LTER Research in Microbial Ecology:

A synopsis of microbial studies at LTER sites

 Introduction:

Research and interest in microbial studies is both extensive and broad within the Long Term Ecological Research (LTER) Network program of the National Science Foundation. The information here is compiled from data provided by LTER sites regarding microbial ecology research within LTER.  A white paper on LTER Microbial Ecology Efforts was also written by and LTER ad-hoc committee.

  Although not an explicit core research area of LTER, observations and investigations of microbial activity are well integrated into the general research conducted at each of the LTER sites. The microbial investigations range from intensive research like that of the Arctic Tundra LTER site in collaboration with research centers at the Woods Hole Marine Biological Laboratory, to individual graduate student investigations such as those of the Niwot Ridge LTER and of the Laboratory of Microbial Ecology at the University of Virginia and Virginia Coast Reserve LTER. The investigations range from identification and DNA sequencing work to biogeochemical measures of microbial processes. Studies range from microbial observations of undisturbed ecosystems such as the microbial mats in the polar Antarctic lakes of the McMurdo (MCM), Taylor Dry Valleys LTER, to perturbation experiments such as the soil warming experiments at the Konza LTER and elevated CO2 studies at the Harvard Forest LTER.  Identification of microbes from the field using sequencing techniques has been one current focus of microbial studies in general. Studies within the LTER program are in general extensive, including microbial function, survival, biogeography, succession throughout the season, rates of grazing on microbes, measurement of activities at low levels, use of probes to identify organisms from the field, active vs. inactive microbes, and gene expression in the field. The studies include bacterial numbers, bacterial productivity, species/sequencing, bacteria/flagellate food webs, protozoans, and phytoplankton.A new initiative in microbial studies could go beyond already developed methods, and use new methods and technologies to answer questions related to microbial ecology itself.

Microbial Observatory efforts within LTER could be enhanced by further collaborations with existing centers, such as the NSF Center for Microbial Ecology at Michigan State University. Collaborations with private industry such as use of identification/ sequencing kits provided by Perkin Elmer offer potential mutual benefits. LTER sites are ideal locations for studies in the spatial extent and distribution of microbes using DNA sequencing techniques and databases integrated with Geographic Information System (GIS) databases. This work is important not only with more conventional studies of the LTER program but also within the new framework and interests of the Phoenix (CAP) and Baltimore (BES) Urban Ecosystem LTER sites. The investigations could include use of information within databases of National Institutes of Health (NIH) and Center for Disease Control (CDC).

The National Science Foundation, Directorate for Biological Sciences (BIO),  has an active program in Microbial Observatories, with a fourth call for proposals due in July, 2002.  The long-term goal of this activity is to develop a network of sites or "microbial observatories" to discover novel microorganisms, microbial consortia, communities, activities and other novel properties, and to study their roles in diverse environments. Individual investigators or teams of investigators are encouraged to develop and conduct research at a site or habitat dedicated to discovery and study of these novel microbes over time and across environmental gradients. Development and application of new experimental approaches to these studies, including the use of genomic and functional genomic methods, is strongly encouraged.

There are currently 8 LTER research project funded by NSF's program on Microbial Ecology:

(MCB-9977933) Diversity of Nitrogen-Cycling Microorganisms at the H.J. Andrews LTER PI: David Myrold

(MCB-9977882) Observing Patterns of Prokaryotic Diversity along Land use Gradients of the CAP LTER PI: Fred Rainey

(MCB-9977907) Spatial Scales of Genetic and Phenotypic Diversity Among Streptomycetes in Native Soils (takes place at Cedar Creek LTER) PI: Linda Kinkel

(MCB-0084223) Microbial Biogeochemistry and Functional Diversity across the Forest-Tundra Ecotone in the Rocky Mountains (Niwot Ridge LTER) PI: Steve Schmidt

(MCB-9977903) A Microbial Observatory for the Northern Temperate Lakes Long Term Ecological Research Site PI: Eric Triplett

(MCB-9977897) Microbial Observatories: Salt Marsh Microbes and Microbial Processes: Sulfur and Nitrogen PI John Hobbie

(MCB-0084164) Prokaryotic Diversity of a Salt Marsh/Estuarine Complex at the University of Georgia Marine Institute, Sapleo Island (Georgia Coastal Ecosystems LTER) PI: Mary Ann Moran

(MCB-0132085) Microbial Observatories: Collaborative Research: A Cold Microbial Observatory: Collaborative Research in an Alaskan Boreal Forest Soil PI: Jo Handelsman

John R. Vande Castle, LTER Network Office, Editor  - Last edited Thursday, July 25, 2002

Index:

AND – H.J. Andrews Experimental Forest, Blue River, Oregon

 

Robert Griffiths [griffitr@ccmail.orst.edu]

 

There is a wide range of microbial observations conducted at the HJA/LTER site in the Central Oregon Cascade Mountains. They can be generally characterized as: (1) studies of microbial activities and microbially mediated processes or (2) studies on fungal ecology. These studies have included the following:

 

The effects of riparian stand management on the microbial characteristics of stream sediment fine particulate matter:

This work has shown a potential link between forest management practices and stream biological productivity. These seasonal studies have shown that headwater stand harvest can alter the microbiology and chemistry of stream sediments passing through these stands. In the future, we would like to continue these studies documenting the influence of riparian vegetation on seasonal patterns of organic particulate matter quality in stream sediments. This will eventually allow us to make direct connections between riparian management and stream productivity as mediated through microbial activity.

 

Studies on ectomycorrhizal fungi:

Since the mid 1980s, there have been a number of studies centering around ectomycorrhizal fungi. Fungi that form ectomycorrhizae establish an essential interface between soil and plants by extending the nutrient-absorbing surface area of the roots, producing extracellular enzymes that increase phosphorous and nitrogen availability, and protecting against pathogens. One set of studies have focused on a specialized group of ectomycorrhizae that form fungal mats. One of these studies addressed the role these mats play in forest soil biogeochemistry and forest productivity as well as factors influencing their distribution. We would like to expand this work to study factors influencing ectomycorrhizal mat inoculation potential and the extent to which these mats are capable of interconnecting different types of plants as well as their possible role in seedling reestablishment. Studies to date suggest that these fungi are capable of transferring both organic and inorganic nutrients from overstory trees to seedlings; increasing the probably of seedling establishment on disturbed sites.

Two major studies have examined the community structure of ectomycorrhizal fungi in and near the HJA/LTER site. Despite the importance of ectomycorrhizal fungi to these ecosystem processes, little is known about their community structure and dynamics. The first of the two studies focused on hypogeous truffle species. While ecologically important, the hypogeous fungi comprise a small fraction of ectomycorrhizal species. In a second study, both epigeous mushroom as well as hypogeous species were included. Permanent plots were established in multiple stands of various age classes of Douglas-fir. This comprehensive four-year study yielded an extensive database providing the backbone for addressing new and critical research topics including: 1) conservation biology of rare fungal species, 2) sustainability of edible, commercially harvested fungi, 3) fungi as important indicators of old-growth legacy and forest community dynamics, 4) development of habitat models for fungi, and 5) implementation of studies employing molecular techniques to more effectively answer questions about belowground community dynamics.

 

Factors influencing respiration and nitrogen fixation rates in decomposing logs:

This ongoing research was established in 1985 and continues today through the efforts of two graduate students. A key finding, determined by remeasuring sporocarp production for 8 years, was that fungal sporocarps were exporting many nutrients (including N and P) during the early stages of log decomposition. This result was total unanticipated, and has caused us to rethink the mechanisms controlling nutrient loss from woody detritus. This work will continue in the future as we collect data to parameterize and validate log decomposition models.

 

The effects of climate on forest soil carbon and nitrogen dynamics:

We have conducted studies of soil processes at the HJA employing the same techniques and experimental design used in similar studies of Oregon Coastal Ranges and southern Oregon forests. These studies have permitted us to compare the effects of regional climate differences on carbon and nitrogen cycling as mediated by microbial transformations and direct effects on vegetation. These studies have shown that moisture is apparently the major driver of soil C and N cycling at this scale. Studies conducted at the HJA have shown other factors being primary drivers of C and N cycling at watershed and stand levels.

At the HJA, two sets of well documented permanent forest floor study sites have been established which have been used by ourselves and others to study the effects of local climate and forest disturbance on forest soil biogeochemistry. One set of 24 permanent plots have been used in monthly measurements of soil respiration at a few selected sites and another set of 184 sites have been established to obtain maximum spatial coverage of the HJA. This ongoing study has allowed us to determine which components of carbon and nitrogen cycling are most influenced by climate by comparing climatic gradients across the HJA with various elements of biogeochemical cycling. These data will be used to parameterize predictive models of landscape-scale climate change scenarios on soil biogeochemistry. The large quantities of soil biogeochemical data have been collected at both sets of permanent sites will also be an invaluable resource to future researchers as they assess long-term changes in forest soils in response to global climate change.

 

The effects of forest disturbance on soil carbon and nitrogen dynamics:

The effects of stand harvest on soil carbon and nitrogen cycling has been studied at both the stand and landscape levels. Since C and N cycling is primarily controlled by microbial processes, these should be considered microbial observations. These studies are being continued under LTER 4 with the assessment of microbial factors influencing successional trajectories of sites after harvest as well as the influence of slow vs. fast regeneration on forest soils. These ongoing studies will be used to parameterize predictive models which will eventually allow us to assess the effects of forest management practices on carbon and nitrogen pools over the landscape.

In the future, we would like to set up manipulated plots at high elevations to evaluate the conditions that lead to the degradation of forest soils after a disturbance. There are a number of sites at high elevations in both the mountains of southern Oregon and the Central Cascades in which repeated efforts to reestablish conifers seedlings after harvest has failed; even 45 years after harvest. These studies will permit us to assess the factor/s responsible for this phenomenon; with this information, management practices could be devised to reduce the generation of degraded forest soils in the future. We are going to need considerable additional resources to maintain and monitor those plots over the coming years.

 

The effects of vegetation on soil biology and chemistry:

This is actually an extension of the forest disturbance studies since disturbances invariably result in the changes in vegetation. There are conditions under which there is a fundamental shift in vegetation that results in much different plant succession that is normally found at the HJA. In these cases, the normal bootstrapping that occurs between vegetation and soils appears to have failed. To better understand this feedback phenomenon, we have been studying and would like to continue studying the basic differences that occur in soil microbiology in response to the establishment of different vegetation types.

There has also been another study recently initiated at the HJA to assess the impact of different organic inputs on forest soil chemical and microbial characteristics. This NSF sponsored study will continue over the next few years as the impact of these manipulations are assessed.

As has traditionally been the case in most microbial studies, most of the studies described above have been of relatively short duration. We see, however, major benefits in continuing many of these measurement programs. Although the function of individual microorganisms is very limited in time and space, their collected effects over large temporal and spatial scales is considerable. In fact, much of our understanding of biogeochemistry on which forest productivity and litter/log decomposition models are based is incomplete. Long-term studies of microbial processes within the context of stand regeneration under different climatic regimes are critical to obtaining this understanding. In addition, long-term studies of fungal distribution patterns are needed to manage for this increasingly valuable commodity for maintenance of both biodiversity and productivity. In summary, the microbial studies currently and recently conducted at the HJA provide an excellent framework on which future long-term studies can be built.

 

 

ARC - Arctic Tundra, Toolik Lake, Brooks Range, Alaska

 

John Hobbie [jhobbie@lupine.mbl.edu]

Michele Bahr [mbahr@lupine.mbl.edu]

MICROBIAL ECOLOGY AT THE ARC LTER SITE

The information on microbial systems in Toolik Lake has developed over the years along the same lines as the developing field of aquatic microbial ecology. The first information came from direct counts (AO, DAPI); bacteria numbers were unexpectedly high in the summer (1-2 x 106/ml, maximum of 3.1) and as low as 0.1 x 106/ml during the winter (Hobbie et al. 1983, Johnston and Kipphut 1988). Radioisotope studies (thymidine, leucine methods) gave an estimate of 3-8 gC/m2 for the annual bacterial production (O’Brien et al. 1997). In this ultraoligotrophic lake, with an annual primary productivity of 15 gC/m2, the DOC from land equals algal production as a source of carbon and energy for bacterial production. Several mesocosm experiments with lake water and DOC leached from tundra vegetation confirm the availability of this DOC to microbial degradation (Hobbie and Kling in prep.). In fact, the input of DOC to Toolik Lake was measured by Whalen and Cornwell (1985) as 98 g DOC/m2/yr so only a few percent are likely used by the bacteria.

The benthos of Toolik Lake is found in the rocky littoral zone and in the soft sediment profundal zone. The sedge Carex aquatilis is found in stands in very shallow water and contributes about 70 g of litter for each square meter of the stands. Microbial decomposition of this litter is rapid and over 60% of the Carex leaf material is lost within the first month after the litter reaches the water (Federle and Vestal 1980). Large fungal-like forms grow rapidly on the surface of the leaves for the first two weeks after which bacteria and pennate diatoms become dominant (Federle and Vestal 1982). Rates of microbial decomposition are adversely affected by reduced pH (McKinley and Vestal 1982) or high turbulence (Federle et al. 1982). Rates of microbial decomposition are stimulated by increased temperature (Federle et al. 1982b) and nutrient enrichment (Federle et al. 1982a). This series of studies showed the importance of physical factors in determining the rate at which plant litter is colonized and degraded in aquatic environments.

The benthos of the Kuparuk River, another ARC LTER site, has been studied throughout a fifteen-year fertilization experiment. The addition of phosphorous has resulted in an overall metabolic shift from heterotrophy to autotrophy (Peterson et al. 1985). Some microbial processes were also stimulated by the increased photosynthesis caused by fertilization. Total respiration of the epilithon, acetate uptake (Hullar and Vestal 1989) and decomposition of lignin monomers were all stimulated but only in light-grown epilithon (Lock et al. 1990, Hershey et al. 1997). When epilithon was grown in the dark in the fertilized region of the river, there was no increased respiration. Also, phosphorous did not stimulate the decomposition of Carex litter (Peterson et al. 1993). Analysis of the micro-algal community structure of epilithon and moss epiphyton is an ongoing Kuparuk River project (Slavik and Peterson in prep.).

With the advent of molecular techniques, it became possible to assess the phylogenetic affinity of cultured and uncultured bacteria from Toolik Lake (Bahr et al. 1996). Ribosomal RNA genes were amplified and sequenced from cultures or DNA extracted from concentrates of bacteria from 40 liters of lake water. All of the genes were closely related to phyla represented in the ribosomal RNA database. Some were related to terrestrial forms or to forms typical of nutrient-rich habitats; these are likely from particles or from land. One group is a relative of the SAR11 cluster previously detected only in marine environments. This initial molecular survey of Toolik Lake microorganisms was not designed to detect Archaea or Eukarya.

In recent years, we have shifted our attention to the heterotrophic nanoflagellates (HNAN, 2-20 m m). An earlier Toolik study (Peterson et al. 1978) was the first to show that bacteria were not grazed to any extent by the larger zooplankton. We found that when parcels of water were confined in large plastic mesocosms (60 m3, 6 m3), it was possible to follow a classic predator-prey cycle between the nanoflagellates and their bacterial prey. This was particularly obvious when the phytoplankton was fertilized with inorganic nutrients. Then a number of cycles could be traced and we even found that the bacterial productivity was at its highest when the bacteria were grazed so much that their numbers fell drastically (Hobbie and Helfrich 1988). The identity of almost all of these nanoflagellates is unknown although we do have numbers of nanoflagellates (a few thousands per ml) and a species list of algae (H. Kling in O’Brien et al. 1996), generated with the inverted microscope, that states that Chrysophyceae are dominant and that the genera Chromulina, Ochromonas, Spiniferomonas, Pseudopedinella, Pseudokephyrion, Paraphysomonas and Kephyrion are present. Some of these have colorless forms or are mixotrophic but we do not have any solid taxonomy for the nanoflagellates. Bahr’s recent experiments (Hobbie et al. in press) solved the problem of measuring grazing rates on bacteria (developed a 14C-labeled bacteria method suggested by Meinhardt Simon) and revealed that the HNAN grazing rate of 16% of the volume per hour removed bacteria at about the same rate (25% of production) as they were produced. Thus in Toolik Lake the nanoflagellates are the dominant grazers of the bacteria.

An additional piece of the puzzle comes from the Rublee and Bettez (1995) finding that the microplankton (20-200 m m, flagellates, ciliates, rotifers, and crustacean nauplii) are the dominant grazers of the nanoflagellates.

The final question concerns the survival of the nanoflagellates in the lake and pond. Do they persist over winter in the water column of the lake or are they reintroduced by streams in the spring. Do they overwinter in the pond and lake sediments?

As a result of the information gathered to date, we conclude that the biggest unknown in our current understanding of the microbial system is the diversity and ecology of nanoflagellates and other protists. We believe this is true for all aquatic environments. For this reason, we have designed a project to use state-of-the-art techniques to study the ecology of nanoflagellates in an arctic system. The information gathered on the identity of the flagellates and their position in molecular based trees (using ribosomal RNA sequences) will provide insights about their evolutionary relationships as well as a data base for designing molecular probes capable of measuring seasonal fluctuations in nanoflagellate populations.

Our goal for future aquatic research is to determine not only the identity, abundance and distribution of all components of the microbial food web, but to understand their physiological capabilities. We want to know what use they are making of these capabilities in the functioning of ecosystems. This will most likely be done via methods, which detect gene expression in the natural environment.

Besides the research being conducted at the lakes and rivers aquatic sites, parallel terrestrial experiments have been ongoing at the ARC LTER. The importance of microbial activity is clearly recognized in the terrestrial environment. However, the soil experiments have been more process-oriented studies of CO2, methane and DOC fluxes and therefore will not be discussed in this brief report.

 

Selected Aquatic ARC References related to Microbial Ecology:

Bahr, M., J.E. Hobbie, and M.L. Sogin. 1996. Bacterial diversity in an arctic lake: a freshwater SAR11 cluster. Aquat. Microb. Ecol. 11:271-277.

Cornwell, J.C. and G.W. Kipphut. 1992. Biogeochemistry of manganese- and iron-rich sediments in Toolik Lake, AK, p. 45-59. In W.J. O’Brien (ed) Toolik Lake: Ecology of an Aquatic Ecosystem in Arctic Alaska.. Hydrobiologia 240:1-269.

Federle, T.W., V.L. McKinley and J.R. Vestal. 1982a. Effects of nutrient enrichment on the colonization and decomposition of plant detritus by the microbiota of an arctic lake. Can. J. Microbiol. 28:1199-1205.

Federle, T.W., V.L. McKinley and J.R. Vestal. 1982b. Physical determinants of microbial colonization and decomposition of plant litter in an arctic lake. Microb. Ecol. 8:128-138.

Federle, T.W. and J. R. Vestal. 1982. Evidence of microbial succession on decaying leaf litter in an arctic lake. Can. J. Microbiol. 28:6856-695.

Federle, T.W. and J. R. Vestal. 1980. Microbial colonization and decomposition of Carex litter in an arctic lake. Appl. Environ. Microbiol. 39:888-893.

Hershey, A.E., W.B. Bowden, L.A. Deegan, J.E. Hobbie, B.J. Peterson, G.W. Kipphut, G.W. Kling, M.A. Lock, R.W. Merritt, M.C. Miller, J.R. Vestal, and J.A. Schuldt. 1997. The Kuparuk River: a long-term study of biological and chemical processes in an arctic river, p. 107-130. In A.M. Milner and M.W. Oswood (eds) Freshwaters of Alaska: ecological synthesis. Springer-Verlag, New York.

Hobbie, J.E., M. Bahr and P. Rublee. 1998. Controls on microbial food webs in oligotrophic arctic lakes. Arch. Hydrobiol. In press.

Hobbie, J.E. 1997. History of limnology in Alaska: expeditions and major projects, p. 45-60. In A.M. Milner and M.W. Oswood (eds) Freshwaters of Alaska: ecological synthesis. Springer-Verlag, New York.

Hobbie, J.E. and J.V.K. Helfrich. 1988. The effect of grazing by microprotozoans on production of bacteria. Arch. Hydrobiol. 31:281-288.

Hobbie, J.E., T.L. Corliss and B.J. Peterson. 1983. Seasonal patterns of bacterial abundance in an arctic lake. Arc. Alp. Res. 15:253-259.

Johnston, C.G. and G.W. Kipphut. 1988. Microbially mediated Mn (II) oxidation in an oligotrophic arctic lake. Appl. Environ. Microbiol. 54:1440-1445.

Kling, G.W., W.J. O’Brien, M.C. Miller and A.E. Hershey. 1992. The biogeochemistry and zoogeography of lakes and rivers in arctic Alaska, p. 1-14. In W.J. O’Brien (ed) Toolik Lake: ecology of an aquatic ecosystem in arctic Alaska. Hydrobiologia 240:1-269.

McKinley, V.L. and J.R. Vestal. 1982. Effect of acid on plant litter decomposition in an arctic lake. Appl. Environ. Microbiol. 43:1188-1195.

Lock, M.A., T.E. Ford, M.A.J. Hullar, M. Kaufman, J.R. Vestal, G.S. Volk and R.M. Ventullo. 1990. Phosphorous limitation in an arctic river biofilm- a whole ecosystem experiment. Water Resources 24: 1545-1549.

O’Brien, W.J.(ed) 1992. Toolik Lake: ecology of an aquatic ecosystem in arctic Alaska. Hydrobiologia 240:1-269

O’Brien, W.J., A.E. Hershey, J.E. Hobbie, M.A. Hullar, G.W. Kipphut, M.C. Miller, B. Moller and J.R. Vestal. 1992. Control mechanisms of arctic lake ecosystems: a limnocorral experiment, p. 143-188. In W.J. O’Brien (ed) Toolik Lake: ecology of an aquatic ecosystem in arctic Alaska. Hydrobiologia 240:1-269

O’Brien, W.J., M Bahr, A.E. Hershey, J.E. Hobbie, G.W. Kipphut, G.W. Kling, H. Kling, M. McDonald, M.C. Miller, P. Rublee, and J.R. Vestal. 1997. The limnology of Toolik Lake, p. 61-106. In A.M. Milner and M.W. Oswood (eds) Freshwaters of Alaska: ecological synthesis. Springer-Verlag, New York.

Peterson, B.J., J.E. Hobbie and J.F. Haney. 1978. Daphnia grazing on natural bacteria. Limnol. Oceanogr. 23:1039-1044.

Peterson, B.J, J.E. Hobbie, A. Hershey, M.A. Lock, T.E. Ford, J.R. Vestal, V.L. McKinley, M.A.J. Hullar, M.C. Miller, R.M. Ventullo and G.S. Volk. 1985. Transformation of a tundra river from heterotrophy to autotrophy by addition of phosphorous. Science 229:1383-1386.

Peterson, B.J, L. Deegan, D.M. Fiebig, T.E. Ford, J. Helfrich, A. Hershey, A. Hiltner, J.E. Hobbie, M. Hullar, G.W. Kipphut, M.A. Lock, V.L. McKinley, M.C. Miller, B. Moller, R. Ventullo, J.R. Vestal and G. Volk. 1993. Biological response of a tundra river to fertilization. Ecology 74: 653-672.

Rublee, P.A., and N. Bettez. 1995. Change in microplankton community structure in response to fertilization of an arctic lake. Hydrobiologia 312:183-190.

Whalen, S.C. and J.C. Cornwell. 1985. Nitrogen, phosphorus and organic carbon cycling in an arctic lake. Can. J. Fish. Aquat. Sci. 42:797-808.

 

 

BES - Baltimore Ecosystem Study LTER Baltimore, Maryland

 

Peter Groffman [groffmanp@ecostudies.org]

Microbial Ecology in the Baltimore LTER:

We are beginning to establish a group of intensive, permanent monitoring sites where there will be lots of vegetation and soil work, including microbial ecology done. The sites will be distributed to represent land use conditions within the study area as follows:

 

Our experimental design has been structured to allow us to compare the influence of natural variation caused by soils with human-induced variation caused by urban air pollution and land use change on microbial nutrient cycling processes.

Specific microbial work that will be done at these sites include routine measurement of nitrogen mineralization, nitrification, denitrification, soil respiration, nitrous oxide flux, methane flux and microbial biomass carbon and nitrogen content. These measurements will be used in simulation models to produce ecosystem, landscape (watershed) and regional scale estimates of microbial processes.

Our sampling design and planned data collection make us well poised to develop our site as a "microbial observatory". We could quickly and easily incorporate the latest techniques for measuring microbial community composition and function into our planned basic microbial monitoring program. Our planned

structure, where microbial data will be collected in the context of a large suite of biogeochemical process measurements, will ensure that new microbial data would immediately be used to help interpret broad patterns of ecosystem structure and function and to stimulate new detailed research on microbial ecology.

BNZ - Bonanza Creek LTER Fairbanks, Alaska

Richard Boone [ffrdb@aurora.alaska.edu]

 

bnz.gif (71078 bytes)Introduction

Work on microbial processes at Bonanza Creek has ranged across a number of scales. Large scale work has focussed on understanding microbial processes at the ecosystem scale. At the opposite extreme, work has focussed on understanding the dynamics of soil microbial and faunal communities and how community structure relates to process dynamics.

Controls over carbon dioxide and methane fluxes across the taiga landscape.

Over four years we monitored CO2 and CH4 fluxes in a range of taiga forests along the Tanana River in interior Alaska. We studied floodplain alder and white spruce sites and upland birch and white spruce sites. Each site had control, fertilized, and sawdust amended plots. CO2 emissions decreased with successional age across the sites (alder & birch > white spruce) regardless of landscape position. When soil temperature was below 17° C, flux rates correlated strongly with temperature but not with moisture. Above 17° C, however, fluxes were lower than predicted by temperature and appeared to be limited by soil moisture. Of the manipulations, only N fertilization had any effect on CO2 production, decreasing flux in the floodplain sites. Landscape position was the best predictor of CH4 flux. The two upland sites consumed CH4 at similar rates (~0.5 mg C m-2 d-1), whereas the floodplain sites had lower consumption rates (0-0.3 mg C m-2 d-1). N fertilization and sawdust both inhibited CH4 consumption in the birch and floodplain spruce sites but not in the upland spruce site. The biological processes driving CO2 fluxes were sensitive to temperature, moisture and vegetation, whereas CH4 fluxes were sensitive primarily to landscape position and biogeochemical disturbances. Hence, climate change effects on C-gas flux in taiga forest soils will depend on the relationship between soil temperature and moisture and the concomitant changes in soil nutrient pools and cycles.

Microbiology of Atmospheric Methane Consumption

Soil consumption of atmospheric CH4 is an important component of the global CH4 cycle, yet it remains poorly understood at the microbial level. We investigated the influences of CH4 supply, soil moisture, dissolved salts, and NH4+-fertilizer on the activity of soil CH4 oxidizers. When starved of CH4, two upland taiga soils lost their capacities to oxidize CH4, indicating that the organisms involved were truly methanotrophic. One of the most intensely investigated controls on soil CH4 consumption has been its inhibition by NH4+ and other factors. Our work on the inhibition of CH4 consumption showed that there are three physiologically different patterns of N-inhibition of CH4 oxidation that occur in different ecosystems. In some soils, inhibition is immediate and results from competitive inhibition by NH3; in other soils inhibition is delayed and appears to result from the inhibition of CH4 oxidizer growth; in yet other ecosystems CH4 oxidizers are simply insensitive to NH4+. On top of these N effects, however, other salts can inhibit CH4 oxidation. In some soils non-NH4+ ions were so toxic that they completely masked the NH4+ effect. These results were the first to demonstrate CH4 oxidizer growth at atmospheric concentrations of CH4. This work also suggested that the primary landscape-level control over the response of soil CH4 consumption to NH4+-fertilization is the distribution of physiologically distinct CH4 oxidizers across sites.

Structure of forest floor soil communities

In the forest floor of Alaskan taiga, annual layers of Equisetum (horsetail) litter demarcate cohorts of birch litter. Forest floor material was separated into each of the three most recent litter cohorts, plus the Oe and Oa layers. Overall, respiration potential decreased with depth of litter (litter age) and over the growing season. Nitrogen mineralization potential increased with depth, and fluctuated over time. Microbial biomass did not vary with depth, but did increase greatly in September in conjunction with increased litter moisture. Litter C:N ratio decreased with time and varied with depth according to the year-to-year variation in litter chemistry. Different groups of soil fauna show distinctly different patterns of activity with depth and time. For example, larval dipterans are limited by moisture conditions to the deeper strata of the litter layer, and thus are important in fragmenting litter, but only after it has been in the forest floor for 2-3 years, though they are capable of feeding on fresh leaves when the environmental conditions are appropriate. Other fauna, such as nematodes and some mites, migrate from deeper to shallower layers of the litter layer over the course of the summer. Microbial and faunal activities on a particular litter cohort is a function of the litter chemistry (N and labile C content), the vertical position of the litter in the forest floor, and the timing within the seasonal cycle.

Plant Secondary Chemical Effects on Soil Microbial Processes

The vegetation mosaic of the Alaskan taiga is produced by patterns of disturbance coupled to well defined successional patterns. In primary succession on river floodplains, one of the critical transitions in succession is that from thinleaf alder (Alnus tenuifolia) to balsam poplar (Populus balsamifera). This is the shift from a N2-fixing shrub to a deciduous tree. Through this transition there are major changes in N cycling including a decrease in N2-fixation, mineralization, and nitrification. Most models of plant effects on soil processes assume that these changes are caused by shifts in litter quality and C/N ratio. However, our studies have shown that balsam poplar secondary chemicals have major effects on soil nutrient cycling. Balsam poplar tannins inhibited N2-fixation in alder nodules. Field studies supported this conclusion by showing the lowest rate of N2 fixation occurs in the successional stage dominated by poplar. Tannins also inhibited decomposition and N-mineralization in alder soils, though different tannin fractions had differing activity, with small tannin molecules (monomers and dimers) acting as microbial substrates. Other poplar compounds, including low-molecular-weight phenolics, were microbial substrates and increased microbial growth and immobilization, thereby reducing net soil N availability. Thus balsam poplar appears to affect nutrient cycling by both inhibiting N2 fixation and mineralization, and by stimulating immobilization. The combination of these activities may function to both reduce N-supply to alder, and to retain N in the ecosystem as alder is replaced and N-inputs decline.

Microbial Stress Ecology

Drying/rewetting and freeze thaw events are common in the taiga, yet we know little about their effects on microbial processes or microbial communities. We have carried out a series of experiments that demonstrate that such stress events cause changes in microbial communities that have long-lasting effects on process dynamics. In an early experiment we showed that a single drying/rewetting event in the lab could reduce respiration by 25% over 2 months following the rewetting. This suggested that stress could reduce the available population of a critical microbial group and that recolonization post-stress could be an important control on microbial activity in litter. To test these ideas, we did a field experiment to evaluate the effect of moisture regime on microbial biomass and activity in birch litter. Litter bags were placed in one of three treatments: continuously moist, watered weekly, and "natural", which experienced two natural dry/wet cycles of two weeks dry followed by rain. There were strong overall correlations between microbial biomass, respiration, and litter moisture content. However, the different treatments had significantly different rates of respiration, biomass, and respiratory quotients (qCO2) that could not be explained by moisture content directly. The natural treatment had lower respiration rates and biomass C and N than the wet or rewet samples, indicating that the 2-week droughts experienced by the natural treatment reduced microbial populations and activity to a greater degree than did shorter droughts. Metabolic diversity of the bacterial community was reduced by multiple episodic drying and rewetting events, as indicated by a dramatic decrease in the number of substrates used in a BIOLOG plate assay. This experiment showed that the size and functioning of the litter microbial community was strongly affected by its prior stress history.

Freeze-thaw cycling caused a flush of microbial C and N during the first cycle, but over three cycles significantly reduced the microbial communities’ abilities to decompose SOM and led to reductions in total C and N mineralized. There differences among soils appear related to differences in the quality of SOM and the native activity of the soil microbial biomass. Multiple stress cycles caused a greater relative reduction in total respiration in high organic-matter-quality than in low quality soils, but recovery of activity is likely faster in high quality soils. The long-term effects of such multiple stresses may depend on the extent of damage to the microbial community and its ability to recover from the stress.

Microbial Activity in frozen soils

We measured microbial respiration in soils from a range of both tundra and taiga communities at temperatures of -2º C and -5º C in a laboratory incubation. For white spruce (Picea glauca), we also measured activity at both 50% and 10% of water holding capacity, as these soils tend to desiccate over the winter. Microbial respiration was significant even down to -5º C. However, C and N mineralization appeared to be disconnected from each other in the frozen soils. The controls on activity in frozen soils are still unclear, but unfrozen water content appears to be important; in the moisture experiment, respiration was significantly greater at 50% WHC than at 10% WHC. It is also possible that the nature of microbial substrates shift as soils freeze, from polymeric soil organic matter to dissolved material and dead microbial biomass.

Mycorrhizal dynamics and mammal browsing

Using an exclosure experiment in the willow stage of primary succession on the floodplain of the Tanana River, we tested the hypothesis that browsing can reduce mycorrhizal infection. We measured the effects winter browsing by moose (Alces alces) and snowshoe hare (Lepus americanus) had on mycorrhizal infection and fine root biomass of willow (Salix spp.) and balsam poplar (Populus balsamifera). We found that protection from winter browsing increased ectomycorrhizal infection by 10% in the top 5 cm of the soil profile, by 23% at 5-10 cm, and by 42% at the 10-15 cm depth. Mammal browsing in taiga forests is now recognized as a major cause of the shift palatable species such as alder and spruce. We suggest that browsing-induced reduction in ectomycorrhizal infection of salicaceous species plays a central role in this shift in plant community composition from palatable deciduous species such as willow and balsam poplar to less palatable species such as alder and spruce. We suggest that browsing-induced reduction in ectomycorrhizal infection of salicaceous species plays a central role in this shift in plant community composition.

Microbial community structure and function in oiled taiga soils.

In 1976 two 7570 liter experimental spills (one in February and one in July) of hot Prudhoe Bay crude oil were conducted in an open black spruce (Picea mariana) forest at the Caribou-Poker Creeks Research Watershed. In 1994 and 1995, a substantial quantity (ca. 0.3 g · g-1 dry soil) of crude oil remained in the soil at the site 18 years after it was spilled, and several microbial parameters showed evidence of long-term oiling effects. Overall, the surviving community in the oiled plot has shifted toward using oil C for growth. Numbers of hydrocarbon degrading microbes, and specific hydrocarbon mineralization potentials, were significantly elevated in the oiled (OIL) plot compared to an adjacent oil-free, reference (REF) plot. Glutamate mineralization potentials and soil C mineralization, on the other hand, were not different between treatments, suggesting that OIL plot heterotrophs were well-acclimated to the oil. Despite the similarity in C mineralization, net N mineralized was lower and net nitrification was absent in OIL soils. Biomasses of total fungi and bacteria, and numbers of protozoa, showed no consistent effects due to oiling, but metabolically active fungal and bacterial biomasses were uniformly lower in OIL samples. Community-level multiple substrate metabolism (Biolog) was assessed using a new technique for extracting kinetic data from the microplates. This analysis suggested that the microbial population diversity in the OIL soils was lower than in REF soils. Further, these data indicated that the surviving populations in the OIL plot may be considered metabolic generalists. Some evidence of crude oil biodegradation was seen in the chemistry data, but enrichment of the oil residue in higher molecular weight components, duration of contact with soil organic material, and slow rates of C mineralization indicate the crude oil will persist at this site for decades. Contamination of Alaskan taiga soil at this site has yielded observable long-term microbial community effects with larger-scale consequences for ecosystem function.

Selected Papers on Microbial Ecology at Bonanza Creek:

  1. Gulledge, J.M. and J.P. Schimel. Effect of CH4–starvation on atmospheric CH4 oxidizers taiga and temperate forest soils. Soil Biology Biochemistry. In press.
  2. Schimel, J.P. and J. Gulledge. Microbial Community Structure and Global Trace Gases. Global Change Biology. In Press.
  3. Schimel, J.P., R.G. Cates, and R. Ruess. The role of balsam poplar secondary chemicals in controlling nutrient dynamics through succession in the Alaskan taiga. Biogeochemistry. In Press.
  4. Gulledge, J. and J.P. Schimel. Moisture Control over Atmospheric CH4 Consumption and CO2 Production in Physically Diverse Soils. Soil Biol. Biochem. In Press.
  5. Wagener, S.M. and J.P. Schimel. 1998. Stratification of soil ecological processes: a study of the birch forest floor in the Alaskan taiga. Oikos. 81: 63-74.
  6. Wagener, S.M., M.W. Oswood, and J.P. Schimel. 1998. River and soil continua: Parallels in carbon and nutrient processing. Bioscience. 48: 104-108.
  7. Lindstrom, J.E., R.P. Barry and J.F. Braddock. 1998. Microbial community analysis: a kinetic approach to constructing potential C source utilization patterns. Soil Biol. Biochem. 30:231-239.
  8. Rossow, L J; Bryant, J P; Kielland, K. 1997. Effects of above-ground browsing by mammals on mycorrhizal infection in an early successional taiga ecosystem. Oecologia 110: 94-98.
  9. Gulledge, J.M., A.P. Doyle, and J.P. Schimel. 1997. Different NH4+–Inhibition patterns of soil CH4 consumption: a result of distinct CH4 oxidizer populations across sites? Soil Biology and Biochemistry. 29: 13-21.
  10. . Schimel, J.P. and J.S. Clein. 1996. Microbial response to freeze-thaw cycles in tundra and taiga soils. Soil Biology and Biochemistry. 28: 1061-1066.
  11. . Schimel, J.P. K. Van Cleve, R. Cates, T. Clausen, and P. Reichardt. 1996. Effects of balsam poplar (Populus balsamifera) tannins and low-molecular-weight phenolics on microbial activity in taiga floodplain soil: changes in N cycling during succession. Can. J. Bot. 74: 84-90.
  12. . Clein, J.S. and J.P. Schimel. 1995. Microbial activity of tundra and taiga soils at sub-zero temperatures. Soil Biol. Biochem. 27: 1231-1234.
  13. . Clein. J.S. and J.P. Schimel. 1995. Nitrogen turnover and availability during succession from alder to poplar in Alaskan taiga forests. Soil Biol. Biochem. 27: 742-752.
  14. Schimel, J. 1995. Ecosystem consequences of microbial diversity and community structure. In: Arctic and Alpine Biodiversity: patterns, causes, and ecosystem consequences. F.S. Chapin and C. Korner (Eds.). Springer-Verlag, Berlin. pp. 239-254.
  15. . Clein, J. and J.P. Schimel. 1994. Reduction in microbial activity in birch litter due to drying and rewetting events. Soil Biology & Biochemistry. 26: 403-406.
  16. . Sugai, S.F. and J.P. Schimel. 1993. Decomposition and biomass incorporation of 14C-labeled glucose and phenolics in taiga forest floor: effect of substrate quality, successional state, and season. Soil Biol. Biochem. 25: 1379-1389.

 

 

CAP Central Arizona - Phoenix LTER Phoenix, Arizona

Nancy Grimm [ngrimm@lternet.edu]

Chris Martin [chris.martin@asu.edu]

 

MICROBIAL ECOLOGY IN THE CENTRAL ARIZONA - PHOENIX LTER

Microbial processes are key elements of changes in organic matter storage and biogeochemistry of both terrestrial and aquatic systems in the urban environment. The Central Arizona - Phoenix (CAP) LTER project will incorporate microbial ecology studies in several of its core monitoring efforts as well as its short-term experiments. Planned measurements of soil respiration and trace gas flux from soils and other urban surfaces on a patch-specific basis will give information on magnitude and spatial distribution of microbial processes. The current scheme of urban patch types includes:

  1. open ground (vacant lot, graded soil, "natural")
  2. water (streams, rivers, canals, lakes)
  3. riparian zones (vegetated, unvegetated)
  4. agriculture (fallow, active)
  5. recreational (golf courses, parks)
  6. residential (mesic, xeric)/commercial/industrial
  7. transportation corridor

Research will characterize these patch types in terms of several ecological variables; those of relevance here include soil organic matter, soil gas fluxes, dissolved organic matter, protist/bacterial biomass of aquatic ecosystems, arbuscular myccorhizal communities, and selected biogeochemical transformations (e.g., denitrification, nitrification, methanogenesis).

Investigations of materials transport and transformation represent an attempt to construct whole-metropolis budgets for nutrients, major ions, salts, and metals. Analysis of input-output balance (using existing data and data collected from the new monitoring project) will give us an idea of where process studies should be focused to understand controls over microbial material transformation in the urban system.

In addition to these planned core elements of research, CAP scientists began a series of short-term studies or "resurveys" in January 1998, several of which have microbial components:

Lichen resurvey with heavy metal analysis

Lichens are a classic bioindicator of air pollution. Twenty-five sites surveyed in Maricopa County in 1974 for the presence or absence of different lichen species will be resurveyed. The new survey will compare the present concentration of trace metals in current lichen tissues with those in samples collected from the original survey.

Comparison of soil respiration among residential patches

Evolution of CO2 from soils in residential lots of four types is being measured as an index of microbial respiration. Permanent soil plots were established in yards of one of four possible residential water-use options (xeriscaped or watered)/patch history (agriculture or desert) combinations.

Biodiversity of AM fungal communities

A preliminary study is being conducted to elucidate arbuscular mycorrhizal (AM) fungal diversity along a temporal gradient (time since landscape establishment) in residential landscaped areas in Phoenix. Rhizosphere soil was collected from ash trees (Fraxinus sp.) at each sampling site in June 1997 and used to establish trap cultures to determine AM species richness and composition. Preliminary findings revealed that AM fungal species richness was greater at more established landscape sites (at least 45 years since planting) compared with recently installed sites (less than 5 years since planting). Species in the Glomaceae predominated at all sites.

Resurvey of urban lakes

Urban lakes of the Phoenix metropolitan area were characterized in the 1970s as to water chemistry, sediment chemistry, and phytoplankton populations. These lakes are currently being resurveyed; surveys will likely be repeated every 10 years.

Rio Salado Town Lake

A pilot project incorporating university classes (field geology, limnology) will examine hydrogeology and limnology of the Tempe Town Lake, an impounded, >300-ha urban lake that will be created in the now dry Salt River bed in the City of Tempe. Pending funding, microbial studies will include: 1) microbial transformations in water column bottom and subsurface substrata; 2) formation and mitigation of noxious algal blooms; 3) microbial biofilm biomass and N and organic C transformations in the 0-1 in stratum of the groundwater recharge zone; and 4) microbially mediated transformations of N and organic C in fringing wetlands receiving surface water inputs (both storm flows and effluent).

 

 

CDR - Cedar Creek Natural History Area - Minnesota

 

Shahid Naeem [naeem001@maroon.tc.umn.edu]

PAST AND CURRENT ACTIVITIES:

More microbial work is needed at Cedar Creek in spite of early studies pointing quite clearly to the strong role of microbes in ecosystem processes at Cedar Creek. Although not directly microbial, (McKone and Biesboer 1986) and (Pastor, et al. 1987, Pastor, et al. 1987) invoked microbial processes in explaining the patterns they observed concerning nitrogen fixation and mineralization and litter decomposition.

Zak (Zak and Grigal 1991) (Zak, et al. 1990) conducted considerable research on microbial biomass in old fields, swamps and forrests. He showed, for example, that N processing can differ by an order of magnitude over short distances in the heterogeneous landscapes of Cedar Creek. Microbial biomass ranged from 80 g C m2 to 165 g C m2 (swamp sites) to as low as 3.4 g N m2 to 9.5 g N m2 (old field, pin oak forest, and savanna sites). Based on relationships between microbial biomass and n-mineralization and denitri, Zak concluded that microbial population dynamics may be exerting control over N availability. Given that most of the hypothetical mechanisms for plant community dynamics at Cedar Creek are nitrogen based, Zak's work clearly points to the potential importance of microbes as the mechanistic agents. His conclusion (Zak, et al. 1990), "Carbon (C) and nitrogen (N) cycles within terrestrial ecosystems are intimately linked through patterns of plant and microbial activity," clearly stated the need to examine both plants and microbes at Cedar Creek.

Johnson, who is again working at Cedar Creek as of this year, examined mycorrhizal fungi at Cedar Creek. Her 1991 paper (Johnson, et al. 1991) showed that VAM mycorrhizal diversity (Shannon-Wierner index, not spp. richness) declines with succession, though density shows modal relationship.

Johnson also tested the hypothesis that nutrient-stress plants release more carbohydrates which leads to selection for mycorrhizae that are less efficient resource provisioners -- turns out to be true in a fertilizer experiment (Johnson 1993).

Although not directly studied, microbes were invoked by Wedin in his 1995 paper on decomposition (Wedin, et al. 1995). Wedin argued that microbes are probably the responsible agents for the increase in litter C which, by stable isotope analysis, proved to originate from soil organic material.

Finally, Terry Chapin and Val Evinger came to Cedar Creek natural history area last summer. They did Biolog plates, short term N15 incubations to estimate microbial N pools, and FAME. We've not heard anything back except that no patterns were observed in Biolog.

CDR is especially interested in getting microbial information (bacterial, protistan, and fungal) from FACE and small biodiversity plots. This involves over 600 plots.

A LTER microbial observatory effort could provide funds to collect bacterial, protistan, and fungal biomass, density, and some measure of diversity from FACE, biodiversity, and smaller, multi-trophic level experiment. A full time technician who would process the samples in collaboration with others at CDR. Data would be collected and dissemination to the investigators and others via Cedar Creek's web page.

Our purpose would be to identify the below ground mechanisms for the biodiversity, CO2, and N fertilization effects we observe and to conduct this work for the next 5 to 10 years. I think this has some very exciting potentials for providing insight into the biodiversity/ecosystem functioning debates.

 

Measurements would include:

Our hypotheses would be that biodiversity effects are mediated by rhizospheric microbial processes. We would get at this by examining microbial responses to the manipulations of plant diversity, CO2, and N in conjunction with measures of soil respiration, n-mineralization, decomposition, and many other variables we keep track of.

 

CDR Microbial Ecology References:

Johnson, N. C. 1993. Can fertilization of soil select less mutualistic mycorrhizae? Ecological Applications 3:749-757.

Johnson, N. C. D., D. R. Zak, D. Tilman and F. L. Pfleger. 1991. Dynamics of vesicular-arbuscular mycorrhizae during old field succession. Oecologia 86:349-358.

McKone, M. J. and D. D. Biesboer. 1986. Nitrogen fixation in association with the root systems of gloldenrods (Solidago L.). Soil Biology and Biochemistry 18:543-545.

Pastor, J., M. A. Stillwell and D. Tilman. 1987. Little bluestem litter dynamics in Minnesota old fields. Oecologia 72:327-330.

Pastor, J., M. A. Stillwell and D. Tilman. 1987. Nitrogen mineralization and nitrification in four Minnesota old fields. Oecologia 71:481-485.

Wedin, D. A., L. L. Tieszen, B. Dewey and J. Pastor. 1995. Carbon isotope dynamics during grass decomposition and soil organic matter formation. Ecology 76:1383-1392.

ZAK, D. R. and D. F. Grigal. 1991. Nitrogen mineralization, nitrification and denitrification in upland and wetland ecosystems. Oecologia 88:189-196.

Zak, D. R., D. F. Grigal, S. Gleeson and D. Tilman. 1990. Carbon and nitrogen cycling during old-field succession: constraints on plant and microbial biomass. Biogeochemistry 11:111-129.

 

 

CWT - Coweeta Hydrologic Laboratory, Otto, North Carolina

David Coleman [Coleman@sparc.ecology.uga.edu]

There are several long-term studies underway which rely extensively upon microbial research and microbial processes. These include measurement of fluxes of nitrogen to the air and to water from dozens of watersheds with varying vegetation covers and of varying successional age, and also studies of human-caused versus natural (hurricane)-caused disturbances.

We have measured the dynamics of carbon, nitrogen, phosphorus and sulfur (Stanko-Golden et al. 1994) into and out of labile organic (microbial) pools (Coleman 1994) in uplands, hillslope/riparian, and streams both within the Coweeta basin (Maxwell and Coleman 1995) and more generally in the 53,000 km2 region of the southern Appalachians. In the Coweeta basin, we have measured small, but significant increases in nitrate export from our upland sites into the > 73 km of upland streams as nitrogenous inputs from the Atlanta metropolitan airshed region moves steadily upward (W.T. Swank, pers. comm.).

In long-term recovery studies after massive windthrows on hillslope sites after Hurricane Opal hit Coweeta in October 1995, there have been large 100-fold increases of nitrate-N in tipup areas into tension lysimeters in our impacted sites (Yeakley, et al., in prep.). In contrast, in a nearby Rhododendron extirpation plot, when all aboveground Rhododendron maximum was removed from the site, but the soil surface was minimally-disturbed, there was only a gradual increase in soil respiration over 2-3 years’ time post-cut (from August 1995 onward). This is attributed to gradually increasing losses of carbon from decaying root systems. No increases in nitrate losses into the lysimeters were noted in the extirpation site.

Coweeta investigators are using a diversity of approaches to assess the trophic significance of bacteria in streams and to assess the role of microbes in nitrogen cycling in streams. Early research investigated the significance of bacteria in the diets of aquatic insects in streams using 3H-thymidine to follow the incorporation of bacteria into animal tissue (Findlay and Meyer 1984, Findlay et al. 1986, Meyer et al. 1987, Meyer 1994). Additional studies used microscopy to examine copepod feeding on bacteria associated with decomposing leaf material (Perlmutter and Meyer 1991) and using fluorescently labeled bacteria to measure the rates at which they were consumed by filter feeders (Hall et al. 1996). Recent microbial work has relied upon stable isotopes to investigate the role of microbes in stream ecosystems.

We have assessed the trophic significance of bacteria in stream food webs by adding tracer quantities of 13C-acetate to streams, one of which had all leaf litter excluded from it for two years (e.g. Hall and Meyer in press). Bacteria in the litter-excluded stream had 7-10 times more labile than those in the reference stream during both summer and winter, showing their higher relative use of streamwater DOC. The fraction of invertebrate carbon derived from bacteria was significantly related to the fraction of amorphous detritus in invertebrate guts, suggesting that the bacterial carbon supporting higher trophic levels was associated with amorphous detritus particles. Invertebrates in the litter-excluded stream did not derive a greater fraction of their carbon from bacteria despite a lower standing crop of detritus in the litter-excluded stream. The standing stock of colloidal carbohydrates was 5 times greater than cellular bacterial biomass; hence the high use of bacterial carbon by invertebrates may be a consequence of the availability of these polymers.

To measure microbial nitrogen in streams, we have also adapted a microbial technique commonly used in soils for estimating microbial biomass nitrogen and carbon. We have measured microbial carbon and nitrogen in fine benthic organic matter, leaves and wood in streams at Coweeta, Hubbard Brook and Walker Branch (Oak Ridge National Lab) using this technique (Sanzone et al. in prep.). We are using this technique to assess the del 15N of microbes in 15NH4 addition experiments in several LTER and other stream sites (e.g. Hall et al. in press).

 

Literature cited:

Coleman,D.C. 1994. The microbial loop concept as used in terrestrial soil ecology studies. Microbial Ecology 28: 245-250.

Findlay, S. G. and J. L. Meyer. 1984. Significance of bacterial biomass and production as an organic carbon source in aquatic detrital systems. Bulletin of Marine Science 35: 3l8-325.

Findlay, S., J.L. Meyer and P.J. Smith. 1986. Incorporation of microbial biomass by Peltoperla sp (Plecoptera) and Tipula sp. (Diptera). Journal of the North American Benthological Society 5:306-310.

Hall, R.O., C. Peredney, and J.L. Meyer. 1996. The effect of invertebrate consumption on bacterial transport in a mountain stream. Limnology and Oceanography 41: 1180-1187.

Hall, R.O. and J.L. Meyer. The trophic significance of bacteria in a detritus-based stream food web. Ecology. In Press.

Hall, R.O., J. Peterson and J.L. Meyer. Testing a nitrogen cycling model of a forested stream using a 15N tracer addition. Ecosystems. In Press.

Maxwell, R.A., and D.C. Coleman. 1995. Seasonal dynamics of nematode and microbial biomass in soils of riparian-zone forests of the southern Appalachians. Soil Biology and Biochemistry 27: 79-84.

Meyer, J.L. The microbial loop in flowing waters. 1994. Microbial Ecology 28:195-199.

Meyer, J.L., C.M. Tate, R.T. Edwards and M.T. Crocker. 1987. The trophic significance of DOC in streams. pp. 269-278 In: W.T. Swank and D.A. Crossley (eds.), Forest Hydrology and Ecology at Coweeta. Springer Verlag.

Perlmutter, D.G. and J.L. Meyer. 1991. The impact of a stream-dwelling harpacticoid copepod upon detritally-associated bacteria. Ecology 72: 2170-2180.

Stanko-Golden, K.M., W.T. Swank, and J.W. Fitzgerald. 1994. Factors affecting sulfate adsorption, organic sulfur formation, and mobilization in forest and grassland spodosols. Biology and Fertility of Soils 17: 289-296.

 

HBR - Hubbard Brook Experimental Forest, West Thornton, New Hampshire 

Peter Groffman [groffmanp@ecostudies.org]

 

Microbial Ecology at the Hubbard Brook LTER

We instituted a long-term monitoring program of microbial biomass and activity at HB in 1993 (see www.hbrook.sr.unh.edu under Data - Soil). The objectives of this program are 1) to provide background data on microbial parameters to help interpret spatial and temporal patterns in our long-term biogeochemical records of stream and soil solution chemistry, litterfall and tree growth and 2) to provide baseline information for more detailed studies of microbial processes (e.g. trace gas fluxes, N mineralization and nitrification, denitrification) and how they respond to ecosystem change (e.g. snow depth and soil freezing, base cation addition).  The microbial program at Hubbard Brook could benefit quickly and greatly from an infusion of funds. A recent NSF site review specifically recommended expansion of the microbial program at Hubbard Brook. Moreover, we have begun collaborating with several individuals who could immediately bring new dimensions to the program. Melany Fisk (Cornell University) is doing novel work with carbon substrate use at Hubbard Brook, Mary Arthur (University of Kentucky) has measured fungal:bacterial ratios, Linda Pardo (USFS) is applying state-of-the-art stable isotope techniques, and Andria Costello, an incoming new   faculty member at Syracuse University is interested in applying the the latest methods of molecular characterization of microbial communities at Hubbard Brook. Sharon Parker (USFS) may also be interested in molecular work at Hubbard Brook. In summary, the HB LTER is well poised to function as a "microbial observatory". We have an existing basic monitoring program and could quickly incorporate the latest techniques into this program. Our existing structure, where microbial data is collected in the context of the long-term biogeochemical record at HB, would ensure that new microbial data would immediately be used to help interpret broad patterns of ecosystem structure and function and to stimulate new detailed research on microbial ecology.

HFR - Harvard Forest, Petersham, Massachusetts

 

John Aber [john.aber@unh.edu]

There are three long-term experiments underway which provide unique opportunities for microbial research, and at least two affiliated projects which use newer techniques. The three projects are the soil warming experiment, the DIRT (or litter input alteration) experiment, and the chronic N addition experiment. Paul Steudler is working with a colleague on a project which used modern biochemical methods to identify changes, and Wric Davidson is working with colleagues at Penn St. on 15N and 13C NMR on the chronic N plots. We would be interested in pursuing the "observatories" path

Unique results have been observed in the Chronic N plots. Over the first 9 years we have added up over 1300 kg N per hectare to a hardwood stand at the Harvard Forest. Only in year 9 did we begin to see measurable nitrate leaching losses, meaning that the system retained nearly 100% of this very large amount of N (annual addition rates of 150kg N/ha/yr are twice the annual internal cycling rate). More than ¾ of this N is retained in soils, suggesting that microbial N immobilization is a major process for N retention. However, two different types of measurements show that there is no significant increase in CO2 evolution from soils accompanying this increased N incorporation. Two alternative hypotheses are being considered: 1) that abiotic (chemical) processes are an important component of the N retention capacity, and 2) that immobilization occurs through Mycorrhizal uptake, assimilation and release, using C from photosynthesis directly. Understanding the capacity of the actual mechanisms responsible for N retention is a critical part of predicting the rate at which forests will approach N saturation and the elevated nitrate leaching losses and potential for forest decline which can result.

References:

Aber, J.D., A. Magill, S.G. McNulty, R. Boone, K.J. Nadelhoffer, M Downsand R.A. Hallett. 1995. Forest biogeochemistry and primary production altered by nitrogen saturation. Water, Air and Soil Pollution85:1665-1670

Magill, A..H., J.D. Aber, J. J. Hendricks, R.D. Bowden, J.M. Melillo and P.A. Steudler. 1997. Biogeochemical response of forest ecosystems to simulated chronic nitrogen deposition . Ecological Applications 7:402-415

Aber, J.D., W.H. McDowell, K.J. Nadelhoffer, A. Magill, G. Berntson, M. Kamakea, S.G. McNulty, W. Currie, L. Rustad and I. Fernandez. Nitrogen saturation in temperate forest ecosystems: hypotheses revisited. BioScience (in press)

 

JRN - Jornada Basin, Las Cruces, New Mexico

 

Peter Herman [rpeter@nmsu.edu]

A large number of microbial investigations are underway by the Jornada research team which can be separated into two types of activities. The first is integrated into the long-term ecological research program itself and part of the core data collection activities of the site. This includes collection of biomass N, viable heterotrophic counts (by MPN) and plate counts of a Nitrogen Efficient Guild (NEG) that is adjacent to the 15 permanent plots (3 in each of 5 vegetation types). The NEG is made up of organisms which are either free living N fixers or very efficient N scavengers. We are also looking at AMF.

The second sort of work involves either site PIs or visitors whose independent projects are either microbial ecology or which have microbial ecology components. For example, there are several projects on the Jornada which are not part of the core research. One looks at microbe distribution associated with shrub invasion and resource island formation (NSF international Programs/Long Term Studies funding) comparing Swedish pasture shrub invasion with that in the Jornada's grassland sites. We (Yossie Steinberger at Bar Elan in Israel and I) just got word that we will be funded for a project which will measure how the total nitrogen, soluble nitrogen and biomass nitrogen pools of in desert soils change with time, particularly what happens after natural wetting events. It will also elucidate how these changes correlate with or control plant productivity and it will follow the population levels of soil organisms including arbuscular mycorrhizal fungi, heterotrophic bacteria and soil nematodes which, because they can be divided into feeding groups, provide a mirror of the soil biota on which they feed. Other researchers like Ross Virginia and Jim Reynolds have looked at various microbial ecology components that impact their work with shrub dynamics, Bill Schlessinger's students have measured microbe mediated processes as part of biogeochemistry work and Curtis Monger has ongoing work which looks at various microbially mediated deposition carbonate processes and cryptogamic crusts as part of soils studies.

Peter Herman is surveying microbial diversity, biomass, and activity in microbial guilds among the various habitats that we study in the Jornada Basin. Several of his papers appear as Herman et al. 1993, 1994, and l995 (all appearing in Applied and Environmental Microbiology). The references are listed in the JRN site bibliography.

Within efforts of Bill Schlesinger to quantify trace gas production at the Jornada, we have measured microbial biomass, nitrogen-fixation and related variables have been measured in different habitats. Several of these papers have appeared as Gallardo and Schlesinger l992, l995 in Biogeochemistry.

On the whole, microbial ecology has a fairly robust presence on the Jornada LTER site and that presence is likely to grow because all of our investigators recognize the connections between macrobiologial and microbiological processes. The limit has been resources (both human and monetary) to make the connections, not a lack of will or interest.

 

KBS - W.K. Kellogg Biological Station, Hickory Corners, Michigan


Michael J. Klug (klug@kbs.msu.edu)
James M. Tiedje (tiedjej@pilot.msu.edu)
Eldor A. Paul (paulea@pilot.msu.edu)
Katherine L. Gross (kgross@kbs.msu.edu)
G. Philip Robertson (robertson@kbs.msu.edu)
Thomas M. Schmidt (tschmidt@pilot.msu.edu)

 

Microbial Research at the KBS LTER Site

soybean landscape.gif (51618 bytes)Soil microbial ecology is a central part of KBS LTER research. Understanding the ecological interactions underlying the productivity of field crop agriculture is the central focus of LTER research at KBS, and microbes comprise one of our most intensively studied taxa, together with vascular plants and insects. Microbial studies at KBS take a variety of forms, with most studies directed towards questions about the patterns, causes, and consequences of microbial diversity and microbial biomass for ecosystem processes in intensively managed ecosystems.

Our studies to date have centered on examinations of microbial growth rates, biomass, and fungal: bacterial ratios.  We have also focused on population-level questions using direct microscopy, classical pure-culture techniques, and, for the multitude of unculturable microbes in soil, molecular analyses of phenotypes and genomes. Many of these latter techniques provide whole-soil signatures of community composition, and have been particularly useful for examaning community-level differences among sites and experimental treatments. For questions related to specific populations we have focused our efforts on examinations of specific functional groups such as denitrifiers, nitrifiers, lignin and 2,4-D degraders, and the rhizobacteria, linking these groups to specific microbial processes.  Much of this research has been collaborative with the NSF Center for Microbial Ecology (www.cme.msu.edu) at Michigan State; a number of   KBS LTER co-PI's are also co-investigators in the CME.

We provide below background information on our current studies of microbial community structure. Other microbial work is also underway at KBS but not described here due to space limitations – including detailed biogeochemical and population-level investigations of microbial processes such as denitrification, trace gas fluxes, and soil organic matter turnover and DOC and DON fluxes. Following this section we highlight specific analytical procedures now in use at KBS.

 


Microbial Community Structure

SEM microbes in soil.jpg (81536 bytes)The diversity and complexity of soil microbial communities present a major challenge to our efforts to understand how biological processes can be managed in agricultural systems. Soil microbial communities are arguably the most diverse communities on earth, and the factors that determine this extraordinarily high diversity are not well understood (Caldwell et al. 1997). Torsvick et al. (1994) have provided evidence that in one gram of soil there are billions of individual organisms and thousands of species. What are the ecological consequences of such high diversity at such a small spatial scale? And how does this change across the range of scales that we consider to be important for other organisms (e.g. plants and consumers) and biogeochemical processes? To determine how to manage the biological processes controlled by soil microbes, it is important to understand the patterns, causes, and consequences of microbial diversity and the scale at which microbial communities are structured. Understanding the link between the scale at which the microbial community is structured and the scale at which ecosystem processes occur may itself tell us a great deal about the role of microbial diversity in ecosystem functioning.

The high spatial heterogeneity of soil in an ecological context is well documented (Robertson and Gross 1994, Paul and Clark 1996). Differences among habitats in the degree of soil heterogeneity may influence the diversity of microbes that occur there and their function (Gross et al. 1995). For example, our results with nitrifiers and soil C dynamics are best interpreted in relation to differences among treatments in soil heterogeneity (reflecting the availability of microhabitats) and soil organic fractions (reflecting resource heterogeneity; Paul et al. 1998a). Spatial heterogeneity in soil microbial communities occurs at a broad range of scales, from soil particles (e.g. soil macroaggregates), to plant rhizospheres, to field plots, and to the ecosystem and global levels (Tiedje 1994).

Old field with trees.jpg (77705 bytes)At KBS we have documented that there is spatially-structured dependence in microbial processes at both a macro- (e.g 10's of meters, Robertson et al.1997) and micro- (cm, Cavigelli et al. 1995) scale. We have shown that microbial activity (measured by short-term microbial respiration) varies among and within plant communities; in some sites samples taken only centimeters apart vary by a factor of >2. The among-community scale component of this nested variation may be attributable to differences in primary productivity and soil physical properties (e.g. depth to the Bt horizon). At the within-community scales, the doubling of microbial activity may be attributable to the distance to the nearest plant. However, we suspect that these differences may also be due to heterogeneity in soil structure, leading to discontinuous resource availability at the millimeter-scale. This small-scale heterogeneity may be driven by the interaction of plant-derived substrates, such as roots and decaying plant particles, and within-aggregate habitats differences due to clay content, pore sizes, and aeration.

To date, our investigations of soil microbial communities have primarily concentrated on the level and pattern of microbial diversity among the different  plant communities that occur on the KBS LTER site. These communities range from the intensively managed row crops under different input intensities to native communities at different successional stages. Our results, generated by a variety of phenotypic and genetic approaches, have documented differences in the apparent diversity of whole-soil microbial communities (patterns of bacterial fatty acids, FAMEs), as well as differences in the diversity of key functional groups, notably denitrifiers (Cavigelli 1998) and nitrifiers (Bruns et al. 1998).

microbial analysis strategies.GIF (53844 bytes)In the past decade we have concentrated on documenting the level and patterns of microbial diversity among ecosystems using strategies such as those outlined in the figure at right. We have now begun more intense investigations of the regulation, maintenance, and consequences of microbial community structure. We hypothesize that the majority of soil microbial diversity is driven by the heterogeneous distribution of resources and habitats in soil. For example, we have found a variety of autotrophic nitrifier genera in our never-tilled successional plots (Bruns et al. 1998), all of which grow in the laboratory only at low NH4+ concentrations. In contrast, in our agronomic plots there is a single dominant genus (Nitrosomonas) that is able to grow across a wider range of NH4+ concentrations. These differences in nitrifier diversity could be due to differences in resource availability, and therefore competitive interactions.  However, this pattern may also be due to greater diversity of protective soil habitats in the never-tilled community.     Heterogeneity in soil structure, which may lead to higher levels of microbial diversity, is affected not only by cultivation regime but also by the presence and activity of plants that create biopores and habitat for the mesofauna that are directly responsible for much of the soil structure (Oades 1993).

To improve our knowledge of how microbial community structure interacts with the functioning of ecosystems we must obtain a more quantitative knowledge of the interaction between microbes, plant residues and disturbance, at a variety of spatial scales. This will require examining the availability of specific resources (at the substrate level) across multiple spatial scales.

We are currently concentrating our research on soil microbial communities at the KBS LTER in three areas:

blue ball.gif (632 bytes) Investigations of the availability of microbial resources (especially substrates) through continued studies on the pools and fluxes of soil organic matter;
blue ball.gif (632 bytes) Examination of the scales at which carbon turns over in soils from the microaggregate (mm) to the landscape (km); and
blue ball.gif (632 bytes) Investigations of the diversity and structure of specific groups of soil microbes across the 11 different communities on the KBS LTER, with an initial emphasis on Basidiomycete fungi, a microbial group that is responsible for significant carbon turnover in soil.
blue ball.gif (632 bytes) Investigations of the linkage between plant diversity, disturbance, soil structure, microbial diversity, and key ecosystem functions such as primary productivity, nitrogen cycling, and nutrient retention in general.

 

Analytical Procedures Used for Microbial Ecology at KBS

Microbial Biomass
Microbial biomass is enumerated at KBS using the chloroform incubation technique calibrated with direct microscopy.  Specific techniques are documented in Howarth et al. (1994, 1996) and Paul et al. (1998). Results are available on the KBS LTER web site.

Fungal Biomass and Fungal: Bacterial Ratios
Ergosterol is a steroid found in most fungi, but absent in other microorganisms. We have found that the concentration of ergosterol in soils (Stahl and Parkin 1996) is directly related to the growth rate of fungi and provides an estimate of the fungal biomass in soil. Comparisons of fungal and bacterial ratios, and the size of bacterial biomass are also useful for documenting changes within soil microbial communities. Computerized fluorescence microscopy has greatly aided our ability to examine these characteristics.

microbes on slide.jpg (74681 bytes)Culturable Microorganisms
During the establishment of our main cropping systems site a culture collection of bacteria was established to provide a benchmark collection. From over 1000 isolates a 100-isolate subset was selected for intensive study ("the KBS 100"). These isolates have been characterized using a variety of polyphasic taxonomic tools (see figure above) and are maintained as a long-term reference collection. Additional collections include lignin decomposing Basidiomycetes from the site (molecular techniques show that many have previously not been described; Thorn et al. 1996) as well as collections of denitrifiers (Cavigelli 1998) and nitrifiers (Bruns 1996).

Non-Culturable Microbes: Community-Level Signatures
We have used a variety of phenotypic tools to characterize soil microbial community composition as related to ecological change (Klug and Tiedje 1993, Sinsabaugh et al. 1998). These include fatty acid methyl ester (FAME) and phospholipid fatty acid (PLFA) analyses (Peterson and Klug 1994, Haack et al. 1994, Cavigelli et al. 1995, Corlew-Newman and Klug 1998), as well as Biolog™ carbon utilization signatures. We are also using G+C analysis to examine the distributions of low G+C populations (e.g. Pseudomonas) vs. high G+C populations (e.g. Arthrobacter), and L-asparaginase activity to resolve differences in rhizosphere populations.

Population-Level Signatures: Gene Probes
We have collaborated with the NSF Center for Microbial Ecology (CME) at MSU in the development and testing of several gene probes for assaying specific soil populations at KBS. Particularly successful has been the deployment of probes for 2,4-D metabolism (Holben et al. 1992, Ka et al. 1994a,b,c,d, 1995), and for nitrifying bacteria (Zhou et al. 1995, Bruns 1996, Bruns et al. 1998). We are beginning to design population-specific rRNA oligonucleotide probes to determine the contribution of these various fractions of rRNA to total prokaryotic community rRNA. The advantage of working with RNA is that it allows detection of the most active (highest ribosome content) populations, which are also probably the most dominant populations.

Population-Level Signatures: Phenotypic Techniques
We have used lipid analysis (fatty acid markers) to track changes in fungal communities in different soils (Stahl and Klug 1996, 1998, Stahl et al. 1998), changes in mycorrhizal associations (Calderon 1997), and differences in denitrifier community composition (Cavigelli 1998). These techniques have been combined with techniques for culturable microbes and community-level signatures (above).

Bacterial Growth Rates
Microbial biomass provides an estimate of the pool size of microorganisms, but not of biomass turnover. We have examined bacterial turnover dynamics using  3H, thymidine, and 14C-leucine incorporation kinetics (Harris 1994, Harris and Paul 1994).

Microbial Process Measurements
Measurements of key microbial processes such as nitrification, carbon mineralization, and carbon and nitrogen gas fluxes are coupled to those of microbial and plant community structure to provide insight into the functional significance of microbial diversity at KBS. Processes examined include CH4 oxidation and N2O production (Robertson 1993, Paustian et al. 1995, Ambus and Robertson 1998a,b), carbon oxidation (Paul et al. 1994, 1998a,b, Paustian et al. 1995), denitrification (Cavigelli 1998), and nitrification (Bruns 1996, Knoke 1997).

Microbial Predators
Nematodes are important fungal and bacterial consumers that can affect the distribution and abundance of microbial populations. We have examined changes in nematode groups among cropping system treatments (Freckman and Ettema 1993) as well as the distribution of various nematode trophic groups  (Robertson and Freckman 1995). These studies, in combination with our data on biomass and biomass turnover measurements, provide evidence on the controls in the distribution of key microbial groups in soils.


References

Ambus, P., and G.P. Robertson. 1998. Automated near-continuous measurement of CO2 and N2O fluxes with a photoacoustic infra-red spectrometer and flow-through soil cover boxes. Soil Science Society of America Journal 62:394-400.

Ambus, P., and G.P. Robertson. 1998. Fluxes of CH4 and N2O from Poplar stands grown under ambient and twice-ambient CO2. Atmospheric Environment (Submitted).

Bruns, M. 1996. Nucleic acid probe analysis of autotrophic ammonia-oxidizer populations in soils. Ph.D. Dissertation, Michigan State University, East Lansing, Michigan.

Bruns, M.A., J.A. Fries, J.M. Tiedje, and E.A. Paul. 1998. Functional gene diversity among terrestrial ammonia oxidizing bacteria. Applied Environmental Microbiology (Submitted).

Calderon, F. 1997. Lipids: Their value as molecular markers and their role in the carbon cycle of arbuscular mycorrihizae. Ph.D. Dissertation, Michigan State University, East Lansing, Michigan.

Caldwell, D.E., G.M. Wolfaardt, D.R. Korber, and J.R. Lawrence. 1997. Do bacterial communities transcend Darwinism? Advances in Microbial Ecology 15: 105-191

Cavigelli, M.A., G.P. Robertson, and M.J. Klug. 1995. Fatty acid methyl ester (FAME) profiles as measures of soil microbial community structure. Pages 99-113 in H.P. Collins, G.P. Robertson, and M.J. Klug, eds. The Significance and Regulation of Soil Biodiversity. Plant and Soil 170. Kluwer Academic Publishing, Dordrecht, Netherlands.

Cavigelli, M. 1998. Ecosystem consequences and spatial variability of microbial soil community structure. Ph.D. Thesis, Michigan State University, East Lansing, Michigan.

Cavigelli, M. A., G. P. Robertson, and M. J. Klug. 1995. Fatty acid methyl ester (FAME) profiles as measures of soil microbial community structure. Pages 99-113 in H. P. Collins, G. P. Robertson, and M. J. Klug, eds. The Significance and Regulation of Soil Biodiversity. Plant and Soil 170. Kluwer Academic Publishers, Dordrecht, Netherlands.

Corlew-Newman, H.L., and M.J. Klug. 1998. Comparison of community and marker bacterial fatty acid profiles for different agricultural management treatments. Plant and Soil. (Submitted).

Collins, H. P., G. P. Robertson, and M. J. Klug, eds. 1995. The Significance and Regulation of Soil Biodiversity. Kluwer Academic Publishers, Dordrecht, The Netherlands. Also published as Plant and Soil 170:1-241.

Freckman, D.W. and C.H. Ettema. 1993. Assessing nematode communities in agroecosystems of varying human intervention. Agriculture, Ecosystems and Envrionment 45:239-261.

Haack, S.K., H. Garchow, D.A. Odelson, L.J. Forney and M.J. Klug. 1994. Microbial community analysis: accuracy, reproducibility and interpretation of fatty acid methyl ester profiles from model bacterial communities. Applied and Environmental Microbiology 60:2483-2493.

Harris, D. 1994. Analyses of DNA extracted from microbial communities. Pages 111-118 in K. Ritz, J. Dighton, and K. Giller, eds. Beyond the Biomass. John Wiley & Sons, Chichester, England.

Harris, D., and E.A. Paul. 1994. Measurement of microbial growth rates in soil. Applied Soil Ecology 1:277-290.

Holben, W.E., B.M. Schroeder, V.G.M. Calabrese, R.H. Olsen, J.K. Kukor, V.O. Biederbeck, A.E. Smith, and J.M. Tiedje. 1992. Gene probe analysis of soil microbial populations selected by amendment with 2,4-dichlorophenoxyacetic acid (2,4-D). Applied Environment Microbiology 58:3941-3948.

Horwath, W.R., and E.A. Paul. 1994. Microbial biomass. Pages 753-774 in R.W. Weaver, J.S. Angle, P.J. Bottomley, D.F. Bezdicek, M.S. Smith, M.A. Tabatabai, and A.G. Wollum, eds. Methods of Soil Analysis Part 2-Microbiological and Biochemical Properties. Soil Science Society of America, Madison, Wisconsin, USA.

Horwath, W.R., E.A. Paul, D. Harris, J. Norton, L. Jagger, and K.A. Horton. 1996. Defining a realistic control for the chloroform-fumigation incubation method using microscopic counting and 14C-substrates. Can. J. Soil Sci. 96:459-467.

Ka, J.O., P. Burauel, J.A. Bronson, W.E. Holben, and J.M. Tiedje. 1995. DNA probe analysis of microbial community selected in field by long-term 2,4-D application. Soil Science Society of America Journal 59:1581-1587.

Ka, J.O., W.E. Holben, and J.M. Tiedje. 1994. Analysis of competition in soil among 2,4-D degrading bacteria. Applied and Environmental Microbiology 60:1121-1128.

Ka, J.O., W.E. Holben, and J.M. Tiedje. 1994. Genetic and phenotypic diversity of 2,4-D degrading bacteria isolated from 2,4-D treated field soils. Applied and Environmental Microbiology 60:1106-1115.

Ka, J.O., W.E. Holben, and J.M. Tiedje. 1994. Integration and excision of a 2,4-dichlorophenoxyacetate acid-degradative plasmid in alcaligenes paradoxus and evidence of its natural intergeneric transfer. Journal Bacteriology 176:5284-5289.

Ka, J.O., W.E. Holben, and J.M. Tiedje. 1994. Use of gene probes to aid recovery and identification of functionally dominant 2,4-D degrading populations in soil. Applied and Environmental Microbiology 60:1116-1120.

Klug, M.J. and J.M. Tiedje. 1993. Response of microbial communities to changing environmental conditions: chemical and physiological approaches. Pages 371-374 in R. Guerrero and C. Pedros-Alio, eds. Trends in Microbial Ecology, Spanish Society for Microbiology, Barcelona, Spain.

Knoke, K.E. 1997. Assessment of the origin and fate of nitrate from soil lysimeters using stable nitrogen isotopes. M.Sc. Thesis, Michigan State University, East Lansing, Michigan.

Oades, J. M. 1993. The role of biology in the formation, stabilization and degradation of soil structure. Geoderma 56: 377-400.

Paul, E.A. and F.E. Clark.  1996. Soil Microbiology and Biochemistry. 2nd edition. Academic Press, Inc., San Diego, CA. 340 pp

Paul, E.A., D. Harris, M. Klug, and R. Ruess. 1998. The determination of microbial biomass. In G.P. Robertson, D.C. Coleman, C.S. Bledsoe, and P. Sollins, eds. Standard Soil Methods for Long-Term Ecological Research, Oxford University Press, New York (In press).

Paul, E.A., H.P. Collins, D. Harris, U. Schulthess, and G.P. Robertson. 1998. The influence of biological management inputs on carbon mineralization in ecosystems. Applied Soil Ecology 327: 1-13.

Paul, E.A., E.T. Elliott, C.V. Cole and K. Paustian (eds.). 1994. Soil Organic Matter Dynamics in Agroecosystems. Lewis CRC Publishers, Boca Raton, Florida. 500 pp.

Paustian, K., G.P. Robertson, and E.T. Elliott. 1995. Management impacts on carbon storage and gas fluxes (CO2, CH4) in mid-latitude cropland ecosystems. Pages 69-84 in R. Lal, J. Kimble, E. Levine, and B.A. Stewart, eds. Soil Management and the Greenhouse Effect, Advances in Soil Science. CRC Press, Boca Raton, Florida.

Paustian, K., H.P. Collins, and E.A. Paul. 1997. Management controls on soil carbon. Pages 15-49 in E.A. Paul, K. Paustian, E.T. Elliot and C.V. Cole, eds. Soil Organic Matter in Temperate Agroecosystems: Long-Term Experiments in North America. CRC Press, Boca Raton, Florida, USA.

Peterson, S.O. and M.J. Klug. 1994. Effects of sieving, storage and incubation temperature on the phospholipid fatty acid profile of a soil microbial community. Applied and Environmental Microbiology 60:2421-2430.

Robertson, G.P. 1993. Fluxes of nitrous oxide and other nitrogen trace gases from intensively managed landscapes: a global perspective. Pages 95-108 in L.A. Harper, A.R. Mosier, J.M. Duxbury, and D.E. Rolston, eds. Agricultural Ecosystem Effects on Trace Gases and Global Climate Change. American Society of Agronomy, Madison, Wisconsin, USA.

Robertson, G.P., and D.W. Freckman. 1995. The spatial distribution of nematode trophic groups across a cultivated ecosystem. Ecology 76:1425-1432.

Robertson, G. P. and E.A. Paul. 1998. Ecological research in agricultural ecosystems: contributions to ecosystem science and to the management of agronomic resources. In M. L. Pace and P. M. Groffman, eds. Successes, Limitations and Frontiers in Ecosystem Science. Cary Conference VII, Springer-Verlag, New York (In press).

Robertson, G. P., K. M. Klingensmith, M. J. Klug, E. A. Paul, J. C. Crum, and B. G. Ellis. 1997. Soil resources, microbial activity, and primary production across an agricultural ecosystem. Ecological Applications 7: 158-170.

Sinsabaugh, R.L., M.J. Klug, H.P. Collins, P.E. Yeager, and S. O. Peterson. 1998. Characterizing soil microbial communities. In G.P. Robertson, C.S. Bledsoe, D.C. Coleman, and P. Sollins, eds. Standard Soil Methods for Long-Term Ecological Research, Oxford University Press, New York (In press).

Stahl, P.D., and T.B. Parkin. 1996. Relationship of soil ergosterol content and fungal biomass. Soil Biology and Biochemistry 28:847-855.

Stahl, P.D., and M.J. Klug. 1998. Lipid comparisons of microfungal communities from soils and on different agricultural management practices. Plant and Soil (In press).

Stahl, P.D., and M.J. Klug. 1996. Characterization and differentiation of filamentous fungi based on fatty acid composition. Applied and Environmental Microbiology 62:4136-4146.

Stahl, P.D., M.J. Klug, and G.P. Robertson. 1998. Soil microfungal diversity under three management regimes. Soil Biology and Biochemistry (submitted).

Tiedje, J.M. 1994. Approaches to the comprehensive evaluation of procaryote diversity of a habitat. In: D.Allsopp, R.R. Colwell, and D.L. Hawksworth (eds) Microbial Diversity and Ecosystem Function, CAB International,Wallingford, U.K.

Torsvik, V., J. Goksoyr, F.L. Daae, R. Sorheim, J. Michelsen, and K. Salte. 1994. Use of DNA analysis to determine the diversity of microbial communities. In: Beyond the Biomass. K. Ritz, J. Dighton, and K.E. Diller (eds.) Wiley Sayce, London, England. pp. 39-48

Zhou, J., M.A. Bruns, and J.M. Tiedje. 1995. Rapid method for recovery of DNA from soils of diverse composition. Applied and Environmental Microbiology 62:316-322.

 

KNZ - Konza Prairie Research Natural Area, Manhattan, Kansas

 

David C. Hartnett [hartnett@lter-konza.konza.ksu.edu]

Walter Dodds [wdodds@lter-konza.konza.ksu.edu]

Chuck Rice [crice@lter-konza.konza.ksu.edu]

The research at Konza Prairie focus on several aspects of microbial ecology. This includes investigation of microbial biomass and activity as affected by fire, water, and nutrients on Konza tallgrass prairie LTER, microbial responses to elevated CO2 on the site, and using Konza Prairie as a benchmark for biological aspects of soil quality.

A major focus has been the ecology and diversity of denitrifiers in the surface and subsurface profiles of tallgrass prairie in comparison with agricultural ecosystems. We have used both molecular techniques and physiological techniques to assess denitrifier diversity, and the results of a recent study on microbial enzymes is now in review.

 

Aquatic Microbial Ecology:                               

We do not monitor aquatic microbes regularly at Konza, but have done some work on this previously. This work includes two main projects on groundwater and several independent projects on periphyton.

The groundwater work involved monitoring bacterial numbers flowing from a spring for two years, and relating those to groundwater invertebrates (Edler, C. and W. K. Dodds 1996. The ecology of a subterranean isopod, Caecidotea tridentata. Freshwater Biol. 35:249-259). A second groundwater project was to compare nitrogen cycling in shallow subsurface below cropland and prairie fields (several references with a basic description in Dodds, W. K. et al. 1996. Biological properties of soil and subsurface sediments under abandoned pasture and cropland. Soil Biol. Biochem. 7:837-846.)

If funds were available for observing (monitoring) aquatic microbes at Konza LTER, the sampling could be prioritized as follows. 1) Algal biomass in streams, 2) Bacterial counts at several groundwater sites, 3) activity measurements of specific groups (e.g. denitrifiers and nitrifiers) in stream and groundwater, and 4) monitoring coliform and giardia from pristine sites.

These initiatives would be based upon using Konza as a pristine baseline for water quality of prairie streams, and helping describe ecosystem function of autotrophic streams. We need data to assess human effects on streams. Even though streams draining grasslands and wooded grasslands are common worldwide, little is known about their ecology.

 

Research on Arbuscular Mycorrhizal Fungi:

1. Mycorrhizal fungal community composition: Spatial and temporal patterns of mycorrhizal fungal community composition and diversity are assessed via AM fungal spore isolation from soil subjected to various treatments, including burning, mowing, phosphorus fertilization, nitrogen fertilization, grazing, and fungicide application (mycorrhizal suppression). In addition, field experiments and greenhouse trap culture experiments are evaluating patterns of host-plant specificity of mycorrhizal fungal species.

Representative Publications:

 

2. Assessment of interspecific variation in mycorrhizal colonization among tallgrass prairie grasses and forbs:

Representative Publications:

 

3. Effects of mycorrhizae on plant population dynamics and community structure:

Ongoing field studies are evaluating the influence of mycorrhizal fungi on plant demography, competitive relationships, and species composition and diversity.

Representative Publications:

 

4. Assessment of AM fungal nutrient uptake in tallgrass prairie:

Representative Publications:

 

Other Related Publications:

Groffman, P. M., C. W. Rice, and J. M. Tiedje. 1993. Denitrificationin a tallgrass prairie landscape. Ecology 74:855-862.

Rice, C. W., and F. O. Garcia. 1994. Biologically active pools of soil C and N in tallgrass prairie. pp. 201- 208. In J. Doran et al. (ed) Defining soil quality for a sustainable environment. Spec. Pub.No. 35. Soil Sci. Soc. Am., Madison, WI.

Garcia, F. O., and C. W. Rice. 1994. Microbial biomass dynamics in tallgrass prairie. Soil Sci. Soc. Am. J. 58:816-823.

Rice, C.W., F.O. Garcia, C.O. Hampton, and C.E. Owensby. 1994. Soil microbial response in tallgrass prairie exposed to increased levels of atmospheric CO2. Plant Soil 165:67-74.

Zak, D.R., D. Tilman, R.R. Parmenter, C.W. Rice, F.M. Fisher, J. Vose, D. Milchunas, and C.W. Martin. 1994. Plant production and the biomass of soil microorganisms in late-successional ecosystems: A continental-scale study. Ecology 75:2333-2347.

Noll, M.C., C.J. Sorenson, and C.W. Rice. 1995. Biological condition of a Midwest soil after six years of conversation reserve. Trans. Kansas Academy of Sci. 98(3-4):102-112.

Dodds, W.D., M.K. Banks, C.S. Clennan, C.W. Rice, D. Sotomayor, E. Strauss, and W. Yu. 1996. Biological properties of soil and subsurface sediments under grassland and cultivation. Soil Biol. Biochem. 28:837-846.

Sotomayor, D., and C.W. Rice. 1996. Denitrification beneath grassland and cultivated soils. Soil Sci. Soc. Am. J. 60:1822-1828.

Rice, C.W., T. Moorman, and M. Beare. 1996. Role of microbial biomass C and N in soil quality. p. 203- 215. In J.W. Doran and A.J. Jones (eds) Methods for assessment of soil quality. Spec. Pub. No. Soil Sci. Soc. Am., Madison, WI

 

 

 

LUQ- Luquillo Experimental Forest, near San Juan, Puerto Rico

Jean Lodge [/S=D.J.LODGE/OU1=S32A@mhs-fswa.attmail.com]

In addition to the LTER researchers, there are two microbial ecologists (Dr. Gary Toranzo & Dr. Arturo Massol) and three mycologists (Drs. Paul Bayman, Steve Rehner & Carlos Betancourt) at the University of Puerto Rico who have contributed to or are planning on initiating studies in the Luquillo forest. They greatly enhance our critical mass and capabilities in microbial ecology research at the LUQ-LTER site.

D.J. Lodge (1993) monitored total fungal biovolume at two-week intervals for one year at El Verde (LUQ-LTER). Fungal biovolume in the litter layer could triple in two weeks in response to frequent rains, and decrease by half during the following two weeks in response to drying cycles. The critical factor forfungal volume in litter was frequency of days with rainfall exceeding 3 mm (sufficient to wet the canopy and reach the forest floor) rather than total rainfall during the previous two weeks. Three consecutive days without rainfall reaching the forest floor was sufficient to dry out the thin litter layer and cause massive crashes in fungal mycelia. Soils were more buffered against fluctuations in rainfall, and six weeks of low rainfall was requiredto reduce the soil moisture sufficiently to cause crashes in the fungal populations in soil. Fungal nutrient stocks in the litter layer were calculated based on the biovolume estimates above and fungal specificdensities and nutrient concentrations obtained from field-collected mycelia at the site (Lodge 1987). Fungal stores represented a large fraction of the potassium and phosphorus in the litter layer, but not much of the nitrogen (Lodge 1993). Pre-fruiting mycelia were found to concentrate so much phosphorus through short-circuited nutrient cycling (translocation of phosphorus from partly decomposed litter to form biomass in fresh leaf litter with low phosphorus content) that they accounted for virtually all of the phosphorus in the litter in some patches of forest floor.

We hypothesize that nutrients are released in pulses from the litter layer, and that nutrients released in pulses are more available to higher plants than if they had been released at a constant rate (Lodge, McDowell & McSwiney 1994). Pulsed nutrient release should result from fungal accumulation and internal recycling of nutrients and subsequent crashes in fungal populations in response to drying cycles, followed by rains that then leach the released nutrients into the soil. Trees cannot compete well against decomposer microorganisms for limiting nutrients if there is an ample supply of labile carbon for the microbes, as demonstrated by a post-hurricane litter removal/fertilization experiment conducted at El Verde (Zimmerman et al. 1995). However, the release of concentrated pulses of nutrients that are synchronized with crashes in microbial biomass should allow windows of opportunity for

plant roots to obtain nutrients under reduced microbial competition (Lodge, McDowell & McSwiney 1994). Microbial nitrogen retention during dry- wet cycles in subtropical evergreen forests was compared between the LEF and southern China by Fu, Zou, Lugo, Yi and Ding (in press). Rootlike structures of basidiomycete fungi that are used to translocate nutrients and colonize new resources in the litter layer also hold litter together in mats which prevents it from being exported down steep slopes (Lodge & Asbury 1988). In addition to preventing the loss of nutrients contained in leaf litter from being exported by streams during heavy rains, the retention of leaf litter on steep slopes apparently protects the soil surface from erosion by pounding rain thereby conserving soil nutrients.

The relationship between soil development, earthworm density and both active and total fungal and bacterial biomass were studied in landslide and adjacent forest soils by Li, Xou and Myster (in press). They found that earthworm density and biomass was positively correlated with total bacterial biomass,leaf litter, and the light-carbon fraction of the soil organic matter pool.

Effects of carbon and phosphorus additions on loss of nitrogen as nitrate and total soil respiration predominantly microbial) in a simulation of post hurricane conditions - (McDowell, Lodge & Massol, in prep.)

 

 

Surveys, Inventories and other Microbial Diversity Studies:

Basidiomycetes of the Greater Antilles, Especially the Luquillo LTER Site: This is a 4-year program funded by NSF Biotic Surveys & Inventories, and is approximately at the half-way point. A monograph of Alboleptonia by T.J. Baroni & D.J. Lodge is in press, as well as a paper describing a new genus,

`Macrocybe' to accommodate a species found in Puerto Rico and other species in the tropics (D.N. Pegler, D.J. Lodge & K.K. Nakasone). Another paper on Calocybe species is also in press (T.J. Baroni, R. Vilgalys, D.J. Lodge & N.V. Legon). A monograph on the Hymenochetaceae and Ganodermataceae by Leif Ryvarden is ready to go to press. Monographs nearing completion are Puteaceae, by E. Horak, and Hygrophoraceae by S.A. Cantrell & D.J. Lodge.

 

Previous fungal surveys:

All historical and recent records of fungi identified from the Luquillo LTER site through 1995 were published by Lodge (1996) according to their substrate and habit in the appendix of the chapter on microorganisms in the book, The Food Web of a Tropical Rain Forest. A review of literature on fungi of Puerto Rico and the Virgin Islands was also published in 1996 by Lodge.

 

Factors important in structuring fungal communities and species diversity

Microfungi in decomposing leaves:

Several hundred species of microfungi were cultured and identified from decomposing leaf litter of two tree species that occurred together at two sites at El Verde in the Luquillo forest (Polishook, Bills & Lodge, 1996). A list of species was included in the publication. As we hypothesized based on studies at the same site in the 1960's, there is a strong preference among microfungal species for different types of leaves. Species composition of fungi isolated from the same leaf species in two widely separated plots was more similar than fungal species composition in leaf litter of different species at the same site.

 

Slime Molds:

Drs. Steven Stephensen and John Landolt cultured slime molds from leaf litter in the 16-ha gridded forest site at El Verde LUQ-LTER site. They found a higher diversity in the part of the grid that was more highly disturbed by clearcutting and agriculture over sixty years previously as compared to the little-disturbed forest in the other part of the grid. This corresponds to higher densities of their prey items, bacteria, in the more disturbed part of the grid (see Willig et al., bacterial functional diversity below). There were also difference in litter entrained in the understory. This led to further studies on canopy slime mold communities along the elevational gradient in the LUQ-LEF, and the degree of isolation of canopy litter communities and modes of slime mold dispersal.

 

Stream fungi and algae:

In the 1960's, Padget identified fungi on decomposing leaves in a stream at El Verde. Recently, Carlos Santana & Carlos Betancourt (1997) published a survey of spores trapped in foam in streams of Puerto Rico, including the LTER site. Pringle (1996) studied the effect of atyid shrimp on spatial heterogeneity of algal communities in montane streams at the LUQ-LTER site. In another study published by Pringle, Blake, Covich, Busby & Finley in 1993, the sediment removal activity of shrimp was found to increase algal biomass as compared to shrimp exclosure plots.

 

Nitrogen fixing cyanobacteria, lichens, and bacteria associated with plants:

Rates of nitrogen fixation by plant epiphyllous microbes were found to be very high by Joe Edmisten in the 1960's using the acetylene reduction technique. Recently, Dr. Li of the US Forest Service research lab in Corvallis Oregon has been culturing and obtaining molecular sequences of nitrogen fixing bacteria that live INSIDE of plant roots at the LEF. These bacteria are abundant, and there is a high