A primer and discussion on DNA-based microbiome data and related bioinformatics analyses

Abstract

The past decade has seen an eruption of interest in profiling microbiomes through DNA sequencing. The resulting investigations have revealed myriad insights and attracted an influx of researchers to the research area. Many newcomers are in need of primers on the fundamentals of microbiome sequencing data types and the methods used to analyze them. Accordingly, here we aim to provide a detailed, but accessible, introduction to these topics. We first present the background on marker-gene and shotgun metagenomics sequencing and then discuss unique characteristics of microbiome data in general. We highlight several important caveats resulting from these characteristics that should be appreciated when analyzing these data. We then introduce the many-faceted concept of microbial functions and several controversies in this area. One controversy in particular is regarding whether metagenome prediction methods (i.e. based on marker gene sequences) are sufficiently accurate to ensure reliable biological inferences. We next highlight several underappreciated developments regarding the integration of taxonomic and functional data types. This is a highly pertinent topic because although these data types are inherently connected, they are often analyzed independently and primarily only linked anecdotally in the literature. We close by providing our perspective on this topic in addition to the issue of reproducibility in microbiome research, which are both crucial data analysis challenges facing microbiome researchers.

Integrating phylogenetic and functional data in microbiome studies

“…While both data types have been leveraged to make valuable observations, they are typically analyzed independently. Yet, linking these data types is clearly required to make coherent interpretations of observed shifts in microbiome data 1,2 . For example, an enrichment of a particular microbial pathway in a group of samples (e.g., a disease or unique environment) could represent selection on multiple independent taxa which possess that pathway, which would be of significant biological interest.…”

A hitchhiker’s guide to DNA-based microbiome analysis

“…A second important point arising from this review is the need for standardization in microbiome data analyses and the complexity of achieving this. As Douglas and Langille [4] state, this has been previously addressed, highlighting the variability in results obtained with different tools. It is often the case that papers describing new bioinformatic tools display their superiority relative to existing alternatives, potentially misleading newcomers to the field that the newest tool is the best and only one to be used.…”

“…Bridging between the two types of data, often extracted from the same dataset, is typically a daunting task that has been addressed by a handful of tools only.The extent of tools and approaches to analyze microbiome data is great and may be overwhelming to researchers new to microbiome or bioinformatic studies. In their paper "A primer and discussion on DNA-based microbiome data and related bioinformatics analyses", Douglas and Langille [4] guide us through the different sequencing approaches useful for microbiome studies. alongside their advantages and caveats and a selection of tools to analyze these data, coupled with examples from their own field of research.Standing out in their primer-style review is the emphasis on the coupling between taxonomic/phylogenetic identification of the organisms and their functionality.…”

“…The extent of tools and approaches to analyze microbiome data is great and may be overwhelming to researchers new to microbiome or bioinformatic studies. In their paper "A primer and discussion on DNA-based microbiome data and related bioinformatics analyses", Douglas and Langille [4] guide us through the different sequencing approaches useful for microbiome studies. alongside their advantages and caveats and a selection of tools to analyze these data, coupled with examples from their own field of research.…”

“…In summary, Douglas and Langille [4] offer both the novice and experienced researcher a detailed guide along the paths of microbiome data analysis, accompanied by informative background information, suggested tools with which analyses can be started, and an insightful view on where the field should be heading.…”