Intricacies of assessing the human microbiome in epidemiologic studies

Abstract

Purpose In the past decade, remarkable relationships have been documented between dysbiosis of the human microbiota and adverse health outcomes. This review seeks to highlight some of the challenges and pitfalls that may be encountered during all stages of microbiota research, from study design and sample collection, to nucleic acid extraction and sequencing, and bioinformatic and statistical analysis. Methods Literature focused on human microbiota research was reviewed and summarized. Results While most studies have focused on surveying the composition of the microbiota, fewer have explored the causal roles of these bacteria, archaea, viruses, and fungi in affecting disease states. Microbiome research is in its relatively early years and many aspects remain challenging, including the complexity and personalized aspects of microbial communities, the influence of exogenous and often confounding factors, the need to apply fundamental principles of ecology and epidemiology, the necessity for new software tools, and the rapidly evolving genomic, technological, and analytical landscapes. Conclusions Incorporating human microbiome research in large epidemiological studies will soon help us unravel the intricate relationships that we have with our microbial partners and provide interventional opportunities to improve human health.

Opportunities and Challenges for Environmental Exposure Assessment in Population-Based Studies

“…Recently, standardized techniques have expanded the study of the microbiome into large-scale studies (52) (Table 1). Establishing norms in large cohorts is an important first step to enable links of multiple exposures to changes in the microbiome and ultimately to long term health outcomes (53).…”

Insights into study design and statistical analyses in translational microbiome studies

“…On the other hand, if the goal is to monitor disease severity or treatmentresponse, an appropriate design would incorporate temporally separated samples and repeated measurements from the same study subject. The frequency of sample collection in temporal study designs is often determined by factors such as the monetary resources budgeted for sample collection and storage, invasiveness of the sampling procedure, subject compliance to study protocol and in the case of retrospective studies, availability of samples from a pre-existing biorepository (40). Samples retrieved from a biorepository might not be uniformly separated in time, an aspect that needs to be accounted for while interpreting study findings.…”

“…A large body of work in the field has addressed day-to-day operations (38) involved in performing microbiome studies in research and epidemiological settings (39,40) including sample collection and storage (41)(42)(43), laboratory protocols for sample processing (40,44), choice of microbiome sequencing protocols (45) and the computational infrastructure required for data processing and storage (46,47). This review builds on these discussions while specifically focusing on the design of translational microbiome studies and development of downstream statistical analysis plans which answer complex questions in a translational setting and inform early and late-phase clinical trials.…”

Maternal–Child Microbiome

“…As with any sample collected for assessing the microbiome, the method of gut microbiome collection should maximize the recovery of fecal microbial DNA while minimizing sampling bias and contamination (Robinson et al, 2016). A self-collected fecal sample is the most common form of specimen collection and either involves a full fecal sample or some variation of swabbing (e.g., swabbing the rectum, swabbing the fecal sample, swiping feces onto a collection card).…”

“…A self-collected fecal sample is the most common form of specimen collection and either involves a full fecal sample or some variation of swabbing (e.g., swabbing the rectum, swabbing the fecal sample, swiping feces onto a collection card). It is recommended that any new swab sampling method be validated in pilot studies before being employed in larger scale studies (Robinson et al, 2016). Participants should be provided the method-specific collection tools (Table 1) and instructed on the self-collection procedure: either self-swab of the rectum or stool collection using disposable trays, ensuring no urine is collected with the stool sample (McInnes & Cutting, 2010).…”