Autor: Georgii Feodoridi

            Kseniia Feodoridi

Place: HYDITEX CORPORATION, North Cyprus

Date: July 2024

Abstract: Hydrogen-oxidizing microorganisms have recently increasingly attracted the attention of microbiologists. The fact is that these microorganisms use molecular hydrogen to produce energy, which, as you know, is considered as the fuel of the future. Hydrogen-oxidizing microorganisms can provide answers to important questions related to the exploration, production, storage and transportation of so-called white-gold (geological) hydrogen. 

Hydrogen-oxidizing microorganisms are not a taxonomic group, but organisms united on the basis of a number of characteristics, their number and diversity are enormous. They are present in various geological environments such as soils, marine muds, sedimentary rocks, salt and soda lake waters and soils containing various engineering industrial sites, etc. and have a great influence on them. 

In this article, we examined in some detail the various areas of application of Hydrogen-oxidizing microorganisms, their diversity and division into groups, the various geological environments in which they are present, as well as the importance that they have for them, including for underground engineering industrial facilities. The main directions requiring further study of these microorganisms were formulated.

Key Words: white hydrogen, geological hydrogen, natural hydrogen, underground hydrogen, hydrogen oxidizing microorganisms, hydrogen exploration, biogeochemistry of hydrogen, biogeochemical, hydrogen.

DOI: 10.13140/RG.2.2.15496.43527

Biogeochemical methods for hydrogen exploration. Hydrogen-oxidizing microorganisms.

Hydrogen-oxidizing microorganisms, in some sources they are called hydrogen bacteria, are a group of microorganisms that include not only bacteria, but archaea that use the oxidation of molecular hydrogen to produce energy. 

The number of microorganisms capable of oxidizing hydrogen and their diversity present in soil, sea mud, sediment, etc. is a good guide to find and determine the size of the hydrogen resource underlying them. This direction is a very promising use of microorganisms, the so-called biogeochemical method of searching for places of generation and release of hydrogen from deep natural accumulations for subsequent production. 

The fact is that natural difficult hydrogen coming from the bowels of the Earth is conditioned because when passing through the medium (soil, marine sediments, etc.) it observes the observations present in them. The method is based not on the search for power itself, but on the search for biomass products. Microorganisms accumulate anywhere, but the microbial biomass is much higher due to the fact that the flow of population passing through the environment significantly increases its possible formation of other microflora. 

For exploration purposes, the most promising geological settings are used where deposits of natural hydrogen are most likely to be found, such as igneous mafic and ultramafic rocks that are rich in hydrogen in the form of dissolved gas and captured by fluid inclusions in ophiolites, rift zones, faults and atmospheric degassing in volcanic gases , geysers, hot springs and surface gas outlets. Coal seams, deep basement faults, which can provide migration and, in favorable places, concentration of diffuse sources of H2. Ring subsidence structures” and some researchers call them “fairy circles”, which themselves cannot be concentrators of natural hydrogen deposits, but indicate that degassing of natural hydrogen occurs in the area. [1]. 

In the above-mentioned places, a selection of media (soils, silt, rocks, etc.) is taken and the biomass production present in them is assessed from a biochemical point of view. Methods for estimating biomass depend on the environment where the microorganisms live. 

The use of biogeochemical methods makes it possible to quickly explore large areas of prospecting work and identify promising areas with a possible release of natural hydrogen flows, including finding hidden objects.  

Thus the number and variety of hydrogen oxidizing microorganisms present in soil, sea mud, sediment, etc. is a good guide to find and determine the size of the hydrogen resource underlying them. 

Geological environments in which hydrogen-oxidising microorganisms are present

Hydrogen is an attractive substrate for growth because it contains a large amount of energy. Hydrogen oxidation is a chemosynthesis reaction and is one of the important inorganic energy sources capable of releasing a relatively large amount of energy (237 kJ/mol-h2). Therefore, hydrogen oxidizing organisms play a key role in the ecosystem [2, 3, 4]. The flow of molecular hydrogen as it leaves the Earth significantly affects the properties of soils, marine silt, sedimentary rocks, etc. 

The mechanism of influence of hydrogen-oxidizing microorganisms on the soil in places where hydrogen is released is as follows: 

Along with molecular hydrogen coming from the bowels of the Earth, it is also one of the most important gaseous microcomponents of the soil, in which it is formed as a result of anaerobic decomposition of various organic residues by microorganisms. According to some data, its content in the soil air of automorphic soils is 1–8–6% [5]. This formation of molecular hydrogen is carried out by strict and facultative anaerobic microorganisms, the main producers being primary anaerobes [6, 7]. 

In normal communities of microorganisms, the concentration of hydrogen is very low due to its interspecies transfer - one species receives energy in a reaction leading to the release of hydrogen, the other oxidizes it using an acceptor inaccessible to the first. The first organism can only exist if the hydrogen concentration is kept vanishingly low. This is a well-organized trophic system in which anaerobic organisms serve as producers of gases from decomposing organic matter, and specific groups of aerobic organisms that oxidize hydrogen do not allow it to escape from the soil air into the atmosphere. This position made it possible to consider the soil both as a kind of bacterial filter and as a gas system of varying degrees of closure depending on moisture conditions [5, 7, 8]. 

Near the places where molecular hydrogen emerges from the bowels of the Earth, the production of biomass of hydrogen bacteria is much greater due to the fact that a current of hydrogen passes through it, significantly exceeding its possible formation by soil microflora. As you know, soil pores are partially filled with water and partially with air. At a sufficiently high concentration of organic matter to ensure the absorption of diffusing O2, water-filled pores become habitats for anaerobic organisms. Since the rate of gas transfer is greater than that of dissolved substances, and the diffusion coefficients in soil for H2 at normal temperatures are 7 times higher than for CO2 and 3 times higher than for O2, it turns out that H2 can serve as a transport vehicle for the removal of electrons from anaerobes, and thus ensure the decomposition of organic matter under anaerobic conditions [6, 7]. 

Soils contain a large and diverse group of hydrogen bacteria capable of using molecular hydrogen. This ability is determined by the presence of hydrogenases in them, which catalyze the oxidation of molecular hydrogen. The energy exchange of hydrogen bacteria carries out the reaction of water synthesis [9, 10]. Most hydrogen bacteria require molecular oxygen to grow. Some can grow on purely mineral media in the presence of carbon monoxide. [7] 

According to [11], hydrogen H2 is an important energy source for seawater communities oxidising hydrogen not only in soil but also in seawater, and those in turn mitigate atmospheric H2 emissions [12] and potentially account for undersaturation of H2 in Antarctic waters [11, 13]. 

Surface layers of the world’s oceans are generally supersaturated with H2 and CO, typically by 2- to 5-fold (up to 15-fold) and 20- to 200-fold (up to 2,000-fold) relative to the atmosphere, respectively [14, 15, 16, 17]. H2 is primarily produced by cyanobacterial nitrogen fixation [18]. High concentrations of H2 are also produced during fermentation in hypoxic sediments, and these high concentrations can diffuse into the overlying water column, especially in coastal waters [19]. As a result, oceans contribute to net atmospheric emissions of these gases [11, 12, 20]. 

According to [11] biogeochemical, metagenomic and thermodynamic modelling analyses together suggest that H2 is oxidized by a diverse but small proportion of community members, but at sufficiently fast cell-specific rates to enable lithotrophic growth. These findings are supported by experimental observations that the ultramicrobacterium Sphingopyxis alaskensis (Strain RB2256) consumes H2 during heterotrophic growth [11]. 

Hydrogen-oxidising microorganisms are also found in groundwater. Moreover, studies [21] have shown that hydrogen is the main energy source ensuring high productivity of old groundwater, which contains biomass-rich microbial communities [21]. 

This study showed that there were significantly more cells in aquifers with older groundwater than in aquifers with younger water.    This suggests that the older aquifers and geochemically evolved groundwaters are in fact productive ecosystems that provide energy for the growth of microorganisms [21]. 

The increase in cell numbers with groundwater age was accompanied by a substantial decrease in archaeal and bacterial diversity  Decreases in diversity and shifts in community structure are common features of microbial bloom situations in productive ecosystems, which often feature elevated abundances of a few key species [21, 22, 23]. 

Also, this study showed that, in general, microbial cell numbers across the entire studied region, did not decrease with depth. Cell counts were on average slightly higher in aquifers in geologic strata that contained shales or coal, suggesting that elevated contents of organic carbon may provide additional energy sources to microbes [21]. 

Microorganisms that oxidize hydrogen are also present in desert soils [24]. Organisms residing in these ecosystems endure severe and prolonged drought interspersed with infrequent hydration pulses, often in combination with other physicochemical pressures [25, 26, 27, 28].  

Drought limits the abundance and productivity of oxygenic photoautotrophs, namely, cyanobacteria, microalgae, and plants, which require water as an electron donor. Low supplies of organic carbon, combined with impaired substrate diffusion and membrane transport, in turn limit the abundance of chemoheterotrophic microorganisms and fauna [28, 29]. The extents of nutrient limitation vary: semiarid deserts experience considerable precipitation and subsequent productivity, albeit counterbalanced by evapotranspiration, whereas hyperarid deserts can be deprived of rainfall over decadal scales [30, 31]. In spite of these pressures, cultivation-independent surveys have shown that microorganisms are moderately abundant in most desert soils and can be as diverse as those of humid soils [24, 32, 33]. 

Traditionally, most bacteria are thought to persist by using organic carbon synthesized by photoautotrophs following transient hydration events. Recent studies focused on Antarctic desert soils have revealed, however, that some bacteria use atmospheric trace gases, such as hydrogen (H2) to conserve energy and fix carbon independently of photosynthesis [24]. 

Recent studies showed that the dominant microorganisms in Antarctic desert soils are metabolically flexible bacteria capable of using both organic and inorganic energy sources, most notably atmospheric hydrogen. Through genome-resolved metagenomics, it was shown that bacteria from phyla such as Actinobacteriota encode specific high-affinity hydrogenases (group 1h and 1l [NiFe]-hydrogenases) to oxidize atmospheric H2. Activity studies have shown that electrons derived from H2 oxidation can support aerobic respiration and, for some cells capable of the Calvin-Benson-Bassham cycle, carbon dioxide fixation [34, 35]. Based on this evidence, it has been inferred that these bacteria are capable of “living on air” [24, 36]. 

Moreover, in contrast to water-soluble substrates, gaseous substrates readily diffuse through cell membranes and are more available to cells in drier soils as a result of diffusion through air-filled soil pores [29, 37, 38]. Thus, atmospheric H2 oxidation may provide a minimalistic way for bacteria to sustain energy and carbon needs in otherwise energy-depleted desert environments [24]. 

Together, these findings highlight that H2 is an important, hitherto-overlooked energy source supporting bacterial communities in desert soils. Contrary to our previous hypotheses, however, H2 oxidation occurs simultaneously rather than alternately with photosynthesis in such ecosystems and may even be mediated by some photoautotrophs. [24]. 

The number of microorganisms that can use hydrogen under aerobic and anaerobic conditions is enormous. To date, researchers have discovered and isolated microorganisms that oxidize hydrogen from ordinary environments such as soil, rivers, lakes, and extreme environments such as deep sea, hydrothermal fluid, volcanic craters. The use of hydrogen as an energy source is widespread in hydrothermal vent symbioses [39]. Hydrotherms populated by bacteria and archaea (hyperthermophiles) create enormous biomass not only on the surface of the bottom, but also in the depths of the ocean [40]. 

Based on a set of data on the growth conditions of hydrogen-oxidising microorganisms (methanogens, homoacetogens and sulphate reducers) in the environment [41], their detection sites (sources) can be divided into 14 species: 

1. Sources associated with oil production, these include, for example, an oil producing well, oil pipeline water, oil-polluted estuarine sediment, etc.; 

2. Deep-sea sources, e.g. microorganisms isolated from a deep-sea hydrothermal vent chimney; 

3. The black smokers; 

4. Marine mud, marine sediment, marine sediment; 

5. Hot spring and thermal spring, and solfataric sea floor sediment; 

6. Rice field soil; 

7. Bacteria isolated from human or animal organisms or waste products, e.g. human and animal faeces, from termite gut, from bovine rumen, etc.; 

8. freshwater sediment, freshwater mud, mud and sediments of lakes and rivers, etc.; 

9. Waste water, sewage sludge, bioreactors, a pond polluted with sewage water; 

10. Soil including compost soil (compost soil); 

11. Sediments from hypersaline soda lakes, mud and water of salt water lakes, etc.; 

12. Brines and permafrost sediments; 

13. Estuarine sediments; 

14. Sources associated with gas and ore deposits, e.g. microorganisms isolated from water extracted from a coal bed, from a uranium mine, from gas-associated formation water from a well of a natural gas field, etc. 

Other sources such as plants, clay, vinegar, peat bogs, algae, etc. are also among the places where bacteria are found.   

All these data suggest that hydrogen-oxidising microorganisms are present in many environments, but at the same time specific studies [42], have been carried out to assess the risk of microbial growth in underground hydrogen storage facilities, which found that environments with temperatures > 122 °C are sterile with respect to hydrogen-consuming microorganisms  in particular to hydrogenotrophic sulfate reducers, that couple H2-oxidation to sulfate reduction to produce hydrogen sulfide (H2S); hydrogenotrophic methanogens that reduce carbon dioxide (CO2) to methane (CH4) by oxidizing H2; and homoacetogens that couple H2 oxidation to carbon dioxide (CO2) reduction to produce acetate [2, 43, 44, 45].

Types of hydrogen-oxidising microorganisms

The study of microorganisms that oxidize hydrogen began in the laboratory of Professor Hans Schlegel, who gained worldwide fame for the publication of the textbook “General Microbiology,” which was published in Germany in the early 1960s. [44]. But their first descriptions were given simultaneously by A.F. Lebedev and H. Kaserer in 1906 [46] from enriched cultures in an atmosphere of hydrogen, oxygen and carbon monoxide., although the biological nature of the process of molecular oxidation hydrogen in soil was established somewhat earlier. 

Microorganisms that oxidize hydrogen are not a taxonomic group, but organisms grouped together on the basis of several physiological characteristics. Hydrogen bacteria include representatives of 20 genera, uniting gram-positive and gram-negative forms of different morphologies, motile and immobile, forming spores and non-spores, reproducing by division and budding [46]. 

Microorganisms that oxidize hydrogen are divided into three main groups, which include: [47] 

facultative autotrophs (facultative chemolithoautotrophic), 

aerobic hydrogen-oxidizing bacteria. 

anaerobic hydrogen-oxidizing bacteria. 

Let's take a closer look at these groups.

Facultative autotrophs (facultative chemolithoautotrophic) 

Compared to the other hydrogen-oxidizing bacteria, those described as facultative autotrophs can fix carbon dioxide. However, they can also use various organic substrates as a source of energy [47]. 

Because of their ability to use organic and inorganic sources, these species are widely distributed in nature. They can be found in aquatic and terrestrial environments including swamps, rice fields, the gut of some animals, deserts, compost, and soils [47]. 

Collectively, these bacteria are also known as Knallgas bacteria, those that can fix carbon dioxide through the oxidation of hydrogen and using oxygen as the terminal electron acceptor [47]. 

Though Knallgas-bacteria need oxygen as the terminal electron acceptor, the majority of species grow well in microaerophilic conditions. This is largely due to the fact that the enzyme hydrogenase required for hydrogen oxidation is affected/inhibited by oxygen [47]. 

Some species like H. Pylori have been shown to grow in elevated oxygen levels in the laboratory. Facultative autotrophs can also grow under completely heterotrophic conditions [47]. 

Generally, carbon dioxide fixation using hydrogen as the electron donor is represented as follows (This process is usually called the Knallgas reaction) [47]: 

 6H2 + 2O2 + CO2 = CH2O + 5H2O   

 The enzyme involved in this process is known as hydrogenase. There are two types of hydrogenase enzyme which include cytoplasmic NAD-specific hydrogenase (also known as hydrogen hydrogenase) and the membrane-bound hydrogenase.  

Some members of this group contain one type of the enzyme while others have been shown to contain both. For species that contain the two types of the enzyme, hydrogen is first oxidized by the membrane-bound hydrogenase. Electrons are then transported through the electron transport chain to the quinones and cytochromes.  

The second hydrogenase enzyme (cytoplasmic hydrogenase) serves to generate reducing power (NADH) that is eventually used to fix carbon dioxide in the Calvin cycle. 

Knallgas bacteria that oxidize hydrogen include [48]: 

Cupriavidus necator (e.g. Strain N-1 and H 1) 

Hydrogenovibrio marinus (e.g. Strain DSM 11271) and others 

One of the best studied facultative autotrophs is the bacterium Aquaspirillum autotrophicum [47] (Herbaspirillum autotrophicum, e.g. Herbaspirillum autotrophicum strain SA 32 and Herbaspirillum autotrophicum strain SA 33), a member of the family Neisseriaceae. Commonly found in eutrophic freshwater environments (e.g. eutrophic lakes), the bacterium is characterized by a rod-shaped morphology and bipolar flagella.  

Other bacteria belonging to this group include: 

Proteobacteria 

Aquificales 

Firmicutes 

Actinobacteria 

This group, Knallgas-bacteria, comprises bacteria from different taxonomic units. For this reason, they display diverse characteristics in morphology, ecology, and nutrition. Whereas Proteobacteria consists of Gram-negative species (pathogens and free-living), the phylum Firmicutes consists of Gram-positive members which are involved in carbohydrate metabolism in the gut [47]. 

Aquificales, on the other hand, are mostly Gram-negative species that can be found in extreme/harsh environments such as hot springs [47]. 

• Proteobacteria 

Proteobacteria is a large phylum of Gram-negative bacteria. Some of the genera within this group include Escherichia, Salmonella, Legionellales, and Vibrio, etc.  Escherichia (e.g. Escherichia coli, Strain CCUG 45711 B and DSM 19683),  Legionellales (e.g. Legionella nagasakiensis CDC-1796-JAP-E, Legionella sainthelensi MSH-4), Salmonella, Vibrio, etc. [47]. 

Most members of the group have flagella used for movement. Others do not have flagella and rely on gliding to move from one point to another. Some of the species (e.g. some members of the class Alphaproteobacteria like Rhizobium species) are free-living [47]. (e.g. Bradyrhizobium japonicum Strain 3I1b6). 

Gammaproteobacteria, a class of the phylum Proteobacteria is an example of hydrogen-oxidizing bacteria. Common in the marine environment, these bacteria have been shown to be capable of producing the enzyme hydrogenase (Fe hydrogenase) that is used for hydrogen oxidation [47]. 

• Aquificales 

Aquificales is an Order of bacteria within the phylum Aquificae. Members of this group are Gram-negative rods that grow in areas with a temperature range between 60 and 90°C (in hot springs, thermal ocean vents, and sulfur pools, etc.) [47]. 

Some species within this group include: 

Thermocrinis ruber (Strain OC 14/7/2 and OC 1/4)  

Calderobacterium hydrogenophilum (Hydrogenobacter hydrogenophilus strain Z-829) and others 

Members of the genus Aquifex (e.g. Aquifex aeolicus) are some of the most popular hydrogen oxidizing bacteria in the group. Like the other hydrogen oxidizing species, they produce hydrogenase (Hydrogenase I) that allows them to fix carbon dioxide (chemolithoautotrophs) [47]. 

• Firmicutes 

The phylum Firmicutes consists of Gram-positive bacteria. However, some of the species (e.g. Pectinatus species) cannot retain the primary stain because of the porous outer membrane [47]. 

They exhibit significant variation in morphology from spherical (cocci) to rod-shaped (straight or curved). They also produce spores that allow them to survive harsh environment conditions, high temperatures or high salinity [47]. 

Some of the genera within this group include: 

Enterococcus  

Ruminicoccus 

Bacillus 

Clostridium 

Paenibacillus (e.g. Paenibacillus sp. DSM 26193) and others 

Members of the phylum Firmicutes can be found on terrestrial environments, in plants, and guts of various animals [47]. 

A good number of species within this group contain Fe-hydrogenase which is used in microaerophilic conditions [47]. 

• Actinobacteria 

Members of this group are Gram-positive rods (slender) that are characterized by high G+C content in their genome. They can be found in aquatic (marine sediments) and terrestrial environments across the globe [47]. 

The majority of species are filamentous and form branched filaments that resemble mycelium. The phylum Actinobacteria consists of a wide variety of organisms that are beneficial for the environment contributing to the carbon cycle [47]. 

They produce different types of products that continue to be used in medicine, agriculture, textile, and other industries [47]. 

Some members of the Phylum Actinobacteria include: 

Micromonospora 

Nocardioform  

Streptomyces 

Nocardia (e.g. Pseudonocardia autotrophica 394) 

Rhodococcus and others 

Like many other hydrogen-oxidizing bacteria, Members of the Phylum Actinobacteria have also been shown to be responsible for hydrogen uptake [47]. 

Mycobacterium smegmatis (e.g. Strain Mycobacterium smegmatis GA 735), a member of the family Mycobacteriaceae, uses two membrane-associated, oxygen-dependent hydrogenases (NiFe hydrogenases) for hydrogen oxidation. Here, oxygen acts as the electron receptor [47]. 

Among the facultative autotrophs, it is worth noting the anaerobic hyperthermophilic bacterium Caldimicrobium rimae (phylum Thermodesulfobacteria). This microorganism is capable of fixing carbon dioxide during growth due to the oxidation of hydrogen associated with the reduction of thiosulfate or sulfur, or due to the disproportionation of sulfur [49, 50, 51, 52].

Aerobes that oxidize hydrogen

Compared to Knallgas-bacteria (which fix carbon dioxide), aerobic hydrogen-oxidizing bacteria use hydrogen without fixing carbon dioxide. Here, however, it's worth noting that some of the bacteria described as Knallgas-bacteria are aerobic (need oxygen as the electron acceptor). As well, not all aerobic hydrogen-oxidizing bacteria are Knallgas-bacteria [47]. 

Some of the bacteria that use hydrogen without fixing carbon dioxide include [47]: 

• Acetobacter species  

The genus Acetobacter consists of acetic acid bacteria. Members of this group are characterized by their ability to convert ethanol to acetic acid aerobically. Morphologically, Acetobacter species may be ovoid or rod-shaped. Rod-shaped species are straight or curved (between 1.0 and 4.0 um in length) [47].  

Depending on the species, they may occur singly or form short chains. Some of the species move by means of flagella (peritrichous bacteria). Across the world, Acetobacter species (Acetobacter hansenii) are widely distributed in vegetation and are responsible for the rot of fruits like pears and apples (e.g.Acetobacter hansenii (Komagataeibacter hansenii, Strain DSM 5602 and 336)) [47].  

Using hydrogenase, species like Acetobacter peroxydans, can take up hydrogen and use it as an electron donor. Reduction of hydrogen peroxide by this bacterium also results in the production of hydrogen which is used to reduce Quinone and oxygen [47]. 

• Azotobacter species 

Azotobacter is a genus of the phylum Proteobacteria. Members of this group are Gram-positive organisms that form thick-walled cysts. They may be oval or spherical in shape; ranging from 2 to 10um in size depending on the species [47]. 

Species like Azotobacter vinelandii move by means of flagella. However, these structures are usually lost during encystment. Though they are ubiquitous in nature, Azotobacter species are commonly found in habitats with neutral to weakly basis pH [47]. 

Examples of species in this group include [47]: 

Azotobacter vinelandii (e.g. Strain 16 and 3a) 

Azotobacter salinestris (e.g. Strain Identifie 184) 

Azotobacter chroococcum (e.g. Strain 43, DSM 369 and Ag) 

Azotobacter beijerinckii (e.g. Strain 3, S 16 and A 17) and others 

In species like Azotobacter vinelandii, there are three types of nitrogenase isoenzymes that produce hydrogen as a by-product. This hydrogen is then re-oxidized [47]. 

Like Knallgas-bacteria, these bacteria belong to different taxonomic groups and therefore exhibit a range of different characteristics. For instance, while the three groups consist of Gram-negative species, Enterobacteriaceae bacteria have a rod-shaped morphology and some of the species can move by means of peritrichouse flagella [47].

Azotobacter species can be oval or spherical in shape and mostly live freely in the soil. Lastly, members of the genus Acetobacter are commonly found in fermented foods, rotting pears, and apples and are characterized by their ability to oxidize lactate and acetate [47].  

Like some of the Knallgas bacteria, Azotobacter vinelandii, a member of the genus Azotobacter, contains a membrane-bound hydrogenase enzyme involved in the reversible oxidation of hydrogen (molecular hydrogen) [47].  

Under aerobic conditions, the bacterium, using the enzyme, oxidizes dihydrogen to produce protons and electrons. These electrons are then transported, through the electron transport chain, to oxygen which acts as the electron acceptor [47]. 

This process has also been observed in Acetic acid bacteria. Here, however, a number of molecules can act as hydrogen acceptors in the absence of oxygen [47]. 

Aerobic microorganisms that oxidize hydrogen also include: 

Wautersia eutropha 335

Acidovorax facilis DSM 649

Roseateles saccharophilus DSM 654 

Afipia carboxidovorans OM5

Hydrogenophaga pseudoflava GA3

Paracoccus denitrificans CCUG 30144

Xantobacter autotrophicus 7C

Mycobacterium Gordonae W-1609

and others

Anaerobes that oxidize hydrogen

Like aerobic species, bacteria that oxidize hydrogen anaerobically do not fix carbon dioxide. Some of the most popular members of this group include [47]: 

Acetobacterium woodii (e.g. Strain DSM 1030) 

Acetobacterium woodii is a member of the genus Acetobacterium. Like the other Acetobacterium, it's an anaerobic, rod-shaped Gram-positive organism. It can be found in several habitats including dust air, aquatic sediments, and various soil habitats. Acetobacterium woodii is anaerobic and does not produce spores. The bacterium produces the enzyme hydrogenase. Here, however, hydrogen utilization occurs anaerobically [47]. 

Clostridium aceticum (e.g. Strain DSM 1496) 

A member of the genus Clostridium, Clostridium aceticum is a Gram-positive organism found in anaerobic habitats (e.g. sewage sludge). Like some of the other members in the genus, Clostridium aceticum is rod-shaped and moves by means of flagella (peritrichous flagella). Chemoorgranotrophically, Clostridium aceticum uses such compounds as fructose and L-glutamate for growth. However, it can also use hydrogen and carbon dioxide to grow under anaerobic conditions; chemolithotrophically [47].  

Also anaerobic microorganisms that oxidize hydrogen include: 

Nitratidesulfovibrio vulgaris

Methanobrevibacter arboriphilus DH-1

Acetobacterium woodii DSM 1030

Acetobacterium wieringae С

Wolinella succinogenes DSM 1740

and others

Use of hydrogen-oxidising microorganisms

Currently, microorganisms that oxidize hydrogen attract much attention due to the use of biotechnologies in different directions: 

To remove carbon dioxide from the atmosphere or from industrial sources in order to implement the “net zero emissions” program. 

For local air regeneration, for example for the exploration of the Moon and Mars. There have been experiments on CO2 consumption in confined spaces - space stations, where the crew constantly releases CO2 when breathing, which is a big problem - supplying the crew with oxygen and removing CO2. One way to solve the problem was to use hydrogen bacteria, since they consume CO2, but also consume hydrogen. Hydrogen and oxygen can be produced by electrolysis of water [44]. 

In the biochemical industry, to produce natural bioplastics. When hydrogen-rich bacteria grow in mineral-poor environments, they form bioplastics, about 10% of the cell. When transplanted onto a carbon substrate, where the carbon and nitrogen content in the environment is changed, which is important for regulating the process, hydrogen bacteria increases the synthesis of bioplastic up to 90% of their weight. Cells growing in media with glucose can form up to 60 g. class/dry weight, which is a high yield of natural plastic  [44]. Such bioplastics are degradable. 

To produce food protein. In 2024, a company in Northern Europe is launching the production of protein made with hydrogen-oxidising bacteria. These are essentially dried microbes created by producing and dehydrating hydrogen-oxidising bacteria. Although there are different types of hydrogen-oxidising bacteria, the particular strain used here is optimal for efficient protein synthesis. These bacteria use a process known as chemosynthesis, in which chemical reactions, rather than light (as in photosynthesis), provide the energy needed for growth. Hydrogen oxidation is a process involving the removal of electrons from hydrogen molecules. This reaction releases energy that bacteria use to fix carbon dioxide, converting it into organic compounds, including proteins. [53] 

Impact of hydrogen and hydrogen-oxidising microorganisms on the underground industry

Hydrogen-oxidising microorganisms not only affect the geological environment in which they are present, but also the sites within that environment. These include engineered industrial facilities such as radioactive waste disposal facilities, storage facilities for a range of fuel gases (e.g. methane, hydrogen or or town gas) and for geological storage of carbon dioxide. 

Typically, these impacts are negative, e.g., leading to increased microbial-influenced corrosion or consumption of stored gases. [54] 

The effect of hydrogen-oxidizing microorganisms on metal corrosion is such that metal corrosion (whether abiotic or microbially influenced) causes the release of hydrogen, and that this hydrogen can then become available for microorganisms to utilize. Hydrogen-consuming microorganisms including methanogens, sulfate reducers, and acetogens have been implicated in causing corrosion, as well as various classes of heterotrophic microorganisms. [54] 

Potential impacts of microbial activity include contamination of the stored gas by hydrogen sulfide, acceleration of corrosion by hydrogen sulfide production, precipitation of iron sulfide, biofilm formation, carbonate precipitation, or the production of methane. [54] 

Hydrogen-oxidizing microorganisms also have an impact during the disposal of radioactive waste. The main concerns with regard to hydrogen generation are the impact on the structural integrity of the repository, the potential for the hydrogen to act as a “carrier” for other radioactive gaseous species (e.g., 222Rn and 14C-bearing species such as CO2 and CH4) present within the waste inventory, and damage to the engineered barrier and adjacent host rock through over-pressuring. [54] 

Studies undertaken in several underground research laboratories in geologies relevant to radioactive waste disposal revealed the presence of native microbial communities heavily dependent on hydrogen metabolism [55, 56, 57, 58]. Hydrogen injection experiments at the granitic Äspö Hard Rock Laboratory indicate hydrogen stimulates the microbial community, increasing microbial growth rates, and increasing sulfide and acetate production [54, 59] 

Hydrogen-oxidising microorganisms in different geological environments

As discussed earlier in this article, hydrogen-oxidising microorganisms can be found in different geological environments, here are examples of some of the bacterial strains found in different environments:

• Sources associated with oil production, these include, for example, an oil producing well, oil pipeline water, oil-polluted estuarine sediment, etc.: 

Desulfosarcina alkanivorans strain PL12

Desulfosarcina cetonica strain 480

Desulfosarcina widdelii strain PP31

• Deep-sea sources, e.g. microorganisms isolated from a deep-sea hydrothermal vent chimney: 

Desulfurobacterium thermolithotrophum strain BSA

Desulfurobacterium thermolithotrophum strain HR11

Caminibacter hydrogeniphilus strain AM1116

Desulfonauticus submarinus strain 6N 

Balnearium lithotrophicum strain 17S

• The black smokers: 

Thermovibrio ammonificans strain HB-1

• Marine mud, marine sediment, marine sediment: 

Desulfobacter hydrogenophilus strain AcRS1

Acidianus infernus strain So4a

Desulfocicer niacini strain NAV-1

Desulfococcus sp strain Hy5 

Desulfonema ishimotonii strain DSM 9680

Desulfonema ishimotonii strain Jade 02

Desulfosarcina ovata strain OXyS1

Desulfosarcina ovata subsp. Sediminis strain 28bB2T

Desulfosarcina variabilis

Escherichia coli strain CCUG 45711 B

Paenibacillus sp strain DSM 26193

Desulforapulum autotrophicum strain HRM2

Hydrogenovibrio marinus strain DSM 11271

• Hot spring and thermal spring, and solfataric sea floor sediment;

Hydrogenobacter Thermophilus strain TK-6

Hydrogenobacter hydrogenophilus strain Z-829

Pyrococcus furiosus strain Vc 1

Aquifex pyrophilus strain Kol5a

Kyrpidia tusciae strain T2

Metallosphaera sedula strain TH2

Thermocrinis ruber strain OC 14/7/2

Thermocrinis ruber strain OC 1/4

Thermocrinis minervae strain CR11

• Bacteria isolated from human or animal organisms or waste products, e.g. human and animal faeces, from termite gut, from bovine rumen, etc.; 

Wolinella succinogenes strain DSM 1740

Mycobacterium Gordonae strain W-1609

• Freshwater sediment, freshwater mud, mud and sediments of lakes and rivers, etc.; 

Acetobacterium woodii strain DSM 1030

Clostridium aceticum strain DSM 1496

Hydrogenophaga flava DSM 619

Roseateles saccharophilus strain DSM 654

Herbaspirillum autotrophicum strain SA 32

Herbaspirillum autotrophicum strain SA 33

Xantobacter autotrophicus strain 7C

Hydrogenibacillus schlegelii strain MA-48

Hydrogenibacillus schlegelii strain MA-51

Legionella nagasakiensis strain CDC-1796-JAP-E

Legionella sainthelensi strain MSH-4

Azotobacter chroococcum strain DSM 369

Thermoanaerobacter kivui strain LKT-1

Cupriavidus necator strain H 1

• Waste water, sewage sludge, bioreactors, a pond polluted with sewage water; 

Acetobacterium wieringae strain С

Methanosarcina barkeri strain MS

Afipia carboxidovorans strain OM5

Desulfococcus multivorans

Desulfococcus biacutus strain KMRActS

Hydrogenibacillus schlegelii strain 71 OMT1

Hydrogenibacillus schlegelii strain 73 OMT4

Hydrogenibacillus schlegelii 74 OMT7

Escherichia coli strain DSM 19683

• Soil including compost soil (compost soil);

Wautersia eutropha strain 335

Paracoccus denitrificans strain CCUG 30144

Acidovorax facilis strain DSM 649

Variovorax paradoxus strain 351

Azotobacter vinelandii strain 3a

Azotobacter salinestris strain 184

Azotobacter chroococcum strain Ag

Azotobacter beijerinckii strain S 16

Azotobacter beijerinckii strain A 17

Cupriavidus necator strain N-1

• Sources associated with gas and ore deposits, e.g. microorganisms isolated from water extracted from a coal bed, from a uranium mine, from gas-associated formation water from a well of a natural gas field, etc. ;

Acidithiobacillus ferrooxidans strain DSM 14882

• Plants;

Pseudomonas palleroniana strain PRM16

Methanobrevibacter arboriphilus strain DH-1

Mycobacterium smegmatis strain GA 735

Bradyrhizobium japonicum strain 3I1b6

Future Research

Interest in hydrogen-oxidising microorganisms is currently growing more and more. Since they have found their application in various fields of biotechnological industry. But first of all the interest to these microorganisms is caused by the fact that they are a reference point for searching and determining the size of the natural hydrogen resource underlying them, because hydrogen is considered as a clean and environmentally friendly fuel of the future, the only by-product of the reaction of conversion of hydrogen into electricity is water. Natural hydrogen has been found in many geological environments, including igneous rocks, ophiolites, rift zones, geysers, hot springs, coal seams, deep faults, "subsidence ring structures", etc.

Considerable evidence is now accumulating on the existence of microbial communities capable of utilising and producing hydrogen in the subsurface. 

Although the world of hydrogen-oxidising microorganisms is fairly well understood to date, some questions require further research, for example: 

• How hydrogen exposure changes microbial community composition in the long term.  

• What impact does an increase in microbial community abundance and activity have on the subsurface hydrogen cycle.  

• The rate of microbial community growth in natural environments.  

• Developing ways of sampling difficult to access environments, e.g. deep water springs, etc. 

Hydrogen concentrations are often kept low by microbial consumption processes, which may hide an active hydrogen economic. Understanding how hydrogen-oxidising microorganisms influence hydrogen concentrations in geological environments will help to answer how much of hydrogen's potential as an energy resource is hidden in the interior of our planet. And answers to the questions of how hydrogen and hydrogen-oxidising microorganisms affect underground subsurface will help in solving problems with the exploration, storage and transport of hydrogen as an energy resource.

HydITEx is developing a geological exploration concept for detailed searches over large areas for hydrogen deposits, where the likelihood of their geological discovery has received sufficient geological justification, by analyzing the biomass of hydrogen bacteria, the presence of noble gases and their isotopes, as well as their composition.

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