Impact of temperature on ladderane lipid distribution in anammox bacteria

Abstract

Anaerobic ammonium-oxidizing (anammox) bacteria have the unique ability to synthesize fatty acids containing linearly concatenated cyclobutane rings, termed "ladderane lipids." In this study we investigated the effect of temperature on the ladderane lipid composition and distribution in anammox enrichment cultures, marine particulate organic matter, and surface sediments. Under controlled laboratory conditions we observed an increase in the amount of C(20) [5]-ladderane fatty acids compared with the amount of C(18) [5]-ladderane fatty acids with increasing temperature and also an increase in the amount of C(18) [5]-ladderane fatty acids compared with the amount of C(20) [5]-ladderane fatty acids with decreasing temperature. Combining these data with results from the natural environment showed a significant (R(2) = 0.85, P = <0.0001, n = 121) positive sigmoidal relationship between the amounts of C(18) and C(20) [5]-ladderane fatty acids and the in situ temperature; i.e., there is an increase in the relative abundance of C(18) [5]-ladderane fatty acids at lower temperatures and vice versa, particularly at temperatures between 12 degrees C and 20 degrees C. Novel shorter (C(16)) and longer (C(22) to C(24)) ladderane fatty acids were also identified, but their relative amounts were small and did not change with temperature. The adaptation of ladderane fatty acid chain length to temperature changes is similar to the regulation of common fatty acid composition in other bacteria and may be the result of maintaining constant membrane fluidity under different temperature regimens (homeoviscous adaptation). Our results can potentially be used to discriminate between the origins of ladderane lipids in marine sediments, i.e., to determine if ladderanes are produced in situ in relatively cold surface sediments or if they are fossil remnants originating from the warmer upper water column.

FIG. 1.

Homologous series of ladderane fatty acid core lipid structures. FA, fatty acid.

FIG. 2.

HPLC-APCI-MS/MS chromatogram illustrating previously unreported protonated molecules and selected product ions for shorter C₁₆ and longer C₂₄ ladderane fatty acids and previously reported C₁₈ to C₂₂ ladderane fatty acids. Panels a to j correspond to structures in Fig. 1a to j, respectively. Ladderane fatty acids were analyzed as methyl esters.

FIG. 3.

NL₅ plotted over time for anammox biomass of “Candidatus Brocadia fulgida” cultured in 16, 25, and 35°C reactors. The control reactor was kept at 35°C, whereas the biomass in the 16°C and 25°C reactors was originally cultured at 35°C. The error bars indicate the standard deviations for the NL₅ values calculated using duplicate or triplicate samples prepared and analyzed separately.

FIG. 4.

HPLC base peak chromatograms showing the differences in the distribution of ladderane fatty acids produced at the in situ temperatures in surface sediment from the northwest African continental slope and POM from the Namibian and Peruvian margins. Ladderane fatty acids were analyzed as methyl esters.

FIG. 5.

The NL₅ calculated using ladderane fatty acid concentrations from sequencing batch reactor enrichment cultures, POM, marine sediments, and microbial mats plotted against temperature. The equation relates to an empirical fourth-order sigmoidal regression. The error bars indicate the standard deviations for samples prepared and analyzed independently, which ranged from 0.01 to 0.04. A full data set is shown in the supplemental material. All data points at 35°C represent laboratory SBR enrichment cultures with a wastewater treatment plant origin.