not to remove all cellular components from the surface. In situ, microscopic methods
often fail because morphology in microbial biofilms may have little relationship to the
metabolic function of the cells. The combination of autoradiography and microscopy
however can provide a powerful tool in the study of biofilms.
Biochemical analysis
Our laboratory has been involved in the development of assays for biofilms of microbial
consortia in which the bias of cultural selection of the classical plate count is eliminated.
The total community is examined in these procedures and by using labeled precursors it is
also possible to determine the metabolic activities of specific components in the microbial
communities. The concept of “signatures” for subsets of the community based on the
limited distribution of specific components has been validated by using antibiotics and
cultural conditions to manipulate the community structure. The resulting changes agreed
both morphologically and biochemically with the expected results (White et al., 1980).
Other validation experiments that involved isolation and analysis of specific organisms
and finding them in appropriate mixtures, utilization of specific inhibitors and noting the
response and changes in the local environment such as the light intensity are summarized
in the review (White, 1983).
With these techniques, we have been able to show succession in marine biofouling films
(Morrison et al., 1977 ; Nickels et al., 1981a), the effects of substratun biodegradability
(Bobbie et al., 1978), the effects of substratum microtopology (Nickels et al, 1981b), the
effects of mechanical disturbance (Nickels et al, 1981c), the effects of amphipod grazing
and resource partitioning (Morrison and White, 1980 ; Smith et al, 1982a), the effects of
sand dollar bioturbation and predation (Findlay and White, 1983a), the effects of
essential elemental chelation (Nickels et al, 1979), the effects of oil and gas well drilling
fluids (Smith et al, 1982b), the effects of light (Bobbie et al, 1981) and the effects of
epibenthic predation (Federle et al., 1983) on microbial films.
In Table 1, the sensitivity of some of the methods by which biomass and community
structure can be determined is listed. These methods utilize fluorimetric detection after
high pressure liquid chromatography (HPLC) (phytanyl glycerol ether phospholipid
assay) or detection by flame ionization after gas-liquid chromatography (GLC). With
these techniques it is possible to detect 10 3 to 10 5 organisms the size of E. coli. The use of
more sensitive assays based on derivatives that give strong responses in electron capture
detection capillary GLC should increase the sensitivity of these assays.
The phospholipids are found in the membranes of all cells. The membrane phospholipids
have a relatively rapid turnover so the assay of these lipids gives a measure of the “viable”
cellular biomass (White et al, 1979a). The phosphate of the phospholipids or the
glycerol-phosphate and acid labile glycerol from phosphatidyl glycerol has been assayed
(Gehron and White, 1983). The ester-linked phospholipid fatty acids are both the most
sensitive and the most useful measures thus far developed ( Bobbie and White, 1980). The
usefulness of this assay has been greatly increased by the determination of the configuration and position of double bonds in monoenoic fatty acids (Nichols et al., 1985). Samples
from marine sediments often yield 300 fatty acids which give deep insight into the
community structure as well as an estimate of the biomass. “Signatures” (components
restricted to subsets of the microbial community with similar physiological functions) for
some of the microbial groups involved in anaerobic fermentations have been developed.
The rate limiting step in fermentations is the degradation of polymers that is carried out
by the anaerobic fermenters. A portion of these organisms contain plasmalogen phospho210
often fail because morphology in microbial biofilms may have little relationship to the
metabolic function of the cells. The combination of autoradiography and microscopy
however can provide a powerful tool in the study of biofilms.
Biochemical analysis
Our laboratory has been involved in the development of assays for biofilms of microbial
consortia in which the bias of cultural selection of the classical plate count is eliminated.
The total community is examined in these procedures and by using labeled precursors it is
also possible to determine the metabolic activities of specific components in the microbial
communities. The concept of “signatures” for subsets of the community based on the
limited distribution of specific components has been validated by using antibiotics and
cultural conditions to manipulate the community structure. The resulting changes agreed
both morphologically and biochemically with the expected results (White et al., 1980).
Other validation experiments that involved isolation and analysis of specific organisms
and finding them in appropriate mixtures, utilization of specific inhibitors and noting the
response and changes in the local environment such as the light intensity are summarized
in the review (White, 1983).
With these techniques, we have been able to show succession in marine biofouling films
(Morrison et al., 1977 ; Nickels et al., 1981a), the effects of substratun biodegradability
(Bobbie et al., 1978), the effects of substratum microtopology (Nickels et al, 1981b), the
effects of mechanical disturbance (Nickels et al, 1981c), the effects of amphipod grazing
and resource partitioning (Morrison and White, 1980 ; Smith et al, 1982a), the effects of
sand dollar bioturbation and predation (Findlay and White, 1983a), the effects of
essential elemental chelation (Nickels et al, 1979), the effects of oil and gas well drilling
fluids (Smith et al, 1982b), the effects of light (Bobbie et al, 1981) and the effects of
epibenthic predation (Federle et al., 1983) on microbial films.
In Table 1, the sensitivity of some of the methods by which biomass and community
structure can be determined is listed. These methods utilize fluorimetric detection after
high pressure liquid chromatography (HPLC) (phytanyl glycerol ether phospholipid
assay) or detection by flame ionization after gas-liquid chromatography (GLC). With
these techniques it is possible to detect 10 3 to 10 5 organisms the size of E. coli. The use of
more sensitive assays based on derivatives that give strong responses in electron capture
detection capillary GLC should increase the sensitivity of these assays.
The phospholipids are found in the membranes of all cells. The membrane phospholipids
have a relatively rapid turnover so the assay of these lipids gives a measure of the “viable”
cellular biomass (White et al, 1979a). The phosphate of the phospholipids or the
glycerol-phosphate and acid labile glycerol from phosphatidyl glycerol has been assayed
(Gehron and White, 1983). The ester-linked phospholipid fatty acids are both the most
sensitive and the most useful measures thus far developed ( Bobbie and White, 1980). The
usefulness of this assay has been greatly increased by the determination of the configuration and position of double bonds in monoenoic fatty acids (Nichols et al., 1985). Samples
from marine sediments often yield 300 fatty acids which give deep insight into the
community structure as well as an estimate of the biomass. “Signatures” (components
restricted to subsets of the microbial community with similar physiological functions) for
some of the microbial groups involved in anaerobic fermentations have been developed.
The rate limiting step in fermentations is the degradation of polymers that is carried out
by the anaerobic fermenters. A portion of these organisms contain plasmalogen phospho210