Microbial populations contribute a lot to the flourishing of particular communities. One of the main components of microbiome studies is the identification of all taxa present in the microbiome and determine their relative abundance. Absolute quantitation of microbial groups is one of the important aspects of microbial ecology. This information aids in understanding functional groups and preparing microbial interaction models in a particular microbial community. DNA Barcoding is a process of identifying organisms up to the species level by means of sequencing short sequences of DNA from specific genes. There are two requirements for establishing DNA barcodes namely, (1) it should be highly conserved meaning that it is present in all species and does not easily change or evolve, and (2) it should possess high levels of variability in order to separate organisms up to the species level.

Phylogenetics studies have revealed that groups of organisms that share a common phylogenetic lineage have conserved sequences that can be used to identify unknown organisms. In microbiology, one of the methods used to barcode microbial groups is by means of sequencing the ribosomal DNA or RNA. DNA barcoding is also done in order to determine the abundance of particular microbial taxa in a metagenome.

Ribosomal RNA sequencing, particularly 16S, 18S, and Internal Transcribe Spacer (ITS) gene sequencing is commonly used for the barcoding of microorganisms in nature. These gene sequences are used in the identification, classification, and quantitation of microbes found in complex biological mixtures such as environmental samples and gut microbiomes in order to determine the relative abundance of different microbial groups.

16S and 18S rRNA are commonly used for phylogenetic studies because these regions are highly conserved and have a very low rate of mutation, and a slow evolution.16S rRNA is a component of the 30S subunit of a prokaryotic ribosome while 18S rRNA is the homolog in microbial eukaryotes. Universal primers for these genes are readily available for phylogenetic studies on bacteria, archaea, and fungi.

16S rRNA gene sequence is one of the most common housekeeping genetic markers present in all bacteria and archaea which long enough to accurately establish taxonomic relationships and identify unknown organisms. In addition to this, it can also be used to characterize non-culturable bacteria.

18S rRNA gene sequence on the other hand is a common DNA barcode for fungi and other eukaryotic microbes. It presents a higher accuracy of identification compared to the 28S rRNA that is why there are more available data for 18S rRNA sequencing. Another DNA barcode for microorganisms is the Internal Transcribed Spacer (ITS). ITS are the regions within the ribosomal transcript that are excised and degraded during maturation. ITS genes are said to have higher hypervariability than ribosomal RNA is commonly used to identify fungi, particularly in environmental samples.

Establishing microbial identity and density in a particular environment gives a preview of the functionality of the community and aids in designing interaction models. Most of the available sequencing technologies can be used for absolute quantitative sequencing and DNA barcoding studies. Different databases are also available for bacteria, archaea, and fungi, to guide researchers in determining the identity of the members of particular microbial groups.

References:

Barlow JT, Bogatyrev SR, Ismagilov RF. A quantitative sequencing framework for absolute abundance measurements of mucosal and luminal microbial communities. Nature Communications, 2020, 11:2590.Banos S, Lentendu G, Kopf A, et al. comprehensive fungi-specific 18S rRNA gene sequence primer toolkit suited for diverse research issues and sequencing platforms. BMC Microbiology, 2018, 18:190.Tkacz A, Hortala M, Poole PS. Absolute quantitation of microbiota abundance in environmental samples. Microbiome, 2018, 6:110.