Bacteriohopanetetrol-x: constraining its application as a lipid biomarker for marine anammox using the water column oxygen gradient of the Benguela upwelling system
Authors: van Kemenade Zoë R. , Villanueva Laura , Hopmans Ellen C. , Kraal Peter , Witte Harry J. , Sinninghe Damsté Jaap S. , Rush Darci 
Published: January 2022

Abstract. Interpreting lipid biomarkers in the sediment archive
requires a good understanding of their application and limitations in modern
systems. Recently it was discovered that marine bacteria performing
anaerobic ammonium oxidation (anammox), belonging to the genus Ca. Scalindua,
uniquely synthesize a stereoisomer of bacteriohopanetetrol (“BHT-x”). The
ratio of BHT-x over total bacteriohopanetetrol (BHT, ubiquitously synthesized
by diverse bacteria) has been suggested as a proxy for water column anoxia.
As BHT has been found in sediments over 50 Myr old, BHT-x has the potential
to complement and extend the sedimentary biomarker record of marine anammox,
conventionally constructed using ladderane lipids. Yet, little is known
about the distribution of BHT-x in relation to the distribution of ladderanes
and to the genetic evidence of Ca. Scalindua in modern marine systems. Here, we
investigate the distribution of BHT-x and the application of the BHT-x ratio in
relation to distributions of ladderane intact polar lipids (IPLs), ladderane
fatty acids (FAs) and Ca. Scalindua 16S rRNA genes in suspended particulate
matter (SPM) from the water column of the Benguela upwelling system (BUS),
sampled across a large oxygen gradient. In BUS SPM, high BHT-x abundances were
restricted to the oxygen-deficient zone on the continental shelf (at
[O2] < 45 µmol L−1, in all but one case). High
BHT-x abundances co-occurred with high abundances of the Ca. Scalindua 16S rRNA
gene (relative to the total number of bacterial 16S rRNA genes) and
ladderane IPLs. At shelf stations with [O2] > 50 µmol L−1, the BHT-x ratio was < 0.04 (in all but one case). In
apparent contradiction, ladderane FAs and low abundances of BHT and BHT-x
(resulting in BHT-x ratios > 0.04) were also detected in oxygenated
offshore waters ([O2] up to 180 µmol L−1), whereas
ladderane IPLs were undetected. The index of ladderane lipids with five
cyclobutane rings (NL5) correlates with in situ temperature. NL5-derived temperatures suggested that ladderane FAs in the offshore waters
were not synthesized in situ but were transported down-slope from warmer shelf
waters. Thus, in sedimentary archives of systems with known lateral organic
matter transport, such as the BUS, relative BHT and BHT-x abundances should be
carefully considered. In such systems, a higher BHT-x ratio may act as a safer
threshold for deoxygenation and/or Ca. Scalindua presence: our results and
previous studies indicate that a BHT-x ratio of ≥ 0.2 is a robust
threshold for oxygen-depleted waters ([O2] < 50 µmol kg−1). In our data, ratios of ≥ 0.2 coincided with Ca. Scalindua 16S
rRNA genes in all samples (n=62), except one. Lastly, when investigating
in situ anammox, we highlight the importance of using ladderane IPLs over BHT-x and/or
ladderane FAs; these latter compounds are more recalcitrant and may derive
from transported fossil anammox bacteria remnants.