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Fate of microbial biomass in soil
R. Kindler, A. Miltner and M. Kästner
Department of Remediation Research, Centre for Environmental Research, UFZ Leipzig-Halle GmbH, Permoserstr. 15, 04318 Leipzig
Tel. +49 341 235-2051; Fax.+49 341 235-2492; Email: [email protected]
Soil organic matter (SOM) consists of more carbon (C) than the sum of C of plants and microorganisms (MO) on and in soil [1]. However,
the origin of SOM is the biomass. Especially the MO are of particular importance because they are involved in the formation of SOM by
two processes. On the one hand the microbial biomass (MB) metabolizes plant residues and builds up the refractory soil organic matter
and on the other hand they are the source material of SOM to a certain extent.
To investigate the fate of MB we incubated Escherichia coli cells with soil under constant conditions in a reactor. These cells are
 isotopically labeled with 13C to trace each microbial C atom and
 genetically labeled with the lux gene to trace the fate of nucleic acids in soil and the survival of the entire cells.
 1. The soil and the added biomass 
The soil
Temperature 20 °C
“Ewiger Roggenbau Halle (Germany) FYM I”
pH (CaCl2)
6
Corg 19 mg/g
d13C -26,3 ‰
Ntot 1 mg/g
d15N 19,9 ‰
microbial biomass
173 µg biomass C/g (Chloroform-fumigation-extraction)
148 µg biomass C/g (Substrate Induced Respiration)
Humid air with O2
7,5*106 cfu/g (R2A-Agar)
CO2 traps
The biomass
reactor 1
reactor 2
reactor 3
soil only
soil +
soil +
12C
E.coli RFM 443
13C
E.coli RFM 443
TM 73 µg/g
 21 % of natural 13C in soil
and natural microbial biomass (CFE)
number of cells 1,3*108 cells/g
E.coli RFM 443
 2. The biomass in the soil 
Survival of E.coli carrying the lux marker gene
Extraction of plasmidic marker-DNA from E.coli RFM 443 (lux-gene)
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soil with C-labeled E.coli cells
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soil with C-labeled E.coli cells
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The quantification of
extracted
lux-DNA
(BIO101 FastDNA SPIN
Kit for Soil, purified with
polyvinylpolypyrrolidone
[4]) based on PCRs with
different primers [5, 6]
points to a decreasing
amount of microbial
marker DNA in soil. This
way of quantification
does not allow to
determine
an
exact
value, but it allows to
confirm the existence of
the marker DNA after 15
weeks. For quantification, real-time PCR
will be performed.
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cell number [cells/g soil]
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extracted cells/g soil
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soil with C-labeled E.coli cells
lowest and
highest concentration
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soil with C-labeled E.coli cells
lowest and
highest concentration
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10
15
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incubation time [weeks]
incubation time [weeks]
Comparison between the amount of extracted DNA and the microbial
biomass in soil
E.coli cells incubated in soil were extracted with sodium pyrophosphate [2] and
quantified by the most propable number (MPN) method based on bioluminescence [3].
The amount of extractable and active E.coli cells decreased during incubation. After 15
weeks no more E.coli luminescence was detectable. No luminescent bacteria were
extractable from the control soil without addition of E.coli.
 3. Mineralization of the biomass 
About 60 % of the added 13C incorporated in the E.coli were mineralized after 15 weeks
of incubation. Thus 40 % of the added 13C corresponding to 40 % of the added microbial
biomass remained in the soil. The remaining carbon may be bound in lifeless E.coli cells
and residues or have been incorporated into soil microbial biomass or have been
transformed to SOM compounds.
13
CO2 evolved [% of added C]
60
13
UFZ CENTRE FOR ENVIRONMENTAL RESEARCH LEIPZIG-HALLE
Introduction
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soil with C-labeled E.coli cells
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incubation time [weeks]
Acknowledgements
This study was supported by the DFG (SPP 1090).
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The comparison of amount and residence time of extractable marker DNA with the amount
and residence time of extractable and luminescent bacteria showed a smaller value and
shorter detectability of E.coli resulting from the MPN method. There are 3 different
explanations:
1. The bacteria are not extractable or culturable.
2. The bacteria exist as non-living compartiments.
3. The DNA is not associated to cells any more, but occurs in free state or bound to humic
substances.
Conclusions
Microbial C added to soil is only temporarily bound in the original
cells (if E.coli is representative for microbial biomass in general).
Viable (luminescent) E.coli cells declined rapidly and were not
detectable after 15 weeks, whereas DNA could still be extracted
after more than 23 weeks. Thus the genetic information seems to
be preserved longer than the entire cells. The DNA may be
 situated in non-living cells/compartiments,
 transformed into other cells or
 stabilized on humic substances.
The added C has a considerably longer residence time in soil than
the cell components as indicated by the mineralization curve. The
remaining C (ca. 40% of the added C) may be
 assimilated by the natural or remaining in residual (E.coli)
biomass (only temporarily) or
 transformed into SOM and stabilized.
References:
[1] Haider, K., 1999, Z. Pflanzenernährung und Bodenkunde, 162, 363-371
[2] Näveke, R., Tepper, K.P., 1982, Einführung in die mikrobiologischen Methoden, Lehrstuhl für Mikrobiologie der TU Braunschweig
[3] Rozen, Y., et. al., 1998, Chemosphere, 38(3), 633-641
[4] Berthelet, M., et. al., 1996, FEMS Microbiol. Lett. 138, 17-22
[5] Kästner, A., 1996, Forschungsbericht, Untersuchungen ökologischer und methodischer Faktoren beim Nachweis rekombinanter
Mikroorganismen im Boden
[6] Jacobsen, C.S., 1995, Applied and environmental microbiology, 61 (9) 3347-3352