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(Special Feature: MDM)”
By Keya

Metagenomics analysis is a revolutionary technique that enables the isolation,
observation, and study of microbial DNA present in various environmental or biological
samples. This innovative approach serves as a valuable tool for assessing microbial
diversity and functional variability within existing microbial communities.
Metagenomics represents a groundbreaking paradigm shift in genomic analysis,
offering the potential to tap into the vast reservoir of novel genes present in diverse
environments, particularly soil. This exploration involves the extraction of DNA from
environmental samples, followed by cloning into suitable vectors and transformation into
competent E. coli cells. The resulting transformants, found in metagenomic libraries, are
screened for novel physiological, metabolic, and genetic traits. Despite its timeconsuming and labor-intensive nature, metagenomics stands as a potent environmental
approach, capable of unveiling novel genes and biomolecules through the expression of
genes sourced from uncultivated and unidentified bacteria in host cells. Ideally, a
comprehensive metagenomic database should encompass DNA sequences for all
genes within the microbial community. However, in practice, many of these genes face
challenges related to expression, folding, or correct excretion within the host system.

MDM(MICROBIAL DARK MATTER): A Repository of
Surprises
The study of Single-Cell Amplified Genomes (SAGs) and Metagenome-Assembled
Genomes (MAGs) has revealed astonishing revelations about microbial diversity in the
natural world, leading to significant discoveries. Over the past decade, researchers
have unearthed numerous MAGs and SAGs representing major microbial lineages from
environmental samples. Several seminal papers have significantly advanced the field of
systematic and evolutionary microbiology. For instance, a study by Hug et al. harnessed
over a thousand MDM genomes to reconstruct the tree of life, unveiling a hyper-diverse
Unlocking the Enigmatic Realm of Microbial Life: Metagenomics (Special Feature: MDM)” 2
clade known as the Candidate Phyla Radiation (CPR), which further subdivided the
bacterial domain. Parks et al. made significant contributions by recovering nearly 8000
MAGs from over 1500 metagenomic datasets, expanding the prokaryotic phylogenetic
diversity by more than 30%, thereby introducing 20 novel phyla of Bacteria and
Archaea. Recent research has leveraged MAG and SAG datasets to explore Asgard
archaea, leading to exciting models explaining the origin of eukaryotic cellular
complexity, such as the entangle-engulf-endogenize model for eukaryogenesis
proposed through the study of 'Candidatus Prometheoarchaeum syntrophicum.'
These endeavors into MDM studies are fundamentally reshaping our understanding of
life's origins and evolution. Another intriguing avenue of research delves into the
metabolic versatility of MDM, revealing significant surprises in recent years. For
example, genomic analysis of MDM has unveiled genes encoding enzymes for
anaerobic methane oxidation and dissimilatory sulfate reduction within Korarchaeota
MAGs, suggesting the coupling of these two functions within a single organism, a
phenomenon typically involving different partners in syntrophic relationships.
Furthermore, the functional diversity of MDM opens new doors for applications in
biomedicine, bioenergy, biotechnology, bioremediation, and more. This versatile aspect
of MDM research surprises us and enhances our understanding of the complex world
we inhabit.

Challenges and Opportunities
Despite the strides made in MDM research, several significant challenges remain on the
horizon. The advent of easily accessible DNA sequencing has generated an abundance
of genomic data for investigating MDM, transitioning the field from a data-poor past to a
data-rich era. As the volume of genomic data in public databases continues to grow,
more MDM discoveries and investigations will ensue. Navigating the post-genomic era
poses challenges, and addressing these challenges is essential.

The first significant challenge is communication. Currently, communicating about MDM
presents difficulties due to the instability and fluidity of their taxonomy and
nomenclature. The International Code of Nomenclature of Prokaryotes (ICNP), which
governs bacterial and archaeal nomenclature, does not encompass uncultivated
microorganisms, creating obstacles in scientific communication. The instability and
synonymy within microbial nomenclature lead to confusion across scientific domains,
affecting fields like agriculture, law, biotechnology, and medicine. Therefore, there is an
Unlocking the Enigmatic Realm of Microbial Life: Metagenomics (Special Feature: MDM)” 3
urgent need for a structured system of rules governing the nomenclature, compilation,
and communication of MDM. Suggestions include using genome sequences as type
material for taxonomic descriptions and establishing an independent nomenclatural
system for MDM. While proposals have been made, community engagement remains
crucial in implementing these recommendations. The Bergey's International Society for
Microbial Systematics (BISMiS) meeting in November 2022 was to address the
nomenclature of uncultivated microorganisms, with the objective of reaching
international consensus.

The Future of MDM Research
The Earth teems with an immense diversity of microbes, with the majority remaining
uncultivated and their ecological functions poorly understood. To further MDM research,
three key approaches are advocated. Firstly, large-scale investigations integrating
various omics approaches, such as metagenomics, metatranscriptomics,
metaproteomics, metabolomics, and culturomics, are imperative to explore a wide range
of natural environments, particularly extreme habitats. These investigations will unveil
the taxonomic and functional diversity of microbial life. Secondly, establishing stronger
connections between species and their in situ functions, which can be studied through a
combination of omics techniques, fluorescence in situ hybridization (FISH), metabolic
activity rate (MAR) measurements, NanoSIMS, and other advanced imaging techniques.

This will provide a better understanding of the ecological roles of MDM and their interactions with other organisms in their natural environments. Lastly, fostering interdisciplinary collaborations among scientists from various fields, including microbiology, ecology, bioinformatics, and computational biology, will promote comprehensive and holistic approaches to MDM research.

Conclusion

In conclusion, the field of Metagenomic Dark Matter (MDM) research has revolutionized
our understanding of microbial diversity and functional variability. Through the study of
Single-Cell Amplified Genomes (SAGs) and Metagenome-Assembled Genomes
(MAGs), we have uncovered astonishing revelations about microbial lineages,
metabolic versatility, and their impact on the environment. However, challenges such as
communication and taxonomy remain, requiring collaborative efforts and international
consensus to address. Moving forward, integrating diverse omics approaches, studying
Unlocking the Enigmatic Realm of Microbial Life: Metagenomics (Special Feature: MDM)” 4
in situ functions, and fostering interdisciplinary collaborations will drive further
advancements in MDM research. By unraveling the mysteries of MDM, we will gain
deeper insights into the complex world of microbial life and its implications for various
scientific fields

(These are the personal views of the author.)