Laura Villanueva:Research

Lipid biosynthetic pathways

Our research is focused on lipid biomarkers of certain organisms either because they are markers of the presence of a specific group (e.g. ladderane lipids of anammox bacteria), physiological condition (e.g. ornithine lipids, thought to be formed under phosphate limitation), or mostly because they have been seen to correlate to growth temperature and thus used to estimate paleotemperature (e.g. long chain alkenones, GDGTs, and long chain diols involved in the organic paleotemperature proxies UK37, TEX86 and LDI).

In order to improve the predictive nature of lipid biomarkers used for microbial ecology and in paleotemperature proxies it is essential to determine the following:

• Their biological source/s, as well as seasonality and spatial distribution of the source/s.
• How changes in physicochemical conditions influence the abundance and distribution of these biomarkers
• How and when these biomarkers have been acquired through evolution and how these capacity has been spread among different biological taxa

For doing so, we combine lipid analysis with molecular techniques based on:

• Analyzing lipid synthetic pathways: evolution and prediction of gene function
• Targeting phylogenetic, metabolic and lipid biosynthetic genes as markers for the presence of lipid biomarker producers.
• Estimating which physicochemical conditions that induce changes in the synthesis of lipid biomarkers.

TOOLS

• Data mining of metagenomic databases
• Detection and quantification of lipids biosynthetic genes and their gene expression by sequencing and quantitative PCR
• 16S and 18S rRNA gene phylogeny (pyrosequencing)
• Protein structure modeling
• Heterologous expression

REFERENCES

• Villanueva, L., Schouten, S., and Sinninghe Damsté, JS. 2014. A Re-evaluation of the Archaeal Membrane Lipid Biosynthetic Pathway. Nat. Rev. Microbiol. 12:438–448. Pubmed

• Villanueva, L., Besseling M, Rampen S., Rodrigo-Gamiz M., and Sinninghe Damste JS. 2014. Potential biological sources of long chain alkyl diols in a lacustrine system. Org. Geochem. 68:27-30. Link

• Villanueva, L., Rijpstra, W.I.C., Schouten, S., and Sinninghe Damsté, J.S. 2014. Genetic biomarkers of the sterol-biosynthetic pathway in microalgae. Environ. Microbiol. Reports 6:35-44. Pubmed

• Villanueva, L., N. Bale, E. C. Hopmans, S. Schouten, and J. S. Sinninghe Damsté. 2014. Diversity and distribution of a key sulpholipid biosynthetic gene in marine microbial assemblages. Environ.Microbiol. 16:774-787. Pubmed

• Villanueva, L., S. Schouten, and J. S. Sinninghe Damsté. 2014. Depth-related distribution of a key gene of the tetraether lipid biosynthetic pathway in marine Thaumarchaeota. Environ.Microbiol. in press. Pubmed

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  Sterol biosynthetic pathway

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  Sulfolipid molecule

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  Phylogeny of archaeal geranylgeranylglyceryl phosphate synthases

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Thaumarchaeota diversity and ecophysiology

Members of the Thaumarchaeota phylum have been found to be ubiquitous in marine, freshwater, soils (and others) environments. So far it has been assumed that all thaumarchaeota are chemolithoautotrophs and ammonia oxidizers based on the presence of a unique carbon fixation pathway and the gene coding for ammonia monooxygenase (amoA gene). These physiological characteristics, as well as the relative high abundance of this group in some environments, suggest an important role of Thaumarchaeota in the carbon and nitrogen cycles. In addition, it has been observed that all cultured representatives of the Thaumarchaeota uniquely synthesize the glycerol dialkyl glycerol tetraether (GDGT) crenarchaeol (with 4 cyclopentane and a cyclohexane moiety), which is considered as a biomarker for the presence of this group. The relative abundance of thaumarchaeotal membrane lipids (GDGTs with zero to 4 cyclopentane moieties, GDGT-0 to GDGT-4, and crenarchaeol) has been shown to be correlated with the temperature at which these organisms are growing. Based on this the TEX86 (TetraEther indeX of tetraethers consisting of 86 carbon atoms) paleotemperature proxy was developed and further tested to reconstruct the temperature in past environments.

TOOLS

• Metagenomics and metatranscriptomics
• Quantification of the abundance and gene expression of 16S rRNA, metabolic genes and genes coding for enzymes involved in the lipid biosynthetic pathways.
• Phylogenetic and evolutionary approaches
• Lipid detection and quantification

REFERENCES

• Pitcher, A.*, Villanueva, L.*, Hopmans, E.C., Schouten, S., Reichart, G-J., and Sinninghe Damsté, J.S. 2011. Niche segregation of ammonia-oxidizing archaea and anammox bacteria in the Arabian Sea oxygen minimum zone. ISME J. 5:1896–1904. *equal contributors Pubmed

• Schouten, S., Pitcher, A., Hopmans, E.C., Villanueva, L., van Bleijswijk, J., and Sinninghe Damsté, J.S. 2012. Intact polar and core glycerol dibiphytanyl glycerol tetraether lipids in the Arabian Sea oxygen minimum zone: I. Selective preservation and degradation in the water column and its consequences for the TEX86. Geochim. Cosmochim. Acta 98: 228–243. Link

• Buckles, L.*, Villanueva, L.*, Weijers, J., Verschuren, D., and Sinninghe Damste, J.S. 2013. Linking isoprenoidal GDGT membrane-lipid distributions with gene abundances of ammonia-oxidising Thaumarchaeota and uncultured crenarchaeotal groups in the water column of a tropical lake (Lake Challa, East Africa). Environ. Microbiol. 15:2445-2462. *equal contributors Pubmed

• Bale, N.*, Villanueva, L.*, Hopmans, E.C., Schouten, S., and Sinninghe Damste, J.S. 2013. Different seasonality of pelagic and benthic Thaumarchaeota in the North Sea. Biogeosciences 10:7195–7206. *equal contributors Link

• Villanueva, L., S. Schouten, and J. S. Sinninghe Damsté. 2014. Depth-related distribution of a key gene of the tetraether lipid biosynthetic pathway in marine Thaumarchaeota. Environ.Microbiol. In press. Pubmed

• Lipsewers, Y.A., Bale, N.J., Hopmans, E.C., Schouten, S., Sinninghe Damsté, J.S., and Villanueva, L. 2014. Seasonality and depth distribution of the abundance and activity of ammonia oxidizing microorganisms in marine coastal sediments (North Sea). Front Microbiol. Sep 5;5:472 Pubmed

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Anaerobic bacteria involved in Nitrogen and Methane cycles

The re-mineralization of organic matter in anoxic sediments is mainly driven by fermentative microorganisms, sulfate reducers, and methanogens. However, there is a general lack of knowledge on the diversity, abundance and activity of the anaerobic microorganisms involved (directly or indirectly) in organic matter recycling in anoxic sediments.

Methanogens (strictly anaerobic archaea) biologically produce methane, a trace greenhouse gas in the earth’s atmosphere the concentration of which has doubled since industrialization. On its way to the atmosphere, methane travels through anaerobic sediments, passing through zones dominated by different regimes of anaerobic respiration before reaching the aerobic sediment or oxic water column. Along this route methane can be oxidized, which significantly decreases/mitigates the effective emission of this greenhouse gas to the atmosphere. However, it is unclear how ecosystems to different physicochemical conditions with respect to methane production, consumption and thus ultimately emissions to the atmosphere.

Our aim is to improve our understanding of the microbial players involved in anaerobic organic matter remineralization and also specifically focus on those involved in the methane cycle. We also want to assess their individual niches, metabolic pathways, environmental significanc, interactions, and their response to environmental changes. Ultimately this information will be key to explore their potential use in biotechnology and to design mitigation strategies for greenhouse gas emission.

This research will be conducted in the framework of the Soehngen Institute for Anaerobic Microbiology (Gravitation grant- Zwaartekracht, from the Dutch Ministry of Education, Culture and Science, read more here) in which the Radboud University, Wageningen University, Delft University of Technology, and NIOZ Royal Netherlands Institute for Sea Research participate.

TOOLS

• Incubation studies and activity rate measurements
• ‘Omic’ approaches (DNA/RNA and lipids)
• Discovery of new lipid biomarkers produced by anaerobic microbes

REFERENCES related to this topic

• Bale NJ*, Villanueva L*, Fan H, Stal LJ, Hopmans EC, Schouten S, Sinninghe Damsté JS. 2014. Occurrence and activity of anammox bacteria in surface sediments of the southern North Sea. FEMS Microbiol Ecol. 89:99-110. *equal contributors Pubmed

• Villanueva, L., Speth, D.R., van Alen, T., Hoischen, A., and Jetten, M.S.M. 2014. Shotgun metagenomic data reveals significant abundance but low diversity of “Candidatus Scalindua” marine anammox bacteria in the Arabian Sea oxygen minimum zone. Front. Microbiol. 5:31. Pubmed

• Moore, E., Hopmans, E.C., Rijpstra, W.I.C., Villanueva, L., Dedysh, E., Kulichevskaya, I., Wienk, H., Schoutsen, F., and Sinninghe Damsté. J.S. 2013. Novel mono-, di-, and trimethylornithine membrane lipids in northern wetland planctomycetes. Appl. Environ. Microbiol. 79:6874-6884. Pubmed

• Lipsewers, Y.A., Bale, N.J., Hopmans, E.C., Schouten, S., Sinninghe Damsté, J.S., and Villanueva, L. 2014. Seasonality and depth distribution of the abundance and activity of ammonia oxidizing microorganisms in marine coastal sediments (North Sea). Front Microbiol. Sep 5;5:472 Pubmed

See also

• Kool, D.M., Zhu, B., Rijpstra, W.I.C., Jetten, M.S.M., Ettwig, K.F., Sinninghe Damsté, J.S. Rare branched fatty acids characterize the lipid composition of the intra-aerobic methane oxidizer Methylomirabilis oxyfera. Applied and Environmental Microbiology 78, 8650-8656. Pubmed

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  Sampling campaign in the Lake Grevelingen (The Netherlands)

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  Anaerobic sediment

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  Phosphatidyl choline (PC)-monoether C20-[3] ladderane molecule

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  Jaap, Laura and Stefan celebrating the Gravity grants