Analyses of microbial communities at biogeochemical interfaces and their interaction with organic pollutants in soil
Kornelia Smalla, Holger Heuer and Newton Gomes
Biologische Bundesanstalt für Land- und Forstwirtschaft, Institut für Pflanzenvirologie, Mikrobiologie und biologische Sicherheit, Messeweg 11-12, D 38104 Braunschweig
ObjectiveIntroduction
This project aims to explore the composition
Microbes are the major drivers of geochemical and biotransformation processes in soils. Inconcert with the inorganic and organic constituents microbes are supposed to actively
Methodological Approach
This project aims to explore the composition
of the microflora and their horizontal gene
pool in response to phenanthrene and
hexadecane and varying interfaces in soil
in an interdisciplinary effort.
Characteristics of biogeochemical interfaces
affect the composition of colonising microbial
communities and the abundance and diversity
of their mobile genetic elements (MGEs)
Hypotheses
Soils: (A) Cambisol,(B) Luvisol, (C) Chernozem
Soils (A-C) + phenanthrene
Soils (A-C) +hexadecane
Sampling of batch experiments at t0, 21, 62
Batch experiments
Artificial soil (Cambisol)
Cambisol
SamplingPercolation water and 10 x 1 cm slices from the receiving layer
Artificial soil +POP
S
R
The fate and the effects of the POPs is influenced by the soil type because the structural and functional diversity of the microbial communityis dependent on the soil type each consisting of soil particle fractions and interfaces
Cambisol + POP
concert with the inorganic and organic constituents microbes are supposed to activelyshape the architecture of the soil matrix by formation and restructuring of aggregates. Thephysical, chemical and biological heterogeneity of the interfaces in soil is the basis of theenormous extent of structural and functional microbial diversity typically found in soil.Despite its important role microbial life in soil remained largely a “black box” until recentlybecause the tools required to study the structural and functional diversity of soil microbesat the various different interfaces in soil and their interaction with POPs were missing.
In collaboration with Kai Totsche (FSU)
Pre-work done (tool box development)
DNA/RNA acid extraction
DGGE of bacterial and fungal communities (16S/18S)Functional gene arraysDGGE of PCR-amplifieddegradative genesqPCR of degradative genes
Capture of MGEs
Transfer frequencies determinedMolecular characterisation of the MGEs captured
Chemical analysisM. Spiteller
Rsoil particle fractions and interfaces
Different interfaces are formed depending on the mineral and organic matter composition of the soils
DNA/RNA extraction
Recovery of soil interfaces
Biparental mating
Exogenous isolation of MGEs from bacteria colonisingdifferent soil interfaces
Recipient, rifr, gfp+ (P. putida
Cupriavidus necator)
Expression of degradative genes in soilbacteria followed by RT-PCR from mRNA andsubsequent Southern blot hybridisation
References
Two additional populations appeared at 21 dpi
DGGE community fingerprints to identify changes in the relative abundance in response to pollution
A: 99% Burkholderia sp. RP007
B: 98.85% similarity to Nocardia vinacea
ndo-DGGE community fingerprints reveal changes in the relative abundance of degradative genes in
response to POP pollution
Control soil
No naphthalene
Soil +
naphthalene (2mg/g)
0 3 210 3 21
Bacterial communities (Donor) with degradative genes on MGEs
Transconjugants, gfp+, rifr, degradative capacity
(I) soil samples amended with naphthalene(II) soil samples amended with naphthalene and P. putida KT2442 (pNF142) inoculated
GOMES, N. C. M., COSTA, R., SMALLA, K. 2004. Rapid simultaneous extraction of DNA and RNA from bulk and rhizosphere soil. In: G. A. Kowalchuk, F. J. de Bruijn; I. M. Head, A. D. Akkermans, J. D. van Elsas (Eds). Molecular Microbial Ecology Manual. 2nd ed. Dordrecht: Kluwer Academic Publishers, pp 159-169.GOMES, N. C. M., KOSHELEVA, I., ABRAHAM, W-R., SMALLA, K. 2005. Effects of the inoculant strain Pseudomonas putida KT2442 (pNF 142) and of naphthalenecontamination on the soil bacterial community. FEMS Microbiol. Ecol. 54, 21-33.GOMES, N. C. M., BORGES, L., PARANHOS, R., PINTO, F. N., KRÖGERRECKLENFORT, E., MENDONÇA-HAGLER, L. C. S., SMALLA, K. 2007. Diversity of ndo
genes in mangrove sediments exposed to different sources of PAH pollution. Appl. Environ. Microbiol. 2007 (after minor revision accepted for publication)