the alpha, beta and gamma proteobacteria, the high mol% G+C gram positive bacteria, as well as the total Bacteria and Eukarya within soil communities. Surprisingly, microbial community structure did not vary significantly across the historically farmed fields of the main site. It is possible that slight, but important differences exist in community structure among the treatments, and that difference were not detected because the number of samples analyzed may have been low relative to the natural variability in the sites. However, differences in the structure of microbial communities in the historically cultivated fields and the uncultivated reference fields were readily detected. This observation suggests that any differences that exist among the historically farmed plots are small in comparison to differences between the historically farmed and undisturbed, reference sites.
Microbial communities in soil samples were also compared by T-RFLP. This analysis provides a different measure of community structure and may better reflect differences in the species-level composition of communities. T-RFLP analyses supported the results from direct rRNA probing: bacterial communities in the historically farmed plots were significantly more similar to each other than to bacterial communities in the reference plots. Additional studies are underway to assess specific factors that may influence the structure of microbial communities in soil and to determine if the observed patterns are constant on a seasonal basis.
E. Effects of plant residue quality on microbial community structure. Plant species (or communities) can have both direct and indirect effects on microbial communities. The indirect effect of a plant community on the composition and structure of the soil microbial communities is manifest by the effect of senescent plant residues. Several authors have suggested that increased plant diversity may have disproportionate effects on ecosystem function suggesting that there are synergistic interactions among species. We tested this hypothesis by examining the response of the soil microbial community to plant residue quality and composition. We found that the response of the microbial community to a five species diverse plant residue mixture was additive, and not synergistic. This study also re-enforced the importance of plant residue quality in driving microbial community composition and dynamics. Further studies of the effects of plant residues will examine the interaction of plant residue quality and quantity on the microbial community, as well as take a more detailed look at the additive effect of plant residue diversity.
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