Estimation of Relative Microbial Activity in Aquatic Habitats
163
1968; Williams 1973; Gocke 1976). Therefore the introduction of this new
version cannot eliminate the above-mentioned drawbacks of methods based
on the use of the same nanomolar concentrations of labeled substrates.
However, the technique of measuring respired CO2 appeared to be applicable
for the study of decomposition rates of organic substrates in natural waters,
so it will be described below (see Sect. 4.3). At the same time, it should be
clearly understood that definite skepticism concerning the prospective of the
use of labeled organic substrates as a useful instrument to even relatively
quantify heterotrophic activity in water bodies is limited only in its relation to
the application of their nanomolar concentrations. When refusing the latter,
we should not throw out the baby with the bath water. This technique could
be adapted not only to estimate the relative distribution of active heterotrophic microflora (or heterotrophic potential), but also to measure the
absolute rates of uptake and decomposition of organic substrates, either
common or specific.
The main practical reason for measuring relative uptake rates of labeled
organic substrates is in the scanning of the spatial distribution of active heterotrophic microflora in a water basin in order to locate zones of most active
processes of decomposition and self-purification (Sorokin 1970a; Banub and
Williams 1972; Novitski 1983). Actually, the goal is to replace the microbial
plate count or dilution methods for the counting of heterotrophic (or saprophytic) bacteria in natural water samples. In both cases, e.g., with the radioisotopic technique and with plate count, in any case we may obtain only relative
values. The problem is to employ the methodology which gives the possibility
to measure relative heterotrophic activity, which will accurately correspond to
a relative abundance of active heterotrophic bacteria in a given sample independently of the organic substrate background in it. This goal proved to be
quite achievable. I had developed this kind of methodology in 1960 and very
often used it successfully for scanning heterotrophic activity in the sea
(Sorokin 1969, 1971, 1972b; Sorokin et al. 1995a). The relative activity of
heterotrophic bacteria expressed as cpm per 50-100ml of water or per 1 g
of wet sediment generally correlates well with the parallel microbiological
colony counts of heterotrophic bacteria on membrane filters, placed at the
surface of peptone-agar medium (Figs 4.1, 4.2). The examples of vertical
profiles of the relative activity of heterotrophic bacteria in deep sea are shown
in Fig. 4.3.
To make the method workable, it is necessary to outline the following
principles:
1. To avoid any possibility of a significant isotopic dilution: the amounts of
labeled substrate added to the sample, taken in a basin of a moderate
trophic level, should be 2-3 f.lmol per 100 ml if it is an individual substance (glutamate) or twice as much if it is the protein hydrolyzate.
The specific radioactivity of such substrates should be correspondingly
0.2-0.4 mCi mmol- 1 •
163
1968; Williams 1973; Gocke 1976). Therefore the introduction of this new
version cannot eliminate the above-mentioned drawbacks of methods based
on the use of the same nanomolar concentrations of labeled substrates.
However, the technique of measuring respired CO2 appeared to be applicable
for the study of decomposition rates of organic substrates in natural waters,
so it will be described below (see Sect. 4.3). At the same time, it should be
clearly understood that definite skepticism concerning the prospective of the
use of labeled organic substrates as a useful instrument to even relatively
quantify heterotrophic activity in water bodies is limited only in its relation to
the application of their nanomolar concentrations. When refusing the latter,
we should not throw out the baby with the bath water. This technique could
be adapted not only to estimate the relative distribution of active heterotrophic microflora (or heterotrophic potential), but also to measure the
absolute rates of uptake and decomposition of organic substrates, either
common or specific.
The main practical reason for measuring relative uptake rates of labeled
organic substrates is in the scanning of the spatial distribution of active heterotrophic microflora in a water basin in order to locate zones of most active
processes of decomposition and self-purification (Sorokin 1970a; Banub and
Williams 1972; Novitski 1983). Actually, the goal is to replace the microbial
plate count or dilution methods for the counting of heterotrophic (or saprophytic) bacteria in natural water samples. In both cases, e.g., with the radioisotopic technique and with plate count, in any case we may obtain only relative
values. The problem is to employ the methodology which gives the possibility
to measure relative heterotrophic activity, which will accurately correspond to
a relative abundance of active heterotrophic bacteria in a given sample independently of the organic substrate background in it. This goal proved to be
quite achievable. I had developed this kind of methodology in 1960 and very
often used it successfully for scanning heterotrophic activity in the sea
(Sorokin 1969, 1971, 1972b; Sorokin et al. 1995a). The relative activity of
heterotrophic bacteria expressed as cpm per 50-100ml of water or per 1 g
of wet sediment generally correlates well with the parallel microbiological
colony counts of heterotrophic bacteria on membrane filters, placed at the
surface of peptone-agar medium (Figs 4.1, 4.2). The examples of vertical
profiles of the relative activity of heterotrophic bacteria in deep sea are shown
in Fig. 4.3.
To make the method workable, it is necessary to outline the following
principles:
1. To avoid any possibility of a significant isotopic dilution: the amounts of
labeled substrate added to the sample, taken in a basin of a moderate
trophic level, should be 2-3 f.lmol per 100 ml if it is an individual substance (glutamate) or twice as much if it is the protein hydrolyzate.
The specific radioactivity of such substrates should be correspondingly
0.2-0.4 mCi mmol- 1 •