Clarisse Marotz

The Microbiome and Human Biology

Over the past few years, microbiome research has dramatically reshaped our understanding of human biology. New insights range from an enhanced understanding of how microbes mediate digestion and disease processes (e.g., in inflammatory bowel disease) to surprising associations with Parkinsons disease, autism, and depression. In this review, we describe how new generations of sequencing technology, analytical advances coupled to new software capabilities, and the integration of animal model data have led to these new discoveries. We also discuss the prospects for integrating studies of the microbiome, metabolome, and immune system, with the goal of elucidating mechanisms that govern their interactions. This systems-level understanding will change how we think about ourselves as organisms.

The Microbiome in Posttraumatic Stress Disorder and Trauma-exposed Controls: An Exploratory Study

Objective Inadequate immunoregulation and elevated inflammation may be risk factors for posttraumatic stress disorder (PTSD), and microbial inputs are important determinants of immunoregulation; however, the association between the gut microbiota and PTSD is unknown. This study investigated the gut microbiome in a South African sample of PTSD-affected individuals and trauma-exposed (TE) controls to identify potential differences in microbial diversity or microbial community structure. Methods The Clinician-Administered PTSD Scale for DSM-5 was used to diagnose PTSD according to Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition criteria. Microbial DNA was extracted from stool samples obtained from 18 individuals with PTSD and 12 TE control participants. Bacterial 16S ribosomal RNA gene V3/V4 amplicons were generated and sequenced. Microbial community structure, &agr;-diversity, and &bgr;-diversity were analyzed; random forest analysis was used to identify associations between bacterial taxa and PTSD. Results There were no differences between PTSD and TE control groups in &agr;- or &bgr;-diversity measures (e.g., &agr;-diversity: Shannon index, t = 0.386, p = .70; &bgr;-diversity, on the basis of analysis of similarities: Bray-Curtis test statistic = –0.033, p = .70); however, random forest analysis highlighted three phyla as important to distinguish PTSD status: Actinobacteria, Lentisphaerae, and Verrucomicrobia. Decreased total abundance of these taxa was associated with higher Clinician-Administered PTSD Scale scores (r = –0.387, p = .035). Conclusions In this exploratory study, measures of overall microbial diversity were similar among individuals with PTSD and TE controls; however, decreased total abundance of Actinobacteria, Lentisphaerae, and Verrucomicrobia was associated with PTSD status.

Impacts of the Human Gut Microbiome on Therapeutics

The human microbiome contains a vast source of genetic and biochemical variation, and its impacts on therapeutic responses are just beginning to be understood. This expanded understanding is especially important because the human microbiome differs far more among different people than does the human genome, and it is also dramatically easier to change. Here, we describe some of the major factors driving differences in the human microbiome among individuals and populations. We then describe some of the many ways in which gut microbes modify the action of specific chemotherapeutic agents, including nonsteroidal anti-inflammatory drugs and cardiac glycosides, and outline the potential of fecal microbiota transplant as a therapeutic. Intriguingly, microbes also alter how hosts respond to therapeutic agents through various pathways acting at distal sites. Finally, we discuss some of the computational and practical issues surrounding use of the microbiome to stratify individuals for drug response, and we envision a future where the microbiome will be modified to increase everyones potential to benefit from therapy.

DNA extraction for streamlined metagenomics of diverse environmental samples

A major bottleneck for metagenomic sequencing is rapid and efficient DNA extraction. Here, we compare the extraction efficiencies of three magnetic bead-based platforms (KingFisher, epMotion, and Tecan) to a standardized column-based extraction platform across a variety of sample types, including feces, oral, skin, soil, and water. Replicate sample plates were extracted and prepared for 16S rRNA gene amplicon sequencing in parallel to assess extraction bias and DNA quality. The data demonstrate that any effect of extraction method on sequencing results was small compared with the variability across samples; however, the KingFisher platform produced the largest number of high-quality reads in the shortest amount of time. Based on these results, we have identified an extraction pipeline that dramatically reduces sample processing time without sacrificing bacterial taxonomic or abundance information.

Improving saliva shotgun metagenomics by chemical host DNA depletion

BackgroundShotgun sequencing of microbial communities provides in-depth knowledge of the microbiome by cataloging bacterial, fungal, and viral gene content within a sample, providing an advantage over amplicon sequencing approaches that assess taxonomy but not function and are taxonomically limited. However, mammalian DNA can dominate host-derived samples, obscuring changes in microbial populations because few DNA sequence reads are from the microbial component. We developed and optimized a novel method for enriching microbial DNA from human oral samples and compared its efficiency and potential taxonomic bias with commercially available kits.ResultsThree commercially available host depletion kits were directly compared with size filtration and a novel method involving osmotic lysis and treatment with propidium monoazide (lyPMA) in human saliva samples. We evaluated the percentage of shotgun metagenomic sequencing reads aligning to the human genome, and taxonomic biases of those not aligning, compared to untreated samples. lyPMA was the most efficient method of removing host-derived sequencing reads compared to untreated sample (8.53 ± 0.10% versus 89.29 ± 0.03%). Furthermore, lyPMA-treated samples exhibit the lowest taxonomic bias compared to untreated samples.ConclusionOsmotic lysis followed by PMA treatment is a cost-effective, rapid, and robust method for enriching microbial sequence data in shotgun metagenomics from fresh and frozen saliva samples and may be extensible to other host-derived sample types.

Culturing: Looking it up in our gut

A new, publicly available, collection of cultured bacterial species from the mouse gut, the Mouse Intestinal Bacterial Collection, opens up opportunities for deepening microbiome research. Cultured isolates allow the functions of specific species and controlled consortia to be determined through in vitro experimentation.

Are microbiome studies ready for hypothesis-driven research?

Hypothesis-driven research has led to many scientific advances, but hypotheses cannot be tested in isolation: rather, they require a framework of aggregated scientific knowledge to allow questions to be posed meaningfully. This framework is largely still lacking in microbiome studies, and the only way to create it is by discovery-driven, tool-driven, and standards-driven research projects. Here we illustrate these issues using several such non-hypothesis-driven projects from our own laboratories, including spatial mapping, the American Gut Project, the Earth Microbiome Project (which is an umbrella project integrating many smaller hypothesis-driven projects), and the knowledgebase-driven tools GNPS and Qiita. We argue that an investment of community resources in infrastructure tasks, and in the controls and standards that underpin them, will greatly enhance the investment in hypothesis-driven research programs.