Microbial Life in Soil

1. Microbial Life in Soil Prof. dr. ir. Willy Vestraete Dr. ir. Tom Van de Wiele Laboratory of Microbial Ecology and Technology (LabMET) Faculty of Bioengineering Ghent University LabMET.Ugent.be 1Laboratory of Microbial Ecology and Technology

2. Topics of Discussion The microbial ecosystem in the soil The most common bacterial soil processes The microbial growth The simulation of the microbial transport in the soil The bioavailability of contaminants 2Laboratory of Microbial Ecology and Technology

3. Topics of Discussion The microbial ecosystem in the soil The most common bacterial soil processes The microbial growth The simulation of the microbial transport in the soil The bioavailability of contaminants 3Laboratory of Microbial Ecology and Technology

4. The Microbial Ecosystem • Ecological importance of soil: • The production of biomass (food,…) • The natural biotope for: • Micro-organisms • The plant-communities • The animal world • To filter or to buffer soil contaminants: • By retaining, transforming, neutralizing… 4Laboratory of Microbial Ecology and Technology

5. The Microbial Ecosystem • The interactions between soil and soil-biotic communities 5Laboratory of Microbial Ecology and Technology

6. The Microbial Ecosystem • “The soil represents a set of physical-chemical conditions in which life develops in all diversity.” • Life: complex communities with ten thousand different species of micro-organisms: • Bacteria • Fungi • Protozoa and macro-organisms Micro-aggregates 6Laboratory of Microbial Ecology and Technology

7. The Microbial Ecosystem • The soil biodiversity: 7Laboratory of Microbial Ecology and Technology

8. The Microbial Ecosystem • The microbial biodiversity: • 35.000 different species • 105-108 per gram soil • Great diversity of ‘genetic capacity’ and ‘biological know-how’ • Participant of a ‘food-web’ in the soil, that develops and grows in complexity until a maximally efficient filling in of the soil functions is obtained 8Laboratory of Microbial Ecology and Technology

9. A0 Deposition of organic material A1: much humus Elution of anorganic and organic compounds from the upper layer A2: less humus B1: humus Depositon of compounds B2: iron Mother-material The Microbial Ecosystem • Soil-profile and micro-organisms:micro-organisms contribute to the profile-development by increasing the solubility of the organic and inorganic material Podzol: number of propagules x 103/g 9Laboratory of Microbial Ecology and Technology

10. Topics of Discussion The microbial ecosystem in the soil The most common bacterial soil processes The microbial growth The simulation of the microbial transport in the soil The bioavailability of contaminants 10Laboratory of Microbial Ecology and Technology

11. Bacterial Soil Processes • Soil bacteria are nutritionally exigent, more than one half of the bacteria requires one or more growth factors 11Laboratory of Microbial Ecology and Technology

12. Bacterial Soil Processes • Organo-heterotrophic bacteria:building organic cell-compounds out of organic materialBacillus: amino-acidsClostridium: carbohydrates + amino-acids • Chemo-lithotrophic bacteria (autotrophic):building organic cell-compounds out of chemical reactions with anorganic materialNitrosomonas: NH4+ + 3/2 O2 NO2-+ 2H+ + H2ONitrobacter: NO2- + 1/2 O2  NO3- 12Laboratory of Microbial Ecology and Technology

13. Bacterial Soil Processes • Microbial respiration: oxygen or other compounds act as hydrogen(=electron)-acceptor • Aerobic: O2 • Facultative aerobic: O2, NO3- • Facultative anaerobic: O2, NO3-, organic acceptors • Anaerobic: Fe3+, Mn4+, SO42-, CO2, organic acceptors • Aerobic conditions: Eh > 0, anaerobic or anoxic conditions: Eh < 0 13Laboratory of Microbial Ecology and Technology

14. 0.77 V 0.74 V -0.23 V -0.24 V STANDARD REDUCTION POTENTIALS substrate product H+ e- Aerobic conditions 0.82 V O2 H2O Anaerobic conditions Fe3+ Fe2+ NO3- N2 SO42- H2S CO2 CH4 14Laboratory of Microbial Ecology and Technology

15. Bacterial Soil Processes • The degradation of organic compounds: • Happens through selective enzymes and delivers energy for the microbial metabolism: metabolic degradation • Happens fortuitously by non selective enzymes and delivers no energy for the metabolism: cometabolic degradation • Reaction kinetics:metabolic > cometabolic 15Laboratory of Microbial Ecology and Technology

16. Bacterial Soil Processes • Degradation of biotic organic material:If favorable conditions are present, every compound will be degraded by the micro-organisms, in a quick (DT50: hours-days) or slow way (DT50: months-years), e.g. • Cellulose (Cellovibrio, Aspergillus, Streptomyces)(DT50-aerobically: 3-4-5 months) • Lignin (Basidiomycetes)(DT50-aerobically: 0,5-1y) • Hydrocarbons e.g. aromatic compounds (Bacillus)(DT50-aerobically-monomers: 0,5-1 month)(DT50-anaerobically-polymers: months-years) 16Laboratory of Microbial Ecology and Technology

17. Bacterial Soil Processes • Example: Aerobic cleavage of the aromatic ring of catechol by oxygenase enzymes 17Laboratory of Microbial Ecology and Technology

18. Bacterial Soil Processes • Degradation of xenobiotic organic material:If favorable conditions are present, some compounds will be degraded, other ones are recalcitrant. • The more a xenobiotic compound resembles a biotic one, so much the more it will be recognized by microbial enzymes and be transformed 18Laboratory of Microbial Ecology and Technology

19. Bacterial Soil Processes • Degradation pathways for the pesticide parathion 19Laboratory of Microbial Ecology and Technology

20. Bacterial Soil Processes • Rules of thumb to judge the biodegradability of an unknown aliphatic chemical compound • The C2-C18 chain length is optimal • CC > C=C > C-C • The more branched, the less the biodegradability • Substitution with –OH or –COOH is positive • Substitution with –Cl, –NO2, –SO3H is negative • The more substituents, the stronger the positive or negative effect • The closer the substituents towards the active group, thegreater its influence 20Laboratory of Microbial Ecology and Technology

21. Bacterial Soil Processes • Rules of thumb to judge the biodegradability of an unknown aromatic chemical compound • Substitution: see aliphatic compounds • Para isomers are more biodegradable than ortho, resp. meta isomers. • Poly aromatic compounds are difficult to degrade, e.g. benzopyrenes 21Laboratory of Microbial Ecology and Technology

22. Bacterial Soil Processes • Environmental factors: • A higher microbial diversity increases the degradation-capacity by proto-coöperation • Water-content: optimal ca. 20% • Temperature: factor 1,5-2 for 10°C • Sorption: through sorption processes, compounds are no longer bio-available (see below), e.g. straws slows down the degradation of atrazin. 22Laboratory of Microbial Ecology and Technology

23. Topics of Discussion The microbial ecosystem in the soil The most common bacterial soil processes The microbial growth The simulation of the microbial transport in the soil The bioavailability of contaminants 23Laboratory of Microbial Ecology and Technology

24. Microbial Growth • Growth: increase in the number of cells • Essential: any given cell has finite life span in nature  species maintains only as result of continued growth of the population • Useful in designing methods to control microbial growth 24Laboratory of Microbial Ecology and Technology

25. Microbial growth ☞ Time required for complete growth cycle is highly variable and dependent on nutritional, environmental and genetic factors 20 21 22 23 24 2n 25Laboratory of Microbial Ecology and Technology

26. Microbial growth • Bacterial growth: cells divide into two new cells by binary fission Bacillus subtilis Dividing streptococci 26Laboratory of Microbial Ecology and Technology

27. 600 500 400 Substrate (mg/l) 300 Log10 viable organisms/ml 200 100 0 Microbial growth ☞ Bacterial population growth: typical growth curve ☞ Growth rate: change in cell number or cell mass per unit time 27Laboratory of Microbial Ecology and Technology

28. Microbial Growth • Most information available resulting from controlled laboratory studies using pure cultures of micro-organisms ☞Compare the complexity of growth in a flask and growth in a soil environment. Although we understand growth in a flask quite well, we stil cannot always predict growth in the environment! 28Laboratory of Microbial Ecology and Technology

29. Topics of Discussion The microbial ecosystem in the soil The most common bacterial soil processes The microbial growth The simulation of the microbial transport in the soil The bioavailability of contaminants 29Laboratory of Microbial Ecology and Technology

30. Microbial transport in the soil • The knowledge about bacterial transport in soil is required: • To protect groundwater sources from microbial contamination • To estimate the influence of rainfall on microbial transport in soil • To design sustainable and safe in situ bioremediation techniques(Can the contact between micro-organisms and the contaminants be realized?) 30Laboratory of Microbial Ecology and Technology

31. Microbial transport in the soil • Determined by: • Dispersion(no straight path by diffusion (concentration gradient and Brownian movement) and mechanical mixing) • Advection(transport of non-reactive components at a rate equal to the average velocity of the percolating water) • Sorption(a part of the bacteria will be sorbed onto the soil particles) • Retention(a part of the bacteria will be retained in the pores in the soil) • Microbial die-off • Modeling this transport requires interdisciplinary research (microbiology + hydrogeology) 31Laboratory of Microbial Ecology and Technology

32. Microbial transport in the soil • Example: The modelling of the evolution of the concentration of the anaerobic micro-organism Desulfitobacterium dichloroeliminans strain DCA1 and the contaminant 1,2-dichloroethane in an in situ bioaugmentation strategy by MOCBAC-3D (Prof. L. Lebbe and K. Smith, UGent) Concentration of Desulfitobacterium dichloroeliminans strain DCA1 32Laboratory of Microbial Ecology and Technology

33. Microbial transport in the soil • Example: The modelling of the evolution of the concentration of the anaerobic micro-organism Desulfitobacterium dichloroeliminans strain DCA1 and the contaminant 1,2-dichloroethane in an in situ bioaugmentation strategy by MOCBAC-3D (Prof. L. Lebbe and K. Smith, UGent) Concentration of the contaminant 1,2-DCA 33Laboratory of Microbial Ecology and Technology

34. Topics of Discussion The microbial ecosystem in the soil The most common bacterial soil processes The microbial growth The simulation of the microbial transport in the soil The bioavailability of contaminants 34Laboratory of Microbial Ecology and Technology

35. Bio-availability • Definition: the fraction of the total concentration of a contaminant that will be taken up by the micro-organisms out of the environment • Generally: the bio-availability to the micro-organisms is directly dependent on the solubility of the contaminant in the aqueous phase • Affecting processes: diffusion of the contaminant in the boundary layer, the macro-pores and the micro-pores, physico-chemical interactions with the particle surface and the desorption velocity of the contaminant out of the sediment which is strongly dependent on the particle size and particle density • Consequence: The degradation efficiency of a contaminant will be reduced as much asthe mass transfer is limited to the micro-organism 35Laboratory of Microbial Ecology and Technology

36. Bio-availability • Processes of bio-availability 36Laboratory of Microbial Ecology and Technology

37. Bio-availability 37Laboratory of Microbial Ecology and Technology

38. Bio-availability • Significance of bio-availability: • The mass transfer limits the bio-availability • The endpoint of bioremediation must be related to the matrix • The concentration of a contaminant in a specific soil must be recalculated to the concentration in a ‘standard soil’ to evaluate the contamination extent • Important for legislation: the line must be drawn, but where? (high ‘grey-value’) 38Laboratory of Microbial Ecology and Technology

39. Take-home message • Great diversity in the ecosystem of the soil • Micro-organisms participate in biogeochemical processes and are able to biodegrade a variety of biotic and xenobiotic compounds • Knowledge about the transport of micro-organisms in soil is required for safely designing clean-up strategies • Bio-availability is determined by the mass-transfer of compounds to the micro-organisms, so the endpoint of bioremediation is not absolute 39Laboratory of Microbial Ecology and Technology