The mysterious archaea discovered by accident can "revive" old oil fields

Methanogens are a type of bacteria that use methane as a specific metabolite under anaerobic conditions. Their metabolic products are methane and carbon dioxide.

Humans have known methanogens for a long time

Microorganisms that can produce methane are called methanogens. These organisms belong to the archaea of ​​prokaryotes. They are anaerobic bacteria that grow and reproduce very slowly and are difficult to culture and isolate. Methanogens can only be found in an environment that is completely lacking in oxygen. Only methanogenesis and fermentation can occur when there are only carbon-containing compounds as electron acceptors. Through methanogenesis, organic waste can be converted into useful, colorless, slightly sour and flammable gas methane (biogas). Methanogenesis also occurs in the intestines of humans and animals.

Methanogens are very old, probably one of the oldest life forms on Earth. In the initial state of the Earth, the special environment allowed methanogens to be born. They do not need oxygen, but can easily breathe and maintain life with simple substances such as carbonates and formates. They have life entities - cells, and began to reproduce naturally. They are the ancestors of life.

Archaea - the "ancestor" of life on Earth

Today, the earth is no longer what it was before, but methanogens still maintain their nature and are still anaerobic. Today, methanogens have a wide range of "food" sources, such as weeds, straw, leaves, leftovers from the canteen, animal feces and urine, and even garbage, which are the best food for methanogens. The bottom of swamps and ponds with dense aquatic plants is generally extremely lacking in oxygen, and methanogens often hide here to eat and exhale at the same time, which is the so-called biogas bubble.

Methane bacteria grow very slowly. Under artificial culture conditions, it takes more than ten days or even dozens of days to grow colonies. Some methane bacteria need to be cultured for 70 to 80 days to grow colonies, and even longer under natural conditions. The colonies are also quite small, especially the colonies of Methanosarcina, which are even smaller and can be easily missed if not carefully observed. The colonies are generally round, transparent, with neat edges, and emit strong fluorescence under a fluorescence microscope.

The reason why methane bacteria grow slowly is that they have very few substances to use, and can only use very simple substances, such as carbon dioxide, hydrogen, formic acid, acetic acid and methylamine. These simple substances must be provided to methane bacteria by other fermentative bacteria after decomposing complex organic matter, so methane bacteria must wait until other bacteria grow in large quantities before they can grow. At the same time, the generation time of methane bacteria is long. Some bacteria reproduce a generation in 20 minutes, while methane bacteria need several days or even dozens of days to reproduce a generation.

Methane bacteria cannot survive in the presence of oxygen, they can only survive in completely oxygen-deficient environments, such as wetland soils, animal digestive tracts, and underwater sediments. Methane production can also occur in places where neither oxygen nor decaying organic matter exists, such as deep underground, deep-sea hot water vents, and oil depots.

Methanogenic bacteria are the final methane producers in biogas fermentation and one of the key bacteria in biogas fermentation. The activity of this group of bacteria and their synergistic relationship with other bacteria participating in biogas fermentation are the key to whether biogas fermentation is normal or not.

The finishing touch to the "revival" of old oil fields

As early as the end of the 20th century, German scientists first reported in the internationally renowned journal Nature that petroleum hydrocarbons can be degraded into methane by anaerobic microorganisms. However, this biodegradation process is similar to traditional biogas fermentation, requiring multiple different types of bacteria and archaea to complete through symbiotic metabolism.

In 2008, Canadian scientists reported in Nature that such mixed bacterial communities also exist in oil reservoirs to degrade crude oil and produce methane.

In the past, it was believed that methanogenic archaea could only produce methane through four pathways: acetic acid fermentation, carbon dioxide reduction, methyl cleavage, and oxygen methyl conversion. The substrates they can use are very simple, mainly one-carbon or two-carbon compounds.

There are many oil extraction technologies that have been put into use, but it is difficult to drive the migration of all the crude oil in underground reservoirs using traditional crude oil extraction technology, and more than half of the crude oil still cannot be extracted. Scientists believe that anaerobic microorganisms that can survive in the reservoir environment may become helpers for humans. Using the principle of biogas fermentation to degrade liquid crude oil into gaseous methane and form oil and gas co-extraction is a path that scientists are committed to exploring.

The Energy Microbiology Innovation Team of the Biogas Science Research Institute of the Ministry of Agriculture and Rural Affairs of my country (hereinafter referred to as the Biogas Science Institute) cooperated with Shenzhen University, the Max Planck Institute for Marine Microbiology in Germany, the Sinopec Key Laboratory of Microbial Oil Recovery and other institutions to discover a new type of methanogenic archaea from oil reservoirs. It can directly oxidize long-chain alkyl hydrocarbons in crude oil to produce methane under anaerobic conditions. This breaks through the traditional perception that methanogenic archaea can only grow using simple compounds and expands the understanding of the carbon metabolic function of methanogenic archaea.

This research has improved the biogeochemical process of carbon cycle and laid a scientific foundation for the biogasification and exploitation of residual crude oil in depleted oil reservoirs - "underground biogas engineering".

Anaerobic microorganisms are the most numerous and species-rich biological resources on Earth. However, due to technical reasons, less than 0.1% of anaerobic microorganisms have been isolated and identified, and most of them are still "microbial dark matter." Scientists know that they exist, but they don't know what kind of existence they are.

my country's Biogas Research Institute has been engaged in basic and applied research on anaerobic microorganisms. For decades, it has preserved the largest number of anaerobic microbial model species in China, nearly 600 species (there are more than 2,000 species in the world). It is the main setter of biogas engineering standards in my country and has developed, designed and built a series of representative household and large and medium-sized biogas projects. This has laid a scientific research foundation for finding microorganisms that can directly degrade petroleum hydrocarbons and also laid the foundation for the "resurrection" of old oil fields.

Mysterious Archaea Discovered by Accident

Methanogenic archaea are unique anaerobic microorganisms that are sensitive to oxygen and usually die after being exposed to air for a few minutes. It is called "archaea" because this unique life existed on Earth as early as 3.5 billion years ago. It has many titles: one of the earliest life forms on Earth, a major contributor to global atmospheric methane emissions, and a key functional microorganism in the biogas fermentation process.

In 2019, after decades of preliminary work, researchers from my country's Institute of Biodiversity Science obtained a culture that produces methane from long-chain petroleum hydrocarbons. It can directly degrade long-chain alkanes from C13 to C34, as well as cyclohexane and cyclohexylbenzene with side chain alkanes greater than 13.

Almost at the same time, foreign scientists proposed the idea that there may be new archaea in nature that can directly degrade alkanes to produce methane, but there is no evidence to support it. When Chinese scientists reanalyzed the culture found in 2019, they did find traces of it, and the abundance was very high. However, there are many kinds of microorganisms in this culture, and evidence from multiple different dimensions is needed.

Scientists first confirmed through stable carbon isotope labeling experiments that the added n-alkanes were almost completely converted into methane and carbon dioxide. Further microbiological analysis revealed a new type of archaea that has a complete metabolic pathway for hydrocarbon degradation and methane production, and has a very high ability to produce methane, which does not require mutual metabolism. This type of archaea completes the decomposition "work" that requires multiple bacteria and archaea to work together in mutual metabolism. The researchers therefore proposed a fifth methane production pathway.

Chinese microbiologists believe that "the fifth methane production pathway has improved our understanding of the global carbon biogeochemical cycle." This also shows that there are abundant unknown microorganisms under reservoir conditions, and they have different functions. Some of these microorganisms degrade crude oil in different ways and convert it into methane or natural gas.

Diversified application prospects

Traditional crude oil extraction technology mainly uses chemical substances or water pressure to drive the migration of crude oil deep underground. This technology of oil extraction using physical and chemical methods still leaves more than half of the crude oil in underground reservoirs, making it difficult to be mined and utilized. Based on this research result related to methanogens, it will be possible to utilize the role of underground anaerobic microorganisms to degrade liquid crude oil into gaseous methane, forming oil and gas co-extraction, and ultimately achieving a relatively high crude oil extraction and utilization rate. It can also extend the development life of oil reservoirs, and is expected to "revive" old oil fields.

This basic research from "0" to "1" has laid a theoretical foundation for people to develop "underground biogas projects". The underground oil does not need to be pumped out, but can be directly turned into gas, and the gas can be collected for methane. This is equivalent to building a biogas pool in an underground oil reservoir several thousand meters deep, forming a huge "underground biogas pool" on a square kilometer scale. Once the technical research based on this achievement is broken through, the total amount of oil and gas produced by the co-production of oil and gas in depleted oil reservoirs will reach hundreds of millions of tons, which will provide scientific support for easing my country's dependence on foreign energy and ensuring national energy security.

Since it eliminates the huge carbon emission process of crude oil extraction, refining and processing, and replaces it with a green and sustainable biological conversion process, it directly obtains methane, a clean energy, and reduces carbon emissions. This is a green, environmentally friendly and low-carbon technology. Of course, this still requires the country to make a top-level design, integrate the country's advantages, and rely on scientists and engineers to make more efforts to complete it. This new type of methanogenic archaea will likely be used as a new basic cell of synthetic biology, with a wider range of application prospects.

Author: Wang Darui (China Petroleum Exploration and Development Research Institute)