Bacterial and fungal growth in soil: The effect of temperature and substrate addition

Bacteria and fungi are the main agents in decomposition of soil organic matter. Their activity is determined by the availability and quality of substrate, especially its content of carbon, but also nitrogen and phosphorus. Other environmental factors, which affect the activity of the microbial community in soil, are temperature, moisture and pH. In this thesis the focus was on how temperature and substrate addition affected the growth of the bacterial and fungal communities in soil. To study the effects of environmental factors on bacteria and fungi in soil with enough sensitivity to allow for a high time resolution, it is necessary to measure their growth rate and not only rely on biomass changes. I have used methods to measure the bacterial and fungal growth rate that rely on incorporation of tracer amounts of radioactive substrates during a short time. With these methods I have investigated: (i) how addition of different glucose-C concentrations affects the simultaneous development of total respiration and bacterial and fungal growth, (ii) how different temperatures affect the balance of bacterial and fungal growth when different substrates are available as a carbon source, (iii) the adaptation of bacterial growth and mineralisation to new temperature regimes, (iv) the temperature sensitivity of the bacterial community in a hot desert soil. The main findings of this thesis were: • Respiration and bacterial growth rate did not coincide in their development over time after glucose-C addition. After glucose addition an initial lag period, with increased respiration but constant bacterial growth, was followed by an exponential phase in both, followed by a drastic decrease in respiration and a much slower decrease in bacterial growth. These findings suggest that only during the exponential period the increase in respiration can be used to estimate the bacterial growth rate. • With the addition of increasing glucose-C concentrations to the soil the community responded by changing from mainly bacterial growth to mainly fungal growth on the added substrate. Respiration measurements could not detect this switch between these two microbial groups. • A threshold glucose-C concentration higher than 200 µg per g soil was needed to induce bacterial growth. This was not reflected in the respiration measurements. • By adding between 500 and 1000 µg glucose-C, no exponential phase in respiration was found, and respiration decreased to low values within 24 h. Still bacterial growth was evident, showing that during periods with constant or decreasing respiration, added glucose-C was not only used for maintenance. • Temperature strongly affected the rate of development of bacterial and fungal growth after adding substrate, but there were only minor differences in the relative importance of the two groups at different temperatures. • A two month exposure of soil to a new temperature led to temperature adaptation of the bacterial community when the incubation temperature was above Topt for growth (30 °C). Incubation below 30 °C did not affect the temperature adaptation. • Even though hot desert soil is characterized by high and seasonally fluctuating temperatures, the temperature sensitivity of the bacterial community in the soil did not change between seasons. Periods with warm temperatures were more important in determining the temperature response of the bacterial community than cold periods. By comparing our Tmin and Topt values with a study from a cold area (Antarctica) we were able to suggest a range of possible values of these cardinal temperatures for soil bacterial communities worldwide. These results show that direct growth measurements of the main agents in the soil, bacteria and fungi, are important to make predictions on the potential of climate change. Respiration alone cannot give this information how substrate availability and substrate quality will affect the development of the microbial community in soil and their effect on higher trophic levels of the food web

The effects of glucose loading rates on bacterial and fungal growth in soil

Microbial activity in soil is usually limited by the availability of carbon (C). Adding an easily available C source, like glucose, has therefore been a common approach to study alleviation of resource limitations. Most such studies have relied on respiration to study microbial dynamics, with few following the explicit growth response. We determined the response in bacterial and fungal growth, as well as respiration, to additions of glucose (0.5-32 mg C g(-1) soil) during up to 6 days, using leucine incorporation for bacterial growth and acetate-in-ergosterol incorporation for fungal growth. A concentration of 2 mg glucose-C g(-1) soil, where the fungal contribution appeared to be small, was also studied with a high time resolution. Adding glucose resulted in an initial lag phase of stable respiration and bacterial growth. Bacterial growth was similar to the unamended control, while respiration was 8 fold higher during this period. The 14-h lag phase was followed by an exponential increase for both respiration and bacterial growth, with a similar intrinsic growth rate (mu) of around 0.25 h(-1). After the exponential phase, bacterial growth decreased exponentially. The respiration initially decreased even more rapidly than bacterial growth. At concentrations exceeding 4 mg glucose-C g(-1) the relative stimulation of fungal growth surpassed that of bacteria, with the highest amendment rates, 32 mg C g(-1), resulting in mainly fungal growth. Lower loading rates than 4 mg glucose-C g(-1) appeared to stimulate mainly bacterial growth. (C) 2013 Elsevier Ltd. All rights reserved

A Sediment-Flow-Through-System to investigate the efficiency of the benthic microbial methane filter

As a consequence of sediment compaction at the forearc of subduction zones, fluids ascend in sediments and are discharged in cold seeps. These fluids include the greenhouse gas methane, which is to a larger proportion depleted by anaerobic oxidization of methane (AOM) next to sediment-water interface. This microbial methane filter consists of archaea (methane oxidizers) and bacteria (sulfate reducers) that are living in syntrophic consortia. During AOM, the microbes produce hydrogen sulfide, which feeds chemoautotrophic “seep-organism”. A new sediment-flow-through-system was developed, to investigate the efficiency of AOM, the range of methane turnover rates, as well as key parameters controlling microbial filter. The system includes two different seawater media. The first medium supplies the microbes with sulfate from the top, (transported by diffusion); the second carries methane and is supplied from the bottom by advective transport. Sampling holes, sealed with silicon, allow the measurement of key parameters such as sulfate, sulfide, pH, redox potential, and total alkalinity as well as sediment sub-sampling to determine the abundance, e.g., of methanotrophic microbial groups. In this presentation the technical details of the sediment-flow-through-system will be introduced. Preliminary results from the first experiments with seep sediments and insights into ongoing studies will be provided

Threshold concentration of glucose for bacterial growth in soil

The activity of heterotrophic soil microorganisms is usually limited by the availability and quality of carbon (C). Adding organic substances will thus trigger a microbial response. We studied the response in bacterial growth and respiration after the addition of low amounts of glucose. First we determined if additions of glucose, at concentrations which did not result in an exponential increase in respiration after the lag phase, still stimulated bacterial growth. The second aim was to determine the threshold concentration of glucose needed to induce bacterial growth. Adding glucose-C at 1000 mu g g(-1) soil resulted in an increased respiration rate, which was stable during 12 h, and then decreased without showing any exponential increase in respiration. Bacterial growth, determined as leucine incorporation, did not change compared to an unamended control during the first 12 h, but then increased to levels 5 times higher than in the control. Thus, after the lag phase, a period with increasing bacterial growth, but at the same time decreasing respiration rates, was found. Similar results, but with a more modest increase in bacterial growth, were found using 500 mu g glucose-C g(-1) soil. Adding 50-700 mu g glucose-C g(-1) resulted in increased respiration during 24 h correlating with the addition rate. In contrast, bacterial growth after 24 h was only stimulated by glucose additions >200 mu g C g(-1) soil. Thus, there was a threshold concentration of added substrate for inducing bacterial growth. Below the threshold concentration growth and respiration appear to be uncoupled. (C) 2014 Elsevier Ltd. All rights reserved

Temperature adaptation of bacterial growth and C-14-glucose mineralisation in a laboratory study

Microbial decomposition of soil organic matter (SOM) is the source of most of the terrestrial carbon dioxide emission. Consequently, our ability to predict how climate warming will affect the global carbon (C) budget relies on our understanding of the temperature relationship and adaptability of microbial processes. We exposed soil microcosms to temperatures between 0 and 54 degrees C for 2 months. After this, bacterial growth (leucine incorporation) and functioning (C-14-glucose mineralisation) were estimated at 8 temperatures in the interval 0-54 degrees C to determine temperature relationships and apparent minimum (T-min) and optimum (T-opt) temperatures for growth and mineralisation. We predicted that incubation at temperatures above the initial T-opt for bacteria would select for a warm-adapted community, i.e. a positive shift in T-min and T-opt for bacterial growth, and that this adaptation of the bacterial community would coincide with a similar shift also for their functioning. As anticipated, we found that exposure to temperatures below T-opt did not change the temperature relationship of bacterial growth or mineralisation. Interestingly, T-opt for glucose mineralisation was >20 degrees C higher than that for growth. For bacterial growth, the temperature relationship for the bacterial community was modulated when soils were incubated at temperature above their initial T-opt (approximate to 30 degrees C). This was shown by an increase in T-min of 0.8 degrees C for every 1 degrees C increase in soil temperature, evidencing a shift towards warm-adapted bacteria. Similarly, the Q-10 (15-25 degrees C) for bacterial growth increased at temperature higher than T-opt. We could not detect a corresponding temperature adaptation of the decomposer functioning. We discuss possible underlying reasons for the temperature-responses of bacterial processes. We note that a temperature adaptation will be rapid when exceeding the T-opt, which initially were >20 degrees C higher for glucose mineralisation than growth. This difference could suggest that different responses to warming exposure should be expected for these microbial processes. (C) 2013 Elsevier Ltd. All rights reserved

Temperature sensitivity of bacterial growth in a hot desert soil with large temperature fluctuations

Hot desert ecosystems are characterized by high soil temperatures with large fluctuations annually and diurnally. Thus, one could hypothesize that not only the microbial community would be adapted to high temperatures, but also have a large temperature range conducive for growth. We determined the temperature sensitivity of the soil bacterial community from the Chihuahuan Desert, Big Bend National Park, Texas, USA, using leucine incorporation as a proxy for bacterial growth. Soil samples were taken during early spring and mid-summer from the surface (0-5 cm) and deeper (15-20 cm) soil layers. Mean winter soil temperature preceding the spring samples was 15 degrees C and in summer 36 degrees C at both depths, but with larger amplitude in the top soil than deeper down. T-min was significantly lower in the top 0-5 cm than at 15-20 cm, -1.2 and 0.0 degrees C, respectively. T-opt also was higher in the top soil than deeper down, 42.9 and 41.4 degrees C, respectively, resulting in a larger temperature range for growth (T-opt - T-min) in the top soil reflecting the larger temperature fluctuations in this layer. There were no significant differences in cardinal temperatures for bacterial growth in soils sampled in early spring and mid-summer despite large seasonal differences in temperatures, showing that long periods of colder temperatures was less important than periods of high temperatures as selection pressure for temperature sensitivity. Comparing with earlier results from Antarctic soils (Rinnan et al., 2009), which in contrast represent an extremely low temperature environment, we suggest that the range of temperature cardinal temperatures for soil bacterial communities globally varies from around -15 to 0 degrees C for T-min and 25 to 45 degrees C for T-opt. (C) 2013 Elsevier Ltd. All rights reserved

Community adaptation to temperature explains abrupt soil bacterial community shift along a geothermal gradient on Iceland

Understanding how and why soil microbial communities respond to temperature changes is important for understanding the drivers of microbial distribution and abundance. Studying soil microbe responses to warming is often made difficult by concurrent warming effects on soil and vegetation and by a limited number of warming levels preventing the detection of non-linear effects. A unique area in Iceland, where soil temperatures have recently increased due to geothermic activity, created a stable warming gradient in both grassland (dominated by Agrostis capillaris) and forest (Picea sitchensis) vegetation. By sampling soils which had been subjected to four years of temperature elevation (ambient (MAT 5.2 °C) to +40 °C), we investigated the shape of the response of soil bacterial communities to warming, and their associated community temperature adaptation. We used 16S rRNA amplicon sequencing to profile bacterial communities, and bacterial growth-based assays (3H-Leu incorporation) to characterize community adaptation using a temperature sensitivity index (SI, log (growth at 40 °C/4 °C)). Despite highly dissimilar bacterial community composition between the grassland and forest, they adapted similarly to warming. SI was 0.6 (equivalent to a minimum temperature for growth of between −6 and −7 °C) in both control plots. Both diversity and community composition, as well as SI, showed similar threshold dynamics along the soil temperature gradient. There were no significant changes up to soil warming of 6–9 °C above ambient, beyond which all indices shifted in parallel, with SI increasing from 0.6 to 1.5. The consistency of these responses provide evidence for an important role for temperature as a direct driver of bacterial community shifts along soil temperature gradients