Sensitive assay, based on hydroxy fatty acids from lipopolysaccharide lipid A, for Gram-negative bacteria in sediments

Abstract

Biochemical measures have provided insight into the biomass and community structure of sedimentary microbiota without the requirement of selection by growth or quantitative removal from the sediment grains. This study used the assay of the hydroxy fatty acids released from the lipid A of the lipopolysaccharide in sediments to provide an estimate of the gram-negative bacteria. The method was sensitive to picomolar amounts of hydroxy fatty acids. The recovery of lipopolysaccharide hydroxy fatty acids from organisms added to sediments was quantitative. The lipids were extracted from the sediments with a single-phase chloroform-methanol extraction. The lipid-extracted residue was hydrolyzed in 1 N HCI, and the hydroxy fatty acids of the lipopolysaccharide were recovered in chloroform for analysis by gas-liquid chromatography. This method proved to be about fivefold more sensitive than the classical phenol-water or trichloroacetic acid methods when applied to marine sediments. By examination of the patterns of hydroxy fatty acids, it was also possible to help define the community structure of the sedimentary gram-negative bacteria. Biochemical methods have proved to be very useful in assessing the microbial biomass of sediments (10, 18, 29, 32). These methods are not dependent upon growth of the organisms for assay or on the quantitative removal of organisms from the sediments (10, 19). Estimations of the total cellular biomass are readily made from the extractable phospholipid (32). The procaryotic biomass can be unequivocally estimated from the muramic acid (10; R. H. Findlay, D. J. W. Moriarty, and D. C. White, Geomicrobiology J., in press). To translate the muramic acid content into cellular carbon, the proportion of cyanophytes, gram-negative, and gram-positive bacteria must be known (10, 18). The pro-MATERIALS AND METHODS Materials. Glass-distilled solvents (Burdick and Jackson, Muskegon, Mich.), freshly distilled chloroform, and derivatizing agents from Pierce Chemical

Detection of the anaerobic dechlorinating microorganism Desulfomonile tiedjei in environmental matrices by its signature lipopolysacchride branched-long-chain hydroxy fatty acids

“…Analysis of LPS-OHFA from the lipid-extracted residue of environmental samples provides a means by which organisms containing unusual compositions of theses fatty acids can be detected. LPS-OHFA analysis has been used to detect Gram-negative organisms in marine and estuarine sediments [5]. An organism such as Planctomyces, which contains unusual LPS-OHFA, could be readily detected using these methods [22].…”

“…We report that D. tiedjei contains characteristic branched, long-chain hydroxy fatty acids in its lipopolysaccharide that can be used as signature biomarkers. Lipopolysaccharide hydroxy fatty acids (LPS-OHFA) can be readily assessed in environmental matrices by gas chromatographic/mass spectrometric analysis of derivatives after hydrolysis and re-extraction of the lipid residue [5]. Analysis of environments containing haloaromatic contaminants for LPS-OHFA patterns like those of D. tiedjei could indicate the presence of related microorganisms with potential aryl reductive dehalogenating activity.…”

Impact of biomass burning on soil microorganisms and plant metabolites: A view from molecular distributions of atmospheric hydroxy fatty acids over Mount Tai

“…Hydroxy fatty acids (FAs) can be used as tracers for microorganisms such as bacteria, fungi as well as for algae, and higher plants during their transport via air [ Tyagi et al , ]. Positional isomers (i.e., α ‐, β ‐, and ω ‐) of these hydroxy FAs have been identified in sediments [ Kawamura and Ishiwatari , ; Parker et al , ; Perry et al , ; Wakeham , ], dust [ Fox et al , ; Hines et al , ; Saraf et al , ], lake waters and sediments [ Kawamura and Ishiwatari , ; Kawamura et al , ; Zhang et al , ], snow [ Tyagi et al , ], and continental [ Graham et al , ; Lee et al , ] and marine aerosols [ Kawamura , ; Tyagi et al , ]. Among the three isomers, β ‐hydroxy FAs (from C 10 to C 18 ) have been reported as chemical markers for the GNB as they are abundant in their cell membrane and are also responsible for their toxic effects (e.g., respiratory problems and skin infections) [ Ratledge and Wilkinson , ].…”

Signature fatty acids in the polar lipids of acid-producingThiobacillusspp.: Methoxy, cyclopropyl, alpha-hydroxy-cyclopropyl and branched and normal monoenoic fatty acids

“…The 2-OTMSiFAME showed the diagnostic M-59 ion [21], as well as major fragments at m/z 103 and 129. The 3-OTMSiFAME show the definitive fragment at m/z 175 plus the rearrangement at m/z 133, and m/z 159 and 131 [17,21]. Mid-chain OTMSiFAME showed major ions attributable to fragmentation either side of the OTMSi group.…”

“…Trimethylsilyl ethers of OHFAME were formed with N, O,-bis-(trimethylsilyl)-trifluoroacetamide (BSFTA) (Pierce) [17].…”