The vocabulary of microbiome research: a proposal

The assemblage of microorganisms present in a defined environment. The term microbiota
was first defined by Lederberg and McCray 1] who emphasized the importance of microorganisms inhabiting the human body in health
and disease. This microbial census is established using molecular methods relying
predominantly on the analysis of 16S rRNA genes, 18S rRNA genes, or other marker genes
and genomic regions, amplified and sequenced from given biological samples. Taxonomic
assignments are performed using a variety of tools that assign each sequence to a
microbial taxon (bacteria, archaea, or lower eukaryotes) at different taxonomic levels
from phylum to species.

Metataxonomics

Metataxonomics is a term we propose and define as the high-throughput process used
to characterize the entire microbiota and create a metataxonomic tree, which shows
the relationships between all sequences obtained. While viruses are an integral part
of the microbiota, no universal viral marker genes are available to perform such taxonomic
assignments.

Metagenome

The collection of genomes and genes from the members of a microbiota. This collection
is obtained through shotgun sequencing of DNA extracted from a sample (metagenomics)
followed by assembly or mapping to a reference database followed by annotation. Metataxonomic
analysis, because it relies on the amplification and sequencing of taxonomic marker
genes, is not metagenomics. Metagenomics is the process used to characterize the metagenome,
from which information on the potential function of the microbiota can be gained.

Metagenomics was first used by Handelsman et al. 2]; however, it was in the context of what the authors called functional metagenomics,
an approach where random fragments of environmental DNA are cloned into a suitable
vector for maintenance in a surrogate host for functional screening, looking for gain
of function in the surrogate host.

Microbiome

This term refers to the entire habitat, including the microorganisms (bacteria, archaea,
lower and higher eurkaryotes, and viruses), their genomes (i.e., genes), and the surrounding
environmental conditions. This definition is based on that of “biome,” the biotic
and abiotic factors of given environments. Others in the field limit the definition
of microbiome to the collection of genes and genomes of members of a microbiota. It
is argued that this is the definition of metagenome, which combined with the environment
constitutes the microbiome. The microbiome is characterized by the application of
one or combinations of metagenomics, metabonomics, metatranscriptomics, and metaproteomics
combined with clinical or environmental metadata.

Metabolomics

This term describes the analytical approaches used to determine the metabolite profile(s)
in any given strain or single tissue. The resulting census of all metabolites present
in any given strain or single tissue is called the metabolome. Most commonly used platforms to characterize the metabolome include nuclear magnetic
resonance (NMR) spectroscopy and mass spectrometry (MS) linked to a liquid chromatography
separation system.

Metabonomics

The term is a variant of the metabolomic approach; however, it describes the approach
used to generate a metabolite profile(s) from complex systems, e.g., mammals in which
more than one strain or tissue has contributed to the total metabolite pool, for example,
fecal water, urine, or plasma. This term avoids the clumsy use of meta-metabolomics
and was first defined by Jeremy Nicholson 3].

Metatranscriptomics

This term refers to the analysis of the suite of expressed RNAs (meta-RNAs) by high-throughput
sequencing of the corresponding meta-cDNAs. This approach provides information on
the regulation and expression profiles of complex microbiomes.

Metaproteomics

First coined by Rodriguez-Valera 4] and refined by Wilmes and Bond 5], this term refers to the large-scale characterization of the entire protein complement
of environmental or clinical samples at a given point in time. The method indiscriminately
identifies proteins from the microbiota and the host/environments (metagenome). Computational
analyses afford assignments of these proteins to their biological origins. It is often
performed using liquid-chromatography-based separation coupled to mass spectrometry
for peptide identification.

Misnomers and correct usage of the terms

Misnomers are often found in studies discussing metataxonomic analyses relying on
sequencing and analysis of 16S rRNA genes. In the literature, one can find the use
of “16S survey,” “16S sequencing,” or “16S analysis,” for example. There is no such
thing as “16S.” The “S” in 16S is a non-SI unit for sedimentation rate and stands
for the Svedberg unit. The Svedberg unit offers a measure of particle size based on
its rate of travel in a tube subjected to high g force. The small subunits of the bacterial and archaeal ribosomes are 30S and comprise
one structural 16S ribosomal RNA (rRNA, ~1540 nucleotides) bound to 21 proteins. Thus,
we would like to argue that the proper terms should be “16S rRNA genes” or “16S rRNA
gene sequencing/analysis.”

Additionally, the word microflora has been used for a long time in the scientific
and medical literature. However, its definition does not justify its use to describe
microbial communities associated with human (i.e., microbiota). Its definition has
evolved over time, but remains “microscopic plants, or the plants or flora of a microhabitat.”
The origin of the definition dates back to the early 1900s. Furthermore, the definition
of the word “flora” further highlights the inappropriateness of the word microflora
in the microbiome scientific literature: “the plants of a particular region or period,
listed by species and considered as a whole” or “a work systematically describing
plants” or “plants, as distinguished from fauna.” The definition of flora dates back
to mid 1600s and has its origin in the Latin name “Flora,” the Roman goddess of flowers
and the Latin word “flor,” meaning flower. These definitions and their origins make
it obvious that “microflora” refers to plants and not microbes. While some dictionaries
are now including a third definition for microflora, “the aggregate of bacteria, fungi,
and other microorganisms normally occurring on or in the bodies of humans and other
animals: intestinal flora,” these newly added definitions are the results of over
one century of misuse of the word, driven by a limited understanding of the microbes
associated with humans. Our knowledge of microbial communities is such that the scientific
community should not continue to use the word in the scientific literature. It is
time to change, and we suggest that to describe the assemblage of microbes living
in a microhabitat we use “microbiota.” Interestingly, microflora is almost exclusively
used in the literature referring to microbial community associated with human or animal,
but rarely in those associated with the environment. We believe that microflora has
still its place in the popular literature or in a yogurt/probiotic advertisement destined
to the general public, but it does not in the scientific and medical literature.

The public, the scientific popular press, medical doctors, and other scientists need
to be educated, but this will come if the scientific community adopts a common language.
The word microbiota is adequate and appropriate to describe the composition and abundance
of microbial communities whether they inhabit the human body or the environment.

This editorial was informed from papers and other communications we have had with
colleagues. We hope that a consensus use of these terms could be adopted in the near
future. This editorial aims at stimulating a discussion and standardizing the vocabulary
of microbiome research. Microbiome will continue to strive toward a standardization of the vocabulary used in this ever-expanding
field of research.