Rocks and their microbes: a co-evolutionary partnership

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Miles beneath our feet, Earth’s rocky crust may seem a cold, dead place. On closer inspection it’s anything but.

Microbes have been making a home on and in rocks since…well, since the beginnings of life, some 3.5 billion years ago. The traditional view of rock-dwelling microbes is one of sparse, energy-starved survivalists eking out an existence on a hostile frontier. These critters are at the mercy of their environments, their numbers dictated by physical factors – things like temperature and pH.

Geobacter, a rock-dwelling bacteria coating iron oxide minerals. Credit: Wikipedia

Geobacter, a rock-dwelling bacteria coating iron oxide minerals. Credit: Eye of Science

A study published in Geomicrobiology Journal, led by graduate student Aaron Jones at the University of Texas at Austin, challenges the assumption that subsurface microbes are passive survivalists. In a series of lab experiments, Jones demonstrates that mineral-dwelling microbial communities are uniquely adapted to their environments on the scale of individual mineral grains. Moreover, his research suggests microbes and minerals share a partnership that stretches back through geologic time.

Jones was interested in how mineralogy- the composition of the minerals that form rocks- influences the abundance and diversity of microbes. In particular, he was interested in sulfur oxidizers- bacteria that strip electrons from sulfur to acquire energy. Sulfur oxidizers are common in other subsurface environments including deep ocean vents. Their metabolism generates strong acids that eat away at certain minerals, forming cracks, pores, even caves.

In the lab, Jones set up a series of “bioreactors”- tanks in which liquid of a specific chemical composition flows over thin sheets of rock. He used a range of rocks that differ in their mineralogy. These included several rocks composed of calcium carbonate (limestone, dolostone and calcite) and several made of silica-rich minerals (albite, microcline and quartz). Jones inoculated his bioreactors with samples of a sulfur-metabolizing biofilm collected from a Wyoming cave spring.

Example of a benchtop bioreactor. Bioreactors are commonly used in microbial biotechnology for cell culturing and fermentation. Credit: Wikimedia Commons

Example of a benchtop bioreactor. Bioreactors are commonly used in microbial biotechnology for cell culturing and fermentation. Credit: Wikimedia Commons

After three weeks of incubation, he measured bacterial growth, extracted DNA and used pyrosequencing to examine the composition and diversity of the microbial communities formed on different rock types.

Different rocks, different microbes

Growth patterns differed substantially across the carbonate and silicate rocks. On carbonates, bacteria grew in thick, filamentous biofilms. By contrast, growth was much sparser on silicates, with bacteria either accumulating in small clumps or spreading out individually.

Community composition also differed among different rocks. The bacterial phyla Alphaproteobacteria dominated on most carbonates. Sulfur oxidizers that thrive at near-neutral pH were common, representing 17-45 % of the community.

A different picture emerged on the silicate rocks. Members of the phyla Firmicutes and Actinobacteria- also known as Gram-positive bacteria– were found here. While community composition was relatively similar among the carbonates, communities that grew on quartz were very different than those found on albite or microcline. Jones observed fewer sulfur oxidizers on silicates, and those he found tended to be more acid-tolerant.

Composition of bacterial communities grown on different mineral surfaces. From Jones & Bennet 2014.

Composition of bacterial communities grown on different mineral surfaces. From Jones & Bennet 2014.

Perhaps the most obvious conclusion here is that microbes do not select rocks at random. Different growth patterns among rocks tell us this. Better microbial growth on carbonates probably reflects a higher density of essential nutrients, particularly phosphorus. Moreover, two of the silicate rocks, albite and microcline, contain high concentrations of aluminum, a toxic element that restricts bacterial growth.

More subtly, however, there appears to be “selective relationship” between mineralogy and microbial ecology. In other words, the microbes making a living on any given rock are uniquely adapted to taking advantage of, or protecting themselves from, the specific chemical properties of that rock.

This relationship bears itself out in many ways. For one Jones found more acid-tolerant microbial communities on silicate rocks. Remember, acid is a byproduct of sulfur metabolism. Carbonate rocks have a high “buffering capacity”, meaning their minerals react with and neutralize acid. By contrast, silicate rocks, especially quartz, are pretty unreactive. Rather than eating away at minerals, acid accumulates at silicate surfaces. The presence of acid-loving bacteria on quartz suggests a feedback loop between sulfur metabolism, mineralogy and microbial community structure.

The presence of Gram-positive bacteria on silicates also underscores the relationship between microbial community composition and the mineral environment. The cell walls of Gram-positives possess unique chemical properties that promote bonding with silicate surfaces. This microbe-mineral binding, known as adhesion, is a key element of survival for most rock-dwelling bacteria. By adhering to a solid surface, bacteria can mine nutrients and form protective biofilms.

Even microbes living on the undesirable aluminum-rich rocks may possess unique adaptations. While few bacteria grew on albite or microcline, these sparse populations were highly diverse. The unique challenges presented by a toxic environment may serve to maintain competition, preventing any single group of organisms from gaining dominance.

Our planet’s rocky subsurface is the largest microbial habitat, but we still know very little about its inhabitants. Most microbial ecologists prefer to study environments they can see, feel and intimately relate to. Jones’s research points to complex ecological interactions between microbes and minerals in the deep, dark depths of the world. Rocks shape microbial communities, but microbes may also shape rocks in profound ways. This study lends evidence to a deep synergy between the geological and biological evolution of our planet.

ResearchBlogging.org

Jones, A., & Bennett, P. (2014). Mineral Microniches Control the Diversity of Subsurface Microbial Populations Geomicrobiology Journal, 31 (3), 246-261 DOI: 10.1080/01490451.2013.809174

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When infection is a good thing: sulfur-eating bacteria enlist viruses to help acquire energy

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“Black smokers”, deep sea vents where hot, sulfur-rich fluids bubble up from beneath the ocean floor, are considered hotspots of microbial activity. They may even be where life originated. Credit: Wikimedia Commons

Life is no cake walk at the ocean floor, where carbon is scarce and light nonexistent.  At least near deep ocean vents, mineral-rich water bubbles up from magma beneath the crust, providing both heat and a source of energy. In these alien environments, lithotrophs– bacteria that eat minerals instead of organic carbon- have staked out a niche by evolving some creative metabolic strategies.

But minerals are a poor source of energy compared to organic matter. Lithotrophs are slow-growing critters, easily outcompeted when carbon is abundant. They need all the help they can get. And it turns out, they might very well get help… from an unlikely source. A study published last week in Science Express reports how viruses may be helping deep marine bacteria eat sulfur.

Viruses are everywhere- in soils, skies, oceans, plants and animals, even deep beneath the ocean floor.  In spite of their ubiquity, it’s not well-known how viruses influence marine bacteria and the nutrient cycles they drive.

To study the impact of viruses on sulfur-eating bacteria, a team of scientists led by Dr. Karthik Anatharaman at the University of Michigan collected samples from five different hydrothermal vents : four in the Western Pacific Ocean and one in the Gulf of California. They used shotgun metagenomic sequencing to look at the genomes of the bacteria and viruses present in these environments.

The researchers found five different virus “types” that carried two genes involved in sulfur metabolism. Known as rdsrA and  rdsrC, these genes encode different pieces of dissimilatory sulfite reductase, an enzyme that breaks down elemental sulfur.

How and why did viruses come to acquire these genes? Not technically considered alive, viruses don’t really need enzymes because they don’t perform metabolism on their own. A virus’s sole purpose is to infect a host with its genetic code so that it can turn the host cell into a virus factory. But during viral replication, pieces of  DNA can be accidentally snipped out of the host and integrated into the viral genome. The scientists found bacteria in the same samples carrying both the rdsrA and rdsrC genes. It’s likely that accidental incorporation of host DNA is how viruses ended up with sulfur genes in the first place.

While acquiring sulfur genes may have been a complete accident, viruses appear to have been actively maintaining those genes for a long time.

When the scientists compared rdsrA and rdsrC from viral and bacterial genomes, they found something interesting: completely different DNA sequences surrounding the genes. If viruses had acquired sulfur genes during recent replication errors, some of the nearby DNA would probably have come along for the ride as well. That completely different DNA sequences surround the sulfur metabolism genes in bacteria and viruses suggests viruses have been passing along rdsr genes, generation to generation, for a very long time. In other words, sulfur metabolism genes are being maintained  in viruses by natural selection.

Why would deep sea viruses bother to maintain sulfur metabolism genes they don’t use?  It turns out these viruses may actually have a use for sulfur genes, albeit an indirect one: helping out their hosts. By carrying genes that bacteria can make use of, viruses may assist their hosts in acquiring energy. The ability of sulfur-eating bacteria to acquire energy is ultimately limited by how quickly they can decode their sulfur genes to build enzymes that can do the work. But if a bacteria were to contain viruses with extra copies of the genes it needs, that could help speed the process along. In essence, when a sulfur virus infects a sulfur bacteria, it donates genes that its host can use.

What’s a virus to gain from all of this? Remember, a virus’s sole purpose is replication. Staying in your host’s good graces has its perks. An “infected” bacteria that can acquire energy faster than its neighbors may be able to grow and reproduce faster- leading to more infected offspring, and more viruses. By supplementing host metabolism, viruses may help ensure their continued survival.

This study sheds light on a potentially widespread, mutually beneficial ecological interaction between bacteria and viruses. Lithotrophs in the deep ocean play an important role in the global sulfur cycle and have done so for billions of years. By giving an adaptive advantage to sulfur-eaters and helping them survive, it’s possible viruses have played an equally important role in the geochemical evolution of our planet.

ResearchBlogging.org

Anantharaman, K., Duhaime, M., Breier, J., Wendt, K., Toner, B., & Dick, G. (2014). Sulfur Oxidation Genes in Diverse Deep-Sea Viruses Science DOI: 10.1126/science.1252229

Crystalline nanotube-forming bacteria help crumble mountains in a tropical rainforest

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Mineral weathering, the slow crumbling, dissolving, and chemical transformation of solid bedrock into smaller fragments and clays, replenishes Earth’s surface with the raw materials needed to make soil and the elements needed to support life.  In the Luquillo mountains of Puerto Rico, hot rain beats incessantly onto the ground and seeps deep into the earth, creating cracks and fissures in solid rock. While a warm, wet climate, frequent hurricanes and mudslides that shake up the landscape all contribute to the rapid weathering of these tropical mountains, a recently published study indicates crystal-forming bacteria may also be important contributors to the most rapid chemical weathering rates recorded on the planet.

Luquillo rainforest in northeast Puerto Rico- a steep, mountainous, tropical forest that has been the site of extensive ecological research for decades

Luquillo rainforest in northeast Puerto Rico- a steep, mountainous, tropical forest that has been the site of extensive ecological research for decades

Historically, scientists have studied weathering as an abiotic (i.e., non-living) process. However, there is growing evidence that living organisms, from bacteria and fungal mycelia  to treescan actively participate in the replenishment of fresh rock material to the soil. In the mid 1980’s, scientists studying geothermal sediments from Yellowstone found the some of the first evidence of biological weathering when they discovered iron-silicate minerals in association with bacterial remains. These bacterial cell fragments, the scientists hypothesized, can act as mineral “nucleation” sites – surfaces to which minerals can condense and grow. If you’ve ever made rock candy, you’ve formed sugar crystals by this same nucleation process.

Snow crystals vizualized by a scanning electron microscope. Like other types of crysals, the formation of snow crystals is promoted by nucleates- things for ice to condense and grow onto. Credit:  Agricultural Research Service, United States Department of Agriculture

Snow crystals vizualized by a scanning electron microscope. Like other types of crysals, the formation of snow crystals is promoted by nucleates- things for ice to condense and grow onto. Credit: Agricultural Research Service, United States Department of Agriculture

Why is crystal nucleation important in weathering? As primary minerals, the stuff coming directly out of rocks, are broken down, they can undergo a series of chemical transformations into a variety of secondary minerals. Many of these secondary minerals, including clays, become important raw materials for the formation of new soils.

Which brings us back to Luquillo. Here, Earth scientists from many different fields- geology, ecology, atmospheric chemistry, and hydrology to name a few, are attempting to build a holistic picture of the processes that shape landscapes. For nearly a decade, a team of scientists now affiliated with the Luquillo Critical Zone Observatory have been drilling holes in the ground to study what makes the rocks crumble. Deep holes. Mineral weathering is so rapid in these mountains that they must drill through five or even ten meters of saprolite- soft, mineral rich material, no longer solid rock but not quite soil yet- before hitting bedrock. What have they found? For one, there are more bacteria living near bedrock– at that contact zone between hard rock and soft saprolite, where weathering is actively occurring-  than higher up. Roughly a hundred times more. This led researchers to ask a simple question- if bacteria are concentrated at the so-called “weathering front”, are they just passive bystanders, or do they have an important role to play in the transformation of rock into soil?

To answer this question, a research team led by Dr. Susan Brantely, a geochemist at Penn State University, drilled several holes to bedrock, collected samples from the contact zone between saprolite and rock, and used high-resolution scanning electron and transmission electron microscopy (SEM and TEM) to investigate the structure and composition of actively weathering minerals. While SEM provides information on the 3-d structure of an object, the higher resolution TEM technique can provide information on crystal bonding structure and help determine the elemental composition of a sample.

Their findings? In most of the samples, the scientists identified halloysite, a secondary mineral that forms long, tube-like crystals. Halloysite is a natural weathering product in young soils and its presence was not entirely surprising.The really exciting part came when they identified small round and rod-shaped cells buried amidst the so-called “crystalline nanotubes”. Bacteria, sheathed in crystals.  Moreover, all of the bacterial cells observed were intact, suggesting that being coated in crystals does not damage these hardy bacteria. The presence of living bacteria within secondary minerals also implicitly suggests longevity:  halloysite crystals can take weeks or months to form.

Scanning electron microscopy image (50,000x) showing a coccus-shaped bacteria surrounded by halloysite nanotubes

Scanning electron microscopy image (50,000x) showing a coccus-shaped bacteria surrounded by halloysite nanotubes. Credit: Minyard et al. 2011.

That bacteria can serve as condensation nuclei for secondary minerals in Luquillo has important ecological implications. For instance, in this  study, halloysite minerals often occurred in higher densities around bacteria than inorganic surfaces. This may be an indication that some bacteria are evolved to be highly efficient nucleating agents. These mineral-formers may have  structures attached to their cell membranes which have a high affinity for particular mineral elements, such as silica.

Scanning electron microscope images of bacteria associated with secondary minerals nearly five meters deep in Luquillo saprolite. From Minyard et al. 2011.

Scanning electron microscope images of bacteria associated with secondary minerals nearly five meters deep in Luquillo saprolite. Credit: Minyard et al. 2011.

Bacteria that serve as mineral nucleation sites may also be able to influence mineral chemistry. Other researchers have found that sulfur, considered to be an “impurity” is more concentrated in halloysite when bacteria are present. Iron, another common impurity in halloysite, was found in many of the samples in this study. The presence of iron could indicate the crystal-forming bacteria are iron oxidizers, organisms that use iron for fuel instead of carbon. Perhaps serving as mineral nucleation sites allows these bacteria to access iron they need for energy.

Scanning electron microscope image of iron oxidizing bacteraia Acidovorax sp. BoFeN1, encrusted in iron minerals. Credit: Eye of Science, Reutlingen

Scanning electron microscope image of iron oxidizing bacteraia Acidovorax sp. BoFeN1, encrusted in iron minerals. Credit: Eye of Science, Reutlingen

Many questions remain to be answered regarding the relationship between microorganisms and weathering. What sorts of organisms can withstand being coated in minerals? What are the physiological adaptations that allow them to do so, how and when did these adaptations evolve? Traditionally, geologists have considered the lithosphere- the rocky part of the earth- to be a cold, dead place. Clearly we need to start rethinking that assumption, and considering what it may mean for our geologic understanding of the planet, if organisms can find an evolutionary space buried amidst the very rocks our biosphere sits on.

Welcome to the deep biosphere

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Remember in Journey to the Center of the Earth when Hans, Lindenbrock and Axel discover giant insects, mastodons and other prehistoric creatures miles beneath the surface? While this was considered science fiction in the mid 19th century, I think most of us today would be more apt to call this story pure fantasy. We know now that such an idea is preposterous. Plants need sunlight to obtain their energy, and animals need plants for food. There is no energy source capable of sustaining complex, multicellular life far away from the sun. The biosphere as we know it can only exist on the surface, where ample sunlight and moderate temperatures prevail.

Single celled life is a bit more creative. Recent drilling expeditions beneath the surface of the continents and the ocean floor have revealed a surprisingly large number of bacteria and archaea living well beneath the surface of the earth- 3 x 10^29 cells according to current estimates!* The scientists behind these projects have coined the term “deep biosphere” to describe the areas of the earth beneath the surface that are still occupied by single-celled life. A great mystery surrounding the deep biosphere is how such communities can survive in environments that seemingly provide minimal energy. In short, how are these critters making a living?

So that we’re all on the same page here, I’ll just briefly review the three basic requirements for life as we know it on planet Earth:

1) A carbon source

2) An energy source

3) A pair of molecules that can donate and accept electrons.

More on the third one later, which relates to how organisms actually break down and harvest energy inside their cells. For now, let’s suffice it to say all life requires an energy source and a carbon source. For us, both of these requirements are satisfied by eating food. The metabolism of our food releases energy. As  we break down the complex organic molecules contained in our food, we are provided with simple carbon molecules which can be rebuilt into different molecules for growth and cellular repair. All animals use organic carbon as both an energy and a carbon source, because of the high energy yields associated with this strategy. By contrast, plants use light as an energy source and carbon dioxide from the atmosphere as a carbon source. Light energy is used to power a series of biochemical transformations, ultimately allowing plants to convert carbon dioxide into sugar: photosynthesis. Thus even in the world we are familiar with there exist two vastly different strategies for meeting life’s basic requirements.

Might the microbes of the deep biosphere choose one of these metabolic strategies? Clearly, the second one is dependent on light, which I probably don’t need to tell you is in rather short supply miles beneath the ocean floor. But what about organic carbon? Is enough organic carbon from, say, dead plant, animal and microbial bodies in the surface ocean or soils making its way into subsurface environments to sustain microbial communities?

At a first approximation, the answer is probably not. Best estimates are that < 1 % of the carbon fixed by plants on the planet’s surface is making it into the “buried organic carbon” pool- the stuff that might be available as food for deep biosphere organisms. Compared with the estimated size of the biomass inhabiting the subsurface, this flux of carbon provides, at best, enough energy to carry out only the most basic life support functions, certainly not enough for growth and reproduction.

What else could be feeding the deep biosphere? The answer is a bit complex, and something I will be exploring in much more detail throughout future posts. Many different options present themselves. Some microbes can use metals, such as iron, as an energy source, allowing them to convert carbon dioxide to sugar without light. This strategies is known as chemolithotrophy.

Dormancy might be another option for some inhabitants of the deep biosphere: in essence, forming a spore and entering a state of metabolic inertia until better conditions present themselves in the future. It is known that only certain groups of bacteria have this capability, but how widespread this strategy might be in the deep biosphere is uncertain. There is an argument to be made against dormancy in the deep biosphere; namely, that it is an evolutionary dead end. Dormancy might make sense in, for instance, a seasonal rainforest, where dry season conditions force microbes to go into hibernation but the onset of the wet season allows them to “wake up” and start growing again. But in a world that is always harsh, always lacking in energy, for thousands and millions of years, would a dormant cell ever wake up? Would it ever reproduce?

Finally, there may be sources of energy available to the deep biosphere that we simply do not understand yet. Perhaps there are microbes that can harvest energy released during the radioactive decay of elements in the earth’s crust? Or perhaps heat energy from even deeper inside the earth fuels the deep biosphere?

In short, there are many questions and many mysteries surrounding life in the deep biosphere. As scientists continue to answer these questions the answers will undoubtedly expand our understanding of what it means to be alive.

*Kallmeyer, J., Pockalny, R., Adhikari, R. R., Smith, D. C. & D’Hondt, S. Global distribution of microbial abundance and biomass in subseafloor sediment. Proc. Natl Acad. Sci. USA 109, 16213–16216 (2012).