Analysis of intact ladderane phospholipids, originating from viable anammox bacteria, using RP-LC-ESI-MS

Since the discovery of the anaerobic ammonium oxidizing (anammox) bacteria, many attempts have been made in order to identify these environmentally important bacteria in natural environments. Anammox bacteria contain a unique class of lipids, called ladderane lipids and here we present a novel method to detect viable anammox bacteria in sediments and waste water treatment plants based on the use of a ladderane lipid biomarker. Intact ladderane phosphatidylcholine (PC) lipids are analyzed using reversed-phase liquid chromatography-electrospray ionization-mass spectrometry. Following extraction from the complex sediment matrix, reversed-phase LC is used to separate ladderane PC lipids based on their tail group hydrophobicity as well as their ether or ester link to the glycerol backbone in the sn-2 position. We investigate the presence of intact ladderane lipids in natural sediments displaying anammox activity and illustrate the use of a specific intact membrane forming PC lipid as a biomarker for viable anammox bacterial cells. The presented method can be used to elucidate the whereabouts of viable anammox bacteria, subsequently enabling an estimation of anammox activity. This will greatly increase the knowledge of anammox bacteria and their importance in the global nitrogen cycle.