Gene-targeted metagenomic analysis of glucan-branching enzyme gene profiles among human and animal fecal microbiota

Abstract

Glycoside hydrolases (GHs), the enzymes that breakdown complex carbohydrates, are a highly diversified class of key enzymes associated with the gut microbiota and its metabolic functions. To learn more about the diversity of GHs and their potential role in a variety of gut microbiomes, we used a combination of 16S, metagenomic and targeted amplicon sequencing data to study one of these enzyme families in detail. Specifically, we employed a functional gene-targeted metagenomic approach to the 1-4-a-glucan-branching enzyme (gBE) gene in the gut microbiomes of four host species (human, chicken, cow and pig). The characteristics of operational taxonomic units (OTUs) and operational glucan-branching units (OGBUs) were distinctive in each of hosts. Human and pig were most similar in OTUs profiles while maintaining distinct OGBU profiles. Interestingly, the phylogenetic profiles identified from 16S and gBE gene sequences differed, suggesting the presence of different gBE genes in the same OTU across different vertebrate hosts. Our data suggest that gene-targeted metagenomic analysis is useful for an in-depth understanding of the diversity of a particular gene of interest. Specific carbohydrate metabolic genes appear to be carried by distinct OTUs in different individual hosts and among different vertebrate species' microbiomes, the characteristics of which differ according to host genetic background and/or diet.

Evaluation of inter-individual differences in gut bacterial isoflavone bioactivation in humans by PCR-based targeting of genes involved in equol formation

“…), 1,4‐ α ‐glucan‐branching enzyme (Lee et al . ), choline trimethyl lyase (Martinez‐del Campo et al . ), lactoyl‐CoA dehydratase, propionaldehyde dehydrogenase and methylmalonyl‐CoA decarboxylase (Reichardt et al .…”

“…Since relationships between phylogeny and activities of bacteria are usually difficult to establish, targeting of functional genes are more meaningful. Targeted metagenomic analysis has been described for a number of important enzymes occurring in the intestinal microbiome of humans, such as 7a-hydroxy-3-oxo-D 4cholenoic acid oxidoreductase (Wells et al 2003;Kang et al 2008), butyryl-CoA:acetate CoA-transferase (Louis and Flint 2007;Louis et al 2010), b-glucuronidases (Dabek et al 2008;McIntosh et al 2012), 1,4-a-glucanbranching enzyme (Lee et al 2014), choline trimethyl lyase (Martinez-del Campo et al 2015), lactoyl-CoA dehydratase, propionaldehyde dehydrogenase and methylmalonyl-CoA decarboxylase (Reichardt et al 2014). The challenge of this approach is to develop appropriate degenerate PCR primers, which essentially target all genes encoding a specific key enzyme in different bacterial species.…”

“…Ideally, this analysis would also capture still unknown equol-forming species. Such function-oriented metagenomic methodologies have been successfully developed for the identification and quantification of key bacterial genes in the human intestine (Wells et al 2003;Louis and Flint 2007;Dabek et al 2008;Kang et al 2008;Louis et al 2010;McIntosh et al 2012;Lee et al 2014;Reichardt et al 2014;Martinez-del Campo et al 2015).…”

Culture-dependent and metagenomic analysis of lesser horseshoe bats’ gut microbiome revealing unique bacterial diversity and signatures of potential human pathogens

“…However, there are limitations in culture-based methods to discover novel and potentially pathogenic bacteria and their interactions with the environment, so the use of culture-independent methods is essential to study the polutation structure, genetic diversity, as well as ecological roles of most microorganisms [18]. Metagenomic studies have used 16S rRNA gene sequencing to characterize gut microbiomes and microbes in the guano of several bat species [19,20] and the feces of other animals to determine their diet and the diversity of metabolic functional genes and enzymes [21,22].…”

Bacterial Glycogen as a Durable Energy Reserve Contributing to Persistence: An Updated Bibliography and Mathematical Model

“…In the context of a study of Agrobacterium tumefaciens, Bains simply stated that short g c glycogen could be utilized slowly due to steric inhibition and promote bacterial survival under starvation (8). Finally, in a targeted metagenomic analysis of glucan-branching enzyme genes in fecal microbiota, Lee et al emphasized that more work should be done on the effects of glycogen average chain length in order to better understand the functions of glycogen-degrading enzymes, i.e., how g c links with degradation rate (9).…”