Previous studies have shown that bacterial membrane lipids are sensitive to changes in environmental parameters such as soil pH (e.g. Weijers et al,. 2007) and have enabled the development of the CBT index, a soil pH proxy based upon the number of cyclopentane rings within these lipids. Although the CBT proxy has evolved in recent years (see De Jonge et al., 2014 for more details), it is also known that archaeal membrane lipids are sensitive to changes in pH (e.g. Weijers et al., 2006). In this edition of Papers ‘n’ Cake, we discussed a recent article by Wei Xie and colleagues (Xie et al., 2015) which explores the distribution of (thaum)archaeal membrane lipids in soils and their potential application as a soil pH proxy.
Before we dig into the details, we’ll plow through a bit of background – why we care about soil pH, what factors control soil pH, and what are the organisms Xie and colleagues studied in such detail?
Why care about soil pH?
1. Plants: Soil “reaction,” also known as soil pH, refers to the acidity (low pH) or alkalinity (high pH) of a soil. Soil reaction is a commonly used indicator of soil fertility and health because it pertains to a number of different soil properties that impact plant growth. Most importantly, soil pH reflects what chemicals are soluble in soil water and thus the nutrients available for plants to take up … or for rain to wash away. This, of course, is critically important for conservation and agriculture as well as home gardening. Indeed, UK agricultural companies apply an estimated 1.6 to 2 million tonnes of lime (CaCO3) per year to raise soil pH, partially to counter the pH-lowering, knock-on effects of nitrogen and sulphur-bearing fertilizers and pollutants.
2. Temperature: In organic geochemistry, we are often interested in soil pH because of its confounding effect on continental temperature proxies (Weijers et al., 2006). For example, the MBT/CBT proxy is based upon the distribution of bacterial membrane lipids, where the degree of methylation, expressed as the MBT ratio (methylation of branched tetraethers), is related to MAT and pH, and the number of cyclopentane rings, expressed as the CBT ratio (cyclisation of branched tetraethers), is related to soil pH (Weijers et al., 2007). As the CBT ratio is related to pH it can be used to remove the pH impact on the MBT record.
3. Biogeochemistry: From a biogeochemical perspective, soil pH is also interesting in that it tends to correlate with soil nitrogen cycling as both respond to similar suites of environmental conditions through impacts on nitrogen species in soil water or adsorbed on clay minerals, and on N-cycling bacteria and archaea.
So, what controls soil reaction and how great a range is observed in natural soils?
Although several processes impact soil reaction, the predominant long-term controls are: 1) the chemical and mineralogical composition of the soil and 2) precipitation and drainage regime. For example, a soil developed on chalk will likely be more alkaline than a soil formed on clay or igneous rock. Similarly, in sites with high precipitation and efficient drainage, rainwater dissolves and re-precipitates carbonate resulting in more acidic conditions at the surface and potentially more alkaline in the lime layer lower in the profile (See the USDA’s Illustrated Soil Guide). These processes combine to result in soil pH that spans the scale from extremely acid (pH < 3.5 to 4.4) to strongly alkaline (pH 8.5 to 9.0), with acidic soils more common in wet regions and alkaline soils in arid. However, this is quite a generalization as soil pH in the UK alone ranges from 3.6 to 9.0 (Emmett et al., 2010). Over timescales of seasons to decades, vegetation type, farming practices (e.g. crop rotation, tilling depth, liming) and pollution can also have large (up to several pH units) impacts on soil reaction (e.g. Emmett et al., 2010). On seasonal or shorter timescales, temperature and respiration can further modify soil pH to the extent of about half a pH unit (e.g. from a pH of 5.5 to 6.0). Importantly, because of interactions and co-variation of contributing factors, soil reaction may also correlate with mineralogy, organic carbon content, temperature, and soil moisture, each of which might also impact the diversity and metabolisms of soil microorganisms.
In sum, soils are tremendously diverse environments and many properties, of which pH is only one, are heterogeneous even on small spatial scales and can vary on centennial to seasonal time scales. Accordingly, soil scientists strongly recommend sampling soils from a large number of cores and conducting analyses at the same time of year. Thus, Xie et al. might expect to see some change in soil reaction on a seasonal basis due to soil moisture and respiration rates. However it might not be a large difference compared to the scale of variability vertically within a soil or laterally among sites.
What are Thaumarchaeota, 16S rDNA and amoA, and why might that matter?
Firstly, Thaumarchaeota are a group within the domain Archaea. For context of what a “domain” means, consider that we, along with everything from roses, Volvox and jaguars to kelp, sponges, ctenophores, yeast and Euglena, are all part of the morphologically diverse but somewhat metabolically restricted domain Eukarya. The archaeal domain is fundamentally different from Eukarya and Bacteria: Archaea are single-celled microorganisms that do not have a nucleus like Eukarya, but have a different cell membrane composition than Bacteria. Although Archaea come in a much smaller range of shapes and sizes than eukaryotes, they have huge genetic diversity, some thrive in conditions that few to no eukaryotes can tolerate, and are able to make a living using a somewhat broader range of chemical reactions than we. It’s a somewhat simplistic comparison, but as a phylum Thaumarchaeota would be comparable to, for example, the eukaryote phylum Arthropoda, which includes all “insects,” that is, all the crunchy, exoskeleton-bearing creatures like lobsters, spiders, barnacles, and dragonflies. So, when we talk about Thaumarchaeota, we mean a group of archaea unequivocally united by genetic commonalities but by no means uniform.
Much of our knowledge of how such very different organisms relate to one another originates with the next mysterious term in this list: 16S rDNA. This portion of the genome is a popular focus for evolution studies because it is highly conserved, that is, 16s rDNA is present in all known organisms and relatively slow to change as the organisms evolve over time. Other sequences, such as the alpha subunit of the gene that encodes for the ammonia monooxygenase enzyme (amoA), are more specific to a particular group of organisms. In this case, archaeal amoA, which controls production of an enzyme essential for ammonia oxidation, is common to many Thaumarchaeota and exists as several different gene variants within that group. So, by analysing the diversity of archaeal 16S rDNA and amoA genes extracted from the soil, Xie and colleagues are able to identify the diversity of all Archaea that are present, and then to look within the archaea that produce amoA genes (i.e. Thaumarchaeota) without having to fully sequence the whole genomes of all the Archaea in the soil. Interestingly, one knock-on consequence of this type of analysis is that it’s not actually possible to say for certain which amoA gene goes with which 16S rDNA sequence. Because the authors use quantitative PCR we can expect some positive relationship between the abundance of Thaumarchaeota (i.e. amoA copy number) in the soil and the abundance of their lipids.
Great, but why look at Thaumarchaeota? Should we expect a relationship with soil reaction?
Until fairly recently, bacteria were thought to dominate prokaryotic life in soils, however culture-independent microbiological tools indicate that archaea are also abundant, and important contributors to terrestrial N-cycling (Leninger et al., 2006). Thaumarchaeota in soils are generally thought to make a living through ammonia oxidation, using O2 to convert NH3 into NO2– in a process akin to sulphide oxidation (see Sabine & Gordon’s post). Thus, Thaumarchaeota are important as one of the key players in the soil nitrogen cycle. The extent to which proxies derived from soil bacteria and archaea also reflect N-cycling is an interesting topic and one that Xie et al.’s combined biomarker and molecular biology approach begins to explore.
Interestingly, recent evidence suggests that not all Thaumarchaeota are obligate ammonia oxidizers; Group 1.1c thaumarchaea derived from acidic forest soils appear able to thrive without evidence for ammonia oxidation (Weber et al., 2015). The 1.1c Thaumarchaeota studied by Weber and colleagues (2015) do not possess an amplifiable copy of amoA, and so would be missed by that part of Xie et al.’s study, but if sufficiently abundant could nonetheless contribute to the soil lipid pool.
There are long-standing environmental (Nicol et al., 2008) and theoretical (summarized by Xie and colleagues) indications of a relationship between Thaumarchaeotal diversity and soil pH. Recent analyses suggest that specialization for different soil pH conditions played an important role in the early diversification of Thaumarchaea (Gubry-Rangin et al., 2015). Gubry-Rangin and colleagues (2015) suggest that Nitrosopumilus and Nitrosotalea in acidic niches and Nitrososphaera – as Xie and colleagues observed – in soils with alkaline reaction share a common neutrophilic stem group thaumarchaea ancestor. Thaumarchaeota from groups 1.1a and 1.1b appear to be associated with higher pH, lower water content, and lower soil organic carbon content (Vico Oton et al., 2015). Given the apparently strong relationships between 1.1a and 1.1b versus 1.1c thaumarchaea and soil pH, one might well expect to see a seasonal difference in diversity accompanying a sufficiently large change in soil reaction. However, the rate of adaptation to seasonal changes, the magnitude of seasonal soil reaction change, and the resolution of the relationship between thaumarchaeal diversity and pH might all cloud this hypothesis.
Lipid-wise, a recent synopsis suggests that isoGDGTs 0, 1-4 and crenarchaeol are all found among the 1.1a and 1.1b Thaumarchaeota (Schouten et al., 2013). Since the number of cultured Thaumarchaeota available for lipid analysis is not very high (25 cultures, none of which are 1.1c, Lehtovirta-Morley et al., 2014) it is difficult to assess whether isoGDGT profiles and relationship to pH differ among these different groups. How the potential diversity of Thaumarchaeota plays out at a single site, whether thaumarchaeal community structure responds coherently and resolvable on a seasonal timescale, and whether turnover of lipids is rapid enough to record such changes are all cogent questions that Xie and colleagues attempt to address.
The paper: Temporal variation in community structure and lipid composition of Thaumarchaeota from subtropical soil: insights into proposing a new soil pH proxy (Xie et al., 2015).
In the paper, Xie et al. (2015) investigate the Archaeal community structure and the membrane lipid GDGT composition in soils from Chongming Island, China. The paper combines molecular biology and organic geochemistry approaches throughout a year on a monthly basis assessment. The authors seek to examine the temporal variability on GDGT and archaeal community and how those respond to seasonally driven environmental changes. Additionally, a new soil pH proxy based on archaeal GDGT is proposed, which is tested against bacterial GDGT soil proxy to assess its applicability and robustness as an alternative to soils pH estimates.
No systematic changes in archaeal community or lipid composition were evident on a seasonal basis, or those were below signal-to-noise (Fig. 2A). Crenarchaeol was the dominant isoprenoid GDGT and Nitrososphaera the most representative archaeal group, based on 16S rDNA sequencing.
In this study, isoGDGTs correlate with changes in soil pH. In particular, crenarchaeol abundance (a proposed biomarker for Thaumarchaea) correlates positively with soil pH as previously demonstrated by Weijers et al. (2006), while isoGDGT-1 and isoGDGT-3 correlate inversely with soil pH. Xie et al. thus propose a new soil pH proxy, the Thaumarchaeota Index (TI, Fig. 1 and 2B), which was tested on a) Chongming Island soils, b) the global soil dataset (Weijers et al., 2007), and 3) Thaumarchaeota enrichment cultures. The new TI proxy successfully reconstructs soil pH on Chongming Island, albeit within a narrow pH range. However on a global basis, the CBT performs better and gives more reliable soil pH correlations.
Archaeal GDGT- vs Bacterial GDGT-based soil pH proxy
The TI (Xie et al., 2015) appears robust on seasonal basis over a short-time scale and in restricted environmental conditions. However, when extrapolated to a global perspective and broad environmental variability, the TI does not reproduce soil pH as accurately as the CBT does. This restricts the application of the TI proxy (Fig 2B). However, TI seems to be more sensitive to short-term changes in soil pH , at least in the local context of Chongming Island. Although still premature, TI could emerge as tracer of local changes in soil pH. Nevertheless, it demands more investigations to secure the application with reliable interpretations
Absence of systematic seasonal changes on Archaeal 16S and amoA sequences and GDGT distribution
The lack of strong evidence of seasonal changes in both GDGT composition and Archaeal community may be biased by the experimental design, which does not cover extensively environmental differences. The study is restricted to one single location, where environmental conditions presents low variability (pH: 7.7 – 8.3; TOC: 0.8 – 1.6%; C/N: 6.4 – 13.3; NH4+: 14.4 – 39.0 µm). Only temperature (7.9 – 29.7 ºC) and soil water content (0.8 – 25.8%) present some variability. Therefore such conclusion may not be representative for a wide soil perspective. This is reflected on low agreement with a global soil database for the new proposed pH soil index. Additionally, the local character of this is study does not explore archaeal growth rates under different environmental conditions (i.e. soil mineralogy, nutrients availability, TOC content, and pH) and seasons, neither if it is similar or variable through time. Consequently, lipids production and turnover time are not fully resolved. Which groups of archaea would be predominant and therefore which isoprenoid GDGTs would be produced under a variety of environmental soil conditions? This is a key point underestimated here, which leads to the final point, the extension of the dataset. Answering questions about seasonal and environmental driven changes requires a wider dataset, which should comprehend more than only one year over a single location. Otherwise, such conclusions are not representative of a wider perspective.
Chongming Island soils present no/low seasonal variability, as indicated by archaeal 16S, amoA sequences and the isoGDGT composition. To what extent this is transferable to other soil environments is debatable. Crenarcheol abundance positive correlation with soil pH allow its use a soil pH proxy by means of the Thaumarchaeota Index. Although TI works well in a narrow pH range (7.6 to 8.4), it has limited applicability over a wider pH range. As such, alternative pH proxies based upon bacterial membrane lipids should be utilised in the first instance (e.g. CBT; Weijers et al., 2007; De Jonge et al., 2015).
De Jonge C, Hopmans E, Zell C, Kim J-H, Schouten S, Sinninghe Damsté J, 2014. Occurrence and abundance of 6-methyl branched glycerol dialkyl glycerol tetraethers in soils: Implications for palaeoclimate reconstruction. GCA. 141. 97-112
Emmett BA, Reynolds B, Chamberlain PM, Rowe E, Spurgeon D, Brittain SA, Frogbrook Z, Hughes S, Lawlor AJ, Poskitt J, Potter E, Robinson DA, Scott A, Wood C, Woods C, 2010. Soils Report from 2007, CS Technical Report No. 9/07, Centre for Ecology and Hydrology, NERC. Revised 23 Feb 2010.
Gubry-Rangin C, Kratsch C, Williams T, McHardy A, Embleyc T, Prosser J, and Macqueen D, 2015. Coupling of diversification and pH adaptation during the evolution of terrestrial Thaumarchaeota. PNAS 112:9370-9375.
Lehtovirta-Morley LE, Ge C, Ross J, et al., 2014. Characterisation of terrestrial acidophilic archaeal ammonia oxidisers and their inhibition and stimulation by organic compounds. FEMS Microbiol Ecol 89:542–52
Leninger S, Urich T, Schloter M, Schwark L, Qi J, Nicol GW, Prosser JI, Schuster SC, Schleper C, 2006. Archaea predominate among ammonia-oxidizing prokaryotes in soils. Nature 442:806–809.
Nicol GW, Leininger S, Schleper C, Prosser JI, 2008. The influence of soil pH on the diversity, abundance and transcriptional activity of ammonia oxidizing archaea and bacteria. Environmental Microbiology 10:2966–2978.
Schouten S, Hopmans EC, Sinninghe Damsté JS, 2013. The organic geochemistry of glycerol dialkyl glycerol tetraether lipids: A review. Organic Geochemistry 54:19-61.
Weber E, Lehtovirta-Morley LE, Prosser JI, Gubry-Rangin C, 2015. Ammonia oxidation is not required for growth of Group 1.1c soil Thaumarchaeota. FEMS Microb Ecol 91.
Weijers JWH, Schouten S, Spaargaren OC, Sinninghe Damsté JS, 2006. Occurrence and distribution of tetraether membrane lipids in soils: Implications for the use of the TEX86 proxy and the BIT index. Organic Geochemistry 37:1680–1693.
Weijers JWH, Schouten S, van Den Donker JC, Hopmans EC, Sinninghe Damsté JS, 2007. Environmental controls on bacterial tetraether membrane lipid distribution in soils. Geochimica et Cosmochimica Acta 71:703–713.
Vico Oton E, Quince C, Nicol GW, Prosser JI, 2015. Phylogenetic congruence and ecological coherence in terrestrial Thaumarchaeota. The ISME Journal.
Soil Analysis: an interpretation manual https://books.google.co.uk/books?id=pWR1vUWbEhEC&pg=PA104&lpg=PA104&dq=soil+pH+measurement+seasonal&source=bl&ots=aYA9pClWWs&sig=l4ypJ5c75erFZXEs21w8Kv1aLo8&hl=en&sa=X&ved=0CD0Q6AEwBGoVChMIrN3Tva-cyQIVC1caCh0MpQYl#v=onepage&q=soil%20pH%20measurement%20seasonal&f=false
Soil orders with descriptions and examples:
US Department of Agriculture soil factsheets: http://www.nrcs.usda.gov/wps/portal/nrcs/detailfull/soils/health/assessment/?cid=stelprdb1237387
Illustrated guide to soil identification:
Soil Survey Staff. 2015. Illustrated guide to soil taxonomy, version 2. U.S. Department of Agriculture, Natural Resources Conservation Service, National Soil Survey Center, Lincoln, Nebraska.