208
Use of Radioisotopic Methodology in Aquatic Microbial Ecology
version factors based on the thymidine incorporation rates into DNA and still
recommended for use (Moriarty 1990) cannot exist or be calculated. The only
reasonable way for the practical application of the thymidine method for this
purpose remains its calibration according to some other maybe old but direct
and proved method for estimation of microbial production, but omitting the
invalid and unnecessary step of calculating the absolute values of TDR incorporation by microbial populations. These values, in principle, cannot be reliable when an indefinite value of CH)-TDR of specific radioactivity is used for
their calculation. In this case, such a calibration may be seen only as an estimation in parallel experiments of bacterial production by the reference
method selected and the cpm (or dpm for the sediments) radioactivity (Rb) of
CH)-TDR incorporated into the macromolecular cold TCA-resistant fraction
of bacterial biomass, both calculated 1-1 day-I.
Experience in the use of empirical conversion factors (Kj ) for calculation
parameters of microbial growth and production started from the very beginning of the practical use of the thymidine methodology (Kirchmann et a1.
1982). The production rates of bacterioplankton as measured during CH)TDR incorporation and calculated with the use of theoretical K j values of
0.2-0.5 x 10 9 cellsnmol- I were found to be significantly and variously lower
than the same values obtained during their parallel estimation with the aid of
other methods like FDC, changes in number of microbial cells, or dark CO2
uptake (Riemann and Sondergaard 1984; Tuomi 1997). Soon it became clear
that to obtain reasonable values of microbial production with the thymidine
method was possible only using some empirical conversion factors obtained
after direct calibration of the thymidine incorporation rates after the values
of microbial production were measured by a reference method. Many such
calibrations were made and hundreds of various K j factors were obtained
(Ducklow and Carlson 1992; Kirchmann and Ducklow 1993). They ranged in
different water basins roughly within 2 orders of values (between 0.5 and over
60) and appeared to be dependent on the level of productivity seasons,
generation time of bacterioplankton (Newell and Fallon 1982; Hanson and
Lowery 1983; Bender 1987; Scavia and Laird 1987; Coveney and Wetzel 1988;
Smith and Riemann 1988; Bell 1990; Leakey et a1. 1996), and also upon how
it was calculated during the calibration experiments (Tuomi 1997), because
as the above parameters change on vertical profiles, the same variability in KI
should be expected at different depths. An example of spatial and seasonal
variability of conversion factors K j is given in Fig. 4.13.
A great deal of confusion and unpredictability in the estimations of the
conversion factor is warranted not only by the above-mentioned indefinite
character of specific radioactivity necessary for the TDR-uptake calculations.
It is caused also by the variability in evaluations of organic carbon content
in 1 mg of bacterial biomass, and of the mean volume of bacterial cells in
their natural populations. The former is evaluated within the range 106 to
350fg Cmm 3 (Bell 1986). Small deviations in linear size of bacterial cells entail
Use of Radioisotopic Methodology in Aquatic Microbial Ecology
version factors based on the thymidine incorporation rates into DNA and still
recommended for use (Moriarty 1990) cannot exist or be calculated. The only
reasonable way for the practical application of the thymidine method for this
purpose remains its calibration according to some other maybe old but direct
and proved method for estimation of microbial production, but omitting the
invalid and unnecessary step of calculating the absolute values of TDR incorporation by microbial populations. These values, in principle, cannot be reliable when an indefinite value of CH)-TDR of specific radioactivity is used for
their calculation. In this case, such a calibration may be seen only as an estimation in parallel experiments of bacterial production by the reference
method selected and the cpm (or dpm for the sediments) radioactivity (Rb) of
CH)-TDR incorporated into the macromolecular cold TCA-resistant fraction
of bacterial biomass, both calculated 1-1 day-I.
Experience in the use of empirical conversion factors (Kj ) for calculation
parameters of microbial growth and production started from the very beginning of the practical use of the thymidine methodology (Kirchmann et a1.
1982). The production rates of bacterioplankton as measured during CH)TDR incorporation and calculated with the use of theoretical K j values of
0.2-0.5 x 10 9 cellsnmol- I were found to be significantly and variously lower
than the same values obtained during their parallel estimation with the aid of
other methods like FDC, changes in number of microbial cells, or dark CO2
uptake (Riemann and Sondergaard 1984; Tuomi 1997). Soon it became clear
that to obtain reasonable values of microbial production with the thymidine
method was possible only using some empirical conversion factors obtained
after direct calibration of the thymidine incorporation rates after the values
of microbial production were measured by a reference method. Many such
calibrations were made and hundreds of various K j factors were obtained
(Ducklow and Carlson 1992; Kirchmann and Ducklow 1993). They ranged in
different water basins roughly within 2 orders of values (between 0.5 and over
60) and appeared to be dependent on the level of productivity seasons,
generation time of bacterioplankton (Newell and Fallon 1982; Hanson and
Lowery 1983; Bender 1987; Scavia and Laird 1987; Coveney and Wetzel 1988;
Smith and Riemann 1988; Bell 1990; Leakey et a1. 1996), and also upon how
it was calculated during the calibration experiments (Tuomi 1997), because
as the above parameters change on vertical profiles, the same variability in KI
should be expected at different depths. An example of spatial and seasonal
variability of conversion factors K j is given in Fig. 4.13.
A great deal of confusion and unpredictability in the estimations of the
conversion factor is warranted not only by the above-mentioned indefinite
character of specific radioactivity necessary for the TDR-uptake calculations.
It is caused also by the variability in evaluations of organic carbon content
in 1 mg of bacterial biomass, and of the mean volume of bacterial cells in
their natural populations. The former is evaluated within the range 106 to
350fg Cmm 3 (Bell 1986). Small deviations in linear size of bacterial cells entail