Spatial and temporal fluctuations in bacteria, microfauna and mineral nitrogen in response to a nutrient impulse in soil

V.V. Zelenev

Research output: Thesisinternal PhD, WU

Abstract

Fluctuations of bacterial populations can be observed when frequent and sufficiently long series of samples are obtained for direct microscopic or plate counts of bacteria. Such fluctuations in time and space have been observed for both bacteria and other soil inhabitants. These fluctuations of bacterial numbers are especially noticeable after some disturbance of soil such as tillage, drying and rewetting, and substrate addition, for example in the form of fresh plant material. However, very seldom were bacterial fluctuations subjected to proper statistical analysis to detect significant periodical components in the analyzed data (Chapter 1). The phenomenon of wave-like bacterial oscillations was investigated in short-term (1 month) controlled experiments for rhizosphere and bulk soil after substrate input from plant roots and fresh plant debris, respectively. Short-term oscillating dynamics of bacterial populations were simulated in a mechanistic model, which may contribute significantly to our understanding of the reasons and consequences of bacterial oscillations after addition of substrate to soil.



To determine the spatial variation in density of different trophic bacterial groups (copiotrophic and oligotrophic) and carbon sources in the rhizosphere, colony-forming units (CFUs) and soluble total organic carbon (TOC) were quantified along the root from rhizosphere and corresponding bulk soil samples at 2 cm intervals along wheat roots two, three, and four weeks after planting (Chapter 2). There was a moderate rhizosphere effect in one experiment with soil rich in fresh plant debris (1% C in soil), and a very pronounced rhizosphere effect in the second experiment with soil low in organic matter (0.7% C). Wave-like patterns of both trophic groups of bacteria as well as TOC could be discerned along the whole root length (60 or 90 cm). Harmonical analysis revealed significant oscillations in bacterial populations and TOC. TOC concentrations were maximal at the root tip and base and minimal in the middle part of the roots. Populations of copiotrophic and oligotrophic bacteria had two maxima close to the root tip and at the root base, or three maxima close to the tip, in the middle section, and at the root base. Phases and periods of the two trophic groups differed slightly. The location and pattern of the waves in bacterial populations changed progressively from week to week, and was not consistently correlated with TOC concentrations or the location of lateral root formation. Thus, the traditional view that patterns in bacterial numbers along the root directly reflect patterns in exudation and rhizodeposition from several fixed sources along the root may not be true. We attributed the observed wave-like patterns in bacterial populations to bacterial growth and death cycles (due to autolysis or grazing by predators). Considering the root tip as a moving nutrient source, temporal oscillations in bacterial populations at any location where the root tip passed would result in moving waves along the root. This change in concept about bacterial populations in the rhizosphere could have significant implications for plant growth promotion and soil health.



To check the hypothesis that the principal mechanism underlying the wave-like distribution of bacteria along the root is a cycle of growth, death, autolysis, and regrowth of copiotrophic bacteria in response to a moving substrate source (root tip) a simulation model was created (Chapter 3). After transformation of observed spatial data to presumed temporal data based on root growth rates, a simulation model was constructed with the Runge-Kutta integration method to simulate the dynamics of colony-forming bacterial biomass, with relative growth and death rates depending on substrate content so that the rate curves crossed over at a substrate concentration within the range of substrate availability. The original source of substrate was the root tip, supplemented with a background flux (BGF) of substrate from soil organic matter and dead root cells. Dead necromass from bacteria was partially recycled into substrate. This model was named "BACWAVE", standing for 'bacterial waves'. The model generated cyclic dynamics of bacteria, which were translated into traveling spatial waves along a moving nutrient source. Parameter values were estimated from calculated initial substrate concentrations and observed microbial distributions along wheat roots by an iterative optimization method. The kinetic parameter estimates fell in the range of values reported in the literature. The model was validated with an independent data set of bacteria along wheat roots in relatively C-rich soil. Calculated microbial biomass values produced spatial fluctuations similar to those obtained for experimental biomass data derived from colony forming units. Concentrations of readily utilizable substrate (RUS) calculated from biomass dynamics did not mimic measured concentrations of TOC, which consists not only of substrate but also various polymers and humic acids. Thus, a moving impulse of nutrients into soil resulting in cycles of growth and death of bacteria can explain the observed phenomenon of moving bacterial waves along roots. This was the first report of wave-like dynamics of micro-organisms in soil along a root resulting from the interaction of a single organism group with its substrate.



The model "BACWAVE" for wave-like dynamics of copiotrophic bacteria (CB) was extended to include dynamics of oligotrophic bacteria (OB) (Chapter 4). CFUs ofOBand CB along wheat roots (24 samples) in a low C soil were transformed to temporal biomass taking root growth rate and cell sizes into account. Growth rates of both groups of bacteria increased with readily utilizable substrate (RUS) according to Monod equations, but each with their own characteristic parameter values. The death rate of CB decreased monotonically with substrate concentration, while the death rate ofOBfirst decreased and then increased with substrate concentration. Model parameters were estimated from literature and with an iterative optimization method. Initial biomass and kinetic parameters were lower forOBthan for CB, and fell in the range of values in the literature. The model was validated with an independent data set of bacteria along wheat roots in relatively C-rich soil, so that BGF and initial microbial populations were higher, but other model parameters were the same for both data sets. A satisfactory fit was obtained between experimental and modeled data. This is the first rhizosphere model in which oligotrophic bacteria are taken into account.



Several microcosm experiments were carried out to investigate the hypothesis that an impulse of fresh substrate into soil would invoke oscillations in bacterial populations (Chapter 5). Soil bacterial populations, mineral nitrogen content, pH, and redox potential (ROP) were monitored daily for one month after incorporation of clover-grass (CG) plant material in soil. Colony-forming units (CFUs) and direct microscopic counts of FDA-stained and FITC-stained bacteria increased immediately after incorporation of the plant material, dropped within 2 days, and fluctuated thereafter. Harmonics analysis demonstrated that there were significant wave-like fluctuations with three or four significant peaks within one month after incorporation of clover-grass material. Ammonium (NH 4+ )concentrations increased from the start of the experiments until nitrification commenced. Nitrate (NO 3−) concentrations dropped immediately after plant incorporation, and then rose monotonically until the end of the experiments. There were no wave-like fluctuations in NH 4+and NO 3−concentrations, so that bacterial fluctuations could not be attributed to alternating mineral N shortage and sufficiency. pH levels rose and declined with NH 4+levels. ROP dropped shortly before NH 4+concentrations rose, and increased before NH 4+concentrations decreased; there were no regular fluctuations in ROP, so that temporary oxygen shortages may not have been responsible for the observed fluctuations in bacterial populations. Thus, for the first time, regular wave-like dynamics were demonstrated for bacterial populations after perturbation by addition of fresh organic matter to soil, and several potential reasons for the death phase of the fluctuations could be excluded from further consideration.



To elucidate possible reasons for the oscillations in bacterial populations, potential interactions with populations of bacterial predators, in particular bacterial-feeding nematodes (BFN), were investigated (Chapter 6). In two microcosm experiments, soil bacteria (CFU's and microscopic counts of stained bacteria) and nematode populations in 22 families were monitored daily for 25 or 30 days after incorporation of clover+grass (CG) plant material into soil. Soil bacterial populations fluctuated significantly after incorporation of the plant material with 2 peaks within the first week and 3 or 4 smaller peaks thereafter. Total nematodes and BFN populations started to increase in the course of the second week after CG incorporation, but the proportion of BFN increased within one week. Inactive juvenile BFN (dauerlarvae) seemed to be activated after two days (as the percentage of Rhabditidae increased and dauerlarvae decreased), followed by step-wise increases in dauerlarvae every four days, indicating that there was a new generation every four days. There were significant wave-like fluctuations in daily population changes of BFN, but not in total nematode communities, over the duration of these experiments. These fluctuations had similar periods (5 days) as those of bacterial populations, but were shifted about 3 days relatively to the bacterial fluctuations.



In another microcosm experiment, dynamics of bacterial populations were monitored in response to gamma-irradiated plant material added to gamma-irradiated soil mixed with filtered bacterial suspensions and to non-irradiated soil. Gamma-irradiation of soil significantly increased the periods and amplitudes of bacterial oscillations compared to untreated field soil. Nematode populations were decimated in gamma-irradiated soils, but a small number of protozoa were accidentally introduced in the irradiated soil, and may have been partially responsible for the delayed regulation of bacterial growth. We conclude that fluctuations in bacterial populations were not directly related to similar fluctuations in populations of BFN, as expected from classical Lotka-Volterra equations for predator-prey relationships, but were related to changes in growth rates of BFN. An alternation in active and inactive stages in a synchronized predator community after a disturbance could allow periods of bacterial growth alternated with periods of death. Fluctuations in bacterial populations were dampened after a much longer period when the soil fauna was largely eliminated.



Findings of regular oscillations in bacterial populations and in the rate of change in numbers of bacterial predators after addition of fresh organic matter to soil stimulated the development of a simulation model to investigate potential mechanisms of those oscillations, and whether they were initiated by bacteria- substrate interactions or predatory regulation of bacteria (Chapter 7). The model could also be used to investigate mineral nitrogen release during short-term organic matter decomposition. A substrate-based food web model was constructed with 3 plant residue and 5 soil organic matter compartments, 3 trophic groups of bacteria (copiotrophic, oligotrophic and hydrolytic), and two predatory groups (BFN and protozoa). Both carbon and nitrogen flows were modeled. Fluctuations in microbial populations in soil after plant residue incorporation could be reproduced with and without participation of predators. The first two peaks in bacterial numbers were mainly related to bacteria-substrate interactions, while predators (particularly protozoa) influenced bacterial dynamics during later stages of bacterial community development. Oligotrophic bacteria had a stabilizing effect on fluctuations of other trophic groups, and were the main source of nutrients for predators. A peak in soil ammonium occurred within one week after residue incorporation. Nitrate increased sigmoidally after a short lag phase. The final nitrate concentration was primarily determined by bacterial dynamics and to a lesser extent by protozoa and nematodes. This model emphasized the importance of substrate-consumer relations for regulation of populations at different trophic levels and nutrient release from fresh organic matter added to soil.



This research has given insight in potential mechanisms underlying oscillations in populations of soil bacteria and their predators after a disturbance. Despite the advances achieved in this thesis, there are still some problems to be solved. Precise regulation of substrate-consumer interactions and mechanisms that initiate growth and death cycles of soil bacteria have to be investigated in detail. Nevertheless, the "BACWAVE-WEB" model has good potential to predict responses of microbial communities to a disturbance, which could be used to characterize soil health. The model could be expanded to include denitrification and nitrate leaching, so that the extent of N losses after soil disturbance could be predicted.
Original languageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • Wageningen University
Supervisors/Advisors
  • van Bruggen, Ariena, Promotor
  • Semenov, A.M., Co-promotor, External person
Award date19 Mar 2004
Place of PublicationWageningen
Print ISBNs9789058089885
Publication statusPublished - 2004

Keywords

  • soil biology
  • soil fauna
  • microorganisms
  • bacteria
  • biological activity in soil
  • rhizosphere
  • variation
  • oscillation
  • population dynamics
  • population distribution
  • soil organic matter
  • organic wastes
  • nutrients
  • mathematical models
  • simulation models

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