The measurement of lipid phosphate is proposed as an indicator of microbial biomass in marine and estuarine sediments. This relatively simple assay can be performed on fresh, frozen or frozen-lyophilized sediment samples with chloroform methanol extraction and subsequent phosphate determination. The sedimentary lipid phosphate recovery correlates with the extractible ATP and the rate of DNA synthesis. Pulse-chase experiments show active metabolism of the sedimentary phospholipids. The recovery of added C-labeled bacterial lipids from sediments is quantitative. Replicate analyses from a single sediment sample gave a standard deviation of 11%. The lipid extract can be fractionated by relatively simple procedures and the plasmalogen, diacyl phospholipid, phosphonolipid and non-hydrolyzable phospholipid content determined. The relative fatty acid composition can be readily determined by gas-liquid chromatography.The lipid composition can be used to define the microbial community structure. For example, the absence of polyenoic fatty acids indicates minimal contamination with benthic micro-eukaryotes. Therefore the high content of plasmalogen phospholipids in these sediments suggests that the anaerobic prokaryotic Clostridia are found in the aerobic sedimentary horizon. This would require anaerobic microhabitats in the aerated zones.
Fatty acids are a widely studied group of lipids of sufficient taxonomic diversity to be useful in defining microbial community structure. The extraordinary resolution of glass capillary gas-liquid chromatography can be utilized to separate and tentatively identify large numbers of fatty acid methyl esters derived from the lipids of estuarine detritus and marine benthic microbiota without the bias of selective methods requiring culture or recovery of the microbes. The gas-liquid chromatographic analyses are both reproducible and highly sensitive, and the recovery of fatty acids is quantitative. The analyses can be automated, and the diagnostic technique of mass spectral fragmentation analysis can be readily applied. Splitless injection on glass capillary gas chromatographic columns detected by mass spectral selective ion monitoring provides an ultrasensitive and definitive monitoring system. Reciprocal mixtures of bacteria and fungi, when extracted and analyzed, showed progressive changes of distinctive fatty acid methyl esters derived from the lipids. By manipulating the environment of an estuarine detrital microbial community with antibiotics and culture conditions, it was possible to produce a community greatly enriched in eucaryotic fungi, as evidenced by scanning electron microscopic morphology. The fatty acid methyl esters from the lipids in the fungus-enriched detritus showed enrichment of the C 18 dienoic and the C 18 and C 20 polyenoic esters. Manipulation of the detrital microbiota that increased the procaryotic population resulted in an absence of large structures typical of fungal mycelia or diatoms, as evidenced by scanning electron microscopy, and a significantly larger proportion of anteiso- and isobranched C 15 fatty acid esters, C 17 cyclopropane fatty acid esters, and the cis -vaccenic isomer of the C 18 monoenoic fatty acid esters. As determined by these techniques, a marine settling community showed greater differences in bacterial as contrasted to microeucaryotic populations when compared with the microbial communities of benthic cores.
Microorganisms colonizing allochthonous detritus (live oak leaves) incubated in a north Florida estuary show metabolic activity that can be assayed reproducibly by the incorporation of [32P]H3PO4 and [1‐14C] sodium acetate into the lipids in a 2‐h period without introducing subculture bias. Relatively uncolonized live oak leaves show only about 1% of the biosynthetic capacity of leaves incubated in the estuary for 1 week. Lipid synthesis is proportional to time for at least 2 h and also to detrital mass. Random sampling from pooled portions of many leaves greatly reduces the variance of activities from individual leaves. Rates of phospholipid synthesis paralleled the total extractable ATP and the alkaline phosphatase activity during a 6‐week incubation of live oak leaves in Apalachicola Bay. Rates of 14C incorporation into lipids paralleled the respiratory and the α‐d‐mannosidase
Multiple biochemical assays of microbial mass and activities were applied to the estuarine detrital microbiota colonizing morphologically similar polyvinyl chloride needles and needles from slash pine (Pinus elliottii). Biodegradable pine needles consistently showed 2to 10-fold higher values of extractable adenosine 5'-triphosphate, rates of oxygen utilization, activities of alkaline phosphatase and phosphodiesterase, and the mucopeptide cell wall component muramic acid than did the polyvinyl chloride needles, during a 14-week incubation in a semitropical estuary. The higher activities by the microbiota of the biodegradable substrate correlated with estimates of the microbial density from scanning electron microscopy. The microbial community associated with the nondegradable substrate showed minimal activity of fl-D-galactosidase, fl-D-glucosidase, and aD -mannosidase in contrast to the biota of the degradable substrate, which showed 10to 100-fold higher activities of these glycoesterases. These enzymes logically could be involved in catabolism of the carbohydrate polymers of the detritus. Assuming equivalent rates of predation, a surface that is also a utilizable substrate supports a threeto fivefold more active microbial population.
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