presence
demethylation
and
Degradation
and
(1988).
acetogenic
(b)
in
syringic
(Colberg,
linked photosynthesis,
degradation
extensive
at
in
CH 4 .
subsequent
least
consortia
subunits
A
Lignocellulose
Metabolism
acid,
of
For
general
degradation
3
1988).
bacteria
Degradation
methanogenic
different
has
then
bacterial
vanillic
of
hydration
with
and
but
to
Anaerobic
the
proceeds
reaction
be
(Daniel
the
lignin
of
fermenting
two
situations:
understood
acid,
under
sulfate
monomers
constitutes
subsequent
methoxy
of
bacteria,
scheme
is
by
and
et
of
cleavage
oxygen
rarely
both
reducers,
fermentation al.,
lignin
cinnamic
(a)
bacteria
in
aerobic
1988).
appears
for more
group
preserved
the
by
degradation
terms
moieties
Microbial
of
anaerobic
the
bacterial
monomers
final
metabolism
these
than
coupled
Volker
to
acid
aromatic
and
Introduction
to
of
of
form
end
take
processes
50
7/22/96
to in
aromatic
anaerobic
the
are
Diversity
sulfate
of
marine
form
percent
products
Bruchert
gallic
to
place
benzene
the
substituted
bacteria
degradation
methanogenesis
of
and
most
hydroxy-substituted
syringic
reducers
monomers
involved
acid.
by
and
of
conditions
1996
of
oligomers
ring
demethylation
the
important
terrestrial
this
Gallic
aromatic
global
to
has
acid
(Kaiser
degradation in
3
has
the
been
acid (Colberg,
acetate
is
(Colberg,
carbon
constituents
sediments
been
thought
initial
rings.
by
and
is
outlined
of
then
a
molecules.
suggested
methoxy
Hanselmann,
fixed
sequence
consortium
cleavage
Derivatives
1988).
to
1988);
further
indicating
proceed
by
of
by
(Fig.
Lignin
Colberg
lignin
substitutes
(c)
to
In
of
are
1).
proceed
by
the
ether-
via
of
1982);
CO 2 of
comparing
syntrophic specific prepared
composition
contain
consortium,
accomplished
methanogenic
also
thought
polymeric
Colberg
the
Furthermore,
whether
are
same
degradation
microorganisms
of
where
stoichiometry
decarboxylated
acetate.
aromatic
also
capable
organisms
In
R
isolates
microbial While
that
and responsible
to
whether
indicates
order
relationships
these
structure
enrich
The
incubations
and
white-rot
of
Young
step
monomers
by little
it
cultures,
of
cleaving
of
to
observed
with
is
concentration
to
that
a
that
the
is
these
group
for
sulfate-reducing,
establish
by
single
R-OCH 3
the
is
form
contingent
of
(1985)
for
same
metabolic
known
are
may
sulfate
now
fungi
lignin
substituted
but
reactions
can
the
to of
the
of
responsible
group
sequence
sequentially
well
bacterial
carry
lignin,
organisms
it
showed
whether
production
be
ether-linked
were
+
about
reducers,
in
is
O.75S0 4 2
of
made
upon
not
products established
of
order
out
may
the
responsible
there
organisms
syntrophic
of
known
methanogenic that
(Hanselmann
sulfate
complete
specific
the
by
metabolic for
to
is
degradation
be
acetogens,
of degrade
is
studying
aromatic
capable
release
preceding
the
oligophenols.
—*
obtained
summarized
ring.
remaining
acetate
whether
reducers
that
HCO
initial
degradation
or
for
degradation
relationships
Pyrogallol
products
many these
the
whether
of
the
the
monomers
during
et
and
by
and
products
breakdown
demethylation
metabolizing
+
aromatic
sulfate-reducing
al.,
were
uncertainty
metabolic
initial
O.25H
groups
aromatic
methanogens.
anaerobic
by
acetogenic
1995).
the
were complete
the
steps.
prepared
is
of
lignolytic
in
suggests
cleavage
finally
could
following
of monomers.
+
aromatic
the
used
reaction.
In
compounds,
O.75HS
products
Inferences
organisms.
bacteria
about
different
the
of
bacteria.
degradation
degradation
also
using
to
broken
Differences
the
bacteria
a
presence
step. of
infer
monomers
consortium
their
reaction:
be
It
+
substituted
the
Until
of
are
inocubations
R-OH
is
aromatic
produced
about
However,
Batch
down
the
isolates
whereby
ability
aromatic
uncertain
capable
or
of
recently,
of
requires
ability
acetogens
possible
in
lignin.
to
sulfate,
cultures
can
of
to
the
acids.
and
aromatic
3
by
of
each
a
ring,
cleave
molecules
be
whether
ofa
that
study
degrading
a
it
the
was
were
are
or
the
ring
by
the 2
inoculum
of
reinoculated
After
roll
in
enrichment
samples
a
methane
bacteria
were
oxygen
Introduction
syringic
changed
and
filled
The
(0.27
K 2 HPO 4
composition
acetogens
final
slightly
the
tubes
vitamin
salt
the
inoculated
secondary
into
tg),
concentration
The
Samples
was
were
were
lml
formation acid,
development
with to solution
containing
(0.26g),
different
serum
Na 2 SO 4
and
ability
of
N 2 /C0 2
in
scrubbed
solution
of
of
transferred
(gil):
incubated
acetogens,
vanillic
vanillic
liquid
methanogens.
an
substrate
were
with
enrichment
vials,
was
NaHCO 3
of
inoculum
NaC1
structure
from
(2.8),
(4:1)
the
this of
enrichments
20mM were
incubated
using
boiled
acid
acid,
of
stoppered,
5mM.
without
same
methanogens
to
colonies,
methanogenic,
consortium
occasionally (1.2),
to
FeSO 4
added
as
a
and
(3.4g),
eliminate
a
after
was
of
Na 2 SO 4
and
secondary
the
2.5%
Samples
concentration
NH 4 C1
EGSB
tannic
at
Na 2
(60
tested
cooled
about
substrate.
after
with
65°C crimped,
two
rezarurin
solution
SO 4 .
mg),
of
solution.
Materials
oxygen
caused granular
by
acid) filtration
(0.3),
were
the
colonies
by
4
anaerobic
enrichment.
and
and
down
Acetogenic
days,
the
vitamin
transfer
same
Finally,
and
of
were
checked
of incubated
sulfate
KC1
(0.5mg),
addition
the
from
Enrichments
to
substrate
Na 2 Slcysteine.
sludge
lml
were
through
autoclaved.
and
substrate
room
introduction
(0.2),
added
microorganisms
solution
of
medium
of
to reducing
Growth
Methods
for
picked bacteria
the
Na 2 S9H 2 O
of
was
test
the
in
temperature
MgCl 2 .2H 2 0
to
and
a
growth.
2-bromoethanesulfonic
syringic
an
concentrations.
liquid
0.2t
the
used
a
(lOmi),
for
Bicarbonate,
anaerobic
and
Na 2 SO 4
from final
occurred
enrichments.
were
ability of
Enrichments
acetogenic
filter.
to
headspace.
enrichment
After
oxygen
acid concentration
(0.2g),
each
to
enrich
trace
enriched
under
as
(0.4),
metabolize
of
Next,
in
medium
enrichment
turbidity
roll
the
these
element
the
to
cysteine
trace
for
N 2 .
and
After
CaC1 2 .2H 2 0
liquid
tube
the
the
for
Substrates
secondary
was
by
sulfate
microorganisms
Aliquots
methanogenic
element
vial.
sulfate
gas
with
suppressing
occurred, of
and
growth
acid
aromatic
transferred
solution
enrichments.
(0.2),
into
10mM.
phase
Excess
reducers,
the
(BRES)
reducers
(i.e.,
an
were
solution,
NiC1 2
(0.1), following
occurred
was
(lml).
acids
into
to 3
introduced precipitation
contained
times
detector
column
authors
3,4-dthydroxy-5-methoxy-benzoic
1995; distinct
acid. that
standards individual
conditions
acetic Waters
equipped methoxybenzoic Analytical
source utilize
was
Similarly,
and
Colberg,
acid
Dissolved
Actetate,
oligomers
with
as and
from
Bondapak
Syringic
RID
distinct
concentrations
1
with
of
an
during
products
it
in methods
mM
monitored
with
known
a
6A.
was
pyrogallol
intermediate
the
Rspak
a
1988;
an
sulfide
proprionate,
UV
from
acid,
The
acid,
ratio
sulfate
not
BaC1 2 .
sampling
of
C18
additional
was
concentrations.
lignin,
detector
KC-G
possible
Phelps
eluent
the
vanillic
protocatechuic
of for
column
and
and
not
was
were
Dissolved
19:76:5
other
metabolic
in
of
the
a
pre-colunm
possible.
gallic
was
and
the
quantified
peak
and
2.5
set
the
to
calibrated
acid,
peaks. (length cultures
anaerobic
separate
a
at
Young,
%
butyrate
adjusted
enrichment.
acid.
occurred
0.2% acid
254
sulfide
Degradation gallic
mixture
products.
Elution
acid
It
3.9
(Fig.
nm.
Based
gravimetrically
were
and
solution
using
is
1996)
gallic
acid,
were
to
were
could
x
degradation tentatively
in
were
The
of
150
1).
a
incubated
times
the
on
standards
pH
Na 2 S/cysteine
and
acid
separated
This
analyzed
flow
of
not
mm,
of
literature
detected
degradation
of
vanillic
pyrogallol,
and
KH 2 PO 4 .
and
be
2.6
compound
rate
identified
125
of
with
after
quantified
concentrations
pyrogallol
of
was
by
by
syringic
was
A).
using
it
known
acid
HPLC tarmic
HPLC
acidification
is
used
Flow
was
A
of
0.2
catechol,
possible
as resulted
has
a
mixture
syringic
used
mi/mm,
Shimadzu
because
acid
as
concentrations.
catechol
rate
acid
so
using
on
been
eluent.
that
a
were
to
(Hanselmann
was
as
that
3,5-dihydroxy-4-
Waters
in
of
recognized
remove to
a acid.
quantification
the
the
the
an
Shodex
or
methanol:H 2 0:
0.5m1/min.
pH=2
calibrated
Under
this
refractive
unidentified
protocatechuic
medium
main
column
This
LC
peak
excess
by
these
Rspak
Module
peak carbon
by
et
represents
was
using
index
al.,
Elution
various
oxygen
of
is
peak
a
these
1 4
constraints
amplification
a
PCR-ampflfication
slides
and
overnight.
reducers,
washed
ii
containing
were
0.1% Reconstituted
establish
In-situ
13000
cofactor
mounts.
Microscopy
culture
of
dried
probe
treated
were
gelatin
A
rpm
1
Isolated
twice
Once
that
6S
Presence
that
more
delta-bacteria,
their
afterwards
The
limited
fluorescent
covered.
was
for
rRNA
ethanol/formaldehyde
was
failed
and
is
turbidity
iwth
pellet
general
next
detailed
5
characteristic
applied
cultures
minutes.
grown
0.01%
hybridization
continuation
of
and deionized
-
morning,
diluted
at
most
methanogenic
phylogenetic
group-specific
37°C.
was
sequencing
identification
to
groups
on
CrK(S0 4 ) 2 12H 2 0
in
The
likely
each
roll
syringic
observed
1:50
Finally,
water
the
of
pellet
tube
of
with
spot
of
methanogenic
because
with
slides
these
roll-tube
of
(90/10
and
bacteria
acid
position.
enrichments
on
low
was
in
of
mounting
isolates
1
PBS.
6S
were the
dried
the
experiments.
organisms
with
the G+C
reconstituted
rRNA
v/v)
slide.
solution.
liquid
enrichments
15
was
cells
washed
from
at
500
Na 2 SO 4
content,
for bacteria.
iii
fluid
37°C.
tested
oligonucleotide
were
Samples
of
.t1
enrichments,
did
5
present
sulfate-reducer
minutes
the
of
Slides
3
was
not
After
and
hybridized
times
in
and
the
by
mixture
PBS
lyse
added
were
using
isolated
in
liquid
archaea
were
drying,
in at
the
with
and
room
bacteria
1
stored
to
the
were
dried
probes
enrichments
X
enrichment
using
centrifuged
each
in
enrichment
the
were
hybridization
SET
blue
temperature,
a
applied
in
at
gene
roll
were
group-specific
spot
characteristic
for
the added
37°C.
fluorescence
tube.
releaser
20
dark
on
was
inspected
was
to
twice.
experiment
Next,
to
minutes
the
slides
However,
mixes
at
centrifuged
the
attempted
and
slide
37°C
used.
cultures
slides.
for
of
then
probes
subbed
using
each,
the
and
sulfate
Time
PCR
F420
40
for
wet
at
to
in 5
Unambiguous
bacteria), primer
in-situ
cultures
tube
thought
16S-rRNA
containing
suggesting
(Fig.
fluorescence
indicating
Phenotypic
forming,
microscopy.
methanogens.
enrichments.
with
roll
and
and
Growth
tubes
diluted
enrichments acetogens
Na 2 SO 4 /syringic
2).
hybridization
labelling
Secondary
The
to
Growth
from
The
rod-shaped
sulfate-reducing
at
that
in-situ outcompete
syringic
sulfate
characterization
enrichments
to
65°C.
same
characteristic
roll
Isolated
identification
2
a
the
after
regions
was
final
colonies
contained
hybridizations
tubes
enrichments
reducers
chosen
wet
After
acid
4-5
(Nierzwicki-Bauer,
observed
bacteria.
concentration
cultures
acid,
methanogens
mounts
enriched
in
days.
and
contained
one
each
media
bacteria,
1
of
and/or
only
6s
and
of
Na 2 SO 4
week,
the
The
1
in
were
rRNA
grew
sulfate-reducing
on also
ml
with
with
were
the
small,
enzyme
Na 2 SO 4 /syringic
acetogenic
secondary
small,
of
and
showed small
of
picked
within
secondary
at
did
BRES/syringic
molecule
syringic
the
not
iO
high
rod-shaped
bacteria
lab
not
liquid
colonies
F420
non-motile
sufficiently
and
from
5
a
protocol).
concentrations
enrich
days,
enrichments
Results
number
bacteria.
acidINa 2 SO 4
each
enrichments
bacteria
indicated
i0 4 .
with
enrichment
the
appeared
and
bacteria
acid,
exclusively
characteristic
roll
lml
acid.
low
rods,
of
selective
A
Thus,
were
was
tubes,
Na 2 SO 4 /vanillic bacteria
universal
of
the
G+C
contained
No
small
of
that
were
the
was
for
in
possible
inspected
presence
the
dissolved
colonies
the
content
and
diluted
for
for
sulfate
did
transferred
secondary
filamentous
that
characterized
of
roll
primer,
one
sulfate
reinoculated
not
a
archaea
for
did
mixture
of
tubes
by
reducers,
grew
(some
sample
group
fluoresce.
sulfate.
the
methanogenic
not
acid,
bright
reducers
and
enrichment
to
roll
in of
fluoresce
(methanogenic
bacteria,
acetogens).
of
a
of was
3 and
using
enrichments
reducers
In
dilution
in
organisms.
light
tube
methanogens
group-specific
these
contrast,
isolated liquid
inoculated
which
1
enrichment
6S
and
organisms
bacteria
enriched
series
rRNA
are
spore-
roll
Blue
for
in 6
place.
residual
the
acid
dihydroxy-5
showed
acid
analysis, HPLC-analysis
dates
not
identification hybridization
sequencing
10
Roll
sulfate
showed however, to
enrichments
observed
putatively
fluoresced ribosomal
30% hybridization
containing
the
mM
substrate
clear
was
where
tube
(Widdel
listed
1
reducers.
Table
6s
for
sequential
weak
sulfate
readily
qualitative
whether
in
picks
it
RNA
RNA
identified
with
syringic/vanillic
an
syringic
above
using
was
samples
methoxy-benzoic
were
is
1
of temperatures
fluorescence
was
and
error
not
can
summarizes
from
the
of
also
degraded
acetogenic
with
In
specific
acetogenic
the
not
Hansen,
decomposition strong
known
sulfate
be
order
occurred
low
acid
analysis
from
as
evident
enrichments
the
columns
pure
used
sulfate
G+C
and
to
other
in
acetogenic
to
reducers.
the
for
with cultures
1992).
other
to
bacteria
the
acid the
better
protocatechuic
of
during
from
Na 2 SO 4
bacteria
probe.
this
secondary
determine
acid,
indicate
reducers
probes
free
analytical
enrichment
low
and
than
of
for
It
sample.
address
fluorescence
and
followed
sulfide, Similarly,
inoculation
would is
Desulfotomaculum
substrate. G+C
20
primers.
at
are acetogenic
37°C
was
therefore
sampling
the
contained
isolated
mM
whether
enrichment
present
results
probes,
the
All
much
have
with
would
dilutions
acid and
for
by
reliability
enrichments
a
Syringic
microscopy
of
possible
weak
gravimetric
Na 2 SO 4
required
in
degradation
vanillic
bacteria
and
using
in weaker,
dates.
breakdown
Archaea
both
be
the
roll
the
with
further
10
necessary.
hybridization
vanillic
tubes
HPLC,
methanogens
enrichments.
The
acid
has
of
with
acid!
that
PCR growing
but
and
syringic
probes,
this
on
using
initial
to
(H-C
analysis
was
to
on
of
the
nevertheless
the
i0
Na 2 SO 4
syringic
acid.
gas amplification
catechol.
technique,
gallic
the
Na 2 SO 4 /syringic
On
low
first
exception
contents
the
on (Fig.
acid
substrate
chromatography
and
aromatic
The
the with
More
of
and
syringic
G+C
F420
acidJpyrogallol.
degraded
(Fig.
acid
probes
and
basis
dissolved 3a).
initial
Concentrations
the
other
control
detectable.
unambiguous
between
probe
Na 2 SO 4 .
fluorescence
and
of
concentration
3b).
Similarly,
and
ring
Archaea
of
acid
for
M4
concentration
bacterial
to
vanillic
these
Hybridization
DNA
also
acid
has
sulfate.
bacterial
samples
3,4-
with
(dilution
for
20
In
taken
probe
Cultures
hybridized
data
Vanillic
also
%
this
methane
vanillic
acid
of
groups.
that
and
The
was
it
and
case,
was
10-i)
is
the
of
of 7
recombined
days,
bacteria
stimulated
which
detected not
probe
and
were
potential
nucleotide
isolates potential
methanogens
intermediate
mmoles
measured,
degradation
pyrogallol.
mmoles
a
degradation
sulfate.
concentration
equivalent
total
extracted
it
hybridized.
the
is
was
led
An
of
Acetate
A
that
Complete
showed
possible
Continuing
in
in
of
liquid
explanation
breakdown
surprising
to 13.75
used
if
additional
probes.
these
these
syringic
4.5
sulfate.
Complete
in
were
products
the
to
cultures
degradation
from
are
one
of
moles
3,4-dihydroxy-5
was
was
to
mmoles
higher hypothesis
conditions
liquid
that
On
responsible
10
thought
hybridize
oxidation
the
liquid
Methane
Degradation
initially
analyzed acid
degradation
mM,
observation
the
of
of
experiment
is
several
are
of
demethylation
roll
concentrations
enrichments.
that
aromatic
sulfate-reducing,
other
of sulfate.
and
enrichment.
present,
products
an
to
tubes.
sulfate
that
are
was
methanogens
not
these
of
be
equivalent
vanillic
different
for
for
hand,
syringic
not
sulfate
production
detected
methoxy
of
For
to
not
was
residual
this
However,
acids,
was
and
samples.
is
are
the
pyrogallol
met.
a
detected
complete
By
required
acid of
the
degradation
After
started
weak
sulfate
colonies
of still
aromatic
reducers
but
vanillic
acid
of
Concentrations
comparison,
late
in
acetate.
methanogenic
aromatic
benzoic
enrichments,
grew
45
present.
after
it
of Individual
3
liquid
fluorescence
to
to
detection
when
days
is
concentrations
mmoles
to
requires
acetate
oxidation
CO 2
were
test
slowly
unclear
acid
based
ring
completely
another
Acetate
step.
acid
enrichments
of
acids.
These
these
whether
requires
to
picked will
inoculation, isolated
depended
of
on
colonies
an
requires
and
of
respectively,.
catechol
whether
of
and
of
week
Na 2 SO 4
No
was
enrichments have
their
was
methane
additional
observations
dissolved
were
syringic
when
degrade
production
acetogenic
are
the
residual
cultures
detected
always
modest
fluorescence
started
from
in
an
on
requires
less
sulfate-reducers,
transfer
not
is
turbidity
the
the
equivalent
in
a
necessary.
acid
detected 7.5
isolated
than
all
another
sulfate
In
inoculae
compounds
roll
detected
roll
when
first
of
amounts
when
do
cultures
of
syringic
the
mmoles
with
of
putative
an
tubes
6.25
tubes
not
acetate
showed
developed.
36
enrichments
with
no equivalent
are
an
group
sulfate
when
of
an
preclude
from
mM
in
electrons,
Complete
intermediate
of
were
Archaea-specific
higher,
were.
were
6.25
acid
of
initial
mixed
16S
were fermenters,
acetogenic
would
acetate
growth.
of
the
and sulfate.
roll
reducers,
very
to
mmoles
rRNA
organisms
After
of
detected,
samples
the
and
12.5
cultures. tube
be
or
were 7.5
small,
Thus,
One
an
5
or
of 8
completely phenols,
vanillic
rapidly
4.
5.
3.
2.
This
1.
with
was
not
compounds
rate
isolated After
not
started
but
identified.
in
sulfate
significant. than
research
BaSO 4
7
The
example
Are
on
Do biomass
Can concentrations
syntrophy
Can
acid
substrate? Are
to
Growth
acetate,
days,
cultures
permitted
aromatic
test
the
exclusive,
these
experiment
thermophilic
sulfate
loss
individual
showed
were
at
project
no
organisms
whether
Gallic
the
tannic
and
was
and
was
However,
organisms
growth were
-
different
pH
reducers
required
acids.
by
HC0 3 ?
that
methane.
observed
relatively
was
furthered
acidlpyrogallol
that
used.
acid?
organisms
able
the
showed
isolates
all
was
anaerobic
accept
Growth
initiated
would
length
determination
from
to
of
capable
carry
to
observed,
It
in
degrade
the
degrade
minor.
the
that
of
similar
also
the
the
account
carry
of
out
was
sulfate-reducing
vanillic
to
growth
bacteria
thermophilic
of
the
intermediate
inoculum
obvious
address
was
the
Methane
usually
the
utilizing
completely
out
and
substrates,
experiment.
complete
of
for
detected
acid
Conclusions
aromatic
of
the
able no
sulfate
either
the
the
that
observed
of
degradation
was products
and
larger-size
to
bacteria
following loss
tannic
products
the
at bacteria
for
aromatic
degradation
utilize
is
converted
acetogenic
ring,
acetate
measurable
individual
of
affected
example
within
acid
had
substrate.
or
enriched
substituted
of
oligomers
or
were
would
questions:
degradation
acids
formed.
after
monomeric
is
within
by
bacteria,
4-5
vanillic
of
a
enrichments,
detected
concentrations,
coprecipitation
consortium
2
completely
aromatic
be
Finally,
in
days.
weeks,
Possibly,
10
able
this
of
aromatic
acid
methanogenic
proceeded
days
these
An
aromatic
in
experiment
to
an
but
acids
this
and
experiment
grow
into
to while
- experiment
aromatic
none
acids
the
possibly
CO 2
enrichment,
syringic
of
and
by
demethylated
on
acids
eluting
at
tannic
not
of
producing
as
and
sulfate
pyrogallol.
a
grew
bacteria,
the
slower
acids,
acting
with
and
acid?
CFT 4 ?
was
acid
loss
were
but
for
in 9
required.
aromatic
between
steps released
acid
skeleton
is unlike
pyrogallol
although observed
aromatic
concentration
sum
aromatic
circumstantial monomers
sediments
bacteria
demethylated
indicating
or
unclear
sulfate
could
of
by
the
the
using
the
during
utilizing
of
ring
pyrogallol
The ring
compounds
over
whether
reducer,
individual
be
that
by (Hanselmann
concentrations
proceeds
tannic
individual
would
present
accomplished
cleavage.
isolates
of
intermediates
the
an
evidence
the
its
the
electron
syringic
acid
course
the
breakdown.
colonies
and
was
strongly
substrate,
in
monomeric
data
is
of
aromatic
was
organisms.
thus
the
Breakdown not
for
detected
et
sulfate
of
balance,
are
of
acid
same
al.,
utilized
catalyzed
the
picked
by
affected
this
compounds
the
shift
insufficient
but
Determination
1995).
produced
GC-MS
breakdown
monomers
reducers
experiment.
reactants
manner
products
in
the
However,
cannot
better
from
as
of
the
the
by
Negative
equilibrium
substrate
tannic
analysis.
chromatographs.
rates
observed
the
the
as
methane
suggests
data
be
to
and
formed
were
of
has
roll
high
assess
However,
explained
removal
acid
of
the
for
of
formed
blank
been
by
utilized.
tubes decomposition.
the
Finally,
the
temperature
aromatic
demonstrates
by by and
that
of
the
the
individual
reported
this
quantification
vanillic organisms
controls
of
were
products
acetate
bacteria
these
potential
by
the
acetate
It
Based
reaction
to
loss
products
ring.
is
not
quantify
organisms
and
also
for
in
of
indicate
of
and
degradation
which
pure
on
that
formed
from
This
Enrichments
for
the
these
enrichments
sulfate.
syringic
towards
possible
the
individual
of
syntrophic
identified
later
degradation
cultures.
the
evidence
the
do
residual
experiments.
structure
that
are
by
individual
stages
No
not
same
acid
the
breakdown not
that
breakdown
products
degradation
add
for
The
monomers
acetate
from
by
responsible
is
relationships
sulfate
inoculum
degradation,
the
of
of
of
based
sulfate-reducing
up
HPLC
sequence
tannic
growth
carbohydrate
degradation
the
lake
There
to
of
side.
are
of
of
the
aromatic
on
tannic
were
was
the
acid,
of
these
the
is
for
initial
of
only
the
10 it Table 1: Concentration of aromatic acids in liquid enrichments (@65°C): n.a. (not analyzed; n.d. (not detected)
enrichment 7/18/96 7/27/96 7/10/96 7/26/96 7/28/96 7/28/ 96 syringic a. interm. gallic a.! syringic a. interm. gallic a.! acetate acetate CH4 H2S S042 syringic acid pyrogal. pyrogal. initial inoculum (10 mM) mM mM mM mM mM mM mM mM ppm mM S4 syringic secondary n.a. n.a. n.a. n.a. n.a. n.a. 0.3 0.6 56.5 + 12.5 acid i0 5.2 0.5 n.d. 0.7 3.6 6.5 n.d. n.d. 21 + 13.8 2.9 1.6 2 0.2 2.1 5.5 n.d 0.6 25 + 15 M4 syringic secondary n.a. n.a. n.a. n.a. n.a. n.a. 0.1 n.d. 98.6 n.a. acid A4 syringic secondary n.a. n.a. n.a. n.a. n.a. n.a. 0.1 n.d. 56.7 + n.a. acid 10 8.8 0.3 n.d. n.a. n.a. n.a. 0.1 n.d. 102 + n.a. S4/S4/A4 syr. i05;i07 n.d. n.d. n.d. 0.2 60 + 18 acid pyrogallol initial inoculum (2.5 mM) S4 pyrogallol 10 syr. 2.5 n.d. n.d. - n.a. S4 pyrogallol syr. 2.5 n.d. n.d. - n.a. vanillic acid vanillic protocatechuic a. vanillic protocatechuic a./ initial inoculum (20 mM) acid /catechol acid catechol S4vanillic acid secondary 7.0 3.7 n.d. 10.8 1.0 0.7 n.d. + (27.7) M4 vanihic secondary (41.5) 2.7 17.8 1.9 0.4 n.d. 1 - n.a. acid A4 vanillic secondary 10.2 2.4 n.d. 15.3 n.d. n.d. n.d - n.a. acid tannic acid initial inoculum (10 mM) - S4 tannic acid secondary unidentified products n.a. n.d. n.a. - 12.3 M4 tannic acid secondary n.a. n.d. n.a. n.d. n.a. A4 tannic acid secondary I’ n.a. n.d. n.a. n.d. n.a.
Daniel
Widdel
Phelps
Kaiser
Hanselmann
Colberg
S.
methoxylated
J.
Schleider),
C.
The
ferulic
Microbiology
monoaromatic
presence
F.
and
Zehnder),
monoaromatic
P.
P.
L.,
D.
and
Procaryotes
3.
conversion
and
Wu
and
K.
(1988)
and
Hansen
Hanselmann
W.,
and
Z.,
Young
Wiley,
vol.1,
syringic
Drake
Kaiser
absence
aromatic
Anaerobic
133,
compounds
T.
lignin
of
(eds.
L.
2nd
A.
pp.333-
methoxylated
185
H.
acids
Y.
J.
(1992)
edition,
K.
derivatives.
of
A.
P.,
L.
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microbial
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W.
194.
Balows,
under
(1988)
Wenk,
372.
by
The
(1982)
FEMS
Springer,
Microbial a
three
bacterial
Microbiological
dissimilatory
Growth
M.,
aromatic
H.
In
degradation
Fermentative
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pp.
Truper,
metabolism
of
community
substrates
thermophilic 583
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of
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of
anaerobic
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-
Bacofen
M.
624.
metabolism
Research
Dworkin,
by
of
from
and
52,
the
Desulfotomaculum
R.
acetogenic
microorganisms
anaerobic
sulfur-reducing
25
Microbial
plant
150,
(1995)
lignin,
W. of
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28.
387
substituted
phenolic
Harder,
Growth
oligolignols, bacteria
-
sediments.
Ecology.
401
K.
compounds
bacteria.
orientis
(ed.
on
L.
on
methanol
A.
Archives
and
J.
in
In the
In the
across
improved tube
wide
into
appear
packed
glass with
added
solution
controlled
an
general
experiment
top.
the
by
N 2 .
tip
Design
tube
a
as
halfway
Further
In
with
pumping
inoculated
dialysis
of
of
Flow
oxygen
design
the
order
by
conditions.
a
NaHCO 3
3
syringe
pressuring
N 2 -flow
cm
of
was
was
design
up
to
membrane
of
indicator.
oxygen
a
of
cultivate
the
glass
introduced
started
the
continuous-flow
connected
glass
and
of
tube. The
was
chemostat
the
tube.
this
into
1mM
beads
to
medium
too
The
or
and
Oxygen
medium
design
setup
the
The
finer
from
high
solution
to
medium
isolate
to
tube
could
Tygone
flow
was
provide
tuning
used
the
permit
a
was
reservoir
continuous
with
sulfide-oxidizing
bottom
not
rate
Na 2 S9H 2 0.
be
was
chemostat
was
introduced
Thiovulum
tubing.
of
oxygenation
achieved
continued
a
surface
was
kept
the
small
filtered
with
through
adjusted
in-
flow
The 0 2 -free
aquarium
for
N 2
lml
from
outflow
either
deoxygenated
because
to
inoculum
driving
apparatus.
growth.
a
of
of
bacteria
cultivate
6mm
such
the
by
the
by
an
to
continuously
pump.
top
passive
0.5M
of
medium
the
that
tube
Medium
the
was
large
through
Beggiatoa
supporting
medium
glass
a
seawater
into
solution
The
Beggiatoa
pink
added
diffusion
fluctuation
in
was
tube.
a
outlet
an
the
coloration
25
purging
through
with
spp.
open
of
introduced
growth
containing
cm
glass
used
rezarurin
of
a
and
long
spp.
6mm
in
Na 2 S
syringe
the
tube.
Teflon
was
the under
would
Thiovulum,
glass
solution
and
glass
into
solution
a
N 2 -flow.
An
a
was
40mM
from
2mm
tubing
tube
the 2
Fellowship
financial
microbiologist,
for
their
I
I
helpful
would
would
support,
Fund
but
also
like
advice
and
participation
at
to
like
the
least
thank
during
to
William
I
acknowledge
have
Ed
this
in
Leadbetter,
become
this
Morton
project.
course
Acknowledgements
support
a
Wheeler
I
more
would
Abigail
would
from
enlightened
not
Family
not
Salyers,
the
claim
have
Arthur
Founders’
that
Kurt
been
geochemist.
this
Kiofein
Hanselmann,
possible.
course
Scholarship.
Scholarship
has
and
turned
Without
Caroline
and
me
into
this
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