144
For leucine uptake, at least, it is possible to do
the incubation in microcentrifuge tubes and to use
centrifugation to concentrate the cells (Smith and
Azam 1993). The extractions (done in this case
with 5% trichloroacetic acid) occur in the centrifuge tube. Best of all, scintillation fluid can be
added directly to the same centrifuge tube which is
then dropped into a dry scintillation vial for the
radioactive counting. This modification is faster,
less expensive, and gives lower variance than earlier methods.
Bacterial Respiration
In soils, bacterial respiration (BR) is more commonly measured than BP is. In planktonic systems,
it is actually difficult to measure BR and it is much
less commonly measured than is BP. In order to get
measurable changes in dissolved oxygen (DO) or
in DIC, the incubation has to be much longer than
that for BP. In oligotrophic waters, the incubation
may have to be 24 hours or more to provide measurable oxygen changes. Further, the total community respiration in a water sample consists of the
respiration of bacteria and other organisms, especially phytoplankton. Typically, differential filtration is used to separate bacteria from these larger
organisms. This physical separation also removes
bacteria from one possible substrate (new phytoplankton growth) and from most grazers. Many researchers estimate BR from literature values ofbacterial growth efficiency (BGE), which have been
measured by a variety of different approaches.
BGE = BP/(BP + BR) * 100% (9.3)
since the term BP + BR is the assimilation of substrate, BGE is analogous to assimilation efficiency
for higher organisms.
The various approaches for measuring BGE
include:
1. measuring the appearance of a representative
14C-Iabeled substrate (glucose, acetate, labeled
phytoplankton exudates) into bacterial biomass
and into DIC or 3H substrates into biomass and
water;
2. measuring the disappearance of ambient DOC
(which equals BP + BR) and the simultaneous
increase in bacterial biomass (which equals BP);
and
Jonathan J. Cole
3. measuring BP (by thymidine or leucine uptake)
and simultaneous consumption of DO or production of DIC.
Unfortunately, values of BGE show a large range
in the literature (table from Jahnke and Craven
1995; Cole and Pace 1995; Roland and Cole in
press). Thus the practice of estimating BR from an
assumed value of BGE is subject to large uncertainty. Interest in directly measuring BR or BGE is
growing and advances in the near future are likely
(del Giorgio and Cole 1998).
Uptake and Turnover
of Specific Substrates
The modem age of aquatic microbiology was ushered in during the mid-1960s when researchers began adding small amounts of radioactive organic
compounds to water samples and examining the uptake (Wright and Hobbie 1966; Wetzel 1967) and
respiration (Hobbie et a1. 1969) of these. The original goal of this research was largely to see how
active bacteria were in various environments (Hobbie et a1. 1969; Hobbie and Rublee 1977) and how
fast specific substrates were being turned over.
These approaches demonstrated early on that many
simple sugars and amino acids were being turned
over by microbes on a time scale of minutes to
hours. A major disappointment of these early measurements of heterotrophic potential was that they
did not provide a measure of total BP. Thus, we
could measure the turnover time and V max for glucose (or any other labeled substrate) but we had no
way of knowing the total array and quantity of substrates in use by the bacterial community. The glucose V max was used as an index of bacterial heterotrophic potential (Hobbie and Rublee 1977; Wetzel
1967).
Substrates Supporting Bacterial Growth
The thymidine and leucine methods (above) provided a means by which total BP could be approached, albeit with some uncertainty. With an estimate of total BP, the use of specific substrates or
suites of compounds takes on a new interest. We
can now begin to ask what fraction of BP is supported by what kind of molecule. In this approach
one adds a tracer amount of a labeled substrate. One
For leucine uptake, at least, it is possible to do
the incubation in microcentrifuge tubes and to use
centrifugation to concentrate the cells (Smith and
Azam 1993). The extractions (done in this case
with 5% trichloroacetic acid) occur in the centrifuge tube. Best of all, scintillation fluid can be
added directly to the same centrifuge tube which is
then dropped into a dry scintillation vial for the
radioactive counting. This modification is faster,
less expensive, and gives lower variance than earlier methods.
Bacterial Respiration
In soils, bacterial respiration (BR) is more commonly measured than BP is. In planktonic systems,
it is actually difficult to measure BR and it is much
less commonly measured than is BP. In order to get
measurable changes in dissolved oxygen (DO) or
in DIC, the incubation has to be much longer than
that for BP. In oligotrophic waters, the incubation
may have to be 24 hours or more to provide measurable oxygen changes. Further, the total community respiration in a water sample consists of the
respiration of bacteria and other organisms, especially phytoplankton. Typically, differential filtration is used to separate bacteria from these larger
organisms. This physical separation also removes
bacteria from one possible substrate (new phytoplankton growth) and from most grazers. Many researchers estimate BR from literature values ofbacterial growth efficiency (BGE), which have been
measured by a variety of different approaches.
BGE = BP/(BP + BR) * 100% (9.3)
since the term BP + BR is the assimilation of substrate, BGE is analogous to assimilation efficiency
for higher organisms.
The various approaches for measuring BGE
include:
1. measuring the appearance of a representative
14C-Iabeled substrate (glucose, acetate, labeled
phytoplankton exudates) into bacterial biomass
and into DIC or 3H substrates into biomass and
water;
2. measuring the disappearance of ambient DOC
(which equals BP + BR) and the simultaneous
increase in bacterial biomass (which equals BP);
and
Jonathan J. Cole
3. measuring BP (by thymidine or leucine uptake)
and simultaneous consumption of DO or production of DIC.
Unfortunately, values of BGE show a large range
in the literature (table from Jahnke and Craven
1995; Cole and Pace 1995; Roland and Cole in
press). Thus the practice of estimating BR from an
assumed value of BGE is subject to large uncertainty. Interest in directly measuring BR or BGE is
growing and advances in the near future are likely
(del Giorgio and Cole 1998).
Uptake and Turnover
of Specific Substrates
The modem age of aquatic microbiology was ushered in during the mid-1960s when researchers began adding small amounts of radioactive organic
compounds to water samples and examining the uptake (Wright and Hobbie 1966; Wetzel 1967) and
respiration (Hobbie et a1. 1969) of these. The original goal of this research was largely to see how
active bacteria were in various environments (Hobbie et a1. 1969; Hobbie and Rublee 1977) and how
fast specific substrates were being turned over.
These approaches demonstrated early on that many
simple sugars and amino acids were being turned
over by microbes on a time scale of minutes to
hours. A major disappointment of these early measurements of heterotrophic potential was that they
did not provide a measure of total BP. Thus, we
could measure the turnover time and V max for glucose (or any other labeled substrate) but we had no
way of knowing the total array and quantity of substrates in use by the bacterial community. The glucose V max was used as an index of bacterial heterotrophic potential (Hobbie and Rublee 1977; Wetzel
1967).
Substrates Supporting Bacterial Growth
The thymidine and leucine methods (above) provided a means by which total BP could be approached, albeit with some uncertainty. With an estimate of total BP, the use of specific substrates or
suites of compounds takes on a new interest. We
can now begin to ask what fraction of BP is supported by what kind of molecule. In this approach
one adds a tracer amount of a labeled substrate. One