Mars Microbiology

I. Microbial Energy Production

Respiration

In the process known as respiration, microorganisms break down organic compounds, such as sugars, as food to produce energy. Another way of saying this is that the organic compounds are oxidized. The organisms use the energy to power cell functions and make cell material. Anaerobic respiration occurs in the absence of oxygen (nevertheless we still say that the the organic compounds used as fuel are oxidized in the process) but produces much less energy per sugar molecule than aerobic respiration, although the latter requires molecular oxygen (O₂) as a substrate.

Because the Martian atmosphere does not contain significant amounts of O₂ astrobiologists do not believe that aerobic respiration occurs on Mars. However, if Martian life forms exist, then some form of anaerobic respiration may be part of their metabolism and if so they would be capable of metabolizing small organic compounds. Therefore, one of the experiments sent to Mars during the 1970s as part of Project Viking was designed to look for evidence of anaerobic respiration in the Martian soil. The experiment, designed by Gilbert Levin was known as the Labeled Release (LR) Experiment, because it employed a solution of test nutrients, labeled with carbon-14 (¹⁴C). In other words each carbon atom of each molecule of nutrient in the LR test solution was of the isotope ¹⁴C, instead of the more common ¹²C and ¹³C. However, unlike the other two carbon isotopes, ¹⁴C is radioactive, so its presence can be detected with a radiation sensor.

At both Viking landing sites when Martian soil was placed within a special container and treated with the LR test solution, radioactivity was detected in the air space above the soil. This means that the some or all of the ¹⁴C-containing liquid nutrients were converted chemically into ¹⁴C-containing gases, because only a gas would rise above the soil sample to produce a radioactive signal in the overhead space. For example, one of the the ¹⁴C-containing nutrients in the LR test solution was formic acid which in many terrestrial microorganisms is oxidized to carbon dioxide (CO₂) during the process of respiration, whether aerobic or anaerobic. Here is the reaction, which in many species requires two enzymes as well as co-factor called NAD⁺.

The radioactivity was detected following treatment with nutrient solution only in those samples that were not heated prior to testing. In samples that were heated to 160 degrees C for three hours before testing, the results were negative, suggesting that microorganisms had been the resson for the positive response in unheated samples but had been killed in the heated samples. To appreciated the value of using heat sterilization as a control for a life detection expetiment, try Classroom Experiment # 1 (see section II).

Initially the Viking scientists were very excited about these results. However, because another instrument designed to look for carbon-containing (organic) compounds produced negative results, most of the scientists began to think that a non-biological oxidizing agent, such as hydrogen peroxide (H₂O₂) had mimicked the effects of life on the LR test solution –like this:

Photosynthesis

Photosynthetic microorganisms: Microcystis aeruginosa strain PCC 7806. Autofluorescence image.

From Cyanobacterial Image Gallery http://www-cyanosite.bio.purdue.edu/images/images.html

In a way, photosynthesis is the opposite of respiration. In photosynthesis, the carbon contained within CO₂ is used to make organic nutrient compounds through a process called reduction, the opposite of oxidation. Photosynthetic organisms can then oxidize the compounds through respiration or they can be saved and/or used by other organisms. Thus, photosynthetic organisms use to CO₂ make their own food! In order to do this, they need sunlight and water and as a by product they produce O₂, which they can use during respiration in the dark or which can be used by non-photosynthetic organisms –humans for example.

Plants are famous for being photosynthetic but did you know that many microorganisms are also photosynthetic? During Project Viking, scientists tested for the possibility that photosynthetic microorganisms might inhabit Mars. They employed the Pyrolytic Release Experiment (PR) developed by Norman Horiwitz. In a way, the PR was the opposite of the LR. While the LR treated Martian soil samples with ¹⁴C-containing liquid nutrients, the PR exposed soil samples to ¹⁴C-containing gases, CO₂ and carbon monoxide (CO), because both of these gases are present in the Martian atmosphere. After exposure to the gases, the soil was studied to see whether it can taken in any radioactivity, which would indicate that the ¹⁴C-containing CO₂ and/or CO had been reduced to organic compounds within the soil.

When the PR experiment was performed, very small amounts of carbon from the gases were found to have become part of the soil, indicating reduction. However this occurred in both unheated samples and those heated prior to testing, suggesting that the effect had been the result of some non-living agent/s process in the soil. Also, the small amount of reduction occurred both in the light and the dark and thus the experiment did not detect photosynthesis.

Electron micrograph of strain Nankai-1 that shows a typical coccoid methanogen

Mikucki, JA, et al. Applied and Environmental Microbiology, June 2003, p. 3311-3316, Vol. 69, No. 6

Methanogenesis

Methanogenic microorganisms produce the gas CH₄ (methane). On Earth, the substrates usually are H₂ and CO₂ both of which are gases at standard temperature and pressure. In the process of synthesizing methane, the carbon atom is changed from its most oxidized state (in CO₂) to its most reduced state (in CH₄) and energy is produced, energy which the organisms use to make food and cell material. Here is the overall reaction:

4H₂ + CO₂ CH₄ + 2H₂O

For methanogenic organisms, the methane is a waste product, released into the atmosphere, just as CO₂ is released as waste by humans. Earth’s atmosphere thus contains some methane, although compared the concentration of nitrogen and oxygen, the levels of methane in our atmosphere is very very tiny. Even so, Earth’s atmospheric methane can be detected from space, using special instruments.

If life forms exists on Mars, it is plausible that some of them might use methanogenesis for energy. It is possible that the gas released during the Viking Labeled Release Experiment (LR) was CH₄ rather than CO₂, but this is not the only reason why a Martian version of methanogen may inhabit the Red Planet. In 2004 a European Space Agency (ESA) probe, Mars Express, detected methane in the Martian atmosphere. Although volcanic activity is a possible non-biological source of methane, there is no evidence for contemporary volcanic activity on Mars. On the other hand, Mars Express also detected trace amounts of HCHO (formaldehyde). Formaldehyde is an intermediate of methanogenesis, a compound produced along the way during the biosynthesis of methane from H₂ and CO₂. It cannot be made through volcanic activity. ESA plans to continue to study the Martian atmosphere and scientists in both ESA and NASA plan additional probes in the near future to confirm the Mars Express findings. But if it turns out that the Martian atmosphere contains methane with a sprinkling of formadehyde, it would mean that methanogenic microorganisms are native to Mars. Stay tuned.

II. Classroom Experiments

1. Take some baker’s yeast and divide it into two portions. Heat one sample for 3 hours at 160 degrees C. For each sample (heated and unheated) place in nutrient solution (which teacher will prepare). The solution will be made from PBS to which sugar or vinegar will be added as food for the yeast. Let the yeast incubate in the nutrient solution, then after 2 hours look for bubbles. Compare the result for the heated and unheated yeast.

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